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Inoculation of dry bean (Phaseolus vulgaris L.) might have potential to increase symbiotic nitrogen fixation (SNF) and to reduce dependency on chemical fertilizers. Peat based inoculant can...

Decomposition of crop residue releases nutrients and preserves soil organic matter dynamics. The decomposition rates of six crop residues namely, corn (Zea mays), dry bean (Phaseolus vulgaris), soy...

Transgenic glyphosate-resistant (GR) soybean (Glycine max L. Merr.) is cultivated throughout the United States. Soybean growth is influenced by the presence of weeds, although managing non-crop vegetation can potentially impact soil arthropods, which are being increasingly recognized for their impacts on soil health, plant growth, and above-ground trophic interactions. We investigated how weed management (weedy controls, hand-weeded, glyphosate herbicide) and soil insecticide (chlorpyrifos) application affected densities of soil arthropods, soil nutrient availability, soybean growth and yield, and densities of an above-ground herbivore on sandy and clayey soils for two consecutive growing seasons. The soil insecticide treatment was intended to lower densities of subterranean arthropods to gain insight into how their presence influenced other factors, although their densities were primarily reduced the first year of the study. Surprisingly, weed management and soil insecticide use had virtually no interactive effects on any response. Weed presence had a positive effect on soil K at the sandy site and on nodule density per unit root. Negative effects of weed management on plant growth and aphids were related to the presence of weeds rather than herbicide use. Reduced soybean aphid density (at the clayey site) and soil P availability (at the sandy site) were associated with insecticide treated plots. Conversely, several measures of plant productivity, including number of nodules per unit root, and root and shoot biomass increased in +insecticide plots compared to other treatments, although effect strength depended on year and location. Collembola were the dominant soil microarthropod, and their densities in 2012 were negatively associated with nodule numbers in 2013. One explanatory hypothesis is that increased plant growth in plots treated with insecticide was caused by altered soil arthropod-microorganism interactions, possibly affecting arbuscular mycorrhizae function. This work highlights the importance of management decisions that affect soil arthropods in annual legume production systems.

Symbiotic nitrogen fixation (SNF) is an environmentally safe source of nitrogen (N) to the crop plants. In total, 12 dry bean (Phaseolus vulgaris L.) cultivars from pinto, navy, black, and kidney m...

Perennial grasses have been promoted to supply cellulose for ethanol production but possible consequences on soil organic carbon (SOC) should be assessed before adoption in a given region. Specific objectives were to (i) compare above-ground biomass C production and depth distribution of SOC content of winter wheat (Triticum aestivum L.) and switchgrass (Panicum virgatum L.), (ii) determine the contribution of switchgrass to SOC, and (iii) characterize soil C inventories under each crop across a wide range of precipitation regimes. A 6-yr (2008-13) field study with winter wheat and switchgrass cv. Blackwell was conducted near Adams, OR, U.S. Solid set sprinkler irrigation was used to create a productivity gradient of low, intermediate, and high precipitation levels. The contribution of switchgrass to SOC was identified with d13C measurements. Winter wheat had significantly higher 5-yr-average aboveground biomass C production than switchgrass across precipitation levels and winter wheat under high precipitation produced significantly higher aboveground biomass C than medium and low precipitation. After 6-yr, SOC did not change under switchgrass at any depth increment. Results of d13C indicated switchgrass contributed profile SOC of 5.99, 5.41, and 9.47 Mg C ha-1 at low, intermediate, and high precipitation levels. Soil organic carbon in dry cropland is resistant to a shift in crop species. Switchgrass does not increase SOC more than winter wheat in dry cropland of the semi-arid Pacific Northwest.

Additions of ammonium sulfate and urease inhibitor with urea might have potential to increase yield and quality due to increased supply of sulfur (S) and reduction in volatilization loss of nitrogen (N), respectively. Treatments consisting, (i) urea alone, (ii) urea with ammonium sulfate (UAS) and (iii) homogeneous granular fertilizer containing urea and ammonium sulfate (HBU), and (iv) urease inhibitor treated urea (UI), at two application rates, 134 and 168 kg N ha−1, were compared for spring wheat (Triticum aestivum L.) and sugar beet (Beta vulgaris L.) production at Glyndon and Ada of Minnesota, USA, during 2014–2015. For both crops, additions of UI had increased yield due to higher soil N availability over urea only in 2015. In 2014, UI also increased the grain protein content over urea and UAS increased sugar content only over HBU in 2015. Spring wheat and sugarbeet yield and quality response to urease inhibitor or supply of S in the form of UAS depends on soil and climatic factors.

Tea is one of the most widely consumed beverages in the world. Black tea, obtained from the leaves of Camellia sinensis is the preferred beverage in India and in most western countries. Epidemiological studies on black tea and cancer are limited. However, preliminary studies indicate a positive correlation between black tea consumption and a lower incidence of breast and ovarian cancer. In the present communication, we wanted to see the effect of black tea extract and the polyphenol theaflavin on cell-ECM interaction, MMP activity etc. to strengthen the anti-cancer effect of black tea.

Sugar beet (Beta vulgaris L.) production leaves very little residue to protect the soil from erosion during late fall and early spring. Biomass from interseeded cover crops provides an opportunity to protect soils from erosion during the fallow period. A three-year field study was conducted to assess cover crop interseeding in sugar beet production systems under rainfed conditions in Ada, MN, USA. Specifically, the impact of cover crop interseeding time, early (mid-June) vs. late (late June or early July), and four species, winter pea (Pisum sativum L.), rye (Secale cereale L.), winter camelina [Camelina sativa (L.) Crantz], and brown mustard (Brassica juncea L.), on cover crop biomass and sugar beet root yield, sucrose concentration, and recoverable sucrose yield. Cover crop biomass accumulation in sugar beet varied across years depending on distribution of rainfall after interseeding. Considering all three years, above ground biomass of cover crops ranged between 28 kg ha−1 to 2347 kg ha−1 at sugar beet harvest. Early interseeded cover crops produced threefold greater cover crop biomass than late interseeded cover crops. Across years, mean aboveground biomass produced by species was from highest to lowest, winter pea > rye > brown mustard > winter camelina. The highest recoverable sucrose yield was observed with early-interseeded brown mustard (13.8 Mg ha-1), late-interseeded winter camelina (11.2 Mg ha-1), and late-interseeded rye (10.46 Mg ha-1) in 2018, 2019, and 2020 growing seasons, respectively. Three years of results have shown that sugar beet grown with interseeded cover crops will produce root yields comparable to the control with no cover crops. Cover crops had no negative impact on root yield and sucrose concentration, but the selection of species and planting time affected growth and cover provided by the cover crops. This study provides evidence that interseeded cover crops offer a good option for protecting the soil during fallow periods in sugar beet production systems, without negatively affecting sugar beet yield or quality.

