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Posts Tagged ‘islet cells of pancreas’


BET Proteins Connect Diabetes and Cancer

Larry H. Bernstein, MD, FCAP, Curator

LPBI

 

New Proteins Discovered That Link Obesity-Driven Diabetes to Cancer

http://www.dddmag.com/news/2016/03/new-proteins-discovered-link-obesity-driven-diabetes-cancer

 Killer T cells surround a cancer cell. Credit: NIH

Killer T cells surround a cancer cell. Credit: NIH

 

For the first time, researchers have determined how bromodomain (BRD) proteins work in type 2 diabetes, which may lead to a better understanding of the link between adult-onset diabetes and certain cancers.

The findings, which appear in PLOS ONE, show that reducing levels in pancreatic beta cells of individual BRDs, called BET proteins, previously shown to play a role in cancer, may also help patients who are obese and diabetic.

The research was led by Gerald V. Denis, PhD, associate professor of pharmacology and medicine at Boston University School of Medicine, who was the first to show that BET protein functions are important in cancer development.

Adult-onset diabetes has been known for decades to increase the risk for specific cancers. The three main members of the BET protein family, BRD2, BRD3 and BRD4, are closely related to each other and often cooperate. However at times, they work independently and sometimes against each other.

According to the researchers new small molecule BET inhibitors have been developed that block all three BET proteins in cancer cells, but they interfere with too many functions.

“The BET proteins provide a new pathway to connect adult-onset diabetes with cancer, so properly targeting BET proteins may be helpful for both,” explained Denis, who is the corresponding author of the study.

He believes this discovery shows the need for deeper analysis of individual BET proteins in all human cell types, starting with boosting insulin and improving metabolism in the pancreas of adults who are obese.

“Without better targeted drugs, some ongoing cancer clinical trials for BET inhibitors are premature. These new results offer useful insight into drug treatments that have failed so far to appreciate the complexities in the BET family.”

 

Epigenetic modulation of type-1 diabetes via a dual effect on pancreatic macrophages and β cells

eLife. 2014; 3: e04631.     doi:  10.7554/eLife.04631

Epigenetic modifiers are an emerging class of anti-tumor drugs, potent in multiple cancer contexts. Their effect on spontaneously developing autoimmune diseases has been little explored. We report that a short treatment with I-BET151, a small-molecule inhibitor of a family of bromodomain-containing transcriptional regulators, irreversibly suppressed development of type-1 diabetes in NOD mice. The inhibitor could prevent or clear insulitis, but had minimal influence on the transcriptomes of infiltrating and circulating T cells. Rather, it induced pancreatic macrophages to adopt an anti-inflammatory phenotype, impacting the NF-κB pathway in particular. I-BET151 also elicited regeneration of islet β-cells, inducing proliferation and expression of genes encoding transcription factors key to β-cell differentiation/function. The effect on β cells did not require T cell infiltration of the islets. Thus, treatment with I-BET151 achieves a ‘combination therapy’ currently advocated by many diabetes investigators, operating by a novel mechanism that coincidentally dampens islet inflammation and enhances β-cell regeneration.

DOI:http://dx.doi.org/10.7554/eLife.04631.001

eLife digest

The DNA inside a cell is often tightly wrapped around proteins to form a compact structure called chromatin. Chemical groups added to the chromatin can encourage nearby genes to either be switched on or off; and several enzymes and other proteins help to read, add, or remove these marks from the chromatin. If these chromatin modifications (or the related enzymes and proteins) are disturbed it can lead to diseases like cancer. It has also been suggested that similar changes may influence autoimmune diseases, in which the immune system attacks the body’s own tissues.

Drugs that target the proteins that read, add, or remove these chromatin modifications are currently being developed to treat cancer. For example, drugs that inhibit one family of these proteins called BET have helped to treat tumors in mice that have cancers of the blood or lymph nodes. However, because these drugs target pathways involved in the immune system they may also be useful for treating autoimmune diseases.

Now Fu et al. have tested whether a BET inhibitor might be a useful treatment for type-1 diabetes. In patients with type-1 diabetes, the cells in the pancreas that produce the insulin hormone are killed off by the immune system. Without adequate levels of insulin, individuals with type-1 diabetes may experience dangerous highs and lows in their blood sugar levels and must take insulin and sometimes other medications.

Using mice that spontaneously develop type-1 diabetes when still relatively young, Fu et al. tested what would happen if the mice received a BET inhibitor for just 2 weeks early on in life. Treated mice were protected from developing type-1 diabetes for the rest of their lives. Specifically, the treatment protected the insulin-producing cells and allowed them to continue producing insulin. The drug reduced inflammation in the pancreas and increased the expression of genes that promote the regeneration of insulin-producing cells.

Diabetes researchers have been searching for drug combinations that protect the insulin-producing cells and boost their regeneration. As such, Fu et al. suggest that these findings justify further studies to see if BET inhibitors may help to treat or prevent type-1 diabetes in humans.

Introduction

Acetylation of lysine residues on histones and non-histone proteins is an important epigenetic modification of chromatin (Kouzarides, 2000). Multiple ‘writers’, ‘erasers’, and ‘readers’ of this modification have been identified: histone acetyltransferases (HATs) that introduce acetyl groups, histone deacetylases (HDACs) that remove them, and bromodomain (BRD)-containing proteins that specifically recognize them. Chromatin acetylation impacts multiple fundamental cellular processes, and its dysregulation has been linked to a variety of disease states, notably various cancers (Dawson and Kouzarides, 2012). Not surprisingly, then, drugs that modulate the activities of HATs or HDACs or, most recently, that block acetyl-lysine:BRD interactions are under active development in the oncology field.

BRDs, conserved from yeast to humans, are domains of approximately 110 amino-acids that recognize acetylation marks on histones (primarily H3 and H4) and certain non-histone proteins (e.g., the transcription factor, NF-κB), and serve as scaffolds for the assembly of multi-protein complexes that regulate transcription (Dawson et al., 2011; Prinjha et al., 2012). The BET subfamily of BRD-containing proteins (BRDs 2, 3, 4 and T) is distinguished as having tandem bromodomains followed by an ‘extra-terminal’ domain. One of its members, Brd4, is critical for both ‘bookmarking’ transcribed loci post-mitotically (Zhao et al., 2011) and surmounting RNA polymerase pausing downstream of transcription initiation (Jang et al., 2005; Hargreaves et al., 2009; Anand et al., 2013; Patel et al., 2013).

Recently, small-molecule inhibitors of BET proteins, for example, JQ1 and I-BET, were found to be effective inhibitors of multiple types of mouse tumors, including a NUT midline carcinoma, leukemias, lymphomas and multiple myeloma (Filippakopoulos et al., 2010; Dawson et al., 2011; Delmore et al., 2011; Zuber et al., 2011). A major, but not the unique, focus of inhibition was the Myc pathway (Delmore et al., 2011; Mertz et al., 2011; Zuber et al., 2011; Lockwood et al., 2012). In addition, BET-protein inhibitors could prevent or reverse endotoxic shock induced by systemic injection of bacterial lipopolysaccharide (LPS) (Nicodeme et al., 2010; Seal et al., 2012; Belkina et al., 2013). The primary cellular focus of action was macrophages, and genes induced by the transcription factor NF-κB were key molecular targets (Nicodeme et al., 2010; Belkina et al., 2013).

Given several recent successes at transposing drugs developed for cancer therapy to the context of autoimmunity, it was logical to explore the effect of BET-protein inhibitors on autoimmune disease. We wondered how they might impact type-1 diabetes (T1D), hallmarked by specific destruction of the insulin-producing β cells of the pancreatic islets (Bluestone et al., 2010). NOD mice, the ‘gold standard’ T1D model (Anderson and Bluestone, 2005), spontaneously and universally develop insulitis at 4–6 weeks of age, while overt diabetes manifests in a subset of individuals beginning from 12–15 weeks, depending on the particular colony. NOD diabetes is primarily a T-cell-mediated disease, but other immune cells—such as B cells, natural killer cells, macrophages (MFs) and dendritic cells (DCs)—also play significant roles. We demonstrate that a punctual, 2-week, treatment of early- or late-stage prediabetic NOD mice with I-BET151 affords long-term protection from diabetes. Mechanistic dissection of this effect revealed important drug influences on both MFs and β cells, in particular on the NF-κB pathway. On the basis of these findings, we argue that epigenetic modifiers are an exciting, emerging option for therapeutic intervention in autoimmune diabetes.

