The type I collagen induction of MT1-MMP-mediated MMP-2 activation is repressed by alphaVbeta3 integrin in human breast cancer cells.

The influence of alphaVbeta3 integrin on MT1-MMP functionality was studied in human breast cancer cells of differing beta3 integrin status. Overexpression of beta3 integrin caused increased cell surface expression of alphaV integrin and increased cellular adhesion to extracellular matrix (ECM) substrates in BT-549, MDA-MB-231 and MCF-7 cells. beta3 integrin expression also enhanced the migration of breast cancer cells on ECM substrates and enhanced collagen gel contraction. In vivo, alphaVbeta3 cooperated with MT1-MMP to increase the growth of MCF-7 cells after orthotopic inoculation in immunocompromised mice, but had no influence on in vitro proliferation. Despite these stimulatory effects, overexpression of beta3 integrin suppressed the type I collagen (Col I) induced MMP-2 activation in all breast cancer cell lines analyzed. This was also evident in extracts from the MCF-7 tumors in vivo, where MMP-2 activation was stimulated by MT1-MMP transfection, but attenuated with beta3 integrin expression. Although our studies confirm important biological effects of alphaVbeta3 integrin on enhancing cell adhesion and migration, ECM remodeling and tumor growth, beta3 integrin caused reduced MMP-2 activation in response to Col I in vitro, which appears to be physiologically relevant, as it was also seen in tumor xenografts in vivo. The reduction of MMP-2 activation (and thus MT1-MMP activity) by alphaVbeta3 in response to Col I may be important in scenarios where cells which are activated for matrix degradation need to preserve some pericellular collagen, perhaps as a substrate for cell adhesion and migration, thus maintaining a balanced level of proteolysis required for efficient tumor growth.     

Rapid expression and activation of MMP-2 and MMP-9 upon exposure of human breast cancer cells (MCF-7) to fibronectin in serum free medium

Interactions between tumour cells and the extracellular matrix (ECM) strongly influence tumour development, affecting cell survival, proliferation and migration. Many of these interactions are mediated through a family of cell surface receptors named integrins. Fibronectin and its integrin receptors play important roles in tumour development. The alpha5beta 1 integrin interacts with the central cell adhesive region of fibronectin and requires both the RGD and synergy sites for maximal binding. Matrix metalloproteinases (MMPs) are a family of zinc dependent endopeptidases. They are capable of digesting the different components of the ECM and basement membrane. The ECM gives structural support to cells and plays a central role in cell adhesion, differentiation, proliferation and migration. Binding of ECM to integrins modulates expression and activity of the different MMPs. Our experimental findings demonstrate that cultivation of human breast cancer cells, MCF-7, in serum free medium in the presence of fibronectin upregulates the activity of MMP-2 and MMP-9. Blocking of alpha5beta 1 integrin with anti-alpha5 monoclonal antibody inhibits the fibronectin-induced MMP activation response appreciably. This strongly indicates alpha5beta 1 mediated signalling events in activation of MMP-2 and MMP-9. Phosphorylation of FAK and PI-3 kinase and the nuclear translocation of ERK and NF-kappaB upon fibronectin binding demonstrate possible participation of the FAK/PI-3K/ERK signalling pathways in the regulation of MMP-2 activity.     

Membrane type-1 matrix metalloproteinase (MT1-MMP) processing of pro-alphav integrin regulates cross-talk between alphavbeta3 and alpha2beta1 integrins in breast carcinoma cells

We have recently demonstrated that in breast carcinoma MCF7 cells MT1-MMP processes the alphav, alpha3, and alpha5 integrin precursors generating the respective mature S-S-linked heavy and light alpha-chains. The precursor of alpha2 integrin subunit was found resistant to MT1-MMP proteolysis. The processing of the alphav subunit by MT1-MMP facilitated alphavbeta3-dependent adhesion, activation of FAK signaling pathway, and migration of MCF7 cells on vitronectin. To elucidate further the effects of MT1-MMP on cellular integrins, we examined the functional activity of alpha5beta1 and alpha2beta1 integrins in MCF7 cells expressing MT1-MMP. Either expression of MT1-MMP alone or its coexpression with alphavbeta3 failed to affect the functionality of alpha5beta1 integrin, and adhesion of cells to fibronectin. MT1-MMP, however, profoundly affected the cross-talk involving alphavbeta3 and alpha2beta1 integrins. In MT1-MMP-deficient cells, integrin alphavbeta3 suppressed the functional activity of the collagen-binding alpha2beta1 integrin receptor and diminished cell adhesion to type I collagen. Coexpression of MT1-MMP with integrin alphavbeta3 restored the functionality of alpha2beta1 integrin and, consequently, the ability of MCF7 cells to adhere efficiently to collagen. We conclude that the MT1-MMP-controlled cross-talk between alphavbeta3 and alpha2beta1 integrins supports binding of aggressive, MT1-MMP-, and alphavbeta3 integrin-expressing malignant cells on type I collagen, the most common substratum of the extracellular matrix.     

