MMP-9 In Bronchoalveolar Lavage Fluid Research Articles

TMT-based quantitative proteomics revealed protective efficacy of Icariside II against airway inflammation and remodeling via inhibiting LAMP2, CTSD and CTSS expression in OVA-induced chronic asthma mice

BackgroundAsthma is a chronic inflammatory disorder in airways with typical pathologic features of airflow limitation, airway inflammation and remodeling. Icariside II (IS), derived from herbal medicine Herba Epimedii, exerts an anti-inflammatory property. However, underlying mechanisms with specifically targeted molecular expression by IS in asthma have not been fully understood, and whether IS could inhibit remodeling and EMT still remains unclear. PurposeThe study aimed to clarify therapeutic efficacy of IS for attenuating airway inflammation and remodeling in asthma, and illustrate IS-regulated specific pathway and target proteins through TMT-based quantitative proteomics. Study design and methodsMurine model of chronic asthma was constructed with ovalbumin (OVA) sensitization and then challenge for 8 weeks. Pulmonary function, leukocyte count in bronchoalveolar lavage fluid (BALF), lung histopathology, inflammatory and fibrotic cytokines, and markers of epithelial-mesenchymal transition (EMT) were evaluated. TMT-based quantitative proteomics were performed on lung tissues to explore IS-regulated proteins. ResultsIS contributed to alleviative airway hyperresponsiveness (AHR) evidenced by declined RL and increased Cdyn. After IS treatment, we observed a remarked down-regulation of leukocyte count, inflammatory cytokines in BALF, and peribronchial inflammation infiltration. Goblet cell hyperplasia, mucus secretion and peribronchial collagen deposition were attenuated, with the level of TGF-β and MMP-9 in BALF declined. Furthermore, IS induced a rise of Occludin and E-cadherin and a decline of N-cadherin and α-SMA in lung tissues. These results proved the protective property of IS against airway inflammation, remodeling and EMT. To further investigate underlying mechanisms of IS in asthma treatment, TMT-based quantitative proteomics were performed and 102 overlapped DEPs regulated by IS were identified. KEGG enrichment exhibited these DEPs were enriched in lysosome, phagosome and autophagy, in which LAMP2, CTSD and CTSS were common DEPs. WB, q-PCR and IHC results proofed expressional alteration of these proteins. Besides, IS could decrease Beclin-1 and LC3B expression with increasing p62 expression thus inhibiting autophagy. ConclusionsThe study demonstrated IS could ameliorate AHR, airway inflammation, remodeling and EMT in OVA-induced chronic asthma mice. Our research was the first to reveal that inhibition of LAMP2, CTSD and CTSS expression in autophagy contributed to the therapeutic efficacy of IS to asthma.

Anti-thymic stromal lymphopoietin antibody suppresses airway remodeling in asthma through reduction of MMP and CTGF.

Thymic stromal lymphopoietin (TSLP) mediates immune reaction in patients with asthma. Matrix metalloproteinase (MMP), connective tissue growth factor (CTGF), and transforming growth factor-β (TGF-β) are inflammatory mediators whose responses to the anti-TSLP antibody are unknown. This study examined the effect of an anti-TSLP antibody on MMP, CTGF, TGF-β, and airway structural changes in airway remodeling in asthma. Mice were randomly divided into phosphate-buffered-saline-challenged (PBS), ovalbumin-challenged (OVA), and ovalbumin-challenged with anti-TSLP antibody (OVA + anti-TSLP) groups. Airway responsiveness and serum ovalbumin-specific immunoglobulin E were measured. Differential cell counts and MMP-2 and MMP-9 were evaluated in bronchoalveolar lavage fluid (BALF). Airway structural changes were quantified using morphometric analysis and presentation by immunohistochemistry staining. Lung CTGF, TGF-β, and TSLP were analyzed using western blot. Airway responsiveness was significantly lower in OVA + anti-TSLP and PBS groups than in OVA group. Airway structural changes exhibited less smooth muscle thickness in OVA + anti-TSLP and PBS groups than in OVA group. MMP-2 and MMP-9 in BALF and CTGF, TGF-β, and TSLP in lungs significantly decreased in OVA + anti-TSLP and PBS groups compared with OVA group. Anti-TSLP antibody exerts the preventive effect of decreasing airway structural changes through reduction of MMP, TGF-β, and CTGF in airway remodeling of asthma.

