title: Oxidative stress causes nuclear factor-kappaB activation in acute hypovolemic hemorrhagic shock.
abstract: Nuclear Factor kappaB (NFkappaB) is an ubiquitous rapid response transcription factor involved in inflammatory reactions and exerts its action by expressing cytokines, chemokines, and cell adhesion molecules. We investigated the role of NF-kappaB in acute hypovolemic hemorrhagic (Hem) shock. Hem shock was induced in male anesthetized rats by intermittently withdrawing blood from an iliac catheter over a period of 20 min (bleeding period) until mean arterial blood pressure (MAP) fell and stabilized within the range of 20-30 mmHg. Hemorrhagic shocked rats died in 26.3 +/- 2.1 min following the discontinuance of bleeding, experienced a marked hypotension (mean arterial blood pressure = 20-30 mmHg) and had enhanced plasma levels of Tumor Necrosis Factor-alpha (200 +/- 15 pg/ml, 20 min after the end of bleeding). Furthermore, aortas taken 20 min after bleeding from hemorrhagic shocked rats showed a marked hypo-reactivity to phenylephrine (PE; 1nM to 10 microM) compared with aortas harvested from sham shocked rats. Hem shocked rats also had increased levels of TNF-alpha mRNA in the liver (15-20 min after the end of bleeding) and enhanced plasma levels of 2,5-dihydroxybenzoic acid (2,5-DHBA; 6 +/- 2.2 microm), 2,3-dihydroxybenzoic acid (2,3-DHBA; 13 +/- 2.1 microm), both studied to evaluate OH(*) production. Electrophoretic mobility shift assay showed that liver NF-kappaB binding activity increased in the nucleus 10 min after the end of hemorrhage and remained elevated until the death of animals. Western blot analysis suggested that the levels of inhibitory IkappaBalpha protein in the cytoplasm became decreased at 5 min after the end of bleeding. IRFI-042, a vitamin E analogue (20 mg/kg intraperitoneally 2 min after the end of bleeding), inhibited the loss of IkappaBalpha protein from the cytoplasm and blunted the increase in NF-kappaB binding activity. Furthermore IRFI-042 increased survival time (117.8 +/- 6.51 min; p <.01) and survival rate (vehicle = 0% and IRFI-042 = 80%, at 120 min after the end of bleeding), reverted the marked hypotension, decreased liver mRNA for TNF-alpha, reduced plasma TNF-alpha (21 +/- 4.3 pg/ml), and restored to control values the hypo-reactivity to PE. Our results suggest that acute blood loss (50% of the estimated total blood volume over a period of 20 min) causes early activation of NF-kappaB, likely through an increased production of reactive oxygen species. This experiment indicates that NF-kappaB-triggered inflammatory cascade becomes early activated during acute hemorrhage even in the absence of resuscitation procedures.
authors.affiliation: Department of Clinical and Experimental Medicine and Pharmacology, Section of Pharmacology, University of Messina, Policlinico Universitario, Messina, Italy.
authors.forename: D
authors.initials: D
authors.lastname: Altavilla
authors.forename: A
authors.initials: A
authors.lastname: Saitta
authors.forename: S
authors.initials: S
authors.lastname: Guarini
authors.forename: M
authors.initials: M
authors.lastname: Galeano
authors.forename: G
authors.initials: G
authors.lastname: Squadrito
authors.forename: D
authors.initials: D
authors.lastname: Cucinotta
authors.forename: L B
authors.initials: LB
authors.lastname: Santamaria
authors.forename: A T
authors.initials: AT
authors.lastname: Mazzeo
authors.forename: G M
authors.initials: GM
authors.lastname: Campo
authors.forename: M
authors.initials: M
authors.lastname: Ferlito
authors.forename: L
authors.initials: L
authors.lastname: Minutoli
authors.forename: C
authors.initials: C
authors.lastname: Bazzani
authors.forename: A
authors.initials: A
authors.lastname: Bertolini
authors.forename: A P
authors.initials: AP
authors.lastname: Caputi
authors.forename: F
authors.initials: F
authors.lastname: Squadrito
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D001011:Aorta / Q000187:drug effects / Q000502:physiology / Q000503:physiopathology; D001572:Benzofurans / Q000494:pharmacology; D001794:Blood Pressure / Q000187:drug effects; D002478:Cells, Cultured; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004731:Endotoxins / Q000097:blood; D006470:Hemorrhage / Q000378:metabolism / Q000503:physiopathology; D017665:Hydroxyl Radical / Q000378:metabolism; D020896:Hypovolemia / Q000378:metabolism* / Q000503:physiopathology; D020672:I-kappa B Proteins; D008099:Liver / Q000378:metabolism; D017737:Macrophages, Peritoneal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D008297:Male; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000819:agonists / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects; D010656:Phenylephrine / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012771:Shock, Hemorrhagic / Q000378:metabolism* / Q000503:physiopathology; D015996:Survival Rate; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism; D014661:Vasoconstriction / Q000187:drug effects; D014662:Vasoconstrictor Agents / Q000494:pharmacology
chemical_list: D000975:Antioxidants; D001572:Benzofurans; D004268:DNA-Binding Proteins; D004731:Endotoxins; D020672:I-kappa B Proteins; C413683:IRFI 042; D016328:NF-kappa B; C000606206:Nfkbia protein, rat; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D014662:Vasoconstrictor Agents; D000072000:NF-KappaB Inhibitor alpha; D010656:Phenylephrine; D017665:Hydroxyl Radical; D004247:DNA