Inhibitory Effects of Metabolite I of Erdosteine on the Generation of Nitric Oxide and Peroxynitrite Chemiluminescence by Human Neutrophils

Abstract

Polymorphonuclear neutrophils (PMNs) can generate superoxide anions and nitric oxide (NO), which is not only an important mediator of various cellular activities, but can also react with superoxide anions to produce peroxynitrite anions (ONOO^–). Peroxynitrite is a potent and potentially toxic oxidant that damages various types of biomolecules. It preferentially mediates the oxidation of thiolic groups in protein and non-protein molecules, thus altering their functions. The aim of this study was to examine whether, in addition to its ability to reduce the respiratory bursts of human PMNs, the SH metabolite I (Met I) of erdosteine, can interfere with NO and NO-derived peroxynitrite production, thus extending its antioxidant activity. This was done by means of the luminol amplified chemiluminescence (LACL), which has been widely used to detect the production of reactive oxidant species (ROS) by PMNs under various conditions. At 5 and 10 µg/ml, Met I significantly reduced LACL after fMLP and PMA stimulation. When L-Arg was added to the reaction medium, as a NO donor, the chemiluminescence of fMLP increased by up to 67% and that of PMA by up to 132%, but was once again significantly reduced by 5 and 10 µg/ml of Met I. In a cell-free system, the use of linsidomine (SIN-1) makes it possible to investigate the behavior of LACL induced by peroxynitrite release, which was significantly reduced by Met I concentrations ranging from 1.25 to 10 µg/ml. Our findings indicate that Met I, a molecule with a SH group, reacts with ROS, NO and NO-derived peroxynitrite, and has both antioxidant and scavenging activity. This is of interest for the strategy of protecting against damage induced by radical species in the pulmonary cell environment, in which they can induce a phlogogenic loop, and suggests that adding exogenous thiols may be useful in antagonizing the toxic effects of reactive molecules on endogenous thiols.