Kehuang capsule inhibits MAPK and AKT signaling pathways to mitigate CCl₄-induced acute liver injury

Abstract

Background and aims: Kehuang (KH) capsule is an herbal medical product approved for the treatment of liver diseases, including liver injury, in China. However, the mechanism is still unclear. This study aimed to elucidate the protective effects of KH capsule against carbon tetrachloride (CCl₄)-induced acute liver injury (ALI) in a murine model.

Methods: Mice were randomly divided into control, model (CCl₄), CCl₄+KH_Low and CCl₄+KH_High group. Liver enzyme levels and histological changes were assessed to evaluate liver injury. Oxidative stress markers and inflammatory cell infiltration in liver tissues were measured. Additionally, network pharmacology was employed to explore the potential mechanisms of KH capsule.

Results: KH capsule significantly reduced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, as well as the necrotic area in liver tissue. KH capsule also decreased the infiltration of macrophages and neutrophils, thereby inhibiting the expression of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and interleukin-1 beta (IL-1β). Furthermore, KH capsule decreased liver malondialdehyde (MDA) levels and increased superoxide dismutase (SOD) activity. The number of terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL)-positive cells in liver tissue was also reduced. The expression of nuclear factor erythroid 2 related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) proteins was significantly elevated, while the protein expression of cytochrome P450 2E1 (CYP2E1) was significantly reduced. Mass spectrometry identified genistein, galangin, wogonin, skullcapflavone II, and hispidulin as potential active ingredients of KH capsule. Network pharmacology analysis revealed enrichment in the mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K)-protein kinase B (AKT) signaling pathways. Western blot analysis confirmed that KH capsule suppressed AKT, extracellular signal-regulated kinase (ERK), and p38 signaling.

Conclusions: These findings suggest that KH capsule could exert protective effects against CCl₄-induced ALI, with the inhibition of MAPK and PI3K-AKT signaling pathways playing a crucial role in its mechanism of action.

Keywords

Kehuang capsule / Acute liver injury (ALI) / Carbon tetrachloride (CCl₄) / Mitogen-activated protein kinase (MAPK) / Protein kinase B (AKT) / Network pharmacology

Data availability statement

The data that support the findings of this study are available on request from the corresponding authors. The data are not publicly available due to privacy or ethical restrictions.

Authors’ contributions

Qinyu Ni and Jiacheng Lin contributed equally to this work and should be considered co-first authors. Qinyu Ni: Investigation. Jiacheng Lin: Methodology. Weifan Huang: Validation. Liu Yang: Methodology. Ran Li: Supervision. Tianzhi Tu: Supervision. Guangfu He: Supervision. Yueqiu Gao: Resources, Funding acquision. Xuehua Sun: Conceptualization. Xiaoni Kong: Concep-tualization. Xiaojun Zhu: Conceptualization. All authors approved the final version of this manuscript.

Declaration of competing interest

The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Ran Li, Tianzhi Tu and Guangfu He are currently employed by the market center of Kexing Biopharm Co., Ltd. The medicines provided by Kexing Biopharm Co., Ltd ensured the smooth conduct of the study. The opinions expressed in this study are those of the authors and do not necessarily represent those of Kexing Biopharm Co., Ltd. All researchers retained complete inde-pendence in the conduct of this study. The company had no influ-ence on the results of the study. Other authors declare that there are no competing interests.

Acknowledgements

This work was supported by the grant from the Shanghai Famous Old Chinese Medicine Academic Research Studio (No. SHGZS-202246).

Appendix A. Supplementary data

Supplementary data to this article can be found online at https://doi.org/10.1016/j.livres.2024.11.006.

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