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Terms modified by ECM Selected AbstractsAbsence of tissue inhibitor of metalloproteinases 3 disrupts alveologenesis in the mouseDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 1 2009Sean E. Gill Tissue inhibitors of metalloproteinases (TIMPs) regulate extracellular matrix (ECM) degradation by matrix metalloproteinases (MMPs) throughout lung development. We examined lungs from TIMP3 null mice and found significant air space enlargement compared with wild type (WT) animals during a time course spanning early alveologenesis (post-partum days 1, 5, 9 and 14). Trichrome staining revealed a similar pattern of collagen distribution in the walls of nascent alveoli; however, the alveolar walls of TIMP3 mutant mice appeared to be thinner than controls. Assessment of MMP2 and MMP9 activities by gelatin zymography demonstrated a significant elevation in the active form of MMP2 at post-partum days 1 and 5. Treatment of null pregnant dams with a broad spectrum synthetic metalloproteinase inhibitor, GM6001, on embryonic day 16.5 enhanced the formation of primitive alveoli during the saccular stage of lung development as evidenced by a partial, but significant, rescue of alveolar size in post-partum day 1 animals. We propose that increased MMP activity in the absence of TIMP3 enhances ECM proteolysis, upsetting proper formation of primitive alveolar septa during the saccular stage of alveologenesis. Therefore, TIMP3 indirectly regulates alveolar formation in the mouse. To our knowledge, ours is the first study to demonstrate that in utero manipulation of the TIMP/MMP proteolytic axis, to specifically inhibit proteolysis, significantly affects lung development. [source] Commitment and response to inductive signals of primary mesenchyme cells of the sea urchin embryoDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 1 2004Masato Kiyomoto In the sea urchin embryo, primary mesenchyme cells (PMC) are committed to produce the larval skeleton, although their behavior and skeleton production are influenced by signals from the embryonic environment. Results from our recent studies showed that perturbation of skeleton development, by interfering with ectoderm,extracellular matrix (ECM) interactions, is linked to a reduction in the gene expression of a transforming growth factor (TGF)-beta growth factor, Pl-univin, suggesting a reduction in the blastocoelic amounts of the protein and its putative involvement in signaling events. In the present study, we examined PMC competence to respond to environmental signals in a validated skeleton perturbation model in Paracentrotus lividus. We found that injection of blastocoelic fluid (BcF), obtained from normal embryos, into the blastocoelic cavity of skeleton-defective embryos rescues skeleton development. In addition, PMC from skeleton-defective embryos transplanted into normal or PMC-less blastula embryos are able to position in correct regions of the blastocoel and to engage spicule elongation and patterning. Taken together, these results demonstrate that PMC commitment to direct skeletogenesis is maintained in skeleton perturbed embryos and confirm the role played by inductive signals in regulating skeleton growth and shape. [source] Presynaptic secretion of mind-the-gap organizes the synaptic extracellular matrix-integrin interface and postsynaptic environmentsDEVELOPMENTAL DYNAMICS, Issue 3 2009Emma Rushton Abstract Mind-the-Gap (MTG) is required during synaptogenesis of the Drosophila glutamatergic neuromuscular junction (NMJ) to organize the postsynaptic domain. Here, we generate MTG::GFP transgenic animals to demonstrate MTG is synaptically targeted, secreted, and localized to punctate domains in the synaptic extracellular matrix (ECM). Drosophila NMJs form specialized ECM carbohydrate domains, with carbohydrate moieties and integrin ECM receptors occupying overlapping territories. Presynaptically secreted MTG recruits and reorganizes secreted carbohydrates, and acts to recruit synaptic integrins and ECM glycans. Transgenic MTG::GFP expression rescues hatching, movement, and synaptogenic defects in embryonic-lethal mtg null mutants. Targeted neuronal MTG expression rescues mutant synaptogenesis defects, and increases rescue of adult viability, supporting an essential neuronal function. These results indicate that presynaptically secreted MTG regulates the ECM-integrin interface, and drives an inductive mechanism for the functional differentiation of the postsynaptic domain of glutamatergic synapses. We suggest that MTG pioneers a novel protein family involved in ECM-dependent synaptic differentiation. Developmental Dynamics 238:554,571, 2009. © 2009 Wiley-Liss, Inc. [source] Expression profiles of the duplicated matrix metalloproteinase-9 genes suggest their different roles in apoptosis of larval intestinal epithelial cells during Xenopus laevis metamorphosisDEVELOPMENTAL DYNAMICS, Issue 8 2007Takashi Hasebe Abstract Matrix metalloproteinases (MMPs) play a pivotal role in development and/or pathogenesis through degrading extracellular matrix (ECM) components. We have previously shown that Xenopus MMP-9 gene is duplicated. To assess possible roles of MMP-9 and MMP-9TH in X. laevis intestinal remodeling, we here analyzed their expression profiles by in situ hybridization and show that their expression is transiently up-regulated during thyroid hormone-dependent metamorphosis. Of interest, MMP-9TH mRNA is strictly localized in the connective tissue and most highly expressed just beneath the larval epithelium that begins to undergo apoptosis. On the other hand, cells