芹菜素诱导胃癌SGC-7901细胞自噬及其对凋亡的影响

doi:10.3969/j.issn.1004-616x.2017.04.009

摘要/Abstract

摘要：

目的：探讨芹菜素（API）是否诱导胃癌SGC-7901细胞发生自噬及其对细胞凋亡的影响。方法：采用不同浓度（0、20、40、60、80 μmol/L）的API处理胃癌SGC-7901细胞24 h，MTT法检测API对细胞生长的抑制情况。分别采用不同浓度（0、20、40、60、80 μmol/L）API处理细胞24 h，及60 μmol/L的API处理不同时间（0、12、24、36、48 h），Western blot检测自噬标志蛋白微管相关蛋白l轻链3（LC3）、Beclin-1和磷脂酰肌醇-3-激酶（PI3K）/Akt/mTOR信号通路中关键分子p-Akt、p-mTOR的表达；电子显微镜下观察细胞自噬情况。细胞经自噬抑制剂氯喹（CQ）或3-甲基腺苷（3-MA）预处理后再加入API，Western blot检测聚二磷酸腺苷-核糖体聚合酶（PARP）的表达；流式细胞术流检测细胞凋亡率。结果：API对胃癌SGC-7901细胞生长具有明显抑制作用并呈剂量-效应关系。随着API浓度和作用时间的增加，SGC-7901细胞LC3Ⅱ与Beclin-1蛋白表达均增加，各剂量组与对照组相比较差异均具有统计学意义（P < 0.05）；API处理后的细胞逐渐出现自噬囊泡和自噬溶酶体；p-Akt、p-mTOR蛋白表达降低。采用自噬抑制剂抑制细胞自噬后，除API组LC3Ⅱ与Beclin-1蛋白表达明显高于对照组外（P < 0.05），各组蛋白表达无显著差异。API+CQ组和API+3-MA组的PARP蛋白剪切片段表达及凋亡率均高于对照组和API组（P < 0.05）。结论：API可以诱导胃癌SGC-7901细胞发生自噬，其机制可能与抑制PI3K/Akt/mTOR信号通路关键分子p-Akt、p-mTOR活性有关；抑制API诱导的自噬能够增强API的凋亡诱导效应，从而增强对胃癌SGC-7901细胞的杀伤作用。

关键词: 芹菜素, 胃癌, 自噬, 细胞凋亡

Abstract:

OBJECTIVE: To study whether apigenin could induce autophagy in human gastric cancer SGC-7901 cells and whether the induced autophagy would affect apoptosis in these cells. METHODS: SGC-7901 cells were treated with different doses (0,20,40,60,80 μmol/L) of apigenin (API) for 24 h. Cell survival rates were measured using the MTT method. In addition,cells were treated with 60 μmol/L API for different hours(0,12,24,36,48 h). Western blot was used to evaluate the autophagy marker protein microtubule-associated protein 1 light chain 3 (LC3),Beclin-1 and the activities of p-Akt and p-mTOR,the critical targets of Akt/mTOR signaling pathway. Electron microscopy was used to observe autophagy. Furthermore, cells were treated with autophagy inhibitor,chloroquine(CQ) and 3-mehtyladenine (3-MA) and 60 μmol/L API. Western blot analyses were performed to detect changes in LC3,Beclin-1 and PARP. Annexin V-FITC/PI was used to detect the apoptotic rate. RESULTS: SGC-7901 cell viability was inhibited under API treatment in a does-response manner. With increasing doses of API,autophagic vesicles and autophagosome began to appear in electron microscopy. Western blot analysis showed that LC3-Ⅱ and Beclin-1 protein levels increased in cells treated with API (P < 0.05). The phosphorylation levels of Akt and mTOR were reduced. After inhibiting autophagy,the expression of LC3 Ⅱ and Beclin-1 proteins in the API group was significantly higher than that in the control group(P < 0.05). There was no significant difference in the other groups. Cleaved PARP and apoptotic rate increased significantly in API+CQ and API+3-MA then control and API. CONCLUSION: API can induce autophagy in human gastric cancer cells SGC-7901 via the Akt/mTOR signaling pathway. The API-induced autophagy could significantly increase API-induced apoptosis in these cells. Furthermore,API-induced autophagy can protect survival of gastric cancer cells.

Key words: apigenin, gastric cancer, autophagy, apoptosis

中图分类号:

R730.2

孙悦, 余洋, 吴坤. 芹菜素诱导胃癌SGC-7901细胞自噬及其对凋亡的影响[J]. 癌变·畸变·突变, 2017, 29(4): 289-294,299.

SUN Yue, YU Yang, WU Kun. Effect of apigenin-induced autophagy on apoptosis in human gastric cancer cells SGC-7901[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2017, 29(4): 289-294,299.

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