转化生长因子-β1对MC3T3-E1小鼠成骨细胞骨形成的影响
Effect of transformation growth factor-β1 on osteoblasts and osteogenesis of MC3T3-E1 cells

目的:探讨转化生长因子-β1(TGF-β1)在成骨细胞骨形成中的负向调节作用及其可能作用机制。方法:离体培养MC3T3-E1小鼠成骨细胞系,以10 μg/L的甲状旁腺激素(PTH)或TGF-β1处理细胞6、12、24 h后,应用荧光定量PCR检测硬骨素(SOST)和TGF-β受体ALK5 mRNA表达变化; 取对数生长期的MC3T3-E1细胞进行ALK5-siRNA转染,荧光定量PCR及Western blot法检测ALK5-siRNA转染对TGF-β1作用下MC3T3-E1细胞中SOST mRNA和Smad2/3蛋白表达的影响。结果:在10 μg/L浓度的TGF-β1诱导下,MC3T3-E1细胞中SOST和ALK5 mRNA表达水平均较对照组升高(P<0.05),且表达水平均随着处理时间的延长而上调(P<0.05)。沉默ALK5可部分抑制TGF-β1诱导的SOST mRNA和Smad2/3蛋白表达上调(P<0.05)。结论:TGF-β1可通过激活Smad2/3信号通路上调SOST表达,从而抑制成骨细胞骨形成。
Aim: To investigate the negative regulation of transforming growth factor-β1(TGF-β1)upon osteoblast formation and the mechanism.Methods: The mice osteoblast cell line MC3T3-E1 were cultured in vitro and stimulated with 10 μg/L parathyroid hormone(PTH)or TGF-β1 for 6, 12, 24 h. The expression of sclerostin(SOST)and TGF-β receptor ALK5 mRNA was detected by qRT-PCR. The effect of ALK5-siRNA transfection on SOST mRNA and Smad2/3 protein expression of the MC3T3-E1 cells were detected by qRT-PCR and Western blot, respectively.Results: The expression of SOST mRNA and ALK5 mRNA were significantly up-regulated after 10 μg/L TGF-β1 treatment(P<0.05); and the expression levels were increased with the culture time(P<0.05). The expression of SOST mRNA was significantly decreased while the protein level of Smad2/3 was increased after ALK5 knockdown(P<0.05).Conclusion: TGF-β1 up-regulates the expression of SOST via activating Smad2/3 signaling pathway, thus inhibits osteoblast formation