Wnt5a/FZD4 Mediates the Mechanical Stretch-Induced Osteogenic Differentiation of Bone Mesenchymal Stem Cells

Background/Aims: Mechanical stimulation and WNT signalling have essential roles in regulating the osteogenic differentiation of bone marrow stromal cells (BMSCs) and bone formation. However, little is known regarding the regulation of WNT signalling molecule expression and therefore the osteogenic differentiation of BMSCs during osteogenesis. Methods: Microarrays of BMSCs from elderly individuals or patients with osteoporosis (GSE35959) from the GEO database were analysed using GeneSight-Lite 4.1.6 (BioDiscovery) and C2 curated gene sets downloaded from Molecular Signatures Database (MSigDB). Realtime PCR and western blotting were used to measure the expression of the indicated genes. ALP and Alizarin red staining were used to evaluate the osteogenesis of BMSCs. Results: In this study, we investigated whether mechanical loading directly regulates the expression of WNT signalling molecules and examined the role of WNT signalling in mechanical loading-triggered osteogenic differentiation and bone formation. We first studied the microarrays of samples from patients with osteoporosis and found downregulation of the GPCR ligand binding gene set in the BMSCs of patients with osteoporosis. Then, we demonstrated that mechanical stimuli can regulate osteogenesis and bone formation both in vivo and in vitro. FZD4 was upregulated during cyclic mechanical stretch (CMS)-induced osteogenic differentiation, and the JNK signalling pathway was activated. FZD4 knockdown inhibited the mechanical stimuli-induced osteogenesis and JNK activity. More importantly, we found an activating effect of WNT5A and FZD4 that regulated bone formation in response to hindlimb unloading in mice, and pretreatment with WNT5A or activation of the expression of FZD4 partly rescued the osteoporosis caused by mechanical unloading. Conclusions: Our results demonstrate, for the first time, that mechanical stimulation alters the expression of genes involved in the osteogenic differentiation of BMSCs via the direct regulation of FZD4 and that therapeutic WNT5A and FZD saRNA may be an efficient strategy for enhancing bone formation under mechanical stimulation.