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Sample records for fibronectin drives focal

  1. Fibronectin is not Present in the Focal Adhesions Formed between Normal Cultured Fibroblasts and Their Substrata

    NASA Astrophysics Data System (ADS)

    Chen, Wen-Tien; Singer, S. J.

    1980-12-01

    Fibronectin is an extracellular matrix protein that has been implicated in the spreading and adhesion of cultured fibroblasts to their substrata. In this paper, double immunoelectron microscopic labeling experiments for fibronectin and for concanavalin A-binding proteins on the cell surface were carried out on ultrathin frozen sections of cultures of embryonic chicken heart fibroblasts. On cross sections through the focal adhesions of the cell to the substratum there was substantial labeling for concanavalin A-binding proteins but no detectable labeling for fibronectin, whereas both the binding proteins and fibronectin were extensively labeled elsewhere on the cell surface and substratum. These results demonstrate that fibronectin is not present within the sites of focal adhesions. Therefore, the functions of fibronectin in cell spreading and adhesion are not directly mediated through its binding at focal adhesion sites. An alternative model is presented which can account for such fibronectin functions.

  2. A small fibronectin-mimicking protein from bacteria induces cell spreading and focal adhesion formation.

    PubMed

    Tegtmeyer, Nicole; Hartig, Roland; Delahay, Robin M; Rohde, Manfred; Brandt, Sabine; Conradi, Jens; Takahashi, Seiichiro; Smolka, Adam J; Sewald, Norbert; Backert, Steffen

    2010-07-23

    Fibronectin, a 250-kDa eukaryotic extracellular matrix protein containing an RGD motif plays crucial roles in cell-cell communication, development, tissue homeostasis, and disease development. The highly complex fibrillar fibronectin meshwork orchestrates the functions of other extracellular matrix proteins, promoting cell adhesion, migration, and intracellular signaling. Here, we demonstrate that CagL, a 26-kDa protein of the gastric pathogen and type I carcinogen Helicobacter pylori, mimics fibronectin in various cellular functions. Like fibronectin, CagL contains a RGD motif and is located on the surface of the bacterial type IV secretion pili as previously shown. CagL binds to the integrin receptor alpha(5)beta(1) and mediates the injection of virulence factors into host target cells. We show that purified CagL alone can directly trigger intracellular signaling pathways upon contact with mammalian cells and can complement the spreading defect of fibronectin(-/-) knock-out cells in vitro. During interaction with various human and mouse cell lines, CagL mimics fibronectin in triggering cell spreading, focal adhesion formation, and activation of several tyrosine kinases in an RGD-dependent manner. Among the activated factors are the nonreceptor tyrosine kinases focal adhesion kinase and Src but also the epidermal growth factor receptor and epidermal growth factor receptor family member Her3/ErbB3. Interestingly, fibronectin activates a similar range of tyrosine kinases but not Her3/ErbB3. These findings suggest that the bacterial protein CagL not only exhibits functional mimicry with fibronectin but is also capable of activating fibronectin-independent signaling events. We thus postulate that CagL may contribute directly to H. pylori pathogenesis by promoting aberrant signaling cross-talk within host cells.

  3. Thrombomodulin promotes focal adhesion kinase activation and contributes to angiogenesis by binding to fibronectin

    PubMed Central

    Hsu, Yun-Yan; Shi, Guey-Yueh; Wang, Kuan-Chieh; Ma, Chih-Yuan; Cheng, Tsung-Lin; Wu, Hua-Lin

