目的探讨针对人端粒酶RNA(hTR)及其催化亚基(hTERT)的小干扰RNA(siRNA)对膀胱移行细胞癌细胞株BIU-87端粒酶活性及其增殖的影响。方法根据人TR mRNA和TERT mRNA的序列,分别设计合成3对不同的且能干扰TR和TERT基因表达的siRNA(TR-siRNA,TERT-siRNA),通过脂质体法将其转染膀胱移行细胞癌BIU-87细胞株,采用荧光定量PCR分析TR、TERT mRNA表达,TRAP-ELISA检测端粒酶活性,MTT法检测细胞增殖。结果3对TERT-siRNA和3对TR-siRNA中,pRNAT-TERT-Ⅲ和pRNAT-TR-Ⅲ可以特异性抑制BIU-87细胞株中TERT和TR表达(分别减少67%和41%,P〈0.05),且BIU-87细胞株的生长和端粒酶活性明显受到抑制。结论TR-siRNA和TERT-siRNA能特异且高效阻断各自基因表达,降低端粒酶活性,进而抑制BIU-87细胞的增殖。
Objective Telomerase activation is correlated to genesis, progression,invasion and metastasis of transitional cell carcinoma of bladder. This study was to evaluate the effects of small interfering RNA ( siRNA ) against either the template RNA (human telomerase RNA, hTR ) or its catalytic subunit (human telomerase reverse transcriptase, hTERT ) of telomerase gene on the telomerase activity and the growth inhibition of human transitional cell carcinoma of bladder cell line BIU - 87. Methods Three human TR - specific small interfering RNAs ( TR - siRNA ) and three human TERT - specific small interfering RNAs ( TERT - siRNA ) were designed based on TR and TERT mRNA sequences. TR - siRNA and TERT - siRNA were transfeeted into transitional cell carcinoma of bladder cell line BIU - 87 by using liposome transfection. The influence on the TR mRNA and TERT mRNA were analyzed by fluorescence quantitative PCR, while telomeric repeat amplification protocol(TRAP ) was applied to detect the telomerase activity, and the growth inhibition of BIU -87 ceils was detected by MTT assay. Results Among 3 pairs of TERT - siRNA and 3 pairs of TR - siRNA, pRNAT - TERT - Ⅲ and pRNAT - TR - Ⅲ could significantly reduce the expression of TERT and TR mRNA in BIU - 87 cells by 63% and 41% ( P 〈 0.05 ) , the growth of BIU - 87 cells were inhibited and telomerase activity were considerably decreased. Conclusions TR - siRNA and TERT - siRNA can specifically and effectively reduce both TR and TERT mRNA expression, inhibit the oroliferation by bloeking telomerase activity of BIU - 87 cells.