Prognostic significance of matrix metalloproteinase 2 and tissue inhibitor of metalloproteinase 2 expression in prostate cancer.

Matrix metalloproteinases (MMPs) are proteolytic enzymes capable of degrading the structural support network for normal and malignant cells, promoting neoplastic cell invasion and metastasis. Tissue inhibitors of metalloproteinases (TIMPs) maintain connective tissue integrity by modulating MMP activity. Formalin-fixed paraffin-embedded tissue sections from 138 prostatic adenocarcinomas (PACs) were immunostained by a combined automated/manual method using monoclonal antibodies against MMP2 and TIMP2. Immunoreactivity was semiquantitatively scored based on stain intensity and distribution, and results were correlated with Gleason grade, pathologic stage, ploidy status, and disease recurrence. One hundred five of 138 (76%) and 113/138 (82%) PACs expressed MMP2 and TIMP2, respectively. Co-expression was observed in 94/138 (68%) of PACs (P =.01), correlated with advanced tumor stage (P =.05), and tended to be associated with disease recurrent cases (P =.07). TIMP2 expression individually correlated with advanced tumor stage (P =.04) and reached near significance with disease recurrence (P =.06). MMP2 expression was also more frequent in recurrent PACs, although this value did not reach statistical significance (P =.07). However, on multivariate analysis, only pathologic stage (P =.009) and ploidy status (P =.03) independently predicted disease recurrence. In conclusion, MMP2 and TIMP2 are co-expressed in a majority of PACs and correlate with prognostic variables. Interestingly, contrary to the previously documented anti-tumor effects of TIMPs, TIMP2 expression appears to have a tumor-promoting role in PACs and warrants further investigation.     

Tissue inhibitor of metalloproteinase 1 regulates matrix metalloproteinase activity during newt limb regeneration.

Matrix metalloproteinase (MMP) activity is important for newt limb regeneration. In most biological processes that require MMP function, MMP activity is tightly controlled by a variety of mechanisms, including the coexpression of natural inhibitors. Here, we show that gene expression of one such inhibitor, tissue inhibitor of metalloproteinase 1 (NvTIMP1), is upregulated during the wound healing and dedifferentiation stages of regeneration when several MMPs are at their maximal expression levels. Newt MMPs and NvTIMP1 also exhibit similar spatial expression patterns during the early stages of limb regeneration. NvTIMP1 inhibits the proteolytic activity of regeneration-related newt MMPs and, like human TIMP1, can induce a weak mitogenic response in certain cell types. These results suggest that NvTIMP1 may be functioning primarily to maintain optimal levels of MMP activity during the early stages of limb regeneration, while possibly serving a secondary role as a mitogen.     

Study of matrix metalloproteinases and their tissue inhibitors in ductal in situ carcinomas of the breast

Aims: To analyse the expression of metalloproteinases (MMPs) and their inhibitors (TIMPs) in ductal carcinoma in situ of the breast (DCIS). Methods and results: An immunohistochemical study was performed in 56 patients with pure DCIS, in 39 with DCIS adjacent to invasive carcinoma (IDC) and 63 patients with T1 IDC, using tissue microarrays and specific antibodies against MMPs and TIMPs. Immunohistochemical results were categorized using a specific software program. The data were analysed by unsupervised hierarchical cluster analysis by each cellular type. IDC showed a higher expression rate of MMP‐7 and TIMP‐1 than pure DCIS, as well as a higher expression rate of MMP‐9 and TIMP‐3 than the DCIS component of mixed cases, whereas pure DCIS showed a higher rate of expression of MMP‐9 and ‐11 and TIMP‐3 than in the DCIS component of mixed cases. Pure DCIS with a periductal inflammatory infiltrate showed significantly higher MMP‐2, ‐14 and TIMP‐1. Dendograms identified two cluster groups with distinct MMP/TIMP expression profiles in neoplastic cells and fibroblastic or mononuclear inflammatory cells surrounding the neoplastic ducts of pure DCIS. Conclusions: The results indicate the distinct variability in MMP/TIMP expression by DCIS, which may be of potential biological and clinical interest in breast cancer.     

