Construction of recombinant adenovirus with survivin shRNA and its effect on lung cancer A549 cells.

Objective To construct the recombinant adenovirus vector expressing survivin shRNA,which mediates siRNA to inhibit the proliferation of human lung cancer A549 cells.Methods Cotransfection of pShuttle-survivin and adenovirus plasmid with LacZ tag into HEK-293 T cells was conducted to get recombinant adenovirus.Virus titer was determined after verification by PCR.Infection efficiency of the adenovirus was investigated by β-gal staining.MTT and FCM were employed to study its biological effects,RT-PCR was used to estimate the effects of Ad-survivin on reducing the expression of survivin in A549 cells.Hoechst staining was used to verify whether cell apoptosis could be induced or not.Results Ad-survivin was constructed successfully and its infection efficiency on A549 cells was evident.By means of Ad-survivin,the proliferation of A549 cells were suppressed and cells were blocked at the G2/M phase.RT-PCR results showed that the expression of survivin mRNA was repressed by Ad-survivin,and Hoechst staining indicated that knockdown of survivin expression by RNAi induced apoptosis in A549 cells.Conclusion Recombinant adenovirus with shRNA targeting survivin has been successfully constructed and its tumor suppression in A549 cells was detected,which may provide an effective way of gene therapy on human lung cancer.