Crystallographic characterization of the radixin FERM domain bound to the cytoplasmic tail of membrane-type 1 matrix metalloproteinase (MT1-MMP)

International Nuclear Information System (INIS)

Terawaki, Shin-ichi; Kitano, Ken; Aoyama, Miki; Hakoshima, Toshio

2008-01-01

The radixin FERM domain was shown to bind the MT1-MMP cytoplasmic peptide and crystals of the complex were obtained. ERM proteins play a role in the cross-linking found between plasma membranes and actin filaments. The N-terminal FERM domains of ERM proteins are responsible for membrane association through direct interaction with the cytoplasmic tails of integral membrane proteins. During cell migration and movement, membrane-type 1 matrix metalloproteinase (MT1-MMP) on plasma membranes sheds adhesion molecule CD44 in addition to degrading the extracellular matrix. Here, the interaction between the radixin FERM domain and the MT1-MMP cytoplasmic tail is reported and preliminary crystallographic characterization of crystals of the radixin FERM domain bound to the cytoplasmic tail of MT1-MMP is presented. The crystals belong to space group P6 1 22, with unit-cell parameters a = b = 122.7, c = 128.3 Å, and contain one complex in the crystallographic asymmetric unit. The diffraction data were collected to a resolution of 2.4 Å

Protein expression of MMP-2 and MT1-MMP in actinic keratosis, squamous cell carcinoma of the skin, and basal cell carcinoma.

Science.gov (United States)

de Oliveira Poswar, Fabiano; de Carvalho Fraga, Carlos Alberto; Gomes, Emisael Stênio Batista; Farias, Lucyana Conceição; Souza, Linton Wallis Figueiredo; Santos, Sérgio Henrique Souza; Gomez, Ricardo Santiago; de-Paula, Alfredo Maurício Batista; Guimarães, André Luiz Sena

2015-02-01

Squamous cell carcinoma (SCC) and basal cell carcinoma (BCC) are 2 skin neoplasms with distinct potentials to invasion and metastasis. Actinic keratosis (AK) is a precursor lesion of SCC. Immunohistochemistry was performed to evaluate the expression of MMP-2 and MT1-MMP in samples of BCC (n = 29), SCC (n = 12), and AK (n = 13). The ratio of positive cells to total cells was used to quantify the staining. Statistical significance was considered under the level P < .05. We found a higher expression of MMP-2 in tumor stroma and parenchyma of SCC as compared to BCC. The expression of this protein was also similar between SCC and its precursor actinic keratosis, and it was higher in the stroma of high-risk BCC when compared to low-risk BCC. MT1-MMP, which is an activator of MMP-2, was similarly expressed in all groups. Our results suggest that MMP-2 expression may contribute to the distinct invasive patterns seen in SCC and BCC. © The Author(s) 2014.

Targeting of Breast Cancer through MT1-MMP/Tetraspanin Complexes

Science.gov (United States)

2011-08-01

182, 765-776. Gordon- Alonso , M., Yanez-Mo, M., Barreiro, O., Alvarez, S., Munoz- Fernandez , M. A., Valenzuela- Fernandez , A. and Sanchez-Madrid, F...Diaz, R., Megias, D., Genis, L., Garcia -Grande, A., Garcia , M. A., Arroyo, A. G., and Montoya, M. C. (2007). MT1-MMP proinvasive activity is regulated by...metalloproteinases as drug targets and anti-targets for cancer therapy. Nat. Rev. Cancer 6, 227–239. Penas, P. F., Garcia -Diez, A., Sanchez-Madrid, F

MT1-MMP and type II collagen specify skeletal stem cells and their bone and cartilage progeny

DEFF Research Database (Denmark)

Szabova, L.; Yamada, S.S.; Wimer, H.

2009-01-01

-expressing cells of the skeleton rescues not only diminished chondrocyte proliferation, but surprisingly, also results in amelioration of the severe skeletal dysplasia associated with MT1-MMP deficiency through enhanced bone formation. Consistent with this increased bone formation, type II collagen was identified...... from nontransgenic MT1-MMP-deficient littermates. These observations show that type II collagen is not stringently confined to the chondrocyte but is expressed in skeletal stem/progenitor cells (able to regenerate bone, cartilage, myelosupportive stroma, marrow adipocytes) and in the chondrogenic...

LIM kinase1 modulates function of membrane type matrix metalloproteinase 1: implication in invasion of prostate cancer cells

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Chakrabarti Ratna

2011-01-01

Full Text Available Abstract Background LIM kinase 1 (LIMK1 is an actin and microtubule cytoskeleton modulatory protein that is overexpressed in a number of cancerous tissues and cells and also promotes invasion and metastasis of prostate and breast cancer cells. Membrane type matrix metalloproteinase 1 (MT1-MMP is a critical modulator of extracellular matrix (ECM turnover through pericellular proteolysis and thus plays crucial roles in neoplastic cell invasion and metastasis. MT1-MMP and its substrates pro-MMP-2 and pro-MMP-9 are often overexpressed in a variety of cancers including prostate cancer and the expression levels correlate with the grade of malignancy in prostate cancer cells. The purpose of this study is to determine any functional relation between LIMK1 and MT1-MMP and its implication in cell invasion. Results Our results showed that treatment with the hydroxamate inhibitor of MT1-MMP, MMP-2 and MMP-9 ilomastat inhibited LIMK1-induced invasion of benign prostate epithelial cells. Over expression of LIMK1 resulted in increased collagenolytic activity of MMP-2, and secretion of pro-MMP2 and pro-MMP-9. Cells over expressing LIMK1 also exhibited increased expression of MT1-MMP, transcriptional activation and its localization to the plasma membrane. LIMK1 physically associates with MT1-MMP and is colocalized with it to the Golgi vesicles. We also noted increased expression of both MT1-MMP and LIMK1 in prostate tumor tissues. Conclusion Our results provide new information on regulation of MT1-MMP function by LIMK1 and showed for the first time, involvement of MMPs in LIMK1 induced cell invasion.

Membrane-type-3 matrix metalloproteinase (MT3-MMP functions as a matrix composition-dependent effector of melanoma cell invasion.

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Olga Tatti

Full Text Available In primary human melanoma, the membrane-type matrix metalloproteinase, MT3-MMP, is overexpressed in the most aggressive nodular-type tumors. Unlike MT1-MMP and MT2-MMP, which promote cell invasion through basement membranes and collagen type I-rich tissues, the function of MT3-MMP in tumor progression remains unclear. Here, we demonstrate that MT3-MMP inhibits MT1-MMP-driven melanoma cell invasion in three-dimensional collagen, while yielding an altered, yet MT1-MMP-dependent, form of expansive growth behavior that phenocopies the formation of nodular cell colonies. In melanoma cell lines originating from advanced primary or metastatic lesions, endogenous MT3-MMP expression was associated with limited collagen-invasive potential. In the cell lines with highest MT3-MMP expression relative to MT1-MMP, collagen-invasive activity was increased following stable MT3-MMP gene silencing. Consistently, MT3-MMP overexpression in cells derived from less advanced superficially spreading melanoma lesions, or in the MT3-MMP knockdown cells, reduced MT1-MMP-dependent collagen invasion. Rather than altering MT1-MMP transcription, MT3-MMP interacted with MT1-MMP in membrane complexes and reduced its cell surface expression. By contrast, as a potent fibrinolytic enzyme, MT3-MMP induced efficient invasion of the cells in fibrin, a provisional matrix component frequently found at tumor-host tissue interfaces and perivascular spaces of melanoma. Since MT3-MMP was significantly upregulated in biopsies of human melanoma metastases, these results identify MT3-MMP as a matrix-dependent modifier of the invasive tumor cell functions during melanoma progression.

Development of a specific affinity-matured exosite inhibitor to MT1-MMP that efficiently inhibits tumor cell invasion in vitro and metastasis in vivo

DEFF Research Database (Denmark)

Botkjaer, Kenneth A; Kwok, Hang Fai; Terp, Mikkel G

2016-01-01

therapeutic target. Here, we report the identification of antibody fragments to MT1-MMP that potently and specifically inhibit its cell surface functions. Lead antibody clones displayed inhibitory activity towards pro-MMP-2 activation, collagen-film degradation and gelatin-film degradation, and were shown......The membrane-associated matrix metalloproteinase-14, MT1-MMP, has been implicated in pericellular proteolysis with an important role in cellular invasion of collagenous tissues. It is substantially upregulated in various cancers and rheumatoid arthritis, and has been considered as a potential...... to bind to the MT1-MMP catalytic domain outside the active site cleft, inhibiting binding to triple helical collagen. Affinity maturation using CDR3 randomization created a second generation of antibody fragments with dissociation constants down to 0.11 nM, corresponding to an improved affinity of 332...

Messenger RNA for membrane-type 2 matrix metalloproteinase, MT2-MMP, is expressed in human placenta of first trimester.

Science.gov (United States)

Bjørn, S F; Hastrup, N; Larsen, J F; Lund, L R; Pyke, C

2000-01-01

An intimately regulated cell surface activation of matrix metalloproteinases (MMPs) is believed to be of critical importance for the control of trophoblast invasion. A histological investigation of the expression and localization of three different MMPs, the membrane-type matrix metalloproteinases 1 and 2 (MT1-MMP, MT2-MMP) and matrix metalloproteinase 2 (MMP-2/gelatinase A) was performed by in situ hybridization on consecutive sections from human placentae of first trimester pregnancies. Cytokeratin immunostaining identified trophoblast cells. Both normal and tubal implantation sites were studied. We observed a high degree of coexpression of MT2-MMP, MT1-MMP and MMP-2 mRNAs in single extravillous cytotrophoblasts that had invaded the endometrium and tubal wall. Furthermore, mRNAs for all three genes were also seen in cytotrophoblasts of cell islands. In contrast to this coexpression pattern, MT2-MMP expression was absent from cell columns and decidual cells, in which signals for MT1-MMP and MMP-2 mRNAs were seen. The present data on the cellular expression of MT2-MMP mRNA in placenta extend our knowledge of the proteolytic events that take place during early pregnancy. The data suggest that MT2-MMP, capable of activating MMP-2 in vitro, is involved in the invasion of extravillous cytotrophoblast, possibly related to the physiological activation of MMP-2. Copyright 2000 Harcourt Publishers Ltd.

Vaginal Lactoferrin Modulates PGE2, MMP-9, MMP-2, and TIMP-1 Amniotic Fluid Concentrations

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Alessandro Trentini

2016-01-01

Full Text Available Inflammation plays an important role in pregnancy, and cytokine and matrix metalloproteases (MMPs imbalance has been associated with premature rupture of membranes and increased risk of preterm delivery. Previous studies have demonstrated that lactoferrin (LF, an iron-binding protein with anti-inflammatory properties, is able to decrease amniotic fluid (AF levels of IL-6. Therefore, we aimed to evaluate the effect of vaginal LF administration on amniotic fluid PGE2 level and MMP-TIMP system in women undergoing genetic amniocentesis. One hundred and eleven women were randomly divided into controls (n=57 or treated with LF 4 hours before amniocentesis (n=54. Amniotic fluid PGE2, active MMP-9 and MMP-2, and TIMP-1 and TIMP-2 concentrations were determined by commercially available assays and the values were normalized by AF creatinine concentration. PGE2, active MMP-9, and its inhibitor TIMP-1 were lower in LF-treated group than in controls (p<0.01, p<0.005, and p<0.001, resp.. Conversely, active MMP-2 (p<0.0001 and MMP-2/TIMP-2 molar ratio (p<0.001 were increased, whilst TIMP-2 was unchanged. Our data suggest that LF administration is able to modulate the inflammatory response following amniocentesis, which may counteract cytokine and prostanoid imbalance that leads to abortion. This trial is registered with Clinical Trial number NCT02695563.

Membrane type 1-matrix metalloproteinase/Akt signaling axis modulates TNF-α-induced procoagulant activity and apoptosis in endothelial cells.

Directory of Open Access Journals (Sweden)

Hiroshi Ohkawara

Full Text Available Membrane type 1-matrix metalloproteinase (MT1-MMP functions as a signaling molecule in addition to a proteolytic enzyme. Our hypothesis was that MT1-MMP cooperates with protein kinase B (Akt in tumor necrosis factor (TNF-α-induced signaling pathways of vascular responses, including tissue factor (TF procoagulant activity and endothelial apoptosis, in cultured human aortic endothelial cells (ECs. TNF-α (10 ng/mL induced a decrease in Akt phosphorylation within 60 minutes in ECs. A chemical inhibitor of MMP, TIMP-2 and selective small interfering RNA (siRNA-mediated suppression of MT1-MMP reversed TNF-α-triggered transient decrease of Akt phosphorylation within 60 minutes, suggesting that MT1-MMP may be a key regulator of Akt phosphorylation in TNF-α-stimulated ECs. In the downstream events, TNF-α increased TF antigen and activity, and suppressed the expression of thrombomodulin (TM antigen. Inhibition of Akt markedly enhanced TNF-α-induced expression of TF antigen and activity, and further reduced the expression of TM antigen. Silencing of MT1-MMP by siRNA also reversed the changed expression of TF and TM induced by TNF-α. Moreover, TNF-α induced apoptosis of ECs through Akt- and forkhead box protein O1 (FoxO1-dependent signaling pathway and nuclear factor-kB (NF-kB activation. Knockdown of MT1-MMP by siRNA reversed apoptosis of ECs by inhibiting TNF-α-induced Akt-dependent regulation of FoxO1 in TNF-α-stimulated ECs. Immunoprecipitation demonstrated that TNF-α induced the changes in the associations between the cytoplasmic fraction of MT1-MMP and Akt in ECs. In conclusion, we show new evidence that MT1-MMP/Akt signaling axis is a key modifier for TNF-α-induced signaling pathways for modulation of procoagulant activity and apoptosis of ECs.

Membrane Type 1–Matrix Metalloproteinase/Akt Signaling Axis Modulates TNF-α-Induced Procoagulant Activity and Apoptosis in Endothelial Cells

Science.gov (United States)

Ohkawara, Hiroshi; Ishibashi, Toshiyuki; Sugimoto, Koichi; Ikeda, Kazuhiko; Ogawa, Kazuei; Takeishi, Yasuchika

2014-01-01

Membrane type 1–matrix metalloproteinase (MT1-MMP) functions as a signaling molecule in addition to a proteolytic enzyme. Our hypothesis was that MT1-MMP cooperates with protein kinase B (Akt) in tumor necrosis factor (TNF)-α-induced signaling pathways of vascular responses, including tissue factor (TF) procoagulant activity and endothelial apoptosis, in cultured human aortic endothelial cells (ECs). TNF-α (10 ng/mL) induced a decrease in Akt phosphorylation within 60 minutes in ECs. A chemical inhibitor of MMP, TIMP-2 and selective small interfering RNA (siRNA)-mediated suppression of MT1-MMP reversed TNF-α-triggered transient decrease of Akt phosphorylation within 60 minutes, suggesting that MT1-MMP may be a key regulator of Akt phosphorylation in TNF-α-stimulated ECs. In the downstream events, TNF-α increased TF antigen and activity, and suppressed the expression of thrombomodulin (TM) antigen. Inhibition of Akt markedly enhanced TNF-α-induced expression of TF antigen and activity, and further reduced the expression of TM antigen. Silencing of MT1-MMP by siRNA also reversed the changed expression of TF and TM induced by TNF-α. Moreover, TNF-α induced apoptosis of ECs through Akt- and forkhead box protein O1 (FoxO1)-dependent signaling pathway and nuclear factor-kB (NF-kB) activation. Knockdown of MT1-MMP by siRNA reversed apoptosis of ECs by inhibiting TNF-α-induced Akt-dependent regulation of FoxO1 in TNF-α-stimulated ECs. Immunoprecipitation demonstrated that TNF-α induced the changes in the associations between the cytoplasmic fraction of MT1-MMP and Akt in ECs. In conclusion, we show new evidence that MT1-MMP/Akt signaling axis is a key modifier for TNF-α-induced signaling pathways for modulation of procoagulant activity and apoptosis of ECs. PMID:25162582

On the role of MT1-MMP, a matrix metalloproteinase essential to collagen remodeling, in murine molar eruption and root growth

NARCIS (Netherlands)

Beertsen, Wouter; Holmbeck, Kenn; Niehof, Anneke; Bianco, Paolo; Chrysovergis, Kaliiopi; Birkedal-Hansen, Henning; Everts, Vincent

2002-01-01

Although the connective tissues of the periodontium are subject to a high turnover rate, no conclusive evidence has yet emerged that periodontal collagen turnover is essential for the eruption of teeth or for root elongation. These processes were studied in mice deficient in MT1-MMP, a membrane type

Developmental expression of membrane type 4-matrix metalloproteinase (Mt4-mmp/Mmp17) in the mouse embryo

Science.gov (United States)

Clemente, Cristina; Montalvo, María Gregoria; Seiki, Motoharu; Arroyo, Alicia G.

2017-01-01

Matrix metalloproteinases (MMPs) constitute a large group of endoproteases that play important functions during embryonic development, tumor metastasis and angiogenesis by degrading components of the extracellular matrix. Within this family, we focused our study on Mt4-mmp (also called Mmp17) that belongs to a distinct subset that is anchored to the cell surface via a glycosylphosphatidylinositol (GPI) moiety and with the catalytic site exposed to the extracellular space. Information about its function and substrates is very limited to date, and little has been reported on its role in the developing embryo. Here, we report a detailed expression analysis of Mt4-mmp during mouse embryonic development by using a LacZ reporter transgenic mouse line. We showed that Mt4-mmp is detected from early stages of development to postnatal stages following a dynamic and restricted pattern of expression. Mt4-mmp was first detected at E8.5 limited to the intersomitic vascularization, the endocardial endothelium and the dorsal aorta. Mt4-mmpLacZ/+ cells were also observed in the neural crest cells, somites, floor plate and notochord at early stages. From E10.5, expression localized in the limb buds and persists during limb development. A strong expression in the brain begins at E12.5 and continues to postnatal stages. Specifically, staining was observed in the olfactory bulb, cerebral cortex, hippocampus, striatum, septum, dorsal thalamus and the spinal cord. In addition, LacZ-positive cells were also detected during eye development, initially at the hyaloid artery and later on located in the lens and the neural retina. Mt4-mmp expression was confirmed by quantitative RT-PCR and western blot analysis in some embryonic tissues. Our data point to distinct functions for this metalloproteinase during embryonic development, particularly during brain formation, angiogenesis and limb development. PMID:28926609

HMEC-1 adopt the mixed amoeboid-mesenchymal migration type during EndMT.

Science.gov (United States)

Kryczka, Jakub; Przygodzka, Patrycja; Bogusz, Helena; Boncela, Joanna

2017-06-01

The contribution of endothelial cells to scar and fibrotic tissue formation is undisputedly connected to their ability to undergo the endothelial-to-mesenchymal transition (EndMT) towards fibroblast phenotype-resembling cells. The migration model of fibroblasts and fibroblast-resembling cells is still not fully understood. It may be either a Rho/ROCK-independent, an integrin- and MMP-correlated ECM degradation-dependent, a mesenchymal model or Rho/ROCK-dependent, integrin adhesion- and MMP activity-independent, an amoeboid model. Here, we hypothesized that microvascular endothelial cells (HMEC-1) undergoing EndMT adopt an intermediate state of drifting migration model between the mesenchymal and amoeboid protrusive types in the early stages of fibrosis. We characterized the response of HMEC-1 to TGF-β2, a well-known mediator of EndMT within the microvasculature. We observed that TGF-β2 induces up to an intermediate mesenchymal phenotype in HMEC-1. In parallel, MMP-2 is upregulated and is responsible for most proteolytic activity. Interestingly, the migration of HMEC-1 undergoing EndMT is dependent on both ECM degradation and invadosome formation associated with MMP-2 proteolytic activity and Rho/ROCK cytoskeleton contraction. In conclusion, the transition from mesenchymal towards amoeboid movement highlights a molecular plasticity mechanism in endothelial cell migration in skin fibrosis. Copyright © 2017 Elsevier GmbH. All rights reserved.

TGF-β1 modulates the homeostasis between MMPs and MMP inhibitors through p38 MAPK and ERK1/2 in highly invasive breast cancer cells

International Nuclear Information System (INIS)

Gomes, Luciana R; Terra, Letícia F; Wailemann, Rosângela AM; Labriola, Leticia; Sogayar, Mari C

2012-01-01

38MAPK, ERK1/2 and MMP inhibitors. Altogether, our results support that TGF-β1 modulates the mRNA and protein levels of MMPs (MMP-2 and MMP-9) as much as their inhibitors (TIMP-2 and RECK). Therefore, this cytokine plays a crucial role in breast cancer progression by modulating key elements of ECM homeostasis control. Thus, although the complexity of this signaling network, TGF-β1 still remains a promising target for breast cancer treatment

3-Phosphoinositide-dependent Protein Kinase-1 (PDK1) promotes invasion and activation of matrix metalloproteinases

International Nuclear Information System (INIS)

Xie, Zhihui; Yuan, Hongyan; Yin, Yuzhi; Zeng, Xiao; Bai, Renkui; Glazer, Robert I

2006-01-01

Metastasis is a major cause of morbidity and mortality in breast cancer with tumor cell invasion playing a crucial role in the metastatic process. PDK1 is a key molecule that couples PI3K to cell proliferation and survival signals in response to growth factor receptor activation, and is oncogenic when expressed in mouse mammary epithelial cells. We now present evidence showing that PDK1-expressing cells exhibit enhanced anchorage-dependent and -independent cell growth and are highly invasive when grown on Matrigel. These properties correlate with induction of MMP-2 activity, increased MT1-MMP expression and a unique gene expression profile. Invasion assays in Matrigel, MMP-2 zymogram analysis, gene microarray analysis and mammary isografts were used to characterize the invasive and proliferative function of cells expressing PDK1. Tissue microarray analysis of human breast cancers was used to measure PDK1 expression in invasive tumors by IHC. Enhanced invasion on Matrigel in PDK1-expressing cells was accompanied by increased MMP-2 activity resulting from stabilization against proteasomal degradation. Increased MMP-2 activity was accompanied by elevated levels of MT1-MMP, which is involved in generating active MMP-2. Gene microarray analysis identified increased expression of the ECM-associated genes decorin and type I procollagen, whose gene products are substrates of MT1-MMP. Mammary fat pad isografts of PDK1-expressing cells produced invasive adenocarcinomas. Tissue microarray analysis of human invasive breast cancer indicated that PDK1pSer241 was strongly expressed in 90% of samples. These results indicate that PDK1 serves as an important effector of mammary epithelial cell growth and invasion in the transformed phenotype. PDK1 mediates its effect in part by MT1-MMP induction, which in turn activates MMP-2 and modulates the ECM proteins decorin and collagen. The presence of increased PDK1 expression in the majority of invasive breast cancers suggests its

MMP2 and MMP9 participate in S1P-induced invasion of follicular ML-1 thyroid cancer cells.

Science.gov (United States)

Kalhori, Veronica; Törnquist, Kid

2015-03-15

The bioactive lipid sphingosine-1-phosphate (S1P) has emerged as a potent inducer of cancer cell migration and invasion. Previously, we have shown that S1P induces invasion of ML-1 follicular thyroid cancer cells via S1P receptors 1 and 3 (S1P1,3). Matrix metalloproteinases (MMPs) are zinc-dependent proteolytic enzymes used by cells for degradation of the extracellular matrix during invasion and migration. In the present study, we examined the role of MMP2 and MMP9 for S1P-induced invasion of ML-1 cells, and found that S1P regulates the secretion and activity of MMP2 and MMP9 via S1P1,3. Both pharmacological inhibitors and siRNA knockdown of MMP2 and MMP9 could attenuate S1P-induced invasion. Additionally, we show that calpains and Rac1 mediate S1P-induced secretion of MMP2 and MMP9. In conclusion, MMP2 and MMP9 participate in S1P-evoked follicular ML-1 thyroid cancer cell invasion. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Extracellular Matrix Regulations of Membrane Type 1 - Matrix Metalloproteinasis (MT1-MMP) and Matrix Metalloproteinase-2 (MMP-2) in Human Breast Fibroblasts

National Research Council Canada - National Science Library

Harnandez-Barrantes, Sonia

2001-01-01

.... Thus, under certain conditions, TIMP-2 is a positive regulator of MMP activity. TIMP-4, a close homologue of TIMP-2 also binds to pro-MMP- 2 and can potentially participate in pro-MMP-2 activation...

Pericellular activation of proMMP-7 (promatrilysin-1) through interaction with CD151.

Science.gov (United States)

Shiomi, Takayuki; Inoki, Isao; Kataoka, Fumio; Ohtsuka, Takashi; Hashimoto, Gakuji; Nemori, Ryoichi; Okada, Yasunori

2005-12-01

Matrix metalloproteinase-7 (MMP-7) (also known as matrilysin-1) is secreted as a proenzyme (proMMP-7) and plays a key role in the degradation of various extracellular matrix (ECM) and non-ECM molecules after activation. To identify the binding proteins related to proMMP-7 activation, a human lung cDNA library was screened by yeast two-hybrid system using proMMP-7 as bait. We identified a candidate molecule CD151, which is a member of the transmembrane 4 superfamily. Complex formation of proMMP-7 with CD151 was demonstrated by immunoprecipitation of the molecules from CaR-1 cells, a human rectal carcinoma cell line, expressing both proMMP-7 and CD151, and CD151 stable transfectants incubated with proMMP-7. Yeast two-hybrid assays using deletion mutants of proMMP-7 and CD151 suggested an interaction between the propeptide of proMMP-7 and the COOH-terminal extracellular loop of CD151. The binding activity of (125)I-labeled proMMP-7 to CD151 on the cell membranes was shown with CD151 stable transfectants. Laser-scanning confocal microscopy demonstrated that proMMP-7 colocalizes with CD151 on the cell membranes of CD151 stable transfectants and CaR-1 cells. In situ zymography using crosslinked carboxymethylated transferrin, a substrate of MMP-7, demonstrated proteinase activity on and around CD151 stable transfectants and CaR-1 cells, while the activity was abolished by their treatment with MMP inhibitors, anti-MMP-7 antibody or anti-CD151 antibody. In human lung adenocarcinoma tissues, colocalization of MMP-7 and CD151 was demonstrated on the carcinoma cells. Metalloproteinase activity was present in these tissues and could be inhibited by antibodies to MMP-7 or CD151. These data demonstrate for the first time that proMMP-7 is captured and activated on the cell membranes through interaction with CD151, and suggest the possibility that similar to the MT1-MMP/MMP-2 system, MMP-7 is involved in the pericellular activation mechanism mediated by CD151, a crucial step in

Stimulation of Alpha7 Nicotinic Acetylcholine Receptor Attenuates Nicotine-Induced Upregulation of MMP, MCP-1, and RANTES through Modulating ERK1/2/AP-1 Signaling Pathway in RAW264.7 and MOVAS Cells

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Liping Liu

2017-01-01

Full Text Available Vagus nerve stimulation through alpha7 nicotine acetylcholine receptors (α7-nAChR signaling had been demonstrated attenuation of inflammation. This study aimed to determine whether PNU-282987, a selective α7-nAChR agonist, affected activities of matrix metalloproteinase (MMP and inflammatory cytokines in nicotine-treatment RAW264.7 and MOVAS cells and to assess the underlying molecular mechanisms. RAW264.7 and MOVAS cells were treated with nicotine at different concentrations (0, 1, 10, and 100 ng/ml for 0–120 min. Nicotine markedly stimulated the phosphorylation of extracellular signal-regulated kinase1/2 (ERK1/2 and c-Jun in RAW264.7 cells. Pretreatment with U0126 significantly suppressed phosphorylation of ERK1/2 and further attenuated nicotine-induced activation of c-Jun and upregulation of MMP-2, MMP-9, monocyte chemotactic protein- (MCP- 1, and regulated upon activation normal T cell expressed and secreted (RANTES. Similarly, nicotine treatment also increased phosphorylation of c-Jun and expressions of MMP-2, MMP-9, MCP-1, and RANTES in MOVAS cells. When cells were pretreated with PNU-282987, nicotine-induced activations of ERK1/2 and c-Jun in RAW264.7 cells and c-Jun in MOVAS cells were effectively inhibited. Furthermore, nicotine-induced secretions of MMP-2, MMP-9, MCP-1, and RANTES were remarkably downregulated. Treatment with α7-nAChR agonist inhibits nicotine-induced upregulation of MMP and inflammatory cytokines through modulating ERK1/2/AP-1 signaling in RAW264.7 cells and AP-1 in MOVAS cells, providing a new therapeutic for abdominal aortic aneurysm.

(-1607 2G/1G) and MMP-3 (-1612 5A/6A)

Indian Academy of Sciences (India)

Hari Om Singh

2017-07-04

Jul 4, 2017 ... (-1612 5A/6A) in development of HAND and modulation .... (Axygen Biosciences, Union City, CA, USA) according to ..... promoter region polymorphisms of MMP-1 and MMP-3 ... have employed the case-only study to find out the risk for ... cytokine network, which may have a role in modulation of. T able. 5.

Platelet-derived growth factor-D modulates extracellular matrix homeostasis and remodeling through TIMP-1 induction and attenuation of MMP-2 and MMP-9 gelatinase activities

Energy Technology Data Exchange (ETDEWEB)

Borkham-Kamphorst, Erawan, E-mail: ekamphorst@ukaachen.de; Alexi, Pascal; Tihaa, Lidia; Haas, Ute; Weiskirchen, Ralf, E-mail: rweiskirchen@ukaachen.de

2015-02-13

Platelet-derived growth factor-D (PDGF-D) is a more recent recognized growth factor involved in the regulation of several cellular processes, including cell proliferation, transformation, invasion, and angiogenesis by binding to and activating its cognate receptor PDGFR-β. After bile duct ligation or in the carbon tetrachloride-induced hepatic fibrosis model{sub ,} PDGF-D showed upregulation comparable to PDGF-B. Moreover, adenoviral PDGF-D gene transfer induced hepatic stellate cell proliferation and liver fibrosis. We here investigated the molecular mechanism of PDGF-D involvement in liver fibrogenesis. Therefore, the GRX mouse cell line was stimulated with PDGF-D and evaluated for fibrotic markers and PDGF-D signaling pathways in comparison to the other PDGF isoforms. We found that PDGF-D failed to enhance Col I and α-smooth muscle actin (α-SMA) production but has capacity to upregulate expression of the tissue inhibitor of metalloprotease 1 (TIMP-1) resulting in attenuation of MMP-2 and MMP-9 gelatinase activity as indicated by gelatinase zymography. This phenomenon was restored through application of a PDGF-D neutralizing antibody. Unexpectedly, PDGF-D incubation decreased both PDGFR-α and -β in mRNA and protein levels, and PDGF-D phosphorylated typrosines specific for PDGFR-α and -β. We conclude that PDGF-D intensifies fibrogenesis by interfering with the fibrolytic activity of the TIMP-1/MMP system and that PDGF-D signaling is mediated through both PDGF-α and -β receptors. - Highlights: • PDGF-D signals through PDGF receptor type α and β. • PDGF-D modulates extracellular matrix homeostasis and remodeling. • Like PDGF-B, PDGF-D triggers phosphorylation of PLC-γ, Akt/PKB, JNK, ERK1/2, and p38. • PDGF-D induces TIMP-1 expression through ERK and p38 MAPK. • PDGF-D attenuates MMP-2 and MMP-9 gelatinase activities.

Platelet-derived growth factor-D modulates extracellular matrix homeostasis and remodeling through TIMP-1 induction and attenuation of MMP-2 and MMP-9 gelatinase activities

International Nuclear Information System (INIS)

Borkham-Kamphorst, Erawan; Alexi, Pascal; Tihaa, Lidia; Haas, Ute; Weiskirchen, Ralf

2015-01-01

Platelet-derived growth factor-D (PDGF-D) is a more recent recognized growth factor involved in the regulation of several cellular processes, including cell proliferation, transformation, invasion, and angiogenesis by binding to and activating its cognate receptor PDGFR-β. After bile duct ligation or in the carbon tetrachloride-induced hepatic fibrosis model , PDGF-D showed upregulation comparable to PDGF-B. Moreover, adenoviral PDGF-D gene transfer induced hepatic stellate cell proliferation and liver fibrosis. We here investigated the molecular mechanism of PDGF-D involvement in liver fibrogenesis. Therefore, the GRX mouse cell line was stimulated with PDGF-D and evaluated for fibrotic markers and PDGF-D signaling pathways in comparison to the other PDGF isoforms. We found that PDGF-D failed to enhance Col I and α-smooth muscle actin (α-SMA) production but has capacity to upregulate expression of the tissue inhibitor of metalloprotease 1 (TIMP-1) resulting in attenuation of MMP-2 and MMP-9 gelatinase activity as indicated by gelatinase zymography. This phenomenon was restored through application of a PDGF-D neutralizing antibody. Unexpectedly, PDGF-D incubation decreased both PDGFR-α and -β in mRNA and protein levels, and PDGF-D phosphorylated typrosines specific for PDGFR-α and -β. We conclude that PDGF-D intensifies fibrogenesis by interfering with the fibrolytic activity of the TIMP-1/MMP system and that PDGF-D signaling is mediated through both PDGF-α and -β receptors. - Highlights: • PDGF-D signals through PDGF receptor type α and β. • PDGF-D modulates extracellular matrix homeostasis and remodeling. • Like PDGF-B, PDGF-D triggers phosphorylation of PLC-γ, Akt/PKB, JNK, ERK1/2, and p38. • PDGF-D induces TIMP-1 expression through ERK and p38 MAPK. • PDGF-D attenuates MMP-2 and MMP-9 gelatinase activities

The melatonin-MT1 receptor axis modulates tumor growth in PTEN-mutated gliomas.

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Ma, Huihui; Wang, Zhen; Hu, Lei; Zhang, Shangrong; Zhao, Chenggang; Yang, Haoran; Wang, Hongzhi; Fang, Zhiyou; Wu, Lijun; Chen, Xueran

2018-02-19

More than 40% of glioma patients have tumors that harbor PTEN (phosphatase and tensin homologue deleted on chromosome ten) mutations; this disease is associated with poor therapeutic resistance and outcome. Such mutations are linked to increased cell survival and growth, decreased apoptosis, and drug resistance; thus, new therapeutic strategies focusing on inhibiting glioma tumorigenesis and progression are urgently needed. Melatonin, an indolamine produced and secreted predominantly by the pineal gland, mediates a variety of physiological functions and possesses antioxidant and antitumor properties. Here, we analyzed the relationship between PTEN and the inhibitory effect of melatonin in primary human glioma cells and cultured glioma cell lines. The results showed that melatonin can inhibit glioma cell growth both in culture and in vivo. This inhibition was associated with PTEN levels, which significantly correlated with the expression level of MT1 in patients. In fact, c-fos-mediated MT1 was shown to be a key modulator of the effect of melatonin on gliomas that harbor wild type PTEN. Taken together, these data suggest that melatonin-MT1 receptor complexes represent a potential target for the treatment of glioma. Copyright © 2018 Elsevier Inc. All rights reserved.

An improved collagen zymography approach for evaluating the collagenases MMP-1, MMP-8, and MMP-13.

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Inanc, Seniz; Keles, Didem; Oktay, Gulgun

2017-10-01

Collagen zymography is an SDS-PAGE-based method for detecting both the proenzyme and active forms of collagenases. Although collagen zymography is used for assessment of the matrix metalloproteinases MMP-1 and MMP-13, it can be difficult to detect these collagenases due to technical issues. Moreover, it remains unclear whether the collagenase activity of MMP-8 can be detected by this method. Here, we present an improved collagen zymography method that allows quantification of the activities of MMP-1, MMP-8, and MMP-13. Activities of recombinant collagenases could be detected in collagen zymogram gels copolymerized with 0.3 mg/mL type I collagen extracted from rat tail tendon. This improved method is sensitive enough to detect the activity of as little as 1 ng of collagenase. We generated standard curves for the three collagenases to quantify the collagenolytic activity levels of unknown samples. To validate our improved method, we investigated MMP-1 activity levels in human thyroid cancer (8505C) and normal thyroid (Nthy-ori-3-1) cell lines, finding that the proenzyme and active MMP-1 levels were greater in 8505C cells than in Nthy-ori-3-1 cells. Taken together, our data show that collagen zymography can be used in both molecular and clinical investigations to evaluate collagenase activities in various pathological conditions.

Plasma MMP1 and MMP8 expression in breast cancer: Protective role of MMP8 against lymph node metastasis

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Christiaens Marie-Rose

2008-03-01

Full Text Available Abstract Background Elevated levels of matrix metalloproteinases have been found to associate with poor prognosis in various carcinomas. This study aimed at evaluating plasma levels of MMP1, MMP8 and MMP13 as diagnostic and prognostic markers of breast cancer. Methods A total of 208 breast cancer patients, of which 21 with inflammatory breast cancer, and 42 healthy controls were included. Plasma MMP1, MMP8 and MMP13 levels were measured using ELISA and correlated with clinicopathological characteristics. Results Median plasma MMP1 levels were higher in controls than in breast cancer patients (3.45 vs. 2.01 ng/ml, while no difference was found for MMP8 (10.74 vs. 10.49 ng/ml. ROC analysis for MMP1 revealed an AUC of 0.67, sensitivity of 80% and specificity of 24% at a cut-off value of 4.24 ng/ml. Plasma MMP13 expression could not be detected. No correlation was found between MMP1 and MMP8 levels. We found a trend of lower MMP1 levels with increasing tumour size (p = 0.07; and higher MMP8 levels with premenopausal status (p = 0.06 and NPI (p = 0.04. The median plasma MMP1 (p = 0.02 and MMP8 (p = 0.007 levels in the non-inflammatory breast cancer patients were almost twice as high as those found in the inflammatory breast cancer patients. Intriguingly, plasma MMP8 levels were positively associated with lymph node involvement but showed a negative correlation with the risk of distant metastasis. Both controls and lymph node negative patients (pN0 had lower MMP8 levels than patients with moderate lymph node involvement (pN1, pN2 (p = 0.001; and showed a trend for higher MMP8 levels compared to patients with extensive lymph node involvement (pN3 and a strong predisposition to distant metastasis (p = 0.11. Based on the hypothesis that blood and tissue protein levels are in reverse association, these results suggest that MMP8 in the tumour may have a protective effect against lymph node metastasis. Conclusion In summary, we observed differences in MMP1

Receptor for advanced glycation end products - membrane type1 matrix metalloproteinase axis regulates tissue factor expression via RhoA and Rac1 activation in high-mobility group box-1 stimulated endothelial cells.

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Koichi Sugimoto

Full Text Available BACKGROUND: Atherosclerosis is understood to be a blood vessel inflammation. High-mobility group box-1 (HMGB-1 plays a key role in the systemic inflammation. Tissue factor (TF is known to lead to inflammation which promotes thrombus formation. Membrane type1 matrix metalloprotease (MT1-MMP associates with advanced glycation endproducts (AGE triggered-TF protein expression and phosphorylation of NF-κB. However, it is still unclear about the correlation of MT1-MMP and HMBG-1-mediated TF expression. In this study, we investigated the molecular mechanisms of TF expression in response to HMGB-1 stimulation and the involvement of MT1-MMP in endothelial cells. METHODS AND RESULTS: Pull-down assays and Western blotting revealed that HMGB-1 induced RhoA/Rac1 activation and NF-kB phosphorylation in cultured human aortic endothelial cells. HMGB-1 increased the activity of MT1-MMP, and inhibition of RAGE or MT1-MMP by siRNA suppressed HMGB-1-induced TF upregulation as well as HMGB-1-triggered RhoA/Rac1 activation and NF-kB phosphorylation. CONCLUSIONS: The present study showed that RAGE/MT1-MMP axis modified HMBG-1-mediated TF expression through RhoA and Rac1 activation and NF-κB phosphorylation in endothelial cells. These results suggested that MT1-MMP was involved in vascular inflammation and might be a good target for treating atherosclerosis.

MMP1 and MMP7 as potential peripheral blood biomarkers in idiopathic pulmonary fibrosis.

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Ivan O Rosas

2008-04-01

Full Text Available Idiopathic pulmonary fibrosis (IPF is a chronic progressive fibrotic lung disease associated with substantial morbidity and mortality. The objective of this study was to determine whether there is a peripheral blood protein signature in IPF and whether components of this signature may serve as biomarkers for disease presence and progression.We analyzed the concentrations of 49 proteins in the plasma of 74 patients with IPF and in the plasma of 53 control individuals. We identified a combinatorial signature of five proteins-MMP7, MMP1, MMP8, IGFBP1, and TNFRSF1A-that was sufficient to distinguish patients from controls with a sensitivity of 98.6% (95% confidence interval [CI] 92.7%-100% and specificity of 98.1% (95% CI 89.9%-100%. Increases in MMP1 and MMP7 were also observed in lung tissue and bronchoalveolar lavage fluid obtained from IPF patients. MMP7 and MMP1 plasma concentrations were not increased in patients with chronic obstructive pulmonary disease or sarcoidosis and distinguished IPF compared to subacute/chronic hypersensitivity pneumonitis, a disease that may mimic IPF, with a sensitivity of 96.3% (95% CI 81.0%-100% and specificity of 87.2% (95% CI 72.6%-95.7%. We verified our results in an independent validation cohort composed of patients with IPF, familial pulmonary fibrosis, subclinical interstitial lung disease (ILD, as well as with control individuals. MMP7 and MMP1 concentrations were significantly higher in IPF patients compared to controls in this cohort. Furthermore, MMP7 concentrations were elevated in patients with subclinical ILD and negatively correlated with percent predicted forced vital capacity (FVC% and percent predicted carbon monoxide diffusing capacity (DLCO%.Our experiments provide the first evidence for a peripheral blood protein signature in IPF to our knowledge. The two main components of this signature, MMP7 and MMP1, are overexpressed in the lung microenvironment and distinguish IPF from other chronic lung

Altered Expression Levels of MMP1, MMP9, MMP12, TIMP1, and IL-1β as a Risk Factor for the Elevated IOP and Optic Nerve Head Damage in the Primary Open-Angle Glaucoma Patients

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Lukasz Markiewicz

2015-01-01

Full Text Available The aim of presented work was to analyze the impact of particular polymorphic changes in the promoter regions of the -1607 1G/2G MMP1, -1562 C/T MMP9, -82 A/G MMP12, -511 C/T IL-1β, and 372 T/C TIMP1 genes on their expression level in POAG patients. Blood and aqueous humor samples acquired from 50 patients with POAG and 50 control subjects were used for QPCR and protein levels analysis by ELISA. In vivo promoter activity assays were carried on HTM cells using dual luciferase assay. All studied subjects underwent ophthalmic examination, including BCVA, intraocular pressure, slit-lamp examination, gonioscopy, HRT, and OCT scans. Patients with POAG are characterized by an increased mRNA expression of MMP1, MMP9, MMP12, and IL-1β genes as compared to the control group (P<0.001. Aqueous humor acquired from patients with POAG displayed increased protein expression of MMP1, MMP9, MMP12, and IL-1β compared to the control group (P<0.001. Allele -1607 1G of MMP1 gene possesses only 42,91% of the -1607 2G allele transcriptional activity and allele -1562 C of MMP9 gene possesses only 21,86% of the -1562 T allele. Increased expression levels of metalloproteinases can be considered as a risk factor for the development of POAG.

Polymorphism of matrix metalloproteinase genes (MMP1 and MMP3) in patients with varicose veins.

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Kurzawski, M; Modrzejewski, A; Pawlik, A; Droździk, M

2009-07-01

Several risk factors for varicose veins have been identified: female gender, combined with obesity and pregnancy, occupations requiring standing for long periods, sedentary lifestyle, history of deep-vein thrombosis and family history. However, no specific gene variants related to a wide prevalence of varicosities in general population have been identified. Extracellular matrix composition, predominantly maintained by matrix metalloproteinases (MMPs), may affect the vein-wall structure, which may lead to dilation of vessels and cause varicosities. MMP-1 (tissue collagenase I) and MMP-3 (stromelysin I) expression was found to be raised in varicose veins compared with normal vessels. Therefore, a study was conducted to evaluate a potential association between MMP1 and MMP3 promoter polymorphisms and a risk of varicose veins. Genotyping for the presence of the polymorphisms -1607dupG (rs1799750) in MMP1 and -1171dupA (rs3025058) in the MMP3 promoter region was performed using PCR and restriction-fragment length polymorphism assays in a group of 109 patients diagnosed with varicose veins and 112 healthy controls. The frequencies of the MMP1 and MMP3 alleles (minor allele frequency 0.440 in patients vs. 0.451 in the controls for MMP1-1607*G and 0.514 vs. 0.469 for MMP3-1171*dupA, respectively) and of genotypes did not differ significantly between patients and controls. The MMP1-1607dupG and MMP3-1171dupA promoter polymorphisms are not valuable markers of susceptibility for varicose veins.

Tiron Inhibits UVB-Induced AP-1 Binding Sites Transcriptional Activation on MMP-1 and MMP-3 Promoters by MAPK Signaling Pathway in Human Dermal Fibroblasts.

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Jing Lu

Full Text Available Recent research found that Tiron was an effective antioxidant that could act as the intracellular reactive oxygen species (ROS scavenger or alleviate the acute toxic metal overload in vivo. In this study, we investigated the inhibitory effect of Tiron on matrix metalloproteinase (MMP-1 and MMP-3 expression in human dermal fibroblast cells. Western blot and ELISA analysis revealed that Tiron inhibited ultraviolet B (UVB-induced protein expression of MMP-1 and MMP-3. Real-time quantitative PCR confirmed that Tiron could inhibit UVB-induced mRNA expression of MMP-1 and MMP-3. Furthermore, Tiron significantly blocked UVB-induced activation of the MAPK signaling pathway and activator protein (AP-1 in the downstream of this transduction pathway in fibroblasts. Through the AP-1 binding site mutation, it was found that Tiron could inhibit AP-1-induced upregulation of MMP-1 and MMP-3 expression through blocking AP-1 binding to the AP-1 binding sites in the MMP-1 and MMP-3 promoter region. In conclusion, Tiron may be a novel antioxidant for preventing and treating skin photoaging UV-induced.

BMP2 induces PANC-1 cell invasion by MMP-2 overexpression through ROS and ERK.

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Liu, Jun; Ben, Qi-Wen; Yao, Wei-Yan; Zhang, Jian-Jun; Chen, Da-Fan; He, Xiang-Yi; Li, Lei; Yuan, Yao-Zong

2012-06-01

The emerging roles of bone morphogenetic proteins (BMPs) in the initiation and progression of multiple cancers have drawn great attention in cancer research. We hypothesized that BMP2 promotes cancer metastasis by modulating MMP-2 secretion and activity through intracellular ROS regulation and ERK activation in human pancreatic cancer. Our data show that stimulation of PANC-1 cells with BMP2 induced MMP-2 secretion and activation, associated with decreased E-cadherin expression, resulting in epithelial-to-mesenchymal transformation (EMT) and cell invasion. Blockade of ROS by the ROS scavenger, 2-MPG, abolished cell invasion, inhibited the EMT process and decreased MMP-2 expression, suggesting ROS accumulation caused an increase in MMP-2 expression in BMP2-stimulated PANC-1 cell invasion. Furthermore, treatment of PANC-1 cells with 2-MPG or ERK inhibitor PD98059 reduced the phosphorylation of ERK, resulting in attenuation of BMP2-induced cell invasion and MMP-2 activation. Taken together, these results suggest that BMP2 induces the cell invasion of PANC-1 cells by enhancing MMP-2 secretion and acting through ROS accumulation and ERK activation.

Bone sialoprotein does not interact with pro-gelatinase A (MMP-2 or mediate MMP-2 activation

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McCulloch Christopher A

2009-04-01

Full Text Available Abstract Background A recent model for activation of the zymogen form of matrix metalloproteinase 2 (MMP-2, also known as gelatinase A has suggested that interactions between the SIBLING protein bone sialoprotein (BSP and MMP-2 leads to conformational change in MMP-2 that initiates the conversion of the pro-enzyme into a catalytically active form. This model is particularly relevant to cancer cell metastasis to bone since BSP, bound to the αvβ3 integrin through its arginine-glycine-aspartic acid motif, could recruit MMP-2 to the cell surface. Methods We critically assessed the relationship between BSP and proMMP-2 and its activation using various forms of recombinant and purified BSP and MMP-2. Gelatinase and collagenase assays, fluorescence binding assays, real-time PCR, cell culture and pull-down assays were employed to test the model. Results Studies with a fluorogenic substrate for MMP-2 showed no activation of proMMP-2 by BSP. Binding and pull-down assays demonstrated no interaction between MMP-2 and BSP. While BSP-mediated invasiveness has been shown to depend on its integrin-binding RGD sequence, analysis of proMMP-2 activation and the level of membrane type 1 (MT1-MMP in cells grown on a BSP substratum showed that the BSP-αvβ3 integrin interaction does not induce the expression of MT1-MMP. Conclusion These studies do not support a role for BSP in promoting metastasis through interactions with pro-MMP-2.

