机械性去负荷后心室明胶酶及其组织型抑制剂与细胞外基质变化

目的：探讨左心室在去除压力和容量负荷下心室组织基质金属蛋白酶-2和-9及金属蛋白酶组织型抑制剂-1和-2表达水平与细胞外基质沉积量的关系。方法：12周龄雄性Lewis大鼠建立Lewis-to-Lewis腹腔异位心脏移植模型,形成左心室去负荷状态,并以同龄雄性Lewis大鼠胸腔原位心脏作为对照。移植后14d采用天狼猩红-偏振光法对移植和对照组心脏的ECM沉积量进行分析。心室组织MMP-2和MMP-9活性检测采用明胶酶谱法。MMP-2、MMP-9、TIMP-1、TIMP-2的mRNA表达水平检测采用荧光定量PCR法;TIMP-1和TIMP-2蛋白含量采用Western blot测定。结果：手术后14d,与原位心脏比较,腹腔移植心脏心肌细胞横截面积减小,并伴有心肌ECM沉积量增多（胶原容积分数5．22％±1．6％VS2．21％±0．9％,P〈0．05）,并且MMP-2、MMP-9明胶酶活性明显增强,MMP-2、MMP-9及其组织型抑制剂T1MP-1、TIMP-2的mRNA表达均增加（P〈0．05）,但TIMP-1、TIMP-2增加幅度较MMP-2、MMP-9高,TIMP-1、TIMP-2的蛋白含量均增加（P〈0．05）,TIMP-1增加幅度更为明显。结论：左心室在去除压力和容量负荷状态下心脏心室组织胶原沉积量增加,伴有MMPs／TIMPs系统失衡,尤其是TIMPs系统的明显上调。     

Tissue inhibitor of metalloproteinase-2 (TIMP-2) regulates neuromuscular junction development via a beta1 integrin-mediated mechanism

Extracellular matrix (ECM) molecules play critical roles in muscle function by participating in neuromuscular junction (NMJ) development and the establishment of stable, cytoskeleton-associated adhesions required for muscle contraction. Matrix metalloproteinases (MMPs) are neutral endopeptidases that degrade all ECM components. While the role of MMPs and their inhibitors, the tissue inhibitor of metalloproteinases (TIMPs), has been investigated in many tissues, little is known about their role in muscle development and mature function. TIMP-2 -/- mice display signs of muscle weakness. Here, we report that TIMP-2 is expressed at the NMJ and its expression is greater in fast-twitch (extensor digitorum longus, EDL) than slow-twitch (soleus) muscle. EDL muscle mass is reduced in TIMP-2-/- mice without a concomitant change in fiber diameter or number. The TIMP-2-/- phenotype is not likely due to increased ECM proteolysis because net MMP activity is actually reduced in TIMP-2-/- muscle. Most strikingly, TIMP-2 colocalizes with beta1 integrin at costameres in the wild-type EDL and beta1 integrin expression is significantly reduced in TIMP-2-/- EDL. We propose that reduced beta1 integrin in fast-twitch muscle may be associated with destabilized ECM-cytoskeletal interactions required for muscle contraction in TIMP-2-/- muscle; thus, explaining the muscle weakness. Given that fast-twitch fibers are lost in muscular dystrophies and age-related sarcopenia, if TIMP-2 regulates mechanotransduction in an MMP-independent manner it opens new potential therapeutic avenues.     

Complex role of matrix metalloproteinases in angiogenesis

Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) play a significant role in regulating angiogenesis,the process of new blood vessel formation.Interstitial collagenase (MMP-1),72kDa gelatinase A/type IV collagenase (MMP-2),and 92 kDA gelatinase B/type IV collagenase (MMP-9) dissolve extracellular matrix (ECM) and may initiate and promote angiogenesis.TIMP-1,TIMP-2,TIMP-3,and possibly,TIMP-4 inhibit neovascularization.A new paradign is emerging that matrilysin (MMP-7),MMP-9,and metalloelastase (MMP-12) may block angiogenesis by converting plasminogen to angiostatin,which is one of the most potent angiogenesis antagonists.MMPs and TIMPs play a complex role in regulating angiogenesis.An understanding of the biochemical and cellular pathways and mechanisms of angiogenesis will provide important information to allow the control of angiogenesis,e.g.the stimulation of angiogenesis for coronary collateral circulation formation;while the inhibition for treating arthritis and cancer.     

