Differential Cytotoxicity and Reactive Oxygen Species Generation in Pulmonary and Aortic Cells Exposed to Inorganic Arsenic and Monomethylarsonous Acid
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Authors
Khalili, Roxana
Issue Date
2013
Type
Thesis
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Keywords
Arsenic , Cytotoxicity , Monomethylarsonous Acid
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Abstract
Chronic ingestion of arsenic (As), a common ground water contaminant, has demonstrated to result in numerous deleterious health outcomes including black foot disease, various cancers, and hyperpigmentation. Chronic arsenic ingestion has also been associated with the development of atherosclerosis and hypertension, though the cellular mechanisms have not been well elucidated. Recent studies have shown that an arsenic metabolite, monomethylarsonous acid (MMAs), causes a higher degree of toxicity than inorganic arsenic (iAs) and may be linked to arsenic-induced vascular diseases. This study explores possible reactive oxygen species that may contribute to atherosclerosis and examines the different cytotoxic effects of iAs and MMAs on thoracic aorta smooth muscle cells (A7r5) and rat pulmonary arterial smooth muscle cells (rPASMC) in culture to look for the generation of malondialdehyde, hydrogen peroxide, and superoxide. Cytotoxicity was determined by cell counts and Trypan Blue exclusion, MTT assay, and light microscopy to study altered smooth muscle cell morphology, cell viability and cytotoxicity. Cells treated with various concentrations of either iAs and MMAs displayed cytotoxic effects and MMAs was significantly more toxic in both A7r5 and rPASMC. After a 24 hour exposure the LC50 in A7r5 treated with iAs was determined to be 11 µM, and 700 nM when treated with MMAs. In rPASMC treated with iAs the LC50 was determined to be 26.5 µM, and 4 µM with MMAs treatment showing higher toxicity in A7r5 cells than in rPASMC. No significant difference in MDA formation or superoxide production was observed with treatment of iAs and MMAs in both A7r5 and rPASMC. Hydrogen peroxide was measured in the extracellular medium of rPASMC and displayed a 12% increase after a 2 hour treatment with 100 nM (p=0.006) iAs and 1 µM iAs (p=0.0002) when compared to untreated cells, a 7% increase when treated with 1 µM MMAs (p=0.03) and a 9% increase when treated with 10 µM MMAs (p=0.0003). In A7r5 cells there was a significant increase in the release of hydrogen peroxide with exposure to 1 µM MMAs displaying a 59% increase (p=0.0169) and 10 µM MMAs displaying a 70% increase (p=0.003) when compared to untreated cells.These results suggest that the increased toxicity of MMAs could be due to increased hydrogen peroxide activity in A7r5 cells, but is not due solely due to oxidative stress when analyzed in vitro. Other mechanisms need to be further explored to understand the association between arsenic and cardiovascular disease.
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In Copyright(All Rights Reserved)