In vivo diabetogenic action of CD4+ T lymphocytes requires Fas expression and is independent of IL-1 and IL-18

CD4(+) T lymphocytes are required to induce spontaneous autoimmune diabetes in the NOD mouse. Since pancreatic β cells upregulate Fas expression upon exposure to pro-inflammatory cytokines, we studied whether the diabetogenic action of CD4(+) T lymphocytes depends on Fas expression on target cells. We assayed the diabetogenic capacity of NOD spleen CD4(+) T lymphocytes when adoptively transferred into a NOD mouse model combining: (i) Fas-deficiency, (ii) FasL-deficiency, and (iii) SCID mutation. We found that CD4(+) T lymphocytes require Fas expression in the recipients' target cells to induce diabetes. IL-1β has been described as a key cytokine involved in Fas upregulation on mouse β cells. We addressed whether CD4(+) T cells require IL-1β to induce diabetes. We also studied spontaneous diabetes onset in NOD/IL-1 converting enzyme-deficient mice, in NOD/IL-1β-deficient mice, and CD4(+) T-cell adoptively transferred diabetes into NOD/SCID IL-1β-deficient mice. Neither IL-1β nor IL-18 are required for either spontaneous or CD4(+) T-cell adoptively transferred diabetes. We conclude that CD4(+) T-cell-mediated β-cell damage in autoimmune diabetes depends on Fas expression, but not on IL-1β unveiling the existing redundancy regarding the cytokines involved in Fas upregulation on NOD β cells in vivo.     

Targeted expression of the anti-apoptotic gene CrmA to NOD pancreatic islets protects from autoimmune diabetes

The activation of apoptosis is a critical mechanism by which pancreatic beta cells are destroyed in type 1 diabetes (T1DM). Strategies aimed at interfering with the apoptotic pathways could therefore be of potential therapeutic value. To this end, we generated NOD transgenic mice with targeted expression of the anti-apoptotic gene Cytokine response modifier A (CrmA) to pancreatic beta cells using the rat insulin promoter and the reverse tetracycline transactivator to express CrmA in a temporally controlled manner. Two lines of transgenic mice were studied whose expression of CrmA occurred only after feeding doxycycline food. Islet expression of CrmA partially protected pancreatic beta cells from the cytokine-mediated cytotoxicity in vitro and reduced modestly the spontaneous development of diabetes in NOD mice in vivo. In addition, beta cells from NOD CrmA mice were significantly protected from the destruction by diabetogenic T cells after adoptive transfer. More strikingly, NODCrmA mice were significantly resistant to the diabetogenic activity of a potent insulin-specific CD8 T-cell clone. Since these adoptive transfer models mainly represent the effector phase rather than the initiation phase of autoimmune diabetes, our data suggest that the latter is more sensitive to CrmA protection. We conclude that anti-apoptotic genes such as CrmA might be potential candidates to enhance islet graft survival in T1DM.     

Mechanisms of PDL1-mediated regulation of autoimmune diabetes

The PD-1-PDL1 pathway plays a critical role in regulating autoimmune diabetes as blockade or deficiency of PD-1 or PDL1 results in accelerated disease in NOD mice. We explored the cellular mechanisms involved in the regulation of these autoimmune responses by investigations involving various gene-deficient mice on the NOD background. Administration of blocking anti-PDL1 antibody to CD4+ T cell-deficient, CD8+ T cell-deficient and B cell-deficient mice demonstrated that PDL1-mediated regulation of autoreactive CD4+ and CD8+ T cells is critical for diabetes development. This concept was confirmed by adoptive transfer studies utilizing lymphocytes from BDC2.5 and 4.1 (CD4+) TCR transgenic mice and 8.3 (CD8+) TCR transgenic mice; efforts showing increased proliferation of both CD4+ and CD8+ T cells following PDL1 blockade in vivo. Furthermore, we observed that anti-PDL1-mediated acceleration is dependent upon events occurring in the pancreatic lymph nodes during early disease stages, but becomes independent of the pancreatic lymph nodes during later disease stages. These data provide strong evidence that PDL1 regulates autoimmune diabetes by limiting the expansion of CD4+ and CD8+ autoreactive T cells, and define the timing and locale of PDL1-mediated regulation of type 1 diabetes.     

