Approaches to Type 1 Diabetes Prevention by Intervention in Cytokine Immunoregulatory Circuits

Type 1 (insulin-dependent) diabetes mellitus, like other organ specific autoimmune diseases, results from a disorder of immunoregulation. T cells specific for pancreatic islet ß cell constituents (autoantigens) exist normally but are restrained by regulatory mechanisms (self-tolerant state). When regulation fails, ß cell-specific autoreactive T cells become activated and expand clonally. Current evidence indicates that islet ß cell-specific autoreactive T cells belong to a T helper 1 (Th1) subset, and these Th1 cells and their characteristic cytokine products, IFNγ and IL-2, are believed to cause islet inflammation (insulitis) and ß cell destruction. Immune-mediated destruction of ß cells precedes hyperglycemia and clinical symptoms by many years because these become apparent only when most of the insulin-secreting ß cells have been destroyed. Therefore, several approaches are being tested or are under consideration for clinical trials to prevent or arrest complete autoimmune destruction of islet ß cells and insulin-dependent diabetes. Approaches that attempt to correct underlying immunoregulatory defects in autoimmune diabetes include interventions aimed at i) deleting ß cell autoreactive Th1 cells and cytokines (IFNγ and IL-2) and/or ii) increasing regulatory Th2 cells and/or Th3 cells and their cytokine products (IL-4, IL-10 and TGFßI).     

Cellular and molecular mechanism for Kilham rat virus-induced autoimmune diabetes in DR-BB rats

Kilham rat virus (KRV) causes autoimmune diabetes in diabetes-resistant BioBreeding (DR-BB) rats; however, the mechanism by which KRV induces autoimmune diabetes without the direct infection of beta cells is not well understood. We first asked whether molecular mimicry, such as a common epitope between a KRV-specific peptide and a beta cell autoantigen, is involved in the initiation of KRV-induced autoimmune diabetes in DR-BB rats. We found that KRV peptide-specific T cells generated in DR-BB rats infected with recombinant vaccinia virus expressing KRV-specific structural and nonstructural proteins could not induce diabetes, indicating that molecular mimicry is not the mechanism by which KRV induces autoimmune diabetes. Alternatively, we asked whether KRV infection of DR-BB rats could disrupt the finely tuned immune balance and activate autoreactive T cells that are cytotoxic to beta cells, resulting in T cell-mediated autoimmune diabetes. We found that both Th1-like CD45RC+CD4+ and cytotoxic CD8+ T cells were up-regulated, whereas Th2-like CD45RC-CD4+ T cells were down-regulated, and that isolated and activated CD45RC+CD4+ and CD8+ T cells from KRV-infected DR-BB rats induced autoimmune diabetes in young diabetes-prone BioBreeding (DP-BB) rats. We conclude that KRV-induced autoimmune diabetes in DR-BB rats is not due to molecular mimicry, but is due to a breakdown of the finely tuned immune balance of Th1-like CD45RC+CD4+ and Th2-like CD45RC-CD4+ T cells, resulting in the selective activation of beta cell-cytotoxic effector T cells.     

Comparing the relative role of perforin/granzyme versus Fas/Fas ligand cytotoxic pathways in CD8+ T cell-mediated insulin-dependent diabetes mellitus.

CD8+ cytotoxic T cells play a critical role in initiating insulin-dependent diabetes mellitus. The relative contribution of each of the major cytotoxic pathways, perforin/granzyme and Fas/Fas ligand (FasL), in the induction of autoimmune diabetes remains controversial. To evaluate the role of each lytic pathway in beta cell lysis and induction of diabetes, we have used a transgenic mouse model in which beta cells expressing the influenza virus hemagglutinin (HA) are destroyed by HA-specific CD8+ T cells from clone-4 TCR-transgenic mice. Upon adoptive transfer of CD8+ T cells from perforin-deficient clone-4 TCR mice, there was a 30-fold increase in the number of T cells required to induce diabetes. In contrast, elimination of the Fas/FasL pathway of cytotoxicity had little consequence. When both pathways of cytolysis were eliminated, mice did not become diabetic. Using a model of spontaneous diabetes, which occurs in double transgenic neonates that express both clone-4 TCR and Ins-HA transgenes, mice deficient in either the perforin or FasL/Fas lytic pathway become diabetic soon after birth. This indicates that, in the neonate, large numbers of autoreactive CD8+ T cells can lead to destruction of islet beta cells by either pathway.     