Cellulosic ethanol commercialization promises to produce energy from agricultural biomass. Available biomass depends on plant net primary productivity (NPP) and crop type, which maintain total soil organic carbon (TOC). Effect of crop-type, residue removal, and NPP on ethanol yield and TOC levels were assessed by means of a three-pool C model derived from long-term soil incubation, acid hydrolysis, and curve fitting of a nonlinear regression model. A 2-yr field study consisting of three input regimes (Low, Medium, or High NPP), three crops [corn (Zea mays L.), wheat (Triticum aestivum, L.), and switchgrass (SG, Panicum virgatum L., cv. Blackwell)], and two harvest levels [residue removed (R) or residue not removed (NR)] was conducted near Prosser, WA, USA. After 2 yr, ethanol yield of all crops were similar under Low NPP while ethanol yield of SG under Medium and High NPP was significantly greater than that of corn or wheat under the same NPP. Switchgrass significantly contributed to active [mean residence time (MRT) 500 yr) soil C pools. Other crops had net zero or significantly reduced C pools. During a transition to cellulosic energy production, SG will contribute to soil C maintenance across a range of potential net productivity.

Increased MMP-1 expression in various tumor cells is significantly correlated with cancer progression. Enhanced secreted and intracellular level of MMP-1 is found in breast cancer cell line MDA-MB-231 in presence of ECM glycoprotein fibronectin. To extrapolate this study into in vivo system, we observed the expression of MMP-1, fibronectin and α5β1 integrin, which is the receptor for fibronectin, in breast cancer tissue samples. Expression of active form of MMP-1 was increased in tumor samples compared with the non-tumor counterpart. In some samples pro-MMP-1 was decreased but active form was increased in tumor part. The difference was more prominent in advanced stage tumor. ELISA showed appreciable increase in expression of α5 and β1integrins in tumor tissue in comparison to the non tumor counterpart in case of advanced stage tumor. Though there is no appreciable difference in fibronectin concentration, enhanced α5, β1 integrin expression may mediate enhanced cell-ECM interaction to upregulate MMP-1 in tumor samples compared with the matched control.

Studies on interaction of tumor cells with ECM components showed increased extracellular protease activity mediated by the family of matrix metalloproteinases (MMPs). Here we studied the effect of human prostate adenocarcinoma PC-3 cells-fibronectin (FN) interaction on MMPs and the underlying signaling pathways. Culturing of PC-3 cells on FN-coated surface upregulated MMP-9 and MMP-1. This response is abrogated by the blockade of α5 integrin. siRNA and inhibitor studies indicate possible involvement of phosphatidyl-inositol-3-kinase (PI-3K), focal adhesion kinase (FAK) and nuclear factor-kappaB (NF-κB) in FN-induced upregulation of MMPs. FN treatment also enhanced phosphorylation of FAK, PI3K, protein kinase B (PKB or Akt), nuclear translocation of NF-κB, surface expression of CD-44, and cell migration. Our findings indicate that, binding of PC-3 cells to FN, possibly via α5β1 integrin, induces signaling involving FAK, PI-3K, Akt, NF-κB followed by upregulation of MMP-9 and MMP-1. CD-44 may have role in modulating MMP-9 activity.

This study aimed to detect the comparative expression and activity of matrix metalloproteinase-9 (MMP-9) and its correlation with known pathological parameters such as estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 (HER2) in 81 malignant breast tumors and adjacent normal breast tissues and in blood sera of these patients from different clinical TNM stages (ductal carcinoma in situ to T4) of breast cancer. MMP-9 was highly expressed in node-positive tumors and the preoperative blood serum of patients, but MMP-9 activity was appreciably inhibited in blood serum samples collected after surgery. The mature form of MMP-9 (84 kD) was expressed only in clinical stage III tumors (T2-4). Appreciable reduction of tissue inhibitor of metalloproteinase 1, phosphorylation of epidermal growth factor receptor, and translocation of nuclear factor-κΒ suggested their possible role in MMP-9 activation in HER2-positive breast cancer Overexpression and activation of MMP-9 predicted a higher stage of hormone-sensitive ductal breast carcinoma. Downregulation of the endogenous inhibitor of MMP-9, tissue inhibitor of metalloproteinase 1, and translocation of the transcription factor nuclear factor-κΒ in tumors may have an appreciable role in the overexpression of MMP-9. However, MMP-9 activation was not correlated with expression of estrogen and progesterone receptors. Evaluation of MMP-9 expression may provide valuable information about breast cancer treatment.

The retinoids, a class of compounds related to vitamin A exert their effects via the retinoic acid (RA) receptors (RARs) and rexinoid receptors and play important roles in a number of physiological processes including vision, tissue maintenance, differentiation and embryonic development. RA, including all-trans RA (ATRA) and its stereoisomer 9-cis RA, have gained importance as chemotherapeutic agents and inhibit development of a number of tumours. ATRA is used, along with chemotherapeutic agents to treat acute promyelocytic leukemia by promoting myeloid differentiation (without destroying the leukemic cells) via modulation of PML-RARα and other transcription factors. The effect of ATRA on solid tumours and the molecular mechanisms by which they exert their effects have not been elucidated in as much detail. Several studies indicate that ATRA promotes apoptosis and downregulates cellular proteases including matrix metalloproteinases and components of the intracellular signalling cascades at various levels. Thus ATRA appears to have good potential as an anti-tumorigenic drug in several carcinomas including breast cancers and melanomas.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms., Microorganisms are vital in mediating the earth's biogeochemical cycles; yet, despite our rapidly increasing ability to explore complex environmental microbial communities, the relationship between microbial community structure and ecosystem processes remains poorly understood. Here, we address a fundamental and unanswered question in microbial ecology: 'When do we need to understand microbial community structure to accurately predict function?' We present a statistical analysis investigating the value of environmental data and microbial community structure independently and in combination for explaining rates of carbon and nitrogen cycling processes within 82 global datasets. Environmental variables were the strongest predictors of process rates but left 44% of variation unexplained on average, suggesting the potential for microbial data to increase model accuracy. Although only 29% of our datasets were significantly improved by adding information on microbial community structure, we observed improvement in models of processes mediated by narrow phylogenetic guilds via functional gene data, and conversely, improvement in models of facultative microbial processes via community diversity metrics. Our results also suggest that microbial diversity can strengthen predictions of respiration rates beyond microbial biomass parameters, as 53% of models were improved by incorporating both sets of predictors compared to 35% by microbial biomass alone. Our analysis represents the first comprehensive analysis of research examining links between microbial community structure and ecosystem function. Taken together, our results indicate that a greater understanding of microbial communities informed by ecological principles may enhance our ability to predict ecosystem process rates relative to assessments based on environmental variables and microbial physiology.

