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目的探讨蛋白激酶C-α(PKCα)对人膀胱癌T24细胞多药耐药的调节作用。方法将载有PKCα基因的表达质粒GFP-PKCα导入人膀胱癌细胞系T24。应用噻唑蓝(MTT)比色法,检测T24/PKCα及亲本细胞对阿霉素的敏感性,以及在激活(10nmol/LPMA2h)和抑制(10nmol/LCalphostinC2h)PKCα的情况下,T24/PKCα细胞对阿霉素敏感性的变化。同时用逆转录-聚合酶链反应(RT-PCR)检测多药耐药基因-1(MDR-1)、多药耐药相关蛋白(MRP)和肺耐药相关蛋白(LRP)基因的表达。结果转染了PKCα基因后,T24细胞对阿霉素的耐药性增强(耐药指数7.52,P<0.05)。PKCα的激活显著提高了T24细胞的耐药性(耐药指数153.78,P<0.01),而抑制PKCα活性则使耐药指数降至1.20(P>0.05)。T24/PKCα的MDR-1表达水平是亲本细胞的2.28倍(P<0.01)。激活PKCα可使MDR-1基因表达水平较亲本细胞升高12.80倍(P<0.01),抑制PKCα则MDR-1表达下降(P>0.05),而MRP和LRP基因表达水平并无明显变化(P>0.05)。结论PKCα可能通过上调MDR-1表达,在人膀胱癌的化疗耐药中起到了重要的作用。
Objective To investigate the regulatory effect of protein kinase C-α (PKCα) on multidrug resistance in human bladder cancer T24 cells. Methods The expression plasmid GFP-PKCα carrying PKCα gene was introduced into human bladder cancer cell line T24. The sensitivity of T24 / PKCαand its parental cells to doxorubicin was detected by MTT assay, and T24 / PKCα cells were detected in the presence of 10nmol / LPMA2h and 10nmol / LCalphostinC2h PKCα Changes in doxorubicin sensitivity. At the same time, the expression of MDR-1, MRP and LRP genes were detected by reverse transcription-polymerase chain reaction (RT-PCR). Results After transfection of PKCα gene, the resistance of T24 cells to doxorubicin was enhanced (resistance index 7.52, P <0.05). Activation of PKCα significantly increased the drug resistance of T24 cells (resistance index 153.78, P <0.01), while inhibition of PKCα activity decreased the resistance index to 1.20 (P> 0.05). The MDR-1 expression level of T24 / PKCα was 2.28 times that of the parental cells (P <0.01). Activation of PKCα could up-regulated the expression of MDR-1 by 12.80-fold (P <0.01) and suppressed the expression of MDR-1 by PKCα (P> 0.05), while the expression of MRP and LRP did not change significantly > 0.05). Conclusion PKCα may play an important role in chemoresistance of human bladder cancer by up-regulating the expression of MDR-1.