MT1-MMP initiates activation of pro-MMP-2 and integrin alphavbeta3 promotes maturation of MMP-2 in breast carcinoma cells

We evaluated cellular mechanisms involved in the activation pathway of matrix prometalloproteinase-2 (pro-MMP-2), an enzyme implicated in the malignant progression of many tumor types. Membrane type-1 matrix metalloproteinase (MT1-MMP) cleaves the N-terminal prodomain of pro-MMP-2 thus generating the activation intermediate that then matures into the fully active enzyme of MMP-2. Our results provide evidence on how a collaboration between MT1-MMP and integrin alphavbeta3 promotes more efficient activation and specific, transient docking of the activation intermediate and, further, the mature, active enzyme of MMP-2 at discrete regions of cells. We show that coexpression of MT1-MMP and integrin alphavbeta3 in MCF7 breast carcinoma cells specifically enhances in trans autocatalytic maturation of MMP-2. The association of MMP-2's C-terminal hemopexin-like domain with those molecules of integrin alphavbeta3 which are proximal to MT1-MMP facilitates MMP-2 maturation. Vitronectin, a specific ligand of integrin alphavbeta3, competitively blocked the integrin-dependent maturation of MMP-2. Immunofluorescence and immunoprecipitation studies supported clustering of MT1-MMP and integrin alphavbeta3 at discrete regions of the cell surface. Evidently, the identified mechanisms appear to be instrumental to clustering active MMP-2 directly at the invadopodia and invasive front of alphavbeta3-expressing cells or in their close vicinity, thereby accelerating tumor cell locomotion.     

Membrane-type 1 matrix metalloproteinase modulates focal adhesion stability and cell migration

Membrane-type 1 matrix metalloproteinase (MT1-MMP) plays an important role in extracellular matrix-induced cell migration and the activation of extracellular signal-regulated kinase (ERK). We showed here that transfection of the MT1-MMP gene into HeLa cells promoted fibronectin-induced cell migration, which was accompanied by fibronectin degradation and reduction of stable focal adhesions, which function as anchors for actin-stress fibers. MT1-MMP expression attenuated integrin clustering that was induced by adhesion of cells to fibronectin. The attenuation of integrin clustering was abrogated by MT1-MMP inhibition with a synthetic MMP inhibitor, BB94. When cultured on fibronectin, HT1080 cells, which endogenously express MT1-MMP, showed so-called motile morphology with well-organized focal adhesion formation, well-oriented actin-stress fiber formation, and the lysis of fibronectin through trails of cell migration. Inhibition of endogenous MT1-MMP by BB94 treatment or expression of the MT1-MMP carboxyl-terminal domain, which negatively regulates MT1-MMP activity, resulted in the suppression of fibronectin lysis and cell migration. BB94 treatment promoted stable focal adhesion formation concomitant with enhanced phosphorylation of tyrosine 397 of focal adhesion kinase (FAK) and reduced ERK activation. These results suggest that lysis of the extracellular matrix by MT1-MMP promotes focal adhesion turnover and subsequent ERK activation, which in turn stimulates cell migration.     

Osteopontin stimulates tumor growth and activation of promatrix metalloproteinase-2 through nuclear factor-kappa B-mediated induction of membrane type 1 matrix metalloproteinase in murine melanoma cells

