| Firstly,literature research show that the research progress about pathogenesis,diagnostic criteria and treatment of chronic kidney disease,providing reference for the deep understanding and clinical treatment of chronic kidney disease.What's more,the main active ingredients of Salvia miltiorrhiza were showed.The pharmacological effects and mechanisms of Salvia miltiorrhiza on chronic kidney disease and its clinical application for the disease were summarized,providing reference for the comprehensive utilization and development of Salvia miltiorrhiza and for the further study on the mechanism on chronic kidney disease.Secondly,Extraction and preparation of Salvia miltiorrhiza stem-leaf and Salvia miltiorrhiza extractsThe extraction method including reflux,cold dip and ultrasonic and extraction solvent were investigated.The results showed that the contents of total salvianolic acid,total tanshinone and total flavonoids were the highest using reflux extraction method,and total tanshinone and total flavonoids were the highest when being extracted by 80%ethanol.The content of total phenolic acid in the water extract of Salvia miltiorrhiza was 16.74%.The content of total phenolic acid in the water extract of Salvia miltiorrhiza stem-leaf was 15.26%,and the total content of rutin and isoquercitrin was 0.41%.The content of total phenolic acid in 80%ethanol extract of Salvia miltiorrhiza was 11.62%and the content of total tanshinone was 3.79%.The total phenolic acid content in the extract of Salvia miltiorrhiza stem-leaf was 11.21%,and the total content of rutin and isoquercitrin was 1.21%.Thirdly,the study about the effect of Salvia miltiorrhiza stem-leaf on diabetic nephropathy rats(a),diabetic rats' model was established by feeding high glucose and high fat diet and 5%glucose water combined with intraperitoneal injection of 30 mg/kg STZ.Then the rats were treated with extract of Salvia miltiorrhiza and stem-leaf of Salvia miltiorrhiza.Serum,urine and kidney were collected to measure biochemical indcators.Western blotting was used to analyze the expression of related proteins in renal tissue.The results showed that the fasting blood glucose of the model group was 29 mmol·L-1,and the obvious proteinuria and renal tubule injury were observed.The fasting blood glucose of administration group rats decreased significantly,the pathological structure of the renal tissue restored to the normal-level.Salvia miltiorrhiza and Salvia miltiorrhiza stem-leaf can inhibite the expression of wnt4,?-catenin,TGF-? in renal tissue and prevent the occurrence of diabetic nephropathy.(b),the metabolic profile of serum,urine and kidney tissue in diabetic nephropathy rats was analyzed by using UPLC-QTOF/MS technique and multivariate statistical analysis.The biological effects and mechanism of the extract of Salvia miltiorrhiza were studied by metabolomics analysis research.Twenty one endogenous markers in serum,sixteen in urine and twenty two in kidney tissues were found,respectively,which involved in phospholipid metabolism,arachidonic acid metabolism,pyrimidine metabolic disorders.Salvia miltiorrhiza and Salvia miltiorrhiza stem-leaf could regulate these disorders.(c),16S rRNA technique was used to study the diversity of fecal flora in normal rats,diabetic nephropathy rats and treated rats.LEfSe analysis was applied to search the potential markers,and the biological function of intestinal flora was predicted by PICRUSt software.The results showed that the richness and diversity of intestinal flora in the feces of diabetic nephropathy were lower than those in the normal group,and that in salvia miltiorrhiza-treated rats was close to the normal group.Potential biomarkers found in the genus-level and family-level include Bifidobacterium,Turicibacter,Peptostreptococcaceae,Desulfovibrio,SMB53,Lactobacillus,Clostridium,Rikenellaceae,Ruminococcaceae.There are 43 metabolic pathways involved in intestinal flora in feces,mainly including membrane transport,amino acid metabolism,nucleotide metabolism,metabolism of coenzymes and vitamins,carbohydrate metabolism,energy metabolism,gene replication and repair,gene transcription and Translation and so on.(d),the rat of glomerular mesangial cell injury model induced by high glucose was established.The effect of extract of Salvia miltiorrhiza and Salvia miltiorrhiza stem and leaf,salvianolic acid B and tanshinone ?A were investigated.The expressions of Wnt-4,?