Curcumin and resveratrol act by different ways on NADPH oxidase activity and reactive oxygen species produced by equine neutrophils.
In neutrophils (PMNs), superoxide anion (O2*-), the first reactive oxygen species (ROS) produced to kill pathogenic agents, is generated by NADPH oxidase, an enzymatic complex formed by the translocation of cytosolic subunits to the membrane flavocytochrome b558. In horses, excessive activation of PMNs is often associated with deadly pathologies and the modulation of their ROS production by acting on NADPH oxidase is a prime target to manage inflammation. We developed a cell-free assay to measure the activity of equine NADPH oxidase assembled in vitro, in order to test the effects of natural or synthetic compounds on the enzyme activity or assembly. The cell-free assay was validated with diphenyleneiodonium chloride and Gp91ds-tat, two inhibitors largely described for human NADPH oxidase. The anti-oxidant effects of curcumin and resveratrol at final concentration ranging from 10(-4) to 10(-6) M were studied on whole cells by chemiluminescence (CL) and by cell-free assay, in which the molecule was added before or after the enzyme assembly. The CL assay demonstrated that curcumin efficiently inhibited the O2(-) production and easily entered into PMNs or interacted with their membrane. Cell-free assay showed that curcumin acted on the reconstitution of NADPH oxidase even at 10(-5)M, while resveratrol appeared to be an O2*- scavenger rather than an inhibitor of NADPH oxidase activity, since it acted from outside the cell in CL and after the complex assembly in cell-free assay. By acting directly on NADPH oxidase, curcumin should be a good candidate for the treatment of acute or inflammatory diseases involving an excessive ROS production.
Center for Oxygen, R&D (CORD), Institute of Chemistry, B6a, University of Liège, Allée de la Chimie 3, B-4000 Liège, Belgium. Electronic address: sderochette@ulg.ac.be.
Sandrine
S
Derochette
Thierry
T
Franck
Ange
A
Mouithys-Mickalad
Justine
J
Ceusters
Ginette
G
Deby-Dupont
Jean-Philippe
JP
Lejeune
Philippe
P
Neven
Didier
D
Serteyn
D000818:Animals; D002474:Cell-Free System; D003474:Curcumin / Q000737:chemistry / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology; D006736:Horses; D006801:Humans; D008163:Luminescent Measurements; D019255:NADPH Oxidases / Q000378:metabolism*; D009504:Neutrophils / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D000077185:Resveratrol; D013267:Stilbenes / Q000737:chemistry / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D017382:Reactive Oxygen Species; D013267:Stilbenes; D019255:NADPH Oxidases; D003474:Curcumin; D000077185:Resveratrol
Cell-free assay; Curcumin; Equine NADPH oxidase; Neutrophils; Resveratrol