High-Dose TGF-β1 Impairs Mesenchymal Stem Cell-Mediated Bone Regeneration via Bmp2 Inhibition.

Transforming growth factor-β1 (TGF-β1) is a key factor in bone reconstruction. However, its pathophysiological role in non-union and bone repair remains unclear. Here we demonstrated that TGF-β1 was highly expressed in both C57BL/6 mice where new bone formation was impaired after autologous bone marrow mesenchymal stem cell (BMMSC) implantation in non-union patients. High doses of TGF-β1 inhibited BMMSC osteogenesis and attenuated bone regeneration in vivo. Furthermore, different TGF-β1 levels exhibited opposite effects on osteogenic differentiation and bone healing. Mechanistically, low TGF-β1 doses activated smad3, promoted their binding to bone morphogenetic protein 2 (Bmp2) promoter, and upregulated Bmp2 expression in BMMSCs. By contrast, Bmp2 transcription was inhibited by changing smad3 binding sites on its promoter at high TGF-β1 levels. In addition, high TGF-β1 doses increased tomoregulin-1 (Tmeff1) levels, resulting in the repression of Bmp2 and bone formation in mice. Treatment with the TGF-β1 inhibitor SB431542 significantly rescued BMMSC osteogenesis and accelerated bone regeneration. Our study suggests that high-dose TGF-β1 dampens BMMSC-mediated bone regeneration by activating canonical TGF-β/smad3 signaling and inhibiting Bmp2 via direct and indirect mechanisms. These data collectively show a previously unrecognized mechanism of TGF-β1 in bone repair, and TGF-β1 is an effective therapeutic target for treating bone regeneration disability.