Membrane-Type 1 Matrix Metal loproteinase Is Regulated by Sp1 through the Differential Activation of AKT, JNK, and ERK Pathways in Human Prostate Tumor Cells

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Isis C. Sroka

2007-05-01

Full Text Available We and other investigators have previously shown that membrane-type 1 matrix metalloproteinase (MT1-MMP is overexpressed in invasive prostate cancer cells. However, the mechanism for this expression is not known. Here, we show that MT1-MMP is minimally expressed in nonmalignant primary prostate cells, moderately expressed in DU-145 cells, and highly expressed in invasive PC-3 and PC-3N cells. Using human MT1-MMP promoter reporter plasmids and mobility shift assays, we show that Spi regulates MT1-MMP expression in DU-145, PC-3, and PC-3N cells and in PC3-N cells using chromatin immunoprecipitation analysis and silencing RNA. Investigation of signaling pathway showed that DU-145 cells express constitutively phosphorylated extracellular stress-regulated kinase (ERK, whereas PC-3 and PC-3N cells express constitutively phosphorylated AKT/PKB and c-Jun NH2 terminal kinase (JNK. We show that MT1-MMP and Spi levels are decreased in PC-3 and PC-3N cells when phosphatidylinositol-3 kinase and JNK are inhibited, and that MT1-MMP levels are decreased in DU-145 cells when MEK is inhibited. Transient transfection of PC-3 and PC-3N cells with a dominant-negative JNK or p85, and of DU-145 cells with a dominant negative ERK, reduces MT1-MMP promoter activity. These results indicate differential signaling control of Spi-mediated transcriptional regulation of MT1-MMP in prostate cancer cell lines.

Membrane Type-1 Matrix Metalloproteinase Expression in Acute Myeloid Leukemia and Its Upregulation by Tumor Necrosis Factor-α

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Anna Janowska-Wieczorek

2012-07-01

Full Text Available Membrane type-1 matrix metalloproteinase (MT1-MMP has been implicated in tumor invasion, as well as trafficking of normal hematopoietic cells, and acts as a physiologic activator of proMMP-2. In this study we examined MT1-MMP expression in primary acute myeloid leukemia (AML cells. Because tumor necrosis factor (TNF-α is known to be elevated in AML, we also investigated the effect of TNF-α on MT1-MMP expression. We found (i MT1-MMP mRNA expression in 41 out of 43 primary AML samples tested; (ii activation of proMMP-2 in co-cultures of AML cells with normal bone marrow stromal cells; and (iii inhibition of proMMP-2 activation and trans-Matrigel migration of AML cells by gene silencing using MT1-MMP siRNA. Moreover, recombinant human TNF-α upregulated MT1-MMP expression in AML cells resulting in enhanced proMMP-2 activation and trans-Matrigel migration. Thus, AML cells express MT1-MMP and TNF-α enhances it leading to increased MMP-2 activation and most likely contributing to the invasive phenotype. We suggest that MT1-MMP, together with TNF-α, should be investigated as potential therapeutic targets in AML.

Membrane Type-1 Matrix Metalloproteinase Expression in Acute Myeloid Leukemia and Its Upregulation by Tumor Necrosis Factor-α

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Marquez-Curtis, Leah A.; Shirvaikar, Neeta; Turner, A. Robert; Mirza, Imran; Surmawala, Amir; Larratt, Loree M.; Janowska-Wieczorek, Anna

2012-01-01

Membrane type-1 matrix metalloproteinase (MT1-MMP) has been implicated in tumor invasion, as well as trafficking of normal hematopoietic cells, and acts as a physiologic activator of proMMP-2. In this study we examined MT1-MMP expression in primary acute myeloid leukemia (AML) cells. Because tumor necrosis factor (TNF)-α is known to be elevated in AML, we also investigated the effect of TNF-α on MT1-MMP expression. We found (i) MT1-MMP mRNA expression in 41 out of 43 primary AML samples tested; (ii) activation of proMMP-2 in co-cultures of AML cells with normal bone marrow stromal cells; and (iii) inhibition of proMMP-2 activation and trans-Matrigel migration of AML cells by gene silencing using MT1-MMP siRNA. Moreover, recombinant human TNF-α upregulated MT1-MMP expression in AML cells resulting in enhanced proMMP-2 activation and trans-Matrigel migration. Thus, AML cells express MT1-MMP and TNF-α enhances it leading to increased MMP-2 activation and most likely contributing to the invasive phenotype. We suggest that MT1-MMP, together with TNF-α, should be investigated as potential therapeutic targets in AML

Membrane Type-1 Matrix Metalloproteinase Expression in Acute Myeloid Leukemia and Its Upregulation by Tumor Necrosis Factor-α

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Marquez-Curtis, Leah A.; Shirvaikar, Neeta [Canadian Blood Services R& D, Edmonton, Alberta T6G 2R8 (Canada); Turner, A. Robert [Departments of Medicine and Oncology, University of Alberta, Edmonton, Alberta T6G 2G3 (Canada); Mirza, Imran [Department of Laboratory Medicine and Pathology, University of Alberta, Edmonton, Alberta T6G 2B7 (Canada); Surmawala, Amir; Larratt, Loree M. [Departments of Medicine and Oncology, University of Alberta, Edmonton, Alberta T6G 2G3 (Canada); Janowska-Wieczorek, Anna, E-mail: anna.janowska@blood.ca [Canadian Blood Services R& D, Edmonton, Alberta T6G 2R8 (Canada); Departments of Medicine and Oncology, University of Alberta, Edmonton, Alberta T6G 2G3 (Canada)

2012-07-25

Membrane type-1 matrix metalloproteinase (MT1-MMP) has been implicated in tumor invasion, as well as trafficking of normal hematopoietic cells, and acts as a physiologic activator of proMMP-2. In this study we examined MT1-MMP expression in primary acute myeloid leukemia (AML) cells. Because tumor necrosis factor (TNF)-α is known to be elevated in AML, we also investigated the effect of TNF-α on MT1-MMP expression. We found (i) MT1-MMP mRNA expression in 41 out of 43 primary AML samples tested; (ii) activation of proMMP-2 in co-cultures of AML cells with normal bone marrow stromal cells; and (iii) inhibition of proMMP-2 activation and trans-Matrigel migration of AML cells by gene silencing using MT1-MMP siRNA. Moreover, recombinant human TNF-α upregulated MT1-MMP expression in AML cells resulting in enhanced proMMP-2 activation and trans-Matrigel migration. Thus, AML cells express MT1-MMP and TNF-α enhances it leading to increased MMP-2 activation and most likely contributing to the invasive phenotype. We suggest that MT1-MMP, together with TNF-α, should be investigated as potential therapeutic targets in AML.

Three-dimensional matrix fiber alignment modulates cell migration and MT1-MMP utility by spatially and temporally directing protrusions

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Fraley, Stephanie I.; Wu, Pei-Hsun; He, Lijuan; Feng, Yunfeng; Krisnamurthy, Ranjini; Longmore, Gregory D.; Wirtz, Denis

2015-10-01

Multiple attributes of the three-dimensional (3D) extracellular matrix (ECM) have been independently implicated as regulators of cell motility, including pore size, crosslink density, structural organization, and stiffness. However, these parameters cannot be independently varied within a complex 3D ECM protein network. We present an integrated, quantitative study of these parameters across a broad range of complex matrix configurations using self-assembling 3D collagen and show how each parameter relates to the others and to cell motility. Increasing collagen density resulted in a decrease and then an increase in both pore size and fiber alignment, which both correlated significantly with cell motility but not bulk matrix stiffness within the range tested. However, using the crosslinking enzyme Transglutaminase II to alter microstructure independently of density revealed that motility is most significantly predicted by fiber alignment. Cellular protrusion rate, protrusion orientation, speed of migration, and invasion distance showed coupled biphasic responses to increasing collagen density not predicted by 2D models or by stiffness, but instead by fiber alignment. The requirement of matrix metalloproteinase (MMP) activity was also observed to depend on microstructure, and a threshold of MMP utility was identified. Our results suggest that fiber topography guides protrusions and thereby MMP activity and motility.

Identification of membrane-type 1 matrix metalloproteinase tyrosine phosphorylation in association with neuroblastoma progression

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Nyalendo, Carine; Sartelet, Hervé; Barrette, Stéphane; Ohta, Shigeru; Gingras, Denis; Béliveau, Richard

2009-01-01

Neuroblastoma is a pediatric tumor of neural crest cells that is clinically characterized by its variable evolution, from spontaneous regression to malignancy. Despite many advances in neuroblastoma research, 60% of neuroblastoma, which are essentially metastatic cases, are associated with poor clinical outcome due to the lack of effectiveness of current therapeutic strategies. Membrane-type 1 matrix metalloproteinase (MT1-MMP, MMP-14), an enzyme involved in several steps in tumor progression, has previously been shown to be associated with poor clinical outcome for neuroblastoma. Based on our recent demonstration that MT1-MMP phosphorylation is involved in the growth of fibrosarcoma tumors, we examined the potential role of phosphorylated MT1-MMP in neuroblastoma progression. Tyrosine phosphorylated MT1-MMP was immunostained on tissue microarray samples from 55 patients with neuroblastoma detected by mass screening (known to be predominantly associated with favourable outcome), and from 234 patients with standard diagnosed neuroblastoma. In addition, the effects of a non phosphorylable version of MT1-MMP on neuroblastoma cell migration and proliferation were investigated within three-dimensional collagen matrices. Although there is no correlation between the extent of tyrosine phosphorylation of MT1-MMP (pMT1-MMP) and MYCN amplification or clinical stage, we observed greater phosphorylation of pMT1-MMP in standard neuroblastoma, while it is less evident in neuroblastoma from mass screening samples (P = 0.0006) or in neuroblastoma samples from patients younger than one year (P = 0.0002). In vitro experiments showed that overexpression of a non-phosphorylable version of MT1-MMP reduced MT1-MMP-mediated neuroblastoma cell migration and proliferation within a three-dimensional type I collagen matrix, suggesting a role for the phosphorylated enzyme in the invasive properties of neuroblastoma cells. Overall, these results suggest that tyrosine phosphorylated MT1-MMP

In vivo imaging of membrane type-1 matrix metalloproteinase with a novel activatable near-infrared fluorescence probe.

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Shimizu, Yoichi; Temma, Takashi; Hara, Isao; Makino, Akira; Kondo, Naoya; Ozeki, Ei-Ichi; Ono, Masahiro; Saji, Hideo

2014-08-01

Membrane type-1 matrix metalloproteinase (MT1-MMP) is a protease activating MMP-2 that mediates cleavage of extracellular matrix components and plays pivotal roles in tumor migration, invasion and metastasis. Because in vivo noninvasive imaging of MT1-MMP would be useful for tumor diagnosis, we developed a novel near-infrared (NIR) fluorescence probe that can be activated following interaction with MT1-MMP in vivo. MT1-hIC7L is an activatable fluorescence probe comprised of anti-MT1-MMP monoclonal antibodies conjugated to self-assembling polymer micelles that encapsulate NIR dyes (IC7-1, λem : 858 nm) at concentrations sufficient to cause fluorescence self-quenching. In aqueous buffer, MT1-hIC7L fluorescence was suppressed to background levels and increased approximately 35.5-fold in the presence of detergent. Cellular uptake experiments revealed that in MT1-MMP positive C6 glioma cells, MT1-hIC7L showed significantly higher fluorescence that increased with time as compared to hIC7L, a negative control probe lacking the anti-MT1-MMP monoclonal antibody. In MT1-MMP negative MCF-7 breast adenocarcinoma cells, both MT1-hIC7L and hIC7L showed no obvious fluorescence. In addition, the fluorescence intensity of C6 cells treated with MT1-hIC7L was suppressed by pre-treatment with an MT1-MMP endocytosis inhibitor (P imaging using probes intravenously administered to tumor-bearing mice showed that MT1-hIC7L specifically visualized C6 tumors (tumor-to-background ratios: 3.8 ± 0.3 [MT1-hIC7L] vs 3.1 ± 0.2 [hIC7L] 48 h after administration, P fluorescence in MCF-7 tumors. Together, these results show that MT1-hIC7L would be a potential activatable NIR probe for specifically detecting MT1-MMP-expressing tumors. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

Expressions of Matrix Metalloproteinases (MMP-2, MMP-7, and MMP-9) and Their Inhibitors (TIMP-1, TIMP-2) in Inflammatory Bowel Diseases.

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Jakubowska, Katarzyna; Pryczynicz, Anna; Iwanowicz, Piotr; Niewiński, Andrzej; Maciorkowska, Elżbieta; Hapanowicz, Jerzy; Jagodzińska, Dorota; Kemona, Andrzej; Guzińska-Ustymowicz, Katarzyna

2016-01-01

Crohn's disease (CD) and ulcerative colitis (UC) belong to a group of inflammatory bowel diseases (IBD). The aim of our study was to evaluate the expression of MMP-2, MMP-7, MMP-9, TIMP-1, and TIMP-2 in ulcerative colitis and Crohn's disease. The study group comprised 34 patients with UC and 10 patients with CD. Evaluation of MMP-2, MMP-7, MMP-9, TIMP-1, and TIMP-2 expression in tissue samples was performed using immunohistochemistry. The overexpression of MMP-9 and TIMP-1 was dominant in both the glandular epithelium and inflammatory infiltration in UC patients. In contrast, in CD subjects the positive expression of MMP-2 and TIMP-1 was in glandular tubes while mainly MMP-7 and TIMP-2 expression was in inflammatory infiltration. Metalloproteinases' expression was associated with the presence of erosions, architectural tissue changes, and inflammatory infiltration in the lamina propria of UC patients. The expression of metalloproteinase inhibitors correlated with the presence of eosinophils and neutrophils in UC and granulomas in CD patients. Our studies indicate that the overexpression of metalloproteinases and weaker expression of their inhibitors may determine the development of IBD. It appears that MMP-2, MMP-7, and MMP-9 may be a potential therapeutic target and the use of their inhibitors may significantly reduce UC progression.

Expressions of Matrix Metalloproteinases (MMP-2, MMP-7, and MMP-9 and Their Inhibitors (TIMP-1, TIMP-2 in Inflammatory Bowel Diseases

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Katarzyna Jakubowska

2016-01-01

Full Text Available Crohn’s disease (CD and ulcerative colitis (UC belong to a group of inflammatory bowel diseases (IBD. The aim of our study was to evaluate the expression of MMP-2, MMP-7, MMP-9, TIMP-1, and TIMP-2 in ulcerative colitis and Crohn’s disease. The study group comprised 34 patients with UC and 10 patients with CD. Evaluation of MMP-2, MMP-7, MMP-9, TIMP-1, and TIMP-2 expression in tissue samples was performed using immunohistochemistry. The overexpression of MMP-9 and TIMP-1 was dominant in both the glandular epithelium and inflammatory infiltration in UC patients. In contrast, in CD subjects the positive expression of MMP-2 and TIMP-1 was in glandular tubes while mainly MMP-7 and TIMP-2 expression was in inflammatory infiltration. Metalloproteinases’ expression was associated with the presence of erosions, architectural tissue changes, and inflammatory infiltration in the lamina propria of UC patients. The expression of metalloproteinase inhibitors correlated with the presence of eosinophils and neutrophils in UC and granulomas in CD patients. Our studies indicate that the overexpression of metalloproteinases and weaker expression of their inhibitors may determine the development of IBD. It appears that MMP-2, MMP-7, and MMP-9 may be a potential therapeutic target and the use of their inhibitors may significantly reduce UC progression.

MMP-1/PAR-1 signal transduction axis and its prognostic impact in esophageal squamous cell carcinoma

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Hong-hua Peng

2012-01-01

Full Text Available The matrix metalloprotease-1 (MMP-1/protease-activated receptor-1 (PAR-1 signal transduction axis plays an important role in tumorigenesis. To explore the expression and prognostic value of MMP-1 and PAR-1 in esophageal squamous cell carcinoma (ESCC, we evaluated the expression of two proteins in resected specimens from 85 patients with ESCC by immunohistochemistry. Sixty-two (72.9% and 58 (68.2% tumors were MMP-1- and PAR-1-positive, respectively, while no significant staining was observed in normal esophageal squamous epithelium. MMP-1 and PAR-1 overexpression was significantly associated with tumor node metastasis (TNM stage and regional lymph node involvement. Patients with MMP-1- and PAR-1-positive tumors, respectively, had poorer disease-free survival (DFS than those with negative ESCC (P = 0.002 and 0.003, respectively. Univariate analysis showed a significant relationship between TNM stage [hazard ratio (HR = 2.836, 95% confidence interval (CI = 1.866-4.308], regional lymph node involvement (HR = 2.955, 95%CI = 1.713-5.068, MMP-1 expression (HR = 2.669, 95%CI = 1.229-6.127, and PAR-1 expression (HR = 1.762, 95%CI = 1.156-2.883 and DFS. Multivariate analysis including the above four parameters identified TNM stage (HR = 2.035, 95%CI = 1.167-3.681, MMP-1 expression (HR = 2.109, 95%CI = 1.293-3.279, and PAR-1 expression (HR = 1.967, 95%CI = 1.256-2.881 as independent and significant prognostic factors for DFS. Our data suggest for the first time that MMP-1 and PAR-1 were both overexpressed in ESCC and are novel predictors of poor patient prognosis after curative resection. The MMP-1/PAR-1 signal transduction axis might be a new therapeutic target for future therapies tailored against ESCC.

MMP-1/PAR-1 signal transduction axis and its prognostic impact in esophageal squamous cell carcinoma

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Peng, Hong-hua; Zhang, Xi; Cao, Pei-guo [Department of Oncology, the Third Xiangya Hospital, Central South University, Changsha, Hunan Province (China)

2011-11-18

The matrix metalloprotease-1 (MMP-1)/protease-activated receptor-1 (PAR-1) signal transduction axis plays an important role in tumorigenesis. To explore the expression and prognostic value of MMP-1 and PAR-1 in esophageal squamous cell carcinoma (ESCC), we evaluated the expression of two proteins in resected specimens from 85 patients with ESCC by immunohistochemistry. Sixty-two (72.9%) and 58 (68.2%) tumors were MMP-1- and PAR-1-positive, respectively, while no significant staining was observed in normal esophageal squamous epithelium. MMP-1 and PAR-1 overexpression was significantly associated with tumor node metastasis (TNM) stage and regional lymph node involvement. Patients with MMP-1- and PAR-1-positive tumors, respectively, had poorer disease-free survival (DFS) than those with negative ESCC (P = 0.002 and 0.003, respectively). Univariate analysis showed a significant relationship between TNM stage [hazard ratio (HR) = 2.836, 95% confidence interval (CI) = 1.866-4.308], regional lymph node involvement (HR = 2.955, 95%CI = 1.713-5.068), MMP-1 expression (HR = 2.669, 95%CI = 1.229-6.127), and PAR-1 expression (HR = 1.762, 95%CI = 1.156-2.883) and DFS. Multivariate analysis including the above four parameters identified TNM stage (HR = 2.035, 95%CI = 1.167-3.681), MMP-1 expression (HR = 2.109, 95%CI = 1.293-3.279), and PAR-1 expression (HR = 1.967, 95%CI = 1.256-2.881) as independent and significant prognostic factors for DFS. Our data suggest for the first time that MMP-1 and PAR-1 were both overexpressed in ESCC and are novel predictors of poor patient prognosis after curative resection. The MMP-1/PAR-1 signal transduction axis might be a new therapeutic target for future therapies tailored against ESCC.

Retroviral gene transfer of an antisense construct against membrane type 1 matrix metalloproteinase reduces the invasiveness of rheumatoid arthritis synovial fibroblasts.

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Rutkauskaite, Edita; Volkmer, Dagmar; Shigeyama, Yukio; Schedel, Jörg; Pap, Geza; Müller-Ladner, Ulf; Meinecke, Ingmar; Alexander, Dorothea; Gay, Renate E; Drynda, Susanne; Neumann, Wolfram; Michel, Beat A; Aicher, Wilhelm K; Gay, Steffen; Pap, Thomas

2005-07-01

Membrane type 1 matrix metalloproteinase (MT1-MMP) is expressed prominently in rheumatoid arthritis synovial fibroblasts (RASFs), but the specific contribution of MT1-MMP to fibroblast-mediated destruction of articular cartilage is incompletely understood. This study used gene transfer of an antisense expression construct to assess the effects of MT1-MMP inhibition on the invasiveness of RASFs. Retroviral gene transfer of a pLXIN vector-based antisense RNA expression construct (MT1-MMPalphaS) to MT1-MMP was used to stably transduce RASFs. Levels of MT1-MMP RNA and protein were determined by quantitative polymerase chain reaction, Western blotting, and immunocytochemistry in MT1-MMPalphaS-transduced RASFs as well as in control cells, with monitoring for 60 days. The effects of MT1-MMPalphaS on the invasiveness of RASFs were analyzed in the SCID mouse co-implantation model of RA. MT1-MMPalphaS-transduced RASFs produced high levels of antisense RNA that exceeded endogenous levels of MT1-MMP messenger RNA by 15-fold and resulted in a down-regulation of MT1-MMP at the protein level. Inhibition of MT1-MMP production was maintained for 60 days and significantly reduced the invasiveness of RASFs in the SCID mouse model. Whereas prominent invasion into cartilage by non-transduced and mock-transduced RASFs was observed (mean invasion scores 3.0 and 3.1, respectively), MT1-MMPalphaS-transduced cells showed only moderate invasiveness (mean invasion score 1.8; P < 0.05). The data demonstrate that an antisense RNA expression construct against MT1-MMP can be generated and expressed in RASFs for at least 60 days. Inhibition of MT1-MMP significantly reduces the cartilage degradation by RASFs.

Radioimmunodetection of membrane type-1 matrix metalloproteinase relevant to tumor malignancy with a pre-targeting method

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Sano, Kohei; Temma, Takashi; Kuge, Yuji; Kudo, Takashi; Kamihashi, Junko; Saji, Hideo; Zhao, Songji

2010-01-01

Since membrane type-1 matrix metalloproteinase (MT1-MMP) is exclusively expressed in tumors and is closely associated with metastasis and invasion, MT1-MMP is a potential target of radiotracers for the evaluation of tumor malignancy. In this study, we planned to visualize MT1-MMP in vivo by a two-step pre-targeting strategy using a streptavidin (SAv)-biotin system combined with anti-MT1-MMP monoclonal immunoglobulin (IgG) (anti-MT1-MMP monoclonal antibody (mAb)). Streptavidinylated anti-MT1-MMP mAb was synthesized by reacting biotinylated anti-MT1-MMP mAb with SAv. In the pre-targeting study, FM3A mouse breast carcinoma-implanted mice were injected with anti-MT1-MMP mAb-SAv, followed 72 h later with radioiodinated biotin, (3-[ 123/125 I]iodobenzoyl)norbiotinamide( 123/125 I-IBB). Biodistribution and imaging (single photon emission computed tomography (SPECT)/CT) data were collected at several time points in the 24 h period following introduction of the tracer. The comparison groups were injected with 125 I-IBB alone or with 125 I-IBB pre-targeted with negative control IgG-SAv. In the pre-targeting study for MT1-MMP, within 1 h of tracer injection, rapid tumor uptake and abrupt clearance from the blood of radioactivity (2.22, 0.87% injected dose/g at 1 h) were observed. The tumor to blood (T/B) radioactivity ratios were significantly higher than those from mice dosed with the pre-targeting negative control (p 125 I-IBB alone did not accumulate in tumors. SPECT/CT image analysis of FM3A bearing mice showed high-contrast tumor images after 3 h with minimal blood-pool activity. The present study that uses a pre-targeting method showed high T/B radioactivity ratios and clear tumor images of MT1-MMP. This imaging method may be useful for the clinical diagnosis of malignant tumors. (author)

EMMPRIN-induced MMP-2 activation cascade in human cervical squamous cell carcinoma

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Sier, Cornelis F. M.; Zuidwijk, Kim; Zijlmans, Henry J. M. A. A.; Hanemaaijer, Roeland; Mulder-Stapel, Adri A.; Prins, Frans A.; Dreef, Enno J.; Kenter, Gemma G.; Fleuren, Gert Jan; Gorter, Arko

2006-01-01

Tumor progression and recurrence of cervical cancer is associated with upregulation of matrix metalloproteinase 2 (MMP-2). We evaluated the location, origin and activity of MMP-2 in cervical squamous cell carcinomas in comparison with MT1-MMP (MMP-14), TIMP-2 and extracellular matrix

Biological activity determination of I-BSP, a potent MMP 2 inhibitor, and its 123I tracer synthesis

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Oltenfreiter, R.; Staelens, L.; Slegers, G.; Lejeune, A.; Frankenne, F.; Dierckx, R.A.

2002-01-01

Aim: Matrix metalloproteinases (MMPs) are a family of at least 18 secreted and membrane-bound zinc endopeptidases. Collectively they function in the degradation of extracellular matrix proteins and play an important role in both normal and pathological tissue remodelling. Increased MMP activity is detected in a wide range of cancers and seems to be correlated to their invasive and metastatic potential. MMPs thus seem an attractive target for both diagnostic (SPECT tracer) and therapeutic purposes. Therefore, we synthesised a 123 I-labelled MMP 2 inhibitor and evaluated it in vitro. Materials and methods: The 123 I-labelled compound was synthesised by a Cu-assisted nucleophilic non-isotopic exchange starting from Br-BSP. After reaction, the mixture was purified by HPLC. IC 50 values were obtained by in vitro enzyme assays. A 1:1 mix between non-radiolabelled inhibitor (concentration range: 300 nM - 0.05 nM) and enzymes (MMP2, cMT1-MMP, cMT3-MMP) was incubated for 15 minutes at 37 0 C. The fluorescent substrate (Mca-Pro-Leu-Gly-Leu-Dap(Dnp)-Ala-Arg-NH2) was added and the increase of fluorescence versus time, due to the hydrolysis of substrate, was measured (GEMINI-XS, λ exc = 328 nm and λ em = 393nm). Initial velocities were calculated for different concentrations of inhibitor and the IC 50 values were then determined. Results: Radiochemical yield was 30% ±3%. Radiochemical purity was >95%. IC 50 values for inhibition of MMP2, cMT3-MMP and cMT1-MMP were 0.5 nM, 7.1 nM and 16.9 nM respectively. Conclusion: 123 I-BSP was synthesised with 30% ±3% yield. After HPLC the radiochemical purity was >95%. In vitro enzyme assays of I-BSP showed an inhibition of MMP2, cMT3-MMP and cMT1-MMP in the low nM range (0.5 nM, 7.1 nM and 16.9 nM respectively). In vivo studies (biodistribution, metabolizing) in mice will be performed in the near future

MMP-1 expression has an independent prognostic value in breast cancer

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Boström, Pia; Söderström, Mirva; Vahlberg, Tero; Söderström, Karl-Ove; Roberts, Peter J; Carpén, Olli; Hirsimäki, Pirkko

2011-01-01

Breast cancer consists of a variety of tumours, which differ by their morphological features, molecular characteristics and outcome. Well-known prognostic factors, e.g. tumour grade and size, Ki-67, hormone receptor status, HER2 expression, lymph node status and patient age have been traditionally related to prognosis. Although the conventional prognostic markers are reliable in general, better markers to predict the outcome of an individual tumour are needed. Matrix metalloproteinase-1 (MMP-1) expression has been reported to inversely correlate with survival in advanced cancers. In breast cancer MMP-1 is often upregulated, especially in basal-type breast tumours. The purpose of this retrospective study was to analyse MMP-1 expression in breast cancer cells and in cancer associated stromal cells and to correlate the results with traditional prognostic factors including p53 and bcl-2, as well as to patient survival in breast cancer subtypes. Immunohistochemical analysis of MMP-1, ER, PR, Ki-67, HER2, bcl-2, p53 and CK5/6 expression was performed on 125 breast cancers. Statistical analyses were carried out using Kruskal-Wallis and Mann-Whitney -tests. In pairwise comparison Bonferroni-adjustment was applied. Correlations were calculated using Spearman rank-order correlation coefficients. Kaplan-Meier survival analyses were carried out to compare breast cancer-specific survival curves. Factors significantly associated with disease-specific survival in univariate models were included in multivariate stepwise. Positive correlations were found between tumour grade and MMP-1 expression in tumour cells and in stromal cells. P53 positivity significantly correlated with MMP-1 expression in tumour cells, whereas HER2 expression correlated with MMP-1 both in tumour cells and stromal cells. MMP-1 expression in stromal cells showed a significant association with luminal A and luminal B, HER2 overexpressing and triple-negative breast cancer subtypes. The most important finding of

IL-1beta signals through the EGF receptor and activates Egr-1 through MMP-ADAM.

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Estella Sanchez-Guerrero

Full Text Available The immediate-early gene Egr-1 controls the inducible expression of many genes implicated in the pathogenesis of a range of vascular disorders, yet our understanding of the mechanisms controlling the rapid expression of this prototypic zinc finger transcription factor is poor. Here we show that Egr-1 expression induced by IL-1beta is dependent on metalloproteinases (MMP and a disintegrin and a metalloproteinase (ADAM. Pharmacologic MMP/ADAM inhibitors and siRNA knockdown prevent IL-1beta induction of Egr-1. Further, IL-1beta activates Egr-1 via the epidermal growth factor receptor (EGFR. This is blocked by EGFR tyrosine kinase inhibition and EGFR knockdown. IL-1beta induction of Egr-1 expression is reduced in murine embryonic fibroblasts (mEFs deficient in ADAM17 despite unbiased expression of EGFR and IL-1RI in ADAM17-deficient and wild-type mEFs. Finally, we show that IL-1beta-inducible wound repair after mechanical injury requires both EGFR and MMP/ADAM. This study reports for the first time that Egr-1 induction by IL-1beta involves EGFR and MMP/ADAM-dependent EGFR phosphorylation.

Correlation Between Th1, Th2 Cells and Levels of Serum MMP-2, MMP-9 in Children with Asthma

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Xuan WANG

2015-12-01

Full Text Available Abstract Objective: To explore the correlation between Th1 and Th2 cells and the levels of serum matrix metalloproteinase-2 (MMP-2 and MMP-9 in children with asthma. Methods: A total of 89 children with asthma were divided into acute group (n=48 and chronic group (n=41 according to the course of disease, and 40 healthy children at the same term were collected as control group. The ratios of Th1 and Th2 cells as well as levels of MMP-2 and MMP-9 were compared in three groups, and the correlation between Th1 and Th2 cells and levels of MMP-2, MMP-9 was analyzed in acute group and chronic group. Results: When compared with control group, the ratios of Th1 and Th2 cells went down in both acute group and chronic group (P<0.01, while the levels of serum MMP-2 and MMP-9 up (P<0.01. The levels of serum MMP-2 and MMP-9 in acute group were dramatically higher than those in chronic group, and there was statistical significance (P<0.01. Pearson correlation analysis revealed that there was no significant correlation between Th1 and Th2 cells and MMP-2 level (r=0.148, P=0.314, r=0.299, P=0.058; r=0.183, P=0.214, r=0.289, P=0.067, whereas both Th1 and Th2 cells were negatively correlated with MMP-9 level in acute group and chronic group (r=-0.489, P=0.000, r=-0.324, P=0.039; r=-0.352, P=0.014, r=-0.357, P=0.022. Conclusion: Aberrant secretion of Th cells can not only damage the immune function of children with asthma, but also decrease the level of serum MMP-9, consequently affecting the collagen degradation and airway remodeling.

Activation of PPARs α, β/δ, and γ Impairs TGF-β1-Induced Collagens' Production and Modulates the TIMP-1/MMPs Balance in Three-Dimensional Cultured Chondrocytes

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Paul-Emile Poleni

2010-01-01

Full Text Available Background and Purpose. We investigated the potency of Peroxisome Proliferators-Activated Receptors (PPARs α, β/δ, and γ agonists to modulate Transforming Growth Factor-β1 (TGF-β1- induced collagen production or changes in Tissue Inhibitor of Matrix Metalloproteinase- (TIMP- 1/Matrix Metalloproteinase (MMP balance in rat chondrocytes embedded in alginate beads. Experimental Approach. Collagen production was evaluated by quantitative Sirius red staining, while TIMP-1 protein levels and global MMP (-1, -2, -3, -7, and -9 or specific MMP-13 activities were measured by ELISA and fluorigenic assays in culture media, respectively. Levels of mRNA for type II collagen, TIMP-1, and MMP-3 & 13 were quantified by real-time PCR. Key Results. TGF-β1 increased collagen deposition and type II collagen mRNA levels, while inducing TIMP-1 mRNA and protein expression. In contrast, it decreased global MMP or specific MMP-13 activities, while decreasing MMP-3 or MMP-13 mRNA levels. PPAR agonists reduced most of the effects of TGF-β1 on changes in collagen metabolism and TIMP-1/MMP balance in rat in a PPAR-dependent manner, excepted for Wy14643 on MMP activities. Conclusions and Implications. PPAR agonists reduce TGF-β1-modulated ECM turnover and inhibit chondrocyte activities crucial for collagen biosynthesis, and display a different inhibitory profile depending on selectivity for PPAR isotypes.

Galectin-3 facilitates cell motility in gastric cancer by up-regulating protease-activated receptor-1 (PAR-1 and matrix metalloproteinase-1 (MMP-1.

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Seok-Jun Kim

Full Text Available BACKGROUND: Galectin-3 is known to regulate cancer metastasis. However, the underlying mechanism has not been defined. Through the DNA microarray studies after galectin-3 silencing, we demonstrated here that galectin-3 plays a key role in up-regulating the expressions of protease-activated receptor-1 (PAR-1 and matrix metalloproteinase-1 (MMP-1 PAR-1 thereby promoting gastric cancer metastasis. METHODOLOGY/PRINCIPAL FINDINGS: We examined the expression levels of Galectin-3, PAR-1, and MMP-1 in gastric cancer patient tissues and also the effects of silencing these proteins with specific siRNAs and of over-expressing them using specific lenti-viral constructs. We also employed zebrafish embryo model for analysis of in vivo gastric cancer cell invasion. These studies demonstrated that: a galectin-3 silencing decreases the expression of PAR-1. b galectin-3 over-expression increases cell migration and invasion and this increase can be reversed by PAR-1 silencing, indicating that galectin-3 increases cell migration and invasion via PAR-1 up-regulation. c galectin-3 directly interacts with AP-1 transcriptional factor, and this complex binds to PAR-1 promoter and drives PAR-1 transcription. d galectin-3 also amplifies phospho-paxillin, a PAR-1 downstream target, by increasing MMP-1 expression. MMP-1 silencing blocks phospho-paxillin amplification and cell invasion caused by galectin-3 over-expression. e Silencing of either galectin-3, PAR-1 or MMP-1 significantly reduced cell migration into the vessels in zebrafish embryo model. f Galectin-3, PAR-1, and MMP-1 are highly expressed and co-localized in malignant tissues from gastric cancer patients. CONCLUSIONS/SIGNIFICANCE: Galectin-3 plays the key role of activating cell surface receptor through production of protease and boosts gastric cancer metastasis. Galectin-3 has the potential to serve as a useful pharmacological target for prevention of gastric cancer metastasis.

Irradiation-induced angiogenesis is associated with an MMP-9-miR-494-syndecan-1 regulatory loop in medulloblastoma cells.

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Asuthkar, S; Velpula, K K; Nalla, A K; Gogineni, V R; Gondi, C S; Rao, J S

2014-04-10

Matrix metalloproteinase-9 (MMP-9) represents one of the most prominent proteins associated with tumorigenesis and is a modulator of the tumor microenvironment during angiogenesis. Recently, syndecan-1 (SDC1), a transmembrane heparan sulfate-bearing proteoglycan, was also speculated to have a critical role in contributing to angiogenesis when associated with MMP-9. However, the mechanism behind their synergistic regulation is not fully understood. In the current study, we report for the first time that ionizing radiation (IR)-induced MMP-9 enhances SDC1 shedding, corroborating to tube-inducing ability of medulloblastoma (MB) cells. Furthermore, we observed that the tumor angiogenesis is associated with higher MMP-9-SDC1 interactions on both the cell surface and extracellular medium. Our results also revealed the existence of a novel regulatory mechanism where MMP-9 drives the suppression of miR-494, resulting in enhanced SDC1 shedding and angiogenesis. From the in situ hybridization analysis, we found that MMP-9-specific shRNA (shMMP-9) treatment of mouse intracranial tumors resulted in elevated expression of miR-494. This negative correlation between MMP-9 and miR-494 levels was observed to be dependent on the methylation status of a miR-494 promoter-associated CpG island region (-186 to -20), which was confirmed by bisulfite-sequencing and methylation-specific PCR (MSP) analysis. Further, validation of MMP-9 and SDC1 3'-untranslated region (3'-UTR) targets with luciferase reporter assay provided a more favorable result for miR-494-mediated regulation of SDC1 but not of MMP-9, suggesting that the 3'-UTR of SDC1 mRNA is a direct target of miR-494. Overall, our results indicate that angiogenesis induced by radiotherapy is associated with an MMP-9-miR-494-SDC1 regulatory loop and that MMP-9-SDC1 activity creates a negative feedback loop by regulating the expression of miR-494.

The CYP2E1 inhibitor DDC up-regulates MMP-1 expression in hepatic stellate cells via an ERK1/2- and Akt-dependent mechanism.

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Liu, Tianhui; Wang, Ping; Cong, Min; Xu, Youqing; Jia, Jidong; You, Hong

2013-06-05

DDC (diethyldithiocarbamate) could block collagen synthesis in HSC (hepatic stellate cells) through the inhibition of ROS (reactive oxygen species) derived from hepatocyte CYP2E1 (cytochrome P450 2E1). However, the effect of DDC on MMP-1 (matrix metalloproteinase-1), which is the main collagen degrading matrix metalloproteinase, has not been reported. In co-culture experiments, we found that DDC significantly enhanced MMP-1 expression in human HSC (LX-2) that were cultured with hepatocyte C3A cells either expressing or not expressing CYP2E1. The levels of both proenzyme and active MMP-1 enzyme were up-regulated in LX-2 cells, accompanied by elevated enzyme activity of MMP-1 and decreased collagen I, in both LX-2 cells and the culture medium. H2O2 treatment abrogated DDC-induced MMP-1 up-regulation and collagen I decrease, while catalase treatment slightly up-regulated MMP-1 expression. These data suggested that the decrease in ROS by DDC was partially responsible for the MMP-1 up-regulation. ERK1/2 (extracellular signal-regulated kinase 1/2), Akt (protein kinase B) and p38 were significantly activated by DDC. The ERK1/2 inhibitor (U0126) and Akt inhibitor (T3830) abrogated the DDC-induced MMP-1 up-regulation. In addition, a p38 inhibitor (SB203580) improved MMP-1 up-regulation through the stimulation of ERK1/2. Our data indicate that DDC significantly up-regulates the expression of MMP-1 in LX-2 cells which results in greater MMP-1 enzyme activity and decreased collagen I. The enhancement of MMP-1 expression by DDC was associated with H2O2 inhibition and coordinated regulation by the ERK1/2 and Akt pathways. These data provide some new insights into treatment strategies for hepatic fibrosis.

Gallic Acid Inhibited Matrix Invasion and AP-1/ETS-1-Mediated MMP-1 Transcription in Human Nasopharyngeal Carcinoma Cells.

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Pang, Jong-Hwei S; Yen, Jia-Hau; Wu, Hsiao-Ting; Huang, Sheng-Teng

2017-06-24

Gallic acid is a trihydroxybenzoic acid found in natural herbal plants. Gallic acid has been reported to inhibit the migration and invasive capability of various cancers. Little is known about the underlying mechanisms of invasion responsible for cancer metastasis via gallic acid. The present study was intended to investigate the anti-invasive effect of gallic acid on human nasopharyngeal carcinoma cells (NPC-BM1) and its related mechanism. Gallic acid inhibited the invasion of NPC-BM1 cells dose- and time-dependently without significant cytotoxic effect. Affymetrix oligonucleotide microarray analysis revealed matrix metalloproteinase-1 (MMP-1) as the most down-regulated gene in NPC-BM1 cells by gallic acid. The cytosolic and secreted MMP-1 levels were both found to be inhibited by gallic acid as demonstrated by western blot analysis and ELISA respectively. The mRNA expression and transcription of MMP-1 gene was also down-regulated as determined by RT/real-time PCR and promoter activity assay. The expression of two major transcription binding factors in the MMP-1 promoter, AP-1 and ETS-1, were demonstrated to be reduced by gallic acid in NPC-BM1 cells. The effect of gallic acid was associated with the inhibition of p38 MAPK signaling pathway. In addition, gallic acid enhanced the gene expression of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) which further suppressed the MMP-1 activity. These findings may be useful to develop a novel chemotherapeutic agent to inhibit the metastasis of nasopharyngeal cancer.

Mechanical stretch induces MMP-2 release and activation in lung endothelium: role of EMMPRIN.

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Haseneen, Nadia A; Vaday, Gayle G; Zucker, Stanley; Foda, Hussein D

2003-03-01

High-volume mechanical ventilation leads to ventilator-induced lung injury. This type of lung injury is accompanied by an increased release and activation of matrix metalloproteinases (MMPs). To investigate the mechanism leading to the increased MMP release, we systematically studied the effect of mechanical stretch on human microvascular endothelial cells isolated from the lung. We exposed cells grown on collagen 1 BioFlex plates to sinusoidal cyclic stretch at 0.5 Hz using the Flexercell system with 17-18% elongation of cells. After 4 days of cell stretching, conditioned media and cell lysate were collected and analyzed by gelatin, casein, and reverse zymograms as well as Western blotting. RT-PCR of mRNA extracted from stretched cells was performed. Our results show that 1) cyclic stretch led to increased release and activation of MMP-2 and MMP-1; 2) the activation of MMP-2 was accompanied by an increase in membrane type-1 MMP (MT1-MMP) and inhibited by a hydroxamic acid-derived inhibitor of MMPs (Prinomastat, AG3340); and 3) the MMP-2 release and activation were preceded by an increase in production of extracellular MMP inducer (EMMPRIN). These results suggest that cyclic mechanical stretch leads to MMP-2 activation through an MT1-MMP mechanism. EMMPRIN may play an important role in the release and activation of MMPs during lung injury.

Gm1-MMP is involved in growth and development of leaf and seed, and enhances tolerance to high temperature and humidity stress in transgenic Arabidopsis.

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Liu, Sushuang; Liu, Yanmin; Jia, Yanhong; Wei, Jiaping; Wang, Shuang; Liu, Xiaolin; Zhou, Yali; Zhu, Yajing; Gu, Weihong; Ma, Hao

2017-06-01

Matrix metalloproteinases (MMPs) are a family of zinc- and calcium-dependent endopeptidases. Gm1-MMP was found to play an important role in soybean tissue remodeling during leaf expansion. In this study, Gm1-MMP was isolated and characterized. Its encoding protein had a relatively low phylogenetic relationship with the MMPs in other plant species. Subcellular localization indicated that Gm1-MMP was a plasma membrane protein. Gm1-MMP showed higher expression levels in mature leaves, old leaves, pods, and mature seeds, as well as was involved in the development of soybean seed. Additionally, it was involved in response to high temperature and humidity (HTH) stress in R7 leaves and seeds in soybean. The analysis of promoter of Gm1-MMP suggested that the fragment from -399 to -299 was essential for its promoter activity in response to HTH stress. The overexpression of Gm1-MMP in Arabidopsis affected the growth and development of leaves, enhanced leaf and developing seed tolerance to HTH stress and improved seed vitality. The levels of hydrogen peroxide (H 2 O 2 ) and ROS in transgenic Arabidopsis seeds were lower than those in wild type seeds under HTH stress. Gm1-MMP could interact with soybean metallothionein-II (GmMT-II), which was confirmed by analysis of yeast two-hybrid assay and BiFC assays. All the results indicated that Gm1-MMP plays an important role in the growth and development of leaves and seeds as well as in tolerance to HTH stress. It will be helpful for us understanding the functions of Gm1-MMP in plant growth and development, and in response to abiotic stresses. Copyright © 2017 Elsevier B.V. All rights reserved.

Induced sputum MMP-1, -3 & -8 concentrations during treatment of tuberculosis.

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Cesar A Ugarte-Gil

Full Text Available Tuberculosis (TB destroys lung tissues and this immunopathology is mediated in part by Matrix Metalloproteinases (MMPs. There are no data on the relationship between local tissue MMPs concentrations, anti-tuberculosis therapy and sputum conversion.Induced sputum was collected from 68 TB patients and 69 controls in a cross-sectional study. MMPs concentrations were measured by Luminex array, TIMP concentrations by ELISA and were correlated with a disease severity score (TBscore. 46 TB patients were then studied longitudinally at the 2nd, 8th week and end of treatment.Sputum MMP-1,-2,-3,-8,-9 and TIMP-1 and -2 concentrations are increased in TB. Elevated MMP-1 and -3 concentrations are independently associated with higher TB severity scores (p<0.05. MMP-1, -3 and -8 concentrations decreased rapidly during treatment (p<0.05 whilst there was a transient increase in TIMP-1/2 concentrations at week 2. MMP-2, -8 and -9 and TIMP-2 concentrations were higher at TB diagnosis in patients who remain sputum culture positive at 2 weeks and MMP-3, -8 and TIMP-1 concentrations were higher in these patients at 2nd week of TB treatment.MMPs are elevated in TB patients and associate with disease severity. This matrix-degrading phenotype resolves rapidly with treatment. The MMP profile at presentation correlates with a delayed treatment response.

Increased expression of metalloproteinase-2 and -9 (MMP-2, MMP-9), tissue inhibitor of metalloproteinase-1 and -2 (TIMP-1, TIMP-2), and EMMPRIN (CD147) in multiple myeloma.

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Urbaniak-Kujda, Donata; Kapelko-Slowik, Katarzyna; Prajs, Iwona; Dybko, Jarosław; Wolowiec, Dariusz; Biernat, Monika; Slowik, Miroslaw; Kuliczkowski, Kazimierz

2016-01-01

Activity of metalloproteinases (MMP) is controlled both by specific tissue inhibitors (TIMP) and activators (extracellular matrix metalloproteinase inducer, EMMPRIN). There are few data available concerning concentration the bone marrow of MMP-2, MMP-9, TIMP-1, and TIMP-2, or EMMPRIM expression by bone marrow mesenchymal stromal cells (BMSCs) in patients with multiple myeloma (MM). We studied 40 newly diagnosed, untreated patients: 18 males and 22 females with de novo MM and 11 healthy controls. Bone marrow was collected prior to therapy. BMSCs were derived by culturing bone marrow cells on MesenCult. Protein concentrations were determined in bone marrow plasma and culture supernatants by ELISA. EMMPRIN expression by BMSCs was assessed by flow cytometry. The median concentrations of MMP-9, TIMP-1, and TIMP-2 in both marrow plasma and culture supernatants were significantly higher in MM patients than controls. EMMPRIN expression and ratios MMP-9/TIMP-1 and MMP-2/TIMP-2 were higher in MM patients, our results demonstrate that in MM patients MMP-2 and MMP-9 are secreted in higher amounts and are not balanced by inhibitors.

Stimulation of MMP-11 (stromelysin-3) expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

International Nuclear Information System (INIS)

Selvey, Saxon; Haupt, Larisa M; Thompson, Erik W; Matthaei, Klaus I; Irving, Michael G; Griffiths, Lyn R

2004-01-01

Matrix metalloproteinases (MMPs) are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14) and stromelysin-3 (MMP-11) are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs) were: a) treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b) grown on collagens I, IV and V; c) treated with fibronectin, con-A and matrigel; and d) co-cultured with a range of HBC (human breast cancer) cell lines of varied invasive and metastatic potential. Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms

Stimulation of MMP-11 (stromelysin-3 expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

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Matthaei Klaus I

2004-07-01

Full Text Available Abstract Background Matrix metalloproteinases (MMPs are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14 and stromelysin-3 (MMP-11 are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. Methods To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs were: a treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b grown on collagens I, IV and V; c treated with fibronectin, con-A and matrigel; and d co-cultured with a range of HBC (human breast cancer cell lines of varied invasive and metastatic potential. Results Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. Conclusion We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms.

Apparent suppression of MMP-9 activity by GD1a as determined by gelatin zymography.

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Hu, Dan; Tan, Xuan; Sato, Toshinori; Yamagata, Sadako; Yamagata, Tatsuya

2006-10-13

Gelatin zymography is widely used to detect and evaluate matrix metalloproteinase-9 (MMP-9) activity. MMP-9 transcription was previously shown to be negatively regulated by ganglioside GD1a [D. Hu, Z. Man, T. Xuan, P. Wang, T. Takaku, S. Hyuga, X.S. Yao, T. Sato, S. Yamagata, T. Yamagata, Ganglioside GD1a regulation of matrix metalloproteinase-9 (MMP-9) expression in mouse FBJ cell Lines: GD1a suppression of MMP-9 expression stimulated by PI3K-Akt and p38 though not by the Erk signaling pathway, 2006, submitted for publication.]. Zymography of MMP-9 of FBJ-M5 cells preincubated with GD1a indicated a greater decrease in activity than expected from mRNA suppression. Incubation of conditioned medium containing MMP-9 with GD1a caused MMP-9 activity to decrease. Examination was thus made to confirm that MMP-9 activity is actually suppressed and/or MMP-9 protein undergoes degradation by GD1a. GD1a was found to have no effect on MMP-9 activity and Western blots indicated GD1a not to diminish MMP-9 during electrophoresis under reducing conditions. GD1a appeared to mediate the binding of a portion of MMP-9 with certain molecules, with consequently greater molecular mass on the gel, to cause decrease in the activity of MMP-9 at the site where it would normally appear. Caution should be used in doing gelatin zymography since molecules other than GD1a may similarly work, causing decrease in MMP-9 activity in zymography.

Age-related modifications of type I collagen impair DDR1-induced apoptosis in non-invasive breast carcinoma cells.