expressing MMP-9 mRNA become first detectable in the connective tissue and then, after the start of epithelial apoptosis, also in the larval epithelium. These results strongly suggest that MMP-9TH is responsible in the larval epithelial apoptosis through degrading ECM components in the basal lamina, whereas MMP-9 is involved in the removal of dying epithelial cells during amphibian intestinal remodeling. Developmental Dynamics 236:2338,2345, 2007. © 2007 Wiley-Liss, Inc. [source] ROCK inhibitor (Y27632) increases apoptosis and disrupts the actin cortical mat in embryonic avian corneal epitheliumDEVELOPMENTAL DYNAMICS, Issue 3 2004Kathy K.H. Svoboda Abstract The embryonic chicken corneal epithelium is a unique tissue that has been used as an in vitro epithelial sheet organ culture model for over 30 years (Hay and Revel [1969] Fine structure of the developing Avian cornea. Basel, Switzerland: S. Karger A.G.). This tissue was used to establish that epithelial cells could produce extracellular matrix (ECM) proteins such as collagen and proteoglycans (Dodson and Hay [1971] Exp Cell Res 65:215,220; Meier and Hay [1973] Dev Biol 35:318,331; Linsenmayer et al. [1977] Proc Natl Acad Sci U S A 74:39,43; Hendrix et al. [1982] Invest Ophthalmol Vis Sci 22:359,375). This historic model was also used to establish that ECM proteins could stimulate actin reorganization and increase collagen synthesis (Sugrue and Hay [1981] J Cell Biol 91:45,54; Sugrue and Hay [1982] Dev Biol 92:97,106; Sugrue and Hay [1986] J Cell Biol 102:1907,1916). Our laboratory has used the model to establish the signal transduction pathways involved in ECM-stimulated actin reorganization (Svoboda et al. [1999] Anat Rec 254:348,359; Chu et al. [2000] Invest Ophthalmol Vis Sci 41:3374,3382; Reenstra et al. [2002] Invest Ophthalmol Vis Sci 43:3181,3189). The goal of the current study was to investigate the role of ECM in epithelial cell survival and the role of Rho-associated kinase (p160 ROCK, ROCK-1, ROCK-2, referred to as ROCK), in ECM and lysophosphatidic acid (LPA) -mediated actin reorganization. Whole sheets of avian embryonic corneal epithelium were cultured in the presence of the ROCK inhibitor, Y27632 at 0, 0.03, 0.3, 3, or 10 ,M before stimulating the cells with either collagen (COL) or LPA. Apoptosis was assessed by Caspase-3 activity assays and visualized with annexin V binding. The ROCK inhibitor blocked actin cortical mat reformation and disrupted the basal cell lateral membranes in a dose-dependent manner and increased the apoptosis marker annexin V. In addition, an in vitro caspase-3 activity assay was used to determine that caspase-3 activity was higher in epithelia treated with 10 ,M Y-27632 than in those isolated without the basal lamina or epithelia stimulated with fibronectin, COL, or LPA. In conclusion, ECM molecules decreased apoptosis markers and inhibiting the ROCK pathway blocked ECM stimulated actin cortical mat reformation and increased apoptosis in embryonic corneal epithelial cells. Developmental Dynamics 229:579,590, 2004. © 2004 Wiley-Liss, Inc. [source] Expression patterns of focal adhesion associated proteins in the developing retinaDEVELOPMENTAL DYNAMICS, Issue 4 2002Ming Li Abstract Adhesive interactions between integrin receptors and the extracellular matrix (ECM) are intimately involved in regulating development of a variety of tissues within the organism. In the present study, we have investigated the relationships between ,1 integrin receptors and focal adhesion associated proteins during eye development. We used specific antibodies to examine the distribution of ,1 integrin ECM receptors and the cytoplasmic focal adhesion associated proteins, talin, vinculin, and paxillin in the developing Xenopus retina. Immunoblot analysis confirmed antibody specificity and indicated that ,1 integrins, talin, vinculin, and paxillin were expressed in developing retina and in the retinal-derived Xenopus XR1 glial cell line. Triple-labeling immunocytochemistry revealed that talin, vinculin, paxillin, and phosphotyrosine proteins colocalized with ,1 integrins at focal adhesions located at the termini of F-actin filaments in XR1 cells. In the retina, these focal adhesion proteins exhibited developmentally regulated expression patterns during eye morphogenesis. In the embryonic retina, immunoreactivities for focal adhesion proteins were expressed in neuroepithelial cells, and immunoreactivity was especially strong at the interface between the optic vesicle and overlying ectoderm. At later stages, these proteins were expressed throughout all retinal layers with higher levels of expression observed in the plexiform layers, optic fiber layer, and in the region of the inner and outer limiting membrane. Strong immunoreactivities for ,1 integrin, paxillin, and phosphotyrosine were expressed in the radially oriented Müller glial cells at later stages of development. These results suggest that focal adhesion-associated proteins are involved in integrin-mediated adhesion and signaling and are likely to be essential in regulating retinal morphogenesis. © 2002 Wiley-Liss, Inc. [source] Laminin and fibronectin modulate inner ear spiral ganglion neurite outgrowth in an in vitro alternate choice assayDEVELOPMENTAL NEUROBIOLOGY, Issue 13 2007Amaretta R. Evans Abstract Extracellular matrix (ECM) molecules have been shown to function as cues for neurite guidance in various populations of neurons. Here we show that laminin (LN) and fibronectin (FN) presented in stripe micro-patterns can provide guidance cues to neonatal (P5) inner ear spiral ganglion (SG) neurites. The response to both ECM molecules