Elevated IL-33 promotes expression of MMP2 and MMP9 via activating STAT3 in alveolar macrophages during LPS-induced acute lung injury

BackgroundPulmonary inflammation and endothelial barrier permeability increase in acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) induced by pro-inflammatory cytokines and matrix metalloproteinases (MMPs). However, the relationship between pro-inflammatory cytokines and MMPs in ALI/ARDS remains poorly understood.MethodsA lipopolysaccharide (LPS)-induced ALI rat model was established through intratracheal instillation. The wet/dry ratios of lung tissues were measured, and bronchoalveolar lavage fluid (BALF) was collected to test protein concentrations, total cell/macrophage numbers, and pro-inflammatory cytokine levels. LPS-treated alveolar macrophages were utilized in in vitro experiments. The expression and secretion of MMPs were respectively detected using quantitative PCR, Western blotting and ELISA assays.ResultsThe levels of IL-33 and MMP2/9 in BALF increased in all the ALI rats with severe lung injury. LPS-induced IL-33 autocrine upregulated the expression of MMP2 and MMP9 through activating STAT3. Neutralizing IL-33 in culture medium with specific antibodies suppressed the expression and secretion of MMP2 and MMP9 in LPS-treated alveolar macrophages. Consistently, eliminating IL-33 decreased the levels of MMP2 and MMP9 in BALF and alleviated lung injury in ALI rats.ConclusionThe IL-33/STAT3/MMP2/9 regulatory pathway is activated in alveolar macrophages during acute lung injury, which may exacerbate the pulmonary inflammation.

Progesterone attenuates airway remodeling and glucocorticoid resistance in a murine model of exposing to ozone

Airway remodeling is a vital component of chronic obstructive pulmonary disease (COPD). Despite the broad anti-inflammation effects of glucocorticoids, they exhibit relatively little therapeutic benefit in COPD, indicating the accelerating demands of new agents for COPD. We aim to explore the effect of progesterone on airway remodeling in a murine modeling of exposing to ozone and to further examine the potential effect of progesterone on glucocorticoid insensitivity. C57/BL6 mice were exposed to ozone for 12 times over 6 weeks, and were administered with progesterone alone or combined with budesonide (BUD) after each exposure until the 10th week. The peribronchial collagen deposition was measured. The protein levels of MMP8 and MMP9 in bronchoalveolar lavage fluid (BALF) and lungs were assessed. Western blot analysis was used to detect the levels of hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF), a-smooth muscle actin (α-SMA), glycogen synthase kinase-3β (GSK-3β). The expression of VEGF and histone deacetylase 2 (HDAC2) in the lung were determined by Immunohistochemical analyses. We observe that progesterone attenuates the peribronchial collagen deposition, as well as the expression of MMP8, MMP9, HIF-1α, VEGF, α-SMA, and GSK-3β in BALF or lung tissues. Progesterone or BUD monotherapy has no effect on HDAC2 production. Progesterone combines with BUD induce dramatically enhanced effects. Thus, these results demonstrate novel roles of progesterone for the pathogenesis and airway remodeling in COPD. Progesterone plus BUD administration exerts more significant inhibition on airway remodeling with dose-independent. Additionally, progesterone may, to some extent, improve the glucocorticoid insensitivity.

Urban particulate matter triggers lung inflammation via the ROS-MAPK-NF-κB signaling pathway.

