Abstract Body: Background: Vascular diseases are a leading cause of mortality worldwide. Epidemiological data suggest significant sex-based differences in vascular disease risk, but the mechanisms underlying these differences remain unclear. Clinical observations indicate that key sex-specific differences may exist in the vessel wall.

Hypothesis: We hypothesize that sex differences in vascular diseases are regulated by sex-specific epigenetic factors/features that alter cellular responses to pathologic triggers in the vessel wall.

Methods: Single cell transcriptomic (scRNA-seq) and epigenetic (scATAC-seq) analyses were performed on male (M) and female (F) ApoE mice at different stages of atherosclerosis progression (0, 16 wks) to uncover sex-specific responses to vascular insult. Comparative analyses were performed on M and F human coronary arteries to identify biologically conserved sex-specific regulatory features shared between humans and mice. These features were integrated with human genetic data to identify the biological processes and regulatory regions driving sex-specific responses. Key findings were subsequently validated in vitro using primary human vascular cells.

Results: Transcriptomic differences between M and F cells were most profound in smooth muscle cells (SMC) and fibroblasts in human and murine arteries. In mouse, the sex-specific transcriptomic differences drastically increased after high fat diet exposures, with number of differentially expressed genes increasing from 29 to 140 in SMC, and 63 to 181 in fibroblasts. Furthermore, we found F SMC to more readily undergo phenotypic modulation despite lower lipid levels. Gene set enrichment analysis showed increased pathways related to extracellular matrix organization and epigenetic modification, as well as key regulators of PDGF and TGFB signaling in mice and human female SMC. Additionally, scATAC-seq identified thousands of sex-specific differentially accessible regions enriching for human genetic signals of vascular diseases. In particular, X-escapee genes Kdm6a and Kdm5c were identified which may work in concert to alter the epigenetic landscape.

Conclusion: This study reveals significant sex-specific, vascular cell-type specific transcriptional and epigenetic mechanisms in vascular disease. These findings provide insights into the genetic and molecular basis of sex differences in vascular diseases and potential therapeutic targets.