    2016-01-01

    Angiogenesis promotes tumor growth and metastasis. Cell adhesion molecules interact with the extracellular matrix (ECM) and increase cell adhesion and migration during angiogenesis. Thrombomodulin (TM) is a cell surface transmembrane glycoprotein expressed in endothelial cells. However, the function and significance of TM in cell-matrix interactions and angiogenesis remain unclear. Here, we first demonstrated that recombinant lectin-like domain of TM interacts with an ECM protein, fibronectin, and identified the N-terminal 70-kDa domain of fibronectin as the TM-binding site. Exogenous expression of TM in TM-deficient A2058 melanoma cells enhanced cell adhesion and migration on fibronectin and invasion on Matrigel. In addition, TM increased focal adhesion kinase (FAK) phosphorylation and matrix metalloproteinase-9 production. In mice bearing subcutaneous B16F10 melanoma tumors, immunofluorescence analysis indicated that TM was highly expressed and co-localized with fibronectin on the tumor vasculature. The interaction between TM and fibronectin in tumor blood vessels was also validated by the proximity ligation assay. In human umbilical vein endothelial cells, up-regulation of TM by vascular endothelial growth factor (VEGF), a tumor angiogenic factor, promoted cell adhesion and tube formation, whereas TM knockdown by RNA interference attenuated VEGF-induced cell adhesion and tube formation. In summary, TM promotes angiogenesis by enhancing cell adhesion, migration, and FAK activation through interaction with fibronectin. TM may represent a novel target for inhibiting tumor angiogenesis. PMID:27602495

  4. Micrometer scale spacings between fibronectin nanodots regulate cell morphology and focal adhesions

    NASA Astrophysics Data System (ADS)

    Horzum, Utku; Ozdil, Berrin; Pesen-Okvur, Devrim

    2014-04-01

    Cell adhesion to extracellular matrix is an important process for both health and disease states. Surface protein patterns that are topographically flat, and do not introduce other chemical, topographical or rigidity related functionality and, more importantly, that mimic the organization of the in vivo extracellular matrix are desired. Previous work showed that vinculin and cytoskeletal organization are modulated by size and shape of surface nanopatterns. However, quantitative analysis on cell morphology and focal adhesions as a function of micrometer scale spacings of FN nanopatterns was absent. Here, electron beam lithography was used to pattern fibronectin nanodots with micrometer scale spacings on a K-casein background on indium tin oxide coated glass which, unlike silicon, is transparent and thus suitable for many light microscopy techniques. Exposure times were significantly reduced using the line exposure mode with micrometer scale step sizes. Micrometer scale spacings of 2, 4 and 8 μm between fibronectin nanodots proved to modulate cell adhesion through modification of cell area, focal adhesion number, size and circularity. Overall, cell behavior was shown to shift at the apparent threshold of 4 μm spacing. The findings presented here offer exciting new opportunities for cell biology research.

  5. Full-Length Fibronectin Drives Fibroblast Accumulation at the Surface of Collagen Microtissues during Cell-Induced Tissue Morphogenesis

    PubMed Central

    Foolen, Jasper; Shiu, Jau-Ye; Mitsi, Maria; Zhang, Yang; Chen, Christopher S.; Vogel, Viola

    2016-01-01

    Generating and maintaining gradients of cell density and extracellular matrix (ECM) components is a prerequisite for the development of functionality of healthy tissue. Therefore, gaining insights into the drivers of spatial organization of cells and the role of ECM during tissue morphogenesis is vital. In a 3D model system of tissue morphogenesis, a fibronectin-FRET sensor recently revealed the existence of two separate fibronectin populations with different conformations in microtissues, i.e. ‘compact and adsorbed to collagen’ versus ‘extended and fibrillar’ fibronectin that does not colocalize with the collagen scaffold. Here we asked how the presence of fibronectin might drive this cell-induced tissue morphogenesis, more specifically the formation of gradients in cell density and ECM composition. Microtissues were engineered in a high-throughput model system containing rectangular microarrays of 12 posts, which constrained fibroblast-populated collagen gels, remodeled by the contractile cells into trampoline-shaped microtissues. Fibronectin’s contribution during the tissue maturation process was assessed using fibronectin-knockout mouse embryonic fibroblasts (Fn-/- MEFs) and floxed equivalents (Fnf/f MEFs), in fibronectin-depleted growth medium with and without exogenously added plasma fibronectin (full-length, or various fragments). In the absence of full-length fibronectin, Fn-/- MEFs remained homogenously distributed throughout the cell-contracted collagen gels. In contrast, in the presence of full-length fibronectin, both cell types produced shell-like tissues with a predominantly cell-free compacted collagen core and a peripheral surface layer rich in cells. Single cell assays then revealed that Fn-/- MEFs applied lower total strain energy on nanopillar arrays coated with either fibronectin or vitronectin when compared to Fnf/f MEFs, but that the presence of exogenously added plasma fibronectin rescued their contractility. While collagen

