Rapid differentiation of regulatory CD4 + T cells in the infarcted myocardium blunts in situ inflammation

Abstract Background Myocardial infarction (MI) is a sterile inflammatory condition associated with tissue injury that results in the activation of T helper cell targeting cardiac antigens. However, the differentiation trajectories and in situ activity of heart-specific CD4 + T cells activated in the MI context remain poorly understood. Methods Herein, we combined T-cell receptor transgenic models targeting myocardial protein, single-cell transcriptomics, and functional phenotyping to elucidate how the myosin-specific CD4 + T cells (TCR-M) differentiate in the murine infarcted myocardium and ultimately influence tissue repair. Furthermore, we adoptively transferred heart-specific T-cells that were pre-differentiated in vitro towards pro-inflammatory versus regulatory phenotypic states to dissect how they differentially regulate post-myocardial infarction (MI) inflammation. Results Flow cytometry and single-cell transcriptomics findings reveled that transferred TCR-M cells rapidly acquired an induced regulatory phenotype (iTreg) in the infarcted myocardium and blunt local inflammation. Myocardial TCR-M cells differentiated into two main lineages enriched with cell activation and pro-fibrotic transcripts (e.g. Tgfb1 ) or with suppressor immune checkpoints (e.g. Pdcd1 ), which we also found in human myocardial tissue. These cells produced high levels of latency-associated peptide (LAP) and inhibited interleukine-17 (IL-17) responses. Notably, TCR-M cells that were pre-differentiated in vitro towards a regulatory phenotype maintained a stable in vivo FOXP3 expression and anti-inflammatory activity when adoptively transferred prior to MI induction. In contrast, TCR-M cells that were pre-differentiated in vitro towards a pro-inflammatory T H 17 phenotype were partially converted towards a regulatory phenotype in the injured myocardium and blunted myocardial inflammation. Conclusions These findings reveal that the myocardial milieu provides a suitable environment for iTreg differentiation and reveals novels mechanisms by which the healing myocardium shapes local immunological processes.