Particulate matter (PM) is a high risk factor for various respiratory diseases and triggers an inflammatory response in lung tissues. However, the molecular mechanism of the PM-induced inflammatory response is incompletely understood. Human bronchial epithelial cells (HBECs) were treated with the urban PM 1649b for assessment of the inflammatory response. The intracellular level of reactive oxygen species (ROS) was measured by flow cytometry. PM-activated signaling pathways were addressed with specific inhibitors. In vivo, the C57 mice model of PM-induced acute lung inflammation was established with intratracheal instillation of PM for 2 consecutive days. The oxidant stress in lung tissues was assessed with dihydroethidium (DHE) staining, and malondialdehyde (MDA) activity and hydrogen peroxide (H2O2) assays. The histopathologic changes in lung tissues and number of inflammatory cells in bronchoalveolar lavage fluid (BALF) were examined. Expression of pro-inflammatory cytokines in BALF was measured by ELISA. PM increased the expression of interleukin (IL)-1β, IL-6, IL-8, matrix metalloproteinase (MMP)-9 and cyclooxygenase (COX)-2 in a dose-dependent manner. ROS generation and activation of MAPK (ERK, JNK, p38 MAPK) and NF-κB pathways were detected in PM-exposed HBECs. Pretreatment with N-acetylcysteine (NAC) led to the inflammatory response, ROS level and activation of the MAPK and NF-κB pathways to be attenuated. Blockade of ERK, JNK or p38 MAPK pathway with specific inhibitor prevented the release of pro-inflammatory cytokines and activation of the NF-κB pathway. Inhibition of the NF-κB pathway reduced the expression of pro-inflammatory cytokines. In vivo, PM exposure increased oxidant stress in lung tissues, infiltration of inflammatory cells around PM in lung tissues, the number of total cells and inflammatory cells in BALF, and the concentrations of IL-1β, IL-6, IL-8 and MMP-9 in BALF, all of which were reversed partially upon NAC treatment. PM exposure enhanced the airway inflammatory response significantly through ROS-mediated activation of MAPK (ERK, JNK, p38 MAPK) and downstream NF-κB signaling pathways. Oxidative stress appeared to be the key regulator for PM-induced lung inflammation. These results suggested the molecular mechanism of lung inflammation caused by PM.

Reversal of TREM-1 ectodomain shedding and improved bacterial clearance by intranasal metalloproteinase inhibitors

Triggering receptor expressed on myeloid cells-1 (TREM-1) is expressed on neutrophils and monocyte/macrophages and amplifies Toll-like receptor-mediated inflammation during infection. TREM-1 also exists in an antagonistic soluble form (sTREM-1) that has been used as a peripheral biomarker in sepsis, though the mechanisms of its release are not entirely clear. The requirement of TREM-1 in single microbial infections is controversial, with some studies showing a protective role and others a contribution to immunopathology. Furthermore, the role of membrane-bound and sTREM-1 in polygenic infections is currently unknown. In a mouse co-infection model where preceding viral infection greatly enhances bacteria co-infection, we now determine a mechanisms for the striking increase in sTREM-1 and the loss of TREM-1 on surface of neutrophils. We identified a matrix metalloproteinase (MMP)-9 cleavage site in TREM-1 and that the increase of MMP-9 in bronchoalveolar lavage fluid mirrors sTREM-1 release. In vitro studies with neutrophils and MMP-9 and the reduction of sTREM-1 in vivo after MMP-9 inhibition verifies that this enzyme cleaves TREM-1. Intriguingly, MMP-9 inhibition significantly reduces bacterial load and ensuing immunopathology in a co-infection model. This highlights MMP-9 inhibition as a potential therapeutic via blocking cleavage of TREM-1.

Hypomethylation of perforin regulatory elements in CD4+T cells from rat spleens contributes to the development of autoimmune emphysema

It is widely accepted that perforin regulatory elements are hypomethylated in CD4+ T cells from patients with active lupus, but whether this is the case in autoimmune emphysema is not known. Twenty rats were randomly divided into a normal control group and an emphysema group. Rat models of emphysema were established by intraperitoneal injection with xenogeneic endothelial cells. The levels of tumour necrosis factor-α, interleukin-8 and matrix metalloproteinase (MMP)-9 in bronchoalveolar lavage fluid (BALF) were measured, lung mean linear intercept and destructive index measured. Mean methylation of perforin gene promoter in CD4+ T cells and the expression of perforin were investigated. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling methods were used to examine the percentage of apoptotic cells in the alveolar septa. The levels of MMP-9 in BALF were higher in emphysema group than in control group (P < 0.05). The mean linear intercept and destructive index were higher in emphysema group than in control group (P < 0.05). The mean perforin gene promotor methylation of emphysema group was significantly decreased as compared with control group, while the expression levels of perforin gene were relatively higher (P < 0.05). There were more terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling-positive cells in the alveolar septa in control group than in emphysema group. Hypomethylation of perforin regulatory elements in CD4+ T cells may result in the lung septal cell apoptosis associated with the development of experimental autoimmune emphysema. MMP-9 may play an important role in the pathogenesis of this kind of disease.