  6. 12-O-tetradecanoylphorbol-13-acetate disrupts actin filaments and focal contacts and enhances binding of fibronectin-coated latex beads to 3T3-L1 cells

    SciTech Connect

    Shiba, Yoshiki; Sasaki, Yasuto; Kanno, Yoshinobu )

    1988-10-01

    The effect of a tumor-promoting phorbol ester on the binding of fibronectin-coated beads to 3T3-L1 cells was studied to clarify the relationship between the binding of fibronectin to the cells, cell adhesion, and the organization of actin filaments. Interference-reflection microscopy revealed focal contacts of 3T3-L1 cells with the substratum. Stress fibers observed after rhodomine-phalloidin staining were well-developed in the cells. Treatment of the cells for 20 min with 12-O-tetradecanoylphorbol-13-acetate (TPA), but not with phorbol, disrupted focal contacts and caused a reorganization of stress fibers to generate actin ribbons. Treatment of the cells with TPA enhanced the binding of beads coated with human plasma fibronectin to the cells, as observed after incubation for 6 h with the beads. The TPA-induced increase in the percentage of cells with bound beads was dependent on the duration of treatment with TPA and on the concentration of TPA. Treatment of the cells with TPA also enhanced proliferation of cells in a dose-dependent manner. Furthermore, treatment of the cells with phorbol did not enhance the binding of beads coated with fibronectin. These results suggest that TPA specifically enhances the binding of fibronectin-coated beads to 3T3-L1 cells, and that TPA-induced binding of the beads may be related to disruption of focal contacts and reorganization of actin filaments.

  7. Focal adhesion kinase and paxillin promote migration and adhesion to fibronectin by swine skeletal muscle satellite cells.

    PubMed

    Wang, Dan; Gao, Chun-Qi; Chen, Rong-Qiang; Jin, Cheng-Long; Li, Hai-Chang; Yan, Hui-Chao; Wang, Xiu-Qi

    2016-05-24

    The focal adhesion kinase (FAK) signaling pathway contributes to the cell migration and adhesion that is critical for wound healing and regeneration of damaged muscle, but its function in skeletal muscle satellite cells (SCs) is less clear. We compared the migration and adhesion of SCs derived from two species of pig (Lantang and Landrace) in vitro, and explored how FAK signaling modulates the two processes. The results showed that Lantang SCs had greater ability to migrate and adhere to fibronection (P < 0.05) than Landrace SCs. Compared to Landrace SCs, Lantang SCs expressed many more focal adhesion (FA) sites, which were indicated by the presence of p-paxillin (Tyr118), and exhibited less F-actin reorganization 24 h after seeding onto fibronectin. Levels of p-FAK (Tyr397) and p-paxillin (Tyr118) were greater (P < 0.05) in Lantang SCs than Landrace SCs after migration for 24 h. Similarly, Lantang SCs showed much higher levels of p-FAK (Tyr397), p-paxillin (Tyr118) and p-Akt (Ser473) than Landrace SCs 2 h after adhesion. Treatment with the FAK inhibitor PF-573228 (5 or 10 μmol/L) inhibited Lantang SC migration and adhesion to fibronectin (P < 0.05), decreased levels of p-paxillin (Tyr118) and p-Akt (Ser473) (P < 0.05), and suppressed the formation of FA sites on migrating SCs. Thus FAK appears to play a key role in the regulation of SC migration and adhesion necessary for muscle regeneration.

  8. Focal adhesion kinase and paxillin promote migration and adhesion to fibronectin by swine skeletal muscle satellite cells

    PubMed Central

    Wang, Dan; Gao, Chun-qi; Chen, Rong-qiang; Jin, Cheng-long; Li, Hai-chang; Yan, Hui-chao; Wang, Xiu-qi

