The expression of proinflammatory cytokines and chemokines in response to T cell receptor (TCR) agonists is regulated by the CARMA1 signalosome through the coordinated assembly of complexes containing the BCL10 adaptor protein. We describe a novel mechanism to negatively regulate the CARMA1 signalosome by the “death” adaptor protein, CRADD/RAIDD. We show that CRADD interacts with BCL10 through its caspase recruitment domain (CARD) and suppresses interactions between BCL10 and CARMA1. TCR agonist-induced interaction between CRADD and BCL10 coincides with reduction of its complex formation with CARMA1 in wild-type, as compared to Cradd-deficient, primary cells. Finally, Cradd-deficient spleen cells, CD4+ T cells, and mice respond to T cell agonists with strikingly higher production of proinflammatory mediators, including γ-interferon, IL-2, TNF-α, and IL-17. These results define a novel role for CRADD as a negative regulator of the CARMA1 signalosome and suppressor of T helper (Th)1- and Th17-mediated inflammatory responses.

Background Interactions between genetic risk factors and the environment drive Type 1 diabetes. The system of Toll-like receptors (TLR) detects these environmental triggers; however, the target cell that intermediates these interactions to drive T1D remains unknown. Methods We investigated the effect of TLR pathway activation (MyD88 vs TRIF) on B cell subsets via flow cytometry including their activation, survival, proliferation and cytoskeletal mobilization. The effect of polyIC on diabetes development was addressed including the B cell dependent activation of diabetes-protective DX5+ cells using genetic models and adoptive transfer. Results NOD B lymphocytes expressed enhanced levels of TLR responsive proteins. Ex vivo analysis of B lymphocyte subsets demonstrated that TLR3 stimulation via TRIF deletes cells displaying a marginal zone phenotype, whereas MyD88 dependent ligands enhance their survival. In vivo, marginal zone B cells were activated by polyIC and were unexpectedly retained in the spleen of NOD mice in contrast to the mobilization of these cells in non-autoimmune mice, a phenotype we traced to defective actin cytoskeletal dynamics. These activated B cells mediated TLR3-induced diabetes protection. Conclusions Immunotherapies must account for both B cell location and activation and these properties may differ in autoimmune and healthy settings. The significant finding of the study is that B lymphocytes respond to TLR ligation in a subset specific manner and are required for TLR-triggered diabetes protection. This study adds new information about the role of TLR ligation in diabetes pathogenesis and further identifies a unique role for B lymphocyte specific trafficking abnormalities in T1D.

T cells are critically dependent on cellular proliferation in order to carry out their effector functions. Autoimmune strains are commonly thought to have uncontrolled T cell proliferation; however, in the murine model of autoimmune diabetes, hypo-proliferation of T cells leading to defective AICD was previously uncovered. We now determine whether lupus prone murine strains are similarly hyporesponsive. Upon extensive characterization of T lymphocyte activation, we have observed a common feature of CD4 T cell activation shared among three autoimmune strains–NOD, MRL, and NZBxNZW F1s. When stimulated with a polyclonal mitogen, CD4 T cells demonstrate arrested cell division and diminished dose responsiveness as compared to the non-autoimmune strain C57BL/6, a phenotype we further traced to a reliance on B cell mediated costimulation, which underscores the success of B cell directed immune therapies in preventing T cell mediated tissue injury. In turn, the diminished proliferative capacity of these CD4 T cells lead to a decreased, but activation appropriate, susceptibility to activation induced cell death. A similar decrement in stimulation response was observed in the CD8 compartment of NOD mice; NOD CD8 T cells were distinguished from lupus prone strains by a diminished dose-responsiveness to anti-CD3 mediated stimulation. This distinction may explain the differential pathogenetic pathways activated in diabetes and lupus prone murine strains.

Overcoming the immune response to establish durable immune tolerance in type 1 diabetes remains a substantial challenge. The ongoing effector immune response involves numerous immune cell types but is ultimately orchestrated and sustained by the hematopoietic stem cell (HSC) niche. We therefore hypothesized that tolerance induction also requires these pluripotent precursors. In this study, we determined that the tolerance-inducing agent anti-CD45RB induces HSC mobilization in nonautoimmune B6 mice but not in diabetes-prone NOD mice. Ablation of HSCs impaired tolerance to allogeneic islet transplants in B6 recipients. Mobilization of HSCs resulted in part from decreasing osteoblast expression of HSC retention factors. Furthermore, HSC mobilization required a functioning sympathetic nervous system; sympathectomy prevented HSC mobilization and completely abrogated tolerance induction. NOD HSCs were held in their niche by excess expression of CXCR4, which, when blocked, led to HSC mobilization and prolonged islet allograft survival. Overall, these findings indicate that the HSC compartment plays an underrecognized role in the establishment and maintenance of immune tolerance, and this role is disrupted in diabetes-prone NOD mice. Understanding the stem cell response to immune therapies in ongoing human clinical studies may help identify and maximize the effect of immune interventions for type 1 diabetes.