Genetic evidence for NAD(P)H:quinone oxidoreductase 1-catalyzed quinone reduction on passage through the mouse pulmonary circulation

Abstract

Lindemer BJ, Bongard RD, Hoffmann R, Baumgardt S, Gonzalez FJ, Merker MP. 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-L780, 2011. First published February 4, 2011 doi:10.1152/ajplung.00394.2010.-The quinones duroquinone (DQ) and coenzyme Q 1 (CoQ1) and quinone reductase inhibitors have been used to identify reductases involved in quinone reduction on passage through the pulmonary circulation. In perfused rat lung, NAD(P)H:quinone oxidoreductase 1 (NQO1) was identified as the predominant DQ reductase and NQO1 and mitochondrial complex I as the CoQ 1 reductases. Since inhibitors have nonspecific effects, the goal was to use Nqo1-null (NQO1 Ϫ / Ϫ ) mice to evaluate DQ as an NQO1 probe in the lung. Lung homogenate cytosol NQO1 activities were 97 Ϯ 11, 54 Ϯ 6, and 5 Ϯ 1 (SE) nmol dichlorophenolindophenol reduced · min Ϫ1 · mg protein Ϫ1 for NQO1 ϩ/ϩ , NQO1 ϩ/Ϫ , and NQO1 Ϫ/Ϫ lungs, respectively. Intact lung quinone reduction was evaluated by infusion of DQ (50 M) or CoQ1 (60 M) into the pulmonary arterial inflow of the isolated perfused lung and measurement of pulmonary venous effluent hydroquinone (DQH2 or CoQ 1H2). DQH2 efflux rates for NQO1 ϩ/ϩ , NQO1 ϩ/Ϫ , and NQO1 Ϫ/Ϫ lungs were 0.65 Ϯ 0.08, 0.45 Ϯ 0.04, and 0.13 Ϯ 0.05 (SE) mol · min Ϫ1 · g dry lung Ϫ1 , respectively. DQ reduction in NQO1 ϩ/ϩ lungs was inhibited by 90 Ϯ 4% with dicumarol; there was no inhibition in NQO1 Ϫ/Ϫ lungs. There was no significant difference in CoQ1H2 efflux rates for NQO1 ϩ/ϩ and NQO1 Ϫ/Ϫ lungs. Differences in DQ reduction were not due to differences in lung dry weights, wet-to-dry weight ratios, perfusion pressures, perfused surface areas, or total DQ recoveries. The data provide genetic evidence implicating DQ as a specific NQO1 probe in the perfused rodent lung. knockout mice; lung metabolism; duroquinone; isolated perfused mouse lung; coenzyme Q THE PULMONARY ENDOTHELIUM and lung tissue participate in altering the redox status and disposition of certain redox-active compounds as they pass through the pulmonary circulation (4 -7, 9). This function is carried out by redox enzymes at the pulmonary endothelial surface and within pulmonary endothelial and other lung cells. Since the redox forms of such compounds have different propensities to permeate tissues and carry out pro-and/or antioxidant functions, passage of these compounds through the pulmonary circulation has the potential to influence their dispositions, concentrations, and bioactivities within the pulmonary vessels and lung itself, as well as in downstream organs and vessels.Quinones comprise a class of redox-active compounds having a wide range of physical and chemical properties, and a number of mammalian enzymes have quinone reductase activity (13). Therefore, quinones are useful for probing lung redox processes that affect their disposition in the blood. We have used the amphipathic quinone duroquinone (DQ) as one model compound for st...

Coenzyme Q1as a probe for mitochondrial complex I activity in the intact perfused hyperoxia-exposed wild-type andNqo1-null mouse lung

“…As a step toward developing nondestructive probes for lung redox function, we have relied on the observation that the pulmonary endothelium and lung tissue participate in alteration of the redox status and disposition of redox-active compounds as they pass through the pulmonary circulation (2)(3)(4)(5)(6)36). Such compounds include certain amphipathic quinones, which are reduced to their two-electron reduction products, or hydroquinones, on passage through the lung.…”

“…Such compounds include certain amphipathic quinones, which are reduced to their two-electron reduction products, or hydroquinones, on passage through the lung. Furthermore, quinones with different physical and chemical properties have different propensities to be reduced via specific target quinone reductases within the lung tissue (2,4,6,36). Identification of the quinone reductases involved in this metabolism was initially based on the use of quinone reductase inhibitors.…”

“…Identification of the quinone reductases involved in this metabolism was initially based on the use of quinone reductase inhibitors. However, using Nqo1-null (NQO1 Ϫ/Ϫ ), NQO1 ϩ/Ϫ , and wild-type (NQO1 ϩ/ϩ ) mice, we recently acquired genetic evidence for the specificity of the quinone duroquinone (DQ) as an NAD-(P)H:quinone oxidoreductase 1 (NQO1) electron acceptor on passage through the mouse lung (36).…”

“…In the isolated perfused rat lung, CoQ 1 reduction to its hydroquinone (CoQ 1 H 2 ) is catalyzed by NQO1 and mitochondrial complex I (4). Initial studies in the mouse lung suggest that CoQ 1 reduction capacity was nearly the same on passage through the NQO1 ϩ/ϩ or NQO1 Ϫ/Ϫ mouse lung (36). Given that rat NQO1 is structurally and catalytically different from the mouse (and human) enzyme, this distinction between the two species may not be surprising (24).…”

“…Nqo1-null (NQO1 Ϫ/Ϫ ) breeder mice were obtained from the colony at the National Cancer Institute, and animals were genotyped as previously described (36). The generation of the original Nqo1-null mice is described elsewhere (45).…”