| Objective: To observe the protein expression of JNK signaling pathways and p-JNK,ERK1/2 signaling pathways and p-ERK1/2 in sufferers with CRF damp-heat syndrome before and after oral administration of Qingshen granules in PBMC.And to inquire whether Qingshen granules can interfere with CKD,The activation of JNK signaling pathway and ERK1/2 signaling pathway in PBMC patients can inhibit renal fibrosis and delay the progression of the disease.Methods: on the basis of the selection criteria and table of random number,70 sufferers with CRF damp-heat syndrome were separated into treatment group and control group,and 30 patients in the normal group were selected.The study was finally completed in 64 cases(32 in the treatment group and 32 in the control group).Both groups of subjects were treated with Western medicine and Chinese medicine retention enema.The treatment group was given Qingshen Granules(30g / day,divided into 3 times,for 12 weeks).To observe the protein expressions of JNK,p-JNK,ERK1/2 and p-ERK1/2 in PBMC,TCM syndrome scores,Scr and eGFR of the two groups before and after treatment,at the same time compare the clinical efficacy and TCM syndrome efficacy of the two groups of patients.Results: 1.Scr: There was no significant difference between the two groups before treatment(P>0.05).After 12 weeks,the Scr of the treatment group was significantly lower than that before treatment(P<0.01),and significantly decreased compared with the control group(P<0.01).There was no significant change before and after treatment in the control group(P>0.05).2.eGFR: There was no significant difference between the two groups before treatment(P>0.05).After 12 weeks,the treatment group was significantly increased compared with that before treatment(P<0.01),and significantly increased compared with the control group(P<0.01).The difference between the patients before and after treatment was small in the control group(P>0.05).3.Clinical efficacy: The total effective rate of the treatment group(81.25%)was significantly higher than that of the control group(53.13%)(P<0.05).4.TCM syndrome efficacy: The total effective rate of the treatment group(84.38%)was significantly higher than that of the control group(56.25%)(P<0.05).5.TCM syndrome scores: There was no significant difference between the two groups before treatment(P>0.05).After 12 weeks,the treatment group was effective(P<0.01),and the control group was effective(P<0.05),and the treatment group score was higher than the control group(P<0.01).6.P-JNK,P-ERK1/2: The difference between the two groups before treatment was not obvious(P>0.05);After 12 weeks,the two groups were significantly decreased(P<0.01);the treatment group was significantly lower than the control group after treatment.(P<0.01);7.JNK,ERK1/2: There was no difference between the two groups before treatment(P>0.05);After 12 weeks,two groups of patients decreased significantly compared with pre-treatment(P<0.01);however,there was no significant difference between the two groups after treatment(P>0.05).8.Safety indicators showed that after 12 weeks,there were no abnormal changes in blood routine,urine routine and fecal routine,ALT and AST,and ECG.Conclusion: 1.The expression levels of JNK,p-JNK,ERK1/2 and p-ERK1/2 protein in PBMC of patients with CRF were higher than those of normal group;2.Qingshen Granule can improve clinical symptoms and renal function in patients with CRF damp-heat syndrome;3.Qingshen Granules can decrease the expression levels of JNK and p-JNK,ERK1/2 and p-ERK1/2 in PBMC of CRF patients;4.There were no abnormal changes in the safety indicators before and after treatment,and there was no adverse drug reaction after taking Qingshen Granules.Objective: The expression of c-Jun N-terminal kinase(JNK)signaling pathway,p-JNK,extracellular regulated protein kinases(ERK1/2)signaling pathway and p-ERK1/2 protein in kidney tissues of 5/6 nephrectomized rats in each dose group of Qingshen Granule were detected to study whether Qingshen Granule could interfere with the activation of ERK1/2 and JNK signaling pathways.To achieve the role of Anti-renal fibrosis.Methods: 72 male Sprague-Dawley rats were randomly divided into normal group,model group,losartan group and Qingshen granules in high,medium and low dose groups.Except the normal group,the other groups were made into a 5/6 nephrectomy model according to standard experimental methods.The normal group and the model group were given standard animal feed and clean water;the losartan group was intragastrically administered with the losartan potassium tablet suspension;the other three groups were sequentially administered with the corresponding dose of Qingshen granule aqueous solution.After 12 weeks of observation,BUN,Scr,Ccr,24-hour Upro and Ucr were measured;Using Western blot analysis the protein expression of JNK,p-JNK,ERK1/2,p-ERK1/2 in kidney tissue;PECAM-1/CD31 was immunofluorescent double-labeled with p-JNK and p-ERK1/2 and then scanned by laser confocal microscopy;The pathological changes of kidney in 6 groups of SD rats were observed by HE,masson and PAS stainingResults:1.Scr and BUN: The model group was significantly higher than the normal group(P<0.01).Compared with the model group,the BUN and Scr of the Qingshen granules group and the losartan group were significantly lower(P<0.01);The difference between the Qingshen granules high,medium and low dose group(P<0.01),and the high dose group was the lowest;the Qingshen granules high dose was significantly lower than that of the losartan group(P<0.01).The middle and low doses of Qingshen granules were not significantly different from the losartan group(P>0.05).2.24 