Effect of Mesenchymal Stem Cells Overexpressing BMP-9 Primed with Hypoxia on BMP Targets, Osteoblast Differentiation and Bone Repair

SIMPLE SUMMARY: Bone formation is regulated by proteins such as bone morphogenetic proteins (BMPs) as well as by oxygen. We previously demonstrated that cell therapy using stem cells genetically modified to express BMP-9 enhances bone formation in skull defects. Here, it was evaluated the effect of these cells submitted to low oxygen tension (hypoxia) on bone cell (osteoblast) differentiation and bone repair. The effect of stem cells expressing BMP-9 on osteoblast differentiation was evaluated using the solution where these cells were grown (conditioned medium). The bone formation induced by stem cells expressing BMP-9 directly injected into rat skull defects was also evaluated. The results demonstrated that the conditioned medium generated under hypoxia favored osteoblast differentiation. Hypoxia-conditioned stem cells expressing BMP-9 did not increase bone repair compared with stem cells expressing BMP-9 under normoxia. Thus, despite the lack of effect of hypoxia on bone formation, the enhancement of osteoblast differentiation can drive further investigations on the regulation of BMP-9 and oxygen availability in the context of therapies to induce bone regeneration, which may improve the clinical treatment of large bone fractures and defects. ABSTRACT: Bone formation is driven by many signaling molecules including bone morphogenetic protein 9 (BMP-9) and hypoxia-inducible factor 1-alpha (HIF-1α). We demonstrated that cell therapy using mesenchymal stem cells (MSCs) overexpressing BMP-9 (MSCs(+BMP-9)) enhances bone formation in calvarial defects. Here, the effect of hypoxia on BMP components and targets of MSCs(+BMP-9) and of these hypoxia-primed cells on osteoblast differentiation and bone repair was evaluated. Hypoxia was induced with cobalt chloride (CoCl(2)) in MSCs(+BMP-9), and the expression of BMP components and targets was evaluated. The paracrine effects of hypoxia-primed MSCs(+BMP-9) on cell viability and migration and osteoblast differentiation were evaluated using conditioned medium. The bone formation induced by hypoxia-primed MSCs(+BMP-9) directly injected into rat calvarial defects was also evaluated. The results demonstrated that hypoxia regulated BMP components and targets without affecting BMP-9 amount and that the conditioned medium generated under hypoxia favored cell migration and osteoblast differentiation. Hypoxia-primed MSCs(+BMP-9) did not increase bone repair compared with control MSCs(+BMP-9). Thus, despite the lack of effect of hypoxia on bone formation, the enhancement of cell migration and osteoblast differentiation opens windows for further investigations on approaches to modulate the BMP-9-HIF-1α circuit in the context of cell-based therapies to induce bone regeneration.