Mechanism of Keke tablets in treating post-infectious cough following influenza A virus infection based on network pharmacology, molecular docking, molecular dynamics and in vivo experiments

Abstract

Purpose: To dissect the therapeutic effect and potential pharmacological mechanisms of Keke tablets (KKT) against post-infectious cough (PIC) following influenza A virus infection based on network pharmacology, molecular docking, molecular dynamics, and in vivo experiments.

Methods: The chemical profiling of KKT was systematically analyzed using ultra-high performance liquid chromatography coupled with mass spectrometry (UPLC-MS). Active ingredients and targets of KKT were then screened through the TCMSP database, while disease-related targets of PIC were collected through the GeneCards database. Common core targets were obtained by drawing a Venn diagram, which were imported to the STRING database for network topology analysis. In addition, protein-protein interaction (PPI) network, Gene Ontology (GO) for core targets, functional analysis and pathway enrichment analysis from Kyoto Encyclopedia of Genes and Genomes (KEGG) were constructed to predict the network of KKT core active ingredients and PIC core targets. Molecular docking and dynamics analysis were performed to evaluate the interactions and movements of the components of KKT with the potential targets of PIC. A PIC model of guinea pigs was established by intranasally administering A/Puerto Rico/8/1934 H1N1/34 (A/PR/8) to verify the efficacy and potential pharmacological mechanism of KKT in treating PIC.

Results: A total of 29 potential active ingredients and 602 related targets in KKT were screened following UPLC-MS analysis. After intersection with 718 potential targets in PIC, 10 core targets were identified through PPI analysis. GO and KEGG analyses indicated that KKT were mainly related to the inflammatory response, NOD-like receptor signaling pathway and Toll-like receptor signaling pathway in the treatment of PIC. Subsequently, the findings of network pharmacology were corroborated by molecular docking and molecular dynamics analysis, suggesting that the main active components in KKT may exert therapeutic effects on PIC by modulating the TLR4/NF-κB/NLRP3 signaling. In vivo experiments demonstrated that KKT reduced cough frequency of A/PR/8-challenged guinea pigs and inhibited the activation of TLR4/NF-κB/NLRP3 signaling pathway in lung tissues, implying that KKT could alleviate PIC by regulating TLR4/ NF-κB/NLRP3 signaling pathway and suppressing pyroptosis.

Conclusion: KKT could inhibit the activation of TLR4/NF-κB/NLRP3 signaling pathway to suppress the assembly of NLRP3 inflammasome and the secretion of cytokines and cough-related neuropeptides in treating influenza A virus-induced PIC.

Keywords: In vivo experimental verification; Keke tablets; Molecular docking; Molecular dynamics; NLRP3 Inflammasome; Network pharmacology; Post-infectious cough.