| Literature researchDiabetic Nephropathy is not only one of the most common microvascular complication of diabetes mellitus,but also the primary cause of end-stage renal disease and the replecement treatment.Therefore,it serious threat to human health.It recording by the research,about 15%～40%diabetic patients will be tend to become Diabetic Nephropathy.In our country,the propotion of end-stage renal disease due to Diabetic Nephropathy rasing year by year.Which predict Diabetic Nephropathy will become the primary cause of end-stage renal disease.However,the pathogenesis of diabetic nephropathy not yet clearly,it may be caused by many synthetic actions.However,it always a difficult clinic problem to take effective prevention and control,due to many bioactivator take part in the pathogenic process of the Diabetic Nephropathy,which make its physiopathologic mechanism with mutiple pathways,the complex features of the interaction of muti ring segments.Therefore,it has important social significance and economic value in exploring the pathogenesis of diabetic nephropathy and research the treatment method of prolong the process of diabetic nephropathy.As western medicine has many limitations,it is necessary to find out more effective treatments and medicines.While Traditional Chinese Medicine has a long history for treating diabetes and some related deseases.In aspects of the intervene and treatment of disease,chinese herb compound has the feasures of multi-path and multi-target point.Therefore,the usage of it has large potent ion in the treatment of diabetic nephropathy.Traditional Chinese Medicine has a long history for treating diabetes.Syndromes of diabetes and its vascular complications had already been recorded in "Huang Di Nei Jing".The understandings of past dynasties docters and The later scholars on the pathogenesis of diabetic get continuous development,the treatment level of it also get constantly implovement.Summarize the ancient and modern literature of traditional chinese medicine in the treatment of diabetic nephropathy,due to the basic pathogenesis of diabetic in the early period is "the deficiency of both qi and yin and the stagnation of heat and blood",the reasonable treatment of it should "tonifying qi and yin and Spilled hot line by stasis." Xiong Manqi,professor of Guangzhou University of Chinese Medicine,with her research team,had developed a more effective decoction called Jiangtang Sanhuang tablet,which was based on traditional Taohi Cheng Qi Decoction.It achieve good clinical efficacy,Since the treatment on diabetic nephropathy has been more than 20 years.In order to further confirm the treatment effect of it,and to further explore its possible mechanism,we carry the animal model of diabetic nephropathy,and observate its early intervation in rats and its effect on the microRNA-21 signal path.Experimental study:ObjectiveIn order to explore the activemechanism of its treatment on diabetic nephropathy,and to provide the experimental basis and surpport of modern biology for its clinical application,we should use model rats of diabetic nephropathy,observe its effect on the glucose metabolism leavel,renal function,renal pathologic and the microRNA-21 signal path.MethodsMale SD rats were firstly fed for 1 week,then according its weight randomly divided it into two groups,ten of them are the normal group,the rest are build module group.The rats in the module group should be forbidden for food,while water is available for 12 hours before injecting STZ by 45 mg/kg dose.The rats in normal group were injected by the same amount of citric acid buffer.After molding,all rats were taken into the metabolism cage for the standard diet,no using insulin and other antidiabetic drugs.By continuous feeding 3 weeks,then measure its blood sugar and urine volume,if blood sugar>16.7 mmol · L-1 and the original of urine>urine volume 150%,the urinary albumin excretion for 24 hours>30mg,we confirmed the rats module of DN was build successfully.During the molding process,there were 3 rats dead,3 rats’ blood glucose returned to normal,8 rats with blood glucose in standard while the urinary albumin excertion rate not up to standard.Therefore,at last there were 56 rats molded successfully.Randomly divided the 56 model rats into module group 12,jiangtangsanhuang tablet High dose group 11,middle dose group 11,low-dose group 11,irbesarta group 11.Borefore and after the model,all of them were fed with clean ordinary feed.The low-dose group were gavaged by 0.675 g·kg-1,the middle dose group by 1.350 g·kg-1,and the High dose group by 2.700 g·kg-1,irbesarta group by 0.0135 g·kg-1.Before