Regulatory B cells (Bregs) have been shown to play a critical role in immune homeostasis and in autoimmunity models. We have recently demonstrated that combined anti-TIM-1 and anti-CD45RB antibody treatment results in tolerance to full MHC-mismatched islet allografts in mice by generating Bregs that are necessary for tolerance. Bregs are antigen-specific, IL-10-dependent, and are capable of transferring tolerance to untreated, transplanted animals. Transplant tolerance induced by anti-TIM-1/anti-CD45RB is both regulatory B cell-dependent and regulatory T cell-dependent. We hypothesize that regulatory B cells, through their production of TGF-β, induce regulatory T cells. Our aim was to determine whether graft survival in our model was dependent on B cell production of TGF-β.

METHODS

Diabetes was induced in recipients with streptozotocin. Recipient mice received 100 ¯o;g anti-mouse CD45RB ip on days 0, 1, 3, 5, and 7 following transplantation. Recipient mice may also receive 500 ¯o;g anti-mouse TIM-1 (RMT1-10) ip on day -1, and 300 ¯o;g on days 0 and 5 following transplantation. To generate B cell-specific TGF-β mutant mice, CD19-cre mice were mated with floxed-TGF-β mice.

Graft survival between experimental groups was compared using Kaplan-Meier survival curves and Wilcoxon statistics. Foxp3-GFP expt was analyzed using ANOVA. P values less than 0.05 were considered statistically significant.

RESULTS

We demonstrate that adoptively transferred Bregs require the presence of Tregs to establish tolerance, and that adoptive transfer of Bregs increases the number of Tregs. Interaction with Bregs in vivo induces significantly more Foxp3 expression in CD4+CD25- T cells than with naive B cells. We also show that Bregs express the TGF-β-associated latency-associated peptide and that Breg-mediated graft prolongation post-adoptive transfer is abrogated by neutralization of TGF-β activity. In contrast to WT recipients treated with anti-TIM-1/anti-CD45RB antibodies, B cell-specific TGF-β-mutant recipients rapidly reject allogeneic islet grafts. These data suggest that TGF-β production by B cells is critical for graft survival and that with IL-10, TGF-β is necessary for regulatory B cell function.

CONCLUSIONS

Collectively, these findings suggest that in this model of antibody-induced transplantation tolerance, Bregs promote graft survival by promoting Treg development via TGF-β production.

« Back to 2015 American Transplant Congress