AUTHOR=Zheng Weiwei , Shi Chao , Meng Yao , Peng Jian , Zhou Yongfei , Pan Tong , Ning Ke , Xie Qiuhong , Xiang Hongyu 

TITLE=Integrated network analysis and metabolomics reveal the molecular mechanism of Yinchen Sini decoction in CCl4-induced acute liver injury

JOURNAL=Frontiers in Pharmacology

VOLUME=14

YEAR=2023

URL=https://www.frontiersin.org/journals/pharmacology/articles/10.3389/fphar.2023.1221046

DOI=10.3389/fphar.2023.1221046

ISSN=1663-9812

ABSTRACT=<p><bold>Objective:</bold> Yinchen Sini decoction (YCSND), a traditional Chinese medicine (TCM) formula, plays a crucial role in the treatment of liver disease. However, the bioactive constituents and pharmacological mechanisms of action remain unclear. The present study aimed to reveal the molecular mechanism of YCSND in the treatment of acute liver injury (ALI) using integrated network analysis and metabolomics.</p><p><bold>Methods:</bold> Ultra-high-performance liquid chromatography coupled with Q-Exactive focus mass spectrum (UHPLC-QE-MS) was utilized to identify metabolites in YCSND, and high-performance liquid chromatography (HPLC) was applied to evaluate the quality of four botanical drugs in YCSND. Cell damage and ALI models in mice were established using CCl<sub>4</sub>. <sup>1</sup>H-NMR metabolomics approach, along with histopathological observation using hematoxylin and eosin (H&amp;E), biochemical measurements, and reverse transcription quantitative real-time PCR (RT-qPCR), was applied to evaluate the effect of YCSND on CCl<sub>4</sub>- induced ALI. Network analysis was conducted to predict the potential targets of YCSND in ALI.</p><p><bold>Result:</bold> Our results showed that 89 metabolites in YCSND were identified using UHPLC-QE-MS. YCSND protected against ALI by reducing the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and malondialdehyde (MDA) contents and increasing those of superoxide dismutase (SOD), and glutathione (GSH) both in vivo and in vitro. The <sup>1</sup>H-NMRmetabolic pattern revealed that YCSND reversed CCl<sub>4</sub>-induced metabolic abnormalities in the liver. Additionally, the Kyoto Encyclopedia of Genes and Genome (KEGG) pathway enrichment analysis identified five pathways related to liver injury, including the PI3K-AKT, MAPK, HIF-1, apoptosis, and TNF signaling pathways. Moreover, RT-qPCR showed YCSND regulated the inflammatory response (<italic>Tlr4</italic>, <italic>Il6</italic>, <italic>Tnfα</italic>, <italic>Nfκb1</italic>, <italic>Ptgs2</italic>, and <italic>Mmp9</italic>) and apoptosis (<italic>Bcl2</italic>, <italic>Caspase3</italic>, <italic>Bax</italic>, and <italic>Mapk3</italic>), and inhibited PI3K-AKT signaling pathway (<italic>Pi3k</italic> and <italic>Akt1</italic>). Combined network analysis and metabolomics showed a link between the key targets (<italic>Tlr4</italic>, <italic>Ptgs2</italic>, and <italic>Mmp9</italic>) and vital metabolites (choline, xanthine, lactate, and 3-hydroxybutyric acid) of YCSND in ALI.</p><p><bold>Conclusion:</bold> Overall, the results contribute to the understanding of the therapeutic effects of YCSND on ALI, and indicate that the integrated network analysis and metabolomics could be a powerful strategy to reveal the pharmacological effects of TCM.</p>