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The History and Creators of Total Parenteral Nutrition

Curator: Larry H. Bernstein, MD, FCAP

 

The History and Creators of Total Parenteral Nutrition

I am a pathologist who became involved in the measurement of acute and chronic malnutrition in hospitalized patients through my working with a burn surgeon, Walter Pleban, in the mid-1980s.  I had already been interested in this as a clinical pathology issue because the most abundant plasma protein, albumin, is markedly decreased, but that protein has a half-life of disappearance on 21 days.  This was problematic because it was inadequate for early recognition, or for response to feeding.  It became of considerable interest that two rapid turnover proteinhttp://www.ncbi.nlm.nih.gov/pubmed/20150597s – transthyretin (TTR)(then referred to as prealbumin) and retinol binding protein (RBP) that are synthesized by the liver have short half-lifes.  The measurement of TTR was then possible by an immunodiffusion assay on agarose overnight, but was not automated.  This changed with the introduction of an immunoassay for research use, and that offered by Beckman was ideal for the automated clinical laboratory.  One could then follow the level of TTR in the recovery phase.  There was some discussion for years about the fact that TTR might be considered an inverted acute phase protein because of a recognition that the liver decreases synthesis of TTR and produces acute phase proteins in the adaptive inflammatory response.  This is not insignificant, but it is also not quite relevant for reasons that have been addressed by Yves Ingenbleek and collaborators.  TTR is a key determinant of protein sufficiency and of sulfur homeostasis in health and disease.  I shall not say more, as the development of total parenteral (TPN), and also enteral (TEN) nutrition are of specific interest here.  However, the evaluation of patients’ nutritional status has widely been carried out by subjective global assessment, which is insufficient in a large population at risk.

 

History of parenteral nutrition.

The concept of feeding patients entirely parenterally by injecting nutrient substances or fluids intravenously was advocated and attempted long before the successful practical development of total parenteral nutrition (TPN) four decades ago. Realization of this 400 year old seemingly fanciful dream initially required centuries of fundamental investigation coupled with basic technological advances and judicious clinical applications. Most clinicians in the 1950’s were aware of the negative impact of starvation on morbidity, mortality, and outcomes, but only few understood the necessity for providing adequate nutritional support to malnourished patients if optimal clinical results were to be achieved. The prevailing dogma in the 1960’s was that, “Feeding entirely by vein is impossible; even if it were possible, it would be impractical; and even if it were practical, it would be unaffordable.” Major challenges to the development of TPN included: (1) formulate complete parenteral nutrient solutions (did not exist), (2) concentrate substrate components to 5-6 times isotonicity without precipitation (not easily done), (3) demonstrate utility and safety of long-term central venous catheterization (not looked upon with favor by the medical hierarchy), (4) demonstrate efficacy and safety of long-term infusion of hypertonic nutrient solutions (contrary to clinical practices at the time), (5) maintain asepsis and antisepsis throughout solution preparation and delivery (required a major culture change), and (6) anticipate, avoid, and correct metabolic imbalances or derangements (a monumental challenge and undertaking). This presentation recounts approaches to, and solution of, some of the daunting problems as really occurred in a comprehensive, concise and candid history of parenteral nutrition.

 

Historical highlights of the development of total parenteral nutrition.
The events and discoveries thought to be the most significant prerequisites to the development of total parenteral nutrition (TPN) dating back to the early 17th century are chronicled. A more detailed description and discussion of the subsequent early modern highlights of the basic and clinical research beginning in the mid-20th century and the advances culminating in the first demonstration of the feasibility and practicality of TPN, and its successful, safe and efficacious applications clinically, are presented. Some of the reasoning, insights, and philosophy of a pioneer clinician-scientist in the field are shared with readers.

 

The History, Principles, and Practice of Parenteral Nutrition in Preterm Neonates

Stanley J. Dudrick , Alpin D. Malkan
Chapter in:  
Nutrition for the Preterm Neonate    27 June 2013   pp 193-213

The history of the successful development of Total Parenteral Nutrition (TPN), first in beagle puppies in the basic science laboratories, and its subsequent clinical translations initially to adults, and shortly thereafter, to a newborn infant, is recounted by the original developer of the techniques, data, and results that have led to its widespread application and acceptance throughout the world. The principles, practices, standards, techniques, observations, technology, and several of the countless details which were so essential in guiding this dream to reality, are woven throughout the narrative. The advances and milestones are traced along this passionate, relentless journey to the present day, when preterm infants are actually expected to live and thrive. The precision and conscientious attention which are essential to the judicious, safe, efficacious use of TPN in preterm neonates throughout all aspects of solution formulation and delivery, together with appropriate monitoring and assessment of outcomes, are described and discussed briefly. The multiple risks and complications associated with this complex life-saving technique are extensively tabulated, with the intention to teach, in order to avoid, prevent, or overcome them. Moreover, attention has been directed toward pointing out many of the persisting shortcomings of the technique which remain to be prevented, overcome, or corrected by future research efforts and experiences. Finally, the costs, philosophy, humanity, and future advancements necessary to apply TPN to the care of preterm infants in developing countries are stated with optimism and hope.