I-BET151 protects NOD mice from development of diabetes

T1D progresses through identifiable phases, which are differentially sensitive to therapeutic intervention (Bluestone et al., 2010). Therefore, we treated NOD mice with the BET-protein inhibitor, I-BET151 (GSK1210151A [Dawson et al., 2011;Seal et al., 2012]) according to three different protocols: from 3–5 weeks of age (incipient insulitis), from 12–14 weeks of age (established insulitis), or for 2 weeks beginning within a day after diagnosis of hyperglycemia (diabetes). Blood-glucose levels of insulitic mice were monitored until 30 weeks of age, after which animals in our colony generally do not progress to diabetes.

I-BET151 prevented diabetes development, no matter whether the treated cohort had incipient (Figure 1A) or established (Figure 1B) insulitis. However, the long-term protection afforded by a 2-week treatment of pre-diabetic mice was only rarely observed with recent-onset diabetic animals. Just after diagnosis, individuals were given a subcutaneous insulin implant, which lowers blood-glucose levels to the normal range within 2 days, where they remain for only about 7 days in the absence of further insulin supplementation (Figure 1C, upper and right panels). Normoglycemia was significantly prolonged in mice treated for 2 weeks with I-BET151; but, upon drug removal, hyperglycemia rapidly ensued in most animals (Figure 1C, lower and right panels). The lack of disease reversal under these conditions suggests that β-cell destruction had proceeded to the point that dampening the autoinflammatory attack was not enough to stem hyperglycemia. However, there was prolonged protection from diabetes in a few cases, suggesting that it might prove worthwhile to explore additional treatment designs in future studies.

I-BET151 inhibits diabetes and insulitis in NOD mice.

…..

BET protein inhibition has a minimal effect on T cells in NOD mice

Given that NOD diabetes is heavily dependent on CD4+ T cells (Anderson and Bluestone, 2005), and that a few recent reports have highlighted an influence of BET-protein inhibitors on the differentiation of T helper (Th) subsets in induced models of autoimmunity (Bandukwala et al., 2012; Mele et al., 2013), we explored the effect of I-BET151 treatment on the transcriptome of CD4+ T cells isolated from relevant sites; that is, the infiltrated pancreas, draining pancreatic lymph nodes (PLNs), and control inguinal lymph nodes (ILNs). Microarray analysis of gene expression revealed surprisingly little impact of the 2-week treatment protocol on any of these populations, similar to what was observed when comparing randomly shuffled datasets (Figure 2A). It is possible that the above protocol missed important effects on T cells because those remaining after prolonged drug treatment were skewed for ‘survivors’. Therefore, we also examined the transcriptomes of pancreas-infiltrating CD4+ T cells at just 12, 24 or 48 hr after a single administration of I-BET151. Again, minimal, background-level, differences were observed in the gene-expression profiles of drug- and vehicle-treated mice (Figure 2B).

Little impact of BET-protein inhibition on CD4+T cells in NOD mice.

I-BET151 induces a regulatory phenotype in the pancreatic macrophage population

I-BET151 treatment promotes an MF-like, anti-inflammatory transcriptional program in pancreatic CD45+ cells.
The NF-κB signaling pathway is a major focus of I-BET151’s influence on NOD leukocytes.

BET-protein inhibition promotes regeneration of NOD β cells

BET-protein inhibition promotes regeneration of islet β cells

The studies presented here showed that treatment of NOD mice with the epigenetic modifier, I-BET151, for a mere 2 weeks prevented the development of NOD diabetes life-long. I-BET151 was able to inhibit impending insulitis as well as clear existing islet infiltration. The drug had a dual mechanism of action: it induced the pancreatic MF population to adopt an anti-inflammatory phenotype, primarily via the NF-κB pathway, and promoted β-cell proliferation (and perhaps differentiation). These findings raise a number of intriguing questions, three of which we address here.

First, why do the mechanisms uncovered in our study appear to be so different from those proposed in the only two previous reports on the effect of BET-protein inhibitors on autoimmune disease? Bandukwala et al. found that I-BET762 (a small-molecule inhibitor similar to I-BET151) altered the differentiation of Th subsets in vitro, perturbing the typical profiles of cytokine production, and reducing the neuropathology provoked by transfer of in-vitro-differentiated Th1, but not Th17, cells reactive to a peptide of myelin oligodendrocyte glycoprotein (Bandukwala et al., 2012). Unfortunately, with such transfer models, it is difficult to know how well the in vitro processes reflect in vivo events, and to distinguish subsidiary effects on cell survival and homing. Mele et al. reported that JQ1 primarily inhibited the differentiation of and cytokine production by Th17 cells, and strongly repressed collagen-induced arthritis and experimental allergic encephalomyelitis (Mele et al., 2013). However, with adjuvant-induced disease models such as these, it is difficult to discriminate influences of the drug on the unfolding of autoimmune pathology vs on whatever the adjuvant is doing. Thus, the very different dual mechanism we propose for I-BET151’s impact on spontaneously developing T1D in NOD mice may reflect several factors, including (but not limited to): pathogenetic differences in induced vs spontaneous autoimmune disease models; our broader analyses of immune target cell populations; and true mechanistic differences between T1D and the other diseases. As concerns the latter, it has been argued that T1D is primarily a Th1-driven disease, with little, or even a negative regulatory, influence by Th17 cells (discussed in [Kriegel et al., 2011]).

Second, how does I-BET151’s effect, focused on MFs and β cells, lead to life-long protection from T1D? MFs seem to play a schizophrenic role in the NOD disease. They were shown long ago to be an early participant in islet infiltration (Jansen et al., 1994), and to play a critical effector role in diabetes pathogenesis, attributed primarily to the production of inflammatory cytokines and other mediators, such as iNOS (Hutchings et al., 1990; Jun et al., 1999a, 1999b; Calderon et al., 2006). More recently, there has been a growing appreciation of their regulatory role in keeping diabetes in check. For example, the frequency of a small subset of pancreatic MFs expressing the complement receptor for immunoglobulin (a.k.a. CRIg) at 6–10 weeks of age determined whether or not NOD diabetes would develop months later (Fu et al., 2012b), and transfer of in-vitro-differentiated M2, but not M1, MFs protected NOD mice from disease development (Parsa et al., 2012).

One normally thinks of immunological tolerance as being the purview of T and B cells, but MFs seem to be playing the driving role in I-BET151’s long-term immunologic impact on T1D. Chronic inflammation (as is the insulitis associated with T1D) typically entails three classes of participant: myeloid cells, in particular, tissue-resident MFs; lymphoid cells, including effector and regulatory T and B cells; and tissue-target cells, that is, islet β cells in the T1D context. The ‘flavor’ and severity of inflammation is determined by three-way interactions amongst these cellular players. One implication of this cross-talk is that a perturbation that targets primarily one of the three compartments has the potential to rebalance the dynamic process of inflammation, resetting homeostasis to a new level either beneficial or detrimental to the individual. BET-protein inhibition skewed the phenotype of pancreatic MFs towards an anti-inflammatory phenotype, whether this be at the population level through differential influx, efflux or death, or at the level of individual cells owing to changes in transcriptional programs. The ‘re-educated’ macrophages appeared to be more potent at inhibiting T cell proliferation. In addition, it is possible that MFs play some role in the I-BET151 influences on β-cell regeneration. The findings on Rag1-deficient mice ruled out the need for adaptive immune cells in the islet infiltrate for I-BET151’s induction of β-cell proliferation, but MFs are not thought to be compromised in this strain. Relatedly, the lack of a consistent I-BET151 effect on cultured mouse and human islets might result from a dearth of MFs under our isolation and incubation conditions (e.g., [Li et al., 2009]). Several recent publications have highlighted a role for MFs, particularly M2 cells, in promoting regeneration of β cells in diverse experimental settings (Brissova et al., 2014; Xiao et al., 2014), a function foretold by the reduced β-cell mass in MF-deficient Csf1op/op mice reported a decade ago (Banaei-Bouchareb et al., 2004).

