Amounts Of Matrix Metalloproteinases Research Articles

Roles of NMDARs in maintenance of the mouse cerebrovascular endothelial cell-constructed tight junction barrier

Glutamate can activate NMDA receptor (NMDAR) and subsequently induces excitotoxic neuron loss. However, roles of NMDARs in the blood-brain barrier (BBB) are little known. This study used a mouse cerebrovascular endothelial cell (MCEC) model to evaluate the effects of NMDAR activation on maintenance of the BBB and its possible mechanisms. Analysis of confocal microscopy revealed expressions of NMDAR subunits, GluN1 and GLUN2B, in MCECs. An immunoblot assay further showed the existence of GluN1 in plasma membranes of MCECs. In brain tissues, a confocal microscopic analysis demonstrated co-localization of GluN1 and factor VIII, a biomarker of MCECs. In addition, GluN1 mRNA was detected in MCECs and the brain. Functional assays showed that exposure of MCECs to NMDA increased calcium influx. Separately, NMDA suppressed transendothelial electrical resistance values, levels of occludin, and occludin tight junctions. As to the mechanism, NMDA stimulated sequential phosphorylations of extracellular signal-regulated kinase (ERK)1/2 and mitogen-activated ERK (MEK)1. Interestingly, amounts of matrix metalloproteinase (MMP)2 and MMP9 in MCECs were augmented by NMDA. The NMDA-induced alterations in ERK1/2 phosphorylation and occludin levels were reversed by pretreatment with PD98059, a MEK inhibitor, and MK-801, a NMDAR antagonist, respectively. Therefore, this study shows the functional presence of NMDARs in MCECs, and NMDAR activation can disrupt the MCEC-constructed tight junction barrier via activation of the MEK1/2-ERK1/2 signaling pathway and upregulation of MMP2/9 expressions.

Prolonged Treatment with Inhaled Corticosteroids does not Normalize High Activity of Matrix Metalloproteinase-9 in Exhaled Breath Condensates of Children with Asthma

The airway remodeling in asthma is associated with increased amount of matrix metalloproteinase (MMP)-9. High levels of MMP-9 were found in mucosal biopsies, sputum and in exhaled breath condensates (EBC) of asthma patients. However, there are no data concerning real in vivo activity. Inhaled corticosteroids are effective in asthma control, but it is unclear, whether they only attenuate inflammation, or also protect against progressive remodeling of respiratory tract. Therefore, the aim of the study was to assess the amount and activity of MMP-9 in context of pro-inflammatory cytokines (IL-6, IL-8 and tumor necrosis factor, TNF), measured in EBC of asthma-suffering children, treated with inhaled steroids. The study involved 27 children with asthma, continuously treated with inhaled fluticasone propionate, and 22 healthy controls. In addition to routine clinical screening, the selected cytokines in EBC were analyzed using Ultrasensitive ELISA, whereas activity of MMP-9 was assessed using a novel immunozymography method. Despite chronic treatment with inhaled steroids mean MMP-9/EBC activity in asthma group was significantly higher than in healthy controls. Moreover, high MMP-9/EBC in asthma-suffering children significantly correlated with IgE serum levels. The IL-6 and IL-8 concentration was below the detection limit in all EBC samples. TNF/EBC levels were similar in both, asthma and healthy children. We hypothesize that MMP-9 hyperactivity in asthma may be closely related to high IgE serum levels. Our results suggest that inhaled steroids may be ineffective to prevent asthma-associated airway remodeling. Finally, we emphasize the necessity of further research focused on MMP-9 inhibition in asthma treatment.

GLP-1R agonist may activate pancreatic stellate cells to induce rat pancreatic tissue lesion