 

Brief History of Parenteral and Enteral Nutrition in the Hospital in the USA
Bruce R. Bistrian
Clinical Nutrition, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA

Elia M, Bistrian B (eds): The Economic, Medical/Scientifi c and Regulatory Aspects of Clinical Nutrition Practice: What Impacts What?
Nestlé Nutr Inst Workshop Ser Clin Perform Program, vol. 12, pp 127–136, Nestec Ltd., Vevey/S. Karger AG, Basel, © 2009.

The meteoric rise in parenteral and enteral nutrition was largely a consequence of the development of total parenteral nutrition and chemically defined diets in the late 1960s and early 1970s and the recognition of the extensive prevalence of protein calorie malnutrition associated with disease in this same period. The establishment of Nutrition Support Services (NSS) using the novel, multidisciplinary model of physician, clinical nurse specialist, pharmacist, and dietitian, which, at its peak in the 1990s, approached 550 well-established services in about 10% of the US acute care hospitals, also fostered growth. The American Society of Parenteral and Enteral Nutrition, a multidisciplinary society reflecting the interaction of these specialties, was established in 1976 and grew from less than 1,000 members to nearly 8,000 by 1990. Several developments in the 1990s initially slowed and then stopped this growth. A system of payments, called diagnosis-related groups, put extreme cost constraints on hospital finances which often limited financial support for NSS teams, particularly the physician and nurse specialist members. Furthermore, as the concern for the nutritional status of patients spread to other specialties, critical care physicians, trauma surgeons, gastroenterologists, endocrinologists, and nephrologists often took responsibility for nutrition support in their area of expertise with a dwindling of the model of an internist or general surgeon with special skills in nutrition support playing the key MD role across the specialties. Nutrition support of the hospitalized patient has dramatically improved in the US over the past 35 years, but the loss of major benefits possible and unacceptable risks of invasive nutritional support if not delivered when appropriate, delivered without monitoring by nutrition experts, or employed where inappropriate or ineffective will require continued attention by medical authorities, hospitals, funding agencies, and industry in the future.

The rapid ascension of parenteral and enteral nutrition into an important component of clinical care in the hospital setting can be traced to three developments that occurred over an about 5-year period in the late 1960s and early 1970s. First and foremost was the first successful use of total parenteral nutrition (TPN), initially in beagle dogs to show the feasibility, and then its successful extension to 30 patients with chronic, complicated gastrointestinal disease by Dudrick et al. [1] at the University of Pennsylvania. At about the same time chemically defined or elemental diets were developed in normal volunteers to be employed in the US Mercury Space Program [2] where storage space and a low residue made these diets very desirable. These novel formulas were subsequently used in clinical conditions in which digestion and/or absorption was impaired and were provided usually through nasoenteric feeding tubes [3]. Both parenteral and enteral nutrition were initially studied in surgical patients in whom protein calorie malnutrition through gut malfunction had long been an often insurmountable problem. The third and final development was the identification of the extraordinary prevalence of malnutrition in hospitalized patients occurring in up to half of those on both surgical [4] and medical [5] services described in 1974 and 1976 respectively, when defined by simple anthropometric tools of weight, height, and upper arm anthropometry and serum albumin levels.

At this point one can view the glass as half full or half empty. From the optimistic or glass half full standpoint the period from 1975 to 1985 after the above advances could be described as a logarithmic phase of growth in clinical nutrition. Nutrition Support Services (NSS) using the novel, multidisciplinary model of physician, nurse specialist, pharmacist, and dietitian initially began in the early 1970s [6, 7] and at their high point probably approached 550 well-established services [8] in about 10% of America’s acute care hospitals by 1990. A number of studies during this early period demonstrated the ability of such groups to dramatically reduce the risk of catheter-related sepsis and to limit the development of electrolyte and metabolic abnormalities with TPN and to reduce complications and increase the adequacy of enteral nutrition [9]. Financial benefits were less certain in part due to difficulties to fully estimate costs and benefits [9], but at the very least were cost neutral in most circumstances [10].

