title: KEAP1-NRF2 complex in ischemia-induced hepatocellular damage of mouse liver transplants.
abstract: OBJECTIVE
  The Keap1-Nrf2 signaling pathway regulates host cell defense responses against oxidative stress and maintains the cellular redox balance.
  
  
  METHODS
  We investigated the function/molecular mechanisms by which Keap1-Nrf2 complex may influence liver ischemia/reperfusion injury (IRI) in a mouse model of hepatic cold storage (20h at 4°C) followed by orthotopic liver transplantation (OLT).
  
  
  RESULTS
  The Keap1 hepatocyte-specific knockout (HKO) in the donor liver ameliorated post-transplant IRI, evidenced by improved hepatocellular function and OLT outcomes (Keap1 HKO→Keap1 HKO; 100% survival), as compared with controls (WT→WT; 50% survival; p<0.01). By contrast, donor liver Nrf2 deficiency exacerbated IRI in transplant recipients (Nrf2 KO→Nrf2 KO; 40% survival). Ablation of Keap1 signaling reduced macrophage/neutrophil trafficking, pro-inflammatory cytokine programs, and hepatocellular necrosis/apoptosis, while simultaneously promoting anti-apoptotic functions in OLTs. At the molecular level, Keap1 HKO increased Nrf2 levels, stimulated Akt phosphorylation, and enhanced expression of anti-oxidant Trx1, HIF-1α, and HO-1. Pretreatment of liver donors with PI3K inhibitor (LY294002) disrupted Akt/HIF-1A signaling and recreated hepatocellular damage in otherwise IR-resistant Keap1 HKO transplants. In parallel in vitro studies, hydrogen peroxide-stressed Keap1-deficient hepatocytes were characterized by enhanced expression of Nrf2, Trx1, and Akt phosphorylation, in association with decreased release of lactate dehydrogenase (LDH) in cell culture supernatants.
  
  
  CONCLUSIONS
  Keap1-Nrf2 complex prevents oxidative injury in IR-stressed OLTs through Keap1 signaling, which negatively regulates Nrf2 pathway. Activation of Nrf2 induces Trx1 and promotes PI3K/Akt, crucial for HIF-1α activity. HIF-1α-mediated overexpression of HO-1/Cyclin D1 facilitates cytoprotection by limiting hepatic inflammatory responses, and hepatocellular necrosis/apoptosis in a PI3K-dependent manner.
authors.affiliation: Dumont-UCLA Transplant Center, Department of Surgery, Division of Liver and Pancreas Transplantation, David Geffen School of Medicine at UCLA, Los Angeles, CA, United States.
authors.forename: Bibo
authors.initials: B
authors.lastname: Ke
authors.forename: Xiu-Da
authors.initials: XD
authors.lastname: Shen
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhang
authors.forename: Haofeng
authors.initials: H
authors.lastname: Ji
authors.forename: Feng
authors.initials: F
authors.lastname: Gao
authors.forename: Shi
authors.initials: S
authors.lastname: Yue
authors.forename: Naoko
authors.initials: N
authors.lastname: Kamo
authors.forename: Yuan
authors.initials: Y
authors.lastname: Zhai
authors.forename: Masayuki
authors.initials: M
authors.lastname: Yamamoto
authors.forename: Ronald W
authors.initials: RW
authors.lastname: Busuttil
authors.forename: Jerzy W
authors.initials: JW
authors.lastname: Kupiec-Weglinski
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000502:physiology*; D000818:Animals; D003598:Cytoskeletal Proteins / Q000502:physiology*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000502:physiology; D000072019:Kelch-Like ECH-Associated Protein 1; D008099:Liver / Q000098:blood supply*; D016031:Liver Transplantation; D008264:Macrophages / Q000502:physiology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D051267:NF-E2-Related Factor 2 / Q000502:physiology*; D009504:Neutrophils / Q000502:physiology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology; D015427:Reperfusion Injury / Q000209:etiology*; D015398:Signal Transduction
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D003598:Cytoskeletal Proteins; C497443:Hif1a protein, mouse; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; C117301:Keap1 protein, mouse; D000072019:Kelch-Like ECH-Associated Protein 1; D051267:NF-E2-Related Factor 2; C495636:Nfe2l2 protein, mouse; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt
keywords: ARE; H(2)O(2); HIF-1; HKO; HO-1; HRE; IRI; Keap1; Keap1-Nrf2 redox system; Kelch-like ECH-associated protein 1; LDH; Liver ischemia/reperfusion injury; Liver transplantation; Nrf2; OLT; PI3K; TUNEL; Trx1; anti-oxidant response element; heme-oxygenase-1; hepatocyte knockout; hydrogen peroxide; hypoxia inducible factor-1; hypoxia response element; ischemia/reperfusion injury; lactate dehydrogenase; nuclear factor erythroid 2-related factor 2; orthotopic liver transplantation; phosphoinositide 3-kinase; sALT; serum alanine aminotransferase; terminal deoxyribonucleotidyl transferase (TdT)-mediated dUTP-digoxigenin nick end labeling; thioredoxin 1