Implication of the intestinal microbiome as a potential surrogate marker of immune responsiveness to experimental therapies in autoimmune diabetes.

Type 1 diabetes (T1D) is an autoimmune proinflammatory disease with no effective intervention. A major obstacle in developing new immunotherapies for T1D is the lack of means for monitoring immune responsiveness to experimental therapies. The LEW1.WR1 rat develops autoimmunity following infection with the parvovirus Kilham rat virus (KRV) via mechanisms linked with activation of proinflammatory pathways and alterations in the gut bacterial composition. We used this animal to test the hypothesis that intervention with agents that block innate immunity and diabetes is associated with a shift in the gut microbiota. We observed that infection with KRV results in the induction of proinflammatory gene activation in both the spleen and pancreatic lymph nodes. Furthermore, administering animals the histone deacetylase inhibitor ITF-2357 and IL-1 receptor antagonist (Anakinra) induced differential STAT-1 and the p40 unit of IL-12/IL-23 gene expression. Sequencing of bacterial 16S rRNA genes demonstrated that both ITF-2357 and Anakinra alter microbial diversity. ITF-2357 and Anakinra modulated the abundance of 23 and 8 bacterial taxa in KRV-infected animals, respectively, of which 5 overlapped between the two agents. Lastly, principal component analysis implied that ITF-2357 and Anakinra induce distinct gut microbiomes compared with those from untreated animals or rats provided KRV only. Together, the data suggest that ITF-2357 and Anakinra differentially influence the innate immune system and the intestinal microbiota and highlight the potential use of the gut microbiome as a surrogate means of assessing anti-inflammatory immune effects in type 1 diabetes.

Genetic polymorphisms of ADRB2 and IL10 may be associated with the risk of IgE sensitization to digestive powders in exposed medical personnel.

BACKGROUND: It has been reported that pancreatic extracts may induce IgE-mediated respiratory allergy in medical personnel. The aim of the study was to identify genetic factors associated with IgE sensitization to digestive powders containing pancreatic extract. METHODS: This case-control study was performed on 153 subjects routinely exposed to digestive powder and on 123 nonexposed controls working in Ajou University Hospital. Skin prick testing was performed using 4 commonly used digestive powders and α-amylase. Serum specific IgE levels were measured by ELISA. Three single nucleotide polymorphisms, ADRB2 46A>G, IL10 -1082A>G and IL4 -589T>C, were genotyped using the single base extension method. RESULTS: The positive rate of serum specific IgE to digestive powder was significantly higher in the 41 (26.8%) exposed personnel with work-related respiratory symptoms than in controls (24.4 vs. 5.4%, p = 0.012). Thirty-nine (25.5%) of the 153 exposed personnel were found to have an allergy to digestive powder, as determined by a positive skin prick test and/or a high serum specific IgE level to digestive powder. The ADRB2 46A>G and IL10 -1082A>G polymorphisms were found to be significantly associated with the development of an allergy to digestive powder in exposed medical personnel by multiple logistic regression analysis after controlling for age, atopy and job type (pharmacist or nurse; p = 0.035 and p = 0.027, respectively). CONCLUSION: These results suggest that the genetic polymorphisms ADRB2 46A>G and IL10 -1082A>G are genetic factors that increase IgE sensitization to pancreatic extracts in medical personnel occupationally exposed to digestive powders.

Caspase-3-dependent beta-cell apoptosis in the initiation of autoimmune diabetes mellitus.

beta-Cell apoptosis is a key event contributing to the pathogenesis of type 1 diabetes mellitus. In addition to apoptosis being the main mechanism by which beta cells are destroyed, beta-cell apoptosis has been implicated in the initiation of type 1 diabetes mellitus through antigen cross-presentation mechanisms that lead to beta-cell-specific T-cell activation. Caspase-3 is the major effector caspase involved in apoptotic pathways. Despite evidence supporting the importance of beta-cell apoptosis in the pathogenesis of type 1 diabetes, the specific role of caspase-3 in this process is unknown. Here, we show that Caspase-3 knockout (Casp3(-/-) mice were protected from developing diabetes in a multiple-low-dose streptozotocin autoimmune diabetes model. Lymphocyte infiltration of the pancreatic islets was completely absent in Casp3(-/-) mice. To determine the role of caspase-3-dependent apoptosis in disease initiation, a defined antigen-T-cell receptor transgenic system, RIP-GP/P14 double-transgenic mice with Casp3 null mutation, was examined. beta-cell antigen-specific T-cell activation and proliferation were observed only in the pancreatic draining lymph node of RIP-GP/P14/Casp3(+/-) mice, but not in mice lacking caspase-3. Together, our findings demonstrate that caspase-3-mediated beta-cell apoptosis is a requisite step for T-cell priming, a key initiating event in type 1 diabetes.

