Human Insulin-dependent Diabetes Mellitus Research Articles

Large Intestinal Endocrine Cells in Non-Obese Diabetic Mice

Colonic endocrine cells from prediabetic and diabetic non-obese diabetic mice as well as of the sister strain, BALB/cJ, were investigated by immunocytochemistry and computer image analysis. In prediabetic mice, enteroglucagon- and serotonin-immunoreactive cells were significantly increased in number, whereas the cell secretory index of these two cell types was significantly reduced. No significant differences were found in numbers or cell secretory index of peptide YY (PYY)-immunoreactive cells. In diabetic mice, PYY-immunoreactive cells were significantly fewer, but there were no significant differences in the numbers of enteroglucagon- and serotonin-immunoreactive cells. Whereas the cell secretory index was reduced in serotonin-producing cells, no significant differences were found between diabetic and control mice regarding the cell secretory index of PYY- and enteroglucagon-immunoreactive cells. Nor was any statistically significant difference found between controls, prediabetic and diabetic non-obese diabetic mice, regarding the thickness of submucosa, of circular and longitudinal-muscle layers, or of the mucosal area/μm baseline. The present study showed that abnormalities in colonic endocrine cells do occur, in both prediabetic and diabetic mice, but they are different in nature and can be divided into primary and secondary to the diabetes onset. The present findings of abnormal colonic endocrine cells in non-obese diabetic mice, an animal model for human insulin-dependent diabetes mellitus, might help explain the gastrointestinal disorders observed in patients with diabetes. The study also showed that the change in the colonic endocrine cells is dynamic and started before the onset of the diabetic condition.

Effect of long-term treatment with a purified micronized flavonoid fraction on pancreatic mononuclear cell infiltration in diabetic BB rats.

Bio Breeding (BB) rats develop a genetically determined insulin-dependent diabetes, because of the early destruction of pancreatic beta cells of Langerhans islets, massively infiltrated by inflammatory mononuclear cells. S 5682, registered as Daflon, 500 mg, is a purified micronized flavonoid fraction (90% diosmin, 10% hesperidin), which has been shown to possess antiinflammatory properties, including anti-free radical activity, effects on vascular permeability, venous tone, and perivenous inflammation. We studied the effect of S 5682 on the course of pancreatic insulitis in diabetic BB rats. All the diabetic BB rats were hyperglycemic, with an increase of plasma levels of fructosamine, alpha-1 acid glycoprotein, and fibrinogen, and a dramatic decrease of C-peptide level. These parameters were not modified by S 5682. Pancreas histologic studies showed that in S 5682-treated diabetic BB rats, lymphocytic infiltration of Langerhans islets was less important and frequent than in untreated diabetic BB rats. By quantitative analysis, a highly significant difference was observed for insulitis, as well as perivasculitis, between S 5682-treated and untreated diabetic BB rats. This inhibitory effect of S 5682 on pancreatic mononuclear cell infiltration may be useful for a complementary treatment to decrease the development of insulitis in human insulin-dependent diabetes mellitus.

Induction of insulitis by glutamic acid decarboxylase peptide-specific and HLA-DQ8-restricted CD4(+) T cells from human DQ transgenic mice.

Insulin-dependent diabetes mellitus in humans is linked with specific HLA class II genes, e.g., HLA-DQA1*0301/ DQB1*0302 (DQ8). To investigate the roles of HLA-DQ8 molecules and glutamic acid decarboxylase (GAD) in disease development, we generated DQ8(+)/I-Abo transgenic mice expressing functional HLA-DQ8 molecules and devoid of endogenous mouse class II. DQ8(+)/I-Abo mice produced antigen-specific antibodies and formed germinal centers after immunization with GAD65 peptides. Two GAD peptide-specific (247-266 and 509-528), DQ8 restricted Th1 CD4(+) T cell lines, were generated from immunized DQ8(+)/I-Abo mice. They induced severe insulitis after adoptive transfer into transgene positive (but not negative) mice who were treated with a very low dose of streptozotocin that alone caused no apparent islet pathology. In addition to CD4, islet mRNA from these mice also showed expression of CD8, IFNgamma, TNFalpha, Fas, and Fas ligand. Our data suggest that a mild islet insult in the presence of HLA-DQ8 bearing antigen-presenting cells promotes infiltration of GAD peptide reactive T cells into the islet.

