Posts Tagged ‘Hypertension’

  1. Lungs can supply blood stem cells and also produce platelets: Lungs, known primarily for breathing, play a previously unrecognized role in blood production, with more than half of the platelets in a mouse’s circulation produced there. Furthermore, a previously unknown pool of blood stem cells has been identified that is capable of restoring blood production when bone marrow stem cells are depleted.


  1. A new drug for multiple sclerosis: A new multiple sclerosis (MS) drug, which grew out of the work of UCSF (University of California, San Francisco) neurologist was approved by the FDA. Ocrelizumab, the first drug to reflect current scientific understanding of MS, was approved to treat both relapsing-remitting MS and primary progressive MS.


  1. Marijuana legalized – research needed on therapeutic possibilities and negative effects: Recreational marijuana will be legal in California starting in January, and that has brought a renewed urgency to seek out more information on the drug’s health effects, both positive and negative. UCSF scientists recognize marijuana’s contradictory status: the drug has proven therapeutic uses, but it can also lead to tremendous public health problems.


  1. Source of autism discovered: In a finding that could help unlock the fundamental mysteries about how events early in brain development lead to autism, researchers traced how distinct sets of genetic defects in a single neuronal protein can lead to either epilepsy in infancy or to autism spectrum disorders in predictable ways.


  1. Protein found in diet responsible for inflammation in brain: Ketogenic diets, characterized by extreme low-carbohydrate, high-fat regimens are known to benefit people with epilepsy and other neurological illnesses by lowering inflammation in the brain. UCSF researchers discovered the previously undiscovered mechanism by which a low-carbohydrate diet reduces inflammation in the brain. Importantly, the team identified a pivotal protein that links the diet to inflammatory genes, which, if blocked, could mirror the anti-inflammatory effects of ketogenic diets.


  1. Learning and memory failure due to brain injury is now restorable by drug: In a finding that holds promise for treating people with traumatic brain injury, an experimental drug, ISRIB (integrated stress response inhibitor), completely reversed severe learning and memory impairments caused by traumatic brain injury in mice. The groundbreaking finding revealed that the drug fully restored the ability to learn and remember in the brain-injured mice even when the animals were initially treated as long as a month after injury.


  1. Regulatory T cells induce stem cells for promoting hair growth: In a finding that could impact baldness, researchers found that regulatory T cells, a type of immune cell generally associated with controlling inflammation, directly trigger stem cells in the skin to promote healthy hair growth. An experiment with mice revealed that without these immune cells as partners, stem cells cannot regenerate hair follicles, leading to baldness.


  1. More intake of good fat is also bad: Liberal consumption of good fat (monounsaturated fat) – found in olive oil and avocados – may lead to fatty liver disease, a risk factor for metabolic disorders like type 2 diabetes and hypertension. Eating the fat in combination with high starch content was found to cause the most severe fatty liver disease in mice.


  1. Chemical toxicity in almost every daily use products: Unregulated chemicals are increasingly prevalent in products people use every day, and that rise matches a concurrent rise in health conditions like cancers and childhood diseases, Thus, researcher in UCSF is working to understand the environment’s role – including exposure to chemicals – in health conditions.


  1. Cytomegalovirus found as common factor for diabetes and heart disease in young women: Cytomegalovirus is associated with risk factors for type 2 diabetes and heart disease in women younger than 50. Women of normal weight who were infected with the typically asymptomatic cytomegalovirus, or CMV, were more likely to have metabolic syndrome. Surprisingly, the reverse was found in those with extreme obesity.





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A Concise Review of Cardiovascular Biomarkers of Hypertension

Curator: Larry H. Bernstein, MD, FCAP


Revised 5/25/2016



While a large body of work had been done on cholesterol synthesis, HDL and LDL cholesterol, triglycerides, and lipoproteins for a quarter century, and the concept of metabolic syndrome was emerging, there was neither a unifying concept nor a sufficient multivariable approach to apply the use of laboratory markers to clinical practice.  The mathematical foundation for such an evaluation of the biological markers and the computational tools were maturing at the turn of the 20th century, and the interest in outcomes research for improved healthcare practice was maturing. In addition, there was now heavy investment in health information systems that would support emerging health networks of a rapidly consolidating patient base.  This has become important for the pharmaceutical industry and for allied health sciences to enable a suitable method of measuring the effectiveness of drug and of lifestyle changes to improve the population health.

The importance of finding biomarkers for hypertension is significant as stated above. I refer to observations in a lecture by Teresa Seeman, Ph.D., Professor, UCLA Geffen School of Medicine (1).
The missed cased of hypertension in the U.S. alone has been examined by the NHANES studies. Table  I
shows the poor identification of this serious chronic condition. The next table (Table II)*, also from NHANES  (Seeman study) looks at Allostatic Load for biomarkers using component biomarker measurement criterion cutpoints.  Table III* gives the odds ratios for mortality by Allostatic Load Score.

An explanatory problem for our difficulty with diagnosis of a number of hypertension disease “subsets” is that there is peripheral hypertension that might be idiopathic, or it might be related to coexisting diseases with both inflammatory and vascular structural dynamics nature.  In addition, this may be concurrent with pulmonary hypertension, systemic hypertension, and progressive renal disease.  This discussion is reserved for later.  As stated, the late or missed diagnosis of systemic or essential idiopathic hypertension is illustrated in the three Seeman tables (1).


Table 1

Table 2

Table 3





Table 1*. Missed cases by “self report”


vs undiagnosed

study NHANES 88-94 NHANES 99-2004 NHANES 2005-08
Hypertension %unaware  BP > 140/90 42.7 43.5 39.06











High cholesterol Chol > 220 g/dl 55.93 49.3 47.05
SR- high
Diabetes HgA1C > 6.4%      

SR- high









*modified from Seeman










Table II* USHANES: Allostatic Load – component cutpoints

Biomarker Total N High Risk Percent (%) Cutpoint
DBP (mm Hg) 15,489 1,180   7.62    90
SBP (mm Hg) 15,491 3,461 22.34  140
Pulse Rate 15,117 1,009   6.67    90
HgA1C (%) 15,441 1,482   9.60    6.4
WHR 14,824 6,778 45.72    0.94
HDL Cholesterol (mg/dl) 15,187 3,440 22.65     40
Total Cholesterol


15,293 3,196  20.90    240

*From  T. Seaman, UCLA Geffen SOM


Table III*. Odds of mortality by Allostatic Load Score.

ALS Odds Ratio
7-8 5
6 2.6
5 2.3
4 2.1
3 1.8
2 1.5
1 1.4


*From  T. Seaman, UCLA Geffen SOM


I refer to cardiovascular diseases in reference to an aggregate of diseases affecting the heart, the circulatory system from large artery to the capillary, the lungs and kidneys, excluding the lymphatics.
These major disease entities are both separate and interrelated, not necessarily found in the same combinations. However, they account for a growing proportion of illness, apart from cancers, that affect the aging population of western societies. In the discussion that follows, I shall construct a picture of the pathophysiology of cardiovascular diseases, describe the major biomarkers for the assessment of these, point out the relationship of these to hypertension, and try to develop a more targeted approach to the assessment of hypertension and related disorders.

Chronic kidney disease (CKD) is defined as persistent kidney damage accompanied by a reduction in the glomerular filtration rate (GFR) and the presence of albuminuria. The rise in incidence of CKD is attributed to an aging populace and increases in hypertension (HTN), diabetes, and obesity within the U.S. population. CKD is associated with a host of complications including electrolyte imbalances, mineral and bone disorders, anemia, dyslipidemia, and HTN. It is well known that CKD is a risk factor for cardiovascular disease (CVD), and that a reduced GFR and albuminuria are independently associated with an increase in cardiovascular and all-cause mortality.

The relationship between CKD and HTN is cyclic, as CKD can contribute to or cause HTN (3). Elevated BP leads to damage of blood vessels within the kidney, as well as throughout the body. This damage impairs the kidney’s ability to filter fluid and waste from the blood, leading to an increase of fluid volume in the blood—thus causing an increase in BP.


A cursory description of the blood circulation

The full circulation involves the heart as a pump, and the arteries and veins, comprising small and large vessels, and capillaries at the point of delivery of oxygen and capture of carbon dioxide, and of transfer of substrates to tissues.  The brain, liver, pancreas and spleen, and endocrines are not further considered here, except for a consideration on neuro-humoral peptides that have emerged in the regulation of blood pressure and are essential to the stress response. The lung and the liver are both important with respect to the exchange of air and metabolites, and both have secondary circulations, the pulmonary and the portal vascular circulations.  In the case of the lungs, the vena cava flows into the right atrium, which delivers unoxygenated blood to the lungs via the right ventricle and right pulmonary artery, which returns to the left atrium by way of the right pulmonary vein.  The blood from the left atrium that flows into the left ventricle is ejected into the aorta.  The coronary arteries that nourish the heart are at the base of the aorta.  The heart muscle is a syncytium, unlike striated muscle, and it is densely packed with mitochondria, suitable for continuous contraction under vasovagal control. This is the anatomical construct, but the physiology is still being clarified because normal function and disease are both a matter of regulatory control.

In order to understand hypertension, we have to view the heart functioning over a long period of time.
In a still frame picture, we envision the left ventricle contracts emptying the oxygenated blood into the circulation. The ejection of blood into the aorta is called systole, by which the blood is delivered by the force of contraction into the circulation.  The filling pressure is called diastole.  So we have a filling and an emptying, and heard by the stethoscope is a lub-dub, synchronously repeated.   A normal systolic blood pressure is below 120. A systolic blood pressure of 120 to 139 means you have prehypertension, or borderline high blood pressure. Even people with prehypertension are at a higher risk of developing heart disease. A systolic blood pressure number of 140 or higher is considered to be hypertension, or high blood pressure. The diastolic blood pressure number or the bottom number indicates the pressure in the arteries when the heart rests between beats. A normal diastolic blood pressure number is less than 80. A diastolic blood pressure between 80 and 89 indicates prehypertension. A diastolic blood pressure number of 90 or higher is considered to be hypertension or high blood pressure. So now we have identified a systolic and a diastolic high blood pressure. Systolic pressure increases with vigorous activity, and becomes normal when the activity resides.  The systolic blood pressure increases with age. Over time, consistently high blood pressure weakens and damages the blood vessels so affected. Moreover, changes in the body’s normal functions may cause high blood pressure, including changes to kidney fluid and salt balances, the renin-angiotensin-aldosterone system, sympathetic nervous system activity, and blood vessel structure and function.


Starling’s Law of the Heart

Two principal intrinsic mechanisms, namely the Frank-Starling mechanism and rate induced regulation, enable the myocardium to adapt to changes in hemodynamic conditions. The Frank-Starling mechanism (also referred to as Starling’s law of the heart), is invoked in response to changes in the resting length of the myocardial fibers. Rate-induced regulation is invoked in response to changes in the frequency of the heartbeat.  (3-9).

Frank and Starling (3, 4) showed that an increase in diastolic volume caused an increase in systolic performance. The stretch effect persists across a range of myocardial contractile states, but during exercise it plays only a lesser role augmenting ventricular function maximal exercise. This is because in healthy human subjects adrenergic reflex mechanisms modulate myocardial performance, heart rate, vascular impedance and coronary flow during exercise and changes in these variables can overshadow the effect of fiber stretch or even prevent an increase in end-diastolic volume during stress (5). (See you- tube (6).

According to Lakatta muscle length modulates the extent of myofilament calcium ion (Ca2+) activation (7-9).   Similarly, the fiber length during a contraction, which is determined in part by the load encountered during shortening, also determines the extent of myofilament Ca2+ activation. Therefore, the terms preload, afterload and myocardial contractile state lose part of their significance in light of current knowledge.


Biology and High Blood Pressure

Researchers continue to study how various changes in normal body functions cause high blood pressure. The key functions affected in high blood pressure include (10):

Kidney Fluid and Salt Balances

The kidneys normally regulate the body’s salt balance by retaining sodium and water and excreting potassium. Imbalances in this kidney function can expand blood volumes, which can cause high blood pressure.

Renin-Angiotensin-Aldosterone System

The renin-angiotensin-aldosterone system makes angiotensin and aldosterone hormones. Angiotensin narrows or constricts blood vessels, which can lead to an increase in blood pressure. Aldosterone controls how the kidneys balance fluid and salt levels. Increased aldosterone levels or activity may change this kidney function, leading to increased blood volumes and high blood pressure.

Sympathetic Nervous System Activity

The sympathetic nervous system has important functions in blood pressure regulation, including heart rate, blood pressure, and breathing rate. Researchers are investigating whether imbalances in this system cause high blood pressure.

Blood Vessel Structure and Function

Changes in the structure and function of small and large arteries may contribute to high blood pressure. The angiotensin pathway and the immune system may stiffen small and large arteries, which can affect blood pressure.

Two or more types of hypertension

Systemic hypertension

Idiopathic hypertension

Hypertension from chronic renal disease

Pulmonary artery hypertension

Hypertension associated with systemic chronic inflammatory disease (rheumatoid arthritis and other collagen vascular diseases)

Genetic Causes of High Blood Pressure

Much of the understanding of the body systems involved in high blood pressure has come from genetic studies. High blood pressure often runs in families. Years of research have identified many genes and other mutations associated with high blood pressure, some in the renal salt regulatory and renin-angiotensin-aldosterone pathways. However, these known genetic factors only account for 2 to 3 percent of all cases. Emerging research suggests that certain DNA changes during fetal development also may cause the development of high blood pressure later in life.

Environmental Causes of High Blood Pressure

Environmental causes of high blood pressure include unhealthy lifestyle habits, being overweight or obese, and medicines.

Other medical causes of high blood pressure include other medical conditions such as chronic kidney disease, sleep apnea, thyroid problems, or certain tumors.

The common complications of hypertension and their signs and symptoms include:


Pulse Pressure and Stroke Volume

The  pulse pressure is the difference between systolic (the upper number) and diastolic (the lower number) (11).

Systemic pulse pressure = Psystolic – Pdiastolic

The pulse pressure is 40 mmHg for a typical blood pressure reading of 120/80 mmHg.

Pulse pressure (PP) is proportional to stroke volume (SV), the amount of blood pumped from the heart in one beat, and inversely proportional to the compliance or flexibility of the blood vessels, mainly the aorta.

A low (also called narrow) pulse pressure means that not much blood is being expelled from the heart, and can be caused by a number of factors, including severe blood loss due to trauma, congestive heart failure, shock, a narrowing of the valve leading from the heart to the aorta (stenosis), and fluid accumulating around the heart (tamponade).

High (or wide) pulse pressures occur during exercise, as stroke volume increases and the overall resistance to blood flow decreases. It can also occur for many reasons, such as hardening of the arteries (which can have numerous causes), various deficiencies in the aorta (mainly) or other arteries, including leaksfistulas, and a usually-congenital condition known as AVM, pain/anxiety, fever, anemia, pregnancy, and more. Certain medications for high blood pressure can widen pulse pressure, while others narrow it. A chronic increase in pulse pressure is a risk factor for heart disease, and can lead to the type of arrhythmia called atrial fibrillation or A-Fib.


Hypertension Background and Definition

The prevalence of CKD has steadily increased over the past two decades, and was reported to affect over 13% of the U.S. population in 2004.  In 2009, more than 570,000 people in the United States were classified as having end-stage renal disease (ESRD), including nearly 400,000 dialysis patients and over 17,000 transplant recipients.  A patient is determined to have ESRD when he or she requires replacement therapy, including dialysis or kidney transplantation. A National Health Examination Survey (NHANES) spanning 2005-2006 showed that 29% of US adults 18 years of age and older were hypertensive, and of those with high blood pressure (BP), 78% were aware they were hypertensive, 68% were being treated with antihypertensive agents, and only 64% of treated individuals had controlled hypertension (12, 13). In addition, data from NHANES 1999-2006 estimated that 30% of adults 20 years of age and older have prehypertension, defined as an untreated SBP of 120-139 mm Hg or untreated DBP of 80-89 mmHg (12, 13).

Hypertension is the most important modifiable risk factor for coronary heart disease (the leading cause of death in North America), stroke (the third leading cause), congestive heart failure, end-stage renal disease, and peripheral vascular disease. The 2010 Institute for Clinical Systems Improvement (ICSI) guideline (14) on the diagnosis and treatment of hypertension indicates that systolic blood pressure (SBP) should be the major factor to detect, evaluate, and treat hypertension In adults aged 50 years and older. The 2013 joint European Society of Hypertension (ESH) (15) and the European Society of Cardiology (ESC) (16) guidelines recommend that ambulatory blood-pressure monitoring (ABPM) be incorporated into the assessment of cardiovascular risk factors and hypertension.

The JNC 7 (17) identifies the following as major cardiovascular risk factors:

  • Hypertension: component of metabolic syndrome
  • Tobacco use, particularly cigarettes, including chewing tobacco
  • Elevated LDL cholesterol (or total cholesterol ≥240 mg/dL) or low HDL cholesterol: component of metabolic syndrome
  • Diabetes mellitus: component of metabolic syndrome
  • Obesity (BMI ≥30 kg/m 2): component of metabolic syndrome
  • Age greater than 55 years for men or greater than 65 years for women: increased risk begins at the respective ages; the Adult Treatment Panel III used earlier age cut points to suggest the need for earlier action
  • Estimated glomerular filtration rate less than 60 mL/min
  • Microalbuminuria
  • Family history of premature cardiovascular disease (men < 55 years; women < 65 years)
  • Lack of exercise

The Eighth Report of the JNC (JNC 8), released in December 2013 no longer recommends just thiazide-type diuretics as initial therapy in most patients. In essence, the JNC 8 recommends treating to 150/90 mm Hg in patients over age 60 years; for everybody else, the goal BP is 140/90 (18).

Biomarkers Associated with Hypertension

The biomarkers associated with hypertension are for the most part derived from features that characterize the disordered physiology. We might first consider the measurement of blood pressure. Then it becomes necessary to analyze the physiological elements that largely contribute to blood pressure. Finally, there are several biomarkers that have loomed large as measures are myocardial function or myocardial cell death, and are also not independent of renal function, that are indicators of short term and long term cardiovascular status. Having already indicated the importance of measurement of pulse, diastolic and systolic blood pressure in the routine examination of physical status, which is related to cardiac output we shall pay attention to the pulse pressure and pulse wave velocity.    These were defined in the preceding discussion.  They are critically related to the development of hypertension and in the long term, they emerge significantly earlier than either congestive heart failure, chronic kidney disease, acute coronary syndrome, stroke, or cardio-renal syndrome.

Even though cardiovascular disease (CVD), the leading cause of death in developed countries, is not predicted by classic risk factors, there are elements of the risk factor association that need further exploration and will be dissected, such as activity level, obesity, lipids, diabetes mellitus, family history and stress.  Further analysis will point to endocrine and/or metabolic factors that drive cardiovascular risk.

In taking into account the blood pressure measurements, we consider the pulse pressure (PP) and the pulse wave velocity (PWV).  If we refer back to the stroke volume and the Law of the Heart, the systolic blood pressure (SBP) is increased with increased left ventricular output that raises the left ventricular (LV) afterload. This coincides with a decrease in diastolic pressure (DBP) that accompanies a change in coronary artery perfusion (CAP).  Thus, many studies point to increased SBP as a strong risk factor for stroke and CVD.  However, there are sufficient studies that indicate the brachial artery pulse pressure (PP) is a strong determinant of CVD and stroke, and these two elements, SBP and brachial artery PP, may be an indicator of increased arterial stiffness in hypertensive patients and the general population. Brachial PP is also a determinant of recurrent events after acute coronary syndrome (ACS) or with left ventricular hypertrophy (LVH), or the risk of CHF in the aging population, and of all-cause-mortality in the general population.  In addition, the aortic PWV calculated from the Framingham equations was a suitable predictor of CVD risk. In a classic study of arterial stiffness and of CVD and all-cause mortality in an essential hypertension cohort at the Broussais Hospital between 1980 and 1996 (19), the carotid-femoral PWV was measured as an indicator of aortic stiffness, and it was found to be significantly associated with all-cause and CVD mortality independent of previous CVD, age, and diabetes. They tested the hypothesis that aortic stiffness is a predictor of cardiovascular and all-cause mortality in hypertensive patients based on the consideration that the elastic properties of the aorta and central arteries are the major determinants of systemic arterial impedance, and the PWV measured along the aortic and aorto-iliac pathway is the most clinically relevant. They assessed arterial stiffness by measuring the PWV using  the Moens-Korteweg equation based on the increase of the square root of the elasticity modulus in stiffer arteries (20).

PWV as a Diagnostic Test

To assess the performance of PWV considered as a diagnostic test, with the use of receiver operating characteristic (ROC) curves, they calculated sensitivities, specificities, positive predictive values, and negative predictive values of PWV at different cutoff values, first to detect the presence of AA in the overall population and second to detect patients with high 10-year cardiovascular mortality risk in the subgroup of 462 patients without AA with age range from 30 to 74 years. Optimal cutoff values of PWV were defined as the maximization of the sum of sensitivity and specificity.

The main finding of the study was that PWV was a strong predictor of cardiovascular risks as determined by the Framingham equations in a population of treated or untreated subjects with essential hypertension (21). They measured the PWV from foot-to-foot transit time in the aorta for a noninvasive evaluation of regional aortic stiffness, which allows an estimate of the distance traveled by the pulse. The presence of a PWV > 13 m/s, taken alone, appeared as a strong predictor of cardiovascular mortality with high performance values (21). Their work and other studies (22, 23) established increased pulse pressure, the major hemodynamic consequence of increased aortic PWV, as a strong independent predictor of cardiac mortality, mainly MI, in populations of normotensive and hypertensive subjects.

In addition to the findings above, the PWV was found to be an independent predictor of future increase in SBP and of incident hypertension in the Baltimore study (21). The authors reported that in a subset of 306 subjects who were normotensive at baseline, hypertension developed in 105 (34%) during a median follow-up of 4.3 years (range 2 to 12 years). PWV was also an independent predictor of incident hypertension (hazard ratio 1.10 per 1 m/s increase in PWV, 95% confidence interval 1.00 to 1.30, p = 0.03) in individuals with a follow-up duration greater than the median. The authors (21) concluded that carotid-femoral PWV measured using nondirectional transcutaneous Doppler probes (model 810A, 9 to 10-Mhz probes, Parks Medical Electronics, Inc., Aloha, Oregon) could be done to identify normotensive individuals who should be targeted for the implementation of interventions aimed at preventing or delaying the progression of subclinical arterial stiffening and the onset of hypertension.  They reported that age, BMI, and MAP were independently associated with higher SBP on the last visit (Table IV); in addition, PWV was also independently associated with higher SBP on the last visit, and explained 4% of its variance. As shown in Table V, age, BMI, and MAP (p = 0.09, p = 0.009, p < 0.0001 respectively for the interaction terms with time) were predictors of the longitudinal changes in SBP. In addition, PWV was also an independent predictor of the longitudinal increase in SBP (p = 0.003 for the interaction term with time).

In addition, they report that in the group with follow-up duration greater than the median (in which all subjects remained normotensive for the first 4.3 years), beyond age (hazard ratio [HR] 1.02 per 1 year, 95% confidence interval [CI] 0.99 to 1.04, p = 0.2) and SBP (HR 1.05 per 1 mm Hg, 95% CI 1.01 to 1.09, p = 0.006), both HDL (HR 0.96 per 1 mg/dl, 95% CI 0.93 to 0.99, p = 0.02) and PWV (HR 1.10 per 1 m/s, 95% CI 1.00 to 1.30, p = 0.03) (Fig. 1) were independent predictors of incident HTN.

Their findings in a longitudinal projection indicate that PWV, a marker of central arterial stiffening, is an independent determinant of longitudinal SBP increase in healthy BLSA volunteers, and an independent risk factor for incident hypertension among normotensive subjects followed up for longer than 4 years. The study was accompanied by a commentary in the same journal that states: “Pulse wave velocity (PWV) is a simple measure of the time taken by the pressure wave to travel over a specific distance. By virtue of its intrinsic relation to the mechanical properties of the artery by the Moens–Kortweg formula (PWV=√(Eh/2)Rρ; where E is the Young’s Modulus of the arterial wall, h the wall thickness, R the end- diastolic radius and ρ is the density of blood)(20), and buoyed a number of longitudinal studies that reported on the independent predictive value of PWV measurement for cardiovascular events and mortality in various populations, PWV is now widely accepted as the ‘gold standard’ measure of arterial stiffness.




Table IV Multiple Regression Analysis Evaluating the Predictors of Last Visit SBP 21

Variable Parameter
p Value
Age (yrs) 0.32 0.06 <0.0001
Gender (men) 0.65 1.78 0.71
Race (white) −1.22 2.00 0.54
Smoking (ever) 2.48 1.61 0.12
BMI (kg/m2)* 0.61 0.22 0.006
MAP (mm Hg)* 0.60 0.08 <0.0001
PWV (m/s)* 1.56 0.38 <0.0001
Heart rate (beats/min) 0.08 0.06 0.20
Total cholesterol (mg/dl) −0.005 0.02 0.83
Triglycerides (mg/dl) −0.009 0.01 0.50
HDL cholesterol (mg/dl) −0.001 0.07 0.98
Glucose (mg/dl) −0.02 0.06 0.75









Table V Predictors of Longitudinal SBP Derived From a Linear Mixed-Effects Regression Model 21

Variable Coefficient Standardized


95% Confidence


p Value
Time (yrs) 3.14 0.14 0.61 to 5.66 0.02
Age (yrs) −0.37 0.25 −0.68 to −0.06 0.02
Age2 (yrs2)* 0.006 0.08 0.002 to 0.008 <0.0001
Gender (men) 0.61 0.03 −1.26 to 2.47 0.52
BMI (kg/m2)* 0.25 0.11 −0.01 to 0.50 0.06
MAP (mmHg)* 1.03 0.47 0.93 to 1.12 <0.0001
PWV (m/s) 0.29 0.12 −0.16 to 0.74 0.21
Time × age* 0.02 0.04 −0.002 to 0.038 0.09
Time × BMI* 0.10 0.06 0.02 to 0.183 0.009
Time × MAP* −0.08 −0.12 −0.11 to −0.05 <0.0001
Time × PWV* 0.22 0.08 0.07 to 0.36 0.003



Figure 1 21

Figure 2.21

The interest in this physiological measure is illustrated by the increasing number and diversity of research publications in this arena related to human hypertension, relating PWV to pathophysiological processes (for example, homocysteine, inflammation and extracellular matrix turnover and disorders related to hypertension, such as sleep apnea). The epidemiology, genetic associations and prognostic implications of PWV (and arterial stiffness) have also been reported as has the relationship to hemodynamics, cardiac structure and function.” (24) Furthermore, arterial stiffening may be “characterized by an increase in (central) PP and changes in the morphology of the arterial waveform, both of which can now be measured non-invasively using tonometers from commercially available devices. Wave reflection is typically characterized by aortic pressure augmentation (ΔP) and the augmentation index (ΔP/PP) (Figure 3)(24). Higher augmented pressure, as an index of wave reflection, has been linked to adverse clinical outcomes in different populations.

Figure 3.24

Analysis of the pressure waveform. The initial systolic pressure is labelled as P1 and augmented pressure ( P) is typically measured as the difference between peak pressure (P2) and P1. Augmentation index is  P/PP. PP, pulse pressure. 24

A review by Payne et al. (25) states that aortic stiffness and arterial pulse wave reflections determine elevated central systolic pressure and are associated with risk of adverse cardiovascular outcomes. This is because an impaired compensatory mechanism through matrix metalloproteinases of remodeling to compensate for changes in wall stress, possibly related to angiotensin II and inhibition of the vascular adhesion protein semicarbazide-sensitive amine oxidase, related to reduced elastin fiber cross-linking. This has implications for pharmacological agents that target age-related advanced glycation end-product cross-links. This also brings into consideration NO playing a considerable role. But they caution that the endogenous NO synthase inhibitors asymmetric dimethylarginine and L-NG-monomethyl arginine associated with clinical atherosclerosis don’t appear to be associated with arterial stiffening. The matter leaves much to be explained.  The mechanisms underlying arterial stiffness could well require insights into inflammation, calcification, vascular growth and remodeling, and endothelial dysfunction. Nevertheless, arterial stiffness is independently associated with cardiovascular outcome in most of the situations where it has been examined.  Given this train of thinking, O’Rourke (26) considers a progressive arterial dilatation with repeated cycles of stress that leads to degeneration of the arterial wall and increases the pressure wave impulse and wave velocity, augmenting the pressure in late systole. Drugs may reduce wave reflection, but have no direct effect on arterial stiffness.  However, reduction in wave reflection decreases aortic systolic pressure augmentation.  DK Arnett (26) depicts the effect of persistently elevated blood pressure in the following diagram (Figure 4).


Figure 4.26  Both transient and sustained stiffening of the artery are likely to be present in hypertension.

An initial elevation in blood pressure may establish a positive feedback in which hypertension biomechanically increases arterial stiffness without any structural change. This elevated blood pressure   might later lead to additional vascular hypertrophy and hyperplasia, collagen deposition, and atherosclerosis, and fixed elevations in arterial stiffness.  As to a genetic factor, she refers to a gene contributing to pulse pressure on chromosome 8 located at 32 cM, which also contains the lipoprotein lipase (LPL) gene which has been associated with hypertension. LPL may be an important candidate gene for pulse pressure.  She specifically identifies a relationship between genetic regions contributing to aortic compliance in African American sibships ascertained for hypertension in Figure 5 (27).  These results suggest there may be influential genetic regions contributing to aortic compliance in African American sibships ascertained for hypertension (27). Collectively, these two studies, the first to our knowledge, indicate the presence of genetic factors influencing hypertension.

Other authors state that PWV has a direct relationship to intrinsic elasticity of the arterial wall, and it is an independent predictor of CVD related morbidity and mortality, but it is not associated with classical risk factors for atherosclerosis (28).  They point out that PWV doesn’t increase during early stages of atherosclerosis, as measured by intima-media thickness and non-calcified atheroma, but it does increase in the presence of aortic calcification that occurs with advanced atherosclerotic plaque. Age-related
PWV measurement. Carotid-to-femoral PWV is calculated by dividing the distance (d) between the two arterial sites by the difference in time of pressure wave arrival between the carotid (t1) and femoral artery (t2) referenced to the R wave of the electrocardiogram.

Figure 5. Linkage of arterial compliance on chromosome 2: HyperGEN27

Widening of the pulse pressure is the major cause of age-related increase in prevalence of hypertension and is related to arterial stiffening. (28)  Commonly used points for measuring the PWV are the carotid and femoral artery because they are superficial and easy to access. Arterial distensibility is measured by the Bramwell and Hill equation (29): PWV = √(V × ΔP/ρ × ΔV), where ρ is blood density. This is shown in Figure 6.


Figure 6 28


View larger version:


Furthermore, these authors (28) report arterial stiffness increases with age by approximately 0.1 m/s/y in East Asian populations with low prevalences of atherosclerosis, but some authors have found accelerated stiffening between 50 and 60 years of age. In contrast, stiffness of peripheral arteries increases less or not at all with increasing age. Again, ageing of the arterial media is associated with increased expression of matrix metalloproteinases (MMP), which are members of the zinc-dependent endopeptidase family and are involved in degradation of vascular elastin and collagen fibers. Several different types of MMP exist in the vascular wall, but in relation to arterial stiffness, much interest has focused on MMP-2 and MMP-9.  This concludes the discussion of PP and PWV in the evolution of hypertension.


Diagnostic Biomarkers of essential hypertension.

Ioannidis and Tzoulaki (30) reviewed the literature on 10 popular ‘‘new’’ biomarkers and found that each one had accrued more than 6000 publications.1 The predictive effects of these popular blood biomarkers for coronary heart disease in the general population are listed in Table VI (31).


Table VI.* Predictive Value of New Biomarkers 30,31

Biomarker Adjusted Relative Risk (95% C.I.)
Triglycerides 0.99 (0.94–1.05)
C-reactive protein 1.39 (1.32–1.47)
Fibrinogen 1.45 (1.34–1.57)
Interleukin 6 1.27 (1.19–1.35)
BNP or NT-proBNP 1.42 (1.24–1.63)
Serum albumin 1.2 (1.1–1.3)
ICAM-1 (0.75–1.64)
Homocysteine 1.05 (1.03–1.07)
Uric acid 1.09 (1.03–1.16)

*Ionnidis and Tzoulaki from Giles
The majority of these biomarkers show small effects, if any, even in combination.  Giles (31) points out that an elevated homocysteine level might be of great importance to a young person with a myocardial infarction and a positive family history of similar occurrences. Emerging biomarkers, eg, asymmetric and symmetric dimethylarginine and galectin-3, are promising more specific biomarkers based on pathophysiologies for cardiovascular disease. Even then, blood pressure remains the biomarker par excellence for hypertension and for many other cardiovascular entities.

The importance of blood pressure was highlighted by the report of the cardiovascular lifetime risk pooling project.(10) Starting at 55 years of age, 61,585 men and women were followed over an average of 14 years, ie, 700,000 person-years. Individuals who maintained or decreased their blood pressure to normal levels had the lowest remaining lifetime risk for cardiovascular disease (22–41%) compared with individuals who had or developed hypertension by 55 years of age (42–69%). The study indicated that efforts should continue to emphasize the importance of lowering blood pressure and avoiding or delaying the incidence of hypertension to reduce the lifetime risk for cardiovascular disease

A small study involving 120 hypertensive patients with or without heart failure tried to establish a multi-biomarker approach to heart failure (HF) in hypertensive patients using N-terminal pro BNP (32). The following biomarkers were included in the study: Collagen III N-terminal propeptide (PIIINP), cystatin C (CysC), lipocalin-2/NGAL, syndecan-4, tumor necrosis factor-α (TNF-α), interleukin 1 receptor type I (IL1R1), galectin-3, cardiotrophin-1 (CT-1), transforming growth factor β (TGF-β) and N-terminal pro-brain natriuretic peptide (NT-proBNP). The highest discriminative value for HF was observed for NT-proBNP (area under the receiver operating characteristic curve (AUC) = 0.873) and TGF-β (AUC = 0.878). On the basis of ROC curve analysis they found that CT-1 > 152 pg/mL, TGF-β < 7.7 ng/mL, syndecan > 2.3 ng/mL, NT-proBNP > 332.5 pg/mL, CysC > 1 mg/L and NGAL > 39.9 ng/mL were significant predictors of overt HF. There was only a small improvement in predictive ability of the multi-biomarker panel including the four biomarkers with the best performance in the detection of HF (NT-proBNP, TGF-β, CT-1, CysC) compared to the panel with NT-proBNP, TGF-β and CT-1 (absent  CysC). The biomarkers with different pathophysiological backgrounds (NT-proBNP, TGF-β, CT-1) give additive prognostic value for incident compared to NT-proBNP alone.

Inflammation has been associated with pathophysiology of hypertension and vascular damage. Resistant hypertensive patients (RHTN) have unfavorable prognosis due to poor blood pressure control and higher prevalence of target organ damage. Endothelial dysfunction and arterial stiffness are involved in such condition. Previous studies showed that RHTN patients have higher arterial stiffness and endothelial dysfunction than controlled hypertensive and normotensive subjects. The relationship between high blood pressure levels and arterial stiffness may be explained in part, by inflammatory pathways. Previous studies also found that hypertensive subjects have higher levels of inflammatory cytokines including TNF-α, IL-10, IL-1β and CRP. Moreover, IL-1β correlates with arterial stiffness and levels of blood pressure, which are particularly high in patients with resistant hypertension. Increased inflammatory cytokines levels might be related to the development of vascular damage and to the higher cardiovascular risk of resistant hypertensive patients. Elevated BP may cause cardiovascular structural and functional alterations leading to organ damage such as left ventricular hypertrophy, arterial and renal dysfunction. TNF-α inhibition reduced systolic BP and endothelial inflammation in SHR [33]. They also found that IL-1β correlates with arterial stiffness and levels of blood pressure, even after adjust for age and glucose [33]. These investigators then demonstrated that isoprostane levels, an oxidative stress marker, were associated with endothelial dysfunction in these patients [33].

Chao et al. carried out studies of kallistatin (34-36). Kallistatin is an endogenous protein in human plasma as a tissue Kallikrein-Binding Protein (KBP). Tissue kallikrein is a serine protease that releases vasodilating kinin peptides from kininogen substrate. The tissue kallikrein-kinin system is involved in mediating beneficial effects in hypertension as well as cardiac, cerebral and renal injury. KBP was later identified as a serine protease inhibitor (serpin) because of its ability to inhibit tissue kallikrein activity, and was subsequently named “kallistatin”. Kallistatin is mainly expressed in the liver, but is also present in the heart, kidney and blood vessel. Kallistatin protein contains two structural elements: an active site and a heparin-binding domain. The active site of kallistatin is crucial for complex formation with tissue kallikrein, and thus tissue kallikrein inhibition.

Kallistatin is expressed in tissues relevant to cardiovascular function, and has consequently been shown to have vasodilating properties.  Kallistatin has pleiotropic effects in vasodilation and inhibition of inflammation, angiogenesis, oxidative stress, fibrosis, and cancer progression. Injection of a neutralizing Kallistatin antibody into hypertensive rats aggravates cardiovascular and renal injury in association with increased inflammation, oxidative stress and tissue remodeling.  Neither the blood pressure-lowering effect nor the vasorelaxation ability of kallistatin is abolished by icatibant (Hoe140, a kinin B2 receptor antagonist), indicating that kallistatin-mediated vasodilation is unrelated to the tissue kallikrein-kinin system.

The findings reported indicate that kallistatin exerts beneficial effects against hypertension and organ damage. Kallistatin levels in circulation, body fluids or tissues were lower in patients with liver disease, septic syndrome, diabetic retinopathy, severe pneumonia, inflammatory bowel disease, and cancer of the colon and prostate. In addition, reduced plasma kallistatin levels are associated with adiposity and metabolic risk in apparently healthy African American youths. Considered a negative acute-phase protein, circulating kallistatin levels as well as hepatic expression are rapidly reduced within 24 hours after Lipopolysaccharide (LPS) induced endotoxemia in mice. Similarly, circulating kallistatin levels are markedly decreased in patients with septic syndrome and liver disease. Taking together, the studies indicate that kallistatin exhibits potent anti-inflammatory activity.

The pathogenesis of hypertension and cardiovascular and renal diseases is tightly linked to increased oxidative stress and reduced NO bioavailability (37-39). Time-dependent elevation of circulating oxygen species are associated with reduced kallistatin levels in animal models of hypertension and cardiovascular and renal injury. Stimulation of NO formation by kallistatin may lead to inhibition of oxidative stress and thus multi-organ damage. On the other hand, endogenous kallistatin depletion by neutralizing antibody increased oxidative stress and aggravated cardiovascular and renal damage.

