| 1.Objective: To investigate the difference of metabolite expression of Mesangial Proliferative Glomerulo-Nephritis(Ms PGN)between patients with spleen-kidney deficiency syndrome and normal people based on urinary metabolomics.To investigate its correlation with TCM syndrome score of spleen-kidney deficiency syndrome Ms PGN and 24 h urinary protein quantification.2.Methods: Mid-morning urine samples from 5 Ms PGN patients and 5 normal persons were collected and analyzed by Liquid Chromatography-Mass Spectrometry(LC-MS)to screen different metabolites and metabolic pathways.To analyze the correlation between different metabolites and TCM syndrome score of spleen-kidney deficiency syndrome Ms PGN and 24 h urinary protein quantitation.3.Results:3.1 32 different metabolites were screened(P < 0.05,VIP > 1),of which 12 were up-reg ulated and 20 down-regulated,mainly involving lipid metabolism,amino acid metabolis m,tricarboxylic acid cycle and other metabolic pathways.Upregulated metabolites inclu de Sulfate、Pyroglutamic acid、Niacinamide、Androsterone sulfate、(R)-beta-Amino isobutyric acid,etc.Down-regulated metabolites include Isocitric acid、 Tetrahydroaldo sterone-3-glucuronide、Citric acid,(Citric acid),11 b,17a,21-trihydroxypregnenolone(11b,17 a,21-Trihydroxypreg-nenolone),lysophosphatidylcholine(Lyso PC(22:1(13Z)/0:0)),phosphatidylacid(PA(0:0/16:0))Isocitric acid)、Tetrahydroaldosterone-3-glucuronid e、Citric acid、11b,17 a,21-Trihydroxypreg-nenolone、Lyso PC(22:1(13Z)/0:0)PA(0:0/16:0),etc.3.2 Ten metabolic pathways(P < 0.05)were screened,including choline metabolism,st eroid hormone biosynthesis,glycerol phospholipid metabolism,citric acid cycle,and int erconversion of pentose and glucuronic acid in cancer.The Choline metabolism in canc er pathway is significantly different(P < 0.01),and the metabolites involved are lysopch oline(Lyso PC(22:1(13Z)/0:0)and phosphatidic acid(PA(0:0/16:0)).3.3 Correlation analysis showed that Lyso PC(22:1(13Z)/0:0)expression level of Ms PGN patients was negatively correlated with TCM syndrome score of spleen-kidney deficiency syndrome,that is,the expression level decreased with the aggravation of clinical symptoms of spleen and kidney deficiency syndrome,while PA(0:0/16:0)expression level was not correlated with TCM syndrome score.Lyso PC(22:1(13Z)/0:0)and PA(0:0/16:0)were negatively correlated with the increase of 24 h urinary protein.4.Conclusion:4.1 There were differences in urine metabolism between Ms PGN patients with spleenkidney deficiency syndrome and healthy control group,mainly involving lipid metabolism,amino acid metabolism,tricarboxylic acid cycle and other metabolic pathways.4.2 Lyso PC(22:1(13Z)/0:0)expression in Ms PGN patients was negatively correlated wit h the clinical manifestations of spleen-kidney deficiency syndrome.Lyso PC(22:1(13Z)/0:0)and PA(0:0/16:0)expression levels were negatively correlated with 24 h urinary pro tein quantity.1.Objective: A rat model of Ms PGN was established,and the relevant mechanism of action of Shendi granule in the treatment of Ms PGN was discussed through the intervention of Shendi Granule,combined with network pharmacology and metabolomics.2.Methods: Thirty rats were randomly divided into normal group,model group and treatment group.The Ms PGN model was replicated in the model and treatment groups by injection of Thy-1 antibody.Seven days after antibody injection,a positive urine protein dipstick was considered a successful model.After 2 weeks of successful modeling,the rats were given Shendi granules(4g/kg·d)by gavage,once a day for 12 weeks.At the 0,4,8,and 12 weeks,the 24-hour urine was collected by metabolic cage,and the lower end of the urine collection tube was placed on the ice or ice bag.After 12 weeks of gavage,blood is taken from the abdominal aorta to measure serum creatinineand renal tissues were obtained for pathological observation.Urine samples were analyzed by liquid chromatography-mass spectrometry(UPLC-MS)to screen differential metabolites and metabolic pathways.To explore the mechanism of Shendi Granule in treating Ms PGN by metabonomics and network pharmacology.3.Results:3.1 24 h urinary protein: Compared with the normal group,the 24 h urinary protein in the model group and the treatment group was significantly increased(P <0.05);After 4weeks,8 weeks and 12 weeks of Shendi granule intervention,compared with the model group,the 24-hour urinary protein quantification in the treatment group was significantly decreased(P<0.05).3.2 Serum creatinine: After 12 weeks,there was no significant change in serum creatinine Scr from baseline in all groups(P > 0.05).3.3 Renal pathological results: The pathological results of HE and PAS suggested showed that the capillaries in the normal group opened well,and there was no mesangial cell proliferation and mesangial matrix increase.In the model group,diffuse mesangial cell proliferation and matrix increased significantly.Mesangial cells and mesangial matrix in the treatment group were significantly less than those in the model group.3.4 After metabolomics screening,a total of 4 differential metabolites were identified th at significantly changed after Shendi particles intervention.After Shendi granule interve ntion,Tetrahydrobiopterin,Thromboxane B2,Lyso PC(17:0/0:0),Lyso PC(18:0/0:0)were significantly up-regulated.3.5 According to the network pharmacological screening,there were 38 potential target s of Shendi granules in the treatment of Ms PGN,and their substance bases included kae mpferol,Myricanone,beta-sitosterol,etc.GO enrichment analysis showed that it mainly involved oxidative stress,cell proliferation and other biological processes.KEGG was enriched to 143 signal pathways,and the metabolomic-related pathways mainly involve d lipid and atherosclerosis signaling pathways,TNF signaling pathways,age-rage signa ling pathways and Choline metabolism in cancer pathways.PPI network was constructe d and gene targets were integrated.AKT1,CASP3,PTGS2,HMOX1 and ICAM1 were the core targets of Shendi granules in the treatment of Ms PGN.3.6 Comprehensive network pharmacology and metabolomics analysis,metabolomics fi nally identified 16 metabolic pathways,and network pharmacology enriched 143 signali ng pathways.Comprehensive network pharmacology and metabolomics analysis identif ied a common Choline metabolism pathway in cancer.The characteristic metabolites in volved were lysophosphatidylcholine Lyso PC(17:0/0:0)and Lyso PC(18:0/0:0).The me chanism by which SDC reduced mesangial cell proliferation may be through up-regulati on of Lyso PC(17:0/0:0)and Lyso PC(18:0/0:0)levels.And intervention of PI3K-AKT si gnaling pathway MAPK signaling pathway.4.Conclusion:4.1 Shendi granules can can reduce urine protein levels in rats and reduce renal pathological damage4.2 Shendi granules may interfere with biological processes such as inflammation,apoptosis and cell proliferation by up-regulating the expression of BH4,TXB2,Lyso PC(17:0/0:0),Lyso PC(18:0/0:0)and other differential metabolites,so as to improve the pathological damage of Ms PGN.4.3 Based on network pharmacology and metabolomics,Shendi granules may play a role in the treatment of Ms PGN by intervening in MAPK signaling pathway,thereby up-regulating Lyso PC(17:0/0:0)and Lyso PC(18:0/0:0),and reducing mesangal cell proliferation. |
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