Background: ACTA2-related aortic disease represents 14% of patients with no syndromic HTAAD. ACTA2 gene encodes a-actin protein, specific protein of vascular smooth muscle cell (VSMC) involved in mechano-transduction through the interaction of actin-myosin filaments with the extracellular matrix (ECM), and the regulation of transcription of others contractile genes. Clinical practice is difficult due to incomplete penetration of ACTA2 pathogenic variants. Among them, one is highly penetrant: ACTA2R179X/+. This variant leads to a rare genetic vascular disease: the multisystemic smooth muscle cell dysfunction syndrome (MSMDS). This syndrome is characterized by a congenital dysfunction of smooth muscle throughout the body, manifesting with aortic, cerebrovascular, intestinal, pulmonary and bladder anomalies. Currently, an increasing number of newly identified variants are being classified as “MSMDS-like”, exhibiting a phenotype that overlaps with the syndrome.

Objective: We hypothesize that the location of the mutated ACTA2 variant and the genetic background may cause distinct functional alterations, leading to varied clinical manifestations in patients.

Methods: To decipher the molecular and cellular mechanisms related to ACTA2 rare variants and provide novel insight into the genotype-phenotype correlations, we used a VSMC model produced by differentiation of human induced pluripotent stem cells (hiPSCs) derived from patient cells and edited cells. Four variants were used in this study: p.R179H (MSMDS); p.G148R (“MSMDS-like”); p.P335R and p.R212Q (nsHTAAD). We performed functional analysis of hiPSC-derived VSMCs to examine the cytoskeleton organization, the proliferation, the migration, the contraction and the transcriptome.

Results: Our results show that ACTA2G148R/+ VSMCs from patient cells have smaller and less contractile compared to control VSMCs. Transcriptomic analysis revealed a downregulation of genes implicated in cell adhesion, differentiation and in actin polymerization. ACTA2R179H/+ VSMCs from patient cells have a normal size but have more globular actin than control cells, along with a reduced contractile capacity compared to controls VSMCs, and a transcriptomic analysis is currently underway.

Conclusion: Our hiPSC-derived VSMC model successfully distinguished phenotypic differences among ACTA2 variants in vitro, allowing to further identification of key regulators involved in diseased VSMCs.