Whether reflecting a cell-intrinsic or -extrinsic impact of the drug, several pro-regenerative pathways appear to be enhanced in β-cells from I-BET151-treated mice. Increased β-cell proliferation could result from up-regulation of the genes encoding Neurod1 (Kojima et al., 2003), GLP-1R (De Leon et al., 2003), or various of the Reg family members (Unno et al., 2002; Liu et al., 2008), the latter perhaps a consequence of higher IL-22R expression (Hill et al., 2013) (see Figure 6B and Supplementary file 4). Protection of β-cells from apoptosis is likely to be an important outcome of inhibiting the NF-κB pathway (Takahashi et al., 2010), but could also issue from enhanced expression of other known pro-survival factors, such as Cntfr (Rezende et al., 2007) and Tox3 (Dittmer et al., 2011) (see Figures 4 and 6B). Lastly, β-cell differentiation and function should be fostered by up-regulation of genes encoding transcription factors such as Neurod1, Pdx1, Pax6, Nkx6-1 and Nkx2-2. The significant delay in re-onset of diabetes in I-BET151-treated diabetic mice suggests functionally relevant improvement in β-cell function. In brief, the striking effect of I-BET151 on T1D development in NOD mice seems to reflect the fortunate concurrence of a complex, though inter-related, set of diabetes-protective processes.

Lastly, why does a drug that inhibits BET proteins, which include general transcription factors such as Brd4, have such circumscribed effects? A 2-week I-BET151 treatment might be expected to provoke numerous side-effects, but this regimen seemed in general to be well tolerated in our studies. This conundrum has been raised in several contexts of BET-inhibitor treatment, and was recently discussed at length (Shi and Vakoc, 2014). The explanation probably relates to two features of BET-protein, in particular Brd4, biology. First: Brd4 is an important element of so-called ‘super-enhancers’, defined as unusually long transcriptional enhancers that host an exceptionally high density of TFs—both cell-type-specific and general factors, including RNA polymerase-II, Mediator, p300 and Brd4 (Hnisz et al., 2013). They are thought to serve as chromatin depots, collecting TFs and coordinating their delivery to transcriptional start-sites via intra-chromosome looping or inter-chromosome interactions. Super-enhancers are preferentially associated with loci that define and control the biology of particular cell-types, notably developmentally regulated and inducible genes; intriguingly, disease-associated, including T1D-associated, nucleotide polymorphisms are especially enriched in the super-enhancers of disease-relevant cell-types (Hnisz et al., 2013;Parker et al., 2013). Genes associated with super-enhancers show unusually high sensitivity to BET-protein inhibitors (Chapuy et al., 2013; Loven et al., 2013;Whyte et al., 2013). Second: although the bromodomain of Brd4 binds to acetyl-lysine residues on histone-4, and I-BET151 was modeled to inhibit this interaction, it is now known to bind to a few non-histone chromosomal proteins as well, notably NF-κB, a liaison also blocked by BET-protein inhibitors (Huang et al., 2009; Zhang et al., 2012; Zou et al., 2014). Abrogating specific interactions such as these, differing according to the cellular context, might be the dominant impact of BET inhibitors, a scenario that would be consistent with the similar effects we observed with I-BET151 and BAY 11–7082 treatment. Either or both of these explanations could account for the circumscribed effect of I-BET151 on NOD diabetes. Additionally, specificity might be imparted by different BET-family members or isoforms—notably both Brd2 and Brd4 are players in MF inflammatory responses (Belkina et al., 2013). According to either of these explanations, higher doses might unleash a broader array of effects.

 

Islet inflammation: A unifying target for diabetes treatment?

In the last decade, islet inflammation has emerged as a contributor to the loss of functional β cell mass in both type 1 (T1D) and type 2 diabetes (T2D). Evidence supports that over-nutrition and insulin resistance result in the production of proinflammatory mediators by β cells. In addition to compromising β cell function and survival, cytokines may recruit macrophages into islets, thus augmenting inflammation. Limited, but intriguing, data implies a role of adaptive immune response in islet dysfunction in T2D. Clinical trials validated anti-inflammatory therapies in T2D, while immune therapy for T1D remains challenging. Further research is required to improve our understanding of islet inflammatory pathways, and to identify more effective therapeutic targets for T1D and T2D.
Islet inflammation: an emerging and unifying target for diabetes treatment

The current epidemic of T2D is closely associated with increases in obesity [1]. Excessive energy balance results in insulin resistance that is compensated for by increasing insulin secretion. However, insufficient compensation results in T2D, which is characterized by the reduction in islet mass and function. In recent years, overwhelming evidence defines insulin resistance as a state of chronic inflammation involving both innate and adaptive immune responses [1]. Although the presence of islet inflammation is acknowledged for autoimmune destruction of β cells in T1D, new data implicates overlapping pathogenesis between T1D and T2D. Epidemiologic studies suggest that obesity modifies the risk of T1D development [2, 3]. Importantly, small but seminal human studies have also provided evidences that anti-inflammatory therapy can improve glycemia and β cell function in T2D [4, 5]. Here, we focus on recent discoveries (past five years) to discuss the contribution of inflammatory pathways to islet dysfunction in T2D, and to provide updates on the pathogenesis of T1D.

What triggers inflammation in islets under insulin resistance?

Ample evidence from rodent and human studies indicates that in obesity, adipose tissue (AT) inflammation is a major source of pro-inflammatory mediators, and a primary response to excessive caloric intake. AT contributes to inflammation in obesity by means of increased mass, modified adipocyte phenotype, and increased infiltration of immune cells, which affects islet function through humoral and neuronal pathways [1, 6, 7]. In addition, it is noteworthy that pancreatic islets are under similar stress as adipocytes in T2D. The chronic inflammatory state of T2D is reflected in the elevation of circulatory cytokines that potentially affect islets as well as adipocytes [6, 8]. Both islets and adipocytes are exposed to excess glucose and lipids, especially free fatty acids (FFA). Over-nutrition forces adipose tissue to remodel and accommodate enlarged adipocytes, which results in endoplasmic reticulum (ER) stress, hypoxia, and mechanical stresses [911]. Under insulin resistance, insulin production increases to meet the high demand, resulting in the expansion of islet mass [12]. Recent findings revealed that obesity is associated with the activation of inflammatory pathway in the hypothalamus, which may alter functions of AT and islets through neuronal regulation [13]. Considering the multiple stressors potentially shared by AT and islets, it is plausible that islets exist also in a chronic inflammatory state, in T2D.

Adipose tissue dysfunction in obesity: a contributor to β cell inflammation in T2D?

