Impaired granzyme B-producing regulatory B cells in systemic lupus erythematosus.

Granzyme B (GrB)-producing B cells are proposed to be a kind of regulatory B cells (Bregs) and have been revealed to be implicated in the pathogenesis of autoimmune diseases. Nevertheless, their role in SLE remains elusive. In this study, the frequencies of GrB-producing Bregs in peripheral blood of heathy control (HC) and systemic lupus erythematosus (SLE) were evaluated by flow cytometry, and their correlation with SLE patient clinical and immunological features were analyzed. The expression of GrB in HC and SLE B cells were also further detected by RT-qPCR analysis and ELISpot. The function of GrB-producing Bregs in HC and SLE patients was further investigated by in vitro CD4+ effector T cells-B cells co-culture assays with GrB blockade. We found that GrB-producing Bregs were significantly decreased in SLE patients and correlated with the clinical and immunological features. Moreover, these cells were functionally impaired under SLE circumstance. The negative correlation between GrB-producing Bregs and CD4+ T cells observed in healthy individuals disappeared in SLE patients. In vitro cell co-culture assay further showed that GrB-producing Bregs from SLE patients failed to suppress the Th1, Th2 and Th17 cell inflammatory responses, partially due to the dampened capacity of down-regulating TCR zeta and inducing T cell apoptosis. Taken together, these results revealed the disturbance of GrB-producing Bregs in SLE that might contribute to the disease initiation and progression.

Inhibition of Glycogen Synthase Kinase 3ß Increases the Proportion and Suppressive Function of CD19+CD24hiCD27+ Breg Cells.

CD19+CD24hiCD27+ memory Breg cells exhibit decreased abundance in patients with chronic graft-versus-host disease (cGVHD) after liver transplantation and produce less IL-10 than those from patients without cGVHD and healthy donors. Due to the lack of Breg cells and the difficulty in expanding them in vitro, in mouse models and early human clinical trials, the adoptive transfer of Breg cells to autoimmune diseases is greatly restricted. Glycogen synthase kinase 3ß (GSK-3ß) is a multifunctional serine/threonine (ser/thr) protein kinase that can participate in B cell growth, metabolic activity, and proliferation. Phosphoprotein array analysis showed that p-GSK-3ß-s9 was highly expressed in mBreg cells. Furthermore, here, we demonstrated that GSK-3ß expression in mBreg cells is lower than that observed in B cells by flow cytometry. We found that the treatment of B cells with the specific GSK-3ß inhibitor SB216763 can significantly increase the proportion and immunosuppressive function of mBreg cells in vitro. Nuclear factor of activated T cells (NFAT) is one of a pivotal regulator of gene expression in adaptive immune system. Here, we observed that inhibition of GSK-3ß by SB216763 results in enhanced expression of NFATc1 in B cells, which is essential in regulating the ability of B cells to secrete IL-10. By constructing a xGVHD mouse model, we observed that SB216763-treated mBreg cells effectively prevent xenogeneic GVHD. Here we propose a novel strategy using SB216763 to inhibit GSK-3ß and then enhance the proportion and immunosuppressive function of mBreg cells by increasing the expression of NFATc1. This approach may be used as a therapy to ameliorate GVHD and inflammatory diseases.

B cell-derived IL-10 suppresses inflammatory disease in Lyn-deficient mice.

Lyn kinase deficient mice represent a well established genetic model of autoimmune/autoinflammatory disease that resembles systemic lupus erythematosus. We report that IL-10 plays a crucial immunosuppressive role in this model, modulating the inflammatory component of the disease caused by myeloid and T-cell activation. Double-mutant lyn(-/-)IL-10(-/-) mice manifested severe splenomegaly and lymphadenopathy, dramatically increased proinflammatory cytokine production, and severe tissue inflammation. Single-mutant lyn(-/-)mice showed expansion of IL-10-producing B cells. Interestingly, WT B cells adoptively transferred into lyn(-/-) mice showed increased differentiation into IL-10-producing B cells that assumed a similar phenotype to endogenous lyn(-/-) IL-10-producing B cells, suggesting that the inflammatory environment present in lyn(-/-) mice induces IL-10-producing B-cell differentiation. B cells, but not T or myeloid cells, were the critical source of IL-10 able to reduce inflammation and autoimmunity in double mutant lyn(-/-)IL-10(-/-) mice. IL-10 secretion by B cells was also crucial to sustain transcription factor Forkhead Box P3 (Foxp3) expression in regulatory T cells during disease development. These data reveal a dominant immunosuppressive function of B-cell-derived IL-10 in the Lyn-deficient model of autoimmunity, extending our current understanding of the role of IL-10 and IL-10-producing B cells in systemic lupus erythematosus.