was dose-dependent. In a LN versus poly- L -lysine (PLL) assay, neurites were more often observed on PLL at low coating concentrations (5 and 10 ,g/mL), while they were more often on LN at a high concentration (80 ,g/mL). In a FN versus PLL assay, neurites were more often on PLL than on FN stripes at high coating concentrations (40 and 80 ,g/mL). In a direct competition between LN and FN, neurites were observed on LN significantly more often than on FN at both 10 and 40 ,g/mL. The data suggest a preference by SG neurites for LN at high concentrations, as well as avoidance of both LN at low and FN at high concentrations. The results also support a potential model for neurite guidance in the developing inner ear in vivo. LN, in the SG and osseus spiral lamina may promote SG dendrite growth toward the organ of Corti. Within the organ of Corti, lower concentrations of LN may slow neurite growth, with FN beneath each row of hair cells providing a stop or avoidance signal. This could allow growth cone filopodia increased time to sample their cellular targets, or direct the fibers upward toward the hair cells. © 2007 Wiley Periodicals, Inc. Develop Neurobiol, 2007 [source] Peroxisome proliferator-activated receptor-, as emerging target in liver diseaseDRUG DEVELOPMENT RESEARCH, Issue 2 2010Bernd Schnabl Abstract Liver fibrosis is characterized by an excessive deposition of extracellular matrix (ECM) proteins that occurs in chronic liver disease of any origin, including nonalcoholic steatohepatitis (NASH), alcohol abuse, and viral hepatitis. Cirrhosis occurs with the development of regenerating nodules of hepatocytes and is a major health burden worldwide. Patients with decompensated liver cirrhosis have a poor prognosis, with liver transplantation often being necessary. The current treatment paradigm for patients with hepatic fibrosis is to treat the underlying liver disease. However, if this cannot be achieved, there are currently no effective antifibrotic treatments for patients with chronic liver diseases. With the advent of basic molecular technology providing insight into the mechanisms of the development of hepatic fibrosis, there is now an opportunity to develop therapeutic interventions for human clinical use. In this review, the function of peroxisome proliferator-activated receptor-, (PPAR ,) will be summarized with a special emphasis on ligand activation as potential use in liver disease. Drug Dev Res 2009. © 2009 Wiley-Liss, Inc. [source] A microfluidic device for characterizing the invasion of cancer cells in 3-D matrixELECTROPHORESIS, Issue 24 2009Tingjiao Liu Abstract A microfluidic device was developed for the study of directed invasion of cancer cells in 3-D matrix with concentration gradient. This device consists of two parallel perfusion channels connected by two cell culture chambers. To mimic extracellular matrix (ECM), gelled basement membrane extract (BME) was used to support 3-D distribution of breast cancer cells (MCF7) in cell culture chambers. A stable linear concentration gradient of epidermal growth factor (EGF) was generated across the chambers by continuous perfusion. Using the device, we investigated MCF7 cell invasion induced by different concentrations of EGF in 3-D matrix. It was found that cancer cells responded to EGF stimulation with forming cellular protrusions and migrating towards high EGF concentration. We further investigated the anti-invasion effect of GM 6001, a matrix metalloproteinase inhibitor. We identified that matrix metalloproteinase inhibition repressed both cellular protrusion formation and cell migration in 3-D matrix. These findings suggest that EGF is able to induce MCF7 cell invasion in 3-D extracellular matrix and this effect is dependent on proteolytic activity. This device is relatively simple to construct and operate. It should be a useful platform for elucidating the mechanism of cancer invasion and screening anti-invasion drugs for cancer therapy. [source] Diversity and expression of nitrogenase genes (nifH) from ectomycorrhizas of Corsican pine (Pinus nigra)ENVIRONMENTAL MICROBIOLOGY, Issue 12 2006Hironari Izumi Summary The diversity of bacterial nitrogenase genes (nifH) and their mRNA transcription in ectomycorrhizas of Corsican pine (Pinus nigra) were examined. DNA and RNA were extracted from surface-sterilized and non-sterilized Corsican pine roots colonized by the ectomycorrhizal (ECM) fungi, Suillus variegatus and Tomentellopsis submollis. DNA-derived nifH polymerase chain reaction (PCR) products were obtained from all samples, but only a few reverse transcription PCRs for nifH mRNA were successful, suggesting that nitrogenase genes were not always transcribed. Several different nifH sequences were detected and the bacteria actively transcribing nifH were different from those whose genes were detected through DNA-based PCR. Putative nitrogenase amino acid sequences revealed that more than half of the nifH products were derived from methylotrophic bacteria, such as Methylocella spp. The next most frequent sequence types were similar to those from Burkholderia. [source] Remodeling of extracellular matrix and epileptogenesisEPILEPSIA, Issue 2010Alexander Dityatev Summary Extracellular matrix (ECM) in the brain is composed of molecules synthesized and secreted by neurons and glial cells, which form stable aggregates of diverse composition in the extracellular space. In the mature brain, ECM undergoes a slow turnover and restrains structural plasticity while supporting multiple physiologic processes, including perisomatic ,-aminobutyric acid (GABA)ergic inhibition, synaptic plasticity, and