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Effects of CircRNA-ITCH on proliferation and apoptosis of hepatocellular carcinoma cells through inhibiting Wnt/β-catenin signaling pathway.

PURPOSE
To explore the effects of circular ribonucleic acid (circRNA)-E3 ubiquitin protein ligase (ITCH) on the proliferation and apoptosis of hepatocellular carcinoma (HCC) cells and its possible molecular mechanism.


METHODS
To study the role of circRNA-ITCH in the occurrence and development of HCC. The relative expression level of circRNA-ITCH was detected via quantitative polymerase chain reaction (qPCR) in four kinds of HCC Huh-7, U251, HB611 and SMMC-7721 cell lines, and human normal liver L-02 cell line. Moreover, the effect of overexpression of circRNA-ITCH on the TCF luciferase activity was detected using β-catenin/TCF-responsive luciferase reporter assay. Finally, the influences of overexpression of circRNA-ITCH on the protein levels of β-catenin and Wnt3α and the mRNA levels of c-myc and cyclinD1 were determined using Western blotting and qPCR, respectively.


RESULTS
Compared with that in human normal liver L-02 cell line, the expression of circRNA-ITCH was significantly down-regulated in HCC Huh-7, U251, HB611 and SMMC-7721 cell lines (p<0.05). According to the results of CCK-8 assay, colony formation assay and flow cytometry, the overexpression of circRNA-ITCH could obviously inhibit cell proliferation, suppress colony formation ability and induce apoptosis (p<0.05). The results of dual-luciferase reporter assay revealed that there was a significant interaction between circRNA-ITCH and miR-7 or miR-214 (p<0.05). CircRNA-ITCH was involved in the regulating of Wnt/β-catenin signal transduction pathway and inhibited the expressions of c-myc and cyclinD1.


CONCLUSIONS
CircRNA-ITCH affects the proliferation and apoptosis of HCC cells through inhibiting the Wnt/β-catenin signal transduction pathway, thereby exerting a carcinogenic effect in the occurrence and development of HCC. The research results provide a new therapeutic target for HCC.