Study On The Effect And Mechanism Of Smilacis Glabrae Rhixoma And Pyrrosiae Folium Medicine On Kidney Senescence

Aim:In this study,the network pharmacology method was used to analyze the effective ingredients,potential targets and mechanisms of Smilacis Glabrae Rhixoma and Pyrrosiae Folium in the treatment of aging-related chronic kidney disease(ARCKD),and cell experiments were used to verify the core targets Point and mechanism,animal experiments verify the efficacy of the drug pair and its common component(quercetin),providing a theoretical basis for its clinical application.Methods:Screen the active ingredients of the drug pair with the aid of the TCM System Pharmacology Database and Analysis Platform(TCMSP),use the TCMSP,STITCH and SWISS databases to obtain the potential targets of the drug pair,and use the UniProt database to standardize the target protein.After screening disease targets using The Human Phenotype Ontology,DisGeNET and Therapeutic Target Database(TTD)databases,the potential targets of ARCKD can be obtained by intersecting with drug targets.Construct "drug-active ingredient-target" network through Cytoscape software,import potential targets into the STRING database and then construct a protein interaction(PPI)network for in-depth analysis.The gene ontology(GO)function enrichment and the Kyoto Encyclopedia of Gene and Genome(KEGG)pathway enrichment analysis were carried out through the DAVID platform.Aristolochic acid(AAI)was used to induce senescence and damage of rat proximal renal tubular epithelial cells(NRK-52E):MTT method was used to detect cell survival rate after AAI modeling and AAI+quercetin intervention;senescence-related β-half Lactosidase(SA-β-gal)staining method was used to detect the condition of senescent cells after AAI+quercetin intervention;Western blot was used to verify the core target p53 and cyclin-dependent kinase inhibitor 1A(p21)protein expression and fibrosis indicators α-smooth muscle agonist(α-SMA)and vimentin protein expression.Intraperitoneal injection of AAI in mice was used to induce renal aging and damage.The degree of renal damage was assessed by blood creatinine,urea nitrogen,uric acid and urine protein.The pathological changes and fibrosis of the kidney were observed by HE and Masson staining.Results:There are 17 active ingredients and 112 potential targets in the drug pair.PPI target screening analysis obtained 6 core targets,27 important targets,and 36 secondary targets,forming a complex network of effects centered on the tumor protein p53 gene(TP53).The GO enrichment function involves 532 items,which mainly affect the processes of aging,redox,inflammatory response,and positive regulation of cell proliferation;KEGG pathway enrichment obtains 137 items,involving such as P53,cancer,NF-κB and AMPK pathways.Cell experiment results confirmed that AAI can significantly reduce the survival rate of NRK-52E cells,induce positive staining of cell senescence,and up-regulate the expression of p53,p21,α-SMA,and Vimentin proteins.Quercetin can improve the above changes after intervention.The results of animal experiments showed that compared with the AAI group,the drug pair and quercetin can significantly improve the mice’s serum creatinine,urea nitrogen,uric acid,urine protein,and the degree of pathological damage and fibrosis.Conclusion:As the "switch" of the progression of ARCKD,the TP53 gene activates reactions such as inflammation,apoptosis,and oxidative stress,forms a complex system network imbalance,and promotes the development of fibrosis.The drug pair can interfere with ARCKD through multiple components,multiple pathways,and multiple targets.Quercetin may be the key effective component of its role,and the P53 signaling pathway may be its potential core pathway.