Islet-infiltrating T cell clones from non-obese diabetic mice that promote or prevent accelerated onset diabetes

In humans and non-obese diabetic mice (NOD), insulin-dependent diabetes mellitus (IDDM) results from a spontaneous T cell-dependent autoimmune destruction of the insulin-producing pancreatic beta cells. Previous data suggest that a delicate balance between autoaggressive T cells and suppressor-type immune phenomena determine whether expression of autoimmunity is limited to insulitis or progresses to IDDM. To resolve the cellular basis of this intricate network of pathogenic CD4+ and CD8+ T cells and the role of T cells in suppressive immune phenomena. T cell clones were propagated directly from islets of NOD mice at the onset of insulitis. Insofar as insulitis, but not IDDM, is universal in NOD mice, we have screened for the in vivo effects of the islet-infiltrating T cell clones upon expression of IDDM, not insulitis. A CD4+ T cell clone, IS-3S7D, proliferates in response to islet antigen(s) and its transfer into prediabetic NOD mice promotes the rapid onset of IDDM. An interleukin 2 (IL 2)-dependent noncytolytic, V beta 11+ CD8+. T cell clones IS-2.15, prevents an accelerated onset diabetes in two distinct models. The present study, which documents the presence of CD4+ diabetogenic T cell clones and CD8+ T cell clones that dampen autoimmunity, gives tangible evidence that opposing autoimmune processes may determine whether an autoimmune-prone host develops frank disease.     

Protection from experimental autoimmune diabetes in the non-obese diabetic mouse with soluble interleukin-1 receptor

We have evaluated the effects of a treatment with soluble interleukin-1 receptor (sIL-1R) in the accelerated model of autoimmune diabetes induced by cyclophosphamide (CY) in the non-obese diabetic (NOD) mouse. Prior to the CY challenge (350 mg/kg body weight), female euglycemic NOD mice were randomly divided into three groups (A–C). Groups B and C were treated daily from 1 day before to 13 days after the CY challenge with sIL-1R at doses of 0.2 and 2 mg/kg body weight. Group A was treated with PBS. By 2 weeks after CY administration, an acute form of autoimmune diabetes with glycosuria, hyperglycemia and severe insulitis occurred in the majority (13/20, 65%) of the control mice (group A). In contrast, repeated injections with sIL-1R protected NOD mice from insulin-dependent diabetes mellitus (IDDM) development in a dose-dependent fashion; the incidence of IDDM was 53.3% (8/15) in the mice treated with 0.2 mg/kg and only 6.7% (1/15) in those treated with 2 mg/kg. However, none of the doses of the sIL-1R reduced the extent of insulitis in NOD mice. Importantly, the anti-diabetogenic property of sIL-1R may not involve major T cell function impairment; accordingly, in parallel experiments, splenic lymphoid cells from NOD mice not challenged with CY, but treated with 2 mg/kg sIL-1R for 5 consecutive days showed a normal distribution of mononuclear cell subsets and maintained their capacity to secrete interferon-γ and IL-2 and to proliferate in response to polyclonal mitogenic stimulation with concanavalin A.     

Intramuscular transfer of naked calcitonin gene-related peptide gene prevents autoimmune diabetes induced by multiple low-dose streptozotocin in C57BL mice

The imbalance of Th1/Th2 subsets is an important pathogenic mechanism for insulin-dependent diabetes mellitus (IDDM). Calcitonin gene-related peptide (CGRP) has been found to play important roles in the regulation of T lymphocytes. We hypothesize that exogenous CGRP administration during insulitis may modulate the balance of Th lymphocytes, thereby providing a therapeutic intervention for IDDM. We established CGRP gene transfer by naked plasmid injection into the skeletal muscles with electroporation enhancement. The effect of CGRP gene transfer on pathogenesis of IDDM was observedin autoimmune diabetic C57BL mice induced by multiple low-dose streptozotocin (MLDS) administration. The treatment significantly decreased morbidity of diabetes, ameliorated hyperglycemia and insulin deficiency, and inhibited lymphocyte infiltration into the islets, indicating the protection of beta cells against autoimmune destruction. CGRP gene transfer significantly inhibited T cell proliferation and secretion of the Th1 cytokine IFN-gamma, increased the level of the Th2 cytokine IL-10, but had no effect on IL-4 and TGF-beta1 secretion. CGRP gene transfer also decreased IL-12 and IFN-gamma levels in peritoneal effusion. Our results demonstrate that CGRP gene transfer selectively suppresses the pro-inflammatory Th1 subsets and promote anti-inflammatory Th2 subsets, resulting in amelioration of beta cell destruction and reduction of IDDM occurrence in mice with MLDS-induced diabetes.     

