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目的:研究禾肾丸对阿霉素肾病大鼠蛋白尿及肾组织足细胞裂孔膜蛋白nephrin表达的影响,为其治疗肾病蛋白尿提供实验依据。方法:SPF级雄性Wistar大鼠随机分为模型组,禾肾丸组(1.8 g·kg~(-1)),贝那普利组(1 mg·kg~(-1))及空白组。采用一次性尾静脉注射阿霉素6 mg·kg~(-1)复制阿霉素肾病大鼠模型。造模成功后,禾肾丸组及贝那普利组分别给予相应药液,模型组及空白组给予等体积蒸馏水,1次/d,连续28 d。给药结束后,收集24 h尿液,检测尿蛋白,腹主动脉采血,检测血中白蛋白(albumin,Alb),血肌酐(serum creatine,SCr),血尿素氮(blood urea nitrogen,BUN)。苏木素-伊红(HE)检查肾脏组织形态变化,透射电镜观察肾小球足细胞足突及基底膜改变,免疫组化及蛋白免疫印迹(Western blot)法检测肾组织nephrin表达。结果:经禾肾丸治疗后,24 h蛋白尿较模型组显著下降(P<0.01),血清SCr,BUN显著降低(P<0.01),Alb显著升高(P<0.01)。组织病理学检查显示,禾肾丸组肾小球上皮细胞肿胀明显减轻,部分肾小球体积略增大,系膜细胞及系膜基质仅见轻微增生,管腔内未见小管有蛋白管型。超微结构研究显示,禾肾丸组肾小球基底膜光滑均匀,略见增厚,足突清晰完整、偶见融合。免疫组化结果显示,禾肾丸组nephrin蛋白阳性面积率显著高于模型组(P<0.01)。Western blot结果显示,nephrin蛋白表达量较模型组显著升高(P<0.01)。结论:禾肾丸可能通过上调nephrin表达水平,修复受损足细胞而减少蛋白尿、保护肾脏。
Objective: To study the effect of HeShen pill on proteinuria and nephrin expression in renal tissue of adriamycin-induced nephropathy rats, and to provide an experimental basis for its treatment of nephropathy proteinuria. Methods: SPF male Wistar rats were randomly divided into model group, HeShen pill group (1.8 g · kg -1), benazepril group (1 mg · kg -1) and blank group. One-time tail vein injection of doxorubicin 6 mg · kg -1 replicate adriamycin nephropathy rat model. After the model was established successfully, the rats in Henshan Pill group and benazepril group were given corresponding liquid respectively, and the model group and the blank group were given equal volume of distilled water once a day for 28 days. Urine was collected 24 h after the administration, urinary protein and abdominal aorta were collected for detection of albumin (ALB), serum creatinine (SCr), blood urea nitrogen (BUN) . The morphology of renal tissue was examined by hematoxylin-eosin (HE) staining. The changes of foot process and basement membrane of glomerular podocyte were observed by transmission electron microscope. The nephrin expression in renal tissue was detected by immunohistochemistry and Western blot. Results: Compared with the model group, proteinuria of 24 h was significantly decreased (P <0.01), serum SCr, BUN and Alb significantly increased (P <0.01). Histopathological examination showed that the swelling of glomerular epithelial cells in He-Shen-Pill group was significantly reduced, the volume of some glomerulus was slightly increased, only slight hyperplasia was seen in mesangial cells and mesangial matrix, and no tubular was found in the lumen. Ultrastructural studies showed that the glomerular basement membrane in Wo sheng pill group was smooth and even, slightly thickening, foot process clear and complete, and occasionally fusion. The results of immunohistochemistry showed that the positive area ratio of nephrin protein in HeShen pill group was significantly higher than that in model group (P <0.01). Western blot results showed that nephrin protein expression was significantly higher than the model group (P <0.01). Conclusion: Wo Shewan Pills may reduce proteinuria and protect kidneys by up-regulating nephrin expression, repairing damaged podocytes.