MMPs are a family of Zn dependent endopeptidases, which can mediate degradation of ECM components during various physiological and pathological processes including cancer. Some ECM components, through interaction with integrin receptor and modulation of downstream signaling, are capable of regulating expression and activity of several MMPs. α₅β₁ integrin is the universally accepted receptor for the ECM component fibronectin (FN). The present review deals with the downstream signaling involved in the α₅β₁ integrin mediated modulation of expression and activity of MMPs and their effector responses in different cellular system.

AIM: 1) To study the participation of Focal Adhesion Kinase (FAK) in regulation of Breast Cancer cell migration in relation with MMP-9 and other signaling proteins. 2) To study the effect of some natural products on FAK. METHODS: Cell culture, Western Blot, Immunoprecipitation, Immunocytochemistry, Zymography, SiRNA transfection, RT-PCR, Real-Time PCR. RESULTS: For our study on FAK, we selected invasive Breast Cancer cell line MDA-MB-231 and treated the cells with Fibronectin (FN). Treatment of FN was found to increase FAK expression, phosphorylation (Tyr 397). FAK was found to be involved in re- gulation of breast cancer cell migration and MMP-9 expression, activity. Fi-bronectin increases association of FAK with integrin α5β1, Paxillin, Actin, ERK, PI3K and localization at Focal Adhesion sites. FAK was found to be involved in modulation of ERK and PI3K phosphorylation. Moreover, FAK signal was found to be transduced through ERK and PI3K, which modulate MMP-9 and thereby cell migration. CONCLUSION: FAK expression, phosphorylation and processing are induced in response to Cell-ECM interactions. Integrin α5β1 is involved in FN induced FAK phosphorylation. FAK is a potent regulator of MMP-9 expression and activity. FAK is involved in regulation of ERK and PI3K phosphorylation. ERK and PI3K are involved in FAK regulated MMP-9 expression & activity. FAK regulates MMP-9 expression and activity and thereby migration of human breast cancer cell. By the regulation of FAK, cell attachment and migration may be regulated by Curcumin, ATRA or EGCG treatment. It may be concluded that invasive potential of breast cancer cells may be modulated by regulation of FAK.

Cell adhesion to extracellular matrix (ECM) initiates signaling cascade regulated by cell surface integrin receptors, which affects the proliferation and invasion of cells. Cells cultured in the presence of ECM ligand fibronectin (FN) stimulate secretion of matrix metalloproteinases (MMPs), facilitating cancer cell invasion and metastasis. Among all the members of the MMP family, MMP-9 is of crucial importance in tumor invasion and metastasis. The present study aims at studying the effects of integrin receptor alpha5beta1 and its ligand FN on expression of MMP-9 in murine melanoma cell line B16F10 and understanding the molecular mechanism(s) involved. The main experimental methods performed in the study were gelatin zymography, immunoblot, real-time RT-PCR, immunocytochemistry, enzyme linked immunosorbent assay (ELISA), transwell chamber assay, and in vivo metastasis assay in syngenic (C57BL6J) mice. The study reports that FN induces the activity, mRNA, and protein expression of MMP-9 and initiates its proteolytic activation in B16F10 cells. Blockage of the alpha5 receptor abrogated the FN-mediated stimulatory response on MMP-9 in B16F10 cells. Inhibitor studies and immunoblot analysis strongly suggest the involvement of focal adhesion kinase (FAK), extracellular regulated kinase (ERK), and phosphatidylinositol-3-kinase (PI-3K) in the FN-mediated responses. Immunocytochemical analysis showed the nuclear localization of nuclear factor-kappaB (NF-kappaB) might lead to activation of MMP-9 gene upon FN treatment. This study demonstrates that integrin receptor alpha5beta1 and FN interaction induces the invasive potential of B16F10 cells and MMP-9 induction is the downstream effectors in the process. This system serves as a novel model system to understand the molecular mechanism of melanoma growth and invasion.

Cricket in Pakistan has always held a significant place as an emblem of national pride and has always been a reliable indicator of the nation's fluctuating bilateral relations with India. In Cricke...

Aims: To identify and estimate salivary VEGF of breast cancer patients using non invasive method. Methods: ELISA, Zymography, Immunoblot. Results: We observed that saliva of breast cancer patients express appreciably high VEGF compare to non breast cancer saliva samples. Conclusion: The findings indicate salivary VEGF as a potential breast cancer marker using non invasive method.

The 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 aimed to study the effect of ATRA on MMP-9 in MDA-MB-231, human breast cancer cells and the probable molecular mechanisms through which ATRA exerts its effect. Results: Our experimental findings demonstrate that ATRA enters into the nucleus and regulates various signaling pathways viz. Integrin, FAK, ERK, PI-3K, NF-κB and also EGFR and down regulates pro-MMP-9 activity as well as its expression. As a result MDA-MB-231 cell migration on fibronectin medium gets retarded in presence of ATRA. ATRA up regulates TIMP-1 expression. Conclusions: Our study may help to understand the role of ATRA as a regulator of MMP-9 and the possible signaling pathways which are involved in this ATRA mediated down regulation of MMP-9.

Interaction between cell surface integrin receptors and extracellular matrix (ECM) components plays an important role in cell survival, proliferation and migration including tumor development and invasion. Matrix metalloproteinases (MMP) are a family of metalloproteinases capable of digesting ECM and facilitate cell migration. Binding of ECM to integrins initiates signaling cascades modulating expression and activity of different MMPs. The present study investigates whether laminin-mediated signaling modulates matrix metalloproteinases (MMP) expression and activity in human cervical cancer cell (SiHa).Western blot, immunocytochemistry, ELISA, zymography, RT-PCR, EMSA and wound-healing assay were used.Culture of SiHa cells on laminin (LN)-coated surface induces MMP-9 expression and activation. Wound-healing assay showed that SiHa cells migrate much faster on laminin-coated surface than that of control. LN-induced MMP-9 expression and activation was appreciably reduced with treatment of extracellular signal-regulated kinase (ERK) inhibitor, phosphatidylinositol-3-kinase (PI-3K) inhibitor and anti-α2 antibody. Phosphorylation of focal adhesion kinase (FAK), ERK, and PI-3K was increased upon LN stimulation. LN induces nuclear translocation of PI-3K and nuclear factor kappa B (NF-κB). LN increases DNA-binding activity of NF-κB and activator protein-1 (AP-1) to MMP-9 promoter.Our findings indicate laminin-induced MMP-9 expression and activation possibly via α2β1 integrin-mediated signaling involving FAK, PI-3K, ERK followed by transcriptional upregulation of MMP-9.

