Protective Role of Matrix Metalloproteinase-9 in Ozone-Induced Airway Inflammation
Publication: Environmental Health Perspectives
Volume 115, Issue 11
Pages 1557 - 1563
Abstract
Background
Exposure to ozone causes airway inflammation, hyperreactivity, lung hyper-permeability, and epithelial cell injury. An early inflammatory response induced by inhaled O3 is characterized primarily by release of inflammatory mediators such as cytokines, chemokines, and airway neutrophil accumulation. Matrix metalloproteinases (MMPs) have been implicated in the pathogenesis of oxidative lung disorders including acute lung injury, asthma, and chronic obstructive pulmonary disease.
Objective
We hypothesized that MMPs have an important role in the pathogenesis of O3-induced airway inflammation.
Methods
We compared the lung injury responses in either Mmp7- (Mmp7−/−) or Mmp9-deficient (Mmp9−/−) mice and their wild-type controls (Mmp7+/+, Mmp9+/+) after exposure to 0.3 ppm O3 or filtered air.
Results
Relative to air-exposed controls, MMP-9 activity in bronchoalveolar lavage fluid (BALF) was significantly increased by O3 exposure in Mmp9+/+ mice. O3-induced increases in the concentration of total protein (a marker of lung permeability) and the numbers of neutrophils and epithelial cells in BALF were significantly greater in Mmp9−/− mice compared with Mmp9+/+ mice. Keratinocyte-derived chemokine (KC) and macrophage inflammatory protein (MIP)-2 levels in BALF were also significantly higher in Mmp9−/− mice than in Mmp9+/+ mice after O3 exposure, although no differences in mRNA expression for these chemokines were found between genotypes. Mean BALF protein concentration and numbers of inflammatory cells were not significantly different between Mmp7+/+ and Mmp7−/− mice after O3 exposure.
Conclusions
Results demonstrated a protective role of MMP-9 but not of MMP-7, in O3-induced lung neutrophilic inflammation and hyperpermeability. The mechanism through which Mmp9 limits O3-induced airway injury is not known but may be via posttranscriptional effects on proinflammatory CXC chemokines including KC and MIP-2.
Ozone is a common urban air pollutant that remains a major health concern. Epidemiologic studies have demonstrated a strong association between high ambient O3 concentration with cardiovascular and respiratory morbidity and mortality (Bell et al. 2005). O3 exposure elicits airway inflammation characterized by neutrophil accumulation (Schelegle et al. 1991; Seltzer et al. 1986) and liberates multiple inflammatory mediators, cytokines, and chemokines as an early inflammatory event (Aris et al. 1993; Koren et al. 1989). O3-induced activation of airway neutrophilic infiltration is likely to produce additional damage through the release of reactive oxygen species and endogenous proteolytic enzymes.
Several different kinds of endogenous proteases participate in the pathogenesis of airway inflammation. Among these, matrix metalloproteinases (MMPs) belong to a family of zinc-dependent extracellular enzymes that share common structural features and have an important role in many physiological and pathological conditions (Parks and Shapiro 2001). MMPs are secreted by inflammatory and noninflammatory cells and have a major role in extracellular matrix (ECM) degradation, proteolytic modulation of biologically active proteins, and cell migration (Goetzl et al. 1996; Kelly et al. 2000; Kheradmand et al. 2002; Mautino et al. 1997; Parks and Shapiro 2001). Elevated levels of MMPs are prominent in inflammatory disorders of the airway and parenchymal lung disease and have been also implicated in several pulmonary diseases characterized by lung tissue damage, alveolar structure alterations, or abnormal repair (Elkington and Friedland 2006; Parks and Shapiro 2001). MMPs can be divided on the basis of their substrate specificity into collagenases (MMP-1, 8, 13), gelatinases (MMP-2, 9), stromelysins (MMP-3, 10, 11), matrilysin (MMP-7), macrophage metalloelastase (MMP-12), and membrane-type (MT)-MMPs (MMP-14, 15, 16, 17) (Visse and Nagase 2003).
Gelatinases degrade many ECM proteins; most important is type IV collagen, a major constituent of lung basement membrane (Nagase and Woessner 1999). Therefore, gelatinases have been thought to have an important role in the pathogenesis of various lung diseases (Nagase and Woessner 1999). Gelatinases also can cleave a variety of non-ECM proteins, including cytokines and chemokines (McQuibban et al. 2000; Yu and Stamenkovic 2000; Zhang et al. 2003). MMP-2 (gelatinase A) is produced by a wide range of cell types in the lung, and it is increased in lung disorders with oxidative stress and inflammation etiologies (e.g., Pardo et al. 1998; Perez-Ramos et al. 1999; Tan et al. 2006). MMP-9 (gelatinase B) has been implicated in the pathogenesis of asthma (Cataldo et al. 2000), idiopathic pulmonary fibrosis (Fukuda et al. 1998), chronic obstructive pulmonary disease (COPD; Russell et al. 2002), and acute lung injury (Lanchou et al. 2003). In the normal lung, constitutive expression of MMP-9 is restricted to neutrophils and eosinophils. However, MMP-9 induced by inflammatory stimuli and proinflammatory cytokines is generated in multiple cell types including alveolar macrophages and other resident pulmonary cells (Atkinson and Senior 2003; Chakrabarti and Patel 2005). MMP-9 is known to be induced by O3 in murine skin and lung (Kenyon et al. 2002; Valacchi et al. 2003).
MMP-7 has homeostatic function and a role in innate immunity in epithelial cells (Parks and Shapiro 2001). In adult human lung, MMP-7 is expressed by epithelial cells lining peribronchial and conducting airway (Dunsmore et al. 1998). Increased expression of MMP-7 has been observed in airway and alveolar cells at sites of lung cancer (Wilson and Matrisian 1996), idiopathic pulmonary fibrosis (Zuo et al. 2002), and cystic fibrosis (Dunsmore et al. 1998). Although it is clear that MMP-7 is associated with chronic lung disease, the role of MMP-7 in O3-induced airway inflammation is still unknown.
In the present studies, we hypothesized that murine MMP-7 and MMP-9 have important functional roles in facilitating the airway inflammatory responses induced by inhaled O3. To address this hypothesis, we used mice genetically deficient in Mmp7 (Mmp7, matrix metalloproteinase 7; Entrez Gene ID 17393, http://www.ncbi.nlm.nih.gov/sites/entrez?db=gene) and Mmp9 (Mmp9; matrix metalloproteinase 9; Entrez Gene ID 17395, http://www.ncbi.nlm.nih.gov/sites/entrez?db=gene) and compared airway injury responses with O3 relative to corresponding wild-type control mice.
