MiR-21 Enhances Melanoma Invasiveness via Inhibition of Tissue Inhibitor of Metalloproteinases 3 Expression: In Vivo Effects of MiR-21 Inhibitor

Metastatic melanoma is the most aggressive form of this cancer. It is important to understand factors that increase or decrease metastatic activity in order to more effectively research and implement treatments for melanoma. Increased cell invasion through the extracellular matrix is required for metastasis and is enhanced by matrix metalloproteinases (MMPs). Tissue inhibitor of metalloproteinases 3 (TIMP3) inhibits MMP activity. It was previously shown by our group that miR-21, a potential regulator of TIMP3, is over-expressed in cutaneous melanoma. It was therefore hypothesized that increased levels of miR-21 expression would lead to decreased expression of TIMP3 and thereby enhance the invasiveness of melanoma cells. miR-21 over-expression in the melanoma cell lines WM1552c, WM793b, A375 and MEL 39 was accomplished via transfection with pre-miR-21. Immunoblot analysis of miR-21-overexpressing cell lines revealed reduced expression of TIMP3 as compared to controls. This in turn led to a significant increase in the invasiveness of the radial growth phase cell line WM1552c and the vertical growth phase cell line WM793b (p < 0.05), but not in the metastatic cell lines A375 or MEL 39. The proliferation and migration of miR-21 over-expressing cell lines was not affected. Reduced expression of TIMP3 was achieved by siRNA knockdown and significantly enhanced invasion of melanoma cell lines, mimicking the effects of miR-21 over-expression. Treatment of tumor cells with a linked nucleic acid antagomir to miR-21 inhibited tumor growth and increased tumor expression of TIMP3 in vivo in 01B74 Athymic NCr-nu/nu mice. Intra-tumoral injections of anti-miR-21 produced similar effects. This data shows that increased expression of miR-21 enhanced the invasive potential of melanoma cell lines through TIMP3 inhibition. Therefore, inhibition of miR-21 in melanoma may reduce melanoma invasiveness.     

Gamma-irradiation enhances RECK protein levels in Panc-1 pancreatic cancer cells

Radiotherapy is an important treatment for many malignant tumors, but there are recent reports that radiation may increase the malignancy of cancer cells by stimulating expression of type IV collagenases. In this study, we examined changes in matrix metalloproteinase (MMP) inhibitors, such as the tissue inhibitors of metalloproteinase (TIMP)-1, TIMP-2 and RECK, in response to irradiation in Panc-1 pancreatic cancer cells. Irradiation increased RECK protein levels but not mRNA levels, whereas no significant changes were found in TIMP-1 and TIMP-2. The enhanced RECK protein levels were associated with an increase in MMP inhibitory activity. However, irradiation slightly but reproducibly increased the invasiveness of the Panc-1 cells. Like irradiation, treatment of Panc-1 cells with transforming growth factor (TGF)-Beta1 led to a 2-fold increase in RECK protein levels. Transient transfection with Smad3 also increased RECK protein levels, but transfection with Smad7 markedly reduced them. Stable expression of Smad7 and treatment with SB431542, an inhibitor of TGF-Beta receptor I kinase, abolished TGF-Beta1- and radiation-mediated effects on RECK. Furthermore, irradiation increased levels of phosphorylated Smad3. We conclude that radiation post-transciptionally enhances RECK protein levels in Panc-1 cells, at least in part, via TGF-Beta signaling, and that irradiation increases Panc-1 invasiveness via a mechanism that may not be linked to MMP-2 activity.     

Docosahexaenoic acid reverses angiotensin II-induced RECK suppression and cardiac fibroblast migration