Control of matrix metalloproteinase activity in cancer

Remodelling of extracellular matrix (ECM) and basement membranes is a key component of the process of tumour cell invasion and metastasis. Matrix metalloproteinases (MMPs) are one of the major classes of enzymes involved in degrading ECM, having different substrate specificities and being inhibited by naturally occurring tissue inhibitors (TIMPs). Elevated levels of MMPs have been associated with poor prognosis for a variety of malignancies. However, the expression and effective action of MMPs are influenced by multiple factors: most are synthesized by stroma rather than tumour cells, suggesting tumour cell–stromal cell co-operation; receptors (MT-MMPs) have to be present on tumour cells for binding and activation of MMP; co-ordination of tissue proteolysis and subsequent integrin binding will aid cell movement through a matrix; integrin receptors can directly moderate the production of MMP. These various components need to be considered when trying to determine the key events regulating matrix proteolysis and hence invasion. © 1997 John Wiley & Sons, Ltd.     

早孕和足月滋养细胞中侵袭相关性基因的表达变化

目的：探讨早孕和足月滋养细胞侵袭能力差异的机制。方法：分离并在体外培养早孕和足月的滋养细胞为，采用RT-PCR法，检测其侵袭相关性基因的表达。结果：早孕滋养细胞表达MMP-2、MMP-9、TIMP-1和UPA；而晚孕滋养细胞表达MMP-2、TIMP-1、TIMP-2和PAI。晚孕滋养细胞TTMP—1的表达显著高于早孕滋养细胞，而MMP-2的表达则无明显变化。结论：MMP-9等蛋白酶及其抑制因子表达的变化，可能是影响滋养细胞侵袭性的重要原因。     

Differential expression of matrix metalloproteinase and tissue inhibitor of matrix metalloproteinase genes in the mouse central nervous system in normal and inflammatory states.

Matrix metalloproteinases (MMPs) are implicated in the pathogenesis of inflammatory disorders of the central nervous system (CNS) whereas the contribution of the major endogenous counter-regulators of MMPs, the tissue inhibitors of the matrix metalloproteinases (TIMPs), is unclear. We investigated the temporal and spatial expression patterns in the CNS of nine MMP genes and three TIMP genes in normal mice, in mice with EAE, and in transgenic mice with astrocyte (glial fibrillary acidic protein)-targeted expression of the cytokines interleukin-3 (macrophage/microglial demyelinating disease), interleukin-6 (neurodegenerative disease), or tumor necrosis factor-alpha (lymphocytic encephalomyelitis). In normal mice, the MMPs MT1-MMP, stromelysin 3, and gelatinase B were expressed at low levels, whereas high expression of TIMP-2 and TIMP-3 was observed predominantly in neurons and in the choroid plexus, respectively. In EAE and the transgenic mice, significant induction or up-regulation of various MMP genes was observed, the pattern of which was somewhat specific for each of the models, and there was significant induction of TIMP-1. In situ localization experiments revealed a dichotomy between MMP expression that was restricted to leukocytes and possibly microglia within inflammatory lesions and TIMP-1 expression that was observed in activated astrocytes circumscribing the lesions. These findings demonstrate specific spatial and temporal regulation in the expression of individual MMP and TIMP genes in the CNS in normal and inflammatory states. The distinct localization of TIMP-1 and MMP expression during CNS inflammation suggests a dynamic state in which the interplay between these gene products may determine both the size and resolution of the destructive inflammatory focus.     

