| Popis: |
Human peripheral blood eosinophils generate superoxide (O2.-) in response to PMA stimulation. These cells are also capable of forming the highly reactive hydroxyl radical (HO.) by a process that is dependent on the presence of active eosinophil peroxidase. To extend this work to tissue-resident cells, we chose to study a murine model of Schistosoma mansoni infection in which parasite ova induce granulomas whose cellular content is 50% eosinophils. In contrast to peritoneal lavage eosinophils, dispersed granuloma cells were unable to reduce ferricytochrome c (as an indicator of O2.-) 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 following PMA stimulation, implying the existence of an inhibitory factor. However, using a 5,5-dimethyl-1-pyrroline-N-oxide spin-trapping system, we were able to demonstrate significant generation of O2.- in response to PMA stimulation, not only in the granuloma cells, but also in the conditioned medium-treated neutrophils, demonstrating that the inhibitory factor was not affecting O2.- generation, but rather was interfering with ferricytochrome c reduction. In addition, using an alpha-(4-pyridyl-1-oxide)-N-tert-butyl-nitrone/ethanol spin-trapping system, we were able to detect HO. formation by these same cells following PMA stimulation. This HO. formation was inhibited by superoxide dismutase, azide, and thiocyanide, and NaSCN, consistent with a mechanism requiring O2.- and enzymatic peroxidase activity. These results demonstrate that tissue eosinophils associated with the schistosome-induced granuloma have the ability to form both O2.- and HO., and point out potential problems associated with the measurement of O2.- in whole tissue preparations. |