A human kallistatin gene polymorphism has been shown to correlate with a decreased risk of developing acute kidney injury during septic shock. Kallistatin levels are markedly reduced in both humans and mice with sepsis syndrome. However, kallistatin administration protects against lethality and organ injury in animal models of toxic septic shock. Moreover, kallistatin levels are decreased in patients with liver disease, septic shock, inflammatory bowel disease, severe pneumonia and acute respiratory distress syndrome. Taken together, the results indicate that kallistatin has the potential to be a molecular biomarker for patients with sepsis, cardiovascular and metabolic disorders.

Pulmonary hypertension (PH) is defined as a mean pulmonary artery pressure of .25 mmHg at rest or .30 mmHg with exercise. Right heart catheterization is required for the definitive diagnosis. Subsequent investigations are instituted to further characterize the disease. The 6-min walk test (6MWT), a measure of exercise capacity, and the New York Heart Association (NYHA)/World Health Organization (WHO) functional classification, a measure of severity, are used to follow the clinical course while receiving treatment, and these both correlate with disease severity and prognosis (43).

Pulmonary arterial hypertension (PAH) is a progressive disease of the pulmonary vasculature that leads to exercise limitation, right heart failure, and death. There is a need for biomarkers that can aid in early detection, disease surveillance, and treatment monitoring in PAH. Several potential molecules have been investigated; however, only brain natriuretic peptide is currently recommended at diagnosis and for follow-up of PAH patients.

ANP is released from storage granules in atrial tissue, while BNP is secreted from ventricular tissue in a constitutive fashion. ANP secretion is stimulated by atrial stretch caused by atrial volume overload; BNP is released in response to ventricular stretch. Natriuretic peptides act on the kidney, causing natriuresis and diuresis, and relax vascular smooth muscle, causing arterial and venous dilatation, leading to reduced blood pressure and ventricular preload. ANP and BNP are released as prohormones and then cleaved into the active peptide and an inactive N-terminal fragment (43).

Natriuretic peptide precursors are released in response to atrial and ventricular stretch, cleaved into active molecules and inactive precursors and convert guanosine 59-triphosphate (GTP) to cyclic guanosine monophosphate (cGMP), leading to their various physiological actions.

There are a number of confounding factors in the interpretation of natriuretic peptide levels, including left heart disease, sex, age and renal dysfunction. Since most studies exclude patients with left heart disease and renal dysfunction, it becomes problematic extrapolating these results to an unselected population (43).

Endothelin-1 (ET-1) is a peptide found in abundance in the human lung and, through action of endothelin receptors (ETA and ETB) on vascular smooth muscle cells, is implicated in the pathogenesis of PAH. Endothelin receptor antagonists are approved for the treatment of PAH. Levels of circulating ET-1 and related molecules are logical biomarkers of interest in PAH. ET-1 is elevated in PAH compared to controls, and correlates with pulmonary hemodynamic parameters. In addition, higher ET-1 levels are associated with increased mortality in patients treated for PAH. ET-1’s precursor, big-ET-1, has a longer half-life and hence is more stable than ET-1.

Endothelin-1 ET-1 is a potent endogenous vasoconstrictor and proliferative cytokine. The ET-1 gene is translated to prepro-ET-1 which is then cleaved, by the action of an intracellular endopeptidase, to form the biologically inactive big ET-1. ET-converting enzymes further cleave this to form functional ET-1 . There are two ET receptor isoforms, termed type A (ETA), located predominantly on vascular smooth muscle cells, and type B (ETB), predominantly expressed on vascular endothelial cells but also on arterial smooth muscle. Activation of both receptor subtypes, when located on vascular smooth muscle, results in vasoconstriction and cell proliferation. In addition, the endothelial ETB receptor mediates vasodilatation and clearance of ET-1 (43).

Prepro-ET-1 is cleaved to inactive big ET-1 and then further cleaved to form active ET-1. This acts on vascular smooth muscle via the ETA and ETB receptors, causing vasoconstriction and cell proliferation, and on endothelial cells via ETB receptors, releasing nitric oxide (NO) and prostacyclin (PGI2), causing vasorelaxation.

As a biomarker, ADMA has been evaluated in several different classes of PH (43, 44). In IPAH, plasma levels are significantly higher than in healthy, matched controls. In such patients, plasma ADMA correlates positively with right atrial pressure, and negatively with mixed venous oxygen saturation, stroke volume, cardiac index and survival. On stepwise multiple regression analysis, ADMA is an independent predictor of mortality and, using Kaplan–Meier survival curves, patients with supramedian ADMA levels have significantly worse survival than those with inframedian levels.

Patients with idiopathic PAH, plasma levels of Ang-1 and Ang-2 were higher in PAH patients as compared to healthy controls.  Moreover, higher plasma levels of Ang-2 were associated with lower CI and mixed venous oxygen saturation (SvO2) and higher PVR, and, with therapy initiation, changes in Ang-2 correlated with changes in hemodynamics (45, 46).

Endostatin is an antiangiogenic peptide. It is synthesized by myocardium, is detectable in the peripheral circulation of patients with decompensated heart failure, and predicts mortality.48 In PAH, reduced RV myocardial oxygen delivery is felt to contribute to a transition from RV adaptation to failure (46).

Cyclic guanosine monophosphate (cGMP) is an intracellular second messenger of nitric oxide and an indirect marker of natriuretic peptide production (46).

Human pentraxin 3 (PTX3) is a protein synthesized by vascular cells that regulates angiogenesis, inflammation, and cell proliferation (46).

N-terminal propeptide of procollagen III (PIIINP), carboxy-terminal telopeptide of collagen I (CITP), matrix metalloproteinase-9 (MMP-9), and tissue inhibitor of metalloproteinase I (TIMP-1)(46).

Osteopontin (OPN) is a matricellular protein that mediates cell migration, adhesion, remodeling, and survival of the vascular and inflammatory cells (46).

F2-isoprostane is a marker of lipid peroxidation of arachidonic acid, which stimulates endothelial cell proliferation and ET-1 synthesis and may play a role in the pathogenesis of PAH (46).

Circulating fibrocytes are bone marrow-derived cells (CD45 /collagen I ) that contribute to organ fibrosis and extracellular matrix deposition (46).

Circulating miRs (46)

Despite many other substances being investigated as potential biomarkers in PAH, more research is needed to validate the results of small studies and assess their clinical utility. Widespread clinical use of current investigational biomarkers will require validated clinical laboratory techniques and increased knowledge of levels in the healthy population as well as other disease states.

Here are important tests in clinical practice (47):


6-min walk distance

Cardiac index



Higher tertiles associated with worse disease

worse renal function

higher right atrial pressure (RAP)

CITP – vascular remodeling


Recent guidelines (17, 18) encourage the use of screening examinations, such as an echocardiogram (UCG), in high-risk populations for the early detection of PAH . To detect PAH in patients with connective tissue disease (CTD), the obvious screening tests are an UCG and spirometry, including assessment of the diffusing capacity of the lung for carbon monoxide (DLCO). Previous studies have suggested that B-type natriuretic peptide (BNP) and its N-terminal prohormone (NT-proBNP) are potential biomarkers for PAH. However, neither BNP nor NT-pro BNP are specific biomarkers of the degeneration of the pulmonary artery; rather, they are biomarkers of cardiac burden resulting from right heart failure.

Human pentraxin 3 (PTX3) is a specific biomarker for PAH, reflecting pulmonary vascular proteins. They are divided into short and long pentraxins on the basis of their primary structure.
C-Reactive protein (CRP) and serum amyloid P are the classic short pentraxins that are produced in the liver in response to systemic inflammatory cytokines (48). In contrast, PTX3 is one of the long pentraxins. It is synthesized by local vascular cells, such as smooth muscle cells, endothelial cells and fibroblasts, as well as innate immunity cells at sites of inflammation. PTX3 plays a key role in the regulation of cell proliferation and angiogenesis (49).

Increased plasma PTX3 levels have been reported in patients with acute myocardial injury in the
24 h after admission to hospital, with levels returning to normal after 3 days. Similarly, PTX3 levels are higher in patients with unstable angina pectoris, with the changes in PTX3 levels found to be independent of other coronary risk factors, such as obesity and diabetes mellitus. Finally, high serum PTX3 levels have been reported in patents with vasculitis, such as small-vessel vasculitis  and Takayasu aortitis.

Mean plasma PTX3 concentrations in the CTD-PAH and CTD patients were 5.02+0.69 ng/mL (range 1.82–12.94 ng/mL) and 2.40+0.14 ng/mL (range 0.70–4.29 ng/mL), respectively (Table 2). Log transformation of the data revealed significantly higher PTX3 levels in CTD-PAH than in CTD patients (1.49+0.12 vs. 0.82+0.06 log ng/mL, respectively; P = 0.001).(not shown)(50)

Figure 1. Serum pentraxin 3 (PTX3) concentrations in 50 patients with pulmonary arterial hypertension (PAH) and 100 healthy controls, and their correlation with serum concentrations of other biomarkers. A: Comparison of PTX3 concentrations in PAH patients and healthy controls. Mean plasma PTX3 concentrations were 4.4060.37 and 1.94+0.09 ng/mL in the controls and PAH patients, respectively. B: Distribution of log-transformed PTX3 concentrations in PAH patients and healthy controls. C: Log-transformed PTX3 concentrations were significantly higher in patients with PAH than in healthy controls (1.34+0.07 vs. 0.55+0.05 log ng/mL, respectively; P,0.001). D, E: There was no correlation between plasma concentrations of PTX3 and either B-type natriuretic peptide (BNP; r=0.33, P=0.02) or C-reactive protein (CRP; r=0.21, P=0.14) in PAH patients. (not shown) (50)


Table 2. Clinical characteristics and biomarkers in patients with connective tissue disease, with or without pulmonary arterial hypertension.

CTD-PAH ( n =17)                CTD alone ( n =34)       P -value

Age (years)                                 56.3+4.6                                 56.3+2.7               0.990

No. women (%)                         15 (88)                                      31(91)                  0.745

No. with SSc (%)                       10 (59)                                      20 (59)                    1

No. with heart failure (%)          1 (6)                                         0                            –

No. being treated for PAH (%)   17 (100)                                  0                           –

Serum PTX3 (mg/dL)                   5.02+0.69                          2.40+0.14             0.001

Serum CRP (mg/dL)                   0.24+0.09                            0.22+0.04             0.936

Serum BNP (pg/mL)                 189.3+74.                            4 49.3+12.1            0.014

…..  CTD, connective tissue disease; PAH, pulmonary arterial hypertension; SSc, scleroderma;

Figure 3. Receiver operating characteristic (ROC) curves for pentraxin 3 (PTX3) and other biomarkers in patients with connective tissue disease (CTD). The areas under the ROC curve (AUCROC) for PTX3 was 0.866 (95% confidence interval (CI) 0.757–0.974). The star indicates the threshold concentration of 2.85 ng/mL PTX3 that maximized true-positive and false-negative results (sensitivity 94.1%, specificity 73.5%). The AUCROC for C-reactive protein (CRP) was 0.518 (95% CI 0.333–0.704), whereas that for B-type natriuretic peptide (BNP) was 0.670 (95% CI 0.497–0.842). (50)

This study was to determine whether PTX3, the regulation of which is independent of that of the systemic inflammatory marker CRP, is a useful biomarker for diagnosing PAH. The investigators found that PTX3 may be a more sensitive biomarker for PAH than BNP, which is, to date, the most established biomarker for PAH, especially in patients with CTD-PAH. Their findings suggest that PTX3 does not reflect the cardiac burden due to the pulmonary hypertension, but rather the activity of pulmonary vascular degeneration because PTX3 levels were significantly decreased after active treatment specifically for PAH (50). PLoS ONE 7(9): e45834.

Pharmacologic treatment for pulmonary arterial hypertension (PAH) remains suboptimal and mortality rates are still high, even with pulmonary vasodilator therapy. In addition, we have only an incomplete understanding of the pathobiology of PAH, which is characterized at the tissue level by fibrosis, hypertrophy and plexiform remodeling of the distal pulmonary arterioles. Novel therapeutic approaches that might target pulmonary vascular remodeling, rather than pulmonary vaso-reactivity, require precise patient phenotyping both in terms of clinical status and disease subtype. However, current risk stratification models are cumbersome and not precise enough for choosing or assessing the results of therapeutic intervention. Biomarkers used in patients with left heart failure, such as troponin-T and N-terminal pro-B-type natriuretic peptide (NT-proBNP) are elevated in PAH patients but tend to simply reflect increased circulating plasma volumes and elevated right heart pressure, rather than conveying information about disease mechanism.

In this issue of Heart, Calvier and colleagues (see page 390) (51)propose galectin-3 as a useful biomarker in PAH. The rationale for this hypothesis is that elevated aldosterone levels induce an increase in serum levels of galectin-3, a β-galactoside-binding lectin expressed by circulating myocytes, endothelial cells and other cardiovascular cell types. Among other effects, activation of the aldosterone/galactin-3 pathway promotes fibrosis (51), suggesting that elevated levels will correlate with the severity of PAH due to increased pulmonary arteriolar remodeling. To test this hypothesis, serum levels were measured in a total of 57 patients – 41 with idiopathic PAH (iPAH) and 16 with PAH associated with a connective tissue disorder (CTD). The magnitude of elevation in serum levels of aldosterone, galectin-3 and NT-proBNP each correlated with the severity of PAH. However, as shown in figure 1, although serum levels of galectin-3 were elevated in both iPAH and PAH-CTD patients, aldosterone was elevated only in those with iPAH.

In addition, elevated vascular cell adhesion molecule 1 (VCAM-1) and proinflammatory, anti-angiogenic interleukin 12 (IL-12) in were elevated only in PAH-CTD patients, not in those in iPAH. These data suggest that aldosterone and galectin-3 can be used as biomarkers “in tandem” that reflect both the severity and cause of PAH (52).

In the accompanying editorial, Maron (see page 335) summarizes the knowledge gaps in PAH and concludes: “Taken together, Calvier and colleagues provide a key contribution to an underdeveloped area of pulmonary vascular medicine and in doing so identify galectin-3/aldosterone as promising biomarker(s) for informing both disease pathobiology and clinical status in PAH. The rationale of this pursuit in PAH was based, in part, on lessons earned from left heart failure in which the importance of systemically circulating vasoactive factors to clinical trajectory is well established. In this regard, the current work not only develops a novel scientific avenue worthy of further investigation, but also adds to the evolving body of evidence implicating a role for neurohumoral activation in the pathophysiology of PAH”.

Rheumatoid arthritis (RA) affects about 1% of the population and is known to be a significant risk factor for cardiovascular disease, with a 3-fold increased risk of myocardial infarction, a 2-fold increased risk of sudden death and a 50% increase in cardiovascular mortality rates. However, outcomes after PCI in RA patients have not been well characterized and there is little data on the possible effects of disease modifying therapy for RA on risk of restenosis after percutaneous coronary intervention (PCI). In a single center retrospective cohort study, Sintek and colleagues (53)(see page 363) compared the primary endpoint of repeat target vessel revascularization (TVR) in 143 RA patients matched to 541 other.

Pathophysiological targets of differing imaging modalities, demonstrate targets for tracers/contrast agents/pharmacotherapy used in SPECT, PET, MRI and echocardiography to assess myocardial viability.  (Not shown. Adapted from Schuster et al., J Am Coll Cardiol 2012; 59:359–70.)

Ischemic cardiomyopathy implies significant left ventricular systolic dysfunction with an underlying pathophysiology that includes myocardial scarring, hibernation and stunning, or a combination of these disease states. The role of imaging in assessment of myocardial viability is emphasized (not shown) (54) with brief summaries of the role of echocardiography, single photon emission computed tomography (SPECT), positron emission tomography (PET), and magnetic resonance imaging (MRI). The effects of revascularization in patients with ischemic cardiomyopathy remain controversial. Instead, the key elements of evidence based therapy for ischemic cardiomyopathy are standard medical therapy for heart failure combined with implantable cardiac defibrillation (ICD) and/or biventricular pacing device therapy in appropriate patients.

The relationship between the heart and the kidney in hypertension and heart failure

Hypertension is undoubtedly a factor in the treatment of chronic kidney disease because of the relationship between kidney function and BP components that have been studied in people with CKD, diabetes, and hypertension.  Cystatin C was used to evaluate the association between kidney function and both SBP and DBP and 24-h creatinine clearance (CrCl) among 906 participants in the Heart and Soul Study.  (56).  The study investigators hypothesized that although both creatinine and cystatin C are freely filtered at the glomerulus, a major difference between them is that creatinine is secreted by renal tubules, whereas cystatin C is metabolized by the proximal tubule and only a small fraction appears in the urine. In addition, Cystatin C has also been shown to be a stronger predictor of adverse outcomes than serum creatinine. Based on the more linear relationship of cystatin C with GFR, they hypothesized that cystatin C would have a stronger association with SBP than conventional measures of kidney function. Their results found that SBP was linearly associated with cystatin C concentrations (1.19 ± 0.55 mm Hg increase per 0.4 mg/L cystatin C, P = .03) across the range of kidney functions, but only in subjects with CrCl <60 mL/min (6.4 ± 2.13 mm Hg increase per 28 mL/min, P = .003), not >60 mL/min. Further, the DBP was not associated with cystatin C or CrCl. However, PP was linearly associated with both cystatin C (1.28 ± 0.55 mm Hg per 0.4 mg/L cystatin, P = .02) and CrCl <60 mL/min (7.27 ± 2.16 mm Hg per 28 mL/min, P = .001). The relationship between SBP and cystatin C by decile is shown in Figure 7 and Table 3.

Figure 7.

Mean systolic blood pressure (SBP) and diastolic blood pressure (DBP) by decile of kidney function measured as cystatin C.



Table 3

Linear regression of systolic blood pressure by kidney function (N = 906)

Age-adjusted Multivariable adjusted*
Measure N β coefficient P β coefficient P
Cystatin-C (per 0.4 mg/L [SD] increase) 1.75 ± 0.72 .01 1.19 ± 0.55 .03
    >1.0 551 2.23 ± 0.07 .03 1.23 ± 0.03 .04
    <1.0 355 1.59 ± 0.04 .71 0.54 ± 0.01 .87
Spline P value for difference in slopes .85
24-h CrCl (per 28 mL/min [SD] decrease)
    Overall 1.96 ± 0.76 .01 0.91 ± 0.61 .14
    <60 222 11.20 ± 2.74 <.001 6.40 ± 2.13 .003
    >60 684 0.31 ± 0.99 .42 0.36 ± 0.77 .64
    Spline P-value for difference in slopes .01

The results for both Cystatin C and for eGFR are in agreement with incidence rates for heart failure (57)categorized by ejection fraction (EF) and kidney function over 1992−2000 in the Cardiovascular Health Study. Estimated glomerular filtration rate (mL/min per 1.73 m2) is labeled as “eGFR”. (

The association of cystatin C with risk for SHF appeared linear across quartiles of cystatin C (57) and slightly stronger at the highest categories of cystatin C, whereas the lower three quartiles of cystatin C had similar risks for DHF. Participants with an estimated GFR ≥ 60 mL/min per 1.73 m2 had an equal likelihood of developing DHF or SHF, whereas participants with an estimated GFR < 60 mL/min per 1.73 m2 had a greater likelihood of developing SHF.

When an interaction term for HF type (SHF or DHF) was inserted into a fully adjusted standard Cox proportional hazards model with HF with either type of EF as the outcome, the association of continuous cystatin C with SHF was significantly greater than the association of cystatin C with DHF ( P value for interaction < 0.001). The association of estimated GFR and SHF compared with DHF was weaker (P value for interaction = 0.06 for the fully adjusted model).

Ascending quartiles of cystatin C were associated with increasing adjusted risk for the development of “unclassified” HF, defined by the absence of a point-of-care EF measurement. The magnitude of the fully adjusted hazard ratios for the association between cystatin C and risk of unclassified HF were intermediate between those described for DHF and SHF [hazard ratios (95% confidence intervals) for each higher quartile of cystatin C 1.00 (reference), 1.12 (0.80−1.57), 1.84 (1.34−2.51), 2.18 (1.58−3.00)]. The authors state that increased left atrial filling pressures trigger the release of atrial natriuretic peptide and inhibition of vasopressin, which leads to decreased renal sympathetic tone and diuresis early in the pathogenesis of HF (57).  They suggest that even relatively small decrements in k58idney function contribute to the risk of SHF.

Aldosterone plays a key role in homeostatic control and maintenance of blood pressure (BP) by regulation of extracellular volume, vascular tone, and cardiac output. Taking this assumption further, a study unrelated to that above explored the magnitude of the effect of relative aldosterone excess in predicting peripheral as well as aortic blood pressure in a cohort of patients undergoing coronary angiography.  (58) They found that mean peripheral systolic blood pressure (SBP) and diastolic blood pressure (DBP) of the entire cohort were 141 ± 24 mm Hg and 81 ± 11 mm Hg, respectively. Median SBP and aortic SBP increased steadily and significantly from aldosterone/renin ratio (ARR), respectively; p < 0.0001 for both) after multivariate adjustment for parameters potentially influencing BP. ARR emerged as the second most significant independent predictor (after age) of mean SBP and as the most important predictor of mean DBP in this patient cohort.  The authors stress the importance of the ARR in modulating BP over a much wider range than is currently appreciated, as it was already known that the ARR was positively associated with pulse wave velocity in young normotensive healthy adults, indicating that relative aldosterone excess might affect arterial remodeling and precede BP rise as a result of increased vascular stiffness. In this study the ARR was calculated as the PAC/PRC ratio (pg/ml/pg/ml). An ARR >50 pg/ml had a sensitivity and specificity of ARR of 89% and 96%, respectively, for primary aldosteronism. The ARR was modeled as a continuous ratio (with log-transformed values).  The study carried out a multivariate stepwise regression analysis for predictors of BP (not shown). They illustrate (not shown) that marked increases in PRC are a major characteristic of lower ARR categories, and that  across a broad range of ARR values, inappropriately elevated aldosterone levels exert a strong effect on BP values and constitute the most important and second-most important predictor of DBP and SBP, respectively.

Cystatin C may be ordered when a health practitioner is not satisfied with the results of other tests, such as a creatinine or creatinine clearance, or wants to check for early kidney dysfunction, particularly in the elderly, and/or wants to monitor known impairment over time. In diverse populations it has been found to improve the estimate of GFR when combined in an equation with blood creatinine. A high level in the blood corresponds to a decreased glomerular filtration rate (GFR) and hence to kidney dysfunction. Since cystatin C is produced throughout the body at a constant rate and removed and broken down by the kidneys, it should remain at a steady level in the blood if the kidneys are working efficiently and the GFR is normal.

Chronic kidney disease (CKD) is defined as the presence of: persistent and usually progressive reduction in GFR (GFR <60 mL/min/1.73 m2) and/or albuminuria (>30 mg of urinary albumin per gram of urinary creatinine), regardless of GFR. Cystatin C is an index of GFR, especially in patients where serum creatinine may be misleading (eg, very obese, elderly, or malnourished patients); for such patients, use of CKD-EPI cystatin C equation is recommended to estimate GFR. Cystatin C eGFR may have advantages over creatinine eGFR in certain patient groups in whom muscle mass is abnormally high or low (for example quadriplegics, very elderly, or malnourished individuals). Blood levels of cystatin C also equilibrate more quickly than creatinine, and therefore, serum cystatin C may be more accurate than serum creatinine when kidney function is rapidly changing (59) (for example amongst hospitalized individuals).

It is a low molecular weight (13,250 kD) cysteine proteinase inhibitor that is produced by all nucleated cells and found in body fluids, including serum. Since it is formed at a constant rate and freely filtered by the kidneys, its serum concentration is inversely correlated with the glomerular filtration rate (GFR); that is, high values indicate low GFRs while lower values indicate higher GFRs, similar to creatinine. While both cystatin C and creatinine are freely filtered by glomeruli, cystatin C is reabsorbed and metabolized by proximal renal tubules. Thus, under normal conditions, cystatin C does not enter the final excreted urine to any significant degree, and the serum concentration is unaffected by infections, inflammatory or neoplastic states, or by body mass, diet, or drugs.  GFR can be estimated (eGFR) from serum cystatin C utilizing an equation which includes the age and gender of the patient (CKD-EPI cystatin C equation, developed by Inker et al. (59) It demonstrated good correlation with measured iothalamate clearance in patients with all common causes of kidney disease, including kidney transplant recipients.

According to the National Kidney Foundation Kidney Disease Outcome Quality Initiative (K/DOQI) classification, among patients with CKD, irrespective of diagnosis, the stage of disease should be assigned based on the level of kidney function:

Table 4

Stage Description GFR mL/min/BSA
1 Kidney damage with normal or  increased GFR 90
2 Kidney damage with mild decrease in  GFR 60-89
3 Moderate decrease in GFR 30-59
4 Severe decrease in GFR 15-29
5 Kidney failure <15 (or dialysis)


In a study to evaluate cystatin C as a measure of renal function in comparison to serum creatinine, 500 patients had cystatin C measured by nephelometry and glomerular filtration rate (GFR) measured by nonradiolabeled iothalamate clearance (59). In addition, serum creatinine was measured and the patients’ medical records reviewed. The correlation of 1/cystatin C with GFR (r=0.90) was significantly superior than 1/creatinine (r=0.82, p<0.05) with GFR. The superior correlation of 1/cystatin C with GFR was observed in the various clinical subgroups of patients studied (ie, subjects with no suspected renal disease, renal transplant patients, recipients of some other transplant, patients with glomerular disease, and patients with non-glomerular renal disease). The findings indicated that cystatin C may be superior to serum creatinine for the assessment of GFR in a wide spectrum of patients (59). Others have similarly found that cystatin C correlates better than serum creatinine for assessment of GFR. (60)

Patients were screened for 3 chronic kidney disease (CKD) studies in the United States (n = 2,980) and a clinical population in Paris, France (n = 438)(61).   GFR was measured by using urinary clearance of iodine125-iothalamate in the US studies and chromium51-EDTA in the Paris study. GFR was calculated using the 4 new equations based on serum cystatin C alone, serum cystatin C, serum creatinine, or both with age, sex, and race. New equations were developed by using linear regression with log GFR as the outcome in two thirds of data from US studies. Internal validation was performed in the remaining one third of data from US CKD studies; external validation was performed in the Paris study.

Mean mGFR, serum creatinine, and serum cystatin C values were 48 mL/min/1.73 m2 (5th to 95th percentile, 15 to 95), 2.1 mg/dL, and 1.8 mg/L, respectively. For the new equations, coefficients for age, sex, and race were significant in the equation with serum cystatin C, but 2- to 4-fold smaller than in the equation with serum creatinine (62, 63). Measures of performance in new equations were consistent across the development and internal and external validation data sets. Percentages of estimated GFR within 30% of mGFR for equations based on serum cystatin C alone, serum cystatin C, serum creatinine, or both levels with age, sex, and race were 81%, 83%, 85%, and 89%, respectively. The equation using serum cystatin C level alone yields estimates with small biases in age, sex, and race subgroups, which are improved in equations including these variables. It is concluded that Serum cystatin C level alone provides GFR estimates not linked to muscle mass, and that an equation including serum cystatin C level in combination with serum creatinine level, age, sex, and race provides the most accurate estimates.
The authors report that absence of urinary excretion has made it difficult to rigorously evaluate cystatin C as a filtration marker and to examine its non-GFR determinants. They also point out that a high level of variation in the cystatin C assay (64, 65), and standardization and calibration of clinical laboratories will be important to obtain accurate GFR estimation using cystatin C, as has been shown for creatinine.

The study reported above was followed by a major study by Inker LA, et al. (59). Their findings are summarized as follows. Mean measured GFRs were 68 and 70 ml per minute per 1.73 m2 of body-surface area in the development and validation data sets, respectively. In the validation data set, the creatinine–cystatin C equation performed better than equations that used creatinine or cystatin C alone. Bias was similar among the three equations, with a median difference between measured and estimated GFR of 3.9 ml per minute per 1.73 m2 with the combined equation, as compared with 3.7 and 3.4 ml per minute per 1.73 m2 with the creatinine equation and the cystatin C equation (P=0.07 and P=0.05), respectively. Precision was improved with the combined equation (interquartile range of the difference, 13.4 vs. 15.4 and 16.4 ml per minute per 1.73 m2, respectively [P=0.001 and P<0.001]), and the results were more accurate (percentage of estimates that were >30% of measured GFR, 8.5 vs. 12.8 and 14.1, respectively [P<0.001 for both comparisons]). In participants whose estimated GFR based on creatinine was 45 to 74 ml per minute per 1.73 m2, the combined equation improved the classification of measured GFR as either less than 60 ml per minute per 1.73 m2 or greater than or equal to 60 ml per minute per 1.73 m2 (net reclassification index, 19.4% [P<0.001]) and correctly reclassified 16.9% of those with an estimated GFR of 45 to 59 ml per minute per 1.73 m2 as having a GFR of 60 ml or higher per minute per 1.73 m2.

Other studies have established the importance of cystatin C levels(66, 67) and the factors influencing cystatin C levels on renal function measurement (68), including an implication that cystatin C, an alternative measure of kidney function, was a stronger predictor of the risk of cardiovascular events and death than either creatinine or the estimated GFR (69). This includes the Dallas Heart Study (30) finding that cystatin C was independently associated with a specific cardiac phenotype of concentric hypertrophy, including increased LV mass, concentricity, and wall thickness, but it was not associated with LV systolic function or volume. This association was particularly robust in hypertensives and blacks. The Cystatin C concentrations within stages of CKD are shown in Table 5 (70).

Table 5

      Cystatin C level
Stage a Description GFR range a (ml/min/1.73 m2) Native kidney disease b Transplant recipient c
1 Normal or increased GFR 90 0.80 0.87
2 Mildly decreased GFR 60 to 89 0.80 to 1.09 0.87 to 1.23
3 Moderately decreased GFR 30 to 59 1.10 to 1.86 1.24 to 2.24
4 Severely decreased GFR 15 to 29 1.87 to 3.17 2.25 to 4.10
5 Kidney Failure <15 >3.17 >4.10

a GFR estimates and CKD stage will be inaccurate if there is a calibration difference with the Dade-Behring BN II Nephelometer assay used in this study.

b Using the prediction equation: GFR=66.8 (cystatin C)-1.30.

c Using the prediction equation: GFR=76.6 (cystatin C)-1.16.


Copeptin, a novel marker

Urinary albumin excretion is a powerful predictor of progressive cardiovascular and renal disease. Copeptin is the inactive C-terminal fragment of the vasopressin precursor. It is a reliable marker of vasopressin secretion serves as a useful substitute for circulating vasopressin concentration. This allows  for the indirect measurement of vasopressin in epidemiological studies. Moreover, it has been shown that copeptin is a candidate biomarker for pneumonia 32), a predictor of outcome in heart failure, and is a powerful predictor of renal disease associated with albumin excretion (71).  Figure 8 shows the association between copeptin and 24-hour urinary volume, 24-h urinary osmolality and osmolality (71).


Figure 8


Association between quintiles of copeptin and median 24-h UAE (upper panel) and prevalence of microalbuminuria (lower panel) for males and females. Differences between the quintiles were tested by Kruskal–Wallis test. UAE, urinary albumin excretion.



Table 6 shows the association between copeptin concentration and urinary albumin excretion (UAE) in a log-log plot (71).


Model Corrected for β 95% CI for β P
 1 − (Crude) 0.25 0.20–0.30 <0.001
 2 As 1+age 0.21 0.16–0.26 <0.001
 3 As 2+MAP, BMI, smoking, glucose, cholesterol, CRP, and eGFR 0.10 0.05–0.16 <0.001
 4 As 3+diuretics and ACEi/ARB. 0.09 0.04–0.15 0.001
 1 − (Crude) 0.19 0.15–0.23 <0.001
 2 As 1+age 0.17 0.14–0.22 <0.001
 3 As 2+MAP, BMI, smoking, glucose, cholesterol, CRP, and eGFR 0.16 0.11–0.21 <0.001
 4 As 3+diuretics and ACEi/ARB. 0.17 0.12–0.21 <0.001

ACEi, angiotensin-converting enzyme inhibitor; ARB, angiotensin-II-receptor blocker; BMI, body mass index; CHD, coronary heart disease; CI, confidence interval; CRP, C-reactive protein; eGFR, estimated glomerular filtration rate; MAP, mean arterial pressure.

Log copeptin concentration was entered in the regression analyses as independent and log UAE as the dependent variable. Copeptin was associated with UAE in all age groups, but this association is the strongest when subjects are older. Twenty-four-hour urinary volume and 24-h urinary osmolarity were significantly different, with 24-h urinary volume being higher and 24-h urinary osmolarity being lower in the oldest age group when compared with the youngest age group. In both males and females, high copeptin concentration (a surrogate for vasopressin) is associated with low 24-h urinary volume and high 24-h urinary osmolarity. However, urinary osmolarity was independently associated with UAE, but it was weaker than that between copeptin and UAE.  This might indicate that induction of specific glomerular hyperfiltration or decreased tubular albumin reabsorption are associated with this relationship. In addition, subjects with higher levels of copeptin had lower renal function.  These investigators concluded that copeptin (a reliable substitute for vasopressin) is associated with UAE and microalbuminuria, consistent with the hypothesis that vasopressin induces UAE (72).  Other studies indicated that copeptin levels are increased in patients with pulmonary artery hypertension (73), and
higher serum copeptin levels, a surrogate for arginine vasopressin (AVP) release, are associated not only with systolic and diastolic blood pressure but also with several components of metabolic syndrome (74) including obesity, elevated concentration of triglycerides, albuminuria, and serum uric acid level.



Natriuretic peptides in the evaluation of heart failure

The brain type natriuretic peptide (BNP) and the N-terminal pro B-type natriuretic peptide (NT proBNP), but not yet the atrial natriuretic peptide have gained prominence in the evaluation of patients with CHF, which may be with or without preserved ejection fraction . Richards et al. (75)  make the following points.


  • Threshold values of B-type natriuretic peptide (BNP) and N-terminal prohormone B-type natriuretic peptide (NT-proBNP) validated for diagnosis of undifferentiated acutely decompensated heart failure (ADHF) remain useful in patients with heart failure with preserved ejection fraction (HFPEF), with minor loss of diagnostic performance.


  • BNP and NT-proBNP measured on admission with ADHF are powerfully predictive of in-hospital mortality in both HFPEF and heart failure with reduced EF (HFREF), with similar or greater risk in HFPEF as in HFREF associated with any given level of either peptide.


  • In stable treated heart failure, plasma natriuretic peptide concentrations often fall below cut-point values used for the diagnosis of ADHF in the emergency department; in HFPEF, levels average approximately half those in HFREF.


  • BNP and NT-proBNP are powerful independent prognostic markers in both chronic HFREF and chronic HFPEF, and the risk of important clinical adverse outcomes for a given peptide level is similar regardless of left ventricular ejection fraction.


  • Serial measurement of BNP or NT-proBNP to monitor status and guide treatment in chronic heart failure may be more applicable in HFREF than in HFPEF.


In addition, they point out the following:


BNP and NT-proBNP fall below ADHF thresholds in stable HFREF in approximately 50% and 20% of cases, respectively. Levels in stable HFPEF are even lower, approximately half those in HFREF.


Whereas BNPs have 90% sensitivity for asymptomatic LVEF of less than 40% in the community (a precursor state for HFREF), they offer no clear guide to the presence of early community based HFPEF.


Guidelines recommend BNP and NT-proBNP as adjuncts to the diagnosis of acute and chronic HF and for risk stratification. Refinements for application to HFPEF are needed.


The prognostic power of NPs is similar in HFREF and HFPEF. Defined levels of BNP and NT-proBNP correlate with similar short-term and long-term risks of important clinical adverse outcomes in both HFREF and HFPEF.


They provide a diagnostic algorithm for suspected heart failure (75)(Figure 9).


Figure 9

Diagnostic algorithm for suspected heart failure presenting either acutely or nonacutely



Diagnostic algorithm for suspected heart failure presenting either acutely or nonacutely. a In the acute setting, mid-regional pro–atrial natriuretic peptide may also be used (cutoff point 120 pmol/L; ie, <120 pmol/L 5 heart failure unlikely). b Other causes of elevated natriuretic peptide levels in the acute setting are an acute coronary syndrome, atrial or ventricular arrhythmias, pulmonary embolism, and severe chronic obstructive pulmonary disease with elevated right heart pressures, renal failure, and sepsis. Other causes of an elevated natriuretic level in the nonacute setting are old age (>75 years), atrial arrhythmias, left ventricular hypertrophy, chronic obstructive pulmonary disease, and chronic kidney disease. c Exclusion cutoff points for natriuretic peptides are chosen to minimize the false-negative rate while reducing unnecessary referrals for echocardiography. Treatment may reduce natriuretic peptide concentration, and natriuretic peptide concentrations may not be markedly elevated in patients with heart failure with preserved ejection fraction.


Patients with acute pulmonary symptoms and with acute myocardial infarct present with dyspnea to the Emergency Department.  The evaluation is made particularly difficult in a patient for whom there is no prior history. Maisel et al. (76) presented the utility of the midregion proadrenomedullin (MR-proADM) in all patients presenting with acute shortness of breath.  They found that MR-proADM was superior to BNP or troponin for predicting 90-day all-cause mortality in patients presenting with acute dyspnea (c index = 0.755, p < 0.0001). Furthermore, MR-proADM added significantly to all clinical variables (all adjusted hazard ratios: HR=3.28), and it was also superior to all other biomarkers.


There is a large body of recent work that has enlarged our view of hypertension, kidney disease, cardiovascular disease, including heart failure with (HFpEF) or without preserved ejection fraction. I shall here refer to my review in Leaders in Pharmaceutical Innovation  (78).  The piece contains a study that I published  (79) with collaborators in Brooklyn, Bridgeport and Philadelphia that is no longer available from the publisher.


The natriuretic peptides, B-type natriuretic peptide (BNP) and NT-proBNP that have emerged as tools for diagnosing congestive heart failure (CHF) are affected by age and renal insufficiency (RI).  NTproBNP is used in rejecting CHF and as a marker of risk for patients with acute coronary syndromes. This observational study was undertaken to evaluate the reference value for interpreting NT-proBNP concentrations. The hypothesis is that increasing concentrations of NT-proBNP are associated with the effects of multiple co-morbidities, not merely CHF,

resulting in altered volume status or myocardial filling pressures.


NT-proBNP was measured in a population with normal trans-thoracic echocardiograms
(TTE) and free of anemia or renal impairment. Exclusion conditions were the following



  • anemia as defined by WHO,
  • atrial fibrillation (AF),
  • elevated troponin T exceeding 0.070 mg/dl,
  • systolic or diastolic blood pressure exceeding 140 and 90 respectively,
  • ejection fraction less than 45%,
  • left ventricular hypertrophy (LVH),
  • left ventricular wall relaxation impairment, and
  • renal insufficiency (RI) defined by creatinine clearance < 60ml/min using
    the MDRD formula .