Collagenase expression and activity is modulated by the interaction of collagen types, hypoxia, and nutrition in human lung cells

Hypoxia not only controls organogenesis, embryogenesis, and wound repair, but also triggers tumor progression and metastasis. Matrix metalloproteinases (MMP), especially gelatinases (MMP-2, MMP-9) regulate the composition and stability of the extracellular matrix (ECM), which affects cell proliferation, migration, and differentiation. This study investigated the effect of hypoxia alone and in combination with ECM compounds and nutrition on MMP-2 and MMP-9 expression, activity, and synthesis in human lung fibroblasts and pulmonary vascular smooth muscle cells (VSMC). We also determined the expression of the tissue inhibitors of MMP (TIMP-1, -2). Cells were grown on plastic, collagen-I, collagen-IV, or gelatin and in either starving medium (0.1% serum) or growth medium (5% serum), and were subjected to normoxia or hypoxia (1% O(2)). Collagenases expression was determined by zymography. TIMP-1, -2 expression was assessed by Western blotting and RT-PCR. Depending on serum concentration human lung cells expressed pro-MMP-2 on all substrates. Hypoxia increased pro-MMP-2 expression, on collagen type I or type IV further via Erk1/2 and p38 MAP kinase signaling. MMP-9 was only expressed when cells were grown on collagen type IV and increased with serum concentration, and by hypoxia. TIMP-1 expression was only expressed when cells were grown on collagen type I and was significantly increased by hypoxia, while TIMP-2 expression was unchanged. We demonstrated that the hypoxia, ECM composition, and nutrition, rather than one of these conditions alone, modulate the expression and activity of collagenases and their inhibitors in primary human lung fibroblasts.     

Expression of matrix metalloproteinase (MMP)-2, MMP-14 and tissue inhibitor of matrix metalloproteinase (TIMP)-2 during bovine placentation and at term with or without placental retention

Matrix metalloproteinases (MMPs) and counteracting tissue inhibitors of metalloproteinases (TIMPs) are balancing extracellular matrix (ECM) formation and degradation. The latter is believed to be an important aspect for the detachment of fetal membranes postpartum when loosening the feto-maternal connection which is a prerequisite to avoid placental retention a common disease in cows leading to considerable economic loss. Membrane-type (MT) MMPs have been suggested as potential activators controlling ECM remodelling. In particular, MT1-MMP (MMP-14) is able to degrade ECM substrates and activate MMP-2 through binding TIMP-2 at the cell surface. Since the connection between the trophoblast and the maternal caruncular epithelium is supported by integrin receptors bound to ECM, we hypothesize that impaired modulation of the ECM by TIMPs/MMPs participates in the aetiology of bovine retained fetal membranes. To analyse this involvement, placentomes were collected from cows after term parturition and timely release of fetal membranes (n = 4) and cows with retained fetal membranes after various treatments for the induction of parturition using progesterone antagonist (aglepristone), PGF_2α analogue, glucocorticoid, and after elective caesarean sections (each group n = 3). The expression of MMP-14, MMP-2 and of TIMP-2 was examined by real-time-PCR, immunohistochemistry, Western blot and zymography. The relative mRNA expression levels of MMP-14 remained unchanged, while the expression levels of TIMP-2 and MMP-2 partly increased in animals with induced parturition and retention of fetal membranes compared to animals without placental retention. MMP-14 protein was expressed in cells of the uninucleated trophoblast, the fetal mesenchyme and maternal stroma. TIMP-2 was present exclusively in trophoblast giant cells, while MMP-2 could be detected in uninucleated trophoblast cells and the fetal mesenchyme. The presence of the activated enzyme was confirmed by zymography. In conclusion, MMP-14, MMP-2 and TIMP-2 are co-localized in the fetal compartment and therefore could influence the timely release of fetal membranes in cattle.     