    2016-01-01

    The focal adhesion kinase (FAK) signaling pathway contributes to the cell migration and adhesion that is critical for wound healing and regeneration of damaged muscle, but its function in skeletal muscle satellite cells (SCs) is less clear. We compared the migration and adhesion of SCs derived from two species of pig (Lantang and Landrace) in vitro, and explored how FAK signaling modulates the two processes. The results showed that Lantang SCs had greater ability to migrate and adhere to fibronection (P < 0.05) than Landrace SCs. Compared to Landrace SCs, Lantang SCs expressed many more focal adhesion (FA) sites, which were indicated by the presence of p-paxillin (Tyr118), and exhibited less F-actin reorganization 24 h after seeding onto fibronectin. Levels of p-FAK (Tyr397) and p-paxillin (Tyr118) were greater (P < 0.05) in Lantang SCs than Landrace SCs after migration for 24 h. Similarly, Lantang SCs showed much higher levels of p-FAK (Tyr397), p-paxillin (Tyr118) and p-Akt (Ser473) than Landrace SCs 2 h after adhesion. Treatment with the FAK inhibitor PF-573228 (5 or 10 μmol/L) inhibited Lantang SC migration and adhesion to fibronectin (P < 0.05), decreased levels of p-paxillin (Tyr118) and p-Akt (Ser473) (P < 0.05), and suppressed the formation of FA sites on migrating SCs. Thus FAK appears to play a key role in the regulation of SC migration and adhesion necessary for muscle regeneration. PMID:27127174

  9. A synthetic peptide from the heparin-binding domain III (repeats III4-5) of fibronectin promotes stress-fibre and focal-adhesion formation in melanoma cells.

    PubMed Central

    Moyano, José V; Maqueda, Alfredo; Albar, Juan P; Garcia-Pardo, Angeles

    2003-01-01

    Cell adhesion to fibronectin results in formation of actin stress fibres and focal adhesions. In fibroblasts, this response requires two co-operative signals provided by interactions of the RGD sequence with alpha5beta1 integrin and the heparin-binding domain II (Hep II) domain with syndecan-4. Within Hep II, this activity was mapped to repeat III13 and to the peptide FN-C/H-V(WQPPRARITGY, repeat III14). We previously described that the synthetic heparin-binding peptide/III5 (HBP/III5) (WTPPRAQITGYRLTVGLTRR, repeat III5) binds heparin and mediates cell adhesion via chondroitin sulphate proteoglycans. We have now studied whether HBP/III5 co-operates with alpha5beta1 and drives a full cytoskeletal response in melanoma cells. SKMEL-178 cells attached and spread on the RGD-containing FNIII7-FNIII10 (FNIII7-10) fragment, but did not form stress fibres or focal adhesions. Co-immobilization of HBP/III5 with FNIII7-10 or adding soluble HBP/III5 to cells prespread on FNIII7-10, effectively induced these structures. Cell transfection with dominant-negative N19RhoA, a member of the small GTPase family, abolished the HBP/III5 effect. Both chondroitinase and heparitinase diminished focal adhesions, indicating that both types of proteoglycans bound HBP/III5 in melanoma cells. We have mapped the active sequence of HBP/III5 to YRLTVGLTRR, which is a novel sequence in fibronectin with focal-adhesion-promoting activity. The last two arginine (R) residues of this sequence are required for activity, since their replacement by alanine completely abrogated the HBP/III5 cytoskeletal effect. Moreover, this sequence is also active in the context of large fibronectin fragments. Our results establish that the Hep III region provides co-operative signals to alpha5beta1 for the progression of the cytoskeletal response and that these include activation of RhoA. PMID:12519080

  10. Fast focal zooming scheme for direct drive fusion implemented by inserting KD2PO4 crystal

    NASA Astrophysics Data System (ADS)

    Zhong, Zheqiang; Hu, Xiaochuan; Zhang, Bin

    2016-06-01

    The highly required uniformity of target in direct-drive fusion is difficult to achieve and maintain during the entire laser fusion implosion. To mitigate the increasing nonuniformity, the fast focal zooming scheme implemented by inserting an electro-optic (EO) crystal in the front end of beamline has been proposed. Functioning as a phase plate, the specifically designed EO crystal may add the induced spherical wavefront to the laser beam and alter its focusing characteristics. However, in order to zoom out the focal spot by half, the required voltage for KD2PO4 (DKDP) with single pair of electrodes is relatively high. In order to decrease the voltage while maintaining the zooming performance, the DKDP cylinder with multi pairs of electrodes has been presented. The continuous phase plate (CPP) is designed according to both the injected beam and the output field. However, the conventional CPP is designed based on the assumption of an injected beam without wavefront distortion, which would zoom in the focal spot variation in the focal zooming scheme. In order to zoom out the focal spot, a redesigned CPP has been proposed by adding a spherical wavefront to the phase variation of the conventional CPP and further optimizing. On the basis, the focusing characteristics of laser beam during the fast focal zooming process have been analyzed. Results indicate that the focal spot size decreases with the increasing voltage on DKDP crystal, meanwhile the uniformity maintains high during the focal zooming process.

