巴豆生物碱抑制卵巢癌细胞增殖和诱导其凋亡的实验研究

Effect of Croton Alkaloid on Cell Cycle,Apoptosis and PCNA Gene Expression in Human Ovarian Cell

目的了解巴豆生物碱诱导人卵巢癌细胞HO-8910细胞周期、凋亡效果,以及凋亡与增殖细胞核抗原(PCNA)表达之间的关系。方法将浓度分别为50、100和150μg/ml的巴豆生物碱作用于人卵巢癌细胞HO-8910细胞后,采用流式细胞术检测巴豆生物碱对人卵巢癌细胞HO-8910细胞的增殖抑制作用和诱导细胞凋亡;采用倒置显微镜及免疫组织化学法对凋亡调节基因PCNA表达进行检测。结果巴豆生物碱作用48h后,倒置显微镜下可见细胞皱缩、出芽及少量凋亡小体形成。应用不同剂量的巴豆生物碱(50～150μg/ml)分别作用于HO-8910细胞24、48、72h后,巴豆生物碱抑制了HO-8910细胞增殖,并呈现剂量、时间依赖性。随着巴豆生物碱的作用时间延长,G0/G1期细胞比例明显下降,而较多的细胞阻滞于G2/M期。巴豆生物碱的作用浓度从50μg/ml增至150μg/ml,则平均细胞凋亡率从2.13%上升为21.89%。结论巴豆生物碱可以通过时间依赖性和剂量依赖性方式,促使细胞G2/M期阻滞和抑制细胞有丝分裂,从而诱导人卵巢癌细胞HO-8910细胞凋亡。由于PCNA在G1～S期进程过度中起着重要作用,PCNA表达的下调也导致细胞周期进程被阻抑。巴豆生物碱作用24h,随浓度的升高,PCNA阳性细胞百分率逐渐降低。

Objective Croton alkaloids induce HO-8910 human ovarian cancer cell cycle,apoptosis effect and apoptosis and PCNA （proliferating cell nuclear antigen） expression relationship.Methods concentrations were 50,100 and 150 μg/ml of croton alkaloids act on the human ovarian cancer HO-8910 cells by flow cytometry （Annexing-V-FITC PI）detection of croton alkaloids on human ovarian cancer HO-8910 cell proliferation and induced apoptosis;using inverted microscopy and immunohistochemical expression of apoptosis regulating gene PCNA were detected.Results role of croton alkaloids 48 h,cell shrinkage was seen under inverted microscope,a small amount of sprouting and the formation of apoptotic bodies.Application of different doses of croton alkaloids （50 μg/ml to 150 μg/ml） were on the HO-8910 cells 24,48,72h after the croton alkaloids HO-8910 inhibited cell proliferation in a dose and time dependent.With the extended role of croton alkaloids,G0/G1 phase cells decreased,while more cells were arrested in G2/M phase.The role of the concentration of croton alkaloids from 50 μg/ml to 150 μg/ml,the mean apoptosis rate increased from 2.13% to 21.89%.Croton alkaloids can reduce the apoptosis-related genes PCNA expression.Conclusion Croton alkaloids can be time-dependent and dose-dependent manner to promote cell G2/M arrest and inhibition of cell mitosis,thereby inducing human ovarian cancer HO-8910 cells.Because of PCNA in the G1 ~ S plays an important role in the process over,so the reduction in the expression of PCNA also resulted in inhibition of cell cycle process is.The role of croton alkaloids 24h,with concentrations,PCNA-positive cell percentage decreased.