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Objective To investigate the inhibitory effect of inhibitor 4 of growth(ING4)delivered by adenovirus on human breast carcinoma cell MDA-MB-231.Methods MDA-MB-231 human breast carcinoma cells were irfected with Ad-ING4.The expression level of ING4 gene was detected by RT-PCR and Western blot;The growth inhibition,cell-cycle alteration,and apoptosis were detected by MTT,Flowcytometry and Hochest33258 staining.respectively.RT-PCR was used to detect the transcription of Bax,Bc1-2,Survivin genes;The expression level of Ang-1 gene was detected by ELISA;Ad-ING4 was given intratumorally in athymic nude mice beating MDA-MB.231 tumors.and then tumor growth was monitored;The expression of Bc1-2,Bax and Caspase-3 was analyzed by immumohistochemistry.Results ING4 was successfully transcribed and translated iU MDA-MB-231 cells:Ad-ING4 significantly inhibited the proliferation and induced G_2/M phase arrest to(24.86±1.24)% and cell apoptosis of MDAMB-231.Intratumoral injection of Ad-ING4 suppressed the tumor growth obviously with a inhibitory rate of 49%;Immumohistochemistry showed that the expression of Bax,Caspase-3 were up-regulated and the expression of Bc1-2.Survivin,CD34 were down-regulated by Ad-ING4.Conclusions Ad-ING4 can inhibit the growth of MDA-MB-231 cells and induce apoptosis in vitro and in vivo.
RESUMO
Objective:To establish the RPB5-mediating protein (RMP)-silenced stable cell lines and study the inhibitory effects of small interfering RNA (siRNA) targeting RMP gene on the proliferation and migration of human hepatoma SMMC-7721 cells. Methods:Three RMPi siRNAs were designed and synthesized in vitro and transfected into SMMC-7721 cells. The inhibitory effect of siRNA on RMP gene expression was measured by RT-PCR to select the best siRNA. The expression vector pGPU6-Neo-RMP-484 was transfected into SMMC-7721 cells by the lipofectamine and the cells stably expressing the siRNA were selected by G418. RT-PCR was used to detect the interference efficacy against RMP gene. Cell proliferation and adhesion were measured by MTT assay. Wound healing test was used to observe the migration ability of cells. Results:The SMMC-7721 cell lines with down-regulated RMP expression were established by using RNA interference technology. Compared with the negative control cells, expression of RMP mRNA was down-regulated by(83.67±2.56)% .The proliferation of stable-transfected cells was inhibited by(74.33±0.58)% . The adhesion capability of stable-transfected cells was enhanced but the migration capacity was decreased compared with the negative control cells. Conclusion:The pGPU6-Neo-RMP-484 cell lines with stable transfection of RMP siRNA recombinant vector are successfully screened,which can be used as a cellular model for studying the molecular mechanism of RMP. Down-regulation of RMP gene expression can effectively inhibit the proliferation, enhance the adhesion, and decrease the migration of SMMC-7721 cells.