Effect of Selective Cysteinyl Leukotriene Receptor Antagonists on Airway Inflammation and Matrix Metalloproteinase Expression in a Mouse Asthma Model

Cysteinyl leukotrienes (CysLTs) play a major role in the pathogenic changes of airway inflammation in asthma treatment. The matrix metalloproteinase (MMP) family, especially MMP-9 and MMP-2 levels, can reflect the status of airway remodeling. This study was undertaken to determine the role of a specific CysLT receptor antagonist in inhibition of airway inflammation and reversal of airway remodeling. Ovalbumin (OVA)-sensitized BALB/c mice were fed with a specific leukotriene receptor antagonist (MK-679), prednisolone or placebo from Days 15 to 27. Airway hyperreactivity, bronchoalveolar lavage fluid (BALF), and sera were analyzed. Pulmonary histology was obtained, and the levels of MMP-2 and MMP-9 in BALF were measured. The OVA-sensitized mice developed significant airway inflammatory responses, including extensive eosinophils trafficking into BALF and lung interstitium, goblet cell hyperplasia, mucus hypersecretion, elevated serum immunoglobulin (Ig) E, and decreased level of serum IgG2a. Administration of MK-679 could reduce airway inflammation but was not as effective as prednisolone. However, MK-679 was more effective than prednisolone for reversing subepithelial fibrotic and myofibrotic reactions of airway remodeling. The levels of MMP-2 and -9 in BALF were proportional to the extent of airway remodeling, which can reflect the effects of treatment. Both prednisolone and MK-679 reverse airway hyperresponsiveness induced by OVA-sensitized mice. Cysteinyl leukotriene receptor plays a more important role than CysLT in the pathogenesis of allergic airway inflammation. MMP-2 and -9 may be more sensitive indicators of airway remodeling.

Quantification of matrix metalloprotease-9 in bronchoalveolar lavage fluid by selected reaction monitoring with microfluidics nano-liquid-chromatography–mass spectrometry

Quantitative protein analysis by liquid chromatography–tandem mass spectrometry (LC–MS/MS) in the selected reaction monitoring (SRM) mode was used to quantify matrix metalloprotease-9 (MMP-9; ∼90kDa) in bronchoalveolar lavage fluid (BALF) from patients having undergone lung transplantation. We developed an SRM assay for microfluidics-based nanoLC–MS/MS on a triple quadrupole mass spectrometer based on two signature peptides. Samples were prepared by chloroform–methanol precipitation followed by trypsin digestion in the presence of stable-isotope-labeled internal peptide standards. The method allows accurate quantification of MMP-9 in BALF with an LLOQ of 2.9ng/mL and an LLOD of 0.25ng/mL without the use of extensive fractionation or antibodies.

MMP-9 Sheds the β2 Integrin Subunit (CD18) from Macrophages

Activated macrophages are essential effectors of immunity and a rich source of matrix metalloproteinase-9 (MMP-9; gelatinase B). To search for cellular substrates of the enzyme, we subjected wild-type macrophages and macrophages expressing an autoactivating form of pro-MMP-9 (M9A macrophages) to proteomics analysis. Two-dimensional liquid chromatography together with tandem mass spectrometry identified 467 proteins in medium conditioned by M9A and/or wild-type macrophages. Subtractive proteomics identified 18 candidate MMP-9 substrates. Biochemical studies confirmed that two transmembrane proteins, beta(2) integrin subunit (CD18) and amyloid protein precursor (APP), were enriched in the medium of M9A macrophages. To identify potential cleavage sites, we synthesized an overlapping library of peptides that spanned 60 residues of the ectodomain and transmembrane domain of beta(2) integrin. Active MMP-9 cleaved a single peptide, ECVKGPNVAAIVGGT, at residues corresponding to Ala(705) and Ile(706) of the beta(2) integrin. Peptides corresponding to this cleavage site were detected by tandem mass spectrometric analysis only in medium from M9A macrophages, strongly supporting the proposal that beta(2) integrin is shed by autoactivating MMP-9. Our observations indicate that subtractive proteomics in concert with peptide substrate mapping is a powerful approach for identifying proteolytic substrates and suggest that MMP-9 plays previously unsuspected roles in the regulation and shedding of beta(2) integrin.

