RNAi-Mediated Knockdown of Skp2 Inhibits Human Bladder Cancer Proliferation and Invasion in T24 Cells

Abstract

Background: Skp2 (S-phase kinase-associated protein 2 is overexpressed in many kinds of cancers, and is related to the occurrence and development of tumors. The molecular mechanism of Skp2 in the regulation of bladder cancer cell biological behavior after Skp2 expression knockdown, however, has remained unknown. Objectives: In our present studies (experimental cytobiological studies, we used an RNAi approach to knock down Skp2 expression, and studied its impact on cell proliferation and invasion of T24 cells. Materials and Methods: The expression of the Skp2 gene was knocked down by RNA interference (RNAi) in T24 cells. The transcription level of Skp2 was detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis. The expression of Skp2, caspase-3, caspase-8, caspase-9, and p27 (p27Kip1) were measured by western blot assay. Cell proliferation and apoptosis were detected by MTT and flow cytometry. Cell invasion analysis was performed by a matrigel transwell assay. We also detected the level of MMP2 (metalloproteinase-2) and MMP9 (metalloproteinase-9) in cell culture medium by ELISA. Results: The levels of Skp2 mRNA in the negative control group (0.911 ± 0.073) and the blank control group (0.940 ± 0.046) was significantly higher than Skp2 RNAi group (0.185 ± 0.033) (P < 0.001). The levels of Skp2 protein in the negative control group (0.907 ± 0.049) and the blank control group (0.925 ± 0.042) was significantly higher than Skp2 RNAi group (0.220 ± 0.047) (P < 0.001). The proliferation and invasion of T24 cells were significantly inhibited in vitro upon Skp2 RNAi treatment. Conclusions: The proliferation and invasion of human bladder cancer cells can be inhibited by RNAi-targeting Skp2. As a result, Skp2 may be a potential target for gene therapy in cases of human bladder cancer.