  11. PPFIA1 drives active α5β1 integrin recycling and controls fibronectin fibrillogenesis and vascular morphogenesis

    PubMed Central

    Mana, Giulia; Clapero, Fabiana; Panieri, Emiliano; Panero, Valentina; Böttcher, Ralph T.; Tseng, Hui-Yuan; Saltarin, Federico; Astanina, Elena; Wolanska, Katarzyna I.; Morgan, Mark R.; Humphries, Martin J.; Santoro, Massimo M.; Serini, Guido; Valdembri, Donatella

    2016-01-01

    Basolateral polymerization of cellular fibronectin (FN) into a meshwork drives endothelial cell (EC) polarity and vascular remodelling. However, mechanisms coordinating α5β1 integrin-mediated extracellular FN endocytosis and exocytosis of newly synthesized FN remain elusive. Here we show that, on Rab21-elicited internalization, FN-bound/active α5β1 is recycled to the EC surface. We identify a pathway, comprising the regulators of post-Golgi carrier formation PI4KB and AP-1A, the small GTPase Rab11B, the surface tyrosine phosphatase receptor PTPRF and its adaptor PPFIA1, which we propose acts as a funnel combining FN secretion and recycling of active α5β1 integrin from the trans-Golgi network (TGN) to the EC surface, thus allowing FN fibrillogenesis. In this framework, PPFIA1 interacts with active α5β1 integrin and localizes close to EC adhesions where post-Golgi carriers are targeted. We show that PPFIA1 is required for FN polymerization-dependent vascular morphogenesis, both in vitro and in the developing zebrafish embryo. PMID:27876801

  12. Focal adhesion kinase and mitogen-activated protein kinases are involved in chondrocyte activation by the 29-kDa amino-terminal fibronectin fragment.

    PubMed

    Gemba, Takefumi; Valbracht, Jean; Alsalameh, Saifeddin; Lotz, Martin

    2002-01-11

    The 29-kDa amino-terminal fibronectin fragment (FN-f) has a potent chondrolytic effect and is thought to be involved in cartilage degradation in arthritis. However, little is known about signal transduction pathways that are activated by FN-f. Here we demonstrated that FN-f induced nitric oxide (NO) production from human articular chondrocytes. Expression of inducible nitric-oxide synthase (iNOS) mRNA and NO production were observed at 6 and 48 h after FN-f treatment, respectively. Interleukin-1beta (IL-1beta) mRNA up-regulation was stimulated by FN-f in human chondrocytes. To address the possibility that FN-f-induced NO release is mediated by IL-1beta production, the effect of IL-1 receptor antagonist (IL-1ra) was determined. IL-1ra partially inhibited FN-f-induced NO release although it almost completely inhibited IL-1beta-induced NO release. Tyrosine phosphorylation of focal adhesion kinase was induced transiently by FN-f treatment. Blocking antibodies to alpha(5) or beta(1) integrin and Arg-Gly-Asp-containing peptides did not inhibit FN-f-induced NO production. PP2, a Src family kinase inhibitor, or cytochalasin D, which selectively disrupts the network of actin filaments, inhibited both FAK phosphorylation and NO production induced by FN-f, but the phosphatidylinositol 3-kinase inhibitor wortmannin had no effect. Analysis of mitogen-activated protein kinases (MAPK) showed activation of extracellular signal-regulated kinase (ERK), c-Jun NH(2)-terminal kinase, and p38 MAPK. High concentrations of SB203580, which inhibit both JNK and p38 MAPK, and PD98059 a selective inhibitor of MEK1/2 that blocks ERK activation, inhibited FN-f induced NO production. These data suggest that focal adhesion kinase and MAPK mediate FN-f induced activation of human articular chondrocytes.

