常春藤皂苷元影响内质网相关途径阻滞骨肉瘤细胞周期的实验研究

目的:研究常春藤皂苷元对骨肉瘤细胞周期的影响及机制。方法:用0、5、10、20、40 μg/ml的常春藤皂苷元处理骨肉瘤细胞，CCK-8法检测细胞增殖情况，细胞克隆实验测定细胞克隆形成能力，计算其半数抑制浓度。用半数抑制浓度的常春藤皂苷元处理骨肉瘤细胞，流式细胞术检测细胞周期和细胞凋亡，荧光探针法测定细胞内Ca2+浓度。用Western blot法检测细胞中结合免疫球蛋白（BIP）、内质网氧化还原酶1-Lα（Erol-Lα）、蛋白质二硫键异构酶（PDI）、剪切型Caspase-12（Cleaved Caspase-12）、细胞周期蛋白D1（Cyclin D1）蛋白水平。结果:5、10、20、40 μg/ml的常春藤皂苷元均可以抑制骨肉瘤细胞的增殖和克隆形成能力，与0 μg/ml常春藤皂苷元作用组比较，差异有统计学意义（P＜0.05）。半数抑制浓度的常春藤皂苷元处理后的骨肉瘤细胞G1期比例升高，细胞凋亡率也升高，细胞内Ca2+浓度升高，细胞中BIP、Erol-Lα、PDI、Cleaved Caspase-12蛋白水平升高，Cyclin D1蛋白水平降低，与0 μg/ml常春藤皂苷元作用组比较，差异有统计学意义（P＜0.05）。结论:常春藤皂苷元通过影响内质网相关途径将骨肉瘤细胞周期阻滞在G1期，诱导细胞凋亡发生。

Effect of hederagenin on the cell cycle of osteosarcoma blocking the endoplasmic reticulum related pathway

Objective:To study the effect and its mechanism of hederagenin on the cell cycle of osteosarcoma.Methods:The osteosarcoma cells were treated with hederagenin of 0,5,10,20 and 40 μg/ml.CCK-8 assay was used to detect cell proliferation.Cell cloning assay was used to determine the ability of cell clone formation.The half inhibitory concentration was calculated.The osteosarcoma cells were treated with a half inhibitory concentration of hederagenin.Cell cycle and apoptosis were detected by flow cytometry,The intracellular Ca2+concentration was measured by fluorescence probe.The levels of BIP,Erol-L alpha,PDI,Cleaved Caspase-12 and protein in the cells were detected by Western blot.Results:The hederagenin of 5,10,20,40 μg/ml could inhibit the proliferation and cloning of osteosarcoma cells.Compared with 0 μg/ml hederagenin,the difference was statistically significant（P＜0.05).The proportion of G1 phase in the osteosarcoma cells treated by a half inhibitory concentration of hederagenin was increased.The rate of apoptosis was also increased,the concentration of Ca2+ in cells increased.The levels of BIP,Erol-Lα,PDI and Cleaved Caspase-12 were increased in the cells,the level of Cyclin D1 protein was reduced.Compared with 0 μg/ml hederagenin,the difference was statistically significant（P＜0.05).Conclusion:Hederagenin blocks cell cycle in G1 phase,and induces apoptosis by the endoplasmic reticulum related pathway.