Role of L-selectin in the development of autoimmune diabetes in non-obese diabetic mice

Autoimmune diabetes is characterized by an early mononuclear infiltration of pancreatic islets and later selective autoimmune destruction of insulin-producing beta cells. Lymphocyte homing receptors have been considered candidate targets to prevent autoimmune diabetes. L-selectin (CD62L) is an adhesion molecule highly expressed in naive T and B cells. It has been reported that blocking L-selectin in vivo with a specific antibody (Mel-14) partially impairs insulitis and diabetes in autoimmune diabetes-prone non-obese diabetic (NOD) mice. In the present study we aimed to elucidate whether genetic blockade of leukocyte homing into peripheral lymph nodes would prevent the development of diabetes. We backcrossed L-selectin-deficient mice onto the NOD genetic background. Surprisingly NOD/L-selectin-deficient mice exhibited unaltered islet mononuclear infiltration, timing of diabetes onset and cumulative incidence of spontaneous diabetes when compared to L-selectin-sufficient animals. CD4, CD8 T cells and B cells were present in islet infiltrates from 9-week-old L-selectin-sufficient and -deficient littermates. Moreover, total splenocytes from wild-type, heterozygous or NOD/L-selectin-deficient donor mice showed similar capability to adoptively transfer diabetes into NOD/SCID recipients. On the other hand, homing of activated, cloned insulin-specific autoaggressive CD8 T cells (TGNFC8 clone) is not affected in NOD/L-selectin-deficient recipients. We conclude that L-selectin plays a small role in the homing of autoreactive lymphocytes to regional (pancreatic) lymph nodes in NOD mice.     

Apoptosis of pancreatic beta-cells detected in accelerated diabetes of NOD mice: no role of Fas-Fas ligand interaction in autoimmune diabetes

Autoreactive T lymphocytes probably cause pancreatic beta-cell death by inducing apoptosis. To visualize apoptotic beta-cells in vivo, we accelerated diabetes of NOD mice with cyclophosphamide (CY) or adoptive transfer. We also studied whether Fas-mediated apoptosis is involved in the development of diabetes by administrating anti-Fas ligand (FasL) Ab and by grafting Fas-deficient neonatal pancreas from NOD-lpr/lpr mice. Apoptotic cells were clearly shown 8 days after CY treatment. Beta-cell apoptosis was also observed after adoptive transfer but in a different kinetic pattern. Anti-FasL Ab administration failed to inhibit diabetes after CY treatment or adoptive transfer, while it inhibited Con A-induced hepatitis. Fas-deficient neonatal pancreata were destroyed by lymphocytic infiltration in diabetic NOD mice. Our results clearly demonstrate apoptosis of beta-cells in accelerated diabetes and indicate that Fas-FasL interaction is not involved in diabetes of NOD mice, contrary to the previous reports.     

Critical roles of CD30/CD30L interactions in murine autoimmune diabetes

CD30/CD30L is a member of tumour necrosis factor (TNF) receptor/TNF superfamily and has been implicated in immune-regulation. A genetic study has also suggested a possible implication of CD30 in spontaneous autoimmune diabetes in NOD mice. In this study, we investigated the involvement of CD30/CD30L in the development of diabetes in NOD mice. Flow cytometric analysis showed that CD30 and CD30L were highly expressed on CD4+ or CD8+ T cells in the spleen and pancreatic lymph node of younger NOD mice. In addition, islet-specific CD4+ or CD8+ T cell lines expressed CD30 and CD30L. Administration of a neutralizing anti-CD30L monoclonal antibody (mAb) from 2 to 10 week of age completely suppressed the development of spontaneous diabetes in NOD mice. In addition, the treatment with anti-CD30L mAb also inhibited the development of diabetes induced by adoptive transfer of spleen cells from diabetic NOD mice or islet-specific CD4+ or CD8+ T cell lines into NOD-SCID mice. Furthermore, anti-CD30L mAb inhibited T cell proliferation in response to islet antigens. These results suggested that CD30/CD30L interaction plays important roles in both induction and effector phases of autoimmune diabetes in NOD mice.     

