Abstract: Anti-cancer Effect of Improved TAT-VP3 Gene on Human Bladder Cancer in VitroChunhuiWANG1, JiansongWANG1, Hui ZHAN1, Hongjun LI2, Mingxia DING1, Ruping YAN1, Changxing KE1, Hongyi XU1Correspondence to: JiansongWANG, E-mail: jiansongwang@yahoo.com1Department of Urology, the Second Affiliated Hospital of Kunming Medical College, Kunming 650101, China2Institute of Medical Biology, Chinese Academy of Medical Science, Peking Union Medical School, Kunming 650118, ChinaThis work was supported by National Natural Science Foundation of China (No. 30760251)Abstract Objective: To explore the expression and apoptosis-inducing effects of improved TAT-VP3 fusion gene on differenthuman bladder cancer cell lines. Methods: The recombinant plasmid PcDNA3-TAT-VP3 was constructed using the prokaryotic expres-sion vector pGEX-6p-1-TAT-VP3. After double enzymatic digestion with BamHⅠand XhoⅠ and verification by sequencing, lipo-some-induced gene transfection was used to transfect BIU-87 and EJ human bladder carcinoma cells. RT-PCR was used to detect theexpression of improved TAT-VP3 in the cells. Transmission electron microscopy was used to observe changes in the ultrastructure ofthe tumor cells at different periods of time after the transfection. MTT assay was conducted to detect the effect of the recombinant plas-mid on proliferative activity of the bladder carcinoma cells, and flow cytometry TUNAL was conducted to detect the apoptotic rate at24, 48 and 72 hours after transfection. Statistical analysis of the experimental results was conducted. Results: The eukaryotic expres-sion vector PcDNA3-TAT-VP3 was successfully constructed. The results of RT-PCR after the transfection confirmed the expression ofimproved TAT-VP3 mRNA in bladder cancer cells at 24 hours after transfection. The typical morphological changes in early apoptosissuch as cell shrinkage, chromatin condensation, nuclear fragmentation and apoptotic bodies were observed using transmission electronmicroscopy. MTT assay showed that the proliferative activity of bladder cancer cells distinctively decreased ( P < 0.01 ) after transfec-tion. The flow cytometry TUNAL detection showed that early apoptosis occurred in BIU-87 and EJ cells with a gradual increase in theapoptotic rate over time. Seventy two hours after the transfection, the apoptotic rate was ( 30.44±1.47 ) % in BIU-87 cells and ( 42.27 ±2.66 ) % in EJ cells. There was statistical significance between the experimental group and the control group ( P < 0.01 ). Conclusion:Expression of improved TAT-VP3 fusion gene can effectively induce apoptosis in 2 different human bladder cancer cell lines. Futurework should include the purification of improved TAT-VP3 fusion protein and the testing of this particular fusion protein as a treatmentfor bladder neoplasms via intravesicular administration.Keywords Apoptosis; VP3 protein; Bladder neoplasm