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dc.contributor.advisorCarter, Dean E.en_US
dc.contributor.authorHatlelid, Kristina Mary, 1967-
dc.creatorHatlelid, Kristina Mary, 1967-en_US
dc.date.accessioned2013-04-18T09:31:32Z
dc.date.available2013-04-18T09:31:32Z
dc.date.issued1996en_US
dc.identifier.urihttp://hdl.handle.net/10150/282124
dc.description.abstractA novel test system using isolated erythrocytes (red blood cells, RBCs) and arsine (AsH₃ gas dissolved in aqueous solution was characterized which allows for the quantitation of RBC exposure to AsH₃ in vitro and for in vitro study of the toxicity of AsH₃. AsH₃ was found to be rapidly and strongly associated with RBCs. Toxicity, measured as hemolysis, was time- and dose-dependent and exhibited a lag phase of about 30 minutes in both dog and rat RBCs. Hemolysis of dog RBCs was completely blocked by carbon monoxide preincubation and was reduced by pure oxygen. Sodium nitrite induction of methemoglobin (metHb) in intact rat RBCs decreased hemolysis, confirming the importance of hemoglobin in the mechanism of AsH₃-induced hemolysis. Spectrophotometric studies of the reaction of AsH₃ with purified dog hemoglobin revealed that AsH₃ reacted with reduced and oxygenated hemoglobin (HbO₂) to produce metHb and degraded Hb, characterized by gross precipitation of the protein. AsH₃ did not alter the spectrum of deoxyHb and did not cause degradation of metHb in oxygen, but bound to and reduced metHb in the absence of oxygen. Oxidative reactions as the putative cause of hemolysis were investigated. AsH₃ caused a slight decrease in cellular glutathione levels in dog RBCs, but only after hemolysis had reached maximum levels. Hydrogen peroxide (H₂O₂) was detected in aqueous solutions containing AsH₃ and HbO₂ or AsH₃ alone but not in intact red blood cells or lysates suggesting that cellular defenses were adequate to detoxify the amount formed. Additionally, high activity catalase or glutathione peroxidase added to solutions of HbO₂ and AsH₃ had little effect against AsH₃-induced damage. The differences between the visible spectra of HbO₂ treated with either AsH₃ or H₂O₂ suggest that two different degradative processes occur. Superoxide anion was not detected in AsH₃ mixtures and superoxide dismutase did not affect AsH₃-induced HbO₂ damage. Other antioxidants, mannitol, DMSO, ascorbate, and glutathione, had no effect against HbO₂ damage. These results indicate that the superoxide anion, the hydroxyl radical, and H₂O₂ are not responsible for AsH₃-induced HbO₂ damage and the subsequent hemolysis. A mechanism by which an arsenic species is the hemolytic agent is proposed.
dc.language.isoen_USen_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.subjectHealth Sciences, Toxicology.en_US
dc.subjectChemistry, Biochemistry.en_US
dc.subjectHealth Sciences, Pathology.en_US
dc.titleIn vitro investigation of the toxic mechanism of action of arsine on the erythrocyteen_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
thesis.degree.grantorUniversity of Arizonaen_US
thesis.degree.leveldoctoralen_US
dc.identifier.proquest9706166en_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.disciplinePharmacology & Toxicologyen_US
thesis.degree.namePh.D.en_US
dc.identifier.bibrecord.b34288016en_US
refterms.dateFOA2018-04-26T12:23:06Z
html.description.abstractA novel test system using isolated erythrocytes (red blood cells, RBCs) and arsine (AsH₃ gas dissolved in aqueous solution was characterized which allows for the quantitation of RBC exposure to AsH₃ in vitro and for in vitro study of the toxicity of AsH₃. AsH₃ was found to be rapidly and strongly associated with RBCs. Toxicity, measured as hemolysis, was time- and dose-dependent and exhibited a lag phase of about 30 minutes in both dog and rat RBCs. Hemolysis of dog RBCs was completely blocked by carbon monoxide preincubation and was reduced by pure oxygen. Sodium nitrite induction of methemoglobin (metHb) in intact rat RBCs decreased hemolysis, confirming the importance of hemoglobin in the mechanism of AsH₃-induced hemolysis. Spectrophotometric studies of the reaction of AsH₃ with purified dog hemoglobin revealed that AsH₃ reacted with reduced and oxygenated hemoglobin (HbO₂) to produce metHb and degraded Hb, characterized by gross precipitation of the protein. AsH₃ did not alter the spectrum of deoxyHb and did not cause degradation of metHb in oxygen, but bound to and reduced metHb in the absence of oxygen. Oxidative reactions as the putative cause of hemolysis were investigated. AsH₃ caused a slight decrease in cellular glutathione levels in dog RBCs, but only after hemolysis had reached maximum levels. Hydrogen peroxide (H₂O₂) was detected in aqueous solutions containing AsH₃ and HbO₂ or AsH₃ alone but not in intact red blood cells or lysates suggesting that cellular defenses were adequate to detoxify the amount formed. Additionally, high activity catalase or glutathione peroxidase added to solutions of HbO₂ and AsH₃ had little effect against AsH₃-induced damage. The differences between the visible spectra of HbO₂ treated with either AsH₃ or H₂O₂ suggest that two different degradative processes occur. Superoxide anion was not detected in AsH₃ mixtures and superoxide dismutase did not affect AsH₃-induced HbO₂ damage. Other antioxidants, mannitol, DMSO, ascorbate, and glutathione, had no effect against HbO₂ damage. These results indicate that the superoxide anion, the hydroxyl radical, and H₂O₂ are not responsible for AsH₃-induced HbO₂ damage and the subsequent hemolysis. A mechanism by which an arsenic species is the hemolytic agent is proposed.


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