Contribution of T cells to the development of autoimmune diabetes in the NOD mouse model

The nonobese diabetic (NOD) mouse spontaneously develops an autoimmune diabetes that shares many immunogenetic features with human insulin-dependent diabetes mellitus (IDDM), type 1 diabetes. The mononuclear cell infiltrates in the islet, which results in the development of insulitis (a prerequisite step for the development of diabetes) are primarily composed of T cells. It is now well accepted that these T cells play important roles in initiating and propagating an autoimmune process, which in turn destroys insulin-producing islet β cells in the pancreas. T cells are subdivided into CD4+ helper T cells and CD8+ cytotoxic T cells. CD4+ T cells are further subdivided into Th1 and Th2 cells based on profiles of cytokine production, and these two T-cell populations counterregulate each other. Because many autoimmune diseases are Th1 T-cell mediated, current studies have focused on manipulating the Th1/Th2 imbalance to suppress the autoimmune process in the NOD model. Furthermore, the incidence of disease is much higher in females than that in males, suggesting an involvement of sex-steroid hormones in the development of diabetes. Understanding insights of the mechanism of immune-mediated islet cell destruction and the interaction between the immune and the neuroendocrine system may, therefore, provide new therapeutic means of preventing this chronic debilitating disease. BioEssays 20:750–757, 1998. © 1998 John Wiley & Sons, Inc.

Diabetes induced by Coxsackie virus: initiation by bystander damage and not molecular mimicry.

Viral induction of autoimmunity is thought to occur by either bystander T-cell activation or molecular mimicry. Coxsackie B4 virus is strongly associated with the development of insulin-dependent diabetes mellitus in humans and shares sequence similarity with the islet autoantigen glutamic acid decarboxylase. We infected different strains of mice with Coxsackie B4 virus to discriminate between the two possible induction mechanisms, and found that mice with susceptible MHC alleles had no viral acceleration of diabetes, but mice with a T cell receptor transgene specific for a different islet autoantigen rapidly developed diabetes. These results show that diabetes induced by Coxsackie virus infection is a direct result of local infection leading to inflammation, tissue damage, and the release of sequestered islet antigen resulting in the re-stimulation of resting autoreactive T cells, further indicating that the islet antigen sensitization is an indirect consequence of the viral infection.

Could myocarditis, insulin-dependent diabetes mellitus, and Guillain-Barré syndrome be caused by one or more infectious agents carried by rodents?

The numbers of small rodents in northern Sweden fluctuate heavily, peaking every 3 or 4 years. We found that the incidence of Guillain-Barré syndrome and insulin-dependent diabetes mellitus, as well as the number of deaths caused by myocarditis, followed the fluctuations in numbers of bank voles, although with different time lags. An environmental factor, such as an infectious agent, has been suggested for all three diseases. We hypothesize that Guillain-Barré syndrome, myocarditis, and insulin-dependent diabetes mellitus in humans in Sweden are caused by one or more infectious agents carried by small rodents. Also, a group of novel picornaviruses recently isolated from these small rodents is being investigated as the possible etiologic agent(s).

Aspects of the involvement of interleukin-1 and nitric oxide in the pathogenesis of insulin-dependent diabetes mellitus.

