Background: widespread maternal microchimerism (MMc) in mice is associated with development of immune regulation, which can be used to predict transplantation tolerance. We recently showed that membrane allo-antigen acquisition (mAQ) by host dendritic cells (DCs) is associated with a type of split tolerance, namely, functionality of semi-direct pathway and anergy of the indirect pathway. Hypothesis: in offspring with MMc, production of exosomes (EXO) by maternal cells leads to mAQ by host DCs and changes in their functional status. This leads to anergy/regulation in the indirect allo-response to non-inherited maternal antigens (NIMAs). Methods: NIMAd–exposed [BDF female x B6 male] F1 backcross H2bxb mice were used. Anti-Kd Ab was used to screen mAQ on splenic myeloid DCs (mDC), while YAe, an Ab specific for the Eα52-68 peptide-IAb complex, was used to look for allo-presentation on plasmacytoid DCs (pDCs). DCs phenotype was determined using Ab to PD-L1, CD11b, CD86. To analyze the regulation of the indirect allo-response using Trans-Vivo Delayed Hypersensitivity Assay, splenocytes were transferred into the footpad of B6 mice, either with PBS, tetanus toxoid (TT), BDF1 Ag or TT+BDF1 Ag. Abs to TGFβ, IL-10, Ebi3 and IL12p35 (subunits of IL-35) were co-injected to assess mechanism of regulation. EXO were isolated from serum of B6, mAQ+ NIMAd, or mAQneg NIMAd mice, then cultured in vitro with B6 splenocytes. After 72 hrs the cells were analyzed to detect mAQ. Results: there was an association between mAQ and linked suppression of TT response [P=0.016]; Abs to IL-10, IL-35 and TGFβ abrogated the regulation. Addition to B6 splenocytes of EXO derived from mAQ+, but not from mAQneg mice, showed that mDCs and pDCs could develop detectable mAQ in vitro. Only mDCs were able to retain surface mAQ expression in vivo. pDCs from mice with mAQ+ mDCs showed higher PD-L1 and lower YAe expressions than pDCs from mAQneg mice. Conclusions: indirect pathway Tregs development was associated with mAQ, with IL-10, IL-35 and TGFβ mediating the regulation. In vivo, pDCs show a higher PD-L1 expression and a decreased capacity of indirect allo-presentation in mice with mAQ+ mDCs. Even though pDCs do not retain the surface allo-antigen, it appears they also receive a tolerogenic influence via EXO. Because the mAQ phenomenon can be generated in vitro without the need of allogeneic cells, but only circulating EXO from mAQ+ mice, we suggest their key role as the “missing link” between MMc and tolerance.

« Back to 2015 American Transplant Congress