ObjectiveTo explore the mechanism of GLP-1R agonist-induced rat pancreatic tissue lesion. MethodsThirty SD male rats were divided into three groups, namely GLP-1R agonist experimental group, diabetes-model experimental group and control group. Diabetes-model rats were induced by streptozotocin and high-sugar high-fat diet. GLP-1R agonist group and diabetes-model group were administered with GLP-1R agonist in dose 5 μg/kg each time, twice a day. After 10 weeks of treatment, the amount of matrix metalloproteinase (MMP)-2 and MMP-9, and expression of α-smooth muscle actin (α-SMA) and type III collagen protein in pancreatic tissue were measured. ResultsThe amount of MMP-2 and MMP-9 in GLP-1R agonist group and diabetes-model group were significantly higher than the control group. Compared with the GLP-1R agonist group, the diabetic model group had more severe pathological changes of pancreatic tissue interstitial edema, inflammatory cell infiltration, glandular atrophy and fibrosis, and significantly increased pancreatic tissue MMP-2 and MMP-9 levels, significantly increased α-SMA and collagen III-positive cell counts, all the differences were statistically significant. α-SMA and type III collagen were expressed in all parts of the lesions of GLP-1R agonist group and diabetes-model group. α-SMA can only be observed in the vessel wall in control group, however, in the other two groups, α-SMA can also be observed in pancreatic acinar cell interstitia, in addition to vessel wall. ConclusionsLong-term subcutaneous GLP-1R agonist injection may activate pancreatic stellate cells, causing the expression of α-SMA and collagen III and the amount of MMP-2 and MMP-9 in pancreatic acinar cell interstitial significantly increasing, and thus inducing chronic inflammatory change.

Regional odontodysplasia: Orthodontic treatment and transplantation of premolars

Regional odontodysplasia is a rare and unique dental anomaly involving both dentitions, but mostly the teeth of 1 quadrant. This report describes the combined surgical and orthodontic treatment of a boy with regional odontodysplasia. The mandibular right central and lateral incisors and the canine (as well as the deciduous predecessors) were affected. In a 2-step procedure, the maxillary right and left second premolars were autotransplanted to the affected area. The extraction sites in the maxilla were closed, and a good functional occlusion was established.

Effect of Sera from Bickerstaff Brainstem Encephalitis and Miller Fisher Syndrome Patients Against Human Blood-Brain Barrier and Blood-Nerve Barrier In Vitro Models (P06.141)

Objective: Antibody against GQ1b is frequently detected in Bickerstaff brainstem encephalitis (BBE) and Miller Fisher syndrome (MFS) and has been considered to be pathogenically important in the development of these two disorders; however, the clinical manifestations of there two disorders are obviously different. We hypothesized that there phenotypic differences may be derived from the difference of blood-brain barrier (BBB)/ blood-nerve barrier(BNB) breakdown. Background We demonstrated the effects of sera from patients with BBE and MFS on the impairment of BBB or BNB function and clarified the roles of humoral factor such as Matrix metalloproteinases(MMP) in the destruction of BBB or BNB. Design/Methods: Both human BBB and BNB-derived endothelial cell lines were recently developed in Yamaguchi University, named TY10 and PnMECs. We evaluated transendothelial electrical resistance (TEER), the amount of tight junction proteins including claudin-5 and occludin, and the amount of MMP including MMP-9 and MMP-2 by western blotting in TY10 and PnMECs after the exposure of the patients9 sera. Results: The amount of claudin-5 protein in TY10, not in PnMECs, decreased after exposure of BBE sera. TEER of TY10 also decreased after application of BBE sera. Sera from MFS patients had no effect in these experiments. The amount of MMP-9 and MMP-2 protein in TY10 increased after exposure of BBE sera. Conclusions: Only sera from BBE patients disrupt BBB. This may partially explain the phenotypic defferences between BBE and MFS. BBE sera break BBB, possibly via autocrine secretion of MMP-9 and MMP-2 from BBB-composing endothelial cells. Disclosure: Dr. Saito has nothing to disclose. Dr. Shimizu has nothing to disclose. Dr. Koga has nothing to disclose. Dr. Sano has nothing to disclose. Dr. Haruki has nothing to disclose. Dr. Maeda has nothing to disclose. Dr. Abe has nothing to disclose. Dr. Tasaki has nothing to disclose. Dr. Suzuki has nothing to disclose. Dr. Kusunoki has nothing to disclose. Dr. Mizusawa has received personal compensation for activities with Tanabemitsubishi Co and Sanofi-Aventis Co. Dr. Mizusawa has received personal compensation in an editorial capacity for No to Shinkei and Clinical Neuroscience. Dr. Kanda has nothing to disclose.