The possible involvement of the cytokine interleukin-1 (IL-1) and nitric oxide (NO) in the pathogenesis of insulin-dependent diabetes mellitus (IDDM) is reviewed and current and potential therapies are discussed. IDDM is a common disorder in the Western world and it is rising in incidence. In IDDM, islet-infiltrating macrophages produce IL-1 which is cytotoxic specifically to beta-cells in vitro. IL-1 increases beta-cell formation of NO, ceramide, prostaglandins, heat-shock proteins, and activates a protease. Additionally, IL-1 depresses beta-cell energy production, insulin gene expression and cyclic AMP synthesis, and impacts negatively on different parts of the insulin stimulus-secretion coupling, actions mimicked by NO. Conversely, blocking NO formation prevented many of these effects in most reports published. Also, changes in cyclic AMP and prostaglandins seem unlikely events in mediating the cytotoxicity of IL-1, while the role of ceramide remains less clear. Peptides capable of blocking beta-cell IL-1 receptors, and agents blocking NO synthesis may prove valuable in preserving beta-cell function in IDDM. Although IDDM causes immense morbidity and expense, uniformly effective preventive or beta-cell protective therapy is not currently available. If IL-1 is causing beta-cell dysfunction in human IDDM through NO production, several processes in the IL-1-NO connection are appropriate targets for agents protecting beta-cells from destruction and functional inhibition in IDDM.

Lack of recurrence of insulin-dependent diabetes mellitus in syngeneic and allogeneic islet-transplanted diabetic biobreeding rats.

Previous reports on experimental islet transplantation in animal models of human insulin-dependent diabetes mellitus show that islet grafts are susceptible to autoimmune destruction similar to that seen in native pancreatic islets. In this study, we demonstrated a lack of disease recurrence in diabetic BioBreeding (BB) rats after syngeneic and allogeneic islet transplantation. Four hundred to 1200 islets from BB (RT1u) and Lewis (RT1(1)) donors, isolated with stationary collagenase digestion and Ficoll density purification, were intraportally transplanted into spontaneously diabetic BB rats. The recipients received no immunologic manipulations before or after islet transplantation. When more than 900 syngeneic islets or when more than 600 allogeneic islets were transplanted, BB recipients remained normoglycemic for over 280 days, irrespective of age at onset, duration of exogenous insulin treatment, or age at transplantation. When at least 500 islets were transplanted, the recipients survived for a long period with normoglycemia or in a noninsulin-dependent diabetic state. Upon histological examination, mononuclear cell infiltration was observed in every islet graft examined, but the severity of infiltration in most of the grafts was mild to moderate. These results indicate that the islet grafts in the BB recipients were destroyed extremely slowly. It is conceivable that in our BB colony, a state of immunologically low responsiveness that allows diabetic animals to accept syngeneic or allogeneic islet grafts, occurs around the onset period and becomes more pronounced with aging. Our BB rat colony can be considered to be a novel substrain with unique immunological characteristics.

Alpha/beta-T cell receptor (TCR)+CD4-CD8- (NKT) thymocytes prevent insulin-dependent diabetes mellitus in nonobese diabetic (NOD)/Lt mice by the influence of interleukin (IL)-4 and/or IL-10.

We have previously shown that nonobese diabetic (NOD) mice are selectively deficient in alpha/beta-T cell receptor (TCR)+CD4-CD8- NKT cells, a defect that may contribute to their susceptibility to the spontaneous development of insulin-dependent diabetes mellitus (IDDM). The role of NKT cells in protection from IDDM in NOD mice was studied by the infusion of thymocyte subsets into young female NOD mice. A single intravenous injection of 10(6) CD4-/lowCD8- or CD4-CD8- thymocytes from female (BALB/c x NOD)F1 donors protected intact NOD mice from the spontaneous onset of clinical IDDM. Insulitis was still present in some recipient mice, although the cell infiltrates were principally periductal and periislet, rather than the intraislet pattern characteristic of insulitis in unmanipulated NOD mice. Protection was not associated with the induction of "allogenic tolerance" or systemic autoimmunity. Accelerated IDDM occurs after injection of splenocytes from NOD donors into irradiated adult NOD recipients. When alpha/beta-TCR+ and alpha/beta-TCR- subsets of CD4-CD8- thymocytes were transferred with diabetogenic splenocytes and compared for their ability to prevent the development of IDDM in irradiated adult recipients, only the alpha/beta-TCR+ population was protective, confirming that NKT cells were responsible for this activity. The protective effect in the induced model of IDDM was neutralized by anti-IL-4 and anti-IL-10 monoclonal antibodies in vivo, indicating a role for at least one of these cytokines in NKT cell-mediated protection. These results have significant implications for the pathogenesis and potential prevention of IDDM in humans.

