Online Issues

ILAR Journal Vol 45(3)

Introduction

Unsung Heroes in the Battle Against Diabetes

Stephen W. Barthold

Stephen W. Barthold, M.D., Ph.D., is a Professor in the Department of Comparative Medicine, University of California, Davis, and a member of the Institute of Medicine.

This issue of ILAR Journal features diabetes as its subject material. Diabetes, including what is now divided into type 1 and type 2 diabetes, is a complex disease, and could not be covered in a single volume. Thus, this issue is focused largely on the subject of type 1 diabetes, or insulin-dependent diabetes mellitus, but also includes material on type 2 diabetes, or non-insulin-dependent diabetes mellitus, which represents an emerging epidemic with pressing need for animal models. The diversity of experimental models and modern research directions are protean, forcing a somewhat arbitrary selection of subjects that touch on this diversity. The various articles within this issue illustrate the international scope of diabetes research, the variety of animal models--mice, rats, pigs, nonhuman primates, and fish (among others not featured)--and the research directions and areas of inquiry that are being pursued by modern science.

In this issue, an overview of the human clinical perspective is provided by Dr. Eiselein et al. (2004), which emphasizes the growing importance of diabetes in the human population. The subsequent articles involving animal model systems will give the reader a perspective on the complexity of this disease, and emphasize the continued need for both human clinical trials and animal-based studies to unravel its mysteries and to discover new therapeutic modalities. The nonobese diabetic (NOD) mouse has become a singularly important model system for deciphering the complexities of diabetes in a genetically-defined rodent. Dr. Homo-Delarche (2004) provides an overview of this model, illustrating that even under the well-defined genetic background of the NOD mouse, the diabetogenic process is extremely complex. Dr. Ikegami and colleagues (2004) also discuss the NOD mouse, but present a case for shared genetic factors in both type 1 and type 2 diabetes. The rat is also genetically defined, but offers other perspectives, with two review articles on rat models provided by Dr. Mordes et al. (2004) and Dr. Tirabassi et al. (2004), who provide overviews of type 1 and type 2 diabetes rat models, respectively. Autoimmunity has been found to be a major factor in type 1 diabetes, and the precipitators of this event include infectious agents, as discussed by several of the authors in this issue. Drs. Jun and Yoon (2002) published an outstanding review on the role of viruses in diabetes and virus-induced models of type 1 diabetes, so we obtained permission to reprint this article from the journal Diabetes/Metabolism Research and Reviews, with an updated Brief Review (Yoon and Jun 2004) for this issue. In addition to genetically defined rodent models, non-inbred species provide useful opportunities for research. One such model is the minipig, as reviewed by Drs. Larsen and Rolin (2004). In addition, nonhuman primates, because of their relatedness to the human, offer other advantages, including islet cell transplantation, as featured in the articles by Drs. Gaur (2004) and Contreras et al. (2004). Transplantation cannot be discussed without consideration of stem cell research, and this topic is reviewed in various animal model systems by Dr. Peck and colleagues (2004). Finally, for emphasis on diversity, Dr. Wright and coauthors (2004) provide a review of the tilapia model. Most gourmands recognize that tilapia are fish, and lest the reader think this article may be delving into the depths of esoterica, it should be emphasized that teleost fish have played a role in understanding diabetes since the very beginning.

None of the articles in this issue provide a history of diabetes research using animal models, so a brief overview is provided in this Introduction. The term diabetes is derived from the Greek roots dia (through) + bainein (to go), meaning a siphon, which aptly describes the polyuria that is a major clinical sign of this disease. The early historical record on diabetes is available in an excellent review (Leibowitz 1972). That reference documents the writings of Aretaeus, who described the clinical syndrome in the second century AD, referring to the disease by its already commonly accepted name in Greek terminology, as well as Galen's classification of diabetes as a disease of the kidney ("hydrops of the chamber pot, or diarrhea of the urine"). Far earlier accounts of polyuria-related diseases exist in the written record of the Ebers papyrus, dating from approximately 1500 BC, and Hindu texts dating between 100 and 700 AD. Hindu medicine gets the credit for the term "honey urine" (from which the moniker mellitus is derived) referring to the clear, colorless nature of diabetic urine (similar to the juice of sugar cane). In contrast to popular myth, this name had nothing to do with awareness of sugar in the urine. It was not until the 17th century that Thomas Willis actually discovered the sweet taste of diabetic urine, and M. Dobson's documentation in 1772 of "saccharine matter" in both the urine and blood. The chemical identification of glucose in the urine was achieved in 1815 by M. E. Chevreul, a French chemist. At that point, the term diabetes mellitus was coined to distinguish "sugar diabetes" from diabetes insipitus (Leibowitz 1972).