CD4+CD25+ regulatory T cells generated in response to insulin B:9-23 peptide prevent adoptive transfer of diabetes by diabetogenic T cells

NOD mice have a relative deficiency of CD4+CD25+ regulatory T cells that could result in an inability to maintain peripheral tolerance. The aim of this study was to induce the generation of CD4+CD25+ regulatory T cells in response to autoantigens to prevent type 1 diabetes (T1D). We found that immunization of NOD mice with insulin B-chain peptide B:9-23 followed by 72 h in vitro culture with B:9-23 peptide induces generation of CD4+CD25+ regulatory T cells. Route of immunization has a critical role in the generation of these cells. Non-autoimmune mice BALB/c, C57BL/6 and NOR did not show up regulation of CD4+CD25+ regulatory T cells. These cells secreted large amounts of TGF-beta and TNF-alpha with little or no IFN-gamma and IL-10. Adoptive transfer of these CD4+CD25+ regulatory T cells into NOD-SCID mice completely prevented the adoptive transfer of disease by diabetogenic T cells. Although, non-self antigenic OVA (323-339) peptide immunization and in vitro culture with OVA (323-339) peptide does result in up regulation of CD4+CD25+ T cells, these cells did not prevent transfer of diabetes. Our study for the first time identified the generation of antigen-specific CD4+CD25+ regulatory T cells specifically in response to immunization with B:9-23 peptide in NOD mice that are capable of blocking adoptive transfer of diabetes. Our results suggest the possibility of using autoantigens to induce antigen-specific regulatory T cells to prevent and regulate autoimmune diabetes.     

CD8+ T lymphocytes specific for glutamic acid decarboxylase 90-98 epitope mediate diabetes in NOD SCID mouse

During the past decade, glutamic acid decarboxylase (GAD) has been considered a crucial beta-cell autoantigen involved in type 1 diabetes in the NOD mouse and human. Recently, the etiological role of GAD has remained controversy. In the NOD mouse, some previous studies argued in favor of a regulatory role for GAD-specific CD4+ T cells, and no diabetogenic CD8+ T cells specific for GAD have been identified so far, discrediting the importance of GAD in beta-cell injury. Here, we identified, in the NOD model, a relevant GAD CD8+ T cell epitope (GAD(90-98)) using immunization with a plasmid encoding GAD, a protocol relying on in vivo processing of peptides from the autoantigenic protein. In pancreatic lymph nodes of na?ve female NOD mice, CD8+ T lymphocytes recognizing GAD(90-98) peptide were detected during the initial phase of invasive insulitis (between 4 and 8 weeks of age), suggesting an important role for these cells in the first stage of the disease. GAD(90-98) specific CD8+ lymphocytes lysed efficiently islet cells in vitro and transferred diabetes into NOD(SCID) mice (100%). Finally, diabetes was accelerated greatly in 3-week-old female NOD mice injected i.p. with GAD(90-98), strengthening the role of GAD-specific CTLs in diabetes pathogenesis.     

Immunomodulation of the Anti-Islet CD8 T Cell Response by B7-2

The absence of diabetes in NOD mice devoid of B7-2 signifies a critical role played by B7-2 in promoting autoimmunity. We asked whether the CD8 T cell compartment is impacted by the absence of B7-2. We found significantly lower expansion of anti-islet CD8 T cells in B7-2KO mice, although their survival and activation states remained unchanged in the pancreatic lymph nodes (PLNs). CD8 T cells from B7-2KO mice exhibited significantly diminished effector function compared to NOD mice. Adoptive transfer experiments using in vitro activated anti-islet CD8 T cells showed that B7-2 does not control the effector phase of the autoreactive CD8 T cell response. Our data indicate that B7-2 promotes pancreatic autoimmunity by controlling CD8 T cell expansion and effector function, but is dispensable for CD8 T cell activation, survival, and the effector phase of anti-islet CD8 T cell response.     