homeostatic regulations. Seizures lead to striking remodeling of ECM, which may be essentially engaged in different aspects of epileptogenesis. This view is supported by human genetic studies linking ECM molecules and epilepsy, by data showing altered epileptogenesis in mice deficient in ECM molecules, and by evidence that ECM may shape seizure-induced sprouting of mossy fibers, granule cell dispersion, and astrogliosis. Therefore, restraining seizure-induced remodeling of ECM or suppressing the signaling triggered by the remodeled ECM might provide effective therapeutic strategies to antagonize the progression of epileptogenesis. [source] Tissue inhibitor of metalloproteinse-1 is a marker of diastolic dysfunction using tissue doppler in patients with type 2 diabetes and hypertensionEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 1 2005M. H. Tayebjee Abstract Background, Tissue inhibitor of metalloproteinase-1 (TIMP-1) is associated with increased fibrosis of the extracellular matrix (ECM). Myocardial stiffness is a feature of diastolic dysfunction. We assessed circulating TIMP-1 as a marker of diastolic dysfunction in patients with type 2 diabetes mellitus (DM) and hypertension, who were compared with healthy controls. Methods, We recruited 54 patients (43 males; mean age 68 ± 5 years) with treated type 2 DM (i.e. controlled glycaemia, hypertension, hyperlipidaemia), 35 (30 males; 69 ± 8 years) treated nondiabetic hypertensives, and 31 healthy controls (18 males; 66 ± 5 years). Circulating TIMP-1 was measured by ELISA. Using transthoracic echocardiography, the early (E) diastolic mitral inflow velocity was measured with pulse wave Doppler, and the early mitral annular velocity (e,), a recognized index of diastolic relaxation, was measured with tissue Doppler. The E/A ratio was also calculated and isovolumic relaxation time measured. Results, Mean e, levels differed significantly between controls, diabetics and hypertensives (P < 0·0001). Circulating TIMP-1 was significantly different between patients and controls (P = 0·006), but there was no statistically significant difference between the DM and hypertension group. In both groups, only e, was negatively correlated with TIMP-1 levels, with a stronger correlation among the hypertensive patients (Spearman r = ,0·544, P = 0·001) when compared with the diabetic group (r = ,0·341, P = 0·011). Conclusion, Diastolic relaxation is impaired in diabetes and hypertensive patients. The relationship between TIMP-1 and e, may reflect increased myocardial fibrosis and consequent diastolic dysfunction, which may be more prominent in hypertension. [source] Immunoexpression of extracellular matrix proteins in human salivary gland developmentEUROPEAN JOURNAL OF ORAL SCIENCES, Issue 6 2004Cristiane Furuse Immunoexpression of the extracellular matrix (ECM) proteins laminin, fibronectin, tenascin and types I, III and IV collagen was analyzed in the major and minor salivary glands of seven human fetuses at different gestational ages. The results showed the presence and localization of laminin, collagen IV and fibronectin around glandular structures at all stages of development. Tenascin was only detectable around excretory ducts. In the earliest stages of development, type I and type III collagen were presented as fine fibers delineating the glandular structures and delimiting the extension of the future lobule. As glandular development proceeded, the lobule was gradually filled with collagens and glandular tissue. [source] mTOR as a potential therapeutic target for treatment of keloids and excessive scarsEXPERIMENTAL DERMATOLOGY, Issue 5 2007C. T. Ong Abstract:, Keloid is a dermal fibroproliferative disorder characterized by excessive deposition of extracellular matrix (ECM) components such as collagen, glycoproteins and fibronectin. The mammalian target of rapamycin (mTOR) is a serine/theronine kinase which plays an important role in the regulation of metabolic processes and translation rates. Published reports have shown mTOR as regulator of collagen expression and its inhibition induces a decrease in ECM deposition. Our aim was to investigate the role of mTOR in keloid pathogenesis and investigate the effect of rapamycin on proliferating cell nuclear antigen (PCNA), cyclin D1, collagen, fibronectin and alpha-smooth muscle actin (, -SMA) expression in normal fibroblasts (NF) and keloid fibroblasts (KF). Tissue extracts obtained from keloid scar demonstrated elevated expression of mTOR, p70KDa S6 kinase (p70S6K) and their activated forms, suggesting an activated state in keloid scars. Serum stimulation highlighted the heightened responsiveness of KF to mitogens and the importance of mTOR and p70S6K during early phase of wound healing. Application of rapamycin to monoculture NF and KF, dose- and time-dependently downregulates the expression of cytoplasmic PCNA, cyclin D1, fibronectin, collagen and , -SMA, demonstrating the anti-proliferative effect and therapeutic potential of rapamycin in the treatment of keloid scars. The inhibitory effect of rapamycin was found to be reversible following recovery in the expression of proteins following the removal of rapamycin from the culture media. These results demonstrate the important role of mTOR in the regulation of cell cycle and the expression of ECM proteins: fibronectin, collagen and , -SMA. [source] Capturing Complex Protein Gradients on Biomimetic Hydrogels for Cell-Based AssaysADVANCED FUNCTIONAL MATERIALS, Issue 21 2009Steffen Cosson Abstract A versatile strategy to rapidly immobilize complex gradients of virtually any desired protein on soft poly(ethylene