ABSTRACT
Rheumatoid arthritis (RA) patients usually exhibit immune complex (IC) deposition and increased neutrophil activation in the joint. In this study, we assessed how four flavonols (galangin, kaempferol, quercetin, and myricetin) modulate the effector functions of healthy individuals' and active RA patients' IC-stimulated neutrophils. We measured superoxide anion and total reactive oxygen species production using lucigenin (CL-luc)- and luminol (CL-lum)-enhanced chemiluminescence assays, respectively. Galangin, kaempferol, and quercetin inhibited CL-lum to the same degree (mean IC50=2.5 µM). At 2.5 µM, quercetin and galangin suppressed nearly 65% CL-lum of active RA patients' neutrophils. Quercetin inhibited CL-luc the most effectively (IC50=1.71±0.36 µM). The four flavonols diminished myeloperoxidase activity, but they did not decrease NADPH oxidase activity, phagocytosis, microbial killing, or cell viability of neutrophils. The ability of the flavonols to scavenge hypochlorous acid and chloramines, but not H2O2, depended on the hydroxylation degree of the flavonol B-ring. Therefore, at physiologically relevant concentrations, the flavonols partially inhibited the oxidative metabolism of IC-stimulated neutrophils without affecting the other investigated effector functions. Using these compounds to modulate IC-mediated neutrophil activation is a promising safe therapeutic strategy to control inflammation in active RA patients.
Subject(s)
Anti-Inflammatory Agents/pharmacology , Arthritis, Rheumatoid/drug therapy , Flavonoids/pharmacology , Kaempferols/pharmacology , Neutrophils/drug effects , Quercetin/pharmacology , Adult , Anti-Inflammatory Agents/chemistry , Antigen-Antibody Complex/immunology , Cell Degranulation/drug effects , Cells, Cultured , Female , Flavonoids/chemistry , Humans , Kaempferols/chemistry , Middle Aged , Neutrophils/immunology , Oxidation-Reduction/drug effects , Peroxidase/metabolism , Quercetin/chemistry , Reactive Oxygen Species/metabolism , Structure-Activity RelationshipABSTRACT
This article shows how six vehicles interfere in the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) by hypochlorous acid (HOCl) and taurine chloramine in vitro. All the tested vehicles inhibited TMB oxidation by HOCl; dimethyl sulfoxide had a remarkable effect at concentrations as low as 0.00005% (v/v). Cremophor EL and ethanol inhibited TMB oxidation by taurine chloramine at concentrations higher than 0.05 and 25% (v/v), respectively; the other vehicles did not affect this reaction. The results will help to guide the choice of solvent for the TMB oxidation assay performed under viable experimental conditions for evaluation of the HOCl and taurine chloramine scavenging ability of drugs.
