Zaoshi Huatan Xiefei formula improved severe pneumonia by regulating MAPK/NF-κB/VCAM1 signaling pathway mediated leukocyte transendothelial migration.

BACKGROUND: Zaoshi Huatan Xiefei Formula (ZHXF) is a Chinese medicine prescription used in the treatment of severe pneumonia (SP). Although clinical studies have confirmed its efficacy in improving pneumonia, the specific bioactive components and their intricate regulatory pathwaysremain incompletely understood. PURPOSE: This research aims to identify the active components of ZHXF, along with its potential targets and molecular mechanisms for treating SP. METHODS: The main active ingredients of ZHXF and its blood chemical components were determined by UPLC-Q-Exactive Orbitrap MS. The core targets and biological immune regulatory mechanisms through which ZHXF improves SP were predicted using an integrated approach of network pharmacology, molecular docking, MD simulation, SPR, and proteomics. The protective effect of ZHXF on SP, along with its potential targets and mechanisms, was validated through experiments. RESULTS: 130 ingredients of ZHXF were identified by UPLC-Q-Exactive Orbitrap MS, five were confirmed as its primary bioactive ingredients in the blood. Network pharmacology analysis showed that modulating the MAPK and NF-κB pathways is a key mechanism through which ZHXF improves SP. Molecular docking, MD simulations, and SPR indicated that the key components of ZHXF bind effectively to ERK1/2, JNK, and P38, thereby suppressing MAPK/NF-κB pathway expression. Proteomic analysis further revealed VCAM1 as the core target of ZHXF, indicating that ZHXF alleviates the impact of lung inflammation by regulating leukocyte transendothelial migration. In vitro study have confirmed the ZHXF inhibitory effect on leukocyte transendothelial migration. In vivo experiments have shown that ZHXF can significantly alleviate pulmonary inflammation symptoms in SP mice, and this protective effect on lung tissue was achieved through the inhibition of leukocyte transendothelial migration. CONCLUSION: This study confirms that ZHXF improves lung inflammation by suppressing the activation of the MAPK/NF-κB pathway and regulating VCAM1 mediated leukocyte transendothelial migration. This study provides a novel perspective for the treatment of SP with ZHXF.