The omega-3 polyunsaturated fatty acids (ω − 3 fatty acids) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have been reported to inhibit or delay the progression of cardiovascular diseases, including myocardial fibrosis. Recently we reported that angiotensin II (Ang II) promotes cardiac fibroblast (CF) migration by suppressing the MMP regulator reversion-inducing-cysteine-rich protein with Kazal motifs (RECK), through a mechanism dependent on AT1, ERK, and Sp1. Here we investigated the role of miR-21 in Ang II-mediated RECK suppression, and determined whether the ω − 3 fatty acids reverse these effects. Ang II induced miR-21 expression in primary mouse cardiac fibroblasts (CFs) via ERK-dependent AP-1 and STAT3 activation, and while a miR-21 inhibitor reversed Ang II-induced RECK suppression, a miR-21 mimic inhibited both RECK expression and Ang II-induced CF migration. Moreover, Ang II suppressed the pro-apoptotic PTEN, and the ERK negative regulator Sprouty homologue 1 (SPRY1), but induced the metalloendopeptidase MMP2, all in a manner that was miR-21-dependent. Further, forced expression of PTEN inhibited Akt phosphorylation, Sp1 activation, and MMP2 induction. Notably, while both EPA and DHA reversed Ang II-mediated RECK suppression, DHA appeared to be more effective, and reversed Ang II-induced miR-21 expression, RECK suppression, MMP2 induction, and CF migration. These results indicate that Ang II-induced CF migration is differentially regulated by miR-21-mediated MMP induction and RECK suppression, and that DHA has the potential to upregulate RECK, and therefore may exert potential beneficial effects in cardiac fibrosis.     

Elevated membrane-type matrix metalloproteinases in gliomas revealed by profiling proteases and inhibitors in human cancer cells

Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) regulate proteolysis of the extracellular matrix and other extracellular proteins, including growth factors and their receptors. The aberrant expression of these genes is common in most cancers. We profiled the RNA levels of every human MMP and TIMP in a variety of cell types (fibroblast, endothelial, hematopoietic, carcinoma, melanoma, and glioma) using quantitative PCR, with the aim of identifying novel expression patterns. Almost all members of the membrane-type (MT-) MMP and TIMP families were elevated in glioma lines compared to carcinomas. In clinical glioma specimens, there were positive correlations between glioma grade and RNA levels of MT-1, MT-2, and MT-6 MMP, TIMP-1 and TIMP-2, and for several growth factors and receptors. These findings suggest that advanced malignant gliomas have elevated levels of membrane-associated MMPs and TIMPs, which may potentially regulate vascularization and invasion. Concurrent elevation of signaling molecules suggests potential bidirectional relationships that enhance tumor aggressiveness.     

Microenvironment-induced TIMP2 loss by cancer-secreted exosomal miR-4443 promotes liver metastasis of breast cancer

We aimed to investigate the role of exosomal miR-4443 in metastasis of breast cancer (BCa). In vitro wound-healing assay and transwell invasion assay were used to investigate effect of miR-4443 on BCa cells. Animal experiments were performed to confirm its effects in vivo. miR-4443 promotes the metastasis of BCa cells through downregulating tissue inhibitors of metalloproteinase 2 (TIMP2) and upregulating matrix metalloproteinases (MMPs). Highly invasive BCa cells have a higher expression of miR-4443 in both cells and exosomes. The exosomes derived from highly invasive BCa cells mainly gather in the primary tumor and liver. In vivo, overexpression of miR-4443 in noninvasive BCa cells induces liver metastasis, accompanied with downregulated TIMP2, and upregulated MMP-2 in both the primary tumor and liver. When we armed MCF-10A exosomes with miR-4443 inhibitors to treat mice bearing high-miR-4443 tumors, exosomes accumulated in the primary tumor, and liver following the upregulation of TIMP2 and downregulation of MMP2, and the metastasis was inhibited. Highly invasive BCa cells destroy natural barriers against metastasis by delivering exosomal miR-4443 to stromal cells of the primary tumor and impairing TIMP2, consequently activating MMP; circulating exosomal miR-4443 might promote BCa cells lodging in future metastatic sites through the similar mechanisms.     

