Disease Markers 20 (2004) 161–165
`IOS Press
`
`161
`
`Update on diabetes mellitus
`
`Murray Korc
`Division of Endocrinology, Departments of Medicine, and Pharmacology and Toxicology, Dartmouth-Hitchcock
`Medical Center and Dartmouth Medical School, Hanover, NH, USA
`
`Abstract. Diabetes mellitus is a complex multi-system disorder that may be classified as autoimmune mediated type 1 diabetes,
`or as insulin resistance associated type 2 diabetes. In type 1 diabetes, there is selective loss of the beta cells within the endocrine
`islets, as a consequence of T-cell and cytokine mediated destruction of these cells, perhaps in conjunction with destruction of
`the peri-islet Schwann cells. In type 2 diabetes, the etiology of the resistance ranges from post-receptor defects in the insulin
`signaling pathway to excessive production of adipocyte derived cytokines that antagonize insulin action to mitochondrial defects
`that interfere with glucose disposal. Proteome based technologies are providing new insights into these defects.
`
`Keywords: Beta cell, insulin resistance, mitochondrial defects, pancreatic cancer
`
`1. Introduction
`
`Diabetes mellitus afflicts 6 to 8% of the population in
`the US. Type 1 diabetes mellitus (T1DM) patients have
`lost their beta cells as a result of aberrant activation
`of cellular immune mechanisms, and they no longer
`produce insulin. They are dependent, therefore, on
`insulin injections for survival. Most individuals with
`diabetes have type 2 diabetes mellitus (T2DM) and are
`resistant to insulin action. T2DM is also associated
`with beta cell dysfunction, and with production of fat-
`derived cytokines that antagonize insulin actions. This
`review will focus on several recent developments in
`T1DM and T2DM with an emphasis on the application
`of proteomic technologies.
`
`2. The beta cell and insulin secretion
`
`The pancreas is a complex tissue consisting of acinar
`cells that produce digestive enzymes, duct cells that
`produce bicarbonate rich fluid, and approximately one
`million endocrine islets which are distributed through-
`
`∗
`Correspondence to: Murray Korc, M.D., Department of
`Medicine, One Medical Center Drive, Lebanon, NH 03756, USA.
`Tel.: +1 603 650 7936; Fax: +1 603 650 6122; E-mail: murray.korc@
`dartmouth.edu.
`
`out the exocrine pancreas, with the greatest density oc-
`curring in the tail of the pancreas. The islets are rich
`in beta cells which secrete insulin, a hormone that is
`packaged into heterogeneous granules with electron-
`dense cores [1]. In addition, there are adjoining cells
`in the islets which secrete glucagon (alpha cells) and
`somatostatin (delta cells), and these hormones coun-
`teract insulin’s hypoglycemic effects and insulin se-
`cretion, respectively. There is also a small population
`of endocrine cells that secrete pancreatic polypeptide,
`whose function is not yet fully elucidated.
`Small amounts of insulin are secreted from the beta
`cells in the fasting state, acting to inhibit hepatic glu-
`coneogenesis [2]. Following a meal, glucose enters the
`beta cell through a process of facilitated diffusion that
`is mediated by the GLUT-2 glucose transporter. The
`same glucose transporter is also found in the liver, renal
`tubules, and the small intestine, but not in other types of
`islet cells [3]. GLUT-2 exhibits a high Km for glucose
`uptake, allowing the beta cell to transport glucose in
`proportion to the extracellular glucose concentration,
`thereby leading to a large increase in insulin secretion.
`Glucose stimulated release of insulin is biphasic,
`with a rapid first phase and a more gradual sec-
`ond phase that lasts for two hours and that is partly
`dependent on the release of newly synthesized in-
`sulin [4]. Glucose acts by generating ATP which in-
`hibits a K+ channel, thereby depolarizing the beta cell
`
`ISSN 0278-0240/04/$17.00  2004 – IOS Press and the authors. All rights reserved
`
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`plasma membrane, activating voltage-dependent cal-
`cium channels, and inducing a rise in intracellular free
`calcium [5–7]. Calcium-calmodulin protein kinases,
`cyclic AMP (cAMP)-dependent protein kinases, phos-
`phatidic acid, lysophospholipids, and arachidonic acid
`and its metabolites also have important roles in modu-
`lating insulin secretion [8–10]. Inter- and intracellular
`signaling by the cells of the pancreatic islet in response
`to glucose, as well as other metabolites, also contribute
`to the integrated response of the beta cell and to the
`controlled release of insulin in response to a meal.
`
`3. Type 1 diabetes mellitus
`
`T1DM is characterized by progressive destruction of
`the beta cells due to the aberrant activation of cellular
`immune mechanisms, as manifested by the presence of
`T-cell infiltrates around and within the islets. There are
`approximately one million T1DM patients in the US,
`all of whom are dependent on insulin therapy for their
`survival. The absence of insulin also makes them prone
`to develop ketoacidosis, a potentially deadly metabolic
`complication.
