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目的:研究新型棉酚衍生物ApoG2在体外应用对鼻咽癌CNE-2细胞凋亡与自噬的影响。方法:CCK-8检测增殖抑制率;Hoechst 33258染色、吖啶橙(AO)染色、透射电镜观察细胞凋亡与自噬形态学变化;FCM检测凋亡率与自噬荧光强度;蛋白质印迹法检测Bcl-2和Beclin1蛋白表达情况。结果:CCK-8法结果显示,不同浓度的ApoG2作用于CNE-2细胞24、48和72h后,均可见对细胞具有生长抑制作用,且呈剂量-时间依赖性。72h的IC50值为23.61μmol/L。Hoechst33258染色显示,ApoG2作用后可观察到更多的核固缩和碎裂等凋亡现象;AO染色显示,ApoG2作用后可观察到更多的亮红色酸性自噬泡。透射电镜可观察到细胞体内大空泡及膜性双层结构增多。FCM检测显示,对照组和ApoG2组凋亡率分别(3.83±0.23)%和(20.06±2.10)%,差异有统计学意义,F=176.823,P=0.000。FCM检测显示,对照组和ApoG2组自噬荧光强度分别为(1.05±0.54)%和(26.91±7.65)%,差异有统计学意义,F=34.019,P=0.004。蛋白质印迹法显示,Bcl-2蛋白表达下调差异有统计学意义,F=68.909,P=0.001;Beclin1蛋白表达上调差异亦有统计学意义,F=497.906,P=0.000。结论:ApoG2在体外可以抑制鼻咽癌细胞CNE-2增殖并诱导细胞凋亡与自噬。
AIM: To study the effect of ApoG2, a novel gossypol derivative, on the apoptosis and autophagy of CNE-2 cells in vitro. Methods: The proliferation inhibition rate of CCK-8 was detected by Hoechst 33258 staining, AO staining and transmission electron microscopy. The morphological changes of apoptosis and autophagy were observed by FCM. The apoptosis rate and autophagic fluorescence intensity were detected by FCM. Bcl-2 and Beclin1 protein expression. Results: The results of CCK-8 assay showed that ApoG2 could inhibit the proliferation of CNE-2 cells in a dose-and time-dependent manner at 24, 48 and 72 hours. 72h IC50 value of 23.61μmol / L. Hoechst33258 staining showed more apoptotic phenomena such as nuclear condensation and fragmentation after ApoG2 treatment; AO staining showed more bright red acidic autophagic vacuoles after ApoG2 treatment. Transmission electron microscopy can be observed in the cell vacuoles and membranous double membrane structure increased. The FCM results showed that the apoptotic rates in control group and ApoG2 group were (3.83 ± 0.23)% and (20.06 ± 2.10)%, respectively, the difference was statistically significant, F = 176.823, P = 0.000. FCM showed that the autophagic fluorescence intensity of control group and ApoG2 group were (1.05 ± 0.54)% and (26.91 ± 7.65)%, respectively, the difference was statistically significant, F = 34.019, P = 0.004. Western blotting showed that the down-regulation of Bcl-2 protein was statistically significant (F = 68.909, P = 0.001). The up-regulation of Beclin1 protein was also significantly different (F = 497.906, P = 0.000). CONCLUSION: ApoG2 can inhibit CNE-2 proliferation and induce apoptosis and autophagy in nasopharyngeal carcinoma cells in vitro.