Single Cell Profiling Reveals Unique Contributions of Senescent Macrophages Across Musculoskeletal Tissues in Posttraumatic Osteoarthritis

Injury to the acute cruciate ligament (ACL) disrupts musculoskeletal tissue architecture and function, leading to atrophy and weakness associated with the development of posttraumatic osteoarthritis (PTOA). The objective of this study was to profile senescent cell burden across skeletal muscle and knee joint capsule compartments and to ascertain the effectiveness of senolytic treatment in mitigation of PTOA. Given the unique features of these tissues, we hypothesized that composition and physiological consequence of senescent cell burden would differ among muscle and joint capsule, and senolytic treatment would prevent cartilage degradation and improve muscle function. We employed a surgical ACL transection model (ACLT) and performed scRNAseq analysis on the quadriceps and joint capsule 14-days post-injury and discerned senescent status by SPiDER-βGal. We also examined the influence of senescent cells on neighboring populations in vitro. We investigated whether senolytic treatment (dasatinib and quercetin; D+Q) improves muscle and joint health 28-d post-injury. scRNAseq revealed that macrophages (MΦ) accounted for 83% of senescent cells in the quadriceps and 48% in the joint capsule. Senescent MΦ exhibited an alternatively activated phenotype (M2) . Gene ontology enrichment analysis in the quadriceps showed downregulation of positive regulation of muscle cell differentiation, and upregulation of secretory granule lumen; in the joint capsule, it revealed upregulation of glycosaminoglycan catabolic process and regulation of ossification. In vitro, we found that mesenchymal progenitors incubated with senescent MΦ conditioned media exhibited upregulation of senescence associated secretory phenotype (SASP) genes and extracellular matrix organization-related genes ( i.e.Col1a1 and Lox). We found upregulation of protein glycosylation and downregulation of chondroitin sulfate proteoglycan biosynthetic process pathways. PTOA mice exhibited significant reductions in quadriceps myofiber cross-sectional area, increased accumulation of M2-like Cd163+ MΦ, and increased fibrosis, which were blunted in D+Q-treated mice. D+Q-treated PTOA mice exhibited a trend of improved peak torque compared to vehicle-treated PTOA mice. Histochemical assessment of whole knee joints revealed compromised articular cartilage and moderate to severe PTOA in vehicle-treated mice, which was blunted in D+Q-treated mice. These findings reveal distinct transcriptional profiles of senescent macrophages isolated from unique musculoskeletal tissues, providing critical insight into the heterogeneity of senescent cell programming following traumatic injury. Further, we show that accumulation of senescent alternatively-activated MΦ likely contributes to musculoskeletal dysfunction and development of PTOA, and that senolytics may improve functional recovery. AR072061. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.