| Objective:By observing the effect of Jin Shuiqing on the expression level of TGF-β1 and NF-κ B in serum and kidney tissues of MsPGN damp-heat syndrome hematuria’s rats,the mechanism of Jin shuiqhing prevention and treatment of the model was discussed,and the changes and relationship of urine,biochemistry,pathology and inflammatory factors in rats were observed partially and dynamically,which is to preliminary explore the occurrence and development of this mode,and to provide basis for clinical treatment of hematuria.Method:75 SPF Healthy Male SD rats(3 weeks old)were randomly divided into 5 groups:normal group,model group(mould group,mould two group),treatment group(jinshuiqing early group,jinshuiqing late group),there is 15 in each group.Mould group on the basis of oral and tail intravenous bovine serum albumin(BSA)and tail intravenous staphylococcal enterotoxin B(SEB),combined with hot humid environment and high sugar and high fat diet,the rat model ofhematuria and damp heat syndrome was prepared.Mould two group,jinshuiqing early group and jinshuiqing late group are to change the tail intravenous SEB of mould group to subcutaneous injection of carbon tetrachloride(CCL4)and tail intravenous lipid polysaccharide(LPS).Jinshuiqing early group and jinshuiqing late group began oral jinShuiqing respectively in the 7th week and 9th week,daily to 12 th weekends.The normal group gives an equal amount of acidification of water,daily to 12 weekends.Observing the rats’ the general situation in each group.Urine was collected weekly,the number of erythrocyte was measured in the odd week of experiment,and the 24 hours’ urine protein was measured in double weeks.Each group of 5 are killed in experiment 8th,10 th,12th week,and their blood biochemistry are tested,the pathological changes of renal tissues in rats were observed under optical microscope,Enzyme-linked immunosorbent assay(Elisa)and immunohistochemical assay detect respectively the content of TGF-β1 and NF-κB in serum and renal tissues.Results:1.General state: In the normal group,there was no obvious abnormality in the mental state,activity,reactivity,weight and so on of rats during the experiment.During the modelling,the model groups and treatment groups rats were tired of lethargy,hair shedding,eating decreased,weight loss,short urination,stool,the tongue is dark red,but the degrees of which are not equal,and some of rats is severe damp-heat symptoms such as subcutaneous edema,anus redness,etc.There was no obvious difference in the damp-heat symptom of mould group and mould two group.The heat-damp symptoms of each treatment group were improved in different degrees,and theheat-damp symptoms were most significant improved in early gold group.2.Detection of RBC and proteins in urine: The normal group’s rats had no or minimal hematuria and proteinuria during the experiment.The model groups had varying degrees of hematuria and proteinuria respectively in the 6th week and 8th week during the experiment.The model groups showed hematuria and a small amount of proteinuria(24Hupro>10mg)in the 8th week,and the hematuria and mass proteinuria(24Hupro>50mg)in the 10 th week,the model groups were statistically significant compared with the normal group(P<0.05).There is no statistically significant difference in the hematuria of mould group and model two groups(P>0.05),and the comparison of proteinuria between mould group and model two groups is statistically significant(P<0.05).The hematuria and proteinuria decreased gradually in each treatment group,the treatment groups were statistically significant compared with the model two groups(P<0.05),jinshuiqing early group compared with mould two group were statistically significant in 12 th week(P<0.01).Jinshuiqing early group compared with Jinshuiqing late group was statistically significant(P<0.05).3.Renal function test: The urea nitrogen and creatinine of the model groups and the treatment groups were higher than the normal group during the preparation of animal models,their comparison don’t have a statistically significant(P>0.05).4.Blood lipid test:The serum total cholesterol and trigly-ceride increased markedly in each model group,and they were statistically significant compared with normal group(P<0.01).The comparison of serum total cholesterol and triglyceride in mouldgroup and mould two group was not statistically significant(P>0.05).The content of CHOL and TG decreased in the treatment groups,the comparison of serum total cholesterol and triglyceride in jinshuiqing early group and mould two group was statistically significant(P < 0.05),and The comparison of serum total cholesterol and triglyceride was not statistically significant in Jinshuiqing late group and mould two group(P>0.05).5.Enzyme-linked immunosorbent assay(ELISA): The model group’s expression level of serum TGF-β1 and NF-κB increased markedly,compared with the normal group,the former was not statistically significant(P > 0.05)and the latter was statistically significant(P<0.05)in 8th week,the model groups were statistically significant in the 10 th week and 12 th week of