半枝莲提取物抑制人恶性胶质瘤U251细胞增殖及机制研究

Inhibitory effects and mechanism of scutellaria barbata extract on proliferation and telomerase activity of human malignant glioma U251 cells

目的 探讨半枝莲提取物（ESB）对人恶性胶质瘤U251细胞体外增殖、凋亡和端粒酶活性的影响. 方法 以50、25、12.5、6.25、3.125、1.5625 mg/mL ESB分别作用体外培养的人恶性胶质瘤U251细胞24、48、72 h,同时设空白对照组,应用MTT法检测ESB对U251细胞增殖的影响,AnnexinV/PI染色检测细胞凋亡率的变化,端粒重复序列扩增法（TRAP-PCR）-酶联免疫吸附试验（ELISA）检测U251细胞端粒酶的活性. 结果 MrT检测结果显示ESB浓度、作用时间因素的交互效应（F=59.908,P=0.000）,50 mg/mL ESB作用72 h时对U251细胞的抑制率最大;AnnexinV/PI染色检测显示浓度、时间因素的交互效应（F=6.548,P=0.000）,25 mg/mL ESB作用72 h时U251细胞的凋亡率最大;TRAP-RCR ELISA法检测显示ESB浓度、时间因素的交互效应（F=138.433,P=0.000）,50 mg/mL浓度ESB作用72 h时U251细胞端粒酶活性最小;细胞端粒酶活性与凋亡率之间呈负相关关系（r=-0.785,P=0.037）,与细胞抑制率之间也呈负相关关系（r=-0.278,P=0.042）. 结论 ESB可能通过下调端粒酶活性抑制U251细胞的增殖、诱导U251细胞凋亡.

Objective To observe the effects of scutellaria barbata extract （ESB） on proliferation, apoptosis and telomerase activity of human malignant glioma U251 cells in vitro.Methods Different concentrations of ESB （50, 25, 12.5, 6.25, 3.125 and 1.5625 mg/mL） were added into the medium cultured human malignant glioma U251 cells for 24, 48 and 72 h, respectively. And blank control group was also established. MTT assay was employed to detect the proliferation of U251cells. AnnexinV/PI staining and low cytometry （FCM） were used to detect the changes of apoptotic rate.And the telomerase activity of the cells was observed under the examination of telomeric repeat amplification protocol-PCR （TRAP-PCR）-ELISA.Results ESB inhibited the proliferation and induced the apoptosis of U251 cells. Interaction effect was found between the concentration of ESB and the treatment time of ESB by MTT assay （F=59.908, P=0.000）; 50 mg/mL ESB for 72 h could most significantly inhibit the proliferation of U251 cells. Interaction effect was found between the concentration of ESB and the treatment time of ESB by AnnexinV/PI staining （F=6.548, P=0.000）; 25mg/mL ESB for 72 h could most significantly induce the apoptosis of U251 cells. Interaction effect was also found between the concentration of ESB and the treatment time of ESB by TRAP-PCR-ELISA（F=138.433, P=0.000）; the telomerase activity of the cells was the lowest by treatment with 50 mg/mL ESB for 72 h; negative correlation was noted between the telomerase activity of the cells and the apoptosis rate （r=-0.785, P=0.037）; so as the telomerase activity of the cells and the inhibition effect （r=-0.278, P=0.042） Conclusion ESB may inhibit the proliferation and induces the apoptosis of U251 cells through down-regulating the telomerase activity.