glycol) (PEG) hydrogel surfaces that are reminiscent of natural extracellular matrices (ECM) is reported. A microfluidic chip is used to generate steady-state gradients of biotinylated or Fc-tagged fusion proteins that are captured and bound to the surface in less than 5,min by NeutrAvidin or ProteinA, displayed on the surface. The selectivity and orthogonality of the binding schemes enables the formation of parallel and orthogonal overlapping gradients of multiple proteins, which is not possible on conventional cell culture substrates. After patterning, the hydrogels are released from the microfluidic chip and used for cell culture. This novel platform is validated by conducting single-cell migration experiments using time-lapse microscopy. The orientation of cell migration, as well as the migration rate of primary human fibroblasts, depends on the concentration of an immobilized fibronectin fragment. This technique can be readily applied to other proteins to address a wealth of biological questions with different cell types. [source] Tissue Engineering: Scaffold-Mediated 2D Cellular Orientations for Construction of Three Dimensionally Engineered Tissues Composed of Oriented Cells and Extracellular Matrices (Adv. Funct.ADVANCED FUNCTIONAL MATERIALS, Issue 7 2009Mater. Novel technology for fabrication of 3D-engineered tissues consisting of oriented cells and extracellular matrices (ECM) is described by Yoshida and co-workers on page 1001. They prepare disulfide-crosslinked hydrogels with oriented pores to induce cellular orientations on and in the scaffolds. Finally, 3D-oriented engineered tissues with 2D-cell and ECM orientation are successfully obtained by decomposing template gel with reductant agent. [source] Scaffold-Mediated 2D Cellular Orientations for Construction of Three Dimensionally Engineered Tissues Composed of Oriented Cells and Extracellular MatricesADVANCED FUNCTIONAL MATERIALS, Issue 7 2009Hiroaki Yoshida Abstract Various hydrogels, such as poly(, -glutamic acid) (, -PGA), gelatin (GT), alginic acid (Alg), and agarose (Aga), with 3D interconnected and oriented fibrous pores (OP gels) are prepared for 3D polymeric cellular scaffolds by using silica fiber cloth (SC) as template. After the preparation of these hydrogels with the SC templates, the latter are subsequently removed by washing with hydrofluoric acid solution. Scanning electron microscopy (SEM) clearly shows OP structures in the hydrogels. These various types of OP gels are successfully prepared in this way, independently of the crosslinking mechanism, such as chemical (, -PGA or GT), coordinate-bonded (Alg), or hydrogen-bonded (Aga) crosslinks. SEM, confocal laser scanning microscopy, and histological evaluations clearly demonstrate that mouse L929 fibroblast cells adhere to and extend along these OP structures on/in , -PGA hydrogels during 3D cell culture. The L929 cells that adhere on/in the oriented hydrogel are viable and proliferative. Furthermore, 3D engineered tissues, composed of the oriented cells and extracellular matrices (ECM) produced by the cells, are constructed in vitro by subsequent decomposition of the hydrogel with cysteine after 14 days of cell culture. This novel technology to fabricate 3D-engineered tissues, consisting of oriented cells and ECM, will be useful for tissue engineering. [source] In the hypoxic central nervous system, endothelial cell proliferation is followed by astrocyte activation, proliferation, and increased expression of the ,6,4 integrin and dystroglycanGLIA, Issue 10 2010Longxuan Li Abstract Cerebral hypoxia induces a profound angiogenic response in the central nervous system (CNS). Using a mouse model of chronic cerebral hypoxia, we previously demonstrated that angiogenic vessels in the hypoxic CNS show marked upregulation of the extracellular matrix (ECM) protein fibronectin, along with increased expression of its major receptor, ,5,1 integrin on brain endothelial cells (BEC). As cerebral hypoxia also leads to glial activation, the aim of the current study was to define the temporal relationship between BEC responses and glial cell activation in this model of cerebral hypoxia. This revealed that BEC fibronectin/,5,1 integrin expression and proliferation both reached maximal level after 4-day hypoxia. Interestingly, up to 4-day hypoxia, all dividing cells were BEC, but at later time-points proliferating astrocytes were also observed. GFAP staining revealed that hypoxia induced marked astrocyte activation that reached maximal level between 7- and 14-day hypoxia. As newly formed cerebral capillaries require ensheathment by astrocyte end-feet to acquire mature brain endothelium characteristics, we next examined how expression of astrocyte end-feet adhesion molecules is regulated by hypoxia. This showed that the astrocyte adhesion receptors ,6,4 integrin and dystroglycan were both markedly upregulated, with a time-course that closely resembled astrocyte activation. Taken together, this evidence shows that cerebral hypoxia promotes first an endothelial response, in which fibronectin promotes BEC proliferation. This is then followed by an astrocyte response, involving astrocyte activation, proliferation, and reorganization of astrocyte end-feet, which correlates with increased expression of astrocyte end-feet adhesion molecules. © 2010 Wiley-Liss, Inc. [source] Elastin-derived peptides: Matrikines critical for glioblastoma cell aggressiveness in a 3-D systemGLIA, Issue 16 2009Bérénice Coquerel Abstract In the most common primary brain tumors, malignant glioma cells invade the extracellular matrix (ECM) and proliferate rapidly in the cerebral