Matrix metalloproteinases (MMPs) degrade the extracellular matrix (ECM) and play critical roles in tissue repair, tumor invasion, and metastasis. MMPs are regulated by different cytokines, ECM proteins, and other factors. However, the molecular mechanisms by which osteopontin (OPN), an ECM protein, regulates ECM invasion and tumor growth and modulates MMP activation in B16F10 cells are not well defined. We have purified OPN from human milk and shown that OPN induces pro-MMP-2 production and activation in these cells. Moreover, our data revealed that OPN-induced membrane type 1 (MT1) MMP expression correlates with translocation of p65 (nuclear factor-kappaB (NF-kappaB)) into the nucleus. However, when the super-repressor form of IkappaBalpha (inhibitor of NF-kappaB) was transfected into cells followed by treatment with OPN, no induction of MT1-MMP expression was observed, indicating that OPN activates pro-MMP-2 via an NF-kappaB-mediated pathway. OPN also enhanced cell migration and ECM invasion by interacting with alpha(v)beta(3) integrin, but these effects were reduced drastically when the MMP-2-specific antisense S-oligonucleotide was used to suppress MMP-2 expression. Interestingly, when the OPN-treated cells were injected into nude mice, the mice developed larger tumors, and the MMP-2 levels in the tumors were significantly higher than in controls. The proliferation data indicate that OPN increases the growth rate in these cells. Both tumor size and MMP-2 expression were reduced dramatically when anti-MMP-2 antibody or antisense S-oligonucleotide-transfected cells were injected into the nude mice. To our knowledge, this is the first report that MMP-2 plays a direct role in OPN-induced cell migration, invasion, and tumor growth and that demonstrates that OPN-stimulated MMP-2 activation occurs through NF-kappaB-mediated induction of MT1-MMP.     

MT1-MMP Initiates Activation of pro-MMP-2 and Integrin αvβ3 Promotes Maturation of MMP-2 in Breast Carcinoma Cells

We evaluated cellular mechanisms involved in the activation pathway of matrix prometalloproteinase-2 (pro-MMP-2), an enzyme implicated in the malignant progression of many tumor types. Membrane type-1 matrix metalloproteinase (MT1-MMP) cleaves the N-terminal prodomain of pro-MMP-2 thus generating the activation intermediate that then matures into the fully active enzyme of MMP-2. Our results provide evidence on how a collaboration between MT1-MMP and integrin αvβ3 promotes more efficient activation and specific, transient docking of the activation intermediate and, further, the mature, active enzyme of MMP-2 at discrete regions of cells. We show that coexpression of MT1-MMP and integrin αvβ3 in MCF7 breast carcinoma cells specifically enhances in trans autocatalytic maturation of MMP-2. The association of MMP-2′s C-terminal hemopexin-like domain with those molecules of integrin αvβ3 which are proximal to MT1-MMP facilitates MMP-2 maturation. Vitronectin, a specific ligand of integrin αvβ3, competitively blocked the integrin-dependent maturation of MMP-2. Immunofluorescence and immunoprecipitation studies supported clustering of MT1-MMP and integrin αvβ3 at discrete regions of the cell surface. Evidently, the identified mechanisms appear to be instrumental to clustering active MMP-2 directly at the invadopodia and invasive front of αvβ3-expressing cells or in their close vicinity, thereby accelerating tumor cell locomotion.     

FAK-mediated src phosphorylation of endophilin A2 inhibits endocytosis of MT1-MMP and promotes ECM degradation

Focal adhesion kinase (FAK) is an important mediator of integrin signaling in the regulation of cell proliferation, survival, migration, and invasion. To understand how FAK contributes to cell invasion, we explored the regulation of matrix metalloproteinases (MMPs) by FAK. We found that v-Src-transformed cells activate a FAK-dependent mechanism that attenuates endocytosis of MT1-MMP. This in turn increases cell-surface expression of MT1-MMP and cellular degradation of extracellular matrix. Further, we identified an interaction between FAK's second Pro-rich motif and endophilin A2's SH3 domain. This interaction served as an autophosphorylation-dependent scaffold to allow Src phosphorylation of endophilin A2 at Tyr315. Tyr315 phosphorylation inhibited endophilin/dynamin interactions, and blockade of Tyr315 phosphorylation promoted endocytosis of MT1-MMP. Together, these results suggest a regulatory mechanism of cell invasion whereby FAK promotes cell-surface presentation of MT1-MMP by inhibiting endophilin A2-dependent endocytosis.     

