EOSINOPHILS FROM SCHISTOSOME-INDUCED HEPATIC GRANULOMAS PRODUCE SUPEROXIDE AND HYDROXYL RADICAL

Human peripheral blood eosinophils generate superoxide (O-2) in respon se to PMA stimulation. These cells are also capable of forming the hig hly reactive hydroxyl radical (HO) by a process that is dependent on t he presence of active eosinophil peroxidase. To extend this work to ti ssue-resident cells, we chose to study a murine model of Schistosoma m ansoni infection in which parasite ova induce granulomas whose cellula r content is 50% eosinophils. In contrast to peritoneal lavage eosinop hils, dispersed granuloma cells were unable to reduce ferricytochrome c (as an indicator of O-2) in response to PMA stimulation. Furthermore , when human neutrophils were pretreated with conditioned medium from the granuloma cells, they also failed to reduce ferricytochrome c foll owing PMA stimulation, implying the existence of an inhibitory factor. However, using a 5,5-dimethyl-1-pyrroline-N-oxide spin-trapping syste m, we were able to demonstrate significant generation of O-2 in respon se to PMA stimulation, not only in the granuloma cells, but also in th e conditioned medium-treated neutrophils, demonstrating that the inhib itory factor was not affecting O-2 generation, but rather was interfer ing with ferricytochrome c reduction. In addition, using an a-(4-pyrid yl-1-oxide)-N-tert-butyl-nitrone/ethanol spin-trapping system, we were able to detect HO formation by these same cells following PMA stimula tion. This HO formation was inhibited by superoxide dismutase, azide, and thiocyanide, and NaSCN, consistent with a mechanism requiring O-2 and enzymatic peroxidase activity. These results demonstrate that tiss ue eosinophils associated with the schistosome-induced granuloma have the ability to form both O-2 and HO; and point out potential problems associated with the measurement of O-2 in whole tissue preparations.