Pharmacological and immunohistochemical evidence for a functional nitric oxide synthase system in rat peritoneal eosinophils

Zanardo, R C O; Costa, Esther Da; Ferreira, Heloisa H.A.; Antunes, Edson; Martins, Antônio Roberto; Murad, Ferid; Nucci, Gilberto De

doi:10.1073/pnas.94.25.14111

Cited by 43 publications

(31 citation statements)

References 32 publications

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“…Previous reports in rat peritoneal eosinophils and rat models of sephadex-induced airway inflammation not only indicated the presence of cNOS and iNOS isoforms in them, but their vital role in eosinophil locomotion by acting through a cyclic GMP transduction mechanism (Zanardo et al 1997) and the role of iNOS in the airway inflammatory responses in allergic inflammation (Birrell et al 2003). NOS has been depicted to play pivotal role in the eosinophil infiltration in airways of actively sensitized rats with ovalbumin, and its importance in in vivo eosinophil migration in both allergic (Ferreira et al 1998) and non-allergic (Ferreira et al 1996) subjects has been demonstrated.…”

Section: Introductionmentioning

confidence: 99%

Ultrastructural immunogold localization of nitric oxide synthase isoforms in rat and human eosinophils

et al. 2010

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The involvement of nitric oxide (NO) as both pro and anti-inflammatory agent in allergic, airway inflammatory, and asthmatic diseases and the active participation of eosinophils in such ailments have been previously suggested. NO modulates eosinophil number, migration and their survival. The microenvironment of NO synthase (NOS) in subcellular organelles determines its rate and efficiency of catalysis, which in turn influences NO generation at distinct intracellular locales. The present study was undertaken to assess the intracellular distribution of NOS isoforms by transmission electron microscopy followed by morphometric analysis in human and rat eosinophils. Rat eosinophils were explored in parallel, and since they are widely used as model systems to mimic human diseases, a comparative study on NOS localization patterns might provide useful information in deciphering NO role in diverse aspects of eosinophil-related inflammatory ailments. The results demonstrated predominance of neuronal NOS (nNOS) in the eosinophilic granules and even distribution of inducible NOS (iNOS) and nNOS in the cytoplasm and nucleus of human eosinophils. In rat eosinophils, however, iNOS was mainly localized in the eosinophilic granules and nucleus, while nNOS was distributed evenly in cytoplasm and nucleus. Distribution of endothelial NOS (eNOS) in eosinophils was scanty. Differences in NOS isoforms and their localization in human and rat cells might have implications in differential mode of catalysis and functional contribution to eosinophil physiology and pathology, warranting detailed investigations. The present study highlights species-specific differences in the relative abundance and distribution pattern of NOS isoforms in rat and human eosinophils, which should be considered cautiously in interpreting the rat data to humans.

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Section: Introductionmentioning

confidence: 99%

Ultrastructural immunogold localization of nitric oxide synthase isoforms in rat and human eosinophils

et al. 2010

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“…The elimination of RNS had no effect on airway eosinophilia in guinea pigs or mice. There have been several reports that nonspecific NOS inhibitors inhibited eosinophil accumulation in the airways and eosinophil activities [20,21]. In other studies, iNOS-specific inhibitor inhibited allergen-induced airway eosinophilia [22,23].…”

Section: Discussionmentioning

confidence: 94%

iNOS depletion completely diminishes reactive nitrogen-species formation after an allergic response

Koarai¹,

Ichinose²,

Sugiura³

et al. 2002

European Respiratory Journal

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Nitric oxide (NO) shows proinflammatory actions mainly via reactive nitrogen species (RNS) formation through superoxide-and peroxidase-dependent mechanisms. The purpose of this study was to examine the role of inducible NO synthase (iNOS) in RNS production, airway hyperresponsiveness, and inflammation after allergen challenge.Ovalbumin (OVA)-sensitised, iNOS-deficient and wild-type mice were used. RNS production was assessed by nitrotyrosine (NT) immunoreactivity in the airways. Airway inflammation and responsiveness were evaluated by eosinophil accumulation and methacholine (i.v.) challenge, respectively.In wild-type mice, OVA-inhalation challenge increased iNOS immunoreactivity in airway epithelial cells as well as iNOS protein measured by Western blotting. The total amounts of nitrite and nitrate in bronchoalveolar lavage (BAL) fluid were increased, and NT immunoreactivity was also observed abundantly in airway inflammatory cells. In iNOS-deficient mice, both iNOS expression and NT formation were completely abolished, and the total amounts of nitrite and nitrate in BAL fluid were significantly decreased. In contrast, OVA-induced airway eosinophil recruitment and hyperresponsiveness were observed almost equally in wild-type and iNOS-deficient mice.These data suggest that reactive nitrogen species production after allergic reaction occurs totally via inducible nitric oxide synthase-dependent pathways. Allergenmediated airway eosinophil recruitment and hyperresponsiveness appear to be independent of reactive nitrogen species production. Eur Respir J 2002; 20: 609-616.