tissue, which is mainly composed of hyaluronan (HA) along with the elastin present in the basement membrane of blood vessels. To determine the role of ECM components in the invasive capacity of glioma cell lines, we developed a 3-D cell-culture system, based on a hydrogel in which HA can be coreticulated with kappa-elastin (HA-,E). Using this system, the invasiveness of cells from four glioma cell lines was dramatically increased by the presence of ,E and a related, specific peptide (VGVAPG)3. In addition, MMP-2 secretion increased and MMP-12 synthesis occurred. Extracellular injections of ,E or (VGVAPG)3 provoked a pronounced and dose-dependent increase in [Ca2+]i. ,E significantly enhanced the expression of the genes encoding elastin-receptor and tropoelastin. We propose the existence of a positive feedback loop in which degradation of elastin generates fragments that stimulate synthesis of tropoelastin followed by further degradation as well as migration and proliferation of the very cells responsible for degradation. All steps in this ECM-based loop could be blocked by the addition of either of the EBP antagonists, lactose, and V-14 peptide, suggesting that the loop itself should be considered as a new therapeutic target. © 2009 Wiley-Liss, Inc. [source] Microglial expression of ,v,3 and ,v,5 integrins is regulated by cytokines and the extracellular matrix: ,5 Integrin null microglia show no defects in adhesion or MMP-9 expression on vitronectinGLIA, Issue 7 2009Richard Milner Abstract As the primary immune effector cells in the CNS, microglia play a central role in regulating inflammation. The extracellular matrix (ECM) protein vitronectin is a strong inducer of microglial activation, switching microglia from a resting into an activated potentially destructive phenotype. As the activating effect of vitronectin is mediated by ,v integrins, the aim of the current study was to evaluate the requirement of the ,v,5 integrin in mediating microglial adhesion and activation to vitronectin, by studying these events in ,5 integrin-null murine microglia. Surprisingly, ,5 integrin null microglia were not defective in adhesion to vitronectin. Further analysis showed that microglia express the ,v,3 integrin, in addition to ,v,5. Flow cytometry revealed that microglial ,v integrin expression is regulated by cytokines and ECM proteins. ,v,3 integrin expression was downregulated by IFN-,, TNF, LPS, and TGF-,1. ,v,5 expression was also reduced by IFN-,, TNF, and LPS, but strongly increased by the antiactivating factors TGF-,1 and laminin. Gel zymography revealed that ,5 integrin null microglia showed no deficiency in their expression of matrix metalloproteinase (MMP)-9 in response to vitronectin. Taken together, these data show that microglia express two different ,v integrins, ,v,3 and ,v,5, and that expression of these integrins is independently regulated by cytokines and ECM proteins. Furthermore, it reveals that the ,v,5 integrin is not essential for mediating microglial adhesion and MMP-9 expression in response to vitronectin. © 2008 Wiley-Liss, Inc. [source] Nanostructured Biomaterials for Regeneration,ADVANCED FUNCTIONAL MATERIALS, Issue 22 2008Guobao Wei Abstract Biomaterials play a pivotal role in regenerative medicine, which aims to regenerate and replace lost/dysfunctional tissues or organs. Biomaterials (scaffolds) serve as temporary 3D substrates to guide neo tissue formation and organization. It is often beneficial for a scaffolding material to mimic the characteristics of extracellular matrix (ECM) at the nanometer scale and to induce certain natural developmental or/and wound healing processes for tissue regeneration applications. This article reviews the fabrication and modification technologies for nanofibrous, nanocomposite, and nanostructured drug-delivering scaffolds. ECM-mimicking nanostructured biomaterials have been shown to actively regulate cellular responses including attachment, proliferation, differentiation, and matrix deposition. Nanoscaled drug delivery systems can be successfully incorporated into a porous 3D scaffold to enhance the tissue regeneration capacity. In conclusion, nanostructured biomateials are a very exciting and rapidly expanding research area, and are providing new enabling technologies for regenerative medicine. [source] Evaluation of the MODFLOW-2005 Conduit Flow ProcessGROUND WATER, Issue 4 2010Melissa E. Hill The recent development of the Conduit Flow Process (CFP) by the U.S. Geological Survey (USGS) provides hydrogeologic modelers with a new tool that incorporates the non-Darcian, multiporosity components of flow characteristic of karst aquifers. CFP introduces new parameters extending beyond those of traditional Darcian groundwater flow codes. We characterize a karst aquifer to collect data useful for evaluating this new tool at a test site in west-central Florida, where the spatial distribution and cross-sectional area of the conduit network are available. Specifically, we characterize: (1) the potential for Darcian/non-Darcian flow using estimates of specific discharge vs. observed hydraulic gradients, and (2) the temporal variation for the direction and magnitude of fluid exchange between the matrix and conduit network during extreme hydrologic events. We evaluate the performance of CFP Mode 1 using a site-scale dual-porosity model and compare its performance with a comparable laminar equivalent continuum model (ECM) using MODFLOW-2005. Based on our preliminary analyses, hydraulic conductivity coupled with conduit wall conductance improved the match between observed and simulated discharges by 12% to 40% over