Ectodomain shedding of EGFR ligands and TNFR1 dictates hepatocyte apoptosis during fulminant hepatitis in mice
Aditya Murthy, … , Carl P. Blobel, Rama Khokha
Aditya Murthy, … , Carl P. Blobel, Rama Khokha
Published July 12, 2010
Citation Information: J Clin Invest. 2010;120(8):2731-2744. https://doi.org/10.1172/JCI42686.
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Research Article Hepatology

Ectodomain shedding of EGFR ligands and TNFR1 dictates hepatocyte apoptosis during fulminant hepatitis in mice

Abstract

The cell death receptor Fas plays a role in the establishment of fulminant hepatitis, a major cause of drug-induced liver failure. Fas activation elicits extrinsic apoptotic and hepatoprotective signals; however, the mechanisms by which these signals are integrated during disease are unknown. Tissue inhibitor of metalloproteinases 3 (TIMP3) controls the critical sheddase a disintegrin and metalloproteinase 17 (ADAM17) and may dictate stress signaling. Using mice and cells lacking TIMP3, ADAM17, and ADAM17-regulated cell surface molecules, we have found that ADAM17-mediated ectodomain shedding of TNF receptors and EGF family ligands controls activation of multiple signaling cascades in Fas-induced hepatitis. We demonstrated that TNF signaling promoted hepatotoxicity, while excessive TNF receptor 1 (TNFR1) shedding in Timp3–/– mice was protective. Compound Timp3–/–Tnf–/– and Timp3–/–Tnfr1–/– knockout conferred complete resistance to Fas-induced toxicity. Loss of Timp3 enhanced metalloproteinase-dependent EGFR signaling due to increased release of the EGFR ligands TGF-α, amphiregulin, and HB-EGF, while depletion of shed amphiregulin resensitized Timp3–/– hepatocytes to apoptosis. Finally, adenoviral delivery of Adam17 prevented acetaminophen-induced liver failure in a clinically relevant model of Fas-dependent fulminant hepatitis. These findings demonstrate that TIMP3 and ADAM17 cooperatively dictate cytokine signaling during death receptor activation and indicate that regulated metalloproteinase activity integrates survival and death signals during acute hepatotoxic stress.

Authors

Aditya Murthy, Virginie Defamie, David S. Smookler, Marco A. Di Grappa, Keisuke Horiuchi, Massimo Federici, Maria Sibilia, Carl P. Blobel, Rama Khokha

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Figure 1

Ablation of TNF signaling delays Fas-mediated hepatotoxicity.

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Ablation of TNF signaling delays Fas-mediated hepatotoxicity. (A) Apopto...
(A) Apoptosis of wild-type primary hepatocytes treated for 12 hours with the indicated doses of Jo-2 in the presence or absence of 1 ng/ml TNF. Caspase-3/7 activity was measured by rate of cleavage of fluorogenic peptide Ac-DEVD-AMC and represented relative to untreated samples. *P < 0.03. Data are mean ± SD (n = 3). (B) Survival curve of wild-type, Tnf–/–, and Tnfr1–/– mice injected i.p. with 0.65 μg/g Jo-2. P < 0.005 versus WT (log-rank test). (C–E) WT, Tnf–/–, and Tnfr1–/– mice were injected i.p. with 0.65 μg/g Jo-2 or PBS. Mice were sacrificed and tissue obtained 3 hours after injection. (C) Liver histology (H&E, active caspase-3 immunohistochemistry) showing absence of tissue damage and caspase-3 activation in Tnf–/– and Tnfr1–/– mice after treatment with Jo-2. Sections are representative of at least 4 mice per genotype and treatment. Scale bars: 100 μm (D) Hepatic toxicity measured by serum ALT levels. *P ≤ 0.04. Data are mean ± SD (n = 5). (E) Immunoblots assaying the processing of caspase-8 and -3 in liver lysates. All data are representative of at least 2 independent experiments.