The American Society of Parenteral and Enteral Nutrition which reflected this unique multidisciplinary membership of the NSS was established and had its first meeting in Chicago in 1976. Membership, initially less than 1,000 grew to nearly 8,000 by 1990 and was composed of approximately 20% physicians, 15% nurses, 15% pharmacists, and 50% dietitians in 1990. The annual ASPEN Clinical Congress, which continues to date, became an important venue to educate and train and provide a forum for the presentation of new research findings.

Finally from a personal perspective when I first became involved with nutrition support during my PhD training in Nutritional Biochemistry and Metabolism at MIT from 1972 to 1975, a period in which we were conducting the early surveys of nutritional status [4, 5], there was a general lack of appreciation for the nutritional status of patients. Protein calorie malnutrition was so widespread and undertreated that we developed a system of measurement of delayed cutaneous hypersensitivity to document cutaneous anergy [11] in order to convince clinicians that their patients required invasive nutritional support to reverse anergy. By 1990 there was a general appreciation that hospital protein calorie malnutrition was common, that invasive feeding could improve outcome, and that lack of feeding for periods of longer than 7–10 days in critically ill patients was an unacceptable practice. During this period from 1975 to 1990 there was a steady increase in the number of converts to better nutritional practices, particularly in surgical patients and in the critically ill in intensive care units, both medical and surgical. Testing for cutaneous anergy was abandoned at our medical center in the mid 1980s [12], principally because prolonged inadequate feeding became so uncommon, and there was little difficulty in convincing the primary physician of the need for invasive feeding when appropriate.

What happened subsequent to 1990? Now we can discuss the glass that is half empty, and this largely relates to medical funding. In the early 1980s the Medicare system in the US began a system of hospital payments based on diagnosis-related groups, where a fixed amount of money was paid according to diagnosis rather than actual costs. Medicare is the government system of reimbursement for patients 65 years or older, the disabled, or those receiving dialysis therapy. But the other source of hospital payments from medical care for the indigent through the government program Medicaid is the joint responsibility of the individual state and the federal government, and private insurance links their payments to government policy. The severe cost-containment pressures brought on by these changes in medical insurance have adversely affected nutrition support team staffing which began to have its greatest impact in the 1990s and was particularly harsh on hyperalimentation nurses and physicians involved in nutrition support. Although there are medical and financial costs associated with the termination of a nutrition support nurse [13], this cost must often be forcefully documented with hospital authorities, and generally can be in terms of unacceptable rates of catheter infection without their presence. With physicians there is no acknowledged medical specialty for clinical nutrition, although there was a split vote of 2–1 against by the American Board of Medical Specialties in the 1990s which would have accomplished this had it passed. Therefore, if the local hospital administrator or chairmen of medicine or surgery cannot be convinced of the value of providing partial financial support to nutrition support physicians for their clinical participation, then either it is done as a free service as an avocation by these individuals or done as a component of their underlying specialty. Thus most intensivists will provide parenteral and enteral nutrition as part of their care, as will many surgical specialists, particularly trauma surgeons, burn surgeons, and general surgeons. Oversight for home parenteral and enteral nutrition is often provided by gastroenterologists. However it is likely in many instances that nutritional care by these specialists is at an acceptable if perhaps not ideal level. For medical patients parenteral and enteral nutritional support is now often delivered under the care of dietitians which is reasonably good vis-à-vis enteral nutrition, but with parenteral nutrition may sometimes be outside their level of clinical competence, particularly for the management of fluid and electrolyte disorders and insulin management in diabetic patients. Dietitians have been less severely impacted by cost considerations, because there is a Joint Commission on Accreditation of Hospital Organizations (JCAHO) requirement that hospitals nutritionally monitor their patients. Pharmacists are also very important in the provision of parenteral nutrition, particularly by determining compatibilities of parenteral nutrition admixtures, checking the stability of orders from day-to-day, and by making certain of the completeness of parenteral regimens. Their continued availability to provide this level of expertise is also mandated by JCAHO as well as by their own professional standards.