Deviation of pancreas-infiltrating cells to Th2 by interleukin-12 antagonist administration inhibits autoimmune diabetes

Nonobese diabetic (NOD) mice develop spontaneous insulin-dependent diabetes mellitus (IDDM), and the pancreas-infiltrating T cells invariably show a Th1 phenotype. We demonstrated here that the interleukin (IL)-12 antagonist (p40)₂ can deviate the default Th1 development of naive T cell receptor (TCR)-transgenic CD4⁺ cells to the Th2 pathway in vitro. Although (p40)₂ does not modify the cytokine profile of polarized Th1 cells, it prevents further recruitment of CD4⁺ cells into the Th1 subset. To study the involvement of Th1 and Th2 cells in the initiation and progression of IDDM, we targeted endogenous IL-12 by administration of (p40)₂ in NOD mice. (p40)₂ administration to NOD mice inhibits interferon-γ but not IL-10 production in response to lipopolysaccharide (LPS) or to the putative autoantigen IA-2. Serum immunoglobulin isotypes determined after (p40)₂ treatment indicate an increase in Th2 and a decrease in Th1 helper activity. Administration of (p40)₂ from 3 weeks of age onwards, before the onset of insulitis, results in the deviation of pancreas-infiltrating CD4⁺ but not CD8⁺ cells to the Th2 phenotype as well as in the reduction of spontaneous and cyclophosphamide-accelerated IDDM. After treating NOD mice with (p40)₂ from 9 weeks of age, when insulitis is well established, few Th2 and a reduced percentage of Th1 cells are found in the pancreas. This is associated with a slightly decreased incidence of spontaneous IDDM, but no protection from cyclophosphamide-accelerated IDDM. In conclusion, deviation of pancreas-infiltrating CD4⁺ cells to Th2 is associated with protection from IDDM. However, targeting IL-12 after the onset of insulitis, when the pancreas contains polarized Th1 cells, is not sufficient to induce an effective immune deviation able to significantly modify the course of disease.     

Pertussigen treatment retards, but fails to prevent, the development of type I, insulin-dependent diabetes mellitus (IDDM) in NOD mice

Current evidence supports an autoimmune etiopathogenesis for Type I, insulin-dependent diabetes mellitus (IDDM) in which the pancreatic beta (beta) cell is the specific target tissue. Recently, the NOD (non-obese diabetic) mouse has become an important model for IDDM, exhibiting many of the pathological features observed in man, including a progressive pancreatic islet leukocytic inflammation referred to as insulitis. The present study was carried out to determine the efficacy of the bacterial-derived bio-product, pertussigen, to retard the progression of insulitis and thereby prevent overt diabetes. Results revealed that (1) the rapid onset of IDDM in female NOD mice is absent if the mothers are treated with pertussigen prior to mating, (2) treatment of young prediabetic NOD mice with repeated injections of pertussigen results in the retardation of onset of IDDM when compared to untreated control NOD mice, and (3) the severity of insulitis in pertussigen-treated NOD mice not developing IDDM was noticeably less severe than age and sex-matched untreated control mice. Since earlier work had shown that pertussis vaccine, which contains pertussigen, could prevent development of IDDM in mice treated with streptozotocin, the present results may indicate basic differences in the inflammatory responses in the genetically-predisposed NOD mice and IDDM-nonsusceptible mice with streptozotocin-induced diabetes.     

Non-obese diabetic mice hemizygous at the T cell receptor α locus are susceptible to diabetes and sialitis

To test the hypothesis that T cells carrying two T cell receptor (TCR)α chains play a role in autoimmunity, we backcrossed the non-obese diabetic (NOD) strain with one carrying a TCRα gene disrupted by homologous recombination. Mice carrying one copy of the disrupted gene are incapable of generating T cells carrying two cell surface TCRα chains. Our early results suggested that either dual TCRα T cells play a role in insulin-dependent diabetes mellitus (IDDM) induction in NOD mice or that a locus co-segregating with the disrupted TCRα locus protected mice from diabetes induction. From the analysis both of mice in which the region co-segregating with the disrupted TCRα locus is minimized and of the F₁ offspring of NOD mice with the 129 strain (TCRα hemizygous mice), the apparent protective effect of the absence of dual TCRα T cells is lost; thus, such cells do not appear to play a critical role in autoimmune disease in NOD mice.     

