Polymeric Three-dimensional Scaffold Promotes Osteogenic Precursor Cells Differentiation through Induction of Periph-eral Macrophages Polarization
Objective:To investigate the effect of three-dimensional scaffold SF-CS-HA on the polarization of mouse macrophages,and explore the impact of bone immunomodulatory microenvironment regulation on the extra-cortical bone formation of mouse calvarial precursor cells.Methods:SF-CS-HA was prepared and co-cultured with mouse macrophages RAW264.7.Flow cytometry was performed to detect the polarization of macrophages into M2 phenotype.RT-qPCR was used to measure the expression of inflammatory cytokine genes,and western blot was conducted to evaluate the expression of TGF-β.After co-incubation of macrophages with SF-CS-HA 3D,condi-tioned medium was prepared.The proliferation,migration,ALP activity,and morphological changes of mouse cal-varial precursor cells MC3T3-E1 were compared between complete medium and conditioned medium.Western blot was performed to detect the expression of bone formation-related proteins.An animal model was used to verify the effect of SF-CS-HA on the increase of skull bone mass in mice.Results:Scanning electron microscopy observations revealed that SF-CS-HA had a uniform porous structure both on the surface and internally.Flow cytometry analysis showed that SF-CS-HA promoted the polarization of peripheral macrophages towards M2 phenotype.RT-qPCR and western blot results demonstrated that SF-CS-HA increased the expression of IL-10 gene and TGF-β protein,while de-creased the expression of IL-1β gene.Additionally,SF-CS-HA promoted the proliferation of MC3T3-E1 cells,signifi-cantly increased ALP activity,enhanced cell migration,and increased the production of cytoskeletal protein Factin.Furthermore,conditioned medium exhibited more significant enhancement in the proliferation and migration capabilities of MC3T3-E1 cells.Moreover,Western blot indicated that under the regulation of the bone immune microenvironment,the expression of bone formation proteins OPN,COLA1,RUNX2,OCN,and TGF-β increased significantly.The results of animal experiments showed that SF-CS-HA could promote new bone formation in the mouse skull in vivo.Conclusion:SF-CS-HA 3D scaffold can promote the polarization of peripheral macrophages and regulate the bone immune microenvironment,thereby promoting os-teogenic differentiation and bone formation in the cortical bone exogenesis.
3D scaffold materialmacrophagespolarizationco-culturecalvarial precursor cells
万方数据
维普
