Studies On The Activety And Mechanism Of Pueraria-coptis And Its Active Components Against Type 2 Diabetic Nephropathy

Diabetic nephropathy refers to microvascular complications and end-stage renal disease in diabetic patients.It is one of the most common and serious complications of diabetes,accounting for about half of diabetic patients,and greatly increases the mortality rate of diabetic patients.The pathogenesis of diabetic nephropathy is very complex,and many mediators are involved in the development of diabetic nephropathy.In order to control the progression of the disease and improve the quality of life of patients,it is urgent to explore the pathogenesis of diabetic nephropathy and develop new treatment strategies.Gegen Qinlian Decoction is one of the classic ancient recipes of traditional Chinese medicine.It comes from the Taiyang disease chapter of "Treatise on Febrile Diseases" written by Zhang zhongjing in the late Eastern Han Dynasty.The theory of compatibility of traditional Chinese medicines is an important means of researching prescriptions,and the research of disassembling prescriptions is an effective way to study the principle for prescriptions compatibility.The dismantling recipes are relatively simple in composition,rigorous in regulations,and clear in compatibility and effectiveness,which can often grasp the key to the disease.Gegen Qinlian Decoction is a combination of Gegen（root of Radix puerariae）and Huanglian（Coptis chinensis）which has been proven to improve diabetes-related biochemical indicators,but there is no relevant research on its complications,diabetic nephropathy.In this study,model rats of 2 type diabetes was established by the feeding of high sugar and fat food and the damage of islet cells with the injection of streptozotocin.Then,by prolonging the course of disease,the diabetic nephropathy was promoted and model rats of diabetic nephropathy were obtained.Diabetic nephropathy model rats were randomly divided into different treating groups.After 6 weeks of administration of the Pueraria-Coptis drug pair and active ingredients,blood glucose,blood lipids,renal function-related indicators and renal pathological changes were detected to explore the in vivo effects of the above drug pairs and active ingredients.Then,the network pharmacology method was combined with the non-targeted metabolomics method to systematically study the mechanism of Pueraria-Coptis medicine pair and its active ingredients in the treatment of diabetic nephropathy.Firstly,through data mining of literature and databases,the targets of traditional Chinese medicine,disease targets and their intersection targets were obtained.Secondly,the protein-protein interaction relationship between the intersecting targets was analyzed and the Pueraria-Coptis drug pair-intersection target-pathway-diabetic nephropathy interaction network diagram was established to explore the potential pharmacodynamic material basis and key targets of the drug pair.Next,based on HPLC-QTOF-MS/MS,non-targeted metabolomic studies were performed on the serum of SD rats in the control group,model group and administration group.The raw data obtained from the HPLC-QTOF-MS/MS were subjected to format conversion,peak identification,deconvolution,peak alignment,normalization,standardization and other preprocessing,and then perform unit statistical analysis and multivariate statistical analysis.Differential metabolites were screened by P value,FC value and VIP value.The MS fragmentation information was combined with databases such as HMDB and Massbank to identify differential metabolites.Finally,the differential metabolites obtained by metabolomics and the key targets obtained by network pharmacology were combined for pathway analysis,and the mechanism of action of the Pueraria-Coptis drug pair and its active ingredients in the treatment of diabetic nephropathy was discussed systematically.The following results were obtained through the study:（1）Compared with the prescription Gegen Qinlian Decoction,the disassembled Gegen-Huanglian medicine can more effectively reduce fasting blood sugar,improve blood lipid-related indexes（TG,HDL-C,CHO）,and improve kidney-related indexes.（CREA,BUN）.Among them,the middle-dose group had the best effect.Berberamine is an active ingredient in the Pueraria-Coptis drug pair.The pharmacological activity study found that berberamine has good anti-diabetic nephropathy activity.（2）Through network pharmacology research,it is speculated that magnolia,quercetin,ferulic acid,berberamine,genistein and other components may be the potential pharmacodynamic material basis for the therapeutic effects of Pueraria-Coptis medicine to diabetic nephropathy.（3）Through non-targeted metabolomics,it was found that the Pueraria-Coptis drug pair can significantly revert the up/down changes of 16 metabolites,such as 2-ketoglutarate,N,N,N-Trimethyl-L-alanyl-L-proline betaine,3-hydroxyisovaleric acid,4-ene-valproic acid,O-methoxycatechol-O-sulfate/4-methoxyphenyl sulfate,p-cresol sulfate,p-cresol glucuronide,L-acetylcarnitine,phenylacetylglycine,taurocholic acid,taurochenodeoxycholic acid,Lyso PC（16:1（9Z）/0:0）,Lyso PC（17:0/0:0）and argininosuccinic acid.Berbamine could significantly callback the changes of 13 metabolites,such as Lyso PA（18:2（9Z,12Z）/0:0）,3-hydroxyisovaleric acid,2-propylglutaric acid,MG（0:0/16:1（9Z）/0:0）,taurochenodeoxycholic acid,azelaic acid,2-methylbutyrylcarnitine,butyrylcarnitine,L-carnitine and hippuric acid.（4）By the combined analysis of network pharmacology and metabolomics,it is speculated that the AGE-RAGE signaling pathway and the arginine and proline metabolic pathways related to diabetic complications may be the key to the treatment of diabetic nephropathy by Pueraria-Coptis and its active ingredient.The small molecule metabolite-reaction-enzyme-gene network relationship,which further speculated that the Pueraria-Coptis drug pair might change NOS1,NOS2 A,NOS3,ARG1,ALDH2 and AOC3,which in turn affect arginine-related metabolic pathways,thereby regulating NO metabolism disorders and improving renal hemodynamics and inflammatory response to play a role in the treatment of diabetic nephropathy.It is extended that the active components in the Pueraria-Coptis drug pair may play therapeutic roles through taurine and hypotaurine metabolism,glycerolipid metabolism,lipid and atherosclerosis,PI3K-Akt signaling pathway and other signaling pathways.