Analysis of the MMP-dependent and independent functions of tissue inhibitor of metalloproteinase-2 on the invasiveness of breast cancer cells

Matrix metalloproteinases (MMPs) are secreted endopeptidases that play an essential role in remodeling the extracellular matrix (ECM). MMPs are primarily active during development, when the majority of ECM remodeling events occurs. In adults, elevated MMP activity has been observed in many pathological conditions such as cancer and osteoarthritis. The proteolytic activity of MMPs is controlled by their natural inhibitors - the tissue inhibitor of metalloproteinases (TIMPs). In addition to blocking MMP-mediated proteolysis, TIMPs have a number of MMP-independent functions including binding to cell surface proteins thereby stimulating signaling cascades. TIMP-2, the most studied member of the family, can both inhibit and activate MMPs directly, as well as inhibit MMP activity indirectly by upregulating expression of RECK, a membrane anchored MMP regulator. While TIMP-2 has been shown to play important roles in breast cancer, we describe how the MMP-independent effects of TIMP-2 can modulate the invasiveness of MCF-7, T47D and MDA-MB-231 breast cancer cells. Using an ALA + TIMP-2 mutant which is devoid of MMP inhibition, but still capable of initiating specific cell signaling cascades, we show that TIMP-2 can differentially affect MMP activity and cellular invasiveness in both an MMP dependent and independent manner. More specifically, MMP activity and invasiveness is increased with the addition of exogenous TIMP-2 in poorly invasive cell lines whereas it is decreased in highly invasive cells lines (MDA-MB-231). Conversely, the addition of ALA + TIMP-2 resulted in decreased invasiveness regardless of cell line.     

RECK‐mediated inhibition of glioma migration and invasion

RECK is an anti‐tumoral gene whose activity has been associated with its inhibitory effects regulating MMP‐2, MMP‐9, and MT1‐MMP. RECK level decreases as gliobastoma progresses, varying from less invasive grade II gliomas to very invasive human glioblastoma multiforme (GBM). Since RECK expression and glioma invasiveness show an inverse correlation, the aim of the present study is to investigate whether RECK expression would inhibit glioma invasive behavior. We conducted this study to explore forced RECK expression in the highly invasive T98G human GBM cell line. Expression levels as well as protein levels of RECK, MMP‐2, MMP‐9, and MT1‐MMP were assessed by qPCR and immunoblotting in T98G/RECK+ cells. The invasion and migration capacity of RECK+ cells was inhibited in transwell and wound assays. Dramatic cytoskeleton modifications were observed in the T98G/RECK+ cells, when compared to control cells, such as the abundance of stress fibers (contractile actin–myosin II bundles) and alteration of lamellipodia. T98G/RECK+ cells also displayed phosphorylated focal adhesion kinase (P‐FAK) in mature focal adhesions associated with stress fibers; whereas P‐FAK in control cells was mostly associated with immature focal complexes. Interestingly, the RECK protein was predominantly localized at the leading edge of migrating cells, associated with membrane ruffles. Unexpectedly, introduced expression of RECK effectively inhibited the invasive process through rearrangement of actin filaments, promoting a decrease in migratory ability. This work has associated RECK tumor‐suppressing activity with the inhibition of motility and invasion in this GBM model, which are two glioma characteristics responsible for the inefficiency of current available treatments. J. Cell. Biochem. 110: 52–61, 2010. © 2010 Wiley‐Liss, Inc.     

Minocycline inhibits PDGF-BB-induced human aortic smooth muscle cell proliferation and migration by reversing miR-221- and -222-mediated RECK suppression

Minocycline, a tetracycline antibiotic, is known to exert vasculoprotective effects independent of its anti-bacterial properties; however the underlying molecular mechanisms are not completely understood. Reversion Inducing Cysteine Rich Protein with Kazal Motifs (RECK) is a cell surface expressed, membrane anchored protein, and its overexpression inhibits cancer cell migration. We hypothesized that minocycline inhibits platelet-derived growth factor (PDGF)-induced human aortic smooth muscle cell (SMC) proliferation and migration via RECK upregulation. Our data show that the BB homodimer of recombinant PDGF (PDGF-BB) induced SMC migration and proliferation, effects significantly blunted by pre-treatment with minocycline. Further investigations revealed that PDGF-BB induced PI3K-dependent AKT activation, ERK activation, reactive oxygen species generation, Nuclear Factor-κB and Activator Protein-1 activation, microRNA (miR)-221 and miR-222 induction, RECK suppression, and matrix metalloproteinase (MMP2 and 9) activation, effects that were reversed by minocycline. Notably, minocycline induced RECK expression dose-dependently within the therapeutic dose of 1–100 μM, and silencing RECK partially reversed the inhibitory effects of minocycline on PDGF-BB-induced MMP activation, and SMC proliferation and migration. Further, targeting MMP2 and MMP9 blunted PDGF-BB-induced SMC migration. Together, these results demonstrate that minocycline inhibits PDGF-BB-induced SMC proliferation and migration by restoring RECK, an MMP inhibitor. These results indicate that the induction of RECK is one of the mechanisms by which minocycline exerts vasculoprotective effects.     

