| Objective:To investigate the effect of ChongCaoYiSheng granule on the membranous nephropathy(MN) induced by cationic bovine serum albumin(C-BSA) in rats,and to explore the mechanism of increased sensitivity to glucocorticoid.Methods:Experiment One:Except12rats were the comparison group,72wistar rats were divided into5groups according to24hUPQ:model group, western medicine group, ChongCaoYiSheng granule low dose group, medium dose group, and high dose group. Rats model were induced by repeatedly tail vein injection C-BSA. The western medicine group were fed with suspension liquid consisted of Prednisone acetate and cyclosporine A. While ChongCaoYiSheng group was fed with traditional Chinese medicine solution, and the normal group was fed with normal saline with the same volume as the western medicine group. Every group was fed one time one day for4weeks. The general conditions of rats were dynamically observed during treatment, and the24hUPQ were determined before and after treatment. ALB, TP, TG, and CHOL level of rats blood serum were measured after treatment which was taken from the anaesthetic rats. All rats were killed after4weeks, and pathological changes were observed by light microscopy and electron microscope. RealTime PCR was used to detect the expression of GRamRNA and MDRlmRNA. Immunohistochemistry and Western Blot method were used to examine the expression of GRa and P-gp170.Experiment Two:48wistar rats were randomly divided into4groups:normal group, pure model group, model group after hormonal intervention (glucocorticoid group), Chinese medicine treatment model after hormonal intervention (glucocorticoid combined with Chinese medicine group), with12rats in every group. The pure model group was founded by the same method as in the experiment one. The glucocorticoid group was founded by feeding the pure model group with Prednisone acetate suspension liquid (6.3mg/kg/d), by which the glucocorticoid C-BSA rats model are founded. The glucocorticoid combined with Chinese medicine group was fed with ChongCaoYiSheng medium dose solution (3.6g/kg/d). The normal group, pure model group and glucocorticoid group was fed with the same volume normal saline. Every group were fed one time each day for4weeks. The general conditions of rats were dynamically observed during treatment, and the24hUPQ were determined before and after treatment. After4weeks, pathological changes were observed by light microscopy and electron microscope, the same indexes (ALB, TP, TG and CHOL level) as experiment one were measured, and the same detecting measures (such as RealTime PCR, Immunohistochemistry and Western Blot method) were also used to examine the expression of GRamRNA, MDR1mRNA, GRa and P-gp170.Results:(1) The fur of the model group rats were loose and lackluster and they showed declining activity, listlessness, and lack of food intakes. There were edema and fur shedding which were observed in the toe slack skin of the model rats. The status of rats in ChongChao YiSheng granule low, medium, and high dose group were ameliorative distinctly, which demonstrated more activity, eating more and their edema were faded. The edema of rats in western medicine group were faded; however their status could not compete with those in Chinese medicine group, and someone were dead.(2) The24hUPQ of rats in ChongChao YiSheng granule group are lower than that model group (P<0.01); the ALB and TP level of blood serum were higher than that of model group (P<0.01); and the TG and CHOL level of rats were less than that model group (P<0.01);(3) The24hUPQ of rats with glucocorticoid combined with Chinese medicine group were lower than that of model group and glucocorticoid group (P<0.01); But the TG, CHOL level of rats were below that of the model group and glucocorticoid group (P<0.01);(4) The results of Real Time PCR and Immunohistochemical results indicated that the GRa and P-gp170expressions level of rats in model group were not obviously discrepancy from normal group (P>0.05);(5) Comparing to normal group, the results of Real Time PCR, Immunohistochemical and Western Blot indicated that the GRa expressions in nail tissue of rats in glucocorticoid group were decreased (P<0.01), whereas the expressions of P-gpl70were raised (P<0.01):(6) Comparing to glucocorticoid group, the results of Real Time PCR, Immunohistochemical and Western Blot indicated that the GRa expressions in nail tissue of rats in glucocorticoid combined with Chinese medicine group were raised (P<0.01), nevertheless the expressions of P-gp170were reduced (P<0.01);Conclusion:(1) ChongCaoYiSheng granule can ameliorate the status of C-BSA rats and improve their quality of life. Its effect in this respect is better than that of western medicine group;(2) ChongCaoYiSheng granule can reduce the albuminuria of C-BSA rats, meliorate Hypoproteinemia, and adjust disturbance of lipid metabolism. At the same time, it can alleviate the nephritic pathological changes of C-BSA rats and protect Glomerulus;(3) The effect of ChongCaoYiSheng granule combined with glucocorticoid treatment on C-BSA rats were fast and significant. It shortens the treating time and has better treatment effect than treatment only with glucocorticoid treatment;(4) Possibly, there were not changes of GRa and P-gp170expressions in renal tissue of the simple C-BSA model rats;(5) Glucocorticoid can reduce the GRa expressions in renal tissue of C-BSA rats and raise the expressions of P-gp170;(6) ChongCaoYiSheng paticle can enhance the glucocorticoid sensitive and improve the GC resistance of RNS. The mechanism may have to do with the up-regulation GRa expressions and down regulation P-gp170expressions of renal tissue. |
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