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“…nNOS, eNOS, and iNOS types. eNOS and iNOS isoforms have been observed in rat peritoneal eosinophils where they are strongly expressed and localized in cytoplasmic granules (48). In human peripheral blood eosinophils, iNOS (49) and eNOS (50) have been detected.…”

Section: Discussionmentioning

confidence: 99%

Post-translational Tyrosine Nitration of Eosinophil Granule Toxins Mediated by Eosinophil Peroxidase

Ulrich

Petre

Youhnovski

et al. 2008

Journal of Biological Chemistry

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Nitration of tyrosine residues has been observed during various acute and chronic inflammatory diseases. However, the mechanism of tyrosine nitration and the nature of the proteins that become tyrosine nitrated during inflammation remain unclear. Here we show that eosinophils but not other cell types including neutrophils contain nitrotyrosine-positive proteins in specific granules. Furthermore, we demonstrate that the human eosinophil toxins, eosinophil peroxidase (EPO), major basic protein, eosinophil-derived neurotoxin (EDN) and eosinophil cationic protein (ECP), and the respective murine toxins, are post-translationally modified by nitration at tyrosine residues during cell maturation. High resolution affinity-mass spectrometry identified specific single nitration sites at Tyr 349 in EPO and Tyr 33 in both ECP and EDN. ECP and EDN crystal structures revealed and EPO structure modeling suggested that the nitrated tyrosine residues in the toxins are surface exposed. Studies in EPO ؊/؊ , gp91 phox؊/؊ , and NOS ؊/؊ mice revealed that tyrosine nitration of these toxins is mediated by EPO in the presence of hydrogen peroxide and minute amounts of NOx. Tyrosine nitration of eosinophil granule toxins occurs during maturation of eosinophils, independent of inflammation. These results provide evidence that post-translational tyrosine nitration is unique to eosinophils.Human eosinophils are bone marrow-derived, non-dividing granulocytes of the innate immune system, which store the highly cationic proteins eosinophil peroxidase (EPO), 3 major basic protein (MBP), eosinophil-derived neurotoxin (EDN), and eosinophil cationic protein (ECP) in secondary granules (1, 2). In rodents, eosinophil proteins with similar structure and activity have been identified (3-6). In response to allergen provocation or parasitic infection, expanded eosinophil populations from the bone marrow are selectively recruited to affected tissues (1). The release of cationic toxins from activated eosinophils by degranulation is regarded as a dominant effector function of these cells, mediating lysis of helminths and protozoae (7-10). The positive net charge conveys tight toxin binding to negatively charged cell surfaces where EPO causes oxidation of membrane components in the presence of hydrogen peroxide (H 2 O 2 ) (8). MBP increases membrane permeability and perturbation following insertion of apolar residues into the membrane (11), whereas ECP creates membrane channels (12). Both EDN and ECP exert ribonuclease activity (13). Due to their unspecific binding to membranes and their cytolytic activities, eosinophil granule proteins also cause host tissue damage during parasite infections and inflammatory disorders such as allergic asthma (14 -16). Despite advances in elucidating the mechanisms of action for the eosinophil granule proteins (1, 2), the cationic nature of these proteins (pI values Ͼ 10) would predict electrostatic repulsion and exclude interaction/cooperation of single granule proteins and among different granule proteins. Because sequen...

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Pharmacological and immunohistochemical evidence for a functional nitric oxide synthase system in rat peritoneal eosinophils

Cited by 43 publications

References 32 publications

Ultrastructural immunogold localization of nitric oxide synthase isoforms in rat and human eosinophils

Ultrastructural immunogold localization of nitric oxide synthase isoforms in rat and human eosinophils

iNOS depletion completely diminishes reactive nitrogen-species formation after an allergic response

Post-translational Tyrosine Nitration of Eosinophil Granule Toxins Mediated by Eosinophil Peroxidase

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