The relationship between the pancreatic islet and AT was thought to be unidirectional, by placing insulin secretion as the major determinant of adipocyte glucose uptake and triglyceride storage. However, several recent studies suggest that insulin resistance in AT significantly contributes to β cell failure, through altered secretion of humoral factors from adipocytes and signals from the adipocyte sensory nerve (Figure. 1) [6, 7]. Of particular interest are adipocytokines that are uniquely produced by adipocytes, such as leptin, adiponectin, omentin, resistin, and visfatin, which may contribute to β cell dysfunction during insulin resistance (Box 1). Circulating cytokines may also connect AT inflammation to β cell dysfunction. Overnight exposure of mouse islets to tumor necrosis factor-alpha (TNFα), Interleukin beta (IL-1β), plus Interferon-gamma (IFNγ), at levels comparable to those seen in human obesity, disrupts the regulation of intracellular calcium [8]. Although glucose stimulated insulin secretion (GSIS) was maintained in this study, circulating cytokines might contribute to islet dysfunction after a prolonged period of exposure and when combined with other stresses [8]. TNFα, a cytokine implicated in insulin resistance, reportedly increased islet amyloid polypeptide (IAPP, amylin) expression in β cells with no concurrent expression of proinsulin. This may lead to amyloid production and β cell death [14]. Recent findings showed that the enzyme dipeptidyl peptidase-4 (DPPIV) is secreted by human adipocytes, and therefore may reduce the half-life of DPPIV substrate glucagon-like peptide-1 (GLP-1) with important implications on the insulinotropic effects of this gut peptide on the β cells [15]. Although it is not clear if obesity is associated with increased levels of DPPIV, inhibition of the latter by sitagliptin in a rodent model of obesity and insulin resistance reduced inflammatory cytokine production both in islets and in AT, and improved glucose-stimulated insulin secretion (GSIS) in islets in vitro [16]. Collectively, dysfunctional AT in obesity produces cytokines and peptides that affect islet health and potentially contribute to islet inflammation in T2D.

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Diabetes Mellitus

Author & Curator: Larry H. Bernstein, MD, FCAP

 

Diabetes mellitus (DM) is a group of metabolic diseases defined by high blood glucose levels, which, depending on the fasting blood glucose, may be pre-diabetes or overt diabetes (110 mg/dl. 124 mg/dl). This blood glucose level reflects a disorder of control of glucose metabolism, which is mediated through the pituitary growth hormone acting on the liver, which produces insulin growth factor 1 (IGF1).  Diabetes is due to either the pancreas not producing enough insulin, or the cells of the body not responding properly to the insulin produced. That said, there is much to be understood about the long term systemic effects of this disorder, a multisystem disease. The presence of pre-diabetes glucose levels is sufficient to proactively take measures to reduce the circulating glucose.

Globally, as of 2013, an estimated 382 million people have diabetes worldwide, with type 2 diabetes making up about 90% of the cases. This is equal to 8.3% of the adults population, with equal rates in both women and men. Worldwide in 2012 and 2013 diabetes resulted in 1.5 to 5.1 million deaths per year, making it the 8th leading cause of death. Diabetes overall at least doubles the risk of death. The number of people with diabetes is expected to rise to 592 million by 2035. The economic costs of diabetes globally was estimated in 2013 at $548 billion and in the United States in 2012 $245 billion.

The observation of symptoms of frequent urination, increased thirst, and increased hunger is symptomatic of overt DM, and is seen with diabetic ketoacidosis, with very high hyperglycemia and glucosuria, particularly in Type 1 DM. Untreated, diabetes leads to serious complications. Acute complications include diabetic ketoacidosis. Serious long-term complications include heart disease, stroke, kidney failure, foot ulcers and damage to the eyes.

There are three main types of diabetes mellitus:

  • Type 1 DM results from the body’s failure to produce enough insulin. This form was previously referred to as “insulin-dependent diabetes mellitus” (IDDM) or “juvenile diabetes”. The cause is unknown.
  • Type 2 DM begins with insulin resistance, a condition in which cells fail to respond to insulin properly. As the disease progresses a lack of insulin may also develop. This form was previously referred to as “non insulin-dependent diabetes mellitus” (NIDDM) or “adult-onset diabetes”. The primary cause is excessive body weight and not enough exercise.
  • Gestational diabetes, the third, occurs when pregnant women without a previous history of diabetes develop a high blood glucose level.

Type 1 DM, which presents suddenly in children or young adults, is possibly an as yet unidentified post-translational or epigenetic form, unrelated to Type 2, which is becoming more common in children.  It results in the destruction of islet beta cells that then have no capacity to produce insulin.  A family history of the disease would be a signal to raise a child with great care to not stress the pancreas.  Even though I raised the possibility of an epigenetic factor, it is important to keep in mind that the regulation of glucose is responsive to a number of stresses, even in a healthy person.  These are:

  • Corticosteroids
  • Glucagon
  • Growth hormone
  • Catecholamines
  • Proinflammatory cytokines
  • Anxiety disorder
  • Eating disorder

Gestational diabetes is perhaps Type 2 diabetes in a pregnant woman initiated by the condition of pregnancy. Whether these women were not diabetic, with a glucose level between 100-110 prior to pregnancy, is an open question. However, the pregnant state is accompanied by large effects by hormone levels.

Type 2 diabetes has been increasing worldwide, not only in western nations.  However, in non-western countries that have large populations of underserved, there is still a major problem with protein energy malnutrition (PEM). Globally, as of 2013, an estimated 382 million people have diabetes worldwide, with type 2 diabetes making up about 90% of the cases. This is equal to 8.3% of the adults population, with equal rates in both women and men. Worldwide in 2012 and 2013 diabetes resulted in 1.5 to 5.1 million deaths per year, making it the 8th leading cause of death. Diabetes overall at least doubles the risk of death. The number of people with diabetes is expected to rise to 592 million by 2035. The economic costs of diabetes globally was estimated in 2013 at $548 billion and in the United States in 2012 $245 billion.

The major long-term complications relate to damage to blood vessels. Diabetes doubles the risk of cardiovascular disease and about 75% of deaths in diabetics are due to coronary artery disease. Other “macrovascular” diseases are stroke, and peripheral vascular disease. The primary microvascular complications of diabetes include damage to the eyes, kidneys, and nerves. Damage to the eyes, known as diabetic retinopathy, is caused by damage to the blood vessels in the retina of the eye, and can result in gradual vision loss and potentially blindness. Damage to the kidneys, known as diabetic nephropathy, can lead to tissue scarring, urine protein loss, and eventually chronic kidney disease, sometimes requiring dialysis or kidney transplant. Damage to the nerves of the body, known as diabetic neuropathy, is the most common complication of diabetes.

Prevention and treatment involves a healthy diet, physical exercise, not using tobacco and being a normal body weight. Blood pressure control and proper foot care are also important for people with the disease. Type 1 diabetes must be managed with insulin injections. Type 2 diabetes may be treated with medications with or without insulin. Insulin and some oral medications can cause low blood sugar. Weight loss surgery in those with obesity is an effective measure in those with type 2 DM. Gestational diabetes usually resolves after the birth of the baby.

A number of articles in http://pharmaceuticalintelligence,com (this journal) have presented the relationship of DM to heart and vascular disease. The complexity of the disease is not to be underestimated, and there havr been serious controversies with adverse consequences over the use of the class of drugs that includes rosiglitazone and piaglitazone, which has opened serious issues about how clinical trials are conducted, and how the data obtained in studies may be compromised.

Pharmaceutical Insights

Management of Diabetes Mellitus: Could Simultaneous Targeting of Hyperglycemia and Oxidative Stress Be a Better Panacea?

Omotayo O. Erejuwa
Int. J. Mol. Sci. 2012, 13, 2965-2972; http://www.mdpi.com/journal/ijms http://dx.doi.org:/10.3390/ijms13032965

The primary aim of the current management of diabetes mellitus is to achieve and/or maintain a glycated hemoglobin level of ≤6.5%. However, recent evidence indicates that intensive treatment of hyperglycemia is characterized by increased weight gain, severe hypoglycemia and higher mortality. Besides, evidence suggests that it is difficult to achieve and/or maintain optimal glycemic control in many diabetic patients; and that the benefits of intensively-treated hyperglycemia are restricted to microvascular complications only. Evidence also indicates that multiple drugs are required to achieve optimal glycemic target in many diabetic patients. In fact, in many diabetic patients in whom optimal glycemic goal is achieved, glycemic control deteriorates even with optimal drug therapy. It does suggest that with the current hypoglycemic or antidiabetic drugs, it is difficult to achieve and/or maintain tight glycemic control in diabetic patients. In many developing countries, the vast majority of diabetic patients have limited or lack access to quality healthcare providers and good therapeutic monitoring.

