Anicequol抑制HepG2细胞自噬的抗肿瘤作用研究

目的 探讨从海洋真菌Penicillium dipodomyicola分离的anicequol抑制肝肿瘤细胞株HepG2增殖与自噬的关系。方法 HepG2分别用 0、2.5、5或10μmol·L?1 的anicequol处理,CCK-8法检测对HepG2的增殖作用,透射电镜观察对自噬囊泡的影响,Western blot检测对LC3II/I表达的影响,mRFP-GFP-LC3腺病毒转染观察对自噬流的影响，吖啶橙(AO)染色检测对溶酶体膜通透性的影响,。结果 Anicequol显著抑制HepG2细胞增殖,透射电镜观察到典型的自噬囊泡,mRFP-GFP-LC3腺病毒转染显示绿色荧光蛋白和红色荧光蛋白同步呈现荧光点状聚集,表明anicequol阻断HepG2的自噬流(P < 0.05)。AO染色表明anicequol引起HepG2溶酶体膜通透性增加,溶酶体膜的完整性被破坏(P < 0.05)。结论 Anicequol可通过调控肿瘤细胞自噬过程，阻断自噬流抑制HepG2细胞增殖。

Anicequol inhibits human hepatocellular cell line HepG2 growth through blocking autophagy

Objective To investigate the inhibition of proliferative effect of anicequol on human hepatocellular cell line HepG2 through blocking autophagy. Methods The inhibition of proliferation on HepG2 cells by anicequol was conducted by CCK8 assay. Typical autophagic vesicles were observed by transmission electron microscopy (TEM). Western blot was used to determine the production of autophagosomes. To evaluate autophagy flux, mRFP-GFP-LC3 adenovirus were transiently transfected to HepG2 cells. AO staining was carried out to investigate lysosomal membrane permeability (LMP). Results CCK-8 assay indicated that anicequol could significantly inhibit the proliferation of HepG2 in a concentration and time-dependent manner. Autophagosomes were mediated by the blockage of autophagic flux, which was bound up with impairing lysosomal function, including LMP as indicated by mRFP-GFP-LC3 adenovirus transfection, TEM and AO staining. Conclusion Anicequol exerted a significant HepG2 cell proliferation inhibition activity in a dose and time-dependent manner by blocking autophagy flux.