肺间质纤维化大鼠肺组织基质金属蛋白酶及其组织抑制因子含量变化

观察肺纤维化形成过程中基质金属蛋白酶（Matrix Metallo proteinas简称MMPs)及其组织抑制因子（Tissue inhibitors of Metallo proteinases简称TIMPs)含量的变化,探讨其在肺纤维化发病中的作用。将Wistar大鼠60只,随机均分为对照组及模型组,气管内注入博莱霉素A5 5mg/kg,制备肺间质纤维化动物模型,观察注药后1、3、7、14及28d肺脏病理变化,利用酶谱法及免疫印记法分析肺组织MMP－2、MMP－9、TIMP－1的含量变化。结果显示各模型组pro-MMP-2、MMP－2、TIMP－1蛋白含量均较对照组增加,尤其7、14及28d组MMP－2较前明显增多。而MMP－9变化不很明显。提示在肺纤维化形成过程中,pro-MMP-2、MMP－2及TIMP－1都有所增高,MMP／TIMP比例失衡是最终导致肺间质纤维化形成的重要因素。     

Matrix metalloproteinases,TIMPs and growth factors regulating ameloblastoma behaviour

Siqueira A S, Carvalho M R D, Monteiro A C D, Freitas V M, Jaeger R G & Pinheiro J J V.
(2010) Histopathology 57 , 128–137
Matrix metalloproteinases, TIMPs and growth factors regulating ameloblastoma behaviour Aims: Ameloblastoma is an odontogenic neoplasm with local invasiveness and recurrence. We have previously suggested that growth factors and matrix metalloproteinases (MMPs) influence ameloblastoma invasiveness ¹ . The aim was to study expression of MMPs, tissue inhibitor of metalloproteinases (TIMPs) and growth factors in ameloblastoma. Methods and results: Thirteen cases of solid/multicystic ameloblastoma were examined. As a control, calcifying cystic odontogenic tumour (CCOT), a non‐invasive odontogenic neoplasm with ameloblastomatous epithelium was also studied. Immunohistochemistry detected MMPs, TIMPs and growth factors in ameloblastoma and CCOT. The labelling index (LI) of MMP‐9 and TIMP‐2 was significantly higher in ameloblastoma compared with CCOT. The LI of epidermal growth factor (EGF), transforming growth factor (TGF)‐α and epidermal growth factor receptor (EGFR) was also increased in ameloblastoma. This neoplasm showed greater expression of MMPs, TIMPs and growth factors compared with CCOT. We then analysed these molecules in ameloblastoma cells and stroma. Ameloblastoma cells exhibited increased LI of MMP‐1, ‐2 and EGFR. We found a positive correlation between EGF and TIMP‐1, and between TGF‐α and TIMP‐2. It is known that signals generated by growth factors are transduced by the ERK pathway. Ameloblastoma stroma exhibited the phosphorylated (activated) form of ERK. Conclusions: These results suggest an interplay involving growth factors MMPs and TIMPs that may contribute to ameloblastoma behaviour. Signals generated by this molecular network would be transduced by ERK 1/2 pathway.     

Platelet-derived growth factor and transforming growth factor-β modulate the expression of matrix metalloproteinases and migratory function of human airway smooth muscle cells

Background Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinase (TIMPs) have been suggested to be involved in the pathogenesis of asthma. Their expression in airway smooth muscle (ASM) cells could be involved in collagen turnover and migration of these cells and thus may contribute to airway remodelling.
Objective To examine the effect of pro-fibrotic growth factors TGF-β and platelet-derived growth factor (PDGF) on the expression of MMPs/TIMPs in cultured human ASM cells and to examine the role of MMP in the migration of ASM cells.
Methods ASM cells were stimulated with TGF-β and/or PDGF. Expression and activity of MMP-1, MMP-2, MMP-3, TIMP-1 and TIMP-2 were evaluated by quantitative RT-PCR, Western blot and zymography. Modified Boyden-chamber migration assay was performed to investigate the effect of secreted MMP-3 and TIMP-1 on ASM-cell migration.
Results PDGF strongly up-regulated the expression of MMP-1 at mRNA and protein levels. PDGF, when combined with TGF-β, caused synergistic up-regulation of MMP-3. TIMP-1 was additively up-regulated by TGF-β and PDGF. These growth factors had no effect on the expression of MMP-2 and TIMP-2. U0126, an extracellular signal-regulated kinase (ERK) pathway inhibitor, inhibited the up-regulation of MMP-1 by PDGF. The synergistic/additive up-regulation of MMP-3 and TIMP-1 was inhibited by U0126 and SB431542, a Smad pathway inhibitor. Supernatant from ASM cells in which MMP-3 production was knocked down by RNA interference showed a decreased migratory effect on ASM cells, whereas supernatant from cells with suppressed TIMP-1 expression resulted in increased migration.
Conclusion Our results suggest that PDGF with/without TGF-β could facilitate migration of ASM cells by modification of MMP–TIMP balance through the ERK pathway.     

