Oxygen-mediated regulation of gelatinase and tissue inhibitor of metalloproteinases-1 expression by invasive cells.

The relative expression of matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) is an important determinant in trophoblast invasion of the uterus and tumor invasion and metastasis. Our previous studies have shown that low oxygen levels increase the in vitro invasiveness of trophoblast and tumor cells. The present study examined whether changes in oxygen levels affect TIMP and MMP expression by cultured trophoblast and breast cancer cells. Reverse zymographic analysis demonstrated reduced TIMP-1 protein secretion by HTR-8/SVneo trophoblast cells as well as MDA-MB-231 and MCF-7 breast carcinoma cells cultured in 1% vs 20% oxygen for 24 h. While gelatin zymography revealed no changes in the levels of MMP-9 secreted by HTR-8/SVneo trophoblasts cultured under various oxygen concentrations for 24 h, human MDA-MB-231 breast carcinoma cells displayed increased MMP-9 secretion and human MCF-7 breast cancer cells exhibited reduced secretion of this enzyme when cultured under similar conditions. In contrast, MMP-2 levels remained unchanged in all cultures incubated under similar conditions. Western blot analysis of MMP-9 protein in cell extracts confirmed the results of zymography. To assess the contribution of enhanced MMP activity to hypoxia-induced invasion, the effect of an MMP inhibitor (llomastat) on the ability of MDA-MB-231 cells to penetrate reconstituted extracellular matrix (Matrigel) was examined. Results showed that MMP inhibition significantly decreased the hypoxic upregulation of invasion by these cells. These findings indicate that the increased cellular invasiveness observed under reduced oxygen conditions may be due in part to a shift in the balance between MMPs and their inhibitors favoring increased MMP activity.     

Matrix metalloproteinases: multifunctional contributors to tumor progression

Matrix metalloproteinases (MMPs) are a family of extracellular matrix degrading proteinases. Owing to their matrix-degrading abilities and high expression in advanced tumors, MMPs were originally implicated in invasion and metastasis during cancer progression. However, recent work extends a role for MMPs during multiple stages of tumor progression to include other functions such as growth, angiogenesis and migration. Based on studies in animal models implicating MMP activity in cancer, synthetic MMP inhibitors are currently being tested in a clinical setting.     

Effect of glycosaminoglycans on matrix metalloproteinases in type II collagen-induced experimental arthritis

Matrix metalloproteinases (MMPs) are a family of neutral proteinases that are involved in tissue remodeling by mediating degradation of extracellular matrix components in both physiology and pathology. As MMPs appear to play a key role in the degradation of cartilage matrix in the progression of arthritic disease, MMPs are considered as potential therapeutic targets. The effect of chondroitin sulfate A (CSA) on MMPs in type II collagen-induced experimental arthritis was studied. The anti-arthritic effect of CSA was evidenced by a decrease in marker activities like lysosomal beta-hexosaminidase and beta-glucuronidase. Arthritic animals showed significantly higher activity of MMP2 and MMP9 and increased levels of other MMPs, including MMP3 and MT-1 MMP in cartilage and serum. Treatment with CSA significantly decreased the activity of MMPs, particularly MMP9 in serum and synovial effusate and cartilage. The effect of CSA was further studied by fragmenting CSA into low-molecular-weight oligosaccharides. The oligosaccharide-treated animals showed considerably lower MMP activity (particularly MMP9) compared with arthritic controls. The CSA (and the oligosaccharides derived from it) not only reduced the activity of MMPs but also decreased the protein level expression of MMPs, indicating that the production of MMPs is affected. These results indicate that the antiarthritic effect of CSA involves down-regulation of MMPs, which are critically involved in the progression of arthritic disease.     

MMP inhibitors: experimental and clinical studies

Matrix metalloproteases (MMPs) are a family of structurally related enzymes that are capable of degrading proteins of the extracellular matrix. These enzymes play a role in tissue remodelling associated with both physiological and pathogenic processes. A high expression of MMPs is associated with cancer malignancy: it is related to the tumor's ability to metastasize and to the process of angiogenesis. Treatment with MMP inhibitors alone or in combination with cytotoxic therapy is an interesting novel approach to control tumor progression. The expected mechanism of action of these compounds and the difference in side effects compared to cytotoxic drugs make the definition of endpoints and the assessment of response difficult. Furthermore, it is not yet clear whether tumor vascularization or, more specifically, MMP expression/activation should be a criterion of eligibility for this kind of treatment. This review provides an overview of the characteristics of MMPs and their role in tumor progression, metastasis and angiogenesis. Preclinical and clinical studies with synthetic MMP inhibitors are described. The presence of MMPs in biological fluids of patients and their use in prognostic evaluation and in determining the efficacy of treatment with MMP inhibitors is discussed.     