While increased weight gain could be due to some component drugs (such as sulphonylureas or insulin) of the intensive therapy regimens, hypoglycemia could be drug-induced or comorbidity-induced. Considering the evidence that associates hypoglycemia with increased mortality, higher incidence of mortality in intensive therapy group could be due to hypoglycemia or too low levels of glycosylated hemoglobin. However, it is difficult to contend that increased mortality was entirely due to hypoglycemia. The possibility of drug-induced or drug-associated toxicities could not be ruled out. For instance, rosiglitazone, which has been prohibited and withdrawn from the market in Europe, was one of the hypoglycemic drugs used to achieve intensive therapy of hyperglycemia in Action to Control Cardiovascular Risk in Diabetes (ACCORD). If these findings are anything to go by, does it not suggest that targeting hyperglycemia as the only therapeutic goal in the management of diabetes mellitus could be detrimental to diabetic patients? In addition, the current hypoglycemic drugs are characterized by limitations and adverse effects. Together with the limitations of intensive glycemic treatment (only beneficial in reducing the risk of microvascular complications, but not macrovascular disease complications), does it not imply that targeting hyperglycemia alone is not only deleterious but also limited and ineffective?

The latest figures predict that the global incidence of diabetes mellitus, which was estimated to be 366 million in 2011, will rise to 522 million by 2030. In view of these frightening statistics on the prevalence of diabetes mellitus and on the lack of adequate healthcare, together with the associated diabetic complications, morbidity and mortality, does it not suggest that there is an urgent need for a better therapeutic management of this disorder? Taken together, with these findings and statistics, it can be contended that it is high time alternative and/or complementary therapies to the currently available hypoglycemic agents (which target primarily hyperglycemia only) were sought.

All these may contribute to the unabated increase in global prevalence of diabetes mellitus and its complications In view of these adverse effects and limitations of intensive treatment of hyperglycemia in preventing diabetic complications, which is linked to oxidative stress,

  • this commentary proposes a hypothesis that “simultaneous targeting of hyperglycemia and oxidative stress” could be more effective than “intensive treatment of hyperglycemia” in the management of diabetes mellitus.

Oxidative stress is defined as

  • an “imbalance between oxidants and antioxidants in favor of the oxidants, potentially leading to damage”.

It is implicated in the pathogenesis and complications of diabetes mellitus. The role of oxidative stress is more definite in the pathogenesis of type 2 diabetes mellitus than in type 1 diabetes mellitus. In regard to diabetic complications, there is compelling evidence in support of the role of oxidative stress in both types of diabetes mellitus. Evidence suggests that elevated reactive oxygen species (ROS), which causes factor of increased ROS production, causes tissue damage or diabetic complications have been identified. These include:

  • hyperglycemia-enhanced polyol pathway;
  • hyperglycemia-enhanced formation of advanced glycation endproducts (AGEs);
  • hyperglycemia-activated protein kinase C (PKC) pathway;
  • hyperglycemia-enhanced hexosamine pathway; and
  • hyperglycemia-activated Poly-ADP ribose polymerase (PARP) pathway.

These pathways are activated or enhanced by hyperglycemia-driven mitochondrial superoxide overproduction.

Even though oxidative stress plays an important role in its pathogenesis and complications,

  • unlike other diseases characterized by oxidative stress, diabetes mellitus is unique.

Its cure (restoration of euglycemia, e.g., via pancreas transplants) does not prevent oxidative stress and diabetic complications. This is very important because hyperglycemia exacerbates oxidative stress which is linked to diabetic complications. Theoretically, restoration of euglycemia should prevent oxidative stress and diabetic complications. However, this is not the case. At present, it remains unclear why restoration of euglycemia does not automatically prevent oxidative stress and diabetic complications. The development of diabetes-related complications (both microvascular and macrovascular) may occur in diabetic patients after normoglycemia has been restored. It is a phenomenon whereby previous hyperglycemic milieu is remembered in many target organs such as heart, eyes, kidneys and nerves. This phenomenon is also documented in diabetic animals. Compelling evidence implicates the role of oxidative stress as an important mechanism by which glycemic memory causes tissue damage and diabetic complications. In view of higher incidence of diabetic complications (of which oxidative stress plays an important role) in conventionally-treated diabetic patients, targeting oxidative stress in these patients might be beneficial. In other words, it is possible that the combination of a conventional therapy of hyperglycemia and antioxidant therapy might be more effective and beneficial than intensive therapy of hyperglycemia alone, which is the gold standard at the moment.

Loss of ACE 2 Exaggerates High-Calorie Diet-Induced Insulin Resistance by Reduction of GLUT4 in Mice

M Takeda, K Yamamoto, Y Takemura, H Takeshita, K Hongyo, et al.  Diabetes 61:1–11, 2012

ACE type 2 (ACE2) functions as

  • a negative regulator of the renin angiotensin system
  • by cleaving angiotensin II (AII) into angiotensin 1–7 (A1–7).

This study assessed the role of

  • endogenous ACE2 in maintaining insulin sensitivity.

Twelve-week-old male ACE2 knockout (ACE2KO) mice had normal insulin sensitivities when fed a standard diet. AII infusion or a high-fat high-sucrose (HFHS) diet impaired glucose tolerance and insulin sensitivity more severely

  • in ACE2KO mice than in their wild-type (WT) littermates.

The strain difference in glucose tolerance

  • was not eliminated by an AII receptor type 1 (AT1) blocker
  • but was eradicated by A1–7 or an AT1 blocker combined with the A1–7 inhibitor (A779).

The expression of GLUT4 and a transcriptional factor, myocyte enhancer factor (MEF) 2A,

  • was dramatically reduced in the skeletal muscles of the standard diet–fed ACE2KO mice.

The expression of GLUT4 and MEF2A was increased

  • by A1–7 in ACE2KO mice and
  • decreased by A779 in WT mice.

A1–7 enhanced upregulation of MEF2A and GLUT4 during differentiation of myoblast cells. In conclusion,

  • ACE2 protects against high calorie diet-induced insulin resistance in mice.

This mechanism may involve the transcriptional regulation of GLUT4 via an A1–7-dependent pathway.
Modulation of the action of insulin by angiotensin-(1–7)
FP. Dominici, V Burghi, MC. Munoz, JF. Giani

Clinical Science (2014) 126, 613–630 http://dx.doi.org:/10.1042/CS20130333

The prevalence of Type 2 diabetes mellitus is predicted to increase dramatically over the coming years and the clinical implications and healthcare costs from this disease are overwhelming. In many cases, this pathological condition is linked to a cluster of metabolic disorders, such as

  1. obesity,
  2. systemic hypertension and
  3. dyslipidaemia,
  • defined as the metabolic syndrome.

Insulin resistance has been proposed as the key mediator of all of these features and contributes to the associated high cardiovascular morbidity and mortality. Although the molecular mechanisms behind insulin resistance are not completely understood, a negative cross-talk between

  • AngII (angiotensin II) and the insulin signalling pathway

has been the focus of great interest in the last decade. Indeed,

substantial evidence has shown that

  • anti-hypertensive drugs that block the RAS (renin–angiotensin system) may also act to prevent diabetes.

Despite its long history, new components within the RAS continue to be discovered.

Among them, Ang-(1–7) [angiotensin-(1–7)] has gained special attention as a counter-regulatory hormone

  • opposing many of the AngII-related deleterious effects.

Specifically, we and others have demonstrated that Ang-(1–7) improves the action of insulin and opposes the negative effect that AngII exerts at this level. In the present review, we provide evidence showing that

  • insulin and Ang-(1–7) share a common intracellular signalling pathway.

We also address the molecular mechanisms behind the beneficial effects of Ang-(1–7) on

  • AngII-mediated insulin resistance.

Finally, we discuss potential therapeutic approaches leading to modulation of the

  • ACE2 (angiotensin-converting enzyme 2)/Ang-(1–7)/Mas receptor axis

as a very attractive strategy in the therapy of the metabolic syndrome and diabetes-associated diseases.