Isolation of self antigen-reactive cells from inflamed islets of nonobese diabetic mice using CD4high expression as a marker.

The low precursor frequency of Ag-reactive CD4+ T cells has been a barrier to the study of CD4+ T cell responses to conventional Ags as well as CD4+ T cell responses to autoantigens recognized during the course of an autoimmune disease. We have recently reported that all "conventional Ag" reactive CD4+ T cells are contained within the subpopulation expressing high levels of the CD4 molecule, termed CD4high. We have identified a CD4high population in the islets of Langerhans of prediabetic nonobese diabetic (NOD) mice that is extremely potent in transferring disease. As few as 500 CD4high islet-infiltrating CD4+ T cells transferred insulin-dependent diabetes mellitus to CD8 reconstituted NOD-SCID mice within 30 days of transfer. In contrast, CD4high T cells isolated from either NOD spleen or salivary glands did not transfer insulin-dependent diabetes mellitus into similar CD8-reconstituted NOD-SCID recipients. These data indicate that the precursor frequency of NOD islet-reactive, pathogenic CD4+ T cells is much higher in the prediabetic NOD pancreas than in these other organs. The islet-infiltrating CD4high T cells displayed selected memory markers, by cell surface analysis, and displayed a Th 1 phenotype by RNase protection assay, but had a marked decrease in IL-4 mRNA determined by quantitative real time PCR when compared with the less pathogenic CD4normal islet-infiltrating T cells. Use of the CD4high marker to select Ag activated T cells represents a tool to isolate and study pathogenic CD4+ T cells from autoimmune lesions in which the Ag has not been previously defined.     

Characterization of peripheral regulatory CD4+ T cells that prevent diabetes onset in nonobese diabetic mice

The period that precedes onset of insulin-dependent diabetes mellitus corresponds to an active dynamic state in which pathogenic autoreactive T cells are kept from destroying beta cells by regulatory T cells. In prediabetic nonobese diabetic (NOD) mice, CD4+ splenocytes were shown to prevent diabetes transfer in immunodeficient NOD recipients. We now demonstrate that regulatory splenocytes belong to the CD4+ CD62Lhigh T cell subset that comprises a vast majority of naive cells producing low levels of IL-2 and IFN-gamma and no IL-4 and IL-10 upon in vitro stimulation. Consistently, the inhibition of diabetes transfer was not mediated by IL-4 and IL-10. Regulatory cells homed to the pancreas and modified the migration of diabetogenic to the islets, which resulted in a decreased insulitis severity. The efficiency of CD62L+ T cells was dose dependent, independent of sex and disease prevalence. Protection mechanisms did not involve the CD62L molecule, an observation that may relate to the fact that CD4+ CD62Lhigh lymph node cells were less potent than their splenic counterparts. Regulatory T cells were detectable after weaning and persist until disease onset, sustaining the notion that diabetes is a late and abrupt event. Thus, the CD62L molecule appears as a unique marker that can discriminate diabetogenic (previously shown to be CD62L-) from regulatory T cells. The phenotypic and functional characteristics of protective CD4+ CD62L+ cells suggest they are different from Th2-, Tr1-, and NK T-type cells, reported to be implicated in the control of diabetes in NOD mice, and may represent a new immunoregulatory population.     

IL-10-dependent suppression of experimental allergic encephalomyelitis by Th2-differentiated, anti-TCR redirected T lymphocytes

We previously showed that transgenically expressed chimeric Ag-MHC-zeta receptors can Ag-specifically redirect T cells against other T cells. When the receptor's extracellular Ag-MHC domain engages cognate TCR on an Ag-specific T cell, its cytoplasmic zeta-chain stimulates the chimeric receptor-modified T cell (RMTC). This induces effector functions such as cytolysis and cytokine release. RMTC expressing a myelin basic protein (MBP) 89-101-IAs-zeta receptor can be used therapeutically, Ag-specifically treating murine experimental allergic encephalomyelitis (EAE) mediated by MBP89-101-specific T cells. In initial studies, isolated CD8+ RMTC were therapeutically effective whereas CD4+ RMTC were not. We re-examine here the therapeutic potential of CD4+ RMTC. We demonstrate that Th2-differentiated, though not Th1-differentiated, CD4+ MBP89-101-IAs-zeta RMTC prevent actively induced or adoptively transferred EAE, and treat EAE even after antigenic diversification of the pathologic T cell response. The Th2 RMTC both Th2-deviate autoreactive T cells and suppress autoantigen-specific T cell proliferation. IL-10 is critical for the suppressive effects. Anti-IL-10R blocks RMTC-mediated modulation of EAE and suppression of autoantigen proliferation, as well as the induction of IL-10 production by autoreactive T cells. In contrast to IL-10, IL-4 is required for IL-4 production by, and hence Th2 deviation of autoreactive T cells, but not the therapeutic activity of the RMTC. These results therefore demonstrate a novel immunotherapeutic approach for the Ag-specific treatment of autoimmune disease with RMTC. They further identify an essential role for IL-10, rather than Th2-deviation itself, in the therapeutic effectiveness of these redirected Th2 T cells.     

