槐杞黄颗粒对阿霉素肾病大鼠肾组织nephrin和podocin表达的影响

Effects of Chinese herbal medicine Huaiqihuang Granule on nephrin and podocin expressions in renal tissues of rats with adriamycin-induced nephrosis

目的:通过观察槐杞黄颗粒对阿霉素肾病大鼠肾组织中nephrin与podocin基因及蛋白表达的影响,探讨槐杞黄颗粒治疗肾病综合征的作用机制。方法:将Sprague-Dawley大鼠20只随机分为5组(即正常对照组、模型组、激素组、槐杞黄组和槐杞黄+激素组),除正常组大鼠外,各组大鼠尾静脉一次性注射阿霉素造模。分别在建模后第7、14、21及28天收集24h尿液,检测大鼠24h尿中的蛋白含量;第29天处死所有大鼠,取肾脏组织,光学显微镜和电子显微镜下观察大鼠肾脏病理改变,实时荧光定量聚合酶链式反应法检测肾组织nephrin与podocin mRNA的表达,蛋白质印迹法检测肾组织nephrin与podocin蛋白的表达。结果:与正常对照组比较,从建模第14天开始模型组大鼠24h尿液中蛋白量增多(P=0.005),并随造模时间延长逐渐增加(P=0.015,P=0.012);从第14天开始,槐杞黄组与正常对照组相比,其尿蛋白减少不明显(P<0.05);第21天时,激素组与正常对照组相比,其尿蛋白明显增加(P=0.045)。光学显微镜下发现:模型组肾间质有灶性炎症细胞浸润,伴少量纤维组织增生;药物治疗组炎症细胞浸润较少。电子显微镜下发现:模型组肾小球足突大部分融合、消失,或出现部分微绒毛;槐杞黄组基底膜基本光滑均匀,足突融合明显减轻;激素组及槐杞黄+激素组足突清晰,甚至接近正常对照组。与正常对照组相比,模型组肾组织中nephrin、podocin mRNA及蛋白表达均明显减少(P<0.05);与模型组比较,药物治疗组nephrin、podocin mRNA及蛋白表达均明显增加(P<0.05)。24h尿蛋白量与nephrin mRNA以及nephrin和podocin蛋白的表达呈负相关。结论:槐杞黄颗粒可通过上调裂孔隔膜上的nephrin及podocin的表达维持足细胞裂孔隔膜的完整性,减轻肾小球滤过屏障的损伤,以此降低尿蛋白漏出。

OBJECTIVE：To investigate the effects of Huaiqihuang Granule,a compound Chinese herbal medicine,on expressions of nephrin and podocin of slit diaphragm of glomerular podocytes in rats with adriamycin-induced nephrosis and to explore the mechanism in reducing the proteinuria.Methods：Twenty SD rats were randomly divided into five groups：control group,model group,glucocorticoid group,Huaiqihuang Granule group and Huaiqihuang Granule plus glucocorticoid group. The 24-hour urine was collected 7,14,21 and 28 days after adriamycin injection respectively to measure 24-hour urinary protein,and all rats were sacrificed after 28-day treatment. Pathological changes in renal tissues were observed under a light microscope and an electron microscope. Expressions of nephrin and podocin mRNAs in renal cortex were determined by real-time polymerase chain reaction,and protein levels of nephrin and podocin were detected by Western blotting.Results：（1） In the model group and the treatment groups,the level of urinary protein increased significantly from the 14th day. （2） Under the light microscope,inflammatory cells and slight fibroplasia were found in renal interstitium of the model group,but there were less inflammatory cells in renal interstitium in the intervention groups than in the model group. Under the electron microscope,29 days after adriamycin injection,extensive fusion of foot processes was observed. （3） The expressions of nephrin and podocin were higher in treatment groups than in the model group. （4） Proteinuria level was negatively correlated with the expressions of nephrin mRNA and nephrin and podocin proteins.Conclusion：The above results indicate that Huaiqihuang Granule can maintain the integrity of the slid diaphragram in podocyte,alleviate the lesion of glomerular filtration membrane,and decrease the proteinuria by up-regulating the expressions of nephrin and podocin. Huaiqihuang Granule plus glucocorticoid maybe has better effects than glucocorticoid alone.