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目的探讨体外淫羊藿素联合阿霉素对人骨肉瘤MG-63细胞的抑制作用及分子机制。方法设置对照组及不同浓度梯度的淫羊藿素组(10、20、40、80、160μmol/L)、阿霉素组(1、2、4、8、16μg/m L)、联合组(淫羊藿素+阿霉素:20μmol/L+1μg/m L、20μmol/L+2μg/m L、20μmol/L+4μg/m L、40 mol/L+1μg/m L、40μmol/L+2μg/m L、40μmol/L+4μg/m L、80μmol/L+1μg/m L、80μmol/L+2μg/m L、80μmol/L+4μg/m L),分别培养骨肉瘤MG-63细胞用倒置相差显微镜观察各组药物作用24、48 h后细胞形态;CCK-8法测定各组的细胞增殖抑制率并计算半数抑制浓度;AnnexinⅤ-FITC/PI双染料流式细胞分析技术测定各组的细胞凋亡率;RT-PCR检测bcl-2、caspase-3及p21表达。结果淫羊藿素及阿霉素可明显抑制MG-63细胞增殖,随着淫羊藿素的浓度从10μmol/L逐渐增加至160μmol/L,细胞增殖抑制率也随之从(9.67±3.62)%逐渐增加至(89.18±9.66)%,二者的半数抑制浓度分别为:46.93μmol/L、3.87μg/m L;淫羊藿素与阿霉素可致细胞早期及晚期凋亡,使bcl-2基因表达下调、caspase-3、p21基因表达上调(P<0.05);以上改变联合组较淫羊藿素组及阿霉素组更明显(P<0.05)。结论淫羊藿素联合阿霉素对抑制人骨肉瘤MG-63细胞增殖具有协同或相加效应,其作用机制可能与调控bcl-2、caspase-3及p21基因有关。
Objective To investigate the inhibitory effect and its molecular mechanism of icaritin combined with doxorubicin on human osteosarcoma MG-63 cells in vitro. Methods The control group, the icariin group (10,20,40,80,160μmol / L), the doxorubicin group (1,2,4,8,16μg / m L), the combination group Icaritin + doxorubicin: 20μmol / L + 1μg / ml, 20μmol / L + 2μg / ml, 20μmol / L + 4μg / ml, 40mol / L + Osteosarcoma MG-63 cells were cultured in different concentrations (2μg / ml, 40μmol / L + 4μg / ml, 80μmol / L + 1μg / ml, 80μmol / L + 2μg / ml and 80μmol / The morphological changes of the cells were observed by inverted phase contrast microscope after 24 and 48 hours. The cell proliferation inhibition rates of each group were measured by CCK-8 method and the half inhibitory concentration was calculated. Flow cytometry analysis of AnnexinⅤ-FITC / PI double staining The apoptosis rates of bcl-2, caspase-3 and p21 were detected by RT-PCR. Results Icaritin and doxorubicin significantly inhibited the proliferation of MG-63 cells. As the concentration of icaritin increased gradually from 10μmol / L to 160μmol / L, the inhibitory rate of cell proliferation decreased from (9.67 ± 3.62) % Increased gradually to (89.18 ± 9.66)%, the half inhibitory concentrations were 46.93μmol / L and 3.87μg / ml, respectively; icariin and doxorubicin induced early and late cell apoptosis, -2 gene was down-regulated while caspase-3 and p21 were up-regulated (P <0.05). Compared with icaritin and adriamycin groups, the combined group was more obvious (P <0.05). Conclusion Icaritin combined with doxorubicin has a synergistic or additive effect on the proliferation of human osteosarcoma MG-63 cells. Its mechanism may be related to the regulation of bcl-2, caspase-3 and p21 genes.