MMP-2 expression precedes the final ripening process of the bovine cervix

Collagen is denatured in the gradual cervical ripening process during late pregnancy, already before the onset of final cervical ripening at parturition. Matrix Metallo Proteinases (MMPs) might be responsible for this process. To investigate the presence and potential function of MMPs at the different stages of the ripening process, serial cervical biopsies were obtained from 10 cows at Days 185 and 275 of pregnancy (approximately 5 days before calving), at parturition and at 30 days after parturition. The mRNA and protein expression of MMP-1, MMP-2, and MMP-9 and of the tissue inhibitors of MMPs (TIMP)-1 and TIMP-2 were semi-quantitatively determined using RT-PCR, respectively, zymography, Westernblot, and ELISA techniques and the localization of MMP-2 protein and presence of granulocytes by immunohistochemistry and Luna staining. At parturition compared to 185 days pregnancy the MMP-1 protein expression and the numbers of granulocytes were significantly increased by 3 and 26-fold respectively. MMP-2 mRNA and protein expression had already increased 2.5 (P < 0.05) and twofold (P < 0.05) at 5 days before parturition, prior to final ripening. At that time, MMP-2 was present in smooth muscle cells and extra cellular matrix. TIMP-1 mRNA expression was significantly increased at parturition and TIMP-2 mRNA expression peaked at 5 days before parturition. The increased expression of MMP-2 at 5 days before parturition, suggests that in the cow MMP-2 is responsible for collagen denaturation in the last part of gradual cervical ripening, while MMP-1 and MMP-9 are only active during the final cervical ripening process at parturition.     

Acute exercise induced changes in rat skeletal muscle mRNAs and proteins regulating type IV collagen content

This experiment tested the hypothesis that running-induced damage to rat skeletal muscle causes changes in synthesis and degradation of basement membrane type IV collagen and to proteins regulating its degradation. Samples from soleus muscle and red and white parts of quadriceps femoris muscle (MQF) were collected 6 h or 1, 2, 4, or 7 days after downhill running. Increased muscle beta-glucuronidase activity indicated greater muscle damage in the red part of MQF than in the white part of MQF or soleus. In the red part of MQF, type IV collagen expression was upregulated at the pretranslational level and the protein concentration decreased, whereas matrix metalloproteinase-2 (MMP-2), a protein that degrades type IV collagen, and tissue inhibitor of metalloproteinase-2 (TIMP-2), a protein that inhibits degradation, were increased in parallel both at mRNA and protein levels. Type IV collagen mRNA level increased in the white part of MQF and soleus muscle. The protein concentration increased in the white part of MQF and was unchanged in soleus muscle. MMP-2 and TIMP-2 changed only slightly in the white part of MQF and soleus muscle. The changes seem to depend on the severity of myofiber injury and thus probably reflect reorganization of basement membrane compounds.     

HPV16 oncoproteins induce MMPs/RECK-TIMP-2 imbalance in primary keratinocytes: possible implications in cervical carcinogenesis

Cervical cancer is the third most common cancer in women worldwide. Persistent infection with high-risk HPV types, principally HPV16 and 18 is the main risk factor for the development of this malignancy. However, the onset of invasive tumor occurs many years after initial exposure in a minority of infected women. This suggests that other factors beyond viral infection are necessary for tumor establishment and progression. Tumor progression is characterized by an increase in secretion and activation of matrix metalloproteinases (MMPs) produced by either the tumor cells themselves or tumor-associated fibroblasts or macrophages. Increased MMPs expression, including MMP-2, MMP-9 and MT1-MMP, has been observed during cervical carcinoma progression. These proteins have been associated with degradation of ECM components, tumor invasion, metastasis and recurrence. However, few studies have evaluated the interplay between HPV infection and the expression and activity of MMPs and their regulators in cervical cancer. We analyzed the effect of HPV16 oncoproteins on the expression and activity of MMP-2, MMP-9, MT1-MMP, and their inhibitors TIMP-2 and RECK in cultures of human keratinocytes. We observed that E7 expression is associated with increased pro-MMP-9 activity in the epithelial component of organotypic cultures, while E6 and E7 oncoproteins co-expression down-regulates RECK and TIMP-2 levels in organotypic and monolayers cultures. Finally, a study conducted in human cervical tissues showed a decrease in RECK expression levels in precancer and cancer lesions. Our results indicate that HPV oncoproteins promote MMPs/RECK-TIMP-2 imbalance which may be involved in HPV-associated lesions outcome.     

