*Purpose: In this study, we were aimed to explore the potential influence of CD19⁺Tim-1⁺ Regulatory B cells (Bregs) on the production of DSAs in a mice skin transplantation model.

*Methods: A mice secondary skin transplantation model was established using the C57BL/6 and Balb/c as the donor and recipients, respectively. Levels of de novo DSAs following the first skin transplant and secondary DSAs after second skin transplant were detected by flow cytometry assay. Levels of Bregs (CD19+Tim-1+), Tregs (CD4+CD25+ Foxp3+), as well as the Th17 cells (CD4+IL-17A+), in peripheral blood monocytes (PBMCs) collected from mice skin transplant recipients were also examined. Various inflammatory cytokines, such as IL-2, IL-4, IL-10, and IL-17, were tested by ELISA assay. To further explore the mechanism involved, primary Bregs were extracted from the spleen of recipients and co-cultured with PBMCs of recipients in vitro. De novo DSAs were detected. Moreover, the Bregs and balance of Th17/Tregs were examined. Finally, the co-culture system was intervened by anti-IL-17 receptor (IL-17R) monoclonal antibody to explore the potential mechanism.

*Results: Along with a higher DSA and a low level of IL-10 after the second skin transplant, Bregs showed significantly lower levels than those after the first skin transplant base on an examination of the allograft. Flow cytometry of splenocytes confirmed the outcomes and showed a decrease of the proportion of Treg cells and an increase of the proportion of Th17 cells after the second skin transplant. Mixed cell culture showed that Bregs infusion could decrease the level of DSAs, as well as lead to the imbalance of Th17/Treg. Importantly, intervention of anti-IL-17R antibody decreased the generation of DSAs.

*Conclusions: In conclusion, our research indicated that Bregs can attenuate the production of DSAs by mediating the imbalance of Th17/Treg through IL-17/IL-17R axis.

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