MicroRNA-146b-5p promotes atrial fibrosis in atrial fibrillation by repressing TIMP4

Alteration of tissue inhibitors of matrix metalloproteinases (TIMP)/matrix metalloproteinases (MMP) associated with collagen upregulation has an important role in sustained atrial fibrillation (AF). The expression of miR-146b-5p, whose the targeted gene is TIMPs, is upregulated in atrial cardiomyocytes during AF. This study was to determine whether miR-146b-5p could regulate the gene expression of TIMP4 and the contribution of miRNA to atrial fibrosis in AF. Collagen synthesis was observed after miR-146b-5p transfection in human induced pluripotent stem cell-derived atrial cardiomyocytes (hiPSC-aCMs)-fibroblast co-culture cellular model in vitro. Furthermore, a myocardial infarction (MI) mouse model was used to confirm the protective effect of miR-146b-5p downregulation on atrial fibrosis. The expression level of miR-146b-5p was upregulated, while the expression level of TIMP4 was downregulated in the fibrotic atrium of canine with AF. miR-146b-5p transfection in hiPSC-aCMs-fibroblast co-culture cellular model increased collagen synthesis by regulating TIMP4/MMP9 mediated extracellular matrix proteins synthesis. The inhibition of miR-146b-5p expression reduced the phenotypes of cardiac fibrosis in the MI mouse model. Fibrotic marker MMP9, TGFB1 and COL1A1 were significantly downregulated, while TIMP4 was significantly upregulated (at both mRNA and protein levels) by miR-146b-5p inhibition in cardiomyocytes of MI heart. We concluded that collagen fibres were accumulated in extracellular space on miR-146b-5p overexpressed co-culture cellular model. Moreover, the cardiac fibrosis induced by MI was attenuated in antagomiR-146 treated mice by increasing the expression of TIMP4, which indicated that the inhibition of miR-146b-5p might become an effective therapeutic approach for preventing atrial fibrosis.     

Activation of the P2X7 receptor induces migration of glial cells by inducing cathepsin B degradation of tissue inhibitor of metalloproteinase 1

The P2X₇ receptor is an ion‐gated channel, which is activated by high extracellular concentrations of adenosine triphosphate (ATP). Activation of P2X₇ receptors has been shown to induce neuroinflammatory changes associated with several neurological conditions. The matrix metalloproteinases (MMPs) are a family of endopeptidases that have several functions including degradation of the extracellular matrix, cell migration and modulation of bioactive molecules. The actions of MMPs are prevented by a family of protease inhibitors called tissue inhibitors of metalloproteinases (TIMPs). In this study, we show that ATP‐treated glial cultures from neonatal C57BL/6 mice release and increase MMP‐9 activity, which is coupled with a decrease in release of TIMP‐1 and an increase in activated cathepsin B within the extracellular space. This process occurs independently of NLRP3‐inflammasome formation. Treatment with a P2X₇ receptor antagonist prevents ATP‐induced MMP‐9 activity, inhibition of active cathepsin B release and allows for TIMP‐1 to be released from the cell. We have shown that cathepsin B degrades TIMP‐1, and inhibition of cathepsin B allows for release of TIMP‐1 and inhibits MMP‐9 activity. We also present data that indicate that ATP or cell damage induces glial cell migration, which is inhibited by P2X₇ antagonism, depletion of MMP‐9 or inhibition of cathepsin B.     

