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[目的]观察穿心莲内酯(Andrographolide,Andro)对不同分化结肠癌细胞克隆形成、迁移及凋亡的影响,检测凋亡蛋白Bax和自噬蛋白LC3-Ⅱ表达改变,探讨其分子机制。[方法]不同浓度Andro分别作用人结肠癌细胞株Caco-2(高分化)和Lovo(低分化)24h后,MTT法检测药物细胞毒性;划痕愈合、transwell观察细胞迁移能力;检测细胞克隆形成能力;流式细胞术检测细胞凋亡;Western blot检测Bax、LC3-Ⅱ及cleaved-caspase-3的蛋白表达水平。[结果](1)Andro以时间和浓度依赖的方式抑制结肠癌细胞生长,Lovo细胞对Andro的敏感性高于Caco-2细胞。(2)Andro明显抑制Caco-2和Lovo细胞的迁移及克隆形成能力,Lovo细胞的效应更为显著。(3)Andro增加Bax和LC3-Ⅱ表达,且在Lovo细胞中表现更明显。(4)凋亡酶caspase-3的激活参与了Andro诱导Caco-2和Lovo细胞的凋亡。[结论]低分化结肠癌细胞Lovo对Andro的敏感性明显高于高分化Caco-2细胞,Andro通过增加Bax的表达,激活caspase-3介导凋亡信号通路,同时通过增强LC3-Ⅱ表达,促进自噬过程,最终抑制结肠癌细胞生长。
[Objective] To observe the effect of Andrographolide (Andro) on colony formation, migration and apoptosis in different differentiated colon cancer cells, and to detect the change of apoptosis protein Bax and autophagy protein LC3-Ⅱ, and to explore its molecular mechanism. [Method] Andro treated with different concentrations of human colon cancer cell lines Caco-2 (well-differentiated) and Lovo (low differentiation) for 24 h respectively. MTT assay was used to determine cytotoxicity. Scratches were healed and transwell was used to observe cell migration. The apoptosis of cells was detected by flow cytometry. The protein expression of Bax, LC3-Ⅱ and cleaved-caspase-3 were detected by Western blot. [Results] (1) Andro inhibited the growth of colon cancer cells in a time and concentration-dependent manner. The sensitivity of Lovo cells to Andro was higher than that of Caco-2 cells. (2) Andro significantly inhibited the migration and colony forming ability of Caco-2 and Lovo cells, and the effect of Lovo cells was more significant. (3) Andro increased the expression of Bax and LC3-Ⅱ, and showed more obvious in Lovo cells. (4) Activation of apoptosis enzyme caspase-3 is involved in the apoptosis of Caco-2 and Lovo cells induced by Andro. [Conclusion] The sensitivity of Lovo to poorly differentiated colon cancer cells is much higher than that of well-differentiated Caco-2 cells. Androxin activates the caspase-3-mediated apoptosis signal pathway by increasing the expression of Bax. At the same time, by increasing the expression of LC3- Promote autophagy and finally inhibit the growth of colon cancer cells.