Richard Morton first noted the now well-known familial (genetic) relation to the disease in 1696 (Leibowitz 1972). Furthermore, G. Harley, a British physician, noted in 1866 that there were at least two distinct forms of the diabetes mellitus; and E. Lancereaux, a French physician, made a distinction between fat (diabete gras) and thin (diabete maigre) forms of diabetes mellitus in 1880 (cited from Gale 2001). Paul Langerhans, a pupil of Rudolph Virchow, described clusters of cells in the pancreas in 1869 that differed from acinar cells, and the French histologist E. Laguesse subsequently named these cell aggregates "ilots de Langerhans" (Laguesse 1893). The pathologist E. L. Opie correlated the presence of lesions in islets with clinical diabetes (Opie 1900a,b). These distantly spaced milestones selected from among many, which are described in far richer detail with Leibowitz' and Gale's reviews, illustrate the slow pace of medical science before the last century, which relied largely on clinical observation.

Although insulin was discovered in the 1920s (see below), physicians continued to be perplexed by the remarkably different forms of diabetes mellitus. W. Falta and R. Boller (1931) noted the difference between insulin-sensitive and insulin-resistant forms of the disease in 1931, but the first direct evidence that early-onset (juvenile) diabetes was due to deficiency in insulin occurred in 1951 with the development of a bioassay for circulating insulin in the blood (Bornstein and Lawrence 1951), followed by the documentation that diabetic pancreata contained nearly undetectable levels of insulin, relative to controls (Wrenshall et al. 1952). Meanwhile, physical anthropologists utilized somatotyping to distinguish two groups (groups I and II) of diabetics (thin juvenile patients and older patients with excess body fat) (Draper et al. 1944). Lister and colleagues also emphasized this distinction (Lister et al. 1951), and designated the two body types as types I and II, but this terminology did not come into general acceptance until 1976, when it was reintroduced by A. G. Cudworth (Cudworth 1976). Immunology emerged as an important new field in the 1950s, and diabetes came under scrutiny as an autoimmune disease in the 1960s and 1970s; yet incrimination of autoimmunity as a critical factor in diabetes pathogenesis has been a tortuous road (reviewed in Gale 2001, and in the articles in this issue), complicated by the inextricable relation of autoimmunity to histocompatibility and immune response genetics and, in many cases, underlying infectious (viral) triggers. To make a long story short, the American Diabetes Association now classifies type 1 diabetes mellitus into type 1A (immune-mediated) and 1B (non-immune-mediated). Type 1 diabetes (T1D) is now also termed insulin-dependent diabetes mellitus (IDDM) in contrast to type 2 diabetes (T2D), or non-insulin-dependent diabetes mellitus (NIDDM).

Arguably, the most significant milestone in understanding diabetes was the discovery of insulin, which has been the subject of numerous historical reviews (including Gale 2001; Groen 1972; Leibowitz 1972; Rosenfeld 2002). The discovery of insulin as a factor in diabetes is often credited to a group of Canadian scientists, Frederick Grant Banting (a surgeon), Charles Herbert Best (a student assistant at the time), and John James Rickard Macleod (a professor of physiology) at the University of Toronto, who published their hallmark discovery in 1922 (Banting and Best 1922; Banting et al. 1922). Precipitated by the importance of the discovery and the distinction of the Nobel Prize in Medicine, vanity and competition besmirched their relationship. The Nobel Prize was awarded in 1923 to only Banting and Macleod, who were already bitter enemies by the time of the award, but not to Best. Nobel Prizes often fail to recognize the equally important work of others, as was the case with the discovery of insulin. Several scientists had actually reported similar findings, which in some cases were published (but not fully recognized by the medical scientific community) many years before the Toronto group performed their studies. Georg Ludwig Zuelzer (Zuelzer 1908), Ernest Lyman Scott (Scott 1912), John Murlin (Murlin and Kramer 1913), Israel S. Kleiner (Kleiner 1919), and Nicolas C. Paulesco (Paulesco 1921) all tested pancreatic extracts and showed reduction of blood or urine sugar levels in animals or humans. Oskar Minkowski should receive major credit because his studies in 1890 served as the foundation of work to follow. He described experiments in animals in which pancreatectomy resulted in diabetes (Mering and Minkowski 1890) more than 20 yr before Banting's work. Several months before Banting's publications in 1922, Paulesco presented a series of lectures, followed by a publication on "pancreatin," which fully documented the discovery of insulin as a water-soluble substance in pancreatic extracts with the ability to reduce blood sugar when given to dogs by intravenous injection (Paulesco 1921). Most notably, Banting's work, at his own admission, was based on prior observations, in 1920, of Moses Barron, who reported the relation of islets of Langerhans with diabetes, and described the experimental approach used to investigate that relation (Barron 1920). Needless to say, awarding the Nobel Prize to Banting and Macleod for the "discovery" of insulin effectively disgruntled several other scientists who felt that they were more deserving of the recognition.

