Presence Of Matrix Metalloproteinase-9 Research Articles

Validation of a rapid collagenase activity detection technique based on fluorescent quenched gelatin with synovial fluid samples

BackgroundMeasuring collagenase activity is crucial in the field of joint health and disease management. Collagenases, enzymes responsible for collagen degradation, play a vital role in maintaining the balance between collagen synthesis and breakdown in joints. Dysregulation of collagenase activity leads to joint tissue degradation and diseases such as rheumatoid arthritis and osteoarthritis. The development of methods to measure collagenase activity is essential for diagnosis, disease severity assessment, treatment monitoring, and identification of therapeutic targets.ResultsThis study aimed to validate a rapid collagenase activity detection technique using synovial fluid samples. Antibody microarray analysis was initially performed to quantify the levels of matrix metalloproteinase-9 (MMP-9), a major collagenase in joints. Subsequently, the developed gelatin-based test utilizing fluorescence measurement was used to determine collagenase activity. There was a significant correlation between the presence of MMP-9 and collagenase activity. In addition, Lower Limit of Detection and Upper Limit of Detection can be preliminary estimated as 8 ng/mL and 48 ng/mL respectively.ConclusionsThe developed technique offers a potential point-of-care assessment of collagenase activity, providing real-time information for clinicians and researchers. By accurately quantifying collagenase activity, healthcare professionals can optimize patient care, improve treatment outcomes, and contribute to the understanding and management of joint-related disorders. Further research and validation are necessary to establish the full potential of this rapid collagenase activity detection method in clinical practice.

Fabrication and Characterization of an Enzyme-Triggered, Therapeutic-Releasing Hydrogel Bandage Contact Lens Material.

The purpose of this study was to develop an enzyme-triggered, therapeutic-releasing bandage contact lens material using a unique gelatin methacrylate formulation (GelMA+). Two GelMA+ formulations, 20% w/v, and 30% w/v concentrations, were prepared through UV polymerization. The physical properties of the material, including porosity, tensile strain, and swelling ratio, were characterized. The enzymatic degradation of the material was assessed in the presence of matrix metalloproteinase-9 (MMP-9) at concentrations ranging from 0 to 300 µg/mL. Cell viability, cell growth, and cytotoxicity on the GelMA+ gels were evaluated using the AlamarBlueTM assay and the LIVE/DEADTM Viability/Cytotoxicity kit staining with immortalized human corneal epithelial cells over 5 days. For drug release analysis, the 30% w/v gels were loaded with 3 µg of bovine lactoferrin (BLF) as a model drug, and its release was examined over 5 days under various MMP-9 concentrations. The 30% w/v GelMA+ demonstrated higher crosslinking density, increased tensile strength, smaller pore size, and lower swelling ratio (p < 0.05). In contrast, the 20% w/v GelMA+ degraded at a significantly faster rate (p < 0.001), reaching almost complete degradation within 48 h in the presence of 300 µg/mL of MMP-9. No signs of cytotoxic effects were observed in the live/dead staining assay for either concentration after 5 days. However, the 30% w/v GelMA+ exhibited significantly higher cell viability (p < 0.05). The 30% w/v GelMA+ demonstrated sustained release of the BLF over 5 days. The release rate of BLF increased significantly with higher concentrations of MMP-9 (p < 0.001), corresponding to the degradation rate of the gels. The release of BLF from GelMA+ gels was driven by a combination of diffusion and degradation of the material by MMP-9 enzymes. This work demonstrated that a GelMA+-based material that releases a therapeutic agent can be triggered by enzymes found in the tear fluid.

