The pulmonary endothelium influences the redox status of blood borne redox active endogenous, pharmacological, toxicological and dietary compounds. It does so by acting as a large chemical reactor surface comprised of cell surface and intracellular redox enzymes situated between the venous and arterial circulations. The impact of the endothelium on the redox status of these compounds affects their bioactivity, including pro-or anti-oxidant effects, tissue permeation, and chemical reactivity. Quinones represent 1 important class of redox-active compounds with a range of physiological, pharmacological, and toxicological activities. The hypothesis underlying this proposal is that redox metabolism of quinones by pulmonary endothelial cells determines the quinone reduction products, and their bioactivity, within the lung and endothelial cells, the plasma, and downstream vessels and organ systems. The goal of the proposed research is to identify and quantify the factors, with regard to properties of both the cells and quinones, that determine the fate of a given quinone that comes in contact with the pulmonary endothelium.
The specific aims are to: 1. Identify and quantify the pulmonary endothelial quinone reductase and hydroquinone oxidase mediated reactions contributing to the fate of a series of quinones with varying physical and chemical properties: 2. Determine the dependence of the activities of quinone reductases contributing to the fate of a given quinone on the pulmonary endothelial redox status: 3. Determine the influence of adaptation of the pulmonary endothelium to hyperoxia on the redox processes contributing to quinone fate. To achieve these aims, the general approaches include: measuring the kinetics of intracellular and cell surface quinone reduction, hydroquinone oxidation and associations with cellular constituents using intact normoxic pulmonary arterial endothelial cells in culture and cells adapted to hyperoxia, as a model of oxidant stress. A panel of inhibitors and kinetic modeling will provide the means for functional and quantitative identification of the contributions of the quinone reductases and hydroquinone oxidases involved. Cell redox status will be manipulated experimentally to determine its role in the fate of the quinones that come in contact with the endothelial cells. Finally, the quinone reductases in the control and hyperoxia-adapted cells will be isolated and identified to further characterize their contributions to the net effect of both cell types to quinone fate. Since quinone reductases and hydroquinone oxidases are important in metabolism of a wide range of other redox active compounds, the results will have implications for endothelial processing of substances as well. The results will reveal pulmonary endothelial mechanisms involved in processing blood borne redox active substances, and their adaptation to oxidative stress, with implications for their pharmacological, physiological and toxicological activities throughout the body.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL065537-08
Application #
7367144
Study Section
Lung Injury, Repair, and Remodeling Study Section (LIRR)
Program Officer
Moore, Timothy M
Project Start
2000-08-20
Project End
2011-02-28
Budget Start
2008-03-01
Budget End
2011-02-28
Support Year
8
Fiscal Year
2008
Total Cost
$209,073
Indirect Cost
Name
Medical College of Wisconsin
Department
Anesthesiology
Type
Schools of Medicine
DUNS #
937639060
City
Milwaukee
State
WI
Country
United States
Zip Code
53226
Gan, Zhuohui; Audi, Said H; Bongard, Robert D et al. (2011) Quantifying mitochondrial and plasma membrane potentials in intact pulmonary arterial endothelial cells based on extracellular disposition of rhodamine dyes. Am J Physiol Lung Cell Mol Physiol 300:L762-72
Lindemer, Brian J; Bongard, Robert D; Hoffmann, Raymond et al. (2011) Genetic evidence for NAD(P)H:quinone oxidoreductase 1-catalyzed quinone reduction on passage through the mouse pulmonary circulation. Am J Physiol Lung Cell Mol Physiol 300:L773-80
Bongard, Robert D; Krenz, Gary S; Gastonguay, Adam J et al. (2011) Characterization of the threshold for NAD(P)H:quinone oxidoreductase activity in intact sulforaphane-treated pulmonary arterial endothelial cells. Free Radic Biol Med 50:953-62
Bongard, Robert D; Lindemer, Brian J; Krenz, Gary S et al. (2009) Preferential utilization of NADPH as the endogenous electron donor for NAD(P)H:quinone oxidoreductase 1 (NQO1) in intact pulmonary arterial endothelial cells. Free Radic Biol Med 46:25-32
Audi, Said H; Merker, Marilyn P; Krenz, Gary S et al. (2008) Coenzyme Q1 redox metabolism during passage through the rat pulmonary circulation and the effect of hyperoxia. J Appl Physiol 105:1114-26
Merker, Marilyn P; Audi, Said H; Lindemer, Brian J et al. (2007) Role of mitochondrial electron transport complex I in coenzyme Q1 reduction by intact pulmonary arterial endothelial cells and the effect of hyperoxia. Am J Physiol Lung Cell Mol Physiol 293:L809-19
Merker, Marilyn P; Audi, Said H; Bongard, Robert D et al. (2006) Influence of pulmonary arterial endothelial cells on quinone redox status: effect of hyperoxia-induced NAD(P)H:quinone oxidoreductase 1. Am J Physiol Lung Cell Mol Physiol 290:L607-19
Audi, Said H; Bongard, Robert D; Krenz, Gary S et al. (2005) Effect of chronic hyperoxic exposure on duroquinone reduction in adult rat lungs. Am J Physiol Lung Cell Mol Physiol 289:L788-97
Merker, Marilyn P; Bongard, Robert D; Krenz, Gary S et al. (2004) Impact of pulmonary arterial endothelial cells on duroquinone redox status. Free Radic Biol Med 37:86-103
Audi, Said H; Zhao, Hongtao; Bongard, Robert D et al. (2003) Pulmonary arterial endothelial cells affect the redox status of coenzyme Q0. Free Radic Biol Med 34:892-907

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