Stable activity of diabetogenic cells with age in NOD mice: dynamics of reconstitution and adoptive diabetes transfer in immunocompromised mice

The non-obese diabetic (NOD) mouse is a prevalent disease model of type 1 diabetes. Immune aberrations that cause and propagate autoimmune insulitis in these mice are being continually debated, with evidence supporting both dominance of effector cells and insufficiency of suppressor mechanisms. In this study we assessed the behaviour of NOD lymphocytes under extreme expansion conditions using adoptive transfer into immunocompromised NOD.SCID (severe combined immunodeficiency) mice. CD4⁺ CD25⁺ T cells do not cause islet inflammation, whereas splenocytes and CD4⁺ CD25⁻ T cells induce pancreatic inflammation and hyperglycaemia in 80–100% of the NOD.SCID recipients. Adoptively transferred effector T cells migrate to the lymphoid organs and pancreas, proliferate, are activated in the target organ in situ and initiate inflammatory insulitis. Reconstitution of all components of the CD4⁺ subset emphasizes the plastic capacity of different cell types to adopt effector and suppressor phenotypes. Furthermore, similar immune profiles of diabetic and euglycaemic NOD.SCID recipients demonstrate dissociation between fractional expression of CD25 and FoxP3 and the severity of insulitis. There were no evident and consistent differences in diabetogenic activity and immune reconstituting activity of T cells from pre-diabetic (11 weeks) and new onset diabetic NOD females. Similarities in immune phenotypes and variable distribution of effector and suppressor subsets in various stages of inflammation commend caution in interpretation of quantitative and qualitative aberrations as markers of disease severity in adoptive transfer experiments.     

A novel mechanism of regulatory T cell-mediated down-regulation of autoimmunity

We have established a novel CD4 and CD8 double-positive CD25+ T regulatory (Treg) clone, MT-5B, from lymph nodes of type 1 diabetes prone non-obese diabetic (NOD) mice immunized with CFA. CFA has previously been shown to prevent the onset of diabetes by inducing Treg cells. In vitro, clone MT-5B was anergic to a panel of antigen stimulations and exerted an immunosuppressive effect in antigen-non-specific and cell contact-independent manners. In vivo, clone MT-5B blocked the adoptive transfer of diabetes. Proteomics and immunoadsorption studies identified the suppressive proteins secreted by clone MT-5B as granzyme B (GrB) and perforin (PFN). GrB-mediated immune suppression was PFN dependent. Removal of GrB or PFN from the culture supernatant (SN) of MT-5B cells or pre-incubation of MT-5B cells with ethyleneglycol-bis(aminoethylether)-tetraacetic acid which blocks PFN activity reduced the immunosuppressive effect in vitro. Pre-incubation of diabetogenic splenocytes from NOD mice with MT-5B SN impaired their ability to transfer disease by inducing T cell apoptosis, and removal of GrB from MT-5B SN by immunoadsorption decreased the effector function of MT-5B SN on diabetogenic splenocytes. Immunization of NOD mice with CFA increased the expression of GrB+ CD4 T cells, indicating that these cells are present in vivo. In conclusion, we describe a novel mechanism of cell contact-independent immune suppression in which Treg cells maintain immune homeostasis by secreting GrB/PFN.     

In vivo blockade of the Fas-Fas ligand pathway inhibits cyclophosphamide-induced diabetes in NOD mice.

There is compelling evidence to show that insulin dependent diabetes ensues from selective apoptosis of pancreatic beta-cells mediated by autoreactive T-lymphocytes. The respective implication in this phenomenon of the various apoptotic pathways driven by Fas, perforin, or tumor necrosis factor is still ill- defined. Here we took advantage of the cyclophosphamide-induced model of accelerated diabetes in NOD mice to explore the physiopathological role of the Fas-Fas Ligand pathway. A single injection of cyclophosphamide (200 mg/kg) to 7-8 week-old prediabetic NOD mice triggered diabetes within 10-15 days in 85-100% of the animals. Cyclophosphamide also induced a significant decrease in spleen T cells, that was most evident by days 6-10 after treatment, and selectively affected the CD3(+)CD62L(+)compartment that includes immunoregulatory T cells. To block the in vivo Fas-Fas ligand (Fas L) interaction we administered a biologically active recombinant fusion protein coupling mouse Fas to the Fc portion of human IgG1 (FAS-Fc). Mice treated with FAS-Fc (10 doses iv of 15 microg) starting on the day of cyclophosphamide injection up to day 22, were fully protected from disease. Unexpectedly this protective effect was not due to blockade of Fas-FasL-mediated beta-cell apoptosis but rather to the inhibition of the cyclophosphamide effect on T cells. Indeed FAS-Fc treatment prevented the drug-induced T cell depletion in general and that of immunoregulatory T cells in particular. Additionally, FAS-Fc administration limited to the phase of beta-cell destruction did not afford any protection.     