There has also been a change in the membership of ASPEN that reflects this trend. After an initial fall of total members through the 1990s, the number has more recently stabilized, but there has been a dramatic decrease in nurses from nearly 1,000 to about 300 in 1999 and less than 200 at present (2006) with a concomitant increase in dietitians to about 60% of a total of 5,000 members, which has been relatively stable for the past 7 years, and a slowly diminishing number of physicians from 1,000 (20%) in 1999 to 735 (15%) in 2006. However both physician and pharmacist numbers have stabilized from 2001 to 2006, at approximately 750 and 620 members. Fellowship opportunities for physicians have also diminished, and there is some concern about what the future holds for physicians principally interested in parenteral and enteral nutrition. The second major American society for clinical nutrition after ASPEN was an independent group of academic physicians and PhD nutritionists interested in this field, the American Society for Clinical Nutrition. Last year by vote of its members it chose to disband and become a component of the American Society of Nutrition. Hopefully this group of individuals will maintain their interest in this field and continue to promote the improvement of parenteral and enteral nutrition for the hospitalized patient. However the likelihood of getting specialty recognition from the American Board of Medical Specialties is dim under the present conditions.

How does this bode for the future? Presumably there will always be some physicians trained in clinical nutrition, but some programs, like the exemplary program at MIT which trained many of the academic clinical nutritionists, have been discontinued and not been replaced. Certainly there is ample evidence for the need for such individuals. For instance one of the most important recent developments in clinical medicine has been the demonstration that tight blood glucose control in the critically ill can dramatically improve the morbidity and mortality of patients [18]. However this was primarily a study in cardiac surgical patients, and a similar study in medical patients by the same group demonstrated that tight blood glucose control improved morbidity but did not affect mortality [19]. In fact in those medical patients who received therapy for less than 3 days, mortality was actually increased. These superb innovative studies were primarily conducted by an endocrinologist who is a specialist in critical care. However an important variable in these two landmark studies, not previously commented on, is that in the surgical study the patients also received hypertonic dextrose initially for the first 24 h and TPN subsequently [18]. The medical patients in the second study received the initial hypertonic dextrose followed by inadequate nasogastric tube feeding for the first 3 days providing substantially less calories and grossly inadequate protein [19]. It may well be that it is the combination with tight glucose control in the setting of adequate feeding that is essential to achieve all the benefits rather than the control of hyperglycemia alone. Similarly a recent study in cardiac surgical patients receiving tight glucose control during their surgery and tight regulation of both treatment and control postoperatively showed no benefit and, in fact, a suggestion of harm in the treatment group [20]. Perhaps lowering blood glucose in cardiac patients not receiving hypertonic dextrose before revascularization may deprive the heart of an essential fuel. Having some physicians thoroughly trained in clinical nutrition to discern these possibilities may be important in the future to design and interpret the results of clinical trials.

 

For Patients Who Can’t Eat, Dr. Stanley Dudrick’s Intravenous Feeding System Is a Lifeline

Nearly 100 patients at the University of Texas Medical Center are undergoing similar nutritional therapy. Each owes his survival to Dr. Dudrick, who in 1972, at the precocious age of 37, became head of the center’s department of surgery.

Dudrick was turned from a fledgling cardiac surgeon into a pioneer nutritionist one day when he was an intern in Philadelphia. “We had three patients who had gone through successful surgery—but they all died,” he recalls. “I was terribly discouraged. Then the chairman of the surgery department said that, if I analyzed it, I’d see they really died of starvation. They couldn’t eat, and they didn’t have enough reserve tissues to draw on. I was too dumb to make that observation myself.”

Dudrick immersed himself in the study of how to provide food for those who can’t eat. From 10 to 40 percent of hospital deaths are still caused, he believes, by malnutrition. Patients with gastrointestinal cancer are especially vulnerable, as well as those with kidney or liver failure or burn trauma.

Sir Christopher Wren experimented with intravenous feeding of dogs as early as the 17th century. In its modern traditional form (most familiar in the glucose drip bottle), it cannot support life for long, however. Dudrick solved the problem by developing a complete nutritive compound. But he faced another obstacle: “We couldn’t put it in through the arm because the mixture was too thick and produced problems in the small veins. We couldn’t thin it down with water either, because that produced edema, or excess fluid in the connective tissue.

“Then,” Dudrick says, “we hit on the idea of putting it into larger veins, where the blood flow is so great that the nutritional substances would be diluted and rushed throughout the body.” Often the compound is pumped into the superior vena cava, through a catheter threaded through a smaller vein near the collarbone.

Dudrick’s nutrient, specially mixed for each patient, is composed of some 40 substances, including amino acids, glucose, vitamins and minerals. In some cases druggists or patients themselves can prepare the mixture.

