Abstract Body: Smooth muscle cells (SMCs) play important roles in arterial homeostasis, disease, and aging through their remarkable phenotypic plasticity. Traditionally, cardiovascular disease has been considered a disease of chronological aging, but emerging evidence suggests that it reflects accelerated biological aging within the arterial wall. With advancing age, SMCs accumulate epigenetic alterations such as DNA methylation (DNAm) that serve as molecular records of cellular aging. Epigenetic clocks based on DNAm patterns provide powerful biomarkers of biological aging, but existing clocks are derived from heterogeneous tissues and therefore, cannot resolve aging dynamics within individual vascular cell populations. Whether SMCs possess an intrinsic epigenetic aging program and how cellular plasticity or transient reprogramming influences this program remain unclear.
To address these questions, we generated genome-wide DNAm profiles from primary aortic SMCs isolated at 11 time points in male and female mice from 0.5 days to 28 months of age and used these data to construct an SMC-specific epigenetic clock. DNAm was measured using the Illumina Mouse Methylation BeadChip, and elastic net penalized regression modeling was used to identify age-associated CpGs to develop predictive aging models. In parallel, primary aortic SMCs isolated from 9-week-old mice were cultured in vitro and profiled for DNAm patterns across passages 2–11 to generate a passage clock. The SMC epigenetic clock accurately predicted chronological age in vivo with strong correlations between predicted and chronological age in training and testing datasets (Figure 1, r = 0.937 and 0.904, respectively), and passage associated aging in culture (Figure 2, r = 0.999 and 0.998, respectively). Thus, SMCs exhibit a robust and measurable cell type-specific epigenetic aging trajectory that is not captured by existing tissue-level clocks.
This study establishes a framework for measuring epigenetic aging within vascular SMCs and provides a platform for investigating mechanisms that regulate vascular aging. Ongoing studies will leverage this clock to evaluate the contributions of SMC differentiation state, transient reprogramming, and atherosclerosis to epigenetic aging dynamics.