足细胞特异性因子nephrin及肿瘤蛋白-1在肾小球硬化过程中表达变化

Expression of nephrin and Wilms’ tumor-1 in process of glomerular sclerosis and its significance

【摘要】目的 应用嘌罗霉素氨基核苷（PAN）模型，通过观察临床、病理指标及足细胞特异性因子nephrin和肿瘤蛋白-1（WT-1）在足细胞的表达，了解足细胞损害在肾小球硬化过程中的重要作用。方法 尾静脉注射PAN制作大鼠肾病模型，测定不同时间点的生化指标，处死大鼠，取其肾脏，石蜡包埋切片，PAS及免疫组化染色，测定肾小球硬化指数（GSI），肾小球面积（GA），nephrin及WT-1的表达，进行统计学分析。结果 PAN组在早期（第8天）表现为典型肾病综合征，后期（20周）表现为慢性肾炎。形态学方面，PAN组的GSI在4，14，20周均明显高于对照组（P＜0.05）。PAN组GA在14周时大于对照组（P＜0.01），但在20周时小于对照组（P＜0.001），并且20周时GSI与GA之间呈强负相关关系。PAN组nephrin表达在4，14，20周时与对照组比较均有所减少（P＜0.05），与此同时，WT-1的表达在14和20周时较对照组明显减少（P＜0.001）。结论 （1）在肾炎发展中，随着时间延长，GSI增高，GA缩小，并且GSI和GA之间呈负相关。（2）在肾小球硬化发展中，nephrin和WT-1的表达进行性减少，说明有明显足细胞数量的减少。因此，足细胞的损伤与肾小球硬化密切相关。

基金项目:国家自然科学基金（81160093）; 宁夏国际合作支撑项目（2011ZYH169）

【Abstract】Objective To investigate the role of podocytes damage in the process of glomerular sclerosis by observing the clinical and pathological indices and the expression of nephrin and Wilms’ tumor-1 (WT-1) in the podocytes of puromycin aminonucleoside (PAN)-induced nephropathy model. Methods A single intravenous injection of PAN via tail vein of rats was carried out to establish nephropathy model, and those with positive urine?protein were identified as successful establishment. Biochemical and pathological parameters were analyzed in the model rats and normal controls at 4 and 8 d, and at 4, 14 and 20 weeks after the model establishment for the chronic renal injuries evaluation. After the rats were sacrificed at 4, 14 and 20 weeks, their kidneys were taken out, embedded in paraffin, and stained with periodic acid-Schiff (PAS) for determination of glomerulosclerosis index (GSI) and glomerular area (GA). Immunohistochemistry assay was performed to detect the expression of nephrin and WT-1. Results The PAN models were successfully established, showing typical nephritic syndrome in the early stage (day 8), and chronic glomerular nephritis in the later stage (week 20). The GSI was significantly higher in PAN group than in the normal controls at 4, 14 and 20 weeks after establishment (P＜0.05). The mean size of GA was significantly larger in PAN rats than in the normal controls at 14 weeks (P＜0.01), but was obviously smaller at 20 weeks (P＜0.01). Significantly positive correlation was found between GSI and GA at 20 weeks in the model group. The expression of nephrin was obviously decreased in PAN group at 4, 14 and 20 weeks (P＜0.05), so was that of WT-1 at 14 and 20 weeks (P＜0.001). Conclusion (1) GSI is increased, while GA is decreased along with the progression of nephritis, and there is a negative correlation between GSI and GA. (2) The expression of nephrin and WT-1 is reduced along with the progression of glomerular sclerosis, indicating the decrease of podocytes. Therefore, the podocyte damage is closely associated with glomerular sclerosis.