17β-estradiol reduces expression of MMP-1, -3, and -13 in human primary articular chondrocytes from female patients cultured in a three dimensional alginate system

Clinical observations have suggested a relationship between osteoarthritis and a changed sex-hormone metabolism, especially in menopausal women. This study analyzes the effect of 17β-estradiol on expression of matrix metalloproteinases-1, -3, -13 (MMP-1, -3, -13) and tissue inhibitors of metalloproteinases-1, -2 (TIMP-1, -2) in articular chondrocytes. An imbalance of matrix metalloproteinases (MMPs) specialized on degradation of articular cartilage matrix over the respective inhibitors of these enzymes (TIMPs) that leads to matrix destruction was postulated in the pathogenesis of osteoarthritis. Primary human articular chondrocytes from patients of both genders were cultured in alginate beads at 5% O(2) to which 10(-11)M-10(-5)M 17β-estradiol had been added and analyzed by means of immunohistochemistry, immunocytochemistry and real-time RT-PCR. Since articular chondrocytes in vivo are adapted to a low oxygen tension, culture was performed at 5% O(2). Immunohistochemical staining in articular cartilage tissue from patients and immunocytochemical staining in articular chondrocytes cultured in alginate beads was positive for type II collagen, estrogen receptor α, MMP-1, and -13. It was negative for type I collagen, MMP-3, TIMP-1 and -2. Using real-time RT-PCR, it was demonstrated that physiological and supraphysiological doses of 17β-estradiol suppress mRNA levels of MMP-3 and -13 significantly in articular chondrocytes of female patients. A significant suppressing effect was also seen in MMP-1 mRNA after a high dose of 10(-5)M 17β-estradiol. Furthermore, high doses of this hormone led to tendentially lower TIMP-1 levels whereas the TIMP-2 mRNA level was not influenced. In male patients, only incubations with high doses (10(-5)M) of 17β-estradiol were followed by a tendency to suppressed MMP-1 and TIMP-1 levels while TIMP-2 mRNA level was decreased significantly. There was no effect on MMP-13 expression of cells from male patients. Taken together, application of 17β-estradiol in physiological doses will improve the imbalance between the amounts of MMPs and TIMPs in articular chondrocytes from female patients. Downregulation of TIMP-2 by 17β-estradiol in male patients would not be articular cartilage protective.

Sex-specific lung remodeling and inflammation changes in experimental allergic asthma

There is evidence that sex and sex hormones influence the severity of asthma. Airway and lung parenchyma remodeling and the relationship of ultrastructural changes to airway responsiveness and inflammation in male, female, and oophorectomized mice (OVX) were analyzed in experimental chronic allergic asthma. Seventy-two BALB/c mice were randomly divided into three groups (n=24/each): male, female, and OVX mice, whose ovaries were removed 7 days before the start of sensitization. Each group was further randomized to be sensitized and challenged with ovalbumin (OVA) or saline. Twenty-four hours after the last challenge, collagen fiber content in airways and lung parenchyma, the volume proportion of smooth muscle-specific actin in alveolar ducts and terminal bronchiole, the amount of matrix metalloproteinase (MMP)-2 and MMP-9, and the number of eosinophils and interleukin (IL)-4, IL-5, and transforming growth factor (TGF)-β levels in bronchoalveolar lavage fluid were higher in female than male OVA mice. The response of OVX mice was similar to that of males, except that IL-5 remained higher. Nevertheless, after OVA provocation, airway responsiveness to methacholine was higher in males compared with females and OVX mice. In conclusion, sex influenced the remodeling process, but the mechanisms responsible for airway hyperresponsiveness seemed to differ from those related to remodeling.

The β1-Integrin–Dependent Function of RECK in Physiologic and Tumor Angiogenesis

Vascular endothelial cells produce considerable amounts of matrix metalloproteinases (MMP), including MMP-2, MMP-9, and membrane type 1 (MT1)-MMP. However, little is known about the regulatory mechanisms of these protease activities exhibited during vascular development. A glycosylphosphatidylinositol-anchored glycoprotein, reversion-inducing cysteine-rich protein with Kazal motifs (RECK), has been shown to attenuate MMP-2 maturation by directly interacting with MT1-MMP. Here, we show that an angiogenic factor angiopoietin-1 induces RECK expression in human umbilical vein endothelial cells (HUVEC), and RECK depletion in these cells results in defective vascular tube formation and cellular senescence. We further observed that RECK depletion downregulates beta1-integrin activation, which was associated with decreased autophosphorylation of focal adhesion kinase and increased expression of a cyclin-dependent kinase inhibitor p21(CIP1). In agreement, significant downregulation of beta1-integrin activity was observed in vascular endothelial cells in Reck-/- mouse embryos. In HUVECs, specific inhibition of MMP-2 significantly antagonized the effect of RECK depletion on beta1-integrin signaling, cell proliferation, and tube elongation. Furthermore, we observed that hypervascular tumor-derived cell lines can induce high RECK expression in convoluted vascular endothelial cells, and this in turn supports tumor growth. Targeting RECK specifically in tumor-associated vascular endothelial cells resulted in tumor regression. Therefore, we propose that RECK in tumor vascular endothelial cells can be an interesting target of cancer treatment via abortion of tumor angiogenesis.