Total Parenteral Nutrition (TPN) is Dudrick’s term for his technique. (Parenteral refers to bypassing the intestines.) In 1964 he astounded a medical convention in Germany with the news that he had raised six beagle puppies entirely on TPN for 287 days. In 1966 he first tried it on six humans with apparently terminal illness; all recovered and four are still alive. Since then Dudrick has used TPN on about 6,000 patients and has received two American Medical Association awards.

Eldest of four children of a Nanticoke, Pa. coal miner turned insurance agent, Dudrick decided on a medical career after watching the family doctor pull his mother through a near-fatal illness. Both his sisters are nurses. Still a crusader, he worries that, while half the nation’s doctors are aware of TPN, only five percent are using it. “It takes time,” he says, “for doctors to accept so much responsibility for dealing with such complex advances in human chemistry, metabolism and nutrition.”

Success will depend on campaigning for the technique, while simplifying it. “Someday we’ll have TPN down so that it will commonly be done in a general practitioner’s office,” Dudrick predicts. “That’s what I’m hoping for. I want to leave something better behind when I go, rather than just practice medicine the way it has always been done.”

Born in Rangoon, Burma on August 26, 1935, Khursheed N. Jeejeebhoy fled seven years later with his family to India to escape the Japanese invaders. He attended medical school in Vellore, India; trained in London, England; married and had three children; and in 1967, accepted a position at the Toronto General Hospital and the University of Toronto.From the beginning of his career, he was always on the forefront of research: he was one of the first to discover lactose intolerance. In 1970, with a surgical colleague, he was experimenting with TPN on post-surgical patients when Judy Ellis Taylor came into his care.

 

Dr. Khursheed Jeejeebhoy received his medical degree from the Christian Medical College Hospital in Vellore, India in 1959 and completed residency in India and the UK. He obtained his PhD from London University in 1963. He became division director of gastroenterology at the University of Toronto and the Toronto General Hospital. Currently, he is directs nutrition support and is a staff physician at St. Michael’s Hospital. He is also a professor of medicine, professor in the department of nutritional sciences and professor in the department of physiology, all at the University of Toronto. He has published over 500 peer-reviewed articles, abstracts and book chapters. He has a CIHR funded research program. He is on the editorial boards of nutritional journals and contributes to the Medical Post. He has received numerous awards throughout his career from Canada, USA and UK. He has been elected senior member of the Canadian Medical Association.

 

This determined young woman intended to live and expected him to save her. He took her up on her challenge and developed first a viable, long-term form of TPN, then a version Judy could use at home.With Judy such a success, Dr. Jeejeebhoy (Jeej to his patients and colleagues) bent his efforts to saving other lives with TPN and to learning more about the nutrients that the human body needs and in what dosages, both orally and intravenously, so that he could better nourish his patients and reduce their suffering. He has written over 350 papers and 100 books and chapters; was made professor of medicine, physiology, and nutrition at the University of Toronto; has lectured in virtually every country; and has taught many graduate students from Europe, North America, Asia, and Australia, as well as the first doctor allowed to leave China to study temporarily after China started opening up to the west.

His patients are intensely loyal to him, for his understanding, listening skills, expertise. In 1990, he moved to St. Michael’s Hospital and built up a TPN program there. He entered the commercial arena when he conducted research in and developed a radical new, nutritional way to improve the function of patients with congestive heart failure. MyLife Requirements “contains a patented combination of three nutrients, which interact synergistically and are needed by the heart to maintain optimal health and to function efficiently.  These nutrients are Coenzyme Q10, and the amino acids Taurine and Carnitine.” Due to the interesting regulation of L-carnitine by Health Canada, this supplement is available only in the US, not here in Canada.

At the end of 2007, he retired, sort of, a few years after becoming Professor Emeritus at the University of Toronto due to mandatory retirement at age 65. He closed his university lab at the end of 2007 when his last grant ran out. That ended a 40-year run of successful research grant applications and groundbreaking research. He embarked on a new role at St. Mike’s at the beginning of 2008, teaching at a Home TPN clinic; he continues to see patients part-time at a private clinic; and he conducts hospital rounds every week. His patients and colleagues would not allow complete retirement! Besides, Jeej is far too curious and interested in exploring new ideas to completely retire either!

 

 

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Metabolomic analysis of two leukemia cell lines. I.