TGF-beta3-induced palatogenesis requires matrix metalloproteinases

Cleft lip and palate syndromes are among the most common congenital malformations in humans. Mammalian palatogenesis is a complex process involving highly regulated interactions between epithelial and mesenchymal cells of the palate to permit correct positioning of the palatal shelves, the remodeling of the extracellular matrix (ECM), and subsequent fusion of the palatal shelves. Here we show that several matrix metalloproteinases (MMPs), including a cell membrane-associated MMP (MT1-MMP) and tissue inhibitor of metalloproteinase-2 (TIMP-2) were highly expressed by the medial edge epithelium (MEE). MMP-13 was expressed both in MEE and in adjacent mesenchyme, whereas gelatinase A (MMP-2) was expressed by mesenchymal cells neighboring the MEE. Transforming growth factor (TGF)-beta3-deficient mice, which suffer from clefting of the secondary palate, showed complete absence of TIMP-2 in the midline and expressed significantly lower levels of MMP-13 and slightly reduced levels of MMP-2. In concordance with these findings, MMP-13 expression was strongly induced by TGF-beta3 in palatal fibroblasts. Finally, palatal shelves from prefusion wild-type mouse embryos cultured in the presence of a synthetic inhibitor of MMPs or excess of TIMP-2 failed to fuse and MEE cells did not transdifferentiate, phenocopying the defect of the TGF-beta3-deficient mice. Our observations indicate for the first time that the proteolytic degradation of the ECM by MMPs is a necessary step for palatal fusion.     

Imbalances between Matrix Metalloproteinases (MMPs) and Tissue Inhibitor of Metalloproteinases (TIMPs) in Maternal Serum during Preterm Labor

Background

Matrix metalloproteinases (MMPs) are involved in remodeling of the extracellular matrix (ECM) during pregnancy and parturition. Aberrant ECM degradation by MMPs or an imbalance between MMPs and their tissue inhibitors (TIMPs) have been implicated in the pathogenesis of preterm labor, however few studies have investigated MMPs or TIMPs in maternal serum. Therefore, the purpose of this study was to determine serum concentrations of MMP-3, MMP-9 and all four TIMPs as well as MMP:TIMP ratios during term and preterm labor.

Methods

A case control study with 166 singleton pregnancies, divided into four groups: (1) women with preterm birth, delivering before 34 weeks (PTB); (2) gestational age (GA) matched controls, not in preterm labor; (3) women at term in labor and (4) at term not in labor. MMP and TIMP concentrations were measured using Luminex technology.

Results

MMP-9 and TIMP-4 concentrations were higher in women with PTB vs. GA matched controls (resp. p = 0.01 and p<0.001). An increase in MMP-9:TIMP-1 and MMP-9:TIMP-2 ratio was observed in women with PTB compared to GA matched controls (resp. p = 0.02 and p<0.001) as well as compared to women at term in labor (resp. p = 0.006 and p<0.001). Multiple regression results with groups recoded as three key covariates showed significantly higher MMP-9 concentrations, higher MMP-9:TIMP-1 and MMP-9:TIMP-2 ratios and lower TIMP-1 and -2 concentrations for preterm labor. Significantly higher MMP-9 and TIMP-4 concentrations and MMP-9:TIMP-2 ratios were observed for labor.

Conclusions

Serum MMP-9:TIMP-1 and MMP-9:TIMP-2 balances are tilting in favor of gelatinolysis during preterm labor. TIMP-1 and -2 concentrations were lower in preterm gestation, irrespective of labor, while TIMP-4 concentrations were raised in labor. These observations suggest that aberrant serum expression of MMP:TIMP ratios and TIMPs reflect pregnancy and labor status, providing a far less invasive method to determine enzymes essential in ECM remodeling during pregnancy and parturition.     

Effects of matrix proteins on the expression of matrix metalloproteinase-2, -9, and -14 and tissue inhibitors of metalloproteinases in human cytotrophoblast cells during the first trimester.