MT1-MMP promotes cell growth and ERK activation through c-Src and paxillin in three-dimensional collagen matrix

Membrane-type 1 matrix metalloproteinase (MT1-MMP) is essential for tumor invasion and growth. We show here that MT1-MMP induces extracellular signal-regulated kinase (ERK) activation in cancer cells cultured in collagen gel, which is indispensable for their proliferation. Inhibition of MT1-MMP by MMP inhibitor or small interfering RNA suppressed activation of focal adhesion kinase (FAK) and ERK in MT1-MMP-expressing cancer cells, which resulted in up-regulation of p21^WAF1 and suppression of cell growth in collagen gel. Cell proliferation was also abrogated by the inhibitor against ERK pathway without affecting FAK phosphorylation. MT1-MMP and integrin α_vβ₃ were shown to be involved in c-Src activation, which induced FAK and ERK activation in collagen gel. These MT1-MMP-mediated signal transductions were paxillin dependent, as knockdown of paxillin reduced cell growth and ERK activation, and co-expression of MT1-MMP with paxillin induced ERK activation. The results suggest that MT1-MMP contributes to proliferation of cancer cells in the extracellular matrix by activating ERK through c-Src and paxillin.     

β1 Integrin-Focal Adhesion Kinase (FAK) Signaling Modulates Retinal Ganglion Cell (RGC) Survival

Extracellular matrix (ECM) integrity in the central nervous system (CNS) is essential for neuronal homeostasis. Signals from the ECM are transmitted to neurons through integrins, a family of cell surface receptors that mediate cell attachment to ECM. We have previously established a causal link between the activation of the matrix metalloproteinase-9 (MMP-9), degradation of laminin in the ECM of retinal ganglion cells (RGCs), and RGC death in a mouse model of retinal ischemia-reperfusion injury (RIRI). Here we investigated the role of laminin-integrin signaling in RGC survival in vitro, and after ischemia in vivo. In purified primary rat RGCs, stimulation of the β1 integrin receptor with laminin, or agonist antibodies enhanced RGC survival in correlation with activation of β1 integrin’s major downstream regulator, focal adhesion kinase (FAK). Furthermore, β1 integrin binding and FAK activation were required for RGCs’ survival response to laminin. Finally, in vivo after RIRI, we observed an up-regulation of MMP-9, proteolytic degradation of laminin, decreased RGC expression of β1 integrin, FAK and Akt dephosphorylation, and reduced expression of the pro-survival molecule bcl-xL in the period preceding RGC apoptosis. RGC death was prevented, in the context of laminin degradation, by maintaining β1 integrin activation with agonist antibodies. Thus, disruption of homeostatic RGC-laminin interaction and signaling leads to cell death after retinal ischemia, and maintaining integrin activation may be a therapeutic approach to neuroprotection.     

Stimulation of pro-MMP-2 production and activation by native form of extracellular type I collagen in cultured hepatic stellate cells

Cultured hepatic stellate cells (HSCs) are known to change their morphology and function with respect to the production of extracellular matrices (ECMs) and matrix metalloproteinases (MMPs) in response to ECM components. We examined the regulatory role of the native form of type I collagen fibrils in pro-MMP-2 production and activation in cultured HSCs. Gelatin zymography of the conditioned media revealed that pro- and active form of MMP-2 was increased in the HSCs cultured on type I collagen gel but not on type I collagen-coated surface, gelatin-coated surface, type IV collagen-coated surface, or Matrigel, suggesting the importance of the native form of type I collagen fibrils in pro-MMP-2 production and activation. The induction of active MMP-2 by extracellular type I collagen was suppressed by the blocking antibody against integrin beta1 subunits, indicating the involvement of integrin signaling in pro-MMP-2 activation. RT-PCR analysis indicated that MMP-2, membrane type-1 MMP (MT1-MMP) and tissue inhibitor of metalloproteinase-2 (TIMP-2) mRNA levels were elevated in HSCs cultured on type I collagen gel. The increased MT1-MMP proteins were localized on the cell surface of HSCs cultured on type I collagen gel. In contrast to the expression of MMP-2, HSCs showed a great decline in MMP-13 expression in HSCs cultured on type I collagen gel. These results indicate that the native fibrillar (polymerized) but not monomeric form of type I collagen induced pro-MMP-2 production and activation through MT1-MMP and TIMP-2 in cultured HSCs, suggesting an important role of HSCs in ECM remodeling in the hepatic perisinusoidal spaces.     