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Charles, Saby; Hassan, Rammal; Kevin, Magnien; Emilie, Buache; Sylvie, Brassart-Pasco; Laurence, Van-Gulick; Pierre, Jeannesson; Erik, Maquoi; Hamid, Morjani

2018-05-07

Type I collagen and DDR1 axis has been described to decrease cell proliferation and to initiate apoptosis in non-invasive breast carcinoma in three-dimensional cell culture matrices. Moreover, MT1-MMP down-regulates these effects. Here, we address the effect of type I collagen aging and MT1-MMP expression on cell proliferation suppression and induced-apoptosis in non-invasive MCF-7 and ZR-75-1 breast carcinoma. We provide evidence for a decrease in cell growth and an increase in apoptosis in the presence of adult collagen when compared to old collagen. This effect involves a differential activation of DDR1, as evidenced by a higher DDR1 phosphorylation level in adult collagen. In adult collagen, inhibition of DDR1 expression and kinase function induced an increase in cell growth to a level similar to that observed in old collagen. The impact of aging on the sensitivity of collagen to MT1-MMP has been reported recently. We used the MT1-MMP expression strategy to verify whether, by degrading adult type I collagen, it could lead to the same phenotype observed in old collagen 3D matrix. MT1-MMP overexpression abrogated the proliferation suppression and induced-apoptosis effects only in the presence of adult collagen. This suggests that differential collagen degradation by MT1-MMP induced a structural disorganization of adult collagen and inhibits DDR1 activation. This could in turn impair DDR1-induced cell growth suppression and apoptosis. Taken together, our data suggest that modifications of collagen structural organization, due to aging, contribute to the loss of the growth suppression and induced apoptosis effect of collagen in luminal breast carcinoma. MT1-MMP-dependent degradation and aging of collagen have no additive effects on these processes.

Tumor cell invasion of collagen matrices requires coordinate lipid agonist-induced G-protein and membrane-type matrix metalloproteinase-1-dependent signaling

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Anthis Nicholas J

2006-12-01

Full Text Available Abstract Background Lysophosphatidic acid (LPA and sphingosine 1-phosphate (S1P are bioactive lipid signaling molecules implicated in tumor dissemination. Membrane-type matrix metalloproteinase 1 (MT1-MMP is a membrane-tethered collagenase thought to be involved in tumor invasion via extracellular matrix degradation. In this study, we investigated the molecular requirements for LPA- and S1P-regulated tumor cell migration in two dimensions (2D and invasion of three-dimensional (3D collagen matrices and, in particular, evaluated the role of MT1-MMP in this process. Results LPA stimulated while S1P inhibited migration of most tumor lines in Boyden chamber assays. Conversely, HT1080 fibrosarcoma cells migrated in response to both lipids. HT1080 cells also markedly invaded 3D collagen matrices (~700 μm over 48 hours in response to either lipid. siRNA targeting of LPA1 and Rac1, or S1P1, Rac1, and Cdc42 specifically inhibited LPA- or S1P-induced HT1080 invasion, respectively. Analysis of LPA-induced HT1080 motility on 2D substrates vs. 3D matrices revealed that synthetic MMP inhibitors markedly reduced the distance (~125 μm vs. ~45 μm and velocity of invasion (~0.09 μm/min vs. ~0.03 μm/min only when cells navigated 3D matrices signifying a role for MMPs exclusively in invasion. Additionally, tissue inhibitors of metalloproteinases (TIMPs-2, -3, and -4, but not TIMP-1, blocked lipid agonist-induced invasion indicating a role for membrane-type (MT-MMPs. Furthermore, MT1-MMP expression in several tumor lines directly correlated with LPA-induced invasion. HEK293s, which neither express MT1-MMP nor invade in the presence of LPA, were transfected with MT1-MMP cDNA, and subsequently invaded in response to LPA. When HT1080 cells were seeded on top of or within collagen matrices, siRNA targeting of MT1-MMP, but not other MMPs, inhibited lipid agonist-induced invasion establishing a requisite role for MT1-MMP in this process. Conclusion LPA is a

Immunohistochemical expression of TGF-β1 and MMP-9 in periapical lesions.

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Álvares, Pâmella Recco; Arruda, José Alcides Almeida de; Silva, Leorik Pereira da; Nascimento, George João Ferreira do; Silveira, Maria Fonseca da; Sobral, Ana Paula Veras

2017-07-03

The objective of this study was to evaluate the expression of matrix metalloproteinase 9 (MMP-9) and transforming growth factor beta (TGF-β1) in periapical lesion samples correlated with the intensity of the inflammatory infiltrate and thickness of the epithelial lining. Forty-five cases of periapical lesions (23 periapical granulomas and 22 radicular cysts) were subjected to morphological and immunohistochemical analyses using anti-MMP-9 and anti-TGF-β1 antibodies. The data were analyzed using the following tests: non-parametric Mann-Whitney, chi-square, Fisher's exact test and Spearman's correlation test (Pperiapical granulomas presented infiltrate grade III, in contrast with 32% of radicular cysts (Pcysts revealed the presence of atrophic epithelium in 86% of the cysts. The immunostaining of MMP-9 was score 2 in 67% of the granulomas and 77% of the cysts. Both lesions were predominantly score 1 for TGF-β1. Significant differences were confirmed between the expression scores of TGF-β1 and MMP-9 in periapical granulomas (p = 0.004) and in radicular cysts (p periapical granulomas and radicular cysts. This immunoregulatory cytokine seems more representative in asymptomatic lesions. The extracellular matrix remodeling process dependent on MMP-9 seems to be similar for both periapical granulomas and radicular cysts. TGF-β1 and MMP-9 may play an important role in the maintenance of periapical lesions.

Quantitative Evaluation of MMP-9 and TIMP-1 Promoter Methylation in Chronic Periodontitis.

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Li, Xiting; Lu, Jiaxuan; Teng, Wei; Zhao, Chuanjiang; Ye, Xiaolei

2018-03-01

In this study, we investigated the promoter DNA methylation (DNAm) status of the MMP-9 and TIMP-1 genes in patients with chronic periodontitis to evaluate disease progression. Using pyrosequencing technology, DNAm levels of MMP-9 and TIMP-1 CpG islands were measured in 88 chronic periodontitis patients and 15 healthy controls. We found a positive correlation between methylation levels of MMP-9 CpG islands and the severity of chronic periodontitis. Methylated CpG islands were also closely associated with the duration of chronic periodontitis. Moreover, female patients exhibited lower methylation levels of MMP-9 but higher methylation levels of TIMP-1 compared with male patients, and the methylation levels of TIMP-1 gradually decreased with age. The findings of gender disparity in the DNAm of MMP-9 and TIMP-1 genes provide novel insights into chronic periodontitis.

Involvement of CD147 in overexpression of MMP-2 and MMP-9 and enhancement of invasive potential of PMA-differentiated THP-1

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Tang Hao

2005-05-01

Full Text Available Abstract Background During infection and inflammation, circulating blood monocytes migrate from the intravascular compartments to the extravascular compartments, where they mature into tissue macrophages. The maturation process prepares the cells to actively participate in the inflammatory and immune responses, and many factors have been reported to be involved in the process. We found in our study that CD147 played a very important role in this process. Results By using PMA-differentiated human monocyte cells line THP-1, we found that CD147 mediated matrix metalloproteinases (MMPs expression of the leukemic THP-1 cells and thus enhanced the invasiveness of THP-1 cells. After 24 hours of PMA-induced monocyte differentiation, the mean fluorescence intensity of CD147 in differentiated THP-1 cells (289.61 ± 31.63 was higher than that of the undifferentiated THP-1 cells (205.1 ± 19.25. There was a significant increase of the levels of proMMP-2, proMMP-9 and their activated forms in the differentiated THP-1 cells. Invasion assays using reconstituted basement membrane showed a good correlation between the invasiveness of THP-1 cells and the production of MMP-2 and MMP-9. The difference in the MMPs expression and the invasive ability was significantly blocked by HAb18G/CD147 antagonistic peptide AP-9. The inhibitory rate of the secretion of proMMP-9 in the undifferentiated THP-1 cells was 45.07%. The inhibitory rate of the secretion of proMMP-9, the activated MMP-9 and proMMP-2 in the differentiated THP-1 cells was 52.90%, 53.79% and 47.80%, respectively. The inhibitory rate of invasive potential in the undifferentiated cells and the differentiated THP-1 cells was 41.82 % and 25.15%, respectively. Conclusion The results suggest that the expression of CD147 is upregulated during the differentiation of monocyte THP-1 cells to macrophage cells, and CD147 induces the secretion and activation of MMP-2 and MMP-9 and enhances the invasive ability of THP-1

TGF-{beta}1 increases invasiveness of SW1990 cells through Rac1/ROS/NF-{kappa}B/IL-6/MMP-2

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Binker, Marcelo G. [Departments of Medicine and Physiology, University of Toronto, Toronto, Ontario, Canada M5S 1A8 (Canada); CBRHC Research Center, Buenos Aires (Argentina); Binker-Cosen, Andres A. [CBRHC Research Center, Buenos Aires (Argentina); Gaisano, Herbert Y. [Departments of Medicine and Physiology, University of Toronto, Toronto, Ontario, Canada M5S 1A8 (Canada); Cosen, Rodica H. de [CBRHC Research Center, Buenos Aires (Argentina); Cosen-Binker, Laura I., E-mail: laura.cosen.binker@utoronto.ca [Departments of Medicine and Physiology, University of Toronto, Toronto, Ontario, Canada M5S 1A8 (Canada); CBRHC Research Center, Buenos Aires (Argentina)

2011-02-04

Research highlights: {yields} Rac1 mediates TGF-{beta}1-induced SW1990 invasion through MMP-2 secretion and activation. {yields} NADPH-generated ROS act downstream of Rac1 in TGF-{beta}1-challenged SW1990 cells. {yields} TGF-{beta}1-stimulated ROS activate NF-{kappa}B in SW1990 cells. {yields} NF{kappa}B-induced IL-6 release is required for secretion and activation of MMP-2 in SW1990 cells. -- Abstract: Human pancreatic cancer invasion and metastasis have been found to correlate with increased levels of active matrix metalloproteinase 2 (MMP-2). The multifunctional cytokine transforming growth factor beta 1 (TGF-{beta}1) has been shown to increase both secretion of MMP-2 and invasion by several pancreatic cancer cell types. In the present study, we investigated the signaling pathway involved in TGF-{beta}1-promoted MMP-2 secretion and invasion by human pancreatic cancer cells SW1990. Using specific inhibitors, we found that stimulation of these tumor cells with TGF-{beta}1 induced secretion and activation of the collagenase MMP-2, which was required for TGF-{beta}1-stimulated invasion. Our results also indicate that signaling events involved in TGF-{beta}1-enhanced SW1990 invasiveness comprehend activation of Rac1 followed by generation of reactive oxygen species through nicotinamide adenine dinucleotide phosphate-oxidase, activation of nuclear factor-kappa beta, release of interleukin-6, and secretion and activation of MMP-2.

TGF-β1 increases invasiveness of SW1990 cells through Rac1/ROS/NF-κB/IL-6/MMP-2

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Binker, Marcelo G.; Binker-Cosen, Andres A.; Gaisano, Herbert Y.; Cosen, Rodica H. de; Cosen-Binker, Laura I.

2011-01-01

Research highlights: → Rac1 mediates TGF-β1-induced SW1990 invasion through MMP-2 secretion and activation. → NADPH-generated ROS act downstream of Rac1 in TGF-β1-challenged SW1990 cells. → TGF-β1-stimulated ROS activate NF-κB in SW1990 cells. → NFκB-induced IL-6 release is required for secretion and activation of MMP-2 in SW1990 cells. -- Abstract: Human pancreatic cancer invasion and metastasis have been found to correlate with increased levels of active matrix metalloproteinase 2 (MMP-2). The multifunctional cytokine transforming growth factor beta 1 (TGF-β1) has been shown to increase both secretion of MMP-2 and invasion by several pancreatic cancer cell types. In the present study, we investigated the signaling pathway involved in TGF-β1-promoted MMP-2 secretion and invasion by human pancreatic cancer cells SW1990. Using specific inhibitors, we found that stimulation of these tumor cells with TGF-β1 induced secretion and activation of the collagenase MMP-2, which was required for TGF-β1-stimulated invasion. Our results also indicate that signaling events involved in TGF-β1-enhanced SW1990 invasiveness comprehend activation of Rac1 followed by generation of reactive oxygen species through nicotinamide adenine dinucleotide phosphate-oxidase, activation of nuclear factor-kappa beta, release of interleukin-6, and secretion and activation of MMP-2.

Changes of MMP-1 and collagen type Ialpha1 by UVA, UVB and IRA are differentially regulated by Trx-1.

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Buechner, Nicole; Schroeder, Peter; Jakob, Sascha; Kunze, Kerstin; Maresch, Tanja; Calles, Christian; Krutmann, Jean; Haendeler, Judith

2008-07-01

Exposure of human skin to solar radiation, which includes ultraviolet (UV) radiation (UVA and UVB) visible light and infrared radiation, induces skin aging. The effects of light have been attributed to irradiation-induced reactive oxygen species (ROS) formation, but the specific signaling pathways are not well understood. Detrimental effects of solar radiation are dermal diseases and photoaging. Exposure of cultured human dermal fibroblasts to UVA, UVB or IRA increased ROS formation in vitro. One important redox regulator is the oxidoreductase thioredoxin-1 (Trx). Trx is ubiquitously expressed and has anti-oxidative and anti-apoptotic properties. Besides its function to reduce H(2)O(2), Trx binds to and regulates transcription factors. The aim of this study was to investigate whether Trx influences the regulation of MMP-1 and collagen Ialpha1 by UVA, UVB and IRA. We irradiated human dermal fibroblasts with UVA, UVB and IRA. UVA, UVB and IRA upregulated MMP-1 expression. Trx inhibited UVA-induced MMP-1 upregulation in a NFkappaB dependent manner. UVA, UVB and IRA reduced collagen Ialpha1 expression. Incubation with Trx inhibited the effects of UVB and IRA on collagen Ialpha1 expression. In conclusion, MMP-1 and collagen Ialpha1, which play important roles in aging processes, seems to be regulated by different transcriptional mechanisms and Trx can only influence distinct signaling pathways induced by UVA, UVB and probably IRA. Thus, Trx may serve as an important contributor to an "anti-aging therapeutic cocktail".

Immunohistochemical expression of TGF-β1 and MMP-9 in periapical lesions

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Pâmella Recco ÁLVARES

2017-07-01

Full Text Available Abstract The objective of this study was to evaluate the expression of matrix metalloproteinase 9 (MMP-9 and transforming growth factor beta (TGF-β1 in periapical lesion samples correlated with the intensity of the inflammatory infiltrate and thickness of the epithelial lining. Forty-five cases of periapical lesions (23 periapical granulomas and 22 radicular cysts were subjected to morphological and immunohistochemical analyses using anti-MMP-9 and anti-TGF-β1 antibodies. The data were analyzed using the following tests: non-parametric Mann-Whitney, chi-square, Fisher’s exact test and Spearman’s correlation test (P<0.05. Analysis of inflammatory infiltrate revealed that 78% of periapical granulomas presented infiltrate grade III, in contrast with 32% of radicular cysts (P<0.001. Morphological evaluation of the epithelial thickness in radicular cysts revealed the presence of atrophic epithelium in 86% of the cysts. The immunostaining of MMP-9 was score 2 in 67% of the granulomas and 77% of the cysts. Both lesions were predominantly score 1 for TGF-β1. Significant differences were confirmed between the expression scores of TGF-β1 and MMP-9 in periapical granulomas (p = 0.004 and in radicular cysts (p < 0.001. Expression of TGF-β1 was different for periapical granulomas and radicular cysts. This immunoregulatory cytokine seems more representative in asymptomatic lesions. The extracellular matrix remodeling process dependent on MMP-9 seems to be similar for both periapical granulomas and radicular cysts. TGF-β1 and MMP-9 may play an important role in the maintenance of periapical lesions.

Expression of the MT1 Melatonin Receptor in Ovarian Cancer Cells

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Karolina Jablonska

2014-12-01

Full Text Available Ovarian cancer (OC is the leading cause of death among women with genital tract disorders. Melatonin exhibits oncostatic properties which it may effect through binding to its membrane receptor, MT1. The aim of this study was to determine the expression of MT1 in OC cells and to correlate this with clinical and pathological data. Immunohistochemistry was performed on 84 cases of OC. Normal ovarian epithelial (IOSE 364 and OC (SK-OV-3, OVCAR-3 cell lines were used to examine the MT1 expression at protein level using the western blot and immunofluorescence technique. The expression of MT1 was observed as cytoplasmic-membrane (MT1CM and membrane (MT1M reactions. A positive correlation between MT1CM and MT1M was found in all the studied cases. There were no significant differences between the expression of MT1CM, MT1M, and histological type, staging, grading, presence of residual disease, or overall survival time. Immunofluorescence showed both MT1M and MT1CM expression in all the tested cell lines. Western blot illustrated the highest protein level of MT1 in IOSE 364 and the lowest in the OVCAR-3. The results indicate the limited prognostic significance of MT1 in OC cells.

Extracellular S100A4(mts1) stimulates invasive growth of mouse endothelial cells and modulates MMP-13 matrix metalloproteinase activity

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Schmidt-Hansen, Birgitte; Ornås, Dorte; Grigorian, Mariam

2004-01-01

with the transcriptional modulation of genes involved in the proteolytic degradation of extracellular matrix (ECM). Treatment of SVEC 4-10 with the S100A4 protein leads to the transcriptional activation of collagenase 3 (MMP-13) mRNA followed by subsequent release of the protein from the cells. Beta-casein zymography...... demonstrates enhancement of proteolytic activity associated with MMP-13. This observation indicates that extracellular S100A4 stimulates the production of ECM degrading enzymes from endothelial cells, thereby stimulating the remodeling of ECM. This could explain the angiogenic and metastasis...

MMP8, MMP9 AND TIMP1 LEVELS IN GCF AND GINGIVAL TISSUE OF PATIENTS WITH GINGIVAL OVERGROWTH DURING ORTHODONTIC TREATMENT

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Petra Surlin

2012-03-01

Full Text Available Aim. Periodontal remodellng produced during dental orthodontic treatment represents a series of biologicallyactive substances, part of them playing some role in the initiation and propagation of inflammatory processes. The present study aims at demonstrating the MMP8, MMP9 and TIMP1 levels intervening in tissular periodontal remodeling produced during orthodontic treatments, accompanied by gingival overgrowth, as a reaction of the marginal periodontium to mechanical stress. Materials and Method. Selected for the study were 21 patients – 13 females and 8 males – with ages between 13 and 32 years (17.6±1.3 years affected with dento-maxillary anomalies, who received orthodontic treatment with fixed apparatus. Sampling from the gingival fluid was performed 6 times, namely: 1 hour prior to the application of the orthodontic apparatus, 4 hours after its application, again after 8 and 24 hours and then 1 and, respectively, 2 weeks later. If gingival hypertrophy was installed (HTG, the hypertrophic gingiva was removed, and an immuno-histo-chemical examination was made. The patient was weekly monitorized in the first 6 weeks – during the initial orthodontic treatment, then monthly, samples being taken over from the gingival sulcus on each visit made in the first 6 weeks. Results. MMP-9 immuno-marking was positive both at corione level and in the deep structures of the covering epithelium. The positive cells at MMP-9 evidenced different intensities at the level of each structure forming the gingival mucous membrane. In four of the cases under analysis, disorganization of the normal layering/stratification of the epithelium was evidenced, along with the presence of numerous red cells in the chorione of the mucous membrane. In such cases, immuno-marking to MMP8 showed a normal intensity, even if few positive cells, dispersed among the extravasated red cells could be observed. Immunologically, MMP8 and MMP9 obey the same pattern, registering maximum

Selenium modulates MMP2 expression through the TGFβ1/Smad signalling pathway in human umbilical vein endothelial cells and rabbits following lipid disturbance.

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Xu, Chenggui; Lu, Guihua; Li, Qinglang; Zhang, Juhong; Huang, Zhibin; Gao, Xiuren

2017-07-01

A high-fat diet is a major risk factor for coronary heart diseases. Matrix metalloprotease (MMP) expression is changed in many cardiovascular diseases. Selenium, which is an important trace element in animals, has a close relationship with cardiovascular diseases. The TGFβ1/Smad signalling pathway is ubiquitous in diverse tissues and cells, and it is also associated with the occurrence and development of cardiovascular diseases. Therefore, in this study, we aimed to determine selenium's effect on lipid metabolism, atherosclerotic plaque formation, and MMP2 expression, as well as the underlying functional mechanism. In vivo tests: 24 male New Zealand white rabbits were randomly divided into 4 groups: regular diet, high-fat diet, high-fat diet+selenium and regular diet+selenium groups. The high-fat diet induced the lipid disturbances of rabbits at week 12. Selenium supplementation lowered total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and triglyceride (TG) levels (pSelenium supplementation also suppressed MMP2 over-expression in thoracic aortas. In vitro tests: Human umbilical vein endothelial cells (HUVECs) were treated with different concentrations of selenium or ox-LDL. Ox-LDL promoted MMP2 expression by increasing TGFβ1, pSmad2, pSmad3 and Smad3 expression (pSelenium attenuated MMP2 over-expression by regulating the TGFβ1/Smad signalling pathway. Selenium suppressed high-fat diet-induced MMP2 over-expression in vivo by improving lipid metabolism. In vitro, selenium attenuated MMP2 over-expression through the TGFβ1/Smad signalling pathway. Copyright © 2017 Elsevier GmbH. All rights reserved.

Mycobacterium tuberculosis Upregulates TNF-α Expression via TLR2/ERK Signaling and Induces MMP-1 and MMP-9 Production in Human Pleural Mesothelial Cells.

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Wei-Lin Chen

Full Text Available Tumor necrosis factor (TNF-α and matrix metalloproteinases (MMPs are elevated in pleural fluids of tuberculous pleuritis (TBP where pleural mesothelial cells (PMCs conduct the first-line defense against Mycobacterium tuberculosis (MTB. However, the clinical implication of TNF-α and MMPs in TBP and the response of PMCs to MTB infection remain unclear.We measured pleural fluid levels of TNF-α and MMPs in patients with TBP (n = 18 or heart failure (n = 18 as controls. Radiological scores for initial effusion amount and residual pleural fibrosis at 6-month follow-up were assessed. In vitro human PMC experiments were performed to assess the effect of heat-killed M. tuberculosis H37Ra (MTBRa on the expression of TNF-α and MMPs.As compared with controls, the effusion levels of TNF-α, MMP-1 and MMP-9 were significantly higher and correlated positively with initial effusion amount in patients with TBP, while TNF-α and MMP-1, but not MMP-9, were positively associated with residual pleural fibrosis of TBP. Moreover, effusion levels of TNF-α had positive correlation with those of MMP-1 and MMP-9 in TBP. In cultured PMCs, MTBRa enhanced TLR2 and TLR4 expression, activated ERK signaling, and upregulated TNF-α mRNA and protein expression. Furthermore, knockdown of TLR2, but not TLR4, significantly inhibited ERK phosphorylation and TNF-α expression. Additionally, both MTBRa and TNF-α markedly induced MMP-1 and MMP-9 synthesis in human PMCs, and TNF-α neutralization substantially reduced the production of MMP-1, but not MMP-9, in response to MTBRa stimulation.MTBRa activates TLR2/ERK signalings to induce TNF-α and elicit MMP-1 and MMP-9 in human PMCs, which are associated with effusion volume and pleural fibrosis and may contribute to pathogenesis of TBP. Further investigation of manipulation of TNF-α and MMP expression in pleural mesothelium may provide new insights into the mechanisms and rational treatment strategies for TBP.

THE ROLE OF MATRIX METALLOPROTEINASE MMP-9, ITS INHIBITOR TIMP-1 AND INTERLEUKINE-1β IN PATHOGENESIS OF TRAUMATIC BRAIN INJURY

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S. V. Ziablitsev

2016-09-01

  Resume Traumatic brain injury (TBI is accompanied by high rates of morbidity and mortality in both developed and undeveloped countries that makes it one of the most actual medical and social problems. In recent years matrix metalloproteinases are in increasing interest while studying TBI pathogenesis because of their ability to increase permeability of the blood-brain barrier and to cause nervous tissue matrix reorganization. The goal of given study was to investigate the role of matrix metalloproteinase MMP-9, its inhibitor TIMP-1 and interleukin IL-1β in pathogenesis of TBI. Methods: The study was performed on 98 mature white rats. Moderate severity TBI was modeled with one blow on the cranial vault by means of free-fall­ing plummet. Control group included 30 rats. Cytokines (IL-1b, IL-6, IL-8, TNF-a, MMP-9 and TIMP-1 levels were investi­gated in animals blood by means of ELISA on 1st, 3rd, 7th, 14th and 21st days after trauma. Results and discussion: MMP-9 levels increased by only 38,2% on the 1st day, but on the 3rd day there was its marked increase to 538%. It is known that metalloproteinases are released from the cells under the influence of various factors, including cytokines. On the 1st day after trauma it was IL-1β which increased by 705% showing the highest rise among other cytokines and exceeding increase in MMP-9 levels. This might indicate regulatory role of IL-1β.  A marked increase in MMP-9 levels in turn lead to TIMP-1 activation. Significant increase in TIMP-1 levels was determined on the 3rd day after trauma. On the 7th day there was a critical period with the highest levels of IL-1β (2147,2%, MMP-9 (720,3% and TIMR-1 (339,3%. Then all research indicators were decreasing with the most pronounced decrease in IL-1β and MMP-9. Conclusion: MMP-9 levels began to increase on the 1st day after trauma due to influence mainly IL-1β. An abrupt increase in MMP-9 in its turn caused an increase in TIMR-1 levels. Conclusion: Identified changes in

In vitro and in vivo MMP gene expression localisation by In Situ-RT-PCR in cell culture and paraffin embedded human breast cancer cell line xenografts

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Haupt, Larisa M; Thompson, Erik W; Trezise, Ann EO; Irving, Rachel E; Irving, Michael G; Griffiths, Lyn R

2006-01-01

Members of the matrix metalloproteinase (MMP) family of proteases are required for the degradation of the basement membrane and extracellular matrix in both normal and pathological conditions. In vitro, MT1-MMP (MMP-14, membrane type-1-MMP) expression is higher in more invasive human breast cancer (HBC) cell lines, whilst in vivo its expression has been associated with the stroma surrounding breast tumours. MMP-1 (interstitial collagenase) has been associated with MDA-MB-231 invasion in vitro, while MMP-3 (stromelysin-1) has been localised around invasive cells of breast tumours in vivo. As MMPs are not stored intracellularly, the ability to localise their expression to their cells of origin is difficult. We utilised the unique in situ-reverse transcription-polymerase chain reaction (IS-RT-PCR) methodology to localise the in vitro and in vivo gene expression of MT1-MMP, MMP-1 and MMP-3 in human breast cancer. In vitro, MMP induction was examined in the MDA-MB-231 and MCF-7 HBC cell lines following exposure to Concanavalin A (Con A). In vivo, we examined their expression in archival paraffin embedded xenografts derived from a range of HBC cell lines of varied invasive and metastatic potential. Mouse xenografts are heterogenous, containing neoplastic human parenchyma with mouse stroma and vasculature and provide a reproducible in vivo model system correlated to the human disease state. In vitro, exposure to Con A increased MT1-MMP gene expression in MDA-MB-231 cells and decreased MT1-MMP gene expression in MCF-7 cells. MMP-1 and MMP-3 gene expression remained unchanged in both cell lines. In vivo, stromal cells recruited into each xenograft demonstrated differences in localised levels of MMP gene expression. Specifically, MDA-MB-231, MDA-MB-435 and Hs578T HBC cell lines are able to influence MMP gene expression in the surrounding stroma. We have demonstrated the applicability and sensitivity of IS-RT-PCR for the examination of MMP gene expression both in vitro and in

Expression of MMP-1/PAR-1 and patterns of invasion in oral squamous cell carcinoma as potential prognostic markers

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Fan HX

2015-07-01

Full Text Available Hai-Xia Fan,1 Yan Chen,1 Bo-Xiong Ni,1 Shan Wang,1 Miao Sun,2 Dong Chen,2 Jin-Hua Zheng11Department of Anatomy, Basic Medical Science College, 2Department of Oral and Maxillofacial Surgery, Stomatological Hospital, Harbin Medical University, Harbin, People’s Republic of ChinaBackground: Matrix metalloproteinase (MMP-1 degrades type I collagen of the extracellular matrix and also activates protease activated receptor (PAR-1 to induce angiogenesis. The aims of this study were to evaluate microvessel density (MVD and the expression of PAR-1 and MMP-1 in oral squamous cell carcinoma (SCC specimens with different patterns of invasion (POI and to evaluate their association with clinical outcomes.Methods: Seventy-four surgically obtained oral SCC samples were classified by POI according to hematoxylin-eosin staining. MVD and the localization and intensity of PAR-1 and MMP-1 expression were detected by immunohistochemistry.Results: Of the 74 oral SCC samples, 18, 5, 34, and 17 showed type I, II, III, and IV POI, respectively. MVD and expression levels of MMP-1 and PAR-1 differed between POI types I–II and POI types III–IV. Patients with low tumor expression of MMP-1 and PAR-1 and low MVD had a longer survival time than those with high tumor expression of MMP-1 and PAR-1. Moreover, the survival time of patients with POI types III–IV was shorter than that of patients with POI types I–II.Conclusion: POI combined with expression levels of MMP-1 and PAR-1 may be a valuable tool for assessing the clinical prognosis of patients with oral SCC.Keywords: oral squamous cell carcinoma, pattern of invasion, immunohistochemistry, clinical outcomes

Pannus invasion and cartilage degradation in rheumatoid arthritis: involvement of MMP-3 and interleukin-1beta.

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Ainola, M M; Mandelin, J A; Liljeström, M P; Li, T F; Hukkanen, M V J; Konttinen, Y T

2005-01-01

Synovial inflammation in rheumatoid arthritis (RA) leads to pannus tissue invasion and destruction of cartilage/bone matrix by proteinases. Our intention was to analyze some of the key matrix metalloproteinases (MMPs) in pannus tissue overlying evolving cartilage erosions in RA. Frozen tissue samples of pannus and synovium from advanced RA and synovium from osteoarthritic patients were used for immunohistochemical, western blotting and quantitative reverse transcriptase polymerase chain reaction (RT-PCR) analysis of MMP-1, -3, -13 and -14. Synovial fibroblast cultures, stimulated with tumour necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1beta), were analyzed with enzyme-linked immunosorbent assays (ELISA) and quantitative RT-PCR. MMP-3 was highly expressed in pannus tissue compared with significantly lower expression levels of MMP-1, -13 and -14. In fibroblast cultures IL-1beta was a potent stimulus for MMP-3, whereas TNF-alpha was more potent for MMP-1. This is the first study to demonstrate quantitatively in real time that MMP-3 mRNA expression is clearly higher in advanced RA pannus tissue compared to parallel RA or osteoarthritic synovium. MMP-3 mRNA levels were also clearly overexpressed in RA pannus compared to MMP-1, -13 and -14. Advanced RA has previously been found to overexpress IL-1beta. The high expression of MMP-3 in pannus and IL-1beta, mediated stimulation of MMP-3 suggest that MMP-3 plays a significant role in the progression of erosions through the proteoglycan-rich cartilage matrix.

Fisetin inhibits epidermal growth factor-induced migration of ARPE-19 cells by suppression of AKT activation and Sp1-dependent MMP-9 expression.

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Lin, Hung-Yu; Chen, Yong-Syuan; Wang, Kai; Chien, Hsiang-Wen; Hsieh, Yi-Hsien; Yang, Shun-Fa

2017-01-01

Proliferative vitreoretinopathy (PVR) can result in abnormal migration of RPE cells. Fisetin is a naturally occurring compound that has been reported to have antitumor effects, but its effects on epidermal growth factor (EGF)-induced cell migration and the underlying mechanisms remain unclear. Effects of fisetin on EGF-induced cell viability and migration were examined with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and in vitro migration assays. Reverse transcription-PCR (RT-PCR) and immunoblotting were performed to evaluate matrix metallopeptidase-9 (MMP-9) expression and activation of specificity protein-1 (Sp1) and protein kinase B (AKT) in ARPE-19 cells treated with EGF and with or without fisetin. Luciferase and chromatin immunoprecipitation (ChIP) assays were performed to examine Sp1 transcription activity and MMP-9 binding activity. Fisetin did not affect ARPE-19 cell viability and significantly inhibited the EGF-induced migration capacity of ARPE-19 cells. Furthermore, fisetin exerted an antimigratory effect and suppressed MMP-9 mRNA and protein expression. Treatment with EGF induced phosphorylation of AKT and expression of MMP-9 and Sp1. Fisetin combined with LY294002 (an inhibitor of AKT) prevented the EGF-induced migration involved in downregulation of Sp1 and MMP-9 expression. Luciferase and ChIP assays suggested that fisetin remarkably decreased the EGF-induced transcription activity of MMP-9 and Sp1 and inhibited EGF-mediated Sp1 from directly binding to the MMP-9 promoter in ARPE-19 cells. Fisetin inhibited EGF-induced cell migration via modulation of AKT/Sp1-dependent MMP-9 transcriptional activity. Therefore, fisetin may be a potential agent in the treatment of migratory PVR diseases.

Ang-(1-7) exerts protective role in blood-brain barrier damage by the balance of TIMP-1/MMP-9.

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Wu, Jitao; Zhao, Duo; Wu, Shuang; Wang, Dan

2015-02-05

Cerebrovascular disease (CVD) ranks as the top three health risks, specially cerebral ischemia characterized with the damage of blood-brain barrier (BBB). The angiotensin Ang-(1-7) was proven to have a protective effect on cerebrovascular diseases. However, its role on blood-brain barrier and the underlying molecular mechanism remains unclear. In this study, Ang-(1-7) significantly relieved damage of ischemia reperfusion injury on blood-brain barrier in cerebral ischemia reperfusion injury (IRI) rats. Furthermore, its treatment attenuated BBB permeability and brain edema. Similarly, Ang-(1-7) also decreased the barrier permeability of brain endothelial cell line RBE4. Further analysis showed that Ang-(1-7) could effectively restore tight junction protein (claudin-5 and zonula occludens ZO-1) expression levels both in IRI-rats and hypoxia-induced RBE4 cells. Furthermore, Ang-(1-7) stimulation down-regulated hypoxia-induced matrix metalloproteinase-9 (MMP-9) levels, whose silencing with (matrix metalloproteinase-9 hemopexin domain) MMP9-PEX inhibitor significantly increased the expression of claudin-5 and ZO-1. Further mechanism analysis demonstrated that Ang-(1-7) might junction protein levels by tissue inhibitor of metalloproteinase 1 (TIMP1)-MMP9 pathway, because Ang-(1-7) enhanced TIMP1 expression, whose silencing obviously attenuated the inhibitor effect of Ang-(1-7) on MMP-9 levels and decreased Ang-(1-7)-triggered increase in claudin-5 and ZO-1. Together, this study demonstrated a protective role of Ang-(1-7) in IRI-induced blood-brain barrier damage by TIMP1-MMP9-regulated tight junction protein expression. Accordingly, Ang-(1-7) may become a promising therapeutic agent against IRI and its complications. Copyright © 2014 Elsevier B.V. All rights reserved.

EFFECTIVENESS OF PNF STRETCHING AND CYCLIC STRETCHING OF CALF TIGHTNESS ON COLLEGE GOING GIRLS

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Ashlesha Sirari

2015-06-01

Full Text Available Background: Flexibility helps with injury prevention, the reduction of soreness following a workout, and a general sense of well-being. There are different stretching techniques and protocols for improvements in calf extensibility and flexibility. The purpose of the study was to investigate the effectiveness of two techniques i.e. CYCLIC and PNF stretching which improves calf flexibility. This study was done to find the effectiveness of calf Cyclic and PNF stretching technique to improve calf flexibility. Methods: 30 subjects with age group 21-22 years were randomly allocated to 2 groups equally. Group 1(n=15 were given CYCLIC and group 2(n=15 were given PNF stretching technique. Plantar flexion was used to measure the calf tightness which was done before and after the treatment. Treatment was given for 7 days and on the 7th day the calf tightness was again measured. Results: The mean difference of the CYCLIC is 4.6 and mean difference of PNF is 4.7 which indicate that CYCLIC and PNF both are effective to improve calf flexibility but PNF is more effective than CYCLIC to improve calf flexibility. Conclusion: The neurophysiological basis of PNF, stating that the excitatory efficient of the neuromuscular spindle or the inhibitory afferent of the Golgi tendon organ (GTO or both are responsible for the effects. During PNF stretch and isometric contraction of stretched agonists for extended period may cause activation of its neuromuscular spindle. The increase in tension created during the isometric contraction of the pre – lengthened agonist contracts concentrically. Both the fascia & the spindle of the agonist adjust to the nearly lengthened position. These impulses travel via causing post synaptic inhibition of the motor neuron to agonist increasing the tension from the GTO. These impulses can override the impulses coming from the neuromuscular spindles arousing the muscle to reflexly resist to the change in length, thus helping in lengthening

Downregulation of reversion-inducing cysteine-rich protein with Kazal motifs in malignant melanoma: inverse correlation with membrane-type 1-matrix metalloproteinase and tissue inhibitor of metalloproteinase 2.

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Jacomasso, Thiago; Trombetta-Lima, Marina; Sogayar, Mari C; Winnischofer, Sheila M B

2014-02-01

The invasive phenotype of many tumors is associated with an imbalance between the matrix metalloproteinases (MMPs) and their inhibitors, tissue inhibitors of metalloproteinases (TIMPs), and the membrane-anchored reversion-inducing cysteine-rich protein with Kazal motifs (RECK). RECK inhibits MMP-2, MMP-9, and MT1-MMP, and has been linked to patient survival and better prognosis in several types of tumors. However, despite the wide implication of these MMPs in melanoma establishment and progression, the role of RECK in this type of tumor is still unknown. Here, we analyzed the expression of RECK, TIMP1, TIMP2, TIMP3, MT1MMP, MMP2, and MMP9 in two publicly available melanoma microarray datasets and in a panel of human melanoma cell lines. We found that RECK is downregulated in malignant melanoma, accompanied by upregulation of MT1MMP and TIMP2. In both datasets, we observed that the group of samples displaying higher RECK levels show lower median expression levels of MT1MMP and TIMP2 and higher levels of TIMP3. When tested in a sample-wise manner, these correlations were statistically significant. Inverse correlations between RECK, MT1MMP, and TIMP2 were verified in a panel of human melanoma cell lines and in a further reduced model that includes a pair of matched primary tumor-derived and metastasis-derived cell lines. Taken together, our data indicate a consistent correlation between RECK, MT1MMP, and TIMP2 across different models of clinical samples and cell lines and suggest evidence of the potential use of this subset of genes as a gene signature for diagnosing melanoma.

Serum Levels of IL-1β, IL-6, TGF-β, and MMP-9 in Patients Undergoing Carotid Artery Stenting and Regulation of MMP-9 in a New In Vitro Model of THP-1 Cells Activated by Stenting

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Rongrong Zhang

2015-01-01

Full Text Available Inflammation plays an important role in the pathophysiological process after carotid artery stenting (CAS. Monocyte is a significant source of inflammatory cytokines in vascular remodeling. Telmisartan could reduce inflammation. In our study, we first found that, after CAS, the serum IL-1β, IL-6, TGF-β, and MMP-9 levels were significantly increased, but only MMP-9 level was elevated no less than 3 months. Second, we established a new in vitro model, where THP-1 monocytes were treated with the supernatants of human umbilical vein endothelial cells (HUVECs that were scratched by pipette tips, which mimics monocytes activated by mechanical injury of stenting. The treatment enhanced THP-1 cell adhesion, migration and invasion ability, and the phosphorylation of ERK1/2 and Elk-1 and MMP-9 expression were significantly increased. THP-1 cells pretreated with PD98095 (ERK1/2 inhibitor attenuated the phosphorylation of ERK1/2 and Elk-1 and upregulation of MMP-9, while pretreatment with telmisartan merely decreased the phosphorylation of Elk-1 and MMP-9 expression. These results suggested that IL-1β, IL-6, TGF-β, and MMP-9 participate in the pathophysiological process after CAS. Our new in vitro model mimics monocytes activated by stenting. MMP-9 expression could be regulated through ERK1/2/Elk-1 pathway, and the protective effects of telmisartan after stenting are partly attributed to its MMP-9 inhibition effects via suppression of Elk-1.

Fisetin inhibits epidermal growth factor–induced migration of ARPE-19 cells by suppression of AKT activation and Sp1-dependent MMP-9 expression

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Lin, Hung-Yu; Chen, Yong-Syuan; Wang, Kai; Chien, Hsiang-Wen

2017-01-01

Purpose Proliferative vitreoretinopathy (PVR) can result in abnormal migration of RPE cells. Fisetin is a naturally occurring compound that has been reported to have antitumor effects, but its effects on epidermal growth factor (EGF)–induced cell migration and the underlying mechanisms remain unclear. Methods Effects of fisetin on EGF-induced cell viability and migration were examined with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and in vitro migration assays. Reverse transcription–PCR (RT–PCR) and immunoblotting were performed to evaluate matrix metallopeptidase-9 (MMP-9) expression and activation of specificity protein-1 (Sp1) and protein kinase B (AKT) in ARPE-19 cells treated with EGF and with or without fisetin. Luciferase and chromatin immunoprecipitation (ChIP) assays were performed to examine Sp1 transcription activity and MMP-9 binding activity. Results Fisetin did not affect ARPE-19 cell viability and significantly inhibited the EGF-induced migration capacity of ARPE-19 cells. Furthermore, fisetin exerted an antimigratory effect and suppressed MMP-9 mRNA and protein expression. Treatment with EGF induced phosphorylation of AKT and expression of MMP-9 and Sp1. Fisetin combined with LY294002 (an inhibitor of AKT) prevented the EGF-induced migration involved in downregulation of Sp1 and MMP-9 expression. Luciferase and ChIP assays suggested that fisetin remarkably decreased the EGF-induced transcription activity of MMP-9 and Sp1 and inhibited EGF-mediated Sp1 from directly binding to the MMP-9 promoter in ARPE-19 cells. Conclusions Fisetin inhibited EGF-induced cell migration via modulation of AKT/Sp1–dependent MMP-9 transcriptional activity. Therefore, fisetin may be a potential agent in the treatment of migratory PVR diseases. PMID:29296070

SIPA1 promotes invasion and migration in human oral squamous cell carcinoma by ITGB1 and MMP7

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Takahara, Toshikazu; Kasamatsu, Atsushi; Yamatoji, Masanobu; Iyoda, Manabu; Kasama, Hiroki; Saito, Tomoaki; Takeuchi, Shin; Endo-Sakamoto, Yosuke; Shiiba, Masashi; Tanzawa, Hideki; Uzawa, Katsuhiro

2017-01-01

Signal-induced proliferation-associated protein 1 (SIPA1) is known to be a GTPase activating protein. Overexpressed SIPA1 is related to metastatic progression in breast and prostate cancers; however, the relevance of SIPA1 in oral squamous cell carcinoma (OSCC) is still unknown. The aim of this study was to examine SIPA1 expression and its functional mechanisms in OSCC. SIPA1 mRNA and protein expressions were analyzed by quantitative reverse transcriptase-polymerase chain reaction, Western blot analysis, and immunohistochemistry. The expressions of SIPA1 were up-regulated significantly in vitro and in vivo. Moreover, SIPA1 expression was correlated with regional lymph node metastasis. We next assessed the cellular functions associated with tumoral metastasis using SIPA1 knockdown (shSIPA1) cells and analyzed the downstream molecules of SIPA1, i.e., bromodomain containing protein 4(BRD4), integrin beta1 (ITGB1), and matrix metalloproteinase 7 (MMP7). The shSIPA1 cells showed decreased invasiveness and migratory activities, however cellular adhesion ability was maintained at a high level. In addition, ITGB1 expression was greater in shSIPA1 cells, whereas MMP7 expression was lower than in control cells. This research is the first to establish that SIPA1 promotes cancer metastasis by regulating the ITGB1 and MMP7. Therefore, SIPA1 might be a novel therapeutic target for patients with lymph node metastasis of OSCC. - Highlights: • SIPA1 expression was up-regulated in oral squamous cell carcinoma (OSCC). • SIPA1-positive OSCCs were correlated with regional lymph node metastasis. • SIPA1 controlled BRD4 and influenced transcription of ITGB1and MMP7. • SIPA1 induced cellular invasion and migration and decreased cellular adhesion. • SIPA1 might be a potential biomarker of cancer metastasis for OSCC.

SIPA1 promotes invasion and migration in human oral squamous cell carcinoma by ITGB1 and MMP7

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Takahara, Toshikazu [Department of Oral Science, Graduate School of Medicine, Chiba University, Chiba (Japan); Kasamatsu, Atsushi, E-mail: kasamatsua@faculty.chiba-u.jp [Department of Dentistry and Oral-Maxillofacial Surgery, Chiba University Hospital, Chiba (Japan); Yamatoji, Masanobu [Department of Dentistry and Oral-Maxillofacial Surgery, Chiba University Hospital, Chiba (Japan); Iyoda, Manabu; Kasama, Hiroki; Saito, Tomoaki [Division of Oral Surgery, Chiba Rosai Hospital, Chiba (Japan); Takeuchi, Shin [Department of Oral Science, Graduate School of Medicine, Chiba University, Chiba (Japan); Endo-Sakamoto, Yosuke [Department of Dentistry and Oral-Maxillofacial Surgery, Chiba University Hospital, Chiba (Japan); Shiiba, Masashi [Department of Medical Oncology, Graduate School of Medicine, Chiba University, Chiba (Japan); Tanzawa, Hideki [Department of Oral Science, Graduate School of Medicine, Chiba University, Chiba (Japan); Department of Dentistry and Oral-Maxillofacial Surgery, Chiba University Hospital, Chiba (Japan); Uzawa, Katsuhiro, E-mail: uzawak@faculty.chiba-u.jp [Department of Oral Science, Graduate School of Medicine, Chiba University, Chiba (Japan); Department of Dentistry and Oral-Maxillofacial Surgery, Chiba University Hospital, Chiba (Japan)

2017-03-15

Signal-induced proliferation-associated protein 1 (SIPA1) is known to be a GTPase activating protein. Overexpressed SIPA1 is related to metastatic progression in breast and prostate cancers; however, the relevance of SIPA1 in oral squamous cell carcinoma (OSCC) is still unknown. The aim of this study was to examine SIPA1 expression and its functional mechanisms in OSCC. SIPA1 mRNA and protein expressions were analyzed by quantitative reverse transcriptase-polymerase chain reaction, Western blot analysis, and immunohistochemistry. The expressions of SIPA1 were up-regulated significantly in vitro and in vivo. Moreover, SIPA1 expression was correlated with regional lymph node metastasis. We next assessed the cellular functions associated with tumoral metastasis using SIPA1 knockdown (shSIPA1) cells and analyzed the downstream molecules of SIPA1, i.e., bromodomain containing protein 4(BRD4), integrin beta1 (ITGB1), and matrix metalloproteinase 7 (MMP7). The shSIPA1 cells showed decreased invasiveness and migratory activities, however cellular adhesion ability was maintained at a high level. In addition, ITGB1 expression was greater in shSIPA1 cells, whereas MMP7 expression was lower than in control cells. This research is the first to establish that SIPA1 promotes cancer metastasis by regulating the ITGB1 and MMP7. Therefore, SIPA1 might be a novel therapeutic target for patients with lymph node metastasis of OSCC. - Highlights: • SIPA1 expression was up-regulated in oral squamous cell carcinoma (OSCC). • SIPA1-positive OSCCs were correlated with regional lymph node metastasis. • SIPA1 controlled BRD4 and influenced transcription of ITGB1and MMP7. • SIPA1 induced cellular invasion and migration and decreased cellular adhesion. • SIPA1 might be a potential biomarker of cancer metastasis for OSCC.

Gelatinase (MMP-2 and -9 expression profiles during gestation in the bovine endometrium

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Sato Takashi

2008-12-01

Full Text Available Abstract Background Various molecules participate in implantation and maintaining endometrial function during gestation. The remodeling of endometrial matrices is a necessary process in the coordination of gestational progress. Matrix-metalloproteinases (MMPs like gelatinases (MMP-2 and -9 and collagenase (MMP-1 are considered to play important roles in this process. We examined MMP-2 and -9 expression using zymography, in situ hybridization, real-time PCR, and microarray analysis to clarify their roles in the bovine endometrium during gestation. Methods Endometria, placentomes, and fetal membranes were collected from Japanese black cows that were killed on day 15 to 252 of gestation or during their estrous cycle. The gene expression of MMP-related molecules (mainly MMP-2 and -9 was examined using a custom-made microarray, real-time RT-PCR, and in-situ hybridization. Gelatinase activity was detected by zymography and film in situ zymography. Results Both gelatinases were expressed in the endometrium and fetal tissues throughout gestation. MMP-2 gene expression declined with the progress of gestation, but its intensity was maintained at a high level during the peri-implantation period and increased in late gestation. The expression level of MMP-9 was stably maintained, but was relatively low compared to that of MMP-2. These gene expression patterns matched those detected by zymography for the proteins. Microarray analysis suggested that the functions of MMP-2 during implantation and the last part of gestation are closely related with those of other molecules such as tissue inhibitors of metalloproteinase (TIMP-2, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS 1, membrane type 1 (MT1-MMP, and extracellular matrix metalloproteinase inducer (EMMPRIN. Conclusion We detected MMP-2 and -9 gene expression in the bovine endometrium and placentome throughout gestation. These data suggest that MMP-2 is one of the main endometrial

Catecholamine up-regulates MMP-7 expression by activating AP-1 and STAT3 in gastric cancer

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Yu Ming

2010-10-01

Full Text Available Abstract Background Stress, anxiety and depression can cause complex physiological and neuroendocrine changes, resulting in increased level of stress related hormone catecholamine, which may constitute a primary mechanism by which physiological factors impact gene expression in tumors. In the present study, we investigated the effects of catecholamine stimulation on MMP-7 expression in gastric cancer cells and elucidated the molecular mechanisms of the up-regulation of MMP-7 level by catecholamine through an adrenergic signaling pathway. Results Increased MMP-7 expression was identified at both mRNA and protein levels in the gastric cancer cells in response to isoproterenol stimulation. β2-AR antigonist effectively abrogated isoproterenol-induced MMP-7 expression. The activation of STAT3 and AP-1 was prominently induced by isoproterenol stimulation and AP-1 displayed a greater efficacy than STAT3 in isoproterenol-induced MMP-7 expression. Mutagenesis of three STAT3 binding sites in MMP-7 promoter failed to repress the transactivation of MMP-7 promoter and silencing STAT3 expression was not effective in preventing isoproterenol-induced MMP-7 expression. However, isoproterenol-induced MMP-7 promoter activities were completely disappeared when the AP-1 site was mutated. STAT3 and c-Jun could physically interact and bind to the AP-1 site, implicating that the interplay of both transcriptional factors on the AP-1 site is responsible for isoproterenol-stimulated MMP-7 expression in gastric cancer cells. The expression of MMP-7 in gastric cancer tissues was found to be at the site where β2-AR was overexpressed and the levels of MMP-7 and β2-AR were the highest in the metastatic locus of gastric cancer. Conclusions Up-regulation of MMP-7 expression through β2-AR-mediated signaling pathway is involved in invasion and metastasis of gastric cancer.