Elevation of long non-coding RNA GAS5 and knockdown of microRNA-21 up-regulate RECK expression to enhance esophageal squamous cell carcinoma cell radio-sensitivity after radiotherapy

ObjectiveLately, lncRNAs have been proposed to function in the radio-sensitivity of tumor cells, yet the role of lncRNA GAS5 in that of esophageal squamous cell carcinoma (ESCC) has scarcely been studied. This study aims to examine GAS5's effects on ESCC cell radio-sensitivity.MethodsGAS5, miR-21 and RECK expression in radiation-sensitive and radiation-resistant ESCC tissues, and TE-1 and TE-1-R cells was determined. TE-1 and TE-1-R cells were treated with pcDNA-GAS5 or miR-21 inhibitors to figure out their roles in ESCC cell proliferation, radio-sensitivity, and apoptosis via gain- and loss-of-function experiments.ResultsWe found underexpressed GAS5 and RECK, and overexpressed miR-21 in ESCC. GAS5 elevation and miR-21 inhibition reduced viability and the colony formation ability, and enhanced the apoptosis of ESCC cells under radiation.ConclusionOur study reveals that GAS5 elevation up-regulates RECK expression by down-regulating miR-21 to increase ESCC cell apoptosis after radiation therapy, thus enhancing cell radio-sensitivity.     

Targeting miR-21 sensitizes Ph+ ALL Sup-b15 cells to imatinib-induced apoptosis through upregulation of PTEN

Philadelphia chromosome positive (Ph+) acute lymphoblastic leukemia (ALL) cells are insensitive to BCR-ABL tyrosine kinase inhibitor imatinib, the underlying mechanisms remain largely unknown. Here, we showed that imatinib treatment induced significant upregulation of miR-21 and downregulation of PTEN in Ph+ ALL cell line Sup-b15. Transient inhibition of miR-21 resulted in increased apoptosis, PTEN upregulation and AKT dephosphorylation, whereas ectopic overexpression of miR-21 further conferred imatinib resistance. Furthermore, knockdown of PTEN protected the cells from imatinib-induced apoptosis achieved by inhibition of miR-21. Additionally, PI3K inhibitors also notably enhanced the effects of imatinib on Sup-b15 cells and primary Ph+ ALL cells similar to miR-21 inhibitor. Therefore, miR-21 contributes to imatinib resistance in Ph+ ALL cells and antagonizing miR-21 demonstrates therapeutic potential by sensitizing the malignancy to imatinib therapy.     

Atrial extracellular matrix remodelling in patients with atrial fibrillation

Atrial fibrillation (AF) is the most frequent clinical arrhythmia. Atrial fibrosis is an important factor in initiating and maintaining AF. However, the collagen turnover and its regulation in AF has not been completely elucidated. We tested the hypothesis that the extracellular matrix changes are more severe in patients with permanent AF in comparison with those in patients in sinus rhythm (SR). Intraoperative biopsies from the right atrial appendages (RAA) and free walls (RFW) from 24 patients with AF undergoing a mini-Maze procedure and 24 patients in SR were investigated with qualitative and quantitative immunofluorescent and Western blot analyses. As compared with SR, all patients with AF exhibited dysregulations in collagen type I and type III synthesis/degradation. Tissue inhibitors of metalloproteinases (TIMP2) was significantly enhanced only in RAA-AF. As compared with SR, collagen VI, matrix metalloproteinases MMP2, MMP9 and TIMP1 were significantly increased while TIMP3 and TIMP4 remained unchanged in all AF groups. Reversion-inducing cysteine-rich protein with Kazal motifs (RECK), a newly discovered MMPs inhibitor, was elevated in RFW as compared to RAA-AF (P<0.05) and RFW-SR (P<0.05). The level of transforming growth factor (TGF)-beta1 was higher in AF than SR. Smad2 and phosphorylated Smad2 showed an elevation in RFW-AF as compared to RFW-SR, RAA-AF, and RAA-SR groups (P<0.05). CONCLUSIONS: Atrial fibrosis in AF is characterized by severe alterations in collagen I and III synthesis/degradation associated with disturbed MMP/TIMP systems and increased levels of RECK. TGF-beta1 contributes to atrial fibrosis via TGF-beta1-Smad pathway by phosphorylating Smad2. These processes culminate in accumulations of fibrillar and non-fibrillar collagens leading to excessive atrial fibrosis and maintainance of AF.     

