Network pharmacology study to reveal active compounds of Qinggan Yin formula against pulmonary inflammation by inhibiting MAPK activation

Pneumonia is common and frequently-occurred disease caused by pathogens which predisposes to lung parenchymal inflammation leading pulmonary dysfunction. To prevent and alleviate the symptoms of pneumonia, Qinggan Yin formula (QGY) was composed based on clinical experience and four classical traditional Chinese medicine prescriptions which frequently applied to treat infectious diseases.Traditional Chinese medicine is a complex mixture and it is difficult to distinguish the effective component molecules. The aim of this study is to identify the compounds of QGY with anti-inflammatory effects and investigate the molecular mechanism.The high-resolution mass spectrometry and molecular networking were performed for comprehensive chemical profiling of QGY. Network pharmacology was used to generate "herbal-target-pathway" network for target predictions. The anti-inflammation effects of QGY were evaluated in mice model of lipopolysaccharide (LPS) induced acute inflammation. Tail transected zebrafish was also employed to validate macrophage migration reversed effect of QGY. Based on the molecular enrichment analysis, the active substances of QGY with anti-inflammatory effects were further identified in cellular model of macrophage activation. The mechanisms of active substances were investigated by testing their effects on the expression of correlated proteins by Western blot.In total, 71 compounds are identified as major substances of QGY. According to the results of network pharmacology, QGY shows moderate anti-inflammatory effects and inhibit pulmonary injury. MAPK signaling pathway was predicted as the most related pathway regulated by QGY. Moreover, QGY significantly inhibit LPS-induced pulmonary inflammation in mice, and reversed macrophage migration toward the injury site in zebrafish. We also validate that some major compounds in QGY significantly attenuated the release of IL-1β, IL-6 and TNF-α in LPS-stimulated macrophage. Those active substances including acacetin and arctiin can inhibit the phosphorylation of ERK/JNK and down-regulated the protein expression of BCL-2.Collectively, QGY possessed pronounced anti-inflammation effects. The integration of network pharmacology and experimental results indicated arctiin, iridin, acacetin, liquiritin, and arctigenin are major active substances of QGY with anti-inflammatory effects. The underlying mechanism of QGY involves MAPK signaling pathway and oxidative stress pathway.