Monocyte-derived dendritic cell subpopulations use different types of matrix metalloproteinases inhibited by GM6001

Matrix metalloproteinases (MMPs) are endopeptidases with the potential to cleave extracellular matrix, support tissue renewal and regulate cell migration. Functional activities of MMPs are regulated by tissue inhibitors of MMPs (TIMPs) and disruption of the MMP–TIMP balance has pathological consequences. Here we studied the expression and secretion of MMPs and TIMPs in CD1a⁻ and CD1a⁺ monocyte-derived dendritic cell (DC) subpopulations. Our results showed that monocytes express TIMPs but lack MMPs, whereas upon differentiation to moDCs and in response to activation signals the expression of MMPs is increased and that of TIMPs is decreased. MMP-9 is expressed dominantly in the CD1a⁻ subpopulation, while MMP-12 is preferentially expressed in CD1a⁺ cells. Experiments performed with the synthetic MMP inhibitor GM6001 revealed that this drug efficiently inhibits the migration of moDCs through inactivation of MMPs. We conclude that modulation of MMP activity by GM6001 emerges as a novel approach to manipulate DC migration under inflammatory conditions.     

Membrane-type-1 matrix metalloproteinase, matrix metalloproteinase 2, and tissue inhibitor of matrix proteinase 2 in prostate cancer: identification of patients with poor prognosis by immunohistochemistry

Overexpression of the matrix metalloproteinase (MMP) 2 is associated with poor prognosis in many tumor types. Membrane-type-1 MMP (MMP14) activates MMP2 using pro-MMP2 specific inhibitor, tissue inhibitor of matrix proteinase 2 (TIMP2), as a receptor. We evaluated, by immunohistochemistry on 189 T3N0-2M0 prostate cancer (Pca) cases, the influence of MMP2, MMP14, and TIMP2 expression, individually and in association, on Pca disease-free survival (DFS). We evaluated marker expression separately in cancer, stromal, and benign epithelial (BE) cells according to a percentage scale (0%, <10%, 10%-50%, and >50%). Median follow-up was 4.61 years. In BE cells, there was an inverse relationship between initial prostate-specific antigen serum level and T3 stage with MMP14 expression (P = .003) and between pN stage and TIMP2 expression (P = .04). The most significant results with survival were obtained by dichotomizing the cases between those with less than 10% and at least 10% of cells expressing the marker, the latter category representing overexpression. TIMP2 overexpression in stromal cells was associated with a longer DFS with a hazard ratio of 0.573 (P = .02) for time to recurrence. MMP2 overexpression by BE cells correlated with a shorter DFS using a multivariate trend test (hazard ratio = 1.46, P = .02). Stromal cells expressing less than 10% TIMP2 and MMP2 overexpression was the only combination that was significantly associated with a shorter DFS (log-rank test, P = .0001). This study suggests that MMP14 is involved mostly in Pca implantation and that MMP2 and TIMP2 expression by reactive stromal cells might be used as predictors of DFS in T3N0-2M0 Pca.     