Enterovirus infections and insulin dependent diabetes mellitus—evidence for causality

Background: Insulin-dependent diabetes mellitus (IDDM) has a long subclinical period characterised by gradually progressing autoimmune damage of insulin producing beta-cells. Clinical IDDM is manifested when 90% of beta-cells have been destroyed. Several studies have indicated that enterovirus infections, coxsackievirus B (CVB) infections especially, are frequent at the manifestation of clinical IDDM suggesting that they can precipitate the symptoms of IDDM in individuals who already have an advanced beta-cell damage. Recently, the first prospective studies have been published suggesting that enterovirus infections can also initiate the process several years before clinical IDDM. This implies that enterovirus infections may have a crucial role in the pathogenesis of human IDDM. Objective: The recent findings have brought up the question whether the time has come when a causal association between enterovirus infections and IDDM could finally be confirmed. This review focuses on this question summarising the current knowledge and the prospects of future research. Study design: Review of the recent progress in studies evaluating the role of enterovirus infections in human IDDM. Conclusions: The currently available information supports the assumption that the role of enterovirus infections may be more important than previously estimated. Enterovirus infections are obviously associated with increased risk of IDDM, but whether this association reflects causal relationship remains to be confirmed in future studies. Prospective birth-cohort studies will be among the most important ones giving important data on the etiologic fraction of enterovirus infections, the properties of diabetogenic virus variants and the mechanisms of beta-cell damage.

Protective and destructive effects of microbial infection in insulin-dependent diabetes mellitus

Insulin-dependent diabetes mellitus (IDDM) is a T-cell mediated autoimmune disease, which results in the destruction of the islet β-cells. The major histocompatibility complex (MHC) encodes the major susceptibility gene in IDDM. The concordance rate for diabetes in identical twins is 30–50sol; and in inbred animal models of disease the incidence rate is 20–80sol;. These results emphasize a role for environmental factors in the disease process. It has long been suggested that IDDM in humans may be caused by viral infections. While considerable progress has been made in defining the genetics of IDDM, our understanding of the role of environmental factors, which might provide a more direct approach to therapy is considerably lacking. We suggest that (1) the density and affinity of epitopes derived from microbial antigens that bind to MHC molecules; (2) their cross-reactivity with β-cell antigens; and (3) the nature of immunoregulatory cytokines induced by the microbial infections are the primary factors in the induction of either effector or protective T cells in IDDM.

Cytokines and IDDM: Implications for etiology and therapy

The evidence that human insulin-dependent diabetes mellitus (IDDM) is a T cell-mediated disease is well substantiated, and the use of transgenic technology to understand the Th1/Th2 paradigm will provide keys to attenuating pathogenic autoimmunity. Insofar as the role of Th1 cytokines in IDDM is concerned, interferon gamma is considered a critical player in the etiology, a proinflammatory role has been determined for IDDM, interleukin-2 is considered an "amplification" factor, and tumor necrosis factor-alpha presents dichotomous effects. Regarding the role of Th2 cytokines in IDDM, interleukin-4 is essential for immunoprotection and counterregulation of IDDM, and interleukin-10 plays immunoprotective and destructive roles. Therefore, Th1 and Th2 cytokines, when expressed individually in islets of Langerhans, have provided surprising results in manipulating the IDDM of transgenic NOD mice. The current data show that the same cytokine can produce either protective or pathological effects, depending upon the timing of its participation in the disease process. Of all the cytokines examined, IL-4 seems to be the likely candidate for preventing IDDM.

