转染Bcl-2基因的膀胱癌细胞多药耐药性研究

Study on multidrug resistance of bladder cancer cells transfected Bcl-2 gene

目的:探讨转染Bcl-2基因后膀胱癌细胞的多药耐药变化。方法:采用基因重组技术构建Bcl-2基因的真核表达载体pcD-NA3.1(+)/Bcl-2,经酶切和基因测序分析对构建结果进行鉴定;通过脂质体将含Bcl-2基因的表达载体转染膀胱癌细胞BIU-87,RT-PCR检测基因转录水平;应用四甲基偶氮唑蓝(MTT)比色法检测细胞在不同化疗药物作用下的细胞耐药性。结果:经酶切鉴定和基因测序分析证明真核表达载体pcDNA3·1(+)/Bcl-2构建成功。将该表达载体转染BIU-87细胞后,Bcl-2基因的表达水平与野生型BIU-87细胞和转染空载体的BIU-87/neo细胞相比显著提高(P<0·01)。与BIU-87细胞和BIU-87/neo细胞相比,BIU-87/Bcl-2细胞对MMC、PHA和DDP等化疗药物的耐药性也明显增强(P<0·01)。结论:Bcl-2基因高表达是膀胱癌多药耐药的原因之一。

OBJECTIVE： To explore the muhidrug resistance after the Bcl-2 gene was transfected into bladder cancer cells. METHODS： The technology of gene recombination was used to con structed the eucaryotic expression vector pcDNA3. 1 （＋）/Bcl-2. Inciding with enzymes and gene sequencing were used to verify the vectors. The human bladder cancer BIU-87 cells were transfected with pcDNA3, 1（＋）/Bcl-2 by liposome introduced transfection and genetic transcription level was evaluated by RT-PCR. The cell survival and drug resistance treated with different chemotherapeutics were detected by MTT method. RESULTS： The recombinant vector pcDNA3. 1 （＋）/Bcl 2 was constructed successfully confirmed by in ciding with enzymes and gene sequencing analysis. The expression level of Bcl-2 gene was higher in pcDNA3. 1 （＋）/Bcl-2 than that in BIU 87 and BIU-87/neo which was transfected with vector pcD- NA3.1（＋）. Compared with BIU 87 and BIU-87/neo, BIU 87/Bcl- 2 showed a higher survival rate and drug resistance. CONCLUSIONS： The high expression of Bcl-2 gene is one of the important reasons of multidrug resistance of bladder cancer.