| Objectives: Diabetic nephropathy ( DN ) is a part of micovasclor complication and one of the common reason of end stage renal disease. The researchers keep working on the reason and treatment of DN. The traditional Chinese herbs play an important role in preventing DN because of its advantage. We invented an effective traditional medicine, according to the pathogenesis which is at the early stage of DN and clinical experience for treating DN. This medicine has the functions of promoting circulation of qi and yin, dispersing blood stasis and dredging collateral which was composed of astragali, figwort root, carapax amydae, angle worm, ect. The clinical data showed that the prescription had good effect on the early stage of DN. This study aim to observe the curative effect of herbs of supplement qi and nourish yin resolve masses and dredge meridian(sZY) and investigate the mechanism in renin angiotensin system in DN rats, besides, a clinical study of ZY in the treatment of diabetic nephropathy was carried out. All the above that have been done was to provide the foundation in order to generalization and exploitation.Motheds:1 A clinical study of ZY in the treatment of diabetic nephropathy. 81 patients with DN of theⅢstage were randomly divided into treatment group(41 cases)and control group(40 cases),The control group received beinazepril, 10mg/d, while the treatment group received ZY herbs with a dose, 2 times a day, for 3 months. The UAER,β2-MG in serum,β2-MG in urine,α1-MG in urine, FIB and D-D were detected before and after the therapy.The symptom, physical sign and integral calculus were recored.2 Eeffct of ZY on the treatment in DN rats: Forty-six male Sprague-Dauley rats were selected in this experiment. According to the weight, the animals were randomly divided into normal group(10 rats)and DM model group(36 rats). Besides the normal group, all the rats were injected with streptozotocin (STZ) in the dose of 60 mg/kg, the normal group were given the corresponding quantity's physiological saline. After 72 hours, measured their blood glucose, took"≥16.7mmol/L"as the DM model standard. According to the blood glucose level, the rats of DM model were randomly divided into three groups: 12 in the model group, 12 in benazepril group and 12 in ZY group. After one week of DM models were made successfully, the drugs started be given to the rats according to adult dosage 20 times, once daily. The rats of ZY group were given the ZY by gavage, 26g/kg, the benazepril group were administrated with benazepril by gavage, 10mg/kg, the normal group and the model group were given physiological saline by gavage. Each group was given the medicine for 23 weeks. After 24 week of DM models made successfully, all rats were put into metabolism cage to obtain 24 hour urine. All animals were killed and the blood and the renal tissue were obtained. 24 hour urinary protein quantitative, glycohemoglobin(HbA1C), total cholesterol(TC), triglyceride (TG), serum creatinine (Scr), urea nitrogen (BUN) were assayed with the automaticbiochemistry analyzer. Pathomorphology change and CollagenⅣof kidney were observed.3 Eeffct of ZY on renin angiotensin system in DN rats:DM model was duplicated the same as the above.After 24 week of DM models made successfully, the renal tissue were obtained. AngⅡand ACE was determined by radioimmunity and ultraviolet spectrophotometry. The protein and genic of AT1R,ACE,ACE2 were measured by immunohistochemical method and RT-PCR.Results:1 A clinical study of ZY in the treatment of diabetic nephropathy1.1 Comparison of syndrome integration between pre-therapy and post-therapy in both groups: There was no difference of syndrome integration between treatment group and control group(P >0.05),After the treatment, There was significangt difference of the syndrome integration between pre-therapy and post-therapy in both groups(P <0.01), and the same as both groups after the treatment(P <0.01).1.2 Comparison of syndrome between pre-therapy and post-therapy in both groups:After the therapy, syndrome in treatment group were improved significantly compared with control group(P <0.01).1.3 Comparison of curative effects between pre-therapy and post-therapy in both groups:After the therapy, curative effects in treatment group were improved significantly compared with control group(P <0.01).1.4 Comparison of UAER,β2-MG in serum,β2-MG in urine andα1-MG in urine between pre-therapy and post-therapy in both groups: Significant difference was found in