Detection of the production of oxygen-centered free radicals by human neutrophils using spin trapping techniques: a critical perspective

Abstract

Human neutrophils generate oxygen reduction products as a consequence of membrane interactions with a number of stimuli. One oxygen-centered free radical (superoxide) has been unequivocally shown to result from this "respiratory burst," and some experimental evidence for another (hydroxyl radical) has been published, although debate remains as to its significance. The role of phagocyte-derived free radicals in microbicidal and tumoricidal activity as well as tissue damage at sites of inflammation has been the focus of extensive investigation in recent years. Of the techniques available to study free radical generation in biological systems, spin trapping has emerged as a powerful tool for detection and identification of these reactive species, owing in part to its ability to measure production of free radicals inside the phagosome. However, interpretation of resulting spectra is extremely complex and filled with pitfalls and limitations. In this communication we review spin-trapping techniques and discuss the application of this system to the identification of free radicals resulting from stimulation of human neutrophils. Criteria are established which are necessary for definitive identification of superoxide and hydroxyl radical when employing this technology. In this context a critical perspective of previous studies of neutrophil-derived free radicals is offered.