Mapping the Cellular Landscape of Calvarial Bone Repair Accelerated by Sympathetic Nerve Inhibition

Understanding the dynamic changes of cells during calvarial bone repair is crucial for identifying novel therapeutic targets to enhance bone regeneration. However, the cellular changes and intercellular communication during calvarial defect repair remain poorly understood. To address this, we performed single-cell RNA sequencing on tissues collected at different time points post-defect. Our analysis revealed a significant enhancement in intercellular communication following defect, particularly among stem cells, endothelial cells, pericytes, and macrophages. Pathways related to neurogenesis were significantly enriched after defect. Furthermore, we found that inhibiting sympathetic nerves promoted calvarial bone repair. Mechanistically, sympathetic nerve inhibition enhanced angiogenesis and osteogenesis by promoting interactions between pericytes and endothelial cells, generating a novel senescenced Arg1+ macrophages, which contributed to bone repair by secreting osteogenesis-related cytokines. Besides, inhibition of sympathetic nerves promotes the generation of Shisa3+ suture cell subpopulation and enhances osteogenic differentiation capacity. Importantly, senolytics abrogated the repair benefits brought about by sympathetic nerve inhibition, underscoring the critical role of senescent macrophages in the repair process. Overall design: To further investigate how sympathetic nerve inhibition promotes calvarial bone defect repair, single-cell RNA sequencing was performed on calvarial bone samples collected 7 and 14 days after frontal bone defect induction with or without sympathetic nerve inhibition.