Study On The Mechanism Of Action Of Volatile Components Of Ginseng In The Treatment Of Gastric Cancer Based On Network Pharmacology

Objective To investigate the chemical constituents and functional analysis of ginseng volatile components,and to analyze the molecular mechanism and key targets of ginseng volatile components in gastric cancer by network pharmacology combined experiment.Methods 1.Solvent-extraction method was used to extract volatile components of ginseng,and Gas chromatography-mass spectrometry(GC-MS)combined with TCMSP analysis platform was used to analyze and screen the specific components of volatile components of ginseng.Swisstargetprediction tool and Batman-TCM tool were used to predict the target of volatile components of ginseng.Network pharmacology was used to analyze the functions,signaling pathways and related diseases of potential targets of ginseng volatile components.2.Network pharmacology combined with bioinformatics methods to analyze the specific mechanism and key targets of ginseng volatile components in gastric cancer intervention,and screen TOP4 core targets;R studio tools were used to analyze the expression of TOP4 core targets in gastric cancer and to conduct survival analy sis.Fifteen cases of cancer and paracancer tissues from patients with clinically confirmed gastric cancer were collected,and the expressions of TOP4 core targets in gastric cancer and paracancer tissues were detected by immunohistochemistry.The SwissDock platform was used for quasi-molecular docking to analyze the binding ability of volatile components of ginseng to core targets.3.Human gastric cancer cell lines(MKN-45,HGC-27)were cultured in vitro and treated with different doses of ginseng volatile components(0μg/mL,50μg/mL,100μg/mL,150μg/mL,200μg/mL and 250μg/mL)for 24 h,48 h,72 h,respectively.CCK-8 assay was used to detect the toxicity of ginseng active ingredients to gastric cancer cells.MKN-45 and HGC-27 cells were treated with low and high doses of appropriate drug concentration and blank control.Annexin V-FITC/PI apoptosis detection kit was used to detect the apoptosis of the three groups.Western blot and RT-PCR was used to detect the protein expression and mRNA of EGFR,MAPK1,MAPK3 and IL-6 in the above three groups.Results 1.Ten active components of ginseng volatile components were identified:Methyl octadeca-8,11-dienoate、oleic acid、Linoleic、panaxydol、Phenylglucoside、(1 aR,4aR,7S,7aR,7bR)-1,1,7-Trimethyl-4-methylenedecahydro-1H-cycl opropaeazulen-7-ol、calarene、(S,Z)-Heptadeca-1,9-dien-4,6-diyn-3-ol、zoomaric acid、(+)-Ledene.529 target genes were screened,and the GO and KEGG enrichment of these targets involved in lipid binding,oxidoreductase activity,regulation of ion migration,regulation of transient receptor potential(TRP)channels by inflammatory mediators,and pathways in cancer.2.A total of 751 genes were obtained from gastric cancer target screening.Venn diagram was used to analyze the potential targets of ginseng volatile components and the above-mentioned gastric cancer targets,and 67 overlapping genes were obtained,which were used as potential targets of ginseng volatile components to intervene in gastric cancer for subsequent analysis.The GO function of the above genes mainly involves the activity of nuclear receptors,kinase activity involved in the positive regulation of cell migration and MAPK cascade regulation,and the signaling pathways involved in cancer pathways,proteoglycans in cancer,and endocrine resistance.Further screening TOP4 core target were EGFR,MAPK1,MAPK3 and IL-6.Bioinformatics analysis showed that MAPK1 were highly expressed in gastric cancer tissues,which were significantly higher than those in the control group,and the difference was statistically significant.Immunohistochemical experiments showed that the positive expressions of EGFR,MAPK1,MAPK3 and IL-6 protein in gastric cancer tissues were significantly increased compared with that in adjacent tissues(P<0.05).EGFR,MAPK1,MAPK3,were significantly correlated with the survival of gastric cancer patients,and the differences were statistically significant.The results of molecular docking showed that the binding capacity of panaxynol,Methyl octadeca-8,11-dienoate,zoomaric acid,oleic acid with the above four targets was less than-5 KJ/mol,and all of them could form a good interaction.Compared with control group(0 μg/mL),ginseng volatile components at 50,100,150,200 and 250 μg/mL significantly inhibited the proliferation of MKN-45 and HGC-27 cells(P<0.05).The activity of MKN-45 and HGC-27 cells decreased with the concentration of ginseng volatile components and treating time increasing,and ginseng volatile components showed a dose-dependent and time-dependent inhibitory effect on the growth of cancer cells.Flow cytometry analysis showed that compared with the Control group(Control),the apoptosis rate of MKN-45 and HGC-27 cells was significantly promoted by both low dose and high dose of Ginseng volatile components(P<0.05),and the apoptosis rate of MKN-45 and HGC-27 cells treated by high dose of ginseng volatile ingredients was more than 2 times of that treated by low dose.Western blot and fluorescence quantitative PCR showed that ginseng volatile components significantly inhibited the expression of EGFR,MAPK1,MAPK3 and IL-6 protein and mRNA levels in 2 gastric cancer cell lines,and the protein and mRNA levels in the high-dose group were significantly lower than those in the low-dose group(P<0.05).Conclusion Ginseng volatile components have a certain anti-gastric cancer effect,and this effect may be realized through the regulation of EGFR,MAPK1,MAPK3 and IL-6 signal pathways,ginseng volatile components have the potential to treat gastric cancer.