活性氧和转化生长因子β1在醛固酮上调纤溶酶原激活物抑制剂1中的作用

目的 探讨活性氧(ROS)、转化生长因子β1 (TGF-β1)在醛固酮促进系膜细胞生成纤溶酶原激活物抑制剂1(PAI-1)途径中的作用。 方法 用醛固酮刺激大鼠肾脏系膜细胞。采用激光共聚焦检测系膜细胞内ROS水平。以貂肺上皮细胞(Mvllu)生长抑制MTT法检测上清中TGF-β1活性。然后用抗氧化剂N-乙酰半胱氨酸(NAC)与抗TGF-β1中和抗体阻断醛固酮刺激肾脏系膜细胞所产生ROS和TGF-β1。用RT-PCR观察系膜细胞PAI-1 mRNA的变化。用Western印迹法测定系膜细胞培养上清液中的PAI-1。 结果 醛固酮刺激大鼠肾脏系膜细胞24 h后，细胞内ROS水平上升到对照组的5倍(P < 0.01)；细胞上清中活性TGF-β1浓度明显升高(P < 0.01)；PAI-1 mRNA与蛋白表达分别上升到对照组的1.8倍(P < 0.01)和1.7倍(P < 0.01)。而采用NAC抑制ROS水平的升高、采用抗TGF-β1中和抗体阻断TGF-β1的作用后，PAI-1 mRNA表达分别下降了32%和30%(P < 0.01)；PAI-1蛋白表达分别下降了21%与11%(P < 0.05)，但仍未下降到正常水平。 结论 ROS、TGF-β1在醛固酮上调系膜细胞的PAI-1表达方面起了重要作用，但醛固酮并非完全依赖ROS、TGF-β1促使PAI-1的表达上调。

Roles of ROS and TGF-β1 in aldosterone-induced production of PAI-1

Objective To explore the roles of reactive oxygen species(ROS) and TGF-β1 in aldosterone-induced PAI-1 production. Methods Quiescent rat mesangial cells (MCs) were treated by aldosterone. The level of ROS in MCs induced by aldosterone was measured by confocal laser scanning microscopy and the TGF-β1 activity in the supernatant of culture was measured by mink lung epithelial cell (Mvllu) proliferation inhibition MTT assay. Then, before the addition of aldosterone, MCs were pretreated with NAC or TGF-β1 neutralizing antibody to decrease cellular ROS or inhibit activity of TGF-β1 induced by aldosterone respectively. PAI-1 mRNA was examined by semi-quantification RT-PCR and PAI-1 protein by Western blotting. Results The intracellular ROS induced by aldosterone increased by 5-fold compared to that of control group, and the activity of TGF-β1 stimulated by aldosterone increased markedly. TGF-β1 neutralizing antibody and NAC effectively decreased aldosterone-induced PAI-1 mRNA expression by 30% and 32%, and PAI-1 protein expression by 21% and 11%, respectively. However, neither TGF-β1 neutralizing antibody nor NAC alone could regulate aldosterone-induced PAI-1 mRNA and protein expression to normal level in 24 hours. Conclusions ROS and TGF-β1 play important roles in up-regulation of aldosterone-induced PAI-1 in MCs. ROS and TGF-β1 are not the exclusive pathway of PAI-1 expression induced by aldosterone in MCs.