Systemic administration of agonist peptide blocks the progression of spontaneous CD8-mediated autoimmune diabetes in transgenic mice without bystander damage

Insulin-dependent diabetes is an autoimmune disease targeting pancreatic beta-islet cells. Recent data suggest that autoreactive CD8+ T cells are involved in both the early events leading to insulitis and the late destructive phase resulting in diabetes. Although therapeutic injection of protein and synthetic peptides corresponding to CD4+ T cell epitopes has been shown to prevent or block autoimmune disease in several models, down-regulation of an ongoing CD8+ T cell-mediated autoimmune response using this approach has not yet been reported. Using CL4-TCR single transgenic mice, in which most CD8+ T cells express a TCR specific for the influenza virus hemagglutinin HA512-520 peptide:Kd complex, we first show that i.v. injection of soluble HA512-520 peptide induces transient activation followed by apoptosis of Tc1-like CD8+ T cells. We next tested a similar tolerance induction strategy in (CL4-TCR x Ins-HA)F1 double transgenic mice that also express HA in the beta-islet cells and, as a result, spontaneously develop a juvenile onset and lethal diabetes. Soluble HA512-520 peptide treatment, at a time when pathogenic CD8+ T cells have already infiltrated the pancreas, very significantly prolongs survival of the double transgenic pups. In addition, we found that Ag administration eliminates CD8+ T cell infiltrates from the pancreas without histological evidence of bystander damage. Our data indicate that agonist peptide can down-regulate an autoimmune reaction mediated by CD8+ T cells in vivo and block disease progression. Thus, in addition to autoreactive CD4+ T cells, CD8+ T cells may constitute targets for Ag-specific therapy in autoimmune diseases.     

MHC class I-restricted determinants on the glutamic acid decarboxylase 65 molecule induce spontaneous CTL activity

CD4(+) T cell responses to glutamic acid decarboxylase (GAD65) spontaneously arise in nonobese diabetic (NOD) mice before the onset of insulin-dependent diabetes mellitus (IDDM) and may be critical to the pathogenic process. However, since both CD4(+) and CD8(+) T cells are involved in autoimmune diabetes, we sought to determine whether GAD65-specific CD8(+) T cells were also present in prediabetic NOD mice and contribute to IDDM. To refine the analysis, putative K(d)-binding determinants that were proximal to previously described dominant Th determinants (206-220 and 524-543) were examined for their ability to elicit cytolytic activity in young NOD mice. Naive NOD spleen cells stimulated with GAD65 peptides 206-214 (p206) and 546-554 (p546) produced IFN-gamma and showed Ag-specific CTL responses against targets pulsed with homologous peptide. Conversely, several GAD peptides distal to the Th determinants, and control K(d)-binding peptides did not induce similar responses. Spontaneous CTL responses to p206 and p546 were mediated by CD8(+) T cells that are capable of lysing GAD65-expressing target cells, and p546-specific T cells transferred insulitis to NOD.scid mice. Young NOD mice pretreated with p206 and p546 showed reduced CTL responses to homologous peptides and a delay in the onset of IDDM. Thus, MHC class I-restricted responses to GAD65 may provide an inflammatory focus for the generation of islet-specific pathogenesis and beta cell destruction. This report reveals a potential therapeutic role for MHC class I-restricted peptides in treating autoimmune disease and revisits the notion that the CD4- and CD8-inducing determinants on some molecules may benefit from a proximal relationship.     