Study participants were seen in acute care for symptoms of shortness of breath suspicious for CHF requiring evaluation with cardiac NTproBNP assay. The median NT-proBNP for patients under 50 years is 60.5 pg/ml with an upper limit of 462 pg/ml, and for patients over 50 years the median was 272.8 pg/ml with an upper limit of 998.2 pg/ml.

We suggested that NT-proBNP levels can be more accurately interpreted only after removal of the major co-morbidities that affect an increase in this  peptide in serum. The PRIDE study guidelines (  should be applied until presence or absence of comorbidities is diagnosed. With no comorbidities, the reference range for normal over 50 years of age remains steady at ~1000 pg/ml. The effect shown in previous papers likely is due to increasing concurrent comorbidity with age.

We observed the following changes with respect to NTproBNP and age:

(i) Sharp increase in NT-proBNP at over age 50

(ii) Increase in NT-proBNP at 7% per decade over 50

(iii) Decrease in eGFR at 4% per decade over 50

(iv) Slope of NT-proBNP increase with age is related to proportion of patients with eGFR less than 90

(v) NT-proBNP increase can be delayed or accelerated based on disease comorbidities

The mean and 95% CI of NTproBNP (CHF removed) by the National Kidney Foundation staging for eGFR interval (eGFR scale: 0, > 120; 1, 90 to 119;2, 60 to 89; 3, 40 to 59; 4, 15 to 39; 5, under 15 ml/min). We created a new variable to minimize the effects of age and eGFR variability by correcting these large effects in the whole sample population.

Adjustment of the NT-proBNP for  both eGFR and for age over 50 differences. We have carried out a normalization to adjust for both eGFR and for age over 50:

(i) Take Log of NT-proBNP and multiply by 1000
(ii) Divide the result by eGFR (using MDRD9 or Cockroft Gault10)
(iii) Compare results for age under 50, 50-70, and over 70 years
(iv) Adjust to age under 50 years by multiplying by 0.66 and 0.56.

Figure 10



NKF staging by GFRe interval and NT-proBNP (CHF removed).



The equation does not require weight because the results are reported normalized

to 1.73 m2 body surface area, which is an accepted average adult surface area.


This is illustrated in Figure 11.

Figure 11


Plot of 1000*log (NT-proBNP)/GFR vs age at  eGFR over 90  and 60 ml/min

Figure 12 compares the reference ranges for NTproBNP before and after adjustment.

  • before adjustment; b) after adjustment. c) the scatterplot for 1000xlog(NT proBNP) versus 1000xlog(NT-proBNP/eGFR). Superimposed scatterplot and regression line with centroid and

confidence interval for 1000*log(NT-proBNP)/eGFR vs age (anemia removed)

at eGFR over 40 and 90 ml/min. (Black: eGFR > 90, Blue:  eGFR > 40)


More recent work is enlightening.  Hijazi et al. (80) studied the incremental value of measuring N-terminal pro–B-type natriuretic peptide (NT-proBNP) levels in addition to established risk factors (including the CHA2DS2VASc [heart failure, hypertension, age 75 years and older, diabetes, and previous stroke or transient ischemic attack, vascular disease, age 65 to 74 years, and sex category) for the prediction of cardiovascular and bleeding events. They concluded that NT-proBNP levels are often elevated in atrial fibrillation (AF) and it is independently associated with an increased risk for stroke and mortality. NT-proBNP improves risk stratification beyond the CHA2DS2VASc score and might be a novel tool for improved stroke prediction in AF. The

efficacy of apixaban compared with warfarin was independent of the NT-proBNP level. Moreover, natriuretic peptides are regulatory hormones associated with cardiac remodeling, namely, left ventricular hypertrophy and systolic/diastolic dysfunction. Another study reported that the risk of death of patients with plasma NT-proBNP 133 pg/mL (third tertile of the distribution) was 3.3 times that of patients with values 50.8 pg/mL (first tertile; hazard ratio: 3.30 [95% CI: 0.90 to 12.29]). This predictive value was independent of, and superior to, that of 2 ECG indexes of left ventricular hypertrophy, the Sokolov-Lyon index and the amplitude of the R wave in lead aVL and it persisted in patients without ECG left ventricular hypertrophy (81).
Many patients presenting with acute dyspnea (including those with ADHF) have multiple coexisting medical disorders that may complicate their diagnosis and management. These patients presenting with acute dyspnea may have longer hospital length of stay and are at high risk for repeat hospitalization or death. In this presentation testing for brain natriuretic peptide (BNP) or NT-proBNP has been shown to be valuable for an accurate and efficient diagnosis and prognostication of HF (82).


The biological activity of BNP, the product of an intracellular peptide (proBNP108) that is converted to NT-proBNP, includes stimulation of natriuresis and vasorelaxation; inhibition of renin, aldosterone, and sympathetic nervous activity; inhibition of fibrosis; and improvement in myocardial relaxation.


Figure 13


Biology of the natriuretic peptide system. BNP indicates brain natriuretic peptide; NT-proBNP, amino-terminal pro-B-type natriuretic peptide; and DPP-IV, dipeptidyl peptidase-4.

The authors remind us that approximately 20% of patients with acute dyspnea have BNP or NT-proBNP levels that are above the cutoff point to exclude HF but too low to definitively identify it (82). Knowledge of the differential diagnosis of non-HF elevation of NP, as well as interpretation of the BNP or NT-proBNP value in the context of a clinical assessment is essential.  Across all stages of HF, elevated BNP or NT-proBNP concentrations are at least comparable prognostic predictors of mortality and cardiovascular events relative to traditional predictors of outcome in this setting, with increasing NP concentrations predicting worse prognosis in a linear fashion. This prognostic value may be used to stratify patients at the highest risk of adverse outcomes (see Figure 2 In this page). Age-adjusted Kaplan-Meier survival curve of mortality at 1 year associated with an elevated amino-terminal pro-B-type natriuretic peptide    (NT-proBNP) concentration at emergency department presentation with dyspnea in those with acutely decompensated heart failure. Reproduced from Januzzi et al22. (82)

The importance of determining diastolic and systolic function and for measurement of pulmonary artery pressure by echocardiography is clear, as NT-proBNP levels may be increased with increase in pulmonary pressure as well as conditions that increase cardiac output. Although Hijazi et al. used the Cockcroft-Gault (CG) equation to determine the glomerular filtration rate (GFR) the CG equation may find higher eGFR in older individuals (80). In addition, elevated NT-proBNP independently predicts all-cause mortality and morbidity of patients with AF. A prominent disease with elevated NT-proBNP is a respiratory system disease, such as chronic obstructive pulmonary disease, pulmonary embolism, and interstitial lung disease, in which B-type natriuretic peptide levels are elevated in response to the pressure of the right side of the heart. The authors conclude that one should keep in mind that NT-proBNP alone may be inadequate.

NT-proBNP level is used for the detection of acute CHF and as a predictor of survival. However, a number of factors, including renal function, may affect the NT-proBNP levels. This study aims to provide a more precise way of interpreting NT-proBNP levels based on GFR, independent of age. This study includes 247 pts in whom CHF and known confounders of elevated NT-proBNP were excluded, to show the relationship of GFR in association with age. The effect of eGFR on NT-proBNP level was adjusted by dividing 1000 x log(NT-proBNP) by eGFR then further adjusting for age in order to determine a normalized NT-proBNP value. The normalized NT-proBNP levels were affected by eGFR independent of the age of the patient. A normalizing function based on eGFR eliminates the need for an age-based reference ranges for NT-proBNP (79).

The routine use of natiuretic peptides in severely dyspneic patients has recently been called into question. We hypothesized that the diagnostic utility of Amino Terminal pro Brain Natiuretic Peptide (NT-proBNP) is diminished in a complex elderly population (83)

We studied 502 consecutive patients in whom NT-proBNP values were obtained to evaluate severe dyspnea in the emergency department (84). The diagnostic utility of NT-proBNP for the diagnosis of congestive heart failure (CHF) was assessed utilizing several published guidelines, as well as the manufacturer’s suggested age dependent cut-off points. The area under the receiver operator curve (AUC) for NT-proBNP was 0.70. Using age-related cut points, the diagnostic accuracy of NT-proBNP for the diagnosis of CHF was below prior reports (70% vs. 83%). Age and estimated creatinine clearance correlated directly with NT-proBNP levels, while hematocrit correlated inversely. Both age > 50 years and to a lesser extent hematocrit < 30% affected the diagnostic accuracy of NT-proBNP, while renal function had no effect. In multivariate analysis, a prior history of CHF was the best predictor of current CHF, odds ratio (OR) = 45; CI: 23-88.

The diagnostic accuracy of NT-proBNP for the evaluation of CHF appears less robust in an elderly population with a high prevalence of prior CHF. Age and hematocrit levels, may adversely affect the diagnostic accuracy off NT-proBNP (85).

Obesity and hypertension.

Obesity is associated with an increased risk of hypertension. In the past 5 years there have been dramatic advances into the genetic and neurobiological mechanisms of obesity with the discovery of leptin and novel neuropeptide pathways regulating appetite and metabolism. In this brief review, we argue that these mounting advances into the neurobiology of obesity have and will continue to provide new insights into the regulation of arterial pressure in obesity. We focus our comments on the sympathetic, vascular, and renal mechanisms of leptin and melanocortin receptor agonists and on the regulation of arterial pressure in rodent models of genetic obesity. Three concepts are proposed (86).

First, the effect of obesity on blood pressure may depend critically on the genetic-neurobiological mechanisms underlying the obesity. Second, obesity is not consistently associated with increased blood pressure, at least in rodent models. Third, the blood pressure response to obesity may be critically influenced by modifying alleles in the genetic background.

Leptin plays an important role in regulation of body weight through regulation of food intake and sympathetically mediated thermogenesis. The hypothalamic melanocortin system, via activation of the melanocortin-4 receptor (MC4-R), decreases appetite and weight, but its effects on sympathetic nerve activity (SNA) are unknown. In addition, it is not known whether sympathoactivation to leptin is mediated by the melanocortin system.

The following study (87) tested the interactions between these systems in regulation of brown adipose tissue (BAT) and renal and lumbar SNA in anesthetized Sprague-Dawley rats. Intracerebroventricular administration of the MC4-R agonist MT-II (200 to 600 pmol) produced a dose-dependent sympathoexcitation affecting BAT and renal and lumbar beds. This response was completely blocked by the MC4-R antagonist SHU9119 (30 pmol ICV). Administration of leptin (1000 m g/kg IV) slowly increased BAT SNA (baseline, 4166 spikes/s; 6 hours, 196628 spikes/s; P50.001) and renal SNA (baseline, 116616 spikes/s; 6 hours, 169626 spikes/s; P50.014).

Intracerebroventricular administration of SHU9119 did not inhibit leptin-induced BAT sympathoexcitation (baseline, 3567 spikes/s; 6 hours, 158634 spikes/s; P50.71 versus leptin alone). However, renal sympathoexcitation to leptin was completely blocked by SHU9119 (baseline, 142617 spikes/s; 6 hours, 146625 spikes/s; P50.007 versus leptin alone). The study (87) demonstrates that the hypothalamic melanocortin system can act to increase sympathetic nerve traffic to thermogenic BAT and other tissues. Our data also suggest that leptin increases renal SNA through activation of hypothalamic melanocortin receptors. In contrast, sympathoactivation to thermogenic BAT by leptin appears to be independent of the melanocortin system.


The introduction of the first generation troponins T and I was an important event leading to the declining use of creatine kinase isoenzyme MB because of the short half-life in the circulation of CKMB and the possibility of missing a late presenting ACS. The situation then would call for the measurement of lactate dehydrogenase isoenzyme 1 (H-type), which had a decline in use.  The troponins T and I are proteins associated with the muscle contractile element with high specificity for the cardiomyocyte apparatus, which increased rapidly after ACS and which had estimated diagnostic cutoffs of 0.08 mg/dl and 1 mg/dl respectively.  The choice of marker was largely dependent of the instrument platform.  These biomarkers went through several generations of improvement to improve the diagnostic sensitivity to a cutoff at 2 SD of the lower limit of detection, magnifying confusion in interpretation that had always existed. These cardiospecific markers are elevated in patients with hypertension and specifically, long term CKD. This was clarified by introducing the terms Type 1 and Type 2 myocardial infarct, designating the classic ACS due to plaque rupture as Type 1.  However, the type 2 class might well be non-homogeneous. In any case, these are the best we have in detecting myocardial ischemic damage with biomarker release.



This discussion has covered a large body of research involving hypertension, the kidney, and cardiovascular humoral mechanisms of control with a broad brush.  The work that has been done is far more than is cited.  There are several biomarkers that we have considered. They are not only laboratory based measurements.  They are: PWV, cystatin C, eGFR, copeptin, BNP or NT-BNP, Midregional prohormone adrenomedullin (MR-ADM), urinary albumin excretion, and the aldosterone/renin ratio.

The preceding discussion reminds us of the story of the blind men palpating an elephant, set in a poem by John Godfrey Saxe. These blind men were asked to tell of their experiences palpating different parts of an elephant, without seeing the entire animal Figure 1. Each of the blind men was able to palpate one part of the elephant, and thus was able to describe it in terms that were “partly in the right.” However, because none of them was able to encompass the entire elephant in their hands, they were also “in the wrong,” in that they failed to identify the whole elephant (88).
The blind men and the elephant. Poem by John Godfrey Saxe (Cartoon originally copyrighted by the authors (88); G. Renee Guzlas, artist).

These authors advanced the “elephant” as the increased oxidative burden in the uremic milieu of patients with chronic kidney disease. I introduce the concept in the diagnostic dilemma about what biomarkers are diagnostically informative in hypertension and ischemic CVD poses a conundrum. In reviewing the full gamut of biomarkers, we have a replay of the Lone Ranger and the silver bullet.  The problem is that there is no “silver” bullet.  We are accustomed to rely on clinical observations that are themselves weak covariates in actual experience.  The studies that have been done to validate the effectiveness of key biomarkers are well designed and show relevance in the populations studied.  However, they are insufficient by themselves in the emergent care population.

Impediments to a solution to the problem

Tests are ordered by physicians based on the findings in a clinical history and physical examination. Test that are ordered are reimbursed by insurance carriers, Medicare and Medicaid based on a provisional diagnosis.  The provisional diagnosis generates an ICD10 code, which has been most recently revised with a weighted input from the insurers that is not in favor of considered clinical evidence.  Moreover, the provider of care is graded based on the number of patients seen and the tests performed on a daily basis over any period.  Given this situation, and in addition, the requirement to interact with an outmoded information system that is more helpful to the insurer and less helpful to the provider, it is not surprising that there is a large burnout of the nursing and physician practitioner workforce.  If the diagnosis is inconclusive at the time of patient examination, then the work is not reimbursable based on ICD10 coding requirements that are disease specific.   This problem breaks down into a workload and a reimbursement inconsistency, neither of which makes sense in terms of the original studies on Diagnosis Related Groups (89) at Yale by Robert Fetter’s group.  The problem is made worse by the design and selection of healthcare information systems.

Many have pointed out the flaws in current EHR design that impede the optimum use of data and hinder workflow. Researchers have suggested that EHRs can be part of a learning health system to better capture and use data to improve clinical practice, create new evidence, educate, and support research efforts. The health care system suffers from both inefficient and ineffective use of data. Data are suboptimally displayed to users, undernetworked, underutilized, and wasted. Errors, inefficiencies, and increased costs occur on the basis of unavailable data in a system that does not coordinate the exchange of information, or adequately support its use (90). Clinicians’ schedules are stretched to the limit and yet the system in which they work exerts little effort to streamline and support carefully engineered care processes. Information for decision-making is difficult to access in the context of hurried real-time workflows(91)



The solution to the problem

The current design of the Electronic Medical Record (EMR) is a linear presentation of portions of the record by services, by diagnostic method, and by date, to cite examples.  This allows perusal through a graphical user interface (GUI) that partitions the information or necessary reports in a workstation entered by keying to icons.  This requires that the medical practitioner finds the history, medications, laboratory reports, cardiac imaging and EKGs, and radiology in different workspaces.  The introduction of a DASHBOARD has allowed a presentation of drug reactions, allergies, primary and secondary diagnoses, and critical information about any patient the care giver needing access to the record.  The advantage of this innovation is obvious.  The startup problem is what information is presented and how it is displayed, which is a source of variability and a key to its success.

Gil David and Larry Bernstein have developed, in consultation with Prof. Ronald Coifman, in the Yale University Applied Mathematics Program, a software system that is the equivalent of an intelligent Electronic Health Records Dashboard (92)( that provides empirical medical reference and suggests quantitative diagnostics options.

The most commonly ordered test used for managing patients worldwide is the hemogram that often incorporates the review of a peripheral smear.  While the hemogram has undergone progressive modification of the measured features over time the subsequent expansion of the panel of tests has provided a window into the cellular changes in the production, release or suppression of the formed elements from the blood-forming organ to the circulation.  In the hemogram one can view data reflecting the characteristics of a broad spectrum of medical conditions.

How we frame our expectations is so important that it determines the data we collect to examine the process.   In the absence of data to support an assumed benefit, there is no proof of validity at whatever cost.   This has meaning for hospital operations, for nonhospital laboratory operations, for companies in the diagnostic business, and for planning of health systems.

In 1983, a vision for creating the EMR was introduced by Lawrence Weed, expressed by McGowan and Winstead-Fry (93)

The data presented has to be comprehended in context with vital signs, key symptoms, and an accurate medical history.  Consequently, the limits of memory and cognition are tested in medical practice on a daily basis.  We deal with problems in the interpretation of data presented to the physician, and how through better design of the software that presents this data the situation could be improved.  The computer architecture that the physician uses to view the results is more often than not presented as the designer would prefer, and not as the end-user would like.

Eugene Rypka contributed greatly to clarifying the extraction of features (94) in a series of articles, which set the groundwork for the methods used today in clinical microbiology.  The method he describes is termed S-clustering, and will have a significant bearing on how we can view hematology data.  He describes S-clustering as extracting features from endogenous data that amplify or maximize structural information to create distinctive classes.  The method classifies by taking the number of features with sufficient variety to map into a theoretic standard. The mapping is done by a truth table, and each variable is scaled to assign values for each: message choice.  The number of messages and the number of choices forms an N-by N table.  He points out that the message choice in an antibody titer would be converted from 0 + ++ +++ to 0 1 2 3.

Bernstein and colleagues had a series of studies using Kullback-Liebler Distance  (effective information) for clustering to examine the latent structure of the elements commonly used for diagnosis of myocardial infarction (95-97)(CK-MB, LD and the isoenzyme-1 of LD),  protein-energy malnutrition (serum albumin, serum transthyretin, condition associated with protein malnutrition (see Jeejeebhoy and subjective global assessment), prolonged period with no oral intake), prediction of respiratory distress syndrome of the newborn (RDS), and prediction of lymph nodal involvement of prostate cancer, among other studies.   The exploration of syndromic classification has made a substantial contribution to the diagnostic literature, but has only been made useful through publication on the web of calculators and nomograms (such as Epocrates and Medcalc) accessible to physicians through an iPhone.  These are not an integral part of the EMR, and the applications require an anticipation of the need for such processing.

Gil David et al. (90, 92) introduced an AUTOMATED processing of the data available to the ordering physician and can anticipate an enormous impact in diagnosis and treatment of perhaps half of the top 20 most common causes of hospital admission that carry a high cost and morbidity.  For example: anemias (iron deficiency, vitamin B12 and folate deficiency, and hemolytic anemia or myelodysplastic syndrome); pneumonia; systemic inflammatory response syndrome (SIRS) with or without bacteremia; multiple organ failure and hemodynamic shock; electrolyte/acid base balance disorders; acute and chronic liver disease; acute and chronic renal disease; diabetes mellitus; protein-energy malnutrition; acute respiratory distress of the newborn; acute coronary syndrome; congestive heart failure; disordered bone mineral metabolism; hemostatic disorders; leukemia and lymphoma; malabsorption syndromes; and cancer(s)[breast, prostate, colorectal, pancreas, stomach, liver, esophagus, thyroid, and parathyroid]. The same approach has also been applied to the problem of hospital malnutrition, but it has not been sufficiently applied to hypertension, cardiovascular diseases, acute coronary syndrome, chronic renal failure.

We have developed (David G, Bernstein L, and Coifman) (92) a software system that is the equivalent of an intelligent Electronic Health Records Dashboard that provides empirical medical reference and suggests quantitative diagnostics options. The primary purpose is to gather medical information, generate metrics, analyze them in realtime and provide a differential diagnosis, meeting the highest standard of accuracy. The system builds its unique characterization and provides a list of other patients that share this unique profile, therefore utilizing the vast aggregated knowledge (diagnosis, analysis, treatment, etc.) of the medical community. The main mathematical breakthroughs are provided by accurate patient profiling and inference methodologies in which anomalous subprofiles are extracted and compared to potentially relevant cases. As the model grows and its knowledge database is extended, the diagnostic and the prognostic become more accurate and precise. We anticipate that the effect of implementing this diagnostic amplifier would result in higher physician productivity at a time of great human resource limitations, safer prescribing practices, rapid identification of unusual patients, better assignment of patients to observation, inpatient beds, intensive care, or referral to clinic, shortened length of patients ICU and bed days.

The main benefit is a real time assessment as well as diagnostic options based on comparable cases, flags for risk and potential problems as illustrated in the following case acquired on 04/21/10. The patient was diagnosed by our system with severe SIRS at a grade of 0.61 .

Method for data organization and classification via characterization metrics.

The database is organized to enable linking a given profile to known profiles. This is achieved by associating a patient to a peer group of patients having an overall similar profile, where the similar profile is obtained through a randomized search for an appropriate weighting of variables. Given the selection of a patients’ peer group, we build a metric that measures the dissimilarity of the patient from its group. This is achieved through a local iterated statistical analysis in the peer group.

This characteristic metric is used to locate other patients with similar unique profiles, for each of whom we repeat the procedure described above. This leads to a network of patients with similar risk condition. Then, the classification of the patient is inferred from the medical known condition of some of the patients in the linked network.

How do we organize the data and linkages provided in the first place?

Predictors: PWV, cystatin C, creatinine, urea, eGFR, copeptin, BNP or NT-BNP, TnI or TnT, Midregional prohormone adrenomedullin (MR-ADM), urinary albumin excretion, and the aldosterone/renin ratio, homocysteine, transthyretin, glucose, albumin, chol/LDL, LD, Na+, K+,  Cl, HCO3, pH.

Conditions: AMI, CRF, ARF, hypertension, HFpEF, HFcEF, ADHF, obesity, PHT, RVHF, pulmonary edema, PEM

Other variables: sex (M,F), age, BMI. …

Conditioning data: take log transform for large ascending values, OR take deciles of variables, if necessary.  This could apply to NT-proBNP, BNP, TnI, TnT, CK and LD.

Arrange predictor variables in columns and patient-sequence in rows.  This is a bidimentional table.  The problem is to assign diagnoses to each patient-in sequence. There can be more than one diagnosis.

In reality the patient-sequence or identifier is not relevant. Only the condition assignment is.  The condition assignments are made in a column adjacent to the patient, and they fall into rows.
The construct appears to be a 2×2, but it is actually an n-dimensional  matrix.  Each patient position has one or more diagnoses.

Multivariate statistical analysis is used to extend this analysis to two or more predictors.   In this case a multiple linear regression or a linear discriminant function would be used to predict a dependent variable from two or more independent variables.   If there is linear association dependency of the variables is assumed and the test of hypotheses requires that the variances of the predictors are normally distributed.  A method using a log-linear model circumvents the problem of the distributional dependency in a method called ordinal regression.    There is also a relationship of analysis of variance, a method of examining differences between the means of  two or more groups.  Then there is linear discriminant analysis, a method by which we examine the linear separation between groups rather than the linear association between groups.  Finally, the neural network is a nonlinear, nonparametric model for classifying data with several variables into distinct classes. In this case we might imagine a curved line drawn around the groups to divide the classes. The focus of this discussion will be the use of linear regression  and explore other methods for classification purposes (98).

The real issue is how a combination of variables falls into a table with meaningful information.  We are concerned with accurate assignment into uniquely variable groups by information in test relationships. One determines the effectiveness of each variable by its contribution to information gain in the system.  The reference or null set is the class having no information.  Uncertainty in assigning to a classification is only relieved by providing sufficient information.  One determines the effectiveness of each variable by its contribution to information gain in the system.  The possibility for realizing a good model for approximating the effects of factors supported by data used for inference owes much to the discovery of Kullback-Liebler distance or “information” (99), and Akaike (100) found a simple relationship between K-L information and Fisher’s maximized log-likelihood function. A solid foundation in this work was elaborated by Eugene Rypka (101).  Of course, this was made far less complicated by the genetic complement that defines its function, which made more accessible the study of biochemical pathways.  In addition, the genetic relationships in plant genetics were accessible to Ronald Fisher for the application of the linear discriminant function.    In the last 60 years the application of entropy comparable to the entropy of physics, information, noise, and signal processing, has been fully developed by Shannon, Kullback, and others,  and has been integrated with modern statistics, as a result of the seminal work of Akaike, Leo Goodman, Magidson and Vermunt, and unrelated work by Coifman. Dr. Magidson writes about Latent Class Model evolution:

The recent increase in interest in latent class models is due to the development of extended algorithms which allow today’s computers to perform LC analyses on data containing more than just a few variables, and the recent realization that the use of such models can yield powerful improvements over traditional approaches to segmentation, as well as to cluster, factor, regression and other kinds of analysis.

Perhaps the application to medical diagnostics had been slowed by limitations of data capture and computer architecture as well as lack of clarity in definition of what are the most distinguishing features needed for diagnostic clarification.  Bernstein and colleagues (102-104) had a series of studies using Kullback-Liebler Distance  (effective information) for clustering to examine the latent structure of the elements commonly used for diagnosis of myocardial infarction (CK-MB, LD and the isoenzyme-1 of LD),  protein-energy malnutrition (serum albumin, serum transthyretin, condition associated with protein malnutrition (see Jeejeebhoy and subjective global assessment), prolonged period with no oral intake), prediction of respiratory distress syndrome of the newborn (RDS), and prediction of lymph nodal involvement of prostate cancer, among other studies.   The exploration of syndromic classification has made a substantial contribution to the diagnostic literature, but has only been made useful through publication on the web of calculators and nomograms (such as Epocrates and Medcalc) accessible to physicians through an iPhone.  These are not an integral part of the EMR, and the applications require an anticipation of the need for such processing.

Gil David et al. introduced an AUTOMATED processing of the data (104) available to the ordering physician and can anticipate an enormous impact in diagnosis and treatment of perhaps half of the top 20 most common causes of hospital admission that carry a high cost and morbidity.  For example: anemias (iron deficiency, vitamin B12 and folate deficiency, and hemolytic anemia or myelodysplastic syndrome); pneumonia; systemic inflammatory response syndrome (SIRS) with or without bacteremia; multiple organ failure and hemodynamic shock; electrolyte/acid base balance disorders; acute and chronic liver disease; acute and chronic renal disease; diabetes mellitus; protein-energy malnutrition; acute respiratory distress of the newborn; acute coronary syndrome; congestive heart failure; disordered bone mineral metabolism; hemostatic disorders; leukemia and lymphoma; malabsorption syndromes; and cancer(s)[breast, prostate, colorectal, pancreas, stomach, liver, esophagus, thyroid, and parathyroid].

Our database organized to enable linking a given profile to known profiles(102-104). This is achieved by associating a patient to a peer group of patients having an overall similar profile, where the similar profile is obtained through a randomized search for an appropriate weighting of variables. Given the selection of a patients’ peer group, we build a metric that measures the dissimilarity of the patient from its group. This is achieved through a local iterated statistical analysis in the peer group.

We then use this characteristic metric to locate other patients with similar unique profiles, for each of whom we repeat the procedure described above. This leads to a network of patients with similar risk condition. Then, the classification of the patient is inferred from the medical known condition of some of the patients in the linked network. Given a set of points (the database) and a newly arrived sample (point), we characterize the behavior of the newly arrived sample, according to the database. Then, we detect other points in the database that match this unique characterization. This collection of detected points defines the characteristic neighborhood of the newly arrived sample. We use the characteristic neighborhood in order to classify the newly arrived sample. This process of differential diagnosis is repeated for every newly arrived point.   The medical colossus we have today has become a system out of control and beset by the elephant in the room – an uncharted complexity.





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  35. Realtime Clinical Expert Support. Pharmaceutical Intelligence.
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  37. Rypka EW and Babb R. Automatic construction and use of an identification scheme. In MEDICAL RESEARCH ENGINEERING Apr 19709; (2):9-19.
  38. Rudolph, R. A., Bernstein, L. H. and Babb, J. Information induction for predicting acute myocardial infarction. Clinical Chemistry 1988; 34: 2031-2038.
  39. Bernstein LH, Qamar A, McPherson C, Zarich S. Evaluating a new graphical ordinal logit method (GOLDminer) in the diagnosis of myocardial infarction utilizing clinical features and laboratory data. Yale J Biol Med 1999; 72:259-268.
  40. Bernstein LH, Good IJ, Holtzman, Deaton ML, Babb J. Diagnosis of acute myocardial infarction from two measurements of creatine kinase isoenzyme MB with use of nonparametric probability estimation. Clin Chem 1989; 35(3):444-447.
  41. Bernstein LH. Regression: A richly textured method for comparison and classification of predictor variables.
  42. Posada D and Buckley TR. Model Selection and Model Averaging in Phylogenetics: Advantages of Akaike Information Criterion and Bayesian Approaches over Likelihood Ratio Tests. Syst. Biol. 200; 53(5):793–808.
  1. Kullback S. and Leibler R. On Information and Sufficiency. Ann Math Statistics. Mar 1951; 22(1):79-86.
  2. Bernstein LH, David G, Rucinski J, Coifman RR. Converting Hematology Based Data Into an Inferential Interpretation. In INTECH Open Access Publisher, 2012.
  3. Bernstein LH, David G, Coifman RR. Generating Evidence Based Interpretation of Hematology Screens via Anomaly Characterization. Open Clin Chem J 2011; 4:10-16
  4. Bernstein LH. Automated Inferential Diagnosis of SIRS, sepsis, septic shock. Medical Informatics View.
  5. Bernstein LH, David G, Coifman RR. The Automated Nutritional Assessment. Nutrition  2013; 29: 113-121


Other related articles published in this Open Access Online Scientific Journal include the following: 

Biomarkers and risk factors for cardiovascular events, endothelial dysfunction, and thromboembolic complications

Commentary on Biomarkers for Genetics and Genomics of Cardiovascular Disease: Views by Larry H Bernstein, MD, FCAP

Summary – Volume 4, Part 2: Translational Medicine in Cardiovascular Diseases

Pathophysiological Effects of Diabetes on Ischemic-Cardiovascular Disease and on Chronic Obstructive Pulmonary Disease (COPD)

Assessing Cardiovascular Disease with Biomarkers

Endothelial Function and Cardiovascular Disease

Adenosine Receptor Agonist Increases Plasma Homocysteine

Inadequacy of EHRs

Innervation of Heart and Heart Rate

Biomarker Guided Therapy


The Union of Biomarkers and Drug Development

Natriuretic Peptides in Evaluating Dyspnea and Congestive Heart Failure

Epilogue: Volume 4 – Translational, Post-Translational and Regenerative Medicine in Cardiology

Atherosclerosis Independence: Genetic Polymorphisms of Ion Channels Role in the Pathogenesis of Coronary Microvascular Dysfunction and Myocardial Ischemia (Coronary Artery Disease (CAD))

Erythropoietin (EPO) and Intravenous Iron (Fe) as Therapeutics for Anemia in Severe and Resistant CHF: The Elevated N-terminal proBNP Biomarker

Biomarkers: Diagnosis and Management, Present and Future

Genetic Analysis of Atrial Fibrillation

Landscape of Cardiac Biomarkers for Improved Clinical Utilization

Fractional Flow Reserve (FFR) & Instantaneous wave-free ratio (iFR): An Evaluation of Catheterization Lab Tools for Ischemic Assessment

Dealing with the Use of the High Sensitivity Troponin (hs cTn) Assays

Cardiotoxicity and Cardiomyopathy Related to Drugs Adverse Effects

Amyloidosis with Cardiomyopathy

Accurate Identification and Treatment of Emergent Cardiac Events

The potential contribution of informatics to healthcare is more than currently estimated

Realtime Clinical Expert Support

Metabolomics is about Metabolic Systems Integration

Automated Inferential Diagnosis of SIRS, sepsis, septic shock

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Calcium Channel Blocker Potential for Angina

Larry H. Bernstein, MD, FCAP, Curator








File:Pranidipine structure.svg

Pranidipine , OPC-13340, FRC 8411


NDA Filing in Japan

A calcium channel blocker potentially for the treatment of angina pectoris and hypertension.


CAS No. 99522-79-9

  • Molecular FormulaC25H24N2O6
  • Average mass 448.468


see dipine series………..




Der Pharmacia Sinica, 2014, 5(1):11-17


IUPAC name

methyl (2E)-phenylprop-2-en-1-yl 2,6-dimethyl-4-(3-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylate
Other names

2,6-dimethyl-4-(3-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylic acid O5-methyl O3-[(E)-3-phenylprop-2-enyl] ester
99522-79-9 Yes
ChEMBL ChEMBL1096842 
ChemSpider 4940726 
Jmol interactive 3D Image
MeSH C048161
PubChem 6436048




Process for the preparation of 1,4 – dihydropyridines and novel 1,4-dihydropyridines useful as therapeutic agents [US2003230478] 2003-12-18
Advanced Formulations and Therapies for Treating Hard-to-Heal Wounds [US2014357645] 2014-08-19 2014-12-04
Protein Carrier-Linked Prodrugs [US2014323402] 2012-08-10 2014-10-30
sGC STIMULATORS [US2014323448] 2014-04-29 2014-10-30
Agonists of Guanylate Cyclase Useful For the Treatment of Gastrointestinal Disorders, Inflammation, Cancer and Other Disorders [US2014329738] 2014-03-28 2014-11-06
ROR GAMMA MODULATORS [US2014343023] 2012-09-18 2014-11-20
High-Loading Water-Soluable Carrier-Linked Prodrugs [US2014296257] 2012-08-10 2014-10-02 



Synthesis, isolation and use of a common key intermediate for calcium antagonist inhibitors

Neelakandan K.a,b, Manikandan H.b , B. Prabhakarana*, Santosha N.a , Ashok Chaudharia *, Mukund Kulkarnic , Gopalakrishnan Mannathusamyb and Shyam Titirmarea
a API Research Centre, Emcure Pharmaceutical Limited, Hinjawadi, Pune, India bDepartment of Chemistry, Annamalai University, Chidhambaram, India cDepartment of Chemistry, Pune University, Pune, India _________________________________________________________________________________

Pelagia Research Library      Der Pharmacia Sinica, 2014, 5(1):11-17


The compound (3) synthesized from Nitrobenzaldehyde, tertiary butyl acetoacetate and piperidine can be used as a common intermediate for the production of calcium channel blockers like Nicardipine hydrochloride (1) and Pranidipine hydrochloride (2) with high purity.


The last twenty years have witnessed discoveries of calcium antagonists associated with multicoated pharmacodynamics potential which include not only antihypertensive and antiarrhythmic effects of the drugs but also action against excessive calcium entry in the cell of cardiovascular system and subsequent cell damage. Among many classes of calcium channel blockers, 1,4-dihydropyrimidine based drug molecules represented by Felodipine, Clevidipine, Benidipine, Nicardipine and Pranidipine are by far the best to reduce systemic vascular resistance and arterial pressure.

The reported synthetic approaches however proceed with complicated work ups, laborious purification procedures, highly expensive chemicals and low overall yields. (Scheme-I).

Synthetic scheme of Nicardipine and Pranidipine In view of the draw backs associated with previous synthetic approaches there is a strong need for environmentfriendly high yielding process applicable to the multi-kilogram production of calcium antagonist inhibitors. Herein, we report a scalable synthesis for Nicardipine hydrochloride (1) and Pranidipine hydrochloride (2) in fairly high overall yield using key intermediate 3-nitro benzylidene acid (3).Compound (3) was synthesized in two steps using 3-nitrobenzaldehyde, tertiary butyl acetoacetate and piperidine as a base to furnish tertiary butyl ester derivative (10). This was followed by hydrolysis of (10) in TFA and DCM to furnish compound (3) which would serve as a precursor for synthesis of versatile calcium antagonist inhibitors (Scheme-II).

Reported routes for synthesis of Benidipine,1,2 Lercanadipine,3-6 Nimodipine,7-11 Barnidipine12-14 and Manidipine15-16 were explored in our laboratory which involve reaction of nitro benzaldehyde with tertiary butyl acetoacetate using piperidine as a base to get tertiary butyl ester derivative (10). This is further treated with respective reagents to get various calcium channel blockers as shown in scheme 4. Since reported procedures involve in-situ generation of intermediate (3) and its reaction with corresponding fragments, it results in the formation of by-products which ultimately decrease the yield and increase the cost of API.

A novel process of manufacturing benzylidine acid derivative (3) was developed. Use of this intermediate was demonstrated by synthesis of Nicardipine and Pranidipine. This protocol may be employed for synthesis of other calcium channel blockers. In conclusion, a highly efficient, reproducible and scalable process for the synthesis of calcium channel blockers has been developed using (3) as the key intermediate.


[1] US 63 365 (Kyowa Hakko; appl.15.4.1982; J-prior.17.4.1981). [2] US 4 448 964 (Kyowa Hakko;15.5.1984; J-prior.17.4.1981). [3] Leonardi, A. et al.: Eur. J. Med.Chem. (EJMCA5) 33,399 (1988). [4] EP 153 016 (Recordati Chem. and Pharm.; appl. 21.1.1985; GB-prior. 14.2.1984). [5] US 4 705 797 (Recordati;10.11.1987; GB-prior. 14.2.1984). [6] WO 9 635 668 (Recordati Chem. and Pharm.; appl. 9.5.1996; I-prior. 12.5.1995). [7] DOS 2 117 571 (Bayer; appl. 10.4.1971). [8] DE 2 117 573 (Bayer; prior.10.4.1971) [9] US 3 799 934 (Bayer;26.3.1974;D-prior.10.4.1971). [10] US 3 932 645 (Bayer;13.1.1976;D-prior.10.4.1971). [11] Meyer, H. et al.: Arzneim.-Forsch. (ARZNAD) 31, 407 (1981); 33, 106 (1983). [12] DE 2 904 552 (Yamanouchi Pharm.; appl. 7.2.1979; J-prior.14.2.1978). [13] US 4 220 649 (Yamanouchi;2.9.1980; J-prior.14.2.1978). [14] CN 85 107 590( Faming Zhuanli Sheqing Gonhali S.; appl. 11.10.1985; J-prior.24.1.1985). [15] EP 94 159 (Takeda; appl. 15.4.1983; J-prior. 10.5.1982). [16] US 4 892 875 (Takeda;9.1.1990; J-prior. 10.5.1982, 11.1.1983).


