A New Osteogenic Neutrophil Subset Maintains Bone Homeostasis

Various immune cells intervene in bone metabolism under physiological or pathological conditions. However, which type of immune cell plays a major role in the maintaining of bone homeostasis and its underlying mechanisms are still unknown. Here, we revealed a positive correlation between neutrophil ratio and human bone mineral density that is not observed in other immune cells. Both neutrophil decline by neutralizing antibody or rudely deletion in Gfi1 −/− mice leaded to apparent bone loss. Single cell RNA-sequencing analysis unveiled a previously unappreciated neutrophil cluster that extensively interacts with osteoblast lineage and facilitate bone homeostasis, namely “osteogenic neutrophil”. Combining single-cell and spatial transcriptomics revealed the aggregation tendency of osteogenic neutrophil to osteo-active areas, indicating its role in maintaining bone homeostasis. Further analysis of secreted cytokines in this novel neutrophil subset and proteomics validation revealed S100a9 as a potential regulatory factor of bone homeostasis. Conditional knock out of S100a9 in neutrophils results in spontaneous bone loss in mice, while supplementary of S100a9 robustly reversed bone density in ovariectomy, neutrophil deletion, and aging-induced bone loss models. Mechanistically, we found that S100a9 binds to Tlr4 and further activates Shp2, thus exerting its role in maintaining bone homeostasis. Knock out of Tlr4 in osteoblast lineage cells abrogated the improved bone phenotype of S100a9 supplementation. Together, these findings reveal a novel osteogenic neutrophil subset maintaining bone homeostasis by secreting S100a9, which refine our understanding of the crosstalk between immune cells and bone metabolism.