New signaling pathways from cancer progression modulators to mRNA expression of matrix metalloproteinases in breast cancer cells

We observed previously that each of seven cancer progression inhibitors suppresses the mRNA expression of some matrix metalloproteinases (MMPs), but stimulates that of others, in breast cancer cells. In the present study we tested the effect of overexpressing other cancer modulators on MMP expression. The MMPs tested are MMP1, MMP2, MMP7, MMP13, MMP14, MMP16, MMP19, and MMP25. The proteins that were overexpressed are cancer inhibitors (NME, DRG1, IL10), enhancers (SOD2, FAK, IL17, and CREB), and proteins that suppress cancer progression in cells of some cancers and promote it in others (FUT1, integrin beta3, serpin E1, TIAM1, and claudin 4). Unexpectedly, all of them only lowered MMP mRNA expression, mainly of MMP16, MMP2, and MMP13, in breast cancer cells. Signaling from SOD2 uncoupled the accumulation of two MMP16 mRNA splice variants, suggesting signaling to a late step in MMP16 mRNA accumulation, such as MMP16 mRNA stabilization or late mRNA processing. Signaling that modulates MMP expression differed widely among the total population of MDA-MB-231 cells and single-cell progenies cloned from that population. It also differed substantially between cells of two metastatic breast basal adenocarcinomas, MDA-MB-231 and MDA-MB-468. The present study detected 37 new signaling pathways from cancer progression modulators located upstream of MMP mRNA expression in human breast cancer cells. Our siRNA-induced MMP knockdown data support the interpretation that signaling from MMP19, MMP1, MMP7, MMP12, MMP14, and MMP11 each stimulates the mRNA expression of other MMPs in breast cancer cells.     

Tumor-stroma interaction: positive feedback regulation of extracellular matrix metalloproteinase inducer (EMMPRIN) expression and matrix metalloproteinase-dependent generation of soluble EMMPRIN

Matrix metalloproteinases (MMPs) are metal-dependent endopeptidases that play pivotal roles in tumor disease progression. In many solid tumors, MMPs are indeed produced by tumor stromal cells, rather than by tumor cells. This expression pattern is, at least in part, regulated by tumor-stroma interaction via tumor cell-associated extracellular matrix metalloproteinase inducer (EMMPRIN). In vitro, recombinant EMMPRIN dose-dependently stimulated MMP-1 production by primary human fibroblast cells. Interestingly, in addition to stimulating MMP expression, EMMPRIN also induced its own gene expression. To further explore this potential positive feedback regulatory mechanism, we generated human breast cancer cells expressing different levels of EMMPRIN. Coculture of EMMPRIN-positive tumor cells with fibroblast cells resulted in a concomitant stimulation of MMP-2, MMP-9, and EMMPRIN production. This induction was EMMPRIN dependent, was further enhanced by overexpression, and was reduced by antisense suppression of EMMPRIN expression in tumor cells. Increased expression of membrane-associated EMMPRIN was accompanied by an MMP-dependent generation of a soluble form of EMMPRIN representing a proteolytic cleavage product lacking the carboxyl terminus. On the basis of these findings, we propose a model in which tumor cell-associated EMMPRIN stimulates MMPs, as well as EMMPRIN expression in tumor stroma. Increased MMP activity in tumor local environment results in proteolytic cleavage of membrane-associated EMMPRIN, releasing soluble EMMPRIN. Soluble EMMPRIN in turn acts in a paracrine fashion on stroma cells that are both adjacent and distant to tumor sites to further stimulate the production of MMPs and additional EMMPRIN, which consequently contributes to tumor angiogenesis, tumor growth, and metastasis.     

