Abstract 13109: Epigenetic Regulation and Function of Pleiotropic GWAS Locus LRP1 in Human Induced Pluripotent Stem Cells Derived Contractile Smooth Muscle Cells

Introduction: Spontaneous coronary artery dissection (SCAD) is an increasingly recognized cause of myocardial infarction in young and middle-aged women. A common genetic variant, rs11172113, located in LRP1 (low density lipoprotein receptor-related protein 1) first intron, was associated with several vascular diseases, including SCAD, and regulation of LRP1 expression in arterial tissues. However, the biological mechanisms through which rs11172113 influence LRP1 function in the context of non-atherosclerotic arterial lesions are not known. Hypothesis: We hypothesized that rs11172113 genotype affects LRP1 expression through specific mechanisms in contractile smooth muscle cells (SMCs), leading to alterations of their physiological function. Methods: We differentiated 4 human induced pluripotent stem cells (iPSCs) lines (2 males, 2 females) towards either contractile or synthetic SMCs using a recently optimized 24-day protocol. We performed RNA-Seq and ATAC-Seq at 6 time points during differentiation. We silenced LRP1 using siRNAs in differentiated SMCs and used CRISPR-Cas9 to induce deletions of the enhancer region containing rs11172113 or frame-shift indels in exons 2 or 5 of LRP1 in iPSCs. Results: iPSCs derived SMCs presented the expected morphology and expression of SMC markers. Transcriptomic analyses showed that contractile SMCs harbored SMC properties during late differentiation and resembled SMCs of artery tissue, whereas genes involved in cell stress were upregulated in synthetic SMCs at the end of differentiation. In particular, LRP1 expression and DNA accessibility at rs11172113 were higher in contractile SMCs. The deletion of rs11172113 enhancer region led to a decrease in the expression of LRP1 in both contractile and synthetic SMCs. LRP1 inactivation decreased the proliferation and increased the migration capacity of contractile SMCs. Detailed characterization of the influence of LRP1 on extracellular matrix maintenance and cellular contraction is ongoing. Conclusions: We confirmed rs11172113 to regulate LRP1 expression in iPSCs derived contractile SMCs. Our results support LRP1 influence on SMCs phenotype as a causal factor in the genetic susceptibility to SCAD and related vascular diseases.