In the mammalian skeletal system, osteogenesis and angiogenesis are closely linked by type H vessels during bone regeneration and repair. Our previous studies confirmed the promotion of these processes by copper-containing metal (CCM) in vitro and in vivo. However, whether and how the coupling of angiogenesis and osteogenesis participates in the promotion of bone regeneration by CCM in vivo is unknown. In this study, M2a macrophages but not M2c macrophages were shown to be immunoregulated by CCM. A CCM, 316L−5Cu, was applied to drilling hole injuries of the tibia of C57/6 mice for comparison. We observed advanced formation of cortical bone and type H vessels beneath the new bone in the 316L−5Cu group 14 and 21 days postinjury. Moreover, the recruitment of CD206-positive M2a macrophages, which are regarded as the primary source of platelet-derived growth factor type BB (PDGF-BB), was significantly promoted at the injury site at days 14 and 21. Under the stimulation of CCM, mitochondria-derived reactive oxygen species were also found to be upregulated in CD206hi M2a macrophages in vitro, and this upregulation was correlated with the expression of PDGF-BB. In conclusion, our results indicate that CCM promotes the evolution of callus through the generation of type H vessels during the process of bone repair by upregulating the expression of PDGF-BB derived from M2a macrophages.

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To investigate the molecular pathogenesis of bone with osteomyelitis, we developed implant-associated osteomyelitis (IAOM) models in mice. An orthopedic stainless pin was surgically placed in the right femoral mid-shaft of mice followed by an inoculation of S. aureus into the medullary cavity. Typical characteristics of IAOM, like periosteal reaction and intra-osseous abscess, occurred by day 14 post-infection. By day 28 post-infection, necrotic abscess, sequestrum formation and deformity of the whole femur were observed. Transcriptional analysis identified 101 and 1,702 differentially expressed genes (DEGs) between groups by days 3 and 14 post-infection, respectively. Analysis of Gene Ontology and Kyoto Encyclopedia of Genes and Genomes revealed the enrichment of pathways in response to bacterium, receptor-ligand activity and chemokine signaling by day 3 post-infection. However, by day 14 post-infection, the enrichment switched to angiogenesis, positive regulation of cell motility and migration, skeletal system development and cytokine-cytokine receptor interaction. Furthermore, protein-protein interaction network analysis identified 4 cytokines (Il6, Cxcl10, IFN-γ and Cxcl9) associated with IAOM at an early stage of infection. Overall, as the pathological changes in this mouse model were consistent with those in human IAOM, our model may be used to investigate the mechanism and treatment of IAOM. Furthermore, the data of transcriptome sequencing and bioinformatic analysis will be an important resource for dissecting the molecular pathogenesis of bone with IAOM. Highlights 1. An implant-associated osteomyelitis (IAOM) mice model was developed. 2. The pathological changes in this mouse model were consistent with those in human IAOM. 3. 101 and 1,702 differentially expressed genes (DEGs) were identified between groups by days 3 and 14 post-infection, respectively. 4. Il6, Cxcl10, IFN-γ and Cxcl9 were screened out as the most important genes associated with early stage IAOM