Skin prick test reactivity to supplemental enzymes in cystic fibrosis and pancreatic insufficiency.

BACKGROUND: Patients with cystic fibrosis and pancreatic insufficiency take supplemental pancreatic enzymes of porcine or plant origin on a daily basis. OBJECTIVE: The frequency of immediate skin prick hypersensitivity reactions to such enzymes and the correlations to gastrointestinal symptoms and nutritional status were assessed in a single center prospective descriptive study. METHODS: Skin prick tests using five different freshly prepared porcine pancreatic enzyme preparations, one enzyme preparation of fungal origin, porcine meat and cat epithelia were performed in patients with cystic fibrosis and pancreatic insufficiency and in healthy control subjects. RESULTS: Positive skin tests in the 48 patients with cystic fibrosis were no more frequent than in 21 control subjects. No correlations with the frequency of gastrointestinal symptoms, nutritional status and lung function were found. CONCLUSION: Although immediate hypersensitivity reactions to pancreatic enzyme preparations in subjects with cystic fibrosis are not common, individuals with possible allergic reactions to supplemental enzymes should be identified by skin prick tests and in unclear cases by double-blind placebo-controlled provocation tests.

A high prevalence of organ-specific autoimmunity in patients with bipolar disorder.

BACKGROUND: In a previous study, we reported an increased prevalence of thyroperoxidase antibodies (TPOA) in patients with bipolar disorder. Here we report the prevalence of other organ-specific autoantibodies: H/K adenosine triphosphatase (ATPA), glutamic acid decarboxylase-65 (GAD65A), and GAD-67 (GAD67A). METHODS: ATPA, GAD65A, and GAD67A were determined (via a commercially available enzyme linked immunosorbent assay for ATPA, and a standardized radio immunoassays for GAD65A and GAD67A)in the sera of 239 patients with DSM-IV bipolar disorder, in 74 patients with DSM-IV schizophrenia, and in 220 healthy control subjects. RESULTS: The positivity prevalences for ATPA and GAD65A (but not GAD67A) were elevated in bipolar patients compared with those in healthy control subjects (11.7 vs. 6.1% and 11.3 vs. 2.6% respectively; p <.05). Schizophrenia patients did not show such statistically higher prevalence. The elevated prevalence of ATPA and GAD65A in bipolar disorder was associated with neither rapid cycling nor the use of lithium. Interestingly, the presence of GAD65A (and not that of TPOA and ATPA) tended to be associated with the activity of bipolar disorder. The level of TPOA was negatively correlated with the serum level of sIL-2R, a measure of T cell activation. CONCLUSION: Bipolar disorder is associated with organ-specific autoimmunity to the antigens TPO, H/K ATPase, and GAD65.

Functional tolerance is maintained despite proliferation of CD4 T cells after encounter with tissue-derived antigen.

Since negative selection in the thymus is incomplete, some self-reactive T cells are able to mature and seed the periphery. To study how these T cells interact following encounter with the self-protein they recognize in the periphery, we have developed an adoptive transfer system in which HEL-specific TCR transgenic CD4 T cells are transferred to mice expressing HEL protein in the pancreas under the control of the rat insulin promoter. Here we show that after adoptive transfer of HEL-specific T cells functional tolerance is maintained despite evidence that the T cells encounter and respond to pancreas-expressed antigen. Even the provision of an additional activation stimulus by peripheral immunization with HEL protein is insufficient to induce the T cells to cause autoimmune tissue injury. However, in the presence of blocking anti-CTLA-4-mAb, immunized adoptive transfer recipients rapidly developed diabetes. These data suggest that the CTLA-4 pathway regulates the pathogenicity of antigen-specific T cells following a peripheral activation stimulus.

Autoimmune insulitis and diabetes in the absence of antigen-specific contact between T cells and islet beta-cells.

Autoimmune diabetes develops following recognition of organ-specific antigens by T cells. The disease begins with peri-islet infiltration by mononuclear cells, proceeds with insulitis and becomes manifest with destruction of insulin-producing islet beta-cells. T cells are necessary to induce insulitis and diabetes, but it is not clear by what mechanisms they can do so, i. e. whether the T cells need to make antigen-specific contact with the beta-cell or whether other interactions are sufficient to induce beta-cell death. In the present study we have constructed chimeric mice in which the bone marrow-derived antigen-presenting cells, but not the islet beta-cells, are capable of presenting antigen to monospecific T cells. We show that both insulitis as well as beta-cell destruction can proceed in the absence of islet beta-cell surface antigen recognition by T cells. Our results support the notion that diabetes can be caused by distinct effector mechanisms.