| ObjectiveIn this experimental research,we investigated the effects of Yiban Decoction of the urine on hematuria,proteinuria,and renal pathology of IgA nephropathy rats by using the animal experiment to establish IgA nephropathy rat model before and after the intervention of Chinese medicine intervention group,and to explore the effect of Yiban Decoction on Wnt signaling in renal tissue ofIgA nephropathy rats.MethodsAfter one week of adaptive feeding,56 SPF wistar male rats were randomly selected,10 in normal group and 46 in the model group.Using Bovine serum albumin(BSA)+ carbon tetrachloride(CCL4)+lipopolysaccharide(LPS)to replicate the rat model of IgA nephropathy,which lasted 12 weeks,with BSA 100 mg/L,400 mg/kg on alternate days of irrigation,0.3 ml castor oil and 0.1 ml CCl4 subcutaneous injection once per week.In the 6th week tail intravenous LPS(with sodium chloride solution prepared into 0.25 g/L concentration)0.2ml.The rats in the normal group were given the same amount of distilled water and the same amount of saline were injected subcutaneously and tail vein,and the frequency was the same as that of the modelgroup.The 43 rats were randomly divided into model group(n=11),Shenyan Kangfu Tablet group(n=10),Yiban Decoction group(n=11)and Erban Decoction group(n=11).The 13 th week began with Chinese herbal medicine intervention treatment,Shenyan Kangfu Tablet group with its suspension,Yiban Decoction group with Yiban Decoction granules decoction,Erban Decoction group with Erban Decoction granules decoction,once a day for 8 weeks,the rats in the model group and normal group were treated with equal volume of distilled water.During the treatment of drug,in the model group two rats died,each from the Yiban Decoction group and the Erban Decoction group,and 50 rats died from the final group: group A(normal group,n=10),group B(model group,n=10),group C(Shenyan Kangfu Tablet group,n=10),group D(Yiban Decoction group,n=10),group E(Erban Decoction group,n=10).The urine erythrocyte count,24h-Upr,TPCR,Alb and Hb were examined and tested before and after treatment in each group of rats.After blood sampling,the left renal tissues were used for electron microscope,light microscope slice,and the right kidney tissues were used for cryopreservation,the expression of Nephrin,Wnt4,Wnt5 a and ?-catenin mRNA in different groups of rats were detected by RT-PCR method and immunohistochemical assay was used to detect the expression of Nephrin,Wnt4,Wnt5 a and ?-catenin proteins.Results1 The results of the general observation: the rats in the Shenyan Kangfu Tablet group,Yiban Decoction group and Erban Decoction group were better than the model group in the feeding drinking water and the activity degree.2 Electron microscopy: at the 8th weekend of the traditional Chinese medicine intervention treatment,the structure of the glomerularmesangial area in the normal group was clearly visible,the mesangial distribution was homogeneous,and there was no significant deposition of electron dense matter in the mesangial area.The model group showed significant proliferation of mesangial matrix,accompanied by cloud-like dendritic electron deposition,podocyte foot segmental fusion.Shenyan Kangfu Tablet group can be seen in the mesangial area of low-density electronic compact deposition,foot structure is more clear.Yiban Decoction group can be seen a small amount of electronic compact deposition,foot fusion Obviously.The structure of the glomerular mesangial area in the Erban Decoction group can be seen a small amount of electron dense material deposition,no foot full of diffuse fusion.3 Light mirror:HE staining showed that,there was no obvious abnormality in the morphology of glomeruli in the normal group at the8 th weekend of the traditional Chinese medicine intervention treatment,and the renal tubulointerstitium was clearly visible.The degree of renal pathological changes in the model group was different,mainly manifested as glomerular swelling,volume increased,glomerular cells increased,mesangial cell hyperplasia mostly mild to moderate,only a few can be seen severe mesangial cell proliferation.The lesion in the Shenyan Kangfu Tablet group,Yiban Decoction group and Erban Decoction group has Improved than the model group.Compared with Erban Decoction group,Yiban Decoction group saw no significant glomerular renal tubular abnormalities.4 In the 12 th weeks of modeling,the number of urine red blood cells and the level of urine protein were significantly elevated in the adult mice(P<0.05).5 The results of the erythrocyte count,24h-Upr,TPCR,Alb and Hbin each group:at the 8th weekend of the treatment,compared with the model group,the erythrocyte count,24h-Upr and TPCR decreased significantly in the Shenyan