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Xin Wang,1,2,* Jie Zhao,1,2,* Rui Zhang,1,* Xinlu Liu,1 Chuanjiang Ma,2 Guangshang Cao,1,2 Yongli Wei,2 Peimin Yang2 1School of Pharmacy, Shandong University of Traditional Chinese Medicine (TCM), Jinan, 250355, Peopleâs Republic of China; 2Grade Three Laboratory of TCM Preparation of National Administration of TCM, Affiliated Hospital of Shandong University of TCM, Jinan, 250014, Peopleâs Republic of China*These authors contributed equally to this workCorrespondence: Peimin Yang, Tel +86-0531-68616607, Email jnypm@hotmail.comObjective: This study aims to investigate the hepatoprotective effect and molecular mechanism of Hedyotis diffusa Willd. ethanol extract (HDWE) against isoniazid (INH)-induced liver injury in the zebrafish model.Methods: INH-induced liver injury model was established by adding an immersion bath of INH in 3 days post-fertilisation (dpf) healthy transgenic zebrafish with liver-specific fluorescence (L-FABP: EGFP). HDWE and INH were given to the zebrafish to observe liver morphology and pathology, fluorescence intensity, and the activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and superoxide dismutase (SOD), as well as the content of glutathione (GSH). The chemical composition of HDWE was analysed using high-performance liquid chromatography coupled with a quadrupole-time-of flight hybrid mass spectrometer (HPLC-Q-TOF-MS). The bioactive compounds, molecular targets and signalling pathways of HDWE were predicted using network pharmacology. Subsequently, molecular docking was adopted to analyze the affinities between the bioactive components and targets by Autodock. Finally, in vitro experiments were conducted to further verify the findings.Results: Our findings showed that HDWE had a remarkable protective effect on INH-induced liver injury in zebrafish. Twenty compounds in HDWE were identified. Nineteen hub targets were identified as possible targets of HDWE, and a compound-target-pathway network was constructed. Nine bioactive compounds, ten molecular targets, and seven key signalling pathways were found to play a pivotal role in the hepatoprotective effect of HDWE against INH-induced liver injury. In vitro studies revealed that the important bioactive compound quercetin-3-O-sambubioside (QSA) could significantly reverse INH-induced cell viability decreases and had a significant effect on the associated targets predicted by network pharmacology and molecular docking.Conclusion: In this study, through the research of hepatoprotective effect of HDWE and bioinformatics analysis, the bioactive compounds, important pathways and key molecular targets were discovered. These findings could provide scientific evidence for the use of HDW in liver injury and prove to help explore its efficacy and the mechanism of action.Keywords: Hedyotis diffusa Willd., hepatoprotective activity, isoniazid-induced liver injury, zebrafish, network pharmacology, molecular docking |
