Resolving ScRNA-Seq Signatures of Antigen-Specific CD4⁺ T Cells in Tolerance across Semi-Allogeneic Transplantation...

Transplantation of allogeneic organs requires lifelong immunosuppression to prevent rejection. Prior sensitization and resultant memory T cells are barriers to achieving successful transplant tolerance. In reproductive immunology by contrast, pregnancy represents a spontaneous model of tolerance where the semi-allogeneic fetus evades rejection even in multiparous or rejection-sensitized mothers. CD8⁺ T cell phenotypes of tolerance and rejection have been previously reported in transplant and pregnancy, but the transcriptional states of donor and fetus-specific CD4⁺ T cells remain poorly defined. Here, we performed Single-cell RNA-sequencing on endogenous, antigen-specific CD4⁺ T cells across models of allogeneic heart transplants and naïve or paternal skin-sensitized pregnancy. We identified expanded T follicular helper (Tfh) and non-follicular effectors in transplant rejection absent in tolerance. Naïve pregnancy resulted in a modest expansion of effector clusters with transcriptional quiescence that mirrored virgin mice. Successful sensitized pregnancy resulted in expanded Tfh clusters consistent with increased fetal-specific antibodies and limited non-Tfh effector responses. Most striking were the extensive changes imposed on donor-specific Foxp3 ^pos regulatory T cells (Tregs) resulting in the co-clustering together with Foxp3 ^neg T conventional cells (Tconvs) in transplant tolerance and the emergence of a Foxp3 ^neg Type I Regulatory cluster observed in pregnancy of sensitized dams. Finally, we showed that these transcriptomes were relevant and enriched in human datasets of health and disease respectively. Thus, the context-dependent signatures of antigen-specific CD4⁺ T cells provide new insights into their divergent responses to allogeneic conflict at the intersection of transplant and reproductive immunology.