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Blood Pressure Lowering

Larry H. Bernstein, MD, FCAP, Curator



Data analysis and publication of landmark NIH blood pressure study confirm that lower blood pressure target can reduce cardiovascular disease, deaths


NIH-supported researchers are reporting more details on a landmark study that announced preliminary findings in September showing a lower blood pressure target can save lives and reduce the risk of cardiovascular disease in a group of non-diabetic adults 50 years and older with high blood pressure. Results of the Systolic Blood Pressure Intervention Trial (SPRINT) appear in the current online issue of the New England Journal of Medicine and were discussed today at the American Heart Association 2015 Scientific Sessions in Orlando.

The study confirms that, in adults 50 years and older with high blood pressure, targeting a systolic blood pressure of less than 120 millimeters of mercury (mm Hg) reduced rates of cardiovascular events, such as heart attack and heart failure, as well as stroke, by 25 percent. Additionally, this target reduced the risk of death by 27 percent—as compared to a target systolic pressure of 140 mm Hg.

“SPRINT is part of a proud legacy of NIH-funded clinical trials that will change clinical practice and save lives for decades to come. These results reinforce the compelling public health importance of enhancing the awareness, treatment and control of hypertension in this country and around the world,” said Gary H. Gibbons, M.D., director of the National Heart, Lung, and Blood Institute (NHLBI), the primary sponsor of SPRINT.

The SPRINT study, which began in the fall of 2009, included more than 9,300 participants age 50 and older, recruited from about 100 medical centers and clinical practices throughout the United States and Puerto Rico. About 36 percent of participants were women, 58 percent were white, 30 percent were African-American, and 11 percent were Hispanic. The SPRINT study did not include patients with diabetes, prior stroke, or polycystic kidney disease, as other NIH trials were studying those particular populations. Approximately 28 percent were 75 or older and 28 percent had chronic kidney disease. The study tested a strategy of using blood pressure medications to achieve the targeted goals of less than 120 mm Hg (intensive treatment group) versus 140 mm Hg (standard treatment group). The NIH stopped the blood pressure intervention in August—a year earlier than planned—after it became apparent that this more intensive intervention was beneficial.

“When the benefits of the stronger intervention became apparent in SPRINT, we made a commitment to rapid public health communication and peer-reviewed publication of the study results,” Dr. Gibbons said. “We are pleased to present the details of the study’s potentially lifesaving findings at this time.”

In their report, investigators provided detailed data showing that both cardiovascular deaths and overall deaths were lower in the intensive treatment group.

Certain types of serious consequences were more common in the intensive group, including low blood pressure, fainting, electrolyte abnormalities, and acute kidney damage. However, other serious adverse events associated with lower blood pressure, such as slow heart rate and falls with injuries, did not increase in the intensive group. In patients with chronic kidney disease, there was no difference in the rate of serious decline in kidney function between the two blood pressure goal groups.

“The benefits of more intensive blood pressure lowering exceeded the potential for harm, regardless of gender or race/ethnicity,” said study co-author Paul Whelton, M.D., of Tulane University School of Public Health and Tropical Medicine in New Orleans, Louisiana. He is chair of the SPRINT Steering Committee.

In addition to its primary cardiovascular outcome, the study continues to examine kidney disease, cognitive function, and dementia among the SPRINT participants; however, these results are not yet available as additional information will be collected and analyzed over the next year.

“Although the study provides strong evidence that a lower blood pressure target saves lives, patients and their health care providers may want to wait to see how guideline groups incorporate this study and other scientific reports into any future hypertension guidelines. In the meantime, patients should talk to their health care providers to determine whether this lower goal is best for their individual care,” said study co-author Lawrence Fine, M.D., Chief, Clinical Applications and Prevention Branch at NHLBI.

“It’s also important to remember that healthy lifestyle changes can make a difference in controlling high blood pressure,” Dr. Fine added. He emphasized the importance of following a healthy diet, being physically active, maintaining a healthy weight, as well as learning to check your blood pressure.

In addition to primary sponsorship by the NHLBI, SPRINT is co-sponsored by the NIH’s National Institute of Diabetes and Digestive and Kidney Diseases, the National Institute of Neurological Disorders and Stroke, and the National Institute on Aging.

Part of the National Institutes of Health, the National Heart, Lung, and Blood Institute (NHLBI) plans, conducts, and supports research related to the causes, prevention, diagnosis, and treatment of heart, blood vessel, lung, and blood diseases; and sleep disorders. The Institute also administers national health education campaigns on women and heart disease, healthy weight for children, and other topics. NHLBI press releases and other materials are available online at

About the National Institutes of Health (NIH): NIH, the nation’s medical research agency, includes 27 Institutes and Centers and is a component of the U.S. Department of Health and Human Services. NIH is the primary federal agency conducting and supporting basic, clinical, and translational medical research, and is investigating the causes, treatments, and cures for both common and rare diseases. For more information about NIH and its programs, visit

NIH…Turning Discovery Into Health®

Landmark NIH study shows intensive blood pressure management may save lives

Embargoed for Release:

September 11, 2015, 10:30 AM EDT

Lower blood pressure target greatly reduces cardiovascular complications and deaths in older adults

More intensive management of high blood pressure, below a commonly recommended blood pressure target, significantly reduces rates of cardiovascular disease, and lowers risk of death in a group of adults 50 years and older with high blood pressure. This is according to the initial results of a landmark clinical trial sponsored by the National Institutes of Health called the Systolic Blood Pressure Intervention Trial (SPRINT). The intervention in this trial, which carefully adjusts the amount or type of blood pressure medication to achieve a target systolicpressure of 120 millimeters of mercury (mm Hg), reduced rates of cardiovascular events, such as heart attack and heart failure, as well as stroke, by almost a third and the risk of death by almost a quarter, as compared to the target systolic pressure of 140 mm Hg.

“This study provides potentially lifesaving information that will be useful to health care providers as they consider the best treatment options for some of their patients, particularly those over the age of 50,” said Gary H. Gibbons, M.D., director of the National Heart, Lung, and Blood Institute (NHLBI), the primary sponsor of SPRINT. “We are delighted to have achieved this important milestone in the study in advance of the expected closure date for the SPRINT trial and look forward to quickly communicating the results to help inform patient care and the future development of evidence-based clinical guidelines.”

High blood pressure, or hypertension, is a leading risk factor for heart disease, stroke, kidney failure, and other health problems. An estimated 1 in 3 people in the United States has high blood pressure.

The SPRINT study evaluates the benefits of maintaining a new target for systolic blood pressure, the top number in a blood pressure reading, among a group of patients 50 years and older at increased risk for heart disease or who have kidney disease. A systolic pressure of 120 mm Hg, maintained by this more intensive blood pressure intervention, could ultimately help save lives among adults age 50 and older who have a combination of high blood pressure and at least one additional risk factor for heart disease, the investigators say.

The SPRINT study, which began in the fall of 2009, includes more than 9,300 participants age 50 and older, recruited from about 100 medical centers and clinical practices throughout the United States and Puerto Rico. It is the largest study of its kind to date to examine how maintaining systolic blood pressure at a lower than currently recommended level will impact cardiovascular and kidney diseases. NIH stopped the blood pressure intervention earlier than originally planned in order to quickly disseminate the significant preliminary results.

The study population was diverse and included women, racial/ethnic minorities, and the elderly.  The investigators point out that the SPRINT study did not include patients with diabetes, prior stroke, or polycystic kidney disease, as other research included those populations.

When SPRINT was designed, the well-established clinical guidelines recommended a systolic blood pressure of less than 140 mm Hg for healthy adults and 130 mm Hg for adults with kidney disease or diabetes. Investigators designed SPRINT to determine the potential benefits of achieving systolic blood pressure of less than 120 mm Hg for hypertensive adults 50 years and older who are at risk for developing heart disease or kidney disease.

Between 2010 and 2013, the SPRINT investigators randomly divided the study participants into two groups that differed according to targeted levels of blood pressure control. The standard group received blood pressure medications to achieve a target of less than 140 mm Hg. They received an average of two different blood pressure medications. The intensive treatment group received medications to achieve a target of less than 120 mm Hg and received an average of three medications.

“Our results provide important evidence that treating blood pressure to a lower goal in older or high-risk patients can be beneficial and yield better health results overall,” said Lawrence Fine, M.D., chief, Clinical Applications and Prevention Branch at NHLBI. “But patients should talk to their doctor to determine whether this lower goal is best for their individual care.”

The study is also examining kidney disease, cognitive function, and dementia among the patients; however, those results are still under analysis and are not yet available as additional information will be collected over the next year.  The primary results of the trial will be published within the next few months.

In addition to primary sponsorship by the NHLBI, SPRINT is co-sponsored by the NIH’s National Institute of Diabetes and Digestive and Kidney Diseases, the National Institute of Neurological Disorders and Stroke, and the National Institute on Aging.

Supplemental Information

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Innervation of Heart and Heart Rate

Writer and Curator: Larry H Bernstein, MD, FCAP



The heart is a four-chambered 350 gm semi-oval muscular organ composed of syncytial myocardium, innervated by the vagus nerve with a sino-atrial (SA) and a atrial ventricular (AV) node.  The blood circulates through it by way of the pulmonary artery and aorta, carrying blood away from the ventricles, to the lungs and the systemic circulation, respectively, and two veins, the vena cava and pulmonary, carrying blood to the atria from the systemic circulation and lungs, respectively.  The coronary arterial supply is the left anterior and left circumflex artery, and posteriorly, the right coronary artery, supplied by the aorta.  Much of the pathology has been referred to in the introduction, except for the molecular pathology of atherosclerosis, which has been well covered in this journal. The chambers are divided centrally by the interventricular septum, which is not completely closed in the blue-baby syndrome, which was repaired surgically by Helen Taussig and Richard Bing.  The piece that follows is primarily directed to the sympathetic innervation of the heart, variation in heart rate, and exercise or reaction to external threats.

What are the common observable events that stimulate or relax the heart:

  1. Running or a treadmill test
  2. Rowing or arm movement exercise
  3. A whole body workout
  4. Yoga or Ayurveda
  5. Sleep – normal or disruptive

Some things that can cause a disruption of balance in integrated circulation, neural innervation, innate immune and hormonal response are:

  1. Traumatic experience and/or Injuries
  2. Climate and seasonal changes
  3. Age
  4. Emotions

The basis for the physiological distress has long been the primary basis for acupuncture, holistic and transcendental medicine, and stress management.

I shall here examine the experimental work that supports such an approach – in principle.

Seattle Heart Watch: Initial Clinical, Circulatory and Electrocardiographic Responses to Maximal Exercise

Robert A Bruce, G0 Gey, Jr., Mn Cooper, Ld Fisher, Dr Peterson
Amer J Cardiol 1974; 33(4): 459-469.

A network of 15 maximal exercise testing facilities in four teaching hospitals, 10 private offices and clinics and an industrial medical department was organized in July 1971 to study prospectively the antecedents of myocardial infarction and sudden cardiac death. Within 18 months 2,332 men were tested: 1,275 healthy “normal” subjects, 97 with prior myocardial infarction, 306 with angina pectoris, 193 with hypertension and 461 with various mutually exclusive combinations of these diagnoses; among these clinical groups were five patients who had had a prior episode of ventricular fibrillation.
Historical, physical and laboratory data were recorded on self-teaching printed forms, with normal, borderline and abnormal responses arranged in three columns. Classification with respect to “unlikely,” “questionable” or “likely” risk of future cardiac events was assessed from the highest tally of items in these columns.
Analysis showed computer-averaged S-T segment responses were more consistent and reliable predictors than visual interpretations. Cardiac manifestations in healthy men varled with age and risk assessment, and in patients with cardiovascular disease varied with diagnosis and natural history of disease. Many significant differences provided insights into mechanisms of impaired cardiac function in relation to type of clinical disease. Testing was responsible for one post-exertional cardiac arrest. Recovery was effected promptly by defibrillation; there was no mortality.

Normal and Abnormal Heart Rate Responses to Exercise

  1. Kirk Hammond and Victor F. Froelicher
    Prog Cardiovasc Dis 1985; XXVII(4) (January/February), pp 27l-296

Of the many factors ultimately important in determining the cardiac output, the heart rate is certainly the easiest to measure. By analysis of the heart rate response to exercise in a variety of disease states we felt that the interrelationships of inotropic state, stroke volume, autonomic dysfunction, and myocardial disease could be clarified. This paper reviews the normal and abnormal heart rate responses to exercise.

The normal heart rate is determined by the frequency of depolarization of specialized cells within the sino-atrial node (S-A node). The S-A node, the vestigal sinus venosus, lies in the posterior portion of the heart near the demarcation between the right atrium and the superior vena cava. In about 80% of humans it receives its primary source of blood from a branch of the right coronary artery. Unlike other myocardial cells, the specialized cells of the S-A node have a slow sodium channel and a low resting potential which give these cells their special property. The slowly rising diastolic depolarization (stage four) leads to a rhythmic slow rising action potential.

The autonomic nervous system plays a key role in the regulation of heart rate (Fig 1). The sympathetic nervous system input to the heart originates in a nucleus in the medulla oblongata. Stimulation of this area with implanted electrodes results in increased heart rate and systemic vascular resistance due to increased sympathetic output. Axons from these nuclei descend to the sympathetic trunk via the intermediolateral columns of the spinal cord. From their synapses in cervical ganglia, postganglionic fibers directly innervate the atrial and ventricular musculature, the S-A node, and the A-V node. The effector neurotransmitter is norepinephrine and the receptors are of the beta adrenergic type. There is evidence from competitive binding studies that the postganglionic fibers are predominantly associated with type I beta receptors. The parasympathetic influence to the S-A node and the myocardium originates from nuclei very near the origin of the sympathetic nerves. From the motor nuclei of the vagus and the nucleus solitarius come fibers that form part of the vagus nerve. These fibers terminate at ganglia in the wall of the heart. The postganglionic cholinergic fibers end mostly near the S-A node and the A-V node; there is little evidence for the distribution of parasympathetic nerves to the ventricular myocardium although cholinergic muscarinic receptors have been characterized. In normal conditions there exists a well balanced autonomic tone influencing the S-A node.

There is a complex interrelation among many systems to determine the autonomic tone at the S-A node (Fig 2). [Arterial mechanoreceptors of the carotid sinus and aortic arch respond to changes in arterial pressure and result in appropriate adjustment in the sympathetic and vagal outflow to the heart and resistance and capacitance vessels. (Reprinted with permission from Shepherd JT, Van Houlte PM: The Human Cardiovascular System, Facts and Concepts. New York, Raven Press, 1979).]

There are cortical inputs to the medullary centers; for example, fear results in tachycardia by this pathway. Visceral afferent inputs increase parasympathetic tone resulting in bradycardia. Several reflexes are present for homeostasis. For example, the baroreflex is important in sensing changes in blood pressure and increasing or decreasing the heart rate via autonomic influences at the S-A node to maintain appropriate cardiac output.

Arterial mechanoreceptors of the carotid sinus and aortic arch respond to changes in arterial pressure and result in appropriate adjustment in the sympathetic and vagal outflow to the heart and resistance and capacitance vessels. (Reprinted with permission from Shepherd JT, Van Houlte PM: The Human Cardiovascular System, Facts and Concepts. New York, Raven Press, 1979).

Although the importance of autonomic influence is well accepted in the usual cardioacceleration to exercise, the role of the recovery or deceleration of heart rate following exercise may not be influenced by autonomic input. Six men were studied after peak treadmill exercise. To assess the contribution of autonomic factors in heart rate recovery, the men were given atropine, propranolol, or both agents. It was found that exponential cardio-deceleration occurred under each experimental condition. They concluded that heart rate recovery after exercise is regulated by changes in venous return mediated through atrial stretch receptors of pacemaker tissue. This study implies that deceleration depends primarily on factors intrinsic to the intact circulation that are independent of autonomic control.

The control of heart rate is complex; autonomic tone, central and peripheral reflexes, hormonal influences, and factors intrinsic to the heart are all important. Although easily measured, the heart rate reflects an integrated physiologic response.

The physiologic response to exercise depends on the type of exercise performed; the two major types are isometric and isotonic. Creating muscle tension with no movement against resistance is a pure form of isometric exercise; this results in increased muscle mass and strength. Isotonic exercise is the repetitive, rhythmic movement of large muscle masses against little resistance, known also as dynamic or aerobic exercise. Although most activities involve degrees of both, running is predominantly dynamic, and weight lifting is predominantly isometric.

Bezucha and colleagues investigated the cardiovascular responses to isometric (static) exercise (leg extension) and compared these to those observed during static-dynamic exercise (one arm cranking) and dynamic exercise (leg cycling) in normal men. Heart rate responses to these three tasks were markedly different with static exercise (holding a 30% of maximum voluntary contraction for 3 minutes) resulting in a mean heart rate of 110 + 6 compared with 164 + 4 beats/min in bicycle exercise at 80% of Vo max. Cardiac outputs were raised in all three activities in a proportional manner: 6.8 + 0.7 for static, 10.8 f 0.7 for arm cranking, and 31.9 + 1.0 L/min for bicycling. Stroke volume did not significantly change in the static or combined static-dynamic exercises. The increases in cardiac output were primarily the result of increases in heart rate. This study demonstrates the predominant pressor response and modest cardio-acceleration of isometric exercise.

Longhurst and coworkers, examined the response to acute and chronic exercise in two groups of athletes who typify the two major types of exercise: long distance runners (dynamic) and weight lifters (isometric). The runners responded to isometric exercise with lower double products than the weight lifters. The end-diastolic volume index (evaluated by echocardiography) in the runners was greater than control subjects both at rest and with exercise. In contrast, the weight lifters’ responses were similar to weight matched controls. Not only is the type of exercise an important determinant of acute physiologic response, but chronic static exercise results in physiologic responses that are no different from the responses of sedentary men.

Dynamic exercise, also called isotonic or aerobic, involves the rapid movement of large muscle masses that results in the need for the body to respond with increased ventilation to increase oxygen consumption. Such exercise is called aerobic since it must be performed by using oxygen. The heart must increase its output and performs flow work rather than pressure work. The response to dynamic muscular exercise consists of a complex series of cardiovascular adjustments designed to:

(1) see that active muscles receive a blood supply appropriate to their metabolic needs;

(2) dissipate the heat generated by active muscles; and,

(3) maintain the blood supply to the brain and the heart.

The regulation of the circulation during exercise involves the four following adaptations?

  • Local
  • Nervous adaptations
  • Humoral adaptations
  • Mechanical adaptations

The relationship of pressure, flow, and resistance in rigid tubes is defined by Poiseuille’s law. This law states that resistance is proportional to pressure divided by flow. Peripheral resistance increases in the tissues that do not function in the performance of the ongoing exercise and decreases in active muscle. The result is a decrease in systemic vascular resistance. While pressure only increases mildly, flow can increase by as much as five times during dynamic exercise. Since flow increases much more than pressure, the result is a decrease in systemic resistance. Another mechanical adaptation occurs when the increasing venous return dilates the left ventricle and cardiac function is enhanced via the Frank-Starling mechanism.

There is a highly predictable relationship between total body oxygen consumption and both the cardiovascular and respiratory responses to exercise (Fig 4). [ (A) The linear relationship between heart rate and oxygen uptake. The data was collected from 86 adult male and female subjects. (B) The linear relationship between cardiac output and oxygen uptake. C The data was collected from 23 adult male and female subjects. (C) The linear relationship between minute ventilation and oxygen uptake. ] The data was collected from 225 subjects.  (Reprinted with permission.) Both parameters increase linearly with increasing oxygen consumption until maximal oxygen consumption is approached.

In summary, the type of exercise is an important determinant of both acute and chronic cardiovascular responses. Isometric exercise can be viewed as a pressure load and dynamic exercise as a volume load to the left ventricle. The acute physiological adjustments to dynamic exercise include peripheral vasodilation in exercising muscle, neural mediated increases in sympathetic tone to the heart and the periphery, the release of catecholamines from the adrenal medulla, and changes in venous return due to mechanical and humoral factors. A linear relationship exists between the consumption of oxygen and cardiac output and minute ventilation such that the work performed is highly correlated with the amount of blood pumped and the oxygen consumed.

An increase in heart rate is a major factor contributing to the exercise-induced increased cardiac output. Bowditch demonstrated that the time interval between beats is a determinant of the force of myocardial contraction. This has been called the frequency-force relationship (Fig 5). [The frequency force relationship is demonstrated by a sudden increase in beat frequency in papillary muscle fixed for isometric contraction. A slow increase in isometric tension results from the change in rate implying in increased contractile state. Each vertical line represents an isometric contraction. (Reprinted with permission of W.B. Saunders.)] The increased tension that accompanies an increased heart rate is the result of increased contractility. Although the mechanism of this phenomenon is not known, it may have to do with calcium availability to contractile elements. Thus an increase in heart rate results in an increase in the force of contraction.

Variations in and Significance of Systolic Pressure During Maximal Exercise (Treadmill) Testing: Relation to Severity of Coronary Artery Disease and Cardiac Mortality

John B. Irving, Robert A. Bruce,, Timothy A. Derouen
Amer J Cardiol 1977; 39: 841-848.

Variations in clinical noninvasive systolic pressure at the point of symptom-limited exercise on a treadmill were examined in six groups of subjects: 5,459 men and 749 women classified into three categories each. Among the men, 2,532 were asymptomatic healthy, 592 were hypertensive and 1,586 had clinical manifestations of coronary heart disease (that is, typical angina pectoris, prior myocardial Infarction or sudden cardiac arrest with resuscitation). Among the women, 244, 158 and 347 were in the corresponding clinical categories. None had had cardiac surgery; all had follow-up status ascertained by periodic mail questionnaires.
Reported deaths were reviewed and classified by three cardiologists; 140 deaths were attributed to coronary heart disease, 118 of them in the men classified as having coronary heart disease. The majority of maximal systolic blood pressure readings were reported to the nearest centimeter rather than millimeter of pressure. Retesting of 156 persons from 1 to 32 months later showed that pressure values agreed within 10 percent in two thirds, the overall mean difference was only 8.6 mm Hg and the correlation at maximal exercise was superior to that of the resting observations just before exercise. Hypertensive patients had a significantly greater body weight than normotensive persons. Among men, the lowest maximal systolic pressure was observed in the group with coronary heart disease; among women, the lowest mean pressure was found in the healthy group. Patients with coronary heart disease were slightly older, and only the women showed a significant correlation in maximal pressure with age. Only 5 percent of the variation in maximal systolic pressure in the patients with coronary heart disease was due to a shortened duration of exercise. Maximal systolic pressures correlated fairly well (r = 0.46 to 0.68 for the various groups) with resting systolic pressure, and this relation was independent of the diagnosis of cardiovascular disease in both men and women. Relations between pressure and the number of stenotic coronary arteries and Impaired ejection fraction at rest were examined in 22 men without and 162 men with coronary artery disease. Lower maximal systolic pressures were often associated with two or three vessel disease or reduced ejection fraction, or both.

The prognostic value of maximal systolic pressure for subsequent death due to coronary heart disease was examined in the men with coronary heart disease. The annual rate of sudden cardiac death decreased from 97.9 per 1,000 men to 25.3 and 6.6 per 1,000 men as the range of maximal systolic pressure increased from less than 140 to 140 to 199 and to 200 mm Hg or more, respectively. Cardiomegaly, Q waves in the resting electrocardiogram and persistent postexertional S-T depression were more common in men with the lowest systolic pressure at maximal exercise.

Circulatory Adjustments to Dynamic Exercise and Effect of Physical Training in Normal Subjects and in Patients With Coronary Artery Disease

Jan Praetorius Clausen
Prog Cardiov Dis 1976; XVIII(6): 459-496

The present paper focuses upon the importance of peripheral circulatory alterations during adjustments to exercise and training. Although training results in central circulatory adaptations and may also improve left ventricular function, the prime importance of such adaptations as regards the circulatory and metabolic response to training will be questioned. The thesis that increased maximal exercise capacity can at least in part be attributed to local alterations in the trained muscles will be presented and analyzed. While it is accepted that maximal oxygen uptake is limited by the blood oxygen transport capacity, it will be postulated that the primary event normally responsible for an enhanced oxygen supply after training is an increased ability to reduce resistance to blood flow in exercising muscles rather than improved performance of the central pump.

adjustment to exercise is limited to factors pertinent to physical training of patients with CAD. More detailed accounts of the normal response to exercise can be found in recent books or reviews.

  1. Astrand, P-O, Rodahl K: Textbook of Work Physiology. New York, McGraw-Hill, 1970
  2. Ekblom B, Hermansen L: Cardiac outputs in athletes. J Appl Physiol 25:619, 1968
  3. Christensen EH: Beitrlge zur Physiologie schwerer kijrperlicher Arbeit. Arbeits physiol 4:470, 1931
  4. Saltin B, Blomqvist G, Mitchell JH, et al: Response to exercise after bed rest and after training. Circulation 38 (Suppl 7): 1, 1968
  5. Clausen JP, Klausen K, Blomqvist G, et al. Central and peripheral circulatory changes after training of the arms or legs. Am J Physiol 225:675, 1973

In connection with patients with CAD, only one type of muscular work is of interest; namely, rhythmic or dynamic exercise, in which a considerable part of the skeletal muscle mass is active. This applies to naturally occurring physical activity. Only these types of activity will be referred to and only at work intensities that can be continued for 3-5 min or more.

Dynamic muscular exercise is characterized by a high metabolic rate in the muscle cells with the skeletal muscle functioning in a manner similar to the myocardium, with regularly alternating contraction and relaxation phases. The mechanical energy expended is grossly proportional to the force and the frequency of contraction, and it is derived from the breakdown of adenosine triphosphate (ATP) and creatine phosphate (CP). Only a limited number of a muscle’s fibers, and thus, of its maximal contractile power, can be used in dynamic work continuing for several minutes. During maximal exercise on a bicycle ergometer with a pedaling frequency of 60 rpm, about 15%-2% of the maximal isometric strength of the quadriceps muscle is mobilized. This is thought related to the fact that skeletal muscle, in contrast to myocardium, is composed of several types of fibers with different enzymatic characteristics.29 Some fibers are similar to cardiac muscle being rich in oxidative intramitochondrial enzymes connected to the citric acid cycle, the fatty acid cycle, and the respiratory chain. These are the classical “red” muscle fibers. At the other end of a continuous spectrum is the typical “white” muscle fiber, with a high content of enzymes necessary for anaerobic glycolysis, but containing few mitochondria. Due to their great capability for aerobic metabolism, red fibers sustain rhythmic contractions for long periods of time, whereas the anaerobic white fibers require longer restitution phases even after short periods of activity.

Oxygen extraction per milliliter of blood perfusing the muscle may increase three- to fourfold, and the enhanced muscle blood flow (MBF) is responsible for the remainder of the augmented oxygen uptake. In human muscle, maximal MBF is in the order of 70-100 ml X 100 g-r X min--1 against a resting value of 2-5 ml X 100 g-r X min--1. The increase in MBF is locally controlled by release of vasodilator metabolites and thereby closely geared to the metabolic demands. Muscle blood flow per unit weight of muscle is closely related to the relative work load; i.e., percentage of maximal work load. The metabolites responsible for the exercise-induced vasodilation and hyperemia in muscle are not yet conclusively identified. The finding that both MBF and ATP-CP depletion are related to the relative work load supports the speculation that split products from high energy phosphates may be involved.

During strenuous exercise, VO2  can attain individually varying maximal values, typically ranging from 2.0 to 6.0 1 02/min. The maximal oxygen consumption (VO2 max) is a highly reproducible measure of a given subject’s capability to perform this type of exercise, and it constitutes a useful physiologic reference standard. The conditions required to obtain VO2 max, and its physiologic implications have recently been reviewed in detail by Rowe and by Hermansen. The VO2 max  for a given type of work is normally achieved at a work intensity that can be sustained for at least 3 min, but will cause complete exhaustion within 5-10 min.  At this intensity of exercise, the cardiovascular functional capacity with respect to increase in cardiac output (Q), widening of systemic arteriovenous oxygen difference (AVDO2), and elevation of heart rate (HR) will be challenged maximally for the given type of exercise. However, the relative contribution of Q and AVDO2.

The above description of the normal central and peripheral circulatory adjustment to exercise can be recapitulated as follows:

During dynamic exercise, Q increases in direct proportion to the augmentation of 30,. The increase in Q is directed to exercising skeletal muscles, to the myocardium and-if exercise is continued for more than approximately 5 min-also to the skin. Blood flow to most “nonexercising” tissues (SBF, RBF,
and noncontracting muscles) is reduced due to a general sympathetic vasoconstriction. At submaximal levels, muscle blood flow per unit tissue,
the degree of peripheral vasoconstriction, the acceleration of HR, and in consequence, the increase in myocardial blood flow and oxygen consumption are all functions of the relative V02 ; i.e., the actual VO2 expressed as a percentage of the highest achievable V02 for the given type of exercise.

Most patients with CAD who have been included in exercise and training studies have had healed myocardial infarction and/or stable angina pectoris and have been between 35 and 65 years of age. Both the aging process and myocardial lesions contribute to the modification of the circulatory response to exercise in this group, as compared to healthy young people. In advanced age-especially after 60 years-the circulation tends to become hypokinetic; i.e., Q/VO2 is reduced.  The decline of Q in l/min is almost the same during submaximal exercise as at rest, and thus the increase in Q with VO2 is essentially the same in older as in younger subjects. Stroke volume is lower at a given VO2 , while arterial blood pressures are higher; Q, HR, and VO2 max decline with aging.

Although patients with angina pectoris often exhibit a more profound impairment of left ventricular function and of working capacity than patients with CAD without angina, there seems not to be any specific differences in their central or peripheral circulatory response to exercise. Accordingly, the abnormalities in hemodynamic adaptations in a patient with angina pectoris are present also at workloads that do not provoke angina pectoris.

From the point of view of an exercise physiologist, the patient with angina pectoris is peculiar in that his capacity for dynamic work is not limited by his total body VO2 max, but by VO2 max in myocardial regions supplied by narrowed coronary arteries. If pain is prevented by prophylactic administration of nitroglycerin, a patient with angina pectoris can exercise longer at a given work load or achieve higher workloads and thus obtain a higher VO2 max.

The circulatory adjustment to exercise in patients with CAD typically differs from that of normal subjects in that the maximal values for Q (and thus for VO2), for HR, and for blood pressures are lower. During submaximal exercise, the relation between Q and VO2 tends to be reduced. Moreover, most of the patients with CAD exhibit signs of left ventricular failure during exercise, including a decrease in SV at higher workloads, reduced myocardial contractility, and increased LVEDp. Nonetheless, the peripheral circulatory regulation in patients with CAD corresponds in principle to that seen in healthy subjects of the same age.

Training changes the different local flows during exercise in such a way that, within the framework of an unchanged or reduced Q, its regional distribution at a given submaximal work load deviates less from that seen at rest: the perfusion of nonworking tissues is relatively greater and the flow to active muscles less elevated. However, this is only valid for exercise performed with trained muscles.

Although the precise mechanism mediating exercise hyperemia is unknown, it seems acceptable that enhanced content of oxidative enzymes enables a reduction in MBF at a given submaximal VO2 . After training, due to the increased capacity for oxidative phosphorylation, ATP and CP in active muscles stabilize at a higher steady state level. At the same time glycolysis occurs at a slower rate, pH is relatively increased, and the concentration of multiple intermediate metabolic products may be lower. In consequence, the intra- and intercellular biochemical milieu-concentrations of electrolytes and osmolality included-is less disturbed as compared to the conditions at rest. Whatever substance or combinations of chemical alterations cause the vasodilation, their extent of change is probably reduced at a given respiratory rate in trained muscle tissue, and the vasodilation is thus diminished.

Training improves exercise tolerance in most patients with angina pectoris. The main part of this effect can be related to the training-induced reduction in HR and SBP that decreases myocardial O2 requirements at a given total body O2 uptake. However, at the same time, higher values for the product of HR and SBP are tolerated before pain is provoked after training, suggesting that training has additional economizing effects on myocardial function or directly improves myocardial O2 supply. As judged from the results obtained in exercise tests, training and nitroglycerin seem almost equally potent in alleviating or preventing angina pectoris on exertion. Beta receptor blockade may be somewhat less efficient, whereas aorto-coronary bypass surgery, when practicable, may be the most efficient treatment of exertional angina available today.

Physical training is efficient in improving exercise capacity in about two thirds of all patients with angina pectoris. Patients with angina pectoris provoked only by exercise will often respond favorably to training, even if their exercise capacity is low.  In contrast, patients who suffer from angina at rest, especially nocturnal attacks, may be less likely to increase their exercise tolerance by training. Accordingly, Hellerstein reports that in patients with more severe coronary arteriosclerosis as assessed from coronary arteriograms and left ventricular function, physical fitness fails to improve from training.

Unfortunately, it appears that the patients who cannot be expected to respond favorably to training are also less likely to improve from other modes of treatment. According to Balcon, only younger patients with normal left ventricular function are prone to achieve substantial improvement in physical working capacity by vein graft surgery. Furthermore, the mortality from the operation is higher in patients with abnormal ventricular function. Thus, the appearance of an apparently efficient surgical intervention has not simplified the selection of treatment.

Characteristics of the Ventilatory Exercise Stimulus

F.M. Bennett and W.E. Fordyce
Respiration Physiology 1985; 59, 55-63

Simple mathematical models were used to quantitatively examine a number of hypotheses concerning the nature of the exercise stimulus. The modelling demonstrated the following for an exercise intensity of 5 times the resting metabolic rate.

(1) During the steady state, a deviation in the coupling between VE and metabolic rate by + 25 % of the value necessary for isocapnia, results in a deviation of Paco2 of + 2 torr from isocapnia.

(2) In the transient phase, a mismatch between VE and Q (and thus CO2 flow) of 50% results in a change of Paco2 of only 1 torr.

(3)When resting Paco2 is changed by 10 torr and it is assumed that the coupling between VE and Paco2 does not change, Paco2 deviates from isocapnia by less than 2 torr.

It is concluded that –

(1) to experimentally test hypotheses of the exercise stimulus requires resolution of small changes in Paco2;

(2)  good regulation of Paco2 does not necessarily imply precise coupling between VE and Vco2;

(3) the ventilatory exercise stimulus need not be a precise function of metabolic rate;

(4) in the steady state, the normal CO2 controller will be very effective in minimizing changes in Paco2 due to a mismatch between ventilation and metabolic rate.

Cardiorespiratory and Metabolic Responses to Positive, Negative and
Minimum-Load Dynamic Leg Exercise

Carl Magnus Hesser, Dag Linnarsson And Hilding Bjurstedt
Respiration Physiology 1977; 30, 5 I-67

Cardiorespiratory and metabolic responses to steady-state dynamic leg exercise were studied in seven male subjects who performed positive and negative work on a modified Krogh cycle ergometer at loads of 0. 16,33,49.98, and 147 W with a pedaling rate of60 rpm.
In positive work, O2 uptake increased with the ergometric load in a parabolic fashion. Net O2 uptake averaged 220 ml*min– 1 at 0 W (loadless pedaling), and was 75 ml* min– 1 lower at the point of physiological minimum load which occurred in negative work at approximately 9 W. The O2 cost of loadless pedaling is for one-third attributed to the work of overcoming elastic and viscous resistance, the remaining part being due mainly to the work of antagonistic muscle contraction in the moving legs. Although at a given Vo2 work rate was much higher in negative than in positive work, corresponding values for VE were similar, suggesting that the mechanical tension in working muscles is of little or no importance in the control of ventilation in steady-state exercise.
Heart rate increased linearly with Vo2 in both positive and negative work, with a steeper slope in negative work. Evidence is presented that none of the current definitions of muscular efficiency yields the true efficiency of muscular contraction in cycle ergometry, net efficiency calculation resulting in too low estimates, and work and delta efficiency calculations in overestimated values in the low-intensity work range, and in underestimated values in the high-intensity range.

The effect of exercise on left ventricular ejection time in patients with hypertension or angina pectoris

James R. Bowlby
Amer Heart J 1979; 97(3): 348-350

Using the method and regression equation of Lewis and associates, the present study confirms their findings in normal men up to the age of 65 years. Despite the significantly higher myocardial oxygen consumption, as measured by the double product, the hypertensive patients responded in a similar fashion. The patients with angina pectoris, however, showed a significantly prolonged post-exercise ejection time.

Cardiac Effects of Prolonged and Intense Exercise Training in Patients With Coronary Artery Disease

Ali A. Ehsani, Wade H. Martin Iii, Gregory W. Heath, Edward F. Coyle
Amer J Cardiol 1982; 50: 246-254

The effects of intense and prolonged exercise training on the heart were studied with echocardiography in eight men with coronary artery disease with a mean age (standard error of the mean) of 52 + 3 years. Training consisted of endurance exercise 3 times/week at 50 to 60 percent of the measured maximal oxygen uptake for 3 months followed by exercise 4 to 5 days/week at 70 to 60 percent of maximal oxygen uptake for 9 months. Maximal oxygen uptake capacity increased by 42 percent (26 + 1 versus 37 + 2 ml/kg per min; p <0.001). Heart rate at rest and submaximal heart rate and systolic blood pressure at a given work rate were significantly lower after training. Systolic blood pressure at the time of maximal exercise increased (145 + 9 before versus 166 + 6 mm Hg after training; probability [p] <0.01). Left ventricular end-diastolic diameter was increased after 12 months of training (from 47 + 1 to 51 + 1 mm; p <0.01. Left ventricular fractional shortening and mean velocity of circumferential shortening decreased progressively in response to graded iisometric handgrip exercise before training but not after training. At comparable levels of blood pressure during static exercise, mean velocity of circumferential shortening was significantly higher after training (0.76 + 0.04 versus 0.96 + 0.07 diameter/set, p <0.01). No improvement in echocardio-graphic or exercise variables was observed over a 12 month period in another group of five patients who did not exercise. Thus the data suggest that prolonged and vigorous exercise training in selected patients with coronary artery disease can elicit cardiac adaptations.

Physical activity and resting pulse rate in older adults: Findings from a randomized controlled trial

Bríain O’Hartaigh, Marco Pahor, Thomas W. Buford, John A. Dodson, et al.
Am Heart J 2014;168:597-604

Background Elevated resting pulse rate (RPR) is a well-recognized risk factor for adverse outcomes. Epidemiological evidence supports the beneficial effects of regular exercise for lowering RPR, but studies are mainly confined to persons younger than 65 years. We set out to evaluate the utility of a physical activity (PA) intervention for slowing RPR among older adults.
Methods A total of 424 seniors (ages 70-89 years) were randomized to a moderate intensity PA intervention or an education-based “successful aging” health program. Resting pulse rate was assessed at baseline, 6 months, and 12 months. Longitudinal differences in RPR were evaluated between treatment groups using generalized estimating equation models, reporting unstandardized β coefficients with robust SEs.
Results Increased frequency and duration of aerobic training were observed for the PA group at 6 and 12 months as compared with the successful aging group (P = 0.001). In both groups, RPR remained unchanged over the course of the 12-month study period (P = .67). No significant improvement was observed (β [SE] = 0.58 [0.88]; P = .51) for RPR when treatment groups were compared using the generalized estimating equation method. Comparable results were found after omitting participants with a pacemaker, cardiac arrhythmia, or who were receiving β-blockers.
Conclusions Twelve months of moderate intensity aerobic training did not improve RPR among older adults. Additional studies are needed to determine whether PA of longer duration and/or greater intensity can slow RPR in older persons.