As forests develop, changes in soil organic matter quantity and quality affect both nutrient dynamics and microbial community structure. Litter decomposition and nitrogen mineralization in association with soil microbial communities were compared between 45- and 135-year-old lodgepole pine ( Pinus contorta var. latifolia (Englem.)) stands in southeastern Wyoming, USA. Compared with the 45-year-old stand, the 135-year-old stand was found to have greater live-tree biomass, litter decomposition rates (264 versus 135 mg·(g litter)–1·year–1), soil nitrification rates (0.38 versus 0.19 µg NO3–·(g soil)–1 after 265 days of field incubation), and total phospholipid fatty acid (PLFA) concentrations (25 versus 9.2 nmol·(g soil)–1 at 0–5 cm depth). Canonical correspondence analysis indicated that variation of PLFA profiles within the 45-year-old stand was explained by soil pH and bulk density, whereas soil process rates explained the distributions of PLFA profiles within the 135-year-old stand. The results of these studies indicate that stand age influences live-tree biomass and soil properties that can lead to changes in litter decomposition rates and soil microbial communities in lodgepole pine forests.

Interaction between cell surface integrin receptors and extracellular matrix (ECM) components plays an important role in cell survival, proliferation, and migration, including tumor development and invasion of tumor cells. Matrix metalloproteinases (MMPs) are a family of metalloproteinases capable of digesting ECM components and are important molecules for cell migration. Binding of ECM to integrins initiates cascades of cell signaling events modulating expression and activity of different MMPs. The aim of this study is to investigate fibronectin-integrin-mediated signaling and modulation of MMPs. Our findings indicated that culture of human cervical cancer cell (SiHa) on fibronectin-coated surface perhaps sends signals via fibronectin-integrin-mediated signaling pathways recruiting focal adhesion kinase (FAK) extracellular signal regulated kinase (ERK), phosphatidyl inositol 3 kinase (PI-3K), integrin-linked kinase (ILK), nuclear factor-kappa B (NF-kappaB), and modulates expression and activation of mainly pro-MMP-9, and moderately pro-MMP-2 in serum-free culture medium.

Forest management practices can have a significant effect on above- and below-ground carbon (C) pools. To better understand the distribution of forest C pools, we evaluated representative forest stands within two dominant Wyoming forest types to assess differences resulting from management practices that have occurred over several decades. Study sites included four ponderosa pine ( Pinus ponderosa Douglas ex Lawson & C.Lawson) treatments (100-year-old unmanaged, 46-year-old even-aged, 110-year-old uneven-aged, and 90-year-old intensively harvested) and two lodgepole pine ( Pinus contorta Engelm. var. latifolia (Engelm. ex Wats.) Critchfield) treatments (145-year-old unmanaged and 45-year-old even-aged). Comparisons of aboveground C pools revealed that distributions of live and dead biomass C pools were different between unmanaged and managed stands; however, belowground soil C pools were similar among stands within the two forest types. Overall, unmanaged stands of both forest types had higher total ecosystem C pools (249 and 247 Mg C·ha–1for ponderosa and lodgepole pine, respectively) compared with managed stands (ponderosa pine: even-aged, 164 Mg C·ha–1; uneven-aged, 170 Mg C·ha–1; intensively harvested, 200 Mg C·ha–1; and lodgepole pine even-aged, 117 Mg C·ha–1). Our results indicate timber harvesting has a major influence on total ecosystem C pools by reducing live tree biomass.

Aims The tumor inhibiting property of green tea polyphenol epigallocatechin-3-gallate (EGCG) is well documented. Studies reveal that matrix-metalloproteinases (MMPs) play pivotal roles in tumor invasion through degradation of basement membranes and extracellular matrix (ECM). We studied the effect of EGCG on matrixmetalloproteinases-2 (MMP-2), the factors involved in activation, secretion and signaling molecules that might be involved in the regulation of MMP-2 in human breast cancer cell line, MCF-7. Main methods MCF-7 was treated with EGCG (20 μM, 24 h), the effect of EGCG on MMP-2 expression, activity and its regulatory molecules were studied by gelatin zymography, Western blot, quantitative and semi-quantitative real time RT-PCR, immunoflourescence and cell adhesion assay. Key findings EGCG treatment reduced the activity, protein expression and mRNA expression level of MMP-2. EGCG treatment reduced the expression of focal adhesion kinase (FAK), membrane type-1-matrix metalloproteinase (MT1-MMP), nuclear factor-kappa B (NF-kB), vascular endothelial growth factor (VEGF) and reduced the adhesion of MCF-7 cells to ECM, fibronectin and vitronectin. Real time RT-PCR revealed a reduced expression of integrin receptors α5, β1, αv and β3 due to EGCG treatment. Significance Down regulation of expression of MT1-MMP, NF-kB, VEGF and disruption of functional status of integrin receptors may indicate decreased MMP-2 activation; low levels of FAK expression might indicate disruption in FAK-induced MMP-2 secretion and decrease in activation of phosphatidyl-inositol-3-kinase (PI-3K), extracellular regulated kinase (ERK) indicates probable hindrance in MMP-2 regulation and induction. We propose EGCG as potential inhibitor of expression and activity of pro-MMP-2 by a process involving multiple regulatory molecules in MCF-7.

Timber harvesting influences both above and belowground ecosystem nutrient dynamics. Impact of timber harvesting on soil organic matter (SOM) mineralization and microbial community structure was evaluated in two coniferous forest species, ponderosa pine (Pinus ponderosa) and lodgepole pine (Pinus contorta). Management of ponderosa pine forests, particularly even-aged stand practices, increased the loss of CO2-C and hence reduced SOM storage potential. Changes in soil microbial community structure were more pronounced in ponderosa pine uneven-aged and heavy harvest stands and in lodgepole pine even-aged stand as compared to their respective unmanaged stands. Harvesting of trees had a negative impact on SOM mineralization and soil microbial community structure in both coniferous forests, potentially reducing coniferous forest C storage potential.

