59: The Immune Regulatory Effect of CpG-ProB Cells on Allograft Survival

Introduction: Previous studies have identified a population of cells emerged when murine bone marrow cells were stimulated with CpG-B (c-kitlowSca-1lowCD19+ B220+CD1dhighIgM-CD5-). The CpG-ProBs were previously reported to be able to prevent diabetes in NOD mice and block progression of experimental autoimmune encephalomyelitis in C57BL/6 mice through immunoregulation. We therefore, hypothesize that CpG-ProBs might be able to suppress allo-immune response and prolong allograft survival in islet transplantation. Materials and Methods: C57BL/6 (CD45.2) STZ diabetic mice were used as recipients, and transplanted with DBA/2 islets. The animals were divided into two groups, in which control group received PBS and the experimental group received C57BL/6 (CD45.1) CpG-ProBs the day of the transplantation. Graft survival was determined by blood glucose monitoring and all animals were sacrificed on day 10 post-transplantation (when all controls were rejected). The lymphoid organs and allografts were harvested and tracking studies were performed to evaluate the CpG-ProBs’ immune suppressive role on allograft survival. Results and Discussion: 50-60,000 CpG-ProBs per transplant were corroborated to be the optimal number of cells that displayed immune suppressive effect. We followed the fate of the CpG-ProBs (CD45.1) into the C57BL/6 mice (CD45.2), and discovered that the progeny of injected cells expanded 2.5 - 3 fold by day 10. Of the recovered cells, 80–90% were TIM-1+, Ki67+, and had various stages of B cell differentiation. Moreover, CpG-ProBs were also recognized to express a certain level of TIM-1 which is a broad marker for Bregs. Ligation of TIM-1 with anti-TIM-1 triggers IL-10 expression, suggesting a link between CpG-ProBs and conventional Bregs. Tracking studies of the CpG-ProBs identified an increased population of CD4+FoxP3+ Treg as well as IL-10-producing cells in lymphoid organs. Proliferation (Ki67+) of CD4+FoxP3+ Tregs increased despite the fact that proliferation of both CD4+ T cells and CD8+ T cells decreased in the lymphoid organs. Histopathology analysis of the grafts in the experimental group showed well identified islets staining positive for insulin. In contrast, grafts from the control group showed no islet cells and polymorphic inflammatory cells at the graft site. Conclusion:Syngeneic CpG-ProBs prolong islets graft survival by immune regulation. The underlying mechanisms possibly involve niche, B progenies, as well as T cell subsets. TIM-1 signaling might participate in CpG-ProBs regulatory effect. NIH UPitt Award Grant (Grant No. 1P01AI129880-01A1). References: 1. Montandon R, Korniotis S, Layseca-Espinosa E, et al. Innate pro-B-cell progenitors protect against type 1 diabetes by regulating autoimmune effector T cells.Proc Natl Acad Sci. 2013; 11;110:E2199-208. 2. Korniotis S, Gras C, Letscher H, et al. Treatment of ongoing autoimmune encephalomyelitis with activated B-cell progenitors maturing into regulatory B cells. Nat Commun. 2016; 11;7:12134. 3. Zavala F, Korniotis S, Montandon R. Characterization and Immunoregulatory Properties of Innate Pro-B-Cell Progenitors. Methods Mol Biol.. 2016;1371:79–88.