Effects of adenoviral-mediated siRNA targeting PNUTS on proliferation and invasion of laryngeal squamous cell carcinomas Hep-2 cells / 上海交通大学学报（医学版）

ABSTRACT

Objective·To explore the effects of adenovirus vector-mediated small interfering RNA (siRNA) targeting phosphatase nuclear targeting subunit (PNUTS) on proliferation,invasion,migration and epithelial-mesenchymal transition (EMT) of laryngeal squamous cell carcinomas Hep-2 cells and its mechanism.Methods· Recombinant adenovirus vector expressing PNUTS siRNA was infected into laryngeal squamous cell carcinomas Hep-2 cells and the experiment was designed into PBS group,Ad-GFP group and Ad-siPNUTS group.Levels ofPNUTS mRNA and protein were detected by real-time PCR and Western blotting respectively.MTT assay was used to detect proliferation abilities of Hep-2 cells.Transwell assays were used to detect invasion and migration abilities of Hep-2 cells.The expression levels of total Rb,phosphorylated Rb (p-Rb),PI3K,phosphorylated AKT (p-AKT),E2FI,E-cadherin,N-cadherin and ZEB1 protein were detected by Western blotting.Results· Compared with Ad-GFP group,in Ad-siPNUTS group,the PNUTS mRNA and protein (both P=0.000) levels were dramatically decreased.The proliferation of Ad-siPNUTS infected Hep-2 cells were inhibited on the second day (P=0.004),the third day (P=0.001) and the fourth day (P=0.000).Meanwhile,the invasion and migration abilities of Ad-siPNUTS infected Hep-2 cells were decreased (both P=0.000).The expression levels of total Rb (P=0.000),p-Rb (P=0.000),PI3K (P=0.023),p-AKT (P=0.000),E2F 1 (P=0.000),N-cadherin (P=0.005) and ZEB 1 (P=0.000) were decreased while the E-cadherin (P=0.003) was increased.Conclusion· Ad-siPNUTS could inhibit the proliferation,invasion and migration abilities of Hep-2 cells and reverse the development of EMT,which may be related to PI3K/AKT signaling pathway and Rb signaling pathway.