Mercury exposure induces cytoskeleton disruption and loss of renal function through epigenetic modulation of MMP9 expression.

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Khan, Hafizurrahman; Singh, Radha Dutt; Tiwari, Ratnakar; Gangopadhyay, Siddhartha; Roy, Somendu Kumar; Singh, Dhirendra; Srivastava, Vikas

2017-07-01

Mercury is one of the major heavy metal pollutants occurring in elemental, inorganic and organic forms. Due to ban on most inorganic mercury containing products, human exposure to mercury generally occurs as methylmercury (MeHg) by consumption of contaminated fish and other sea food. Animal and epidemiological studies indicate that MeHg affects neural and renal function. Our study is focused on nephrotoxic potential of MeHg. In this study, we have shown for the first time how MeHg could epigenetically modulate matrix metalloproteinase 9(MMP9) to promote nephrotoxicity using an animal model of sub chronic MeHg exposure. MeHg caused renal toxicity as was seen by increased levels of serum creatinine and expression of early nephrotoxicity markers (KIM-1, Clusterin, IP-10, and TIMP). MeHg exposure also correlated strongly with induction of MMP9 mRNA and protein in a dose dependent manner. Further, while induction of MMP9 promoted cytoskeleton disruption and loss of cell-cell adhesion (loss of F-actin, Vimentin and Fibronectin), inhibition of MMP9 was found to reduce these disruptions. Mechanistic studies by ChIP analysis showed that MeHg modulated MMP9 by promoting demethylation of its regulatory region to increase its expression. Bisulfite sequencing identified critical CpGs in the first exon of MMP9 which were demethylated following MeHg exposure. ChIP studies also showed loss of methyl binding protein, MeCP2 and transcription factor PEA3 at the demethylated site confirming decreased CpG methylation. Our studies thus show how MeHg could epigenetically modulate MMP9 to promote cytoskeleton disruption leading to loss of renal function. Copyright © 2017 Elsevier B.V. All rights reserved.

IGF-1-induced MMP-11 expression promotes the proliferation and invasion of gastric cancer cells through the JAK1/STAT3 signaling pathway.

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Su, Chao; Wang, Wenchang; Wang, Cunchuan

2018-05-01

The present study aimed to investigate the association between insulin-like growth factor-1 (IGF-1) and matrix metalloproteinase-11 (MMP-11) expression in gastric cancer (GC) and the underlying mechanisms in SGC-7901 cells. Reverse transcription-quantitative polymerase chain reaction analysis revealed that the expression of IGF-1 and MMP-11 was significantly upregulated in GC tissues compared with normal gastric tissue. Furthermore, IGF-1 significantly and dose-dependently promoted MMP-11. Western blotting revealed that the addition of IGF-1 to SGC-7901 cells led to an evident enhancement in signal transducer and activator of transcription 3 (STAT3), IGF-1R and Janus kinase 1 (JAK1) phosphorylation at 20 and 40 min. A decrease in the extent of the elevated expression of MMP-11 and the enhanced phosphorylation of STAT3, JAK1 and IGF-1 receptor (IGF-1R) induced by IGF-1 in SGC-7901 cells were observed following treatment with NT157 (an IGF-1R inhibitor). Furthermore, piceatannol (a JAK1 inhibitor) or small interfering RNA against STAT3 reduced the extent of the increased expression of MMP-11 induced by IGF-1 in SGC-7901 cells. Piceatannol treatment induced the dose-dependent decline in the enhancement of STAT3 phosphorylation induced by IGF-1, indicating that the JAK1/STAT3 pathway may be implicated in the elevated expression of MMP-11 induced by IGF-1 in SGC-7901 cells. Finally, IGF-1 treatment significantly promoted the proliferation and invasion of SGC-7901 cells, which was inhibited following NT157, piceatannol or si-STAT3 treatment. The present study therefore demonstrated that IGF-1-induced MMP-11 may have facilitated the proliferation and invasion of SGC-7901 cells via the JAK1/STAT3 pathway.

MMP-7 and TIMP-1, new targets in predicting poor wound healing in apical periodontitis.

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Letra, Ariadne; Ghaneh, Ghazaleh; Zhao, Min; Ray, Herbert; Francisconi, Carolina Favaro; Garlet, Gustavo Pompermaier; Silva, Renato Menezes

2013-09-01

Matrix metalloproteinases (MMPs) and the tissue inhibitors of metalloproteinases (TIMPs) are strongly associated with tissue destruction because of inflammation. In this study, we investigated the expression of MMPs and TIMPs messenger RNA and protein levels in apical periodontitis lesions. Tissue samples from patients presenting clinical signs of chronic apical abscess (CAA) or asymptomatic apical periodontitis (AAP) were collected postoperatively and used for gene expression analysis of MMP-2, -3, -7, -9, -14, -16, and -25; TIMP-1; and TIMP-2 in real-time polymerase chain reaction. Immunohistochemistry was also performed to detect the expression of MMP-7 and TIMP-1 proteins. Lastly, U-937 cells were induced to terminal differentiation into macrophages, infected with purified Escherichia coli lipopolysaccharide, and assessed for the expression of MMP-7 and TIMP-1 using immunocytochemistry and confocal microscopy. Significantly higher messenger RNA levels were found for all genes in AAP and CAA samples when compared with healthy control samples (P < .001). AAP cases exhibited significantly higher TIMP-1 when compared with CAA cases, whereas CAA cases showed higher MMP-2, MMP-7, and MMP-9 messenger RNA levels (P < .05). We also detected positive the expression of MMP-7 and TIMP-1 proteins in the tissue samples. The expression of both MMP-7 and TIMP-1 were increased in lipopolysaccharide-stimulated cells compared with nonstimulated cells and appear to colocalize in the Golgi apparatus. MMPs appear to have an influential role in CAA cases in which ongoing tissue destruction is observed. TIMPs are preferentially associated with AAP, perhaps as a subsequent defense mechanism against excessive destruction. Taken together, our findings implicate MMP and TIMP molecules in the dynamics of inflammatory periapical lesion development. Copyright © 2013 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Effects of acute static, ballistic, and PNF stretching exercise on the muscle and tendon tissue properties.

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Konrad, A; Stafilidis, S; Tilp, M

2017-10-01

The purpose of this study was to investigate the influence of a single static, ballistic, or proprioceptive neuromuscular facilitation (PNF) stretching exercise on the various muscle-tendon parameters of the lower leg and to detect possible differences in the effects between the methods. Volunteers (n = 122) were randomly divided into static, ballistic, and PNF stretching groups and a control group. Before and after the 4 × 30 s stretching intervention, we determined the maximum dorsiflexion range of motion (RoM) with the corresponding fascicle length and pennation angle of the gastrocnemius medialis. Passive resistive torque (PRT) and maximum voluntary contraction (MVC) were measured with a dynamometer. Observation of muscle-tendon junction (MTJ) displacement with ultrasound allowed us to determine the length changes in the tendon and muscle, respectively, and hence to calculate stiffness. Although RoM increased (static: +4.3%, ballistic: +4.5%, PNF: +3.5%), PRT (static: -11.4%, ballistic: -11.5%, PNF: -13,7%), muscle stiffness (static: -13.1%, ballistic: -20.3%, PNF: -20.2%), and muscle-tendon stiffness (static: -11.3%, ballistic: -10.5%, PNF: -13.7%) decreased significantly in all the stretching groups. Only in the PNF stretching group, the pennation angle in the stretched position (-4.2%) and plantar flexor MVC (-4.6%) decreased significantly. Multivariate analysis showed no clinically relevant difference between the stretching groups. The increase in RoM and the decrease in PRT and muscle-tendon stiffness could be explained by more compliant muscle tissue following a single static, ballistic, or PNF stretching exercise. © 2017 The Authors Scandinavian Journal of Medicine & Science In Sports Published by John Wiley & Sons Ltd.

Activation of Nrf2 Reduces UVA-Mediated MMP-1 Upregulation via MAPK/AP-1 Signaling Cascades: The Photoprotective Effects of Sulforaphane and Hispidulin

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Chaiprasongsuk, Anyamanee; Lohakul, Jinaphat; Soontrapa, Kitipong; Sampattavanich, Somponnat; Akarasereenont, Pravit

2017-01-01

UVA irradiation plays a role in premature aging of the skin through triggering oxidative stress-associated stimulation of matrix metalloproteinase-1 (MMP-1) responsible for collagen degradation, a hallmark of photoaged skin. Compounds that can activate nuclear factor E2-related factor 2 (Nrf2), a transcription factor regulating antioxidant gene expression, should therefore serve as effective antiphotoaging agents. We investigated whether genetic silencing of Nrf2 could relieve UVA-mediated MMP-1 upregulation via activation of mitogen-activated protein kinase (MAPK)/activator protein 1 (AP-1) signaling using human keratinocyte cell line (HaCaT). Antiphotoaging effects of hispidulin (HPD) and sulforaphane (SFN) were assessed on their abilities to activate Nrf2 in controlling MMP-1 and collagen expressions in association with phosphorylation of MAPKs (extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38), c-Jun, and c-Fos, using the skin of BALB/c mice subjected to repetitive UVA irradiation. Our findings suggested that depletion of Nrf2 promoted both mRNA expression and activity of MMP-1 in the UVA-irradiated HaCaT cells. Treatment of Nrf2 knocked-down HaCaT cells with MAPK inhibitors significantly suppressed UVA-induced MMP-1 and AP-1 activities. Moreover, pretreatment of the mouse skin with HPD and SFN, which could activate Nrf2, provided protective effects against UVA-mediated MMP-1 induction and collagen depletion in correlation with the decreased levels of phosphorylated MAPKs, c-Jun, and c-Fos in the mouse skin. In conclusion, Nrf2 could influence UVA-mediated MMP-1 upregulation through the MAPK/AP-1 signaling cascades. HPD and SFN may therefore represent promising antiphotoaging candidates. PMID:28011874

Association of MMP-9 Haplotypes and TIMP-1 Polymorphism with Spontaneous Deep Intracerebral Hemorrhage in the Taiwan Population.

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Wei-Min Ho

Full Text Available Spontaneous deep intracerebral hemorrhage (SDICH is a devastating stroke subtype. The causes of SDICH are heterogeneous. Matrix metalloproteinase-9 (MMP-9, Gelantinase B has been shown to relate to stroke and the development of aneurysm and may increase risks of intracerebral hemorrhage. MMP activities are modulated by their endogenous inhibitors, tissue inhibitors of metalloproteinases (TIMPs. We analyzed the genetic variants of MMP-9 and TIMP-1 and SDICH susceptibility.Associations were tested by logistic regression or general linear models with adjusting for multiple covariables. Multiplicative terms between genes were applied to detect the interaction effects on SDICH. Permutation testing of 1,000 replicates was performed for empirical estimates.In the group of ≥65 years old (y/o, we found associations of SDICH with rs3787268 (Odds ratio [OR] = 0.48, 95% confidence interval [CI] 0.27 to 0.86, P = 0.01 and haplotype1 (Hap1 (OR = 0.48, 95% CI 0.26 to 0.86, P = 0.014. For TIMP1 gene, rs4898 was associated with SDICH in the elder male group (OR = 0.35, 95% CI 0.15 to 0.81, P = 0.015. In contrast, in the younger male group, there were associations of SDICH with rs2250889 (OR = 0.48, 95% CI 0.27 to 0.84, P = 0.01 and Hap3 (OR = 0.61, 95% CI 0.38 to 0.97, P = 0.04. We found significant genetic interaction between TIMP-1 and MMP-9 in SDICH susceptibility among younger male subjects (P = 0.004. In subjects carrying rs4898 minor allele, carriers with Hap3 had lower SDICH risk than non-carriers (OR = 0.19, 95% CI 0.07 to 0.51, P = 0.001. In addition, this study showed that when young males were exposed to alcohol, Hap3 was a protective factor of SDICH (OR = 0.06, 95% CI 0.01 to 0.27, P = 0.0002. In contrast, when they were exposed to smoke, Hap2 carriers had increased risk of SDICH (OR = 2.45, 95% CI 1.05 to 5.73, P = 0.04.This study showed modest to moderate effects of MMP-9 and TIMP-1 polymorphisms on SDICH risks with significant age differences

Proteomic analysis identifies MMP-9, DJ-1 and A1BG as overexpressed proteins in pancreatic juice from pancreatic ductal adenocarcinoma patients

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Tian, Mei; Cui, Ya-Zhou; Song, Guan-Hua; Zong, Mei-Juan; Zhou, Xiao-Yan; Chen, Yu; Han, Jin-Xiang

2008-01-01

There is an urgent need to discover more sensitive and specific biomarkers to improve early diagnosis and screen high-risk patients for pancreatic ductal adenocarcinoma (PDAC). Pancreatic juice is an ideal specimen for PDAC biomarkers discovery, because it is an exceptionally rich source of proteins released from pancreatic cancer cells. To identify novel potential biomarkers for PDAC from pancreatic juice, we carried out difference gel electrophoresis (DIGE) and tandem mass spectrometry (MS/MS) to compare the pancreatic juice profiling from 9 PDAC patients and 9 cancer-free controls. Of the identified differently expressed proteins, three up-regulated proteins in pancreatic cancer juice, matrix metalloproteinase-9 (MMP-9), oncogene DJ1 (DJ-1) and alpha-1B-glycoprotein precursor (A1BG), were selected for validation by Western blot and immunohistochemistry. Serum MMP-9 levels were also detected by enzyme linked immunosorbent assay (ELISA). Fourteen proteins were up-regulated and ten proteins were down-regulated in cancerous pancreatic juice compared with cancer-free controls. Increased MMP-9, DJ-1 and A1BG expression in cancerous pancreatic juice were confirmed by Western blot. Immunohistochemical study showed MMP-9, DJ-1 and A1BG positively expressed in 82.4%, 72.5% and 86.3% of pancreatic cancer tissues, significantly higher than that in normal pancreas tissues. Up-regulation of DJ-1 was associated with better differentiation (p < 0.05). Serum MMP-9 levels were significantly higher in PDAC (255.14 ng/ml) than those in chronic pancreatitis (210.22 ng/ml, p = 0.009) and healthy control (203.77 ng/ml, p = 0.027). The present proteome analysis revealed MMP-9, DJ-1 and A1BG proteins as elevated in pancreatic juice from PDAC, which suggest their further utility in PDAC diagnosis and screening. This is the first time A1BG was identified as a potential biomarker in pancreatic cancer associated samples. The measurement of serum MMP-9 might be clinically useful for PDAC

The dimer interface of the membrane type 1 matrix metalloproteinase hemopexin domain: crystal structure and biological functions.

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Tochowicz, Anna; Goettig, Peter; Evans, Richard; Visse, Robert; Shitomi, Yasuyuki; Palmisano, Ralf; Ito, Noriko; Richter, Klaus; Maskos, Klaus; Franke, Daniel; Svergun, Dmitri; Nagase, Hideaki; Bode, Wolfram; Itoh, Yoshifumi

2011-03-04

Homodimerization is an essential step for membrane type 1 matrix metalloproteinase (MT1-MMP) to activate proMMP-2 and to degrade collagen on the cell surface. To uncover the molecular basis of the hemopexin (Hpx) domain-driven dimerization of MT1-MMP, a crystal structure of the Hpx domain was solved at 1.7 Å resolution. Two interactions were identified as potential biological dimer interfaces in the crystal structure, and mutagenesis studies revealed that the biological dimer possesses a symmetrical interaction where blades II and III of molecule A interact with blades III and II of molecule B. The mutations of amino acids involved in the interaction weakened the dimer interaction of Hpx domains in solution, and incorporation of these mutations into the full-length enzyme significantly inhibited dimer-dependent functions on the cell surface, including proMMP-2 activation, collagen degradation, and invasion into the three-dimensional collagen matrix, whereas dimer-independent functions, including gelatin film degradation and two-dimensional cell migration, were not affected. These results shed light on the structural basis of MT1-MMP dimerization that is crucial to promote cellular invasion.

Plasma levels of the MMP-9:TIMP-1 complex as prognostic biomarker in breast cancer: a retrospective study

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Thorsen, Stine B; Møller, Susanne; Brünner, Nils; Schrohl, Anne-Sofie; Stenvang, Jan; Christensen, Sarah LT; Würtz, Sidse Ø; Lundberg, Martin; Nielsen, Birgitte S; Vinther, Lena; Knowles, Mick; Gee, Nick; Fredriksson, Simon

2013-01-01

Worldwide more than one million women are annually diagnosed with breast cancer. A considerable fraction of these women receive systemic adjuvant therapy; however, some are cured by primary surgery and radiotherapy alone. Prognostic biomarkers guide stratification of patients into different risk groups and hence improve management of breast cancer patients. Plasma levels of Matrix Metalloproteinase-9 (MMP-9) and its natural inhibitor Tissue inhibitor of metalloproteinase-1 (TIMP-1) have previously been associated with poor patient outcome and resistance to certain forms of chemotherapy. To pursue additional prognostic information from MMP-9 and TIMP-1, the level of the MMP-9 and TIMP-1 complex (MMP-9:TIMP-1) was investigated in plasma from breast cancer patients. Detection of protein:protein complexes in plasma was performed using a commercially available ELISA kit and, for the first time, the highly sensitive in-solution proximity ligation assay (PLA). We screened plasma from 465 patients with primary breast cancer for prognostic value of the MMP-9:TIMP-1 complex. Both assays were validated and applied for quantification of MMP-9:TIMP-1 concentration. In this retrospective study, we analyzed the association between the concentration of the MMP-9:TIMP-1 complex and clinicopathological data and disease free survival (DFS) in univariate and multivariate survival analyses. Following successful validation both assays were applied for MMP-9:TIMP-1 measurements. Of the clinicopathological parameters, only menopausal status demonstrated significant association with the MMP-9:TIMP-1 complex; P = 0.03 and P = 0.028 for the ELISA and PLA measurements, respectively. We found no correlation between the MMP-9:TIMP-1 protein complex and DFS neither in univariate nor in multivariate survival analyses. Despite earlier reports linking MMP-9 and TIMP-1 with prognosis in breast cancer patients, we here demonstrate that plasma levels of the MMP-9:TIMP-1 protein complex hold no

Biochemical properties of the matrix metalloproteinase NtMMP1 from Nicotiana tabacum cv. BY-2 suspension cells.

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Mandal, Manoj K; Fischer, Rainer; Schillberg, Stefan; Schiermeyer, Andreas

2010-09-01

A zinc-dependent matrix metalloproteinase (NtMMP1) found in the plasma membrane of Nicotiana tabacum cv. Bright Yellow 2 (BY-2) suspension cells is thought to be responsible for the degradation of recombinant proteins secreted into the culture supernatant. We have characterized the proteolytic activity of NtMMP1 by expressing a recombinant derivative lacking the C-terminal transmembrane domain in yeast. After purifying the protein by affinity chromatography, its autocatalytic activity was analyzed using monoclonal antibodies raised against its N-terminal and C-terminal portions. Both the unprocessed and processed forms of NtMMP1 displayed caseinolytic activity and N-terminal sequencing identified an autocatalytic cleavage site within the sequence motif HFSFFP, which is similar to the corresponding sequences of the human matrix metalloproteinases stromelysin-1 (MMP-3) and stromelysin-2 (MMP-10). Unlike all other matrix metalloproteinases investigated so far, NtMMP1 contains a disulfide bond within its propeptide thus rendering the proenzyme catalytically active. Kinetic analysis of NtMMP1 with a synthetic substrate revealed a K(m) of 10.55 +/- 0.9 microM, a k(cat) of 0.6 +/- 0.01 s(-1) and maximum activity at pH 7.5. We found that NtMMP1 degrades Desmodus rotundus salivary plasminogen activator alpha 1 (DSPAalpha1), a biopharmaceutical protein, that has proven difficult to produce in tobacco BY-2 cells. This provides a likely explanation for the frequent instability of secreted recombinant biopharmaceuticals produced in plant suspension cell cultures. Our data suggest new avenues that can be explored to improve the production of pharmaceutical proteins in plants and plant cells.

Resveratrol Targeting of Carcinogen-Induced Brain Endothelial Cell Inflammation Biomarkers MMP-9 and COX-2 is Sirt1-Independent

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Borhane Annabi

2012-01-01

Full Text Available The occurrence of a functional relationship between the release of metalloproteinases (MMPs and the expression of cyclooxygenase (COX-2, two inducible pro-inflammatory biomarkers with important pro-angiogenic effects, has recently been inferred. While brain endothelial cells play an essential role as structural and functional components of the blood-brain barrier (BBB, increased BBB breakdown is thought to be linked to neuroinflammation. Chemopreventive mechanisms targeting both MMPs and COX-2 however remain poorly investigated. In this study, we evaluated the pharmacological targeting of Sirt1 by the diet-derived and antiinflammatory polyphenol resveratrol. Total RNA, cell lysates, and conditioned culture media from human brain microvascular endothelial cells (HBMEC were analyzed using qRT-PCR, immunoblotting, and zymography respectively. Tissue scan microarray analysis of grade I–IV brain tumours cDNA revealed increased gene expression of Sirt-1 from grade I–III but surprisingly not in grade IV brain tumours. HBMEC were treated with a combination of resveratrol and phorbol 12-myristate 13-acetate (PMA, a carcinogen known to increase MMP-9 and COX-2 through NF-κB. We found that resveratrol efficiently reversed the PMA-induced MMP-9 secretion and COX-2 expression. Gene silencing of Sirt1, a critical modulator of angiogenesis and putative target of resveratrol, did not lead to significant reversal of MMP-9 and COX-2 inhibition. Decreased resveratrol inhibitory potential of carcinogen-induced IκB phosphorylation in siSirt1-transfected HBMEC was however observed. Our results suggest that resveratrol may prevent BBB disruption during neuroinflammation by inhibiting MMP-9 and COX-2 and act as a pharmacological NF-κB signal transduction inhibitor independent of Sirt1.

Effects of ultrasound therapy with taping PNF training and PNF training with taping in treatment and rehabilitation of sports injuries of high ankle sprain

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D L Charly Daniel

2017-01-01

Conclusions: It was concluded that combining ultrasound with taping and PNF training with taping were found to be more beneficial in the treatment and rehabilitation of high ankle sprain injury. The combined effect of UT, PNF training, and taping may be explored by future researchers.

Downregulation of MMP1 in MDS-derived mesenchymal stromal cells reduces the capacity to restrict MDS cell proliferation.

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Zhao, Sida; Zhao, Youshan; Guo, Juan; Fei, Chengming; Zheng, Qingqing; Li, Xiao; Chang, Chunkang

2017-03-06

The role of mesenchymal stromal cells (MSCs) in the pathogenesis of myelodysplastic syndromes (MDS) has been increasingly addressed, but has yet to be clearly elucidated. In this investigation, we found that MDS cells proliferated to a greater extent on MDS-derived MSCs compared to normal MSCs. Matrix metalloproteinase 1(MMP1), which was downregulated in MDS-MSCs, was identified as an inhibitory factor of MDS cell proliferation, given that treatment with an MMP1 inhibitor or knock-down of MMP1 in normal MSCs resulted in increased MDS cell proliferation. Further investigations indicated that MMP1 induced apoptosis of MDS cells by interacting with PAR1 and further activating the p38 MAPK pathway. Inhibition of either PAR1 or p38 MAPK can reverse the apoptosis-inducing effect of MMP1. Taken together, these data indicate that downregulation of MMP1 in MSCs of MDS patients may contribute to the reduced capacity of MSCs to restrict MDS cell proliferation, which may account for the malignant proliferation of MDS cells.

Medicago truncatula copper transporter 1 (MtCOPT1) delivers copper for symbiotic nitrogen fixation.

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Senovilla, Marta; Castro-Rodríguez, Rosario; Abreu, Isidro; Escudero, Viviana; Kryvoruchko, Igor; Udvardi, Michael K; Imperial, Juan; González-Guerrero, Manuel

2018-04-01

Copper is an essential nutrient for symbiotic nitrogen fixation. This element is delivered by the host plant to the nodule, where membrane copper (Cu) transporter would introduce it into the cell to synthesize cupro-proteins. COPT family members in the model legume Medicago truncatula were identified and their expression determined. Yeast complementation assays, confocal microscopy and phenotypical characterization of a Tnt1 insertional mutant line were carried out in the nodule-specific M. truncatula COPT family member. Medicago truncatula genome encodes eight COPT transporters. MtCOPT1 (Medtr4g019870) is the only nodule-specific COPT gene. It is located in the plasma membrane of the differentiation, interzone and early fixation zones. Loss of MtCOPT1 function results in a Cu-mitigated reduction of biomass production when the plant obtains its nitrogen exclusively from symbiotic nitrogen fixation. Mutation of MtCOPT1 results in diminished nitrogenase activity in nodules, likely an indirect effect from the loss of a Cu-dependent function, such as cytochrome oxidase activity in copt1-1 bacteroids. These data are consistent with a model in which MtCOPT1 transports Cu from the apoplast into nodule cells to provide Cu for essential metabolic processes associated with symbiotic nitrogen fixation. © 2018 The Authors. New Phytologist © 2018 New Phytologist Trust.

Peroxisome proliferator-activated receptor δ modulates MMP-2 secretion and elastin expression in human dermal fibroblasts exposed to ultraviolet B radiation.

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Ham, Sun Ah; Yoo, Taesik; Hwang, Jung Seok; Kang, Eun Sil; Paek, Kyung Shin; Park, Chankyu; Kim, Jin-Hoi; Do, Jeong Tae; Seo, Han Geuk

2014-10-01

Changes in skin connective tissues mediated by ultraviolet (UV) radiation have been suggested to cause the skin wrinkling normally associated with premature aging of the skin. Recent investigations have shown that peroxisome proliferator-activated receptor (PPAR) δ plays multiple biological roles in skin homeostasis. We attempted to investigate whether PPARδ modulates elastin protein levels and secretion of matrix metalloproteinase (MMP)-2 in UVB-irradiated human dermal fibroblasts (HDFs) and mouse skin. These studies were undertaken in primary HDFs or HR-1 hairless mice using Western blot analyses, small interfering (si)RNA-mediated gene silencing, and Fluorescence microscopy. In HDFs, UVB irradiation induced increased secretion of MMP-2 and reduced levels of elastin. Activation of PPARδ by GW501516, a ligand specific for PPARδ, markedly attenuated UVB-induced MMP-2 secretion with a concomitant increase in the level of elastin. These effects were reduced by the presence of siRNAs against PPARδ or treatment with GSK0660, a specific inhibitor of PPARδ. Furthermore, GW501516 elicited a dose- and time-dependent increase in the expression of elastin. Modulation of MMP-2 secretion and elastin levels by GW501516 was associated with a reduction in reactive oxygen species (ROS) production in HDFs exposed to UVB. Finally, in HR-1 hairless mice, administration of GW501516 significantly reduced UVB-induced MMP-2 expression with a concomitant increase in elastin levels, and these effects were significantly reduced by the presence of GSK0660. Our results suggest that PPARδ-mediated modulation of MMP-2 secretion and elastin expression may contribute to the maintenance of skin integrity by inhibiting ROS generation. Copyright © 2014 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

Joint effect of MCP-1 genotype GG and MMP-1 genotype 2G/2G increases the likelihood of developing pulmonary tuberculosis in BCG-vaccinated individuals.

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Malathesha Ganachari

2010-01-01

Full Text Available We previously reported that the -2518 MCP-1 genotype GG increases the likelihood of developing tuberculosis (TB in non-BCG-vaccinated Mexicans and Koreans. Here, we tested the hypothesis that this genotype, alone or together with the -1607 MMP-1 functional polymorphism, increases the likelihood of developing TB in BCG-vaccinated individuals. We conducted population-based case-control studies of BCG-vaccinated individuals in Mexico and Peru that included 193 TB cases and 243 healthy tuberculin-positive controls from Mexico and 701 TB cases and 796 controls from Peru. We also performed immunohistochemistry (IHC analysis of lymph nodes from carriers of relevant two-locus genotypes and in vitro studies to determine how these variants may operate to increase the risk of developing active disease. We report that a joint effect between the -2518 MCP-1 genotype GG and the -1607 MMP-1 genotype 2G/2G consistently increases the odds of developing TB 3.59-fold in Mexicans and 3.9-fold in Peruvians. IHC analysis of lymph nodes indicated that carriers of the two-locus genotype MCP-1 GG MMP-1 2G/2G express the highest levels of both MCP-1 and MMP-1. Carriers of these susceptibility genotypes might be at increased risk of developing TB because they produce high levels of MCP-1, which enhances the induction of MMP-1 production by M. tuberculosis-sonicate antigens to higher levels than in carriers of the other two-locus MCP-1 MMP-1 genotypes studied. This notion was supported by in vitro experiments and luciferase based promoter activity assay. MMP-1 may destabilize granuloma formation and promote tissue damage and disease progression early in the infection. Our findings may foster the development of new and personalized therapeutic approaches targeting MCP-1 and/or MMP-1.

Joint effect of MCP-1 genotype GG and MMP-1 genotype 2G/2G increases the likelihood of developing pulmonary tuberculosis in BCG-vaccinated individuals.

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Ganachari, Malathesha; Ruiz-Morales, Jorge A; Gomez de la Torre Pretell, Juan C; Dinh, Jeffrey; Granados, Julio; Flores-Villanueva, Pedro O

2010-01-25

We previously reported that the -2518 MCP-1 genotype GG increases the likelihood of developing tuberculosis (TB) in non-BCG-vaccinated Mexicans and Koreans. Here, we tested the hypothesis that this genotype, alone or together with the -1607 MMP-1 functional polymorphism, increases the likelihood of developing TB in BCG-vaccinated individuals. We conducted population-based case-control studies of BCG-vaccinated individuals in Mexico and Peru that included 193 TB cases and 243 healthy tuberculin-positive controls from Mexico and 701 TB cases and 796 controls from Peru. We also performed immunohistochemistry (IHC) analysis of lymph nodes from carriers of relevant two-locus genotypes and in vitro studies to determine how these variants may operate to increase the risk of developing active disease. We report that a joint effect between the -2518 MCP-1 genotype GG and the -1607 MMP-1 genotype 2G/2G consistently increases the odds of developing TB 3.59-fold in Mexicans and 3.9-fold in Peruvians. IHC analysis of lymph nodes indicated that carriers of the two-locus genotype MCP-1 GG MMP-1 2G/2G express the highest levels of both MCP-1 and MMP-1. Carriers of these susceptibility genotypes might be at increased risk of developing TB because they produce high levels of MCP-1, which enhances the induction of MMP-1 production by M. tuberculosis-sonicate antigens to higher levels than in carriers of the other two-locus MCP-1 MMP-1 genotypes studied. This notion was supported by in vitro experiments and luciferase based promoter activity assay. MMP-1 may destabilize granuloma formation and promote tissue damage and disease progression early in the infection. Our findings may foster the development of new and personalized therapeutic approaches targeting MCP-1 and/or MMP-1.

Correlation of MMP-9, GA, HbA1c, and adipokines levels with DR

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Cheng Qian

2017-12-01

Full Text Available AIM: To investigate the correlation of matrix metalloproteinase -9(MMP-9, glycated albumin(GA, glycosylated hemoglobin(HbA1cand adipokines(including visfatin, resistin and leptinwith diabetic retinopathy(DR. METHODS: From March 2015 to March 2017, 74 patients with DR were treated in our hospital, including 40 patients(80 eyeswith non proliferative diabetic retinopathy(NPDRand 34 patients(68 eyeswith proliferative diabetic retinopathy(PDR, and diabetes mellitus 40 patients(80 eyeswith non DR(NDRand 40 healthy volunteers(80 eyeswere selected as controls, the levels of MMP-9, GA, HbA1c, visfatin, resistin and leptin in each group were detected. RESULTS: PDR group visfatin was 4.41±0.82ng/mL, was significantly lower than the NPDR group, NDR group and control group(PPPPrs=0.523, 0.461 and 0.414, Prs=-0.433, Prs=0.401 and 0.460, PCONCLUSION: MMP-9, GA, HbA1c, and adipokines may play a role in the development and progression of DR, in which MMP-9 is associated with adipokines, both are not significantly related to the levels of GA and HbA1c.

A novel cell line derived from pleomorphic adenoma expresses MMP2, MMP9, TIMP1, TIMP2, and shows numeric chromosomal anomalies.

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Aline Semblano Carreira Falcão

Full Text Available Pleomorphic adenoma is the most common salivary gland neoplasm, and it can be locally invasive, despite its slow growth. This study aimed to establish a novel cell line (AP-1 derived from a human pleomorphic adenoma sample to better understand local invasiveness of this tumor. AP-1 cell line was characterized by cell growth analysis, expression of epithelial and myoepithelial markers by immunofluorescence, electron microscopy, 3D cell culture assays, cytogenetic features and transcriptomic study. Expression of matrix metalloproteinases (MMPs and their tissue inhibitors (TIMPs was also analyzed by immunofluorescence and zymography. Furthermore, epithelial and myoepithelial markers, MMPs and TIMPs were studied in the tumor that originated the cell line. AP-1 cells showed neoplastic epithelial and myoepithelial markers, such as cytokeratins, vimentin, S100 protein and smooth-muscle actin. These molecules were also found in vivo, in the tumor that originated the cell line. MMPs and TIMPs were observed in vivo and in AP-1 cells. Growth curve showed that AP-1 exhibited a doubling time of 3.342 days. AP-1 cells grown inside Matrigel recapitulated tumor architecture. Different numerical and structural chromosomal anomalies were visualized in cytogenetic analysis. Transcriptomic analysis addressed expression of 7 target genes (VIM, TIMP2, MMP2, MMP9, TIMP1, ACTA2 e PLAG1. Results were compared to transcriptomic profile of non-neoplastic salivary gland cells (HSG. Only MMP9 was not expressed in both libraries, and VIM was expressed solely in AP-1 library. The major difference regarding gene expression level between AP-1 and HSG samples occurred for MMP2. This gene was 184 times more expressed in AP-1 cells. Our findings suggest that AP-1 cell line could be a useful model for further studies on pleomorphic adenoma biology.

Ancient mtDNA genetic variants modulate mtDNA transcription and replication.

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Sarit Suissa

2009-05-01

Full Text Available Although the functional consequences of mitochondrial DNA (mtDNA genetic backgrounds (haplotypes, haplogroups have been demonstrated by both disease association studies and cell culture experiments, it is not clear which of the mutations within the haplogroup carry functional implications and which are "evolutionary silent hitchhikers". We set forth to study the functionality of haplogroup-defining mutations within the mtDNA transcription/replication regulatory region by in vitro transcription, hypothesizing that haplogroup-defining mutations occurring within regulatory motifs of mtDNA could affect these processes. We thus screened >2500 complete human mtDNAs representing all major populations worldwide for natural variation in experimentally established protein binding sites and regulatory regions comprising a total of 241 bp in each mtDNA. Our screen revealed 77/241 sites showing point mutations that could be divided into non-fixed (57/77, 74% and haplogroup/sub-haplogroup-defining changes (i.e., population fixed changes, 20/77, 26%. The variant defining Caucasian haplogroup J (C295T increased the binding of TFAM (Electro Mobility Shift Assay and the capacity of in vitro L-strand transcription, especially of a shorter transcript that maps immediately upstream of conserved sequence block 1 (CSB1, a region associated with RNA priming of mtDNA replication. Consistent with this finding, cybrids (i.e., cells sharing the same nuclear genetic background but differing in their mtDNA backgrounds harboring haplogroup J mtDNA had a >2 fold increase in mtDNA copy number, as compared to cybrids containing haplogroup H, with no apparent differences in steady state levels of mtDNA-encoded transcripts. Hence, a haplogroup J regulatory region mutation affects mtDNA replication or stability, which may partially account for the phenotypic impact of this haplogroup. Our analysis thus demonstrates, for the first time, the functional impact of particular mt

PER, a Circadian Clock Component, Mediates the Suppression of MMP-1 Expression in HaCaT Keratinocytes by cAMP.

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Yeom, Miji; Lee, HansongI; Shin, Seoungwoo; Park, Deokhoon; Jung, Eunsun

2018-03-23

Skin circadian clock system responds to daily changes, thereby regulating skin functions. Exposure of the skin to UV irradiation induces the expression of matrix metalloproteinase-1 (MMP-1) and causes DNA damage. It has been reported both DNA repair and DNA replication are regulated by the circadian clock in mouse skin. However, the molecular link between circadian clock and MMP-1 has little been investigated. We found PERIOD protein, a morning clock component, represses the expression of MMP-1 in human keratinocytes by using a PER-knockdown strategy. Treatment with siPer3 alleviated the suppression of MMP-1 expression induced by forskolin. Results revealed PER3 suppresses the expression of MMP-1 via cAMP signaling pathway. Additionally, we screened for an activator of PER that could repress the expression of MMP-1 using HaCaT cell line containing PER promoter-luciferase reporter gene. Results showed Lespedeza capitate extract (LCE) increased PER promoter activity. LCE inhibited the expression of MMP-1 and its effect of LCE was abolished in knockdown of PER2 or PER3, demonstrating LCE can repress the expression of MMP-1 through PER. Since circadian clock component PER can regulate MMP-1 expression, it might be a new molecular mechanism to develop therapeutics to alleviate skin aging and skin cancer.

PER, a Circadian Clock Component, Mediates the Suppression of MMP-1 Expression in HaCaT Keratinocytes by cAMP

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Miji Yeom

2018-03-01

Full Text Available Skin circadian clock system responds to daily changes, thereby regulating skin functions. Exposure of the skin to UV irradiation induces the expression of matrix metalloproteinase-1 (MMP-1 and causes DNA damage. It has been reported both DNA repair and DNA replication are regulated by the circadian clock in mouse skin. However, the molecular link between circadian clock and MMP-1 has little been investigated. We found PERIOD protein, a morning clock component, represses the expression of MMP-1 in human keratinocytes by using a PER-knockdown strategy. Treatment with siPer3 alleviated the suppression of MMP-1 expression induced by forskolin. Results revealed PER3 suppresses the expression of MMP-1 via cAMP signaling pathway. Additionally, we screened for an activator of PER that could repress the expression of MMP-1 using HaCaT cell line containing PER promoter-luciferase reporter gene. Results showed Lespedeza capitate extract (LCE increased PER promoter activity. LCE inhibited the expression of MMP-1 and its effect of LCE was abolished in knockdown of PER2 or PER3, demonstrating LCE can repress the expression of MMP-1 through PER. Since circadian clock component PER can regulate MMP-1 expression, it might be a new molecular mechanism to develop therapeutics to alleviate skin aging and skin cancer.

Extracellular matrix proteins matrix metallopeptidase 9 (MMP9) and soluble intercellular adhesion molecule 1 (sICAM-1) and correlations with clinical staging in euthymic bipolar disorder.

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Reininghaus, Eva Z; Lackner, Nina; Birner, Armin; Bengesser, Susanne; Fellendorf, Frederike T; Platzer, Martina; Rieger, Alexandra; Queissner, Robert; Kainzbauer, Nora; Reininghaus, Bernd; McIntyre, Roger S; Mangge, Harald; Zelzer, Sieglinde; Fuchs, Dietmar; Dejonge, Silvia; Müller, Norbert

2016-03-01

Matrix metallopeptidase 9 (MMP9) and soluble intercellular adhesion molecule 1 (sICAM-1) are both involved in the restructuring of connective tissues. Evidence also implicates MMP9 and sICAM in cardiovascular and neoplastic diseases, where blood levels may be a marker of disease severity or prognosis. In individuals with bipolar disorder (BD), higher risk for cardiovascular illness has been extensively reported. The aim of this investigation was to measure and compare peripheral levels of serum MMP9 and sICAM in adults with euthymic BD and healthy controls (HC). Furthermore, we focussed on correlations with illness severity and metabolic parameters. MMP9 levels among the BD sample (n = 112) were significantly higher than among the HC (n = 80) (MMP9: F = 9.885, p = 0.002, η(2)  = 0.058) after controlling for confounding factors. Patients with BD in a later, progressive stage of disease showed significantly higher MMP9 as well as sICAM-1 levels compared to patients with BD in an earlier stage of disease (MMP9: F = 5.8, p = 0.018, η(2)  = 0.054; sICAM-1: F = 5.6, p = 0.020, η(2)  = 0.052). Correlation analyses of cognitive measures revealed a negative association between performance on the d2 Test of Attention and MMP9 (r = -0.287, p = 0.018) in the BD sample. Despite the sample being euthymic (i.e., according to conventional criteria) at the time of analysis, we found significant correlations between MMP9 as well as sICAM-1 and subthreshold depressive/hypomanic symptoms. A collection of disparate findings herein point to a role of MMP9 and cICAM-1 in the patho-progressive process of BD: the increased levels of serum MMP9 and sICAM-1, the correlation between higher levels of these parameters, progressive stage, and cognitive dysfunction in BD, and the positive correlation with subthreshold symptoms. As sICAM-1 and MMP9 are reliable biomarkers of inflammatory and early atherosclerotic disease, these markers may provide indications of the

Bmi-1 promotes the aggressiveness of glioma via activating the NF-kappaB/MMP-9 signaling pathway

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Jiang, Lili; Wu, Jueheng; Yang, Yi; Liu, Liping; Song, Libing; Li, Jun; Li, Mengfeng

2012-01-01

The prognosis of human glioma is poor, and the highly invasive nature of the disease represents a major impediment to current therapeutic modalities. The oncoprotein B-cell-specific Moloney murine leukemia virus integration site 1 protein (Bmi-1) has been linked to the development and progression of glioma; however, the biological role of Bmi-1 in the invasion of glioma remains unclear. A172 and LN229 glioma cells were engineered to overexpress Bmi-1 via stable transfection or to be silenced for Bmi-1 expression using RNA interfering method. Migration and invasiveness of the engineered cells were assessed using wound healing assay, Transwell migration assay, Transwell matrix penetration assay and 3-D spheroid invasion assay. MMP-9 expression and activity were measured using real-time PCR, ELISA and the gelatin zymography methods. Expression of NF-kappaB target genes was quantified using real-time PCR. NF-kappaB transcriptional activity was assessed using an NF-kappaB luciferase reporter system. Expression of Bmi-1 and MMP-9 in clinical specimens was analyzed using immunohistochemical assay. Ectopic overexpression of Bmi-1 dramatically increased, whereas knockdown of endogenous Bmi-1 reduced, the invasiveness and migration of glioma cells. NF-kappaB transcriptional activity and MMP-9 expression and activity were significantly increased in Bmi-1-overexpressing but reduced in Bmi-1-silenced cells. The reporter luciferase activity driven by MMP-9 promoter in Bmi-1-overexpressing cells was dependent on the presence of a functional NF-kappaB binding site, and blockade of NF-kappaB signaling inhibited the upregulation of MMP-9 in Bmi-1 overexpressing cells. Furthermore, expression of Bmi-1 correlated with NF-kappaB nuclear translocation as well as MMP-9 expression in clinical glioma samples. Bmi-1 may play an important role in the development of aggressive phenotype of glioma via activating the NF-kappaB/MMP-9 pathway and therefore might represent a novel therapeutic

Bmi-1 promotes the aggressiveness of glioma via activating the NF-kappaB/MMP-9 signaling pathway

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Jiang Lili

2012-09-01

Full Text Available Abstract Background The prognosis of human glioma is poor, and the highly invasive nature of the disease represents a major impediment to current therapeutic modalities. The oncoprotein B-cell-specific Moloney murine leukemia virus integration site 1 protein (Bmi-1 has been linked to the development and progression of glioma; however, the biological role of Bmi-1 in the invasion of glioma remains unclear. Methods A172 and LN229 glioma cells were engineered to overexpress Bmi-1 via stable transfection or to be silenced for Bmi-1 expression using RNA interfering method. Migration and invasiveness of the engineered cells were assessed using wound healing assay, Transwell migration assay, Transwell matrix penetration assay and 3-D spheroid invasion assay. MMP-9 expression and activity were measured using real-time PCR, ELISA and the gelatin zymography methods. Expression of NF-kappaB target genes was quantified using real-time PCR. NF-kappaB transcriptional activity was assessed using an NF-kappaB luciferase reporter system. Expression of Bmi-1 and MMP-9 in clinical specimens was analyzed using immunohistochemical assay. Results Ectopic overexpression of Bmi-1 dramatically increased, whereas knockdown of endogenous Bmi-1 reduced, the invasiveness and migration of glioma cells. NF-kappaB transcriptional activity and MMP-9 expression and activity were significantly increased in Bmi-1-overexpressing but reduced in Bmi-1-silenced cells. The reporter luciferase activity driven by MMP-9 promoter in Bmi-1-overexpressing cells was dependent on the presence of a functional NF-kappaB binding site, and blockade of NF-kappaB signaling inhibited the upregulation of MMP-9 in Bmi-1 overexpressing cells. Furthermore, expression of Bmi-1 correlated with NF-kappaB nuclear translocation as well as MMP-9 expression in clinical glioma samples. Conclusions Bmi-1 may play an important role in the development of aggressive phenotype of glioma via activating the NF-kappaB/MMP

1-L-MT, an IDO inhibitor, prevented colitis-associated cancer by inducing CDC20 inhibition-mediated mitotic death of colon cancer cells.

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Liu, Xiuting; Zhou, Wei; Zhang, Xin; Ding, Yang; Du, Qianming; Hu, Rong

2018-04-01

Indoleamine 2,3-dioxygenase 1 (IDO1), known as IDO, catabolizes tryptophan through kynurenine pathway, whose activity is correlated with impaired clinical outcome of colorectal cancer. Here we showed that 1-L-MT, a canonical IDO inhibitor, suppressed proliferation of human colorectal cancer cells through inducing mitotic death. Our results showed that inhibition of IDO decreased the transcription of CDC20, which resulted in G2/M cycle arrest of HCT-116 and HT-29. Furthermore, 1-L-MT induced mitochondria injuries and caused apoptotic cancer cells. Importantly, 1-L-MT protected mice from azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced colon carcinogenesis, with reduced mortality, tumor number and size. What is more, IDO1-/- mice exhibited fewer tumor burdens and reduced proliferation in the neoplastic epithelium, while, 1-L-MT did not exhibit any further protective effects on IDO-/- mice, confirming the critical role of IDO and the protective effect of 1-L-MT-mediated IDO inhibition in CRC. Furthermore, 1-L-MT also alleviated CRC in Rag1-/- mice, demonstrating the modulatory effects of IDO independent of its role in modulating adaptive immunity. Taken together, our findings validated that the anti-proliferation effect of 1-L-MT in vitro and the prevention of CRC in vivo were through IDO-induced cell cycle disaster of colon cancer cells. Our results identified 1-L-MT as a promising candidate for the chemoprevention of CRC. © 2018 UICC.

Expression of melatonin receptor MT1 in cells of human invasive ductal breast carcinoma.

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Jablonska, Karolina; Pula, Bartosz; Zemla, Agata; Owczarek, Tomasz; Wojnar, Andrzej; Rys, Janusz; Ambicka, Aleksandra; Podhorska-Okolow, Marzena; Ugorski, Maciej; Dziegiel, Piotr

2013-04-01

In humans, two main types of membrane melatonin receptors have been identified, MT1 and MT2. Expression of MT1 in neoplastic cells seems to increase the efficacy of melatonin's oncostatic activity. The purpose of this study was to determine the distribution and the intensity of MT1 expression in breast cancer cells and to correlate it with clinicopathological factors. Immunohistochemical studies (IHC) were conducted on 190 cases of invasive ductal breast carcinomas (IDC) and molecular studies were performed on 29 cases of frozen tumor fragments and selected breast cancer cell lines. Most of the studied tumors manifested a membranous/cytoplasmic IHC expression of MT1. In IDC, the MT1 expression was higher than in fibrocystic breast disease. MT1 expression was higher in estrogen receptor positive (ER+) and HER2 positive (HER2+) tumors. Triple negative tumors (TN) manifested the lowest MT1 expression level. The lowest MT1 protein expression level was noted in the TN breast cancer cell line MDA-MB-231 compared with ER+ cell lines MCF-7 and SK-BR-3. MT1 mRNA expression was negatively correlated with the malignancy grade of the studied IDC cases. Moreover, higher MT1 expression was associated with patients' longer overall survival (OS) in the group of ER+ breast cancers and treated with tamoxifen. Multivariate analysis indicated that MT1 was an independent prognostic factor in the ER+ tumors for OS and event-free survival in the ER+ tumors. The results of this study may point to a potential prognostic and therapeutic significance of MT1 in IDC. © 2013 John Wiley & Sons A/S. Published by Blackwell Publishing Ltd.

Regeneration of soft tissues is promoted by MMP1 treatment after digit amputation in mice.

