Smooth Muscle Cell Stress Response: Essential in Atherosclerotic Pathogenesis

Atherosclerosis remains the leading cause of death worldwide. Recent research using lineage tracing and single-cell RNA sequencing unveiled a previously undescribed and prominent role of smooth muscle cells (SMCs) in atherosclerotic plaque formation. SMCs undergo complex phenotypic modulation characterized by de-differentiation of SMCs and heterogeneously increased expression of markers for macrophages, fibroblasts, chondrocytes, and mesenchymal stem cells as plaques form. Cultured SMCs exposed to exogenous cholesterol mimic many of the phenotypic changes associated with plaque formation, and our lab determined this SMC modulation is driven by cholesterol entering the endoplasmic reticulum (ER) and triggering ER stress and PERK signaling. Further, our lab discovered that heterozygous missense mutations in ACTA2, which encodes smooth muscle-specific α-actin (SM α-actin), predispose to premature coronary artery disease (CAD). In this dissertation, I conducted an in-depth analysis on the SMC stress response in atherosclerosis formation in vivo. We found that SMC-specific deletion of Perk in hyperlipidemic mice reduces atherosclerotic plaque formation by 80% when compared to wild-type mice, emphasizing the role of SMC PERK signaling in plaque formation. We also found that SMCs deficient in Perk had suppressed modulation and migration, and histology confirmed increased medial SMCs density with Perk deficiency. Furthermore, we found a 2.5 times increased burden of Acta2R149C/+Apoe-/- aortas. Our studies identified a novel mechanism by which a mutation in an SMC contractile protein augments plaque burden. SM α-actin with the R149C mutation is misfolded and retained in the cytoplasmic chaperonin containing T-complex polypeptide-1 folding complex in Acta2R149C/+ SMCs. This retention activates heat shock factor 1, which drives endogenous cholesterol synthesis through increased HMG CoA reductase levels, leading to increased cellular cholesterol, ER stress, and PERK signaling in Acta2R149C/+ SMCs. Consequently, Acta2R149C/+ SMCs de-differentiate, migrate, proliferate, and upregulate modulation markers with little to no cholesterol exposure. Additionally, in a hyperhomocysteinemia and hypercholesterolemia mouse model, we demonstrated the pivotal role of SMC involvement in augmenting atherosclerotic plaque burden through a similar pathway. These findings collectively expand our understanding of the molecular mechanisms underlying the role of SMC in the formation of atherosclerosis, presenting novel therapeutic targets that could mitigate the development of atherosclerotic disease.