Signal-off electrochemical sensor for matrix metalloproteinase 9 detection based on sacrificial FeMOF and host-guest strategy

Matrix metalloproteinase-9 (MMP-9) has been implicated in various tumor cell invasions and metastases. In light of the limitations of traditional methods for MMP-9 detection, we have constructed a novel biosensor depending on cucurbit[8]uril (CB[8]) -mediated host-guest interactions and a sacrificial iron metal-organic framework (FeMOF). Herein, MMP9-specific peptides modified on the gold bare electrode are bonded to the FeMOF@AuNPs@peptide complex through CB[8] addition. The connection between MMP9-specific peptides and signal peptides via CB[8] provides stability as well as enables the immobilization of FeMOF on the electrode surface. When Fe3+ from the FeMOF interacts with electrochemical buffer K4Fe(CN)6, Prussian blue will be generated on the gold electrode surface, and a significantly enlarged current response can be detected. However, in the presence of MMP-9, their peptide substrates are specifically cleaved at the site between serine (S) and Leucine (L), which causes an abrupt decrease in the electrochemical signal. The change of signal can reflect MMP-9 concentration. This sensor can reach an ultrahigh sensitivity with a wide detection range of 0.5 pg⋅mL−1 to 500 ng⋅mL−1 and a low detection limit of 1.30 pg⋅mL−1. Importantly, this sensor is very simple, relying solely on self-sacrificial label of FeMOF, rather than complex functional materials. Additionally, it has been well used in serum samples, showing attractive potential for practical applications.

MP03-02 ASSESSMENT OF MATRIX METALLOPROTEINASE-9 AND TISSUE INHIBITOR OF METALLOPROTEINASE-1 VIA M2 MACROPHAGE-SEEDED COLLAGEN MESH FOR TREATMENT OF STRESS URINARY INCONTENCE

You have accessJournal of UrologyCME1 Apr 2023MP03-02 ASSESSMENT OF MATRIX METALLOPROTEINASE-9 AND TISSUE INHIBITOR OF METALLOPROTEINASE-1 VIA M2 MACROPHAGE-SEEDED COLLAGEN MESH FOR TREATMENT OF STRESS URINARY INCONTENCE Ilaha Isali, Phillip McClellan, Thomas R. Wong, James M. Anderson, Ozan Akkus, and Adonis Hijaz Ilaha IsaliIlaha Isali More articles by this author , Phillip McClellanPhillip McClellan More articles by this author , Thomas R. WongThomas R. Wong More articles by this author , James M. AndersonJames M. Anderson More articles by this author , Ozan AkkusOzan Akkus More articles by this author , and Adonis HijazAdonis Hijaz More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003214.02AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Over the past decade, there have been several innovations with respect to macrophage-based therapies and biomaterials. Patients suffering from stress urinary incontinence (SUI) exhibit a significant increase in matrix metalloproteinase-9 (MMP-9) and a significant decrease in tissue inhibitor of metalloproteinase-1 (TIMP-1) expression. These findings are consistent with elevated rates of collagen breakdown and may play a critical role in the onset and development of SUI. Our recent study demonstrated improved biomechanical and histological outcomes following incorporation of M2 macrophages into a collagen mesh developed for tissue repair and offers future strategies focused on connective tissue regeneration. This study aims to assess the regulation of MMP-9 and TIMP-1 by M2 macrophages in terms of regenerating connective tissue in a rat model. METHODS: Macrophages were harvested from rat bone marrow, and macrophage subtypes were identified by flow cytometry. Electrochemically aligned collagen threads were filament wound to produce small collagen meshes and seeded with macrophages and cultured for up to 3 days. Macrophage subtype-seeded meshes were implanted subcutaneously in rats and harvested at 3 weeks and 3 months, respectively. Explanted meshes were analyzed using histology and immunohistochemistry. Specimens stained separately for MMP-9 and TIMP-1, and quantified to determine overall presence of MMP-9 and TIMP-1 in the harvested tissues. RESULTS: Immunohistochemical staining demonstrated a reduction of MMP-9 with addition of M2 macrophages, whereas the M1-seeded group continued to exhibit elevated MMP-9 after 3 months. TIMP-1 presence was elevated with the addition of M2 macrophages, whereas the M1-seeded group showed low TIMP-1 after 3 months (Figure 1). CONCLUSIONS: Delivery of M2 macrophages via collagen mesh can reduce MMP-9 while increasing TIMP-1, which may limit collagen breakdown associated with the development of SUI. This study shows the potential for improved restoration of extracellular matrix following the incorporation of M2 macrophages into a genipin-crosslinked collagen mesh for tissue repair and remodeling. Source of Funding: NIH grant: R21HD095439 © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e21 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Ilaha Isali More articles by this author Phillip McClellan More articles by this author Thomas R. Wong More articles by this author James M. Anderson More articles by this author Ozan Akkus More articles by this author Adonis Hijaz More articles by this author Expand All Advertisement PDF downloadLoading ...

