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Characterization of the threshold for NAD(P)H:quinone oxidoreductase activity in intact sulforaphane-treated pulmonary arterial endothelial cells. Free Radic Biol Med 2011 Apr 15;50(8):953-62

Date

01/18/2011

Pubmed ID

21238579

Pubmed Central ID

PMC3851029

DOI

10.1016/j.freeradbiomed.2011.01.009

Scopus ID

2-s2.0-79952443629 (requires institutional sign-in at Scopus site)   10 Citations

Abstract

Treatment of bovine pulmonary arterial endothelial cells in culture with the phase II enzyme inducer sulforaphane (5μM, 24h; sulf-treated) increased cell-lysate NAD(P)H:quinone oxidoreductase (NQO1) activity by 5.7 ± 0.6 (mean ± SEM)-fold, but intact-cell NQO1 activity by only 2.8 ± 0.1-fold compared to control cells. To evaluate the hypothesis that the threshold for sulforaphane-induced intact-cell NQO1 activity reflects a limitation in the capacity to supply NADPH at a sufficient rate to drive all the induced NQO1 to its maximum activity, total KOH-extractable pyridine nucleotides were measured in cells treated with duroquinone to stimulate maximal NQO1 activity. NQO1 activation increased NADP(+) in control and sulf-treated cells, with the effect more pronounced in the sulf-treated cells, in which the NADPH was also decreased. Glucose-6-phosphate dehydrogenase (G-6-PDH) inhibition partially blocked NQO1 activity in control and sulf-treated cells, but G-6-PDH overexpression via transient transfection with the human cDNA alleviated neither the restriction on intact sulf-treated cell NQO1 activity nor the impact on the NADPH/NADP(+) ratios. Intracellular ATP levels were not affected by NQO1 activation in control or sulf-treated cells. An increased dependence on extracellular glucose and a rightward shift in the K(m) for extracellular glucose were observed in NQO1-stimulated sulf-treated vs control cells. The data suggest that glucose transport in the sulf-treated cells may be insufficient to support the increased metabolic demand for pentose phosphate pathway-generated NADPH as an explanation for the NQO1 threshold.

Author List

Bongard RD, Krenz GS, Gastonguay AJ, Williams CL, Lindemer BJ, Merker MP

Author

Carol L. Williams PhD Professor in the Pharmacology and Toxicology department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Animals
Blotting, Western
Cattle
Endothelium, Vascular
Isothiocyanates
NAD(P)H Dehydrogenase (Quinone)
Pulmonary Artery
Sulfoxides
Thiocyanates