Mechanisms of Allergy
p38 Mitogen-activated protein kinase–induced glucocorticoid receptor phosphorylation reduces its activity: Role in steroid-insensitive asthma,☆☆,

https://doi.org/10.1067/mai.2002.122465Get rights and content

Abstract

Background: Although glucocorticoids are the most effective treatment for chronic inflammatory diseases, such as asthma, some patients show a poor response. IL-2 combined with IL-4 can alter glucocorticoid receptor (GR) ligand-binding affinity and modulate glucocorticoid function. Objective: We sought to confirm the altered ligand-binding affinity in a distinct group of steroid-dependent asthmatic subjects and examine the mechanism by which IL-2 and IL-4 modify the ligand-binding affinity of the GR. Methods: We examined PBMCs from healthy subjects, subjects with mild asthma, and steroid-dependent subjects with severe asthma using dexamethasone-binding assays and Western blot analysis of GR and phosphorylated activated transcription factor 2 expression. GR phosphorylation was measured after orthophosphate labeling and immunoprecipitation and cytokine production by means of ELISA. Results: GR ligand-binding affinity was reduced in the nucleus but not in the cytoplasm of steroid-dependent asthmatic subjects compared with that seen in healthy subjects (dissociation constant, 39.8 ± 4.6 vs 6.79 ± 0.8 nmol/L). This difference in ligand-binding affinity could be mimicked by IL-2 and IL-4 cotreatment and was blocked by the p38 mitogen-activated kinase (MAPK) inhibitor SB203580. Activation of p38 MAPK by IL-2 and IL-4, as shown by means of phosphorylation of activated transcription factor 2, resulted in GR phosphorylation and reduced dexamethasone repression of LPS-stimulated GM-CSF release. p38 MAPK phosphorylation of CD2+ T cells occurred on serine residues. The ability of dexamethasone to modulate IL-10 release was also inhibited by IL-2 and IL-4 cotreatment. These effects were also inhibited by SB203580. Conclusion: These data show that p38 MAPK inhibitors may have potential in reversing glucocorticoid insensitivity and reestablishing the beneficial effects of glucocorticoids in patients with severe asthma. (J Allergy Clin Immunol 2002;109:649-57.)

Section snippets

Patients

Nine asthmatic subjects whose symptoms were controlled with ICSs (0.8 μg/d [95% CI, 0.2-1.3 μg/d], mean fluticasone equivalents), 19 steroid-dependent asthmatic subjects (poorly controlled on inhaled and oral steroids), and 6 healthy nonasthmatic subjects were studied. All subjects were matched in terms of age and atopic status (Table I).

. Characteristics of asthmatic and healthy subjects

Empty CellSteroid-dependent subjectsICS subjectsHealthy subjects
No. of subjects1996
Age (y), mean (95% CI)46 (40-52)48

GR ligand binding of PBMCs

There was no significant difference in the cell profile of the PBMCs isolated from any subject group (data not shown). The ability of dexamethasone to bind to GR was decreased in the nucleus in subjects with steroid-depend-ent asthma. Steroid-dependent subjects (Kd, 39.8 ± 4.6 nmol/L) had a greater Kd compared with those with ICS-controlled asthma symptoms (Kd, 25 ± 0.9 nmol/L; P < .05) or healthy subjects (Kd, 6.79 ± 0.8 nmol/L; P < .001). In addition, there was a significant difference in Kd

Discussion

The ability of dexamethasone to bind to GR was decreased according to the degree of asthma severity. This difference was detected only in the nucleus and not in the cytoplasm, possibly reflecting an effect of a nuclear protein masking the GR ligand-binding site or in an altered conformation of the activated GR. This altered affinity of dexamethasone for GR may reflect either an intrinsic defect in the GR within these patients or may relate to changes in the receptor induced by the increased

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    *These authors contributed equally to this work.

    ☆☆

    Supported by the Wellcome Trust, the National Asthma Campaign, the Clinical Research Committee (Brompton Hospital), and Glaxo-Wellcome. E.I. is supported by the South African Pulmonology Society through a Grant from Boehringer Ingelheim (S.A.).

    Reprint requests: Ian M. Adcock, PhD, Thoracic Medicine, Imperial College School of Medicine, National Heart and Lung Institute, Dovehouse St, London SW3 6LY, UK.

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