Abstract PS16-26: The prognostic role of MMP 9 in early breast cancer

Abstract Background: Matrix metalloproteinase 9 (MMP9) is involved in the extracellular matrix degradation during physiological and pathological conditions including tumorigenesis. This translational study was aimed to evaluate the prognostic role of the intratumoral MMP9 expression and correlate it with presence of CTCs in early breast cancer. Methods: A total of 318 primary breast cancer (PBC) patients were enrolled into this study. Surgical specimens were processed by the tissue microarray method and subjected to immunohistochemistry using the MMP9 monoclonal antibody. The MMP9 expression was evaluated in tumor cell as well as in tumor associated stroma. Quantitative real-time polymerase chain reaction -based assays was applied for identification of CTCs. Results: Significantly increased expression of MMP9 was found in breast cancer cells when compared to tumor associated stroma. A positive correlation was determined between MMP9 expression and hormone positive status as well as low proliferation index of analysed breast cancer tumour cells. Additionally, in tumor associated stroma was confirmed only the association with hormone receptor status. The univariate survival analysis of whole tested population detected no prognostic role of MMP9 expression neither in tumor cells (HR = 0.96, 95% CI 0.58-1.59, P = 0.864) nor in tumor associated stroma (HR = 1.29, 95% CI 0.60-2.78, P = 0.547). However, the subgroup of in hormone receptor negative and triple negative patients with absence of MMP9 expression in tumor cells and stroma had significantly better disease-free survival (DFS) (HR = 0.33, 95% CI 0.12-0.93, P = 0.025, and (HR = 0.14, 95% CI 0.00-4.81, P = 0.002, respectively) and (HR = 0.17, 95% CI 0.05-0.57, P = 0.003); (HR = 0.12, 95% CI 0.00-4.89, P = 0.001) compared with patients with presence of MMP9. Moreover, while tumor MMP9 was prognostic in CTC_EMT positive subgroup (HR = 0.40, 95% CI 0.16-0.95, P = 0.047), absence of stromal MMP9 had protective role in CTC_EP positive patients (HR = 0.18, 95% CI 0.01-2.75, P = 0.053). Conclusions: Our data suggest that the increased expression of MMP9 in PBC was related with favorable tumor characteristics. However, it´s prognostic value was limited only to hormone receptor negative, triple negative, CTC_EMT and CTC_EP positive subgroups. Therefore, we can suppose that evaluating of MMP9 tumor expression could help identify patients with increased risk of disease recurrence in these subgroups of patients. Citation Format: Michal Mego, Zuzana Cierna, Marian Karaba, Gabriel Minarik, Juraj Benca, Tatiana Sedlackova, Ivana Mrvova, Daniel Pindak, Katarina Rejlekova, Jozef Mardiak, Katarina Kalavska. The prognostic role of MMP 9 in early breast cancer [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS16-26.

Intracerebral matrix metalloproteinase 9 in fatal diabetic ketoacidosis

There is increasing awareness that in addition to the metabolic crisis of diabetic ketoacidosis (DKA) caused by severe insulin deficiency, the immune inflammatory response is likely an active multicomponent participant in both the acute and chronic insults of this medical crisis, with strong evidence of activation for both the cytokine and complement system. Recent studies report that the matrix metalloproteinase enzymes and their inhibitors are systemically activated in young Type 1 diabetes mellitus (T1D) patients during DKA and speculate on their involvement in blood-brain barrier (BBB) disruption. Based on our previous studies, we address the question if matrix metalloproteinase 9 (MMP9) is expressed in the brain in the fatal brain edema (BE) of DKA. Our data show significant expression of MMP9 on the cells present in brain intravascular areas. The presence of MMP9 in intravascular cells and that of MMP+ cells seen passing the BBB indicates a possible role in tight junction protein disruption of the BBB, possibly leading to neurological complications including BE. We have also shown that MMP9 is expressed on neurons in the hippocampal areas of both BE/DKA cases investigated, while expression of tissue inhibitor of metalloproteinases 1 (TIMP1) was reduced in the same areas. We can speculate that intraneuronal MMP9 can be a sign of neurodegeneration. Further studies are necessary to determine the role of MMP9 in the pathogenesis of the neurologic catastrophe of the brain edema of DKA. Inhibition of MMP9 expression might be helpful in preserving neuronal function and BBB integrity during DKA.