Differential expression of matrix metalloproteinases and their inhibitors in non-small cell lung cancer

In a comprehensive immunohistochemical study of the expression of ten metalloproteinases (MMPs) and their four inhibitors (TIMPs) in 115 non-small cell lung carcinomas (NSCLCs), the findings have been correlated with the histological and clinical features of the tumours. All MMPs and TIMPs were expressed in tumours, with frequencies ranging from 41% for MMP-2 to 68% for MMP-13. Stromal immunoreactivity ranged from 6% for TIMP-4 to 87% for MMP-13. In some tumours, an overexpression of these proteins, as revealed by stronger staining in cancer cells than in adjacent normal bronchial epithelium, was also observed. The frequency ranged from 1% for MMP-3 to 28% for MMP-13. Compared with squamous cell carcinoma (SqCC), adenocarcinoma (AdC) more frequently overexpressed MMP-1, -11, -13, -14, and TIMP-2, and TIMP-1 and/or TIMP-2 overexpression positively correlated with more advanced stage disease. None of the MMP or TIMP expression correlated with the ras genotype of the tumours. The higher frequency of MMP overexpression in AdC than in SqCC may relate to the greater tendency of the former for systemic metastasis. The association of TIMP-1 overexpression with more advanced disease may suggest a role in prognosis.     

Serum matrix metalloproteinase and tissue inhibitor of metalloproteinase concentrations in infertile women achieved pregnancy following IVF-ET

PROBLEM: Matrix metalloproteinase (MMP) and tissue inhibitor of metalloproteinase (TIMP) play important roles throughout various stages of pregnancy, including embryo implantation, trophoblastic invasion, placentation in early gestation, and cervical dilatation in later gestation, and feto-maternal membrane lysis. It would be beneficial if assessment of serum concentrations of MMP and TIMP could predict successful implantation following in vitro fertilization-embryo transfer (IVF-ET). This study was performed to compare serum MMP and TIMP concentrations between patients with and without the establishment of pregnancy following ET. METHOD OF STUDY: Ten patients who conceived and 10 patients who did not after IVF-ET were entered in the present study. Only intra-uterine single pregnancies with uneventful courses to term were included in the study subjects. Blood samples were obtained at 7, 14 and 21 days after oocyte retrieval. Serum concentrations of MMP-1, MMP-2, MMP-3, and TIMP-1 were measured using enzyme-linked immunosorbent assay. These variables were compared with estradiol (E(2)), progesterone (P(4)), and betahCG levels in the patients' sera. Clinical pregnancies were diagnosed only when fetal heartbeat was visualized on ultrasound. RESULTS: There were no significant differences in serum MMP concentrations between the pregnant group and the non-pregnant group. However, serum TIMP-1 concentrations on Days 14 and 21 in the pregnant group were significantly higher than those in non-pregnant group [Day 14: 223.1 +/- 11.9 versus 177.5 +/- 20.6 ng/mL (P = 0.004); Day 21: 215.4 +/- 27.8 versus 181.5 +/- 27.4 ng/mL (P = 0.03)]. Serum TIMP-1 concentrations were also correlated with serum E(2) and P(4) levels (P < 0.0001), but not with those of the MMPs. None of MMP nor TIMP-1 were correlated with serum betahCG level. CONCLUSIONS: It was demonstrated that the patients who successfully conceived after IVF-ET showed significantly higher levels of TIMP-1 at 14 and 21 days after IVF-ET, but not at day 7; further work will be required to determine if serum TIMP-1 can be used to improve prediction of pregnancy outcome in these patients.     