T cell receptor beta chain polymorphisms are associated with insulin-dependent diabetes.

We investigated the T cell receptor constant beta chain (TCR C beta) genes of patients with insulin dependent diabetes mellitus (IDDM) using DNA restriction fragment length polymorphism (RFLP) analysis. Genomic DNA from patients and controls was digested with the restriction endonuclease Bg1 II and transferred to nylon filters using the Southern blot procedure. This enzyme identifies a polymorphic site between the two TCR C beta chain genes. We have found a highly significant increase in the frequency of the 10.0; 9.2 kilobase heterozygous phenotype in patients with IDDM (62.7% versus 42.0% in normal controls; P = 0.0006). In identical twin pairs, this association was most striking in those concordant for IDDM (79.2% versus 42.0% in controls; P = 0.0006).     

Suppression of insulitis and diabetes in B cell-deficient mice treated with streptozocin: B cells are essential for the TCR clonotype spreading of islet-infiltrating T cells

In order to clarify the role of B cells in the development of insulitis and diabetes, B cell-deficient (B(-)) mice treated with streptozocin (STZ) were studied. The extent of insulitis and the cumulative incidence of diabetes were significantly suppressed in B(-) mice (P < 0.0001), indicating that B cells are crucial for the progression of insulitis and diabetes. Accumulation of both CD4(+) T cells and B cells was observed in islets of B(+) mice, while CD4(+) T cells but not B cells were found in B(-) mice. A few CD8(+) T cells and macrophages were detectable in both types of mice. The immunohistochemical study did not reveal any change in the subpopulations of infiltrating lymphocytes except for the absence of B cells in the B(-) mice. TCR V(beta) gene repertoire usage of islet-infiltrating T cells was restricted to some extent in the B(+) or B(-) mice, but there was no significant difference between the B(+) and B(-) mice, suggesting that the initial islet-reactive T cell response can occur in the absence of B cells. In contrast, TCR clonotype spreading of islet-infiltrating T cells was significantly suppressed in B(-) mice compared with B(+) mice (P < 0.0001). These data suggest that initial priming of T cells is not impaired and TCR V(beta) repertoire usage is not limited by the lack of B cells, while B cells are important essentially for the spreading of islet-infiltrating clonal T cells in autoimmune diabetic mice induced with STZ.     

Prevention of diabetes in NOD mice treated with antibody to murine IFN gamma

The NOD mouse is studied as an animal model of human insulin-dependent diabetes mellitus (IDDM). To evaluate the role of IFN gamma in the pathogenesis of the disease, we have studied the effect of anti-IFN gamma mAb on the expression of insulitis and clinical diabetes. Treatment of mice with anti-IFN gamma mAb prevented the induction of early IDDM by cyclophosphamide as well as the adoptive transfer of diabetes by spleen cells from diabetic NOD mice. The protection against induction of diabetes by cyclophosphamide was observed in animals treated with the anti-IFN gamma mAb within 24 h following the first cyclophosphamide injection but not in animals in which mAb treatment was started 7 days later. Transfer of disease was prevented both in adult irradiated and in newborn recipients. The absence of clinical signs in these mice was corroborated by a significant reduction of both the extent and severity of insulitis. Over-expression of Ia antigen on endothelial cells lining the islets was also considerably reduced in mice treated with mAb. These data strongly suggest a role for IFN gamma during the autoimmune process leading to beta cell destruction in diabetes and prompt further investigation of the use of such antibodies in the immunoprevention of IDDM.     

The Effect of Cytokines on Expression of Glutamic Acid Decarboxylase-65 in Cultured Islets

Insulin dependent diabetes mellitus (IDDM) is an autoimmune disease characterized by lymphocytic infiltration of the pancreatic islets (insulitis). Cytokines released as part of the insulitis process have been suggested to play an important role in the beta cell lesion of IDDM. A possible diabetogenic effect of cytokines may be mediated by their inducing abnormal expression of islet cell autoantigens. Since glutamic acid decarboxylase-65 (GAD-65) is a target autoantigen in IDDM, we investigated whether the cytokines IL-1β, TNFα IFNγ altered islet cell expression of GAD-65 and whether the effect of cytokines on GAD-65 expression was similar to their effect on insulin secretion.