A novel multimeric form of FasL modulates the ability of diabetogenic T cells to mediate type 1 diabetes in an adoptive transfer model

Activation induced cell death (AICD) via Fas/FasL is the primary homeostatic molecular mechanism employed by the immune system to control activated T-cell responses and promote tolerance to self-antigens. We herein investigated the ability of a novel multimeric form of FasL chimeric with streptavidin (SA-FasL) having potent apoptotic activity to induce apoptosis in diabetogenic T cells and modulate insulin-dependent type 1 diabetes (IDDM) in an adoptive transfer model. Diabetogenic splenocytes from NOD/Lt females were co-cultured in vitro with SA-FasL, SA control protein, or alone without protein, and adoptively transferred into NOD/Lt-Rag1(null) recipients for diabetes development. All animals receiving control (Alone: n=16 or SA: n=17) cells developed diabetes on average by 6 weeks, whereas animals receiving SA-FasL-treated (n=25) cells exhibited significantly delayed progression (p<.001) and decreased incidence (70%). This effect was associated with an increase in CD4(+)CD25(+) T cells and correlated with FoxP3 expression in pancreatic lymph nodes. Extracorporeal treatment of peripheral blood lymphocytes using SA-FasL during disease onset represents a novel approach that may alter the ability of pathogenic T cells to mediate diabetes and have therapeutic utility in clinical management of IDDM.     

Diabetes in non-obese diabetic mice is not associated with quantitative changes in CD4+ CD25+ Foxp3+ regulatory T cells

The role of regulatory T cells (Tregs) in maintaining self tolerance has been intensively researched and there is a growing consensus that a decline in Treg function is an important step towards the development of autoimmune diseases, including diabetes. Although we show here that CD25+ cells delay diabetes onset in non-obese diabetic (NOD) mice, we found, in contrast to previous reports, neither an age-related decline nor a decline following onset of diabetes in the frequency of CD4+ CD25+ Foxp3+ regulatory T cells. Furthermore, we demonstrate that CD4+ CD25+ cells from both the spleen and pancreatic draining lymph nodes of diabetic and non-diabetic NOD mice are able to suppress the proliferation of CD4+ CD25- cells to a similar extent in vitro. We also found that pretreatment of NOD mice with anti-CD25 antibody allowed T cells with a known reactivity to islet antigen to proliferate more in the pancreatic draining lymph nodes of NOD mice, regardless of age. In addition, we demonstrated that onset of diabetes in NOD.scid mice is faster when recipients are co-administered splenocytes from diabetic NOD donors and anti-CD25. Finally, we found that although diabetic CD4+ CD25+ T cells are not as suppressive in cotransfers with effectors into NOD.scid recipients, this may not indicate a decline in Treg function in diabetic mice because over 10% of CD4+ CD25+ T cells are non-Foxp3 and the phenotype of the CD25- contaminating population significantly differs in non-diabetic and diabetic mice. This work questions whether onset of diabetes in NOD mice is associated with a decline in Treg function.     

Absence of significant Th2 response in diabetes-prone non-obese diabetic (NOD) mice.

Accumulating evidence suggests that Th1 T cells play a pivotal role in the development of autoimmune diabetes. Conversely, promoting a Th2 response inhibits disease progression. However, it has not been determined whether Th2 cells are regulatory T cells that fail at the time of diabetes development in naive non-diabetic NOD mice. Therefore, in order to evaluate cytokine secretion by spleen and islet infiltrating T cells in NOD mice at different stages of the autoimmune process, we developed an ELISPOT assay that detects IL-2, IL-4, and interferon-gamma (IFN-gamma) secretion in vitro at the single-cell level. We showed that, whatever the age considered, IFN-gamma is predominantly secreted, and that no IL-4-secreting cells are detected in the islets of male and female NOD mice. Spleen cells from 8-week-old female NOD mice, which include regulatory suppressor T cells, do not secrete IL-4, either upon presentation of islet cell antigens in vitro, or after transfer in vivo, but do secrete IFN-gamma. IFN-gamma secretion by T cells from diabetic mice results from CD4 but not CD8 T cells in transfer experiments into NOD/severe combined immunodeficient (SCID) recipients. These results suggest that (i) detection of regulatory CD4 T cells in NOD mice is not paralleled by a Th2 response; (ii) beta cell destruction does not depend on a switch from a Th2 to a Th1-type response; and (iii) CD8 T cells do not participate in induction of diabetes by secreting IFN-gamma.     

