Study and Verification of Anti-inflammatory Mechanism of Jinhua Qinggan Granules Based on Network Pharmacology
Objective To evaluate and explore the anti-inflammatory activity and mechanisms of Jinhua Qinggan Granules based on network pharmacology and in vitro cellular experiments,in order to provide theoretical basis for its clinical application.Methods TCMSP,Pubchem database and related literature were utilized to screen out the active ingredients and targets of Jinhua Qinggan Granules.The associated targets of influenza,headache,fever,and cough were screened through NCBI and GenCards databases,and comparative analyses with the above ingredient-targets were carried out in order to screen out the core targets.Cytoscape 3.7.2 software was used to generate the network diagram,and finally,GO and KEGG enrichment analyses of the key targets were conducted by Metascape database.In vitro cellular assays were performed using the lipopolysaccharide(LPS)-induced RAW264.7 inflammatory cell model.The effect of the extract of Jinhua Qinggan Granules on the secretion of nitric oxide(NO)was detected by Griess method,and the effects of the extract on the RNA and protein levels of key targets in RAW264.7 cells were determined by quantitative real-time polymerase chain reaction(qRT-PCR)and Western blotting,respectively.Results A total of 278 active ingredients and 1316 potential targets were identified,including 1065 disease-related targets,which were mainly related to the mitogen-activated protein kinase(MAPK)signaling pathway,the nuclear factor-κB(NF-κB)signaling pathway,and the neuroactive ligand-receptor interaction signaling pathway.The results of in vitro cellular experiments showed that compared with the LPS-stimulated group,the aqueous extract of Jinhua Qinggan Granules significantly reduced the secretion of NO in LPS-induced RAW264.7 cells at the concentration of 200,400 and 800 μg/ml(P<0.01).The qRT-PCR assay revealed that Jinhua Qinggan Granules extract could obviously inhibit the mRNA levels of inducible nitric oxide synthase(iNOS),cyclooxygenase-2(COX-2),and interleukin 6(IL-6)in LPS-induced RAW264.7 cells(P<0.01).Western blotting assay revealed that the extract of Jinhua Qinggan Granules could inhibit the expression of iNOS and the activation of key proteins in the NF-κB and MAPK signaling pathways.Conclusion Based on the network pharmacology study,Jinhua Qinggan Granules has the characteristics of multi-target and multi-pathway synergistic effect,and further in vitro cellular experiments show that Jinhua Qinggan Granules exert anti-inflammatory activity by inhibition of the expression of iNOS and the activation of NF-κB and MAPK signaling pathways to intervene in the release of inflammatory cytokines in the inflammatory cell model.The results provide a theoretical basis for the clinical application of Jinhua Qinggan Granules.
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