turbulent flow alone (less than 1%). [source] Temperature-Induced Hydrogels Through Self-Assembly of Cholesterol-Substituted Star PEG- b -PLLA Copolymers: An Injectable Scaffold for Tissue Engineering,ADVANCED FUNCTIONAL MATERIALS, Issue 8 2008Koji Nagahama Abstract Partially cholesterol-substituted 8-arm poly(ethylene glycol)- block -poly(L -lactide) (8-arm PEG- b -PLLA-cholesterol) has been prepared as a novel star-shaped, biodegradable copolymer derivative. The amphiphilic 8-arm PEG- b -PLLA-cholesterol aqueous solution (polymer concentration, above 3,wt%) exhibits instantaneous temperature-induced gelation at 34,°C, but the virgin 8-arm PEG- b -PLLA does not, irrespective of concentration. Moreover, an extracellular matrix (ECM)-like micrometer-scale network structure has been created with favorable porosity for three-dimensional proliferation of cells inside the hydrogel. This network structure is mainly attributed to specific self-assembly between cholesterol groups. The 10 and 20,wt% hydrogels are eroded gradually in phosphate buffered saline at 37,°C over the course of a month, and after that the gel becomes completely dissociated. Moreover, L929 cells encapsulated into the hydrogel are viable and proliferate three-dimensionally inside the hydrogels. Thus, in-vitro cell culture studies demonstrate that 8-arm PEG- b -PLLA-cholesterol is a promising candidate as a novel injectable cellular scaffold. [source] Thirty-kilodalton Tat-interacting protein suppresses tumor metastasis by inhibition of osteopontin transcription in human hepatocellular carcinoma,HEPATOLOGY, Issue 1 2008Jian Zhao It has been previously demonstrated that the 30-kDa Tat-interacting protein (TIP30) plays an important role in the suppression of hepatocarcinogenesis by acting as a tumor suppressor. Here we report that TIP30 suppresses metastasis of hepatocellular carcinoma (HCC) through inhibiting the transcription of osteopontin (OPN), a key molecule in the development of tumor metastasis. The expression of TIP30 messenger RNA was reverse to that of OPN messenger RNA in HCC cell lines. Ectopic expression of TIP30 greatly suppressed OPN expression, inhibited invasion of HCC cells through extracellular matrix (ECM) and adhesion with fibronectin in vitro, whereas down-regulation of TIP30 by RNA-mediated interference enhanced OPN expression and promoted metastatic abilities of HCC cells in vitro. Moreover, overexpression of TIP30 significantly inhibited the growth and lung metastases of HCC cells in nude mice. In contrast, down-regulation of TIP30 greatly promoted tumor cell growth and metastases in vivo. TIP30 repressed OPN transcription through interaction with Ets-1 and suppressed the transcriptional activity of Ets-1 and synergistic actions of Ets-1 and alkaline phosphatase-1. Thus, TIP30 may act as an Ets-1 modulator and inhibit tumor metastasis through abrogating Ets-1,dependent transcription. Moreover, expression of TIP30 was inversely associated with OPN expression in HCC tissue samples as detected by immunohistochemistry assay. Conclusion: Our results reveal a novel pathway by which OPN and possibly other Ets-1 target genes involved in tumor metastasis are regulated by TIP30 and elucidate a mechanism for metastasis promoted by TIP30 deficiency. (HEPATOLOGY 2008.) [source] Laminin-5 stimulates hepatocellular carcinoma growth through a different function of ,6,4 and ,3,1 integrins,HEPATOLOGY, Issue 6 2007Carlo Bergamini Hepatocellular carcinoma (HCC) growth severely affects prognosis. Ki-67, a known marker of cell proliferation, is a negative prognostic factor in HCC. Growth factors such as the epidermal growth factor (EGF) induce HCC cell proliferation but do not explain the great heterogeneity of HCC growth. Laminin-5 (Ln-5) is an extracellular matrix protein (ECM) present in the tissue microenvironment of HCC. The two main receptors for Ln-5, integrins ,3,1 and ,6,4, are expressed on the cell surface of HCC cells. The aim of this study is to investigate an alternative mechanism of HCC growth whereby Ln-5 promotes HCC cell proliferation through ,3,1 and ,6,4. HCC tissues containing Ln-5 display a larger diameter and higher number of positive cells for Ki-67, a well known proliferative index, as determined by double immunofluorescence staining and real-time PCR on microdissected tissues. In vitro, Ln-5, but not collagen I, collagen IV or fibronectin, induces proliferation as much as EGF does, via Erk phosphorylation as a consequence of ,4 integrin phosphorylation. However, the two HCC cell lines do not proliferate in presence of Ln-5 despite ,4 integrin and Erk1/2 activation. After transfection with ,3 integrin, in the presence of Ln-5 one of these HCC cell lines acquires a proliferative activity whereas one of the proliferative HCC cell lines, knocked-down for ,3 integrin, loses its proliferative activity. Conclusions: Our study suggests a new mechanism of HCC growth whereby Ln-5 stimulates proliferation via a different function of ,6,4 and ,3,1. (HEPATOLOGY 2007.) [source] Fibroblast activation protein increases apoptosis, cell adhesion, and migration by the LX-2 human stellate cell line,HEPATOLOGY, Issue 4 2005Xin Maggie Wang Injury and repair in chronic liver disease involve cell adhesion, migration, apoptosis, proliferation, and a wound healing response. In liver, fibroblast activation protein (FAP) has both collagenase and dipeptidyl peptidase IV (DPIV) activities and is expressed only by activated hepatic stellate cells (HSC) and myofibroblasts, which produce and degrade extracellular matrix (ECM). FAP was colocalized with collagen fibers, fibronectin, and collagen