LncRNA HULC shRNA disinhibits miR-377-5p to suppress the growth and invasion of hepatocellular carcinoma in vitro and hepatocarcinogenesis in vivo

Background: Aberrant expression of up-regulated long non-coding RNA [LncRNA highly upregulated in liver cancer (HULC)] has been observed to play an important regulatory role in the development of multiple human diseases. However, the molecular mechanism underlying the role of HULC and miR-377-5p in HCC needs to be urgently explored.

Methods: The mRNA and protein expression levels of HULC were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot in hepatocellular carcinoma (HCC) cell line HB611, HepG2 and H22, respectively. HULC-shRNA was transfected into HepG-2 cells, which were randomly divided into the control, shRNA-NC, and sh-HULC groups. The correlation between HULC and miR-377-5p was analyzed by performing a luciferase reporter assay. The targeting relationship between miR-377-5p and hypoxia-inhibitory factor-1α (HIF-1α) was also investigated using a luciferase reporter assay. Sh-HULC and miR-377-5p inhibitors were transfected either alone or together into HepG2 cells, and which were divided into the control group, the sh-HULC group, the miR-377-5p inhibitor, and the sh-HULC + inhibitor group for subsequent experiments. HepG2 cell proliferation and invasion were measured by 5-Ethynyl-2-Deoxyuridine (EdU) staining and Transwell invasion assay, respectively. Western plot was carried out to detect the protein expression levels of Ki67, PCNA, E-cadherin, and N-cadherin. Tumor xenograft mouse models were established to confirm the effect of HULC down-regulation on the development of HCC in vivo.

Results: The mRNA and protein expression levels of HULC were markedly increased, whereas the mRNA expression levels of miR-377-5p were decreased in HCC cell lines. HepG2 cell proliferation and invasion were suppressed in the Sh-HULC group, while miR-377-5p showed the opposite. Further experiments exhibited that miR-377-5p was targeted by HULC, and an negative correlation between HULC and miR-377-5p was observed. Importantly, the in vivo experiments indicated that down-regulation of HULC could inhibit tumor growth. Taken together, our research demonstrated that down-regulation of HULC plays an anti-cancer role through restrainingHepG2 cell proliferation and invasion.

Conclusions: In summary, our in vitro and in vivo findings confirmed HULC to play a role in the progression of HCC, with the underlying mechanism possibly involving the miR-377-5p/HIF-1α pathway.