T-cell reactivity to wheat gluten in patients with insulin-dependent diabetes mellitus.

Dietary wheat gluten has been associated with the risk of diabetes in animal models of human insulin-dependent diabetes mellitus (IDDM). To evaluate the role of wheat gluten as a T cell antigen in human IDDM, we studied the cell-mediated immune response to wheat gluten in patients with IDDM and in control subjects. The cellular response to gluten was measured by the peripheral blood mononuclear cell (PBMC) proliferation test, and the results were expressed as a stimulation index (SI). We observed an enhanced cellular immune response to gluten (SI > or = 3) in seven of 29 patients with newly diagnosed IDDM (24.1%), in six of 39 patients with a longer duration of IDDM (15.4%), and in two of 37 non-diabetic controls (5.4%). Reactivity of T cells to gluten was associated with IDDM at diagnosis (P = 0.03), whereas patients with longer duration of IDDM did not differ from controls (P = 0.16). Responses of T cells to gluten were low in general: the median SI (range) was 2.0 (1-8.6) in patients with newly diagnosed IDDM and 1.5 (1-5.8) in control subjects (P = 0.03). Cellular responsiveness to gluten was not associated with HLA-DQB1 risk alleles for IDDM in patients. Although T cell responses to gluten were slightly increased in newly diagnosed patients the responsiveness was rare, and thus our results do not support a major role of gluten in the pathogenesis of human IDDM.

Autoimmune Diabetes: The Role of T Cells, MHC Molecules and Autoantigens

Type 1 diabetes (IDDM) is a T cell mediated autoimmune disease which in part is determined genetically by its association with major histocompatibility complex (MHC) class II alleles. The major role of MHC molecules is the regulation of immune responses through the presentation of peptide epitopes of processed protein antigens to the immune system. Recently it has been demonstrated that MHC molecules associated with autoimmune diseases preferentially present peptides of other endogenous MHC proteins, that often mimic autoantigen-derived peptides. Hence, these MHC-derived peptides might represent potential targets for autoreactive T cells. It has consistently been shown that humoral autoimmunity to insulin predominantly occurs in early childhood. The cellular immune response to insulin is relatively low in the peripheral blood of patients with IDDM. Studies in NOD mice however have shown, that lymphocytes isolated from pancreatic islet infiltrates display a high reactivity to insulin and in particular to an insulin peptide B 9-23. Furthermore we have evidence that cellular autoimmunity to insulin is higher in young pre-diabetic individuals, whereas cellular reactivity to other autoantigens is equally distributed in younger and older subjects. This implicates that insulin, in human childhood IDDM and animal autoimmune diabetes, acts as an important early antigen which may target the autoimmune response to pancreatic fi cells. Moreover, we observed that in the vast majority of newly diagnosed diabetic patients or individuals at risk for IDDM, T cell reactivity to various autoantigens occurs simultaneously. In contrast, cellular reactivity to a single autoantigen is found with equal frequency in (pre)-type 1 diabetic individuals as well as in control subjects. Therefore the autoimmune response in the inductive phase of IDDM may be targeted to pancreatic islets by the cellular and humoral reactivity to one β-cell specific autoantigen, but spreading to a set of different antigens may be a prerequisite for progression to destructive insulitis and clinical disease. Due to mimic epitopes shared by autoantigen(s), autologous MHC molecules and environmental antigens autoimmunity may spread, intramolecularly and intermolecularly and amplify upon repeated reexposure to mimic epitopes of environmental triggers.

Gamma interferon prevents diabetes in the BB rat.

