Document Detail

Evidence for NAD(P)H:quinone oxidoreductase 1 (NQO1)-mediated quinone-dependent redox cycling via plasma membrane electron transport: A sensitive cellular assay for NQO1.
MedLine Citation:
PMID:  19932748     Owner:  NLM     Status:  MEDLINE    
2,3-Dimethoxy 1,4-naphthoquinone (DMNQ), which redox cycles via two-electron reduction, mediates reduction of the cell-impermeative tetrazolium dye WST-1 in kidney epithelial cells (MDCK), which express high levels of NQO1, but not in HL60 or CHO cells, which are NQO1 deficient. DMNQ-dependent WST-1 reduction by MDCK cells was strongly inhibited by low concentrations of the NQO1 inhibitor dicoumarol and was also inhibited by diphenyleneiodonium, capsaicin, and superoxide dismutase (SOD), but not by the uncoupler FCCP or the complex IV inhibitor cyanide. This suggests that DMNQ-dependent WST-1 reduction by MDCK cells is catalyzed by NQO1 via redox cycling and plasma membrane electron transport (PMET). Interestingly, we observed an association between DMNQ/WST-1 reduction and extracellular H(2)O(2) production as determined by Amplex red. Exposure of MDCK cells to DMNQ for 48 h caused cellular toxicity that was extensively reversed by co-incubation with dicoumarol or exogenous SOD, catalase, or N-acetylcysteine. No effects were observed in NQO1-deficient CHO and HL60 cells. In conclusion, we have developed a simple real-time cellular assay for NQO1 and show that PMET plays a significant role in DMNQ redox cycling via NQO1, leading to cellular toxicity in cells with high NQO1 levels.
An S Tan; Michael V Berridge
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2009-11-22
Journal Detail:
Title:  Free radical biology & medicine     Volume:  48     ISSN:  1873-4596     ISO Abbreviation:  Free Radic. Biol. Med.     Publication Date:  2010 Feb 
Date Detail:
Created Date:  2010-02-03     Completed Date:  2010-04-13     Revised Date:  2010-09-07    
Medline Journal Info:
Nlm Unique ID:  8709159     Medline TA:  Free Radic Biol Med     Country:  United States    
Other Details:
Languages:  eng     Pagination:  421-9     Citation Subset:  IM    
Copyright Information:
Copyright 2009 Elsevier Inc. All rights reserved.
Malaghan Institute of Medical Research, Wellington, New Zealand.
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MeSH Terms
Biological Assay
Blotting, Western
Cell Membrane / drug effects*,  enzymology*
Cell Proliferation / drug effects
Cells, Cultured
Dicumarol / pharmacology
Electron Transport*
Enzyme Inhibitors / pharmacology
Hela Cells
Hydrogen Peroxide / metabolism
Kidney / cytology,  drug effects,  metabolism
NAD(P)H Dehydrogenase (Quinone) / antagonists & inhibitors,  metabolism*
Oxidants / metabolism
Quinones / pharmacology*
Superoxides / metabolism
Reg. No./Substance:
0/Enzyme Inhibitors; 0/Oxidants; 0/Quinones; 11062-77-4/Superoxides; 66-76-2/Dicumarol; 7722-84-1/Hydrogen Peroxide; EC Dehydrogenase (Quinone)
Erratum In:
Free Radic Biol Med. 2010 Apr 1;48(7):978

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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