h Upro: The model group was significantly higher than the normal group(P<0.01).Compared with the model group,the other groups was significantly decreased(P<0.01);the Qingshen granules were significantly different in different dose groups(P<0.01).The high dose group was the lowest;the Qingshen granule high dose group was significantly lower than the losartan group(P<0.01);the Qingshen granules in the middle and low dose groups were not significantly different from the losartan group(P>0.05).3.Ccr : The model group was significantly lower than the normal group(P<0.01).Compared with the model group,the Qingshen granule high,medium and low dose group and the losartan group were increased significantly(P<0.01);the Qingshen granules high dose group was highest(P<0.01);the Qingshen granule high dose group was significantly higher than the losartan group(P<0.01);There was no significant difference among the Qingshen granule middle dose group and the Qingshen granule low dose group and losartan group(P>0.05).4.ERK1/2,p-ERK1/2 protein expression(Western blot method)(1)ERK1/2: The expression of ERK1/2 protein in the model group was significantly higher than that in the normal group(P<0.05).Compared with the model group,the expression of ERK1/2 protein in the each Qingshen granule group and the losartan group were decreased significantly(P<0.01).There was no significant difference between the Qingshen granules in each dose group(P>0.05).There was no significant difference between the Qingshen granules group and the losartan group(P>0.05).(2)p-ERK1/2: The expression of p-ERK1/2 protein in the model group was significantly higher than that in the normal group(P<0.01).Compared with the model group,the expression of p-ERK1/2 protein in the each Qingshen granule group and the losartan group was significantly decreased(P<0.01);the Qingshen granules were significantly different in different dose groups(P<0.01),The Qingshen granule high dose group was the lowest;compared with the losartan group,the Qingshen granule middle dose group and the Qingshen granule low dose group was increased significantly(P<0.01);There was no significant difference between the high-dose Qingshen group and the losartan group(P>0.05).5.The expression of JNK and p-JNK protein(Western blot method)(1)JNK: The expression of JNK protein in the model group was significantly higher than that in the normal group(P<0.05).Compared with the model group,the expression of JNK protein in the each Qingshen granule group and the losartan group were decreased significantly(P<0.01).There was no significant difference between the Qingshen granules in each dose group(P>0.05).There was no significant difference between the Qingshen granules group and the losartan group(P>0.05).(2)p-JNK: The expression of p-ERK1/2 protein in the model group was significantly higher than that in the normal group(P<0.01).Compared with the model group,the expression of p-ERK1/2 protein in the each Qingshen granule group and the losartan group was significantly decreased(P<0.01);the Qingshen granules were significantly different in different dose groups(P<0.01),The Qingshen granule high dose group was the lowest;compared with the losartan group,the Qingshen granule high and medium dose group was significantly decreased(P<0.01);the Qingshen granule low group was significantly increased(P>0.05).6.Immunofluorescence double staining was used to detect the expression of CD31 and p-ERK1/2,CD31 and p-JNK(laser confocal microscopy scanning imaging)(1)CD31 and p-ERK1/2: The expression of p-ERK1/2 was not found in the normal group.Compared with the model group,the expression of p-ERK1/2 of the Qingshen Granule each dose group and the Losartan group was lower.Compared with every group,the lowest expression was in the high dose group.(2)CD31 and p-JNK: The expression of p-ERK1/2 was not found in the normal group.Compared with the model group,the expression of p-ERK1/2 of the Qingshen Granule each dose group and the Losartan group was lower.Compared with every group,the lowest expression was in the high dose group.7.Pathomorphological changes of kidneys in rats of each group: There was no abnormal change in the pathological structure of the kidney in the normal group;the model group showed that the renal tubular lumen was obviously dilated,the epithelial cells of the renal tubules were exfoliated and vacuolated,a large number of inflammatory cells in the renal interstitium were infiltrated and the deposition of blue-stained collagen fibers was obvious,the index of renal interstitial injury was significantly increased,the size and shape of the glomeruli were different,mesangial cell proliferation and matrix increase Focal glomerulosclerosis were observed in some cases.Compared with the model group,the above pathological changes of rats in the high,middle and low dose groups of Qingshen Granules and losartan groups were alleviated to varying degrees.Conclusion: 1.Activation of JNK signaling pathway and ERK1/2 signaling pathway is involved in the formation of RIF,especially p-JNK and p-ERK1/2 play a major role in the intermediate process.2.Qingshen Granule can reduce Scr and BUN,decrease proteinuria and increase Ccr in rats with chronic renal failure.3.Qingshen Granule can alleviate the pathological degree of kidney tissue in rats with chronic renal failure.4.Qingshen Granule can reduce the expression of ERK1/2,p-ERK1/2,JNK1/2 and p-JNK1/2 in renal tissue of rats with chronic renal failure,and achieve the effect of renal interstitial fibrosis by inhibiting the activation of JNK and ERK1/2 signaling pathways. |
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