Increased Skeletal Muscle Capillarization After Aerobic Exercise Training and Weight Loss Improves Insulin Sensitivity in Adults With IGT

Prior, JB. Blumenthal, LI. Katzel, AP. Goldberg, AS. Ryan. Diabetes Care 2014;37:1469–1475
http://dx.doi.org:/10.2337/dc13-2358

Transcapillary transport of insulin is one determinant of glucose uptake by skeletal muscle; thus,

  • a reduction in capillary density (CD) may worsen insulin sensitivity.

Skeletal muscle CD is lower in older adults with impaired glucose tolerance (IGT) compared with those with normal glucose tolerance and

  • may be modifiable through aerobic exercise training and weight loss (AEX+WL).

Insulin sensitivity (M) and 120-min postprandial glucose (G120) correlated with CD at baseline (r = 0.58 and r = 20.60, respectively, P < 0.05).

AEX+WL increased maximal oxygen consumption (VO2max) 18%(P = 0.02) and reduced weight and fat mass 8% (P < 0.02).

Regression analyses showed that the AEX+WL-induced increase in CD

  • independently predicted the increase in M (r = 0.74, P < 0.01)
  • as well as the decrease in G120 (r = 20.55, P < 0.05).

AEX+WL increases skeletal muscle CD in older adults with IGT. This represents one mechanism by which AEX+WL improves insulin sensitivity in older adults with IGT.

Glycaemic durability with dipeptidyl peptidase-4 inhibitors in type 2 diabetes: a systematic review and meta-analysis of long-term randomised controlled trials.

K Esposito, P Chiodini, MI Maiorino, G Bellastella, A Capuano, D Giugliano. BMJ Open 2014;4:e005442.
http://dx.doi.org:/10.1136/bmjopen-2014-005442

A systematic review and meta-analysis of longterm randomised trials of DPP-4 inhibitors (sitagliptin, vildagliptin, saxagliptin, linagliptin and alogliptin). on haemoglobin A1c (HbA1c) was conducted. The difference between final and intermediate HbA1c assessment was the primary outcome. All trials were of 76 weeks duration at least. The difference in HbA1c changes between final and intermediate points averaged 0.22% (95% CI 0.15% to 0.29%), with high heterogeneity (I2=91%, p<0.0001). Estimates
of differences were not affected by the analysis of six extension trials (0.24%, 0.02 to 0.46), or five trials in which a DPP-4 inhibitor was added to metformin (0.24%, 0.16 to 0.32).

  • The effect of DPP-4 inhibitors on HbA1c in type 2 diabetes significantly declines during the second year of treatment.

Overcoming Diabetes Mellitus & Borderline Diabetes
By Max Stanley Chartrand, Ph.D. (Behavioral Medicine)

The over-arching biomarker that has more to do with the ability to restore normal metabolic processes is in achieving a cellular pH 7.45 (via the Kreb’s Cycle). To say the least, getting one’s cellular pH to 7.45 and A1C score below 6.0 can be a daunting task!

SIRCLE®: Naturally Achieved Targets

 Cellular pH 7.35-7.45

 Oxygen 99-100% @55-65 bpm

 Resting Blood Pressure: 110-135/ 65-80

mmHg (differs male vs female)

 Fasting blood sugar consistently <70-99

mg/dL or 3.5-5.5 mmol/L

 HgA1C score: .04-5.8

 HDL: 40-60 mg/dL; LDL: 100 -140 mg/dL;

triglycerides: <85 mg/dL

 C-Reactive Protein (CRP) Score <.5

 Galectin-3 Assay <17.8 ng/mL

Antidiabetic Activity of Hydroalcoholic Extracts of Nardostachys jatamansi in Alloxan-induced Diabetic Rats

M.A. Aleem, B.S. Asad, T Mohammed, R.A. Khan, M.F. Ahmed, A. Anjum, M. Ibrahim. Brit J Med & Medical Res 4(28): 4665-4673, 2014. http://www.sciencedomain.org/review-history.php?iid=579&id=12&aid=5024

The antidiabetic study was carried out to estimate the anti hyperglycemic potential of Nardostachys Jatamansi rhizome’s hydroalcoholic extracts in alloxan induced diabetic rats over a period of two weeks. The hydroalcoholic extract HAE1 at a dose (500mg/kg) exhibited significantly greater antihyperglycemic activity than extract HAE2 at a dose (500mg/kg) in diabetic rats. The hydroalcoholic extracts showed improvement in different parameters associated with diabetes, like body weight, lipid
profile and biochemical parameters. Extracts also showed improvement in

  • regeneration of β-cells of pancreas in diabetic rats.

Histopathological studies support the healing of pancreas by hydro alcoholic extracts (HAE1& HAE2) of Nardostachys Jatamansi, as a probable mechanism of their antidiabetic activity.

Antidiabetic and Antihyperlipidemic Effect of Parmelia Perlata. Ach. in Alloxan Induced Diabetic Rats.
Jothi G and Brindha P
Internat J of Pharmacy and Pharmaceut Sciences 2014; 6(suppl 1)

The aqueous extract of the selected plant was administered at dose levels of 200mg and 400mg/kg body weight for 60 days. After the experimental period the blood and tissue samples were collected and subjected to various biochemical and enzymic parameters. There were profound alteration in

  • fasting blood glucose,
  • serum insulin,
  • glycosylated hemoglobin (HbA1C) and
  • liver glycogen levels in alloxanized rats.
  1. Glucose-6-phosphatase,
  2. glucokinase, and
  3. fructose 1-6 bisphosphatase activity
  • were also altered in diabetic rats.

Administration of plant extract significantly (P<0.05)

  • reduced the fasting blood glucose and HbA1C level and increased the level of plasma insulin.

The activities of glucose metabolizing enzymes were also resumed to normal. There was a profound improvement in serum lipid profiles by

  • reducing serum triglyceride, cholesterol, LDL, VLDL, free fatty acids, phospholipids and increasing the HDL level in a dose dependent manner.

The effects of leaf extract were compared with standard drug glibenclamide (600μg/Kg bw). The results indicate that Parmelia perlata. Ach., Linn. could be a good natural source for developing an antidiabetic drug that can effectively maintained the blood glucose levels and lipid profile to near normal values.

Pathophysiological Insights
Diabetic glomerulosclerosis

Reviewers: Nikhil Sangle, M.D.
Revised: 21 February 2014,
Copyright: (c) 2003-2012, PathologyOutlines.com, Inc.

General

==================================================

  • Diffuse capillary basement membrane thickening, diffuse and nodular glomerulosclerosis
  • Causes glomerular disease, arteriolar sclerosis, pyelonephritis, papillary necrosis; similar between type I and II patients
  • Accounts for 30% of long term dialysis patients in US; causes 20% of deaths in patients with diabetes < age 40
  • Changes may be related to nephronectin, which functions in the assembly of extracellular matrix (Nephrol Dial Transplant 2012;27:1889)

Clinical features

==================================================

  • Proteinuria occurs in 50%, usually 12-22 years after onset of diabetes
  • End stage renal disease occurs in 30% of type I patients
  • Early increased GFR and microalbuminemia (30-300 mg/day) are predictive of future diabetic nephropathy
  • Renal disease reduced by tight diabetic control; may recur with renal allografts; ACE inhibitors may reduce progression

Micro description

==================================================

  • Basement membrane thickening and increased mesangial matrix in ALL patients
  • Diffuse glomerulosclerosis: increase in mesangial matrix associated with PAS+ basement membrane thickening, eventually obliterates mesangial cells
  • Nodular glomerulosclerosis: also called intercapillary glomerulosclerosis or Kimmelstiel-Wilson disease; ovoid, spherical, laminated hyaline masses in peripheral of glomerulus, PAS+, eventually obliterates glomerular tuft; specific for diabetes and membranoproliferative glomerulonephritis, light-chain disease and amyloidosis (Hum Pathol 1993;24:77 (pathogenesis of Kimmelstiel-Wilson nodule))
  • Profound hyalinization of afferent arterioles (insudative lesion-intramural): specific for diabetes in afferent arterioles, but non-specific if in periphery of glomerular loop, Bowman’s capsule or mesangium; insudative material composed of proteins, lipids and mucopolysaccharides
  • Organizing fibroepithelial crescents: associated with aggressive clinical course
  • Diffuse thickening of tubular basement membrane, tubular atrophy and interstitial fibrosis
  • Isolated thickened glomerular basement membrane and proteinuria may be an early predictor of diabetic disease (Mod Pathol 2004;17:1506)

Nodular glomerulosclerosis, Kidney

 Glomeruli:

  1.     Acellular, homogeneous, eosinophilic, globular nodules in the mesangial orintercapillary region of a glomerular tuft with capillary displaced to the periphery.
  2.     Diffuse intercapillary glomerulosclerosis: increasing eosinophilic mesangial matrix materials.
  3.     Capsular drop: eosinophilic small nodules on Bowman’s capsule.
  4.     Fibrin cap: eosinophilic, waxy, fatty structure within the lumen of one or more capillary loops of glomerular tufts.
nodular glomeruloschlerosis

nodular glomeruloschlerosis

http://www.kidneypathology.com/Imagenes/Diabetes/Imagen.Hial.jul.w.jpg

Islet amyloid polypeptide, islet amyloid, and diabetes mellitus.

