Superfund Research Program
Mechanisms Underlying Arsenic-Induced Vascular Disease
Drinking water in many countries, including the United States, can contain levels of arsenic that are associated with an increased incidence of occlusive vascular diseases. However, the mechanisms by which arsenic, especially its most toxic form, arsenite or arsenic(III), promotes the narrowing of blood vessels are relatively unknown. The current working hypothesis to explain how arsenite causes inward expansion of blood vessels is that it stimulates the production of reactive oxygen species in the membranes of the endothelial and smooth muscle cells that make up the vessel wall.
Research at Dartmouth College suggests that reactive oxygen formation has the dual effects of initiating cell signaling and decreasing the release of vessel dilators in endothelial cells exposed to arsenite. Reactive oxygen species also promote growth in endothelial and smooth muscle cells; however, this growth depends on the concentration of arsenite and occurs only below a toxic threshold.
Recent studies have focused on defining the cellular and molecular events involved in the conversion of vascular cells into an abnormal proliferative state. These studies used primary cells that were freshly isolated from blood vessels rather than using established cell lines. The primary cells retain many of the attributes of the vessel wall that are lost as cells are continually maintained in culture. The most important finding in these recent studies is that low, environmentally relevant levels of arsenite activated certain endothelial cell signaling and proliferation events without activating stress-related signaling events.
Interestingly, the geological research group in the Dartmouth College Superfund Basic Research Program previously demonstrated that some well water in New Hampshire contains as much as 300-400 ppb (parts per billion) of arsenic. In 1996, the first group of researchers reported that 35 ppb of arsenite was the threshold for activating the transcription factor NF-kB in cultured endothelial cells. NF-kB activates genes that promote cell survival and growth. Maximal activation of this factor in endothelial cells occurred in response to 175 ppb of arsenite. This activation required oxidant formation and was associated with an increase in DNA synthesis and cell proliferation.
More recent data from the laboratory demonstrated that 400 ppb of arsenite is the threshold for toxicity in this cell model. However, activation of the important stress signaling pathways in cultured endothelial cells required addition of more than 2000 ppb of arsenite. These stress pathways require signaling enzymes called MAP (mitogen-activated protein) kinases. Activation of these kinases occurred only when lethal amounts of arsenite were given to the cells. Overall, these findings indicate that arsenite can activate vascular cells when present at concentrations that are environmentally relevant, but not stressful or lethal at the cellular level.
Another important aspect of this research has been identification of the source of reactive oxygen that is stimulated by arsenite and required for activating cell signaling. These studies have relied on electron paramagnetic resonance with highly selective compounds to trap and identify types of reactive oxygen species produced in response to less than 400 ppb of arsenite. The primary species produced in response to arsenite has been identified as superoxide. Experiments with selective enzyme inhibitors indicate that the source of the superoxide produced following exposure of endothelial cells to arsenite is the NADPH oxidase in surface membranes of the cells. This enzyme generates superoxide anion from the oxidation of NADPH to NADP+. Once produced, superoxide is rapidly converted to hydrogen peroxide and it is this oxidant that activates cell signaling involving tyrosine kinase activity and NF-kB-dependent gene expression.
Current studies are focused on defining how arsenite activates NADPH oxidase and the causal links between the stimulation of tyrosine kinase activity, NF-kB activation and cell proliferation. Finally, in vivo studies are being planned to demonstrate how these changes in oxidant formation and cell signaling affect expansion of the walls of blood vessels in mice chronically exposed to arsenite in their drinking water.
These studies are defining the cellular and molecular signaling pathways that promote pathological changes in vascular cells following exposure to relatively low environmental levels of arsenite. These levels are not lethal to the vascular cells, but cause aberrant cell proliferation that eventually lead to reduced flow in blood vessels. The data generated will greatly improve the understanding of how low arsenite exposure causes vascular disease. In addition to defining the mechanisms for this non-cancer endpoint, these studies will provide fundamental insight into mechanisms for arsenite-induced signaling that could lead to other proliferative diseases, including cancer.
For More Information Contact:
University of Pittsburgh
Department of Environmental and Occupational Health
Bridgeside Point, 100 Technology Drive
Pittsburgh, Pennsylvania 15219-3130
To learn more about this research, please refer to the following sources:
- Barchowsky A, Dudek EJ, Treadwell MD, Wetterhahn KE. 1996. Arsenic induces oxidant stress and NF-ÇB activation in cultured aortic endothelial cells. Free Radic Biol Med 21(6):783-790. PMID:8902524
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