Deletion of IKK2 in hepatocytes does not sensitize these cells to TNF-induced apoptosis but protects from ischemia/reperfusion injury
Tom Luedde, … , Manolis Pasparakis, Christian Trautwein
Tom Luedde, … , Manolis Pasparakis, Christian Trautwein
Published April 1, 2005
Citation Information: J Clin Invest. 2005;115(4):849-859. https://doi.org/10.1172/JCI23493.
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Article Hepatology

Deletion of IKK2 in hepatocytes does not sensitize these cells to TNF-induced apoptosis but protects from ischemia/reperfusion injury

Abstract

The inhibitor of NF-κB (I-κB) kinase (IKK) complex consists of 3 subunits, IKK1, IKK2, and NF-κB essential modulator (NEMO), and is involved in the activation of NF-κB by various stimuli. IKK2 or NEMO constitutive knockout mice die during embryogenesis as a result of massive hepatic apoptosis. Therefore, we examined the role of IKK2 in TNF-induced apoptosis and ischemia/reperfusion (I/R) injury in the liver by using conditional knockout mice. Hepatocyte-specific ablation of IKK2 did not lead to impaired activation of NF-κB or increased apoptosis after TNF-α stimulation whereas conditional NEMO knockout resulted in complete block of NF-κB activation and massive hepatocyte apoptosis. In a model of partial hepatic I/R injury, mice lacking IKK2 in hepatocytes displayed significantly reduced liver necrosis and inflammation than wild-type mice. AS602868, a novel chemical inhibitor of IKK2, protected mice from liver injury due to I/R without sensitizing them toward TNF-induced apoptosis and could therefore emerge as a new pharmacological therapy for liver resection, hemorrhagic shock, or transplantation surgery.

Authors

Tom Luedde, Ulrike Assmus, Torsten Wüstefeld, Andreas Meyer zu Vilsendorf, Tania Roskams, Mark Schmidt-Supprian, Klaus Rajewsky, David A. Brenner, Michael P. Manns, Manolis Pasparakis, Christian Trautwein

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Activation of NF-κB and induction of NF-κB target genes after I/R is dep...
Activation of NF-κB and induction of NF-κB target genes after I/R is dependent on functional IKK2. (A) Nuclear protein extracts (5 μg) from livers of Ikk2f/f and Ikk2Δhepa mice at different time points after I/R were subjected to a gel-retardation assay with an NF-κB consensus oligonucleotide. In lanes 9 and 10, antibodies for the NF-κB subunits p50 or p65 were added as supershift control. (B) Immunohistochemical staining for iNOS at 6 hours after I/R in Ikk2f/f and Ikk2Δhepa mice. Original magnification, ×40. Results are representative of those obtained in mice (n = 4). (C) Immunohistochemical staining for TNF-α at 6 hours after I/R. Original magnification, ×40. The number of TNF-α–positive cells was quantified (right panel). Values are mean ± SD for independent animals (n = 4). The asterisk indicates statistical significance: P < 0.01 versus Ikk2f/f control mice. (D) Kupffer cells were isolated from livers of Ikk2f/f and Ikk2Δhepa mice and stained with an antibody against the F4/80 antigen, which is a macrophage-specific marker, to verify the specificity of the preparation procedure. Equal expression of IKK2 was verified by staining with a polyclonal antibody against IKK2. Cells were stimulated with 1 μg/ml LPS for 1 hour and TNF-α expression examined by immunohistochemical staining to prove that Kupffer cells in the livers of Ikk2Δhepa mice are functionally active. Original magnification, ×400. For negative control, no primary antibody was added.