  13. Modeling of pattern development during fibronectin nanofibril formation.

    PubMed

    Pompe, Tilo; Starruss, Jörn; Bobeth, Manfred; Pompe, Wolfgang

    2006-09-01

    Formation of fibrillar fibronectin networks is a major process during embryogenesis and tissue formation, but the molecular details of fibril assembly remain poorly understood. Based on current ideas of fibronectin fibrillogenesis, a stochastic model was developed to enlighten the mechanism of the formation of paired fibronectin nanofibrils by adherent endothelial cells, which has been observed recently. The development of fibronectin clusters and fibrils was investigated by means of Monte Carlo simulations, including diffusion-controlled aggregation and myosin-driven transport of fibronectin-integrin complexes in the vicinity of a focal adhesion. Different evolving growth patterns were summarized in a morphological diagram as a function of the fibronectin substrate and fibronectin-fibronectin interaction energies. The formation of paired nanofibrils was found to occur in a certain region of interaction parameters. Beyond this region branched fibronectin clusters as well as tear-off of fibronectin fibrils were observed.

  14. Focal waveforms for various source waveforms driving a prolate-spheroidal impulse radiating antenna (IRA)

    NASA Astrophysics Data System (ADS)

    Altunc, Serhat; Baum, Carl E.; Christodoulou, Christos G.; Schamiloglu, Edl; Buchenauer, C. Jerald

    2008-08-01

    Impulse radiating antennas (IRAs) are designed to radiate very fast pulses in a narrow beam with low dispersion and high field amplitude. For this reason they have been used in a variety of applications. IRAs have been developed for use in the transient far-field region using parabolic reflectors. However, in this paper we focus in the near field region and develop the field waveform at the second focus of a prolate-spheroidal IRA. Certain skin cancers can be killed by the application of a high-amplitude electric field pulse. This can be accomplished by either inserting electrodes near the skin cancer or by applying fast, high-electric field pulses without direct contact. We investigate a new manifestation of an IRA, in which we use a prolate spheroid as a reflector instead of a parabolic reflector and focus in the near-field region instead of the far-field region. This technique minimizes skin damage associated with inserting electrodes near the tumor. Analytical and experimental behaviors for the focal waveforms of two and four-feed arm prolate-spheroidal IRAs are explored. With appropriate choice of the driving waveform we maximize the impulse field at the second focus. The focal waveform of a prolate-spheroidal IRA has been explained theoretically and verified experimentally.

  15. Haematopoietic focal adhesion kinase deficiency alters haematopoietic homeostasis to drive tumour metastasis.

    PubMed

    Batista, Silvia; Maniati, Eleni; Reynolds, Louise E; Tavora, Bernardo; Lees, Delphine M; Fernandez, Isabelle; Elia, George; Casanovas, Oriol; Lo Celso, Cristina; Hagemann, Thorsten; Hodivala-Dilke, Kairbaan

    2014-10-01

    Metastasis is the main cause of cancer-related death and thus understanding the molecular and cellular mechanisms underlying this process is critical. Here, our data demonstrate, contrary to established dogma, that loss of haematopoietic-derived focal adhesion kinase (FAK) is sufficient to enhance tumour metastasis. Using both experimental and spontaneous metastasis models, we show that genetic ablation of haematopoietic FAK does not affect primary tumour growth but enhances the incidence of metastasis significantly. At a molecular level, haematopoietic FAK deletion results in an increase in PU-1 levels and decrease in GATA-1 levels causing a shift of hematopoietic homeostasis towards a myeloid commitment. The subsequent increase in circulating granulocyte number, with an increase in serum CXCL12 and granulocyte CXCR4 levels, was required for augmented metastasis in mice lacking haematopoietic FAK. Overall our findings provide a mechanism by which haematopoietic FAK controls cancer metastasis.