Materials and Methods
Animals and inhalation exposure
Male (6–8 weeks old, 20–25 g) C57BL/6J (Mmp7+/+) and B6.129-Mmp7tm1Lmm/J (Mmp7−/−), FVB/NJ (Mmp9+/+), and FVB.Cg-Mmp9tm1Tvu/J (Mmp9−/−) mice were purchased from The Jackson Laboratory (Bar Harbor, ME). Inhalation exposures were performed in a barrier facility at Alion Science and Technology, Inc. (Research Triangle Park, NC). After acclimation, mice were placed in individual stainless-steel wire cages within a Hazelton 1000 chamber (Lab Products, Maywood, NJ) equipped with a charcoal and high-efficiency particulate air-filtered air supply. Mice had free access to water and pelleted open-formula rodent diet NIH-31 (Zeigler Bros, Inc., Gardners, PA). Mice were exposed continuously to 0.3 ppm O3 for 6, 24, 48, or 72 hr. O3 was generated from ultra-high purity air (< 1 ppm total hydrocarbons; National Welders, Inc., Raleigh, NC) using a silent arc discharge O3 generator (Model L-11; Pacific Ozone Technology, Benecia, CA). Constant chamber air temperature (72 ± 3°F) and relative humidity (50 ± 15%) were maintained. O3 concentration was continually monitored (Dasibi model 1008-PC, Dasabi Environmental Corp., Glendale, CA). Parallel exposure to filtered air was conducted in a separate chamber for 48 hr. All animal use was approved by the National Institute of Environmental Health Sciences Animal Care and Use Committee, and all animals were treated humanely and with regard for alleviation of suffering.
Lung bronchoalveolar lavage fluid (BALF) and cell preparation
Immediately after exposure to air or O3, mice were euthanized with sodium pentobarbital (104 mg/kg). After 24-or 48-hr exposure, BAL was performed in situ 4 times with Hanks balanced salt solution (35 mL/kg; pH 7.2–7.4). These time points were chosen for BAL procedures on the basis of previous investigations with multiple inbred strains of mice that indicated inflammation and injury peaked between 24 and 48 hr during continuous exposure to 0.3 ppm O3 (e.g., Cho et al. 2007; Dahl et al. 2007; Kleeberger et al. 1993). Recovered BALF was immediately cooled in ice and centrifuged (1,000 × g, 10 min). The cell pellets were resuspended in 1 mL of Hanks balanced salt solution, and the cells were counted with a hemacytometer. Aliquots (150 μL) were cyto-centrifuged, and the cells were stained with Wright-Giemsa stain solutions (Fisher Scientific, Pittsburgh, PA) for differential cell analysis (markers of cellular inflammation and epithelial injury). The supernatants from the first BALF return were assayed for total protein (a marker of lung permeability) with the Bradford assay (Bradford 1976).
Lung tissue preparation for histopathology and immunohistochemistry
Left lung lobes from mice exposed to either 0.3 ppm O3 or filtered air for 48 hr were inflated gently with zinc formalin, fixed at a constant pressure (25 cm) of the same fixative for 30 min, tied off at the trachea, and immersed in the fixative for 1 day. Proximal and distal intrapulmonary axial airway was excised, embedded in paraffin, and sectioned (5 μm). Tissue sections were histochemically stained with hematoxylin and eosin (H&E) for morphological comparison of pulmonary injury between genotypes. The terminal bronchioles and alveoli were the primary focus of study because 48-hr exposure to 0.3 ppm O3 causes histologically evident inflammation and epithelial lesions in these regions of the mouse lung. Separate tissue sections from Mmp9+/+ or Mmp9−/− mice were immunologically stained using a goat anti-mouse MMP-9 antibody (R&D Systems, Inc., Minneapolis, MN) or a rat anti-mouse Ki-67 antibody (Dako North American, Inc., Carpinteria, CA) to localize MMP-9 and Ki-67, respectively, using the standard peroxidase–diaminobenzidine (DAB) method.
Western blot analysis
Equal amounts of total lung protein (45 μg) isolated from mouse lung homogenates (air or 24, 48, and 72 hr of O3) in radioimmunoprecipitation (RIPA) buffer (1% Nonidet P-40, 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulfate (SDS), 10 g phenylmethylsulfonyl fluoride/mL, 1 mM sodium orthovanadate, 1× protease inhibitor cocktail in 1× phosphate-buffered saline) were separated on 10–20% SDS gel electrophoresis gels. The 24-, 48-, and 72-hr time points were chosen for investigation to characterize the kinetics of lung MMP activity changes induced by O3 exposure. Proteins were then transferred to a polyvinylidene fluoride (PVDF) membrane. The membrane was blocked for 2 hr at room temperature with 5% nonfat milk in Tris-buffered saline–Tween buffer (20 mmol/L Tris, 500 mmol/L NaCl, 0.01% Tween 20). The blot was then incubated at 4°C overnight with anti-MMP-9 (R&D Systems), anti-MMP-2 (R&D Systems), or an anti-actin antibody (SantaCruz Biotechnology Inc., Santa Cruz, CA) as an internal control, followed by incubation for 1–2 hr with the proper secondary horseradish peroxidase-conjugated antibodies. The immunoblot was visualized through enhanced chemiluminescence.
Total RNA isolation and real-time quantitative reverse transcriptase–polymerase chain reaction (RT-PCR)
Total RNA from frozen lungs (air or 6, 24, and 48 hr of O3) was isolated using the RNeasy Midiprep kit (QIA-GEN, Valencia, CA). The 6-hr time point was included in this protocol to optimize the likelihood of capturing early inflammatory cytokine/chemokine gene expression on the basis of previous experience with this model (Cho et al. 2007). One microgram total RNA was reverse transcribed into cDNA in a volume of 50 μL containing 1× PCR buffer [50 mM KCl and 10 mM Tris (pH 8.3)], 5 mM MgCl2, 1 mM each dNTPs, 125 ng oligo (dT)15, and 50 U of Moloney murine leukemia virus reverse transcriptase (Invitrogen Life Technologies, Carlsbad, CA), at 45°C for 15 min and 95°C for 5 min using the Gene Amp PCR System 9700 (Applied Biosystems, Foster City, CA).
An aliquot (2–5 μL) of the reverse transcriptase product (equivalent to 40–100 ng RNA) was amplified using PowerSYBR gene expression assays on the 7700 Prism sequence detection system (Applied Biosystems). The PCR conditions and data analysis were performed according to the manufacturer’s protocol in User Bulletin no. 2, “Applied Biosystems Prism 7700 Sequence Detection System” (Applied Biosystems 2007). Quantification of gene expression was determined by the number of cycles to threshold (CT) of fluorescence detection. Relative gene expression was determined using the comparative CT method as described previously (Chakrabarti and Patel 2005). Briefly, the ΔCT value was obtained by subtracting the 18s rRNA CT value from the CT value of the gene tested in the same sample. Message levels of each gene were expressed as fold changes relative to those in Mmp9+/+ air controls.
