Abstract
The aim of the current study was to investigate the underlying mechanism of S-phase kinase associated protein 2 (Skp2) gene inhibition by lentivirus-mediated RNA interference (RNAi) on the cell cycle, apoptosis and proliferation of endometrial carcinoma HEC-1-A cells. A lentivirus shRNA vector targeting Skp2 was constructed and transfected into HEC-1-A cells. HEC-1-A cells transfected with a scramble sequence were used as negative controls. The mRNA and protein expression of Skp2, p27, cyclin D1 and caspase-3 were detected via reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. The effects of Skp2 inhibition on the cell cycle, apoptosis and proliferation of HEC-1-A cells were detected using flow cytometry and a cell counting kit-8. Skp2 co-expression data was analyzed using Oncomine and TCGA databases. The positive recombinant viral clones were identified via PCR and confirmed via sequencing. The mRNA and protein expression of Skp2 were significantly decreased in HEC-1-A cells transfected with the lentiviral vectors compared with the negative control. In addition, there were no significant changes in the mRNA expression of p27 and cyclin D1; however, the protein levels of p27 and cyclin D1 were upregulated and downregulated, respectively, in HEC-1-A cells transfected with lentiviral vectors compared with negative controls. RNAi-induced Skp2 inhibition exerted an anti-proliferative effect by inducing cell cycle arrest, however cell apoptosis was not significantly affected. In the TCGA database, Skp2 expression positively associated with IGF2R, IGF2BP3, IGFBP1 and CCNF, while Skp2 expression negatively associated with IGF2, IGFBP6, IGFBP7 and IGFBP3. RNAi-induced Skp2 inhibition upregulated the protein expression of p27 and downregulated the protein expression of cyclin D1. The expression of Skp2 in endometrial cancer may therefore be regulated by the insulin-like growth factor 1 receptor signaling pathway.