We found that: 1) IL-1β at low dose (1 U/ml) which stimulated insulin secretion, had no effect on GAD-65 expression, whereas higher doses of IL-1β (10, 100, 1000 U/ml) which inhibited insulin secretion, decreased GAD-65 expression. 2) TNFα at doses of 10, 100, 1000 U/ml which stimulated insulin secretion had no effect on GAD-65 expression. 3) IFNγ at doses of 10, 100, 1000 U/ml had no effect on insulin secretion or on GAD-65 expression. 4) In combination, IL-1β plus TNFα and IFNγ showed a similar inhibitory effect on GAD-65 expression as IL-1β alone.

In summary: 1) IL-1β dramatically inhibits GAD-65 expression. 2) TNFα and IFNγ have no effect on GAD-65 expression. Of these three cytokines, IL-1β is the primary cytokine affecting GAD-65 expression.     

The role of CD8+ T cells in the initiation of insulin-dependent diabetes mellitus

While it is generally accepted that T cells are critical for the development of diabetes in the non-obese diabetic (NOD) mouse, the precise functions of the CD4⁺ and CD8⁺ subsets remain ill-defined. Transfer experiments have provided evidence that CD4⁺ cells are the disease initiators, provoking massive mononuclear leukocyte infiltration into the pancreatic islets, while CD8⁺ cells play an effector role, responsible for the final destruction of islet beta cells. It was surprising, then, to find that NOD mice carrying a null mutation at the β2-microglobulin (β2-μ) locus, and thereby lacking major histocompatibility complex class I molecules and CD8⁺ T cells, developed neither insulitis nor diabetes. Here, we argue that the absence of insulitis in these animals results from their lack of CD8⁺ cells because islet infiltration is also absent when NOD mice are treated with an anti-CD8 monoclonal antibody (mAb) at a young age. Interestingly, the anti-CD8 effect is only observed when the mAb is injected during a discrete age window – 2 to 5 weeks after birth. Transfer experiments indicate that the lack of CD8⁺ cells during this period somehow alters the phenotype of CD4⁺ cells, preventing them from expressing their insulitic potential. This is not because they are generally immuno-incompetent nor because they are generally more prone to differentiating into cells with Th2 characteristics. Given that neither the β2-μ mutation nor anti-CD8 treatment affect insulitis in a T cell receptor transgenic (tg) mouse strain with a CD4⁺ T cell repertoire highly skewed for an anti-islet cell reactivity, the most straight-forward interpretation of these observations is that CD8⁺ cells are required for effective priming and expansion of autoreactive CD4⁺ cells.     

Inhibition of autoimmune diabetes in NOD mice with serum from streptococcal preparation (OK-432)-injected mice.

We have recently reported that systemic and chronic administration of recombinant tumour necrosis factor alpha (TNF-alpha), as well as streptococcal preparation (OK-432), inhibits development of insulin-dependent diabetes mellitus (IDDM) in NOD mice and BB rats, models of IDDM. In this study we examined whether serum containing endogenous TNF induced by OK-432 injection could inhibit IDDM in NOD mice. Treatment twice a week from 4 weeks of age with OK-432-injected mouse serum, which contained endogenous TNF (75U), but not IL-1, IL-2 and interferon-gamma (IFN-gamma) activity, reduced the intensity of insulitis and significantly inhibited the cumulative incidence of diabetes by 28 weeks of age in NOD mice, as compared with the incidence in non-treated mice (P less than 0.01) and in mice treated with control serum (P less than 0.02). This inhibitory effect of the serum was diminished, although not significantly, by neutralization of serum TNF activity with anti-mouse TNF antibody. In the mice treated with the serum from OK-432-injected mice, Thy-1.2+ or CD8+ spleen cells decreased (P less than 0.01) and surface-Ig+ (S-Ig+) cells increased (P less than 0.05), whereas the proliferative response of spleen cells to concanavalin A (P less than 0.01) and lipopolysaccharide (P less than 0.05) increased. The results indicate that the inhibition by OK-432 treatment of IDDM in NOD mice was partially mediated by serum factors including endogenous TNF.     

A shared TCR CDR3 sequence in NOD mouse autoimmune diabetes.

