The TRPC Mechanism Of Diaphoresis And Consolidation Therapy For Proteinuria In Rats With PAN Nephropathy Based On The Xuanfu Theory

Objective:To study the therapeutic effects and molecular mechanisms of the methods of diaphoresis and consolidation in Traditional Chinese Medicine(TCM)for Puromycin aminonucleoside(PAN)nephropathy rats based on the similarity between Xuanfu and glomerular podocyte by targeting transient receptor potential channel 5(TRPC5)and TRPC6.The molecular basis of the treatment for nephropathy proteinuria was explored according to the Xuanfu theory.Methods:Eighty male wistar rats were randomized into four subgroups according to weights,consisting of model group,Yuebi decoction group,Gujing decoction group and control group.PAN nephropathy models were established by a single intravenous injection of PAN 90mg/kg for minimal change disease(MCD),and followed dosage of 9mg/kg at day 13,16,19,22,25 for focal segmental glomerulosclerosis(FSGS).The periods were 2 weeks and 12 weeks,respectively.The levels of serum creatinine,urea nitrogen,uric acid,albumin,total cholesterol,triglyceride and urinary protein creatinine ratio were detected.The expression and distribution of TPRC5 and TRPC6 were analyzed by westernblots,real-time PCR and immunofluorescence staining.The structural molecules of podocyte were detected by Western blots.The expression of F-actin was observed by confocal microscopy.The pathological changes of kidneys were observed by light microscopy after HE,Mssson and PAS staining.The foot processes effacements of podocytes were observed by electron microscopy.Results:1.Compared with the control group,hypoalbuminemia,proteinuria,hyperlipidemia,and ascites were showed in the model group of MCD.Hypoalbuminemia,proteinuria,and focal segmental glomerulosclerosis were observed in the model group of FSGS,the expressions of TRPC5,TRPC6,RHoA,and Rac1 increased(P<0.05),while Nephrin,Podocin,Synaptopodin and F-actin decreased(P<0.05,P<0.05),and extensive foot processes effacements of podocyte were observed in rats with MCD and FSGS.2.Compared with the model group of MCD,the total cholesterol,triglyceride and urinary protein creatinine ratio decreased in Yuebi decoction group and Gujing decoction group,albumin increased,and the expressions of TRPC5 and TRPC6 decreased(P<0.05).The levels of TRPC5 mRNA in Yuebi decoction group were lower than those in Gujing decoction group,while TRPC6 mRNA was higher(P<0.05).The expressions of Nephrin,Synaptopodin and F-actin were increased(P<0.05),foot processes effacements were alleviated in Yuebi decoction group and Gujing decoction group.However,the expressions of RHoA,Racl,Podocin had no significant differences between Yuebi decoction group,Gujing decoction group and MCD model group(P>0.05).3.Compared with the model group of FSGS,the urinary protein creatinine ratios were decreased in Gujing decoction group(P<0.05),urinary protein creatinine ratio also decreased in Yuebi decoction group,but had no significant differences comparing to model group(P>0.05).The serum albumin and the levels of Synaptopodin in renal tissue increased(P<0.05),the expressions of TRPC5,TRPC6,RHoA and Racl decreased in Gujing decoction group and Yuebi decoction group.The expressions of Nephrin and F-actin increased in Gujing decoction group(P<0.05).No significant differences of the expressions of Podocin were found between Gujing decoction group,Yuebi decoction group and FSGS model group(P>0.05),foot processes effacements were alleviated in both TCM groups.Conclusions:Yuebi decoction and Gujing decoction can decrease the proteinuria and increase serum albumin of rats with PAN nephropathy,the mechanism might be associated with regulating different subtypes of TRPC5 and TRPC6 and alleviating podocyte injury.The TRP channels might be an important regulatory factor of Xuanfu,and the targets of TCM methods of diaphoresis and consolidation.