The activity of matrix metalloproteinases (MMPs) specifies the ability of the trophoblast cell to degrade extracellular matrix (ECM) substrates. Usually the process of normal human placentation involves a coordinated interaction between the fetal-derived trophoblast cells and their microenvironment in the uterus. In this study, the effects of ECM proteins on the expression of MMP-2, -9, and -14 (membrane-type MMP-1); and the production of tissue inhibitors of metalloproteinase (TIMP) types -1, -2, and -3 have been investigated. Cytotrophoblast cells at 9 or 10 wk of gestation were cultured on various ECM coated dishes under serum-free conditions. Gelatin zymography analysis showed that cells grown on fibronectin (FN), laminin (LN), and vitronectin (VN) secreted more MMP-9 (about 1.5- to 3-fold more) than cells cultured on collagen I (Col I), whereas the secretion of MMP-9 by cells cultured on collagen IV (Col IV) was only half that by the cells on Col I. Northern Blot analysis gave the same results as zymography, indicating that expression of the MMP-9 gene in cytotrophoblast cells can be affected by matrix proteins. There was no significant difference in the expression of MMP-2 either at protein or mRNA levels among the cells cultured on the different matrix substrates. The expression of MMP-14 was regulated in a manner similar to that of MMP-2. Using ELISA, we detected higher levels of TIMP-1 in the culture medium of cells grown on VN, LN, and FN compared with that grown on Col I. But the expression of TIMP-3 mRNA was remarkably inhibited by VN, and ECM proteins had no effect on TIMP-1 and TIMP-2 mRNA expression. It was also observed that cultured cytotrophoblast cells expressed the corresponding receptors for the tested matrix proteins, such as integrins alpha(1), alpha(5), alpha(6), beta(1), and beta(4). Furthermore, the adhesiveness of cytotrophoblast cells on Col I, Col IV, FN, and LN was increased by 62%, 45%, 21%, and 22%, respectively, when compared with adhesiveness on VN. Isolated cytotrophoblast cells remained stationary when cultured on dishes coated with Col I and Col IV, but they assumed a more motile morphology and aggregated into a network when cultured on LN and VN. These data indicate that human trophoblast cells interact with their microenvironment to control their behavior and function.     

MMP-9 and MMP-2 gelatinases and TIMP-1 and TIMP-2 inhibitors in breast cancer: correlations with prognostic factors

The goal of our study was to analyse the prognostic values for some matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) in breast cancer. We evaluated the activity and the expression levels of MMP-9, MMP-2, TIMP-1 and TIMP-2 in malignant versus benign fresh breast tumor extracts. For this purpose, gelatinzymography, immunoblotting and ELISA were used to analyse the activity and expression of MMPs and TIMPs. We found that MMP-9 expression level and activity are increased in malignant tumors. In addition, MMP-9/TIMP-1 and MMP-2/TIMP-2 ratio values obtained by us were significantly different in malignant tumors compared to benign tumors. We suggest that the abnormal MMP-9/TIMP-1 balance plays a role in the configuration of breast invasive carcinoma of no special type and also in tumor growth, while altered MMP-2/TIMP-2 ratio value could be associated with lymph node invasion and used as a prognostic marker in correlation with Nottingham Prognostic Index. Finally, we showed that in malignant tumors high expression of estrogen receptors is associated with enhanced activity of MMP-2 and increased bcl- 2 levels, while high expression of progesterone receptors is correlated with low TIMP-1 protein levels.     

Plaque oxysterols induce unbalanced up-regulation of matrix metalloproteinase-9 in macrophagic cells through redox-sensitive signaling pathways: Implications regarding the vulnerability of atherosclerotic lesions

An imbalance in the matrix metalloproteinases/tissue inhibitors of metalloproteinases (MMPs/TIMPs) contributes to atherosclerotic plaque destabilization and rupture. Here we determined whether oxysterols accumulating in advanced atherosclerotic lesions play a role in plaque destabilization. In human promonocytic U937 cells, we investigated the effects of an oxysterol mixture of composition similar to that in advanced human carotid plaques on the expression and synthesis of MMP-9 and its endogenous inhibitors TIMP-1 and TIMP-2. A marked increment of MMP-9 gene expression, but not of its inhibitors, was observed by real-time RT-PCR; MMP-9 gelatinolytic activity was also found increased by gel zymography. Consistently, a net increment of MMP-9 protein level was also observed by immunoblotting. Using antioxidants or specific inhibitors or siRNAs, we demonstrated that the oxysterol mixture induces MMP-9 expression through: (i) overproduction of reactive oxygen species, probably by NADPH-oxidase and mitochondria; (ii) up-regulation of mitogen-activated protein kinase signaling pathways via protein kinase C; and (iii) up-regulation of activator protein-1- and nuclear factor-κB-DNA binding. These results suggest, for the first time, that oxysterols accumulating in advanced atherosclerotic lesions significantly contribute to plaque vulnerability by promoting MMP-9/TIMP-1/2 imbalance in phagocytic cells.     

