Effect of GARP on osteogenic differentiation of bone marrow mesenchymal stem cells via the regulation of TGFβ1 in vitro

PeerJ. 2019 May 23:7:e6993. doi: 10.7717/peerj.6993. eCollection 2019.

Abstract

Mesenchymal stem cells (MSCs), which have multipotential differentiation and self-renewal potential, are possible cells for tissue engineering. Transforming growth factor β1 (TGFβ1) can be produced by MSCs in an inactive form, and the activation of TGFβ1 functions as an important regulator of osteogenic differentiation in MSCs. Recently, studies showed that Glycoprotein A repetitions predominant (GARP) participated in the activation of latent TGFβ1, but the interaction between GARP and TGFβ1 is still undefined. In our study, we successfully isolated the MSCs from bone marrow of rats, and showed that GARP was detected in bone mesenchymal stem cells (BMSCs). During the osteogenic differentiation of BMSCs, GARP expression was increased over time. To elucidate the interaction between GARP and TGFβ1, we downregulated GARP expression in BMSCs to examine the level of active TGFβ1. We then verified that the downregulation of GARP decreased the secretion of active TGFβ1. Furthermore, osteogenic differentiation experiments, alkaline phosphatase (ALP) activity analyses and Alizarin Red S staining experiments were performed to evaluate the osteogenic capacity. After the downregulation of GARP, ALP activity and Alizarin Red S staining significantly declined and the osteogenic indicators, ALP, Runx2, and OPN, also decreased, both at the mRNA and protein levels. These results demonstrated that downregulated GARP expression resulted in the reduction of TGFβ1 and the attenuation of osteoblast differentiation of BMSCs in vitro.

Keywords: Bone marrow mesenchymal stem cell; Bone regeneration; GARP; TGFβ1.

Grants and funding

This work was supported by Science Researching Project of the National Science Foundation of China “Study on the effect of GARP-TGF-beta1 complex in inhibiting the inflammatory alveolar absorption by bone marrow mesenchymal stem cells”, 2017/01-2020/12. (No. 8167040094), Science Researching Project of the National Science Foundation of China “The effect of AdipoRon in improving the osseointegration of dental implant in diabetic patients and its relative mechanisms”, 2019/01-2022/12.(No.81870793), and the Shanghai Science Commission project “Biological effects on osteoblast of the structure of femtosecond laser produced micro-grooves on the surface of zirconia dental implants”, 2016/07-2019/06.(No.16520710400). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.