gavaged,all the drugs were mixed with pure water,and make it become 2 ml suspension.Gavaged them once a day,continue to gavage 8 w.both the normal group and the model group gavaged them the same amount of distilled water.During the experiment,make the rats free to eat and drink water,and observe the normal situation in the same time.Every 4 week measure its weight and FBG of the tip of the tail.After the last gavaged,measure urine volume and water consumption in 24 h.When the experiment finished,measure its renal function(BUN、Scr),blood fat(TC、TG、LDL、HDL),glycosylated hemoglobin(HbA1c),24 hurine protein(Upro).Take the rats kidney,calculate it and its relative weight.Observed the pathological changes of rennal tissue by light microscope and electron microscope.Observe the kidney dye situation of TGF-β1、Smad3、Smad7 by using the immunohistochemical method.Detect each rat kidney gene expression of TGF-β1、Smad3、Smad7 by using Western blotting method.And its expression of TGF-β1、Smad3、Smad7、microRNA-21 mRNA by the RTQ-PCR method.results1.In terms of body weight,water consumption and urine volume:after the drug intervene 4 and 8 weeks,compared to the model group,body weight of the drug intervene group be markedly increased(P<0.01).among them,the high dose group had the best effect,followed by the middle dose group and low dose group and irbesarta group had no markedly differe nce(P>0.05).Compared to model group,water comsumption of high dose group and irbesarta had decreased(P<0.01),but the low dose group has no obviously difference(P>0.05).Compared to irbesarta group,low dose group and the middle dose group had no markedly difference(P>0.05).But the high dose group obviously lower than irbesarta group(P<0.01).Urine volume of middle dose group,the high dose group and the irbesarta group are all decreased.but its low dose group has no significant diffe rence(P>0.05);compared to irbesarta group,urine volume of the high dose group had decreased,but the low dose group higher than it.2.As to glucose and lipid metabolism,after 4 and 8 w treatment,high dose group,the middle dose group and low dose group are all lower than the model and the irbesarta group group(P<0.01).and the latter two group had no diffierence(P>0.05).As to glycosylated hemoglobin,after 8 weeks treatment,the high dose group,the middle and the low are all lower than the model and the irbesarta group group(P<0.01).and the latter two group had no diffierence(P>0.05).AS to TC、TG、LDL-C,aft er 8 weeks treatment,the high dose group,the middle dose group and the low dose group are all lower than the model and the irbesarta group(P<0.01),and the latter two group had no diffierence(P>0.05).As to HDL-C,compared to model group,both the middle dose group and the high dose group are obviously increased(P<0.01),but the low dose,the mo del and the irbesarta group had no significant diffierence(P>0.05).3.In terms of renal weight,relative renal weight,renal function and proteinuria,the middle dose,the high dose and the irbesarta group obviouly lower than model group(P<0.01).But the low dose and the mode 1 dose group had no significant diffierence(P>0.05).The middle dose,the high dose and the irbesarta group had no obviously diffierence(P>0.05).But the low dose group obviously low than the irbesarta group(P<0.01).As to relative renal weight,the drug intervene group obviously lower than the model group(P<0.01).The low dose,the high dose and the irbesarta group had no obviouly diffierence(P>0.05).But the low dose group obviously higher than the irbesarta group(P<0.01).As to renal function(BUN、Scr),the drug intervene group obviously lower than the model group(P<0.01).the middle dose and the irbesarta group had no obviously diffierence(P>0.05)The high dose obviously lower than the irbesarta group(P<0.01).But the low dose group obviously higher than the irbesarta group(P<0.01).AS to 24 h proteinuria,before treatment,the model group and the drug intervene had no obviously diffierence(P>0.05).After 8 weeks treatment,compared to model group,the low dose group decreased(P<0.05).The high dose,the middle dose and the irb esarta group obviously decreased(P<0.01).Compared to irbesarta group,the middle dose group had no obviously diffierence(P>0.05).The high dose obviously lower than the irbesarta group(P<0.01).the low dose group obviously higher than the irbesarta group(P<0.01).4.Under the light microscope and electron microscope,observing the rental pathological morphology,the rats rental of normal group had no pathological changes,he rats rental of model group had serious pathologi cal changes,it can be observed that the glomerular volume obviouly