`The mechanisms that lead to beta cell destruction in
`T1DM are still not completely understood. It is widely
`accepted, however, that the selective assault on the beta
`cell is mediated by cytotoxic T-cells and by certain cy-
`tokines [11]. For example, interleukin-1β (IL-1β) can
`suppress beta cell function and survival [12]. Proteome
`based studies of islets and islet derived cell lines that
`were exposed to IL-1β has revealed a complex pat-
`tern of protein alterations, including increased and de-
`creased protein expression and de novo protein induc-
`tion, underscoring the wide range of responses that can
`be elicited in the these cells and that can modulate their
`responsiveness and susceptibility to cytokine mediated
`cell death [13–15].
`In addition to in vitro studies, investigators have ad-
`dressed the issue of beta cell destruction in numer-
`ous in vivo studies. For example, in a recent study
`with the non-obese diabetic (NOD) mouse, which is
`an excellent model of T1DM, the Schwann cells that
`surround the islets were noted to be targeted at an
`early, pre-diabetic stage by the aberrantly activated T
`cells [16]. These peri-islet Schwann cells (pSC) ex-
`press glial fibrillary acidic protein (GFAP), and the
`authors noted that the diabetes-prone NOD mice de-
`velop pSC-autoreactive T and B cell responses that are
`associated with progressive pSC death that precedes
`beta cell death. They used surface-enhanced laser des-
`
`orption/ionization (SELDI) with time-of-flight (TOF)
`mass spectrometry to detect auto-antibodies against
`GFAP in the sera of NOD mice and newly diagnosed
`diabetic children with T1DM [16]. Thus, an important
`component of the immune mediated destruction of the
`beta cell may be mediated by aberrant targeting of pSC.
`In another study, peptide epitopes from naturally pro-
`cessed proinsulin were delineated by TOF mass spec-
`trometry, and used to establish very sensitive enzyme-
`linked immunosorbent assays to assess the nature of
`auto-reactive T-cells in T1DM [17]. These T-cells ex-
`hibited a pro-inflammatory Th1 response in T1DM pa-
`tients, but a T-regulatory cell response leading to the
`preferential production of the anti-inflammatory cy-
`tokine interleukin-10 (IL-10) in non-diabetic individu-
`als [17]. These observations suggest that the immune
`system actively protects against potential beta cell de-
`struction, and that this protective mechanism is lost in
`T1DM.
`
`4. Type 2 diabetes mellitus
`
`T2DM is characterized by insulin resistance, a fail-
`ure of the beta cell to produce enough insulin to over-
`come the resistance, inappropriate hepatic glucose re-
`lease, and production of cytokines by adipose tissue
`that interfere with insulin action. There are approxi-
`mately 16 million T2DM patients in the US, some of
`whom may require insulin therapy to achieve adequate
`blood glucose control. The presence of normal, in-
`creased, or slightly decreased circulating insulin levels
`assures that they only rarely develop ketoacidosis.
`Insulin resistance in T2DM may be the consequence
`of abnormal production of anti-insulin receptor an-
`tibodies [18], but is generally due to post-receptor
`defects [19]. Some examples of post-receptor de-
`fects include silent polymorphisms and naturally oc-
`curring amino acid substitutions in the insulin recep-
`tor substrate-1 (IRS-1) signaling protein which may
`contribute to insulin resistance [20,21].
`In support
`of this hypothesis, IRS-1 knockout mice exhibit in-
`sulin resistance and glucose intolerance [22], and IRS-
`2 knockout mice exhibit decreased insulin-stimulated
`glucose transport in conjunction with decreased beta
`cell mass and overt diabetes [23]. Similarly, there
`is homology between the ataxia-telangiectasia (AT)
`gene product and phosphatidylinositol 3-kinase (PI 3-
`kinase). The relatively high frequency of T2DM in pa-
`tients with AT raises the possibility that perturbations
`in PI 3-kinase function may represent one of the post-
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`
`receptor defects that contributes to insulin resistance
`in some T2DM [24]. Mutations in the peroxisome
`proliferator-activated receptor (PPARγ) may lead to al-
`tered adipocyte differentiation and energy storage and
`may also contribute to insulin resistance in T2DM [25].
`Furthermore, excessive secretion of certain hormones
`such as cortisol, as seen in Cushing’s disease, or growth
`hormone, as seen in acromegaly, may antagonize in-
`sulin action sufficiently to induce a T2DM-like state.
`T2DM is often associated with obesity, which is a
`major cause for insulin resistance. Increased visceral
`obesity is especially deleterious in this regard because
`visceral fat is prone to release free fatty acids (FFA)
`which directly interfere with efficient insulin signaling,
`partly as a result of enhanced protein kinase C isoforms
`activity and increased hepatic glucose release [26,27].