The individuals mentioned above are but a few of the "heroes" worth mention in the diabetes mellitus story, but the unsung heroes are the human patients and experimental animals who have contributed enormously to understanding this disease and that continue to play a vital role in unraveling the complex issues of pathogenesis and treatment. Before the discovery of insulin, type 1 diabetes had an extremely grave prognosis, and patients were understandably willing to subject themselves to experimentation. However, these early human clinical subjects suffered considerably from fever, abscesses, and pain related to crude extracts of insulin, not to mention frequent failures of the trials. A variety of animals likewise contributed to the early diabetes story. Banting's work involved pancreatic duct ligation, pancreatectomy, and insulin treatment of dogs; Minkowski performed similar studies in dogs, pigs, and rabbits; and Paulesco used dogs. Early studies that demonstrated the first direct evidence of insulin's effects utilized alloxan-induced diabetic, hypophysectomized, and adrenalectomized rats (Bornstein and Lawrence 1951). Best developed a method to measure insulin levels with a rat diaphragm bioassay (cited from Groen 1972). Mice were used to prove the association of viral infection (Coxsackie virus) with induction of diabetes (Pappenheimer et al. 1951), and mice were used in convulsion bioassays to test batches of commercial insulin (Wrenshall et al. 1952). Much of the early research on the role of the pancreas in diabetes revolved around methods to separate the endocrine from the exocrine components of the organ. Based on Barron's and Paulesco's work, Banting used pancreatic duct ligation to induce acinar atrophy in dogs, thereby reducing the proteolytic effects of acinar products in isolating insulin. In an elegantly simple observation of comparative anatomy using teleost fishes, Macleod prepared extracts of islet tissues, which are anatomically separate from the exocrine portion of the pancreas in fish, and definitively demonstrated that islets were the source of insulin (Macleod 1922). James Bertram Collip joined Mcleod's laboratory and developed methods for purification of insulin, but he also analyzed the potency of insulin batches by monitoring the blood sugar of normal rabbits given injections of these extracts, in lieu of depancreatized dogs (cited from Rosenfeld 2002).

In the early years of diabetes research, conditions of experimental animals, as well as human clinical trial subjects, were vastly different from those of contemporary research institutions. Banting acquired dogs off the streets of Toronto for $1 to $3 each, no questions asked. Following the success of his research, Banting demanded a modest salary (he was not paid until that point), a "laboratory boy" to look after his dogs, and repairs to the floor of his operating room. These requests were met only after he threatened to leave and go to the Mayo Clinic. Mortality among Banting's dogs in these early experiments was quite high. Banting had no previous experience in animal work, which resulted in the death of seven of his original 10 dogs, despite his training as a surgeon. Banting, by his own admission on more than one occasion, was not familiar with the literature before beginning his research. As a result, his work repeated that of others (reviewed in Rosenfeld 2002). Without demeaning the significance of his work, these observations make one wonder how a Nobel Prize-winning research protocol, submitted by a volunteer scientist without a faculty appointment, no previous animal experience, and unfamiliarity with the literature, would have fared through scrutiny of contemporary institutional animal care and use committees and institutional review boards (human subjects), and how his operating room and animal husbandry practices would be judged by current guidelines. Fortunately, laboratory animal and human studies standards have progressed as much as science.

In summary, diabetes research has depended on parallel and intersecting advances provided by human clinical observations and animal-related research, which continue today. Notably, diabetes not only afflicts humans, but is also a significant clinical problem in domestic animals. The incidence of type 1 diabetes mellitus is estimated to be one in 200 dogs and one in 800 cats, with strong breed predisposition, and appears to be less frequent in horses and cattle (Jones et al. 1996). The marvelous outcome of this comparative medical story is that research advances on diabetes involving human and animal subjects has benefited both humans and animals to an equal degree.

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