Real time visualization of the differential contribution of medullary thymic epithelial cells versus dendritic cells to central tolerance in the thymus (HEM7P.233)

Abstract

Abstract Tolerance for tissue-restricted antigens (TRAs) is established through interactions of thymocytes with medullary thymic epithelial cells (mTECs) and dendritic cells (DCs) in the thymic medulla, though their relative contributions remain unclear. Previous studies using genetic models to impair antigen presentation by mTECs or DCs showed that either subset could contribute to negative selection; however, because interactions between thymocytes and stromal cells are necessary for maintenance of a differentiated stromal compartment, results from these models may not reflect physiologic contributions of mTECs or DCs. Thus, we adopted a two-photon fluorescence microscopy approach to directly quantify interactions between thymocytes and mTECs or DCs at the outset of negative selection in an intact thymic environment. We find that both mTECs and DCs activate TCR signaling in CD8+ and CD4+ thymocytes in the context of secreted and membrane-bound forms of a negatively selecting TRA. mTECs account for more selection than DCs, regardless of the TRA form, and their contribution is greatest with a membrane-bound TRA. Strikingly, while interactions with mTECs and DCs account for all antigen-driven signaling of CD8+ thymocytes, our data suggest other cells also present TRAs to CD4+ thymocytes. Altogether, we find that mTECs make the most significant contribution to negative selection against Aire-driven TRAs, though DCs also contribute in the context of a physiologic thymic microenvironment.