UAER,β2-MG in serum,β2-MG in urine andα1-MG in urine between pre-therapy and post-therapy in both groups(P <0.05 or P <0.01). After the treatmens, UAER,β2-MG in serum,β2-MG in urine andα1-MG in urine in treatment group decreased significantly compared with control group(P <0.05). In treatment group, FIB and D-D decreased significantly after treatment(sP <0.01). There were great difference in FIB and D-D between control and treatment group after the therapy(P <0.01).There was no difference in control group between pre-therapy and post-therapy(P >0.05).1.5 Comparison of blood fat between pre-therapy and post-therapy in both groups:Compared with pre-therapy, TC, TG, LDL of treatment group decreased while the HDL increased after the therapy(P <0.01). After the therapy, TC, LDL of treatment group decreased while the HDL increased compared with control group(P <0.01). In control group, TG decreased significantly after the therapy(P <0.01)while there was no difference in TC, LDL, HDL between pre-therapy and post-therapy(P >0.05).1.6 Comparison of blood glucose, glycosylated hemoglobin between pre-therapy and post-therapy in both groups:There was no difference in FBG and HbAIc between pre-therapy and post-therapy in both groups(P >0.05).2 Eeffct of ZY on the treatment in DN rats:2.1 The 24 hour urinary protein quantitative of each groupThe 24 hour urinary protein quantitative of the rats in model group, benazepril group and ZY group was significantly higher than that of normal group (P <0.01), compared with model group, the 24 hour urinary protein quantitative of benazepril group and ZY group was obviously lower (P <0.01).There was no difference between benazepril group and ZY group (P >0.05).2.2 The HbA1C of each groupCompared with normal group, the HbA1C of the rats in model group, benazepril group and ZY group was significantly higher (P <0.01), but there was no difference among model group, benazepril group and ZY group (P >0.05).2.3 The TC and TG of each groupThe TC of the rats in model group, benazepril group and ZY group was significantly higher than that of normal group (P <0.01), compared with model group, the TC of ZY group was obviously lower (P <0.01), there was no difference between benazepril group and model group (P >0.05). The TG of the rats in normal group, benazepril group and ZY group was significantly lower than that of model group (P <0.01), there was no difference among normal group, benazepril group and ZY group (P>0.05).2.4 The Scr and BUN of each groupCompared with model group, the Scr of the rats in normal group, benazepril group and ZY group was significantly lower (P<0.01), and benazepril group was obviously higher than that of normal group (P<0.05). There was no difference between normal group and ZY group (P>0.05). The BUN of the rats in normal group, benazepril group and ZY group was significantly lower than that of model group (P<0.01). Compared with normal group, the BUN of benazepril group and ZY group was significantly higher (P<0.01), and there was no difference between benazepril group and ZY group (P>0.05).2.5 The pathomorphology change of kidney by Light microscope observationIn the normal group: the structure of renal glomerulus was integrated, glomerular capillary basement membrane, mesenterium and base material were normal. In the model group: glomerular hypertrophy, thickening of glomerular capillary basement membrane, mesangial cell proliferation, mesangial matrix increased, mesangial area broadening and tubulointerstitial fibrosis were seen. vacuolar degeneration, analosis and hyaline cast were seen in some renal tubule. The benazepril group and ZY group also showed different degree of pathological changes, but the pathological degree was obviously lighter than the changes of the model group.2.6 Observations under transmission electron microscopyIn normal group: the structure of glomerular capillary basement membrane was clear and complete, microcirculatary endothelial cell, foot processes was normal. In the model group: most of the glomerular capillary basement membrane showed significant homogeneous thicking, partial basement membrane showed elevation in the shape of hills; microcirculatary endothelial cell significantly confluence and the most of window structure disappeared; the fusion of the foot processes was extensive. The benazepril group and ZY group also showed the similar pathological changes, but the pathological degree was lighter than that in model group.2.7 Expression of CollegeⅣexamined