目的 观察植物雌激素香豆雌酚对成骨细胞增殖分化的作用并探讨其作用机制.方法 从小鼠颅盖骨获得成骨细胞并用0,10-9～10-5 M香豆雌酚孵育48 h,以17β雌二醇为阳性对照,用酶消化法测定碱性磷酸酶及Ⅰ型胶原含量,放免法测定骨钙素含量,RT-PCR法测定OPG及RANKL mRNA表达情况,Western Blot测定OPG蛋白含量.结果 干预48 h,不同浓度香豆雌酚呈剂量依赖性增加碱性磷酸酶及Ⅰ型胶原含量,10-6 M时达到最大效应(P<0.05),但10-5 M效应有所降低,香豆雌酚轻度增加成骨细胞骨钙素含量,各组间无统计学差异.香豆雌酚呈剂量依赖性增加OPG基因及蛋白的表达(P<0.05),轻度降低RANKL基因的表达.结论 香豆雌酚能增加成骨细胞增殖及分化,可能其部分通过OPG/RANKL发挥作用.%Objective To observe the effect of phytoestrogen coumestrol on the function of osteoblast including ALP and osteocalcin type I collagen, OPG and RANKL. Methods Osteoblasts obtained from fetal mouse calvaria and were incubated with coumestrol (0, 10-9-10-5 M) or 17β-Estradiol (10-8 M) for 48 hours. Alkaline phosphatase (ALP) activity and type I collagen contents was measured by enzyme digestion. Osteocalcin production was measured by radioimmunity. OPG and RANKL mRNA level were detected by RT-PCR and OPG protein was determined by Western Blot. Results Treated with coumestrol for 48 hours, ALP activity and type I collagen contents was dose-dependently increased by all kinds concentration of coumestrol and reached peak at 10-6 M (P<0.05) and then decreased slightly when cotreated with 10-5 M coumestrol. Coumestrol increased osteocalcin expression slightly, but there were no significant differences among groups. Coumestrol increased OPG mRNA expression and protein production in dose-dependently manner (P<0.05) and slightly decreased RANKL mRNA expression. Conclusions Coumestrol has an enhancing effect on the proliferation and differentiation of osteoblasts, at least in part, by stimulating OPG/RANKL expression in osteoblasts.
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