Kangfu Tablet group,Yiban Decoction group and Erban Decoction group(P<0.05),the levels of Hb increased significantly in the Shenyan Kangfu Tablet group,Yiban Decoction group and Erban Decoction group(P < 0.05),the levels of Alb did not increased significantly(P > 0.05).Compared with the Shenyan Kangfu Tablet group,24h-Upr and TPCR decreased significantly in the Yiban Decoction group(P < 0.05),the erythrocyte count decreased and the levels of Alb and Hb increased in the Yiban Decoction group,but the difference was not significant(P > 0.05),the difference that the erythrocyte count,24h-Upr and TPCR decreased and the levels of Alb and Hb increased was not significant(P > 0.05).Compared with the Erban Decoction group,the erythrocyte count,24h-Upr and TPCR decreased,the levels of Alb and Hb increased,but neither of the difference was significant(P>0.05).6 Results of Nephrin,Wnt4,Wnt5 a and ?-catenin mRNA expression in each group of rats:compared with the normal group,the relative expression of Nephrin mRNA significantly decreased(P<0.05)in the model group at the 8th weekend of the traditional Chinese medicine intervention treatment,and the relative expression of Wnt4,Wnt5 a and ?-catenin mRNA was significantly up-regulated in the model group(P < 0.05).Compared with the model group,the relative expression of Nephrin mRNA in the Shenyan Kangfu Tablet group,the Yiban Decoction group and the Erban Decoction group was significantly up-regulated(P<0.05),and the relative expression of Wnt4,Wnt5 a and?-catenin mRNA in the Shenyan Kangfu Tablet group,the Yiban Decoction group and the Erban Decoction group significantly decreased(P < 0.05).Compared with the Shenyan Kangfu Tablet group,the increase of Nephrin mRNA relative expression in the Yiban Decoction group was significant(P<0.05),the increase of Nephrin mRNA relative expression in the Erban Decoction group was not(P > 0.05),and the relative expression of Wnt4,Wnt5 a and ?-catenin mRNA was not significantly down-regulated in the Yiban Decoction group and Erban Decoction group(P>0.05).Compared with the Erban Decoction group,the relative expression of Nephrin mRNA in the Yiban Decoction group was significantly up-regulated(P <0.05),the relative expression of Wnt4,Wnt5 a and ?-catenin mRNA in the Yiban Decoction group were not significantly down-regulated(P>0.05).7 The immunohistochemical results of Nephrin,Wnt4,Wnt5 a and?-catenin in each group:compared with the normal group,the average integral optical density expression of Nephrin in the model group decreased significantly(P<0.05),and the average integral optical density expression of Wnt4,Wnt5 a and ?-catenin in the model group was significantly increased(P<0.05).Compared with the model group,the average integral optical density expression of Nephrin increased significantly in the Shenyan Kangfu Tablet group,the Yiban Decoction group and the Erban Decoction group(P<0.05),and the average integral optical density expression of Wnt4,Wnt5 a and ?-catenin significantly decreased(P<0.05)in the Shenyan Kangfu Tablet group,the Yiban Decoction group and the Erban Decoction group.Compared with the Shenyan Kangfu Tablet group,the average integral optical density expression of Nephrin in the Yiban Decoction group significantly increased(P<0.05),the difference in the Erban Decoction group was not significant(P>0.05),the average integral optical density expression of Wnt4,Wnt5 a and ?-catenin in the Erban Decoction groupwas not significantly decreased(P>0.05).Compared with the Erban Decoction group,the average integral optical density expression of Nephrin in the Yiban Decoction group significantly increased(P<0.05),the average integral optical density expression of Wnt4,Wnt5 a and ?-catenin in the Yiban Decoction group did not decreased significantly(P>0.05).Conlusions1.The rat model of IgA nephropathy was successfully induced by bovine serum albumin,carbon tetrachloride and lipopolysaccharide.IgA nephropathy model rats showed proteinuria,hematuria.Shenyan Kangfu Tablet,Yiban Decoction,Erban Decoction can reduce hematuria,proteinuria,delay the progress of IgA nephropathy,and the effect of Yiban Decoction on treating proteinuria was better than Erban Decoction.2.Compared with Erban Decoction,Yiban Decoction has better curative effect on treatment of proteinuria,such speculation could be increased Nephrin and its mRNA expression to reduce podocyte damage,reducing Wnt4,Wnt5 a,?-catenin and its mRNA expression,inhibition of Wnt signaling pathways of abnormal activation,so as to reduce proteinuria,further delay the progress of IgA nephropathy,provide a certain modern medicine theoretical basis for the clinical application of Yiban Decoction. |
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