BACKGROUND: Chinese herbs have been shown to affect osteoblasts through Wnt/β-catenin signaling pathway. Sclerostin (Sost) is an inhibitor of Wnt signaling pathway and an inhibitor of osteogenesis, and it is also an antagonistic factor that regulates the differentiation and proliferation of osteoblasts. Bushen Jianpi Huoxue decoction (BJHD) is the empirical formula in the prevention and treatment of osteoporosis at the Affiliated Orthopedics Hospital, Guangzhou University of Chinese Medicine. BJHD can promote osteogenesis and effectively treat osteoporosis. OBJECTIVE: To observe the effect of BJHD on the cell proliferation, alkaline phosphatase activity and expression levels of related proteins in Sost-overexpressed adenovirus transfected osteoblasts. METHODS: The osteoblasts were divided into two groups: blank control group (empty adenovirus transfection) and Sost overexpressed group (Sost overexpression and adenovirus transfection). Two groups were cultured with blank serum and medicine serum. The cell proliferation was tested by cell counting kit-8 assay, and alkaline phosphatase activity was tested by alkaline phosphatase kit. Expression levels of bone regulation-related proteins were tested by western blot assay. RESULTS AND CONCLUSION: Compared with the blank serum, medicine serum could increase the cell viability at 24, 48 and 72 hours in both groups (P ＜ 0.01). Compared with the blank serum, medicine serum increased the alkaline phosphatase activity (P ＜ 0.01). Compared with the blank serum, medicine serum in the blank control group decreased the expression of low-density lipoprotein receptor-related protein 5 and osteoprotegerin (P ＜ 0.01), but showed no effect on the expression of osteopontin and tumor necrosis factor α. Medicine serum in the Sost-overexpressed group decreased the expression of low-density lipoprotein receptor-related protein 5, osteoprotegerin and osteopontin, increased the expression of tumor necrosis factor α (P ＜ 0.01). These results indicate that BJHD can increase the proliferation of recombinant adenovirus transfected Sost-overexpressed osteoblasts and alkaline phosphatase activity, and regulate the expression of low-density lipoprotein receptor-related protein 5, osteoprotegerin, osteopontin, and tumor necrosis factor α.%背景:有研究发现中药可以通过Wnt/β-catenin 信号通路影响成骨细胞,而Sost是Wnt信号通路的抑制因子和骨形成中的抑制蛋白,也是一种调控骨分化的拮抗因子,可调控成骨细胞的分化和增殖.补肾健脾活血方是广州中医药大学附属骨伤科医院临床防治骨质疏松症的经验方,对骨形成有促进作用,治疗骨质疏松症疗效显著.目的:应用补肾健脾活血方含药血清干预过表达骨硬化蛋白 Sclerostin(Sost)重组腺病毒转染的成骨细胞,观察其对细胞增殖、碱性磷酸酶活性和相关蛋白的影响.方法:将培养好的成骨细胞分为空白对照组、过表达Sost组(Ad-Sost组)2组,空白对照组给予空载腺病毒转染,Ad-Sost组予过表达Sost重组腺病毒转染,两组分别给予空白血清和含药血清干预,采用CCK-8检测细胞增殖、碱性磷酸酶试剂盒检测碱性磷酸酶活性,Western Blot检测骨调节蛋白表达情况.结果与结论:①与空白血清干预比较,两组含药血清在24,48,72 h均可以提高细胞活性(P均＜ 0.01);②与空白血清干预比较,两组含药血清均可以提高碱性磷酸酶活性(P ＜ 0.01);③与空白血清干预比较,空白对照组含药血清可以降低低密度脂蛋白受体相关蛋白5、骨保护蛋白的表达(P均＜ 0.01),对骨桥蛋白、肿瘤坏死因子α蛋白的表达无明显影响,Ad-Sost组含药血清也可以降低低密度脂蛋白受体相关蛋白5、骨保护蛋白、骨桥蛋白蛋白的表达(P均＜ 0.01),提高肿瘤坏死因子α蛋白的表达(P ＜ 0.01);④结果说明,补肾健脾活血方含药血清可以提高过表达Sost转染成骨细胞增殖和碱性磷酸酶活性,影响低密度脂蛋白受体相关蛋白5、骨保护蛋白、骨桥蛋白和肿瘤坏死因子α蛋白的表达.
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