Autonomic regulation and maze-learning performance in older and younger dults

Karen J. Mathewson, J Dywan, PJ Snyder, WJ Tays, SJ Segalowitz
Biological Psychology 88 (2011) 20– 27

There is growing evidence that centrally modulated autonomic regulation can influence performance on complex cognitive tasks but the specificity of these influences and the effects of age-related decline in these systems have not been determined. We recorded pre-task levels of respiratory sinus arrhythmia (RSA; an index of phasic vagal cardiac control) and rate pressure produce (RPP; an index of cardiac workload) to determine their relationship to performance on a cumulative maze learning task. Maze performance has been shown to reflect executive error monitoring capacity and non-executive visuomotor processing speed. Error monitoring was predicted by RSA in both older and younger adults but by RPP only in the older group. Non-executive processes were unrelated to either measure. These data suggest that vagal regulation is more closely associated with executive than nonexecutive aspects of maze performance and that, in later life, pre-task levels of cardiac workload also influence executive control.

Sympathovagal Imbalance Contributes to Prehypertension Status and Cardiovascular Risks Attributed by Insulin Resistance, Inflammation, Dyslipidemia and Oxidative Stress in First Degree Relatives of Type 2 Diabetics

Gopal Krushna Pal, C Adithan, P Hariharan Ananthanarayanan, Pravati Pal, et al.
PLoS OME 2013; 8(11), e78072

Background: Though cardiovascular (CV) risks are reported in first-degree relatives (FDR) of type 2 diabetics, the pathophysiological mechanisms contributing to these risks are not known. We investigated the association of sympathovagal imbalance (SVI) with CV risks in these subjects.
Subjects and Methods: Body mass index (BMI), basal heart rate (BHR), blood pressure (BP), rate-pressure product (RPP), spectral indices of heart rate variability (HRV), autonomic function tests, insulin resistance (HOMA-IR), lipid profile, inflammatory markers, oxidative stress (OS) marker, rennin, thyroid profile and serum electrolytes were measured and analyzed in subjects of study group (FDR of type 2 diabetics, n = 72) and control group (subjects with no family history of diabetes, n = 104).
Results: BMI, BP, BHR, HOMA-IR, lipid profile, inflammatory and OS markers, renin, LF-HF (ratio of low-frequency to high frequency power of HRV, a sensitive marker of SVI) were significantly increased (p,0.0001) in study group compared to the control group. SVI in study group was due to concomitant sympathetic activation and vagal inhibition. There was significant correlation and independent contribution of markers of insulin resistance, dyslipidemia, inflammation and OS to LF-HF ratio. Multiple-regression analysis demonstrated an independent contribution of LF-HF ratio to prehypertension status (standardized beta 0.415, p,0.001) and bivariate logistic-regression showed significant prediction (OR 2.40, CI 1.128–5.326, p = 0.002) of LF-HF ratio of HRV to increased RPP, the marker of CV risk, in study group.
Conclusion: SVI in FDR of type 2 diabetics occurs due to sympathetic activation and vagal withdrawal. The SVI contributes to prehypertension status and CV risks caused by insulin resistance, dyslipidemia, inflammation and oxidative stress in FDR of type 2 diabetics.

Exercise prescription for patients with type 2 diabetes and pre-diabetes: A position statement from Exercise and Sport Science Australia

Matthew D. Hordern, DW Dunstan, JB Prins, MK Baker, et al.
Journal of Science and Medicine in Sport 15 (2012) 25–31

Type 2 diabetes mellitus (T2DM) and pre-diabetic conditions such as impaired fasting glucose (IFG) and/or impaired glucose tolerance (IGT) are rapidly increasing in prevalence. There is compelling evidence that T2DM is more likely to develop in individuals who are insufficiently active. Exercise training, often in combination with other lifestyle strategies, has beneficial effects on preventing the onset of T2DM and improving glycaemic control in those with pre-diabetes. In addition, exercise training improves cardiovascular risk profile, body composition and cardiorespiratory fitness, all strongly related to better health outcomes. Based on the evidence, it is recommended that patients with T2DM or pre-diabetes accumulate a minimum of 210 min per week of moderate-intensity exercise or 125 min per week of vigorous intensity exercise with no more than two consecutive days without training. Vigorous intensity exercise is more time efficient and may also result in greater benefits in appropriate individuals with consideration of complications and contraindications. It is further recommended that two or more resistance training sessions per week (2–4 sets of 8–10 repetitions) should be included in the total 210 or 125 min of moderate or vigorous exercise, respectively. It is also recommended that, due to the high prevalence and incidence of comorbid conditions in patients with T2DM, exercise training programs should be written and delivered by individuals with appropriate qualifications and experience to recognise and accommodate comorbidities and complications.

Estimation of the Ejection Fraction in Patients with Myocardial Infarction Obtained from the Combined Index of Systolic and Diastolic Left Ventricular Function: A New Method

Jorge A. Lax, Alejandra M. Bermann, Tomás F. Cianciulli, Luis A. Morita, et al.
J Am Soc Echocardiogr 2000;13:116-23.

The index of myocardial performance combining systolic and diastolic time intervals (Index) is a useful method, already explained in past studies, that offers new values that have not been widely known among clinical cardiologists. The aim of this study is to obtain from this Index a measurement of the ejection fraction (EF), which is a very well-known value.
The study involved 97 patients with myocardial infarction, 55 of whom were studied retrospectively (group A, aged 46-62 years, 50 men) to obtain and test the formula EF = 60 – (34 × Index). The second group (group B, aged 47-63 years, 40 men) included 42 patients who were evaluated prospectively. The EF obtained was compared with that reached through the use of radionuclide angiography (EF-RNA).
The Index was obtained through the use of the formula (a – b)/b, where a is the interval between cessation and onset of the mitral inflow, and b is the ejection time. In group A the EF obtained by the Index (EF-Index) was 37.5% ± .8%, and the EF-RNA was 37.7% ± 11% (r = 0.76). In group B the EF-Index was 41.6% ± 7%, and the EF-RNA was 41.2% ± 10% (r = 0.75).
Conclusion: Through the new formula described here it is possible to obtain a reliable measurement of the EF in patients with myocardial infarction, a well known and extremely useful value, especially for those patients with poor acoustic windows.

HCN channels: new roles in sinoatrial node function

Christian Wahl-Schott, Stefanie Fenske and Martin Biel
Current Opinion in Pharmacology 2014, 15:83–90

Hyperpolarization-activated cyclic nucleotide gated (HCN) channels pass a cationic current (Ih/If) that crucially contributes to the slow diastolic depolarization (SDD) of sinoatrial pacemaker cells and, hence, is a key determinant of cardiac automaticity and the generation of the heart beat. There is growing evidence, that HCN channel functions in the sinoatrial node (SAN) are not restricted to impulse formation but are also required for impulse propagation. In addition, HCN channels are involved in coordination and maintenance of sinoatrial network activity and, hence, are crucial for stabilizing cardiac rhythmicity. In the present review we will outline these new concepts.

In this review we will focus on HCN channel functions in the sinoatrial node beyond the established concepts described above. We will outline recent advances involving the characterization of the HCN1-deficient mouse line (HCN1-/- mouse) which have provided evidence that HCN channels are required for impulse propagation and the precision of the heart beat [19**]. Furthermore, we show how these properties can be generalized across the other HCN channel subtypes in the sinoatrial node.

19** Fenske S, Krause SC, Hassan SI, Becirovic E, Auer F, Bernard R, Kupatt C, Lange P, Ziegler T, Wotjak CT et al.: Sick sinus syndrome in HCN1-deficient Mice. Circulation 2013. Epub 2013 Nov 11.
First demonstration of a functional relevance of HCN1 channels in the murine sinoatrial node. The authors demonstrate that mice lacking the pacemaker channel HCN1 display congenital sinoatrial node dysfunction characterized by bradycardia, sinus dysrhythmia, prolonged sinoatrial node recovery time, increased sinoatrial conduction time and recurrent sinus pauses. As a consequence of sinoatrial node dysfunction HCN1-deficient mice display a severely reduced cardiac output.

Recent studies indicate that the role of cardiac HCN channels extends well beyond generation of pacemaker potentials. In addition to being merely ‘pacemaker channels’, HCN channels are important for sinoatrial impulse propagation, cardiac excitability and for the precision of the heartbeat. Furthermore, cardiac HCN channels are involved in the repolarization process of heart ventricles [56**,57]. It will be important to consider the full spectrum of these diverse cardiac functions when exploring agents acting on HCN channels for a specific clinical purpose such as reduction of heart rate.

56.** Fenske S, Mader R, Scharr A, Paparizos C, Cao-Ehlker X, et al.: HCN3 contributes to the ventricular action potential waveform in the murine heart. Circ Res 2011, 109:1015-1023.
First study demonstrating a functional role of HCN3 channels in the heart. Using HCN3-deficient mouse line the authors show that HCN3 together with other members of the HCN channel family confers a depolarizing background current that regulates ventricular resting potential and counteracts the action of hyperpolarizing potassium currents in late repolarization.
57. Fenske S, Krause S, Biel M, Wahl-Schott C: The role of HCN channels in ventricular repolarization. Trends Cardiovasc Med 2011, 21:216-220.

Roles of HCN1 channels for sinoatrial impulse conduction (source-sink relation) The primary impulse initiating the heart beat is generated in the leading pacemaker cell(s) of the sinoatrial node. Once the leading pacemaker cell(s) reaches the threshold for L-type Ca2+ channels an action potential is generated. Since pacemaker cells are interconnected via gap junctions, the impulse is conducted through the sinoatrial network and to the atrium. During impulse propagation the source cell (the cell which first reached AP threshold and is firing the action potential) charges the neighboring cell (sink), in which the membrane potential is below threshold (Figure 1) [24*]. Impulse propagation depends on the source-sink relation [24*, 25–29]. HCN1 deletion increases the sinoatrial conduction time suggesting the existence of a source sink mismatch in the HCN1-deficient mouse [19**].

Role of HCN1 channels for impulse formation and impulse conduction in the sinoatrial node. Schematic pacemaker potential in sinoatrial node cells of wild type (a) and HCN1-/- mice.
(b) HCN channels contribute to the slow diastolic depolarization. In the absence of HCN1 the slope of SDD isdecreased and the time to threshold for an action potential increased. HCN channels decrease the maximal diastolic potential (MDP). In the absence of HCN1 the MDP is increased. This results in an increased distance and time to threshold for an action potential and a decrease in impulse propagation.  [SDD: slow diastolic depolarization; MDD: maximal diastolic depolarization; Vthr: threshold potential for the generation of an action potential.]
(c) Direction of intracellular and extracellular current flow during propagation of an action potential from depolarized (source) to resting cells (sink).
(d)Source sink relationship in propagation. Charge from excited cells (source) flows into unexcited cell (sink) and provides the charge to depolarize them to activation threshold. Arrows and dotted lines indicate changes observed in HCN1-/- mice of parameter indicated leading to source sink mismatch and prolonged sinoatrial conduction. Modified from [24*].

24.* Spector P: Principles of cardiac electric propagation and their implications for re-entrant arrhythmias. Circ Arrhythm Electrophysiol 2013, 6:655-661.
The authors provide an excellent review of the principles of impulse propagation in relation to arrhythmia.

HCN1 channels increase the temporal and spatial precision of impulse formation in sinoatrial node

HCN1 channels increase the temporal and spatial precision of impulse formation in sinoatrial node

HCN1 channels increase the temporal and spatial precision of impulse formation in sinoatrial node.
(a) Schematic of the sinoatrial node. Atrial cells invaginate into the central sinoatrial node. Putative localization of HCN1 channels at contact interface between strands of atrial myocytes which extend into the central SAN and sinoatrial node pacemaker cells. Green: autonomous innervation. HCN1 channels dampen network noise generated by neighboring pacemaker cells in the sinoatrial network, by invading hyperpolarization of atrial cells and by autonomous regulation. SAN: sinoatrial node, RA: right atrium, CT: crista terminalis.
(b) Model of sinoatrial node function (for detail see text). Note that individual cells display different phases and slightly different periods.

Pharmacological inhibition of cardiac HCN channels

HCN channels have emerged as interesting targets for the development of drugs that lower the heart rate. Ivabradine is the first and currently the only clinically approved compound that specifically targets HCN channels. The therapeutic indication of ivabradine is the symptomatic treatment of chronic stable angina pectoris in patients with coronary artery disease with a normal sinus rhythm (for details see [48], the international trial on the treatment of angina with ivabradine vs. atenolol (INITIATIVE) trial (n = 939) [49] and the antianginal efficacy and safety of the association of the Ih/If current inhibitor ivabradine with a beta-blocker (ASSOCIATE) study (n = 889) [50]).

The Role of HCN Channels in Ventricular Repolarization

Stefanie Fenske, Stefanie Krause, Martin Biel, and Christian Wahl-Schott
Trends Cardiovasc Med 2011; 21:216-220
PII S1050-1738(12)00143-0

Hyperpolarization-activated cyclic nucleotide gated (HCN) channels pass a cationic current (Ih/If) that crucially contributes to the slow diastolic depolarization (SDD) of sinoatrial pacemaker cells and, hence, is a key determinant of cardiac automaticity and the generation of the heartbeat. However, there is growing evidence that HCN channels are not restricted to the spontaneously active cells of the sinoatrial node and the conduction system but are also present in ventricular cardiomyocytes that produce an action potential lacking SDD. This observation raises the question of the principal function(s) of HCN channels in working myocardium. Our recent analysis of an HCN3-deficient (HCN3–/–) mouse line has shed new light on this central question.

We propose that HCN channels contribute to the ventricular action potential waveform, specifically during late repolarization. In this review, we outline this new concept.

In the late 1970s, the hyperpolarization activated current (Ih/If) was discovered and characterized in sinoatrial node cells (Brown and Difrancesco 1980). This current displays several unique biophysical properties: activation upon hyperpolarization and deactivation by depolarization, with a small but substantial degree of activation at resting potentials typically observed in sinoatrial node pacemaker cells (–60 to –50 mV) and ventricular cells (–85 to –75 mV); shift of the activation curve to more positive potentials by cAMP;  block by millimolar concentrations of external Cs+; and permeability for Na+ and K+ions with a reversal potential near –35 mV.

  • HCN3 Is a Component of Ventricular Ih
  • HCN Channels Prolong Action Potentials During Late Repolarization
  • HCN3 Forms Background Channels That Do Not Deactivate During the Action Potential
  • HCN channels need to be open at the resting membrane potential;
    (2) HCN channels remain open during the entire time course of the action potential—de novo opening of HCN channels during the AP does not occur because these channels are activated by hyperpolarization and depolarization decreases open probability; and
    (3) a driving force is needed to sustain an HCN-mediated current during the AP. A detailed analysis of the functional properties of heterologously expressed HCN3 channels revealed that these three prerequisites are met.

Neurophysiology of HCN channels: From cellular functions to multiple regulations

Chao He, Fang Chen, Bo Li, Zhian Hu
Progress in Neurobiology 112 (2014) 1–23

Hyperpolarization-activated cyclic nucleotide-gated (HCN) cation channels are encoded by HCN1-4 gene family and have four subtypes. These channels are activated upon hyperpolarization of membrane potential and conduct an inward, excitatory current Ih in the nervous system. Ih acts as pacemaker current to initiate rhythmic firing, dampen dendritic excitability and regulate presynaptic neurotransmitter release. This review summarizes recent insights into the cellular functions of Ih and associated behavior such as learning and memory, sleep and arousal. HCN channels are excellent targets of various cellular signals to finely regulate neuronal responses to external stimuli. Numerous mechanisms, including transcriptional control, trafficking, as well as channel assembly and modification, underlie HCN channel regulation. In the next section, we discuss how the intracellular signals, especially recent findings concerning protein kinases and interacting proteins such as cGKII, Ca2+/CaMKII and TRIP8b, regulate function and expression of HCN channels, and subsequently provide an overview of the effects of neurotransmitters on HCN channels and their corresponding intracellular mechanisms. We also discuss the dysregulation of HCN channels in pathological conditions. Finally, insight into future directions in this exciting area of ion channel research is provided.

The hyperpolarization-activated current, Ih, was first observed in sino-atrial node tissue in 1976 and later was identified in rod photoreceptors and hippocampal pyramidal neurons (Noma and Irisawa, 1976). Due to its unique properties, particularly the activation upon hyperpolarization of the membrane potential, Ih has been also termed If (f for funny) or Iq (q for queer). The hyperpolarization-activated cyclic nucleotide-gated (HCN) cation ion channels underlying Ih were discovered in the late 1990s and subsequently, the genes encoding these channels were identified, which enable the expression of HCN channels in heterologous systems.

HCN channels belong to the superfamily of voltage-gated pore loop channels with four pore-forming subunits (HCN1-4) encoded by the HCN1-4 gene family in mammals (Robinson and Siegelbaum, 2003). Each subunit has six transmembrane helices (S1–S6), with the positively charged voltage sensor (S4) and the pore region carrying the GYG motif between S5 and S6, which forms the ion selectivity filter (Macri et al., 2012). Following S6 is the 80-residue C-linker comprising six a-helices (A0–F0) and the cyclic nucleotide binding domain (CNBD). The CNBD consists of three a-helices (A–C) and a b-roll between the A- and B-helices (Fig. 1) (Biel et al., 2009; Wahl-Schott and Biel, 2009; Wicks et al., 2011). Together, the C-linker and CBND can be referred to as the ‘‘cAMP-sensing domain’’ (CSD) because they are of functional importance for the cAMP-induced positive shift of the voltage-dependent activation of HCN channels. The crystal structure of CSD has been elucidated at an atomic resolution, but a high-resolution structure of the transmembrane core remains unsolved.

Structure of HCN channels

Structure of HCN channels

Structure of HCN channels. Left: one subunit is composed of six transmembrane segments (S1–S6), with the positive charged voltage sensor (S4) and the pore region carrying the GYG motif between S5 and S6. The C-terminal of HCN channels is composed of the C-linker and the cyclic nucleotide-binding domain (CNBD) which mediates their responses to cAMP. The C-linker consists of six a-helices: A0 to F0 . The CNBD follows the C-linker domain and consists of a-helices A–C with a b-roll between the A- and B-helices. Right: the four subunits assemble in homomeric or heteromeric tetramer configurations in vivo.

Regulatory mechanisms of Ih function by the small molecules, protein kinases and interacting proteins.

Regulatory mechanisms of Ih function by the small molecules, protein kinases and interacting proteins.

Regulatory mechanisms of Ih function by the small molecules, protein kinases and interacting proteins. Black arrows indicate known sites of HCN channels interaction with small molecules, protein kinases and interacting proteins. Broken lines indicate the speculated interaction sites. Filamin A interacts with HCN1 via a region of 22 amino acids located downstream from the CNBD. Tamalin and Mint2 bind to the CNBD-downstream sequence of HCN2. The binding of the PDZ domain of S-SCAM occurs at the cyclic nucleotide-binding domain (CNBD) and the CNBD downstream sequence of the carboxy-terminal tail. CNBD, cyclic nucleotide binding domain; SNL, C-terminal tripeptide of HCN1, HCN2 and HCN4.

modulation of HCN channels by neurotransmitters and associated intracellular signal pathways

modulation of HCN channels by neurotransmitters and associated intracellular signal pathways

The modulation of HCN channels by neurotransmitters and associated intracellular signal pathways. Glutamate (Glu) activates N-methyl-D-aspartate receptors (NMDARs) and a-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) which results in the Ca2+ influx and subsequently activates calcium calmodulin kinase (CaMKII). CaMKII increases channels surface expression through the interacting protein TRIP8b (1a-4) or reduces the HCN1 gene transcription via Neuronal Restrictive Silencing Factor (NRSF) in pathological conditions. Glu, norepinephrine (NE, in rats), 5-hydroxytryptamine (5-HT) and triphosphate (ATP) bind to specific G-coupled receptors and modulate the activity of HCN channels via the PLC-PKC or p38-MAPK signaling pathways. Activation of PKC suppresses the activation of HCN channels, whereas p38-MAPK causes a positive shift of HCN channels voltage-dependent activation. Adenosine, NE (in monkey), 5-HT, dopamine (DA) and Ach (acetylcholine) bind to Gs- or Gi coupled receptors. Gs or Gi oppositely control the activity of adenylate cyclase (AC), which catalyzes the ATP to cAMP. cAMP could shift the HCN channels voltage-dependent activation to positive direction and accelerate the kinetics of channels activation. Nitric oxide (NO) interacts with soluble guanylyl cyclase (GC) and thus increases the intracellular concentration of cGMP, which induces a positive shift of HCN channels voltage-dependent activation. Sharp and blunted arrows represent the positive and negative regulation, respectively. Broken lines indicate the speculated signal pathway.

Ultimately, the study of the HCN channels will provide an overall picture underlying the real-time in vivo regulation of the function and expression of HCN channels to fulfill complex functions in different contexts.

Oxygen uptake kinetics during high-intensity arm and leg exercise

Katrien Koppo, Jacques Bouckaert, Andrew M. Jones
Respiratory Physiology & Neurobiology 133 (2002) 241-250
PII: S1569 – 9048 ( 02 ) 00184 – 2

The purpose of the present study was to examine the oxygen uptake kinetics during heavy arm exercise using appropriate modelling techniques, and to compare the responses to those observed during heavy leg exercise at the same relative intensity. We hypothesized that any differences in the response might be related to differences in muscle fiber composition that are known to exist between the upper and lower body musculature. To test this, ten subjects completed several bouts of constant-load cycling and arm cranking exercise at 90% of the mode specific ˙VO2 peak. There was no difference in plasma [lactate] at the end of arm and leg exercise. The time constant of the fast component response was significantly longer in arm exercise compared to leg exercise (mean ­+ S.D., 489 +12 vs. 219 + 5 sec; P < 0.01), while the fast component gain was significantly greater in arm exercise (12.19 + 1.0 vs. 9.29 + 0.5 ml min-1 W-1; P < 0.01). The ˙VO2 slow component emerged later in arm exercise (1269 + 27 vs. 959 + 20 sec; P < 0.01) and, in relative terms, increased more per unit time (5.5 vs. 4.4% min-1; P < 0.01). These differences between arm crank and leg cycle exercise are consistent with a greater and/or earlier recruitment of type II muscle fibers during arm crank exercise.

Probability and magnitude of response to cardiac resynchronization therapy according to QRS duration and gender in nonischemic cardiomyopathy and LBBB

Niraj Varma, Mahesh Manne, Dat Nguyen, …, Patrick Tchou
Heart Rhythm 2014; 11: 1139–1147

BACKGROUND QRS morphology and QRS duration (QRSd) determine cardiac resynchronization therapy (CRT) candidate selection but criteria require refinement.
OBJECTIVE To assess CRT effect according to QRSd, treated by dichotomization vs a continuous function, and modulation by gender.
METHODS Patients selected were those with New York Heart Association classIII/IV heart failure and with left bundle branch block and nonischemic cardiomyopathy (totest “pure” CRT effect) with pre-and post- implant echocardiographic evaluations. Positive response was defined as increased left ventricular ejection fraction (LVEF) post-CRT.
RESULTS In 212 patients (LVEF 19% +  7.1%; QRSd 160 + 23 ms; 105 (49.5%) women), CRT increased LVEF to 30% + 15% (P < .001) during a median follow-up of 2 years. Positive response occurred in 150 of 212 (71%) patients. Genders did not differ for QRSd, pharmacotherapy, and comorbidities, but response to CRT among women was greater: incidence 84% (88of105) in women vs 58% (62of107) in men (P < .001); increase in LVEF 15%+ 14% vs 7.2% + 13%, respectively (P < .001). Overall, the response rate was 58% when QRSd <150 ms and 76% when QRSd > 150 ms (P <.009). This probability differed between genders: 86% in women vs 36% in men (P < .001) when QRSd <150 ms and 83% vs 69%, respectively, when QRSd >150 ms (P < .05). Thus, female response rates remained high whether QRSd was < 150 ms >150 ms (86% vs 83%; P = .77) but differed in men (36% vs 69%; P < .001). With QRSd as a continuum, the CRT-response relationship was nonlinear and significantly different between genders. Female superiority at shorter QRSd inverted with prolongation > 180 ms.
CONCLUSION The QRSd-CRT response relationship in patients with heart failure and with left bundle branch block and non-ischemic cardiomyopathy is better  described by a sex-specific continuous function and not by dichotomization by 150ms, which excludes a large proportion of women with potentially favorable outcome.

Comparison of eterminants Myocardial Oxygen Consumption During Arm and Leg Exercise in Normal Persons

Gary J. Balady, et al.  Am J Cardiol 1985; 57: 1385-87.

The effects of arm exercise on myocardiai oxygen consumption are not well understood; they may differ from the effects of leg exercise. Previous studies have shown that the ischemic threshold is higher in patients performing arm exercise and leg exercise at the same heart rate-blood pressure product. The contribution of other determinants of myocardiai oxygen consumption-left ventricular (LV) peak meridional systolic wail stress and contractility-to these observed differences were studied.
Thirty healthy subjects exercised to the same peak rate-pressure product during dynamic upper- and lower-extremity exercise. Peak workload was lower
during arm exercise (100 + 16 W) leg exercise (170 + 21 W, p < 0.001). LV wail stress did not differ during either form of exercise (197 + 44 vs 204 + 33 dynes/cm2 X 103, arm vs leg, respectively). This was also true of contractility as assessed by the velocity of circumferential fiber shortening (2.6 + 0.6 vs 2.5 + 0.4 circ/s, arm vs leg, respectively) and the preejection period/LV ejection time ratio (0.33 + 0.11 vs 0.31 + 0.07, arm vs leg, respectively). Normal subjects exercising to a similar rate-pressure product showed the same levels at LV wail stress and contractility for arm and leg exercise despite the lower rkioad performed with arm exercise.

Anti-hypertensive effect of radiofrequency renal denervation in spontaneously hypertensive rats

Takeshi Machino, N Murakoshi, A Sato, …, T Hoshi, T Kimura, K Aonuma
Life Sciences 110 (2014) 86–92

Aims: We aimed to investigate the anti-hypertensive effect of radiofrequency (RF) renal denervation (RDN) in an animal model of hypertension.           Materials and methods: RF energy was delivered to bilateral renal arteries through a 2 Fr catheter with opening abdomen in 8 spontaneously hypertensive rats (SHRs) and 8 Wistar–Kyoto rats (WKYs). Sham operation was performed in other 8 SHRs and 8 WKYs. Blood pressure (BP), heart rate (HR), and urinary norepinephrine excretion were followed up for 3 months. Plasma and renal tissue concentrations of norepinephrine and plasma renin activity were measured 3 months after the procedure. The RDN was confirmed by a decrease in renal tissue norepinephrine.
Key findings: RF-RDN restrained a spontaneous rise in systolic BP (46 ± 12% increase from 158 ± 8 to 230 ± 14 mmHg vs. 21 ± 18% increase from 165 ± 9 to 197 ± 20 mmHg, p= 0.01) and diastolic BP (55 ± 27% increase from 117 ± 9 to 179 ± 23 mmHg vs. 28 ± 13% increase from 120 ± 7 to 154 ± 13 mm Hg, p= 0.04) in SHRs; however, WKYs were not affected. Although there were no changes in HR and systemic norepinephrine, the renal tissue norepinephrine was decreased by RF-RDN in both SHR (302±41 vs. 159±44 ng/g kidney, p b 0.01) and WKY (203 ± 33 vs. 145 ± 26 ng/g kidney, p= 0.01). Plasma renin activity was reduced by the RF-RDN only in SHR (35.3 ± 9.5 vs. 21.4 ±  8.6 ng/mL/h, p < 0.01).
Significance: RF-RDN demonstrated an anti-hypertensive effect with a reduction of renal tissue norepinephrine and plasma renin activity in SHR.

Effectiveness of Renal Denervation Therapy for Resistant Hypertension: A Systematic Review and Meta-Analysis

Mark I. Davis, KB Filion, D Zhang, MJ Eisenberg, …, EL Schiffrin, D Joyal
J Am Coll  Cardiol 2013; 62(3): 231-241.

Objectives This study sought to determine the current effectiveness and safety of sympathetic renal denervation (RDN) for resistant hypertension.               Background RDN is a novel approach that has been evaluated in multiple small studies.
Methods We performed a systematic review and meta-analysis of published studies evaluating the effect of RDN in patients with resistant hypertension. Studies were stratified according to controlled versus uncontrolled design and analyzed using random-effects meta-analysis models.                                    Results We identified 2 randomized controlled trials, 1 observational study with a control group, and 9 observational studies without a control group. In controlled studies, there was a reduction in mean systolic and diastolic blood pressure (BP) at 6 months of –28.9 mm Hg (95% confidence interval [CI]: –37.2 to –20.6 mm Hg) and –11.0 mm Hg (95% CI: –16.4 to –5.7 mm Hg), respectively, compared with medically treated patients (for both, p < 0.0001). In uncontrolled studies, there was a reduction in mean systolic and diastolic BP at 6 months of –25.0 mm Hg (95% CI: –29.9 to –20.1 mm Hg) and –10.0 mm Hg (95% CI: –12.5 to –7.5 mm Hg), respectively, compared with pre-RDN values (for both, p < 0.00001). There was no difference in the effect of RDN according to the 5 catheters employed. Reported procedural complications included 1 renal artery dissection and 4 femoral pseudoaneurysms.
Conclusions RDN resulted in a substantial reduction in mean BP at 6 months in patients with resistant hypertension. The decrease in BP was similar irrespective of study design and type of catheter employed. Large randomized controlled trials with long-term follow-up are needed to confirm the sustained efficacy and safety of RDN.

Effects of renal denervation on the development of post-myocardial infarction heart failure and cardiac autonomic nervous system in rats

Jialu Hu, Yan Yan, Qina Zhou, Meng Ji, Conway Niu, Yuemei Hou, Junbo Ge
Intl J Cardiol 172 (2014) e414–e416

Prior studies indicated that radiofrequency renal denervation (RD) had beneficial effects on post-myocardial infarction (MI) heart failure (HF) in rats. In this study we aimed to assess its effects on cardiac autonomic nervous system (CANS) which might be one of the most important mechanisms of RD’s therapeutic effect on post-MI HF and determine the best timing for RD.

One hundred Wistar rats were randomly assigned into five experimental groups: MI group (n = 20), RD group (n = 20), MI-1d + RD group (RD performed one day post-MI, n = 20), MI-4w + RD group (RD performed four weeks post-MI, n = 20), and N group (control group, n = 20).MI was produced through ligation of the anterior descending artery. RD was performed through stripping of the renal nerves. The experimental design and implementation were conducted in accordance with animal welfare guidelines.

Eight weeks post-MI, significant improvements were observed in both MI-1d + RD and MI-4w + RD groups compared to the MI group, that include

(1) improved left ventricular (LV) function and hemodynamics with increased water and sodium excretion;
(2) decreased plasma and renal tissue norepinephrine levels while tissue norepinephrine content increased in myocardium;
(3) increased β1-receptor in myocardium and improved heart rate variability;  (4) decreased plasma renin, angiotensin II, aldosterone, BNP and endothelin levels.

More therapeutic effects were found in the MI-1d + RD group than the MI-4w + RD group.

Firstly, our study showed that RD attenuated the remodeling of CANS and modulated its activities. RD leads to preservation of β1 receptors content along with the β1 mRNA expression in noninfarcted cardiac tissue in this HF model (Fig. 1). This correlated with an improvement in heart function and cardiac remodeling. HRV is a sensitive marker for the CANS. RD led to a slower HR and higher SDNN in both intervention groups.

Secondly, we found that RD blocked both peripheral and central RAAS and sympathetic nervous system (SNS) at the same time. And this may answer the question how RD exerted effect on CANS. In our study RD restores renin, angiotensin II, and aldosterone to near normal levels. This not only explains the increase in sodium and water excretion, but also confirms that RD blocks renal RAAS via blockage of the efferent renal sympathetic nerves which is consistent with our previous study.

Thirdly, early RD, performed one day post-MI, resulted in greater excretion of urinary sodium, lower circulating BNP and ET-1 levels compared to late interventions (four weeks post-MI). This suggests that RD performed in the acute phase of MI may not only reverse cardiac remodeling but also has a preventive effect against the development of HF, as what was observed with β-blockers. RD alleviated cardiac preload and afterload by increasing water and sodium retention, blocking cardiac sympathetic activation and decreasing a variety of vasomotor factors which may lead to alleviated acute and chronic ischemia of the heart.

RD improves hemodynamics, decreases neuro-hormonal activations, modulates cardiac autonomic activities, and attenuates LV remodeling in HF. Early intervention appears to have greater beneficial effects on cardiac functional recovery and reverse remodeling after myocardial injury. Circulating neuro-hormones may be effective indicators to evaluate the therapeutic effect of RD on HF. Our data suggested that RD is a safe, non-pharmaceutical treatment of HF after cardiac injury, with unique benefits in stabilizing cardiac autonomic activity and remodeling post-MI.

The cardiac pacemaker current

Mirko Baruscotti, Andrea Barbuti, Annalisa Bucchi
Journal of Molecular and Cellular Cardiology 48 (2010) 55–64

In mammals cardiac rate is determined by the duration of the diastolic depolarization of sinoatrial node (SAN) cells which is mainly determined by the pacemaker If current. f-channels are encoded by four members of the hyperpolarization-activated cyclic nucleotide-gated gene (HCN1–4) family. HCN4 is the most abundant isoform in the SAN, and its relevance to pacemaking has been further supported by the discovery of four loss-of-function mutations in patients with mild or severe forms of cardiac rate disturbances. Due to its selective contribution to pacemaking, the If current is also the pharmacological target of a selective heart rate-reducing agent (ivabradine) currently used in the clinical practice. Albeit to a minor extent, the If current is also present in other spontaneously active myocytes of the cardiac conduction system (atrioventricular node and Purkinje fibres). In working atrial and ventricular myocytes f-channels are expressed at a very low level and do not play any physiological role; however in certain pathological conditions over-expression of HCN proteins may represent an arrhythmogenic mechanism. In this review some of the most recent findings on f/HCN channels contribution to pacemaking are described.

Cardiac pacemaking originates in the sinoatrial node (SAN) as a consequence of spontaneous firing of rhythmic action potentials generated by specialized myocytes. Although the electrical behavior of a typical SAN cell differs in several aspects from that of a working myocyte, the functional hallmark can be precisely identified in the events that take place during the diastolic interval. During this phase atrial and ventricular myocytes rest in a standby-like condition at a stable voltage (∼−80 mV); a quite different situation characterizes SAN cells, where the cell potential slowly creeps up from the
maximum diastolic potential of about −60 mV to the threshold for the ignition of a new action potential. Since this time interval sets the pace of the heart, this phase is named “pacemaker depolarization”. Given the large spectrum of heart rates observed in mammals the duration of this phase can vary substantially, however the voltage range encompassed is extremely constant and roughly extends from −60 to−40 mV . To sustain this phase several ionic currents and pumps enter in action at variable times and voltages, and this complexity allows for a highly flexible system since the chronotropic fine tuning operated by neuro-hormonal regulators can target different effectors.

In this review we will focus on the If current which is responsiblefor initiating the diastolic depolarization of SAN cells. Due to its fundamental role and its unusual characteristics of being activated in hyperpolarization, this current was named “pacemaker current” or “funny” (If) current. The unique property of a reverse voltage dependence, together with the inward nature of the current at diastolic potentials, makes this current apt to initiate and support the diastolic depolarization. In addition, the direct modulation of the current operated by the second messenger cAMP, represents one of the main pathways by which the autonomic nervous system controls cardiac chronotropism. Two recent clinical findings further confirm the role of f-channels in setting the cardiac rate: one is the evidence of a causative link between the presence of loss-of-function mutations found in these channels and the arrhythmic state of individuals carrying the mutations, and the other is the specific heart rate reduction observed in patients treated with ivabradine, a drug that at therapeutic doses selectively reduces the If current (see specific sections in this review).

Although originally discovered in the heart, the If current is also abundantly present in a large fraction of neuronal elements, where it contributes to rhythmic firing, synaptic integration, and dendritic integration.

Molecular and functional properties of SAN myocytes

Molecular and functional properties of SAN myocytes

Molecular and functional properties of SAN myocytes. (A) Spontaneous action potentials (left) and If current traces (right) recorded from typical rabbit SANmyocytes; currents were elicited by hyperpolarizing voltage steps in the range−45 to −75 mV. (B) Immunofluorescence analysis of rabbit SAN tissue slice labelled with anti-connexin 43 (Cx43, red) and anti-HCN4 (green) antibodies. HCN4 is strongly expressed in the central region of the SAN, while the opposite staining is observed for Cx43; crista terminalis (CT), interatrial septum (IS). (C) HCN4 labelling of single myocytes isolated from CT, SAN and IS (top), and  representative current traces recorded at−125mV frommyocytes isolated from the same regions (bottom). Both If current density and HCN4 labelling are more abundant in the central nodal area. (Panels B and C from [61] with permission).

[61] Brioschi C, Micheloni S, Tellez JO, Pisoni G, Longhi R, Moroni P, et al. Distribution of the pacemaker HCN4 channel mRNA and protein in the rabbit sinoatrial node. J Mol Cell Cardiol 2009;47:221–7.

The search of new therapeutic tools consisting of gene- and/or cell-based intervention aimed to restore compromised cardiac functions has prompted researchers to exploit the use of HCN channels to alter cellular electrical activity in order to generate, in normally quiescent substrates, stable rhythmic activity similar to that of native pacemaker myocytes. The specific features of pacemaker channels and in particular the fact that they are activated only at diastolic potentials and do not contribute to other phases of the action potentials, make them particularly suitable for such purpose. Early in vitro studies demonstrated that virus-mediated over-expression of HCN2 channels induced a significant increase in the rate of spontaneously beating neonatal ventricular myocytes by causing an If-mediated increase of the diastolic depolarization slope. This approach was later confirmed in vivo by showing that direct injection of the HCN2-adenovirus in the left atrium or into the ventricular conduction system of dogs, was able to induce ectopic regular spontaneous activity after AV block. Similarly, adenovirus-mediated over-expression of HCN1 or HCN4 was sufficient to induce a regular rhythm in quiescent cardiomyocyte. Alternative cell-based strategies, aimed to avoid the use of viruses, have been developed by engineering cells in order to express high levels of HCN channels. Engineered human mesenchymal stem cells (hMSCs) expressing either HCN2 or HCN4 have been shown in vitro to properly connect to neonatal cardiomyocytes and to increase their intrinsic spontaneous rhythm. HCN2-expressing hMSCs have also been successfully transplanted in canine left ventricular wall where they were able to induce stable ectopic beats.