Disruption of the homeostatic balance between autoaggressive (CD4+CD40+) and regulatory (CD4+CD25+FoxP3+) T cells promotes diabetes

Although regulatory T cells (Tregs) are well described, identifying autoaggressive effector T cells has proven more difficult. However, we identified CD4loCD40+ (Th40) cells as being necessary and sufficient for diabetes in the NOD mouse model. Importantly, these cells are present in pancreata of prediabetic and diabetic NOD mice, and Th40 cells but not CD4+CD40(-) T cells transfer progressive insulitis and diabetes to NOD.scid recipients. Nonobese-resistant (NOR) mice have the identical T cell developmental background as NOD mice, yet they are diabetes-resistant. The seminal issue is how NOR mice remain tolerant to diabetogenic self-antigens. We show here that autoaggressive T cells develop in NOR mice and are confined to the Th40 subset. However, NOR mice maintain Treg numbers equivalent to their Th40 numbers. NOD mice have statistically equal numbers of CD4+CD25+forkhead box P3+intrinsic Tregs compared with NOR or nonautoimmune BALB/c mice, and NOD Tregs are equally as suppressive as NOR Tregs. A critical difference is that NOD mice develop expanded numbers of Th40 cells. We suggest that a determinant factor for autoimmunity includes the Th40:Treg ratio. Mechanistically, NOD Th40 cells have low susceptibility to Fas-induced cell death and unlike cells from NOR and BALB/c mice, have predominantly low Fas expression. CD40 engagement of Th40 cells induces Fas expression but further confers resistance to Fas-mediated cell death in NOD mice. A second fundamental difference is that NOD Th40 cells undergo much more rapid homeostatic expansion than Th40 cells from NOR mice.     

Mechanisms of regulatory T-cell induction by antigen-IgG-transduced splenocytes

Our previous studies have demonstrated that splenocytes, transduced with glutamate decarboxylate 65 (GAD) and IgG fusion construct, protect non-obese diabetes (NOD) mice from diabetes. However, the mechanism by which this strategy prevents diabetes is not well understood. Here, we found that CD4(+)Foxp3(+)Treg cells, in vitro induced by GAD-IgG-transduced splenocytes, after transfer, were responsible for prevention of diabetes in NOD mice. Further studies suggested that GAD-IgG-transduced B cells could secrete high level of TGF-beta and stimulated CD4(+)T cells to secrete high level of IFN-gamma. Finally, we found that when TGF-beta and/or IFN-gamma were blocked, CD4(+)Foxp3(-)T cells were not converted into CD4(+)Foxp3(+)Treg cells. The results suggest that GAD-IgG-transduced B cells via TGF-beta and IFN-gamma in vitro induce the CD4(+)Foxp3(+)Treg cells which are responsible for prevention of diabetes in NOD mice by GAD-IgG-gene transfer.     

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.     

Detection of vasostatin‐1‐specific CD8+ T cells in non‐obese diabetic mice that contribute to diabetes pathogenesis

Chromogranin A (ChgA) is an antigenic target of pathogenic CD4⁺ T cells in a non‐obese diabetic (NOD) mouse model of type 1 diabetes (T1D). Vasostatin‐1 is a naturally processed fragment of ChgA. We have now identified a novel H2‐K^d‐restricted epitope of vasostatin‐1, ChgA 36‐44, which elicits CD8⁺ T cell responses in NOD mice. By using ChgA 36‐44/K^d tetramers we have determined the frequency of vasostatin‐1‐specific CD8⁺ T cells in pancreatic islets and draining lymph nodes of NOD mice. We also demonstrate that vasostatin‐1‐specific CD4⁺ and CD8⁺ T cells constitute a significant fraction of islet‐infiltrating T cells in diabetic NOD mice. Adoptive transfer of T cells from ChgA 36‐44 peptide‐primed NOD mice into NOD/severe combined immunodeficiency (SCID) mice led to T1D development. These findings indicate that vasostatin‐1‐specific CD8⁺ T cells contribute to the pathogenesis of type 1 diabetes in NOD mice.