Westermark P1, Andersson A, Westermark GT.
Physiol Rev. 2011 Jul;91(3):795-826.
http://dx.doi.org:/10.1152/physrev.00042.2009.

Islet amyloid polypeptide (IAPP), or amylin, was named for its tendency to

  • aggregate into insoluble amyloid fibrils, features typical of islets of most individuals with type 2 diabetes.

This pathological characteristic is most probably of

  • great importance for the development of the β-cell failure in this disease,
  • but the molecule also has regulatory properties in normal physiology.

In addition, it possibly contributes to the diabetic condition. This review deals with both these facets of IAPP.

Islet amyloid polypeptide (IAPP, or amylin) is one of the major secretory products of β-cells of the pancreatic islets of Langerhans. It is

  • a regulatory peptide with putative function
  • both locally in the islets, where it inhibits insulin and glucagon secretion, and at distant targets.

It has binding sites in the brain, possibly contributing also to satiety regulation and inhibits gastric emptying. Effects on several other organs have also been described.

IAPP was discovered through its ability to

  • aggregate into pancreatic islet amyloid deposits,

which are seen particularly in association with type 2 diabetes in humans and with diabetes in a few other mammalian species, especially monkeys and cats.

Aggregated IAPP has cytotoxic properties and is believed to be

  • of critical importance for the loss of β-cells in type 2 diabetes

and also in pancreatic islets transplanted into individuals with type 1 diabetes. This review deals both with physiological aspects of IAPP and with the

  • pathophysiological role of aggregated forms of IAPP,
  • including mechanisms whereby human IAPP forms toxic aggregates and amyloid fibrils.

Islet amyloid, initially named “islet hyalinization,” was described in 1901 by two researchers independently and for a long time was considered an enigma. It was found to occur in association with diabetes mellitus, particularly in elderly individuals, but its possible pathogenetic importance was often denied. The similarity of the hyaline substance to amyloid was noted at an early date, and some researchers reported staining reactions typical of amyloid. It had been shown in 1959 that

  • amyloid of several types has a characteristic ultrastructure,
  • and islet deposits were found to share this appearance.

When biochemical analyses of amyloid fibrils from systemic primary and secondary amyloidoses showed that

  • these consisted of distinctive proteins,
  • it was suspected that the islet deposits might also be a polymerized protein.

The chemical composition of islet amyloid did not attract much attention even after the characteristics of other amyloid fibrils had been elucidated. The finding that the amyloid in C cell-derived medullary thyroid carcinoma is of polypeptide hormonal origin was an important indication that amyloid in other endocrine tissues also comes from the local secretory products, and it was believed that

  • insulin, or proinsulin, or split products thereof constitute the islet amyloid fibrils.

Immunological trials to characterize the amyloid yielded equivocal results. Only when concentrated formic acid was used on amyloid,

  • extracted from an amyloid-rich insulinoma, was it possible to purify the major fibril protein
  • and characterize it by NH2-terminal amino acid sequence analysis,

which very unexpectedly revealed a novel peptide,

  • not resembling any part of proinsulin
  • but with partial identity to the neuropeptide calcitonin gene-related peptide (CGRP).

Further characterization of the peptide purified from an insulinoma and from islet amyloid of human and feline origin proved it to be a 37-amino acid (aa) residue peptide. The peptide was initially named “insulinoma amyloid peptide” , later diabetes-associated peptide (DAP), and finally islet amyloid polypeptide (IAPP), or “amylin”.

IAPP is a 37-aa residue long peptide, but by the application of molecular biological methods it was quickly shown that IAPP is expressed initially as

  • part of an 89-aa residue preproprotein containing a 22-aa signal peptide and
  • two short flanking peptides, the latter cleaved off at double basic aa residues similar to proinsulin.

IAPP is expressed by one single-copy gene on the short arm of chromosome 12,

  • in contrast to insulin and the other members of the calcitonin family, including
  • CGRP,
  • adrenomedullin, and
  • calcitonin,

all of which are encoded by genes on the evolutionary related chromosome 11.

The preproIAPP gene contains three exons, of which

  • the last two encode the full prepromolecule.

The signal peptide is cleaved

  • off in the endoplasmic reticulum (ER), and
  • conversion of proIAPP to IAPP takes place in the secretory vesicles.

ProIAPP and proinsulin are both processed by the two endoproteases

  • prohormone convertase 2 (PC2) and
  • prohormone convertase 1/3 (PC1/3) and
  • by carboxypeptidase E (CPE) (Figure 1).
amylin

amylin

A: the amino acid sequence of human pro-islet amyloid polypeptide (proIAPP) with the cleavage site for PC2 at the NH2 terminus and the cleavage site for PC1/3 at the COOH terminus, indicated by arrows. The KR residues (blue) that remain at the COOH terminus after PC1/3 processing are removed by carboxypeptidase E. This event exposes the glycine residue that is used for COOH-terminal amidation.
Below is a cartoon of IAPP in blue with the intramolecular S-S bond between residues 2–7 and the amidated COOH terminus.

B: the amino acid sequence of human proinsulin with the basic residues at the B-chain/C-peptide junction and the A-chain/C-peptide/junction indicated in blue and the processing sites indicated by arrows. PC1/3 does almost exclusively process proinsulin at the B-chain/C-peptide junction while PC2 preferentially processes proinsulin at the A-chain/C-peptide junction. The basic residues (RR) (position 31, 32) that remain at the COOH terminus of the B-chain is removed by the carboxypeptidase CPE. Below is a cartoon of insulin A-chain and B-chain in red with intermolecular SS bonds between cystein residues 7 in the A and B chains, between cystein residues at position 19 in the B-chain and 20 in the A-chain and the intermolecular SS bond between cystein residues at position 6 and 11 of the A-chain.

http://physrev.physiology.org/content/physrev/91/3/795/F1.large.jpg

  1. IAPP and insulin genes contain similar promoter elements,
  2. and the transcription factor PDX1 regulates the effects of glucose on both genes.
  3. Glucose stimulated β-cells respond with a parallel expression pattern of IAPP and insulin in the rat.

However, this parallel secretion of IAPP and insulin is altered in experimental diabetes models in rodents. Perfused rat pancreas secreted relatively

  • more IAPP than insulin when exposed to dexamethasone, whereas
  • high doses of streptozotocin or alloxan reduced insulin secretion more than that of IAPP.

Oleat and palmitate increased the expression of IAPP but not of insulin in MIN6 cells. In mice fed a diet high in fat for 6 mo, plasma IAPP increased 4.5 times more than insulin compared with mice fed standard food containing 4% fat.

In human recipients who had become insulin-independent by intrahepatically transplanted islets, there was disproportionately

  • more IAPP than normal secreted during hyperglycemia.

These examples show that the strictly parallel expression of IAPP and insulin may be disturbed under certain conditions.

The crystalline structure of insulin in granules is well characterized.

