论文部分内容阅读
目的探讨纤溶酶原激活物抑制剂(PAI)的发卡样小干扰RNA(siRNA)真核表达载体对人肝癌HpG2细胞组织型纤溶酶原激活物(tPA)和尿激酶PA(uPA)表达的影响。方法设计短发夹状PAI siRNA的寡核苷酸链,将其插入到真核细胞表达载体pSilencerTM 2.1-U6 neo载体中构建重组载体后,采用脂质体转染方法将构建的重组载体导入人肝癌细胞HpG2中,采用逆转录聚合酶链反应(RT-PCR)、Western印迹法分别检测转染细胞PAI mRNA和蛋白对tPA和uPA表达的影响。结果转染后细胞中PAI含量降低,tPA和uPA的表达减少(均P<0.01)。结论测序结果表明发卡样的PAI siRNA真核表达载体转染HpG2细胞后获得稳定表达,并可特异性封闭PAI的表达,PAI siRNA能有效阻断PAI的蛋白合成,同时抑制细胞中tPA和uPA的表达。
Objective To investigate the expression of tissue plasminogen activator (tPA) and urokinase PA (uPA) in human hepatocellular carcinoma HpG2 cells by plasmids inhibitor (PAI) Impact. Methods The oligonucleotide of short hairpin PAI siRNA was designed and inserted into the eukaryotic expression vector pSilencerTM 2.1-U6 neo vector to construct a recombinant vector. The constructed recombinant vector was introduced into human The effect of PAI mRNA and protein on the expression of tPA and uPA in transfected cells was detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting respectively in HpG2 cells. Results After transfection, the content of PAI in the cells decreased and the expression of tPA and uPA decreased (all P <0.01). Conclusion The sequencing results showed that the hairpin-like PAI siRNA eukaryotic expression vector transfected into HpG2 cells was stably expressed and specifically blocked the expression of PAI. PAI siRNA could effectively block the PAI protein synthesis and inhibit the expression of tPA and uPA expression.