In vivo expression of perforin by CD8+ lymphocytes in autoimmune disease. Studies on spontaneous and adoptively transferred diabetes in nonobese diabetic mice

A potent cytolytic pore-forming protein (perforin or cytolysin) has previously been found to be associated with the cytoplasmic granules of CTL and NK cells. Inasmuch as all previous studies on perforin have been conducted with cultured CTL and NK cell lines, it is not clear whether perforin may play a role in the cytotoxicity mediated by CTL that have been primed in vivo. In this study, we investigated the presence of perforin in pancreata from nonobese diabetic (NOD) mice, which have been studied as a model of autoimmune, insulin-dependent (type I) diabetes mellitus. Whereas adult NOD mice spontaneously develop diabetes, it is possible to induce diabetes in young, irradiated NOD mice by adoptive transfer of splenocytes obtained from diabetic donors. By means of immunohistochemical analysis, we were able to detect perforin Ag in a small subpopulation of CD8+/Thy-1+/asialo GM1-/CD4- lymphocytes in the pancreatic islets of animals undergoing both spontaneous and adoptive transfer-mediated insulitis. Perforin+/CD8+ lymphocytes were found in small clusters and were observed to display the morphology of large granular lymphocytes. These observations show for the first time the presence of perforin-containing CD8+ lymphocytes in tissues of animals undergoing autoimmune disease.     

Microenvironment-dependent requirement of STAT4 for the induction of P-selectin ligands and effector cytokines on CD4+ T cells in healthy and parasite-infected mice

T effector cells require selectin ligands to migrate into inflamed regions. In vitro, IL-12 promotes induction of these ligands as well as differentiation of CD4+ T cells into IFN-gamma-producing Th1 but not Th2 cells. STAT4 is strongly involved in these processes. However, the presence of selectin ligands on various T effector cell subsets in vivo points to more complex regulatory pathways. To clarify the role of the IL-12/STAT4 signaling pathway, we analyzed the impact of STAT4 deficiency on the expression of P-selectin ligands (P-lig) on CD4+ T cells in vitro and in vivo, including conditions of infection. In vitro, we found significant expression of P-lig upon activation not only in the presence, but also in the absence, of IL-12, which was independent of STAT4. TGF-beta, an alternative inducer of selectin ligands in human T cells, was not effective in murine CD4+ T cells, suggesting a role of additional signaling pathways. In vivo, a significant impact of STAT4 for the generation of P-lig+CD4+ T cells was observed for cells from peripheral lymph nodes, but not for those from spleen or lung. However, upon infection with the Th2-inducing parasite Nippostrongylus brasiliensis, P-lig expression became dependent on STAT4 signaling. Interestingly, also the frequency of IL-4-producing cells was greatly diminished in absence of STAT4. These data reveal a hitherto unknown contribution of STAT4 to the generation of Th2 cells in parasite infection and suggest that signals inducing inflammation-seeking properties in vivo vary depending on environmental conditions, such as type of organ and infection.     

T cell-intrinsic and -extrinsic contributions of the IFNAR/STAT1-axis to thymocyte survival

STAT1 is an essential part of interferon signaling, and STAT1-deficiency results in heightened susceptibility to infections or autoimmunity in both mice and humans. Here we report that mice lacking the IFNα/β-receptor (IFNAR1) or STAT1 display impaired deletion of autoreactive CD4(+)CD8(+)-T-cells. Strikingly, co-existence of WT T cells restored thymic elimination of self-reactive STAT1-deficient CD4(+)CD8(+)-T cells. Analysis of STAT1-deficient thymocytes further revealed reduced Bim expression, which was restored in the presence of WT T cells. These results indicate that type I interferons and STAT1 play an important role in the survival of MHC class I-restricted T cells in a T cell intrinsic and non-cell intrinsic manner that involves regulation of Bim expression through feedback provided by mature STAT1-competent T cells.     

IFN-alpha is not sufficient to drive Th1 development due to lack of stable T-bet expression

During inflammatory immune responses, the innate cytokine IL-12 promotes CD4+ Th-1 development through the activation of the second messenger STAT4 and the subsequent expression of T-bet. In addition, type I IFN (IFN-alphabeta), secreted primarily during viral and intracellular bacterial infections, can promote STAT4 activation in human CD4+ T cells. However, the role of IFN-alphabeta in regulating Th1 development is controversial, and previous studies have suggested a species-specific pathway leading to Th1 development in human but not mouse CD4+ T cells. In this study, we found that although both IFN-alpha and IL-12 can promote STAT4 activation, IFN-alpha failed to promote Th1 commitment in human CD4+ T cells. The difference between these innate signaling pathways lies with the ability of IL-12 to promote sustained STAT4 tyrosine phosphorylation, which correlated with stable T-bet expression in committed Th1 cells. IFN-alpha did not promote Th1 development in human CD4+ T cells because of attenuated STAT4 phosphorylation, which was insufficient to induce stable expression of T-bet. Further, the defect in IFN-alpha-driven Th1 development was corrected by ectopic expression of T-bet within primary naive human CD4+ T cells. These results indicate that IL-12 remains unique in its ability to drive Th1 development in human CD4+ T cells and that IFN-alpha lacks this activity due to its inability to promote sustained T-bet expression.     