The BB rat model of human insulin-dependent diabetes mellitus (IDDM) spontaneously develops diabetes through an autoimmune process. Gamma interferon (IFN-gamma) is thought to play an important pathogenic role. This study examined if IFN-gamma administration can, paradoxically, prevent diabetes in BB rats. Diabetes-prone BB rats were initially injected intraperitoneally with murine recombinant IFN-gamma (rIFN-gamma) at doses of 0.5 x 10(4) to 40 x 10(4) U three times a week for 6 weeks beginning at 35 days of age. The effects of altering the duration of treatment (2 to 6 weeks) and the age at which injections were initiated (45 through 65 days) were also assessed. rIFN-gamma administration prevented the development of diabetes in a dose-dependent manner. The optimal treatment condition resulted in a 9.1% incidence of diabetes versus a 90% incidence in control rats. This diabetes-sparing effect was long lasting and continued to 7 months of age. A 4- to 6-week course resulted in maximal inhibition. Treatment initiated as late as 55 days of age, when insulitis is already present, was effective in preventing diabetes. Islet inflammation was dramatically lower in rIFN-gamma- versus saline-injected rats (P < 0.01). Total leukocyte count and subpopulations of peripheral mononuclear cells were unaltered by rIFN-gamma. In summary, rIFN-gamma paradoxically and potently prevents diabetes in BB rats in a dose-dependent fashion by inhibiting islet inflammation. This diabetes-sparing effect occurs even when injections are initiated after evidence of the diabetic process is already present.

In vitro survival and RT6 expression kinetics in peripheral T cells of diabetes prone BB and Lewis rats

THE DIABETES PRONE BB (BB-DP) rat is one of the classical animal models of spontaneous auto-immune insulin-dependent diabetes mellitus (IDDM). In contrast to IDDM in the non-obese diabetic mouse and to human juvenile IDDM, disease susceptibility of BB-DP rats is tightly correlated to T cell lymphopenia that, moreover, coincides with an RT6 expression defect. 1 These experiments study comparatively the in vitro survival of T cells of different maturational stages of Lewis (LEW) and BB-DP rats, as well as investigate the kinetics of Thy-1 and RT6 expression. In the light of these data and those published previously, a hypothesis regarding the possible relations between lymphopenia and RT6 expression defect in BB-DP rats is presented.

Induction of insulin-dependent diabetes mellitus in PVG.RT1 u rats

THE BIOBREEDING (BB) rat strain is used to model human autoimmune insulin-dependent diabetes mellitus (IDDM). 1 The diabetes-resistant (DR) subline of the BB rat does not develop diabetes spontaneously, but is genetically predisposed to the disorder. 1 More than 90% of BB-DR rats become diabetic after combined treatment with a depleting monoclonal antibody (MAb) to remove RT6.1 + regulatory T cells and polyinosinic:polycytidylic acid (poly I:C), an inducer of interferons. 2 Lymph node T cells from diabetic BB-DR rats adoptively transfer IDDM to untreated histocompatible (RTI u) WAG- nu/nu rats. 3,4 Thymocytes from either normal or diabetic BB-DR rats also transfer IDDM provided that recipients are treated with anti-RT6.1 MAb to preclude maturation of RT6 + regulatory T cells from the transfused thymocytes. 5 We have previously observed that other rat strains that express the RT6.1 alloantigen, among them the YO (RTI u) and LEW (RT1 1) strains do not develop autoimmune diabetes when treated in the same way as BB-DR rats. 6 To investigate further if combination treatment with anti-RT6.1 MAb and poly I:C can induce autoimmunity in strains other than the BB, we studied major histocompatibility complex (MHC) congenic PVG.RTI u rats. These animals are phenotypically normal and free of spontaneous disease but reportedly develop IDDM after irradiation and thymectomy. 7

Alleviation of insulitis in NOD mice is associated with expression of transgenic MHC E molecules on primary antigen-presenting cells.