T cells involved in autoimmune diseases have been characterized by the genetic elements used to construct their autoimmune TCR. In the present study, we sequenced the alpha and beta chains of the TCR expressed by a CD4(+) T cell clone, C9, functional in NOD mouse diabetes. Clone C9 can adoptively transfer diabetes or, when attenuated, C9 can be used to vaccinate NOD mice against diabetes. Clone C9 recognizes a peptide epitope (p277) of the 60 kDa heat shock protein (hsp60) molecule. We now report that the C9 TCR beta chain features a CDR3 peptide sequence that is prevalent among NOD mice. This CDR3 element is detectable by 2 weeks of age in the thymus, and later in the spleen and in the autoimmune insulitis. Thus, a TCR CDR3beta sequence appears to be a common idiotope associated with mouse diabetes.     

Anti-suppressor effect of cyclophosphamide on the development of spontaneous diabetes in NOD mice

In the NOD mouse, an autoimmune process beginning by 5 weeks of age with lymphocyte infiltration and destruction of insulin-secreting beta cells leads to overt diabetes which begins to appear by 11 weeks of age. Although there is a high incidence of insulitis by 10 weeks of age (greater than 80%) in both males and females, by 30 weeks of age diabetic symptoms have occurred in 53-80% of females and in 12-40% of males. Intraperitoneal injection of a high dose (200 mg/kg) of cyclophosphamide (CY) consistently induces the onset of diabetes in male and female NOD mice at an age when spontaneous diabetes rarely occurs. Spleen T cells from CY-induced diabetic mice are capable of transferring the disease into irradiated nondiabetic syngeneic recipients. This indicates that the diabetogenic effect of CY is not mediated by direct toxicity on pancreatic beta cells but is mediated by abrogation of a suppressor mechanism which may prevent activation of T cells responsible for the development of diabetes in the NOD mouse. Additionally, CY is only effective in NOD mice and not in F1 hybrids between NOD and other strains of mice. Thus, the potential beta cell aggressor mechanism is not present in these hybrids as it is in homozygous mice, which indicates that it is not under the control of dominant genes.     

BETA-CELL APOPTOSIS IS RESPONSIBLE FOR THE DEVELOPMENT OF IDDM IN THE MULTIPLE LOW-DOSE STREPTOZOTOCIN MODEL

Although insulin-dependent diabetes mellitus (IDDM) results from irreversible loss of beta cells, the mode of cell death responsible for this loss has not previously been categorized. In this study, the multiple low-dose streptozotocin (stz) model (intraperitoneal injection of stz at a concentration of 40 mg/kg body weight per day for five consecutive days) was used to investigate beta-cell death during the development of IDDM in male C57B1/6 mice. Apoptotic cells were evident by light microscopy within the islets of Langerhans of treated animals from day 2 (the day of the second stz injection) until day 17. Immunohistochemical localization of insulin to the dying cells confirmed the beta-cell origin of the apoptosis. Two peaks in the incidence of beta-cell apoptosis occurred: the first at day 5, which corresponded to an increase in blood glucose concentration, and the second at day 11, when lymphocytic infiltration of the islets (insulitis) was maximal. Insulitis did not begin until day 9, by which time treated animals had developed overt diabetes as revealed by blood glucose and pancreatic immunoreactive insulin (IRI) measurements. Beta-cell apoptosis preceded the appearance of T-cells in the islets and continued throughout the period of insulitis. Thus, whether induced by stz or a subsequent immune response, apoptosis is the mode of cell death responsible for beta-cell loss in the multiple low-dose stz model of IDDM.     

Peptide-specific amelioration of T cell-mediated pathogenesis in murine type 1 diabetes

NOD mice spontaneously develop insulitis and type 1 diabetes (T1D) mellitus similar to humans. Insulitis without overt disease occurs in the BDC2.5 TCR-transgenic NOD mice that express the rearranged TCR alpha- and beta-chain genes of a diabetogenic T cell clone reactive to an unknown beta cell autoantigen. A previous study identified an extensive panel of peptides that are highly active in stimulating T cells from transgenic BDC2.5 mice in culture. However, none of these peptides cause active disease in NOD and BDC2.5 animals or in NOD recipients of adoptively transferred BDC2.5 T cells following direct immunization in vivo. We show that direct immunization of transgenic BDC2.5 mice causes many BDC2.5 T cells to become activated and apoptotic. Strikingly, soluble peptides administered to recipients of activated, highly pathogenic BDC2.5 T cells results in protection from disease. These results suggest that high affinity peptide analogues of autoimmune epitopes might be useful as therapeutic modulators in active autoimmune disease.     

Viral infection induces cytokine release by beta islet cells.