基质金属蛋白酶MMP-26能有效促进人绒毛膜上皮癌细胞JEG-3浸润能力

胚胎植入过程中,滋养层细胞浸润与肿瘤的迁移过程非常相似,但显著的区别在于前者是受严格调控的有节制的浸润,基质金属蛋白酶(MMPs)的许多成员在其中起重要的作用.MMP-26是近年来发现的MMPs家族的新成员,它在滋养层细胞中的作用所知甚少.利用国际常用的人滋养层细胞模型——人绒毛膜上皮癌细胞系(JEG-3)作为体外实验模型,探讨MMP-26在人滋养层细胞浸润调节中的作用.将含有MMP-26全长cDNA的pCR3.1质粒转染到JEG-3细胞中,获得过量表达MMP-26基因的稳定细胞系JEG-3/MMP-26;细胞浸润分析表明JEG/MMP-26细胞的浸润能力较母本细胞明显增强;RT-PCR和明胶酶谱分析显示JEG-3/MMP-26细胞中MMP-9的表达和分泌水平提高;双荧光免疫细胞化学进一步显示MMP-26和MMP-9蛋白在细胞中有共定位现象.上述结果表明MMP-26能有效促进人滋养层细胞浸润,其作用可能是通过与其他MMP分子(如MMP-9)的协调来实现的.     

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.     

Matrix metalloproteinase 1: role in sarcoma biology

In carcinomas stromal cells participate in cancer progression by producing proteases such as MMPs. The expression MMP1 is a prognostic factor in human chondrosarcoma, however the role in tumor progression is unknown. Laser capture microdissection and In Situ hybridization were used to determine cellular origin of MMP1 in human sarcomas. A xenogenic model of tumor progression was then used and mice were divided in two groups: each harboring either the control or a stably MMP1 silenced cell line. Animals were sacrificed; the neovascularization, primary tumor volumes, and metastatic burden were assessed. LCM and RNA-ISH analysis revealed MMP1 expression was predominantly localized to the tumor cells in all samples of sarcoma (p = 0.05). The percentage lung metastatic volume at 5 weeks (p = 0.08) and number of spontaneous deaths secondary to systemic tumor burden were lower in MMP1 silenced cell bearing mice. Interestingly, this group also demonstrated a larger primary tumor size (p<0.04) and increased angiogenesis (p<0.01). These findings were found to be consistent when experiment was repeated using a second independent MMP1 silencing sequence. Prior clinical trials employing MMP1 inhibitors failed because of a poor understanding of the role of MMPs in tumor progression. The current findings indicating tumor cell production of MMP1 by sarcoma cells is novel and highlights the fundamental differences in MMP biology between carcinomas and sarcomas. The results also emphasize the complex roles of MMP in tumor progression of sarcomas. Not only does metastasis seem to be affected by MMP1 silencing, but also local tumor growth and angiogenesis are affected inversely.     

Role of matrix metalloproteinases in melanoma cell invasion

Cutaneous melanomas are notorious for their tendency to metastasize. Essential steps in this process are the degradation of basement membranes and remodeling of the extracellular matrix (ECM) by proteolytic enzymes such as matrix metalloproteinases (MMPs), which are regulated by their tissue inhibitors (TIMPs). An MMP expression is not restricted to tumor cells but is also found in stromal cells, indicating that stroma-derived proteases may contribute to melanoma progression. The MMPs have been shown to interact with a broad range of non-matrix proteins including adhesion molecules, growth factors and mediators of angiogenesis and apoptosis. In this review, we evaluate new insights into the interplay of MMPs and their molecular partners in melanoma progression.     

Tetraspanin CD9 Promotes the Invasive Phenotype of Human Fibrosarcoma Cells via Upregulation of Matrix Metalloproteinase-9

Tumor cell metastasis, a process which increases the morbidity and mortality of cancer patients, is highly dependent upon matrix metalloproteinase (MMP) production. Small molecule inhibitors of MMPs have proven unsuccessful at reducing tumor cell invasion in vivo. Therefore, finding an alternative approach to regulate MMP is an important endeavor. Tetraspanins, a family of cell surface organizers, play a major role in cell signaling events and have been implicated in regulating metastasis in numerous cancer cell lines. We stably expressed tetraspanin CD9 in an invasive and metastatic human fibrosarcoma cell line (CD9-HT1080) to investigate its role in regulating tumor cell invasiveness. CD9-HT1080 cells displayed a highly invasive phenotype as demonstrated by matrigel invasion assays. Statistically significant increases in MMP-9 production and activity were attributed to CD9 expression and were not due to any changes in other key tetraspanin complex members or MMP regulators. Increased invasion of CD9-HT1080 cells was reversed upon silencing of MMP-9 using a MMP-9 specific siRNA. Furthermore, we determined that the second extracellular loop of CD9 was responsible for the upregulation of MMP-9 production and subsequent cell invasion. We demonstrated for the first time that tetraspanin CD9 controls HT1080 cell invasion via upregulation of an integral member of the MMP family, MMP-9. Collectively, our studies provide mounting evidence that altered expression of CD9 may be a novel approach to regulate tumor cell progression.     