Early and Late Changes of MMP-2 and MMP-9 in Bleomycin-Induced Pulmonary Fibrosis

PurposeMatrix metalloproteinases (MMPs) have been implicated in the pathogenesis of pulmonary fibrosis. To understand the role of MMP-2 and MMP-9 in pulmonary fibrosis, we evaluated the sequential dynamic change and different cellular sources of the 2 MMPs along the time course and their differential expression in the bronchoalveolar lavage (BAL) fluid and in the lung parenchyma of the bleomycin-induced pulmonary fibrosis models in rats.Materials and MethodsThe level of MMPs in BAL fluid of 54 bleomycin-treated rats was assessed by zymography from 1 to 28 days after intratracheal bleomycin instillation. The level of MMPs in lung parenchyma was evaluated by immunohistochemistry.ResultsMMP-2 and MMP-9 were markedly increased in both the BAL fluid and in the lung parenchyma of the bleomycin-treated rats, especially in the early phase with the peak on the 4th day. The levels of both MMPs in the BAL fluid correlated generally well to those in lung parenchyma, although the level of MMP-9 in BAL fluid was higher than MMP-2. In the lung parenchyma, the 2 MMPs, in early stage, were predominantly expressed in the inflammatory cells. In late stage, type II pneumocytes and alveolar epithelial cells at the periphery of the fibrotic foci retained MMP expression, which was more prominent in the cells showing features of cellular injury and/or repair.ConclusionIn bleomycin-induced pulmonary fibrosis, MMP-2 and MMP-9 may play important roles, especially in the early phase. In the late stage, the MMP-2 and MMP-9 may play a role in the process of repair.

Matrix metalloproteinases and matrix metalloproteinase inhibitors in acute lung injury

The objective of this study was to assess matrix metalloproteinase (MMP) and MMP inhibitor expression in the airspace of patients with acute lung injury (ALI) or acute respiratory distress syndrome (ARDS) and to determine the prognostic significance of MMP expression in this patient population. Twenty-eight patients with ALI or ARDS were prospectively enrolled in this study; bronchoalveolar lavage (BAL) fluid obtained from these patients was examined for expression of MMP-1 (interstitial collagenase), MMP-2 (gelatinase A), MMP-3 (stromelysin-1), MMP-8 (neutrophil collagenase), and MMP-9 (gelatinase B). Levels of MMP inhibitors (ie, tissue inhibitor of metalloproteinases-1 and -2 [TIMP-1 and TIMP-2]) were examined in parallel. Expression of MMPs was correlated with relevant clinical outcomes in patients with ALI/ARDS. In nearly all specimens obtained from patients with ALI/ARDS, there were high levels of MMP-2, MMP-8, MMP-9, and TIMP-1, but in only a small subset of patients (6/28) were there detectable levels of MMP-1 and/or MMP-3. In the patients with elevated MMP-1 and/or MMP-3, the mortality rate was higher (83%) than in the group without detectable levels of these enzymes (32%). Likewise, the overall severity of disease as indicated by Acute Physiology and Chronic Health Evaluation III scores was higher in this group (98 +/- 30) than in the group without detectable MMP-1 or MMP-3 (78 +/- 28). The percentage of individuals in whom lung disease was complicated by multiorgan failure was also higher in the group with detectable MMP-1 and/or MMP-3 (83%) than in the group without (64%), as was the number of organs that failed. In contrast, there was no correlation between MMP-1 and/or MMP-3 expression and impairment in gas exchange, as determined by the ratio of partial pressure of oxygen to fraction of inspired oxygen (Pao(2)/Fio(2)) on the day of BAL sample. Based on these findings, we conclude that elevated MMP-2, MMP-8, and MMP-9 in BAL fluid is a marker of acute lung injury (and, perhaps, a contributor to ALI) but is not necessarily an indicator of a poor outcome. On the other hand, the presence of detectable MMP-1 and/or MMP-3 is an indicator of more ominous disease progression.

A rise of MMP-2 and MMP-9 in bronchoalveolar lavage fluid is associated with acute lung injury after cardiopulmonary bypass in a swine model.

Postoperative acute lung injury (ALI) contributes to the morbidity and mortality following cardiopulmonary bypass (CPB). To determine whether the presence of matrix metalloproteinases (MMPs) is associated with ALI after CPB, MMP-2 and MMP-9 activities in bronchoalveolar lavage fluid (BALF) were compared with parameters indicating impaired gas exchange. In a prospective study, 17 minipigs were subjected to CPB for 60 min. Before and at five and 180 min after CPB, MMP-2 and MMP-9 were assayed in BALF and the arterial-alveolar gradient of oxygen tension (AaDO2), the pulmonary capillary wedge pressure (PCWP) and the water content of lung tissue samples (Wt) were evaluated and compared with baseline values. MMP-2 and MMP-9 increased significantly 5 minutes (2.1- and 6.2-fold, respectively) and 180 minutes (3.4- and 14.3-fold, respectively) post-CPB. AaDO2 and Wt, but not PCWP, increased significantly 180 minutes after CPB and only AaDO2, but not PCWP or Wt, was significantly correlated with MMP-2 (r = 0.66, p = 0.006) and MMP-9 (r = 0.62, p = 0.01). In conclusion, high levels of MMP-2 and MMP-9 in the pulmonary compartment are associated with ALI after CPB.