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.     

Cutting edge: An in vivo requirement for STAT3 signaling in TH17 development and TH17-dependent autoimmunity

STAT3 activation has been observed in several autoimmune diseases, suggesting that STAT3-mediated pathways promote pathologic immune responses. We provide in vivo evidence that the fundamental role of STAT3 signaling in autoimmunity relates to its absolute requirement for generating T(H)17 T cell responses. We show that STAT3 is a master regulator of this pathogenic T cell subtype, acting at multiple levels in vivo, including T(H)17 T cell differentiation and cytokine production, as well as induction of RORgamma t and the IL-23R. Neither naturally occurring T(H)17 cells nor T(H)17-dependent autoimmunity occurs when STAT3 is ablated in CD4 cells. Furthermore, ablation of STAT3 signaling in CD4 cells results in increased T(H)1 responses, indicating that STAT3 signaling skews T(H) responses away from the T(H)1 pathway and toward the T(H)17 pathway. Thus, STAT3 is a candidate target for T(H)17-dependent autoimmune disease immunotherapy that could selectively inhibit pathogenic immune pathways.     

Nondepleting anti-CD4 has an immediate action on diabetogenic effector cells, halting their destruction of pancreatic beta cells

The induction of tolerance in a primed immune system is a major aim for therapy in autoimmunity and transplant rejection. In this paper, we investigate the action of the nondepleting anti-CD4 Ab, YTS 177. Although this Ab is nondepleting, we have demonstrated a direct action in vivo on activated effector cells. We show that the Ab inhibits transfer of insulin-dependent diabetes mellitus by the CD4+ Th1 clone BDC2.5 to nonobese diabetic mice. Furthermore, we show that this Ab acts directly on diabetogenic effector cells because it prevented BDC2.5-induced insulin-dependent diabetes mellitus in nonobese diabetic-scid recipients in the absence of other T cells. The Ab halts the diabetic process even when it is administered after the BDC2.5 cells have infiltrated the pancreas and destruction of islets is already underway. This is accompanied by an immediate decrease in proinflammatory cytokine production with cessation of beta cell destruction and disappearance of infiltrating cells from the pancreas, leaving any remaining beta cells intact. These data suggest that Abs such as this may be effective not only because they induce regulatory T cells but also because they are able to directly prevent effector cell function.     

Development of memory-like autoregulatory CD8+ T cells is CD4+ T cell dependent

Progression of spontaneous autoimmune diabetes is associated with development of a disease-countering negative-feedback regulatory loop that involves differentiation of low-avidity autoreactive CD8(+) cells into memory-like autoregulatory T cells. Such T cells blunt diabetes progression by suppressing the presentation of both cognate and noncognate Ags to pathogenic high-avidity autoreactive CD8(+) T cells in the pancreas-draining lymph nodes. In this study, we show that development of autoregulatory CD8(+) T cell memory is CD4(+) T cell dependent. Transgenic (TG) NOD mice expressing a low-affinity autoreactive TCR were completely resistant to autoimmune diabetes, even after systemic treatment of the mice with agonistic anti-CD40 or anti-4-1BB mAbs or autoantigen-pulsed dendritic cells, strategies that dramatically accelerate diabetes development in TG NOD mice expressing a higher affinity TCR for the same autoantigenic specificity. Furthermore, whereas abrogation of RAG-2 expression, hence endogenous CD4(+) T cell and B cell development, decelerated disease progression in high-affinity TCR-TG NOD mice, it converted the low-affinity TCR into a pathogenic one. In agreement with these data, polyclonal CD4(+) T cells from prediabetic NOD mice promoted disease in high-affinity TCR-TG NOD.Rag2(-/-) mice, but inhibited it in low-affinity TCR-TG NOD.Rag2(-/-) mice. Thus, in chronic autoimmune responses, CD4(+) Th cells contribute to both promoting and suppressing pathogenic autoimmunity.     

