Hypoxia-inducible factor 1. may regulate the commitment of mesenchymal stromal cells toward angio-osteogenesis by mirna-675-5P

Background aims. During bone formation, angiogenesis and osteogenesis are regulated by hypoxia, which is able to induce blood vessel formation, as well as recruit and differentiate human mesenchymal stromal cells (hMSCs). The molecular mechanisms involved in HIF-1 alpha response and hMSC differentiation during bone formation are still unclear. This study aimed to investigate the synergistic role of hypoxia and hypoxia-mimetic microRNA miR-675-5p in angiogenesis response and osteo-chondroblast commitment of hMSCs. Methods. By using a suitable in vitro cell model of hMSCs (maintained in hypoxia or normoxia), the role of HIF-1 alpha and miR-675-5p in angiogenesis and osteogenesis coupling was investigated, using fluorescence-activated cell sorting (FACS), gene expression and protein analysis. Results. Hypoxia induced miR-675-5p expression and a hypoxia-angiogenic response, as demonstrated by increase in vascular endothelial growth factor messenger RNA and protein release. MiR-675-5p overexpression in normoxia promoted the down-regulation of MSC markers and the up-regulation of osteoblast and chondroblast markers, as demonstrated by FACS and protein analysis. Moreover, miR-675-5p depletion in a low-oxygen condition partially abolished the hypoxic response, including angiogenesis, and in particular restored the MSC phenotype, demonstrated by cytofluorimetric analysis. In addition, current preliminary data suggest that the expression of miR-675-5p during hypoxia plays an additive role in sustaining Wnt/beta-catenin pathways and the related commitment of hMSCs during bone ossification. Discussion. MiR-675-5p may trigger complex molecular mechanisms that promote hMSC osteoblastic differentiation through a dual strategy: increasing HIF-1 alpha response and activating Wnt/beta-catenin signaling.