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Xiaodong Mu

Full Text Available The ratio of matrix metalloproteinases (MMPs to the tissue inhibitors of metalloproteinases (TIMPs in wounded tissues strictly control the protease activity of MMPs, and therefore regulate the progress of wound closure, tissue regeneration and scar formation. Some amphibians (i.e. axolotl/newt demonstrate complete regeneration of missing or wounded digits and even limbs; MMPs play a critical role during amphibian regeneration. Conversely, mammalian wound healing re-establishes tissue integrity, but at the expense of scar tissue formation. The differences between amphibian regeneration and mammalian wound healing can be attributed to the greater ratio of MMPs to TIMPs in amphibian tissue. Previous studies have demonstrated the ability of MMP1 to effectively promote skeletal muscle regeneration by favoring extracellular matrix (ECM remodeling to enhance cell proliferation and migration. In this study, MMP1 was administered to the digits amputated at the mid-second phalanx of adult mice to observe its effect on digit regeneration. Results indicated that the regeneration of soft tissue and the rate of wound closure were significantly improved by MMP1 administration, but the elongation of the skeletal tissue was insignificantly affected. During digit regeneration, more mutipotent progenitor cells, capillary vasculature and neuromuscular-related tissues were observed in MMP1 treated tissues; moreover, there was less fibrotic tissue formed in treated digits. In summary, MMP1 was found to be effective in promoting wound healing in amputated digits of adult mice.

Comparison of circulating MMP-9, TIMP-1 and CA19-9 in the detection of pancreatic cancer

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Jørgensen, Maiken Thyregod; Brunner, Nils; Schaffalitzky de Muckadell, Ove B.

2010-01-01

, TIMP-1 and CA19-9 in detecting pancreatic ductal adenocarcinoma were 58.82%, 47.1% and 86%, respectively, with specificities of 34.6%, 69.2% and 73%. The AUCs of MMP-9, TIMP-1 and CA19-9 were 0.50, 0.64 and 0.84, respectively. Combining the three markers did not significantly improve detection......Background/Aim: The performance of the circulating tumor markers carbohydrate antigen 19-9 (CA19-9), matrix metalloproteinase 9 (MMP-9) and tissue inhibitor of metalloproteinase 1 (TIMP-1) were evaluated separately and in combination for their potential value in detecting pancreatic ductal...... adenocarcinoma. PATIENTS AND METHODS: The patients had symptoms of pancreatic cancer. The discriminative strength of MMP-9 and TIMP-1 were compared to that of CA19-9 using receiver operating characteristics curves, area under the curves (AUC), specificity and sensitivity. RESULTS: The sensitivities of MMP-9...

Matrix metalloproteinase-10 (MMP-10) interaction with tissue inhibitors of metalloproteinases TIMP-1 and TIMP-2: binding studies and crystal structure.

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Batra, Jyotica; Robinson, Jessica; Soares, Alexei S; Fields, Alan P; Radisky, Derek C; Radisky, Evette S

2012-05-04

Matrix metalloproteinase 10 (MMP-10, stromelysin-2) is a secreted metalloproteinase with functions in skeletal development, wound healing, and vascular remodeling; its overexpression is also implicated in lung tumorigenesis and tumor progression. To understand the regulation of MMP-10 by tissue inhibitors of metalloproteinases (TIMPs), we have assessed equilibrium inhibition constants (K(i)) of putative physiological inhibitors TIMP-1 and TIMP-2 for the active catalytic domain of human MMP-10 (MMP-10cd) using multiple kinetic approaches. We find that TIMP-1 inhibits the MMP-10cd with a K(i) of 1.1 × 10(-9) M; this interaction is 10-fold weaker than the inhibition of the similar MMP-3 (stromelysin-1) catalytic domain (MMP-3cd) by TIMP-1. TIMP-2 inhibits the MMP-10cd with a K(i) of 5.8 × 10(-9) M, which is again 10-fold weaker than the inhibition of MMP-3cd by this inhibitor (K(i) = 5.5 × 10(-10) M). We solved the x-ray crystal structure of TIMP-1 bound to the MMP-10cd at 1.9 Å resolution; the structure was solved by molecular replacement and refined with an R-factor of 0.215 (R(free) = 0.266). Comparing our structure of MMP-10cd·TIMP-1 with the previously solved structure of MMP-3cd·TIMP-1 (Protein Data Bank entry 1UEA), we see substantial differences at the binding interface that provide insight into the differential binding of stromelysin family members to TIMP-1. This structural information may ultimately assist in the design of more selective TIMP-based inhibitors tailored for specificity toward individual members of the stromelysin family, with potential therapeutic applications.

Oral administration of curcumin suppresses production of matrix metalloproteinase (MMP)-1 and MMP-3 to ameliorate collagen-induced arthritis: inhibition of the PKCdelta/JNK/c-Jun pathway.

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Mun, Se Hwan; Kim, Hyuk Soon; Kim, Jie Wan; Ko, Na Young; Kim, Do Kyun; Lee, Beob Yi; Kim, Bokyung; Won, Hyung Sik; Shin, Hwa-Sup; Han, Jeung-Whan; Lee, Hoi Young; Kim, Young Mi; Choi, Wahn Soo

2009-09-01

We investigated whether oral administration of curcumin suppressed type II collagen-induced arthritis (CIA) in mice and its effect and mechanism on matrix metalloproteinase (MMP)-1 and MMP-3 production in CIA mice, RA fibroblast-like synoviocytes (FLS), and chondrocytes. CIA in mice was suppressed by oral administration of curcumin in a dose-dependent manner. Macroscopic observations were confirmed by histological examinations. Histological changes including infiltration of immune cells, synovial hyperplasia, cartilage destruction, and bone erosion in the hind paw sections were extensively suppressed by curcumin. The histological scores were consistent with clinical arthritis indexes. Production of MMP-1 and MMP-3 were inhibited by curcumin in CIA hind paw sections and tumor necrosis factor (TNF)-alpha-stimulated FLS and chondrocytes in a dose-dependent manner. As for the mechanism, curcumin inhibited activating phosphorylation of protein kinase Cdelta (PKCdelta) in CIA, FLS, and chondrocytes. Curcumin also suppressed the JNK and c-Jun activation in those cells. This study suggests that the suppression of MMP-1 and MMP-3 production by curcumin in CIA is mediated through the inhibition of PKCdelta and the JNK/c-Jun signaling pathway.

Regulation of Membrane-Type 4 Matrix Metalloproteinase by SLUG Contributes to Hypoxia-Mediated Metastasis12

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Huang, Chi-Hung; Yang, Wen-Hao; Chang, Shyue-Yih; Tai, Shyh-Kuan; Tzeng, Cheng-Hwei; Kao, Jung-Yie; Wu, Kou-Juey; Yang, Muh-Hwa

2009-01-01

The hypoxic tumor environment has been shown to be critical to cancer metastasis through the promotion of angiogenesis, induction of epithelial-mesenchymal transition (EMT), and acquisition of invasive potential. However, the impact of hypoxia on the expression profile of the proteolytic enzymes involved in invasiveness is relatively unknown. Membrane-type 4 matrix metalloproteinase (MT4-MMP) is a glycosyl-phosphatidyl inositol-anchored protease that has been shown to be overexpressed in human cancers. However, detailed mechanisms regarding the regulation and function of MT4-MMP expression in tumor cells remain unknown. Here, we demonstrate that hypoxia or overexpression of hypoxia-inducible factor-1α (HIF-1α) induced MT4-MMP expression in human cancer cells. Activation of SLUG, a transcriptional factor regulating the EMT process of human cancers, by HIF-1α was critical for the induction of MT4-MMP under hypoxia. SLUG regulated the transcription of MT4-MMP through direct binding to the E-box located in its proximal promoter. Short-interference RNA-mediated knockdown of MT4-MMP attenuated in vitro invasiveness and in vivo pulmonary colonization of tumor cells without affecting cell migratory ability. MT4-MMP promoted invasiveness and pulmonary colonization through modulation of the expression profile of MMPs and angiogenic factors. Finally, coexpression of HIF-1α and MT4-MMP in human head and neck cancer was predictive of a worse clinical outcome. These findings establish a novel signaling pathway for hypoxia-mediated metastasis and elucidate the underlying regulatory mechanism and functional significance of MT4-MMP in cancer metastasis. PMID:20019845

Regulation of Membrane-Type 4 Matrix Metalloproteinase by SLUG Contributes to Hypoxia-Mediated Metastasis

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Chi-Hung Huang

2009-12-01

Full Text Available The hypoxic tumor environment has been shown to be critical to cancer metastasis through the promotion of angiogenesis, induction of epithelial-mesenchymal transition (EMT, and acquisition of invasive potential. However, the impact of hypoxia on the expression profile of the proteolytic enzymes involved in invasiveness is relatively unknown. Membrane-type 4 matrix metalloproteinase (MT4-MMP is a glycosyl-phosphatidyl inositol-anchored protease that has been shown to be overexpressed in human cancers. However, detailed mechanisms regarding the regulation and function of MT4-MMP expression in tumor cells remain unknown. Here, we demonstrate that hypoxia or overexpression of hypoxia-inducible factor-1α (HIF-1α induced MT4-MMP expression in human cancer cells. Activation of SLUG, a transcriptional factor regulating the EMT process of human cancers, by HIF-1α was critical for the induction of MT4-MMP under hypoxia. SLUG regulated the transcription of MT4-MMP through direct binding to the E-box located in its proximal promoter. Short-interference RNA-mediated knockdown of MT4-MMP attenuated in vitro invasiveness and in vivo pulmonary colonization of tumor cells without affecting cell migratory ability. MT4-MMP promoted invasiveness and pulmonary colonization through modulation of the expression profile of MMPs and angiogenic factors. Finally, coexpression of HIF-1α and MT4-MMP in human head and neck cancer was predictive of a worse clinical outcome. These findings establish a novel signaling pathway for hypoxia-mediated metastasis and elucidate the underlying regulatory mechanism and functional significance of MT4-MMP in cancer metastasis.

MMP-1 is a (pre-invasive factor in Barrett-associated esophageal adenocarcinomas and is associated with positive lymph node status

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Otto Christoph

2010-10-01

Full Text Available Abstract Background Esophageal adenocarcinomas (EACs arise due to gastroesophageal reflux, with Barrett's esophagus (BE regarded as precancerous lesion. Matrix metalloproteinases (MMPs might play a role during the multistep carcinogenetic process. Methods Expression of MMP-1 and -13 was analyzed in esophageal cancer (n = 41 EAC with BE, n = 19 EAC without BE, and n = 10 esophageal squamous-cell carcinomas, ESCC, furthermore in BE without intraepithelial neoplasia (IN (n = 18, and the cell line OE-33. MMP-1 was co-labelled with Ki-67 (proliferation, Cdx-2 (marker for intestinal metaplasia, BE and analyzed on mRNA level. MMP-1 staining results were correlated with clinicopatholocical parameters. Results On protein level, MMP-1 expression was found in 39 of 41 (95% EAC with BE, in 19 of 19 (100% EAC without BE, in 6 of 10 (60% ESCC, and in 10 of 18 (56% BE without IN. No expression of MMP-13 was found in these specimens. Quantification showed 48% MMP-1 positive cells in EAC with BE, compared to 35% in adjacent BE (p Conclusions Our findings suggest that MMP-1 plays a role as preinvasive factor in BE-associated EAC. Expression of MMP-1 in proliferating BE and EAC cells suggest malignant proliferation following the clonal expansion model.

MRI growth patterns of plexiform neurofibromas in patients with neurofibromatosis type 1

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Mautner, V.F.; Hartmann, M.; Kluwe, L.; Friedrich, R.E. [University Hospital Eppendorf, Section for Phakomatoses, Department of Maxillofacial Surgery, Hamburg (Germany); Fuensterer, C. [MRI Institute Hamburg-Othmarschen, Hamburg (Germany)

2006-03-15

Neurofibromatosis type 1 (NF1) is an autosomal dominant disorder with an incidence of 1:3000. Approximately 30% of NF1 patients develop plexiform neurofibromas (PNF) which often cause severe clinical deficits. We studied the growth patterns of 256 plexiform neurofibromas (PNF) by magnetic resonance imaging (MRI) and associated disfigurement and functional deficits to determine whether there are definable growth types of these tumors. Retrospectively, we evaluated MRI scans obtained during 1997 to 2003 of 256 plexiform neurofibromas from 202 patients with NF1. Clinical investigation was carried out at the same time as the MRI scans. We identified three growth patterns: superficial in 59, displacing in 76, and invasive growth in 121 tumors. The majority (52%) of invasive PNF were found in the face, head and neck area. While superficial PNF primarily caused aesthetic problems, displacing PNF led in most cases to aesthetic problems and pain, while invasive PNF led mainly to functional deficits and disfigurement. Our study demonstrates that PNF have different growth patterns that are associated with specific clinical features. Classification of PNF may open new opportunities in clinical management, especially regarding decisions and options associated with surgical intervention. (orig.)

Fraxinus rhynchophylla ethanol extract attenuates carbon tetrachloride-induced liver fibrosis in rats via down-regulating the expressions of uPA, MMP-2, MMP-9 and TIMP-1.

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Peng, Wen-Huang; Tien, Yun-Chen; Huang, Chih-Yang; Huang, Tai-Hung; Liao, Jung-Chun; Kuo, Chao-Lin; Lin, Ying-Chih

2010-02-17

To investigate the effect of Fraxinus rhynchophylla ethanol extract (FR(EtOH)) on liver fibrosis induced by carbon tetrachloride (CCl(4)) in rats. Rat hepatic fibrosis was induced by oral administration of CCl(4). Sixty SD rats were divided randomly into 6 groups: control, CCl(4) group, silymarin group and three FR(EtOH)-treated groups. Except for the rats in control group, all rats were administered orally with CCl(4) (20%, 0.2 mL/100g body weight) twice a week for 8 weeks. Rats in FR(EtOH) groups were treated daily with FR(EtOH) (0.1, 0.5 and 1.0 g/kg, p.o.) throughout the whole experimental period. Liver function parameters (such as activities of serum GOT and GPT levels), activities of liver anti-oxidant enzymes (such as catalase, SOD, GPx) and expressions of uPA, tPA, MMP-2, MMP-9 and TIMP-1, -2, -3, -4 in the liver fibrosis pathway were detected. The results showed that FR(EtOH) (0.1, 0.5 and 1.0 g/kg BW) significantly reduced the elevated activities of sGOT and sGPT caused by CCl(4). FR(EtOH) (0.1 and 0.5 g/kg BW) and significantly increased the activities of GSH-Px. The histopathological study showed that FR(EtOH) (0.1 and 0.5 g/kg BW) reduced the incidence of liver lesions, including hepatic cells cloudy swelling, lymphocytes infiltration, cytoplasm vacuolization hepatic necrosis and fibrous connective tissue proliferated induced by CCl(4) in rats. In our study it was showed that CCl(4)-treated group significantly increased the protein levels of uPA, MMP-2, MMP-9 and TIMP-1. FR(EtOH) (0.1 and 0.5 g/kg BW) could inhibit the protein levels of uPA, MMP-2, MMP-9 and TIMP-1. Finally, the amount of esculetin in the FR(EtOH) was 33.54 mg/g extract. Oral administration of FR(EtOH) significantly reduces CCl(4)-induced hepatic fibrosis in rats, probably by exerting a protective effect against hepatocellular fibrosis by its free radical scavenging ability. FR(EtOH) down-regulated the expressions of uPA, MMP-2 and MMP-9 in CCl(4)-induced liver fibrosis in rats

New Radioligands for Describing the Molecular Pharmacology of MT1 and MT2 Melatonin Receptors

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Olivier Nosjean

2013-04-01

Full Text Available Melatonin receptors have been studied for several decades. The low expression of the receptors in tissues led the scientific community to find a substitute for the natural hormone melatonin, the agonist 2-[125I]-iodomelatonin. Using the agonist, several hundreds of studies were conducted, including the discovery of agonists and antagonists for the receptors and minute details about their molecular behavior. Recently, we attempted to expand the panel of radioligands available for studying the melatonin receptors by using the newly discovered compounds SD6, DIV880, and S70254. These compounds were characterized for their affinities to the hMT1 and hMT2 recombinant receptors and their functionality in the classical GTPS system. SD6 is a full agonist, equilibrated between the receptor isoforms, whereas S70254 and DIV880 are only partial MT2 agonists, with Ki in the low nanomolar range while they have no affinity to MT1 receptors. These new tools will hopefully allow for additions to the current body of information on the native localization of the receptor isoforms in tissues.

Immunohistochemical analysis of MMP-9, MMP-2 and TIMP-1, TIMP-2 expression in the central nervous system following infection with viral and bacterial meningitis.

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Lech Chyczewski

2009-01-01

Full Text Available Matrix metalloproteinases (MMPs are capable of degrading components of the basal lamina of cerebral vessels, thereby disrupting the blood-brain barrier and inducing leukocyte recruitment. This study provides comprehensive information regarding the cell specificity of matrix metalloproteinases (MMP-2, MMP-9 and their binding tissue inhibitors (TIMP-1, TIMP-2 in the central nervous system during viral and bacterial meningitis. Specifically, we evaluated the immunoreactivity of MMPs and TIMPs in various cell types in brain parenchyma and meninges obtained from autopsy tissues. We found that a higher proportion of endothelial cells were positive for MMP-9 during meningitis when compared to controls. In addition, the immunoreactivity of MMP-9 decreased and the immunoreactivity of TIMP-1 increased in astrocytes upon infection. Furthermore, the results of this study revealed that mononuclear cells were highly immunoreactive for TIMP-1, TIMP-2 and MMP-9 during viral meningitis and that the expression of TIMPs in polymorphonuclear cells was even higher during bacterial meningitis. Taken together the results of this study indicated that the central nervous system resident cells and inflammatory infiltrates contribute to MMPs activity and that the expression patterns vary between cell types and in response to viral and bacterial meningitis.

DNA methyltransferase 1 mutations and mitochondrial pathology: is mtDNA methylated?

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Alessandra eMaresca

2015-03-01

Full Text Available Autosomal dominant cerebellar ataxia, deafness and narcolepsy (ADCA-DN and Hereditary sensory neuropathy with dementia and hearing loss (HSN1E are two rare, overlapping neurodegenerative syndromes that have been recently linked to allelic dominant pathogenic mutations in the DNMT1 gene, coding for DNA (cytosine-5-methyltransferase 1. DNMT1 is the enzyme responsible for maintaining the nuclear genome methylation patterns during the DNA replication and repair, thus regulating gene expression. The mutations responsible for ADCA-DN and HSN1E affect the replication foci targeting sequence domain, which regulates DNMT1 binding to chromatin. DNMT1 dysfunction is anticipated to lead to a global alteration of the DNA methylation pattern with predictable downstream consequences on gene expression. Interestingly, ADCA-DN and HSN1E phenotypes share some clinical features typical of mitochondrial diseases, such as optic atrophy, peripheral neuropathy and deafness, and some biochemical evidence of mitochondrial dysfunction. The recent discovery of a mitochondrial isoform of DNMT1 and its proposed role in methylating mitochondrial DNA (mtDNA suggests that DNMT1 mutations may directly affect mtDNA and mitochondrial physiology. On the basis of this latter finding the link between DNMT1 abnormal activity and mitochondrial dysfunction in ADCA-DN and HSN1E appears intuitive, however mtDNA methylation remains highly debated. In the last years several groups demonstrated the presence of 5-methylcytosine in mtDNA by different approaches, but, on the other end, the opposite evidence that mtDNA is not methylated has also been published. Since over 1500 mitochondrial proteins are encoded by the nuclear genome, the altered methylation of these genes may well have a critical role in leading to the mitochondrial impairment observed in ADCA-DN and HSN1E. Thus, many open questions still remain unanswered, such as why mtDNA should be methylated, and how this process is

Assessment of apoptosis and MMP-1, MMP-3 and TIMP-2 expression in tibial hyaline cartilage after viable medial meniscus transplantation in the rabbit.

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Zwierzchowski, Tomasz J; Stasikowska-Kanicka, Olga; Danilewicz, Marian; Fabiś, Jarosław

2012-12-20

The porpuse of this animal study was to assess chondrocyte apoptosis and MMP-1, MMP-3 and TIMP-2 expression in rabbit tibial cartilage 6 months after viable medial meniscal autografts and allografts. Twenty white male New Zealand rabbits were chosen for the study. The medial meniscus was excised from 14 animals and stored under tissue culture conditions for 2 weeks, following which t of them were implantated as autografts and 7 as allografts. The control group consisted of 6 animals which underwent arthtrotomy. When the animals were eutanized, the tibial cartilage was used for immunohisochemical examination. Apoptosis (TUNEL method) and MMP-1, MMP-3 and TIMP-2 expression were estimated semiquantatively. An increased level of chodrocyte apoptosis in the tibail cartilage was observed after both kinds of transplants (p hyaline cartilage against excessive apoptosis. The results of experimantal studies on humans indicate the need to device a method of apoptosis inhibition in the hyaline cartilage to improve long-term results of meniscal transplantation.

MMP-2 participates in the sclera of guinea pig with form-deprivation myopia via IGF-1/STAT3 pathway.

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Liu, Y-X; Sun, Y

2018-05-01

To investigate the expression changes of MMP-2 (matrix metalloproteinases-2) mediated by IGF-1 (insulin-like growth factors-1) STAT3 (signal transducer and activator of transcription 3) pathway in the sclera of the form-deprivation myopia guinea pigs. Twenty-four three-week-old guinea pigs were randomly divided into 4 groups: group A (Control), B, C and D. Guinea pigs in group A were sacrificed after 21 days without any special treatment. Guinea pigs in group B were sacrificed 7 days after receiving stitch in the right eye. Guinea pigs in group C were sacrificed 14 days after receiving stitch in the right eye. Guinea pigs in group D were sacrificed 21 days after receiving stitch in the right eye. Eyeball refraction and axial length of guinea pigs were measured before sacrifice. Eyeballs of guinea pigs were enucleated after sacrifice. The expressions of IGF-1, STAT3 and MMP-2 in scleral tissue were detected by Western blot. Axial length extension and myopia appeared in the right eye of guinea pigs in group B. The expressions of IGF-1, STAT3 and MMP-2 in the sclera significantly increased after 7 days of occlusion compared with that in control group A (pIGF-1, STAT3 and MMP-2 in sclera significantly increased compared with that in group A (pIGF-1, STAT3 and MMP-2 in scleral significantly upregulated 21 days after occlusion (pIGF-1 in sclera were positively correlated (r = 0.962, pIGF-1, STAT3 and MMP-2 in the sclera and myopia of guinea pigs. The expressions of IGF-1, STAT3 and MMP-2 increased progressively over the time of deprivation. Additionally, overexpression of MMP-2 mediated by IGF-1/STAT3 pathway in sclera might promote the formation of myopia.

PNF and manual therapy treatment results of patients with cervical spine osteoarthritis.

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Maicki, Tomasz; Bilski, Jan; Szczygieł, Elżbieta; Trąbka, Rafał

2017-09-22

The aim of this study was to evaluate the effectiveness of PNF and manual therapy methods in the treatment of patients with cervical spine osteoarthritis, especially their efficacy in reducing pain and improving functionality in everyday life. Long-term results were also compared in order to determine which method of treatment is more effective. Eighty randomly selected females aged 45-65 were included in the study. They were randomly divided into two groups of 40 persons. One group received PNF treatment and the other received manual therapy (MAN.T). To evaluate functional capabilities, the Functional Rating Index was used. To evaluate changes in pain, a shortened version of the McGill Questionnaire was used. The PNF group achieved a greater reduction in pain than the MAN.T group. The PNF group showed a greater improvement in performing daily activities such as sleeping, personal care, travelling, work, recreation, lifting, walking and standing as well as decreased intensity and frequency of pain compared to the MAN.T group. The PNF method proved to be more effective in both short (after two weeks) and long (after three months) term.

Increasing extracellular matrix collagen level and MMP activity induces cyst development in polycystic kidney disease.

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Liu, Bin; Li, Chenghai; Liu, Zijuan; Dai, Zonghan; Tao, Yunxia

2012-09-11

Polycystic Kidney Disease (PKD) kidneys exhibit increased extracellular matrix (ECM) collagen expression and metalloproteinases (MMPs) activity. We investigated the role of these increases on cystic disease progression in PKD kidneys. We examined the role of type I collagen (collagen I) and membrane bound type 1 MMP (MT1-MMP) on cyst development using both in vitro 3 dimensional (3D) collagen gel culture and in vivo PCK rat model of PKD. We found that collagen concentration is critical in controlling the morphogenesis of MDCK cells cultured in 3D gels. MDCK cells did not form 3D structures at collagen I concentrations lower than 1 mg/ml but began forming tubules when the concentration reaches 1 mg/ml. Significantly, these cells began to form cyst when collagen I concentration reached to 1.2 mg/ml, and the ratios of cyst to tubule structures increased as the collagen I concentration increased. These cells exclusively formed cyst structures at a collagen I concentration of 1.8 mg/ml or higher. Overexpression of MT1-MMP in MDCK cells significantly induced cyst growth in 3D collagen gel culture. Conversely, inhibition of MMPs activity with doxycycline, a FDA approved pan-MMPs inhibitor, dramatically slowed cyst growth. More importantly, the treatment of PCK rats with doxycycline significantly decreased renal tubule cell proliferation and markedly inhibited the cystic disease progression. Our data suggest that increased collagen expression and MMP activity in PKD kidneys may induce cyst formation and expansion. Our findings also suggest that MMPs may serve as a therapeutic target for the treatment of human PKD.

Increasing extracellular matrix collagen level and MMP activity induces cyst development in polycystic kidney disease

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Liu Bin

2012-09-01

Full Text Available Abstract Background Polycystic Kidney Disease (PKD kidneys exhibit increased extracellular matrix (ECM collagen expression and metalloproteinases (MMPs activity. We investigated the role of these increases on cystic disease progression in PKD kidneys. Methods We examined the role of type I collagen (collagen I and membrane bound type 1 MMP (MT1-MMP on cyst development using both in vitro 3 dimensional (3D collagen gel culture and in vivo PCK rat model of PKD. Results We found that collagen concentration is critical in controlling the morphogenesis of MDCK cells cultured in 3D gels. MDCK cells did not form 3D structures at collagen I concentrations lower than 1 mg/ml but began forming tubules when the concentration reaches 1 mg/ml. Significantly, these cells began to form cyst when collagen I concentration reached to 1.2 mg/ml, and the ratios of cyst to tubule structures increased as the collagen I concentration increased. These cells exclusively formed cyst structures at a collagen I concentration of 1.8 mg/ml or higher. Overexpression of MT1-MMP in MDCK cells significantly induced cyst growth in 3D collagen gel culture. Conversely, inhibition of MMPs activity with doxycycline, a FDA approved pan-MMPs inhibitor, dramatically slowed cyst growth. More importantly, the treatment of PCK rats with doxycycline significantly decreased renal tubule cell proliferation and markedly inhibited the cystic disease progression. Conclusions Our data suggest that increased collagen expression and MMP activity in PKD kidneys may induce cyst formation and expansion. Our findings also suggest that MMPs may serve as a therapeutic target for the treatment of human PKD.

Matrix metalloproteinase 2 and membrane type 1 matrix metalloproteinase co-regulate axonal outgrowth of mouse retinal ganglion cells

DEFF Research Database (Denmark)

Gaublomme, Djoere; Buyens, Tom; De Groef, Lies

2014-01-01

regenerative therapies, an improved understanding of axonal outgrowth and the various molecules influencing it, is highly needed. Matrix metalloproteinases (MMPs) constitute a family of zinc-dependent proteases that were sporadically reported to influence axon outgrowth. Using an ex vivo retinal explant model......, but not MMP-9, are involved in this process. Furthermore, administration of a novel antibody to MT1-MMP that selectively blocks pro-MMP-2 activation revealed a functional co-involvement of these proteinases in determining RGC axon outgrowth. Subsequent immunostainings showed expression of both MMP-2 and MT1...... nervous system is lacking in adult mammals, thereby impeding recovery from injury to the nervous system. Matrix metalloproteinases (MMPs) constitute a family of zinc-dependent proteases that were sporadically reported to influence axon outgrowth. Inhibition of specific MMPs reduced neurite outgrowth from...

The Regulatory Mechanism of MLT/MT1 Signaling on the Growth of Antler Mesenchymal Cells

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Feifei Yang

2017-10-01

Full Text Available Melatonin (MLT plays an important role in regulating the physiological cycle of seasonal breeding animals. Melatonin receptor I (MT1 is effectively expressed in the cambium layer of deer antler. However, the function and metabolic mechanism of MLT/MT1 signaling in the mesenchymal cells of sika deer remain to be further elucidated. In this work, we detected the effects of MLT/MT1 signaling on mesenchymal cells proliferation and the interaction between MLT/MT1 and IGF1/IGF1-R signaling. The results show that (1 deer antler mesenchymal cells actually express MT1; (2 exogenous melatonin significantly promotes mesenchymal cells proliferation, while MT1 knock-down significantly impairs the positive effects of melatonin; and (3 melatonin significantly enhanced IGF1/IGF1-R signaling, as both the expression of IGF1 and IGF-1R increased, while MT1 knock-down significantly decreased IGF1-R expression and IGF1 synthesis. In summary, these data verified that MLT/MT1 signaling plays a crucial role in antler mesenchymal proliferation, which may be mediated by IGF1/IGF1-R.

Canis mtDNA HV1 database: a web-based tool for collecting and surveying Canis mtDNA HV1 haplotype in public database.

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Thai, Quan Ke; Chung, Dung Anh; Tran, Hoang-Dung

2017-06-26

Canine and wolf mitochondrial DNA haplotypes, which can be used for forensic or phylogenetic analyses, have been defined in various schemes depending on the region analyzed. In recent studies, the 582 bp fragment of the HV1 region is most commonly used. 317 different canine HV1 haplotypes have been reported in the rapidly growing public database GenBank. These reported haplotypes contain several inconsistencies in their haplotype information. To overcome this issue, we have developed a Canis mtDNA HV1 database. This database collects data on the HV1 582 bp region in dog mitochondrial DNA from the GenBank to screen and correct the inconsistencies. It also supports users in detection of new novel mutation profiles and assignment of new haplotypes. The Canis mtDNA HV1 database (CHD) contains 5567 nucleotide entries originating from 15 subspecies in the species Canis lupus. Of these entries, 3646 were haplotypes and grouped into 804 distinct sequences. 319 sequences were recognized as previously assigned haplotypes, while the remaining 485 sequences had new mutation profiles and were marked as new haplotype candidates awaiting further analysis for haplotype assignment. Of the 3646 nucleotide entries, only 414 were annotated with correct haplotype information, while 3232 had insufficient or lacked haplotype information and were corrected or modified before storing in the CHD. The CHD can be accessed at http://chd.vnbiology.com . It provides sequences, haplotype information, and a web-based tool for mtDNA HV1 haplotyping. The CHD is updated monthly and supplies all data for download. The Canis mtDNA HV1 database contains information about canine mitochondrial DNA HV1 sequences with reconciled annotation. It serves as a tool for detection of inconsistencies in GenBank and helps identifying new HV1 haplotypes. Thus, it supports the scientific community in naming new HV1 haplotypes and to reconcile existing annotation of HV1 582 bp sequences.

A novel Drosophila injury model reveals severed axons are cleared through a Draper/MMP-1 signaling cascade

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Purice, Maria D; Ray, Arpita; Münzel, Eva Jolanda; Pope, Bernard J; Park, Daniel J; Speese, Sean D; Logan, Mary A

2017-01-01

Neural injury triggers swift responses from glia, including glial migration and phagocytic clearance of damaged neurons. The transcriptional programs governing these complex innate glial immune responses are still unclear. Here, we describe a novel injury assay in adult Drosophila that elicits widespread glial responses in the ventral nerve cord (VNC). We profiled injury-induced changes in VNC gene expression by RNA sequencing (RNA-seq) and found that responsive genes fall into diverse signaling classes. One factor, matrix metalloproteinase-1 (MMP-1), is induced in Drosophila ensheathing glia responding to severed axons. Interestingly, glial induction of MMP-1 requires the highly conserved engulfment receptor Draper, as well as AP-1 and STAT92E. In MMP-1 depleted flies, glia do not properly infiltrate neuropil regions after axotomy and, as a consequence, fail to clear degenerating axonal debris. This work identifies Draper-dependent activation of MMP-1 as a novel cascade required for proper glial clearance of severed axons. DOI: http://dx.doi.org/10.7554/eLife.23611.001 PMID:28825401

Safety and Usage of C1-Inhibitor in Hereditary Angioedema

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Riedl, Marc A; Bygum, Anette; Lumry, William

2016-01-01

, international patient registry documented widespread implementation of pnfC1-INH self-administration outside of a health care setting consistent with current HAE guidelines. These real-world data revealed pnfC1-INH usage for a variety of reasons in patients with HAE and showed a high level of safety regardless...... of this study was to describe safety and usage patterns of pnfC1-INH. METHODS: A multicenter, observational, registry was conducted between 2010 and 2014 at 30 United States and 7 European sites to obtain both prospective (occurring after enrollment) and retrospective (occurring before enrollment) safety...... and usage data on subjects receiving pnfC1-INH for any reason. RESULTS: Of 343 enrolled patients, 318 received 1 or more doses of pnfC1-INH for HAE attacks (11,848 infusions) or for prophylaxis (3142 infusions), comprising the safety population. Median dosages per infusion were 10.8 IU/kg (attack treatment...

Elevated expression of MMP-13 and TIMP-1 in head and neck squamous cell carcinomas may reflect increased tumor invasiveness

International Nuclear Information System (INIS)

Culhaci, Nil; Metin, Kubilay; Copcu, Eray; Dikicioglu, Emel

2004-01-01

Matrix metalloproteinases [MMPs], which degrade the extracellular matrix, play an important role in the invasion and metastasis of squamous cell carcinomas. One MMP, MMP-13, is thought to play a central role in MMP activation. The purpose of this study was to investigate MMP-13 and TIMP-1 expression in squamous cell carcinomas of the head and neck and to relate these levels of expression to histologic patterns of invasion. This study included T1 lesions obtained via biopsy from the larynx, tongue, and skin/mucosa of 78 patients with head and neck squamous cell carcinomas. The relationship between expression of MMP-13 and TIMP-1 and the mode of tumor invasion [MI] was evaluated immunohistochemically, using breast carcinoma tissue as a positive control. Increased expression was observed in highly invasive tumors, as reflected by the significant correlation between the degree of staining for MMP-13 or TIMP-1 and MI grade [p < 0.05]. There was no significant relationship between the degree of staining for MMP-13 or TIMP-1 and patient age, sex, tumor site, or tumor histologic grade. In addition, levels of staining for MMP-13 did not correlate with levels of staining for TIMP-1. The expression of MMP-13 and TIMP-1 appears to play an important role in determining the invasive capacity of squamous cell carcinomas of the head and neck. Whereas additional studies are needed to confirm these findings, evaluating expression of these MMPs in small biopsy samples may be useful in determining the invasive capacity of these tumors at an earlier stage

Lung Adenocarcinomas and Lung Cancer Cell Lines Show Association of MMP-1 Expression With STAT3 Activation

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Alexander Schütz

2015-04-01

Full Text Available Signal transducer and activator of transcription 3 (STAT3 is constitutively activated in the majority of lung cancer. This study aims at defining connections between STAT3 function and the malignant properties of non–small cell lung carcinoma (NSCLC cells. To address possible mechanisms by which STAT3 influences invasiveness, the expression of matrix metalloproteinase-1 (MMP-1 was analyzed and correlated with the STAT3 activity status. Studies on both surgical biopsies and on lung cancer cell lines revealed a coincidence of STAT3 activation and strong expression of MMP-1. MMP-1 and tyrosine-phosphorylated activated STAT3 were found co-localized in cancer tissues, most pronounced in tumor fronts, and in particular in adenocarcinomas. STAT3 activity was constitutive, although to different degrees, in the lung cancer cell lines investigated. Three cell lines (BEN, KNS62, and A549 were identified in which STAT3 activitation was inducible by Interleukin-6 (IL-6. In A549 cells, STAT3 activity enhanced the level of MMP-1 mRNA and stimulated transcription from the MMP-1 promoter in IL-6–stimulated A549 cells. STAT3 specificity of this effect was confirmed by STAT3 knockdown through RNA interference. Our results link aberrant activity of STAT3 in lung cancer cells to malignant tumor progression through up-regulation of expression of invasiveness-associated MMPs.

Matrix metalloproteinase 9 (MMP-9) mediated release of MMP-9 resistant stromal cell-derived factor 1α (SDF-1α) from surface modified polymer films.

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Steinhagen, Max; Hoffmeister, Peter-Georg; Nordsieck, Karoline; Hötzel, Rudi; Baumann, Lars; Hacker, Michael C; Schulz-Siegmund, Michaela; Beck-Sickinger, Annette G

2014-04-23

Preparation of smart materials by coatings of established surfaces with biomolecules will lead to the next generation of functionalized biomaterials. Rejection of implants is still a major problem in medical applications but masking the implant material with protein coatings is a promising approach. These layers not only disguise the material but also equip it with a certain biological function. The anti-inflammatory chemokine stromal cell-derived factor 1α (SDF-1α) is well suited to take over this function, because it efficiently attracts stem cells and promotes their differentiation and proliferation. At least the initial stem cell homing requires the formation of a concentration gradient. Thus, a reliable and robust release mechanism of SDF-1α from the material is essential. Several proteases, most notably matrix metalloproteinases, are upregulated during inflammation, which, in principle, can be exploited for a tightly controlled release of SDF-1α. Herein, we present the covalent immobilization of M-[S4V]-SDF-1α on novel biodegradable polymer films, which consist of heterobifunctional poly(ethylene glycol) and oligolactide-based functionalized macromers. A peptidic linker with a trimeric matrix metalloproteinase 9 (MMP-9) cleavage site (MCS) was used as connection and the linkage between the three components was achieved by combination of expressed protein ligation and Cu(I) catalyzed azide/alkyne cycloaddition. The MCS was used for MMP-9 mediated release of M-[S4V]-SDF-1α from the biomaterial and the released SDF-1α derivative was biologically active and induced strong cell migration, which demonstrates the great potential of this system.

The ERK1/2 Inhibitor U0126 Attenuates Diabetes-Induced Upregulation of MMP-9 and Biomarkers of Inflammation in the Retina

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Ghulam Mohammad

2013-01-01

Full Text Available This study was conducted to determine the expression of matrix metalloproteinase-9 (MMP-9 and tissue inhibitor of metalloproteinase-1 (TIMP-1 in a time-dependent manner and the effect of extracellular-signal-regulated kinases-1/2 (ERK1/2 inhibition on the expressions of MMP-9, TIMP-1, and inflammatory biomarkers in the retinas of diabetic rats. The expression of MMP-9 was quantified by zymography, and the mRNA level of MMP-9 and TIMP-1 was quantified by RT-PCR. The expression of inducible nitric oxide synthase (iNOS, interleukin-6 (IL-6, and tumor necrosis factor-alpha (TNF-α was examined by Western blot analysis. MMP-9 expression was significantly higher in diabetic rat retinas compared to controls at all time points.TIMP-1 expression was nonsignificantly upregulated at 1week of diabetes and was significantly downregulated at 4 and 12 weeks of diabetes. Intravitreal administration of the ERK1/2 inhibitor U0126 prior to induction of diabetes decreased ERK1/2 activation, attenuated diabetes-induced upregulation of MMP-9, iNOS, IL-6, and TNF-α and upregulated TIMP-1 expression. In MMP-9 knockout mice, diabetes had no effect on retinal iNOS expression and its level remained unchanged. These data provide evidence that ERK1/2 signaling pathway is involved in MMP-9, iNOS, IL-6, and TNF-α induction in diabetic retinas and suggest that ERK1/2 can be a novel therapeutic target in diabetic retinopathy.

Detection of the matrix metalloproteinases MMP-2 and MMP-9 and tissue inhibitors of metalloproteinases TIMP-1 and TIMP-2 in llama (Lama glama) oviduct.

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Zampini, R; Argañaraz, M E; Miceli, D C; Apichela, S A

2014-06-01

Matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) are involved in several reproductive events like oocyte-spermatozoa interaction and semen liquefaction. In order to study their role in the llama oviductal reproductive process, MMP activity in oviductal fluid (OF) was assayed. Considering that llama genome sequences are partially known, a strategy to procure cDNA sequences of MMP-2, MMP-9, TIMP-1 and TIMP-2 was designed. Afterwards, their expression patterns in the different llama oviductal segments were assayed. Gelatine zymograms detected 62 and 94 kDa protease activities that matched MMP-2 and pro-MMP-9, respectively. Expression pattern analysis showed that MMP and TIMP mRNAs were present in ampulla, isthmus, utero-tubal junction (UTJ) and papilla. Altogether, these findings support the argument that MMPs/TIMPs are produced in the oviduct and secreted into the oviductal lumen. Our results encourage further studies to elucidate the role of these proteins in reproductive oviductal events. © 2014 Blackwell Verlag GmbH.

MMP3 and TIMP1 variants contribute to chronic periodontitis and may be implicated in disease progression.

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Letra, Ariadne; Silva, Renato M; Rylands, Ryan J; Silveira, Elcia M; de Souza, Ana P; Wendell, Steven K; Garlet, Gustavo P; Vieira, Alexandre R

2012-08-01

Matrix metalloproteinases (MMPs) play a key role in the tissue destruction characteristic of chronic periodontitis. The purpose of this study was to investigate the association of MMP and TIMP polymorphisms with chronic periodontitis in two populations. A total of 34 polymorphisms spanning 12 MMP and 2 TIMP genes were genotyped in 401 individuals from Brazil (99 cases with chronic periodontitis and 302 controls), and 274 individuals from the US (70 cases and 204 controls). Individuals were considered cases if presenting at least three teeth exhibiting sites of clinical attachment loss ≥ 5 mm in two different quadrants. Controls were characterized by absence of clinical attachment loss and no sites with probing depth >3 mm. MMP3 and TIMP1 mRNA expression was evaluated in healthy and diseased periodontal tissues. TIMP1 showed association with chronic periodontitis in the Brazilian population (for rs5906435, p = 0.0004), whereas MMP3 showed association in the US population (for rs679620, p = 0.0003; and rs650108, p = 0.002) and in the Brazilian population (for rs639752, p = 0.005). MMP3 and TIMP1 mRNA expression was significantly higher in diseased tissues when compared to control tissues. Our results further support a role for variations in MMP3 in chronic periodontitis and report a novel association with TIMP1. These genes may be considered additional candidate genes for chronic periodontitis. © 2012 John Wiley & Sons A/S.

VANGL2 regulates membrane trafficking of MMP14 to control cell polarity and migration.

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Williams, B Blairanne; Cantrell, V Ashley; Mundell, Nathan A; Bennett, Andrea C; Quick, Rachel E; Jessen, Jason R

2012-05-01

Planar cell polarity (PCP) describes the polarized orientation of cells within the plane of a tissue. Unlike epithelial PCP, the mechanisms underlying PCP signaling in migrating cells remain undefined. Here, the establishment of PCP must be coordinated with dynamic changes in cell adhesion and extracellular matrix (ECM) organization. During gastrulation, the membrane type-1 matrix metalloproteinase (MT1-MMP or MMP14) is required for PCP and convergence and extension cell movements. We report that the PCP protein Vang-like 2 (VANGL2) regulates the endocytosis and cell-surface availability of MMP14 in manner that is dependent on focal adhesion kinase. We demonstrate that zebrafish trilobite/vangl2 mutant embryos exhibit increased Mmp14 activity and decreased ECM. Furthermore, in vivo knockdown of Mmp14 partially rescues the Vangl2 loss-of-function convergence and extension phenotype. This study identifies a mechanism linking VANGL2 with MMP14 trafficking and suggests that establishment of PCP in migrating gastrula cells requires regulated proteolytic degradation or remodeling of the ECM. Our findings implicate matrix metalloproteinases as downstream effectors of PCP and suggest a broadly applicable mechanism whereby VANGL2 affects diverse morphogenetic processes.

Cross talk between MMP2-Spm-Cer-S1P and ERK1/2 in proliferation of pulmonary artery smooth muscle cells under angiotensin II stimulation.

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Chowdhury, Animesh; Sarkar, Jaganmay; Pramanik, Pijush Kanti; Chakraborti, Tapati; Chakraborti, Sajal

2016-08-01

The aim of the present study is to establish the mechanism associated with the proliferation of PASMCs under ANG II stimulation. The results showed that treatment of PASMCs with ANG II induces an increase in cell proliferation and 100 nM was the optimum concentration for maximum increase in proliferation of the cells. Pretreatment of the cells with AT1, but not AT2, receptor antagonist inhibited ANG II induced cell proliferation. Pretreatment with pharmacological and genetic inhibitors of sphingomyelinase (SMase) and sphingosine kinase (SPHK) prevented ANG II-induced cell proliferation. ANG II has also been shown to induce SMase activity, SPHK phosphorylation and S1P production. In addition, ANG II caused an increase in proMMP-2 expression and activation, ERK1/2 phosphorylation and NADPH oxidase activation. Upon inhibition of MMP-2, SMase activity and S1P level were curbed leading to inhibition of cell proliferation. SPHK was phosphorylated by ERK1/2 during ET-1 stimulation of the cells. ANG II-induced ERK1/2 phosphorylation and proMMP-2 expression and activation in the cells were abrogated upon inhibition of NADPH oxidase activity. Overall, NADPH oxidase plays an important role in proMMP-2 expression and activation and that MMP-2 mediated SMC proliferation occurs through the involvement of Spm-Cer-S1P signaling axis under ANG II stimulation of PASMCs. Copyright © 2016 Elsevier Inc. All rights reserved.

Effects of Stretching by P.N.F and Harmonic Techniques on Hamstring Flexibility

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Hassan Shakeri

2006-01-01

Full Text Available Objective: Improving of muscle flexibility is an important issue in physiotherapy and sport sciences. There are many methods for increasing muscle length and decreasing muscle stiffness. In research findings, PNF method has been found to be better than static and ballistic methods. There is another method named Harmonic technique (introduced by E. Lederman 1997 that has been claimed to be more effective, but there is not enough documentation about this claim. Aim of this study was to compare effects of stretching by PNF and harmonic techniques on hamstring flexibility. Materials & Methods: This research is a RCT study in that 45 colledge students aged 18-35 years were arranged in three groups (Harmonic, P.N.F, and control. Subjects haven’t had any painful pathology in low-back and lower extremities for last six months. Subjects had limited hamstring length (20 degrees deficiency in Active-Knee-Extension test and hadn’t professional sport activities. Dependent variablies were muscle stiffness and hamstring length which popliteal angle in AKE test was its indirect index. In pilot study, reliability of measurement of these variables were approved. Then hamstring muscle of subjects in harmonic and PNF groups were stretched by harmonic and PNF methods for six weeks, 5 minute per day and 3d/wks, whereas control group hadn’t any exercise. Results: Findings of this study showed that in both used techniques, changes of hamstring length were significant (P=0.000, but in control group there wasn’t significant change. There wasn’t significant differences between changes of hamstring length in PNF and Harmonic groups. Only in harmonic group, muscle stiffness had significant changes (P<0.03. Conclusion: According to findings of this research, both harmonic and PNF methods equally increased length of hamstring, and harmonic technique can be used as an alternative stretching method for other techniques. Maybe harmonic technique is better than PNF

EFFECT OF PROPRIOCEPTIVE NEUROMUSCULAR FACILITATION (PNF IN IMPROVING SENSORIMOTOR FUNCTION IN PATIENTS WITH DIABETIC NEUROPATHY AFFECTING LOWER LIMBS

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Kamaljeet Singh

2016-06-01

Full Text Available Background: Diabetic Mellitus is a group of metabolic disease characterized by hyperglycaemia resulting from defects in insulin secretion, insulin action or both. Distal Sensorimotor Polyneuropathy is the most common complication of diabetes which mainly affects the lower limbs. Most of the studies aimed at individually increasing muscle strength or sensation but not on overall performance enhancements of the diabetic lower limbs. The evidence supporting the effectiveness of PNF in diabetic neuropathic patients is scarce. Methods: 30 patients, with age between 50 to 70 years, diagnosed with Diabetic Sensorimotor Polyneuropathy (DSP were selected from the department of Medicine and department of Neurosurgery Guru Gobind Singh Medical College and Hospital. Patients were evaluated at the beginning and at the end of the intervention using Diabetic Neuropathy Examination scores. Patients received 3 sets of exercises one hour/day with 3 days/week for 3 months. Each set of exercises consists of 5 repetitions of PNF patterns (alternate day and techniques. Results: D1 & D2 patterns of PNF are effective in improving both motor and sensory functions of diabetic patients with neuropathic symptoms. Improvement in muscle strength, reflex and sensations occurred to a greater extent after the treatment of three months in these subjects. This study shows that PNF patterns were effective at enhancing sensorimotor problems of lower limbs. Conclusion: This study concluded that PNF is found to be effective in improving sensorimotor functions of diabetic neuropathic patients affecting lower limbs.

The ERK MAP kinase-PEA3/ETV4-MMP-1 axis is operative in oesophageal adenocarcinoma

LENUS (Irish Health Repository)

Keld, Richard

2010-12-09

Abstract Background Many members of the ETS-domain transcription factor family are important drivers of tumourigenesis. In this context, their activation by Ras-ERK pathway signaling is particularly relevant to the tumourigenic properties of many ETS-domain transcription factors. The PEA3 subfamily of ETS-domain transcription factors have been implicated in tumour metastasis in several different cancers. Results Here, we have studied the expression of the PEA3 subfamily members PEA3\\/ETV4 and ER81\\/ETV1 in oesophageal adenocarcinomas and determined their role in oesophageal adenocarcinoma cell function. PEA3 plays an important role in controlling both the proliferation and invasive properties of OE33 oesophageal adenocarcinoma cells. A key target gene is MMP-1. The ERK MAP kinase pathway activates PEA3 subfamily members and also plays a role in these PEA3 controlled events, establishing the ERK-PEA3-MMP-1 axis as important in OE33 cells. PEA3 subfamily members are upregulated in human adenocarcinomas and expression correlates with MMP-1 expression and late stage metastatic disease. Enhanced ERK signaling is also more prevalent in late stage oesophageal adenocarcinomas. Conclusions This study shows that the ERK-PEA3-MMP-1 axis is upregulated in oesophageal adenocarcinoma cells and is a potentially important driver of the metastatic progression of oesophageal adenocarcinomas.