Purpose: Several studies indicate that integrin receptors are involved in the regulation of matrix metalloproteinase (MMP) expression. Integrin–ECM ligand interaction leads to phosphorylation of focal adhesion kinase (FAK) and activation of mitogen activated protein kinase pathways. In this present communication, we cultured human cervical cancer cells, SiHa, in the presence of fibronectin to study fibronectin–integrin mediated modulation of MMP activity. Methods: SiHa cells were cultured in serum-free medium (SFCM) in the presence of fibronectin, SFCM was collected and gelatin zymography was performed. Western blot, RT-PCR and immunocytochemistry were performed with SiHa cells cultured in the presence of fibronectin. Results: The culture of SiHa cells in the presence of 50 μg/1.5 ml fibronectin led to expression of pro-MMP-9 and activation of MMP-2 within 2 h. When cells were treated with ERK inhibitor (PD98059) and grown in the presence of fibronectin MMP-2 activation was partially inhibited, but when cells were treated with PI-3K inhibitor (LY294002) and grown in the presence of fibronectin MMP-2 activation was appreciably reduced. Tyrosine phosphorylation of FAK, PI-3K and ERK and nuclear trafficking of ERK were increased in SiHa cells grown in the presence of fibronectin. Increased MT1-MMP mRNA expression and processing of MT1-MMP were also observed in SiHa cells grown in the presence of fibronectin. Conclusions: Our findings indicate that the culture of SiHa cells in SFCM in the presence of fibronectin perhaps generates a signalling cascade which leads to the expression of pro-MMP-9 and the activation of MMP-2 within 2 h. The signalling pathways activated seem to be the FAK/ERK/PI-3K pathway.

UNLABELLED Among the soluble MMPs, MMP-2 (gelatinase A) is particularly important in the invasive property of tumor cells. Cell membrane-associated MMP-2 activation is one of the challenging areas in tumor biology. In the present communication, we studied the membrane dependent activation of MMP-2 in SiHa cells. METHODS Activation of pro-MMP-2 by membrane fraction, membrane extract, and live SiHa cells was studied by gelatin zymography. The role of MT1-MMP in MMP-2 activation was studied by incubating SiHa cells and cell membrane fractions with anti-MT1-MMP antibody. RESULTS Activation of purified pro-MMP-2 by membrane fraction isolated from SiHa cells, by SiHa cell membrane extract and by SiHa cells, pro-MMP-2 from Con A treated HT-1080 conditioned medium by SiHa cells, and pro-MMP-2 from serum free culture medium of SiHa cells and cervical tissue homogenate by SiHa cell membrane fraction was shown by gelatin zymography. SiHa membrane fraction activated only pro-MMP-2 from purified MMP-9/MMP-2 mixture, indicating that the activation is specific for MMP-2. Inhibition of MMP-2 activation in the presence of anti-MT1-MMP antibody strongly indicated that the cell membrane mediated MMP-2 activation is MT1-MMP dependent. Immunocytochemistry of SiHa cells demonstrated expression of MT1-MMP at focal points. Invasion assay showed that invasiveness of anti-MT1-MMP antibody treated SiHa cells through Matrigel was drastically reduced compared to control SiHa cells. CONCLUSIONS Our findings furnish an example of the cell membrane-associated MT1-MMP mediated MMP-2 activation in SiHa cells and suggest that this MT1-MMP mediated MMP-2 activation is of importance in tumor invasion and metastasis. This MT1-MMP mediated MMP-2 activation on tumor cell surface could be a realistic target for managing metastatic diseases.

UNLABELLED Curcumin (diferuloylmethane) has been widely studied for its tumor inhibiting and anticarcino-genic properties. In the present communication, we studied the effect of curcumin on matrix-metalloproteinase-2 (MMP-2), integrin receptors, and focal adhesion kinase (FAK) in human laryngeal cancer cells, HEp2. METHODS HEp2 cells were treated with curcumin (5 microM) for 30 days and then grown without curcumin for 28 days. Effect of curcumin on MMP-2 expression and activity and on membrane type matrixmetalloproteinase-1 (MT1-MMP), FAK, and integrin receptors was studied by zymography, Western blot, ELISA, RT-PCR, and cell adhesion assay. RESULTS Treatment of HEp2 cells with curcumin downregulated MMP-2 expression and activity and expression of integrin receptors, FAK, and MT1-MMP to almost background levels. MMP-2 (but not MMP-9) mRNA expression was abolished on curcumin treatment, indicating specific inhibition of MMP-2. Invasive potential of HEp2 cells was also significantly reduced. After drug withdrawal, expression of MMP-2, integrin receptors, MT1-MMP, and FAK returned to control levels. However, MMP-2 activity in serum free medium remained low. CONCLUSIONS Downregulation of integrin receptors and low levels of FAK may hinder integrin-mediated signal transduction, preventing upregulation of MMP-2 activity. Reduction of MMP-2 activity and inhibition of HEp2 cell invasion by curcumin strongly indicate the potential of curcumin as an inhibitor of tumor cell invasion and metastasis.

UNLABELLED Matrix metalloproteinases (MMPs), a family of zinc-dependent endopeptidases, can degrade extracellular matrix components under physiological conditions and during cancer invasion and metastasis. Among the MMPs, the 72 kDa type IV collagenase MMP-2 (gelatinase A) is activated in a membrane-associated manner by an activation complex composed of membrane type 1 matrix metalloproteinase (MT1-MMP), tissue inhibitor of matrixmetalloproteinase-2 (TIMP-2), and pro-MMP-2 in the presence of alphavbeta3 integrin receptor. The activation of pro-MMP-2 correlates with increased occurrence of metastases. Increased MMP-2 activity has been demonstrated in many human tumors. In the present communication, we studied cell surface-associated activation of MMP-2 (72 kDa collagenase type IV) in the moderately metastatic human melanoma cell line A375. RESULTS Activation of purified 72 kDa collagenase type IV, pro-MMP-2 from cervical cancer tissue homogenate and from serum-free culture medium of HT1080 cells grown in presence of concanavalin A, by A375 cells, was shown by gelatin zymography. A375 cells activated only pro-MMP-2 from purified MMP-9/MMP-2 mixture indicating that the activation is specific for MMP-2. Activation of MMP-2 and purified collagenase type IV by A375 membrane fraction and membrane extract was also demonstrated by gelatin zymography. When A375 cells were first incubated with anti-MT1-MMP polyclonal antibody, activation of collagenase type IV was significantly decreased, indicating that membrane-associated MMP-2 activation is MT1-MMP-mediated. Immunocytochemistry showed MT1-MMP localization at focal adhesion sites. The presence of the components of activation complex-MT1-MMP and integrin alphavbeta3-were confirmed by Western blot, cell adhesion assay, and integrin subunit assay. CONCLUSION Our experimental findings furnish another example of the unique membrane-associated MMP-2 activation mechanism in A375 melanoma cells and clearly indicate the role of MT1-MMP in MMP-2 activation. The information could help in developing new therapies designed to interfere with MMP activation and management of cancer and metastases.