Currently, ivabradine is marketed for treatment of chronic stable angina in patients with normal sinus rhythm who have a contraindication or intolerance to β-blockers; clinical studies of patients with chronic stable angina have shown that ivabradine acts as a pure heart rate-reducing agent and has anti-ischemic and anti-anginal properties equivalent to β-blockers and Ca2+ channel blockers and presents a good safety and tolerability profile even during long-term treatment. Mild visual symptoms (phosphenes) were occasionally reported, but were generally well tolerated. Additional information comes from results from a recent large clinical trial (BEAUTIFUL) which indicate that ivabradine treatment of patients with stable coronary artery disease (CAD) and heart rate ≥70 bpm can reduce the incidence of some CAD outcomes such as hospitalization for myocardial infarction and coronary revascularization.

The beat goes on: Cardiac pacemaking in extreme conditions

Christopher M.Wilson, Georgina K. Cox, Anthony P. Farrell
Comparative Biochemistry and Physiology, Part A xxx (2014) xxx–xxx

In order for an animal to survive, the heart beat must go on in all environmental conditions, or at least restart its beat. This review is about maintaining a rhythmic heartbeat under the extreme conditions of anoxia (or very severe hypoxia) and high temperatures. It starts by considering the primitive versions of the protein channels that are responsible for initiating the heartbeat, HCN channels, divulging recent findings from the ancestral craniate, the Pacific hagfish (Eptatretus stoutii). It then explores how a heartbeat can maintain a rhythm, albeit slower, for hours without any oxygen, and sometimes without autonomic innervation. It closes with a discussion of recent work on fishes, where the cardiac rhythm can become arrhythmic when a fish experiences extreme heat.

Sympathetic renal denervation: Hypertension beyond SYMPLICITY

Israel M. Barbash, Ron Waksman
Cardiovascular Revascularization Medicine 14 (2013) 229–235

Despite a wide range of drug treatment for hypertension, resistant hypertension rates remain high. The Symplicity™ Renal Denervation System (Medtronic, Santa Rosa, CA), which creates renal nerve denervation, has shown initial success in lowering blood pressure among patients with resistant  hypertension. Given the enormous market for this treatment approach, an estimated two dozen other companies are pursuing technologies with alternative approaches. Despite this fact, very little has been published on preclinical and clinical experience with these new devices. The current review summarizes the most prominent technologies in the pipeline and provides insight into the mechanism of action, preclinical, and clinical experience with these new devices

A large body of evidence has established the central role of the kidneys in hypertension, both as an affector and effector of the central sympathetic system [9]. Renal efferent sympathetic activity initiates processes towards fluid retention, such as the release of renin and increased tubular sodium reabsorption. Moreover, afferent sympathetic activity increases central sympathetic drive, which plays a major role in sustaining hypertension. In fact, historic studies of surgical sympathectomy in patients with resistant hypertension or malignant hypertension uncontrolled by pharmacotherapy were shown to be effective in reducing blood pressure, albeit with severe side effects. Thus, with the introduction of more effective medications, this procedure was abandoned. Renal sympathetic nerves run alongside the renal artery adventitia to enter the hilus of the kidney. Thereafter, they divide into smaller nerve bundles following the anatomic course of the renal blood vessels, penetrating the cortical and juxtamedullary areas inside the kidneys. Based on these anatomic features, it was postulated that creating local nerve injury along the renal arteries may achieve effective denervation.

A key issue in accomplishing effective RDN is to target the sympathetic nerve bundles lying in the adventitia of the renal arteries. Because the vast majority of devices currently under development are percutaneous, RDN is performed from within the vessel lumen. Thus, one of the most important features of such a device is the ability to minimize the damage to the renal artery wall.

Ultrasound energy consists of high-frequency sound waves emitted by a transducer within the catheter. This high energy can pass through surrounding fluids and can generate frictional heating in tissues resulting in a temperature increase that is sufficient to cause injury to the surrounding tissue, specifically the renal nerves. Based on these principles, several systems were developed and are currently being evaluated. ReCor Medical’s (Ronkonkoma, NY) PARADISE™ Percutaneous Renal Denervation System is based on delivery of high ultrasonic energy to induce nerve tissue injury. The PARADISE system is composed of two components: a 6 F-compatible balloon catheter with a cylindrical ultrasound transducer that emits ultrasound energy circumferentially (Fig. 2A)[ Ultrasound based renal denervation systems: (A) Percutaneous Renal Denervation System (PARADISE™); (B) TIVUS system]  and a portable generator which controls automated balloon inflation and deflation, and energy delivery. Energy is delivered in 3 different locations along the artery with 50 s inflation and delivery of ultrasound energy at each site. This device received CE mark in February 2012. For RDN, the PARADISE balloon catheter is positioned inside the renal artery and the generator automatically inflates the balloon, delivers the ultrasonic energy, and deflates the balloon. Endothelial thermal damage is prevented by cooled fluid in the balloon.

Radiofrequency based renal denervation systems

Radiofrequency based renal denervation systems: (A) Symplicity Renal Denervation System; (B) EnligHTN Renal Denervation System; (C) V2 bipolar balloon catheter; (D) OneShot Balloon catheter

Sample Entropy and Traditional Measures of Heart Rate Dynamics Reveal Different Modes of Cardiovascular Control During Low Intensity Exercise

Matthias Weippert, Martin Behrens, Annika Rieger and Kristin Behrens
Entropy 2014, 16, 5698-5711;

Biological time series like the normal heartbeat-to-heartbeat fluctuation demonstrate complex dynamics. Based on their potential to give additional information beyond traditional heart rate variability (HRV) indices, nonlinear parameters have been applied for investigating short and long term effects of exercise on heart rate (HR) control. However, despite their diagnosticity and their clinical significance, the physiological background of their behavior is not very well established. It is assumed that complexity and regularity measures are fundamentally different from traditional HRV indices and show no correlation to these measures. However, many researchers found at least modest correlations for some nonlinear measures and traditional HRV indices under different conditions. It has also been shown that complexity of short-term HRV is under control of the autonomic nervous system. Currently, there are only few studies available that compared the cardiovascular response pattern to different exercise modes at similar HR. Lindquist et al. found a stronger increase of systolic (SBP) and diastolic arterial blood pressure (DBP) during isometric handgrip compared to cycling at comparable HR of 90 bpm.

Nonlinear parameters of heart rate variability (HRV) have proven their prognostic value in clinical settings, but their physiological background is not very well established. We assessed the effects of low intensity isometric (ISO) and dynamic (DYN) exercise of the lower limbs on heart rate matched intensity on traditional and entropy measures of HRV. Due to changes of afferent feedback under DYN and ISO a distinct autonomic response, mirrored by HRV measures, was hypothesized. Five-minute inter-beat interval measurements of 43 healthy males (26.0 ± 3.1 years) were performed during rest, DYN and ISO in a randomized order. Blood pressures and rate pressure product were higher during ISO vs. DYN (p < 0.001). HRV indicators SDNN as well as low and high frequency power were significantly higher during ISO (p < 0.001 for all measures). Compared to DYN, sample entropy (SampEn) was lower during ISO (p < 0.001). Concluding, contraction mode itself is a significant modulator of the autonomic cardiovascular response to exercise. Compared to DYN, ISO evokes a stronger blood pressure response and an enhanced interplay between both autonomic branches. Non-linear HRV measures indicate a more regular behavior under ISO. Results support the view of the reciprocal antagonism being only one of many modes of autonomic heart rate control. Under different conditions; the identical “end product” heart rate might be achieved by other modes such as sympathovagal co-activation as well.

ANOVA revealed a significant effect of experimental condition on all cardiovascular measures and autonomic indices. Average HR raised moderately from 65 ± 9 bpm at baseline to 85 ± 9 bpm during both types of exercise. HR during the first exercise perfectly matched HR of the subsequent exercise; average difference was only 0.3 ± 1.5 bpm (range: −2.6 to 4.3 bpm). Accordingly, HR and average R-R interval did not differ between DYN and ISO. The traditional vagal modulation HRV measure RMSSD was also not affected by the exercise mode, whereas SDNN was. Natural log-transformed HRV spectral indices HFP and LFP, the normalized powers LF n. u. and HF n. u. as well SampEn (Figure 1) were significantly different between DYN and ISO. Interestingly, SampEn did not differ between REST and DYN. There was no difference of the LF/HF ratio between REST and ISO, whereas comparison of REST vs. DYN showed a statistical trend (p = 0.077). Further, there was a small effect of condition on the HF peak frequency (F(2; 84) = 4.959, p < 0.01, η² = 0.106). While HF peak significantly shifted from 0.22 ± 0.07 Hz during REST to 0.26 ± 0.09 Hz during DYN (p < 0.05), no difference was found between REST and ISO (0.23 ± 0. 07 Hz). Post-hoc pair wise comparison between DYN and ISO showed a statistical trend for the HF peak shift (p = 0.063). SBP and RPP were moderately, DBP and MAP largely affected by the type of exercise. In comparison to DYN, myocardial oxygen consumption, reflected by RPP, was about 5% higher under ISO. Correlation analysis revealed only modest associations between traditional HRV indices and entropy measures during the different experimental conditions. Consistent correlation coefficients across all conditions were found for SampEn and R-R length only.

Mean ± SD of sample entropy during REST, ISO, and DYN; N = 43.

Mean ± SD of sample entropy during REST, ISO, and DYN; N = 43.
*** = significantly different from rest on a p-level < 0.001;
§§§ = significantly different from the respective exercise condition on a p-level < 0.001.

Role of neurotensin and opioid receptors in the cardiorespiratory effects of [Ile9]PK20, a novel antinociceptive chimeric peptide

Katarzyna Kaczynska, M Szereda-Przestaszewska, P Kleczkowska, AW Lipkowski European Journal of Pharmaceutical Sciences 63 (2014) 8–13

Ile9PK20 is a novel hybrid of opioid–neurotensin peptides synthesized from the C-terminal hexapeptide of neurotensin and endomorphin-2 pharmacophore. This chimeric compound shows potent central and peripheral antinociceptive activity in experimental animals, however nothing is known about its influence on the respiratory and cardiovascular parameters.

The present study was designed to determine the cardiorespiratory effects exerted by an intravenous injection (i.v.) of [Ile9]PK20. Share of the vagal afferentation and the contribution of NTS1 neurotensin and opioid receptors were tested.

Intravenous injection of the hybrid at a dose of 100 lg/kg in the intact, anaesthetized rats provoked an increase in tidal volume preceded by a prompt short-lived decrease. Immediately after the end of injection brief acceleration of the respiratory rhythm appeared, and was ensued by the slowing down of breathing. Changes in respiration were concomitant with a bi-phasic response of the blood pressure: an immediate increase was followed by a sustained hypotension. Midcervical vagotomy eliminated the increase in tidal volume and respiratory rate responses. Antagonist of opioid receptors – naloxone hydrochloride eliminated only [Ile9]PK20-evoked decline in tidal volume response. Blockade of NTS1 receptors with an intravenous dose of SR 142,948, lessened the remaining cardiorespiratory effects. This study depicts that [Ile9]PK20 acting through neurotensin NTS1 receptors augments the tidal component of the breathing pattern and activates respiratory timing response through the vagal pathway. Blood pressure effects occur outside vagal afferentation and might result from activation of the central and peripheral vascular NTS1 receptors. In summary the respiratory effects of the hybrid appeared not to be profound, but they were accompanied with unfavorable prolonged hypotension.

Integrative regulation of human brain blood flow

Christopher K.Willie, Yu-Chieh Tzeng, Joseph A. Fisher and Philip N. Ainslie
J Physiol 2014; 592(5): pp 841–859

Herein, we review mechanisms regulating cerebral blood flow (CBF), with specific focus on humans. We revisit important concepts from the older literature and describe the interaction of various mechanisms of cerebrovascular control. We amalgamate this broad scope of information into a brief review, rather than detailing any one mechanism or area of research. The relationship between regulatory mechanisms is emphasized, but the following three broad categories of control are explicated:

  • the effect of blood gases and neuronal metabolism on CBF;
  • buffering of CBF with changes in blood pressure, termed cerebral autoregulation; and
  • the role of the autonomic nervous system in CBF regulation.

With respect to these control mechanisms, we provide evidence against several canonized paradigms of CBF control. Specifically, we corroborate the following four key theses:

(1) that cerebral autoregulation does not maintain constant perfusion through a mean arterial pressure range of 60–150 mmHg;
(2) that there is important stimulatory synergism and regulatory interdependence of arterial blood gases and blood pressure on CBF regulation;

(3) that cerebral autoregulation and cerebrovascular sensitivity to changes in arterial blood gases are not modulated solely at the pial arterioles; and
(4) that neurogenic control of the cerebral vasculature is an important player in autoregulatory function and, crucially, acts to buffer surges in perfusion pressure.
Finally, we summarize the state of our knowledge with respect to these areas, outline important gaps in the literature and suggest avenues for future research.

Integrative physiological and computational approaches to understand autonomic control of cerebral autoregulation

Can Ozan Tan and J. Andrew Taylor
Exp Physiol 99.1 (2014) pp 3–15

New Findings

  1. What is the topic of this review?

This review focuses on the autonomic control of the cerebral vasculature in health and disease from an integrative physiological and computational perspective.

  1. What advances does it highlight?

This review highlights recent studies exploring autonomic effectors of cerebral autoregulation as well as recent advances in experimental and analytical approaches to understand cerebral autoregulation.

The brain requires steady delivery of oxygen and glucose, without which neurodegeneration occurs within minutes. Thus, the ability of the cerebral vasculature to maintain relatively steady blood flow in the face of changing systemic pressure, i.e. cerebral autoregulation, is critical to neurophysiological health. Although the study of autoregulation dates to the early 20th century, only the recent availability of cerebral blood flow measures with high temporal resolution has allowed rapid, beat-by-beat measurements to explore the characteristics and mechanisms of autoregulation. These explorations have been further enhanced by the ability to apply sophisticated computational approaches that exploit the large amounts of data that can be acquired. These advances have led to unique insights. For example, recent studies have revealed characteristic time scales wherein cerebral autoregulation is most active, as well as specific regions wherein autonomic mechanisms are prepotent. However, given that effective cerebral autoregulation against pressure fluctuations results in relatively unchanging flow despite changing pressure, estimating the pressure–flow relationship can be limited by the error inherent in computational models of autoregulatory function. This review focuses on the autonomic neural control of the cerebral vasculature in health and disease from an integrative physiological perspective. It also provides a critical overview of the current analytical approaches to understand cerebral autoregulation.

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Biomarker Guided Therapy

Writer and Curator: Larry H. Bernstein, MD, FCAP

Novel serum protein biomarker panel revealed by mass spectrometry and its prognostic value in breast cancer

Liping Chung, K Moore, L Phillips, FM Boyle, DJ Marsh and RC Baxter
Breast Cancer Research 2014, 16:R63

Introduction: Serum profiling using proteomic techniques has great potential to detect biomarkers that might improve diagnosis and predict outcome for breast cancer patients (BC). This study used surface-enhanced laser desorption/ionization time-of-flight (SELDI-TOF) mass spectrometry (MS) to identify differentially expressed  proteins in sera from BC and healthy volunteers (HV), with the
goal  of developing a new prognostic biomarker panel.
Methods: Training set serum samples from 99 BC and 51 HV subjects were applied to four adsorptive chip surfaces (anion-exchange, cation-exchange, hydrophobic, and metal affinity) and analyzed by time-of-flight MS. For validation, 100 independent BC serum samples and 70 HV samples were analyzed similarly. Cluster analysis of protein spectra was performed to identify protein patterns related to BC and HV groups. Univariate and multivariate statistical analyses were used to develop a protein panel to distinguish breast cancer sera from healthy sera, and its prognostic potential was evaluated.
Results: From 51 protein peaks that were significantly up- or downregulated in BC patients by univariate analysis, binary logistic regression yielded five protein peaks that together classified BC and HV with a receiver operating characteristic (ROC) area-under-the-curve value of 0.961. Validation on an independent patient cohort confirmed the five-protein parameter (ROC value 0.939). The five-protein parameter showed positive association with large tumor size (P = 0.018) and lymph node involvement (P = 0.016). By matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS, immunoprecipitation and western blotting the proteins were identified as a fragment of apolipoprotein H (ApoH), ApoCI, complement C3a, transthyretin, and ApoAI. Kaplan-Meier analysis on 181 subjects after median follow-up of >5 years demonstrated that the panel significantly predicted disease-free survival (P = 0.005), its efficacy apparently greater in women with estrogen receptor (ER)-negative tumors (n = 50, P = 0.003) compared to ER-positive (n = 131, P = 0.161), although the influence of ER status needs to be confirmed after longer follow-up.
Conclusions: Protein mass profiling by MS has revealed five serum proteins which, in combination, can distinguish between serum from women with breast cancer and healthy control subjects with high sensitivity and specificity. The five-protein panel significantly predicts recurrence-free survival in women with ER-negative tumors and may have value in the management of these patients.

Variants of uncertain significance in BRCA: a harbinger of ethical and policy issues to come?

Jae Yeon Cheon, Jessica Mozersky and Robert Cook-Deegan
Genome Medicine 2014, 6:121

After two decades of genetic testing and research, the BRCA1 and BRCA2 genes are two of the most well-characterized genes in the human genome. As a result, variants of uncertain significance (VUS; also called variants of unknown significance) are reported less frequently than for genes that have been less thoroughly studied. However, VUS continue to be uncovered, even for BRCA1/2. The increasing use of multi-gene panels and whole-genome and whole-exome sequencing will lead to higher rates of VUS detection because more genes are being tested, and most genomic loci have been far less intensively characterized than BRCA1/2. In this article, we draw attention to ethical and policy-related issues that will emerge. Experience garnered from BRCA1/2 testing is a useful introduction to the challenges of detecting VUS in other genetic testing contexts, while features unique to BRCA1/2 suggest key differences between the BRCA experience and the current challenges of multi-gene panels in clinical care. We propose lines of research and policy development, emphasizing the importance of pooling data into a centralized open-access database for the storage of gene variants to improve VUS interpretation. In addition, establishing ethical norms and regulated practices for sharing and curating data, analytical algorithms, interpretive frameworks and patient re-contact are important policy areas.

The Significance of Normal Pretreatment Levels of CA125 (<35 U/mL) in Epithelial Ovarian Carcinoma

Joseph Menczer,  Erez Ben-Shem,  Abraham Golan, and Tally Levy
Rambam Maimonides Med J 2015;6 (1):e0005.

Objective: To assess the association between normal CA125 levels at diagnosis of epithelial ovarian carcinoma (EOC) with prognostic factors and with outcome.
Methods: The study group consisted of histologically confirmed EOC patients with normal pretreatment CA125 levels, and the controls consisted of EOC patients with elevated (≥35 U/mL) pretreatment CA125 levels, diagnosed and treated between 1995 and 2112. Study and control group patients fulfilled the following criteria: 1) their pretreatment CA125 levels were assessed; 2) they had full standard primary treatment, i.e. cytoreductive surgery and cisplatin-based chemotherapy; and 3) they were followed every 2–4 months during the first two years and every 4–6 months thereafter.
Results: Of 114 EOC patients who fulfilled the inclusion criteria, 22 (19.3%) had normal pretreatment CA125 levels. The control group consisted of the remaining 92 patients with ≥35 U/mL serum CA125 levels pretreatment. The proportion of patients with early-stage and low-grade disease, with optimal cytoreduction, and with platin-sensitive tumors was significantly higher in the study group than in the control group. The progression-free survival (PFS) and overall survival (OS) were significantly higher in the study group than in the control group on univariate analysis but not on multivariate analysis.

Higher gene expression variability in the more aggressive subtype of chronic lymphocytic leukemia

Simone Ecker, Vera Pancaldi, Daniel Rico and Alfonso Valencia
Genome Medicine (2015) 7:8

Background: Chronic lymphocytic leukemia (CLL) presents two subtypes which have drastically different clinical outcomes, IgVH mutated (M-CLL) and IgVH unmutated (U-CLL). So far, these two subtypes are not associated to clear differences in gene expression profiles. Interestingly, recent results have highlighted important roles for heterogeneity, both at the genetic and at the epigenetic level in CLL progression.
Methods: We analyzed gene expression data of two large cohorts of CLL patients and quantified expression variability across individuals to investigate differences between the two subtypes using different measures and statistical tests. Functional significance was explored by pathway enrichment and network analyses. Furthermore, we implemented a random forest approach based on expression variability to classify patients into disease subtypes.
Results: We found that U-CLL, the more aggressive type of the disease, shows significantly increased variability of gene expression across patients and that, overall, genes that show higher variability in the aggressive subtype are related to cell cycle, development and inter-cellular communication. These functions indicate a potential relation between gene expression variability and the faster progression of this CLL subtype. Finally, a classifier based on gene expression variability was able to correctly predict the disease subtype of CLL patients.
Conclusions: There are strong relations between gene expression variability and disease subtype linking significantly increased expression variability to phenotypes such as aggressiveness and resistance to therapy in CLL.

The Emerging Roles of Thyroglobulin

Yuqian Luo, Yuko Ishido, Naoki Hiroi, Norihisa Ishii, and Koichi Suzuki
Advances in Endocrinology 2014, Article ID 189194, 7 pages

Thyroglobulin (Tg), the most important and abundant protein in thyroid follicles, is well known for its essential role in thyroid hormone synthesis. In addition to its conventional role as the precursor of thyroid hormones, we have uncovered a novel function of Tg as an endogenous regulator of follicular function over the past decade. The newly discovered negative feedback effect of Tg on follicular function observed in the rat and human thyroid provides an alternative explanation for the observation of follicle heterogeneity. Given the essential role of the regulatory effects of Tg, we consider that dysregulation of normal Tg function is associated with multiple human thyroid diseases including autoimmune thyroid disease and thyroid cancer. Additionally, extrathyroid Tg may serve a regulatory function in other organs. Further exploration of Tg action, especially at the molecular level, is needed to obtain a better understanding of both the physiological and pathological roles of Tg.

The GUIDE-IT trial will help doctors find a new standard of care for heart failure.

Heart failure affects more than 25 million people worldwide, including 5.8 million in the United States and 6.9 million in Europe. About one to two percent of adults in developed countries have been diagnosed with heart failure; this increases to more than 10 percent in people over age 70. Moreover, heart failure accounts for more than 17 percent of Medicare spending and about 5 percent of total US healthcare spending. The cost to society in the US is about 30 billion dollars a year—and rising.

For people hospitalized due to heart failure, the outlook isn’t encouraging. Following discharge, one in four patients is likely to be back in the hospital in less than a month. With every acute heart failure event that requires readmission, the chances of dying from the disease increase.

Heart failure occurs when the heart is unable to fill with or pump sufficient blood to meet the needs of the body. Some heart failure symptoms—shortness of breath, fatigue and fluid buildup—which are present in other health problems. Heart failure may develop from coronary artery disease, high blood pressure, cardiomyopathy, heart valve disease, arrhythmias, viral or bacterial infections, and congenital heart defects. As a consequence, these patients often have additional diseases (comorbidities) and managing heart failure can be extremely challenging.

There have been no new drugs for heart failure in more than a decade. The last breakthrough was cardiac resynchronization therapy, a device and not a drug. The goals of therapy are to treat heart failure’s underlying causes, reduce symptoms, improve the patient’s quality of life and keep the disease from getting worse.

More than a pump

The heart isn’t just a muscle pumping blood through the body. It is also an endocrine gland that secretes peptides and hormones. When the heart is failing, its stressed cells release larger amounts of substances known as natriuretic peptides, including N-terminal prohormone brain natriuretic peptide, or NT-proBNP.

Roche’s NT-proBNP test measures the levels of this peptide and helps doctors to determine whether patients are suffering from heart failure and to assess their prognosis. Most recently, NT-proBNP has also been shown to help physicians guide and adjust the patient’s drug therapy. The objective of the pivotal GUIDE-IT trial is to demonstrate the efficacy and safety of NT-proBNP guided heart failure therapy.

Sponsored by the National Institutes of Health (NIH), the GUIDE-IT trial will help doctors answer important questions about NT-proBNP’s impact on medical care. About 1100 patients are enrolled in this robustly powered, randomized controlled trial comparing NT-proBNP guided therapy on top of standard care versus standard care alone in high-risk heart failure patients. Its primary endpoint is time to cardiovascular death or first heart failure hospitalization.

With the NT-proBNP biomarker, doctors can create personalized treatment plans for patients to substantially reduce mortality and morbidity. It can be viewed as a companion diagnostic that works with all the drugs recommended by the major guidelines.

Finding new answers

GUIDE-IT will last five years and involve approximately 45 trial sites in the United States. The first group of patients will be enrolled by the end of 2012.

“We need to take a more strategic approach if we are going to meet the AHA/ASA’s 2020 goal of reducing heart failure hospitalizations by 20 percent,” Dr. O’Connor, Chief of the Division of Cardiovascular Medicine at Duke Heart Center in Durham, North Carolina, said at a media briefing held in October at Roche Diagnostics International in Rotkreuz, Switzerland.
The relative and combined ability of: high-sensitivity cardiac troponin T, and N-terminal pro-B-type natriuretic Peptide – to predict cardiovascular events and death in patients with type 2 diabetes.

Hillis GS; Welsh P; Chalmers J; Perkovic V; Chow CK; Li Q; Jun M; Neal B; et al.

OBJECTIVE Current methods of risk stratification in patients with type 2 diabetes are suboptimal. The current study assesses the ability of N-terminal pro-B-type natriuretic peptide (NT-proBNP) and high-sensitivity cardiac troponin T (hs-cTnT) to improve the prediction of cardiovascular events and death in patients with type 2 diabetes.
RESEARCH DESIGN AND METHODS A nested case-cohort study was performed in 3,862 patients who participated in the Action in Diabetes and Vascular Disease: Preterax and Diamicron Modified Release Controlled Evaluation (ADVANCE) trial. RESULTS Seven hundred nine (18%) patients experienced a major cardiovascular event (composite of cardiovascular death, nonfatal myocardial infarction, or nonfatal stroke) and 706 (18%) died during a median of 5 years of follow-up. In Cox regression models, adjusting for all established risk predictors, the hazard ratio for cardiovascular events for NT-proBNP was 1.95 per 1 SD increase (95% CI 1.72, 2.20) and the hazard ratio for hs-cTnT was 1.50 per 1 SD increase (95% CI 1.36, 1.65). The hazard ratios for death were 1.97 (95% CI 1.73, 2.24) and 1.52 (95% CI 1.37, 1.67), respectively. The addition of either marker improved 5-year risk classification for cardiovascular events (net reclassification index in continuous model, 39% for NT-proBNP and 46% for hs-cTnT). Likewise, both markers greatly improved the accuracy with which the 5-year risk of death was predicted. The combination of both markers provided optimal risk discrimination.
CONCLUSIONS NT-proBNP and hs-cTnT appear to greatly improve the accuracy with which the risk of cardiovascular events or death can be estimated in patients with type 2 diabetes.

Genetics and Heart Failure: A Concise Guide for the Clinician

Cécile Skrzynia, Jonathan S. Berg, Monte S. Willis and Brian C. Jensen
Current Cardiology Reviews, 2013; 9.

Abstract: The pathogenesis of heart failure involves a complex interaction between genetic and environmental factors. Genetic factors may influence the susceptibility to the underlying etiology of heart failure, the rapidity of disease progression, or the response to pharmacologic therapy. The genetic contribution to heart failure is relatively minor in most multifactorial cases, but more direct and profound in the case of familial dilated cardiomyopathy. Early studies of genetic risk for heart failure focused on polymorphisms in genes integral to the adrenergic and renin-angiotensin-aldosterone system. Some of these variants were found to increase the risk of developing heart failure, and others appeared to affect the therapeutic response to neurohormonal antagonists. Regardless, each variant individually confers a relatively modest increase in risk and likely requires complex interaction with other variants and the environment for heart failure to develop. Dilated cardiomyopathy frequently leads to heart failure, and a genetic etiology increasingly has been recognized in cases previously considered to be “idiopathic”. Up to 50% of dilated cardiomyopathy cases without other cause likely are due to a heritable genetic mutation. Such mutations typically are found in genes encoding sarcomeric proteins and are inherited in an autosomal dominant fashion. In recent years, rapid advances in sequencing technology have improved our ability to diagnose familial dilated cardiomyopathy and those diagnostic tests are available widely. Optimal care for the expanding population of patients with heritable heart failure involves counselors and physicians with specialized training in genetics, but numerous online genetics resources are available to practicing clinicians.

Cardiac Troponin Testing Is Overused after the Rule-In or Rule-Out of Myocardial Infarction

Olaia Rodriguez Fraga, Y Sandoval, SA Love, ZJ McKinney, MAM Murakami, SW Smith, FS Apple
Clinical Chemistry 2015; 61:2

No good studies have systematically evaluated appropriate clinical utilization of cardiac troponin testing in the clinical setting of the rule-in and rule-out of myocardial infarction (MI). Our collective 100-plus years of clinical and laboratory experience suggested that provider test ordering and use of cardiac troponin has been excessive after a diagnosis of MI or no MI has been determined. There is no evidence that supports continuation of cardiac troponin testing after a diagnosis is made.

Number of cTnI results demonstrating excessive orders by diagnosis

Number of cTnI results demonstrating excessive orders by diagnosis

Time and Frequency Domain Analysis of Heart Rate Variability and their orrelations in Diabetes Mellitus
T. Ahamed Seyd, V. I. Thajudin Ahamed, Jeevamma Jacob, Paul Joseph K
Intl J Biolog and Life Sciences 2008; 4(1)

Diabetes mellitus (DM) is frequently characterized by autonomic nervous dysfunction. Analysis of heart rate variability (HRV) has become a popular noninvasive tool for assessing the activities of autonomic nervous system (ANS). In this paper, changes in ANS
activity are quantified by means of frequency and time domain analysis of R-R interval variability. Electrocardiograms (ECG) of 16 patients suffering from DM and of 16 healthy volunteers were recorded. Frequency domain analysis of extracted normal to normal interval (NN interval) data indicates significant difference in very low frequency (VLF) power, low frequency (LF) power and high frequency (HF) power, between the DM patients and control group. Time domain measures, standard deviation of NN interval (SDNN), root mean square of successive NN interval differences (RMSSD), successive NN intervals differing more than 50 ms (NN50 Count), percentage value of NN50 count (pNN50), HRV triangular index and triangular interpolation of NN intervals (TINN) also show significant difference between the DM patients and control group.

Power Spectral Density of the RR interval of a 55 year old healthy volunteer

Power Spectral Density of the RR interval of a 55 year old healthy volunteer

Power Spectral Density of the RR interval of a 55 year old healthy volunteer

Power Spectral Density of the RR interval of a 62 year old woman suffering

Power Spectral Density of the RR interval of a 62 year old woman suffering

Power Spectral Density of the RR interval of a 62 year old woman suffering
from diabetes for the last 15 years

HRV analysis has gained much importance in recent years, as a technique employed to explore the activity of ANS, and as an important early marker for identifying different pathological conditions. DM is a disease in which the cardiac autonomic activity is progressively compromised. Our investigation indicates that different time domain and frequency domain measures of HRV would be able to provide valuable information regarding the autonomic dysfunction to DM.

Time domain and frequency domain analysis of the RR interval variability of diabetic and normal subjects shows that there is significant difference in these measures for DM patients with respect to normal subjects. Variation of the HRV parameters indicates changes in ANS activity of DM patients. This can provide valid information regarding autonomic neuropathy in people with diabetes. It may be noted that these methods can detect changes before clinical signs appear. So we can expect that these measures enable early detection and treatment/subsequent management of patients and thus can avoid acute and chronic complications.

Multiparametric diagnostics of cardiomyopathies by microRNA signatures

Christine S. Siegismund & Maria Rohde & Uwe Kühl & Dirk Lassner
Microchim Acta 2014

The diagnosis of cardiomyopathies by endomyocardial biopsy analysis is the gold standard for confirmation of causative reasons but is failing if a sample does not contain the area of interest due to focal pathology. Biopsies are revealing an extract of the current situation of the heart muscle only, and the need for global organ-specific or systemic markers is obvious in order to minimize sampling errors. Global markers like specific gene expression signatures in myocardial tissue may therefore reflect the focal situation or condition of the whole myocardium. Besides gene expression profiles, microRNAs (miRNAs) represent a new group of stable biomarkers that are detectable both in tissue and body fluids. Such miRNAs may serve as cardiological biomarkers to characterize inflammatory processes, to confirm viral infections, and to differentiate various forms of infection.
The predictive power of single miRNAs for diagnosis of complex diseases may be further increased if several distinctly deregulated candidates are combined to form a specific miRNA signature. Diagnostic systems that generate disease related miRNA profiles are based on microarrays, bead-based oligo sorbent assays, or on assays based on real-time polymerase chain reactions and placed on microfluidic cards or nanowell plates. Multiparametric diagnostic systems that can measure differentially expressed miRNAs may become the diagnostic tool of the future due to their predictive value with respect to clinical course, therapeutic decisions, and therapy monitoring. We discuss here specific merits, limitations and the potential of currently available analytical platforms for diagnostics of heart muscle diseases based on miRNA profiling.

Predictive value of plasma galectin-3 levels in heart failure with reduced and preserved ejection fraction

Rudolf A. de Boer, DJA Lok, T Jaarsma, P van der Meer, AA Voors, et al.
Annals Med, 2011; 43: 60–68

We studied the prognostic value of base-line galectin-3 in a large HF cohort, with preserved and reduced left ventricular ejection fraction (LVEF), and compared this to other biomarkers.
Methods. We studied 592 HF patients who had been hospitalized for HF and were followed for 18 months. The primary end-point was a composite of all-cause mortality and HF hospitalization.
Results. A doubling of galectin-3 levels was associated with a hazard ratio (HR) of 1.97 (1.62–2.42) for the primary outcome (P= 0.001). After correction for age, gender, BNP, eGFR, and diabetes the HR was 1.38 (1.07–1.78; P= 0.015). Galectin-3 levels were correlated with higher IL -6 and CRP levels (P= 0.002). Changes of galectin-3 levels after 6 months did not add prognostic information to the base-line value (n= 291); however, combining plasma galectin-3 and BNP levels increased prognostic value over either biomarker alone (ROC analysis, P = 0.05). The predictive value of galectin-3 was stronger in patients with preserved LVEF (n= 114) compared to patients with reduced LVEF (P= 0.001).
Conclusions. Galectin-3 is an independent marker for outcome in HF and appears to be particularly useful in HF patients with preserved LVEF.

Criteria for the use of omics-based predictors in clinical trials

Lisa M. McShane, MM Cavenagh, TG Lively, DA Eberhard, et al.
Nature  17 Oct 2013; 502: 317-320.

The US National Cancer Institute (NCI), in collaboration with scientists representing multiple areas of expertise relevant to ‘omics’-based test development, has developed a checklist of criteria that can be used to determine the readiness of omics-based tests for guiding patient care in clinical trials. The checklist criteria cover issues relating to specimens, assays, mathematical modelling, clinical trial design, and ethical, legal and regulatory aspects. Funding bodies and journals are encouraged to consider the checklist, which they may find useful for assessing study quality and evidence strength. The checklist will be used to evaluate proposals for NCI-sponsored clinical
trials in which omics tests will be used to guide therapy.

M-Atrial Natriuretic Peptide and Nitroglycerin in a Canine Model of Experimental Acute Hypertensive Heart Failure: Differential Actions of 2 cGMP Activating Therapeutics.

Paul M McKie, Alessandro Cataliotti, Tomoko Ichiki, S Jeson Sangaralingham, Horng H Chen, John C Burnett
J Am Heart Assoc 01/2014; 3(1):e000206.

Systemic hypertension is a common characteristic in acute heart failure (HF). This increasingly recognized phenotype is commonly associated with renal dysfunction and there is an unmet need for renal enhancing therapies. In a canine model of HF and acute vasoconstrictive hypertension we characterized and compared the cardiorenal actions of M-atrial natriuretic peptide (M-ANP), a novel particulate guanylyl cyclase (pGC) activator, and nitroglycerin, a soluble guanylyl cyclase (sGC) activator.
HF was induced by rapid RV pacing (180 beats per minute) for 10 days. On day 11, hypertension was induced by continuous angiotensin II infusion. We characterized the cardiorenal and humoral actions prior to, during, and following intravenous M-ANP (n=7), nitroglycerin (n=7), and vehicle (n=7) infusion. Mean arterial pressure (MAP) was reduced by M-ANP (139±4 to 118±3 mm Hg, P<0.05) and nitroglycerin (137±3 to 116±4 mm Hg, P<0.05); similar findings were recorded for pulmonary wedge pressure (PCWP) with M-ANP (12±2 to 6±2 mm Hg, P<0.05) and nitroglycerin (12±1 to 6±1 mm Hg, P<0.05). M-ANP enhanced renal function with significant increases (P<0.05) in glomerular filtration rate (38±4 to 53±5 mL/min), renal blood flow (132±18 to 236±23 mL/min), and natriuresis (11±4 to 689±37 mEq/min) and also inhibited aldosterone activation (32±3 to 23±2 ng/dL, P<0.05), whereas nitroglycerin had no significant (P>0.05) effects on these renal parameters or aldosterone activation.
Our results advance the differential cardiorenal actions of pGC (M-ANP) and sGC (nitroglycerin) mediated cGMP activation. These distinct renal and aldosterone modulating actions make M-ANP an attractive therapeutic for HF with concomitant hypertension, where renal protection is a key therapeutic goal.

Genome-Wide Association Study of a Heart Failure Related Metabolomic Profile Among African Americans in the Atherosclerosis Risk in Communities (ARIC) Study

Bing Yu, Y Zheng, D Alexander, TA Manolio, A Alonso, JA Nettleton, & E Boerwinkle
Genet Epidemiol 2013; 00:1–6,

Both the prevalence and incidence of heart failure (HF) are increasing, especially among African Americans, but no large-scale, genome-wide association study (GWAS) of HF-related metabolites has been reported. We sought to identify novel genetic variants that are associated with metabolites previously reported to relate to HF incidence. GWASs of three metabolites identified previously as risk factors for incident HF (pyroglutamine, dihydroxy docosatrienoic acid, and X-11787, being either hydroxy-leucine or hydroxy-isoleucine) were performed in 1,260 African Americans free of HF at the baseline examination of the Atherosclerosis Risk in Communities (ARIC) study. A significant association on chromosome 5q33 (rs10463316, MAF = 0.358, P-value = 1.92 × 10−10) was identified for pyroglutamine. One region on chromosome 2p13 contained a nonsynonymous substitution in N-acetyltransferase 8 (NAT8) was associated with X-11787 (rs13538, MAF = 0.481, P-value = 1.71 × 10−23). The smallest P-value for dihydroxy docosatrienoic acid was rs4006531 on chromosome 8q24 (MAF = 0.400, P-value = 6.98 × 10−7). None of the above SNPs were individually associated with incident HF, but a genetic risk score (GRS) created by summing the most significant risk alleles from each metabolite detected 11% greater risk of HF per allele. In summary, we identified three loci associated with previously reported HF-related metabolites. Further use of metabolomics technology will facilitate replication of these findings in independent samples.