Larry H. Bernstein, MD, FCAP, Reviewer and Curator

Leaders in Pharmaceutical Intelligence

 

I have just posted a review of metabolomics.  In the last few weeks, the Human Metabolome was published.  I am hopeful that my decision has taken the right path to prepare my readers adequately if they will have read the articles that preceded this.  I pondered how I would present this massive piece of work, a study using two leukemia cell lines and mapping the features and differences that drive the carcinogenesis pathways, and identify key metabolic signatures in these differentiated cell types and subtypes.  It is a culmination of a large collaborative effort that required cell culture, enzymatic assays, mass spectrometry, the full measure of which I need not present here, and a very superb validation of the model with a description of method limitations or conflicts.  This is a beautiful piece of work carried out by a small group by today’s standards.

I shall begin this by asking a few questions that will be addressed in the article, which I need to beak up into parts, to draw the readers in more effectively.

Q 1. What metabolic pathways do you expect to have the largest role in the study about to be presented?

Q2. What are the largest metabolic differences that one expects to see in compairing the two lymphoblastic cell lines?

Q3. What methods would be used to extract the information based on external metabolites, enzymes, substrates, etc., to create the model for the cell internal metabolome?

 

 

Abstract

Metabolic models can provide a mechanistic framework to analyze information-rich omics data sets, and are increasingly being used

  • to investigate metabolic alternations in human diseases.

An expression of the altered metabolic pathway utilization is

  • the selection of metabolites consumed and released by cells.

However, methods for the inference of intracellular metabolic states from extracellular measurements in the context of metabolic models

  • remain underdeveloped compared to methods for other omics data.

Herein, we describe a workflow for such an integrative analysis

  • extracting the information from extracellular metabolomics data.

We demonstrate, using the lymphoblastic leukemia cell lines Molt-4 and CCRF-CEM, how

  • our methods can reveal differences in cell metabolism.

Our models explain metabolite uptake and secretion by

  • predicting a more glycolytic phenotype for the CCRF-CEM model and
  • a more oxidative phenotype for the Molt-4 model, which
  • was supported by our experimental data.

Gene expression analysis revealed altered expression of gene products at

  • key regulatory steps in those central metabolic pathways,

and literature query emphasized

  • the role of these genes in cancer metabolism.

Moreover, in silico gene knock-outs identified

  • unique control points for each cell line model, e.g., phosphoglycerate dehydrogenase for the Molt-4 model.

Thus, our workflow is well suited to the characterization of cellular metabolic traits based on

  • extracellular metabolomic data, and
  • it allows the integration of multiple omics data sets into a cohesive picture based on a defined model context.

Keywords Constraint-based modeling _ Metabolomics _Multi-omics _ Metabolic network _ Transcriptomics

 

Reviewer Summary:

  1. A model is introduced to demonstrate a lymphocytic integrated data set using to cell lines.
  2. The method is required to integrate extracted data sets from extracellular metabolites to an intracellular picture of cellular metabolism for each cell line.
  3. The method predicts a more glycolytic or a more oxidative metabolic framework for one or the othe cell line.
  4. The genetic phenotypes differ with a unique control point for each cell line.
  5. The model presents an integration of omics data sets into a cohesive picture based on the model context.

Without having seen the full presentation –

  1. Is the method a snapshot of the neoplastic processes described?
  2. Does the model give insight into the cellular metabolism of an initial cell state for either one or both cell lines?
  3. Would one be able to predict a therapeutic strategy based on the model for either or both cell lines?

Before proceeding further into the study, I would conjecture that there is no way of knowing the initial state ( consistent with what is described by Ilya Prigogine for a self-organizing system) because the model is based on the study of cultured cells that had an unknown metabolic control profile in a host proliferating bone marrow that is likely B-cell origin.  So this is a snapshot of a stable state of two incubated cell lines.  Then the question that is raised is whether there is not only a genetic-phenotypic relationship between the cells in culture and the external metabolites produced, but also whether differences can be discerned between the  internal metabolic constructions that would fit into a family tree.

 

Introduction

Modern high-throughput techniques

  • have increased the pace of biological data generation.

Also referred to as the ‘‘omics avalanche’’, this wealth of data

  • provides great opportunities for metabolic discovery.

Omics data sets contain a snapshot of almost the entire repertoire of

  • mRNA, protein, or metabolites at a given time point or
  • under a particular set of experimental conditions.

Because of the high complexity of the data sets,

  • computational modeling is essential for their integrative analysis.

