Cytochemical localization of H2O2 in biological tissues.

Department of Pharmacology and Therapeutics, College of Medicine, University of Florida, Gainesville, FL, USA.
Methods in Molecular Biology (Impact Factor: 1.29). 02/2002; 196:3-12. DOI: 10.1385/1-59259-274-0:03
Source: PubMed
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    ABSTRACT: The effects of l-Arg, vitamin C (VC), and vitamin E (VE) on xanthine- (XO) and NAD(P)H-oxidase (NOX) activities, and nitric oxide (NO) availability of hypoxic broilers were evaluated. Chickens were kept in wire cages with free access to feed and water. One-day-old chicks were assigned to 1 of 3 diets: control (CTL; ME 3,200 kcal/kg, CP 23%), high Arg (HA; CTL + Arg 0.8%), or high Arg plus VE and VC (AEC; HA + 200 IU of VE/kg of feed + 500 mg of VC/L of water), and grown under hypobaric hypoxia (HYP) from d 7 to 30. A fourth group of birds was fed the CTL diet and grown under normoxia (CTL-NOR). At d 30, chickens were euthanized, their lungs fixed in vivo, excised, and processed for cyto- and histochemistry. The enzymes XO and NOX were localized and activities assessed histochemically and in lung homogenates. The NO depletion was assessed through nitrotyrosine immunocytochemistry colloidal gold particles (NTY). The XO and NOX localized in cell membranes and within vesicles of pulmonary vessel endothelial cells. The XO activity was higher in CTL-NOR birds (586 ± 43 reflectance units) than in both AEC-HYP (456 ± 39) and HA-HYP birds (394 ± 31), whereas CTL-HYP birds had the lowest XO activity (313 ± 27). The NO depletion was not affected by dietary or hypoxia conditions in clinically healthy birds; nevertheless, hypoxic birds that developed pulmonary hypertension had higher NTY levels (less NO, 145 ± 19) than hypoxic but clinically healthy birds (56 ± 11). Thus, the concurrent supplementation of Arg, VE, and VC restored XO activity without affecting NOX activity or NO availability. The dual role of XO, which produces superoxide and uric acid, may have buffered the effects of superoxide in broiler chickens grown under hypobaric hypoxia.
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