  16. Fibronectin and atherosclerosis.

    PubMed

    Stenman, S; von Smitten, K; Vaheri, A

    1980-01-01

    Fibronectin is a polymorphic glycoprotein of plasma, other body fluids and connective tissue, and it occurs in an insoluble and a soluble form. Insoluble fibronectin is found associated with basement membranes and in loose connective tissue matrix as well as in the pericellular matrix formed around cultured adherent cells, such as endothelial, fibroblastic and smooth muscle cells. In these positions fibronectin apparently functions as a substrate for cell attachment and as a scaffold for cell migration and movement. Soluble fibronectin, present e.g. in the circulation (300 micronm/ml) exhibits some important interations with other proteins. It is covalently cross-linked to fibrin during thrombus formation and binds to collagen. Fibronectin is released from platelets during their aggregation and soluble fibronectin potentiates the action of plasminogen activator. We have detected fibronectin in the sub-endothelium, in the matrix of smooth muscle cells of the media and in the adventitia of arteries. By using immunohistological techniques we have further found that fibronectin is prominent in atherosclerotic lesions of the intima, especially in developing fibrous plaques. Fibronectin was also prominent in experimentally induced atherosclerotic lesions. These findings suggest that fibronectin is an indicator of connective tissue formation in atherosclerotic processes and that the protein can have a role in their pathogenesis.

  17. MIEN1 drives breast tumor cell migration by regulating cytoskeletal-focal adhesion dynamics

    PubMed Central

    Van Treuren, Timothy; Vishwanatha, Jamboor K.

    2016-01-01

    Migration and invasion enhancer 1 (MIEN1) is an important regulator of cell migration and invasion. MIEN1 overexpression represents an oncogenic event that promotes tumor cell dissemination and metastasis. The underlying mechanism by which MIEN1 regulates migration and invasion has yet to be deciphered. Here, we demonstrate that MIEN1 acts as a cytoskeletal-signaling adapter protein to drive breast cancer cell migration. MIEN1 localization is concentrated underneath the actin-enriched protrusive structures of the migrating breast cancer cells. Depletion of MIEN1 led to the loss of actin-protrusive structures whereas the over-expression of MIEN1 resulted in rich and thick membrane extensions. Knockdown of MIEN1 also decreased the cell-substratum adhesion, suggesting a role for MIEN1 in actin cytoskeletal dynamics. Our results show that MIEN1 supports the transition of G-actin to F-actin polymerization and stabilizes F-actin polymers. Additionally, MIEN1 promotes cellular adhesion and actin dynamics by inducing phosphorylation of FAK at Tyr-925 and reducing phosphorylation of cofilin at Ser-3, which results in breast cancer cell migration. Collectively, our data show that MIEN1 plays an essential role in maintaining the plasticity of the dynamic membrane-associated actin cytoskeleton, which leads to an increase in cell motility. Hence, targeting MIEN1 might represent a promising means to prevent breast tumor metastasis. PMID:27462783

  18. Fibronectin fibril pattern displays the force balance of cell-matrix adhesion.

    PubMed

    Pompe, Tilo; Keller, Kristin; Mitdank, Claudia; Werner, Carsten

    2005-11-01

    Formation of fibrillar patterns of fibronectin on polymer substrates with gradated physicochemical surface properties was analysed during early stages of endothelial cell adhesion. Fibronectin was pre-adsorbed onto three maleic anhydride copolymer thin films with distinct differences in the protein adsorption strength as verified by heteroexchange experiments. The evolved micrometer scale fibrillar patterns of fibronectin on the compared polymer surfaces were characterized after 50 min of cellular reorganization by an auto-correlation analysis using fluorescence microscopy data. Statistical analysis revealed a decrease of the typical spacings of the fibronectin fibrils from 2.6 to 1.8 mum with decreasing fibronectin adsorption strength to the substrate. Size and density of focal adhesions correlated with this dependence of the fibronectin fibril pattern. From these data a model was developed relating the fibronectin fibril pattern to the fibronectin-substrate adsorption strength through the cytoskeletal force regulation mechanism of the cell.

  19. Nanoscale features of fibronectin fibrillogenesis depend on protein-substrate interaction and cytoskeleton structure.