Gelatin zymography analysis
Equal volumes of BALF (10 μL) samples were loaded on 10% Tris–glycine gel with 0.1% gelatin (Invitrogen Life Technologies) for electrophoresis. To eliminate the SDS content, the gels were washed with 2.7% (wt/vol) Triton X-100 at room temperature for 30 min and incubated for 16 hr in a buffer containing 50 mM Tris base, 40 mM HCl, 200 mM NaCl, 5 mM CaCl2, and 0.02% (wt/vol) Brij 35 for gelatinolytic enzymes to act. The digested gels were stained in Colloidal Blue Staining kit (Bio-Rad Laboratories, Hercules, CA) for 4 hr to visualize the gelatin activity. Gelatinolytic bands were analyzed by a Gel Doc 2000 System (Bio-Rad).
Cytokine enzyme-linked immunosorbent assay (ELISA)
Immunoreactive macrophage inflammatory protein-2 [MIP-2, human interleukin-8 (IL-8) analogue, similar to chemokine (C-X-C motif) ligand (CXCL) 1 and 2] and keratinocyte-derived chemokine [KC, human growth-regulated oncogene (GRO)-α analogue, similar to CXCL1, 2, and 3] were quantified in aliquots of BALF (50 μL) using commercially available ELISA kits (R&D Systems), according to the manufacturer’s instructions. Each cytokine quantity was calculated from absorbance at 450 nm using a standard curve.
Statistics
All data are expressed as group means ± SE. Two-way analysis of variance (ANOVA) was used to evaluate the effects of subacute O3 exposure on lung injury parameters between Mmp7+/+ and Mmp7−/− or Mmp9+/+ and Mmp9−/− mice. The factors in the analysis were exposure (air vs. O3) and genotype (Mmp7+/+ vs. Mmp7−/−, Mmp9+/+ vs. Mmp9−/−). The dependent variables were protein level, cell number, MMP-2 activity, and cytokine concentrations in BALF, and lung protein and mRNA levels. One-way ANOVA was used to evaluate the effect of O3 on MMP-9 production and expression in Mmp9+/+ mice. The Student–Newman–Keuls test was used for a posteriori comparisons of means. All analyses were performed using a commercial statistical analysis package (SigmaStat; SPSS Inc., Chicago, IL). Statistical significance was accepted at p < 0.05.
Results
Roles of MMP-7 and MMP-9 in O3-induced airway inflammation
The roles of MMP-7 and MMP-9 in O3-induced airway inflammation were assessed by exposing Mmp7+/+, Mmp7−/−, Mmp9+/+, and Mmp9−/− mice continuously to air or O3 for 24 and 48 hr. No significant differences in mean BAL protein concentration or cell numbers were found between respective +/+ and −/− genotypes after exposure to air (Figure 1, Table 1). After 48 hr of O3 exposure, BAL protein concentration was significantly (p < 0.05) increased in all four genotypes (Figure 1, Table 1). The BALF protein concentration was significantly greater (68%) in Mmp9−/− mice compared with Mmp9+/+ mice (Figure 1A); however, no significant differences in protein concentration were found between Mmp7+/+ and Mmp7−/− mice after exposure to O3 (Table 1). O3 caused significant increases in the mean numbers of BAL neutrophils in Mmp9+/+ and Mmp9−/− mice after 48 hr of O3 exposure (p < 0.05; Figure 1B). However, compared with Mmp9+/+ mice, O3-enhanced BAL neutrophil numbers were significantly higher in Mmp9−/−mice (p < 0.05). Mean numbers of epithelial cells were also significantly greater in Mmp−/− mice relative to those in Mmp+/+ mice after 24- and 48-hr O3 exposure (p < 0.05; Figure 1C). No significant differences in the number of BALF alveolar macrophages were found between Mmp9+/+ and Mmp9−/− mice (data not shown). No significant differences in mean numbers of BALF inflammatory cells were found between Mmp7−/− and Mmp7+/+ mice after exposure to O3 (Table 1).
Figure 1 Concentration of total protein (A) and numbers of neutrophils (B) and epithelial cells (C) in BALF recovered from Mmp9+/+ and Mmp9−/− mice after exposure to 0.3 ppm O3 or filtered air. Data are presented as means ± SE (n = 8–10/group).
*Significantly different from genotype-matched air control mice (p < 0.05). #Significantly different from exposure-matched Mmp9+/+ mice (p < 0.05).
| Genotype | Proteins (μg/mL) | Macrophages (×103/mL) | Neutrophils (×103/mL) | Epithelial cells (×103/mL) |
|---|---|---|---|---|
| Mmp7+/+ (Air) | 107.0 ± 6.7 | 21.8 ± 2.8 | 0.1 ± 0.1 | 1.7 ± 0.2 |
| Mmp7−/− (Air) | 140.0 ± 4.7 | 23.5 ± 1.9 | 0.02 ± 0.02 | 1.5 ± 0.3 |
| Mmp7+/+ (O3) | 540.8 ± 32.1* | 47.6 ± 2.2* | 5.9 ± 1.5* | 3.0 ± 0.5 |
| Mmp7−/− (O3) | 707.5 ± 73.1* | 49.7 ± 4.4* | 10.0 ± 3.3* | 7.1 ± 1.2* |
Results are the means ± SE for 5–8 mice in each group.
*
Significantly different from genotype-matched air control mice (p < 0.05).
Effect of O3 on MMP-2 and MMP-9 expression in the lung
We performed gelatin zymogram and Western blot analyses on BALF and lung protein extracts from Mmp9+/+ and Mmp9−/− mice to determine whether production of MMP-2 and MMP-9 was augmented by O3 exposure. Both gelatinases have been implicated in the pathogenesis of inflammation-related lung diseases (e.g., Pardo et al. 1998; Tan et al. 2006); we also reasoned that MMP-2 expression might be enhanced to compensate for MMP-9 deletion. Minimal gelatinase activity was found in BALF from either genotype after air exposure. Exposure to O3 (48 and 72 hr) caused significant (p < 0.05) increase in MMP-9 activity at 92 kDa (latent form) and 88 kDa (active form) in BALF from Mmp9+/+ mice (Figure 2A). Western blot analyses confirmed O3-induced increase in MMP-9 protein levels in lung homogenates from Mmp9+/+ mice (Figure 2B). Furthermore, lung MMP-9 mRNA expression in Mmp9+/+ mice was significantly increased compared with air controls after 6 hr of O3 and remained increased by 48-hr exposure (p < 0.05; Figure 2C). MMP-9 proteins were localized primarily in alveolar epithelial cells, infiltrating inflammatory cells, endothelial cells, and cells in the injured parenchyma of Mmp9+/+ mice after O3 exposure (Figure 3).
