Research On The Effect Of Yishen Ning Granule On NF-κB Signaling Pathway And TGF-β1/CTGF Of Chronic Glomerulonephritis Rats

Objective: To investigate the effects of Yishen Ning granule on NF-κB signaling pathway and TGF-β1/CTGF of chronic glomerulonephritis rats by reference the improved border method to establish the rat model of chronic nephritis,so as to study on Yishen Ning in the treatment mechanism from the micro level.Method: 50 healthy SPF grade male SD rats were selected, adaptive feeding for 1weeks. According to the random grouping method, 10 rats were selected as normal group, and the other 40 rats were established C-BSA model. Carry on pre immunization on first days, C-BSA soluble in PBS(phosphate buffer solution), and the same amount of the incomplete adjuvant mixed into C-BSA’s 2.5mg/ml milk white solution, each rat multi-point subcutaneous injection by 1ml. On eighth days,Carry on pre immunization again with the same method. Carry on formal immunity on Tenth days, intravenous injection of C-BSA 16 mg / kg every other day, dissolved in PBS. Carry on the immunization for 9 times. The rats in the normal control group were injected with normal saline every other day. After the completion of the immune,model rats were divided into model group, Yishen Ning group, benazepril group,Yishen Ning + benazepril group. Animals in each group were given medicine for 8weeks after the model finished. After 8 weeks of treatment, measured levels of 24 h quantitative urinary protein, bun and creatinine.Left kidney specimens to make paraffin sections.The changes of renal tissue were observed by HE staining, PAS staining and Masson staining. The expression of TGF-βl/CTGF and NF-κB in renal tissues were detected by immunohistochemistry.Result:1)Body weight and kidney index of rats After modeling, the weight of modeling rats were lower than that of normal rats(P < 0.05); after 2 weeks of treatment, the body weight of the rats in the treatment group gradually increased to 4 weeks of treatment, the difference had no statistical significance between the treatment group and normal group(P > 0.05). The body weight of the rats in the model group compared with that of normal rats(P < 0.05).After treatment, the model rats with bilateral renal weight significantly heavier than normal rats bilateral kidney weight(P < 0.05). The treatment group with bilateral renal weight and normal rats with bilateral renal weight no statistical significance(P > 0.05).2)24h urinary protein quantitation in rats After modeling, 24 h urinary protein in model group, Yishen Ning group,benazepril group, Yishen Ning + benazepril group were significantly higher than those in normal group(P < 0.05).After 8 weeks of treatment, 24 h urinary protein of the normal group and the model group compared with before treatment, the difference had no statistical significance(P > 0.05), 24 h urine protein of the model group rats was significantly higher than that in the normal group(P < 0.05); compared with before treatment, 24 h urine protein of Yishen Ning group, benazepril group, Yishen Ning + benazepril group decreased significantly(P < 0.05).3)Creatinine and urea nitrogen in rats After 8 weeks of treatment, the levels of creatinine and blood urea nitrogen in the model group were higher than those of other groups(P < 0.05);the difference had no statistical significance between Yishen Ning group and benazepril group(P >0.05), the levels of creatinine and blood urea nitrogen in Yishen Ning + benazepril group were lower than that of Yishen Ning group and benazepril group(P < 0.05).4) Renal biopsy tissue Under the microscope, in HE staining, PAS and Masson staining, normal group basement membrane thickening, epithelial cell morphology normal, mesangial cells and matrix proliferation and no abnormal renal tubules, renal interstitium, PAS and Masson staining without exception.In the model group, the glomerular enlargement, the number of cells increased,the matrix increased, basement membrane thickening, a large number of inflammatory cell infiltration around the glomerular, glomerular appearance.Compared with the model group, glomerular enlargement was not obvious of benazepril group and Yishen Ning group, the proliferation of glomerular mesangial cells was not obvious. The expression level of Yishen Ning + benazepril group was better than that of Yishen Ning group and benazepril group.5)The results of immunohistochemical method Under the light microscope, there was almost no expression of TGF-βl/CTGF and NF-κB in the kidney tissue of normal rats. The positive staining was mainly in the cytoplasm of renal tubular epithelial cells which appeared at the junction of skin and marrow. In the model group, the expression of glomerular cells was stronger than that of mesangial cells, epithelial cells and peripheral renal tubular epithelial cells,which were stronger than the normal group and the treatment group(P<0.05). In the Benazepril group and Yishen Ning group, positive staining intensity and range were reduce compared with the model group(P < 0.05); In the kidney tissue of Yishen Ning + benazepril group, the positive staining intensity and range decreased significantly compared other groups(P < 0.05).Conclusion:1) Yi Shen Ning Granule can effectively improve the symptoms of nephritis in rats, reduce the urine protein of experimental rats, reduce the level of creatinine and urea nitrogen, and have protective effect on renal function;2) To a certain extent, Yi Shen Ning Granule can effectively improve the pathological damage of the kidney of the experimental rats, and slow down the thickness of the basement membrane.3) Yi Shen Ning Granule in a certain extent can play an effective role in the anti renal fibrosis, the mechanism may be down-regulated the expression level of TGF-βl/CTGF and NF-κB, which can provide theoretical support for Yi Shen Ning Granule clinical treatment mechanism.