increased,swelling,hyperemia,and the narrowed rental capsule.The mesangial cell of the glomerular be proliferated,particial tubular cell hypertrophy,which can see some vacuolar degeneration,capillary lumens stenosis,pedal flat,unclear structure,the mesangial matrix increased,rental tubular epithelial shedding,decrease and shrink.But the The high dose,the low,the middle and the irbesarta group,all of them can implove the rental tissues pathological changes of Diabetic Nephropathy.and among them,the The high dose group had best effect.5.Rental TGF-β1、Smad3、Smad7 protein expression:according to the immunohistochemical staining method,the result display that,compared to normal group,the TGF-β1、Smad3 protein expression of the model group obviouly increased(P<0.01).Which display the brownish yellow particles,and the color getting deeply.The drug intervention group was between the normal group and the model group,and obviously lower than the model group(P<0.01).Compared to the irbesarta group,the The high dose group had no significant diffierence(P>0.05),the low dose group and the middle group higher than the irbesarta group(P<0.05).Compared to model group,the Smad7 protein expression of The drug intervention group obv iously increased(P<0.01).The Smad7 protein expression of the irbesarta group obviously higher than the high dose,the low dose and the middle group(P<0.01).Measured the Rental TGF-β1、Smad3、Smad7 protein expression by using the Western blotting method displayed that compared normal group,the TGF-β1、Smad3 protein expression of model group obviously increased(P<0.01).The low dose group lower than the model group(P<0.05).the high dose,the middle group and the irbesarta group obviously lower than model group(P<0.01).the TGF-β1、Smad3 protein expression of the high dose group compared to the irbesarta group had no obviously diffierence(P<0.01).And the irbesarta group is superior to the low dose and the middle group(P<0.01 or P<0.05).Compared to norma 1 group,Smad7 protein expression of model group obviously decreased(P<0.01).The drug intervention group obviously higher than the model group(P<0.01).Smad7 protein expression of the irbesarta group obviously higher the low dose and the middle group(P<0.01).But compared to the high dose group that had no significant difierence(P<0.05).6.Detect the TGF-β1、Smad3、Smad7、microRNA-21 mRNA expression of the rental by using the RTQ-PCR method,which can displayed:refer to gene expression in normal group,the TGF-β1、Smad3、Smad7、microRNA-21 mRNA gene expression of the model group obviously increased(P<0.01).Compared to the model group,the drug intervene group obviously decreased(P<0.01).the high dose group and the irbesarta group had no obviously diff ierence(P>0.05),but the low dose and the middle group obviously higher than the irbesarta group(P<0.01).refer to gene expression in norma 1 group,Smad7 mRNA gene expressionof model group obviously decreased(P<0.01).Compared to model group,the the drug intervene group obviously increased(P<0.01)the high dose group and the irbesarta group had no obviously diffierence(P>0.05),but the low dose and the middle group obviously lower than the irbesarta group(P<0.01).Conclusion1.In summaring up the pre physicians on the basis of diabetes pathogenesis and treatment,and it comes to a decision that Diabetic Nephropathy pathogenesis intermingled xu and shi,the formal is the essence and the latter is appearance.Only by using the treatment methods of strengthening the body resistence and eliminating evil,and combine with tonifying qi and yin,Spilled hot line by stasis,activate yang and promoting blood,and through relieving drain turbidity four methods can we prevent and cure this disease.2.Jiangtangsanhuang tablet had the effection of improve the normal situation of the rats and reduce its bloods sugar,and regulate blood fat.3.Jiangtangsanhuang tablet had the effection of protect the rental function of the Diabetic Nephropathy rats,which can reduce its urine protein and the serum BUN and Scr.4.Jiangtangsanhuang tablet can improve the pathological damage of rental tissue in the Diabetic Nephropathy rats,and delay the pathological damage of kidney.5.Jiangtangsanhuang tablet had two-way regulation:TGF-β1、Smad3 protein expression was down-regulated,Smad7 protein expression was up-regulated;TGF-β1、Smad3、microRNA-21 mRNA expression of rats rental was down-regulated,Smad7 mRNA expression was up-regulated.Which control the signal-path transmission of microRNA-21 and TGF-β1/Smad of the Diabetic Nephropathy,and delayed the development of the course of disease. |
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