title: In vivo siRNA delivery of Keap1 modulates death and survival signaling pathways and attenuates concanavalin-A-induced acute liver injury in mice.
abstract: Oxidative stress contributes to the progression of acute liver failure (ALF). Transcription factor nuclear factor-erythroid 2-related factor (Nrf2) serves as an endogenous regulator by which cells combat oxidative stress. We have investigated liver damage and the balance between death and survival signaling pathways in concanavalin A (ConA)-mediated ALF using in vivo siRNA delivery targeting Keap1 in hepatocytes. For that goal, mice were injected with Keap1- or luciferase-siRNA-containing liposomes via the tail vein. After 48 hours, ALF was induced by ConA. Liver histology, pro-inflammatory mediators, antioxidant responses, cellular death, and stress and survival signaling were assessed. Keap1 mRNA and protein levels significantly decreased in livers of Keap1-siRNA-injected mice. In these animals, histological liver damage was less evident than in control mice when challenged with ConA. Likewise, markers of cellular death (FasL and caspases 8, 3 and 1) decreased at 4 and 8 hours post-injection. Nuclear Nrf2 and its target, hemoxygenase 1 (HO1), were elevated in Keap1-siRNA-injected mice compared with control animals, resulting in reduced oxidative stress in the liver. Similarly, mRNA levels of pro-inflammatory cytokines were reduced in livers from Keap1-siRNA-injected mice. At the molecular level, activation of c-jun (NH2) terminal kinase (JNK) was ameliorated, whereas the insulin-like growth factor I receptor (IGFIR) survival pathway was maintained upon ConA injection in Keap1-siRNA-treated mice. In conclusion, our results have revealed a potential therapeutic use of in vivo siRNA technology targeted to Keap1 to combat oxidative stress by modulating Nrf2-mediated antioxidant responses and IGFIR survival signaling during the progression of ALF.
authors.affiliation: Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Instituto de Salud Carlos III, Spain Instituto de Investigaciones Biomédicas "Alberto Sols" (Consejo Superior de Investigaciones Científicas/Universidad Autónoma de Madrid), 28029 Madrid, Spain aguedagr@iib.uam.es avalverde@iib.uam.es.
authors.forename: Águeda
authors.initials: Á
authors.lastname: González-Rodríguez
authors.affiliation: Roche Kulmbach GmbH, Kulmbach, D 95326, Germany.
authors.forename: Bjorn
authors.initials: B
authors.lastname: Reibert
authors.affiliation: Roche Kulmbach GmbH, Kulmbach, D 95326, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Amann
authors.affiliation: Roche Kulmbach GmbH, Kulmbach, D 95326, Germany.
authors.forename: Rainier
authors.initials: R
authors.lastname: Constien
authors.affiliation: Metabolic Diseases, Hoffmann-la Roche Inc., Nutley, NJ 07110-1199, USA.
authors.forename: Cristina M
authors.initials: CM
authors.lastname: Rondinone
authors.affiliation: Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Instituto de Salud Carlos III, Spain aguedagr@iib.uam.es avalverde@iib.uam.es.
authors.forename: Ángela M
authors.initials: ÁM
authors.lastname: Valverde
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000502:physiology*; D000818:Animals; D002470:Cell Survival / Q000502:physiology*; D003208:Concanavalin A / Q000187:drug effects / Q000494:pharmacology / Q000633:toxicity*; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000502:physiology*; D017931:DNA Primers; D000072019:Kelch-Like ECH-Associated Protein 1; D008099:Liver / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D003598:Cytoskeletal Proteins; D017931:DNA Primers; C117301:Keap1 protein, mouse; D000072019:Kelch-Like ECH-Associated Protein 1; D034741:RNA, Small Interfering; D003208:Concanavalin A
keywords: Acute liver failure; Apoptosis; In vivo siRNA; Keap1; Nrf2