Objective(s) . The purpose was to study the interrelationship between cell surface integrin receptor (α2β1) and matrixmetalloproteinases. Methods . Immunoprecipitation and cell adhesion assay were done to assay α2β1 and α3β1 on SiHa cell surface. Zymogram was developed to assay secreted MMP activity of cells grown in presence of α2 monoclonal antibody. Immunoblot was developed to assay expression of MMP-2, FAK, and p-FAK. Plasma membrane-dependent activation of MMP2 was performed by incubating pure MMP-2 with membrane-enriched fraction isolated from SiHa cells. Results . Immunoprecipitation and cell adhesion assay results confirmed the presence of α2β1 receptor on SiHa cells. Zymographic analysis of serum-free media collected at different time points from SiHa cells grown on α2 monoclonal antibody-coated culture dishes showed the expression and activation of MMP-2 within 2–4 h, confirmed by immunoblot. Western blot of cells grown on α2-coated dishes for 30 min–4 h showed increased phosphorylation of FAK. Membrane-enriched fraction isolated from SiHa cells was found to specifically activate proMMP-2 to its activated forms within 30 min. Conclusion(s) . The experimental findings strongly indicate that SiHa cell surface α2β1 regulates MMP-2 expression. Increased phosphorylation of focal adhesion kinase (FAK) strongly indicates the possible role of FAK in signaling cascade. Incubation of SiHa cell membrane fraction with pure MMP-2 strongly confirms the cell membrane-dependent activation of proMMP2.

Curcumin (diferuloyl methane), the major pigment from the rhizome of Curcuma longa L., has been widely studied for its tumor-inhibiting properties. Recent studies indicate that curcumin can modify cell receptor binding, it also affects intracellular signalling reactions. Curcumin-treated B16F10 melanoma cells formed eight-fold fewer lung metastases in C57BL6 mice. In the cell adhesion assays, curcumin-treated cells showed a dose-dependent reduction in their binding to four extracellular matrix (ECM) proteins. The binding to fibronectin, vitronectin, and collagen IV decreased by over 50% in 24 hours, and by 100% after 48 hours of curcumin treatment, it persisted at this level even after 15 days of cultivating cells in curcumin-free medium. Curcumin-treated cells showed a marked reduction in the expression of alpha5beta1 and alpha(v)beta3 integrin receptors. In addition, curcumin treatment inhibited pp125 focal adhesion kinase (FAK), tyrosine phosphorylation of a 120 kD protein, and collagenase activity. Curcumin enhances the expression of antimetastatic proteins, tissue inhibitor metalloproteinase (TIMP)-2, nonmetastatic gene 23 (Nm23), and E-cadherin. In this article we report on the effect of curcumin on the expression of integrin, TIMP-2, Nm23, E-cadherin, adhesion, and metalloproteinase activity.

Integrins are transmembrane heterodimeric molecules that mediate cellular adhesion and are involved in different biological processes, such as tumor development and invasion of tumor cells. Matrixmetalloproteases (MMP) are a family of secreted or membrane proteins capable of digesting extracellular matrix. It has been shown that MMP-2 binds to alphavbeta3 integrin. Recent evidence suggests that a complex of membrane-type MMP (MT1-MMP) and tissue inhibitor of metalloptroteinase-2 (TIMP-2) participate in the activation of alphavbeta3-associated MMP-2. We investigated whether alphavbeta3 and MMP-2 are associated on the membranes of a human cell line, SiHa, and the possible involvement of MT1-MMP and TIMP-2 in the modulation of MMP-2 activity.Immunoprecipitation of SiHa membrane extracts with monoclonal antibodies against alphav or MMP-2, and western blots of immunoprecipitates and serum-free conditioned media were performed. TIMP-2 in conditioned medium and MT1-MMP in the membrane fraction was assayed by western blot. Zymography of anti-alphav antibody immunoprecipitates and conditioned media were used to show gelatinolytic activity.The coprecipitation of MMP-2 with alphavbeta3 by anti-alphav antibody is a strong indication that SiHa cell surface alphavbeta3 integrin is a receptor for MMP-2. Immunoblot assays show the expression of MT1-MMP on SiHa cell membranes and secreted TIMP-2 and pro-MMP-2 in the medium.SiHa cells express all the molecules which are reported to form a complex to activate pro-MMP-2. Active MMP-2 associated with alphavbeta3 may regulate matrix degradation and thereby modulate directed motility of SiHa cells.

Studies on interaction of tumor cells with extracellular matrix (ECM) components showed increased extracellular protease activity mediated by the family of matrix metalloproteinases (MMPs). Here we studied the effect of human breast cancer cell line MCF-7-laminin (LM) interaction on MMPs and the underlying signaling pathways. Culturing of MCF-7 cells on LM coated surface upregulated MMP-9 expression as well as reduced tissue inhibitor of metalloproteinases-1 (TIMP-1) expression. LM induced MMP-9 expression is abrogated by the blockade of α2 integrin. Inhibitor studies indicate possible involvement of phosphatidyl-inositol-3-kinase (PI3K), extracellular signal regulated kinase (ERK) and nuclear factor-kappaB (NF-κB) in LM induced signaling. LM treatment also enhanced phosphorylation of FAK (focal adhesion kinase), PI3K, ERK; nuclear translocation of ERK, pERK, NF-κB and cell migration. Our findings indicate that, binding of MCF-7 cells to LM, possibly via α2β1 integrin, induces signaling involving FAK, PI3K, ERK, NF-κB followed by upregulation of MMP-9 and cell migration.

