熊果酸对小鼠成牙骨质细胞系OCCM-30细胞分化的促进作用

Promotion effect of ursolic acid on differentiation of mouse cementoblast cell line OCCM-30 cells

目的:探讨熊果酸对小鼠成牙骨质细胞系OCCM-30细胞分化的影响,为牙根吸收的修复提供理论依据。方法:将对数生长期的OCCM-30细胞分为对照组和不同浓度熊果酸组。其中熊果酸组OCCM-30细胞以3种浓度(0.625、1.250和2.500μmol·L^-1)的熊果酸处理,对照组不做任何处理。采用MTT法检测不同时间点(24、48和72h)各组OCCM-30细胞增殖抑制率,碱性磷酸酶(ALP)法检测细胞体外分化和矿化情况,实时定量PCR法检测24 h内骨桥蛋白(OPN)mRNA表达水平,Westernblotting法检测给药3和5d时OPN蛋白表达水平。结果:不同浓度熊果酸组与对照组OCCM-30细胞增殖抑制率比较差异无统计学意义(P>0.05)。与对照组比较,2.500 μmol·L^-1熊果酸组熊果酸处理3、5和7d后,OCCM-30细胞中ALP活性明显升高(P<0.05),不同浓度熊果酸组OCCM-30细胞中OPNmRNA和蛋白表达水平明显升高(P<0.01)。结论:一定浓度熊果酸对小鼠成牙骨质细胞系OCCM-30细胞增殖无明显抑制作用,但可促进其分化并上调OPNmRNA和蛋白表达水平。

Objective : To study the effects of ursolic acid on the differentiation of cementoblast cell line OCCM-30 cells, and to provide the theoretical basis for the repair of root restoration. Methods :The OCCM-30 cells in logarithmic growth phase were divided into control group and different concentrations of ursolic acid groups. The OCCM-30 cells in ursolic acid groups were treated with different concentrations(0.625, 1.250 and 2.500 μmol·L^-1 ) of ursolic acid and the cells in control group did not receive any treatment. MTT method was performed to detect the inhibitory rates of proliferation of the OCCM-30 cells in various groups at different time points (24, 48, and 72 h);ALP (alkaline phosphatase) assay was performed to detect the cell differentiation and mineralization;real-time quantitative PCR was used to determine the expression levels of osteopontin(OPN) mRNA during 24 h;the expression levels of OPN protein at 3 and 5 d after administration were detected by Western blotting method. Results : There were no significant differences in the inhibitory rates of proliferation of OCCM-30 cells between control group and different concentrations of ursolic acid groups( P >0.05). Compared with control group, the ALP activities in the OCCM-30 cells in 2.500 μmol·L^-1 ursolic acid group at 3, 5, and 7 d after administration were significantly increased( P <0.05).Compared with control group,the expression levels of OPN mRNA and protein in the OCCM -30 cells in different concentrations of ursolic acid groups were significantly increased ( P <0.01). Conclusion : Appropriate concentration of ursolic acid has no effect on the inhibitory rate of proliferation of OCCM-30 cells, but it can promote the differentiation of OCCM-30 cells and up-regul ate the expression levels of OPN mRNA and protein.