the experiment(P<0.05).The comparison of expression level of serum TGF-β1 and NF-κB in mould group and mould two group was not statistically significant(P>0.05).Expression level of TGF-β1and NF-κB decreased in treatment groups,and the comparison of expression level of serum TGF-β1 and NF-κB in Jingshuiqing early group and mould two group was statistically significant in the 12 th week(P<0.05),the comparison of them in Jingshuiqing late group and mould two group were not statistically significant(P>0.05).The comparison of expression level of NF-κB in Jingshuiqing early group and Jingshuiqing late group was statistically significant(P<0.05),the comparison of expression level of TGF-β1 was not statistically significant(P>0.05).6.Immunohistochemistry: There was no or occasionally slight TGF-β 1’s and NF-κB’s brown positive expression in rat glomerular and renal tubules of normal group;TGF-β1 and NF-κBhas strong expression in the rat kidney tissues of model groups,brown positive expression is mainly distributed in the Mesangial area,renal tubule epithelial cells and so on;there were positive expressions in the treatment groups and the site was the same as the model group,but the positive expressions’ degree was decreased.OD value of image analysis displayed: the OD values of each model group are higher than the normal group,the comparison of expression level of kidney tissues TGF-β1 OD value in both was no statistically significant(P>0.05)and NF-κB OD value was statistically significant(P<0.05)in the experimental 8th week,the comparison of expression level of kidney tissues’ TGF-β1 and NF-κB OD value in both were statistically significant in the 10 th week and 12 th week of the experiment(P < 0.05).There is no statistically significant difference between mould group and model two groups(P>0.05).Comparison with the mould two group’s OD values,the treatment groups’ s decreased,and in the 12 th week the comparison of expression level of kidney tissues’ OD value in Jingshuiqing early group and the mould two group were statistically significant(P < 0.05).There was no statistically significant difference between Jingshuiqing late group and the mould two group(P > 0.05).Jingshuiqing early group compared with Jingshuiqing late group was no statistically significant(P>0.05).7.Light microscopic: The normal group’s renal tissues don’t have mesangial cell hyperplasia,extracellular matrix aggregation,capillary loops open good,normal glomerular morphology,renal tubule structure rules,renal interstitial non-inflammatory cell infiltration.There were obvious hyperplasia of mesangial cells andmesangial matrices in each model group,diffuse glomerular mesangial area is divided into leaf-shaped,partial glomerular visible atrophy,capillary lumen is very narrow or very occlusion,renal interstitial with a large number of inflammatory cell infiltration,renal tubular epithelial cells is swollen,denatured and severely deciduous,renal tubule lumen is also occluded in model groups;the pathological changes of the 8th week,10 th week and 12 th week were gradually aggravated,and there was no significant difference between the model group and the model two groups.Glomerular mesangial cells,extracellular matrix mild hyperplasia,renal glomerulus without atrophy,capillary lumen stenosis,renal tubular epithelial cells of even degeneration swelling,interstitial mild inflammatory cell infiltration in jinshuiqing early group,which improved markedly in 12 th week of the experiment.Glomerular mesangial cells,extracellular matrix light medium hyperplasia,renal glomerulus occasionally atrophy,capillary tube cavity light medium narrow,renal tubular epithelial cell degeneration swelling,renal tubular cavity occasionally occlusion,interstitial mild medium-size inflammatory cell infiltration in jinshuiqing late group,which improvement was not obvious compared with jinshuiqing early group in 12 th week of the experiment.Conlusion:According to the etiology and pathology theory of TCM heat-damp syndrome,the comprehensive factors were used to establish a more economical and practical the rat model of damp-heat syndrome hematuria of mesangial proliferative nephritis.JinShuiqing can significantly improve the damp-heat syndrome of the damp-heat syndrome mesangial hyperplasia nephritis hematuria rats,andthrough the reduction of early inflammatory marker factor NF-κB’s and renal factor markers TGF-β1’s expression,thus reducing the renal inflammatory immune response and the synthesis of stimulating factors,reducing the MC hyperplasia and ECM aggregation,reducing hematuria,proteinuria and improve blood lipid disorders,to achieve the goal of preventing and curing MSPGN disease progress,worthy of further popularization of clinical use.The changes of hematuria and proteinuria in rats were consistent with biochemical,renal pathology,NF-κB and TGF-β1 in the 8th week and 10 th week,which could be key points in the development of MsPGN damp-heat syndrome hematuria model,and which are significant for the clinical treatment of hematuria. |
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