Adventitial Fibroblast Phenotypic Modulation in Atherosclerosis

New avenues of therapy for atherosclerotic diseases are needed as they remain the leading killers worldwide. Therapies that target each of the different cell types that influence the formation of atherosclerotic plaque have translated into lifesaving therapies. Characterizing a novel key cell population that plays a pathogenic role in atherosclerosis will open entirely new avenues for therapies. Our preliminary single cell transcriptomic data and histology of human and murine atherosclerotic plaque have identified an understudied population of cells that we define as Adventitial Fibroblasts (AdvFib). Enabled by an innovative genetic tool, our preliminary data has unveiled a previously unrecognized non-cell-autonomous role of AdvFib in plaque biology. AdvFib ablation or modification of AdvFib through GWAS gene TCF21 significantly influences atherosclerotic plaque formation and calcification. Identification and validation of previously unrecognized outside-in signals affecting atherosclerotic plaque using state-of-the-art techniques form the central focus of this proposal and will lead to novel therapies. Our study is highly innovative by focusing on an overlooked population of vascular cells. No current therapeutic targets these cells. Previous studies of adventitial cells were limited by the inability to deconvolute the causal cell type and relevant disease-related pathology due to cellular heterogeneity of the adventitia and lack of a specific tool to track the fibroblast lineage. Our innovative approach overcomes the shortcomings of previous adventitial studies by using single cell genomics tools, a new and unique murine model that we designed specifically to study vascular adventitial fibroblasts, and complementary in vitro human primary cell models. Aim 1 will determine the mechanism behind the observation that even though AdvFib do not contribute to the intimal lesion, targeted genetic ablation of AdvFib significantly decreases plaque formation. We will use our novel AdvFib-specific inducible CreERT2 murine model, inducible diphtheria toxin receptor, and single cell transcriptomics with mechanistic validation using human AdvFib co-culture experiments. Aim 2 will determine why conditional AdvFib deletion of Tcf21, a GWAS gene for coronary disease, alters plaque calcification. We will accomplish this through a combination of single cell transcriptomic, epigenetics and in vitro human AdvFib functional epigenetic mapping and detailed mechanistic validations. Aim 3 will test the potential therapeutic potential of AdvFib-specific Tcf21 overexpression on plaque composition and vascular calcification. We will use a combination of single cell transcriptomics using a novel murine model and in vitro human AdvFib studies. Our rigorous team science approach synergizes expertise from diverse disciplines, including coronary artery development, human genetics, epigenomics, and the innate/adaptive immune system. Completion of this project will define the role of AdvFib in atherosclerosis, solidify AdvFib as a new target for therapy, and identify new AdvFib disease-related pathways and genes that may also play a role in other vascular diseases. Project Number: 5R01HL179083-02 | Fiscal Year: 2026 | NIH Institute/Center: National Heart Lung and Blood Institute (NHLBI) | Principal Investigator: Paul cheng | Institution: STANFORD UNIVERSITY, STANFORD, CA | Award Amount: $686,252 | Activity Code: R01 | Study Section: Atherosclerosis and Vascular Inflammation Study Section[AVI] View on NIH RePORTER: https://reporter.nih.gov/project-details/5R01HL17908302