Cell surface receptors have been the subject of intensive investigations over the past few decades. One very important group of receptors on the cell surface is the "integrin" receptors which bind to extracellular matrix (ECM) proteins. Because of integrin's importance in cellular growth, development, and morphology the role of integrin receptors in cellular transformation, malignant growth, and metastasis has received wide attention. In this article we report on the effect of all-trans-retinoic acid (ATRA) on (a) the integrin family of cell surface receptors, (b) collagenase enzyme activity, and (c) invasive potential in human cervical cancer (SiHa) cells. A comparative cell adhesion assay clearly showed that ATRA affects the cell surface integrin receptors against different ECM proteins in a dose- and time-dependent manner. The binding of SiHa cells to ECM proteins (fibronectin, vitronectin, laminin, collagen IV) was drastically reduced when cells were treated with ATRA at 10 microM for 96 h in culture. Interestingly, when ATRA-treated (10 microM, 96 h) SiHa cells were allowed to grow for 15 days in ATRA-free complete medium the binding of SiHa cells to fibronectin and vitronectin was inhibited, even after 15 days of drug withdrawal, whereas cell adhesion to laminin and collagen IV returned to normal within 3-7 days. The comparative immunoprecipitation of two cell surface integrin receptors (alpha5beta1 and alphavbeta3) shows the effect of ATRA on the expression of alpha5, alphav, and beta1 subunits. In ATRA-treated SiHa cells the cell surface expression of the alphav subunit (in alphavbeta3 receptor) is much less than in untreated SiHa cells. In the case of the alpha5beta1 integrin receptor ATRA treatment caused a significant reduction in the expression of both alpha5 and beta1 subunits on the cell surface. Comparative zymography clearly demonstrated the inhibitory effect of ATRA on collagenase enzyme activity. Interestingly, the effect was irreversible, even after 15 days of culture in ATRA-free medium. The assay of the invasive potential of ATRA-treated and untreated SiHa cells in Boyden's invasion chamber demonstrated that ATRA treatment (10 microM, 96 h) inhibits the invasive potential of SiHa cells. The effect was not reversible even after 15 days of culture in ATRA-free medium. In conclusion, our observations indicate that ATRA has an inhibitory effect on the expression of SiHa cell surface integrin receptors and collagenase enzyme activity. The effect of ATRA on cell surface integrin receptors and collagenase enzyme activity may affect the invasive potential of SiHa cells.

Metastasis is a combination of biological events that makes the difference between cancer and other diseases. Metastasis requires flow of erroneous but precisely coordinated basic cellular activities like cell migration–invasion, cell survival–apoptosis, cell proliferation, etc. All of these processes require efficient regulation of cell attachment and detachment, which recruit integrin receptors in this flow of events. World literatures show several aspects of interrelation of integrins and metastasis. Integrin molecules are being used as prime target to battle metastasis. In this review we are collating the observations showing importance of integrin biology in regulation of metastasis and the strategies where integrin receptors are being used as targets to regulate metastasis.

Epidermal growth factor receptor (EGFR/ErbB1) is a transmembrane protein with tyrosine kinase activity activated mainly by ligand, EGF. Matrix metalloproteinases (MMPs) are a family of proteinases that catalyses the destruction of ECM, among which MMP-9 has important role in tumor cell invasion. Secretion of MMP-9 is stimulated by a variety of factors, EGFR being significant. Epigallocatechin-3-gallate (EGCG) is a major polyphenol of green tea that inhibits cell proliferation and invasion. Here, we study the effect of EGFR alone and in collaboration with fibronectin on the status of MMP-9 in human breast cancer cell MDA-MB-231 and its molecular mechanism; study the role of EGCG on the induced MMP-9; and elucidate the signaling molecules involved in the process.We performed zymography, immunoblots, real-time RT-PCR, cell adhesion assay, siRNA studies, and electrophoretic mobility shift assay to demonstrate the findings.EGF induces MMP-9 activity and expression; FAK, PI3 K, and ERK are mainly involved in the process. EGF also causes the transactivation of MMP-9 gene by increasing the DNA binding activity of the transcription factors. EGCG downregulates EGF-induced MMP-9 expression by inhibiting the involved regulatory kinases. EGF collaborates with fibronectin to create a synergistic response, and EGCG inhibits the synergistic response in MDA-MB-231.The study demonstrates the requirement of cross talk between cell matrix adhesion molecules and growth factor receptors to improve biological responses and shows FAK/ERK as the pivotal point of this convergence in human breast carcinoma cell line MDA-MB-231. We also establish EGCG as the potential anti-tumor agent in human breast carcinoma.

The tumor-inhibiting property of black tea polyphenol, theaflavin, is well documented. Matrix metalloproteinases (MMPs) play a pivotal role in tumor invasion through degradation of extracellular matrix (ECM). In the present study, we observed the effect of theaflavin on MMP-2, which is upregulated in most tumor types, and its regulatory molecules, in human melanoma cell line, A375. The treatment of theaflavin downregulated the gelatinolytic activity, mRNA and protein expression of MMP-2. It reduced the mRNA and protein expression of membrane type-1 MMP (MT1-MMP) and induced mRNA and protein expression of tissue inhibitor of MMP-2 (TIMP-2), suggesting theaflavin's inhibitory effect on MMP-2 activation. Theaflavin reduced the binding of A375 cell to ECM ligands demonstrating that theaflavin treatment hinders cell-ECM adhesion, cell motility, and integrin-mediated MMP-2 activation. Theaflavin treatment inhibited the protein expression FAK EGFR and ERK, suggesting that, theaflavin treatment downregulates the molecules participating in MMP-2 secretion and regulation. The downregulation of NFchiB suggests downregulation of MMP-2 transactivation. Theaflavin also reduced the tumor volume in syngenic black mice. Thus, we report that theaflavin causes an inhibition of the expression and activity of pro-MMP-2 by a process involving multiple regulatory molecules in human melanoma cells, A375.

Epigallocatechin-3-gallate (EGCG) is effective against the initiation, progression, and invasion of carcinogenesis.Matrix-metalloproteinases (MMPs) are a family of endopeptidases that hydrolyze the majority of extracellular proteins. MMP-9 is one of the most important members of the family and we observed the effect of EGCG on MMP-9 in the human breast cancer cell line, MDA-MB-231.The effect of EGCG on MMP-9 was studied by gelatin zymography, western blot, quantitative and semiquantitative real-time RT-PCR, immunoflourescence, cell adhesion assay, enzyme-linked immunosorbent assay,and electrophoretic mobility shift assay. EGCG treatment reduced the activity, protein, and mRNA expression ofMMP-9 and enhanced the expression of the tissue inhibitor of MMP 1 (TIMP-1). EGCG downregulated the activation of focal adhesion kinase (FAK) and extracellular regulated kinase (ERK), reduced the adhesion of MDA-MB-231 cells to fibronectin and vitronectin, and reduced the mRNA expression of the integrin receptors alpha5beta1 and alphavbeta3. The expression of the nuclear factor kappa B (NFjB), and the DNA binding activity of NFjB and activator protein 1 (AP1)to MMP-9 promoter were noticeably reduced on EGCG treatment. Upregulation of TIMP-1 and disruption of the functional status of integrin receptors may indicate decreased MMP-9 activation; inhibition of FAK andERK activation might indicate disruption in the FAK/ERK-induced MMP-9 secretion and induction. Decreased DNA binding activity of NFjB and AP1 to MMP-9 promoter might indicate transcriptional deregulation of MMP-9 gene on EGCG treatment. We propose EGCG as a potential inhibitor of the expression and activity of MMP-9 by a process involving FAK/ERK and transcription factorsin MDA-MB-231.

