Conditioned Medium from Bone Marrow Mesenchymal Stem Cells Transiently Retards Osteoblast Differentiation by Downregulating Runx2

Abstract

Mesenchymal stem cells (MSCs) are attractive candidates for cell therapy and regenerative medicine because of their potential for proliferation and multilineage differentiation. MSCs secrete various cytokines, acting as trophic mediators to regulate neighboring cells. Osteoblasts are the cells directly responsible for forming new bone, and they are the final target of many osteogenic regulators. However, the induction effect of MSCs on osteoblasts is still unknown. In this study, we isolated osteoblasts from rat calvariae and investigated their proliferation and differentiation under induction of varied concentrations of MSC-conditioned medium (MSC-CM). Cells in the MSC-CM groups showed a reduction in cell proliferation at 3–6 days, and a decrease in the expression of osteocalcin and osteopontin at 3 days, with low levels of alkaline phosphatase activity. The expression of osteogenic markers went back to normal at 7 days. In order to evaluate the molecular mechanisms underlying this suppression, levels of two osteoblastic transcription factors, runt-related transcription factor 2 (Runx2) and osterix (Osx), were detected at both mRNA and protein levels. The results indicated that MSC-CM significantly inhibited Runx2 expression in a concentration-dependent manner. However, the effect was not due to the inhibition of Osx, for Osx was not significantly altered. This work demonstrates that MSCs may suppress osteoblast proliferation and transiently retard osteoblast differentiation by downregulating Runx2. These results highlight the need to take into account the paracrine effect of MSCs when using them in regenerative medicine for the repair of bone defects.