Clinical Findings, Bronchoalveolar Lavage Fluid Cytology and Matrix Metalloproteinase-2 and -9 in Canine Pulmonary Eosinophilia

We characterized clinical and clinicopathological features, and the involvement of gelatinolytic matrix metalloproteinases (MMP-2 and -9) in canine pulmonary eosinophilia (PE). Study material consisted of 20 PE dogs and 16 healthy beagles. All dogs underwent a similar clinical examination and bronchoalveolar lavage (BAL). Analysis for cell count and differential cell count of BAL fluid (BALF), arterial blood gas analysis before and after BAL, and thoracic radiographs before BAL and after treatment were obtained. Twelve dogs were re-evaluated and six relavaged. MMP-2 and MMP-9 in BALF were analysed by zymography, Western immunoblotting and immunocytochemistry.In the PE dogs, BALF, cell count, number and percentage of eosinophils, and numbers of macrophages, lymphocytes, neutrophils, mast cells and epithelial cells were all significantly elevated. Blood eosinophilia was detected in half of the PE dogs. Three PE dogs had mild hypoxaemia. The BAL procedure had an equal effect on PE and healthy dogs' arterial blood gas values. Bronchointerstitial densities were seen in PE dogs' radiographs. Treatment of PE decreased BALF cell count, eosinophil count and percentage and diminished radiographic changes. Gelatinolytic activity was higher in PE dogs' BALF. BALF macrophages and epithelial cells were the principal sources of the MMP-9.

Increased matrix metalloproteinase-9 in the airway after allergen challenge.

Persistent asthma is associated with airway inflammation, tissue damage, and deposition of extracellular matrix (ECM) proteins, which may be mediated, in part, through release of matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinase-1 (TIMP-1). To investigate the role of allergen in the induction of MMP-9 and TIMP-1, bronchoscopy and segmental bronchoprovocation (SBP) with saline (SAL) and antigen (AG) were performed in 17 allergic subjects. Bronchoalveolar lavage (BAL) was done 5 min and 48 h after challenge and concentrations of MMP-9 and TIMP-1 in BAL fluid (BALF) were measured by ELISA. Forty-eight hours after AG challenge, concentrations of MMP-9 and TIMP-1 were increased in the airway, but not in serum. Zymography demonstrated that MMP-9 was the predominant metalloproteinase and was present in a latent proform. MMP-9 immunoreactivity was associated primarily with neutrophils, and concentrations of MMP-9 in BALF correlated with airway neutrophils and, to a lesser extent, with alveolar macrophages. These data suggest that AG challenge leads to generation of factors, including MMP-9, that may be associated with the initiation of bronchial injury, which may then lead to remodeling in asthma.

Tissue inhibitor of metalloproteinase-1 levels in bronchoalveolar lavage fluid from asthmatic subjects.

Airway remodeling is a well-recognized feature in patients with chronic asthma. The accumulation in the submucosa of fibrous proteins that are substrates of matrix metalloproteinases (MMP), and the demonstration of increased levels of MMP-9 in bronchoalveolar lavage fluid, prompted us to determine whether there was an imbalance between MMPs and tissue inhibitors of metalloproteinase (TIMP) in such patients. We investigated the presence of TIMPs and other MMPs. TIMP levels were compared with those of all MMPs and inflammatory cytokines. Adults with stable asthma, either untreated or treated with glucocorticoids (GCs), were enrolled. Healthy nonsmokers served as a control population. MMPs and TIMPs were identified through zymography or immunoblotting. TIMPs, MMPs, and cytokines were measured with enzyme immunoassays. TIMP-1 levels were significantly higher in untreated asthmatic subjects than in GC-treated subjects or controls (p < 0.0001), and were far greater than those of MMP-1, MMP-2, MMP-3, and MMP-9 combined. TIMP-2 was undetectable. TIMP-1 levels were correlated with levels of interleukin-6 (p < 0.012) and the number of alveolar macrophages recovered (p < 0.005). This observation has important implications, since an excess of TIMP-1 could lead to airway fibrosis, a hallmark of airway remodelling in patients with chronic asthma.