Remodeling the model organism: matrix metalloproteinase functions in invertebrates

The matrix metalloproteinase (MMP) family of extracellular proteases is conserved throughout the animal kingdom. Studies of invertebrate MMPs have demonstrated they are involved in tissue remodeling. In Drosophila, MMPs are required for histolysis, tracheal growth, tissue invasion, axon guidance, and dendritic remodeling. Recent work demonstrates that MMPs also participate in Drosophila tumor invasion. In Caenorhabditis elegans an MMP is involved in anchor cell invasion; a Hydra MMP is important for regeneration and maintaining cell identity; and a sea urchin MMP degrades matrix to allow hatching. In worms and in flies, MMPs are regulated by the JNK pathway.     

MMP-11 promotes papillary thyroid cell proliferation and invasion via the NF-κB pathway

Papillary thyroid carcinoma (PTC) is the most common form of thyroid cancer, and its incidence is on the rise. It has been reported that some matrix metalloproteinases (MMPs) are abnormally expressed in PTC and can be used as diagnostic markers. However, few studies have explored the underlying mechanisms by which MMPs promote tumor progression. In this study, we used microarray analysis to compare the variations of gene expression within the PTC cell populations and their adjacent normal tissues and found that MMP-11 was the most differentially expressed MMP. To investigate the role of MMP-11 in the mediation of thyroid cancer cell development, pEnter-MMP-11 plasmid, and MMP-11 small interfering RNA were applied to up- and downregulate MMP-11 expression of in cultured PTC cell lines K1 and BCPAP. The results suggested that the levels of proliferation and migration of cells transfected with MMP-11 siRNA were significantly reduced, while the levels in MMP-11-plasmid-transfected cells were increased. In terms of the mechanism, experimental data showed that the change in cyclin D1 is consistent with MMP-11 expression, which may explain the changes in proliferation. In addition, Western blot assay was conducted to analyze the p65 and activated (phospho-) p65 protein levels concomitant with MMP-11 adjustments. Variations in intracellular MMP-11 significantly altered the amount of phospho-p65 in thyroid cells, while p65 knockdown did not affect MMP-11 expression. These results suggest that MMP-11 is located upstream of p65 and regulates its activity. Interestingly, the data for the Transwell assay suggested that MMP-11 regulatory migration is also associated with the NF-κB p65 signaling pathway. In conclusion, this report describes the important role of MMP-11 in the regulation of thyroid cell proliferation and migration. Mechanistic studies have shown that cyclin D1 and p65 are important mediators in the processes, which provides a new way to study the mechanism of MMPs promoting the progression of thyroid cancer.     

Beneficial and detrimental influences of tissue inhibitor of metalloproteinase-1 (TIMP-1) in tumor progression

Tissue inhibitor of metalloproteinase-1 (TIMP-1) is one representative of the natural matrix metalloproteinase (MMP) inhibitor family, encompassing four members. It inhibits all MMPs, except several MT-MMPs, and a disintegrin with a metalloproteinase domain (ADAM)-10 with Kis < nM. Unexpectedly, its upregulation was associated to poor clinical outcome for several cancer varieties. Such finding might be related to the growth-promoting and survival activities of TIMP-1 for normal and cancer cells. In most cases, such properties are MMP-independent and binding of TIMP-1 to an unknown receptor system can trigger JAK (or FAK)/PI3 kinase/Akt/bad-bclX2 (erythroid, myeloid, epithelial cell lines) or Ras/Raf1/FAK (osteosarcoma cell line) signaling pathways. The relationship between viral infection and TIMP-1 expression is here underlined. Thus, TIMP-1 might display a dual influence on tumor progression; either beneficial by inhibiting MMPs as MMP-9 and by impairing angiogenesis or detrimental by favoring cancer cells growth or survival. We consider that the proMMP-9/TIMP-1 balance is of critical importance in early events of tumor progression, and might show promise as diagnostic and prognostic marker of malignancy.