Compatibility Mechanisms of Liangge San (凉膈散) and Its Qingshang (清上) or Xiexia (泄下) Decomposed Prescription Based on Its Anti-Inflammatory Effects
Objective:To investigate the anti-inflammatory effects and compatibility mechanisms of Liangge San (凉膈散) and its Qingshang (清上) or Xiexia (泄下) decomposed prescription based on network pharmacology and zebrafish inflammation model.Methods:Three zebrafish inflammation models,survival analysis,and histopathological section experiment were used to verify the anti-inflammatory effects of Liangge San and its Qingshang or Xiexia decomposed prescription.The TCMSP database was used to screen out the effective chemical constituents and corresponding targets of Liangge San and its Qingshang or Xiexia decomposed prescription.Inflammation-related targets were searched in da-tabases such as GeneCards and DisGeNET,and the intersection of the effective chemical constituent targets of Liangge San and its Qingshang or Xiexia decomposed prescription and inflammation targets was selected.The STRING database was used to construct a protein-protein inter-action (PPI) network for the intersecting targets.Gene ontology (GO) enrichment analysis and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis were performed on potential targets using the David database.Real-time fluorescence quantification polymerase chain reaction (PCR) was used to detect the gene expression levels of key targets.Results:The experimental results of the ze-brafish inflammation model showed that compared with the normal control group,neutrophils of the zebrafish migrated to the injured site signif-icantly,the survival time was shortened after microinjection of LPS into the yolk sac,and the mortality rate was significantly increased in the model group (P<0.01).Compared with the model group,dexamethasone and different concentrations of Liangge San and its Qingshang or Xiexia decomposed prescription could significantly inhibit the inflammatory response,reduce the migration of neutrophils,prolong the survival time of zebrafish after microinjection of LPS into yolk sac,and improve the survival rate (P<0.05 or P<0.01).Compared with the Liangge San prescription,the inhibitory effect of its Qingshang or Xiexia decomposed prescription on neutrophil migration was weakened (P<0.05 or P<0.01),and both decomposed prescriptions showed synergistic effects (CI<1).The protective effect of Qingshang or Xiexia decomposed pre-scription on survival of zebrafish yolk sac injected with LPS was weakened (P<0.01),while Liangge San had the best effect.The results of network pharmacological analysis showed that there were 176 effective chemical constituents in Liangge San,such as phillyrin,quercetin,kaempferol,and luteolin,corresponding to 266 targets.A total of 74 effective chemical constituents in Qingshang prescription were detected,such as geniposide,quercetin,wogonin,and locution,corresponding to 233 targets.There were also 109 effective chemical constituents in Xiexia prescription,such as emodin,rhein,and sitosterol,corresponding to 234 targets.A total of 10933 inflammatory disease-related targets and 168 intersecting targets were found.Through the PPI network diagram,core targets were identified,such as protein kinase B (AKT1),tumor protein p53 (TP53),tumor necrosis factor (TNF),interleukin-6 (IL-6),and vascular endothelial growth factor (VEGF).GO func-tional analysis suggested that the active ingredients in Liangge San and its Qingshang or Xiexia decomposed prescription were closely related to biological processes such as positive regulation of RNA polymerase Ⅱ transcription and regulation of gene expression and drug response,cell composition such as plasma membrane,extracellular region,and cytosol,and molecular functions such as protein binding and enzyme binding.KEGG pathway enrichment analysis revealed that Liangge San and its Qingshang or Xiexia decomposed prescription showed anti-inflammatory effects through the cancer pathway,AGE-RAGE pathway,lipids and atherosclerosis,fluid shear stress and atherosclerosis,chemical carcino-gene-receptor activation,and other pathways.The results of real-time quantitative PCR showed that compared with the normal control group,the mRNA expression of inflammatory factors Tnfα and Il6 in the model group was significantly up-regulated (P<0.01),and the mRNA ex-pression of Akt was significantly down-regulated (P<0.01).Compared with the model group,Liangge San and its Qingshang or Xiexia decom-posed prescription could significantly reduce the mRNA expression of Tnfα and Il6 (P<0.01),while Liangge San significantly up-regulated the mRNA expression of Akt (P<0.05).Conclusion:Liangge San and its Qingshang or Xiexia decomposed prescription have significant anti-inflammatory effects,and both decomposed prescriptions show a synergistic effect.The mechanism may be related to the key targets TNF,IL-6,and AKT.
Liangge San (凉膈散)InflammationTherapeutic methodNetwork pharmacologyZebrafish
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