乌骨藤粗提物对大鼠胶质瘤C6细胞的体外实验研究

Effect of Marsdenia tenacissima extract on proliferation of rat C6 glioma cells in vitro

目的观察乌骨藤粗提取物(MTE)不同组分对大鼠C6胶质瘤细胞增殖情况的影响,探讨其作用机制。方法采用新型四氮唑盐法(MTS)检测MTE的Fr1、Fr2、Fr3组分对C6细胞增殖的影响;采用流式细胞仪检测Fr2对C6细胞凋亡和周期的影响;采用荧光定量PCR(qPCR)技术检测不同浓度Fr2对C6细胞的Bax、Bcl-2表达的影响。结果 MTE的Fr1、Fr2、Fr3均能显著抑制C6细胞的增殖(P<0.05),与阳性对照组5-氟尿嘧啶(5-FU)相比抑制率也较高(P<0.05),其半数抑制浓度分别为:60.32、52.30、83.83μg/ml。经80、160μg/ml的Fr2处理24 h后,可有效促进C6细胞早期和晚期凋亡(P<0 05),其周期被阻滞在G_2期;Fr2相同方式处理后,C6细胞中Bax表达水平分别提高到(124.80±7.68)%和(380.30±6.91)%;Bcl-2表达水平分别降低到(84.20±3.10)%和(34.20±4.80)%(P<0.05)。结论 MTE能够显著抑制C6胶质瘤细胞的增殖,其中主要有效成分Fr2能够诱导C6细胞发生G_2期周期阻滞,并通过影响Bax和Bcl-2的表达,促进其凋亡。

Objective To investigate the effect of Marsdenia tenacissima extract （MTE） on proliferation of rat glioma C6 cells. Methods A new tetrazolium salt reduction method （MTS） was used to measure the effect of Fr1, Fr2, Fr3 component from MTE on C6 cell proliferation, flow cytometry was applied to measure cell apoptosis and cell cycle, real-time quantitative PCR （qPCR） was used to detect Bax and Bcl-2 gene expression in C6 ceils treated by Fr2 with different concentrations. Results Fr1, Fr2 and Fr3 of MTE significantly inhibited the proliferation of C6 cells （P〈0.05）, the inhibition rate was higher than that of 5-fluorouracil （5-FU）（P〈0.05）, and the 50% inhibitory concentrations were 60.32μ g/ml, 52.30 μ g/ml, and 83.83 μg/ml, respectively. After treated with 80 μ g/ml and 160 μg/ml of Fr2 for 24h, early and late apoptosis of C6 cells were both promoted （P〈0.05）, and the cell cycle was arrested at G2 phase; Bax gene expression was increased to （124.80 ±7.68）% and （380.30 ±6.91）%; Bcl-2 gene expression was reduced to （84.20 ±3.1）% and（34.20±4.80）%, respectively （P〈0.05）. Conclusion This study suggests that MTE can significantly inhibit the proliferation of C6 glioma cells, and the main active ingredient Fr2 can induce G2 phase cycle arrest and promote apoptosis, which were associated with changes of Bax and Bcl-2 gene expression.