Figure 2 MMP-9 mRNA, protein, and activity after exposure to 0.3 ppm O3 or filtered air. (A) Representative images of MMP-9 activity in BALF from Mmp9+/+ mice determined by gelatin zymogram after exposure to 0.3 ppm O3 (24, 48, 72 hr) or filtered air (n = 3/group). (B) Representative Western blots of MMP-9 in pooled lung homogenates from Mmp9+/+ (+/+) and Mmp9−/− (−/−) mice after exposure to 0.3 ppm O3 (24, 48, 72 hr) or filtered air (n = 3/group). (C) MMP-9 mRNA expression in lung homogenates from Mmp9+/+ mice after exposure to 0.3 ppm O3 (6, 24, 48 hr) or filtered air. Quantitative real-time PCR-determined fold increase of normalized expression to 18s rRNA compared with air-exposed mice (n = 3/group). Data are presented as means ± SE.
*Significantly different from air control mice (p < 0.05).
Basal levels of MMP-2 protein and activity were detectable in lung protein extract and BALF, respectively, from Mmp9+/+ and Mmp9−/− mice, although no differences were detected between genotypes. O3 increased MMP-2 activity and production in both genotypes compared with corresponding baseline (Figure 4A,B), but the increases were significantly (p < 0.05) greater in Mmp9−/− compared with Mmp9+/+ mice.
Figure 4 Measurement of MMP-2 in BALF and lung protein extracts from Mmp9+/+ and Mmp9−/− mice after exposure to filtered air or 0.3 ppm O3. (A) Representative Western blot image of MMP-2 protein expression in pooled lung homogenates from Mmp9+/+ (+/+) and Mmp9−/− (−/−) mice after exposure to 0.3 ppm O3 (24, 48, 72 hr) or filtered air (n = 3/group). (B) Representative gelatin zymogram image for MMP-2 activity in BALF from Mmp9+/+ (+/+) and Mmp9−/− (−/−) mice after exposure to 0.3 ppm O3 (48 hr) or filtered air. Relative optical density of MMP-2 activity quantified from zymogram blots. Data are presented as means ± SE (n = 3/group).
*Significantly different from genotype-matched air control mice (p < 0.05). #Significantly different from O3 (48 hr)-exposed Mmp9+/+ mice (p < 0.05).
Effects of Mmp9 deficiency on O3-induced pulmonary pathology.
H&E-stained lung sections from Mmp9+/+ and Mmp9−/− mice exposed to air were normal, and no histological differences were found between them (Figure 5A,5C). After 48 hr of O3 exposure, significant peribronchiolar inflammation and proliferation as well as epithelial hyperplasia in terminal bronchioles were detected in Mmp9+/+ and Mmp9−/− mice (Figure 5B,D). However, O3 caused more severe epithelial thickening, inflammation, and alveolar vacuolization in the terminal bronchioles of Mmp9−/− compared with Mmp9+/+ mice (Figure 5D). Enhanced airway cellular proliferation was also more severe in the absence of MMP-9 as determined by immunohistochemistry of Ki-67–positive cells in G/S phases of the cell cycle (Figure 6). Nuclear localization of Ki-67 was mainly found in inflammatory cells and in cells of the terminal bronchioles that were injured by O3.
Effect of Mmp9 deficiency on protein levels and mRNA expression of the neutrophil chemokines KC and MIP-2.
To investigate the mechanism for increased neutrophil recruitment to the airway in the absence of MMP-9, we used ELISA to measure BALF levels of chemokines known to be involved in neutrophil chemotaxis (KC and MIP-2). No significant genotype-specific differences in mean concentrations of KC or MIP-2 were found after exposure to air (Figure 7). Mean KC and MIP-2 concentrations were significantly increased in BALF by O3 exposure in both genotypes (p < 0.05, Figure 7). However, consistent with BALF inflammatory parameters and pathology (Figures 1, 5, and 6), KC and MIP-2 levels were significantly (p < 0.05) greater in Mmp9−/− mice compared with Mmp9+/+ mice after 24- and 48-hr exposure to O3 (Figure 7). KC and MIP-2 mRNA levels were not different between genotypes after exposure to air. After 24- and 48-hr exposure to O3, KC and MIP-2 mRNA was increased significantly (p < 0.05) over the air controls in both genotypes but, interestingly, no differences were found between genotypes (Figure 8).
Figure 7 Protein levels of neutrophil chemokines KC (A) and MIP-2 (B) in BALF from Mmp9+/+ and Mmp9−/− mice exposed to air and 0.3 ppm O3. Data are presented as means ± SE (n = 7–8/group).
*Significantly different from genotype-matched air control mice (p < 0.05). #Significantly different from exposure-matched Mmp9+/+ mice (p < 0.05).
Figure 8 mRNA expression of neutrophilic chemokines KC (A) and MIP-2 (B) in Mmp9+/+ and Mmp9−/− mice exposed to air and 0.3 ppm O3. Quantitative real time PCR determined fold increase of message expression normalized to18s rRNA compared with air exposed Mmp9+/+ mice (n = 3/group). Data are presented as means ± SE (n = 3/group).
*Significantly different from genotype-matched air control mice (p < 0.05).
Discussion
O3-induced airway inflammation is characterized by early neutrophilic infiltration followed by a mononuclear cell dominated inflammation (Chitano et al. 1995). Molecular changes during the pulmonary pathogenesis induced by O3 include increased production of prostaglandins, proinflammatory cytokines, and chemokines such as IL-6, IL-8, granulocyte macrophage colony-stimulating factor, KC, and MIP-2 (Devlin et al. 1991; Johnston et al. 2005a). In humans, neutrophil influx begins in peripheral airways as early as 6 hr after O3 exposure and is known to be mediated by neutrophilic chemokines such as IL-8 and GRO-α (Krishna et al. 1998). Functional roles of several inflammatory mediators such as tumor necrosis factor-α (TNF-α) (Cho et al. 2001; Kleeberger et al. 1997), IL-6 (Johnston et al. 2005b), and IL-1β (Arsalane et al. 1995), and the transcription factors nuclear factor kappa B (Cho et al. 2007; Laskin et al. 2002) and activator protein (AP)-1 (Cho et al. 2007) have also been determined in pulmonary inflammation of laboratory rodents exposed to O3.
The primary functions of MMPs include degradation and turnover of ECM, tissue repair and remodeling, and leukocyte migration from peripheral circulation to inflammatory sites. Furthermore, recent findings suggested that MMPs can modulate inflammation and innate immunity by affecting the activity of various nonmatrix proteins (Parks et al. 2004). MMP-9 has been thought to be particularly important in the pathogenesis of inflammatory lung diseases, including acute lung injury, asthma, and COPD (Atkinson and Senior 2003). Higher than normal MMP-9 levels in the patients with these disorders may promote destruction of normal tissue architecture and increased migration of inflammatory cells to the disease sites (Lemjabbar et al. 1999a, 1999b).
