| Objective: In this study,the network pharmacology method was used to analyze and predict the effective active ingredients,core targets and key signaling pathways of Sini Powder in the treatment of Acute myeloid leukemia(AML),and combined with network pharmacology results,the mechanism of action of Sini Powder against AML was verified and analyzed by in vitro cell experiments.to reveal the molecular mechanism of Sini powder in treating AML,aiming to provide experimental basis for the clinical application of Sini powder.Methods:1.Network pharmacology analysis and prediction: The active ingredients of the four Chinese herbs in Sini Power were screened through the TCMSP database,and then the Swiss Target Prediction database was used to obtain information on the targets relative to the ingredients,while "Acute myeloid leukemia" was used as a keyword in the The targets related to AML disease were retrieved from Gene Cards and OMIM databases,and the common targets of Sini Power and AML were obtained by online Venn diagram,and the component-target regulatory network of Sini Power in AML was established and visualized by cytoscape 3.7.2 software.In order to further obtain the key targets of Tetraconjugate in AML,the same targets of both were imported into String database to build the protein interaction network,and then the saved TSV files were uploaded to cytoscape software for analysis and screening,and the shared targets were uploaded to DAVID database for GO and KEGG enrichment analysis,and the top results were screened according to the P-value through the microbiology platform.The results were visualized and displayed in bar and bubble plots to identify the key signaling pathways for the treatment of AML with Sini Power.2.Cell experiments to study the effect of Sini Powder on AML: by preparing Sini Powder liquid medicine,SD rats were divided into groups and intragastrically administered for 7 consecutive days,and different doses of Sini Powder containing medicine were extracted after blood was collected from the abdominal aorta Serum,use Sini Powder containing drug serum to intervene KG1 a cells,detect cell viability level by CCK-8,calculate proliferation inhibition rate,flow cytometry was used to analyze the effect of different doses of Sini Powercontaining serum on apoptosis of KG1 a cells detect Sin by Western blot Effects of powdered drug-containing serum on protein expression of PI3 K,AKT,and m TOR in KG1 a cells.In order to verify the accuracy of network pharmacology enrichment analysis prediction.Results:1.Pharmacological results of online drugs: A total of 71 active ingredients and 288 corresponding targets of Sini Power were obtained through network pharmacological analysis,984 AML disease-related targets were collected,and the overlapping targets of Sini Power and AML were obtained using online Venn diagrams,resulting in a total of61 potential targets of Sini Power acting on AML.A PPI network screening of potential targets was constructed to obtain 15 core targets,namely AKT1,HRAS,STAT3,EGFR,HSP90AA1,SRC,MTOR,ESR1,ERBB2,PIK3 CA,MAPK1,MCL1,KIT,MAPK14,IL2.GO and KEGG enrichment analysis obtained 419 GO enrichment analysis items and 137 signaling pathways.The analysis results indicated that Sinisan can regulate protein phosphorylation,cell Proliferation,cell apoptosis,protein autophosphorylation,tyrosine phosphorylation and other biological processes affect molecular functions such as ATP binding,same protein binding,protein kinase activity,protein serine/threonine kinase activity,enzyme binding,etc,so as to achieve therapeutic The role of AML.According to the KEGG pathway enrichment analysis results,the PI3K/AKT pathway has the most enriched targets with the highest significance.Therefore,this study selected the PI3K/Akt pathway for subsequent cell experiment verification.2.Results of in vitro cell experiments:(1)Cell proliferation inhibition rate: compared with the blank serum control group,the cell activity of low,medium and high dose groups of Sini powder decreased significantly,and the inhibition rate of each group increased with the action time and drug dose,and the inhibition rate of each group reached the highest at 72 hours(P < 0.01).(2)Flow cytometry detection of cell apoptosis rate: Compared with the blank serum control group,the low-,medium-,and high-dose groups of Sini Powder could increase the apoptosis rate of KG1 a cells,which was closely related to the dose,and the apoptosis rates were 13.037± 0.375%,18.670±0.507%,37.933±1.305%(P<0.05,P<0.01).(3)Western blot detection of protein expression: Compared with the blank serum control group,the Sini Powder administration group could significantly inhibit the expression of PI3 K,Akt,and m TOR proteins(P<0.01).Conclusion:1.Sini Powder achieves the purpose of treating AML through multiple channels and multiple targets.The core targets of Sini Powder on AML are AKT1,HRAS,STAT3,EGFR,HSP90AA1,and SRC,and the key pathway is the PI3K/AKT signaling pathway.2.Sini powder can inhibit the proliferation of KG1 a cells and induce apoptosis of KG1 a cells,which is related to time and dose.3.Sini Powder can down-regulate the relative expression of PI3 K,Akt and m TOR proteins,and it is speculated that the mechanism of action of Sini Powder in treating AML may be related to the regulation of PI3K/Akt signaling pathway. |
