The inhibition of proliferation of hepatocellular carcinoma Hep3B cells induced by RNA interference targeting eEF1A1

Objective: To investigate the effects of RNA interference targeting eukaryotic translation elongation factor 1A1 (eEF1A1) expression on the proliferation of hepatocellular carcinoma Hep3B cells, and to explore its possible mechanism. Methods: A recombinant vector pGPU6/GFP/Neo-eEF1A1-shRNA containing short hairpin RNA (shRNA) targeting eEF1A1 was constructed, and then it was transfected into the Hep3B cells. After tansfection with pGPU6/GFP/Neo-eEF1A1-shRNA, the expressions of eEF1A1 mRNA and protein were examined by real time fluorescence quantitative PCR and Western blotting, respectively, the cellular growth ability was examined by cell counting kit 8 (CCK-8) assay, the colony formation ability was detected by colony formation assay, and the cell cycle distribution of Hep3B cells was detected by flow cytometry (FCM). The expression levels of cyclin D1 and cyclin-dependent kinase 4 (CDK4) proteins in Hep3B cells were examined by Western blotting. Results: The recombinant vector pGPU6/GFP/Neo-eEF1A1-shRNA was successfully constructed, and the Hep3B cells with stable expession of eEF1A1 were established. The expression levels of eEF1A1 mRNA and protein in Hep3B cells after transfection with pGPU6/GFP/Neo-eEF1A1-shRNA were lower than those in the negative control cells (Hep3B cells transfected with a negative control vector pGPU6/GFP/Neo-NC) and the blank control cells (Hep3B cells without any transfection) (P < 0.01, P < 0.05). As compared with the negative control, the cellular growth ability and the colony formation ability of Hep3B cells after tranfection with pGPU6/GFP/Neo-eEF1A1-shRNA were obviously decreased (P < 0.05, P < 0.01); the percentage of the cells in G1 phase was increased (P < 0.01), and which in S-phase was decreased (P < 0.05); the expression levels of cyclin D1 and CDK4 proteins were down-regulated (both P < 0.05). Conclusion: Down-regulation of eEF1A1 expression can inhibit the proliferation of Hep3B cells. DOI:10.3781/j.issn.1000-7431.2014.11.471