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Shc is implicated in calreticulin-mediated sterile inflammation in alcoholic hepatitis

Open AccessPublished:September 16, 2022DOI:https://doi.org/10.1016/j.jcmgh.2022.09.005
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      Abstract

      Background

      &Aims: Src homology and collagen (Shc) proteins are major adapters to extra-cellular signals however the regulatory role of Shc isoforms in sterile inflammatory responses in alcoholic hepatitis (AH) has not been fully investigated. We hypothesized that in an isoform-specific manner Shc modulates pre-apoptotic signals, calreticulin (CRT) membrane exposure and recruitment of inflammatory cells.

      Methods

      Liver biopsy samples from patients with AH vs. healthy subjects were studied for Shc expression using DNA microarray data and immunohistochemistry. ShcKD (hypomorph) and age-matched wild type (WT) mice were pair-fed according to the chronic-plus-binge alcohol (NIAAA) diet. To analyze hepatocyte-specific effects, AAV8-TBG-Cre (ShcHepKO)-mediated deletion was performed in fl/fl Shc mice. Lipid peroxidation, proinflammatory signals, redox radicals, NADH/NAD+ ratio, as well as cleaved caspase 8, BAP31, Bax, and Bak, in vivo. CRT translocation was studied in ethanol-exposed p46ShcẟSH2-transfected hepatocytes by membrane biotinylation in conjunction with p-Eif2α, BAP31, caspase 8, Bax/Bak. The effects of idebenone, a novel Shc inhibitor was studied in alcohol/pair-fed mice.

      Results

      Shc was significantly induced in patients with AH (p<0.01). ALT, NADH/NAD+ ratios, production of redox radicals, lipid peroxidation improved (p<0.05), and IL-1β, MCP-1, and CXCL10 were reduced in ShcKD and ShcHepKO mice. In vivo, Shc-dependent induction, and in hepatocytes, p46Shc-dependent increase in pre-apoptotic proteins Bax/Bak, caspase 8, BAP31 cleavage and membrane translocation of CRT/Erp57 were seen. Idebenone protected against alcohol-mediated liver injury.

      Conclusion

      Alcohol induces p46Shc-dependent activation of pre-apoptotic pathways and translocation of CRT to the membrane, where it acts as a DAMP, instigating immunogenicity. Shc inhibition could be a novel treatment strategy in AH.

      Graphical abstract

      Keywords

      Abbreviations:

      4-HNE (4-hydroxynonenal), AAV (adeno-associated virus), ACAA2 (3-ketoacyl CoA thiolase), AH (alcoholic hepatitis), ALD (alcoholic liver disease), ALT (alanine aminotransferase), AST (aspartate aminotransferase), Bak (Bcl-2 homologous antagonist killer), BAP31 (B-cell receptor-associated protein 31), Bax (Bcl-2-associated X protein), CRT (calreticulin), CXCL10 (C-X-C chemokine ligand 10), DAMP (damage-associated molecular pattern), EIF2S1 (eukaryotic transcription factor 2 subunit 1), ER (endoplasmic reticulum), IL-1 β (interleukin 1 beta), MCP1 (monocyte chemoattractant protein 1), MCS-F (macrophage colony-stimulating factor), NAD+ (oxidized nicotinamide adenine dinucleotide), NADH (reduced nicotinamide adenine dinucleotide), ROS (reactive oxygen species), RTK (receptor tyrosine kinase), Shc (Src homology and collagen), ShcKD (Shc knockdown/Shc hypomorph), ShcHepKO (hepatocyte-specific Shc knockout), TG (triglyceride)