type I in human liver. FAP function was examined in vitro by expressing green fluorescent protein FAP fusion protein in cell lines cultured on collagen-I, fibronectin, and Matrigel. Glutamates at 203 and 204 as well as serine624 of FAP were essential for peptidase activity. Human embryonic kidney 293T cells overexpressing FAP showed reduced adhesion and migration. FAP overexpression in the human HSC line LX-2 caused increased cell adhesion and migration on ECM proteins as well as invasion across transwells in the absence or presence of transforming growth factor beta-1. FAP overexpression enhanced staurosporine streptomyces,stimulated apoptosis in both cell lines. Interestingly, the enzyme activity of FAP was not required for these functions. Overexpressing FAP increased the expression of matrix metalloproteinase-2 and CD44 and reduced integrin-,1 expression in 293T cells, suggesting potential pathways of FAP-mediated impairment of cell adhesion and migration in this epithelial cell line. In conclusion, these findings further support a pro-fibrogenic role for FAP by indicating that, in addition to its enzymatic functions, FAP has important nonenzymatic functions that in chronic liver injury may facilitate tissue remodeling through FAP-mediated enhancement of HSC cell adhesion, migration, and apoptosis. Supplementary material for this article can be found on the HEPATOLOGY website (http://www.interscience.wiley.com/jpages/0270-9139/suppmat/index.html). (HEPATOLOGY 2005;42:935,945.) [source] Dynamic Hydrogels: Switching of 3D Microenvironments Using Two-Component Naturally Derived Extracellular Matrices (Adv. Mater.ADVANCED MATERIALS, Issue 6 20106/2010) The front cover image depicts a two-component extracellular matrix (ECM) in which one component acts as a stable structural element (which supports cell attachment and migration) and another component gels or dissolves reversibly (a modulatory component). Samuel K. Sia and co-workers show on p. 686 that by dynamically adding or removing crosslinks in the modulatory component, properties of the composite ECM, such as the ability of cells to migrate and the rate of diffusive transport, can be altered. [source] Dynamic Hydrogels: Switching of 3D Microenvironments Using Two-Component Naturally Derived Extracellular MatricesADVANCED MATERIALS, Issue 6 2010Brian M. Gillette This article describes fabrication of a two-component extracellular matrix (ECM) in which one component acts as a stable structural element and another component gels or dissolves reversibly (a modulatory component). Using a composite collagen-alginate ECM, reversible crosslinking of the alginate (the modulatory component) via application of calcium or citrate modulates cell mobility in a 3D collagen matrix (the structural component). [source] Progress in the Field of Electrospinning for Tissue Engineering ApplicationsADVANCED MATERIALS, Issue 32-33 2009Seema Agarwal Abstract Electrospinning is an extremely promising method for the preparation of tissue engineering (TE) scaffolds. This technique provides nonwovens resembling in their fibrillar structures those of the extracellular matrix (ECM), and offering large surface areas, ease of functionalization for various purposes, and controllable mechanical properties. The recent developments toward large-scale productions combined with the simplicity of the process render this technique very attractive. Progress concerning the use of electrospinning for TE applications has advanced impressively. Different groups have tackled the problem of electrospinning for TE applications from different angles. Nowadays, electrospinning of the majority of biodegradable and biocompatible polymers, either synthetic or natural, for TE applications is straightforward. Different issues, such as cell penetration, incorporation of growth and differentiating factors, toxicity of solvents used, productivity, functional gradient, etc. are main points of current considerations. The progress in the use of electrospinning for TE applications is highlighted in this article with focus on major problems encountered and on various solutions available until now. [source] Immunolocalization of bone extracellular matrix proteins (type I collagen, osteonectin and bone sialoprotein) in human dental pulp and cultured pulp cellsINTERNATIONAL ENDODONTIC JOURNAL, Issue 6 2003J. M. Q. Garcia Abstract Aim, To simultaneously analyse the expression of type I collagen, osteonectin and bone sialoprotein (BSP) in human dental pulp of different ages. Methodology, Cultured dental pulp fibroblasts (FP1 cell line), pulps from dental germs with incomplete root formation (n = 4) and pulps of erupted teeth with total root formation (n = 4) were used. Bone proteins were searched by immunohistochemistry and immunofluorescence using polyclonal antibodies and compared among the three groups assessed. Results, Immunohistochemistry detected the three proteins in dental pulp tissue, as it labelled extracellular matrix, predentine and odontoblasts. The BSP label was weaker, when compared to both type I collagen and osteonectin. The presence of type I collagen was more evident in pulps from erupted teeth, when compared to germ dental pulps. On the other hand, a strong expression of osteonectin in germ dental pulps was observed. Conclusions, Regardless of the degree of maturation, dental pulps present type I collagen, osteonectin and BSP in the extracellular matrix (ECM) and in the odontoblastic layer. Thus, the results suggest that these proteins are related to the production and mineralization of dentine. [source] | |||||||||||||||||||||||||||||||||||||