地塞米松通过稳定podocin的表达和分布抑制嘌呤霉素对小鼠肾小球足细胞的损伤  

Dexamethasone inhibits puromycin aminonucleoside-induced injury of mouse glomerular podocytes via stabilizing the expression and distribution of podocin in vitro

作　　者：温跃强[1] 于力[1] 温捷[1] 郝志宏[1] 陈蓉燕[1] 王丽娜[1] 

机构地区：[1]广州医学院附属广州市第一人民医院儿科,510180

出　　处：《中华肾脏病杂志》2009年第7期509-513,共5页Chinese Journal of Nephrology

基　　金：广东省自然科学基金（5000424）;广东省科技计划项目（2005B36001019）;广州市科技攻关计划项目（200623-E0231）志谢 衷心感谢美国Mundel教授和北京大学第一医院儿科丁洁教授赠送足细胞;感谢丁洁教授在实验中给予的技术指导;感谢赵丹师姐在足细胞培养中给予的帮助

摘　　要：目的观察嘌呤霉素（PAN）和地塞米松（DEX）对足细胞分子podocin表达和分布的影响，探讨DEX改善蛋白尿的机制。方法体外培养小鼠足细胞（MPC5），分为对照组、PAN组和DEX组。对照组用含0．02％DMSO的RPMI-1640培养液培养；PAN组加入PAN；DEX组同时加入PAN和DEX。处理后8h、24h和48h，观察细胞形态并摄像，用图像处理软件分析各组细胞形态及胞体面积的差异。用RT—PCR、Western印迹和间接免疫荧光染色检测各时间点podocin mRNA和蛋白的表达及分布。结果正常足细胞呈星形，胞体大并有树样突起，细胞相互连接。PAN刺激8h，足细胞胞体面积缩小，为对照组的43％；24h为10％；48h为5．7％（P〈0．01）；部分足细胞足突及细胞连接消失。DEX组在8h、24h和48h，足细胞胞体面积显著大于PAN组，分别为对照组的43．9％、26．2％及29．6％（P〈0．05），足突形态正常。PAN组podoein mRNA表达量呈下降趋势，蛋白表达量显著降低（P〈0．01），分布异常。DEX组podocin mRNA和蛋白的表达量及分布与对照组相似，48h时mRNA和蛋白的表达量均显著高于PAN组（P〈0．05）。结论DEX直接作用于足细胞，稳定podocin mRNA和蛋白的表达量及分布，该作用可能与其能保护足细胞及改善蛋白尿有关。Objective To observe the effects of puromycin aminonucleoside （PAN） and dexamethasone （DEX） on the expression and distribution of podocin in vitro, and to explore the possible mechanism of DEX in improving proteinuria. Methods Mouse podoeyte cells （MPCs） in control group were cuhured with RPMI-1640 plus 0.02% DMSO, and were subjected to PAN treatment alone （PAN group） or PAN plus DEX （DEX group） for 8, 24,48 hours respectively. The podocyte morphology was observed by phase-contrast microscope, and was analyzed by Image J. The distribution, mRNA and protein expression of podocin were detected by indirect immunoeytofluorescence, semi-quantitative RT-PCR and Western blot, respectively. Results The well-developed arborization and interconnection of podocytes were found in control group. PAN treatment led to significant shrinkage of podoeytes with decreased distribution at 43% of control group at 8 h, 10% at 24 h and 5.7% at 48 h （P〈0.01）, respectively, together with podocyte foot process retraction as well as effacement and loss of cell contact. RT-PCR revealed podocin mRNA expression prone to decrease. Western blot showed podocin protein expression was significantly decreased and immunocytochemistry revealed podocin expression was disappeared in the cellular membrane after PAN treatment. DEX significantly prevented the shrinkage of podcytes, with decreased area at 43.9% of control at 8 h, 26.2% at 24 h and 29.6% at 48 h （P〈0.05）, respectively, and up-regulated the mRNA and protein expression of podocin at 48 h （P〈0.05）. The abnormal distribution of podocin was also alleviated by DEX. Conclusion DEX exerts a direct action on podocyte via stabilizing mRNA, protein expression and distribution of podocin, which may be associated with the improvement of proteinuria.

关 键 词：足细胞 地塞米松 嘌呤霉素 PODOCIN 

分 类 号：R692[医药卫生—泌尿科学] R277.52[医药卫生—外科学]