Effect of compressive force on the expression of MMPs, PAs, and their inhibitors in osteoblastic Saos-2 cells

Bone matrix turnover is regulated by matrix metalloproteinases (MMPs), tissue inhibitors of matrix metalloproteinases (TIMPs), and the plasminogen activation system, including tissue-type plasminogen activator (tPA), urokinase-type plasminogen activator (uPA), and plasminogen activator inhibitor type-1 (PAI-1). We previously demonstrated that 1.0g/cm(2) of compressive force was an optimal condition for inducing bone formation by osteoblastic Saos-2 cells. Here, we examined the effect of mechanical stress on the expression of MMPs, TIMPs, tPA, uPA, and PAI-1 in Saos-2 cells. The cells were cultured in Dulbecco's modified Eagle's medium containing 10% fetal bovine serum and with or without continuously compressive force (0.5-3.0g/cm(2)) for up to 24h. The levels of MMPs, TIMPs, uPA, tPA, and PAI-1 gene expression were estimated by determining the mRNA levels using real-time PCR, and the protein levels were determined using ELISA. The expression levels of MMP-1, MMP-2, MMP-14, and TIMP-1 markedly exceeded the control levels at 1.0g/cm(2) of compressive force, whereas the expression levels of MMP-3, MMP-13, TIMP-2, TIMP-3, TIMP-4, tPA, uPA, and PAI-1 markedly exceeded the control levels at 3.0g/cm(2). These results suggest that mechanical stress stimulates bone matrix turnover by increasing these proteinases and inhibitors, and that the mechanism for the proteolytic degradation of bone matrix proteins differs with the strength of the mechanical stress.     

Matrix metalloprotease-9 dysregulation in lower airway secretions of cystic fibrosis patients

Matrix metalloproteases (MMPs) are proteolytic enzymes that regulate extracellular matrix turnover and aid in restoring tissue architecture following injury. There is an emerging role for extracellular matrix destruction in the pathogenesis of chronic neutrophilic lung diseases. In this study, we examined the expression and activity profiles of MMPs in lower airway secretions from cystic fibrosis (CF) patients, patients with acute respiratory failure (ARF), and normal controls. A discrete repertoire of MMP isoforms was found in the CF samples, with robust MMP-9 expression compared with normal controls and ARF. CF samples possessed increased levels of active MMP-9, as well as decreased amounts of tissue inhibitor of metalloprotease-1 (TIMP-1), a natural inhibitor of MMP-9. The CF inpatient samples demonstrated fully active MMP-9 activity compared with CF outpatients, ARF, and normal controls. CF samples also demonstrated increased human neutrophil elastase (HNE) levels compared with ARF and normal controls. To examine potential mechanisms for the protease dysregulation seen in the CF clinical samples, in vitro studies demonstrated that HNE could activate pro-MMP-9 and also degrade TIMP-1; this HNE-based activation, however, was not seen with MMP-8. A strong correlation was seen between HNE and MMP-9 activity in CF inpatient samples. Finally, the dysregulated MMP-9 activity seen in CF inpatient sputum samples could be significantly reduced by the use of MMP-9 inhibitors. Collectively, these findings further emphasize the proposed protease/antiprotease imbalance in chronic neutrophilic lung disease, providing a potential mechanism contributing to this proteolytic dysregulation.     

Gelatinases and their inhibitors in tumor metastasis: from biological research to medical applications

The involvement of matrix metalloproteinase (MMPs)-2 and -9, also known as gelatinases, in cancer cell migration and invasion has been well documented, although it is not yet clear how they facilitate metastasis formation in the course of malignancies. The idea that gelatinases are responsible for degradation of extracellular matrix (ECM) components and breakdown of basement membrane (BM) tissue boundaries has turned out not to be entirely correct. An action by remodelling the ECM components of the BM exposing new cryptic sites, or releasing growth factors, cytokines, or active ECM proteolysed fragments seems to be nearer to the truth. On the other hand, tissue inhibitors of gelatinase activity (TIMP-2), are involved both in the MMP-2 activation process; in concert with membrane type 1-MMP (MT1-MMP), and in the inhibition of gelatinolytic activity. Therefore proteolysis, the central step for cancer metastasis, should occur as a result of an imbalance between MMP-2 and TIMP-2. Many studies have reported the importance of this balance in patients with different malignancies, and considerable effort is currently being spent on the study of molecules that can shift the balance in favour of inhibition of MMP proteolytic activity. In this review we focus on the role of gelatinase activity in cancer invasion, addressing the following issues: how and where proteolysis occurs in cancer tissues, how it can be regulated, what the clinical implications are of the studies reported in literature so far, and finally what the future developments in this field that could have an impact on the management of patients affected by malignancies may be.