Mesenchymal stem cells inhibit both endogenous and exogenous MMPs via secreted TIMPs

Mesenchymal stem cells (MSCs) have been shown to be perivascular, occupying a prime location for regulating vessel stability. Here, we focused on the MSC‐contribution of key regulators of the perivascular niche, the matrix metalloproteinases (MMPs) and their inhibitors, the TIMPs. Despite secretion of active forms of MMPs by MSCs, MMP enzyme activity was not detected in MSC‐conditioned medium (MSC‐CM) due to TIMP‐mediated inhibition. By means of bifunctional‐crosslinking to probe endogenous MMP:TIMP interactions, we showed MMP‐2‐inhibition by TIMP‐2. MSCs also inhibited high levels of exogenous MMP‐2 and MMP‐9 through TIMP‐2 and TIMP‐1, respectively. Furthermore, MSC‐CM protected vascular matrix molecules and endothelial cell structures from MMP‐induced disruption. MSCs remained matrix‐protective when exposed to pro‐inflammatory cytokines and hypoxia, countering these stresses with increased TIMP‐1 expression and augmented MMP‐inhibition. Thus, MSCs are revealed as robust sources of TIMP‐mediated MMP‐inhibition, capable of protecting the perivascular niche from high levels of MMPs even under pathological conditions. J. Cell. Physiol. 226: 385–396, 2011. © 2010 Wiley‐Liss, Inc.     

A biochemical model of matrix metalloproteinase 9 activation and inhibition

Matrix metalloproteinases (MMPs) are a class of extracellular and membrane-bound proteases involved in an array of physiological processes, including angiogenesis. We present a detailed computational model of MMP9 activation and inhibition. Our model is validated to existing biochemical experimental data. We determine kinetic rate constants for the processes of MMP9 activation by MMP3, MMP10, MMP13, and trypsin; inhibition by the tissue inhibitors of metalloproteinases (TIMPs) 1 and 2; and MMP9 deactivation. This computational approach allows us to investigate discrepancies in our understanding of the interaction of MMP9 with TIMP1. Specifically, we find that inhibition due to a single binding event cannot describe MMP9 inhibition by TIMP1. Temporally accurate biphasic inhibition requires either an additional isomerization step or a second lower affinity isoform of MMP9. We also theoretically characterize the MMP3/TIMP2/pro-MMP9 and MMP3/TIMP1/pro-MMP9 systems. We speculate that these systems differ significantly in their time scales of activation and inhibition such that MMP9 is able to temporarily overshoot its final equilibrium value in the latter. Our numerical simulations suggest that the ability of pro-MMP9 to complex TIMP1 increases this overshoot. In all, our analysis serves as a summary of existing kinetic data for MMP9 and a foundation for future models utilizing MMP9 or other MMPs under physiologically well defined microenvironments.     

Tissue and serum loss of metalloproteinase inhibitors in high grade soft tissue sarcomas

The activity of matrix metalloproteinases (MMPs) in degrading extracellular matrix is controlled by activation of pro-enzymes and inhibition of MMP tissue inhibitors (TIMPs). To assess proteolytic cascade imbalance in malignancy progression, the enzymatic activity of MMP2 and MMP9 and the expression and serum level of their inhibitors, TIMP2 and TIMP1 respectively, was evaluated in selected patients with high-risk soft tissue sarcoma (STS). Gelatinase activity and inhibitor expression was evaluated on 69 biopsies by zymography and immunohistochemistry. TIMP1 and TIMP2 serum concentration was tested in 53 STS patients and in 56 controls using a sandwich enzyme immunoassay. Clinical and biological variables were related to clinical outcome of the patients. A significant gelatinolytic activity was seen in a high percentage of STS. TIMP expression was weak or negative in the majority of samples. The difference between disease-free (p=0.001) and overall survival (p=0.007) curves based on TIMP2 immunoreactivity was statistically significant. TIMP plasma concentration of 53 STS revealed significantly lower levels compared to those of 56 controls (p=0.0001). In conclusion, low levels of negative regulators of proteolysis may be related to tumor biological aggressiveness and used to select patients with poor prognosis to improve cure.     