Biochanin-A antagonizes the interleukin-1β-induced catabolic inflammation through the modulation of NFκB cellular signaling in primary rat chondrocytes

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Oh, Ji-Su [Department of Oral and Maxillofacial Surgery, Chosun University, Gwangju, 61452 (Korea, Republic of); Cho, In-A; Kang, Kyeong-Rok [Department of Dental Bioengineering, Chosun University, Gwangju, 61452 (Korea, Republic of); You, Jae-Seek [Department of Oral and Maxillofacial Surgery, Chosun University, Gwangju, 61452 (Korea, Republic of); Yu, Sang-Joun [Department of Periodontology, Chosun University, Gwangju, 61452 (Korea, Republic of); Lee, Gyeong-Je [Department of Prosthodontics, Chosun University, Gwangju, 61452 (Korea, Republic of); Seo, Yo-Seob [Department of Oral and Maxillofacial Radiology, Chosun University, Gwangju, 61452 (Korea, Republic of); Kim, Chun Sung; Kim, Do Kyung [Pre-Dentistry, School of Dentistry, Chosun University, Gwangju, 61452 (Korea, Republic of); Kim, Su-Gwan [Department of Oral and Maxillofacial Surgery, Chosun University, Gwangju, 61452 (Korea, Republic of); Seo, Young-Woo [Korea Basic Science Institute, Gwangju Center, Chonnam National University, Gwangju, 61186 (Korea, Republic of); Im, Hee-Jeong [Department of Biochemistry, Rush University Medical Center, Chicago, IL, 60612 (United States); Kim, Jae-Sung, E-mail: js_kim@chosun.ac.kr [Pre-Dentistry, School of Dentistry, Chosun University, Gwangju, 61452 (Korea, Republic of)

2016-09-02

Biochanin-A, a phytoestrogen derived from herbal plants, protected from the IL-1β-induced loss of proteoglycans through the suppression of matrix degrading enzymes such as matrix metalloproteinase (MMP)-13, MMP-3, MMP-1, and ADAMTS-5 in primary rat chondrocytes and the knee articular cartilage. It also suppressed the expression of IL-1β-induced catabolic factors such as nitric oxide synthase 2, cyclooxygenase-2, prostaglandin E{sub 2}, and inflammatory cytokines. Furthermore, biochanin-A suppressed the IL-1β-induced phosphorylation of NFκB, and inhibited its nuclear translocation in primary rat chondrocytes. These results indicate that biochanin-A antagonizes the IL-1β-induced catabolic effects through its anti-inflammatory activity that involves the modulation of NFκB signaling. - Highlights: • Biochanin-A is a phytoestrogen derived from medicinal plants. • It suppressed the IL-1β-induced matrix degrading enzymes and catabolic factors. • It inhibited IL-1β-induced proteoglycan loss in chondrocytes and cartilage tissues. • Its anti-catabolic effects were mediated by modulation of NFκB signaling. • It may be used as a potential anti-catabolic biomaterial for osteoarthritis.

Correlation of matrix metalloproteinase (MMP)-1, -2, -3, and -9 expressions with demographic and radiological features in primary lumbar intervertebral disc disease.

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Basaran, Recep; Senol, Mehmet; Ozkanli, Seyma; Efendioglu, Mustafa; Kaner, Tuncay

2017-07-01

Degeneration of IVD is a progressive and irreversible process and can be evaluated with immunohistochemical examination or radiological grading. MMPs are a family of proteolytic enzymes and involved in the degradation of the matrix components of the IVD. We aimed to compare MMP-1, -2, -3, and -9 expressions with demographic features, visual analogue scale (VAS), Oswestry Disability Index (ODI) and radiological (MRI) grades. The study involved 60 participants. We recorded data about age, complaint, radiological imaging, expression levels of MMP-1, -2, -3, and -9, ODI and VAS for back pain retrospectively. Intervertebral disc degeneration was graded on a 0-5 scale according to the Pfirrmann classification. As a result of the study, the median age was 52.09±12.74years. There were statistical significances between age and MMP-1, and MMP-2. There was a close correlation between grade and MMP-9. We found correlation between the VAS and the MMP-9 expression. In addition, there was relationship between expression of MMP-2 and MMP-1, MMP-3, MMP-9. In conclusion, the expressions of MMP-1 and -2 are increased with aging. There was no relationship between radiological evaluation of IVDD and aging. Increased expression of MMPs affected IVDD positively. The relationship with MMPs is not explained. This study adds to our understanding of the interaction between MMPs and IVDD. Copyright © 2017 Elsevier Ltd. All rights reserved.

Studies on Multifunctional Effect of All-Trans Retinoic Acid (ATRA on Matrix Metalloproteinase-2 (MMP-2 and Its Regulatory Molecules in Human Breast Cancer Cells (MCF-7

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Anindita Dutta

2009-01-01

Full Text Available Background. Vitamin A derivative all-trans retinoic acid (ATRA is considered as a potent chemotherapeutic drug for its capability of regulating cell growth and differentiation. We studied the effect of ATRA on MMP-2 in MCF-7, human breast cancer cells, and the probable signaling pathways which are affected by ATRA on regulating pro-MMP-2 activity and expression. Methods. Gelatin zymography, RT-PCR, ELISA, Western blot, Immunoprecipitation, and Cell adhesion assay are used. Results. Gelatin zymography showed that ATRA caused a dose-dependent inhibition of pro-MMP-2 activity. ATRA treatment downregulates the expression of MT1-MMP, EMMPRIN, FAK, NF-kB, and p-ERK. However, expression of E-cadherin, RAR, and CRABP increased upon ATRA treatment. Binding of cells to extra cellular matrix (ECM protein fibronectin reduced significantly after ATRA treatment. Conclusions. The experimental findings clearly showed the inhibition of MMP-2 activity upon ATRA treatment. This inhibitory effect of ATRA on MMP-2 activity in human breast cancer cells (MCF-7 may result due to its inhibitory effect on MT1-MMP, EMMPRIN, and upregulation of TIMP-2. This study is focused on the effect of ATRA on MMP, MMP-integrin-E-cadherin interrelationship, and also the effect of the drug on different signaling molecules which may involve in the progression of malignant tumor development.

Exercise Intolerance and Myoglobinuria Associated with a Novel Maternally Inherited MT-ND1 Mutation

DEFF Research Database (Denmark)

Rafiq, Jabin; Duno, Morten; Østergaard, Elsebet

2016-01-01

The most common clinical phenotype caused by a mtDNA mutation in complex I of the mitochondrial respiratory chain is Leber hereditary optic neuropathy. We report a family with a novel maternally inherited homoplasmic mtDNA m.4087A>G mutation in the ND1 gene (MT-ND1) associated with isolated...... myopathy, recurrent episodes of myoglobinuria, and rhabdomyolysis. DNA from blood in seven family members and muscle from four family members were PCR amplified and sequenced directly and assessed for the m.4087A>G variation in MT-ND1. Mitochondrial enzyme activity in all muscle biopsies was measured. PCR...... myoglobinuria is a rare phenotype of mitochondrial myopathies. We report this phenotype in a family affected by a novel homoplasmic mutation in MT-ND1. It is the first time such a phenotype has been associated with complex I gene mutations and a homoplasmic mutation of mtDNA....

Sphingosine-1-phosphate promotes extravillous trophoblast cell invasion by activating MEK/ERK/MMP-2 signaling pathways via S1P/S1PR1 axis activation.

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Yang, Weiwei; Li, Qinghua; Pan, Zhifang

2014-01-01

Successful placentation depends on the proper invasion of extravillous trophoblast (EVT) cells into maternal tissues. Previous reports demonstrated that S1P receptors are expressed in the EVT cells and S1P could regulate migration and function of trophoblast cells via S1P receptors. However, little is known about roles of S1P in the invasion of EVT cells. Our study was performed to investigate S1P effect on the invasion of EVT cells. We used the extravillous trophoblast cell line HTR8/SVneo cells to evaluate the effect. In vitro invasion assay was employed to determine the invasion of HTR8/SVneo cells induced by S1P. MMP-2 enzyme activity and relative level in the supernatants of HTR8/SVneo was assessed by gelatin zymography and western blot. Based on the above, siRNA and specific inhibitors were used for the intervention and study of potential signal pathways, and Real-time qPCR and western blot were used to test the mRNA and protein level of potential signal targets. We found that S1P could promote HTR8/SVneo cell invasion and upregulates activity and level of MMP-2. The promotion requires activation of MEK-ERK and is dependent on the axis of S1P/S1PR1. Our investigation of S1P may provide new insights into the molecular mechanisms of EVT invasion.

Overexpression of TIMP-1 under the MMP-9 promoter interferes with wound healing in transgenic mice

OpenAIRE

Salonurmi, T.; Parikka, M.; Kontusaari, S.; Pirila, E.; Munaut, Carine; Salo, T.; Tryggvason, K.

2004-01-01

We have generated transgenic mice harboring the murine matrix metalloproteinase 9 (MMP-9) promoter cloned in front of human TIMP-1 cDNA. The transgenic mice were viable and fertile and exhibited normal growth and general development. During wound healing the mice were shown to express human TIMP-1 in keratinocytes that normally express MMP-9. However, the healing of skin wounds was significantly retarded with slow migration of keratinocytes over the wound in transgenic mice. In situ zymograph...

IGF-1R Regulates the Extracellular Level of Active MMP-2, Pathological Neovascularization, and Functionality in Retinas of OIR Mouse Model.

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Lorenc, Valeria E; Subirada Caldarone, Paula V; Paz, María C; Ferrer, Darío G; Luna, José D; Chiabrando, Gustavo A; Sánchez, María C

2018-02-01

In ischemic proliferative diseases such as retinopathies, persistent hypoxia leads to the release of numerous neovascular factors that participate in the formation of abnormal vessels and eventually cause blindness. The upregulation and activation of metalloproteinases (MMP-2 and MMP-9) represent a final common pathway in this process. Although many regulators of the neovascular process have been identified, the complete role of the insulin-like growth factor 1 (IGF-1) and its receptor (IGF-1R) appears to be significantly more complex. In this study, we used an oxygen-induced retinopathy (OIR) mouse model as well as an in vitro model of hypoxia to study the role of MMP-2 derived from Müller glial cells (MGCs) and its relation with the IGF-1/IGF-1R system. We demonstrated that MMP-2 protein expression increased in P17 OIR mice, which coincided with the active phase of the neovascular process. Also, glutamine synthetase (GS)-positive cells were also positive for MMP-2, whereas IGF-1R was expressed by GFAP-positive cells, indicating that both proteins were expressed in MGCs. In addition, in the OIR model a single intravitreal injection of the IGF-1R blocking antibody (αIR3) administered at P12 effectively prevented pathologic neovascularization, accelerated physiological revascularization, and improved retinal functionality at P17. Finally, in MGC supernatants, the blocking antibody abolished the IGF-1 effect on active MMP-2 under normoxic and hypoxic conditions without affecting the extracellular levels of pro-MMP-2. These results demonstrate, for the first time, that the IGF-1/IGF-1R system regulates active MMP-2 levels in MGCs, thus contributing to MEC remodeling during the retinal neovascular process.

Transcription factor CgMTF-1 regulates CgZnT1 and CgMT expression in Pacific oyster (Crassostrea gigas) under zinc stress

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Meng, Jie; Zhang, Linlin [Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, Shandong (China); Li, Li, E-mail: lili@qdio.ac.cn [Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, Shandong (China); Li, Chunyan; Wang, Ting [Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, Shandong (China); University of Chinese Academy of Sciences, Beijing 100039 (China); Zhang, Guofan, E-mail: gfzhang@qdio.ac.cn [Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, Shandong (China)

2015-08-15

Highlights: • CgMTF-1 and CgZnT1 were first identified in oysters. • CgMTF-1 localized in cell nucleus under unstressed conditions. • CgMTF-1 proteins could bind with the typical MRE motif. • CgMTF-1 activated CgZnT1, CgMT1 and CgMT4 promoters and regulated their expressions under zinc exposure. - Abstract: Oysters accumulate zinc at high tissue concentrations, and the metal response element (MRE)-binding transcription factor (MTF) functions as the cellular zinc sensor that coordinates the expression of genes involved in zinc efflux and storage, as well as those that protect against metal toxicity. In this study, we cloned MTF-1 in oysters and examined its regulation mechanism for its classic target genes, including MTs and ZnT1 under zinc exposure conditions. We cloned CgMTF-1 and determined the subcellular locations of its protein product in HEK293 cells. CgMTF-1 has a 2826 bp open reading frame that encodes a predicted polypeptide with 707 amino acid residues, showing six well-conserved zinc finger domains that are required for metal binding. In HEK293 cell lines, CgMTF-1 primarily localizes in the cell nucleus under unstressed conditions and nuclear translocation was not critical for the activation of this gene. We searched for CgMTF-1-regulated genes in oysters using RNA interference. Decreased expression levels of CgMT1, CgMT4, and CgZnT1 were observed after CgMTF-1 interference (>70% inhibition) under zinc exposure, indicating the critical role of CgMTF-1 in the regulation of these genes. We searched for a direct regulation mechanism involving CgMTF-1 for CgMT1, CgMT4, and CgZnT1 in vitro. EMSA experiments indicated that CgMTF-1 can bind with the MREs found in the CgZnT1, CgMT1 and CgMT4 promoter regions. Additionally, luciferase reporter gene experiments indicated that CgMTF-1 could activate the CgMT1, CgMT4, and CgZnT1 promoters. Overall, our results suggest that CgMTF-1 directly coordinates the regulation of CgMTs and CgZnT1 expression and plays

Transcription factor CgMTF-1 regulates CgZnT1 and CgMT expression in Pacific oyster (Crassostrea gigas) under zinc stress

International Nuclear Information System (INIS)

Meng, Jie; Zhang, Linlin; Li, Li; Li, Chunyan; Wang, Ting; Zhang, Guofan

2015-01-01

Highlights: • CgMTF-1 and CgZnT1 were first identified in oysters. • CgMTF-1 localized in cell nucleus under unstressed conditions. • CgMTF-1 proteins could bind with the typical MRE motif. • CgMTF-1 activated CgZnT1, CgMT1 and CgMT4 promoters and regulated their expressions under zinc exposure. - Abstract: Oysters accumulate zinc at high tissue concentrations, and the metal response element (MRE)-binding transcription factor (MTF) functions as the cellular zinc sensor that coordinates the expression of genes involved in zinc efflux and storage, as well as those that protect against metal toxicity. In this study, we cloned MTF-1 in oysters and examined its regulation mechanism for its classic target genes, including MTs and ZnT1 under zinc exposure conditions. We cloned CgMTF-1 and determined the subcellular locations of its protein product in HEK293 cells. CgMTF-1 has a 2826 bp open reading frame that encodes a predicted polypeptide with 707 amino acid residues, showing six well-conserved zinc finger domains that are required for metal binding. In HEK293 cell lines, CgMTF-1 primarily localizes in the cell nucleus under unstressed conditions and nuclear translocation was not critical for the activation of this gene. We searched for CgMTF-1-regulated genes in oysters using RNA interference. Decreased expression levels of CgMT1, CgMT4, and CgZnT1 were observed after CgMTF-1 interference (>70% inhibition) under zinc exposure, indicating the critical role of CgMTF-1 in the regulation of these genes. We searched for a direct regulation mechanism involving CgMTF-1 for CgMT1, CgMT4, and CgZnT1 in vitro. EMSA experiments indicated that CgMTF-1 can bind with the MREs found in the CgZnT1, CgMT1 and CgMT4 promoter regions. Additionally, luciferase reporter gene experiments indicated that CgMTF-1 could activate the CgMT1, CgMT4, and CgZnT1 promoters. Overall, our results suggest that CgMTF-1 directly coordinates the regulation of CgMTs and CgZnT1 expression and plays

Expression of matrix metalloproteinase 9 (MMP-9) and tissue inhibitor of metalloproteinases 1 (TIMP-1) by colorectal cancer cells and adjacent stroma cells - associations with histopathology and patients outcome

DEFF Research Database (Denmark)

Jensen, Søren Astrup; Vainer, Ben; Bartels, Annette

2010-01-01

AIM: To elucidate cellular features accountable for colorectal cancers' (CRC) capability to invade normal tissue and to metastasize, we investigated the level of the collagenase matrix metalloproteinase 9 (MMP-9) and its physiological inhibitor tissue inhibitor of metalloproteinases 1 (TIMP-1) in...... cells is associated with poor prognosis independent of its function as inhibitor of MMP-9. MMP-9 and TIMP-1 are important mediators of the host-cancer cell interaction in the tumour microenvironment with significant influence on the histopathology and on prognosis of CRC....

Chrysin inhibits tumor promoter-induced MMP-9 expression by blocking AP-1 via suppression of ERK and JNK pathways in gastric cancer cells.

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Yong Xia

Full Text Available Cell invasion is a crucial mechanism of cancer metastasis and malignancy. Matrix metalloproteinase-9 (MMP-9 is an important proteolytic enzyme involved in the cancer cell invasion process. High expression levels of MMP-9 in gastric cancer positively correlate with tumor aggressiveness and have a significant negative correlation with patients' survival times. Recently, mechanisms suppressing MMP-9 by phytochemicals have become increasingly investigated. Chrysin, a naturally occurring chemical in plants, has been reported to suppress tumor metastasis. However, the effects of chrysin on MMP-9 expression in gastric cancer have not been well studied. In the present study, we tested the effects of chrysin on MMP-9 expression in gastric cancer cells, and determined its underlying mechanism. We examined the effects of chrysin on MMP-9 expression and activity via RT-PCR, zymography, promoter study, and western blotting in human gastric cancer AGS cells. Chrysin inhibited phorbol-12-myristate 13-acetate (PMA-induced MMP-9 expression in a dose-dependent manner. Using AP-1 decoy oligodeoxynucleotides, we confirmed that AP-1 was the crucial transcriptional factor for MMP-9 expression. Chrysin blocked AP-1 via suppression of the phosphorylation of c-Jun and c-Fos through blocking the JNK1/2 and ERK1/2 pathways. Furthermore, AGS cells pretreated with PMA showed markedly enhanced invasiveness, which was partially abrogated by chrysin and MMP-9 antibody. Our results suggest that chrysin may exert at least part of its anticancer effect by controlling MMP-9 expression through suppression of AP-1 activity via a block of the JNK1/2 and ERK1/2 signaling pathways in gastric cancer AGS cells.

Insulin-like growth factor binding protein-3 induces angiogenesis through IGF-I- and SphK1-dependent mechanisms.

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Granata, R; Trovato, L; Lupia, E; Sala, G; Settanni, F; Camussi, G; Ghidoni, R; Ghigo, E

2007-04-01

Angiogenesis is critical for development and repair, and is a prominent feature of many pathological conditions. Based on evidence that insulin-like growth factor binding protein (IGFBP)-3 enhances cell motility and activates sphingosine kinase (SphK) in human endothelial cells, we have investigated whether IGFBP-3 plays a role in promoting angiogenesis. IGFBP-3 potently induced network formation by human endothelial cells on Matrigel. Moreover, it up-regulated proangiogenic genes, such as vascular endothelial growth factor (VEGF) and matrix metalloproteinases (MMP)-2 and -9. IGFBP-3 even induced membrane-type 1 MMP (MT1-MMP), which regulates MMP-2 activation. Decreasing SphK1 expression by small interfering RNA (siRNA), blocked IGFBP-3-induced network formation and inhibited VEGF, MT1-MMP but not IGF-I up-regulation. IGF-I activated SphK, leading to sphingosine-1-phosphate (S1P) formation. The IGF-I effect on SphK activity was blocked by specific inhibitors of IGF-IR, PI3K/Akt and ERK1/2 phosphorylation. The disruption of IGF-I signaling prevented the IGFBP-3 effect on tube formation, SphK activity and VEGF release. Blocking ERK1/2 signaling caused the loss of SphK activation and VEGF and IGF-I up-regulation. Finally, IGFBP-3 dose-dependently stimulated neovessel formation into subcutaneous implants of Matrigel in vivo. Thus, IGFBP-3 positively regulates angiogenesis through involvement of IGF-IR signaling and subsequent SphK/S1P activation.

Increased expression of HIF-1α, VEGF-A and its receptors, MMP-2, TIMP-1, and ADAMTS-1 at the venous stenosis of arteriovenous fistula in a mouse model with renal insufficiency

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Misra, Sanjay; Shergill, Uday; Yang, Binxia; Janardhanan, Rajiv; Misra, Khamal D.

2010-01-01

Purpose A mouse model of renal insufficiency with arteriovenous fistula (AVF) and venous stenosis was created. We tested the hypothesis that there is increased gene expression of hypoxia inducible factor-1 alpha (HIF-1α), vascular endothelial growth factor- A (VEGF-A) and its receptors (VEGFR-1, -2), matrix metalloproteinase-2 (MMP-2), -9 (MMP-9), tissue inhibitor of metalloproteinase-1, -2 (TIMP-1, -2), and a disintegrin and metalloproteinase thrombospondin-1 (ADAMTS-1) at the venous stenosis. Materials and methods Nineteen male C57BL/6 mice underwent a left nephrectomy and a surgical occlusion of the right upper pole to induce renal insufficiency and characterized in eight mice. Twenty eight days later, an AVF (n=11) was created from the right carotid artery to ipsilateral jugular vein and the mice were sacrificed at day 7 (n=4) and day 14 (n=4). The outflow and control veins were removed for gene expression. Three mice were sacrificed at day 28 for histologic analysis. Results The mean serum blood urea nitrogen remained significantly elevated for 8 weeks when compared to baseline (P<0.05). By day 7, there was a significant increase in the expression of HIF-1α, VEGF-A, VEGFR-1, VEGFR-2, MMP-2, TIMP-1, and ADAMTS-1 at the outflow vein with HIF-1α and TIMP-1 being significantly elevated at day 14 (P<0.05). By day 28, the venous stenosis was characterized by a thickened vein wall and neointima. Conclusions A mouse model of renal insufficiency with AVF was developed which had increased expression of HIF-1α, VEGF-A, VEGFR-1, VEGFR-2, MMP-2, TIMP-1, and ADAMTS-1 at the outflow vein with venous stenosis by day 28. PMID:20598569

Increased expression of HIF-1alpha, VEGF-A and its receptors, MMP-2, TIMP-1, and ADAMTS-1 at the venous stenosis of arteriovenous fistula in a mouse model with renal insufficiency.

Science.gov (United States)

Misra, Sanjay; Shergill, Uday; Yang, Binxia; Janardhanan, Rajiv; Misra, Khamal D

2010-08-01

A mouse model of renal insufficiency with arteriovenous fistula (AVF) and venous stenosis was created. The authors tested the hypothesis that there is increased gene expression of hypoxia-inducible factor-1 alpha (HIF-1alpha); vascular endothelial growth factor-A (VEGF-A) and its receptors (VEGFR-1, -2); matrix metalloproteinase-2 (MMP-2), -9 (MMP-9); tissue inhibitor of metalloproteinase-1, -2 (TIMP-1, -2); and a disintegrin and metalloproteinase thrombospondin-1 (ADAMTS-1) at the venous stenosis. Nineteen male C57BL/6 mice underwent a left nephrectomy and a surgical occlusion of the right upper pole to induce renal function characterized in eight animals. Twenty eight days later, an AVF (n = 11) was created from the right carotid artery to ipsilateral jugular vein, and the mice were killed at day 7 (n = 4) and day 14 (n = 4). The outflow and control veins were removed for gene expression. Three mice were killed at day 28 for histologic analysis. The mean serum blood urea nitrogen level remained significantly elevated for 8 weeks when compared with baseline (P < .05). By day seven, there was a significant increase in the expression of HIF-1alpha, VEGF-A, VEGFR-1, VEGFR-2, MMP-2, TIMP-1, and ADAMTS-1 at the outflow vein, with HIF-1alpha and TIMP-1 levels significantly elevated at day 14 (P < .05). By day 28, the venous stenosis was characterized by a thickened vein wall and neointima. A mouse model of renal insufficiency with AVF was developed that had increased expression of HIF-1alpha, VEGF-A, VEGFR-1, VEGFR-2, MMP-2, TIMP-1, and ADAMTS-1 at the outflow vein with venous stenosis by day 28. Copyright (c) 2010 SIR. Published by Elsevier Inc. All rights reserved.

Role of Spm-Cer-S1P signalling pathway in MMP-2 mediated U46619-induced proliferation of pulmonary artery smooth muscle cells: protective role of epigallocatechin-3-gallate.

Science.gov (United States)

Chowdhury, Animesh; Sarkar, Jaganmay; Chakraborti, Tapati; Chakraborti, Sajal

2015-10-01

During remodelling of pulmonary artery, marked proliferation of pulmonary artery smooth muscle cells (PASMCs) occurs, which contributes to pulmonary hypertension. Thromboxane A2 (TxA2) has been shown to produce pulmonary hypertension. The present study investigates the inhibitory effect of epigallocatechin-3-gallate (EGCG) on the TxA2 mimetic, U46619-induced proliferation of PASMCs. U46619 at a concentration of 10 nM induces maximum proliferation of bovine PASMCs. Both pharmacological and genetic inhibitors of p(38)MAPK, NF-κB and MMP-2 significantly inhibit U46619-induced cell proliferation. EGCG markedly abrogate U46619-induced p(38)MAPK phosphorylation, NF-κB activation, proMMP-2 expression and activation, and also the cell proliferation. U46619 causes an increase in the activation of sphingomyelinase (SMase) and sphingosine kinase (SPHK) and also increase sphingosine 1 phosphate (S1P) level. U46619 also induces phosphorylation of ERK1/2, which phosphorylates SPHK leading to an increase in S1P level. Both pharmacological and genetic inhibitors of SMase and SPHK markedly inhibit U46619-induced cell proliferation. Additionally, pharmacological and genetic inhibitors of MMP-2 markedly abrogate U46619-induced SMase activity and S1P level. EGCG markedly inhibit U46619-induced SMase activity, ERK1/2 and SPHK phosphorylation and S1P level in the cells. Overall, Sphingomyeline-Ceramide-Sphingosine-1-phosphate (Spm-Cer-S1P) signalling axis plays an important role in MMP-2 mediated U46619-induced proliferation of PASMCs. Importantly, EGCG inhibits U46619 induced increase in MMP-2 activation by modulating p(38)MAPK-NFκB pathway and subsequently prevents the cell proliferation. Copyright © 2015 John Wiley & Sons, Ltd.

MtCAS31 Aids Symbiotic Nitrogen Fixation by Protecting the Leghemoglobin MtLb120-1 Under Drought Stress in Medicago truncatula

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Xin Li

2018-05-01

Full Text Available Symbiotic nitrogen fixation (SNF in legume root nodules injects millions of tons of nitrogen into agricultural lands and provides ammonia to non-legume crops under N-deficient conditions. During plant growth and development, environmental stresses, such as drought, salt, cold, and heat stress are unavoidable. This raises an interesting question as to how the legumes cope with the environmental stress along with SNF. Under drought stress, dehydrin proteins are accumulated, which function as protein protector and osmotic substances. In this study, we found that the dehydrin MtCAS31 (cold-acclimation-specific 31 functions in SNF in Medicago truncatula during drought stress. We found that MtCAS31 is expressed in nodules and interacts with leghemoglobin MtLb120-1. The interaction between the two proteins protects MtLb120-1 from denaturation under thermal stress in vivo. Compared to wild type, cas31 mutants display a lower nitrogenase activity, a lower ATP/ADP ratio, higher expression of nodule senescence genes and higher accumulation of amyloplasts under dehydration conditions. The results suggested that MtCAS31 protects MtLb120-1 from the damage of drought stress. We identified a new function for dehydrins in SNF under drought stress, which enriches the understanding of the molecular mechanism of dehydrins.

Effect of Agmatine Sulfate on Modulation of Matrix Metalloproteinases via PI3K/Akt-1 in HT1080 Cells.

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Kim, Hyejeong; Kim, Moon-Moo

2017-11-01

The purpose of this study was to investigate the mechanism by which agmatine sulfate induces an anti-metastatic effect in human HT1080 fibrosarcoma cells, by affecting matrix metalloproteinases (MMPs). For the experiments, we used a non-toxic concentration of agmatine, below 512 μM, that was determined using an MTT assay. The effect of agmatine sulfate on metastasis was gelatin zymography, western blot, immunofluorescence staining and cell invasion assay. Agmatine sulfate inhibited MMP-2 activity stimulated by phenazine methosulfate (PMS). Furthermore, the expression level of MMP-2 stimulated by PMS, was decreased, but the expression level of TIMP-1 was increased in the presence of agmatine sulfate. Moreover, it was observed that the expression levels of ERK and p38 were increased, but those of PI3K and Akt-1 associated with the modulation of MMP-2 were decreased in this study. Furthermore, agmatine sulfate decreased the invasion level of human fibrosarcoma cells stimulated by VEGF. These results suggest that agmatine sulfate could inhibit metastasis through inhibition of MMP-2 via the PI3K/Akt-1 signaling pathway. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

The Vip1 inositol polyphosphate kinase family regulates polarized growth and modulates the microtubule cytoskeleton in fungi.

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Jennifer Pöhlmann

2014-09-01

Full Text Available Microtubules (MTs are pivotal for numerous eukaryotic processes ranging from cellular morphogenesis, chromosome segregation to intracellular transport. Execution of these tasks requires intricate regulation of MT dynamics. Here, we identify a new regulator of the Schizosaccharomyces pombe MT cytoskeleton: Asp1, a member of the highly conserved Vip1 inositol polyphosphate kinase family. Inositol pyrophosphates generated by Asp1 modulate MT dynamic parameters independent of the central +TIP EB1 and in a dose-dependent and cellular-context-dependent manner. Importantly, our analysis of the in vitro kinase activities of various S. pombe Asp1 variants demonstrated that the C-terminal phosphatase-like domain of the dual domain Vip1 protein negatively affects the inositol pyrophosphate output of the N-terminal kinase domain. These data suggest that the former domain has phosphatase activity. Remarkably, Vip1 regulation of the MT cytoskeleton is a conserved feature, as Vip1-like proteins of the filamentous ascomycete Aspergillus nidulans and the distantly related pathogenic basidiomycete Ustilago maydis also affect the MT cytoskeleton in these organisms. Consistent with the role of interphase MTs in growth zone selection/maintenance, all 3 fungal systems show aspects of aberrant cell morphogenesis. Thus, for the first time we have identified a conserved biological process for inositol pyrophosphates.

Non-coding RNA ANRIL and the number of plexiform neurofibromas in patients with NF1 microdeletions

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Mußotter Tanja

2012-10-01

Full Text Available Abstract Background Neurofibromatosis type-1 (NF1 is caused by mutations of the NF1 gene at 17q11.2. In 95% of non-founder NF1 patients, NF1 mutations are identifiable by means of a comprehensive mutation analysis. 5-10% of these patients harbour microdeletions encompassing the NF1 gene and its flanking regions. NF1 is characterised by tumours of the peripheral nerve sheaths, the pathognomonic neurofibromas. Considerable inter- and intra-familial variation in expressivity of the disease has been observed which is influenced by genetic modifiers unrelated to the constitutional NF1 mutation. The number of plexiform neurofibromas (PNF in NF1 patients is a highly heritable genetic trait. Recently, SNP rs2151280 located within the non-coding RNA gene ANRIL at 9p21.3, was identified as being strongly associated with PNF number in a family-based association study. The T-allele of rs2151280, which correlates with reduced ANRIL expression, appears to be associated with higher PNF number. ANRIL directly binds to the SUZ12 protein, an essential component of polycomb repressive complex 2, and is required for SUZ12 occupancy of the CDKN2A/CDKN2B tumour suppressor genes as well as for their epigenetic silencing. Methods Here, we explored a potential association of PNF number and PNF volume with SNP rs2151280 in 29 patients with constitutional NF1 microdeletions using the exact Cochran-Armitage test for trends and the exact Mann–Whitney–Wilcoxon test. Both the PNF number and total tumour volume in these 29 NF1 patients were assessed by whole-body MRI. The NF1 microdeletions observed in these 29 patients encompassed the NF1 gene as well as its flanking regions, including the SUZ12 gene. Results In the 29 microdeletion patients investigated, neither the PNF number nor PNF volume was found to be associated with the T-allele of rs2151280. Conclusion Our findings imply that, at least in patients with NF1 microdeletions, PNF susceptibility is not associated with

TNF-α promotes human retinal pigment epithelial (RPE) cell migration by inducing matrix metallopeptidase 9 (MMP-9) expression through activation of Akt/mTORC1 signaling

International Nuclear Information System (INIS)

Wang, Cheng-hu; Cao, Guo-Fan; Jiang, Qin; Yao, Jin

2012-01-01

Highlights: ► TNF-α induces MMP-9 expression and secretion to promote RPE cell migration. ► MAPK activation is not critical for TNF-α-induced MMP-9 expression. ► Akt and mTORC1 signaling mediate TNF-α-induced MMP-9 expression. ► SIN1 knockdown showed no significant effect on MMP-9 expression by TNF-α. -- Abstract: Tumor necrosis factor-alpha (TNF-α) promotes in vitro retinal pigment epithelial (RPE) cell migration to initiate proliferative vitreoretinopathy (PVR). Here we report that TNF-α promotes human RPE cell migration by inducing matrix metallopeptidase 9 (MMP-9) expression. Inhibition of MMP-9 by its inhibitor or its neutralizing antibody inhibited TNF-α-induced in vitro RPE cell migration. Reversely, exogenously-added active MMP-9 promoted RPE cell migration. Suppression Akt/mTOR complex 1(mTORC1) activation by LY 294002 and rapamycin inhibited TNF-α-mediated MMP-9 expression. To introduce a constitutively active Akt (CA-Akt) in cultured RPE cells increased MMP-9 expression, and to block mTORC1 activation by rapamycin inhibited its effect. RNA interference (RNAi)-mediated silencing of SIN1, a key component of mTOR complex 2 (mTORC2), had no effect on MMP-9 expression or secretion. In conclusion, this study suggest that TNF-α promotes RPE cell migration by inducing MMP-9 expression through activation of Akt/ mTORC1, but not mTORC2 signaling.

TNF-{alpha} promotes human retinal pigment epithelial (RPE) cell migration by inducing matrix metallopeptidase 9 (MMP-9) expression through activation of Akt/mTORC1 signaling

Energy Technology Data Exchange (ETDEWEB)

Wang, Cheng-hu; Cao, Guo-Fan [The Affiliated Eye Hospital of Nanjing Medical University, Nanjing 210029 (China); Jiang, Qin, E-mail: Jqin710@vip.sina.com [The Affiliated Eye Hospital of Nanjing Medical University, Nanjing 210029 (China); Yao, Jin, E-mail: dryaojin@yahoo.com [The Affiliated Eye Hospital of Nanjing Medical University, Nanjing 210029 (China)

2012-08-17

Highlights: Black-Right-Pointing-Pointer TNF-{alpha} induces MMP-9 expression and secretion to promote RPE cell migration. Black-Right-Pointing-Pointer MAPK activation is not critical for TNF-{alpha}-induced MMP-9 expression. Black-Right-Pointing-Pointer Akt and mTORC1 signaling mediate TNF-{alpha}-induced MMP-9 expression. Black-Right-Pointing-Pointer SIN1 knockdown showed no significant effect on MMP-9 expression by TNF-{alpha}. -- Abstract: Tumor necrosis factor-alpha (TNF-{alpha}) promotes in vitro retinal pigment epithelial (RPE) cell migration to initiate proliferative vitreoretinopathy (PVR). Here we report that TNF-{alpha} promotes human RPE cell migration by inducing matrix metallopeptidase 9 (MMP-9) expression. Inhibition of MMP-9 by its inhibitor or its neutralizing antibody inhibited TNF-{alpha}-induced in vitro RPE cell migration. Reversely, exogenously-added active MMP-9 promoted RPE cell migration. Suppression Akt/mTOR complex 1(mTORC1) activation by LY 294002 and rapamycin inhibited TNF-{alpha}-mediated MMP-9 expression. To introduce a constitutively active Akt (CA-Akt) in cultured RPE cells increased MMP-9 expression, and to block mTORC1 activation by rapamycin inhibited its effect. RNA interference (RNAi)-mediated silencing of SIN1, a key component of mTOR complex 2 (mTORC2), had no effect on MMP-9 expression or secretion. In conclusion, this study suggest that TNF-{alpha} promotes RPE cell migration by inducing MMP-9 expression through activation of Akt/ mTORC1, but not mTORC2 signaling.

Food-induced reinforcement is abrogated by the genetic deletion of the MT1 or MT2 melatonin receptor in C3H/HeN mice.

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Clough, Shannon J; Hudson, Randall L; Dubocovich, Margarita L

2018-05-02

Palatable food is known for its ability to enhance reinforcing responses. Studies have suggested a circadian variation in both drug and natural reinforcement, with each following its own time course. The goal of this study was to determine the role of the MT 1 and MT 2 melatonin receptors in palatable snack food-induced reinforcement, as measured by the conditioned place preference (CPP) paradigm during the light and dark phases. C3H/HeN wild-type mice were trained for snack food-induced CPP at either ZT 6 - 8 (ZT: Zeitgeber time; ZT 0 = lights on), when endogenous melatonin levels are low, or ZT 19 - 21, when melatonin levels are high. These time points also correspond to the high and low points for expression of the circadian gene Period1, respectively. The amount of snack food (chow, Cheetos®, Froot Loops® and Oreos®) consumed was of similar magnitude at both times, however only C3H/HeN mice conditioned to snack food at ZT 6 - 8 developed a place preference. C3H/HeN mice with a genetic deletion of either the MT 1 (MT 1 KO) or MT 2 (MT 2 KO) receptor tested at ZT 6 - 8 did not develop a place preference for snack food. Although the MT 2 KO mice showed a similar amount of snack food consumed when compared to wild-type mice, the MT 1 KO mice consumed significantly less than either genotype. We conclude that in our mouse model snack food-induced CPP is dependent on time of day and the presence of the MT 1 or MT 2 receptors, suggesting a role for melatonin and its receptors in snack food-induced reinforcement. Copyright © 2018 Elsevier B.V. All rights reserved.

Use of a C1 Inhibitor Concentrate in Adults ≥65 Years of Age with Hereditary Angioedema

DEFF Research Database (Denmark)

Bygum, Anette; Martinez-Saguer, Inmaculada; Bas, Murat

2016-01-01

BACKGROUND: Treatment of hereditary angioedema (HAE) in 'older adults' (those aged ≥65 years) has not been well studied. The international Berinert Patient Registry collected data on the use of intravenous plasma-derived, pasteurized, nanofiltered C1-inhibitor concentrate (pnfC1-INH; Berinert......(®)/CSL Behring) in patients of any age, including many older adults. METHODS: This observational registry, conducted from 2010 to 2014 at 30 US and seven European sites, gathered prospective (post-enrollment) and retrospective (pre-enrollment) usage and adverse event (AE) data on subjects treated with pnfC1-INH....... RESULTS: The registry documented 1701 pnfC1-INH infusions in 27 older adults. A total of 1511 HAE attacks treated with pnfC1-INH administration were reported among 25 of the 27 (92.6 %) older adults. Among the older adults, mean (standard deviation [SD]) (8.8 [4.1] IU/kg) and median (6.4 IU/kg) pnfC1-INH...

Modulation of MMP-2 and MMP-9 in Churg-Strauss syndrome respiratory mucosa: potential monitoring parameters.

Science.gov (United States)

Leone, A; Uzzo, M L; Gerbino, A; Tortorici, S; Tralongo, P; Cappello, F; Incandela, S; Spatola, G F; Jurjus, A R

2014-01-01

Churg-Strauss (CSS) syndrome is rare and of unknown etiology. It is associated with vasculitis, blood eosinophilia and granulomatosis, and affects multiple organs and systems at various stages of the disease. Specific diagnostic and monitoring tests are not yet available. This study aims to assess the changes in MMP-2 and MMP-9 along with the histopathological alterations in two cases of CSS, as possible potential diagnostic and monitoring criteria. Two adult male patients were diagnosed with CSS in the otorhinolaryngology clinic in the University of Palermo, based on multiple clinical and histopathologic criteria. Biopsies of respiratory mucosa were taken after the consent of the patients, processed for routine histopathology and immunohistochemistry as well as quantitative polymerase chain reaction (qPCR). Similar biopsies were also taken from a non- CSS patient. The Assessment of MMP-2 and MMP-9 was performed using both immunohistochemistry and qPCR techniques. Histopathological alterations in the respiratory mucosa were consistent with vasculitis and granulomatous tissue formation, in addition to inflammatory cell infiltration with abundance of eosinophils. Immunohistochemistry assay performed on the samples derived from the two CSS patients showed a relative and remarkable increase of both MMP-2 and MMP-9 compared to controls. Such an increase was consistent with the qPCR results which depicted a significant increase between 20 and 30% for both MMP-2 and MMP-9, respectively. Since the secretion of MMPs is an essential step in angiogenesis, could these enzymatic factors be used as parameters to diagnose or monitor the evolution of CSS? The small number of samples analyzed in this study does not allow us to suggest a general statement correlating the increase in expression of MMP-2 and MMP-9 to the appearance or evolution of vasculitis; it is only speculative.

MT1 melatonin receptors and their role in the oncostatic action of melatonin

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Karolina Danielczyk

2009-09-01

Full Text Available Melatonin, the main hormone produced by the pineal gland, strongly inhibits the growth of cancer cells [i]in vitro[/i] and [i]in vivo[/i]. Some publications indicate that the addition of melatonin to culture medium slows the proliferation of some cancer cell lines. It is also suggested that melatonin used as an adjuvant benefits the effectiveness and tolerance of chemotherapy. The mechanisms of this are not fully understood, but melatonin receptors might be one of the most important elements. Two distinct types of membrane-bound melatonin receptors have been identified in humans: MT1 (Mel1a and MT2 (Mel1b receptors. These subtypes are 60�0homologous at the amino-acid level. MT1 receptors are G-protein-coupled receptors. Through the α subunit of G protein, melatonin receptors stimulate an adenylate cyclase and decrease the level of cAMP. This has a significant influence on cell proliferation and has been confirmed in many tests on different cell lines, such as S-19, B-16 murine melanoma cells, and breast cancer cells. It seems that expression of the MT1 melatonin receptors benefits the efficacy of melatonin treatment. Melatonin and its receptors may provide a promising way to establish new alternative therapeutic approaches in human cancer prevention.

Association between promoter -1607 polymorphism of MMP1 and Lumbar Disc Disease in Southern Chinese

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Leong John CY

2008-04-01

Full Text Available Abstract Background Matrix metalloproteinases (MMPs are involved in the degradation of the extracellular matrix of the intervertebral disc. A SNP for guanine insertion/deletion (G/D, the -1607 promoter polymorphism, of the MMP1 gene was found significantly affecting promoter activity and corresponding transcription level. Hence it is a good candidate for genetic studies in DDD. Methods Southern Chinese volunteers between 18 and 55 years were recruited from the population. DDD in the lumbar spine was defined by MRI using Schneiderman's classification. Genomic DNA was isolated from the leukocytes and genotyping was performed using the Sequenom® platform. Association and Hardy-Weinberg equilibrium checking were assessed by Chi-square test and Mann-Whitney U test. Results Our results showed substantial evidence of association between -1607 promoter polymorphism of MMP1 and DDD in the Southern Chinese subjects. D allelic was significantly associated with DDD (p value = 0.027, odds ratio = 1.41 with 95% CI = 1.04–1.90 while Genotypic association on the presence of D allele was also significantly associated with DDD (p value = 0.046, odds ratio = 1.50 with 95% CI = 1.01–2.24. Further age stratification showed significant genotypic as well as allelic association in the group of over 40 years (genotypic: p value = 0.035, odds ratio = 1.617 with 95% CI = 1.033–2.529; allelic: p value = 0.033, odds ratio = 1.445 with 95% CI = 1.029–2.029. Disc bulge, annular tears and the Schmorl's nodes were not associated with the D allele. Conclusion We demonstrated that individuals with the presence of D allele for the -1607 promoter polymorphism of MMP1 are about 1.5 times more susceptible to develop DDD when compared with those having G allele only. Further association was identified in individuals over 40 years of age. Disc bulge, annular tear as well as Schmorl's nodes were not associated with this polymorphism.

Increased expressions of MMP-2 and MMP-9 in lung following 12 Gy local irradiation

International Nuclear Information System (INIS)

Yang Kunyu; Liu Li; Zhang Tao; Wu Gang; Hu Yu; Ruebe, C.; Ruebe, C.

2006-01-01

Objective: To measure expressions of metalloproteinases and tissue inhibitors of metalloproteinases in the lung following thoracic irradiation of 12 Gy, and explore its possible role in the development of radiation-induced lung damage. Methods: C57BL/6J mice at age of 8 weeks were thoracically irradiated with 12 Gy X-rays (10 MV, 2.4 Gy/min, single exposure), and the control mice were sham-irradiated. The mice were sacrificed at 4 or 8 weeks after thoracic irradiation by decapitation. Lung tissues samples were collected. Expressions of MMP-2, MMP-9, MMP-3, MMP-13, TIMP-1, TIMP-2, and TIMP-3 in lung samples were measured. Results: There was no significant difference in expressions of MMP-3, MMP-13, TIMP-1 TIMP-2, and TIMP-3 in the lung between the two groups at 4 and 8 weeks after thoracic irradiation (or sham-irradiation). However, the expressions of MMP-2 were enhanced by 1.7 and 1.9 folds, and MMP-9 by 2.7 and 2.6 folds at 4 and 8 weeks after thoracic irradiation, respectively. Conclusion: Enhanced expressions of MMP-2 and MMP-9 in the lung were involved in the development of acute lung injury after thoracic irradiation, leading to a disruption of the structure and fibrosis. (authors)

Inactivation of Mycobacterium tuberculosis l,d-Transpeptidase LdtMt1 by Carbapenems and Cephalosporins

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Dubée, Vincent; Triboulet, Sébastien; Mainardi, Jean-Luc; Ethève-Quelquejeu, Mélanie; Gutmann, Laurent; Marie, Arul; Dubost, Lionel

2012-01-01

The structure of Mycobacterium tuberculosis peptidoglycan is atypical since it contains a majority of 3→3 cross-links synthesized by l,d-transpeptidases that replace 4→3 cross-links formed by the d,d-transpeptidase activity of classical penicillin-binding proteins. Carbapenems inactivate these l,d-transpeptidases, and meropenem combined with clavulanic acid is bactericidal against extensively drug-resistant M. tuberculosis. Here, we used mass spectrometry and stopped-flow fluorimetry to investigate the kinetics and mechanisms of inactivation of the prototypic M. tuberculosis l,d-transpeptidase LdtMt1 by carbapenems (meropenem, doripenem, imipenem, and ertapenem) and cephalosporins (cefotaxime, cephalothin, and ceftriaxone). Inactivation proceeded through noncovalent drug binding and acylation of the catalytic Cys of LdtMt1, which was eventually followed by hydrolysis of the resulting acylenzyme. Meropenem rapidly inhibited LdtMt1, with a binding rate constant of 0.08 μM−1 min−1. The enzyme was unable to recover from this initial binding step since the dissociation rate constant of the noncovalent complex was low (carbapenem side chains affected both the binding and acylation steps, ertapenem being the most efficient LdtMt1 inactivator. Cephalosporins also formed covalent adducts with LdtMt1, although the acylation reaction was 7- to 1,000-fold slower and led to elimination of one of the drug side chains. Comparison of kinetic constants for drug binding, acylation, and acylenzyme hydrolysis indicates that carbapenems and cephems can both be tailored to optimize peptidoglycan synthesis inhibition in M. tuberculosis. PMID:22615283

Tumor necrosis factor-α-accelerated degradation of type I collagen in human skin is associated with elevated matrix metalloproteinase (MMP)-1 and MMP-3 ex vivo

DEFF Research Database (Denmark)

Ågren, Magnus S; Schnabel, Reinhild; Christensen, Lise H

2015-01-01

Tumor necrosis factor (TNF)-α induces matrix metalloproteinases (MMPs) that may disrupt skin integrity. We have investigated the effects and mechanisms of exogenous TNF-α on collagen degradation by incubating human skin explants in defined serum-free media with or without TNF-α (10ng/ml) in the a......Tumor necrosis factor (TNF)-α induces matrix metalloproteinases (MMPs) that may disrupt skin integrity. We have investigated the effects and mechanisms of exogenous TNF-α on collagen degradation by incubating human skin explants in defined serum-free media with or without TNF-α (10ng...... tissue-derived collagenolytic activity with TNF-α exposure was blocked by neutralizing MMP-1 monoclonal antibody and was not due to down-regulation of tissue inhibitor of metalloproteinase-1. TNF-α increased production (pendogenous MMP-1...