Viral infection has been suggested to play a triggering role in the pancreatic beta cell destruction which occurs in insulin-dependent diabetes (IDDM). However, the underlying mechanism of this phenomenon is unknown. In this study a human insulinoma cell line has been infected with measles, mumps and rubella viruses since a temporal association is reported between the clinical onset of IDDM and diseases caused by these viruses. The infection with measles and mumps viruses induced the release of interleukin-1 (IL-1) and interleukin-6 (IL-6) by the cell line as assessed by a bioassay and up-regulated the expression of human leucocyte antigen (HLA) class I and class II antigens as evaluated by cytofluorimetric analysis. Stimulation with rubella virus induced the release of IL-6 only and had no effect on HLA antigen expression. These data show for the first time that IL-1 and IL-6 secretion by an insulinoma cell line may occur after viral infection and suggest that cytokine release and increased expression of HLA molecules by beta cells may act to induce the immune response towards beta cells in IDDM.     

Effects of ginkgolide B, a platelet-activating factor inhibitor on insulitis in the spontaneously diabetic BB rat.

The BB rat spontaneously develops insulin-dependent diabetes mellitus (IDDM) in association with marked insulitis in the islet of Langerhans. Since platelet-activating factor (PAF-acether) is involved in allergic and inflammatory reactions, we tested a PAF antagonist, Ginkgolide B (BN 52021) for potential effects on islet inflammation and diabetes. Diabetes prone BB/Wor rats were treated daily from weaning at 25 days until 105 days of age with either saline (n = 30, controls), 10 (n = 25, low dose) or 20 (n = 30, high dose) mg/kg body weight of BN 52021. The overall incidence of IDDM was unaffected by treatment. Quantitative analysis of insulin area showed a dose-dependent protection of beta cells by Ginkgolide B, reflected in a 6- (low dose) to 8-fold (high dose) (P less than 0.01-0.005) increase in the insulin/glucagon cell ratio compared to the saline treated rats. Ginkgolide B reduced severe insulitis from 84% in the saline rats developing IDDM to 59% (n.s.) in the low and to 33% (P less than 0.001) in the high dose group. These data suggest that PAF inhibitors may prove useful in immunomodulator therapy of IDDM since beta cells are preserved.     

Intra-islet proliferation of cytotoxic T lymphocytes contributes to insulitis progression

Infiltration of pancreatic islets by immune cells, termed insulitis, increases progressively once it begins and leads to clinical type 1 diabetes. But even after diagnosis some islets remain unaffected and infiltration is patchy rather than uniform. Traffic of autoreactive T cells into the pancreas is likely to contribute to insulitis progression but it could also depend on T-cell proliferation within islets. This study utilizes transgenic NOD mice to assess the relative contributions of these two mechanisms. Progression of insulitis in NOD8.3 TCR transgenic mice was mildly reduced by inhibition of T-cell migration with the drug FTY720. In FTY720-treated mice, reduced beta cell MHC class I expression prevented progression of insulitis both within affected islets and to previously unaffected islets. CTL proliferation was significantly reduced in islets with reduced or absent beta cell expression of MHC class I protein. This indicates that intra-islet proliferation, apparently dependent on beta cell antigen presentation, in addition to recruitment, is a significant factor in progression of insulitis.     

Escape from self-tolerance leads to neonatal insulin-dependent diabetes mellitus.

Double transgenic (dTg) mice expressing the hemagglutinin (HA) of influenza virus under the insulin promoter and the TCR specific for the immunodominant CD4 T cell epitope of HA (HA110-120) develop insulin-dependent diabetes mellitus (IDDM). In order to gain information on the breaking down of neonatal self-tolerance we studied the occurrence of IDDM after birth. Our results showed that newborn mice develop fulminant IDDM characterized by occurrence of insulitis as early as 3 days after birth, followed by hyperglycemia by 7 days, and significant hypoinsulinemia by 28 days. The neonatal breakdown of self-tolerance of T cells positively selected in the thymus is supported by the facts that: (i) peripheral HA110-120 specific T cells from neonates are fully functional and proliferated upon stimulation with the nominal peptide, and (ii) peptide-specific T cells were accumulated in the pancreas of dTg mice as early as 3 days after birth. Our results demonstrate that diabetes occurring in young dTg mice is due to early activation of self-reactive T cells immediately after birth. Accumulation of specific T cells in the target organ leads to destruction of pancreatic beta-cells and IDDM. These mice may provide a useful model to evaluate new strategies for the prevention of diabetes.