The Intervention Of Huayu Tongluo Huoxue Traditional Chinese Medicine To Diabetic Nephropathy Rats And Its Effect On Kidney AT₂R Expression

Objective: Diabetic nephropathy(diabetic kindey disease, DKD) is one of the common chronic microvascular complications of diabetes, and it is the main reasons which leading to end-stage renal disease(ESRD). So searching for its pathogenesis and effective preventing and controlling measures is of great significance. Modern medicine has confirmed that the activation of renin-angiotensin system(RAS) is the main mechanism involved in DKD. Based on the cognition of over-activation of the classic pathway of RAS, the angiotensin-converting enzyme-angiotensin Ⅱ- angiotensin type 1 receptor(ACE-AngⅡ-AT1) pathway, in the development of DKD, angiotensin converting enzyme inhibitor(ACEI) and angiotensin receptor blocker(ARB) have been used in clinic. This experiment adopted high fat diet combination with small dose of streptozotocin(STZ, streptozotocin) intraperitoneal injection to develop DKD, huoxue huayue tongluo traditional medicine was used to interveneted, searched for intervention of Huayu Tongluo Huoxue traditional Chinese medicine to diabetic nephropathy rats and its effect on kidney AT2 R expression.Methods: 50 adult male SD rats weighing 60- 80 gm were bred one week. Then they were divided into 2 groups: 9 in normal group(N) and others in model group. Except the normal group, the rats of another groups were bred for high-fat diet 4 weeks. The model groups which fasted for 12 hours were injected STZ with 35mg/kg intraperitoneally. After 72 hours, the rats whose tail vein random blood glucose were measured≥16.7umol/l were successful as DKD model. Then the rats were divided into 3 groups: model group(M), ZY group(ZY) and irbesartan group(I). Four weeks later, the different drugs started to be lavaged to the rats according their different groups every day. 16 weeks later, the blood of rats were drew under anesthesia condition for testing serum biochemical markers, and specimens from renal tissue for electron microscopy, light microscopy, immunohistochemistry and RT-PCR detection.Results:1 The general state of ratsNormal control rats were generally in good condition. The rats whichwere injected STZ were polydipsia, polyphagia, polyuriau, and the modelgroup is the most significant. 2 Body weight, kidney hyper-trophy indexCompared with normal group, the body weight of the rats in model groupwas obviously lighter(P<0.05), there was no significant difference amongthe M, ZY and I groups(P>0.05). The hypertrophy index of the rats inmodel group was obviously higher than normal group(P<0.05), there wasno significant difference among the M, ZY and I groups(P>0.05).3 The 24-hour urinary proteinAfter successful modeling, there was no difference between the fourgroups(P>0.05). In the first 4 weeks, compared with the normal groups, M,I and ZY groups were significantly increased(P<0.05), no significantdifference(P>0.05) between M, ZY and I groups. In the 16 weeks modelgroup was increased significantly compared with the normal group, I groupand ZY group were significantly reduced compared with the model group(P<0.05). No significant difference between I group and ZY group(P>0.05).4 HamegglutinationCompared with the normal group, the PT of model group wassignificantly decreased(P<0.05). Compared with the model group, the PTof ZY group was significantly increased(P<0.05). Compared with thenormal group, the INR of model group was significantly decreased(P<0.05),there was no significant difference among the M, ZY and I groups(P>0.05). 5 Biochemical indicatorsCompared with the normal group, the TP of model group was significantly decreased(P<0.05). Compared with the model group, the TP of treatment groups were significantly increased(P > 0.05), No significant difference between I group and ZY group(P>0.05).Compared with the normal group, the TG and TC of model group wassignificantly increased(P<0.05). Compared with the model group, the TGof treatment groups were significantly decreased(P>0.05), No significantdifference between I group and ZY group(P>0.05), there was no significantdifference among the M, ZY and I groups of TC(P>0.05).Compared with the normal group, the BUN of model group was significantly increased(P<0.05). Compared with the model group, the BUN of treatment groups were significantly decreased(P<0.05). No significant difference between I group and ZY group(P>0.05). Compared with the normal group, the Scr of model group was significantly increased(P<0.05). Compared with the model group, the Scr of treatment groups were slightly decreased(P>0.05).6 Morphological changes in renal tissue 6.1 Photomicrographs of renal cortex from rats in different groupsThe glomerular structure of normal group was complete and clear, andthere were no obviously hypertrophy. The model group showed that,glomerular hypertrophy, the basement membrane of glomerular thickened,the cell of mesangial proliferation, mesangial matrix increased, mesangialarea broadened. The pathological changes of the treatment groups werelighter than the model group.6.2 Ultrastructural changes of glomeruli from rats in different groups undertransmission electron microscopeThe glomerular structure of normal group was complete and the basement membrane of glomerular was clear. Endothelial cell, foot process, and mesangial area did not show significant change. The model group showed that: the glomerular basement membrane was thickened, endothelial cell and the foot process were extensive fusion. The treatment groups showed the similar changes, but the degree was lighter than model group. 7 AT2 R expressed in renal tissue by immunohistochemistryAll groups showed that AT2 R expressed in renal tissue. AT2 R expressedmainly located in vascular endothelial cell and glomerular endothelial cell.The result showed that compared with the normal group, the AT2 R of modelgroup was significantly decreased(P<0.05). Compared with the modelgroup, the AT2 R of treated groups were significantly increased(P>0.05). Nosignificant difference between I group and ZY group(P>0.05). 8 AT2 Rm RNA expressed in renal tissue by RT-PCRCompared with the normal group, the AT2 R of model group andtreatment groups were significantly decreased(P<0.05). Compared with themodel group, the AT2 R of treatment groups were significantly increased(P>0.05). No significant difference between I group and ZY group(P>0.05).Conclusions:1 The huoxue huayu tongluo medicine can reduce the diabetic rats of urinary protein.2 The huoxue huayu tongluo medicine can extend PT.3 The huoxue huayu tongluo medicine can lower level of triglyceride.4 The huoxue huayu tongluo medicine can lower level of BUN.5 The huoxue huayu tongluo medicine can enhance the expression of AT2 R in renal tissues.