by immunohistochemistry:In normal group, Col-Ⅳhas the basal expression. Compared with normal group, the expression of Col-Ⅳin model group was stronger(P<0.01). Compared with model group, the expression of Col-Ⅳin benazepril group and ZY group was significantly lighter (P <0.01), and there was no difference between benazepril group and ZY group (P >0.05).3 Eeffct of ZY on renin angiotensin system in DN rats: 3.1 The cotent of AngⅡin renal tissue:The cotent of AngⅡin model and treatment groups were higher than that in normal group (P<0.01);Compared with model group, the cotent of AngⅡin benazepril group and ZY group was significantly lower (P <0.05), and there was no difference between benazepril group and ZY group (P >0.05).3.2 Expression of AT1R in renal tissue:The results detected by immunohistochemistry showed: The expression of AT1R in model and treatment groups were higher than that in normal group (P<0.01);Compared with model group, the expression of AT1R in benazepril group and ZY group was significantly decreased (P <0.01), and there was no difference between benazepril group and ZY group (P >0.05).The results detected by RT-PCR showed: The expression of AT1R in model and treatment groups were higher than that in normal group (P<0.01);Compared with model group, the expression of AT1R in benazepril group and ZY group was significantly decreased (P <0.01), and there was no difference between benazepril group and ZY group (P >0.05).3.3 The activity of ACE:The results shows that: The activity of ACE in model group was higher than that in normal group (P<0.01);Compared with control group, the activity of ACE in benazepril group and ZY group was significantly decreased (P <0.05); Compared with model group, the activity of ACE in benazepril group and ZY group was significantly decreased (P <0.01), and there was no difference between benazepril group and ZY group(P>0.05).3.4 Expression of ACE in renal tissue:The results detected by immunohistochemistry showed: Compared with normal group, the expression of ACE in model and treatment groups were decreased significantly (P<0.01),there was no difference between model,benazepril group and ZY group (P>0.05).The results detected by RT-PCR showed: Compared with normal group, the expression of ACE2 in model and treatment groups were decreased significantly (P<0.01),there was no difference between model,benazepril group and ZY group (P>0.05)。3.5 Expression of ACE2 in renal tissue:The results detected by immunohistochemistry showed: Compared with normal group, the expression of ACE2 in model and treatment groups were decreased significantly (P<0.01),the expression of ACE2 in ZY group were higher than that in model group(P<0.01), while there was no difference between benazepril group and model group (P>0.05); the expression of ACE2 in ZY group were higher than that in benazepril group (P<0.05).The results detected by RT-PCR showed: Compared with normal group, the expression of ACE2 in model and treatment groups were decreased significantly (P<0.01),the expression of ACE2 in ZY group were higher than that in model group(P<0.01), while there was no difference between benazepril group and model group (P>0.05); the expression of ACE2 in ZY group were higher than that in benazepril group (P<0.05).Conclusions:1 Herbs of supplement qi and nourish yin resolve masses and dredge meridians could improve the clinical symptom and physical sign of DN patients.2 Herbs of supplement qi and nourish yin resolve masses and dredge meridians could regulate lipid metabolism, improve hypercoagulabale state, decrease the UAER,β2-MG in serum,β2-MG in urine andα1-MG in urine of DN patients which could prevent DN from aggravating.3 Herbs of supplement qi and nourish yin resolve masses and dredge meridians could decreased the DN rat's proteinuria, improve the lipid metabolism, decrease Scr and BUN, lessen the damage of nephridial tissue. It also could decrease the ColIV deposition in order to slow down the progress of glomerulosclerosis.4 Herbs of supplement qi and nourish yin resolve masses and dredge meridians could decrease the content of AngⅡwhich could protect the renal function.5 Herbs of supplement qi and nourish yin resolve masses and dredge meridians could decrease the activity of ACE in the kidney of DN rats, reduce the production of AngⅡand promote the expression of ACE2 which could slow down the progress of glomerulosclerosis and protect the renal function.
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