Modulation of nicotinamide adenine dinucleotide phosphate oxidase expression and function by 3',4'-dihydroxyflavonol in phagocytic and vascular cells.
Previously we have demonstrated that 3',4'-dihydroxyflavonol (DiOHF), a novel synthetic flavonol, protects against ischemia reperfusion injury in both heart and brain. In this study, we characterized the pharmacological effects of DiOHF on phagocytic and vascular NADPH oxidase. Superoxide release (lucigenin-enhanced chemiluminescence or cytochrome c reduction), NADPH oxidase activation (membrane translocation of p47phox), and subunit expression (real-time polymerase chain reaction and Western blot) were examined in differentiated HL-60 cells, human neutrophils, vascular endothelial and smooth muscle cells, and mouse aorta. DiOHF concentration dependently suppressed superoxide accumulation (EC(50) = 8.4 +/- 1.7 microM) in vascular smooth muscle cells, which appears to be attributable to its superoxide scavenging activity (EC(50) = 6.1 +/- 1.1 microM measured in a cell-free system). DiOHF had similar effects in HL-60 cells and isolated aortic rings. In HL-60 cells, but not endothelial or smooth muscle cells, DiOHF and quercetin (10 and 30 microM) significantly reduced the protein expression of p47phox, whereas p67phox was not altered. DiOHF did not affect phorbol ester-induced membrane translocation of either p47phox or protein kinase C in leukocytes. Our results suggest that suppression of NADPH oxidase-dependent superoxide accumulation may contribute to the cytoprotective actions of DiOHF during ischemia-reperfusion injury.
Jiang
Fan
F
Bernard O'Brien Institute of Microsurgery, 42 Fitzroy Street, Fitzroy, Victoria 3065, Australia. fjiang@unimelb.edu.au
Guo
Nancy
N
Dusting
Gregory J
GJ
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D001013:Aorta, Thoracic / Q000378:metabolism; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D044948:Flavonols / Q000494:pharmacology*; D018922:HL-60 Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000378:metabolism; D019255:NADPH Oxidases / Q000378:metabolism*; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013481:Superoxides / Q000378:metabolism
D000975:Antioxidants; D044948:Flavonols; D010750:Phosphoproteins; C079789:neutrophil cytosol factor 67K; D013481:Superoxides; D006861:Hydrogen Peroxide; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1