CAJ | 학술논문

目的观察醛固酮(ALD)及其受体拮抗剂螺内酯(SPI)对足细胞活性氧(ROS)产生及凋亡的影响,并探讨其可能机制.方法体外培养条件的永生化小鼠足细胞系,分为空白对照组、ALD组、SPI组、ALD+SPI组;用荧光分光光度计检测足细胞内ROS水平;间接免疫荧光检测nephrin表达;流式细胞仪检测足细胞凋亡率;RT-PCR、Western印迹法检测bax、bcl-2 mRNA及蛋白表达.同时观察抗氧化剂N-乙酰半胱氨酸(NAC)对上述效应的阻断作用.结果与对照组相比,ALD诱导足细胞ROS产生增多(P＜0.05),该作用可被SPI阻断(P＜0.05).ALD可诱导足细胞nephrin表达降低及足细胞凋亡(P＜0.05),同时伴有bax mRNA、蛋白表达升高及bcl-2 mRNA、蛋白表达降低(P＜0.05),SPI及NAC可阻断这一变化(P＜0.05).结论 ALD通过ROS途径作用于盐皮质激素受体上调促凋亡因子bax表达,下调抑凋亡因子bcl-2表达,进而诱导足细胞凋亡.
목적 관찰철고동(ALD)급기수체길항제라내지(SPI)대족세포활성양(ROS)산생급조망적영향,병탐토기가능궤제.방법 체외배양조건적영생화소서족세포계,분위공백대조조、ALD조、SPI조、ALD+SPI조;용형광분광광도계검측족세포내ROS수평;간접면역형광검측nephrin표체;류식세포의검측족세포조망솔;RT-PCR、Western인적법검측bax、bcl-2 mRNA급단백표체.동시관찰항양화제N-을선반광안산(NAC)대상술효응적조단작용.결과 여대조조상비,ALD유도족세포ROS산생증다(P＜0.05),해작용가피SPI조단(P＜0.05).ALD가유도족세포nephrin표체강저급족세포조망(P＜0.05),동시반유bax mRNA、단백표체승고급bcl-2 mRNA、단백표체강저(P＜0.05),SPI급NAC가조단저일변화(P＜0.05).결론 ALD통과ROS도경작용우염피질격소수체상조촉조망인자bax표체,하조억조망인자bcl-2표체,진이유도족세포조망.
Objective To access the effects of aldosterone (ALD) and its receptor antagonist- spironolactone (SPI) on the production of reactive oxygen species (ROS) and apoptosis in podocytes and to explore the possible mechanism involved. Methods Conditionally immortalized mouse podocytes were divided into control group, ALD group, SPI group and SPI +ALD group. The level of ROS production and the expression of nephrin protein were assayed by fluorescent spectrophotometry and indirect immunofluorescence technology. The apoptosis rate of podocytes was monitored by flow cytometry. The expression of bax and bcl-2 mRNA and protein was detected by RT-PCR and Western blotting methods. The anti-oxidant N-acetylcysteine (NAC)was applied to determine whether the effects of ALD on podocytes were mediated by ROS pathway.Results Compared with the control group, ALD significantly increased ROS production in podocytes (P＜0.05). SPI completely abolished the above-mentioned effect of ALD (P＜0.05). ALD induced the down-regulation of the expression of nephrin and the up-regulation of podocytes apoptosis (P＜0.05), which was accompanied with the elevated expression of bax mRNA and protein and the reduced expression of bcl-2 mRNA and protein (P＜0.05). SPI or NAC prevented the above-mentioned changes induced by ALD (P＜0.05). Conclusion ALD increases theexpression of pro-apoptotic factor (bax) but decreases the expression of anti-apoptotic factor (bcl-2)to induce podocytes apoptosis through the mineralocorticoid receptor (MR) possibly via the mechanisms involving ROS pathway.