Major histocompatibility complex (MHC) class II genes are important in the pathogenesis of insulin-dependent diabetes mellitus (IDDM) both in the mouse and in man. The non-obese diabetic (NOD) mouse, which is a good model for human IDDM, has a particular MHC class II with an A complex consisting of A alpha d and the unique A beta g7 chain, as well as an absent E molecule due to a deletion in the Ea promoter region. Transgenic insertion of a functional Ea gene protects against insulitis and diabetes, but when the transgene expression is restricted to certain compartments of the immune system by deleting parts of the promoter region, the protection against insulitis is disrupted. We have analysed three promoter-mutated lines where one lacks expression on B cells and has a reduced expression on approximately 1/3 of the dendritic cells and macrophages (Sma), one lacks thymic cortical expression and has a slightly reduced B-cell expression (delta X), and one lacks expression in the thymic medulla, on macrophages, dendritic cells and about half of the B cells (delta Y). None of these lines is protected against insulitis, but Sma and delta X display a reduced intensity of insulitis, with an average of 10-15% of the islets infiltrated in each mouse, while delta Y resembles non-transgenic mice with 30-35% infiltrated islets. Bone-marrow chimeras between Sma and delta Y mice demonstrate that peripheral cells of Sma origin reduce insulitis significantly when developed in the delta Y host, while insulitis is enhanced when delta Y bone marrow is given to Sma mice. This shows that E expression on the primary antigen-presenting macrophages and dendritic cells is of crucial importance to the alleviation of insulitis.

A peptide-binding motif for I-A(g7), the class II major histocompatibility complex (MHC) molecule of NOD and Biozzi AB/H mice.

The class II major histocompatibility complex molecule I-A(g7) is strongly linked to the development of spontaneous insulin-dependent diabetes mellitus (IDDM) in non obese diabetic mice and to the induction of experimental allergic encephalomyelitis in Biozzi AB/H mice. Structurally, it resembles the HLA-DQ molecules associated with human IDDM, in having a non-Asp residue at position 57 in its beta chain. To identify the requirements for peptide binding to I-A(g7) and thereby potentially pathogenic T cell epitopes, we analyzed a known I-A(g7)-restricted T cell epitope, hen egg white lysozyme (HEL) amino acids 9-27. NH2- and COOH-terminal truncations demonstrated that the minimal epitope for activation of the T cell hybridoma 2D12.1 was M12-R21 and the minimum sequence for direct binding to purified I-A(g7) M12-Y20/K13-R21. Alanine (A) scanning revealed two primary anchors for binding at relative positions (p) 6 (L) and 9 (Y) in the HEL epitope. The critical role of both anchors was demonstrated by incorporating L and Y in poly(A) backbones at the same relative positions as in the HEL epitope. Well-tolerated, weakly tolerated, and nontolerated residues were identified by analyzing the binding of peptides containing multiple substitutions at individual positions. Optimally, p6 was a large, hydrophobic residue (L, I, V, M), whereas p9 was aromatic and hydrophobic (Y or F) or positively charged (K, R). Specific residues were not tolerated at these and some other positions. A motif for binding to I-A(g7) deduced from analysis of the model HEL epitope was present in 27/30 (90%) of peptides reported to be I-A(g7)-restricted T cell epitopes or eluted from I-A(g7). Scanning a set of overlapping peptides encompassing human proinsulin revealed the motif in 6/6 good binders (sensitivity = 100%) and 4/13 weak or non-binders (specificity = 70%). This motif should facilitate identification of autoantigenic epitopes relevant to the pathogenesis and immunotherapy of IDDM.

Dendritic cells in the autoimmune insulitis in NOD mouse models of diabetes.

Dendritic cells (DC) are thought to play important roles as antigen presenting cells in the autoimmune insulitis preceding insulin dependent diabetes mellitus (IDDM). Not only are DC the first cells appearing in the pancreatic inflammatory process in animal models of diabetes (NOD mouse1.2 and BB rat3.4), DC are also abundantly present in progressed human insulitis5. Although insulitis in the NOD mouse starts around 3 weeks, overt diabetes will not develop until an age of 20 weeks. The lag between occult insulitis and diabetes, which can also be observed in human IDDM, was always considered to reflect the time needed for accumulation of a sufficient number of destructive T cells in the islets.