In the present study, we hypothesized that MMP-9 is essential for O3-induced airway inflammation because MMP-9 is one of the most predominant MMPs found in inflammatory airway diseases (Atkinson and Senior 2003). Contrary to predictions, we found that O3-induced neutrophilic airway inflammation and injury were markedly increased in Mmp9−/− mice compared with Mmp9+/+ mice, indicating a protective role of MMP-9 in O3-exposed airways. The results from the current study suggest that MMP-9 may not act primarily on ECM degradation that facilitates leukocyte migration to pulmonary inflammatory sites as indicated by another investigation (Betsuyaku et al. 1999). Heightened neutrophilic inflammation was also associated with enhanced levels of KC and MIP-2, which are important chemokines for neutrophil recruitment to the lung.
MMP-9 has diverse effects on neutrophilic inflammation in experimental animal models. It enhances neutrophil chemotaxis in response to certain chemokines (D’Haese et al. 2000) and increases neutrophil influx in glomerulonephritis (Sternlicht and Werb 2001). Conversely, MMP-9 also has inhibitory effects on BALF neutrophilia (Lanone et al. 2002); increased tissue neutrophil and inflammatory cell infiltration have been shown in Mmp9−/−mice in response to epithelial injury and chemokine administration (D’Haese et al. 2000; Mohan R et al. 2002). Several lines of molecular evidence have determined that proteolytic function of MMP-9 affects cytokine and chemokine levels as well as their activities. For example, MMP-9 processes and activates pro-IL-1β (Schonbeck et al. 1998), inactive membrane bound forms of TNF-α (Mohan MJ et al. 2002), and transforming growth factor-β (TGF-β) (Yu and Stamenkovic 2000). MMP-9 also processes CXC chemokines, which exert potent chemoattractant activities on leukocytes and alters their specific activities, but not CC chemokines (e.g., RANTES and MCP-2) (Van den Steen et al. 2000). MMP-9 truncates IL-8 (1–77) into IL-8 (7–77), which enhances neutrophil activation more than 10-fold (Van den Steen et al. 2000). In contrast, neutrophilic chemoattractant activity was decreased by MMP-9 degradation of the other CXC chemokines GRO-α and connective tissue–activating peptide (CTAP)-III (Van den Steen et al. 2000). Our current observation that O3 induced higher levels of BAL CXC chemokines KC and MIP-2 in Mmp9−/− mice than in Mmp9+/+ mice is consistent with, but does not prove, the notion that these chemokines were key effectors of pulmonary MMP-9. Interestingly, we also found no differences in the steady state mRNA levels of the O3-increased KC and MIP-2 between Mmp9+/+ and Mmp9−/− mice. Together these results suggest that differences in these chemokine levels in Mmp9−/− mice compared with those in Mmp9+/+ mice were caused by translational or posttranslational processes, which may include degradation/cleavage by MMP-9.
Significantly greater elevation of MMP-2 level and activity were observed in Mmp9−/− mice compared with Mmp9+/+ mice after O3 exposure. We cannot rule out the possibility that increased MMP-2 is a compensatory response in Mmp9−/− mice and may be involved in increased neutrophilic airway inflammation. However, MMP-9 has been reported to be the dominant airway MMP controlling inflammatory cell egression (Corry et al. 2004), and there is no evidence that MMP-2 has a modulating effect on inflammatory chemokines such as KC and MIP-2 (Parks et al. 2004). In addition, neutrophil concentrations in BALF were not changed in Mmp2−/− mice in a model of allergic asthma (Corry et al. 2002). We therefore postulate that MMP-2 does not have a critical role in heightened neutrophilic inflammation in Mmp9−/− mice, although it may be involved with other mechanisms of lung injury induced by O3 exposure (e.g., lung hyperpermeability).
The role of MMP-7 in acute lung injury has been studied in mouse models of interstitial pulmonary diseases such as fibrosis: Mmp7−/− mice had suppressed pulmonary fibrosis caused by bleomycin (Li et al. 2002; Zuo et al. 2002), and it was accompanied by decreased neutrophilic inflammation and chemokines (e.g., KC) in the alveolar fluid (Li et al. 2002). We predicted that O3-induced airway inflammation would be attenuated in Mmp7−/− mice. However, the current findings suggest that MMP-7 is not significantly associated with O3-induced airway inflammation and injury in mice. In addition, KC concentrations in BALF were not significantly different between Mmp7−/− and Mmp7+/+ mice in the current model (data not shown). No studies have investigated the role of MMP-7 in oxidative lung injury such as O3. Different from MMP-9, matrilysin is produced by alveolar epithelial cells in injured lungs, and it has been thought to contribute to alveolar epithelial injury and re-epithelialization. Inhaled O3 mostly affects centriacinar legions of the airway, but not alveoli, which may partially explain why MMP-7 deficiency failed to alter airway inflammation by O3.
In summary, our results showed that deficiency in MMP-9 was associated with enhanced airway epithelial injury, neutrophil recruitment, and permeability following O3 exposure, but a deficiency in MMP-7 did not significantly affect O3-induced airway injury. The aberrant neutrophil recruitment was correlated with increased levels of KC and MIP-2 protein, but not mRNA expression, in the Mmp9−/− mice relative to Mmp9+/+ mice. Results are consistent with the hypothesis that enhanced O3-induced injury in Mmp−/− mice is related to a difference in posttranscriptional processing of these CXC chemokines in the airway. These findings increase our understanding of the pathophysiological process of O3-induced lung injury and suggest that MMP-9 produced in the lung in response to oxidative stimuli may have a beneficial function.
Article Notes
The authors thank D. Morgan and H. Price for coordinating the inhalation exposures. D. Brass and D. Cook at the NIEHS provided excellent critical review of the manuscript.
This research was supported by the Intramural Research Program of the NIH, and NIEHS. Ozone exposures were conducted at the NIEHS Inhalation Facility under contract to Alion Science and Technology, Inc.
References
Applied Biosystems. 2007. User Bulletin no. 2: Applied Biosystems Prism 7700 Sequence Detection System Foster City, CA Applied Biosystems Available: http://www3.appliedbiosystems.com/cms/groups/mcb_support/documents/generaldocuments/cms_040980.pdf [accessed 1 October 2007].
Aris RM, Christian D, Hearne PQ, Kerr K, Finkbeiner WE, Balmes JR. 1993. Ozone-induced airway inflammation in human subjects as determined by airway lavage and biopsy. Am Rev Respir Dis 148(5):1363-1372 https://pubmed.ncbi.nlm.nih.gov/8239177/.
Arsalane K, Gosset P, Vanhee D, Voisin C, Hamid Q, Tonnel ABet al. 1995. Ozone stimulates synthesis of inflammatory cytokines by alveolar macrophages in vitro. Am J Respir Cell Mol Biol 13(1):60-68 https://pubmed.ncbi.nlm.nih.gov/7598938/.