-catenin and TGF-? were determined by Weatern blotting,and Salvia miltiorrhiza stem-leaf and Salvia miltiorrhiza can significantly regulate the expressions of the protein.Fourthly,the study about renal protection effects of Salvia miltiorrhiza stem-leaf extracts on chronic renal failure rats(a),the effects of different doses of Salvia miltiorrhiza extracts on adenine-induced chronic renal failure were studied.It was found that Salvia miltiorrhiza extract could regulate the content of serum creatinine,blood urea nitrogen,and urine protein and regulate the expression of related proteins in renal tissue in order to improve its renal function.(b),UPLC-QTOF/MS technique combined with multivariate statistical analysis was applied for analyzing the metabolic profile of serum,urine and kidney tissues in normal group,model rats and treatment rats,which was used to evaluate the biological effects and mechanism of Salvia miltiorrhiza.The results showed that normal group and treatment group were distinguished from model group in the PLS-DA map.13,27,19 endogenous biomarkers were identified in serum,urine and kidney,respectively.The metabolic pathways involved in these markers include tryptophan metabolism,purine metabolism,pyrimidine metabolism and phenylalanine metabolism.These metabolic pathways in chronic renal failure rats were disordered,which were regulated after administration.(c),16S rRNA technique was applied to study the diversity of fecal flora in adenine-induced chronic renal failure rats.Multiple potential microbial markers were found in genus and family level by LEfSe analysis,including Mucispirillum,Kurthia,Clostridium,Blautia,Butyrivibrio,Shuttleworthia,Peptococcus,Ruminococcus,Bradyrhizobium,Methylobacterium,Azospirillum,Thalassospira,Methylophilus,Pseudomonas,Peptostreptococcaceae and S24-7.Salvia miltiorrhiza water extract and ethanol extract can improve the diversity of fecal flora in adenine-induced chronic renal failure rats.Salvia miltiorrhiza ethanol extract could significantly regulate the relative abundance of Clostridium,Pseudomonas and Peptostreptococcaceae,and Salvia miltiorrhiza water extract could significantly regulate relative abundance of Peptococcus,Ruminococcus and Peptostreaceococcaceae.(d),after being cultured with the intestinal flora from feces of normal rats and chronic kidney failure rats,the major metabolites of active ingredients in Salvia miltiorrhiza were identified by UPLC-QTOF/MS technique.The metabolic pathways of tanshinone IIA,cryptotanshinone,tanshinone ? and dihydrotanshinone ? contains Methylation,demethylation,oxidation,hydrogenation and hydroxylation.Four tanshinones can be mutually transformed in the role of intestinal flora.The metabolic pathways of salvianolic acid B,salvianolic acid C and rosmarinic acid mainly include glucuronidation,methylation and hydrogenation.(e),indoxyl sulfate with the same concentration as its blood concentration in chronic renal failure rats was slected to induce renal tubular epithelial cells(HK-2)injury.Salvia miltiorrhiza water and ethnol extract were used to treat the cells.Weatem blotting was applied for analyzing the expression of ?-SMA,Fibronectin and p-ERK.The results showed that Salvia miltiorrhiza water and ethnol extract can significantly regulate the expressions of?-SMA,Fibronectin and p-ERK,NOX1,NOX2,NOX4,TGF-?,TGF-?R?,TGF-?R?,Smad2 and Smad3,and in further to regulate the NADPH/ROS/ERK and TGF-?/smad signaling pathways in HK-2 cells.Fivethly,the study on the action mechanisms of Salvia miltiorrhiza multicomponent for the treatment of chronic kidney disease based on the network pharmacology showed that ten main compounds in Salvia miltiorrhiza were slected as research object and Network pharmacology research methods was used to study the correlation among the compounds,multi-target and signal pathway,providing a basis for further clarifying the molecular mechanism of salvia miltiorrhiza in treating chronic kidney disease.As the network pharmacological analysis showed,main salvianolic acid and tanshinones in salvia miltiorrhiza related to renal disease signaling pathways includ purine metabolism,Wnt signaling pathway and so on. |
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