摘要
To modify the splicing pattern of Bcl-x and compare the effect of this approach with that of the antisense gene therapy in BIU-87 cell line of bladder cancer, by using 5′-Bcl-x AS to target downstream alternative 5′-Bcl-x splice site to shift splicing from Bcl-xL to Bcl-xS and 3′-Bcl-x AS antisense to the 3′-splice site of exon Ⅲ in Bcl-x pre-mRNA to down regulation of Bcl-xL expression, the inhibitory effects on cancer cells by modification of alternative splicing and antisense gene therapy were observed and compared by microscopy, MTT Assay, RT-PCR, FACS, Westhern bloting and clone formation. The growth of cells BIU-87 was inhibited in a dose- and time-dependent man- ner. Its inhibitory effect began 12 h after the exposure, reaching a maximum value after 72h. The number of cells decreased in S phase and the number increased in G1 phase. The ability to form loci was reduced and the antisense gene therapy was approximately half as efficient as modification of alternative splicing in inducing apoptosis. It is concluded that modification of splicing pattern of Bcl-x pre-mRNA in bladder cancer cell BIU-87 is better than antisense gene therapy in terms of tumor inhibition.
To modify the splicing pattern of Bcl-x and compare the effect of this approach with that of the antisense gene therapy in BIU-87 cell line of bladder cancer, by using 5′-Bcl-x AS to target downstream alternative 5′-Bcl-x splice site to shift splicing from Bcl-xL to Bcl-xS and 3′-Bcl-x AS antisense to the 3′-splice site of exon Ⅲ in Bcl-x pre-mRNA to down regulation of Bcl-xL expression, the inhibitory effects on cancer cells by modification of alternative splicing and antisense gene therapy were observed and compared by microscopy, MTT Assay, RT-PCR, FACS, Westhern bloting and clone formation. The growth of cells BIU-87 was inhibited in a dose- and time-dependent man- ner. Its inhibitory effect began 12 h after the exposure, reaching a maximum value after 72h. The number of cells decreased in S phase and the number increased in G1 phase. The ability to form loci was reduced and the antisense gene therapy was approximately half as efficient as modification of alternative splicing in inducing apoptosis. It is concluded that modification of splicing pattern of Bcl-x pre-mRNA in bladder cancer cell BIU-87 is better than antisense gene therapy in terms of tumor inhibition.
基金
This project was supported by a grant the Natural SciencesFoundation of Hubei Province (No .2000J048)