Atkinson JJ, Senior RM. 2003. Matrix metalloproteinase-9 in lung remodeling. Am J Respir Cell Mol Biol 28(1):12-24 https://pubmed.ncbi.nlm.nih.gov/12495928/.
Bell ML, Dominici F, Samet JM. 2005. A meta-analysis of time-series studies of ozone and mortality with comparison to the national morbidity, mortality, and air pollution study. Epidemiology 16(4):436-445 https://pubmed.ncbi.nlm.nih.gov/15951661/.
Betsuyaku T, Shipley JM, Liu Z, Senior RM. 1999. Neutrophil emigration in the lungs, peritoneum, and skin does not require gelatinase B. Am J Respir Cell Mol Biol 20(6):1303-1309 https://pubmed.ncbi.nlm.nih.gov/10340950/.
Bradford MM. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248-254 https://pubmed.ncbi.nlm.nih.gov/942051/.
Cataldo D, Munaut C, Noel A, Frankenne F, Bartsch P, Foidart JMet al. 2000. MMP-2- and MMP-9-linked gelatinolytic activity in the sputum from patients with asthma and chronic obstructive pulmonary disease. Int Arch Allergy Immunol 123(3):259-267 https://pubmed.ncbi.nlm.nih.gov/11112863/.
Chakrabarti S, Patel KD. 2005. Matrix metalloproteinase-2 (MMP-2) and MMP-9 in pulmonary pathology. Exp Lung Res 31(6):599-621 https://pubmed.ncbi.nlm.nih.gov/16019990/.
Chitano P, Hosselet JJ, Mapp CE, Fabbri LM. 1995. Effect of oxidant air pollutants on the respiratory system: insights from experimental animal research. Eur Respir J 8(8):1357-1371 https://pubmed.ncbi.nlm.nih.gov/7489804/.
Cho HY, Morgan D, Bauer AK, Kleeberger SR. 2007. Signal trans-duction pathways of tumor necrosis factor-mediated lung injury induced by ozone in mice. Am J Respir Crit Care Med 175(8):829-839 https://pubmed.ncbi.nlm.nih.gov/17255564/.
Corry DB, Kiss A, Song LZ, Song L, Xu J, Lee SHet al. 2004. Overlapping and independent contributions of MMP2 and MMP9 to lung allergic inflammatory cell egression through decreased CC chemokines. FASEB J 18(9):995-997 https://pubmed.ncbi.nlm.nih.gov/15059974/.
Corry DB, Rishi K, Kanellis J, Kiss A, Song Lz LZ, Xu Jet al. 2002. Decreased allergic lung inflammatory cell egression and increased susceptibility to asphyxiation in MMP2-deficiency. Nat Immunol 3(4):347-353 https://pubmed.ncbi.nlm.nih.gov/11887181/.
Dahl M, Bauer AK, Arredouani M, Soininen R, Trygvasson K, Kleeberger SRet al. 2007. Protection against inhaled oxidants through scavenging of oxidized lipids by macrophage receptors MARCO and SR-AI/II. J Clin Invest 117:757-764 https://pubmed.ncbi.nlm.nih.gov/17332894/.
D’Haese A, Wuyts A, Dillen C, Dubois B, Billiau A, Heremans Het al. 2000. In vivo neutrophil recruitment by granulocyte chemotactic protein-2 is assisted by gelatinase B/MMP-9 in the mouse. J Interferon Cytokine Res 20(7):667-674 https://pubmed.ncbi.nlm.nih.gov/10926210/.
Devlin RB, McDonnell WF, Mann R, Becker S, House DE, Schreinemachers Det al. 1991. Exposure of humans to ambient levels of ozone for 6.6 hours causes cellular and biochemical changes in the lung. Am J Respir Cell Mol Biol 4(1):72-81 https://pubmed.ncbi.nlm.nih.gov/1846079/.
Dunsmore SE, Saarialho-Kere UK, Roby JD, Wilson CL, Matrisian LM, Welgus HGet al. 1998. Matrilysin expression and function in airway epithelium. J Clin Invest 102(7):1321-1331 https://pubmed.ncbi.nlm.nih.gov/9769324/.
Elkington PT, Friedland JS. 2006. Matrix metalloproteinases in destructive pulmonary pathology. Thorax 61(3):259-266 https://pubmed.ncbi.nlm.nih.gov/16227332/.
Fukuda Y, Ishizaki M, Kudoh S, Kitaichi M, Yamanaka N. 1998. Localization of matrix metalloproteinases-1, -2, and -9 and tissue inhibitor of metalloproteinase-2 in interstitial lung diseases. Lab Invest 78(6):687-698 https://pubmed.ncbi.nlm.nih.gov/9645759/.
Goetzl EJ, Banda MJ, Leppert D. 1996. Matrix metalloproteinases in immunity. J Immunol 156(1):1-4 https://pubmed.ncbi.nlm.nih.gov/8598448/.
Johnston RA, Mizgerd JP, Shore SA. 2005a. CXCR2 is essential for maximal neutrophil recruitment and methacholine responsiveness after ozone exposure. Am J Physiol Lung Cell Mol Physiol 288(1):L61-L67 https://pubmed.ncbi.nlm.nih.gov/15361358/.
Johnston RA, Schwartzman IN, Flynt L, Shore SA. 2005b. Role of interleukin-6 in murine airway responses to ozone. Am J Physiol Lung Cell Mol Physiol 288(2):L390-397 https://pubmed.ncbi.nlm.nih.gov/15516495/.
Kelly EA, Busse WW, Jarjour NN. 2000. Increased matrix metalloproteinase-9 in the airway after allergen challenge. Am J Respir Crit Care Med 162(3 Pt 1):1157-1161 https://pubmed.ncbi.nlm.nih.gov/10988146/.
Kenyon NJ, van der Vliet A, Schock BC, Okamoto T, McGrew GM, Last JA. 2002. Susceptibility to ozone-induced acute lung injury in iNOS-deficient mice. Am J Physiol Lung Cell Mol Physiol 282(3):L540-L545 https://pubmed.ncbi.nlm.nih.gov/11839550/.
Kheradmand F, Rishi K, Werb Z. 2002. Signaling through the EGF receptor controls lung morphogenesis in part by regulating MT1-MMP-mediated activation of gelatinase A/MMP2. J Cell Sci 115(Pt 4):839-848 https://pubmed.ncbi.nlm.nih.gov/11865039/.
Kleeberger SR, Levitt RC, Zhang L-Y. 1993. Susceptibility to ozone-induced inflammation. I. Genetic control of the response to subacute exposure. Am J Physiol 264(1 pt 1):L15-L20 https://pubmed.ncbi.nlm.nih.gov/8430812/.
Kleeberger SR, Levitt RC, Zhang LY, Longphre M, Harkema J, Jedlicka Aet al. 1997. Linkage analysis of susceptibility to ozone-induced lung inflammation in inbred mice. Nat Genet 17(4):475-478 https://pubmed.ncbi.nlm.nih.gov/9398854/.