A PEGylated hyaluronic acid conjugate for targeted cancer immunotherapy

The cell-free approach to foreignizing tumor cells with non-self antigens has received increasing attention as a method to induce cytotoxic T lymphocyte (CTL)-mediated immunological rejection of tumors, because the clinical translation of the conventional CTL-based cancer immunotherapies has been limited by a complicated manufacturing process and autotransplantation. In this study, we prepared matrix metalloproteinase 9 (MMP9)-responsive polymeric conjugates consisting of PEGylated hyaluronic acid (HA) as the targeting moiety and ovalbumin (OVA) as the model foreign antigen. The MMP9-cleavable linker was introduced between PEG and the HA backbone to facilitate the detachment of the PEG corona from the conjugate at the tumor site. From the in vitro cellular uptake study, it was revealed that the conjugate was effectively taken up by the CD44-expressing TC-1 cancer cells in the presence of MMP9 via receptor-mediated endocytosis. When the conjugate was systemically administered into the tumor-bearing mice with endogenous OVA-specific CTLs, the tumor growth was markedly inhibited, which was attributed to the significant antigen presentation on the tumor cells. Overall, the MMP9-responsive conjugates bearing foreign antigens might have the potential as an alternative to CTL-based cancer immunotherapeutics.

Expression of uterine lipocalin 2 and its receptor during early- to mid-pregnancy period in mares.

From previous cDNA subtraction studies analyzing gene expression in equine endometrium, high lipocalin 2 (LCN2) mRNA expression was found in the gravid endometrium. In the uterus, LCN2 may transport hydrophobic molecules and siderophores with iron, or may form a complex with another protein, however, the expression of uterine LCN2 beyond day 20 of equine pregnancy and its receptor has not been characterized. To study the expression and potential roles of uterine LCN2 from pre-implantation to mid-gestation period, stage-specific endometrial samples were obtained from day 13 (day 0 = ovulation) cyclic and days 13, 19, 25, and 60 to 131 pregnant mares. Expression of LCN2 mRNA increased in day 19 gravid endometrium and was abundant from day 60 onward. The expression of LCN2 mRNA was localized to the glandular epithelium. LCN2 protein was detected in day 25 gravid endometrium and luminal fluid, and the protein was localized to the glandular epithelium and luminal cavity, whereas LCN2 receptor expression was found in luminal and glandular epithelium and trophectoderm throughout the experimental period. The presence of matrix metalloproteinase-9 (MMP9) was also examined because MMP9 is known to form a complex with LCN2. Although MMP9 and LCN2 were both found in luminal fluid from day 25 pregnant uterus, the complex of these proteins was not detected. Localization of the receptor in the trophectoderm suggests that endometrial LCN2 could play a role in carrying small substances from the mother to fetus in the equine species.

Nanoparticle‐Bound Cytolytic Peptide Prodrugs for Cancer Therapy

ObjectiveTo develop a nanoparticle platform which promotes the delivery of cytolytic peptide prodrugs to tumors and their activation by locally produced proteases.Introduction: Melittin is a peptide toxin derived from bee venom which has been proposed as a cancer therapy due to its cytolytic activity. Yet, its therapeutic applications have been limited by its nonspecific cytotoxicity and rapid clearance from the circulation. We have previously developed a prodrug form of melittin which is activated upon cleavage by matrix metalloproteinase 9 (MMP‐9), a protease closely linked with tumor growth and metastasis. Here, we present a platform for delivery of this prodrug based on a perfluorocarbon nanoparticle system designed to enhance intratumoral accumulation while permitting prodrug activation by local MMP‐9 activity.Methods and Results: Incubation of the melittin prodrug with perfluorocarbon nanoparticles (NPs) resulted in stable loading of 10,250 peptides per NP with no significant change in NP size. Addition of 20 nM MMP‐9 led to activation of 57.8% of NP‐bound prodrug within 3 hours. B16F10 melanoma cell viability was virtually eliminated following incubation with 50 μM free or NP‐bound prodrug in the presence of MMP‐9 (p &lt; 0.001 compared to control). In a mouse model of B16F10 lung metastasis, NP accumulation in lung tumors was demonstrated by both fluorescence microscopy (Figure A; NPs in red) and quantitative 19F magnetic resonance spectroscopy (Figure B). Gelatin zymography confirmed production of active MMP‐9 by these tumors.ConclusionPerfluorocarbon nanoparticles accumulate within tumors and permit activation of bound prodrugs, making them promising delivery vehicles for cytolytic peptide agents.Research Support: NIH R01 HL073646 image image

