Connective tissue growth factor enhances TGF-β1-induced osteogenic differentiation via activation of p38 MAPK in mesenchymal stem cells

Objectives

Cellular differentiation is based on the effects of various growth factors. Transforming growth factor (TGF)-β1 plays a pivotal role in inducing osteogenic differentiation of mesenchymal stem cells (MSCs). In this study, we investigated the influence of connective tissue growth factor (CTGF), known to function synergistically with TGF-β1, on osteogenic differentiation in MSCs.

Methods

UE7T-13 cells were treated with TGF-β1 and/or CTGF. Subsequently, protein levels of intracellular signaling pathway molecules were determined through western blot analysis. The mRNA expression levels of osteogenic differentiation markers were investigated using reverse transcription-quantitative polymerase chain reaction. Bone matrix mineralization was evaluated through alizarin red staining.

Results

Co-treatment with TGF-β1 and CTGF resulted in the suppression of TGF-β1-induced phosphorylation of extracellular signal-regulated kinase 1/2, an intracellular signaling pathway molecule in MSCs, while significantly enhancing the phosphorylation of p38 mitogen-activated protein kinase (MAPK). In MSCs, co-treatment with CTGF and TGF-β1 led to increased expression levels of alkaline phosphatase and type I collagen, markers of osteogenic differentiation induced by TGF-β1. Osteopontin expression was observed only after TGF-β1 and CTGF co-treatment. Notably, bone sialoprotein and osteocalcin were significantly upregulated by treatment with CTGF alone. Furthermore, CTGF enhanced the TGF-β1-induced mineralization in MSCs, with complete suppression observed after treatment with a p38 MAPK inhibitor.

Conclusions

CTGF enhances TGF-β1-induced osteogenic differentiation and subsequent mineralization in MSCs by predominantly activating the p38 MAPK-dependent pathway.