Transcellular Invasion of MM1 Rat Ascites Hepatoma Cells Requires Matrix Metalloproteinases Derived from Host Mesothelium

Degradation of the extracellular matrix by proteases secreted by invading tumor cells is thought to be essential for metastasis. Using an in vitro transcellular migration assay model, we examined the requirement of matrix metalloproteinases (MMP) in the invasion of MM1 rat hepatoma cells through normal mesothelial cell monolayers. Here we show that MM1 cells transmigrate using MMPs not expressed in the tumor cells but secreted by host mesothelial cells. Additionally, the amount of MMP secreted by mesothelial cells was increased by co-culture with hepatoma cells. Our results point to the role of normal host cells in the tumor microenvironment as a source of invasion factors necessary for the metastatic process.

Airway remodeling and lack of bronchodilator response in steroid-resistant asthma

Steroid-resistant (SR) asthma is characterized by airway inflammation that fails to resolve despite treatment with corticosteroids, raising concerns that resistance to steroid therapy in asthma could lead to airway remodeling. We sought to determine whether SR asthma is accompanied by decreased airflow reversibility and could lead to airway remodeling. Spirometric results were evaluated for 40 asthmatic patients defined as having SR or steroid-sensitive (SS) asthma on the basis of a 1-week course of oral prednisone. Twenty-three asthmatic patients underwent bronchoscopy with collection of bronchoalveolar lavage (BAL) fluid to analyze markers of airway remodeling in BAL fluid and cells. Prednisone significantly improved FEV(1) percent predicted in SS asthma (62.0% +/- 10.9% [mean +/- SD] to 79.4% +/- 11.3%, P < .001) but not in SR asthma (66.9% +/- 10.0% to 65.9% +/- 12.1%). The bronchodilator response was significantly greater in the SS than in the SR group (Delta FEV(1) percent predicted, 33.5% +/- 22.5% vs 15.2% +/- 7.9%; P = .001), regardless of inhaled corticosteroid use. No difference in amounts of matrix metalloproteinase (MMP) 9, PMN elastase, or vascular endothelial growth factor was found in BAL fluid from both groups. Tissue inhibitor of metalloproteinases (TIMP) 1 levels were, however, significantly less in BAL fluid of patients with SR asthma compared with those in patients with SS asthma (921.9 +/- 313.4 vs 2267.0 +/- 456.8 pg/mL, P < .05), resulting in significantly higher MMP-9/TIMP-1 ratios in the BAL fluid of patients with SR asthma (0.24 +/- 0.04 vs 0.11 +/- 0.03, P < .01). Finally, dexamethasone treatment induced TIMP-1 mRNA in BAL fluid cells from patients with SS asthma (P < .01) but not in cells from patients with SR asthma. Bronchodilator reversibility is impaired in SR asthma and is associated with a shift in MMP-9/TIMP-1 ratio caused by inability of steroids to enhance TIMP-1 production, potentially promoting proteolytic activity in airways of patients with SR asthma and contributing to chronic airway remodeling. SR asthma might lead to irreversible airways disease.

Expression of MMP-13 in osteoblast cells and rat tibia after exposure to gamma rays or accelerated carbon ions

In past research, we found that carbon ion irradiation increased bone volume in rats, and a significant amount of cartilage remained inside the carbon ion-irradiated trabeculae. The amounts of matrix metalloproteinase 13 (MMP-13) mRNA in osteoblast-like MC3T3-E1 cells tended to decrease after carbon ion irradiation. The level of MMP-13 mRNA in non-irradiated cells was stable during the experimental period, but in gamma ray-irradiated cells it tended to increase. When localization of MMP-13 in locally irradiated experimental rats was investigated, it was found in the marginal trabeculae in both non-irradiated and gamma ray-irradiated animals. MMP-13 was detected in osteoid and neogenetic bone in the trabeculae surface. The trabeculae in carbon ion-irradiated bone remained cartilaginous. Carbon ion-irradiated rats exhibited weak expression of MMP-13 around the cartilage inside the trabeculae. We conclude that carbon ion irradiation reduced expression of MMP-13, thus suppressing both chondrocyte maturation and cartilage resorption. Increases in hyperplasia of the bone trabeculae and of bone volume were caused by ongoing bone addition and calcification in the absence of sufficient cartilage resorption.