  • Hexameric insulin, together with zinc, constitutes the core of the mature granules, while
  • IAPP, together with a large number of additional components, including the C peptide, is found in the halo region.

The highly fibrillogenic human IAPP has to be protected in some way from aggregation, which otherwise would take place spontaneously. The fact that very fibril-prone proteins can be kept in solution at high concentrations is known from studies of arthropod silk. The composition of the β-cell granule is extremely complex, and it has many components in addition to insulin and C peptide, in micromolar concentrations.

It is probable that IAPP is protected from aggregation by interaction with other components. Plausible candidates are

  • proinsulin, insulin, or their processing intermediates.

Insulin has been found to be

  • a strong inhibitor of IAPP fibril formation.

This finding has been verified in a number of subsequent studies, which have also shown the potency of the inhibition. The inhibition seems to depend

  • solely on the B-chain,
  • which binds specifically to a short segment of IAPP.

An insulin-to-IAPP ratio of between 1:5 and 1:100 had a strong inhibitory effect. The molar ratio between IAPP and insulin in the granule as a whole is ∼1–2:50.

Type 2 Diabetes, APOE Gene, and the Risk for Dementia and Related Pathologies. The Honolulu-Asia Aging Study

Rita Peila, Beatriz L. Rodriguez and Lenore J. Launer
Diabetes Apr 2002; 51(4): 1256-1262
http://dx.doi.org:/10.2337/diabetes.51.4.1256

Type 2 diabetes may be a risk factor for dementia, but the associated pathological mechanisms remains unclear. We evaluated the association of diabetes

  • alone or combined with the apolipoprotein E (APOE) gene
  • with incident dementia and neuropathological outcomes

in a population-based cohort of 2,574 Japanese-American men enrolled in the Honolulu-Asia Aging Study, including 216 subjects who underwent autopsy. Type 2 diabetes was ascertained by interview and direct glucose testing. Dementia was assessed in 1991 and 1994 by clinical examination and magnetic resonance imaging and was diagnosed according to international guidelines. Logistic regression was used to assess the RR of developing dementia, and log-linear regression was used to estimate the incident rate ratio (IRR) of neuropathological outcomes.

Diabetes was associated with

  1. total dementia (RR 1.5 [95% CI 1.01–2.2]),
  2. Alzheimer’s disease (AD; 1.8 [1.1–2.9]), and
  3. vascular dementia (VsD; 2.3 [1.1–5.0]).

Individuals with both type 2 diabetes and the APOE ε4 allele

  • had an RR of 5.5 (CI 2.2–13.7) for AD compared with those with neither risk factor.

Participants with type 2 diabetes and the ε4 allele had

  • a higher number of hippocampal neuritic plaques (IRR 3.0 [CI 1.2–7.3]) and
  • neurofibrillary tangles in the cortex (IRR 3.5 [1.6–7.5]) and hippocampus (IRR 2.5 [1.5–3.7]), and
  • they had a higher risk of cerebral amyloid angiopathy (RR 6.6, 1.5–29.6).

Type 2 diabetes is a risk factor for AD and VsD. The association between diabetes and AD is particularly strong among carriers of the APOE ε4 allele. The neuropathological data are consistent with the clinical results.

Role of insulin signaling impairment, adiponectin and dyslipidemia in peripheral and central neuropathy in mice

  1. Anderson, MR. King, L Delbruck, CG. Jolivalt
    Dis. Model. Mech. June 2014; 7(6): 625-633
    http://dx.doi.org:/10.1242/dmm.015750

One of the tissues or organs affected by diabetes is the nervous system,

  • predominantly the peripheral system (peripheral polyneuropathy and/or painful peripheral neuropathy)
  • but also the central system with impaired learning, memory and mental flexibility.

The aim of this study was to test the hypothesis that the pre-diabetic or diabetic condition caused by a high-fat diet (HFD) can damage both the peripheral and central nervous systems. Groups of C57BL6 and Swiss Webster mice were fed a diet containing 60% fat for 8 months and compared to control and streptozotocin (STZ)-induced diabetic groups that were fed a standard diet containing 10% fat. Aspects of peripheral nerve function (conduction velocity, thermal sensitivity) and central nervous system function (learning ability, memory) were measured at assorted times during the study. Both strains of mice on HFD developed impaired glucose tolerance, indicative of insulin resistance, but

  • only the C57BL6 mice showed statistically significant hyperglycemia.

STZ-diabetic C57BL6 mice

  • developed learning deficits in the Barnes maze after 8 weeks of diabetes, whereas
  • neither C57BL6 nor Swiss Webster mice fed a HFD showed signs of defects at that time point.

By 6 months on HFD, Swiss Webster mice developed

  • learning and memory deficits in the Barnes maze test,
  • whereas their peripheral nervous system remained normal.

In contrast, C57BL6 mice fed the HFD developed peripheral nerve dysfunction,

  • as indicated by nerve conduction slowing and thermal hyperalgesia,
  • but showed normal learning and memory functions.

Our data indicate that STZ-induced diabetes or a HFD can damage

  • both peripheral and central nervous systems,
  • but learning deficits develop more rapidly in insulin-deficient than in insulin-resistant conditions
  • and only in Swiss Webster mice.

In addition to insulin impairment, dyslipidemia or adiponectinemia might determine the neuropathy phenotype.

Neuroinflammation and neurologic deficits in diabetes linked to brain accumulation of amylin

S Srodulski, S Sharma, AB Bachstetter, JM Brelsfoard, et al.
Molecular Neurodegeneration  2014; 9(30):
http://dx.doi.org:/10.1186/1750-1326-9-30

Background: We recently found that brain tissue from patients with type-2 diabetes (T2D) and cognitive impairment

  • contains deposits of amylin, an amyloidogenic hormone synthesized and co-secreted with insulin by pancreatic β-cells.

Amylin deposition is promoted by

  • chronic hypersecretion of amylin (hyperamylinemia), which is common in humans with obesity or pre-diabetic insulin resistance.

Human amylin oligomerizes quickly when oversecreted, which is toxic,

  • induces inflammation in pancreatic islets and
  • contributes to the development of T2D.

Here, we tested the hypothesis that accumulation of oligomerized amylin affects brain function.

Methods: In contrast to amylin from humans,

  • rodent amylin is neither amyloidogenic nor cytotoxic.

We exploited this fact by comparing

  • rats overexpressing human amylin in the pancreas (HIP rats) with their littermate rats

which express only wild-type (WT) non-amyloidogenic rodent amylin. Cage activity, rotarod and novel object recognition tests were performed on animals nine months of age or older. Amylin deposition in the brain was documented by immunohistochemistry, and western blot. We also measured neuroinflammation by immunohistochemistry, quantitative real-time PCR and cytokine protein levels.

Results: Compared to WT rats, HIP rats show

i) reduced exploratory drive,
ii) impaired recognition memory and
iii) no ability to improve the performance on the rotarod.

The development of neurological deficits is

  • associated with amylin accumulation in the brain.

The level of oligomerized amylin in supernatant fractions and pellets from brain homogenates

  • is almost double in HIP rats compared with WT littermates (P < 0.05).

Large amylin deposits (>50 μm diameter) were also occasionally seen in HIP rat brains. Accumulation of oligomerized amylin

  • alters the brain structure at the molecular level.

Immunohistochemistry analysis with an ED1 antibody indicates possible activated microglia/macrophages which

  • are clustering in areas positive for amylin infiltration.

Multiple inflammatory markers are expressed in HIP rat brains as opposed to WT rats, confirming that

  • amylin deposition in the brain induces a neuroinflammatory response.

Conclusions:

  1. Hyperamylinemia promotes accumulation of oligomerized amylin in the brain
  2. leading to neurological deficits through an oligomerized amylin-mediated inflammatory response.

Additional studies are needed to determine

  • whether brain amylin accumulation may predispose to diabetic brain injury and cognitive decline.

Keywords: Diabetes, Alzheimer’s Disease, Amylin, Pre-diabetes, Insulin Resistance, Inflammation, Behavior

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