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目的观察TIEG基因沉默对AGEs介导肾小管上皮细胞PAI-1表达的影响。方法以pshRNA-copGFP-lentivector作为载体,构建含有TIEG基因的慢病毒载体psiRNA-TIEG,转染至正常大鼠近端肾小管上皮细胞(NRK52E),然后给予AGEs刺激24 h和48 h,采用Western blot方法测定PAI-1蛋白的表达水平。结果 AGEs以时间依赖方式上调NRK52E细胞TIEG mRNA和PAI-1蛋白表达。TGF-β1中和抗体可有效降低AGEs刺激NRK52E细胞48 h后PAI-1的表达水平(1.150±0.089vs0.707±0.015,P<0.05)。TIEGsiRNA转染后可显著下调AGEs刺激NRK52E细胞48 h后TIEG mRNA表达水平(0.852±0.019vs0.448±0.028,P<0.01),及PAI-1蛋白表达水平(0.396±0.042vs0.245±0.063,P<0.05)。结论 TIEG基因沉默能有效抑制AGEs诱导NRK52E细胞PAI-1的表达。
Objective To observe the effect of TIEG gene silencing on AGEs-mediated PAI-1 expression in renal tubular epithelial cells. METHODS: The lentiviral vector psiRNA-TIEG containing TIEG gene was constructed and transfected into proximal tubular epithelial cells (NRK52E) of normal rats with pshRNA-copGFP-lentivector, then stimulated with AGEs for 24 h and 48 h, blot method to determine PAI-1 protein expression level. Results AGEs up-regulated the expression of TIEG mRNA and PAI-1 protein in NRK52E cells in a time-dependent manner. The neutralizing antibody of TGF-β1 could effectively reduce the expression of PAI-1 in AGKs stimulated NRK52E cells 48 h (1.150 ± 0.089 vs0.707 ± 0.015, P <0.05). After transfected with TIEG siRNA, the expression of TIEG mRNA in NRK52E cells induced by AGEs for 48 h was significantly lower (0.852 ± 0.019 vs. 0.448 ± 0.028, P <0.01) and PAI-1 protein expression (0.396 ± 0.042 vs 0.245 ± 0.063, P <0.05). Conclusion TIEG gene silencing can effectively inhibit AGEs-induced PAI-1 expression in NRK52E cells.