Interaction between cell surface integrin receptors with extracellular matrix (ECM) plays an important role in cell survival, proliferation, and migration including tumor development and invasion. Binding of ECM to integrins initiates intracellular signaling cascades, modulating expression and activity of different matrix metalloproteinases (MMPs) which is important in ECM degradation. The present study investigates fibronectin–integrin-mediated signaling and thereby modulation of MMPs expression and activity in human breast cancer cell line, MDA-MB-231. Culture of MDA-MB-231 cells on fibronectin (FN) induced expression and activity of pro-matrixmetalloproteinase-9 (MMP-9). Appreciable reduction of FN-induced pro-MMP-9 activity was observed in anti-α5 antibody treated cells. Inhibitor studies revealed that inhibitors of phosphatidyl inositiol-3-kinase (PI-3K), and nuclear factor kappa B (NF-κB) inhibited FN-induced pro-MMP-9 activity. FN increased tyrosine phosphorylation of focal adhesion kinase (FAK), integrin linked kinase (ILK), and PI-3K in MDA-MB-231 cells. FN-induced the transactivation of MMP-9 promoter by enhancing DNA binding activity of NF-κB and Sp1. Wound healing assay showed faster migration of MDA-MB-231cells grown on fibronectin-coated as surface as compared to control. Our findings indicated that culture of MDA-MB-231 on fibronectin perhaps send signals via fibronectin–integrin-mediated signaling pathways recruiting FAK, PI-3K, ILK, NF-κB, and modulate expression and activation of pro-MMP-9. These observations may enrich fundamental aspects of cancer biology especially role of α5β1 integrin in regulation of MMPs expression and activity.

Cell adhesion to extracellular matrix initiates intracellular signaling cascade regulated by integrin family of receptors. Evidences show that cultured cells in presence of extracellular matrix adhesion molecule Fibronectin (FN) stimulates secretion of matrix metalloproteinases (MMPs), facilitating cancer cell invasion. Amongst all MMPs, MMP-9 is often reported to play crucial role in tumor cell growth and metastasis. The present study aims at examining the effects of FN on MMP-9 in laryngeal carcinoma cell line, HEp-2, and understand the molecular mechanism(s) involved. The study reports that FN induces the gelatinolytic activity, mRNA and protein expression of MMP-9 in HEp-2 cells. This effect appears to be mediated mainly by integrin receptor α5β1, since, the blockade of α5 abrogated the FN-mediated stimulatory response on MMP-9. siRNA and inhibitor studies suggested involvement of Focal adhesion kinase (FAK), Phosphatidyl-inositol-3-kinase (PI-3K), Extracellular regulated kinase (ERK) and nuclear factor-kappaB (NFκB) in FN-mediated MMP-9 induction. Immunocytochemical analysis demonstrated the nuclear localization of ERK, PI-3K and NFκB; immunoblot showed enhanced expression of p-FAK, p-PI-3K, p-ERK and nuclear-NF-κB and indicated involvement of ILK in the FN-mediated response. FN-induced transactivation of MMP-9 gene by enhanced DNA binding activity of transcription factors NFκB, Activator protein-1 (AP-1) and Specificity protein-1 (Sp1) to the MMP-9 promoter. Thus, this study suggests that extracellular matrix protein FN induces MMP-9 in HEp-2 cells mainly by involving integrin receptor α5β1 and involves activation of multiple signaling pathways which independently or in "cross-talk" to each other finally leads to the transactivation of the MMP-9 gene.

Interactions between tumour cells and the extracellular matrix (ECM) strongly influence tumour development, affecting cell survival, proliferation and migration. Many of these interactions are mediated through a family of cell surface receptors named integrins. Fibronectin and its integrin receptors play important roles in tumour development. The alpha5beta 1 integrin interacts with the central cell adhesive region of fibronectin and requires both the RGD and synergy sites for maximal binding. Matrix metalloproteinases (MMPs) are a family of zinc dependent endopeptidases. They are capable of digesting the different components of the ECM and basement membrane. The ECM gives structural support to cells and plays a central role in cell adhesion, differentiation, proliferation and migration. Binding of ECM to integrins modulates expression and activity of the different MMPs. Our experimental findings demonstrate that cultivation of human breast cancer cells, MCF-7, in serum free medium in the presence of fibronectin upregulates the activity of MMP-2 and MMP-9. Blocking of alpha5beta 1 integrin with anti-alpha5 monoclonal antibody inhibits the fibronectin-induced MMP activation response appreciably. This strongly indicates alpha5beta 1 mediated signalling events in activation of MMP-2 and MMP-9. Phosphorylation of FAK and PI-3 kinase and the nuclear translocation of ERK and NF-kappaB upon fibronectin binding demonstrate possible participation of the FAK/PI-3K/ERK signalling pathways in the regulation of MMP-2 activity.

UNLABELLED The importance of tumor cell surface integrin receptors in regulation of matrix metalloproteinase (MMP) expression and function has been reported. Integrin-ECM ligand interaction leads to phosphorylation of focal adhesion kinase (FAK) and activation of mitogen activated protein kinase (MAPK) pathways. In this present study, we cultured human fibrosarcoma cells, HT-1080, in presence of fibronectin to study fibronectin-integrin mediated modulation of MMP activity. METHODS HT-1080 cells were cultured in serum free medium (SFCM) in presence of fibronectin, SFCM was collected, and gelatin zymography was performed. Western blot and immunocytochemistry were performed with HT-1080 cells cultured in presence of fibronectin. RESULTS Culture of HT-1080 cells in presence of 50 microg/1.5 ml fibronectin led to expression of pro-MMP-9 and activation of MMP-2 within 1 hr. When HT-1080 cells were treated with PI-3K inhibitor (LY294002) and grown in presence of fibronectin, MMP-2 activation was partially inhibited, but when cells were treated with ERK inhibitor (PD98059) and grown in presence of fibronectin, MMP-2 activation was almost completely inhibited. Tyrosine phosphorylation of FAK and ERK were increased in HT-1080 cells grown in presence of fibronectin. Processing of MT1-MMP was also observed in HT-1080 cells grown in presence of fibronectin. The reorganization of actin filaments in fibronectin treated HT-1080 cells was also noticeable. CONCLUSIONS Our findings indicate that culture of HT-1080 cells in SFCM in presence of fibronectin perhaps generates a signaling cascade that leads to expression of pro-MMP-9 and activation of MMP-2 within 1 hr. The signaling pathway activated seems to be the FAK/ERK pathway.