Abstract: Objective ClC-3 chloride channels express in osteogenic lineage cells and regulate osteoblast differentiation.This study is designed to observe the effect of TGF-β1 receptor (TβR) on the expression of ClC-3 chloride channels in osteoblasts, and to explore the role of TβR in the ClC-3 chloride channels regulating osteogenic differentiation. Methods CCK-8 kit method and Real-Time PCR method were used to screen the optimal inhibition mode of TβR expression in MC3T3-E1 cells. Then inhibiting TβRI and TβRII expression to observed ClC-3 chloride channels，and down-regulated ClC-3 chloride channels expression to detected the impact on TβRI and TβRII by the siRNA transfection technology. Finally, exploring the effect of TβR on expression of Alp and Runx2 via Real-Time PCR. Results It was found that with the concentration of 2μM and the experimental time of 24h was the optimal TβR inhibition mode, and under this circumstances, the cytotoxicity of TβR inhibitors was within safe levels. Inhibiting TβR expression can promote ClC-3 expression. On the contrary, knocking down ClC-3 chloride channels can promote the expression of TβRI and TβRII. TβR inhibitor can promote the expression of osteogenesis-related genes. Conclusions TβR can inhibit the expression of ClC-3 chloride channel in osteoblasts, and TRβII is more sensitive to the expression of ClC-3 chloride channel.

Key words: ClC-3 chloride channel, TGF-β1 receptor, Osteogenic differentiation, MC3T3-E1 cells