Predominant Role of PDGF Receptor Transactivation in Wnt3a-Induced Osteoblastic Cell Proliferation

Previous studies have shown that Wnt3a enhances the proliferation and inhibits the osteogenic differentiation of human mesenchymal stem cells (hMSCs). In this study, we investigated the signaling pathways involved in Wnt3a-induced osteoblastic cell proliferation. Experiments with DKK1, a natural antagonist of Lrp5/6, indicated that Wnt/beta-catenin did not play a major role in Wnt3a-induced osteoblastic cell proliferation. The use of selective inhibitors of known mitogenic pathways implicates Src family kinases (SFKs) and a protein kinase C (PKC) in this cellular response. Time-dependent analysis of signaling molecules activated by Wnt3a in MC3T3-E1 cells revealed parallel activation of the canonical pathway and of several tyrosine kinases, including SFKs and PDGF receptors (PDGF-Rs). Functional analysis with specific inhibitors suggested a major role of PDGF-Rs in mediating Wnt3a-induced cell proliferation. Further investigation with an si-RNA approach confirmed a predominant role of this receptor in this cellular response. The use of soluble decoy PDGF-Rs that can sequester extracellular PDGFs excluding that part of the increased PDGF receptor phosphorylation by Wnt3a was the result of autocrine production of PDGFs. A selective SFK inhibitor blunted the enhanced PDGF-R phosphorylation and cell proliferation induced by Wnt3a. Studies of initial events involved in the regulation of this pathway suggest a role of dishevelled. In conclusion, data presented in this study indicate that cell proliferation induced by Wnt3a in osteoblastic cells is mediated by a dishevelled-dependent and beta-catenin-independent pathway, which involves the transactivation of PDGF receptors. (C) 2013 American Society for Bone and Mineral Research.