Inhibition of β1 integrin induces its association with MT1-MMP and decreases MT1-MMP internalization and cellular invasiveness

Integrin signaling plays a fundamental role in the establishment of focal adhesions and the subsequent formation of invadopodia in malignant cancer cells. Invadopodia facilitate localized adhesion and degradation of the extracellular matrix (ECM), which promote tumour cell invasion and metastasis. Degradation of ECM components is often driven by membrane type-1 matrix metalloproteinase (MT1-MMP), and we have recently shown that regulation of enzyme internalization is dependent on signaling downstream of β1 integrin. Phosphorylation of the cytoplasmic tail of MT1-MMP is required for its internalization and delivery to Rab5-marked early endosomes, where it is then able to be recycled to new sites of invadopodia formation and promote invasion. Here we found that inhibition of β1 integrin, using the antibody AIIB2, inhibited the internalization and recycling of MT1-MMP that is necessary to support long-term cellular invasion. MT1-MMP and β1 integrin were sequestered at the cell surface when β1-integrin was inhibited, and their association under these conditions was detected using immunoprecipitation and mass spectrometry analyses. Sequestration of β1 integrin and MT1-MMP at the cell surface resulted in the formation of large invadopodia and local ECM degradation; however, the impaired internalization and recycling of MT1-MMP and β1 integrin ultimately led to a loss of invasive behaviour.     

Curcumin, demethoxycurcumin and bisdemethoxycurcumin differentially inhibit cancer cell invasion through the down-regulation of MMPs and uPA

Curcumin (Cur), a component of turmeric (Curcuma longa), has been reported to exhibit antimetastatic activities, but the mechanisms remain unclear. Other curcuminoids present in turmeric, demethoxycurcumin (DMC) and bisdemethoxycurcumin (BDMC) have not been investigated whether they exhibit antimetastatic activity to the same extent as curcumin. The regulation of matrix metalloproteinases (MMPs) and urokinase plasminogen activator (uPA) play important role in cancer cell invasion by cleavage of extracellular matrix (ECM). In this line, we comparatively examined the influence of Cur, DMC and BDMC on the expressions of uPA, MMP-2, MMP-9, membrane Type 1 MMP (MT1-MMP), tissue inhibitor of metalloproteinases (TIMP-2), and in vitro invasiveness of human fibrosarcoma cells. The results indicate that the differential potency for inhibition of cancer cell invasion was BDMC> or =DMC>Cur, whereas the cell migration was not affected. Zymography analysis exhibited that curcumin, DMC and BDMC significantly decreased uPA, active-MMP-2 and MMP-9 but not pro-MMP-2 secretion from the cells in a dose-dependent manner, in which BDMC and DMC show higher potency than curcumin. The suppression of active MMP-2 level correlated with inhibition of MT1-MMP and TIMP-2 protein levels involved in pro-MMP-2 activation. Importantly, BDMC and DMC at 10 microM reduced MT1-MMP and TIMP-2 protein expression, but curcumin slightly reduced only MT1-MMP but not TIMP-2. In addition, three forms of curcuminoids significantly inhibited collagenase, MMP-2, and MMP-9 but not uPA activity. In summary, these data demonstrated that DMC and BDMC show higher antimetastasis potency than curcumin by the differentially down-regulation of ECM degradation enzymes.     

3-D collagen-dependent cell surface expression of MT1-MMP and MMP-2 activation regardless of integrin β1 function and matrix stiffness

Matrix metalloproteinases (MMPs) play roles in spatially dynamic processes, including morphogenesis, wound healing, and tumor invasion. Three-dimensional (3-D) type I collagen stimulates cellular activation of MMP-2, however, the mechanisms underlying this are controversial. The present study investigated mechanisms for 3-D collagen-induced MMP-2 activation in highly invasive human malignant mesothelioma cells. MMP-2 was effectively activated by cells cultured in 3-D collagen but not in 2-D collagen, whereas MMP-2 activation was not regulated by the flexibility of collagen. The 3-D collagen did not largely increase the gene expression of MMP-2 and MT1-MMP. However, MT1-MMP exposed to the cell surface was much increased by 3-D collagen, and loss of MT1-MMP abolished MMP-2 activation in response to 3-D collagen. MT1-MMP and integrin β1 translocated to pericellular regions interacting with collagen-coated microbeads, however their localization was different. Importantly, inhibition of integrin β1 function and expression did not affect 3-D collagen-induced cell surface localization of MT1-MMP and MMP-2 activation. Our results strongly suggest that 3-D collagen scaffolding may provide opportunity for direct and multivalent interaction with MT1-MMP, by which MMP-2 activation occur in abundant cell surface MT1-MMP-dependent manner, rather than a manner regulated by matrix stiffness and integrin β1 function.     

