title: Bilirubin mediated oxidative stress involves antioxidant response activation via Nrf2 pathway.
abstract: Unconjugated bilirubin (UCB) is responsible for neonatal jaundice and high level of free bilirubin (Bf) can lead to kernicterus. Previous studies suggest that oxidative stress is a critical component of UCB-induced neurotoxicity. The Nrf2 pathway is a powerful sensor for cellular redox state and is activated directly by oxidative stress and/or indirectly by stress response protein kinases. Activated Nrf2 translocates to nucleus, binds to Antioxidant Response Element (ARE), and enhances the up-regulation of cytoprotective genes that mediate cell survival. The aim of the present study was to investigate the role of Nrf2 pathway in cell response to bilirubin mediated oxidative stress in the neuroblastoma SH-SY5Y cell line. Cells exposed to a toxic concentration of UCB (140 nM Bf) showed an increased intracellular ROS levels and enhanced nuclear accumulation of Nrf2 protein. UCB stimulated transcriptional induction of ARE-GFP reporter gene and induced mRNA expression of multiple antioxidant response genes as: xCT, Gly1, γGCL-m, γGCL-c, HO-1, NQO1, FTH, ME1, and ATF3. Nrf2 siRNA decreased UCB induced mRNA expression of HO1 (75%), NQO1 (54%), and FTH (40%). The Nrf2-related HO-1 induction was reduced to 60% in cells pre-treated with antioxidant (NAC) or specific signaling pathway inhibitors for PKC, P38α and MEK1/2 (80, 40 and 25%, respectively). In conclusion, we demonstrated that SH-SY5Y cells undergo an adaptive response against UCB-mediated oxidative stress by activation of multiple antioxidant response, in part through Nrf2 pathway.
authors.affiliation: Fondazione Italiana Fegato ONLUS, Italian Liver Foundation ONLUS, Bldg Q AREA Science Park - Basovizza Campus, 34149 Trieste, Italy. Electronic address: m.qaisiya@csf.units.it.
authors.forename: Mohammed
authors.initials: M
authors.lastname: Qaisiya
authors.affiliation: Fondazione Italiana Fegato ONLUS, Italian Liver Foundation ONLUS, Bldg Q AREA Science Park - Basovizza Campus, 34149 Trieste, Italy. Electronic address: carlos.zabetta@csf.units.it.
authors.forename: Carlos Daniel
authors.initials: CD
authors.lastname: Coda Zabetta
authors.affiliation: Fondazione Italiana Fegato ONLUS, Italian Liver Foundation ONLUS, Bldg Q AREA Science Park - Basovizza Campus, 34149 Trieste, Italy. Electronic address: cristina.bellarosa@csf.units.it.
authors.forename: Cristina
authors.initials: C
authors.lastname: Bellarosa
authors.affiliation: Fondazione Italiana Fegato ONLUS, Italian Liver Foundation ONLUS, Bldg Q AREA Science Park - Basovizza Campus, 34149 Trieste, Italy; Department of Medical Sciences, University of Trieste, 34100 Trieste, Italy. Electronic address: ctliver@csf.units.it.
authors.forename: Claudio
authors.initials: C
authors.lastname: Tiribelli
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D051700:Activating Transcription Factor 3 / Q000096:biosynthesis; D061745:Antioxidant Response Elements / Q000235:genetics*; D000975:Antioxidants / Q000494:pharmacology*; D001052:Apoferritins / Q000096:biosynthesis; D001663:Bilirubin / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D016166:Free Radical Scavengers / Q000494:pharmacology; D051547:Heme Oxygenase-1 / Q000096:biosynthesis; D056945:Hep G2 Cells; D006801:Humans; D048369:MAP Kinase Kinase 1 / Q000037:antagonists & inhibitors; D048370:MAP Kinase Kinase 2 / Q000037:antagonists & inhibitors; D048308:Mitogen-Activated Protein Kinase 14 / Q000037:antagonists & inhibitors; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000096:biosynthesis; D051267:NF-E2-Related Factor 2 / Q000235:genetics / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D034622:RNA Interference; D012333:RNA, Messenger / Q000096:biosynthesis; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics; D015533:Transcriptional Activation / Q000187:drug effects / Q000235:genetics
chemical_list: C490449:ATF3 protein, human; D051700:Activating Transcription Factor 3; D000975:Antioxidants; D016166:Free Radical Scavengers; D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D001052:Apoferritins; C496541:HMOX1 protein, human; D051547:Heme Oxygenase-1; D016660:NAD(P)H Dehydrogenase (Quinone); C447479:NQO1 protein, human; C482128:MAP2K2 protein, human; D011493:Protein Kinase C; D048308:Mitogen-Activated Protein Kinase 14; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; C482125:MAP2K1 protein, human; D001663:Bilirubin; D000111:Acetylcysteine
keywords: ARE; ATF3; Antioxidant Response Element; Bf; Bilirubin neurotoxicity; FTH; GSH; Gly1; HO-1; Heme oxygenase 1; JNK; MAPK; ME1; N-acetylcystein; NAC; NADPH quinone oxidoreductase 1; NF-E2 related factor 2; NQO1; Nrf2; Nrf2 pathway; OS; Oxidative stress; PERK; PI3K; PKC; ROS; Stress response protein kinases; UCB; activating transcription factor 3; c-Jun NH2-terminal kinases; cystine/glutamate exchanger system; cytosolic malic enzyme 1; eIF2α; eukaryotic translation initiation factor 2α; ferritin heavy chain; free bilirubin; glutathione; glycine up-take transporter; heme oxygenase 1; mitogen-activated protein kinase; oxidative stress; phosphatidylinositide 3-kinases; protein kinase C; protein kinase-like endoplasmic reticulum kinase; reactive oxygen species; tBHQ; tertiary-butylhydroquinone; unconjugated bilirubin; xCT; γ-GCL-c; γ-GCL-m; γ-glutamylcysteine synthetase, catalytic subunit; γ-glutamylcysteine synthetase, modulatory subunit