Acidic environments may be able to help wounds to heal faster

It is assumed that all wounds go through a series of processes such as inflammation, proliferation and remodelling in what is known as the healing cascade. These phases normally result in a healed wound but sometimes one or more of these phases will become 'stuck' so the wound is unable to progress and so becomes chronic. In some cases this may be because of an uncontrolled amount of matrix metalloproteinases (MMPs) in the wound bed. MMPs are enzymes present in wound exudates. In the inflammatory and proliferation stages, they break down dead tissue making it easier for macrophages to do their work of removing dead cells. MMP activity is normally closely controlled by enzyme inhibitors but sometimes this control fails so that MMPs continue to break down new tissues. Laboratory experiments have suggested that this situation is more likely to occur where the wound has a high pH. Application of a weak acid reduces the activity of these out-of-control MMPs.

Thrombospondin 2 deficiency in pregnant mice results in premature softening of the uterine cervix.

The gradual disorganization of collagen fibers in the stromal connective tissue of the uterine cervix is characteristic of progressive cervical softening during pregnancy. A lack of thrombospondin (TSP) 2 has been shown to be associated with altered collagen fibril morphology of connective-tissue-rich organs such as skin and tendon. The goal of this study was to determine the role of TSP2 in cervical softening by studying a TSP2-null mouse line. Creep testing showed that, in the nonpregnant animal and on Day 10 of pregnancy, there was no difference between the cervical extensibility of the wild-type and the TSP2-deficient mice. However, by Day 14 of pregnancy, the TSP2-null mice showed 4.5-fold increase in cervical extensibility, and by Day 18, a 6.1-fold increase, when compared with wild-type mice. A further indicator of compromised cervical integrity was that, on Days 14 and 18 of pregnancy, the cervix of TSP2-null mice broke rapidly under standard loading conditions that did not break the cervix of wild-type mice. Western blotting showed that TSP2 was expressed in the cervix of mice on Days 14 and 18 of pregnancy but not on Day 10 or in the nonpregnant animal. As determined by immunohistochemistry, the amount of matrix metalloproteinase 2 (MMP2) in the cervix of TSP2-null mice increased 11-fold on Day 14 of pregnancy and 19-fold on Day 18. Thus, TSP2-null mice provide an animal model to assist in the understanding of the molecular basis of spontaneous, premature softening of the uterine cervix.

Effect of estrogen on the expression of matrix metalloproteinase (MMP)-1, MMP-3, and MMP-13 and tissue inhibitor of metalloproternase-1 in osteoarthritis chondrocytes.

The aim of this study was to evaluate the effect of estrogen on matrix metalloproteinase (MMP)-1, MMP-3, MMP-13, and tissue inhibitor of metalloproteinase (TIMP)-1 in osteoarthritic chondrocytes. Chondrocytes from the knee cartilage of 25 postmenopausal osteoarthritic (OA) patients were cultured under various conditions: 0 pg/mL, 50 pg/mL, 500 pg/mL, and 5,000 pg/mL of 17beta-estradiol, with or without 10-1,000 pg/mL of either interleukin (IL)-1beta or tumor necrosis factor alpha (TNFalpha). MMP-1, MMP-3, MMP-13, and TIMP-1 in the conditioned media were analyzed with immunoblot or enzyme-linked immunosorbent assay (ELISA). Type II collagenolytic activity was measured by fluorogenic type II collagenolytic activity assay. Real-time reverse transcriptase polymerase chain reaction (RT-PCR) using SYBR Green I dye was performed for the quantification of mRNA. Without cytokine stimulation, the secretion of MMP-1 was significantly reduced by 50 pg/mL of 17beta-estradiol (in immunoblot by a median of 12.3%, P=0.007; in ELISA by a median of 18.4%, P=0.001), and 500 pg/mL (in immunoblot by a median of 23.1%, P=0.001; in ELISA by a median of 21.0%, P=0.001). Additionally, under 10 pg/mL TNFalpha, 17beta-estradiol also significantly suppressed the secretion of MMP-1 (in immunoblot by a median of 39.0%, P=0.016; in ELISA by a median of 38.4%, P=0.041). Estrogen did not exert any significant effect on MMP-3, MMP-13, or TIMP-1 expression. With IL-1beta or TNFalpha above 10 pg/mL stimulation, 17beta-estradiol demonstrated no effect on MMP-1, MMP-3, MMP-13, or TIMP-1 secretion. Type II collagenolytic activity in the 50 pg/mL estradiol group decreased by 9.6% (-51.5-5.5%, P>0.05). 17beta-estradiol showed a tendency to decrease in MMP-1 mRNA. Estrogen may improve the imbalance between the amounts of MMPs and TIMP in chondrocytes, and these results suggest that hormone replacement therapy may provide some chondroprotective effect.