Increased cyclooxygenase activity impairs apoptosis of inflammatory neutrophils in mice lacking gelatinase B/matrix metalloproteinase‐9

Matrix metalloproteinase-9 (MMP-9)/gelatinase B plays an important role in neutrophil infiltration during inflammation and cyclooxygenases (COX-1 and COX-2) and their products are important regulators of inflammation. Recently, we reported that a genetic lack of MMP-9 impairs neutrophil infiltration during early zymosan-induced peritonitis but at later stages (> 24 hr) neutrophils persist in the peritoneal cavity. Here we show that this is the result of impaired apoptosis of MMP-9(-/-)-derived leucocytes. As enhanced COX-1 expression was reported in MMP-9(-/-) mice, we evaluated the hypothesis that altered COX expression induced the above phenomenon as COX-dependent prostaglandins can act either anti-apoptotically (PGE(2)) or pro-apoptotically (PGD(2)). The current data demonstrate that messenger RNA and protein expression of both COX isoforms and their activities are increased in MMP-9(-/-) mice during late peritonitis. Application of selective COX inhibitors revealed enhanced COX-1-dependent PGE(2) production and impaired COX-2-dependent PGD(2) synthesis in MMP-9(-/-) mice. Most importantly, inhibition of COX-1 abolished prolonged neutrophil accumulation in the peritoneal cavity of MMP-9(-/-) mice and increased apoptosis of inflammatory leucocytes. Similarly, weaker apoptosis of MMP-9(-/-) bone marrow neutrophils treated in vitro with zymosan was reversed by COX-1 inhibition. In conclusion, enhanced COX-1 expression is responsible for persistent neutrophil presence in the peritoneum of MMP-9(-/-) mice because of increased synthesis of anti-apoptotic PGE(2). In non-transgenic mice, however, inflammatory leucocytes die apoptotically in the late stages of peritonitis as a result of COX-2-dependent PGD(2) activity. Overall, we show a dependence of COX expression on the presence of MMP-9.

Immunohistological Evaluation of the Healing Response at the Flap Interface in Patients With LASIK Ectasia Requiring Penetrating Keratoplasty

To evaluate the healing response at the flap interface in corneas with LASIK ectasia that required penetrating keratoplasty (PK). Corneas of five patients who developed corneal ectasia after LASIK (range: 2.5 to 5 years postoperative) were collected after corneal transplant surgery. The corneas were bisected and processed for conventional histologic analysis and immunofluorescence. Light microscopy showed a hypocellular fibrotic scar at the wound margin compared with the adjacent corneal stroma in all eyes. All corneas had positive staining for alpha-smooth muscle actin (SMA), a myofibroblast marker. In one eye, alpha-SMA cells were located in the fibrotic scar region in the area of the semicircular ring of haze along the margin of the LASIK flap corresponding to an area of epithelial ingrowth. In all other eyes, alpha-SMA positive cells were fewer and mainly located in the superficial stroma under the epithelial wound margin surface. Type III collagen was minimal or absent in the central zone and wound margin of all corneas except for the cornea with epithelial ingrowth present in the hypercellular fibrotic scar region. Chondroitin sulfate was stronger in the periphery of the flap wound coinciding with a higher presence of alpha-SMA-positive cells in that region. Positive staining for matrix metalloproteinase 9 (MMP-9) in the paracentral wound margin scar was seen. A wound-healing process characterized by absence of significant fibrosis and myofibroblasts at the wound edge in the flap interface was noted in all keratectatic eyes. However, changes in the composition of collagen and the presence of MMP-9 at the wound edge several years after LASIK indicates active wound remodeling that may explain the ongoing loss of tissue and tendency of the cornea to bulge.