Synthesis and preliminary biological evaluation of new radioiodinated MMP inhibitors for imaging MMP activity in vivo

International Nuclear Information System (INIS)

Kopka, Klaus; Breyholz, Hans-Joerg; Wagner, Stefan; Law, Marilyn P.; Riemann, Burkhard; Schroeer, Sandra; Trub, Monika; Guilbert, Benedicte; Levkau, Bodo; Schober, Otmar; Schaefers, Michael

2004-01-01

Non-invasive measurement of matrix metalloproteinase (MMP) activity in vivo is a clinical challenge in many disease processes such as inflammation, tumor metastasis and atherosclerosis. Therefore, radioiodinated analogues of the non-peptidyl broad-spectrum MMP inhibitor (MMPI) CGS 27023A 1a were synthesized for non-invasive detection of MMP activity in vivo using single photon emission computed tomography (SPECT). The compounds Br-CGS 27023A 1b and HO-CGS 27023A 1d were synthesized from the amino acid D-valine and used as precursors for radioiodinated derivatives of CGS 27023A and their non-radioactive references I-CGS 27023A 1c and HO-I-CGS 27023A 1e. Radioiodination of the precursors with [ 123 I]NaI or [ 125 I]NaI produced the no-carrier-added MMP inhibitors [ 123 I]I-CGS 27023A 1f, [ 125 I]I-CGS 27023A 1g, HO-[ 123 I]I-CGS27023A 1h, and HO-[ 125 I]I-CGS 27023A 1i. In vitro studies showed that the non-radioactive analogues of the MMP inhibitors exhibited affinities against gelatinase A (MMP-2) and gelatinase B (MMP-9) in the nanomolar range, comparable to the parent compound CGS 27023A. In vivo biodistribution using HO-[ 125 I]I-CGS 27023A 1i in CL57 Bl6 mice showed rapid blood and plasma clearance and low retention in normal tissues. The preliminary biological evaluation warrant further studies of these radioiodinated MMP inhibitors as potential new radiotracers for imaging MMP activity in vivo

Matrix metalloproteinase-10/TIMP-2 structure and analyses define conserved core interactions and diverse exosite interactions in MMP/TIMP complexes.

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Batra, Jyotica; Soares, Alexei S; Mehner, Christine; Radisky, Evette S

2013-01-01

Matrix metalloproteinases (MMPs) play central roles in vertebrate tissue development, remodeling, and repair. The endogenous tissue inhibitors of metalloproteinases (TIMPs) regulate proteolytic activity by binding tightly to the MMP active site. While each of the four TIMPs can inhibit most MMPs, binding data reveal tremendous heterogeneity in affinities of different TIMP/MMP pairs, and the structural features that differentiate stronger from weaker complexes are poorly understood. Here we report the crystal structure of the comparatively weakly bound human MMP-10/TIMP-2 complex at 2.1 Å resolution. Comparison with previously reported structures of MMP-3/TIMP-1, MT1-MMP/TIMP-2, MMP-13/TIMP-2, and MMP-10/TIMP-1 complexes offers insights into the structural basis of binding selectivity. Our analyses identify a group of highly conserved contacts at the heart of MMP/TIMP complexes that define the conserved mechanism of inhibition, as well as a second category of diverse adventitious contacts at the periphery of the interfaces. The AB loop of the TIMP N-terminal domain and the contact loops of the TIMP C-terminal domain form highly variable peripheral contacts that can be considered as separate exosite interactions. In some complexes these exosite contacts are extensive, while in other complexes the AB loop or C-terminal domain contacts are greatly reduced and appear to contribute little to complex stability. Our data suggest that exosite interactions can enhance MMP/TIMP binding, although in the relatively weakly bound MMP-10/TIMP-2 complex they are not well optimized to do so. Formation of highly variable exosite interactions may provide a general mechanism by which TIMPs are fine-tuned for distinct regulatory roles in biology.

Changes of MMP-9 and IL-1β in serum and cerebrospinal-fluid in children with central nervous system infection

Institute of Scientific and Technical Information of China (English)

Shu-Qin Jiao

2016-01-01

Objective:To provide a new basis for the detection of the central nervous system infection cases, we explored and compared the role of cerebrospinal-fluid (CSF), matrix metalloproteinases 9 (MMP-9) and interleukin 1 (the level of IL -1β) in the central nervous system (CNS).Methods:Sixty cases of children acute central nervous system infection were selected, including 30 cases of viral encephalitis children (VE) and 30 cases of purulent meningitis children (PM). Forty cases of non-central nervous system infection children were control group. The serum albumin (SA1b) of each group was detected by full-automatic analysis instrument, and the CSF albumin (CA1b) was detected by immunoephelometry and the albumin index (AQ) was accounted. ELISE was used to detect the levels of MMP-9 and IL-1β in serum and cerebrospinal-fluid.Results:The level of MMP-9 in the serum of groups of VE, PM and control were (267.84 ± 91.88) μg/L, (488.98 ± 159.07) μg/L and (133.04 ± 31.68) μg/L, while in the CSF were (37.18 ± 17.78) μg/L, (117.9 ± 42.87) μg/L and (10.36 ± 5.43) μg/L; The level of IL-1β in serum of groups of PM, VE and control were (19.69 ± 11.12) ng/L, (24.37 ± 4.13) ng/L and (15.01 ± 3.89) ng/L, while in the CSF were (66.94 ± 10.65) ng/L, (106.27 ± 12.79) ng/L and (49.98 ± 12.59) ng/L; The level of CAlb were (0.53 ± 0.15) g/L, (1.05 ± 0.27) g/L and (0.17 ± 0.07) g/L and AQ were (13.75 ± 3.44), (26.99 ± 7.28) and (4.63 ± 2.04). The PM, VE were respectively compared with the control, the levels of IL-1β and MMP-9 in serum and CSF all increased, with statistically significant difference.The VE, compared to the PM, the level of IL-1β in serum and CSF all decreased, with statistically significant difference; The level of MMP-9 in serum and CSF all decreased, with statistically significant difference. The level of CA1b and AQ in the VE and PM all increased, with a statistically significant difference. The level of MMP-9 and IL-1β in serum and CSF of the

The effect of adjunctive chlorhexidine mouthrinse on GCF MMP-8 and TIMP-1 levels in gingivitis: a randomized placebo-controlled study

Science.gov (United States)

2014-01-01

Background The aim of the present study was to evaluate the effect of adjunctive chlorhexidine (CHX) mouthrinse on gingival crevicular fluid (GCF) MMP-8 and TIMP-1 levels in plaque-associated gingivitis. Methods A total of 50 gingivitis patients were included in the present study. In addition to daily plaque control, CHX group rinsed with CHX, while placebo group rinsed with placebo mouthrinse for 4 weeks. GCF samples were collected, and clinical parameters including plaque index, papillary bleeding index, calculus index and pocket depth were recorded at baseline and 4 weeks. GCF MMP-8 and TIMP-1 levels were determined by immunofluorometric assay (IFMA) and enzyme-linked immunosorbent assay (ELISA), respectively. Results In both groups, GCF MMP-8 levels of anterior and posterior sites at four weeks were not different from baseline (p > 0.05). There were no significant differences in GCF MMP-8 levels between the study groups at four weeks (p > 0.05). GCF TIMP-1 levels of anterior and posterior sites at four weeks were higher compared to baseline in both groups (p  0.05). Conclusions CHX usage had no significant effects on the GCF MMP-8 and TIMP-1 levels in plaque-associate gingivitis. However, daily plaque control resulted in the increase of GCF TIMP-1 levels regardless of CHX usage. PMID:24886536

Synthesis and preliminary biological evaluation of new radioiodinated MMP inhibitors for imaging MMP activity in vivo

Energy Technology Data Exchange (ETDEWEB)

Kopka, Klaus E-mail: kopka@uni-muenster.de; Breyholz, Hans-Joerg; Wagner, Stefan; Law, Marilyn P.; Riemann, Burkhard; Schroeer, Sandra; Trub, Monika; Guilbert, Benedicte; Levkau, Bodo; Schober, Otmar; Schaefers, Michael

2004-02-01

Non-invasive measurement of matrix metalloproteinase (MMP) activity in vivo is a clinical challenge in many disease processes such as inflammation, tumor metastasis and atherosclerosis. Therefore, radioiodinated analogues of the non-peptidyl broad-spectrum MMP inhibitor (MMPI) CGS 27023A 1a were synthesized for non-invasive detection of MMP activity in vivo using single photon emission computed tomography (SPECT). The compounds Br-CGS 27023A 1b and HO-CGS 27023A 1d were synthesized from the amino acid D-valine and used as precursors for radioiodinated derivatives of CGS 27023A and their non-radioactive references I-CGS 27023A 1c and HO-I-CGS 27023A 1e. Radioiodination of the precursors with [{sup 123}I]NaI or [{sup 125}I]NaI produced the no-carrier-added MMP inhibitors [{sup 123}I]I-CGS 27023A 1f, [{sup 125}I]I-CGS 27023A 1g, HO-[{sup 123}I]I-CGS27023A 1h, and HO-[{sup 125}I]I-CGS 27023A 1i. In vitro studies showed that the non-radioactive analogues of the MMP inhibitors exhibited affinities against gelatinase A (MMP-2) and gelatinase B (MMP-9) in the nanomolar range, comparable to the parent compound CGS 27023A. In vivo biodistribution using HO-[{sup 125}I]I-CGS 27023A 1i in CL57 Bl6 mice showed rapid blood and plasma clearance and low retention in normal tissues. The preliminary biological evaluation warrant further studies of these radioiodinated MMP inhibitors as potential new radiotracers for imaging MMP activity in vivo.

In vitro progesterone modulation on bacterial endotoxin-induced production of IL-1β, TNFα, IL-6, IL-8, IL-10, MIP-1α, and MMP-9 in pre-labor human term placenta.

Science.gov (United States)

Garcia-Ruíz, G; Flores-Espinosa, P; Preciado-Martínez, E; Bermejo-Martínez, L; Espejel-Nuñez, A; Estrada-Gutierrez, G; Maida-Claros, R; Flores-Pliego, A; Zaga-Clavellina, Veronica

2015-10-07

During human pregnancy, infection/inflammation represents an important factor that increases the risk of developing preterm labor. The purpose of this study was to determine if pre-treatment with progesterone has an immunomodulatory effect on human placenta production of endotoxin-induced inflammation and degradation of extracellular matrix markers. Placentas were obtained under sterile conditions from pregnancies delivered at term before the onset of labor by cesarean section. Explants from central cotyledons of 10 human placentas were pre-treated with different concentrations of progesterone (0.01, 01, 1.0 μM) and then stimulated with 1000 ng/mL of LPS of Escherichia coli. Cytokines TNFα, IL-1β, IL-6, IL-8, MIP-1α, IL-10 concentrations in the culture medium were then measured by specific ELISA. Secretion profile of MMP-9 was evaluated by ELISA and zymogram. Statistical differences were determined by one-way ANOVA followed by the appropriate ad hoc test; P progesterone significantly blunted (73, 56, 56, 75, 25, 48 %) the secretion of TNF-α, IL-1β, IL-6, IL-8, MIP-1α, IL-10, respectively. The MMP-9 induced by LPS treatment was inhibited only with the highest concentration of progesterone. Mifepristone (RU486) blocked the immunosuppressive effect of progesterone. The present results support the concept that progesterone could be part of the compensatory mechanism that limits the inflammation-induced cytotoxic effects associated with an infection process during gestation.

BubR1 Acts as a Promoter in Cellular Motility of Human Oral Squamous Cancer Cells through Regulating MMP-2 and MMP-9

Directory of Open Access Journals (Sweden)

Chou-Kit Chou

2015-07-01

Full Text Available BubR1 is a critical component of spindle assembly checkpoint, ensuring proper chromatin segregation during mitosis. Recent studies showed that BubR1 was overexpressed in many cancer cells, including oral squamous cell carcinomas (OSCC. However, the effect of BubR1 on metastasis of OSCC remains unclear. This study aimed to unravel the role of BubR1 in the progression of OSCC and confirm the expression of BubR1 in a panel of malignant OSCC cell lines with different invasive abilities. The results of quantitative real-time PCR showed that the mRNA level of BubR1 was markedly increased in four OSCC cell lines, Ca9-22, HSC3, SCC9 and Cal-27 cells, compared to two normal cells, normal human oral keratinocytes (HOK and human gingival fibroblasts (HGF. Moreover, the expression of BubR1 in these four OSCC cell lines was positively correlated with their motility. Immunofluorescence revealed that BubR1 was mostly localized in the cytosol of human gingival carcinoma Ca9-22 cells. BubR1 knockdown significantly decreased cellular invasion but slightly affect cellular proliferation on both Ca9-22 and Cal-27 cells. Consistently, the activities of metastasis-associated metalloproteinases MMP-2 and MMP-9 were attenuated in BubR1 knockdown Ca9-22 cells, suggesting the role of BubR1 in promotion of OSCC migration. Our present study defines an alternative pathway in promoting metastasis of OSCC cells, and the expression of BubR1 could be a prognostic index in OSCC patients.

Rac1/β-Catenin Signalling Pathway Contributes to Trophoblast Cell Invasion by Targeting Snail and MMP9

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Minghua Fan

2016-03-01

Full Text Available Background/Aims: Preeclampsia is an idiopathic and serious complication during gestation in which placental trophoblast cells differentiate into several functional subtypes, including highly invasive extravillous trophoblasts (EVTs. Although the cause and pathogenesis of preeclampsia have remained unclear, numerous studies have suggested that the inadequacy of EVT invasion leads to imperfect uterine spiral artery remodelling, which plays a crucial role in the development of preeclampsia. Rac1, or Ras-related C3 botulinum toxin substrate 1, was found to be a key regulator of the migration, invasion uand apoptosis of various tumour cells. Because EVTs share similar invasive and migratory biological behaviours with malignant cells, this study aimed to determine whether the Rac1 signalling pathway affects trophoblast invasion and is thus involved in the pathogenesis of preeclampsia. Methods: We measured the activity of Rac1 and its downstream targets, β-catenin, Snail and MMP9 in placental tissues from patients experiencing a normal pregnancy and those with preeclampsia. Furthermore, we treated HTR-8/SVneo cells with a shRNA Rac1 vector and the β-catenin inhibitor IWP-2 and explored Rac1 signalling pathway activation as well as the effects of Snail and β-catenin on trophoblast invasion. Results: In placental samples from patients experiencing a normal pregnancy and those with preeclampsia, active Rac1 levels and MMP9 protein and mRNA levels were significantly decreased in term pregnancy samples compared to early pregnancy samples. Lower levels were found in preeclampsia samples than in normal term pregnancy samples, and these levels significantly declined in severe preeclampsia samples compared with mild preeclampsia samples. Further analyses demonstrated that both Rac1 shRNA and the β-catenin inhibitor significantly suppressed MMP9 and Snail activation in trophoblasts, thus impairing trophoblast invasion. Notably, silencing Rac1 down

Effects of Mutations on Structure-Function Relationships of Matrix Metalloproteinase-1.

Science.gov (United States)

Singh, Warispreet; Fields, Gregg B; Christov, Christo Z; Karabencheva-Christova, Tatyana G

2016-10-14

Matrix metalloproteinase-1 (MMP-1) is one of the most widely studied enzymes involved in collagen degradation. Mutations of specific residues in the MMP-1 hemopexin-like (HPX) domain have been shown to modulate activity of the MMP-1 catalytic (CAT) domain. In order to reveal the structural and conformational effects of such mutations, a molecular dynamics (MD) study was performed of in silico mutated residues in the X-ray crystallographic structure of MMP-1 complexed with a collagen-model triple-helical peptide (THP). The results indicate an important role of the mutated residues in MMP-1 interactions with the THP and communication between the CAT and the HPX domains. Each mutation has a distinct impact on the correlated motions in the MMP-1•THP. An increased collagenase activity corresponded to the appearance of a unique anti-correlated motion and decreased correlated motions, while decreased collagenase activity corresponded both to increased and decreased anti-correlated motions.

Effects of Mutations on Structure–Function Relationships of Matrix Metalloproteinase-1

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Warispreet Singh

2016-10-01

Full Text Available Matrix metalloproteinase-1 (MMP-1 is one of the most widely studied enzymes involved in collagen degradation. Mutations of specific residues in the MMP-1 hemopexin-like (HPX domain have been shown to modulate activity of the MMP-1 catalytic (CAT domain. In order to reveal the structural and conformational effects of such mutations, a molecular dynamics (MD study was performed of in silico mutated residues in the X-ray crystallographic structure of MMP-1 complexed with a collagen-model triple-helical peptide (THP. The results indicate an important role of the mutated residues in MMP-1 interactions with the THP and communication between the CAT and the HPX domains. Each mutation has a distinct impact on the correlated motions in the MMP-1•THP. An increased collagenase activity corresponded to the appearance of a unique anti-correlated motion and decreased correlated motions, while decreased collagenase activity corresponded both to increased and decreased anti-correlated motions.

PNF 2.0? Initial evidence that gamification can increase the efficacy of brief, web-based personalized normative feedback alcohol interventions.

Science.gov (United States)

Boyle, Sarah C; Earle, Andrew M; LaBrie, Joseph W; Smith, Daniel J

2017-04-01

Gamified interventions exploit the motivational characteristics of a game in order to provide prevention information and promote behavior change. Despite the modest effect sizes observed in increasingly popular web-based personalized normative feedback (PNF) alcohol interventions for college students, previous research has yet to consider how gamification might be used to enhance efficacy. This study examines whether a novel, gamified PNF intervention format, which includes a point-based reward system, the element of chance, and personal icons to visually represent users, is more effective in reducing short-term alcohol use than the standard web-based style of PNF currently used on college campuses. Two-hundred and thirty-seven college students were randomly assigned to receive either a standard brief, web-based PNF alcohol intervention or the same alcohol intervention components delivered within a Facebook-connected social game called CampusGANDR (Gamified Alcohol Norm Discovery and Readjustment). In both study conditions participants answered identical questions about their perceptions of peer drinking norms and own drinking and then received the same PNF slides. Two weeks following PNF delivery, participants again reported their perceptions of peers' alcohol use and own drinking. Students in the CampusGANDR condition reported significantly reduced peer drinking norms and alcohol use at the two-week follow-up relative to students who received identical PNF delivered by standard online survey. Further, a mediation model demonstrated that this effect was driven by larger reductions in perceived drinking norms among participants assigned to receive CampusGANDR, relative to control. As web-based PNF is becoming an increasingly universal prevention strategy, findings from this study suggest gamification may represent one method by which intervention efficacy could be substantially improved. The potential methodological and economic benefits associated with gamified

Optimization of Beam Transmission of PAL-PNF Electron Linac

Energy Technology Data Exchange (ETDEWEB)

Shin, S. G.; Kim, S. K.; Kim, E. A. [Pohang University of Science and Technology, Pohang (Korea, Republic of)

2012-05-15

The PNF (Pohang Neutron Facility) electron Linac is providing converted neutrons and photons from electron beams to users for nuclear physics experiments and high energy gamma-ray exposures. This linac is capable of producing 100 MeV electron beams with a beam current of pulsed 100 mA. The pulse length is 2 {mu}s and the pulse repetition rate is typically 30 Hz. This linac consists of two SLAC-type S-band accelerating columns and the thermionic RF gun. They are powered by one klystron and the matching pulse modulator. The electron beams emitted from the RF gun are bunched as they pass through the alpha magnet and are injected into the accelerating column thereafter. In this paper, we discuss procedures and results of the beam transmission optimization with technical details of the accelerator system. We also briefly discuss the future upgrade plan to obtain short-pulse or electron beams for neutron TOF experiments by adopting a triode type thermionic DC electron gun

The Involvement of Mutual Inhibition of ERK and mTOR in PLCγ1-Mediated MMP-13 Expression in Human Osteoarthritis Chondrocytes

Directory of Open Access Journals (Sweden)

Zejun Liu

2015-08-01

Full Text Available The issue of whether ERK activation determines matrix synthesis or degradation in osteoarthritis (OA pathogenesis currently remains controversial. Our previous study shows that PLCγ1 and mTOR are involved in the matrix metabolism of OA cartilage. Investigating the interplays of PLCγ1, mTOR and ERK in matrix degradation of OA will facilitate future attempts to manipulate ERK in OA prevention and therapy. Here, cultured human normal chondrocytes and OA chondrocytes were treated with different inhibitors or transfected with expression vectors, respectively. The levels of ERK, p-ERK, PLCγ1, p-PLCγ1, mTOR, p-mTOR and MMP-13 were then evaluated by Western blotting analysis. The results manifested that the expression level of ERK in human OA chondrocytes was lower than that in human normal articular chondrocytes, and the up-regulation of ERK could promote matrix synthesis, including the decrease in MMP-13 level and the increase in Aggrecan level in human OA chondrocytes. Furthermore, the PLCγ1/ERK axis and a mutual inhibition of mTOR and ERK were observed in human OA chondrocytes. Interestingly, activated ERK had no inhibitory effect on MMP-13 expression in PLCγ1-transformed OA chondrocytes. Combined with our previous study, the non-effective state of ERK activation by PLCγ1 on MMP-13 may be partly attributed to the inhibition of the PLCγ1/mTOR axis on the PLCγ1/ERK axis. Therefore, the study indicates that the mutual inhibition of ERK and mTOR is involved in PLCγ1-mediated MMP-13 expression in human OA chondrocytes, with important implication for the understanding of OA pathogenesis as well as for its prevention and therapy.

Matrix metalloproteinase-10/TIMP-2 structure and analyses define conserved core interactions and diverse exosite interactions in MMP/TIMP complexes.

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Jyotica Batra

Full Text Available Matrix metalloproteinases (MMPs play central roles in vertebrate tissue development, remodeling, and repair. The endogenous tissue inhibitors of metalloproteinases (TIMPs regulate proteolytic activity by binding tightly to the MMP active site. While each of the four TIMPs can inhibit most MMPs, binding data reveal tremendous heterogeneity in affinities of different TIMP/MMP pairs, and the structural features that differentiate stronger from weaker complexes are poorly understood. Here we report the crystal structure of the comparatively weakly bound human MMP-10/TIMP-2 complex at 2.1 Å resolution. Comparison with previously reported structures of MMP-3/TIMP-1, MT1-MMP/TIMP-2, MMP-13/TIMP-2, and MMP-10/TIMP-1 complexes offers insights into the structural basis of binding selectivity. Our analyses identify a group of highly conserved contacts at the heart of MMP/TIMP complexes that define the conserved mechanism of inhibition, as well as a second category of diverse adventitious contacts at the periphery of the interfaces. The AB loop of the TIMP N-terminal domain and the contact loops of the TIMP C-terminal domain form highly variable peripheral contacts that can be considered as separate exosite interactions. In some complexes these exosite contacts are extensive, while in other complexes the AB loop or C-terminal domain contacts are greatly reduced and appear to contribute little to complex stability. Our data suggest that exosite interactions can enhance MMP/TIMP binding, although in the relatively weakly bound MMP-10/TIMP-2 complex they are not well optimized to do so. Formation of highly variable exosite interactions may provide a general mechanism by which TIMPs are fine-tuned for distinct regulatory roles in biology.

Transcriptional Inhibition of Matrix Metal loproteinase 9 (MMP-9 Activity by a c-fos/Estrogen Receptor Fusion Protein is Mediated by the Proximal AP-1 Site of the MMP-9 Promoter and Correlates with Reduced Tumor Cell Invasion

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David L. Crowe

1999-10-01

Full Text Available Tumor cell invasion of basement membranes is one of the hallmarks of malignant transformation. Tumor cells secrete proteolytic enzymes known as matrix metalloproteinases (MMPs which degrade extracellular matrix molecules. Increased expression of MMP-9 has been associated with acquisition of invasive phenotype in many tumors. However, multiple mechanisms for regulation of MMP-9 gene expression by tumor cell lines have been proposed. A number of transcription factor binding sites have been characterized in the upstream regulatory region of the MMP-9 gene, including those for AP-1. To determine how a specific AP-1 family member, c-fos, regulates MMP-9 promoter activity through these sites, we used an expression vector containing the c-fos coding region fused to the estrogen receptor (ER ligand binding domain. This construct is activated upon binding estradiol. Stable expression of this construct in ER negative squamous cell carcinoma (SCC lines produced an estradiol dependent decrease in the number of cells that migrated through a reconstituted basement membrane. This decreased invasiveness was accompanied by estradiol dependent downregulation of MMP-9 activity as determined by gelatin zymography. Estradiol also produced transcriptional downregulation of an MMP-9 promoter construct in cells transiently transfected with the c-fosER expression vector. This downregulation was mediated by the AP-1 site at —79 by in the MMP-9 promoter. We concluded that the proximal AP-1 site mediated the transcriptional downregulation of the MMP-9 promoter by a conditionally activated c-fos fusion protein.

Tissue inhibitor of metalloproteinase-2 (TIMP-2) regulates myogenesis and β1 integrin expression in vitro

International Nuclear Information System (INIS)

Lluri, Gentian; Langlois, Garret D.; Soloway, Paul D.; Jaworski, Diane M.

2008-01-01

Myogenesis in vitro involves myoblast cell cycle arrest, migration, and fusion to form multinucleated myotubes. Extracellular matrix (ECM) integrity during these processes is maintained by the opposing actions of matrix metalloproteinase (MMP) proteases and their inhibitors, the tissue inhibitor of metalloproteinases (TIMPs). Here, we report that TIMP-2, MMP-2, and MT1-MMP are differentially expressed during mouse myoblast differentiation in vitro. A specific role for TIMP-2 in myogenesis is demonstrated by altered TIMP-2 -/- myotube formation. When differentiated in horse serum-containing medium, TIMP-2 -/- myotubes are larger than wild-type myotubes. In contrast, when serum-free medium is used, TIMP-2 -/- myotubes are smaller than wild-type myotubes. Regardless of culture condition, myotube size is directly correlated with MMP activity and inversely correlated with β1 integrin expression. Treatment with recombinant TIMP-2 rescues reduced TIMP-2 -/- myotube size and induces increased MMP-9 activation and decreased β1 integrin expression. Treatment with either MMP-2 or MMP-9 similarly rescues reduced myotube size, but has no effect on β1 integrin expression. These data suggest a specific regulatory relationship between TIMP-2 and β1 integrin during myogenesis. Elucidating the role of TIMP-2 in myogenesis in vitro may lead to new therapeutic options for the use of TIMP-2 in myopathies and muscular dystrophies in vivo

Periodic disruptions in the MT-1 tokamak

International Nuclear Information System (INIS)

Zoletnik, S.

1988-11-01

Disruptive instabilities are common phenomena in toroidal devices, especially in tokamaks. Three types can be distinguished: internal, minor and major disruptions. Periodic minor disruptions in the MT-1 tokamak were measured systematically with values of the limiter safety factor between 4 and 10. The density limit as a function of plasma current and horizontal displacement was investigated. Precursor oscillations always appear before the instability with increasing amplitude but can be observed at the density limit with quasi-stationary amplitude. Phase correlation between precursor oscillations were measured with Mirnov coils and x-ray detectors, and they show good agreement with a simple magnetic island model. (R.P.) 11 refs.; 6 figs

Germacrane sesquiterpenes isolated from the rhizome of Curcuma xanthorrhiza Roxb. inhibit UVB-induced upregulation of MMP-1, -2, and -3 expression in human keratinocytes.

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Park, Ji-Hae; Mohamed, Mohamed Antar Aziz; Jung, Ye-Jin; Shrestha, Sabina; Lee, Tae Hoon; Lee, Chang-Ho; Han, Daeseok; Kim, Jiyoung; Baek, Nam-In

2015-10-01

Four sesquiterpenes were isolated from the rhizome of Curcuma xanthorrhiza Roxb.: furanodiene (1), germacrone (2), furanodienone (3), and 13-hydroxygermacrone (4). Importantly, this was the first time compounds 1 and 4 were isolated from this plant. The chemical structures of these compounds were determined using 1D- and 2D-nuclear magnetic resonance, infrared spectroscopy, and electron ionization mass spectrometry analyses. Among the isolated compounds, compounds 2 and 4 inhibited UVB-induced upregulation of the mRNA and protein expression levels of MMP-1, MMP-2, and MMP-3 in human keratinocytes (HaCaT). Moreover, this upregulation occurred in a dose-dependent manner over the range of 1-10 μM for each compound.

Sella turcica measurements on lateral cephalograms of patients with neurofibromatosis type 1

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Friedrich, Reinhard E.

2017-03-01

Full Text Available The aim of this study was to measure line segments and areas of sella turcica on lateral cephalograms with respect to the clinical diagnosis of facial phenotype of patients with neurofibromatosis type 1 (NF1. Special attention was given to correlate the measured values with certain tumour types that are typical for this disease.Material and methods: Lateral cephalograms of 194 individuals were investigated. Patients with NF1 were further divided according to the detection and topography of facial plexiform neurofibromas (PNF taking into account the distribution pattern of the trigeminal nerve. All patients with PNF showed unilateral tumour localisation. Patients without any facial PNF constituted a separate group. Healthy volunteers with ideal occlusion and no history of any intervention in the maxillofacial region served as a control group. The following items were determined on the radiographs: sella entrance, sella width, sella depths, sella diagonal, and sella area.Results: Patients with PNF of the first and second trigeminal nerve branch or affected in all branches showed highly statistically significant enlarged sella tucica measurement values. On the other hand, patients with PNF restricted to one branch only or simultaneously in the second and third branches showed measurement values that were not different to those obtained in NF1 patients devoid of facial PNF. The latter group also showed no difference of sella turcica parameters obtained in the control group.Conclusion: This study provides evidence for the association of a certain NF1 phenotype with distinct skeletal alterations of the skull base, shown here using the example of the representation of the sella turcica in the lateral radiograph. These findings are also relevant in the discussion of NF1 as a disease of bones and in the assessment of brain development in NF1. Both items are discussed in relationship to a facial plexiform neurofibroma. Furthermore, the knowledge of this

A novel role of BELL1-like homeobox genes, PENNYWISE and POUND-FOOLISH, in floral patterning.

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Yu, Lifeng; Patibanda, Varun; Smith, Harley M S

2009-02-01

Flowers are determinate shoots comprised of perianth and reproductive organs displayed in a whorled phyllotactic pattern. Floral organ identity genes display region-specific expression patterns in the developing flower. In Arabidopsis, floral organ identity genes are activated by LEAFY (LFY), which functions with region-specific co-regulators, UNUSUAL FLORAL ORGANS (UFO) and WUSCHEL (WUS), to up-regulate homeotic genes in specific whorls of the flower. PENNYWISE (PNY) and POUND-FOOLISH (PNF) are redundant functioning BELL1-like homeodomain proteins that are expressed in shoot and floral meristems. During flower development, PNY functions with a co-repressor complex to down-regulate the homeotic gene, AGAMOUS (AG), in the outer whorls of the flower. However, the function of PNY as well as PNF in regulating floral organ identity in the central whorls of the flower is not known. In this report, we show that combining mutations in PNY and PNF enhance the floral patterning phenotypes of weak and strong alleles of lfy, indicating that these BELL1-like homeodomain proteins play a role in the specification of petals, stamens and carpels during flower development. Expression studies show that PNY and PNF positively regulate the homeotic genes, APETALA3 and AG, in the inner whorls of the flower. Moreover, PNY and PNF function in parallel with LFY, UFO and WUS to regulate homeotic gene expression. Since PNY and PNF interact with the KNOTTED1-like homeodomain proteins, SHOOTMERISTEMLESS (STM) and KNOTTED-LIKE from ARABIDOPSIS THALIANA2 (KNAT2) that regulate floral development, we propose that PNY/PNF-STM and PNY/PNF-KNAT2 complexes function in the inner whorls to regulate flower patterning events.

Regulation of MMP2 and MMP9 metalloproteinases by FSH and growth factors in bovine granulosa cells

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Valerio M. Portela

2009-01-01

Full Text Available Matrix metalloproteinases (MMP are key enzymes involved in tissue remodeling. Within the ovary, they are believed to play a major role in ovulation, and have been linked to follicle atresia. To gain insight into the regulation of MMPs, we measured the effect of hormones and growth factors on MMP2 and MMP9 mRNA levels in non-luteinizing granulosa cells in serum-free culture. FSH and IGF1 both stimulated estradiol secretion and inhibited MMP2 and MMP9 mRNA abundance. In contrast, EGF and FGF2 both inhibited estradiol secretion but had no effect on MMP expression. At physiological doses, none of these hormones altered the proportion of dead cells. Although we cannot link MMP expression with apoptosis, the specific down regulation by the gonadotropic hormones FSH and IGF1 in vitro suggests that excess MMP2 and MMP9 expression is neither required nor desired for follicle development.

The mycorrhiza-dependent defensin MtDefMd1 of Medicago truncatula acts during the late restructuring stages of arbuscule-containing cells.

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Marian Uhe

Full Text Available Different symbiotic and pathogenic plant-microbe interactions involve the production of cysteine-rich antimicrobial defensins. In Medicago truncatula, the expression of four MtDefMd genes, encoding arbuscular mycorrhiza-dependent defensins containing an N-terminal signal peptide and exhibiting some differences to non-symbiotic defensins, raised over the time of fungal colonization. Whereas the MtDefMd1 and MtDefMd2 promoters were inactive in cells containing young arbuscules, cells with fully developed arbuscules displayed different levels of promoter activities, indicating an up-regulation towards later stages of arbuscule formation. MtDefMd1 and MtDefMd2 expression was absent or strongly down-regulated in mycorrhized ram1-1 and pt4-2 mutants, known for defects in arbuscule branching or premature arbuscule degeneration, respectively. A ~97% knock-down of MtDefMd1/MtDefMd2 expression did not significantly affect arbuscule size. Although overexpression of MtDefMd1 in arbuscule-containing cells led to an up-regulation of MtRam1, encoding a key transcriptional regulator of arbuscule formation, no morphological changes were evident. Co-localization of an MtDefMd1-mGFP6 fusion with additional, subcellular markers revealed that this defensin is associated with arbuscules in later stages of their life-cycle. MtDefMd1-mGFP6 was detected in cells with older arbuscules about to collapse, and ultimately in vacuolar compartments. Comparisons with mycorrhized roots expressing a tonoplast marker indicated that MtDefMd1 acts during late restructuring processes of arbuscule-containing cells, upon their transition into a post-symbiotic state.

Role of MMP-3 and MMP-9 and their haplotypes in risk of bladder cancer in North Indian cohort.

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Srivastava, Priyanka; Mandhani, Anil; Kapoor, Rakesh; Mittal, Rama D

2010-11-01

Matrix metalloproteinases (MMPs) play critical roles in cancer development and progression. Nonsynonymous single nucleotide polymorphisms (SNPs) in functional domain of MMP-3 and MMP-9 contribute appreciably to cancer predisposition and aggression. To test this proposition we examined whether six SNPs of the MMP-3 and MMP-9 genes are associated with risk of bladder cancer (BC) in a North Indian population. Six SNPs of MMP-3 and MMP-9 were analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in a case-control study including 200 BC patients and 200 age/gender/ethnicity-matched controls. Increased risk for BC susceptibility was observed in MMP-3 (1171) 5A/5A [P = 0.022; odds ratio (OR), 3.46; 95% confidence interval (CI), 1.20-9.98], MMP-9 (Q279R) QQ (P = 0.048; OR, 1.92; 95%CI, 1.01-3.66), MMP-9 (P574R) PR (P BCG)-treated non-muscle-invasive BC (NMIBC) patients (log-rank P = 0.025). Our data suggested that MMP-3-1171 5A/5A and MMP-9 (Q279R) QQ, MMP-9 (P574R) PR, PR + RR, and R allele are associated with high risk of BC.

MR-1S Interacts with PET100 and PET117 in Module-Based Assembly of Human Cytochrome c Oxidase

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Sara Vidoni

2017-02-01

Full Text Available The biogenesis of human cytochrome c oxidase (COX is an intricate process in which three mitochondrial DNA (mtDNA-encoded core subunits are assembled in a coordinated way with at least 11 nucleus-encoded subunits. Many chaperones shared between yeast and humans are involved in COX assembly. Here, we have used a MT-CO3 mutant cybrid cell line to define the composition of assembly intermediates and identify new human COX assembly factors. Quantitative mass spectrometry analysis led us to modify the assembly model from a sequential pathway to a module-based process. Each module contains one of the three core subunits, together with different ancillary components, including HIGD1A. By the same analysis, we identified the short isoform of the myofibrillogenesis regulator 1 (MR-1S as a new COX assembly factor, which works with the highly conserved PET100 and PET117 chaperones to assist COX biogenesis in higher eukaryotes.

MR-1S Interacts with PET100 and PET117 in Module-Based Assembly of Human Cytochrome c Oxidase.

Science.gov (United States)

Vidoni, Sara; Harbour, Michael E; Guerrero-Castillo, Sergio; Signes, Alba; Ding, Shujing; Fearnley, Ian M; Taylor, Robert W; Tiranti, Valeria; Arnold, Susanne; Fernandez-Vizarra, Erika; Zeviani, Massimo

2017-02-14

The biogenesis of human cytochrome c oxidase (COX) is an intricate process in which three mitochondrial DNA (mtDNA)-encoded core subunits are assembled in a coordinated way with at least 11 nucleus-encoded subunits. Many chaperones shared between yeast and humans are involved in COX assembly. Here, we have used a MT-CO3 mutant cybrid cell line to define the composition of assembly intermediates and identify new human COX assembly factors. Quantitative mass spectrometry analysis led us to modify the assembly model from a sequential pathway to a module-based process. Each module contains one of the three core subunits, together with different ancillary components, including HIGD1A. By the same analysis, we identified the short isoform of the myofibrillogenesis regulator 1 (MR-1S) as a new COX assembly factor, which works with the highly conserved PET100 and PET117 chaperones to assist COX biogenesis in higher eukaryotes. Copyright © 2017 Elsevier Inc. All rights reserved.

Matriptase/MT-SP1 is required for postnatal survival, epidermal barrier function, hair follicle development, and thymic homeostasis

DEFF Research Database (Denmark)

List, Karin; Haudenschild, Christian C; Szabo, Roman

2002-01-01

of Matriptase/MT-SP1 also seriously affected hair follicle development resulting in generalized follicular hypoplasia, absence of erupted vibrissae, lack of vibrissal hair canal formation, ingrown vibrissae, and wholesale abortion of vibrissal follicles. Furthermore, Matriptase/MT-SP1-deficiency resulted...... in dramatically increased thymocyte apoptosis, and depletion of thymocytes. This study demonstrates that Matriptase/MT-SP1 has pleiotropic functions in the development of the epidermis, hair follicles, and cellular immune system....

TGF-beta1 modulates matrix metalloproteinase-13 expression in hepatic stellate cells by complex mechanisms involving p38MAPK, PI3-kinase, AKT, and p70S6k.

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Lechuga, Carmen G; Hernández-Nazara, Zamira H; Domínguez Rosales, José-Alfredo; Morris, Elena R; Rincón, Ana Rosa; Rivas-Estilla, Ana María; Esteban-Gamboa, Andrés; Rojkind, Marcos

2004-11-01

Transforming growth factor-beta1 (TGF-beta1), the main cytokine involved in liver fibrogenesis, induces expression of the type I collagen genes in hepatic stellate cells by a transcriptional mechanism, which is hydrogen peroxide and de novo protein synthesis dependent. Our recent studies have revealed that expression of type I collagen and matrix metalloproteinase-13 (MMP-13) mRNAs in hepatic stellate cells is reciprocally modulated. Because TGF-beta1 induces a transient elevation of alpha1(I) collagen mRNA, we investigated whether this cytokine was able to induce the expression of MMP-13 mRNA during the downfall of the alpha1(I) collagen mRNA. In the present study, we report that TGF-beta1 induces a rapid decline in steady-state levels of MMP-13 mRNA at the time that it induces the expression of alpha1(I) collagen mRNA. This change in MMP-13 mRNA expression occurs within the first 6 h postcytokine administration and is accompanied by a twofold increase in gene transcription and a fivefold decrease in mRNA half-life. This is followed by increased expression of MMP-13 mRNA, which reaches maximal values by 48 h. Our results also show that this TGF-beta1-mediated effect is de novo protein synthesis-dependent and requires the activity of p38MAPK, phosphatidylinositol 3-kinase, AKT, and p70(S6k). Altogether, our data suggest that regulation of MMP-13 by TGF-beta1 is a complex process involving transcriptional and posttranscriptional mechanisms.

Recombinant human melatonin receptor MT1 isolated in mixed detergents shows pharmacology similar to that in mammalian cell membranes.

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Christel Logez

Full Text Available The human melatonin MT1 receptor-belonging to the large family of G protein-coupled receptors (GPCRs-plays a key role in circadian rhythm regulation and is notably involved in sleep disorders and depression. Structural and functional information at the molecular level are highly desired for fine characterization of this receptor; however, adequate techniques for isolating soluble MT1 material suitable for biochemical and biophysical studies remain lacking. Here we describe the evaluation of a panel of constructs and host systems for the production of recombinant human MT1 receptors, and the screening of different conditions for their solubilization and purification. Our findings resulted in the establishment of an original strategy using a mixture of Fos14 and CHAPS detergents to extract and purify a recombinant human MT1 from Pichia pastoris membranes. This procedure enabled the recovery of relatively pure, monomeric and ligand-binding active MT1 receptor in the near-milligram range. A comparative study based on extensive ligand-binding characterization highlighted a very close correlation between the pharmacological profiles of MT1 purified from yeast and the same receptor present in mammalian cell membranes. The high quality of the purified MT1 was further confirmed by its ability to activate its cognate Gαi protein partner when reconstituted in lipid discs, thus opening novel paths to investigate this receptor by biochemical and biophysical approaches.

Recombinant human melatonin receptor MT1 isolated in mixed detergents shows pharmacology similar to that in mammalian cell membranes.

Science.gov (United States)

Logez, Christel; Berger, Sylvie; Legros, Céline; Banères, Jean-Louis; Cohen, William; Delagrange, Philippe; Nosjean, Olivier; Boutin, Jean A; Ferry, Gilles; Simonin, Frédéric; Wagner, Renaud

2014-01-01

The human melatonin MT1 receptor-belonging to the large family of G protein-coupled receptors (GPCRs)-plays a key role in circadian rhythm regulation and is notably involved in sleep disorders and depression. Structural and functional information at the molecular level are highly desired for fine characterization of this receptor; however, adequate techniques for isolating soluble MT1 material suitable for biochemical and biophysical studies remain lacking. Here we describe the evaluation of a panel of constructs and host systems for the production of recombinant human MT1 receptors, and the screening of different conditions for their solubilization and purification. Our findings resulted in the establishment of an original strategy using a mixture of Fos14 and CHAPS detergents to extract and purify a recombinant human MT1 from Pichia pastoris membranes. This procedure enabled the recovery of relatively pure, monomeric and ligand-binding active MT1 receptor in the near-milligram range. A comparative study based on extensive ligand-binding characterization highlighted a very close correlation between the pharmacological profiles of MT1 purified from yeast and the same receptor present in mammalian cell membranes. The high quality of the purified MT1 was further confirmed by its ability to activate its cognate Gαi protein partner when reconstituted in lipid discs, thus opening novel paths to investigate this receptor by biochemical and biophysical approaches.

Requirement of matrix metalloproteinase-1 for intestinal homeostasis in the adult Drosophila midgut

International Nuclear Information System (INIS)

Lee, Shin-Hae; Park, Joung-Sun; Kim, Young-Shin; Chung, Hae-Young; Yoo, Mi-Ae

2012-01-01

Stem cells are tightly regulated by both intrinsic and extrinsic signals as well as the extracellular matrix (ECM) for tissue homeostasis and regenerative capacity. Matrix metalloproteinases (MMPs), proteolytic enzymes, modulate the turnover of numerous substrates, including cytokine precursors, growth factors, and ECM molecules. However, the roles of MMPs in the regulation of adult stem cells are poorly understood. In the present study, we utilize the Drosophila midgut, which is an excellent model system for studying stem cell biology, to show that Mmp1 is involved in the regulation of intestinal stem cells (ISCs). The results showed that Mmp1 is expressed in the adult midgut and that its expression increases with age and with exposure to oxidative stress. Mmp1 knockdown or Timp-overexpressing flies and flies heterozygous for a viable, hypomorphic Mmp1 allele increased ISC proliferation in the gut, as shown by staining with an anti-phospho-histone H3 antibody and BrdU incorporation assays. Reduced Mmp1 levels induced intestinal hyperplasia, and the Mmp1depletion-induced ISC proliferation was rescued by the suppression of the EGFR signaling pathway, suggesting that Mmp1 regulates ISC proliferation through the EGFR signaling pathway. Furthermore, adult gut-specific knockdown and whole-animal heterozygotes of Mmp1 increased additively sensitivity to paraquat-induced oxidative stress and shortened lifespan. Our data suggest that Drosophila Mmp1 is involved in the regulation of ISC proliferation for maintenance of gut homeostasis. -- Highlights: ► Mmp1 is expressed in the adult midgut. ► Mmp1 is involved in the regulation of ISC proliferation activity. ► Mmp1-related ISC proliferation is associated with EGFR signaling. ► Mmp1 in the gut is required for the intestinal homeostasis and longevity.

Requirement of matrix metalloproteinase-1 for intestinal homeostasis in the adult Drosophila midgut

Energy Technology Data Exchange (ETDEWEB)

Lee, Shin-Hae; Park, Joung-Sun [Department of Molecular Biology, College of Natural Science, Pusan National University, Busan 609-735 (Korea, Republic of); Kim, Young-Shin [Research Institute of Genetic Engineering, Pusan National University, Busan 609-735 (Korea, Republic of); Chung, Hae-Young [Molecular Inflammation Research Center for Aging Intervention (MRCA), College of Pharmacy, Pusan National University, Busan 609-735 (Korea, Republic of); Yoo, Mi-Ae, E-mail: mayoo@pusan.ac.kr [Department of Molecular Biology, College of Natural Science, Pusan National University, Busan 609-735 (Korea, Republic of)

2012-03-10

Stem cells are tightly regulated by both intrinsic and extrinsic signals as well as the extracellular matrix (ECM) for tissue homeostasis and regenerative capacity. Matrix metalloproteinases (MMPs), proteolytic enzymes, modulate the turnover of numerous substrates, including cytokine precursors, growth factors, and ECM molecules. However, the roles of MMPs in the regulation of adult stem cells are poorly understood. In the present study, we utilize the Drosophila midgut, which is an excellent model system for studying stem cell biology, to show that Mmp1 is involved in the regulation of intestinal stem cells (ISCs). The results showed that Mmp1 is expressed in the adult midgut and that its expression increases with age and with exposure to oxidative stress. Mmp1 knockdown or Timp-overexpressing flies and flies heterozygous for a viable, hypomorphic Mmp1 allele increased ISC proliferation in the gut, as shown by staining with an anti-phospho-histone H3 antibody and BrdU incorporation assays. Reduced Mmp1 levels induced intestinal hyperplasia, and the Mmp1depletion-induced ISC proliferation was rescued by the suppression of the EGFR signaling pathway, suggesting that Mmp1 regulates ISC proliferation through the EGFR signaling pathway. Furthermore, adult gut-specific knockdown and whole-animal heterozygotes of Mmp1 increased additively sensitivity to paraquat-induced oxidative stress and shortened lifespan. Our data suggest that Drosophila Mmp1 is involved in the regulation of ISC proliferation for maintenance of gut homeostasis. -- Highlights: Black-Right-Pointing-Pointer Mmp1 is expressed in the adult midgut. Black-Right-Pointing-Pointer Mmp1 is involved in the regulation of ISC proliferation activity. Black-Right-Pointing-Pointer Mmp1-related ISC proliferation is associated with EGFR signaling. Black-Right-Pointing-Pointer Mmp1 in the gut is required for the intestinal homeostasis and longevity.

Expression of MMP-2 and TIMP-1 in cerebrospinal fluid and the correlation with dynamic changes of serum PCT in neonatal purulent meningitis

Science.gov (United States)

Chen, Huilan; Wu, Fei; Fu, Rong; Feng, Xiangchun

2018-01-01

Matrix metalloproteinase 2 (MMP-2) and tissue inhibitor of metalloproteinase-1 (TIMP-1) levels in cerebrospinal fluid of pediatric patients with neonatal purulent meningitis were observed to analyze changes in serum procalcitonin (PCT) and the correlation among the three factors (MMP-2, TIMP-1 and PCT). Sixty pediatric patients with neonatal purulent meningitis from April 2015 to December 2016 were enrolled as the purulent meningitis group and 60 pediatric patients with viral encephalitis treated during the same period were enrolled as the viral encephalitis group. Additionally, 60 healthy newborns who underwent physical examinations in our hospital during the same period were enrolled as the control group. The levels of MMP-2 were 136.73±25.42 ng/ml in the purulent meningitis group, 45.32±6.57 ng/ml in the viral encephalitis group and 1.32±0.51 ng/ml in the control group and the differences between the three groups were statistically significant (F=15.052, pfluid were 374.55±36.04 ng/ml in the purulent meningitis group, 176.61±21.06 ng/ml in the viral encephalitis group and 7.72±2.44 ng/ml in the control group. The serum levels of PCT were 14.56±2.21 ng/ml in the purulent meningitis group, 9.04±1.17 ng/ml in the viral encephalitis group and 0.38±0.14 ng/ml in the control group. The level of MMP-2 in cerebrospinal fluid of pediatric patients in the purulent meningitis group was positively correlated with the level of serum PCT (r=0.582, pfluid of pediatric patients in the viral encephalitis group was positively correlated with the level of serum PCT (r=0.635, p<0.05). In conclusion, MMP-2 and TIMP-1 were positively correlated with the levels of serum PCT, suggesting that MMP-2, TIMP-1 and PCT were involved in the occurrence and development of neonatal purulent meningitis. PMID:29399119

Surgery-induced hippocampal angiotensin II elevation causes blood-brain barrier disruption via MMP/TIMP in aged rats

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Zhengqian eLi

2016-04-01

Full Text Available Reversible BBB disruption has been uniformly reported in several animal models of postoperative cognitive dysfunction (POCD. Nevertheless, the precise mechanism underlying this occurrence remains unclear. Using an aged rat model of POCD, we investigated the dynamic changes in expression of molecules involved in BBB disintegration, matrix metalloproteinase-2 (MMP-2 and -9 (MMP-9, as well as three of their endogenous tissue inhibitors (TIMP-1, -2, -3, and tried to establish the correlation between MMP/TIMP balance and surgery-induced hippocampal BBB disruption. We validated the increased hippocampal expression of angiotensin II (Ang II and Ang II receptor type 1 (AT1 after surgery. We also found MMP/TIMP imbalance as early as 6 h after surgery, together with increased BBB permeability and decreased expression of Occludin and zonula occludens-1 (ZO-1, as well as increased basal lamina protein laminin at 24 h postsurgery. The AT1 antagonist candesartan restored MMP/TIMP equilibrium and modulated expression of Occludin and laminin, but not ZO-1, thereby improving BBB permeability. These events were accompanied by suppression of the surgery-induced canonical nuclear factor-κB (NF-κB activation cascade. Nevertheless, AT1 antagonism did not affect nuclear receptor peroxisome proliferator-activated receptor-γ expression. Collectively, these findings suggest that surgery-induced Ang II release impairs BBB integrity by activating NF-κB signaling and disrupting downstream MMP/TIMP balance via AT1 receptor.