Global Left Atrial Strain Correlates with CHADS2 Risk Score in Patients with Atrial Fibrillation

SK Saha, PL Anderson, G Caracciolo, A Kiotsekoglou, S Wilansky, S Govind, et al.
J Am Soc Echocardiogr 2011; 24(5): 506-512.

Background: The aim of this cross-sectional study was to explore the association between echocardiographic parameters and CHADS2 score in patients with nonvalvular atrial fibrillation (AF).
Methods: Seventy-seven subjects (36 patients with AF, 41 control subjects) underwent standard twodimensional, Doppler, and speckle-tracking echocardiography to compute regional and global left atrial (LA) strain.
Results: Global longitudinal LA strain was reduced in patients with AF compared with controls (P < .001) and was a predictor of high risk for thromboembolism (CHADS2 score $ 2; odds ratio, 0.86; P = .02). LA strain indexes showed good interobserver and intraobserver variability. In sequential Cox models, the prediction of hospitalization and/or death was improved by addition of global LA strain and indexed LA volume to CHADS2 score (P = .003).
Conclusions: LA strain is a reproducible marker of dynamic LA function and a predictor of stroke risk and cardiovascular outcomes in patients with AF.

Gene Expression and Genetic Variation in Human Atria

Honghuang Lin, EV Dolmatova, MP Morley, KL Lunetta, et al.
Heart Rhythm, HRTHM5533. PII: S1547-5271(13)01226-5

Background— The human left and right atria have different susceptibilities to develop atrial fibrillation (AF). However, the molecular events related to structural and functional changes that enhance AF susceptibility are still poorly understood.
Objective— To characterize gene expression and genetic variation in human atria.
Methods— We studied the gene expression profiles and genetic variations in 53 left atrial and 52 right atrial tissue samples collected from the Myocardial Applied Genomics Network (MAGNet) repository. The tissues were collected from heart failure patients undergoing transplantation and from unused organ donor hearts with normal ventricular function. Gene expression was profiled using the Affymetrix GeneChip Human Genome U133A Array. Genetic variation was profiled using the Affymetrix Genome-Wide Human SNP Array 6.0.
Results— We found that 109 genes were differentially expressed between left and right atrial tissues. A total of 187 and 259 significant cis-associations between transcript levels and genetic variants were identified in left and right atrial tissues, respectively. We also found that a SNP at a known AF locus, rs3740293, was associated with the expression of MYOZ1 in both left and right atrial tissues. Conclusion— We found a distinct transcriptional profile between the right and left atrium, and extensive cis-associations between atrial transcripts and common genetic variants. Our results implicate MYOZ1 as the causative gene at the chromosome 10q22 locus for AF.

Atrial Natriuretic Peptide Single Nucleotide Polymorphisms in Patients with Nonfamilial Structural Atrial Fibrillation

Pietro Francia, A Ricotta, A Frattari, R Stanzione, A Modestino, et al.
Clinical Medicine Insights: Cardiology 2013:7 153–159

Background: Atrial natriuretic peptide (ANP) has antihypertrophic and antifibrotic properties that are relevant to AF substrates. The −G664C and rs5065 ANP single nucleotide polymorphisms (SNP) have been described in association with clinical phenotypes, including hypertension and left ventricular hypertrophy. A recent study assessed the association of early AF and rs5065 SNPs in low-risk subjects. In a Caucasian population with moderate-to-high cardiovascular risk profile and structural AF, we conducted a case-control study to assess whether the ANP −G664C and rs5065 SNP associate with nonfamilial structural AF.
Methods: 168 patients with nonfamilial structural AF and 168 age- and sex-matched controls were recruited. The rs5065 and −G664C ANP SNPs were genotyped.
Results: The study population had a moderate-to-high cardiovascular risk profile with 86% having hypertension, 23% diabetes, 26% previous myocardial infarction, and 23% left ventricular systolic dysfunction. Patients with AF had greater left atrial diameter (44 ± 7
vs. 39 ± 5 mm; P , 0.001) and higher plasma NTproANP levels (6240 ± 5317 vs. 3649 ± 2946 pmol/mL; P , 0.01). Odds ratios (ORs)
for rs5065 and −G664C gene variants were 1.1 (95% confidence interval [CI], 0.7–1.8; P = 0.71) and 1.2 (95% CI, 0.3–3.2; P = 0.79), respectively, indicating no association with AF. There were no differences in baseline clinical characteristics among carriers and noncarriers of the −664C and rs5065 minor allele variants.
Conclusions: We report lack of association between the rs5065 and −G664C ANP gene SNPs and AF in a Caucasian population of patients with structural AF. Further studies will clarify whether these or other ANP gene variants affect the risk of different subphenotypes of AF driven by distinct pathophysiological mechanisms.

N-terminal proBNP and mortality in hospitalized patients with heart failure and preserved vs. reduced systolic function: data from the prospective Copenhagen Hospital Heart Failure Study (CHHF)

Kirk, M. Bay, J. Parnerc, K. Krogsgaard, T.M. Herzog, S. Boesgaard, et al.
Eur Journal Heart Failure 6 (2004) 335–341

Preserved systolic function among heart failure patients is a common finding, a fact that has only recently been fully appreciated. The aim of the present study was to examine the value of NT-proBNP to predict mortality in relation to established risk factors among consecutively hospitalised heart failure patients and secondly to characterise patients in relation to preserved and reduced systolic function. Material: At the time of admission 2230 consecutively hospitalised patients had their cardiac status evaluated through determinations of NT-proBNP, echocardiography, clinical examination and medical history. Follow-up was performed 1 year later in all patients. Results: 161 patients fulfilled strict diagnostic criteria for heart failure (HF). In this subgroup of patients 1-year mortality was approximately 30% and significantly higher as compared to the remaining non-heart failure population (approx. 16%). Using univariate analysis left ventricular ejection fraction (LVEF), New York Heart Association classification (NYHA) and plasma levels of NT-proBNP all predicted mortality independently. However, regardless of systolic function, age and NYHA class, risk-stratification was provided by measurements of NT-proBNP. Having measured plasma levels of NT-proBNP, LVEF did not provide any additional prognostic information on mortality among heart failure patients (multivariate analysis).
Conclusion: The results show that independent of LVEF, measurements of NT-proBNP add additional prognostic information. It is concluded that NT-proBNP is a strong predictor of 1-year mortality in consecutively hospitalised patients with heart failure with preserved as well as reduced systolic function.

N-terminal pro-B-type natriuretic peptide and the prediction of primary cardiovascular events: results from 15-year follow-up of WOSCOPS

Paul Welsh, Orla Doolin, Peter Willeit, Chris Packard, Peter Macfarlane, et al.
Eur Heart Journal 2014.

Aims: To test whether N-terminal pro-B-type natriuretic peptide (NT-proBNP) was independently associated with, and improved the prediction of, cardiovascular disease (CVD) in a primary prevention cohort.
Methods and results:  In the West of Scotland Coronary Prevention Study (WOSCOPS), a cohort of middle-aged men with hypercholesterolemia at a moderate risk of CVD, we related the baseline NT-proBNP (geometric mean 28 pg/mL) in 4801 men to the risk of CVD over 15 years during which 1690 experienced CVD events. Taking into account the competing risk of non-CVD death, NT-proBNP was associated with an increased risk of all CVD [HR: 1.17 (95% CI: 1.11–1.23) per standard deviation increase in log NT-proBNP] after adjustment for classical and clinical cardiovascular risk factors plus C-reactive protein. N-terminal pro-B-type natriuretic peptide was more strongly related to the risk of fatal [HR: 1.34 (95% CI: 1.19–1.52)] than non-fatal CVD [HR: 1.17 (95% CI: 1.10–1.24)] (P ¼ 0.022). The addition of NT-proBNP to traditional risk factors improved the C-index (+0.013; P , 0.001). The continuous net reclassification index improved with the addition of NT-proBNP by 19.8% (95% CI: 13.6–25.9%) compared with 9.8% (95% CI: 4.2–15.6%) with the addition of C-reactive protein. N-terminal pro-B-type natriuretic peptide correctly reclassified 14.7% of events, whereas C-reactive protein correctly reclassified 3.4% of events. Results were similar in the 4128 men without evidence of angina, nitrate prescription, minor ECG abnormalities, or prior cerebrovascular disease.
Conclusion: N-terminal pro-B-type natriuretic peptide predicts CVD events in men without clinical evidence of CHD, angina, or history of stroke, and appears related more strongly to the risk for fatal events. N-terminal pro-B-type natriuretic peptide also provides moderate risk discrimination, in excess of that provided by the measurement of C-reactive protein.

Effect of B-type natriuretic peptide-guided treatment of chronic heart failure on total mortality and hospitalization: an individual patient meta-analysis

Richard W. Troughton, Christopher M. Frampton, Hans-Peter Brunner-La Rocca,
Matthias Pfisterer, Luc W.M. Eurlings, Hans Erntell, Hans Persson, et al.
Eur Heart J 2014; 35: 1559–1567

Aims Natriuretic peptide-guided (NP-guided) treatment of heart failure has been tested against standard clinically guided care in multiple studies, but findings have been limited by study size. We sought to perform an individual patient data metaanalysis to evaluate the effect of NP-guided treatment of heart failure on all-cause mortality.
Methods and results
Eligible randomized clinical trials were identified from searches of Medline andEMBASEdatabases and the Cochrane Clinical
Trials Register. The primary pre-specified outcome, all-cause mortality was tested using a Cox proportional hazards regression model that included study of origin, age (< 75 or ≥75 years), and left ventricular ejection fraction (LVEF, ≤45 or .45%) as covariates. Secondary endpoints included heart failure or cardiovascular hospitalization. Of 11 eligible studies, 9 provided individual patient data and 2 aggregate data. For the primary endpoint individual data from 2000 patients were included, 994 randomized to clinically guided care and 1006 to NP-guided care. All-cause mortality was significantly reduced by NP-guided treatment [hazard ratio = 0.62 (0.45–0.86);
P = 0.004] with no heterogeneity between studies or interaction with LVEF. The survival benefit from NP-guided therapy was seen in younger ( <75 years) patients [0.62 (0.45–0.85); P = 0.004] but not older (≥75 years) patients [0.98 (0.75–1.27); P = 0.96]. Hospitalization due to heart failure [0.80 (0.67–0.94); P = 0.009] or cardiovascular disease [0.82 (0.67–0.99); P = 0.048]was significantly lower in NP-guided patients with no heterogeneity between studies and no interaction with age or LVEF.
Conclusion: Natriuretic peptide-guided treatment of heart failure reduces all-cause mortality in patients aged < 75 years and overall reduces heart failure and cardiovascular hospitalization.

Diagnostic and prognostic evaluation of left ventricular systolic heart failure by plasma N-terminal pro-brain natriuretic peptide concentrations in a large sample of the general population

B A Groenning, I Raymond, P R Hildebrandt, J C Nilsson, M Baumann, F Pedersen
Heart 2004;90:297–303.

Objective: To evaluate N-terminal pro-brain natriuretic peptide (NT-proBNP) as a diagnostic and prognostic marker for systolic heart failure in the general population.
Design: Study participants, randomly selected to be representative of the background population, filled in a heart failure questionnaire and underwent pulse and blood pressure measurements, electrocardiography, echocardiography, and blood sampling and were followed up for a median (range) period of 805 (6021171) days.
Setting: Participants were recruited from four randomly selected general practitioners and were examined in a Copenhagen university hospital.
Patients: 382 women and 290 men in four age groups (50259 (n = 174); 60269 (n = 204); 70279 (n = 174); > 80 years (n = 120)).
Main outcome measures: Value of NT-proBNP in evaluating patients with symptoms of heart failure and impaired left ventricular (LV) systolic function; prognostic value of NT-proBNP for mortality and hospital admissions.
Results: In 38 (5.6%) participants LV ejection fraction (LVEF) was (40%. NT-proBNP identified patients with symptoms of heart failure and LVEF (40% with a sensitivity of 0.92, a specificity of 0.86, positive and negative predictive values of 0.11 and 1.00, and area under the curve of 0.94. NT-proBNP was the strongest independent predictor of mortality (hazard ratio (HR) = 5.70, p = 0.0001), hospital admissions for heart failure (HR = 13.83, p = 0.0001), and other cardiac admissions (HR = 3.69, p = 0.0001). Mortality (26 v 6, p = 0.0003), heart failure admissions (18 v 2, p = 0.0002), and admissions for other cardiac causes (44 v 13, p = 0.0001) were significantly higher in patients with NTproBNP above the study median (32.5 pmol/l). Conclusions: Measurement of NT-proBNP may be useful as a screening tool for systolic heart failure in the general population.

Copeptin—Marker of Acute Myocardial Infarction

Martin Möckel & Julia Searle
Curr Atheroscler Rep 2014; 16:421

The concentration of copeptin, the C-terminal part of pro-arginine vasopressin, has been shown to increase early after acute and severe events. Owing to complementary pathophysiology and kinetics, the unspecific marker copeptin, in combination with highly cardio-specific troponin, has been evaluated as an early-rule-out strategy for acute myocardial infarction in patients presenting with signs and symptoms of acute coronary syndrome. Overall, most studies have reported a negative predictive value between 97 and 100 % for the diagnosis of acute myocardial infarction in low- to intermediate-risk patients with suspected acute coronary syndrome. Additionally, a recent multicenter, randomized process study, where patients who tested negative for copeptin and troponin were discharged from the emergency department, showed that the safety of the new process was comparable to that of the current standard process. Further interventional trials and data from registries are needed to ensure the effectiveness and patient benefit of the new strategy.

The role of copeptin as a diagnostic and prognostic biomarker for risk stratification in the emergency department

Christian H Nickel1, Roland Bingisser and Nils G Morgenthaler
BMC Medicine 2012, 10:7

The hypothalamic-pituitary-adrenal axis is activated in response to stress. One of the activated hypothalamic hormones is arginine vasopressin, a hormone involved in hemodynamics and osmoregulation. Copeptin, the C-terminal part of the arginine vasopressin precursor peptide, is a sensitive and stable surrogate marker for arginine vasopressin release. Measurement of copeptin levels has been shown to be useful in a variety of clinical scenarios, particularly as a prognostic marker in patients with acute diseases such as lower respiratory tract infection, heart disease and stroke. The measurement of copeptin levels may provide crucial information for risk stratification in a variety of clinical situations. As such, the emergency department appears to be the ideal setting for its potential use. This review summarizes the recent progress towards determining the prognostic and diagnostic value of copeptin in the emergency department.

Variability of the Transferrin Receptor 2 Gene in AMD

Daniel Wysokinski, Janusz Blasiak, Mariola Dorecka, Marta Kowalska, et al.
Disease Markers 2014, Article ID 507356, 8 pages

Oxidative stress is a major factor in the pathogenesis of age-related macular degeneration (AMD). Iron may catalyze the Fenton reaction resulting in overproduction of reactive oxygen species. Transferrin receptor 2 plays a critical role in iron homeostasis and variability in its gene may influence oxidative stress and AMD occurrence. To verify this hypothesis we assessed the association between  polymorphisms of the TFR2 gene and AMD. A total of 493AMDpatients and 171matched controls were genotyped for the two polymorphisms of the TFR2 gene: c.1892C>T (rs2075674) and c.−258+123T>C (rs4434553). We also assessed the modulation of some AMD risk factors by these polymorphisms.The CC and TT genotypes of the c.1892C>T were associated with AMD occurrence but the latter only in obese patients. The other polymorphism was not associated with AMD occurrence, but the CC genotype was correlated with an increasing AMD frequency in subjects with BMI < 26. The TT genotype and the T allele of this polymorphism decreased AMD occurrence in subjects above 72 years, whereas the TC genotype and the C allele increased occurrence of AMD in this group.The c.1892C>T and c.−258+123T>C polymorphisms of the TRF2 gene may be associated with AMD occurrence, either directly or by modulation of risk factors.

Urinary N-Acetyl-beta-D-glucosaminidase as an Early Marker for Acute Kidney Injury in Full-Term Newborns with Neonatal Hyperbilirubinemia

Bangning Cheng, Y Jin, G Liu, Z Chen, H Dai, and M Liu
Disease Markers 2014, Article ID 315843, 6 pages

Purpose. To investigate renal function estimated by markers in full-term newborns with hyperbilirubinemia.
Methods. A total of 332 full-term newborns with hyperbilirubinemia and 60 healthy full-term newborns were enrolled. Total serum bilirubin, serum creatinine (Cr), serum blood urea nitrogen (BUN), serum cystatin C (Cys-C), urinary beta-2-microglobulin (𝛽2MG) index, and urinary N-acetyl-beta-D-glucosaminidase (NAG) index were measured before and after treatment. All newborns were divided into three groups according to total serum bilirubin levels: group 1 (221-256), group 2 (256-342), and group 3 (>342). Results. The control group and group 1 did not differ significantly in regard to serum Cr, serum BUN, serum Cys-C, urinary 𝛽2MG index, and urinary NAG index. Urinary NAG index in group 2 was significantly higher than that in control group (𝑃 < 0.001). Between control group and group 3, serum Cys-C, urinary 𝛽2MG index, and urinary NAG index differed significantly. The significant positive correlation between total serum bilirubin and urinary NAG index was found in newborns when total serum bilirubin level was more than 272 𝜇mol/L.
Conclusions. High unconjugated bilirubin could result in acute kidney injury in full-term newborns. Urinary NAG might be the suitable marker for predicting acute kidney injury in full-term newborns with hyperbilirubinemia.

Urinary C-peptide creatinine ratio detects absolute insulin deficiency in Type 2 diabetes.

S V Hope, A G Jones, E Goodchild, M Shepherd, R E J Besser, B Shields, T McDonald, B A Knight, A Hattersley

Department of Geriatrics, Royal Devon and Exeter NHS Foundation Trust; NIHR Exeter Clinical Research Facility, University of Exeter.

Diabetic Medicine (impact factor: 2.9). 05/2013;

Source: PubMed

ABSTRACT AIMS: To determine the prevalence and clinical characteristics of absolute insulin deficiency in long-standing Type 2 diabetes, using a strategy based on home urinary C-peptide creatinine ratio measurement.
METHODS: We assessed the urinary C-peptide creatinine ratios, from urine samples taken at home 2 h after the largest meal of the day, in 191 insulin-treated subjects with Type 2 diabetes (diagnosis age ≥45 years, no insulin in the first year). If the initial urinary C-peptide creatinine ratio was ≤0.2 nmol/mmol (representing absolute insulin deficiency), the assessment was repeated. A standardized mixed-meal tolerance test with 90-min stimulated serum C-peptide measurement was performed in nine subjects with a urinary C-peptide creatinine ratio ≤ 0.2 nmol/mmol (and in nine controls with a urinary C-peptide creatinine ratio >0.2 nmol/mmol) to confirm absolute insulin deficiency.
RESULTS: A total of 2.7% of participants had absolute insulin deficiency confirmed by a mixed-meal tolerance test. They were identified initially using urinary C-peptide creatinine ratio: 11/191 subjects (5.8%) had two consistent urinary C-peptide creatinine ratios ≤ 0.2 nmol/mmol; 9/11 subjects completed a mixed-meal tolerance test and had a median stimulated serum C-peptide of 0.18nmol/l. Five out of nine subjects had stimulated serum C-peptide <0.2 nmol/l and 9/9 subjects with urinary C-peptide creatinine ratio >0.2 had endogenous insulin secretion confirmed by the mixed-meal tolerance test. Compared with subjects with a urinary C-peptide creatinine ratio >0.2 nmol/mmol, those with confirmed absolute insulin deficiency had a shorter time to insulin treatment (median 2.5 vs. 6 years, P=0.005) and lower BMI (25.1 vs. 29.1kg/m(2) , P=0.04). Two out of five patients were glutamic acid decarboxylase autoantibody-positive.
CONCLUSIONS: Absolute insulin deficiency may occur in long-standing Type 2 diabetes, and cannot be reliably predicted by clinical features or autoantibodies. Its recognition should help guide treatment, education and management. The urinary C-peptide creatinine ratio is a practical non-invasive method to aid detection of absolute insulin deficiency, with a urinary C-peptide creatinine ratio > 0.2 nmol/mmol being a reliable indicator of retained endogenous insulin secretion.

Unlocking Biomarkers’ Full Potential

David Daniels, Ph.D.     genengnews  Feb 1, 2013 (Vol. 33, No. 3)

Biomarker research and development has evolved over the past years from looking for a single marker (e.g., PSA) linked to a disease state to looking for a panel of markers that can capture the heterogeneity inherent in both the disease and the impacted patient population.

That is one of the key messages to be delivered at GTC’s “Biomarkers Summit” next month. Across the board, resources are being focused on the delivery of more precise, quantifiable biomarkers with predictive value in therapeutic decisions and for the prognosis of illness.

“Our focus on biomarker development is the recognition that the new products need to provide cost savings for the already strapped healthcare systems rather than just be cost effective,” shares Paul Billings, M.D., Ph.D., CMO at Life Technologies.

“We have built a new medical sciences group to address the needs of the multiple delivery systems in the world—from the sophisticated medical clinics in the developed world to the nurse-run shanty clinic in the third world. Providing tools for equitable access to quality diagnosis, on assay platforms that can provide care for all patients, is our goal.”

Life Tech’s medical sciences division has been built by acquisition of Pinpoint Genomics, Navigenics, and Compendia, and collaborations with partners such as Ingenuity Systems and CollabRx. The division is focused on taking the tools that have been used in the life science laboratories and providing molecular diagnostic data to the clinic. The intent is to deliver data in a valuable format that can be used by the molecular pathologist or the treating physician.

The division is developing the Pervenio™ Lung RS assay, a 14-gene expression profile that serves as a risk stratifier that uses a weighted algorithm for the expressed biomarkers within the tumor biopsy, a first-of-its-kind prognostic test for lung cancer, the firm reports.

Initially, tests will be offered as a service through Life Tech’s CLIA laboratory. Then, from the performance lessons learned, Life Tech’s will develop a simpler assay platform, with FDA approval, that can be dispersed globally without reduction of the essential content in the biomarker panel. The focus is on the workflow—screening for known mutations using established easy-to-use assay platforms, like RT-PCR. Should the screen not produce useful results, clinicians can search for new mutations via discovery platforms like next-gen sequencing (NGS).

Sequenom’s LungCarta panel of 214 somatic mutations in 26 tumor suppressors and oncogenes covers highly mutated pathways in lung adenocarcinomas.

At Sequenom, the company provides both the tools (DNA mass spectrometry and reagents) for confirmatory biomarker development as well as serving on the front lines as a diagnostic service provider (CLIA lab). The beauty of DNA mass spec is that it can process multiplexed PCR samples (10–60 loci) in a method that is quantitative when used for profiling tumor biopsies that are either archival or fresh tissue.

Given a tumor sample with multiple somatic mutations, the instrument enables the determination of the homogeneity of the cells, in which case the mutations will have the same allele frequency. Accuracy, as measured by coefficient of variance, is less than 2%. Despite this level of sensitivity, the mass spec can only be used as a confirmatory tool looking for known mutations. Discovery is best done using DNA sequencing. DNA mass spec can also be used to study methylation in tumor samples.

“In the not-too-distant future, we will be looking for mutations in plasma samples rather than biopsies,” predicts Charles Cantor, Ph.D., CSO at Sequenom.

“The key is to look noninvasively for mutations within plasma samples such that we can potentially catch the disease state earlier, rather than after tumor formation. Regardless of the tumor type, this approach will enable us to monitor therapeutic response and metastatic potential noninvasively. DNA mass spec is an ultrasensitive detection product that can detect somatic mutations at levels of 1 per 1,000. This level of sensitivity is critical for the future of plasma screening. NGS technology is not that sensitive.”

Sequenom’s CLIA lab is using automated DNA mass spec to provide three different test protocols: (1) carrier screening for cystic fibrosis looking at more than 100 different mutations, (2) adult macular degeneration progression using an SNP test with 13 loci, and (3) a noninvasive test for Rh compatibility between a mother and her unborn fetus.

Scientists are using Illumina’s HiSeq system to discover molecular biomarkers that may provide opportunities for early detection of a range of diseases.

Sequenom has also set up an NGS facility within a CLIA lab in San Diego using Illumina’s HiSEQ platform. The NGS platform has been set up for noninvasive aneuploidy detection of maternal plasma (10 cc sample) looking at chromosomes 13, 18, and 21. The lab says it has analyzed more than 40,000 samples this year and is planning to increase that volume up to 100,000 samples per year. Most of these samples come from the U.S., but given the development of a new blood collection tube that allows for 72-hour ambient shipping, the lab is looking to increase the number of samples from outside the U.S.

Drug Development

During drug development, biomarkers function as pharmacodynamic markers to help assess the mechanism of action of a drug candidate, to define the downstream biological pathway, and to determine whether the drug is engaging the target with the anticipated biological effect. Later, biomarkers help determine whether a drug is effective using the tested regime (route of delivery, dosage level, and length of exposure time).

Following early development, the second stage is to use biomarkers to help segment patients for clinical trials. Part of the consideration here is how heterogeneous the disease is; are there homogeneous subsets of patients that will respond differentially to the drug based on different mechanisms of the disease?

“Biomarker research is focused on on- target effects,” says Nick Dracopoli, Ph.D., vp, head of oncology biomarkers at Janssen Research and Development, a J&J company.

“We look at indications and at patients with those indications that are most likely to respond to the drug candidates we’re developing. For oncology biomarkers, germ-line effects are weaker indicators than somatic changes in the tumor. As a consequence, SNP-based, genome-wide association studies are not very useful. It is better to focus on molecular changes within the tumor and define gene expression profiles and epigenetic modifications that correlate with the tumor phenotype. We are increasingly tracking patient immune response, particularly as more immuno-oncology products are moving into the drug development pipeline.”

The number of biomarkers being developed varies from project to project. But it is very clear that to be successful in the clinic, the biomarkers and the assays need to be of low complexity. Of the 10 to 12 companion diagnostics that have been approved by the FDA to date, all measure the status of the drug target (on-target markers). For example, EGFR measures the level of receptor expression; Braf and Kras markers measure the presence of the mutation and translocation in the ALK gene measures gene knockout.

It is important to realize that molecular profiles for first-in-class drugs are not optimal because they are based on only a few patients. Consequently they have weak predictive value overall.

“Aside from that rule of thumb, if you have a greater than 50% response rate for your drug, it is unlikely that you need a biomarker to predict response. Biomarker utility is best for drugs that would have a difficult road to approval, where it is critical to enrich for the subpopulation of responders. For example, Pfizer’s crizontinib was approved for non-small-cell lung patients but is only effective for 5% of all patients. If Pfizer was unable to demonstrate the relationship between activation of the ALK gene and disease, this inhibitor would not have been approved,” says Dr. Dracopoli.

“Drugs that are more broadly active can come to market without a companion diagnostic test. There is always a balance between the predictive values of the biomarker test and the response rates to treatment. That is, we should not treat if the chance of response is only 3–5%, rather than if it were 50% where the patient would want to take the chance if the drug were safe.”

An important take-home message is that mutations are not unique to an indication. So if you find a driver mutation in indications for which the drug has not been approved, you could discover new indications for the drug.

“At the end of the day, this is what cancer is—heterogeneous,” says Dr. Dracopoli. “We’d all love to treat one cancer with one drug and at one dose, but the story is more complex. The future of oncology is around understanding the molecular heterogeneity or underlying molecular pathology of the disease and the diversity of it, and then treat each patient accordingly.”

Clinical Considerations

“Given the complexity of biology,” says Achim Plum, Ph.D., principal consultant, Siemens, “whether is it cancer, metabolic disease, or any other disease state, we have been forced to move away from the idea that a single biomarker can capture the entire ‘story’ or mechanistic view of any disease. Hence newly developed biomarkers will be made up of a panel of markers that serve as a profile. In addition, with the sheer volume of DNA and protein analytics data, the clinic will need to employ software tools and algorithms to help the decision making.”

The task of getting broad profiling technologies that are analytical into a clinical setting and making them routine is difficult but not insurmontable. This will take a collaborative effort, something that Siemens among others are looking to develop. The key is to avoid technology hype and to establish good reliable software to process the data for decision making. “Data is not knowledge, and knowledge is not automatically decision making.”

As an academic, Daniel Chan, Ph.D., has a view of the whole value chain for biomarkers from discovery to development to use in the clinic. Dr. Chan holds the titles of professor in pathology, oncology, radiology, and urology, and is the director of the clinical chemistry division lab at Johns Hopkins Hospital.

Given his perspective from discovery to clinical use, Dr. Chan indicated that from the clinical point of view, “we need more markers.” He oversees the discovery of new biomarkers in his research lab, their validation in his translational research lab, and finally their utility in practice in his clinical chemistry lab. He is a strong advocate for collaboration of biomarker development from discovery to verification and validation to incorporation within the clinical practice.

Beyond the use of biomarkers for patient stratification and correlation between marker and therapeutic choice, as is the focus of the biopharma industry, for the clinic the use of biomarkers is for prevention and early detection. The earlier the detection, the better the outcome. That is, provide the “cure” before you need to initiate treatment.

To be successful in the future of biomarkers, we need to look beyond the biopharma focus and expand the horizon for early detection and monitor therapy later, says Dr. Chan. He describes a roadmap of developing bridges (to bridge the knowledge gaps), gates (decision gates for go/no go decisions as to whether a development path is viable), and partnerships (to collaborate with different points of view) for efficient new biomarker development.

According to Dr. Chan, we must define the intended use of the biomarker, which identifies the specific application and sets up the clinical study and study population to meet the clinical needs. We need to define specific assays to monitor biomarkers that will work within a clinical setting, not a research lab setting that uses disease models (tissue culture cells or small animals) and not real patient samples.

“The days when single markers are sufficient (PSA for prostate cancer or troponin for cardiovascular disease) are behind us. We need to develop a panel of markers or a profile pattern to address patient population heterogeneity and disease complexity that will guide our decision-making process,” remarks Dr. Chan. “Molecular biomarkers are giving way to protein biomarkers,” he adds.

Prevention and early detection will require the use of whole-body scans, so the sampling technology and analytical tools to be developed are critical to realize this goal. Assay ease of use, automation, and analytical performance that is suitable for the clinical lab are fundamental.

“An important future goal for biomarkers,” says Dr. Billings, “is to sample circulating tumor cells or circulating DNA in blood or plasma samples as a noninvasive measure of patient status. A decline in tumor biomarkers during chemotherapy, for example, could reflect the efficacy of the therapy. In contrast, an increase in tumor biomarkers, in a patient who had previously undergone surgery and therapy, might indicate disease recurrence, and is likely to do so before a tumor mass is detectable by imaging methods.”

STAT4 Gene Polymorphisms Are Associated with Susceptibility and ANA Status in Primary Biliary Cirrhosis

Satoru Joshita, T Umemura, M Nakamura, Y Katsuyama, S Shibata, et al.
Disease Markers  2014, Article ID 727393, 8 pages

Recent genome-wide association studies suggest that genetic factors contribute to primary biliary cirrhosis (PBC) susceptibility. Although several reports have demonstrated that the interleukin (IL) 12 signaling pathway is involved in PBC pathogenesis, its precise genetic factors have not been fully clarified. Here, we performed an association analysis between IL12A, IL12RB, and signal transducer and activator of transcription 4 (STAT4) genetic variations and susceptibility to PBC. Single nucleotide polymorphisms (SNPs) were genotyped in 395 PBC patients and 458 healthy subjects of Japanese ethnicity and evaluated for associations with PBC susceptibility, anti-nuclear antibody (ANA) status, and anti-mitochondrial antibody (AMA) status. We detected significant associations with PBC susceptibility for several STAT4 SNPs (rs10168266; p = 9.4 × 10−3, rs11889341; p = 1.2 × 10−3, rs7574865; p = 4.0 × 10−4, rs8179673; p = 2.0 × 10−4, and rs10181656; p = 4.2 × 10−5). Three risk alleles (rs7574865; p = 0.040, rs8179673; p = 0.032, and rs10181656; p = 0.031) were associated with ANA status, but not with AMA positivity. Our findings confirm that STAT4 is involved in PBC susceptibility and may play a role in ANA status in the Japanese population.

Serum Omentin-1 as a Disease Activity Marker for Crohn’s Disease

Yan Lu, Li Zhou, L Liu, Yan Feng, Li Lu, X Ren, X Dong, & W Sang
Disease Markers  2014, Article ID 162517, 5 pages

Background and Aim. It remains challenging to determine the inflammatory activity in Crohn’s disease (CD) for lack of specific laboratory markers. Recent studies suggest that serum omentin-1 is associated with inflammatory response. We aimed to assess the potential of serum omentin-1 as a marker of disease activity in CD patients.
Methods. Serum omentin-1 concentrations were determined by enzyme-linked immunosorbent assay (ELISA) in patients with CD (n = 240), functional gastrointestinal disorders (FGDs, n = 120), and healthy controls (HC, n = 60) and evaluated for correlation with disease activity. Expression of omentin-1 in colonic tissues from patients with CD was also analyzed by real-time PCR and Western blotting. Serum omentin-1 levels as an activity index were evaluated using a receiver operating characteristic (ROC) curve.
Results. Serum omentin-1 concentrations were significantly decreased in active CD patients compared with patients in remission, FGDs, and HC (all p < 0.001). Expression of omentin-1 was decreased at mRNA and protein levels in inflamed colonic tissues in active CD than that in noninflamed colonic tissues. Serum omentin-1 levels were negatively correlated with disease activity in CD, better than C-reactive protein (CRP).
Conclusion. Our results indicate that serum and colonic omentin-1 expressions are decreased in active CD patients. The correlation of serum omentin-1 with disease activity in CD is superior to that of CRP. Serum omentin-1 is a potential marker for CD disease activity.
Serum Levels of Resistin, Adiponectin, and Apelin in Gastroesophageal Cancer Patients

Dorota Diakowska, K Markocka-Mdczka, P Szelachowski, and K Grabowski
Disease Markers 2014, Article ID 619649, 8 pages

The aim of the study was the investigation of relationship between cachexia syndrome and serum resistin, adiponectin, and apelin in patients with gastroesophageal cancer (GEC).
Material and Methods. Adipocytokines concentrations were measured in sera of 85 GEC patients and 60 healthy controls. They were also evaluated in tumor tissue and appropriate normal mucosa of 38 operated cancer patients.
Results. Resistin and apelin concentrations were significantly higher in GEC patients than in the controls. The highest resistin levels were found in cachectic patients and in patients with distant metastasis. Serum adiponectin significantly decreased in GEC patients with regional and distant metastasis. Serum apelin was significantly higher in cachectic patients than in the controls. Apelin was positively correlated with hsCRP level. Resistin and apelin levels increased significantly in tumor tissues. Weak positive correlations between adipocytokines levels in serum and in tumor tissue were observed.
Conclusions. Resistin is associated with cachexia and metastasis processes of GEC. Reduction of serum adiponectin reflects adipose tissue wasting in relation to GEC progression. Correlation of apelin with hsCRP can reflect a presumable role of apelin in systemic inflammatory response in esophageal and gastric cancer.

Serum Level of HER-2 Extracellular Domain in Iranian Patients with Breast Cancer: A Follow-up Study

Mehrnoosh Doroudchi, Abdolrasoul Talei, Helmout Modjtahedi, et al.
IJI 2005; 2(4): 191-200

Background: A soluble form of HER-2/neu extracellular domain (sHER-2) is reported to be released in the sera of metastatic breast cancer patients.
Objective: To measure the level of sHER-2 in sera of 115 breast cancer patients. Methods: Serial samples of 27 patients with metastasis, 18 non-metastatic patients, 15 patients in stage 0/I and 14 patients with accompanying benign breast disease were also included in this study.
Results: No significant difference was observed between sHER-2 level in the pre-operative sera of breast cancer patients and that of healthy individuals. Only 8 out of 27 patients whom later developed metastasis showed elevated levels of sHER-2 in their first serum sample. However, a trend of increase in the level of sHER-2 was observed in 14 (51.8%) of 27 metastatic sera before clinical diagnosis of the metastasis. A significant association between sHER-2 positive status and vascular invasion of the tumor was observed (P = 0.02). In addition, significant correlation of sHER-2 level with CEA (highest r = 0.74) and CA 15.3 (highest r = 0.74) tumor marker levels in the serial sera were observed. The mean time from sHER-2 positivity to tumor metastasis was calculated to be 98 days (range = 29-174).  Conclusion: Our results indicate that a relatively high percentage of Iranian patients with breast cancer show an elevated level of sHER-2 in their sera before clinical diagnosis of the tumor metastasis. Therefore, measuring the level of this oncoprotein, not only helps physicians in monitoring the patients during HERCEPTIN therapy, but also can be helpful in choosing more aggressive treatments at the early satges of tumor metastasis.
B-type natriuretic peptide is a biomarker for pulmonary hypertension in preterm infants with bronchopulmonary dysplasia

Alain Cuna, Jegen Kandasamy, Naomi Fineberg, Brian Sims
Research and Reports in Neonatology 2013:3 33–36

Background: B-type natriuretic peptide (BNP) is a cardiac biomarker useful in screening for pulmonary hypertension (PH) in adults. It is possible that BNP may also be useful in detecting PH among preterm infants with bronchopulmonary dysplasia (BPD).
Objective: To determine the utility of BNP for identification of PH among preterm infants with BPD.
Methods: We retrospectively identified preterm infants with BPD who underwent screening echocardiography for suspected PH and had serum BNP levels measured within 10 days before or after echocardiography. Eligible infants were classified based on echocardiographic diagnosis of either PH or no PH. Median and interquartile ranges (IQR) of BNP values were compared, and area under the curve (AUC) of receiver operator characteristic (ROC) analysis was used to determine the optimum threshold value for detection of PH.
Results: Twenty-five preterm infants with BPD (mean gestational age 26.5 ± 1.7 weeks, mean birth weight 747 ± 248 g) were identified. The median difference in days between echocardiography and BNP measurement was 1 day (IQR 0–3, range 0–10 days). Based on echocardiography, 16 were diagnosed with PH and nine without PH. No significant difference in terms of gestational age, birth weight, sex, race, or respiratory support was found between the two groups. Median (IQR) BNP values of those with PH were higher than those without PH (413 [212–1178] pg/mL versus 55 [21–84] pg/mL, P , 0.001). AUC of ROC analysis showed that a BNP value of 117 pg/mL had 93.8% sensitivity and 100% specificity for detecting PH.
Conclusion: BNP estimation may be useful for screening of PH in infants with BPD.