The expression of matrix metalloproteinases and inhibitors in acute rupture of the anterior cruciate ligament

Rapid degeneration of the anterior cruciate ligament (ACL) may occur after ligament rupture, making primary repair of the anterior cruciate ligament difficult. Nine completely ruptured anterior cruciate ligaments were collected by arthroscopic surgery performed within 6 months of injury. The authors studied the localization of matrix metalloproteinases and tissue inhibitors of matrix metalloproteinases in ruptured anterior cruciate ligament using immunohistochemistry, and measured messenger ribonucleic acid expression using the reverse transcriptase polymerase chain reaction. Cells in residual ligament tissue seemed to contain matrix metalloproteinases 1 and 3 and tissue inhibitors of matrix metalloproteinases 1 and 2. Promatrix metalloproteinase-9 positive cells were observed in the perivascular area. Promatrix metalloproteinase-2 positive cells frequently were seen between irregular collagen bundles in stumps of ruptured ligaments. Tissue inhibitors of matrix metalloproteinase-2 positive cells commonly were observed in ruptured ligaments. Matrix metalloproteinase-1 and matrix metalloproteinase-3 messenger ribonucleic acid were highly expressed compared with matrix metalloproteinase-2 messenger ribonucleic acid. Tissue inhibitor of matrix metalloproteinase-2 messenger ribonucleic acid was highly expressed compared with tissue inhibitor of matrix metalloproteinase 1. The authors could not identify whether these intrinsic reactions mediated by anterior cruciate ligament cells were the causes of rapid degradation or the results of the degradation process. Various amounts of matrix metalloproteinase and inhibitor production of intrinsic ligament cells were observed in the ruptured anterior cruciate ligament. The biological reaction reported in this study may suggest that pronounced metabolism is undertaken in ruptured ACL cells, and provide useful insight concerning the possiblitiy of achieving the primary repair of ruptured ACL.

Excitatory GABA function and its involvement in the NMDA-induced Ca 2+ mobilization in the suprachiasmatic nucleus

Glutamate can activate NMDA receptor (NMDAR) and subsequently induces excitotoxic neuron loss. However, roles of NMDARs in the blood–brain barrier (BBB) are little known. This study used a mouse cerebrovascular endothelial cell (MCEC) model to evaluate the effects of NMDAR activation on maintenance of the BBB and its possible mechanisms. Analysis of confocal microscopy revealed expressions of NMDAR subunits, GluN1 and GLUN2B, in MCECs. An immunoblot assay further showed the existence of GluN1 in plasma membranes of MCECs. In brain tissues, a confocal microscopic analysis demonstrated co-localization of GluN1 and factor VIII, a biomarker of MCECs. In addition, GluN1 mRNA was detected in MCECs and the brain. Functional assays showed that exposure of MCECs to NMDA increased calcium influx. Separately, NMDA suppressed transendothelial electrical resistance values, levels of occludin, and occludin tight junctions. As to the mechanism, NMDA stimulated sequential phosphorylations of extracellular signal-regulated kinase (ERK)1/2 and mitogen-activated ERK (MEK)1. Interestingly, amounts of matrix metalloproteinase (MMP)2 and MMP9 in MCECs were augmented by NMDA. The NMDA-induced alterations in ERK1/2 phosphorylation and occludin levels were reversed by pretreatment with PD98059, a MEK inhibitor, and MK-801, a NMDAR antagonist, respectively. Therefore, this study shows the functional presence of NMDARs in MCECs, and NMDAR activation can disrupt the MCEC-constructed tight junction barrier via activation of the MEK1/2-ERK1/2 signaling pathway and upregulation of MMP2/9 expressions.