Vascular Cell Adhesion Molecule 1 Positive Fibroadipogenic Progentiors Promote Adipogenesis In Chronic Limb Threatening Ischemia Patients

Introduction: Chronic limb threatening ischemia (CLTI) is associated with a severe myopathy. A hallmark of skeletal muscle disorders is fatty infiltration. Fatty infiltration leads to progressive muscle dysfunction. The primary cell type implicated as the source of fatty infiltration are fibroadipogenic progenitors (FAPs). Prevention of FAP conversion to adipocytes may represent a novel approach to ameliorating the CLTI myopathy. However, the molecular mechanisms that govern FAP cell fate decisions in the ischemic limb remain unknown. Herein, we sought to identify the mechanisms that promote fatty infiltration in the ischemic limb. Methods: We quantified fatty infiltration in human CLTI patients and in BALB/c (failed regeneration) and C57BL/6 (successful regeneration) mice with Oil Red O staining. To elucidate the mechanisms responsible for this phenotype, we applied single cell RNA-sequencing (scRNA-seq) to C57BL/6 and BALB/c mice at day 10 and 14 following hindlimb ischemia. Next, we FACS isolated candidate FAP sub-populations and assessed apidogenesis in-vitro. Results: We show that ischemic human CLTI skeletal muscle displays increased adipogenesis. Furthermore, BALB/c mice recapitulate the human CLTI adipogenic phenotype. Next, we identified 10 distinct FAP populations in the ischemic limb. We also discovered a candidate FAP subpopulation, VCAM1+ FAPs, that promotes adipogenesis. Finally, we FACS isolated this sub-population and demonstrated in an in-vitro assay that it controls adipogenesis. Conclusions: Our results shed new light on skeletal muscle repair in the CLTI limb. First, we demonstrate that fatty infiltration is a component of the CLTI phenotype. Second, we identify an injury-induced FAP population that is responsible for the phenotype. Taken together, these observations may aid in the development of future therapeutic strategies to block adipogenesis and improve outcomes for CLTI patients.