Koren HS, Devlin RB, Graham DE, Mann R, McGee MP, Horstman DHet al. 1989. Ozone-induced inflammation in the lower airways of human subjects. Am Rev Respir Dis 139(2):407-415 https://pubmed.ncbi.nlm.nih.gov/2913889/.
Krishna MT, Madden J, Teran LM, Biscione GL, Lau LC, Withers NJet al. 1998. Effects of 0.2 ppm ozone on biomarkers of inflammation in bronchoalveolar lavage fluid and bronchial mucosa of healthy subjects. Eur Respir J 11(6):1294-1300 https://pubmed.ncbi.nlm.nih.gov/9657569/.
Lanchou J, Corbel M, Tanguy M, Germain N, Boichot E, Theret Net al. 2003. Imbalance between matrix metalloproteinases (MMP-9 and MMP-2) and tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) in acute respiratory distress syndrome patients. Crit Care Med 31(2):536-542 https://pubmed.ncbi.nlm.nih.gov/12576963/.
Lanone S, Zheng T, Zhu Z, Liu W, Lee CG, Ma Bet al. 2002. Overlapping and enzyme-specific contributions of matrix metalloproteinases-9 and -12 in IL-13-induced inflammation and remodeling. J Clin Invest 110(4):463-474 https://pubmed.ncbi.nlm.nih.gov/12189240/.
Laskin DL, Fakhrzadeh L, Heck DE, Gerecke D, Laskin JD. 2002. Upregulation of phosphoinositide 3-kinase and protein kinase B in alveolar macrophages following ozone inhalation. Role of NF-kappaB and STAT-1 in ozone-induced nitric oxide production and toxicity. Mol Cell Biochem 234–235(1–2):91-98.
Lemjabbar H, Gosset P, Lamblin C, Tillie I, Hartmann D, Wallaert Bet al. 1999a. Contribution of 92 kDa gelatinase/type IV collagenase in bronchial inflammation during status asthmaticus. Am J Respir Crit Care Med 159(4 pt 1):1298-1307 https://pubmed.ncbi.nlm.nih.gov/10194181/.
Lemjabbar H, Gosset P, Lechapt-Zalcman E, Franco-Montoya ML, Wallaert B, Harf Aet al. 1999b. Overexpression of alveolar macrophage gelatinase B (MMP-9) in patients with idiopathic pulmonary fibrosis: effects of steroid and immunosuppressive treatment. Am J Respir Cell Mol Biol 20(5):903-913 https://pubmed.ncbi.nlm.nih.gov/10226060/.
Li Q, Park PW, Wilson CL, Parks WC. 2002. Matrilysin shedding of syndecan-1 regulates chemokine mobilization and transepithelial efflux of neutrophils in acute lung injury. Cell 111(5):635-646 https://pubmed.ncbi.nlm.nih.gov/12464176/.
Mautino G, Oliver N, Chanez P, Bousquet J, Capony F. 1997. Increased release of matrix metalloproteinase-9 in bronchoalveolar lavage fluid and by alveolar macrophages of asthmatics. Am J Respir Cell Mol Biol 17(5):583-591 https://pubmed.ncbi.nlm.nih.gov/9374109/.
McQuibban GA, Gong JH, Tam EM, McCulloch CA, Clark-Lewis I, Overall CM. 2000. Inflammation dampened by gelatinase A cleavage of monocyte chemoattractant protein-3. Science 289(5482):1202-1206 https://pubmed.ncbi.nlm.nih.gov/10947989/.
Mohan MJ, Seaton T, Mitchell J, Howe A, Blackburn K, Burkhart Wet al. 2002. The tumor necrosis factor-alpha converting enzyme (TACE): a unique metalloproteinase with highly defined substrate selectivity. Biochemistry 41(30):9462-9469 https://pubmed.ncbi.nlm.nih.gov/12135369/.
Mohan R, Chintala SK, Jung JC, Villar WV, McCabe F, Russo LAet al. 2002. Matrix metalloproteinase gelatinase B (MMP-9) coordinates and effects epithelial regeneration. J Biol Chem 277(3):2065-2072 https://pubmed.ncbi.nlm.nih.gov/11689563/.
Nagase H, Woessner JF. 1999. Matrix metalloproteinases. J Biol Chem 274(31):21491-21494 https://pubmed.ncbi.nlm.nih.gov/10419448/.
Pardo A, Barrios R, Maldonado V, Melendez J, Perez J, Ruiz Vet al. 1998. Gelatinases A and B are up-regulated in rat lungs by subacute hyperoxia: pathogenetic implications. Am J Pathol 153(3):833-844 https://pubmed.ncbi.nlm.nih.gov/9736032/.
Parks WC, Shapiro SD. 2001. Matrix metalloproteinases in lung biology. Respir Res 2(1):10-19 https://pubmed.ncbi.nlm.nih.gov/11686860/.
Parks WC, Wilson CL, Lopez-Boado YS. 2004. Matrix metalloproteinases as modulators of inflammation and innate immunity. Nat Rev Immunol 4(8):617-629 https://pubmed.ncbi.nlm.nih.gov/15286728/.
Perez-Ramos J, Segura-Valdez MD, Vanda B, Selman M, Pardo A. 1999. Matrix metalloproteinases 2, 9, and 13, and tissue inhibitors of metalloproteinases 1 and 2 in experimental lung silicosis. Am J Respir Crit Care Med 160:1274-1282 https://pubmed.ncbi.nlm.nih.gov/10508819/.
Russell RE, Culpitt SV, DeMatos C, Donnelly L, Smith M, Wiggins Jet al. 2002. Release and activity of matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 by alveolar macrophages from patients with chronic obstructive pulmonary disease. Am J Respir Cell Mol Biol 26(5):602-609 https://pubmed.ncbi.nlm.nih.gov/11970913/.
Schelegle ES, Siefkin AD, McDonald RJ. 1991. Time course of ozone-induced neutrophilia in normal humans. Am Rev Respir Dis 143(6):1353-1358 https://pubmed.ncbi.nlm.nih.gov/2048824/.
Schonbeck U, Mach F, Libby P. 1998. Generation of biologically active IL-1 beta by matrix metalloproteinases: a novel caspase-1-independent pathway of IL-1 beta processing. J Immunol 161(7):3340-3346 https://pubmed.ncbi.nlm.nih.gov/9759850/.
Seltzer J, Bigby BG, Stulbarg M, Holtzman MJ, Nadel JA, Ueki IFet al. 1986. O3-induced change in bronchial reactivity to methacholine and airway inflammation in humans. J Appl Physiol 60(4):1321-1326 https://pubmed.ncbi.nlm.nih.gov/3084448/.
Sternlicht MD, Werb Z. 2001. How matrix metalloproteinases regulate cell behavior. Annu Rev Cell Dev Biol 17:463-516 https://pubmed.ncbi.nlm.nih.gov/11687497/.
