醉茄素A对非小细胞肺癌A549细胞增殖、凋亡及PI3K／Akt信号通路的影响

目的 探讨醉茄素A（WFA） 对非小细胞肺癌（NSCLC）A549细胞增殖、凋亡及PI3K／Akt信号通路的影响。方法 采用0、2.5、5.0、10.0、20.0μmol／L WFA 处理A549细胞，采用四甲基偶氮唑盐（MTT）比色法检测上述浓度处理24、48、72和96h的细胞增殖抑制率，Hoechst染色和磷酯酰丝氨酸结合蛋白 异硫氢酸荧光素／碘化丙啶双染法（Annexin V-FITC／PI）检测各浓度组48h的细胞凋亡情况，流式细胞仪检测各浓度组48h的细胞周期分布情况，免疫印迹检测各浓度组48h凋亡相关基因（Bcl-2、Bax和Cleaved caspase-3）和PI3K／Akt信号通路重要蛋白Akt及其磷酸化形式p-Akt的蛋白水平。结果 WFA 可抑制细胞增殖，并呈剂量和时间依赖性（P＜0.05）；0、2.5、5.0、10.0、20.0μmol／L WFA作用48h后A549细胞的凋亡指数分别为2.75±0.64、4.61±1.36、9.75±2.78、12.92±3.42和18.68±4.31，组间差异有统计学意义（P＜0.05）。除2.5μmol／L外，其余浓度组的早、晚期凋亡率、凋亡促进基因（Bax和Cleaved caspase-3）水平及G0／G1期细胞比例均高于0μmol／L，凋亡抑制基因Bcl-2水平及S期和G2／M期细胞比例均低于0μmol／L（P＜0.05）； 2.5、5.0、10.0、20.0μmol／L的组间差异有统计学意义（P＜0.05）。随着浓度升高，p-Akt／Akt值呈降低趋势，差异有统计学意义（P＜0.05）。结论 WFA能够抑制A549细胞的增殖及凋亡，可能通过抑制PI3K／Akt通路激活实现。

Influences of withaferin A on proliferation,apoptosis and PI3K/Akt signaling pathway of non-small cell lung cancer A549 cell

Objective To explore the influences of withaferin A （WFA） on proliferation, apoptosis and PI3K/Akt signaling pathway of non-small cell lung cancer （NSCLC） A549 cell. Methods The A549 cells were treated with different concentrations of WFA （0, 2. 5, 5.0, 10. 0, 20. 0 p.mol/L）. The MTT assay was used to measure the proliferation inhibition rates at 24, 48, 72 and 96h treated with different concentrations of WFA. The Hoechst staining and Annexin V-FITC/PI double staining were employed to de- tect cell apoptosis after treatment with different concentrations of WFA. The cycle distribution at 48h after treatment with WFA was de- tected by flow cytometry. The Western blotting was used to measure the protein levels of apoptosis-related genes （Bcl-2, Bax and Cleaved caspase-3）, Akt and its phosphorylated form （P-Akt）. Results WFA could increase the proliferation inhibition rates in a dose- and time-dependent manner（P〈O. 05）. The apoptotic indices of A549 cells after treatment with different concentrations of WFA （0, 2.5, 5.0, 10.0, 20.0 p, mol/L） were 2.75-＋0.64, 4.6±1.36, 9.75±2.78, 12.92±3.42 and I8.68±4.31 with significant differences （P〈O. 05 ）. In addition to 2. 5μmol/L group, the early and late apoptosis rates, protein levels of apoptosis-promoting genes （Bax and Cleaved caspase-3） and the cell proportion in G0/Glphase of the remaining concentration groups were higher than those of 0μmo/L group（ P〈O. 05）.The p-Akt/Akt value decreased with increasing concentration and the differences were statistically significant among concentrations （P〈O. 05）. Conclusion WFA can inhibit the proliferation and apoptosis of A549 cell possibly by inhibiting the activation of PI3K/Akt pathway activation.