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Background & AimsThe transcription factor nuclear factor-κB (NF)-κB promotes survival of hepatic myofibroblasts and fibrogenesis through poorly defined mechanisms. We investigated the activities of angiotensin II and IκB kinase (IKK) in regulation of NF-κB activity and the role of these proteins in liver fibrosis in rodents and humans. MethodsPhosphorylation of the NF-κB subunit RelA at serine 536 (P-Ser536-RelA) was detected by immunoblot and immunohistochemical analyses. P-Ser536-RelA function was assessed using vectors that expressed mutant forms of RelA, cell-permeable blocking peptides, and assays for RelA nuclear transport and apoptosis. Levels of P-Ser536-RelA were compared with degree of fibrosis in liver sections from chronically injured rats and patients with hepatitis C virus-mediated fibrosis who had been treated with the AT1 antagonist losartan. ResultsConstitutive P-Ser536-RelA is a feature of human hepatic myofibroblasts, both in vitro and in situ in diseased livers. Autocrine angiotensin II stimulated IKK-mediated phosphorylation of RelA at Ser536, which was required for nuclear transport and transcriptional activity of NF-κB. Inhibition of angiotensin II, the angiotensin II receptor type 1 (AT1), or IKK blocked Ser536 phosphorylation and stimulated myofibroblast apoptosis. Treatment of fibrotic rodent liver with the angiotensin converting enzyme (ACE) inhibitor captopril or the IKK inhibitor sulphasalazine resulted in loss of P-Ser536-RelA-positive myofibroblasts and fibrosis regression. In human liver samples, increased numbers of P-Ser536-RelA-positive cells were associated with fibrosis that regressed following exposure to losartan. ConclusionsAn autocrine pathway that includes angiotensin II, IKK, and P-Ser536-RelA regulates myofibroblast survival and can be targeted to stimulate therapeutic regression of liver fibrosis. Abbreviations used in this paper: ACE, angiotensin converting enzyme, α-SMA, smooth muscle α-actin, AT1, angiotensin type 1 receptor, IκB, inhibitor of NF-κB, IKK, IκB kinase, NF-κB, nuclear factor-κB, P-Ser536-RelA, RelA phosphorylated on serine 536 ⁎ Liver Research Group, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom ‡ Liver Group, University of Southampton, Southampton, United Kingdom § Liver Unit, Institut Clínic de Malalties Digestives i Metabòliques, Hospital Clínic, Institut d'Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain ∥ University of Queensland, School of Medicine, Southern Division, Princess Alexandra Hospital, Queensland, Australia ¶ Laboratory of Molecular and Cellular Biology, Division of Basic Sciences, the University of Crete Medical School, Crete, Greece
Conflicts of interest The authors disclose no conflicts. Funding Supported by grants from the British Liver Trust and the UK Medical Research Council (to D.A.M.). PII: S0016-5085(09)00369-2 doi:10.1053/j.gastro.2009.02.081 © 2009 AGA Institute. Published by Elsevier Inc. All rights reserved.
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ABSTRACT
Angiotensin II Activates IκB Kinase Phosphorylation of RelA at Ser