Currently, such data analysis

  • is a bottleneck in the research process and
  • methods are needed to facilitate the use of these data sets, e.g.,
  1. through meta-analysis of data available in public databases
    [e.g., the human protein atlas (Uhlen et al. 2010)
  2. or the gene expression omnibus (Barrett  et al.  2011)], and
  3. to increase the accessibility of valuable information
    for the biomedical research community.

Constraint-based modeling and analysis (COBRA) is

  • a computational approach that has been successfully used
  • to investigate and engineer microbial metabolism through
    the prediction of steady-states (Durot et al.2009).

The basis of COBRA is network reconstruction: networks are assembled

  1. in a bottom-up fashion based on genomic data and
  2. extensive organism-specific information from the literature.

Metabolic reconstructions

  1. capture information on the known biochemical transformations
    taking place in a target organism
  2. to generate a biochemical, genetic and genomic knowledge base
    (Reed et al. 2006).

Once assembled, a metabolic reconstruction

  • can be converted into a mathematical model
    (Thiele and Palsson 2010), and
  • model properties can be interrogated using a great variety of methods
    (Schellenberger et al. 2011).

The ability of COBRA models to represent

  • genotype–phenotype and environment–phenotype relationships
  • arises through the imposition of constraints,
  • which limit the system to a subset of possible network states
    (Lewis et al. 2012).

Currently, COBRA models exist for more than 100 organisms, including humans
(Duarte et al. 2007; Thiele et al. 2013).

Since the first human metabolic reconstruction was described
[Recon 1 (Duarte et al. 2007)],

  • biomedical applications of COBRA have increased
    (Bordbar and Palsson 2012).

One way to contextualize networks is to

  • define their system boundaries
  • according to the metabolic states of the system,
    e.g., disease or dietary regimes.

The consequences of the applied constraints

  • can then be assessed for the entire network
    (Sahoo and Thiele 2013).

Additionally, omics data sets have frequently been used

  • to generate cell-type or condition-specific metabolic models.

Models exist for specific cell types, such as

  • enterocytes (Sahoo and Thiele2013),
  • macrophages (Bordbar et al. 2010), and
  • adipocytes (Mardinoglu et al. 2013), and
  • even multi-cell assemblies that represent
    the interactions of brain cells (Lewis et al. 2010).

All of these cell type specific models,

  • except the enterocyte reconstruction
  • were generated based on omics data sets.

Cell-type-specific models have been used

  • to study diverse human disease conditions.

For example, an adipocyte model was generated using

  • transcriptomic,
  • proteomic, and
  • metabolomics data.

This model was subsequently used to investigate

  • metabolic alternations in adipocytes
  • that would allow for the stratification of obese patients
    (Mardinoglu et al. 2013).

One highly active field within the biomedical applications of COBRA is

  • cancer metabolism (Jerby and Ruppin, 2012).

Omics-driven large-scale models have been used

  • to predict drug targets (Folger et al. 2011; Jerby et al. 2012).

A cancer model was generated using

  • multiple gene expression data sets and
  • subsequently used to predict synthetic lethal gene pairs
  • as potential drug targets selective for the cancer model,
  • but non-toxic to the global model (Recon 1),
  • a consequence of the reduced redundancy in the
    cancer specific model (Folger et al. 2011).

In a follow up study, lethal synergy between

  • FH and enzymes of the heme metabolic pathway
    were experimentally validated and
  • resolved the mechanism by which FH deficient cells,
    e.g., in renal-cell cancer cells
  • survive a non-functional TCA cycle (Frezza et al. 2011).

Contextualized models, which contain only 

  • the subset of reactions active in 
  • a particular tissue (or cell-) type,
  • can be generated in different ways
    (Becker and Palsson, 2008; Jerby et al. 2010).

However, the existing algorithms mainly consider

  • gene expression and proteomic data to define the reaction sets
  • that comprise the contextualized metabolic models.

These subset of reactions are usually defined based on

  • the expression or absence of expression of the genes or proteins
    (present and absent calls), or
  • inferred from expression values or differential gene expression.

Comprehensive reviews of the methods are available
(Blazier and Papin, 2012; Hyduke et al. 2013).

Only the compilation of a large set of omics data sets

  • can result in a tissue (or cell-type) specific metabolic model, whereas

the representation of one particular experimental condition is achieved through

  • the integration of omics data set generated from one experiment only
    (condition-specific cell line model).

Recently, metabolomic data sets

  • have become more comprehensive and using these data sets allow
  • direct determination of the metabolic network components (the metabolites).

Additionally, metabolomics has proven to be

  1. stable,
  2. relatively inexpensive, and
  3. highly reproducible
    (Antonucci et al. 2012).