Diffusion of MMPs on the Surface of Collagen Fibrils: The Mobile Cell Surface – Collagen Substratum Interface

Science.gov (United States)

Collier, Ivan E.; Legant, Wesley; Marmer, Barry; Lubman, Olga; Saffarian, Saveez; Wakatsuki, Tetsuro; Elson, Elliot; Goldberg, Gregory I.

2011-01-01

Remodeling of the extracellular matrix catalyzed by MMPs is central to morphogenetic phenomena during development and wound healing as well as in numerous pathologic conditions such as fibrosis and cancer. We have previously demonstrated that secreted MMP-2 is tethered to the cell surface and activated by MT1-MMP/TIMP-2-dependent mechanism. The resulting cell-surface collagenolytic complex (MT1-MMP)2/TIMP-2/MMP-2 can initiate (MT1-MMP) and complete (MMP-2) degradation of an underlying collagen fibril. The following question remained: What is the mechanism of substrate recognition involving the two structures of relatively restricted mobility, the cell surface enzymatic complex and a collagen fibril embedded in the ECM? Here we demonstrate that all the components of the complex are capable of processive movement on a surface of the collagen fibril. The mechanism of MT1-MMP movement is a biased diffusion with the bias component dependent on the proteolysis of its substrate, not adenosine triphosphate (ATP) hydrolysis. It is similar to that of the MMP-1 Brownian ratchet we described earlier. In addition, both MMP-2 and MMP-9 as well as their respective complexes with TIMP-1 and -2 are capable of Brownian diffusion on the surface of native collagen fibrils without noticeable dissociation while the dimerization of MMP-9 renders the enzyme immobile. Most instructive is the finding that the inactivation of the enzymatic activity of MT1-MMP has a detectable negative effect on the cell force developed in miniaturized 3D tissue constructs. We propose that the collagenolytic complex (MT1-MMP)2/TIMP-2/MMP-2 represents a Mobile Cell Surface – Collagen Substratum Interface. The biological implications of MT1-MMP acting as a molecular ratchet tethered to the cell surface in complex with MMP-2 suggest a new mechanism for the role of spatially regulated peri-cellular proteolysis in cell-matrix interactions. PMID:21912660

Diffusion of MMPs on the surface of collagen fibrils: the mobile cell surface-collagen substratum interface.

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Ivan E Collier

Full Text Available Remodeling of the extracellular matrix catalyzed by MMPs is central to morphogenetic phenomena during development and wound healing as well as in numerous pathologic conditions such as fibrosis and cancer. We have previously demonstrated that secreted MMP-2 is tethered to the cell surface and activated by MT1-MMP/TIMP-2-dependent mechanism. The resulting cell-surface collagenolytic complex (MT1-MMP(2/TIMP-2/MMP-2 can initiate (MT1-MMP and complete (MMP-2 degradation of an underlying collagen fibril. The following question remained: What is the mechanism of substrate recognition involving the two structures of relatively restricted mobility, the cell surface enzymatic complex and a collagen fibril embedded in the ECM? Here we demonstrate that all the components of the complex are capable of processive movement on a surface of the collagen fibril. The mechanism of MT1-MMP movement is a biased diffusion with the bias component dependent on the proteolysis of its substrate, not adenosine triphosphate (ATP hydrolysis. It is similar to that of the MMP-1 Brownian ratchet we described earlier. In addition, both MMP-2 and MMP-9 as well as their respective complexes with TIMP-1 and -2 are capable of Brownian diffusion on the surface of native collagen fibrils without noticeable dissociation while the dimerization of MMP-9 renders the enzyme immobile. Most instructive is the finding that the inactivation of the enzymatic activity of MT1-MMP has a detectable negative effect on the cell force developed in miniaturized 3D tissue constructs. We propose that the collagenolytic complex (MT1-MMP(2/TIMP-2/MMP-2 represents a Mobile Cell Surface-Collagen Substratum Interface. The biological implications of MT1-MMP acting as a molecular ratchet tethered to the cell surface in complex with MMP-2 suggest a new mechanism for the role of spatially regulated peri-cellular proteolysis in cell-matrix interactions.

Suppressed invasive and migratory behaviors of SW1353 chondrosarcoma cells through the regulation of Src, Rac1 GTPase, and MMP13.

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Xu, Wenxiao; Wan, Qiaoqiao; Na, Sungsoo; Yokota, Hiroki; Yan, Jing-Long; Hamamura, Kazunori

2015-12-01

Chondrosarcoma is the second frequent type of primary bone cancer. In response to stress to the endoplasmic reticulum, activation of eIF2α-mediated signaling is reported to induce apoptosis. However, its effects on invasive and migratory behaviors of chondrosarcoma have not been understood. Focusing on potential roles of Src kinase, Rac1 GTPase, and MMP13, we investigated eIF2α-driven regulation of SW1353 chondrosarcoma cells. In particular, we employed two chemical agents (salubrinal, Sal; and guanabenz, Gu) that elevate the level of eIF2α phosphorylation. The result revealed that both Sal and Gu reduced invasion and motility of SW1353 chondrosarcoma cells in a dose dependent manner. Live imaging using a fluorescent resonance energy transfer (FRET) technique showed that Sal and Gu downregulated activities of Src kinase as well as Rac1 GTPase in an eIF2α dependent manner. RNA interference experiments supported an eIF2α-mediated regulatory network in the inhibitory role of Sal and Gu. Partial silencing of MMP13 also suppressed malignant phenotypes of SW1353 chondrosarcoma cells. However, MMP13 was not regulated via eIF2α since administration of Sal but not Gu reduced expression of MMP13. In summary, we demonstrate that eIF2α dependent and independent pathways regulate invasion and motility of SW1353 chondrosarcoma cells, and inactivation of Src, Rac1, and MMP13 by Sal could provide a potential adjuvant therapy for combating metastatic chondrosarcoma cells. Copyright © 2015 Elsevier Inc. All rights reserved.

Heterogeneity of serum gelatinases MMP-2 and MMP-9 isoforms and charge variants

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Rossano, Rocco; Larocca, Marilena; Riviello, Lea; Coniglio, Maria Gabriella; Vandooren, Jennifer; Liuzzi, Grazia Maria; Opdenakker, Ghislain; Riccio, Paolo

2014-01-01

The matrix metalloproteinases (MMPs) gelatinase A (MMP-2) and gelatinase B (MMP-9) are mediators of brain injury in multiple sclerosis (MS) and valuable biomarkers of disease activity. We applied bidimensional zymography (2-DZ) as an extension of classic monodimensional zymography (1-DZ) to analyse the complete pattern of isoforms and post-translational modifications of both MMP-9 and MMP-2 present in the sera of MS patients. The enzymes were separated on the basis of their isoelectric points (pI) and apparent molecular weights (Mw) and identified both by comparison with standard enzyme preparations and by Western blot analysis. Two MMP-2 isoforms, and at least three different isoforms and two different states of organization of MMP-9 (the multimeric MMP-9 and the N-GAL-MMP-9 complex) were observed. In addition, 2-DZ revealed for the first time that all MMP-9 and MMP-2 isoforms actually exist in the form of charge variants: four or five variants in the N-GAL complex, more charge variants in the case of MMP-9; and five to seven charge variants for MMP-2. Charge variants were also observed in recombinant enzymes and, after concentration, also in sera from healthy individuals. Sialylation (MMP-9) and phosphorylation (MMP-2) contributed to molecular heterogeneity. The detection of charge variants of MMP-9 and MMP-2 in MS serum samples illustrates the power of 2-DZ and demonstrates that in previous studies MMP mixtures, rather than single molecules, were analysed. These observations open perspectives for better diagnosis and prognosis of many diseases and need to be critically interpreted when applying other methods for MS and other diseases. PMID:24616914

TIMP-1 resistant matrix metalloproteinase-9 is the predominant serum active isoform associated with MRI activity in patients with multiple sclerosis.

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Trentini, Alessandro; Manfrinato, Maria C; Castellazzi, Massimiliano; Tamborino, Carmine; Roversi, Gloria; Volta, Carlo A; Baldi, Eleonora; Tola, Maria R; Granieri, Enrico; Dallocchio, Franco; Bellini, Tiziana; Fainardi, Enrico

2015-08-01

The activity of matrix metalloproteinase-9 (MMP-9) depends on two isoforms, an 82 kDa active MMP-9 modulated by its specific tissue inhibitor (TIMP-1), and a 65 kDa TIMP-1 resistant active MMP-9. The relevance of these two enzymatic isoforms in multiple sclerosis (MS) is still unknown. To investigate the contribution of the TIMP-1 modulated and resistant active MMP-9 isoforms to MS pathogenesis. We measured the serum levels of the 82 kDa and TIMP-1 resistant active MMP-9 isoforms by activity assay systems in 86 relapsing-remitting MS (RRMS) patients, categorized according to clinical and magnetic resonance imaging (MRI) evidence of disease activity, and in 70 inflammatory (OIND) and 69 non-inflammatory (NIND) controls. Serum levels of TIMP-1 resistant MMP-9 were more elevated in MS patients than in OIND and NIND (p < 0.05, p < 0.02, respectively). Conversely, 82 kDa active MMP-9 was higher in NIND than in the OIND and MS patients (p < 0.01 and p < 0.00001, respectively). MRI-active patients had higher levels of TIMP-1 resistant MMP-9 and 82 kDa active MMP-9, than did those with MRI inactive MS (p < 0.01 and p < 0.05, respectively). Our findings suggested that the TIMP-1 resistant MMP-9 seem to be the predominantly active isoform contributing to MS disease activity. © The Author(s), 2015.

Nonoperative Management May Be a Viable Approach to Plexiform Neurofibroma of the Porta Hepatis in Patients with Neurofibromatosis-1

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Natesh Yepuri

2018-01-01

Full Text Available Background. Plexiform neurofibroma (PNF in the porta hepatis (PH is an unusual manifestation of neurofibromatosis-1 (NF-1. Resection is often recommended given the risk of malignant transformation. We encountered a challenging case in clinical practice which prompted us to report our findings and perform a systematic review on the management of these tumors. Methods. We reported the case of a 31-year-old woman with NF-1 and PNF of the PH. PRISMA 2009 guidelines were followed for systematic review. Results. Our patient was found to have unresectable disease at exploration. After >5 years of follow-up, she continued to have stable disease on imaging. We identified 12 studies/case reports including 10 adult and 6 pediatric patients with PNF of PH. None of the 7 adult patients with NF-1 and PNF of PH underwent a successful tumor resection. All pediatric patients were managed with surveillance alone. All but one pediatric patient had NF-1. None of the reported cases of PNF of PH had malignant transformation. Conclusion. Our findings suggest that PNFs of PH in the setting of NF-1 are often unresectable and may have an indolent course. Surveillance alone may be a reasonable option in some patients; however, further studies are needed.

Resection of small plexiform neurofibromas in neurofibromatosis type 1 children

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Fünsterer Carsten

2005-01-01

Full Text Available Abstract Background Plexiform neurofibromas (PNF are benign tumors of the peripheral nerve which mostly develop in patients with neurofibromatosis type 1 (NF1. Surgical interventions are usually not applied to children with small tumors. These are rather restricted to debulking of larger tumors in adults that cause clinical complications or aesthetic disfigurement. In most cases, a total resection of PNF is not possible due to the network-like growth of the tumors. Patients and methods Early surgical intervention was carried out for 9 small PNFs in 7 NF1 children. Tumor resection was performed following the graphical delineation of the affected skin and according the MRI findings. Results Total resection was achieved for all 9 PNF without causing any neurological or organic deficit. Annual magnetic resonance tomography over a period of four years did not reveal any relapse of the tumors. Conclusions Early surgical intervention for small superficial PNFs in NF1 children have various advantages and may especially be considered a strategy to prevent progression.

Use of matrix metalloproteinase-9 (MMP-9 and its tissue inhibitor (TIMP-1 in the pathomorphological diagnosis of carotid pathology: literature review and own observations

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Yu. I. Kuzyk

2016-04-01

Full Text Available Matrix metalloproteinases (MMPs are the degradative enzymes of the extracellular matrix. Currently, the role of MMP-2 and MMP-9 in the progression of atherosclerosis (AS is proved. The question of possible involvement of MMP-9 into elastin degradation in fibromuscular dysplasia (FMD and pathological tortuosity (PT remains open and insufficiently explored. The aim of the study – analysis of the current literature on the role of degradative enzymes in the development of carotid pathology and study of the expression of type I, III, IV collagens, MMP-9 and TIPM-1 in the wall of the carotid arteries in FMD, PT and AS. Materials and methods included literature review and own research. Immunohistochemical study of type I, III and IV collagens, TIMP-1 and MMP-9 was carried out on surgical material of patients with main carotid diseases: three observations with AS, two – with FMD, two – with PT. The level of expression was assessed by semiquantitative method. Results. Own observations showed that in FMD types I and III collagen content in the media and in the adventitia remains unchanged. MMP-9 expression level reached the highest level of intensity in atherosclerotic plaques, particularly in macrophages, constituting the main part of the atheromatous mass. Moderate intensity of expression is noted in FMD and PT. In PT expression prevailed in the lower third of the media on the border with adventitia, including the adventitia, in FMD – mainly in the media. The level of TIMP-1 is weakly positive in PT and FMD, negative in AS. Conclusions. These results demonstrate the possibility of using MMP-9 and TIMP-1 as a morphological marker determining pathological processes in carotid pathology. Data of immunohistochemical study of type I, II, IV collagens indicate moderate expression of collagen type I in FMD and PT, severe expression of collagen III in FMD, moderate in PT. Type IV collagen is highly expressed in atherosclerotic plaques. For AS high

MT71x: Multi-Temperature Library Based on ENDF/B-VII.1

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Conlin, Jeremy Lloyd [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Parsons, Donald Kent [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Gray, Mark Girard [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Lee, Mary Beth [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); White, Morgan Curtis [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

2015-12-16

The Nuclear Data Team has released a multitemperature transport library, MT71x, based upon ENDF/B-VII.1 with a few modifications as well as additional evaluations for a total of 427 isotope tables. The library was processed using NJOY2012.39 into 23 temperatures. MT71x consists of two sub-libraries; MT71xMG for multigroup energy representation data and MT71xCE for continuous energy representation data. These sub-libraries are suitable for deterministic transport and Monte Carlo transport applications, respectively. The SZAs used are the same for the two sub-libraries; that is, the same SZA can be used for both libraries. This makes comparisons between the two libraries and between deterministic and Monte Carlo codes straightforward. Both the multigroup energy and continuous energy libraries were verified and validated with our checking codes checkmg and checkace (multigroup and continuous energy, respectively) Then an expanded suite of tests was used for additional verification and, finally, verified using an extensive suite of critical benchmark models. We feel that this library is suitable for all calculations and is particularly useful for calculations sensitive to temperature effects.

Matrix metalloproteinase-9 (MMP9) is involved in the TNF-α-induced fusion of human M13SV1-Cre breast epithelial cells and human MDA-MB-435-pFDR1 cancer cells.

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Weiler, Julian; Mohr, Marieke; Zänker, Kurt S; Dittmar, Thomas

2018-04-10

In addition to physiological events such as fertilisation, placentation, osteoclastogenesis, or tissue regeneration/wound healing, cell fusion is involved in pathophysiological conditions such as cancer. Cell fusion, which applies to both the proteins and conditions that induce the merging of two or more cells, is not a fully understood process. Inflammation/pro-inflammatory cytokines might be a positive trigger for cell fusion. Using a Cre-LoxP-based cell fusion assay we demonstrated that the fusion between human M13SV1-Cre breast epithelial cells and human MDA-MB-435-pFDR1 cancer cells was induced by the pro-inflammatory cytokine tumour necrosis factor-α (TNF-α). The gene expression profile of the cells in the presence of TNF-α and under normoxic and hypoxic conditions was analysed by cDNA microarray analysis. cDNA microarray data were verified by qPCR, PCR, Western blot and zymography. Quantification of cell fusion events was determined by flow cytometry. Proteins of interest were either blocked or knocked-down using a specific inhibitor, siRNA or a blocking antibody. The data showed an up-regulation of various genes, including claudin-1 (CLDN1), ICAM1, CCL2 and MMP9 in M13SV1-Cre and/or MDA-MB-435-pFDR1 cells. Inhibition of these proteins using a blocking ICAM1 antibody, CLDN1 siRNA or an MMP9 inhibitor showed that only the blockage of MMP9 was correlated with a decreased fusion rate of the cells. Likewise, the tetracycline-based antibiotic minocycline, which exhibits anti-inflammatory properties, was also effective in both inhibiting the TNF-α-induced MMP9 expression in M13SV1-Cre cells and blocking the TNF-α-induced fusion frequency of human M13SV1-Cre breast epithelial cells and human MDA-MB-435-pFDR1 cancer cells. The matrix metalloproteinase-9 (MMP9) is most likely involved in the TNF-α-mediated fusion of human M13SV1-Cre breast epithelial cells and human MDA-MB-435-pFDR1 cancer cells. Likewise, our data indicate that the tetracycline

EFFECTIVENESS OF PNF STRETCHING VERSUS STATIC STRETCHING ON PAIN AND HAMSTRING FLEXIBILITY FOLLOWING MOIST HEAT IN INDIVIDUALS WITH KNEE OSTEOARTHRITIS

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Meena .V

2016-10-01

Full Text Available Background: Osteoarthritis (OA is a degenerative joint disease and one of the major public health problem that causesfunctional impairment and reduced quality of life. To compare the effectiveness of PNF Hold relax stretching versus Static stretching on pain and flexibility of hamstring following moist heat in individuals with knee osteoarthritis. Hamstring tightness is the major problem in knee osteoarthritis individuals. Therefore the need of study is comparing the effectiveness of PNF Hold relax stretching versus static stretching on pain and flexibility of hamstrings following moist heat in knee osteoarthritis participants. Determining the effects of PNF Hold relax stretching versus Static stretching along with moist heat on pain and hamstring flexibility by VAS and Active knee extension range of motion in knee osteoarthritis individuals. Methods: 30 subjects with symptoms of knee osteoarthritis were randomly distributed into 2 groups 15 in each group. PNF Hold relax stretching along with moist heat is compared to Static stretching along with moist heat. Pain was measured by Visual Analogue Scale (VAS and hamstring flexibility by Active knee Extension Range of Motion (AKEROM by universal goniometer. Measurements are taken pre and post intervention. Results: The results indicated PNF Hold relax stretching along with moist heat showed a statistically significant improvement in pain (p<0.05 and improvement in hamstring flexibility (p<0.05 when compared to Static stretching along with moist heat. Conclusion: Subjects with PNF Hold relax stretching along with moist heat showed significant improvement in pain reduction and improving hamstring flexibility than Static stretching along with moist heat.

PRL-3 promotes the motility, invasion, and metastasis of LoVo colon cancer cells through PRL-3-integrin β1-ERK1/2 and-MMP2 signaling

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Wu Jian

2009-11-01

Full Text Available Abstract Background Phosphatase of regenerating liver-3 (PRL-3 plays a causative role in tumor metastasis, but the underlying mechanisms are not well understood. In our previous study, we observed that PRL-3 could decrease tyrosine phosphorylation of integrin β1 and enhance activation of ERK1/2 in HEK293 cells. Herein we aim to explore the association of PRL-3 with integrin β1 signaling and its functional implications in motility, invasion, and metastasis of colon cancer cell LoVo. Methods Transwell chamber assay and nude mouse model were used to study motility and invasion, and metastsis of LoVo colon cancer cells, respectively. Knockdown of integrin β1 by siRNA or lentivirus were detected with Western blot and RT-PCR. The effect of PRL-3 on integrin β1, ERK1/2, and MMPs that mediate motility, invasion, and metastasis were measured by Western blot, immunofluorencence, co-immunoprecipitation and zymographic assays. Results We demonstrated that PRL-3 associated with integrin β1 and its expression was positively correlated with ERK1/2 phosphorylation in colon cancer tissues. Depletion of integrin β1 with siRNA, not only abrogated the activation of ERK1/2 stimulated by PRL-3, but also abolished PRL-3-induced motility and invasion of LoVo cells in vitro. Similarly, inhibition of ERK1/2 phosphorylation with U0126 or MMP activity with GM6001 also impaired PRL-3-induced invasion. In addition, PRL-3 promoted gelatinolytic activity of MMP2, and this stimulation correlated with decreased TIMP2 expression. Moreover, PRL-3-stimulated lung metastasis of LoVo cells in a nude mouse model was inhibited when integrin β1 expression was interfered with shRNA. Conclusion Our results suggest that PRL-3's roles in motility, invasion, and metastasis in colon cancer are critically controlled by the integrin β1-ERK1/2-MMP2 signaling.

Profile of MMP and TIMP Expression in Human Pancreatic Stellate Cells: Regulation by IL-1α and TGFβ and Implications for Migration of Pancreatic Cancer Cells

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Vegard Tjomsland

2016-07-01

Full Text Available Pancreatic ductal adenocarcinoma is characterized by a prominent fibroinflammatory stroma with both tumor-promoting and tumor-suppressive functions. The pancreatic stellate cell (PSC is the major cellular stromal component and the main producer of extracellular matrix proteins, including collagens, which are degraded by metalloproteinases (MMPs. PSCs interact with cancer cells through various factors, including transforming growth factor (TGFβ and interleukin (IL-1α. The role of TGFβ in the dual nature of tumor stroma, i.e., protumorigenic or tumor suppressive, is not clear. We aimed to investigate the roles of TGFβ and IL-1α in the regulation of MMP profiles in PSCs and the subsequent effects on cancer cell migration. Human PSCs isolated from surgically resected specimens were cultured in the presence of pancreatic cancer cell lines, as well as IL-1α or TGFβ. MMP production and activities in PSCs were quantified by gene array transcripts, mRNA measurements, fluorescence resonance energy transfer–based activity assay, and zymography. PSC-conditioned media and pancreatic cancer cells were included in a collagen matrix cell migration model. We found that production of IL-1α by pancreatic cancer cells induced alterations in MMP and tissue inhibitors of matrix metalloproteinase (TIMP profiles and activities in PSCs, upregulated expression and activation of MMP1 and MMP3, and enhanced migration of pancreatic cancer cells in the collagen matrix model. TGFβ counteracted the effects of IL-1α on PSCs, reestablished PSC MMP and TIMP profiles and activities, and inhibited migration of cancer cells. This suggests that tumor TGFβ has a role as a suppressor of stromal promotion of tumor progression through alterations in PSC MMP profiles with subsequent inhibition of pancreatic cancer cell migration.

Molecular characterization and expression profile of the melatonin receptor MT1 in the ovary of Tianzhu white yak (Bos grunniens).

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Hu, Jun Jie; Zhang, Xiao Yu; Zhang, Yong; Zhao, Xing Xu; Li, Fa Di; Tao, Jin Zhong

2017-02-01

Melatonin plays crucial roles in a wide range of ovarian physiological functions via the melatonin receptors (MRs). Structure and function of MRs have been well studied in sheep, cattle, and humans, but little information exists on the genetic characterization and function of these receptors in the ovary of the white yak. In the present study, the melatonin receptor MT1 was cloned by RT-PCR in the ovary of white yak; the MT1 cDNA fragment obtained (843bp) comprised an open reading frame (827bp) encoding a protein containing 275 residues, characterized by seven transmembrane regions and an NRY motif, two distinct amino acid replacements were found. The white yak MT1 had a 83.9-98.6% protein sequence identity with that of nine other mammals. Using RT-PCR, the expression levels of MT1, MT2, and LHR in the ovary of pregnant and non-pregnant white yaks were compared, revealing higher levels of all genes in pregnant yaks: 3.83-fold increase for MT1 (Pmelatonin and MT1 are associated with the corpus luteum function of pregnancy maintenance and follicular development during oocyte maturation in the white yak. Copyright © 2015 Elsevier Inc. All rights reserved.

Clinical significance of determination the changes of serum CGRP, MMP-9 and TIMP-1 levels both before and after treatment in pediatric patients with bronchial asthma

International Nuclear Information System (INIS)

Chen Guijin

2011-01-01

Objective: To explore the changes of serum CGRP, MMP-9 and TIMP-1 levels after treatment in pediatric patients with bronchial asthma. Methods: Serum CGRP(with RIA), MMP-9, TIMP-1 (with ELISA) levels were measured in 32 patients with bronchial asthma both before and after treatment as well as in 35 controls. Results: Before treatment, the serum CGRP levels was significantly lower in patients than those in controls (P 0.05). Conclusion: Abnormal lower CGRP and high MMP-9, TIMP-1 levels might play an important role in the pathogenesis and development of bronchial asthma in children. (authors)

PAI-1, a target gene of miR-143, regulates invasion and metastasis by upregulating MMP-13 expression of human osteosarcoma.

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Hirahata, Mio; Osaki, Mitsuhiko; Kanda, Yusuke; Sugimoto, Yui; Yoshioka, Yusuke; Kosaka, Nobuyoshi; Takeshita, Fumitaka; Fujiwara, Tomohiro; Kawai, Akira; Ito, Hisao; Ochiya, Takahiro; Okada, Futoshi

2016-05-01

Despite recent improvements in the therapy for osteosarcoma, 30-40% of osteosarcoma patients die of this disease, mainly due to its lung metastasis. We have previously reported that intravenous injection of miR-143 significantly suppresses lung metastasis of human osteosarcoma cells (143B) in a mouse model. In this study, we examined the biological role and mechanism of miR-143 in the metastasis of human osteosarcoma cells. We identified plasminogen activator inhibitor-1 (PAI-1) as a direct target gene of miR-143. To determine the role of PAI-1 in human osteosarcoma cells, siRNA was transfected into 143B cells for knockdown of PAI-1 expression. An in vitro study showed that downregulation of PAI-1 suppressed cell invasion activity, but not proliferation. Moreover, injection of PAI-1 siRNA into a primary lesion in the osteosarcoma mouse model inhibited lung metastasis compared to control siRNA-injected mice, without influencing the proliferative activity of the tumor cells. Subsequent examination using 143B cells revealed that knockdown of PAI-1 expression resulted in downregulation of the expression and secretion of matrix metalloproteinase-13 (MMP-13), which is also a target gene of miR-143 and a proteolytic enzyme that regulates tumor-induced osteolysis. Immunohistochemical analysis using clinical samples showed that higher miR-143 expressing cases showed poor expression of PAI-1 in the primary tumor cells. All such cases belonged to the lung metastasis-negative group. Moreover, the frequency of lung metastasis-positive cases was significantly higher in PAI-1 and MMP-13 double-positive cases than in PAI-1 or MMP-13 single-positive or double-negative cases (P target gene of miR-143, regulates invasion and lung metastasis via enhancement of MMP-13 expression and secretion in human osteosarcoma cells, suggesting that these molecules could be potential therapeutic target genes for preventing lung metastasis in osteosarcoma patients. © 2016 The Authors. Cancer

Repeated cadmium nebulizations induce pulmonary MMP-2 and MMP-9 production and enphysema in rats

International Nuclear Information System (INIS)

Kirschvink, Nathalie; Vincke, Gregoire; Fievez, Laurence; Onclinx, Cecile; Wirth, Delphine; Belleflamme, Michele; Louis, Renaud; Cataldo, Didier; Peck, Michael J.; Gustin, Pascal

2005-01-01

This study describes induction of pulmonary inflammation, production of matrix metalloprotease of type 2 (MMP-2) and type 9 (MMP-9), and emphysema in cadmium (Cd)-exposed rats. Sprague-Dawley rats were randomly distributed into two groups: one placebo-exposed group undergoing saline (NaCl 0.9%) inhalation (n = 30) and one Cd-exposed group undergoing cadmium (CdCl 2 0.1%) inhalation (n = 30). The animals of the placebo- and Cd-exposed groups were divided in five subgroups (n = 6). Subgroups underwent either a single exposure of 1 h or repeated exposures three times weekly for 1 h during 3 weeks (3W), 5 weeks (5W), 5 weeks followed by 2 weeks without exposure (5W + 2) or 5 weeks followed by 4 weeks without exposure (5W + 4). Each animal underwent determination of enhanced pause (Penh) as index of airflow limitation prior to the first exposure as well as before sacrifice. The animals were sacrificed the day after their last exposure. The left lung was fixed for histomorphometric analysis (determination of median interwall distance (MIWD)), whilst bronchoalveolar lavage fluid (BALF) was collected from the right lung. BALF was analyzed cytologically, and MMP-2 and MMP-9 levels were determined by gelatine zymography. Twelve rats previously instilled with pancreatic elastase were used as positive emphysema controls and underwent the same investigations. Cd-exposure induced a significant increase of BALF macrophages, neutrophils and MMP-9 up to 5W + 4, whereas MMP-2 gelatinolytic activity returned to baseline levels within 5W. MIWD was significantly increased in all repeatedly Cd-exposed groups and elastase-treated rats. Penh was increased in Cd-exposed rats after a single exposure and after 3W. MMP gelatinolytic activity was significantly correlated with macrophages, neutrophils and Penh. In repeatedly exposed rats, MIWD was positively and significantly correlated with MMP gelatinolytic activity, suggesting that increased MMP-2 and MMP-9 production favours the development

Salvia miltiorrhiza extract inhibits TPA-induced MMP-9 expression and invasion through the MAPK/AP-1 signaling pathway in human breast cancer MCF-7 cells.

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Kim, Jeong-Mi; Noh, Eun-Mi; Song, Hyun-Kyung; Lee, Minok; Lee, Soo Ho; Park, Sueng Hyuk; Ahn, Chan-Keun; Lee, Guem-San; Byun, Eui-Baek; Jang, Beom-Su; Kwon, Kang-Beom; Lee, Young-Rae

2017-09-01

Cancer cell invasion is crucial for metastasis. A major factor in the capacity of cancer cell invasion is the activation of matrix metalloproteinase-9 (MMP-9), which degrades the extracellular matrix. Salvia miltiorrhiza has been used as a promotion for blood circulation to remove blood stasis. Numerous previous studies have demonstrated that S. miltiorrhiza extracts (SME) decrease lipid levels and inhibit inflammation. However, the mechanism behind the effect of SME on breast cancer invasion has not been identified. The inhibitory effects of SME on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced MMP-9 expression were assessed using western blotting, reverse transcription-quantitative polymerase chain reaction and zymography assays. MMP-9 upstream signal proteins, including mitogen-activated protein kinases and activator protein 1 (AP-1) were also investigated. Cell invasion was assessed using a matrigel invasion assay. The present study demonstrated the inhibitory effects of the SME ethanol solution on MMP-9 expression and cell invasion in TPA-treated MCF-7 breast cancer cells. SME suppressed TPA-induced MMP-9 expression and MCF-7 cell invasion by blocking the transcriptional activation of AP-1. SME may possess therapeutic potential for inhibiting breast cancer cell invasiveness.

Increase in IL-6, TNF-a, and MMP-9, but not sICAM-1, concentrations depends on exercise duration

DEFF Research Database (Denmark)

Reihmane, Dace; Jurka, Antra; Tretjakovs, Peteris

2013-01-01

), tumour necrosis factor-α (TNF-α), soluble form of intercellular adhesion molecule-1 (sICAM-1), and matrix metalloproteinase-9 (MMP-9) was studied in 22 half-marathon (HM) and 18 marathon (M) male amateur runners who completed their exercise task in 1.8 ± 0.2 (mean ± standard deviation) and 3.6 ± 0.4 h...

Expression of matrix metalloproteinase 9 (MMP-9) and tissue inhibitor of metalloproteinases 1 (TIMP-1) by colorectal cancer cells and adjacent stroma cells--associations with histopathology and patients outcome

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Jensen, Søren Astrup; Vainer, Ben; Bartels, Annette

2010-01-01

To elucidate cellular features accountable for colorectal cancers' (CRC) capability to invade normal tissue and to metastasize, we investigated the level of the collagenase matrix metalloproteinase 9 (MMP-9) and its physiological inhibitor tissue inhibitor of metalloproteinases 1 (TIMP-1) in canc...

Possible Mechanisms of Di(2-ethylhexyl Phthalate-Induced MMP-2 and MMP-9 Expression in A7r5 Rat Vascular Smooth Muscle Cells

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Mei-Fen Shih

2015-12-01

Full Text Available Proliferation and migration of vascular smooth muscle cells (VSMC are important in the development and/or progression of many cardiovascular diseases, including atherosclerosis. Evidence shows that matrix metalloproteinase (MMP-2 and MMP-9 are related to the pathogenesis of atherosclerosis. The expressions of MMP-2 and MMP-9 in atherosclerosis are regulated via various pathways, such as p38 mitogen activated protein kinase (MAPK, extracellular signal regulated kinase 1 and 2 (ERK1/2, Akt, and nuclear factor kappa (NF-κB. Di(2-ethylhexyl phthalate (DEHP has been shown to induce atherosclerosis by increasing tumor necrosis factor (TNF-α, interleukin (IL-6, and intercellular adhesion molecule (ICAM productions. However, whether DEHP poses any effects on MMP-2 or MMP-9 expression in VSMC has not yet been answered. In our studies, rat aorta VSMC was treated with DEHP (between 2 and 17.5 ppm and p38 MAPK, ERK1/2, Akt, NF-κB, and MMP-2 and MMP-9 proteins and activities were measured. Results showed that the presence of DEHP can induce higher MMP-2 and MMP-9 expression than the controls. Similar results on MMP-regulating proteins, i.e., p38 MAPK, ERK1/2, Akt, and NF-κB, were also observed. In summary, our current results have showed that DEHP can be a potent inducer of atherosclerosis by increasing MMP-2 and MMP-9 expression at least through the regulations of p38 MAPK, ERK1/2, Akt, and NF-κB.

Overexpression of MMP-3 and uPA with Diminished PAI-1 Related to Metastasis in Ductal Breast Cancer Patients Attending a Public Hospital in Mexico City

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Luis Miguel Barajas-Castañeda

2016-01-01

Full Text Available Extracellular matrix metalloproteases and the fibrinolytic system are important protease systems interacting with each other in charge of remodeling and recycling of tissues. Their role in tumor invasion and metastasis is often discussed. In this study several metalloproteases such as MMP-1, MMP-3, MMP-9, and TIMP-1 together with molecules from the fibrinolytic system like uPA, its receptor uPAR, and its inhibitor, PAI-1, were studied by immune-histochemistry to establish a comparison with and without metastasis. From the (118 primary tumors of Mexican patients with ductal breast cancer studied, 56% were grade II and 69% were size T2; the group with metastatic ganglia included 64 samples (54.3%. In patients with metastasis the estimated expression of MMP-3 and uPA (resp., 28% and 45% was higher than that from no metastatic tumors; it means there is higher expression of both markers in metastatic tumors (p<0.05. At the same time, metastatic tumors showed statistically significant lower signal of PAI-1 (24% than tumors without metastasis (p<0.05. We concluded that overexpression of MMP-3 and uPA, altogether with diminished expression of PAI-1 from metastatic tumors, might be a crucial step towards metastasis in ductal breast cancer. Nevertheless, additional studies in different populations are necessary to establish a pattern.

An economical mtDNA SNP assay detecting different mitochondrial haplogroups in identical HVR 1 samples of Caucasian ancestry.

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Köhnemann, Stephan; Hohoff, Carsten; Pfeiffer, Heidi

2009-09-01

We had sequenced 329 Caucasian samples in Hypervariable Region 1 (HVR 1) and found that they belong to eleven different mitochondrial DNA (mtDNA) haplotypes. The sample set was further analysed by an mtDNA assay examining 32 single nucleotide polymorphisms (SNPs) for haplogroup discrimination. In a validation study on 160 samples of different origin it was shown that these SNPs were able to discriminate between the evolved superhaplogroups worldwide (L, M and N) and between the nine most common Caucasian haplogroups (H, I, J, K, T, U, V, W and X). The 32 mtDNA SNPs comprised 42 different SNP haplotypes instead of only eleven haplotypes after HVR 1 sequencing. The assay provided stable results in a range of 5ng genomic DNA down to virtually no genomic DNA per reaction. It was possible to detect samples of African, Asian and Eurasian ancestry, respectively. The 32 mtDNA SNP assay is a helpful adjunct to further distinguish between identical HVR 1 sequences of Caucasian origin. Our results suggest that haplogroup prediction using HVR 1 sequencing provides instable results. The use of coding region SNPs for haplogroup assignment is more suited than using HVR 1 haplotypes.

Poly(I:C) induces expressions of MMP-1, -2, and -3 through various signaling pathways including IRF3 in human skin fibroblasts.

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Yao, Cheng; Lee, Dong Hun; Oh, Jang-Hee; Kim, Min-Kyoung; Kim, Kyu Han; Park, Chi-Hyun; Chung, Jin Ho

2015-10-01

Ultraviolet (UV) irradiation can result in premature skin aging (photoaging) which is characterized by decreased expression of collagen and increased expression of matrix metalloproteinases (MMPs). Double-stranded RNAs (dsRNAs) can be generated at various conditions including virally infected cells or UV-damaged skin cells. Recent studies have shown that a synthetic dsRNA, polyinosinic-polycytidylic acid (poly(I:C)), can reduce procollagen expression in human skin fibroblasts. However, little is known about the effect of poly(I:C) on the expression of MMPs in skin fibroblasts and its underlying mechanisms. We examined the effect of poly(I:C) on MMP-1, -2, and -3 expressions in human skin fibroblasts. Then, we further explored the underlying signaling pathways involved in the processes. Human skin fibroblasts were treated with poly(I:C) for the indicated times in the presence or the absence of various chemical inhibitors or small interfering RNAs (siRNAs) at the indicated concentrations. Protein and mRNA levels of various target molecules were examined by Western blotting and quantitative real-time PCR, respectively. Poly(I:C) induced MMP-1, -2, and -3 expressions, which were dependent on TLR3. Poly(I:C) also induced activations of the mitogen-activated protein kinases (MAPKs), the nuclear factor-kappaB (NF-κB) and the interferon regulatory factor 3 (IRF3) pathways. By using specific inhibitors, we found that poly(I:C)-induced expressions of MMP-1, -2, and -3 were differentially regulated by these signaling pathways. In particular, we found that the inhibition of IRF3 signaling pathways attenuated poly(I:C)-induced expressions of all the three MMPs. Our data show that the expressions of MMP-1, -2, and -3 are induced by poly(I:C) through various signaling pathways in human skin fibroblasts and suggest that TLR3 and/or IRF3 may be good targets for regulating the expressions of MMP-1, -2, and -3 induced by dsRNAs. Copyright © 2015 Elsevier Ireland Ltd. All rights

The Relationship Between Transcript Expression Levels of Nuclear Encoded (TFAM, NRF1 and Mitochondrial Encoded (MT-CO1 Genes in Single Human Oocytes During Oocyte Maturation

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Ghaffari Novin M.

2015-06-01

Full Text Available In some cases of infertility in women, human oocytes fail to mature when they reach the metaphase II (MII stage. Mitochondria plays an important role in oocyte maturation. A large number of mitochondrial DNA (mtDNA, copied in oocytes, is essential for providing adenosine triphosphate (ATP during oocyte maturation. The purpose of this study was to identify the relationship between transcript expression levels of the mitochondrial encoded gene (MT-CO1 and two nuclear encoded genes, nuclear respiratory factor 1 (NRF1 and mitochondrial transcription factor A (TFAM in various stages of human oocyte maturation. Nine consenting patients, age 21-35 years old, with male factors were selected for ovarian stimulation and intracytoplasmic sperm injection (ICSI procedures. mRNA levels of mitochondrial- related genes were performed by singlecell TaqMan® quantitative real-time polymerase chain reaction (qRT-PCR. There was no significant relationship between the relative expression levels in germinal vesicle (GV stage oocytes (p = 0.62. On the contrary, a significant relationship was seen between the relative expression levels of TFAM and NRF1 and the MT-CO1 genes at the stages of metaphase I (MI and MII (p = 0.03 and p = 0.002. A relationship exists between the transcript expression levels of TFAM and NRF1, and MT-CO1 genes in various stages of human oocyte maturation.

Interleukin-1β modulates endochondral ossification by human adult bone marrow stromal cells

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M Mumme

2012-09-01

Full Text Available Inflammatory cytokines present in the milieu of the fracture site are important modulators of bone healing. Here we investigated the effects of interleukin-1β (IL-1β on the main events of endochondral bone formation by human bone marrow mesenchymal stromal cells (BM-MSC, namely cell proliferation, differentiation and maturation/remodelling of the resulting hypertrophic cartilage. Low doses of IL-1β (50 pg/mL enhanced colony-forming units-fibroblastic (CFU-f and -osteoblastic (CFU-o number (up to 1.5-fold and size (1.2-fold in the absence of further supplements and glycosaminoglycan accumulation (1.4-fold upon BM-MSC chondrogenic induction. In osteogenically cultured BM-MSC, IL-1β enhanced calcium deposition (62.2-fold and BMP-2 mRNA expression by differential activation of NF-κB and ERK signalling. IL-1β-treatment of BM-MSC generated cartilage resulted in higher production of MMP-13 (14.0-fold in vitro, mirrored by an increased accumulation of the cryptic cleaved fragment of aggrecan, and more efficient cartilage remodelling/resorption after 5 weeks in vivo (i.e., more TRAP positive cells and bone marrow, less cartilaginous areas, resulting in the formation of mature bone and bone marrow after 12 weeks. In conclusion, IL-1β finely modulates early and late events of the endochondral bone formation by BM-MSC. Controlling the inflammatory environment could enhance the success of therapeutic approaches for the treatment of fractures by resident MSC and as well as improve the engineering of implantable tissues.

Human iPSC-Derived Neural Progenitors Are an Effective Drug Discovery Model for Neurological mtDNA Disorders.

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Lorenz, Carmen; Lesimple, Pierre; Bukowiecki, Raul; Zink, Annika; Inak, Gizem; Mlody, Barbara; Singh, Manvendra; Semtner, Marcus; Mah, Nancy; Auré, Karine; Leong, Megan; Zabiegalov, Oleksandr; Lyras, Ekaterini-Maria; Pfiffer, Vanessa; Fauler, Beatrix; Eichhorst, Jenny; Wiesner, Burkhard; Huebner, Norbert; Priller, Josef; Mielke, Thorsten; Meierhofer, David; Izsvák, Zsuzsanna; Meier, Jochen C; Bouillaud, Frédéric; Adjaye, James; Schuelke, Markus; Wanker, Erich E; Lombès, Anne; Prigione, Alessandro

2017-05-04

Mitochondrial DNA (mtDNA) mutations frequently cause neurological diseases. Modeling of these defects has been difficult because of the challenges associated with engineering mtDNA. We show here that neural progenitor cells (NPCs) derived from human induced pluripotent stem cells (iPSCs) retain the parental mtDNA profile and exhibit a metabolic switch toward oxidative phosphorylation. NPCs derived in this way from patients carrying a deleterious homoplasmic mutation in the mitochondrial gene MT-ATP6 (m.9185T>C) showed defective ATP production and abnormally high mitochondrial membrane potential (MMP), plus altered calcium homeostasis, which represents a potential cause of neural impairment. High-content screening of FDA-approved drugs using the MMP phenotype highlighted avanafil, which we found was able to partially rescue the calcium defect in patient NPCs and differentiated neurons. Overall, our results show that iPSC-derived NPCs provide an effective model for drug screening to target mtDNA disorders that affect the nervous system. Copyright © 2016 Elsevier Inc. All rights reserved.

Differential inhibition of activity, activation and gene expression of MMP-9 in THP-1 cells by azithromycin and minocycline versus bortezomib: A comparative study.

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Jennifer Vandooren

Full Text Available Gelatinase B or matrix metalloproteinase-9 (MMP-9 (EC 3.4.24.35 is increased in inflammatory processes and cancer, and is associated with disease progression. In part, this is due to MMP-9-mediated degradation of extracellular matrix, facilitating influx of leukocytes into inflamed tissues and invasion or metastasis of cancer cells. MMP-9 is produced as proMMP-9 and its propeptide is subsequently removed by other proteases to generate proteolytically active MMP-9. The significance of MMP-9 in pathologies triggered the development of specific inhibitors of this protease. However, clinical trials with synthetic inhibitors of MMPs in the fight against cancer were disappointing. Reports on active compounds which inhibit MMP-9 should be carefully examined in this regard. In a considerable set of recent publications, two antibiotics (minocycline and azythromycin and the proteasome inhibitor bortezomib, used in cancers, were reported to inhibit MMP-9 at different stages of its expression, activation or activity. The current study was undertaken to compare and to verify the impact of these compounds on MMP-9. With exception of minocycline at high concentrations (>100 μM, the compounds did not affect processing of proMMP-9 into MMP-9, nor did they affect direct MMP-9 gelatinolytic activity. In contrast, azithromycin specifically reduced MMP-9 mRNA and protein levels without affecting NF-κB in endotoxin-challenged monocytic THP-1 cells. Bortezomib, although being highly toxic, had no MMP-9-specific effects but significantly upregulated cyclooxygenase-2 (COX-2 activity and PGE2 levels. Overall, our study clarified that azithromycin decreased the levels of MMP-9 by reduction of gene and protein expression while minocycline inhibits proteolytic activity at high concentrations.

In vitro effects of infrared A radiation on the synthesis of MMP-1, catalase, superoxide dismutase and GADD45 alpha protein.

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Costa, Adilson; Eberlin, Samara; Clerici, Stefano P; Abdalla, Beatrice M Z

2015-01-01

Harmful influences in the process of photoaging and skin damage are associated with infrared A (IRA) radiation, such as, disturbance of dermal extracellular matrix by up regulation of matrix metalloproteinase-1 (MMP1). Furthermore, DNA damage, induction of cytotoxicity and oxidative stress by decreasing natural antioxidant ability has been reported after acute exposure to IRA. The present study provides additional evidence that IRA radiation response in human skin fibroblasts produces deleterious effects to the cell, such as accelerating aging and weakening of their antioxidant defense mechanism. Human skin fibroblasts were exposed to a non-cytotoxic dose of IRA radiation and cultured for different periods for further collection of cell-free supernatants and lysates, and quantification of MMP-1, catalase, superoxide dismutase, and GADD45a. Our results corroborate previous published data and strongly indicate a negative impact of IRA radiation on the skin physiological by mechanisms involving reduced endogenous antioxidant enzymatic defense, increased MMP-1 and decreased repair process of DNA by reducing GADD45a protein, in cultured human fibroblasts. From a clinical perspective, IRA radiation acts by mechanisms distinct from those observed in ultraviolet radiation indicating the need for developing and making available cosmetics for skin care with properties beyond protection exerted by traditional sunscreens.

Chronic cerebral hypoperfusion-induced impairment of Aβ clearance requires HB-EGF-dependent sequential activation of HIF1α and MMP9.

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Ashok, Anushruti; Rai, Nagendra Kumar; Raza, Waseem; Pandey, Rukmani; Bandyopadhyay, Sanghamitra

2016-11-01

Chronic cerebral hypoperfusion (CCH) manifests Alzheimer's Disease (AD) neuropathology, marked by increased amyloid beta (Aβ). Besides, hypoxia stimulates Heparin-binding EGF-like growth factor (HB-EGF) mRNA expression in the hippocampus. However, involvement of HB-EGF in CCH-induced Aβ pathology remains unidentified. Here, using Bilateral Common Carotid Artery Occlusion mouse model, we explored the mechanism of HB-EGF regulated Aβ induction in CCH. We found that HB-EGF inhibition suppressed, while exogenous-HB-EGF triggered hippocampal Aβ, proving HB-EGF-dependent Aβ increase. We also detected that HB-EGF affected the expression of primary Aβ transporters, receptor for advanced glycation end-products (RAGE) and lipoprotein receptor-related protein-1 (LRP-1), indicating impaired Aβ clearance across the blood-brain barrier (BBB). An HB-EGF-dependent loss in BBB integrity supported impaired Aβ clearance. The effect of HB-EGF on Amyloid Precursor Protein pathway was relatively insignificant, suggesting a lesser effect on Aβ generation. Delving into BBB disruption mechanism demonstrated HB-EGF-mediated stimulation of Matrix metalloprotease-9 (MMP9), which affected BBB via HB-EGF-ectodomain shedding and epidermal growth factor receptor activation. Examining the intersection of HB-EGF-regulated pathway and hypoxia revealed HB-EGF-dependent increase in transcription factor, Hypoxia-inducible factor-1alpha (HIF1α). Further, via binding to hypoxia-responsive elements in MMP9 gene, HIF1α stimulated MMP9 expression, and therefore appeared as a prominent intermediary in HB-EGF-induced BBB damage. Overall, our study reveals the essential role of HB-EGF in triggering CCH-mediated Aβ accumulation. The proposed mechanism involves an HB-EGF-dependent HIF1α increase, generating MMP9 that stimulates soluble-HB-EGF/EGFR-induced BBB disintegration. Consequently, CCH-mediated hippocampal RAGE and LRP-1 deregulation together with BBB damage impair Aβ transport and clearance