Tan RJ, Fattman CL, Niehouse LM, Tobolewski JM, Hanford LE, Li Qet al. 2006. Matrix metalloproteinases promoted inflammation and fibrosis in asbestosis-induced lung injury in mice. Am J Respir Cell Mol Biol 35:289-297 https://pubmed.ncbi.nlm.nih.gov/16574944/.
Valacchi G, Pagnin E, Okamoto T, Corbacho AM, Olano E, Davis PAet al. 2003. Induction of stress proteins and MMP-9 by 0.8 ppm of ozone in murine skin. Biochem Biophys Res Commun 305(3):741-746 https://pubmed.ncbi.nlm.nih.gov/12763055/.
Van den Steen PE, Proost P, Wuyts A, Van Damme J, Opdenakker G. 2000. Neutrophil gelatinase B potentiates interleukin-8 tenfold by aminoterminal processing, whereas it degrades CTAP-III, PF-4, and GRO-alpha and leaves RANTES and MCP-2 intact. Blood 96(8):2673-2681 https://pubmed.ncbi.nlm.nih.gov/11023497/.
Visse R, Nagase H. 2003. Matrix metalloproteinases and tissue inhibitors of metalloproteinases: structure, function, and biochemistry. Circ Res 92(8):827-839 https://pubmed.ncbi.nlm.nih.gov/12730128/.
Wilson CL, Matrisian LM. 1996. Matrilysin: an epithelial matrix metalloproteinase with potentially novel functions. Int J Biochem Cell Biol 28(2):123-136 https://pubmed.ncbi.nlm.nih.gov/8729000/.
Yu Q, Stamenkovic I. 2000. Cell surface-localized matrix metalloproteinase-9 proteolytically activates TGF-beta and promotes tumor invasion and angiogenesis. Genes Dev 14(2):163-176 https://pubmed.ncbi.nlm.nih.gov/10652271/.
Zhang K, McQuibban GA, Silva C, Butler GS, Johnston JB, Holden Jet al. 2003. HIV-induced metalloproteinase processing of the chemokine stromal cell derived factor-1 causes neurodegeneration. Nat Neurosci 6(10):1064-1071 https://pubmed.ncbi.nlm.nih.gov/14502291/.
Zuo F, Kaminski N, Eugui E, Allard J, Yakhini Z, Ben-Dor Aet al. 2002. Gene expression analysis reveals matrilysin as a key regulator of pulmonary fibrosis in mice and humans. Proc Natl Acad Sci USA 99(9):6292-6297 https://pubmed.ncbi.nlm.nih.gov/11983918/.
Information & Authors
Information
Published In
Environmental Health Perspectives
Volume 115 • Issue 11 • November 2007
Pages: 1557 - 1563
PubMed: 18007984
License Information
EHP is an open-access journal published with support from the National Institute of Environmental Health Sciences, National Institutes of Health. All content is public domain unless otherwise noted.
History
Received: 21 March 2007
Accepted: 24 August 2007
Published online: 24 August 2007
Keywords
Authors
Metrics & Citations
Metrics
Citations
Download citation
If you have the appropriate software installed, you can download article citation data to the citation manager of your choice. Simply select your manager software from the list below and click DOWNLOAD.
Cited by
- Lin C, Chen Y, Lin M, Chang H, Yang X, Lin C, ACE2 and a Traditional Chinese Medicine Formula NRICM101 Could Alleviate the Inflammation and Pathogenic Process of Acute Lung Injury, Medicina, 10.3390/medicina59091554, 59, 9, (1554), (2023).
- Hunter R, Baird B, Garcia M, Begay J, Goitom S, Lucas S, Herbert G, Scieszka D, Padilla J, Brayer K, Ottens A, Suter M, Barrozo E, Hines C, Bleske B, Campen M, Gestational ozone inhalation elicits maternal cardiac dysfunction and transcriptional changes to placental pericytes and endothelial cells, Toxicological Sciences, 10.1093/toxsci/kfad092, 196, 2, (238-249), (2023).
- Bougault V, Turmel J, Boulet L, Serum and sputum MMP-9/TIMP-1 in winter sports athletes and swimmers: relationships with airway function, Biomarkers, 10.1080/1354750X.2021.2020902, 27, 2, (127-137), (2022).
- Raeeszadeh-Sarmazdeh M, Coban M, Mahajan S, Hockla A, Sankaran B, Downey G, Radisky D, Radisky E, Engineering of tissue inhibitor of metalloproteinases TIMP-1 for fine discrimination between closely related stromelysins MMP-3 and MMP-10, Journal of Biological Chemistry, 10.1016/j.jbc.2022.101654, 298, 3, (101654), (2022).
- Tong Y, Bao C, Xu Y, Tao L, Zhou Y, Zhuang L, Meng Y, Zhang H, Xue J, Wang W, Zhang L, Pan Q, Shao Z, Hu T, Guo Q, Xue Q, Lu H, Luo Y, The β3/5 Integrin-MMP9 Axis Regulates Pulmonary Inflammatory Response and Endothelial Leakage in Acute Lung Injury, Journal of Inflammation Research, 10.2147/JIR.S331939, Volume 14, (5079-5094), (2021).
- Smith G, Tovar A, Kanke M, Wang Y, Deshane J, Sethupathy P, Kelada S, Ozone‐induced changes in the murine lung extracellular vesicle small RNA landscape, Physiological Reports, 10.14814/phy2.15054, 9, 18, (2021).
- Tong Y, Yu Z, Chen Z, Zhang R, Ding X, Yang X, Niu X, Li M, Zhang L, Billiar T, Pitt B, Li Q, The HIV protease inhibitor Saquinavir attenuates sepsis-induced acute lung injury and promotes M2 macrophage polarization via targeting matrix metalloproteinase-9, Cell Death & Disease, 10.1038/s41419-020-03320-0, 12, 1, (2021).
- Gao Y, Wang F, Liu Q, Qi Y, Wang Q, Liu H, Comparison of anti-inflammatory effects of Lonicerae Japonicae Flos and Lonicerae Flos based on network pharmacology, Chinese Herbal Medicines, 10.1016/j.chmed.2021.06.005, 13, 3, (332-341), (2021).
- Sarkar J, Chakraborti T, Chakraborti S, Role of NADPH Oxidase-Induced Oxidative Stress in Matrix Metalloprotease-Mediated Lung Diseases, Oxidative Stress in Lung Diseases, 10.1007/978-981-32-9366-3_4, (75-101), (2019).
- Shaffo F, Grodzki A, Fryer A, Lein P, Mechanisms of organophosphorus pesticide toxicity in the context of airway hyperreactivity and asthma, American Journal of Physiology-Lung Cellular and Molecular Physiology, 10.1152/ajplung.00211.2018, 315, 4, (L485-L501), (2018).