`Furthermore, visceral fat expresses the β3-adrenergic
`receptor, which participates in the regulation of ther-
`mogenesis and lipolysis [28], and mutations in this re-
`ceptor are associated with insulin resistance, an earlier
`onset of T2DM and attenuated insulin secretion [29,
`30]. The adipocyte is also the source of several cy-
`tokines such as tumor necrosis factor alpha (TNF-α),
`which induces insulin resistance by down-regulating
`GLUT4 and increasing free fatty acid release [31],
`and interleukin-6 (IL-6), which induces insulin resis-
`tance in fat cells and hepatocytes [32,33]. By con-
`trast, adiponectin and IL-10 exert anti-diabetogenic ef-
`fects [34,35], and reduced levels of IL-10 increase the
`risk for the development of the dysmetabolic syndrome
`in women [36]. In addition to obesity and diabetes, pa-
`tients with the dysmetabolic syndrome exhibit hyper-
`tension, hyperlipidemia, heart disease and peripheral
`vascular disease, and a tendency towards elevated uric
`acid levels.
`The role of adipose tissue in the pathogenesis of in-
`sulin resistance has been investigated in several model
`cell lines, including the mouse 3T3-L1 fibroblastic cell
`line which differentiates rapidly into an adipocyte phe-
`notype in response to insulin and is useful,therefore, for
`studying insulin action and adipogenic differentiation.
`Thus, two-dimensional gel electrophoresis of 3T3-L1
`cell lysates revealed that insulin does not stimulate
`calmodulin phosphorylation under conditions in which
`it stimulates the phosphorylation of other proteins [37],
`and indicated that altered expression of several cellu-
`lar proteins contributes to the differentiation process in
`these cells [38]. More recently, a proteome based ap-
`proach using velocity gradient centrifugation to achieve
`the initial separation of proteins revealed that there was
`a marked increase in mitochondrial proteins in these
`
`cells during the differentiation process [39]. The au-
`thors also examined the effects of rosiglitazone, an ag-
`onist that activates the gamma isoform of the peroxi-
`some proliferator-activated receptor (PPARγ), which is
`a nuclear receptor that modulates adipocyte differentia-
`tion. In addition to altering mitochondrial morphology,
`rosiglitazone increased the levels of proteins involved
`in fatty acid oxidation, such as acyl-CoA synthetase and
`dehydrogenase [40]. Rosiglitazone also increases car-
`boxypeptidase B expression in mouse islets [41], and
`modulates components of the peroxisomal fatty acid
`metabolism pathway in adipose tissue [42], raising the
`possibility that it may also ameliorate glucose home-
`ostasis by improving insulin processing and fatty acid
`metabolism.
`Another site of insulin resistance is the skeletal mus-
`cle. A proteome based analysis of human skeletal
`muscle identified 8 potential markers of T2DM [43].
`The levels of two proteins that have a crucial role in
`ATP synthesis, creatine kinase B and ATP synthase β-
`subunit, were decreased in the muscle tissue of T2DM
`patients. Genetic muscular disorders may also be as-
`sociated with a high incidence of T2DM. For example,
`patients with mitochondrial myopathies may present
`with muscle weakness, symmetric paralysis of the ex-
`traocular muscles, drooping eyelids (ptosis), T2DM
`and cardiomyopathy [44]. Other patients with T2DM
`exhibit a maternal pattern of inheritance in conjunc-
`tion with inherited deafness and mitochondrial gene de-
`fects [45]. The importance of mitochondrial defects in
`T2DM was highlighted in a recent study which revealed
`that insulin-resistant children of T2DM parents exhib-
`ited increased muscle cell lipid content in conjunction
`with decreased mitochondrial phosphorylation, as de-
`termined by magnetic resonance spectroscopy [46]. In-
`terestingly, rates of lipolysis and plasma levels of IL-6,
`TNF-a, and adiponectin were not different in the two
`groups, further underscoring the potential importance
`of the mitochondrial defect in the pathogenesis of in-
`sulin resistance.
`Disorders of the pancreas that are neither T1DM nor
`T2DM may also be associated with glucose intolerance
`and diabetes. These include chronic pancreatitis, cystic
`fibrosis, pancreatic cancer, sequelae of partial pancrea-
`tectomy, hemochromatosis, and transfusion associated
`iron overload. It has been suggested that increased iron
`stores in general may lead to an increased propensity
`to T2DM [47], and that both chronic pancreatitis and
`pancreatic cancer may be associated with a significant
`component of insulin resistance that contributes to the
`hyperglycemia that may occur in these conditions [48,
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`49]. Future proteome based studies in these conditions
`may therefore shed new light on novel mechanisms for
`insulin resistance.
`
`5. Conclusion
`
`T1DM is caused by immune mediated beta cell de-
`struction. T2DM is caused by insulin resistance in con-
`junction with variable degrees of a defective beta cell
`response to hyperglycemia [50]. Proteome based stud-
`ies are likely to yield additional insight into the mech-
`anisms involved in pathophysiology of both disorders.
`Already, proteome based studies are contributing infor-
`mation about beta cell responses to insulin treatment
`and to inflammatory cytokines, as well as potential
`biomarkers of T2DM in skeletal muscle.
`
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`MPI EXHIBIT 1074 PAGE 5
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`MPI EXHIBIT 1074 PAGE 5
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