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  3. Targeting GCS regulates the NRF2 axis to inhibit hepatocyte endoplasmic reticulum stress-ferroptosis improving immune-mediated liver injury

Targeting GCS regulates the NRF2 axis to inhibit hepatocyte endoplasmic reticulum stress-ferroptosis improving immune-mediated liver injury

  • Int Immunopharmacol. 2025 Sep 13:165:115532. doi: 10.1016/j.intimp.2025.115532.
Zibing Qian 1,  Junfeng Li 2,  Xiaorong Mao 3,  Jian Gan 4,  Aipin Tian 3,  Xuebin Peng 3
Affiliations

Affiliations

  • 1 The First Hospital of Lanzhou University, Lanzhou, 730000, China; The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China; Department of Infectious Disease, The First Hospital of Lanzhou University, Lanzhou 730000, China.
  • 2 The First Hospital of Lanzhou University, Lanzhou, 730000, China; The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China; Department of Hepatology, The First Hospital of Lanzhou University, Lanzhou 730000, China; Institute of Infectious Diseases, The First Hospital of Lanzhou University, Lanzhou 730000, China. Electronic address: ldyy_lijf@lzu.edu.cn.
  • 3 Department of Infectious Disease, The First Hospital of Lanzhou University, Lanzhou 730000, China.
  • 4 Department of Gastroenterology, Yantai Affiliated Hospital of Binzhou Medical University, Yantai 264000, China.
PMID: 40946670 DOI: 10.1016/j.intimp.2025.115532
Abstract

Background: Ferroptosis is a regulated, iron-dependent form of cell death triggered by toxic accumulation of lipid peroxides on cell membranes. Although it is established that glycosphingoid metabolites can activate the Ferroptosis signaling pathway, whether they are involved in regulating endoplasmic reticulum stress (ERS)-ferroptosis in immune-mediated liver injury requires further investigation.

Methods: This study used the GCS inhibitor GENZ-123346 (GENZ) to treat ConA-induced immune liver injury in mice. Inflammatory factor levels and liver histology were analyzed to evaluate GENZ's effects. Additionally, the antioxidant and lipid peroxidation capacity of GENZ was tested in vivo and in vitro. Based on the regulation of NFE2-related factor 2 (NRF2) signaling pathway expression (including si-NRF2, NRF2 inducer sulforaphane (SFN), and inhibitor ML385), the regulatory effect of GENZ on ERS-ferroptosis was investigated.

Results: GENZ administration improved the survival rate of mice with immune-mediated liver injury, decreased serum transaminase activity, inhibited inflammatory factor production, reduced hepatocyte Ferroptosis. In in vitro experiments, Ferroptosis in AML12 cells was accompanied by the activation of ERS-related proteins Glucose-regulated protein 78 (GRP78) and C/EBP homologous protein (CHOP) and mild alterations in endoplasmic reticulum ultrastructure. Inhibition of ERS by 4-phenylbutyric acid (4-PBA) attenuated Ferroptosis in AML12 cells. However, when the cells were treated with the Ferroptosis inducer RSL3 in combination with the ERS inducer thapsigargin (Tg), GENZ could not effectively suppress Ferroptosis. Silencing NRF2 exacerbated cellular Ferroptosis and ERS while diminishing the expression of downstream effectors heme oxygenase-1 (HO-1) and NAD(P)H quinone oxidoreductase 1 (NQO1). Conversely, SFN treatment alleviated these effects and upregulated NRF2 signaling, increasing HO-1 and NQO1 expression. Furthermore, ML385 administration reversed the hepatoprotective and antioxidant effects of GENZ on immune-mediated liver injury in mice.

Conclusions: GCS inhibition exerts hepatoprotective effects by modulating the NRF2 antioxidant pathway to suppress ERS and hepatocyte Ferroptosis, revealing a novel role of glycosphingolipid metabolism in immune-mediated liver injury and programmed hepatocyte death.

Keywords

Endoplasmic reticulum stress; Ferroptosis; Glucosylceramide synthase; Immune-mediated liver injury; NRF2.

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