Mechanistic Exploration Of Insensitivity Of Lung Cancer Cells To Vesicular Stomatitis Virus(VSV) Treatment

Objective: Vesicular stomatitis virus (VSV) is a promising biological therapy,which inducing apoptosis specifically in tumor cells. A major of previousstudies was focusing on the IFN system、 P53、 BCL-1. Livin-beta is aendogenous anti-apoptotic molecule, but the mechanism underling its ability ofmodulating the pro-apoptotic effect of VSV is still unknown. The purpose ofour study is to investigate modulation of the pro-apoptotic effect of VSV inA549lung cancer cells by Livin-beta and its mechanism.Methods: The pro-apoptotic effect of VSV on A549cells was verified by flowcytometry and TUNEL. A549cells treated with VSV was verified by flowcytometry and the DEVDaseactivity were detected as an indicator of apoptosisrate or caspase-3activity. The expression of Livin were analyzed bywestern-blot. Making sure that the Knock-down or overexpression ofLivin-beta was successfully transmitted into the A549cells,we used real-timePCR, RT-PCR and western-blot. The apoptosis rate were observed throughcytometry. Caspase-3activity in these cells was detested. Moreover, weverified the effects of Smac using the same method as Livin. The bindingconsequence betwen Livin and Smac was verified using immunoprecipitation.Results: VSV improves the apoptosis of A549, and the effects istime-dependent. the apoptosis rate was detected by flow cytometry. There was asignificant difference in apoptosis rates between VSV group and UT group (P＜0.001). When treated with zVAD-fmk, the apoptosis rate and caspase-3activity were much lower than those of the control. The expression of Livin inA549cells obviously decreased when treated with0.1PFU/ml VSV, and thenincreased gradually in a dose-dependent manner. Apoptosis rate and caspase-3activity increased when the cells was treated with Livin-siRNA. And the Apoptosis rate、 caspase-3activity decreased when the cells were treated withSmac-siRNA. The binding between Livin and Smac was much stronger inLivin overexpression cell line.Conclusion: Livin-beta regulates the caspase activity in a direct or indirectmanner, leading to the modulation of the pro-apoptotic effect of VSV in A549lung cancer cells. it may contribute to the resistance to VSV in A549cells.