M1 macrophage-derived CXCL12 drives neurogenic heterotopic ossification following spinal cord injury

Patients with spinal cord injury frequently develop neurogenic heterotopic ossification, whose pathogenesis remains incompletely understood. Existing research models struggle to accurately simulate the complex pathological process. To establish a reliable neurogenic heterotopic ossification research model, elucidate its pathogenesis, and explore early intervention strategies, this study successfully developed a spinal cord injury-induced neurogenic heterotopic ossification mouse model. Significant ectopic bone formation, restricted hip and knee joint mobility, and motor dysfunction were observed, accompanied by elevated expression of the osteogenic markers alkaline phosphatase, runt-related transcription factor 2, sex-determining region Y-box 9, and osteocalcin . Proteomics and quantitative polymerase chain reaction analysis revealed upregulation of chemokine (C-X-C motif) ligand (CXCL)12, C-X-C chemokine receptor type 4 (CXCR4), and LYN proto-oncogene, whereas CXCL1 was downregulated in the neurogenic heterotopic ossification group. In vivo experiments confirmed abnormal accumulation of M1 macrophages in muscles surrounding early ectopic bone tissue, with markedly elevated CXCL12 expression. In vitro studies further revealed that M1 macrophages are the primary source of CXCL12 secretion, and their cell culture supernatants promote the proliferation, migration, and osteoblastic differentiation potential of bone marrow mesenchymal stem cells. Mechanistically, CXCL12 activates the phosphatidylinositol 3-kinase/protein kinase B pathway by binding to the CXCR4 receptor, thereby driving the osteogenic differentiation of bone marrow mesenchymal stem cells. These findings indicate a causal relationship between the pathological process of neurogenic heterotopic ossification following spinal cord injury and the activation of the CXCL12-CXCR4-phosphatidylinositol 3-kinase-protein kinase B signaling axis, which modulates bone marrow mesenchymal stem cell function through M1 macrophage polarization. This study reveals a key mechanism driving the osteogenic differentiation of bone marrow mesenchymal stem cells, providing new directions for early warning and targeted treatment of neurogenic heterotopic ossification following spinal cord injury.