Fate determination of msc by inflammatory niche in HO [spatial transcriptomics]

Heterotopic ossification (HO), usually following traumatic challenges, is initiated by the aberrant osteochondral differentiation of mesenchymal stem cell (MSC). However, the role of trauma-induced inflammatory exposure in MSC fate determination remains unclear. Here, we found that quiescent MSC transited into cycling MSC and later gave rise to chondrogenic (cMSC) and/or osteogenic (oMSC) commitment following either muscle or tendon injury via single-cell RNA sequencing and lineage tracing analysis. Further spatial transcriptome analysis revealed that cycling MSC, cMSC and oMSC resided in the inflammatory niche comprised of monocytes/macrophages, neutrophils, dendritic cells (DCs), NK cells, T and B lymphocytes. We uncovered the diversity of each type of immune cells during MSC development and demonstrated that macrophages, including M1 and M2 subtypes, dynamically promotes proliferation and osteochondral differentiation of MSCs. Additionally, neutrophils and NK cells are essential for cycling MSC transition from quiescent MSC. CD4+ and CD8+ T lymphocytes promotes chondrogenesis. CellchatDB analysis and regulon analysis identify several gene networks, which could be essential for aberrant MSC proliferation and osteochondral differentiation respectively. Collectively, our findings indicate that inflammation is necessary to regulate MSC fate and uncovered the molecular landscape of osteoimmunological interactions that occur during aberrant osteochondral differentiation that could be useful for HO treatment. Single cell RNA-sequencing and spatial transcriptome assays were used to examine the strucutre of inflammatory niche in the HO and depletion of each type immune cells via chemical or genetic approaches was applied to assess the function of inflammatory niche in MSC fate determination.