关键词: 肝细胞性肝癌, miR?365, 自噬, 自噬相关基因3

Abstract:

Objective To investigate the effect of microRNA（mi）⁃365 on autophagy in hepatocellularcarcinoma（HCC）cells by regulating autophagy related gene 3（ATG3）. Methods Collected the cancerous tissuesand adjacent tissues of HCC patients，purchased human liver cancer SMMC⁃7721 cells，liver cancer HepG2 cells，human normal liver cells L02，transfected SMMC⁃7721 cells，and randomly divide them into control group and NCgroup（transfected with miR⁃365⁃NC）and overexpression group（transfected with miR⁃365 mimics）. RT⁃qPCRwas used to detect the expression levels of miR⁃365 and ATG3 in different tissues，different liver cancer cell linesand normal hepatocytes，MTT method was used to detect the proliferation of SMMC⁃7721 cells after transfection，flow cytometry was used to detect cell apoptosis，transmission electron microscopy and MDC staining was used toobserve cell autophagy，Western blot was used to observe the relative expression of ATG3，microtubule⁃associatedprotein1 light chain3⁃II（LC3⁃II），p62，Beclin1 protein in cells，dual luciferase experiment was used to detect thetargeting of miR⁃365 and ATG3. Results The expressions of miR⁃365 and ATG3 in cancer tissues of HCC patientswere lower than those in adjacent tissues（P < 0.05）. The expressions of miR⁃365 and ATG3 in liver cancer SMMC⁃7721 cells and HepG2 cells were lower than those of normal liver cells L02（P < 0.05）. After transfection of miR⁃365，compared with the control group and NC group，the expression of miR⁃365 and ATG3 in the overexpressiongroup increased，cell proliferation decreased，and the apoptosis rate increased（P < 0.05）. In the overexpressiongroup，autophagic vesicles were formed and the fluorescence intensity of MDC staining increased. Compared withthe control group and the NC group，the overexpression group ATG3，LC3⁃II，Beclin1 protein expression increased，and the p62 protein expression decreased（P < 0.05）. The dual luciferase experiment showed that ATG3 is thetarget gene of miR⁃365. Conclusions Overexpression of miR⁃365 can up⁃regulate the expression of ATG3，in⁃duce autophagy，inhibit the proliferation of SMMC⁃7721 cells，and promote cell apoptosis.

Key words: hepatocellular carcinoma, microRNA, autophagy, autophagy related gene 3