LGI1, a putative tumor metastasis suppressor gene, controls in vitro invasiveness and expression of matrix metalloproteinases in glioma cells through the ERK1/2 pathway

Gliomas take a number of different genetic routes in the progression to glioblastoma multiforme, a highly invasive variant that is mostly unresponsive to current therapies. Gliomas express elevated levels of matrix metalloproteinases (MMPs), which have been implicated in the control of proliferation and invasion as well as neovascularization. Progressive loss of LGI1 expression has been associated with the development of high grade gliomas. We have shown previously that the forced re-expression of LGI1 in different glioma cells inhibits proliferation, invasiveness, and anchorage-independent growth in cells null for its expression. Here, using Affymetrix gene chip analysis, we show that reexpression of LGI1 in T98G cells results in the down-regulation of several MMP genes, in particular MMP1 and MMP3. LGI1 expression also results in the inhibition of ERK1/2 phosphorylation but not p38 phosphorylation. Inhibition of the MAPK pathway using the pharmacological inhibitors PD98059, U0126, and SB203580 in T98G LGI1-null cells inhibits MMP1 and MMP3 production in an ERK1/2-dependent manner. Treatment of LGI1-expressing cells with phorbol myristate acetate prevents the inhibition of MMP1/3 and restores invasiveness and ERK1/2 phosphorylation, suggesting that LGI1 acts through the ERK/MAPK pathway. Furthermore, LGI1 expression promotes phosphorylation of AKT, which leads to phosphorylation of Raf1(Ser-259), an event shown previously to negatively regulate ERK1/2 signaling. These data suggest that LGI1 plays a major role in suppressing the production of MMP1/3 through the phosphatidylinositol 3-kinase/ERK pathway. Loss of LGI1 expression, therefore, may be an important event in the progression of gliomas that leads to a more invasive phenotype in these cells.     

miR-25 promotes gastric cancer cells growth and motility by targeting RECK

Gastric cancer (GC) is the second leading cause of cancer-related death worldwide. Recently, accumulating evidence suggests that microRNAs (miRNAs) play prominent roles in tumorigenesis and metastasis. Here, we confirmed that miR-25 was significantly increased in human GC tissues and cell lines. Forced expression of miR-25 remarkably enhanced cell proliferation, migration, and invasion in GC cells, whereas inhibition of miR-25 by inhibitor caused significant suppression of proliferation and significant increase of apoptosis. Moreover, inhibition of miR-25 significantly decreased migration and invasion of GC cells. Finally, reversion-inducing-cysteine-rich protein with kazal motifs (RECK) was found to be a target of miR-25. Overexpression of RECK could significantly reverse the oncogenic effect of miR-25. Taken together, miR-25 might promote GC cells growth and motility partially by targeting RECK.     

Matrix metalloproteinases and their inhibitors,modulators of neuro-immune interactions and of pathophysiological processes in the nervous system

The matrix metalloproteinases (MMPs) belong to a growing family of Zn2+-dependent endopeptidases, secreted or membrane-bound (MT-MMP), that regulate or degrade by proteolytic cleavage protein components of the extracellular matrix, cytokines, chemokines, cell adhesion molecules and a variety of membrane receptors. MMP activity is counterbalanced by their physiological inhibitors, the tissue inhibitors of MMPs (TIMPs), a family of 4 secreted multifunctional proteins that have growth promoting activities. In physiological conditions MMP activity is tightly regulated and altered MMP regulation is associated with pathological processes including inflammation, cell proliferation, cell death and tissue remodeling. The MMP/TIMP system is involved in the development and function of cells of the immune system by promoting their differentiation, activation, migration across basement membranes and tissues. In the last years, data has accumulated indicating that the MMP/TIMP system is expressed in the nervous system where it regulates neuro-immune interactions and plays a major role in pathophysiological processes. In this review, we present recent in vivo and in vitro studies that highlight the contribution of the MMP/TIMP system to various diseases of the nervous system, involving blood brain barrier breakdown, neuroinflammation, glial reactivity, neuronal death, reactive plasticity, and to developmental and physiological processes including cell migration, axonal sprouting and neuronal plasticity. This review also alludes to the beneficial effects of synthetic MMP inhibitors in different animal models of neuropathology. In all, a further understanding of the role of MMPs and TIMPs in the nervous system should contribute to unravel mechanisms of neuronal plasticity and pathology and set the basis of new therapeutic strategies in nervous system disorders based on the development of synthetic MMP inhibitors.