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目的:研究重楼总皂苷(Paridis Rhizoma total saponins,PRTS)诱导人胃癌MKN-45细胞凋亡的机制。方法:在前期研究基础上,将PRTS设定3个不同质量浓度(5,10,20 mg·L~(-1))处理人胃癌细胞12 h,采用荧光染色法观察MKN-45细胞凋亡形态学变化;流式细胞术(Annexin V/PI双染色法)测定细胞凋亡率;ELISA测定MKN-45细胞caspase-3,caspase-8酶活性;Western blotting法检测Fas,FasL蛋白表达水平。结果:在荧光显微镜下,经PRTS处理的MKN-45细胞,可见典型的凋亡形态学特征;PRTS可明显增加细胞凋亡率,且PRTS 20 mg·L~(-1)与对照组相比凋亡率有显著差异(P<0.01);PRTS作用于MKN-45细胞12 h,与对照组比较,其caspase-3,caspase-8活性明显增高(P<0.01),Fas,FasL蛋白表达明显增高(P<0.01)。结论:PRTS能显著诱导人胃癌细胞MKN-45凋亡,其机制可能与增加caspase-3,caspase-8活性,上调Fas和FasL的表达有关。
Objective: To investigate the mechanism of Paritis Rhizoma total saponins (PRTS) inducing apoptosis in human gastric cancer MKN-45 cells. Methods: Based on the previous studies, human gastric cancer cells were treated with PRTS at concentrations of 5, 10 and 20 mg · L -1 for 12 h, and the apoptosis of MKN-45 cells was observed by fluorescence staining Morphological changes were observed. The apoptotic rate was determined by flow cytometry (Annexin V / PI double staining). The activity of caspase-3 and caspase-8 in MKN-45 cells was detected by ELISA. The expressions of Fas and FasL protein were detected by Western blotting. Results: Under the fluorescence microscope, PRTS treated MKN-45 cells showed typical apoptotic morphological features. PRTS increased the apoptotic rate significantly, and PRTS 20 mg · L -1 compared with the control group (P <0.01). Compared with the control group, the activity of caspase-3 and caspase-8 of PRTS group was significantly increased (P <0.01), while the expression of Fas and FasL protein was significantly increased Increased (P <0.01). Conclusion: PRTS can significantly induce the apoptosis of human gastric cancer cell line MKN-45. The mechanism may be related to increasing the activity of caspase-3 and caspase-8 and up-regulating the expression of Fas and FasL.