Effect Of LncRNA HULC Regulating Autophagy On Invasion And Metastasis Of Hepatocellular Carcinoma

Objective: To investigate the effect of Lnc RNA HULC regulating auophagy on invasion and metastasis of hepatocellular carcinoma.Methods: The expression of HULC was down-regulated by HULC Smart Silencer in SMMC-7721 and Huh-6.GFP-RFP-LC3 double-labeled adenovirus were used to observe the changes of autophagy activity.Western blot was used to detect the expression of Atg7?p62?LC3?E-cadherin and ?-Catenin in hepatocellular carcinoma cells.After reduction of HULC expression,autophagy was induced by starvation.Western blot and cellular immunofluorescence were used to detect the expression of E-cadherin??-Catenin?Vimentin and MMP2 in hepatocellular carcinoma cells.Transwell detects its invasion and metastasis ability.Immunohistochemistry was used to detect the expression of Atg7?E-cadherin and N-cadherin in HULC(L)and HULC(H)groups.q RT-PCR was used to detect the expression and correlation of HULC ?mi R-137 and Atg7 m RNA in clinical hepatocellular carcinoma tissues;confirm that Lnc RNA HULC specifically binds mi R-137 by mi RNA sponge.q RT-PCR and Western blot were used to detect the effect of mi R-137 on Atg7,autophagic activity and epithelial mesenchymal transition.Nude mice sub-skin transplanted tumor model was used to observe the tumor growth.PCR was used to detect the expression of HULC and mi R-137 in subcutaneous xenografts.Western blot was used to detect the expression of Atg7 and LC3.Immunohistochemistry was used to detect the expression of Ki67,N-cadherin and Atg7 in subcutaneous xenografts.The orthotopic transplanted tumor model was used to observe the intrahepatic metastasis and the expression of Vimentin and Atg7 was detected by immunohistochemistry in the two groups.Results: After knocking down the expression of HULC in SMMC-7721 and Huh-6,Atg7 expression and autophagic activity decreased,while epithelial transformation ability decreased.After reduction of HULC expression,autophagy was induced by starvation.Autophagic activity,invasion and metastasis were significantly enhanced. In clinical specimens,HULC was positively correlated with Atg7 expression and negatively correlated with mi R-137 expression.Compared with HULC(L)group,the expression of Atg7 and N-cadherin increased and the expression of E-cadherin decreased in HULC(H)group.Knocking down of HULC expression increases the expression of mi R-137 in SMMC-7721 and Huh-6.Transfected with mi R-137 mimics,HULC expression decreased;and transfected with mi R-137 inhbitor,HULC expression increased.PCR and Western blot analysis showed that mi R-137 negatively regulates the expression of Atg7,inhibits autophagic activity and reduces the ability of hepatocarcinoma cells EMT.Nude mice subcutaneously transplanted tumor models showed that the reduction of HULC expression leaded to inhibit the of transplanted tumors growth,increase expression of mi R-137,decrease expression of Atg7 m RNA and autophagic activity.Immunohistochemistry showed that the expression of Ki67,Atg7 and N-cadherin decreased in the experimental group.Orthotopic transplanted tumor model results showed the reduction of HULC expression leaded to reduce intrahepatic metastasis.Immunohistochemical results showed that the expression of Atg7 and Vimentin decreased in the experimental group.Conclusion: Lnc RNA HULC specifically binds mi R-137 through mi RNA sponge,alleviates its inhibitory effect on Atg7,increases the autophagy activity of hepatocarcinoma cells,and promotes the invasion and metastasis of hepatocellular carcinoma.