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Writer and Curator: Larry H. Bernstein, MD, FCAP


Implementation and utilization of genetic testing in personalized medicine

NS Abul-Husn, AO Obeng, SC Sanderson, O Gottesman, S A Scott
Pharmacogenomics and Personalized Medicine 2014:7 227–240

Clinical genetic testing began over 30 years ago with the availability of mutation detection for sickle cell disease diagnosis. Since then, the field has dramatically transformed to include gene sequencing, high-throughput targeted genotyping, prenatal mutation detection, preimplantation genetic diagnosis, population-based carrier screening, and now genome-wide analyses using microarrays and next-generation sequencing. Despite these significant advances in molecular technologies and testing capabilities, clinical genetics laboratories historically have been centered on mutation detection for Mendelian disorders. However, the ongoing identification of deoxyribonucleic acid (DNA) sequence variants associated with common diseases prompted the availability of testing for personal disease risk estimation, and created commercial opportunities for direct-to-consumer genetic testing companies that assay these variants. This germline genetic risk, in conjunction with other clinical, family, and demographic variables, are the key components of the personalized medicine paradigm, which aims to apply personal genomic and other relevant data into a patient’s clinical assessment to more precisely guide medical management. However, genetic testing for disease risk estimation is an ongoing topic of debate, largely due to inconsistencies in the results, concerns over clinical validity and utility, and the variable mode of delivery when returning genetic results to patients in the absence of traditional counseling. A related class of genetic testing with analogous issues of clinical utility and acceptance is pharmacogenetic testing, which interrogates sequence variants implicated in interindividual drug response variability. Although clinical pharmacogenetic testing has not previously been widely adopted, advances in rapid turnaround time genetic testing technology and the recent implementation of preemptive genotyping programs at selected medical centers suggest that personalized medicine through pharmacogenetics is now a reality. This review aims to summarize the current state of implementing genetic testing for personalized medicine, with an emphasis on clinical pharmacogenetic testing.

Pharmacogenomic knowledge gaps and educational resource needs among physicians in selected specialties

Katherine A Johansen Taber, Barry D Dickinson
Pharmacogenomics and Personalized Medicine 2014:7 145–162

Background: The use of pharmacogenomic testing in the clinical setting has the potential to improve the safety and effectiveness of drug therapy, yet studies have revealed that physicians lack knowledge about the topic of pharmacogenomics, and are not prepared to implement it in the clinical setting. This study further explores the pharmacogenomic knowledge deficit and educational resource needs among physicians.
Materials and methods: Surveys of primary care physicians, cardiologists, and psychiatrists were conducted.
Results: Few physicians reported familiarity with the topic of pharmacogenomics, but more reported confidence in their knowledge about the influence of genetics on drug therapy. Only a small minority had undergone formal training in pharmacogenomics, and a majority reported being unsure what type of pharmacogenomic tests were appropriate to order for the clinical situation. Respondents indicated that an ideal pharmacogenomic educational resource should be electronic and include such components as how to interpret pharmacogenomic test results, recommendations for prescribing, population subgroups most likely to be affected, and contact information for laboratories offering pharmacogenomic testing.
Conclusion: Physicians continue to demonstrate pharmacogenomic knowledge gaps, and are unsure about how to use pharmacogenomic testing in clinical practice. Educational resources that are clinically oriented and easily accessible are preferred by physicians, and may best support appropriate clinical implementation of pharmacogenomics.

Developing genomic knowledge bases and databases to support clinical management: current perspectives

Vojtech Huser, Murat Sincan, James J Cimino
Pharmacogenomics and Personalized Medicine 2014:7 275–283

Personalized medicine, the ability to tailor diagnostic and treatment decisions for individual patients, is seen as the evolution of modern medicine. We characterize here the informatics resources available today or envisioned in the near future that can support clinical interpretation of genomic test results. We assume a clinical sequencing scenario (germline whole-exome sequencing) in which a clinical specialist, such as an endocrinologist, needs to tailor patient management decisions within his or her specialty (targeted findings) but relies on a genetic counselor to interpret off-target incidental findings. We characterize the genomic input data and list various types of knowledge bases that provide genomic knowledge for generating clinical decision support. We highlight the need for patient-level databases with detailed lifelong phenotype content in addition to genotype data and provide a list of recommendations for personalized medicine knowledge bases and databases. We conclude that no single knowledge base can currently support all aspects of personalized recommendations and that consolidation of several current resources into larger, more dynamic and collaborative knowledge bases may offer a future path forward.


Tumor Heterogeneity: Mechanisms and Bases for a Reliable Application of Molecular Marker Design

Salvador J. Diaz-Cano
Int. J. Mol. Sci. 2012, 13, 1951-2011;

Tumor heterogeneity is a confusing finding in the assessment of neoplasms, potentially resulting in inaccurate diagnostic, prognostic and predictive tests. This tumor heterogeneity is not always a random and unpredictable phenomenon, whose knowledge helps designing better tests. The biologic reasons for this intratumoral heterogeneity would then be important to understand both the natural history of neoplasms and the selection of test samples for reliable analysis. The main factors contributing to intratumoral heterogeneity inducing gene abnormalities or modifying its expression include: the gradient ischemic level within neoplasms, the action of tumor microenvironment (bidirectional interaction between tumor cells and stroma), mechanisms of intercellular transference of genetic information (exosomes), and differential mechanisms of sequence-independent modifications of genetic material and proteins. The intratumoral heterogeneity is at the origin of tumor progression and it is also the byproduct of the selection process during progression. Any analysis of heterogeneity mechanisms must be integrated within the process of segregation of genetic changes in tumor cells during the clonal expansion and progression of neoplasms. The evaluation of these mechanisms must also consider the redundancy and pleiotropism of molecular pathways, for which appropriate surrogate markers would support the presence or not of heterogeneous genetics and the main mechanisms responsible. This knowledge would constitute a solid scientific background for future therapeutic planning.

Systematic evaluation of connectivity map for disease indications

Jie Cheng, Lun Yang, Vinod Kumar and Pankaj Agarwal
Genome Medicine 2014, 6:95

Background: Connectivity map data and associated methodologies have become a valuable tool in understanding drug mechanism of action (MOA) and discovering new indications for drugs. One of the key ideas of connectivity map (CMAP) is to measure the connectivity between disease gene expression signatures and compound-induced gene expression profiles. Despite multiple impressive anecdotal validations, only a few systematic evaluations have assessed the accuracy of this aspect of CMAP, and most of these utilize drug-to-drug matching to transfer indications across the two drugs.
Methods: To assess CMAP methodologies in a more direct setting, namely the power of classifying known drug-disease relationships, we evaluated three CMAP-based methods on their prediction performance against a curated dataset of 890 true drug-indication pairs. The disease signatures were generated using Gene Logic BioExpress system and the compound profiles were derived from the Connectivity Map database (CMAP, build 02,
Results: The similarity scoring algorithm called eXtreme Sum (XSum) better than the standard Kolmogorov-Smirnov (KS) statistic in terms of the area under curve and can achieve a four-fold enrichment at 0.01, false positive rate level, with AUC = 2.2E-4, P value = 0.0035.
Conclusion: Connectivity map can significantly enrich true positive drug-indication pairs given an effective matching algorithm.

Pharmacogenetics of Statin-Induced Myopathy: A Focused Review of the Clinical Translation of Pharmacokinetic Genetic Variants

Jasmine A Talameh and Joseph P Kitzmiller
J Pharmacogenomics Pharmacoproteomics 2014, 5:2

Statins are the most commonly prescribed drugs in the United States and are extremely effective in reducing major cardiovascular events in the millions of Americans with hyperlipidemia. However, many patients (up to 25%) cannot tolerate or discontinue statin therapy due to statin-induced myopathy (SIM). Patients will continue to experience SIM at unacceptably high rates or experience unnecessary cardiovascular events (as a result of discontinuing or decreasing their statin therapy) until strategies for predicting or mitigating SIM are identified. A promising strategy for predicting or mitigating SIM is pharmacogenetic testing particularly of pharmacokinetic genetic variants as SIM is  related to statin exposure. Data is emerging on the association between pharmacokinetic genetic variants and SIM.
A current, critical evaluation of the literature on pharmacokinetic genetic variants and SIM for potential translation to clinical practice is lacking. This review focuses specifically on pharmacokinetic genetic variants and their association with SIM clinical outcomes. We also discuss future directions, specific to the research on pharmacokinetic genetic variants, which could speed the translation into clinical practice. For simvastatin, we did not find sufficient evidence to support the clinical translation of pharmacokinetic genetic variants other than SLCO1B1. However, SLCO1B1 may also be clinically relevant for pravastatin- and pitavastatin-induced myopathy, but additional studies assessing SIM clinical outcome are needed. CYP2D6*4 may be clinically relevant for atorvastatin-induced myopathy, but mechanistic studies are needed. Future research efforts need to incorporate statin-specific analyses, multi-variant analyses, and a standard definition of SIM. As the use of statins is extremely common and SIM continues to occur in a significant number of patients, future research investments in pharmacokinetic genetic variants have the potential to make a profound impact on public health.

Benefits of Pharmacogenetics in the Management of Hypertension

Clara Torrellas, Juan Carlos Carril and Ramón Cacabelos
J Pharmacogenomics Pharmacoproteomics 2014, 5:2

Introduction: Hypertension, suffered by 35% of the population, stands out as the main risk factor for cardiovascular disorders with the highest death rate worldwide. Only a small number of patients with hypertension gets efficient control over blood pressure (BP) with appropriate drug therapy.  harmacogenetics, as a tool to identify antihypertensive therapeutic response-associated polymorphisms, could help to reduce this problem.
Objectives: We present here an epidemiological study of the prevalence of hypertension and its pharmacological treatment to demonstrate the error rate that physicians can commit when the patient´s pharmacogenetic profile is unknown.
Method: The sample consisted of 1115 individuals of which 332 met criteria for hypertension. We recorded each patient´s drug prescription prior to their visit to EuroEspes Biomedical Research Center, and analyzed their pharmacogenetic profile.
Results: About 30% of patients were hypertensive, of whom only 40.4% were receiving an active ingredient for hypertension control. Among them, CYP3A4/5 and CYP2C9 were the major metabolizing enzymes. Antagonists of angiotensin II receptors, followed by calcium-blocking agents and beta-adrenergic antagonists were the most commonly-prescribed drug categories. However, 61% of hypertensive patients were not taking suitable antihypertensive agents for their metabolism according to their genetic idiosyncrasy. Furthermore, the highest error rate was determined for CYP2C9.
Conclusion: The introduction of changes in the management of hypertension in the Spanish population could be useful to promote the prevention and treatment of high blood pressure in a more efficient way. The integration of pharmacogenetic testing into routine clinical procedures could optimize the therapeutic response, guiding the physician in the choice of the correct antihypertensive drug and the correct dose. The control of BP arises as an area of particular interest in assessing the validity and utility of pharmacogenetic testing/intervention.

Pharmacogenomics Study of Clopidogrel by RFLP based Genotyping of CYP2C19 in Cardiovascular Disease Patients in North-East Population of India

Prasanthi SV, Vinayak S Jamdade, Nityanand B Bolshette, Ranadeep Gogoi and Mangala Lahkar
J Pharmacogenomics Pharmacoproteomics 2014, 5:3

Introduction and Objective: Pharmacogenetics is a genetically determined variability in drug responses. The genes and their allelic variants which affect our response to drugs are the main routes in development of pharmacogenetics. Clopidogrel is an antiplatelet drug, used against athero-thrombotic events in cardiovascular patients. The objective of our study was to identify the CYP2C19 Single Nucleotide Polymorphisms, responsible for altering the metabolism of clopidogrel, at gene level. And to document the prevalence of CYP2C19 gene mutations in clopidogrel treated cardiovascular disease patients in Assam population, Guwahati Medical College & Hospital, in North- East India.
Patients and Methods: We have studied 60 patients who received clopidogrel from Gauhati medical college and hospital Assam. Genomic DNA was extracted by using Hipura blood genomic DNA extracting mini preparation kit by following the manufacturer’s instructions.RFLP analysis was done by DNA amplification which was carried out by using set of primers and resulting ampicons of CYP2C19*2;CYP2C19*3 and CYP2C19*17 were subjected for Restriction digestion with SmaI, BamHI and Lwe0I respectively.
Results: We found that CYP2C19*2 had allelic frequency of ~40% in Gauhati Medical College and Hospital, Assam, North East India. None of the samples were mutated with CYP2C19*3 andCYP2C19*17 allele. Other CYP2C19 variant alleles with reduced or absent enzymatic activity have been identified. Conclusion: We found that loss of functional allele CYP2C19*2 had higher carriage frequency; whereas, CYP2C19*3 and *17 alleles were not found in cardiovascular patients who were taking clopidogrel. Personalized therapy targeting patients who carry these genetic variants might help to improve the clinical outcome.

Role of cytochrome P450 genotype in the steps toward personalized drug therapy

Larisa H Cavallari, Hyunyoung Jeong, Adam Bress
Pharmacogenomics and Personalized Medicine 2011:4 123–136

Genetic polymorphism for cytochrome 450 (P450) enzymes leads to interindividual variability in the plasma concentrations of many drugs. In some cases, P450 genotype results in decreased enzyme activity and an increased risk for adverse drug effects. For example, individuals with the CYP2D6 loss-of-function genotype are at increased risk for ventricular arrhythmia if treated with usual does of thioridazine. In other cases, P450 genotype may influence the dose of a drug required to achieve a desired effect. This is the case with warfarin, with lower doses often necessary in carriers of a variant CYP2C9*2 or *3 allele to avoid supratherapeutic anticoagulation. When a prodrug, such as clopidogrel or codeine, must undergo hepatic biotransformation to its active form, a loss-of-function P450 genotype leads to reduced concentrations of the active drug and decreased drug efficacy. In contrast, patients with multiple CYP2D6 gene copies are at risk for opioid-related toxicity if treated with usual doses of codeine-containing analgesics. At least 25 drugs contain information in their US Food and Drug Administration-approved labeling regarding P450 genotype. The CYP2C9, CYP2C19, and CYP2D6 genes are the P450 genes most often cited. To date, integration of P450 genetic information into clinical decision making is limited. However, some institutions are beginning to embrace routine P450 genotyping to assist in the treatment of their patients. Genotyping for P450 variants may carry less risk for discrimination compared with genotyping for disease-associated variants. As such, P450 genotyping is likely to lead the way in the clinical implementation of pharmacogenomics. This review discusses variability in the CYP2C9, CYP2C19, and CYP2D6 genes and the implications of this for drug efficacy and safety.

Asthma pharmacogenetics and the development of genetic profiles for personalized medicine

Victor E Ortega, Deborah A Meyers, Eugene R Bleecker
Pharmacogenomics and Personalized Medicine 2015:8 9–22

Human genetics research will be critical to the development of genetic profiles for personalized or precision medicine in asthma. Genetic profiles will consist of gene variants that predict individual disease susceptibility and risk for progression, predict which pharmacologic therapies will result in a maximal therapeutic benefit, and predict whether a therapy will result in an adverse response and should be avoided in a given individual. Pharmacogenetic studies of the glucocorticoid, leukotriene, and β2-adrenergic receptor pathways have focused on candidate genes within these pathways and, in addition to a small number of genome-wide association studies, have identified genetic loci associated with therapeutic responsiveness. This review summarizes these pharmacogenetic discoveries and the future of genetic profiles for personalized medicine in asthma. The benefit of a personalized, tailored approach to health care delivery is needed in the development of expensive biologic drugs directed at a specific biologic pathway. Prior pharmacogenetic discoveries, in combination with additional variants identified in future studies, will form the basis for future genetic profiles for personalized tailored approaches to maximize therapeutic benefit for an individual asthmatic while minimizing the risk for adverse events.

Clinical application of high throughput molecular screening techniques for pharmacogenomics

Arun P Wiita, Iris Schrijver
Pharmacogenomics and Personalized Medicine 2011:4 109–121

Genetic analysis is one of the fastest-growing areas of clinical diagnostics. Fortunately, as our knowledge of clinically relevant genetic variants rapidly expands, so does our ability to detect these variants in patient samples. Increasing demand for genetic information may necessitate the use of high throughput diagnostic methods as part of clinically validated testing. Here we provide a general overview of our current and near-future abilities to perform large-scale genetic testing in the clinical laboratory. First we review in detail molecular methods used for high throughput mutation detection, including techniques able to monitor thousands of genetic variants for a single patient or to genotype a single genetic variant for thousands of patients simultaneously. These methods are analyzed in the context of pharmacogenomic testing in the clinical laboratories, with a focus on tests that are currently validated as well as those that hold strong promise for widespread clinical application in the near future. We further discuss the unique economic and clinical challenges posed by pharmacogenomic markers. Our ability to detect genetic variants frequently outstrips our ability to accurately interpret them in a clinical context, carrying implications both for test development and introduction into patient management algorithms. These complexities must be taken into account prior to the introduction of any pharmacogenomic biomarker into routine clinical testing.

Clinical implementation of RNA signatures for pharmacogenomic decision-making

Weihua Tang, Zhiyuan Hu, Hind Muallem, Margaret L Gulley
Pharmacogenomics and Personalized Medicine 2011:4 95–107

RNA profiling is increasingly used to predict drug response, dose, or toxicity based on analysis of drug pharmacokinetic or pharmacodynamic pathways. Before implementing multiplexed RNA arrays in clinical practice, validation studies are carried out to demonstrate sufficient evidence of analytic and clinical performance, and to establish an assay protocol with quality assurance measures. Pathologists assure quality by selecting input tissue and by interpreting results in the context of the input tissue as well as the technologies that were used and the clinical setting in which the test was ordered. A strength of RNA profiling is the array-based measurement of tens to thousands of RNAs at once, including redundant tests for critical analytes or pathways to promote confidence in test results. Instrument and reagent manufacturers are crucial for supplying reliable components of the test system. Strategies for quality assurance include careful attention to RNA preservation and quality checks at pertinent steps in the assay protocol, beginning with specimen collection and proceeding through the variousphases of transport, processing, storage, analysis, interpretation, and reporting. Specimen quality is checked by probing housekeeping transcripts, while spiked and exogenous controls serve as a check on analytic performance of the test system. Software is required to manipulate abundant array data and present it for interpretation by a laboratory physician who reports results in a manner facilitating therapeutic decision-making. Maintenance of the assay requires periodic documentation of personnel competency and laboratory proficiency. These strategies are shepherding genomic arrays into clinical settings to provide added value to patients and to the larger health care system.

Dysregulation of the homeobox transcription factor gene HOXB13: role in prostate cancer

Brennan Decker, Elaine A Ostrander
Pharmacogenomics and Personalized Medicine 2014:7 193–201

Prostate cancer (PC) is the most common noncutaneous cancer in men, and epidemiological studies suggest that about 40% of PC risk is heritable. Linkage analyses in hereditary PC families have identified multiple putative loci. However, until recently, identification of specific risk alleles has proven elusive. Cooney et al used linkage mapping and segregation analysis to identify a putative risk locus on chromosome 17q21-22. In search of causative variant(s) in genes from the candidate region, a novel, potentially deleterious G84E substitution in homeobox transcription factor gene HOXB13 was observed in multiple hereditary PC families. In follow-up testing, the G84E allele was enriched in cases, especially those with an early diagnosis or positive family history of disease. This finding was replicated by others, confirming HOXB13 as a PC risk gene. The HOXB13 protein plays diverse biological roles in embryonic development and terminally differentiated tissue. In tumor cell lines, HOXB13 participates in a number of biological functions, including coactivation and localization of the androgen receptor and FOXA1. However, no consensus role has emerged and many questions remain. All HOXB13 variants with a proposed role in PC risk are predicted to damage the protein and lie in domains that are highly conserved across species. The G84E variant has the strongest epidemiological support and lies in a highly conserved MEIS protein-binding domain, which binds cofactors required for activation. On the basis of epidemiological and biological data, the G84E variant likely modulates the interaction between the HOXB13 protein and the androgen receptor, as well as affecting FOXA1-mediated transcriptional programming. However, further studies of the mutated protein are required to clarify the mechanisms by which this translates into PC risk.

Patient selection and targeted treatment in the management of platinum-resistant ovarian cancer

Christopher P Leamon, Chandra D Lovejoy, Binh Nguyen
Pharmacogenomics and Personalized Medicine 2013:6 113–125

Ovarian cancer (OC) has the highest mortality rate of any gynecologic cancer, and patients generally have a poor prognosis due to high chemotherapy resistance and late stage disease diagnosis. Platinum-resistant OC can be treated with cytotoxic chemotherapy such as paclitaxel, topotecan, pegylated liposomal doxorubicin, and gemcitabine, but many patients eventually relapse upon treatment. Fortunately, there are currently a number of targeted therapies in development for these patients who have shown promising results in recent clinical trials. These treatments often target the vascular endothelial growth factor pathway (eg, bevacizumab and aflibercept), DNA repair mechanisms (eg, iniparib and olaparib), or they are directed against folate related pathways (eg, pemetrexed, farletuzumab, and vintafolide). As many targeted therapies are only effective in a subset of patients, there is an increasing need for the identification of response predictive biomarkers. Selecting the right patients through biomarker screening will help tailor therapy to patients and decrease superfluous treatment to those who are biomarker negative; this approach should lead to improved clinical results and decreased toxicities. In this review the current targeted therapies used for treating platinum-resistant OC are discussed. Furthermore, use of prognostic and response predictive biomarkers to define OC patient populations that may benefit from specific targeted therapies is also highlighted.

Pharmacogenetics in breast cancer: steps toward personalized medicine in breast cancer management

Sarah Rofaiel, Esther N Muo1, Shaker A Mousa
Pharmacogenomics and Personalized Medicine 2010:3 129–143

There is wide individual variability in the pharmacokinetics, pharmacodynamics, and tolerance to anticancer drugs within the same ethnic group and even greater variability among different ethnicities. Pharmacogenomics (PG) has the potential to provide personalized therapy based on individual genetic variability in an effort to maximize efficacy and reduce adverse effects. The benefits of PG include improved therapeutic index, improved dose regimen, and selection of optimal types of drug for an individual or set of individuals. Advanced or metastatic breast cancer is typically treated with single or multiple combinations of chemotherapy regimens including anthracyclines, taxanes, antimetabolites, alkylating agents, platinum drugs, vinca alkaloids, and others. In this review, the PG of breast cancer therapeutics, including tamoxifen, which is the most widely used therapeutic for the treatment of hormone-dependent breast cancer, is reviewed. The pharmacological activity of tamoxifen depends on its conversion by cytochrome P450 2D6 (CYP2D6) to its abundant active metabolite, endoxifen. Patients with reduced CYP2D6 activity, as a result of either their genotype or induction by the coadministration of other drugs that inhibit CYP2D6 function, produce little endoxifen and hence derive limited therapeutic benefit from tamoxifen; the same can be said about the different classes of therapeutics in breast cancer. PG studies of breast cancer therapeutics should provide patients with breast cancer with optimal and personalized therapy

Novel treatment strategies in triple-negative breast cancer: specific role of poly(adenosine diphosphate-ribose) polymerase inhibition

M William Audeh
Pharmacogenomics and Personalized Medicine 2014:7 307–316

Inhibitors of the poly(adenosine triphosphate-ribose) polymerase (PARP)-1 enzyme induce synthetic lethality in cancers with ineffective DNA (DNA) repair or homologous repair deficiency, and have shown promising clinical activity in cancers deficient in DNA repair due to germ-line mutation in BRCA1 and BRCA2. The majority of breast cancers arising in carriers of BRCA1 germ-line mutations, as well as half of those in BRCA2 carriers, are classified as triple-negative breast cancer (TNBC). TNBC is a biologically heterogeneous group of breast cancers characterized by the lack of immunohistochemical expression of the ER, PR, or HER2 proteins, and for which the current standard of care in systemic therapy is cytotoxic chemotherapy. Many “sporadic” cases of TNBC appear to have indicators of DNA repair dysfunction similar to those in BRCA-mutation carriers, suggesting the possible utility of PARP inhibitors in a subset of TNBC. Significant genetic heterogeneity has been observed within the TNBC cohort, creating challenges for interpretation of prior clinical trial data, and for the design of future clinical trials. Several PARP inhibitors are currently in clinical development in BRCA-mutated breast cancer. The use of PARP inhibitors in TNBC without BRCA mutation will require biomarkers that identify cancers with homologous repair deficiency in order to select patients likely to respond. Beyond mutations in the BRCA genes, dysfunction in other genes that interact with the homologous repair pathway may offer opportunities to induce synthetic lethality when combined with PARP inhibition.

Clinical potential of novel therapeutic targets in breast cancer: CDK4/6, Src, JAK/STAT, PARP, HDAC, and PI3K/AKT/mTOR pathways

Sarah R Hosford, Todd W Miller
Pharmacogenomics and Personalized Medicine 2014:7 203–215

Breast cancers expressing estrogen receptor α, progesterone receptor, or the human epidermal growth factor receptor 2 (HER2) proto-oncogene account for approximately 90% of cases, and treatment with antiestrogens and HER2-targeted agents has resulted in drastically improved survival in many of these patients. However, de novo or acquired resistance to antiestrogen and HER2-targeted therapies is common, and many tumors will recur or progress despite these treatments. Additionally, the remaining 10% of breast tumors are negative for estrogen receptor α, progesterone receptor, and HER2 (“triple-negative”), and a clinically proven tumor-specific drug target for this group has not yet been identified. Therefore, the identification of new therapeutic targets in breast cancer is of vital clinical importance. Preclinical studies elucidating the mechanisms driving resistance to standard therapies have identified promising targets including cyclin-dependent kinase 4/6, phosphoinositide 3-kinase, poly adenosine diphosphate–ribose polymerase, Src, and histone deacetylase. Herein, we discuss the clinical potential and status of new therapeutic targets in breast cancer.

Overview of diagnostic/targeted treatment combinations in personalized medicine for breast cancer patients

Anna Tessari, Dario Palmieri, Serena Di Cosimo
Pharmacogenomics and Personalized Medicine 2014:7 1–19

Breast cancer includes a body of molecularly distinct subgroups, characterized by different presentation, prognosis, and sensitivity to treatments. Significant advances in our understanding of the complex architecture of this pathology have been achieved in the last few decades, thanks to new biotechnologies that have recently come into the research field and the clinical practice, giving oncologists new instruments that are based on biomarkers and allowing them to set up a personalized approach for each individual patient. Here we review the main treatments available or in preclinical development, the biomolecular diagnostic and prognostic approaches that changed our perspective about breast cancer, giving an overview of targeted therapies that represent the current standard of care for these patients. Finally, we report some examples of how new technologies in clinical practice can set in motion the development of new drugs.

Human ABC transporter ABCG2/BCRP expression in chemoresistance: basic and clinical perspectives for molecular cancer therapeutics

Kohji Noguchi, Kazuhiro Katayama, Yoshikazu Sugimoto
Pharmacogenomics and Personalized Medicine 2014:7 53–64

Adenosine triphosphate (ATP)-binding cassette (ABC) transporter proteins, such as ABCB1/P-glycoprotein (P-gp) and ABCG2/breast cancer resistance protein (BCRP), transport various structurally unrelated compounds out of cells. ABCG2/BCRP is referred to as a “half-type” ABC transporter, functioning as a homodimer, and transports anticancer agents such as irinotecan, 7-ethyl-10-hydroxycamptothecin (SN-38), gefitinib, imatinib, methotrexate, and mitoxantrone from cells. The expression of ABCG2/BCRP can confer a multidrug-resistant phenotype on cancer cells and affect drug absorption, distribution, metabolism, and excretion in normal tissues, thus modulating the in vivo efficacy of chemotherapeutic agents. Clarification of the substrate preferences and structural relationships of ABCG2/BCRP is essential for our understanding of the molecular mechanisms underlying its effects in vivo during chemotherapy. Its single-nucleotide polymorphisms are also involved in determining the efficacy of chemotherapeutics, and those that reduce the functional activity of ABCG2/BCRP might be associated with unexpected adverse effects from normal doses of anticancer drugs that are ABCG2/BCRP substrates. Importantly, many recently developed molecular-targeted cancer drugs, such as the tyrosine kinase inhisbitors, imatinib mesylate, gefitinib, and others, can also interact with ABCG2/BCRP. Both functional single-nucleotide polymorphisms and inhibitory agents of ABCG2/BCRP modulate the in vivo pharmacokinetics and pharmacodynamics of these molecular cancer treatments, so the pharmacogenetics of ABCG2/BCRP is an important consideration in the application of molecular-targeted chemotherapies.

Bosutinib: a SRC–ABL tyrosine kinase inhibitor for treatment of chronic myeloid leukemia

Fuad El Rassi, Hanna Jean Khoury
Pharmacogenomics and Personalized Medicine 2013:6 57–62

Bosutinib is one of five tyrosine kinase inhibitors commercially available in the United States for the treatment of chronic myeloid leukemia. This review of bosutinib summarizes the mode of action, pharmacokinetics, efficacy and safety data, as well as the patient-focused perspective through quality-of-life data. Bosutinib has shown considerable and sustained efficacy in chronic myeloid leukemia, especially in the chronic phase, with resistance or intolerance to prior tyrosine kinase inhibitors. Bosutinib has distinct but manageable adverse events. In the absence of T315I and V299L mutations, there are no absolute contraindications for the use of bosutinib in this patient population.

Toward precision medicine with next-generation EGFR inhibitors in non-small-cell lung cancer
Timothy A Yap, Sanjay Popat
Pharmacogenomics and Personalized Medicine 2014:7 285–295

The use of genomics to discover novel targets and biomarkers has placed the field of oncology at the forefront of precision medicine. First-generation epidermal growth factor receptor (EGFR) inhibitors have transformed the therapeutic landscape of EGFR mutant non-small-cell lung carcinoma through the genetic stratification of tumors from patients with this disease. Somatic EGFR mutations in lung adenocarcinoma are now well established as predictive biomarkers of response and resistance to small-molecule EGFR inhibitors. Despite early patient benefit, primary resistance and subsequent tumor progression to first-generation EGFR inhibitors are seen in 10%–30% of patients with EGFR mutant non-small-cell lung carcinoma. Acquired drug resistance is also inevitable, with patients developing disease progression after only 10–13 months of antitumor therapy. This review details strategies pursued in circumventing T790M-mediated drug resistance to EGFR inhibitors, which is the most common mechanism of acquired resistance, and focuses on the clinical development of second-generation EGFR inhibitors, exemplified by afatinib (BIBW2992). We discuss the rationale, mechanism of action, clinical efficacy, and toxicity profile of afatinib, including the LUX-Lung studies. We also discuss the emergence of third-generation irreversible mutant-selective inhibitors of EGFR and envision the future management of EGFR mutant lung adenocarcinoma.

ALK-driven tumors and targeted therapy: focus on crizotinib

Carlos Murga-Zamalloa, Megan S Lim
Pharmacogenomics and Personalized Medicine 2014:7 87–94

Receptor tyrosine kinases have emerged as promising therapeutic targets for a diverse set of tumors. Overactivation of the tyrosine kinase anaplastic lymphoma kinase (ALK) has been reported in several types of malignancies such as anaplastic large cell lymphoma, inflammatory myofibroblastic tumor, neuroblastoma, and non-small-cell lung carcinoma. Further characterization of the molecular role of ALK has revealed an oncogenic signaling signature that results in tumor dependence on ALK. ALK-positive tumors display a different behavior than their ALK-negative counterparts; however, the specific role of ALK in some of these tumors remains to be elucidated. Although more studies are required to establish selective targeting of ALK as a definitive therapeutic option, initial trials have shown extraordinary results in the majority of cases.

Non-small-cell lung cancer: molecular targeted therapy and personalized medicine – drug resistance, mechanisms, and strategies

Marybeth Sechler, AD Cizmic, S Avasarala, M Van Scoyk, C Brzezinski, et al.
Pharmacogenomics and Personalized Medicine 2013:6 25–36

Targeted therapies for cancer bring the hope of specific treatment, providing high efficacy and in some cases lower toxicity than conventional treatment. Although targeted therapeutics have helped immensely in the treatment of several cancers, like chronic myelogenous leukemia, colon cancer, and breast cancer, the benefit of these agents in the treatment of lung cancer remains limited, in part due to the development of drug resistance. In this review, we discuss the mechanisms of drug resistance and the current strategies used to treat lung cancer. A better understanding of these drug-resistance mechanisms could potentially benefit from the development of a more robust personalized medicine approach for the treatment of lung cancer.

ERCC1 and XRCC1 as biomarkers for lung and head and neck cancer

Alec Vaezi, Chelsea H Feldman, Laura J Niedernhofer
Pharmacogenomics and Personalized Medicine 2011:4 47–63

Advanced stage non-small cell lung cancer and head and neck squamous cell carcinoma are both treated with DNA damaging agents including platinum-based compounds and radiation therapy. However, at least one quarter of all tumors are resistant or refractory to these genotoxic agents. Yet the agents are extremely toxic, leading to undesirable side effects with potentially no benefit. Alternative therapies exist, but currently there are no tools to predict whether the first-line genotoxic agents will work in any given patient. To maximize therapeutic success and limit unnecessary toxicity, emerging clinical trials aim to inform personalized treatments tailored to the biology of individual tumors. Worldwide, significant resources have been invested in identifying biomarkers for guiding the treatment of lung and head and neck cancer. DNA repair proteins of the nucleotide excision repair pathway (ERCC1) and of the base excision repair pathway (XRCC1), which are instrumental in clearing DNA damage caused by platinum drugs and radiation, have been extensively studied as potential biomarkers of clinical outcomes in lung and head and neck cancers. The results are complex and contradictory. Here we summarize the current status of single nucleotide polymorphisms, mRNA, and protein expression of ERCC1 and XRCC1 in relation
to cancer risk and patient outcomes.

Optimizing response to gefitinib in the treatment of non-small-cell lung cancer

Pietro Carotenuto, Cristin Roma, Anna Maria Rachiglio, Raffaella Pasquale, et al.
Pharmacogenomics and Personalized Medicine 2011:4 1–9

The epidermal growth factor receptor (EGFR) is expressed in the majority of non-small-cell lung cancer (NSCLC). However, only a restricted subgroup of NSCLC patients respond to treatment with the EGFR tyrosine kinase inhibitor (EGFR TKI) gefitinib. Clinical trials have demonstrated that patients carrying activating mutations of the EGFR significantly benefit from treatment with gefitinib. In particular, mutations of the EGFR TK domain have been shown to increase the sensitivity of the EGFR to exogenous growth factors and, at the same time, to EGFR TKIs such as gefitinib. EGFR mutations are more frequent in patients with particular clinical and pathological features such as female sex, nonsmoker status, adenocarcinoma histology, and East Asian ethnicity. A close correlation was found between EGFR mutations and response to gefitinib in NSCLC patients. More importantly, randomized Phase III studies have shown the superiority of gefitinib compared with chemotherapy in EGFR mutant patients in the first-line setting. In addition, gefitinib showed a good toxicity profile with an incidence of adverse events that was significantly lower compared with chemotherapy. Therefore, gefitinib is a major breakthrough for the management of EGFR mutant NSCLC patients and represents the first step toward personalized treatment of NSCLC.

Pharmacogenomics of drug metabolizing enzymes and transporters: implications for cancer therapy

Jing Li, Martin H Bluth
Pharmacogenomics and Personalized Medicine 2011:4 11–33

The new era of personalized medicine, which integrates the uniqueness of an Individual with respect to the pharmacokinetics and pharmacodynamics of a drug, holds promise as a means to provide greater safety and efficacy in drug design and development. Personalized medicine is particularly important in oncology, whereby most clinically used anticancer drugs have a narrow therapeutic window and exhibit a large interindividual pharmacokinetic and pharmacodynamics variability. This variability can be explained, at least in part, by genetic variations in the genes encoding drug metabolizing enzymes, transporters, or drug targets. Understanding of how genetic variations influence drug disposition and action could help in tailoring cancer therapy based on individual’s genetic makeup. This review focuses on the pharmacogenomics of drug metabolizing enzymes and drug transporters, with a particular highlight of examples whereby genetic variations in the metabolizing enzymes and transporters influence the pharmacokinetics and/or response of chemotherapeutic agents.

Transcriptome-wide signatures of tumor stage in kidney renal clear cell carcinoma: connecting copy number variation, methylation and transcription factor activity
Qi Liu, Pei-Fang Su, Shilin Zhao and Yu Shyr
Genome Medicine 2014, 6:117

Background: Comparative analysis of expression profiles between early and late stage cancers can help to understand cancer progression and metastasis mechanisms and to predict the clinical aggressiveness of cancer. The observed stage-dependent expression changes can be explained by genetic and epigenetic alterations as well as transcription dysregulation. Unlike genetic and epigenetic alterations, however, activity changes of transcription factors, generally occurring at the post-transcriptional or post-translational level, are hard to detect and quantify.
Methods: Here we developed a statistical framework to infer the activity changes of transcription factors by simultaneously taking into account the contributions of genetic and epigenetic alterations to mRNA expression variations.
Results: Applied to kidney renal clear cell carcinoma (KIRC), the model underscored the role of methylation as a significant contributor to stage-dependent expression alterations and identified key transcription factors as potential drivers of cancer progression.
Conclusions: Integrating copy number, methylation, and transcription factor activity signatures to explain stage-dependent expression alterations presented a precise and comprehensive view on the underlying mechanisms during KIRC progression.

Developments in renal pharmacogenomics and applications in chronic kidney disease

Ariadna Padullés, Inés Rama, Inés Llaudó, Núria Lloberas
Pharmacogenomics and Personalized Medicine 2014:7 251–266

Chronic kidney disease (CKD) has shown an increasing prevalence in the last century. CKD encompasses a poor prognosis related to a remarkable number of comorbidities, and many patients suffer from this disease progression. Once the factors linked with CKD evolution are distinguished, it will be possible to provide and enhance a more intensive treatment to high-risk patients. In this review, we focus on the emerging markers that might be predictive or related to CKD progression physiopathology as well as those related to a different pattern of response to treatment, such as inhibitors of the renin–angiotensin system (including angiotensin-converting enzyme inhibitors and angiotensin II receptor blockers; the vitamin D receptor agonist; salt sensitivity hypertension; and progressive kidney-disease markers with identified genetic polymorphisms). Candidate-gene association studies and genome-wide association studies have analyzed the genetic basis for common renal diseases, including CKD and related factors such as diabetes and hypertension. This review will, in brief, consider genotype-based pharmacotherapy, risk prediction, drug target recognition, and personalized treatments, and will mainly focus on findings in CKD patients. An improved understanding will smooth the progress of switching from classical clinical medicine to gene-based medicine.








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