基于Hedgehog信号通路的熊果酸诱导结直肠癌SW480细胞凋亡的机制研究

目的研究熊果酸通过经典Hedgehog信号通路影响人结直肠癌SW480细胞增殖和凋亡的作用及其机制。方法通过显微镜观察熊果酸对SW480细胞形态的影响;采用MTT比色法检测熊果酸对细胞活力的影响;细胞划痕实验检测熊果酸对细胞迁移的影响;Hoechest/PI染色法观察熊果酸对细胞凋亡的影响;荧光探针法检测熊果酸对细胞线粒体膜电位的影响;Caspase活性检测试剂盒检测熊果酸对细胞凋亡相关因子Caspase-3、Caspase-9的影响;蛋白免疫印迹法检测Hedgehog信号通路相关蛋白SHh、Gli1、Ptch1、Smo、c-Myc、Su Fu及细胞凋亡相关蛋白Bax、Bcl-2的表达水平。结果 SW480细胞经熊果酸给药后形态发生改变;细胞迁移能力显著下降;线粒体膜电位降低;细胞核出现致密浓染;Caspase-3、Caspase-9活性升高;SW480细胞发生凋亡;Hedgehog信号通路相关蛋白Su Fu的表达水平升高,SHh、Gli1、Ptch1、Smo、c-Myc的表达水平下降;细胞凋亡相关蛋白Bax的表达水平升高,Bcl-2的表达水平下降。结论熊果酸对SW480细胞的生长、增殖具有抑制作用,通过抑制Hedgehog信号通路的激活促进SW480细胞凋亡。

Mechanism study on ursolic acid induced apoptosis of colorectal cancer SW480 cells based on Hedgehog signaling pathway

Objective To investigate the effects and mechanisms of ursolic acid on proliferation and apoptosis of human colorectal cancer cell SW480 through canonical Hedgehog signaling pathway. Methods The effect of ursolic acid on cell morphology was observed by microscope. MTT colorimetric method, cell scratch experiment and Hoechest/PI staining were used to detect the effect of ursolic acid on cell viability, cell migration and cell apoptosis. Fluorescent probe method was used to detect the effect of ursolic acid on mitochondrial membrane potential. The effects of ursolic acid on Caspase-3 and Caspase-9 were detected by Caspase activity detection kit. The expressions of Hedgehog signaling pathway-related proteins SHh, Gli1, Ptch1, Smo, c-Myc, SuFu and apoptosis-related proteins Bax and Bcl-2 were detected by Western blotting. Results Ursolic acid significantly affected the morphology of human colorectal cancer cell SW480; The migration ability of SW480 cells was decreased markedly; Mitochondrial membrane potential was decreased; The nuclei were densely stained; The activity of Caspase-3 and Caspase-9 was increased; SW480 cell apoptosis was found; The expression levels of Hedgehog signaling pathway related protein SuFu and apoptosis-related protein Bax were increased; The expression levels of apoptosis-related protein Bcl-2 and Hedgehog signaling pathway-related proteins SHh, Gli1, Ptch1, Smo c-Myc were decreased. Conclusion Ursolic acid enable to inhibit the growth and proliferation and promote apoptosis of SW480 cells by inactivating Hedgehog signaling pathway.