Profiling site specific TCR diversity of CD4+ T Cells in mice undergoing cardiac dysfunction

Purpose: Despite the well-established association between T cell-mediated inflammation and non-ischemic heart failure (HF), the specific mechanisms triggering T cell activation during the progression of HF and the antigens involved are poorly understood. We hypothesized that clonal expansion of CD4+ T cell in response to cardiac neoantigens promote the progression of HF. Methods: We used transverse aortic constriction (TAC) in mice to trigger HF and profiled the TCR repertoire by mRNA sequencing of CD4+ T cells from 3 pooled Nur77GFP reporter mice, which transiently express GFP upon TCR engagement. We performed TCR clonal analysis of 5000 TCR-activated GFP+CD4+ T cells sorted from the LV, and 5000 CD4+ T cells sorted from the thymus, the inguinal lymph nodes (iLNs) and the heart-draining mediastinal lymph nodes (mLNs) by bulk RNA sequencing of the TCR beta chain after 8wks of TAC surgery. Paired‐end 300 base pair reads were obtained by Illumina sequencing and aligned and assembled using MiXCR software. Results: As expected, the greatest number of unique TCR beta clonotypes (4412 clones) – and therefore the highest TCR diversity – was identified in the thymus. The heart draining mLNs showed a much lower degree of TCR clonal diversity relative to the peripheral iLNs, suggesting that cardiac antigen-driven clonal expansion occurs at the mLNs that directly drain the heart. TCR-activated GFP+CD4+ T cells in the LV represented even lower TCR diversity relative to other sites. Conclusion: Collectively, our study demonstrates that in response to cardiac pressure overload, CD4+ T cells clonally expand in the heart-draining mLN; whilst a limited repertoire of CD4+ T cell clones engage endogenous antigens in the heart. TCR profiling of CD4+ T cells in 3 pooled Nur77GFP reporter mice after 8 weeks of TAC surgery.