Pancreatic IL-4 expression results in islet-reactive Th2 cells that inhibit diabetogenic lymphocytes in the nonobese diabetic mouse.

When immunological tolerance breaks down, autoimmune destruction of insulin-producing beta cells in the pancreas can cause insulin-dependent diabetes mellitus. We previously showed that transgenic nonobese diabetic (NOD) mice expressing IL-4 in the pancreas (NOD-IL-4 mice) were protected from insulitis and diabetes. Here we have characterized the avoidance of pathological autoimmunity in these mice. The absence of disease did not result from a lack of T cell priming, because T cells responding to dominant islet Ags were present. These islet Ag-specific T cells displayed a Th2 phenotype, indicating that Th2 responses could account for the observed tolerance. Interestingly, islet Ag-specific Th1 T cells were present and found to be functional, because neutralization of the Th2 effector cytokines IL-4 and IL-10 resulted in diabetes. Histological examination revealed that NOD-IL-4 splenocytes inhibited diabetogenic T cells in cotransfer experiments by limiting insulitis and delaying diabetes. Neutralization of IL-4 in this system abrogated the ability of NOD-IL-4 splenocytes to delay the onset of diabetes. These results indicate that IL-4 expressed in the islets does not prevent the generation of pathogenic islet responses but induces islet Ag-specific Th2 T cells that block the action of diabetogenic T cells in the pancreas.     

Amelioration of experimental autoimmune encephalomyelitis by plumbagin through down-regulation of JAK-STAT and NF-κB signaling pathways

Plumbagin (PL), a herbal compound derived from roots of the medicinal plant Plumbago zeylanica, has been shown to have immunosuppressive properties. Present report describes that PL is a potent novel agent in control of encephalitogenic T cell responses and amelioration of mouse experimental autoimmune encephalomyelitis (EAE), through down-regulation of JAK-STAT pathway. PL was found to selectively inhibit IFN-γ and IL-17 production by CD4(+) T cells, which was mediated through abrogated phosphorylation of JAK1 and JAK2. Consistent with IFN-γ and IL-17 reduction was suppressed STAT1/STAT4/T-bet pathway which is critical for Th1 differentiation, as well as STAT3/ROR pathway which is essential for Th17 differentiation. In addition, PL suppressed pro-inflammatory molecules such as iNOS, IFN-γ and IL-6, accompanied by inhibition of IκB degradation as well as NF-κB phosphorylation. These data give new insight into the novel immune regulatory mechanism of PL and highlight the great value of this kind of herb compounds in probing the complex cytokine signaling network and novel therapeutic targets for autoimmune diseases.     

Pancreas-infiltrating Th1 cells and diabetes develop in IL-12-deficient nonobese diabetic mice.

IL-12 and IL-12 antagonist administration to nonobese diabetic (NOD) mice accelerates and prevents insulin-dependent diabetes mellitus (IDDM), respectively. To further define the role of endogenous IL-12 in the development of diabetogenic Th1 cells, IL-12-deficient NOD mice were generated and analyzed. Th1 responses to exogenous Ags were reduced by approximately 80% in draining lymph nodes of these mice, and addition of IL-12, but not IL-18, restored Th1 development in vitro, indicating a nonredundant role of IL-12. Moreover, spontaneous Th1 responses to a self Ag, the tyrosine phosphatase-like IA-2, were undetectable in lymphoid organs from IL-12-deficient, in contrast to wild-type, NOD mice. Nevertheless, wild-type and IL-12-deficient NOD mice developed similar insulitis and IDDM. Both in wild-type and IL-12-deficient NOD mice, approximately 20% of pancreas-infiltrating CD4+ T cells produced IFN-gamma, whereas very few produced IL-10 or IL-4, indicating that IDDM was associated with a type 1 T cell infiltrate in the target organ. T cell recruitment in the pancreas seemed favored in IL-12-deficient NOD mice, as revealed by increased P-selectin ligand expression on pancreas-infiltrating T cells, and this could, at least in part, compensate for the defective Th1 cell pool recruitable from peripheral lymphoid organs. Residual Th1 cells could also accumulate in the pancreas of IL-12-deficient NOD mice because Th2 cells were not induced, in contrast to wild-type NOD mice treated with an IL-12 antagonist. Thus, a regulatory pathway seems necessary to counteract the pathogenic Th1 cells that develop in the absence of IL-12 in a spontaneous chronic progressive autoimmune disease under polygenic control, such as IDDM.