Tissue inhibitor of metalloproteinase-2 (TIMP-2) regulates myogenesis and beta1 integrin expression in vitro

Myogenesis in vitro involves myoblast cell cycle arrest, migration, and fusion to form multinucleated myotubes. Extracellular matrix (ECM) integrity during these processes is maintained by the opposing actions of matrix metalloproteinase (MMP) proteases and their inhibitors, the tissue inhibitor of metalloproteinases (TIMPs). Here, we report that TIMP-2, MMP-2, and MT1-MMP are differentially expressed during mouse myoblast differentiation in vitro. A specific role for TIMP-2 in myogenesis is demonstrated by altered TIMP-2(-/-) myotube formation. When differentiated in horse serum-containing medium, TIMP-2(-/-) myotubes are larger than wild-type myotubes. In contrast, when serum-free medium is used, TIMP-2(-/-) myotubes are smaller than wild-type myotubes. Regardless of culture condition, myotube size is directly correlated with MMP activity and inversely correlated with beta1 integrin expression. Treatment with recombinant TIMP-2 rescues reduced TIMP-2(-/-) myotube size and induces increased MMP-9 activation and decreased beta1 integrin expression. Treatment with either MMP-2 or MMP-9 similarly rescues reduced myotube size, but has no effect on beta1 integrin expression. These data suggest a specific regulatory relationship between TIMP-2 and beta1 integrin during myogenesis. Elucidating the role of TIMP-2 in myogenesis in vitro may lead to new therapeutic options for the use of TIMP-2 in myopathies and muscular dystrophies in vivo.     

An alternative processing of integrin alpha(v) subunit in tumor cells by membrane type-1 matrix metalloproteinase.

Membrane type-1 matrix metalloproteinase (MT1-MMP) and alpha(v)beta(3) integrin are both essential to cell invasion. Maturation of integrin pro-alpha(v)chain (pro-alpha(v)) involves its cleavage by proprotein convertases (PC) to form the disulfide-bonded 125-kDa heavy and 25-kDa light alpha chains. Our report presents evidence of an alternative pathway of pro-alpha(v) processing involving MT1-MMP. In breast carcinoma MCF7 cells deficient in MT1-MMP, pro-alpha(v) is processed by a conventional furin-like PC, and the mature alpha(v) integrin subunit is represented by the 125-kDa heavy chain and the 25-kDa light chain commencing from the N-terminal Asp(891). In contrast, in cells co-expressing alpha(v)beta(3) and MT1-MMP, MT1-MMP functions as an integrin convertase. MT1-MMP specifically cleaves pro-alpha(v), generating a 115-kDa heavy chain with the truncated C terminus and a 25-kDa light chain commencing from the N-terminal Leu(892). PC-cleavable alpha(3) and alpha(5) but not the PC-resistant alpha(2) integrin subunit are also susceptible to MT1-MMP cleavage. These novel mechanisms involved in the processing of integrin alpha subunits underscore the significance and complexity of interactions between MT1-MMP and adhesion receptors and suggest that regulation of integrin functionality may be an important role of MT1-MMP in migrating tumor cells.     

Modulation of membrane type 1 matrix metalloproteinase by LPS and gamma interferon bound to extracellular matrix in intestinal crypt cells

Membrane type 1 matrix metalloproteinase (MT1-MMP) is an integral membrane protein that participates in the processing and degradation of cell surface proteins and the extracellular matrix (ECM). This enzyme regulates ECM turnover in wound repair, promotes cell migration and activates other MMPs, such as MMP-2, which is involved in angiogenesis, cell migration and tumoral metastasis. An increase in pro-inflammatory cytokine expression, such as gamma interferon (IFN-gamma), has been associated with chronic wounds in inflammatory bowel diseases. However, the extent to which cytokines modulate MT1-MMP has not been totally defined. In this report, the effects of the bacterial lipopolysaccharide (LPS) and ECM-bound IFN-gamma on MT1-MMP expression and MMP-2 activity were evaluated by Western blot, RT-PCR and zymography in isolated intestinal epithelial and cultured HT-29 cells. In the presence of LPS, ECM-bound IFN-gamma, but not soluble IFN-gamma, reduced the enterocyte MT1-MMP protein expression. In addition, the active form of MMP-2 was also decreased in the presence of both LPS and IFN-gamma, indicating that lower MMP-2 activity accompanied the decrease in MT1-MMP expression. These results suggest the possibility that endotoxin and ECM-bound IFN-gamma may affect matrix remodeling by modulating matrix metalloproteinase in enterocytes during wound healing.