These factors make metabolomic data sets

  •  particularly valuable for interrogation of metabolic phenotypes. 

Thus, the integration of these data sets is now an active field of research
(Li et al. 2013; Mo et al. 2009; Paglia et al. 2012b; Schmidt et al. 2013).

Generally, metabolomic data can be incorporated into metabolic networks as

  1. qualitative,
  2. quantitative, and
  3. thermodynamic constraints
    (Fleming et al. 2009; Mo et al. 2009).

Mo et al. used metabolites detected in the spent medium
of yeast cells to determine

  • intracellular flux states through a sampling analysis (Mo et al. 2009),
  • which allowed unbiased interrogation of the possible network states
    (Schellenberger and Palsson 2009)
  • and prediction of internal pathway use.

Such analyses have also been used

  • to reveal the effects of enzymopathies on red blood cells (Price et al. 2004),
  • to study effects of diet on diabetes (Thiele et al. 2005) and
  • to define macrophage metabolic states (Bordbar et al. 2010).

This type of analysis is available as a function in the COBRA toolbox
(Schellenberger et al. 2011).

 

 

 

In this study, we established a workflow for the generation and analysis of

  • condition-specific metabolic cell line models that
  • can facilitate the interpretation of metabolomic data.

Our modeling yields meaningful predictions regarding

  • metabolic differences between two lymphoblastic leukemia cell lines
    (Fig. 1A).
Differences in the use of the TCA cycle by the CCRF-CEM

Differences in the use of the TCA cycle by the CCRF-CEM

 

 

 

http://link.springer.com/static-content/images/404/art%253A10.1007%252
Fs11306-014-0721-3/MediaObjects/11306_2014_721_Fig1_HTML.gif

Fig. 1

A  Combined experimental and computational pipeline to study human metabolism.
Experimental work and omics data analysis steps precede computational modeling. Model

  • predictions are validated based on targeted experimental data.

Metabolomic and transcriptomic data are used for

  • model refinement and submodel extraction.

Functional analysis methods are used to characterize

  • the metabolism of the cell-line models and compare it to additional experimental
    data.

The validated models are subsequently 

  • used for the prediction of drug targets.

B Uptake and secretion pattern of model.
All metabolite uptakes and secretions that were mapped during model
generation are shown.
Metabolite uptakes are depicted on the left, and

  • secreted metabolites are shown on the right.

A number of metabolite exchanges mapped to the model

  • were unique to one cell line.

Differences between cell lines were used to set

  • quantitative constraints for the sampling analysis.

C Statistics about the cell line-specific network generation.

 Quantitative constraints.
For the sampling analysis, an additional

  • set of constraints was imposed on the cell line specific models,
  • emphasizing the differences in metabolite uptake and secretion between cell lines.

Higher uptake of a metabolite was allowed in the model of the cell line

  • that consumed more of the metabolite in vitro, whereas
  • the supply was restricted for the model with lower in vitro uptake.

This was done by establishing the same ratio between the models bounds as detected in vitro.
X denotes the factor(slope ratio) that

  1. distinguishes the bounds, and
  2. which was individual for each metabolite.
  • (a) The uptake of a metabolite could be x times higher in CCRF-CEM cells,
    (b) the metabolite uptake could be x times higher in Molt-4,
    (c) metabolite secretion could be x times higher in CCRF-CEM, or
    (d) metabolite secretion could be x times higher in Molt-4 cells. LOD limit of detection.

The consequence of the adjustment was, in case of uptake, that  one model

  1. was constrained to a lower metabolite uptake (A, B), and the difference
  2. depended on the ratio detected in vitro.

In case of secretion,

  • one model had to secrete more of the metabolite, and again

the difference depended on

  • the experimental difference detected between the cell lines.

Q5. What is your expectation that this type of integrative approach could be used for facilitating medical data interpretations?

The most inventive approach was made years ago by using data constructions from the medical literature by a pioneer in the medical record development, but the technology was  not what it is today, and the cost of data input was high.  Nevertheless, the data acquisition would not be uniform across institutions, except for those that belong to a consolidated network with all of the data in the cloud, and the calculations would be carried out with a separate engine.  However, whether the uniform capture of the massive amount of data needed is not possible in the near foreseeable future.  There is no accurate way of assessing the system cost, and predicting the benefits.  In carrying this model forward there has to be a minimal amount of insufficient data.  The developments in the regulatory sphere have created a high barrier.

This concludes a first portion of this presentation.

 

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