Elucidating the Role of Mechanotransduction in SMC Dedifferentiation in Peripheral Artery Disease

About This Grant

PROJECT SUMMARY Atherosclerosis-associated PAD is a leading cause of cardiovascular mortality, driven by SMC dedifferentiation into proliferative and migratory phenotypes. Despite its role in numerous vascular diseases, the mechanisms regulating SMC dedifferentiation remain unclear, limiting effective treatment development. Mechanotransduction, the process by which cells convert mechanical stimuli into biochemical signals, is altered in dedifferentiated SMCs in PAD. In this K99/R00 project, I propose to leverage (i) human induced pluripotent stem cell-derived smooth muscle cells (iPSC-SMCs), (ii) multiomics, (iii) biomaterials with tunable stiffness, and (iv) lineage tracing mouse models, to test the hypothesis that impaired mechanotransduction contributes to SMC dysfunction by translating ECM alterations into transcriptional and epigenetic changes, ultimately increasing susceptibility to atherosclerosis and PAD. In Aim 1, I will utilize soft and stiff hydrogels to explore the effects of impaired mechanotransduction mediators on iPSC-SMCs phenotype by assessing SMC function, transcriptional profile, and downstream mediators (RhoA/ROCK signaling, YAP transcription factor, αβ-integrins, FAK). In Aim 2, I will investigate how dysfunctional mechanotransduction affects SMC dynamics using an SMC-specific lineage tracing mouse model, inducing either atherosclerosis or hind limb ischemia. I will evaluate (i) plaque area (ii) blood flow restoration, (iii) SMC dedifferentiation (single nucleus RNA-seq) at different timepoints. Dysregulated targets will be prioritized by cross-integration of the mouse with the human dataset, and validation will be performed in human PAD “early” and “advanced” plaques. In Aim 3, I will identify upstream noncoding genetic regulators controlling SMC fate in response to vascular stiffness, by single nucleus ATAC-seq of human PAD and control tissues. I will conduct an arrayed CRISPRi screen targeting noncoding regions enriched in PAD compared to controls, followed by mechanistic validation. The proposed research builds upon my prior training in vascular biology and single cell analysis, while providing new opportunities for training on iPSC-SMC technology, animal models and large-scale data integration. My mentor, Dr. Joseph Wu, is a pioneer in iPSC and cardiovascular biology, and my co-mentor, Dr. Thomas Quertermous, is a renowned expert in vascular diseases and genetics, whose mentorship complements Dr. Wu’s expertise. To further strengthen my training, I have assembled an Advisory Committee comprising Dr. Chiara Giannarelli (atherosclerosis and multiomics), Dr. Ronald L. Dalman (peripheral artery disease), and Dr. Michael Snyder (large-scale functional genomics), who will provide specialized guidance and expertise across key aspects of my research. In conclusion, my path to independence is supported by rigorous training, outstanding mentorship, and exceptional research environment at Stanford. These elements will provide me with the expertise and resources needed to achieve my long-term goal of studying the genetics of mechanotransduction regulating stromal cell function in vascular diseases.