Progressive cellular response in the lamina propria of the colorectal adenoma–carcinoma sequence

Aims:  The lamina propria is inevitably involved in epithelial transformation. The aim was to evaluate the dynamic cellular changes in the tumour lamina propria throughout the colorectal adenoma–carcinoma sequence.
Methods and results:  Using immunohistochemistry and double immunohistochemistry, we examined lamina propria cellular changes in 41 colorectal adenomas, 25 colorectal cancers and 15 control tissues. The results showed that the proliferation labelling index in lamina propria cells began to increase in the precancerous lesions (adenomas) and became even higher in the colorectal cancers; these proliferative cells were primarily identified as myofibroblasts and lymphocytes. Phenotypic analysis revealed gradually increasing lymphocytic infiltration in both the lamina propria and adenomatous epithelium, as well as myofibroblasts in the lamina propria. However, the intraepithelial macrophage density also showed a tendency to increase gradually. Furthermore, cyclooxygenase-2-expressing cell density and microvessel density gradually increased in the tumour lamina propria throughout the adenoma–carcinoma sequence.
Conclusions:  Progressive cellular responses in the lamina propria could be involved in the adenoma–carcinoma transition.     

Progesterone receptor modulator CDB-2914 induces extracellular matrix metalloproteinase inducer in cultured human uterine leiomyoma cells

Effects of progesterone receptor modulator CDB-2914 on the expression of the extracellular matrix (ECM) components were examined in cultured human uterine leiomyoma and myometrial cells. ECM metalloproteinase inducer (EMMPRIN), matrix metalloproteinases (MMPs), tissue inhibitors of MMP (TIMPs) and collagen levels were assessed by Western blot analysis, MMP activity assay and real-time RT-PCR. RNA interference (RNAi) of EMMPRIN was performed using small interfering mRNA. In cultured leiomyoma cells, CDB-2914 treatment at concentrations greater than or equal to 10(-8) M significantly increased EMMPRIN, MMP-1 and MMP-8 protein contents and MMP-1, MMP-2, MMP-3 and MMP-9 mRNA levels, and activity of MMP-1, MMP-2, MMP-3 and MMP-9 in the medium. TIMP-1 and TIMP-2 were significantly decreased at mRNA and protein levels by CDB-2914 treatment at concentrations > or =10(-7) M in these cells. CDB-2914 treatment decreased types I and III collagen protein contents. However, CDB-2914 treatment did not affect the ECM component expression in cultured myometrial cells. RNAi of EMMPRIN abrogated CDB-2914-mediated both induction of MMPs and reduction of TIMPs and collagens in cultured leiomyoma cells. These results suggest that CDB-2914 modulates the expression of EMMPRIN, MMPs, TIMPs and collagens in cultured leiomyoma cells without comparable effects on myometrial cells.     

Expression profiling of metalloproteinases and inhibitors in cartilage

Objective   To profile the expression of all known members of the matrix metalloproteinase ( MMP ), a disintegrin and metalloproteinase with thrombospondin motifs ( ADAMTS ), and tissue inhibitor of metalloproteinases ( TIMP s) gene families in normal cartilage and that from patients with osteoarthritis (OA).
Methods   Human cartilage was obtained from femoral heads at joint replacement for either osteoarthritis or following fracture to the neck of femur. Total RNA was purified and expression of genes assayed using quantitative real-time PCR.
Results   Several members of the above gene families were regulated in OA. Genes increasing in expression in OA were: at P  < 0.001, MMP-13 , MMP-28 , ADAMTS-16 ; at P  < 0.01, MMP-9 , MMP-16 , ADAMTS-2 , ADAMTS-14 and at P  < 0.05, MMP-2 , TIMP-3 , ADAMTS-12 . Genes decreasing in expression in OA were: at P  < 0.001, MMP-1 , MMP-3 , ADAMTS-1 ; at P  < 0.01, MMP-10 , TIMP-1 , ADAMTS-9 and at P  < 0.05, TIMP-4 , ADAMTS-5 , ADAMTS-15 . Correlation analysis revealed that groups of genes across the gene families are co-expressed in cartilage.
Conclusion   This is the first comprehensive expression profile of all known MMP , ADAMTS and TIMP genes in cartilage. Patterns of expression provide a foundation on which to understand mechanisms of gene regulation in OA and potentially for refining the specificity of anti-proteolytic therapies.