γ-Aminbuturic Acid A Receptor Mitigates Homocysteine-Induced Endothelial Cell Permeability

Many cerebrovascular disorders are accompanied by an increased homocysteine (Hcy) levels. We have previously shown that acute hyperhomocysteinemia (HHcy) leads to an increased microvascular permeability in the mouse brain. Hcy competitively binds to gamma -aminbuturic acid (GABA) receptors and may increase vascular permeability by acting as an excitatory neurotransmitter. However, the role of GABA-A (GABA(A)) receptor in Hcy-induced endothelial cell (EC) permeability remains unclear. In the present study we attempted to determine the role of GABA(A) receptor and the possible mechanisms involved in Hcy-induced EC layer permeability. Mouse aortic and brain ECs were grown in Transwells and treated with 50 mu M Hcy in the presence or absence of GABA(A)-specific agonist muscimol. Role of matrix metalloproteinase-9 (MMP-9) was determined using its activity inhibitor GM-6001. Involvement of extracellular signal-regulated kinase (ERK) signaling was assessed using its kinase activity inhibitors PD98059 or U0126. EC permeability to the known content of bovine serum albumin (BSA)-conjugated with Alexa Flour-488 was assessed by measuring fluorescence intensity of the solutes in the Transwell's lower chambers. It was found that Hcy induced the formation of filamentous actin (F-actin). Hcy-induced EC permeability to BSA was significantly decreased by GABA and muscimol treatments. Presence of MMP-9 or ERK kinase activity inhibitors restored the Hcy-induced EC permeability to its baseline level. The mediation BSA leakage through the ECs was further confirmed in the experiments where Hcy-induced alterations in transendothelial electrical resistance of confluent ECs were assessed. The data suggest that Hcy increases EC layer permeability through inhibition of GABA(A) receptor and F-actin formation, in part, by transducing ERK and MMP-9 activation.

Matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1: expression in the lung of fetal rats with nitrofen-induced diaphragmatic hernia.

The surrounding extracellular matrix of airway wall tissues changes in response to mechanical stresses and hypoxia. The presence of matrix metalloproteinase-9 (MMP-9) and its inhibitor, tissue inhibitor of metalloproteinase-1 (TIMP-1), is correlated with collagen degradation and tissue repair in lung disorders. The aim of this study was to evaluate the expression of MMP-9 and TIMP-1 in the lung of fetal rats with nitrofen-induced congenital diaphragmatic hernia (CDH). Administering 100 mg of nitrofen dissolved in 1 ml olive oil to pregnant Wistar rats on day 9 of gestation induced left-sided CDH in fetal rats. In control animals, the same dose of olive oil was given without nitrofen. Cesarean section was performed on day 21 of gestation. The fetuses were divided into two groups: normal controls (n = 10) and nitrofen-induced left-sided CDH (n = 10). Immunoreactivity of the staining for MMP-9 and TIMP-1 in the lung tissues was semiquantitatively analyzed using the staining scores. The relative amount of MMP-9 or TIMP-1 divided by the amount of beta-actin for each lung sample was measured by using the real-time reverse-transcriptase polymerase chain reaction. The immunoreactivity of MMP-9 was significantly increased in the CDH group (n = 5) compared with the control group (n = 5) (p = 0.031). On the other hand, the immunoreactivity of TIMP-1 in the two groups was not significantly different (n = 0.134). The relative amount of MMP-9 (or TIMP-1) in the CDH group (n = 5) does not differ significantly from that in the control group (n = 5) (p = 0.059, 0.596, respectively), but the relative amount of MMP-9 is higher in the CDH group, although it is not significantly higher. On the other hand, the ratios of MMP-9 to TIMP-1 were significantly higher in the CDH group (p = 0.028). In conclusion, fetal rats with nitrofen-induced CDH, a model of respiratory disorders, manifested the excess of MMP-9 activity due to the absence of TIMP-1 that would suggest a trend toward disruption of the extracellular matrix in the CDH lung tissues.