Neutrophil-derived matrix metalloproteinase-9 is increased in severe asthma and poorly inhibited by glucocorticoids

doi:10.1016/j.jaci.2003.08.013

Journal of Allergy and Clinical Immunology

Volume 112, Issue 6, December 2003, Pages 1064-1071

https://doi.org/10.1016/j.jaci.2003.08.013 Get rights and content

Abstract

Background

Matrix metalloproteinase (MMP)-9 levels are increased in bronchoalveolar lavage (BAL) fluid from patients with severe asthma on high doses of glucocorticoids (GCs).

Objective

We sought to identify neutrophils as the source of increased BAL fluid MMP-9 in severe asthma and to evaluate the effects of GCs on this MMP-9.

Methods

MMP-9 protein, activity, and mRNA were measured in BAL fluid and cells at baseline, and after in vitro GCs in patients with severe asthma and controls using enzyme immunoassays, zymography, Western blotting, and real-time PCR.

Results

The high molecular weight (HMW) form of MMP-9 was significantly increased in severe asthma (P = .02). Western blotting confirmed a heterodimer of MMP-9 and neutrophil gelatinase–associated lipocalin. The HMW MMP-9 correlated with BAL neutrophils (r = .65, P < .0001). BAL cell supernatant MMP-9 protein levels also tended to be higher in patients with severe asthma (overall, P = .09), whereas the HMW activity form was increased (P = .03). MMP-9 protein (and HMW activity) correlated with neutrophils in the cell pellet (r = .75, P < .0001). In contrast to protein and activity, BAL cell mRNA levels were marginally lower in patients with severe asthma than in control subjects (overall, P = .06). Although GCs decreased BAL cell MMP-9 protein and mRNA in vitro, the effect was significantly smaller in severe asthma (P < .01 for both). GCs decreased the pro–MMP-9 activity in patients with severe asthma and normal control subjects, while having no effects on the HMW form (P = .22). Peripheral blood neutrophil MMP-9 protein was not affected by GCs.

Conclusions

BAL neutrophils contribute to BAL fluid MMP-9 protein and activity and are poorly inhibited by GCs.

Section snippets

Subjects

Subjects with asthma were divided into severity categories by use of previous criteria.¹³ Patients with severe asthma were all referred for severe asthma. Despite treatment with oral and high-dose inhaled steroids, with at least 1 other controller, they had symptoms and used urgent health care. Normal control subjects had normal pulmonary function and no history of respiratory illness. The study was approved by the National Jewish Institutional Review Board, and all subjects gave informed

Subject characteristics

Thirty-nine patients with severe asthma, 9 patients with moderate asthma, 13 patients with mild asthma, and 17 normal control subjects underwent bronchoscopy with BAL for evaluation of cell counts, differentials, and MMP-9 levels (Table I). Subgroups were involved in studies of MMP-9 production and activity as described in the individual sections. As reported previously, MMP-9 protein levels were highest in the severe asthmatics (overall, P = .001). Similarly, the group of patients with severe

Discussion

This study suggests that the increased MMP-9 protein in BAL fluid from patients with severe asthma is derived primarily from neutrophils, with minor contribution from monocyte/macrophages or other cells. Although it has been suggested that increases in MMP-9 after allergen challenge might be neutrophil derived, this is the first study to detail the relationship of BAL fluid MMP-9 to neutrophils and disease severity.18, 19 In addition, in severe asthma, a significantly higher amount of the MMP-9

Acknowledgements

We thank Dr Niels Borregaard for the initial supply of NGAL antibody.

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^☆

Supported by NIH grants HL-64087, AI-40600, RR-00051 and ALA of Colo-rado, Oklahoma, and Alaska.

The first 2 authors contributed equally to the development of this manuscript.

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Asthma, Rhinitis, Other Respiratory DiseasesNeutrophil-derived matrix metalloproteinase-9 is increased in severe asthma and poorly inhibited by glucocorticoids☆

Abstract

Background

Objective

Methods

Results

Conclusions

Section snippets

Subjects

Subject characteristics

Discussion

Acknowledgements

J Biol Chem

J Allergy Clin Immunol

J Allergy Clin Immunol

J Allergy Clin Immunol

Anal Biochem

Chest

Blood

J Allergy Clin Immunol

Bronchoscopic evaluation of severe asthma: Persistent inflammation associated with high dose glucocorticoids

Am J Respir Crit Care Med

Neutrophilic inflammation in severe persistent asthma

Am J Respir Crit Care Med

The relationship between airways inflammation and asthma severity

Am J Respir Crit Care Med

Glucocorticoid treatment inhibits apoptosis in human neutrophils

Separation of survival and activation outcomes. J Immunol

The human neutrophil lipocalin supports the allosteric activation of matrix metalloproteinases

Eur J Biochem

Sputum metalloproteinase-9/tissue inhibitor of metalloproteinase-1 ratio correlates with airflow obstruction in asthma and chronic bronchitis

Am J Respir Crit Care Med

Asthma, Rhinitis, Other Respiratory Diseases
Neutrophil-derived matrix metalloproteinase-9 is increased in severe asthma and poorly inhibited by glucocorticoids☆