    PubMed

    Pompe, Tilo; Renner, Lars; Werner, Carsten

    2005-01-01

    Cell-reorganized fibronectin layers on polymer films providing a gradation of the binding strength between protein and substrate were analyzed by combined fluorescence and scanning force microscopy. The nanoscale fibronectin patterns exhibited paired parallel fibrils with characteristic spacings of 156, 233, 304, and 373 nm. These spacings depend on the interaction of fibronectin with the substrate: at enhanced fibronectin-substrate anchorage the cells form larger stress fibers, which are assembled by alpha-actinin cross-linked pairs of actin filaments subunits at the focal adhesions. A ubiquitous repeating unit of approximately 71 nm was found within these characteristic distances. We conclude that the dimensions of the actin stress fibers reflect the binding strength of fibronectin to the polymer substrate and act--in turn--as a template for the reorganization of fibronectin into surface-bound nanofibrils with characteristic spacings. This explanation was confirmed by data showing the alpha-actinin/fibronectin colocalization.

  20. Fibrillin Assembly Requires Fibronectin

    PubMed Central

    Sabatier, Laetitia; Chen, Daliang; Fagotto-Kaufmann, Christine; Hubmacher, Dirk; McKee, Marc D.; Annis, Douglas S.; Mosher, Deane F.

    2009-01-01

    Fibrillins constitute the major backbone of multifunctional microfibrils in elastic and nonelastic extracellular matrices. Proper assembly mechanisms are central to the formation and function of these microfibrils, and their properties are often compromised in pathological circumstances such as in Marfan syndrome and in other fibrillinopathies. Here, we have used human dermal fibroblasts to analyze the assembly of fibrillin-1 in dependence of other matrix-forming proteins. siRNA knockdown experiments demonstrated that the assembly of fibrillin-1 is strictly dependent on the presence of extracellular fibronectin fibrils. Immunolabeling performed at the light and electron microscopic level showed colocalization of fibrillin-1 with fibronectin fibrils at the early stages of the assembly process. Protein-binding assays demonstrated interactions of fibronectin with a C-terminal region of fibrillin-1, -2, and -3 and with an N-terminal region of fibrillin-1. The C-terminal half of fibrillin-2 and -3 had propensities to multimerize, as has been previously shown for fibrillin-1. The C-terminal of all three fibrillins interacted strongly with fibronectin as multimers, but not as monomers. Mapping studies revealed that the major binding interaction between fibrillins and fibronectin involves the collagen/gelatin-binding region between domains FNI6 and FNI9. PMID:19037100

  1. Genetics Home Reference: fibronectin glomerulopathy

    MedlinePlus

    ... tiny blood vessels in the kidneys that filter waste products from blood . The waste products are then released in urine. The fibronectin- ... the body attempts to filter it out as waste . Even though there is an abundance of fibronectin- ...

  2. Fibronectin regulates calvarial osteoblast differentiation

    NASA Technical Reports Server (NTRS)

    Moursi, A. M.; Damsky, C. H.; Lull, J.; Zimmerman, D.; Doty, S. B.; Aota, S.; Globus, R. K.

    1996-01-01

    The secretion of fibronectin by differentiating osteoblasts and its accumulation at sites of osteogenesis suggest that fibronectin participates in bone formation. To test this directly, we determined whether fibronectin-cell interactions regulate progressive differentiation of cultured fetal rat calvarial osteoblasts. Spatial distributions of alpha 5 integrin subunit, fibronectin, osteopontin (bone sialoprotein I) and osteocalcin (bone Gla-protein) were similar in fetal rat calvaria and mineralized, bone-like nodules formed by cultured osteoblasts. Addition of anti-fibronectin antibodies to cultures at confluence reduced subsequent formation of nodules to less than 10% of control values, showing that fibronectin is required for normal nodule morphogenesis. Anti-fibronectin antibodies selectively inhibited steady-state expression of mRNA for genes associated with osteoblast differentiation; mRNA levels for alkaline phosphatase and osteocalcin were suppressed, whereas fibronectin, type I collagen and osteopontin were unaffected. To identify functionally relevant domains of fibronectin, we treated cells with soluble fibronectin fragments and peptides. Cell-binding fibronectin fragments (type III repeats 6-10) containing the Arg-Gly-Asp (RGD) sequence blocked both nodule initiation and maturation, whether or not they contained a functional synergy site. In contrast, addition of the RGD-containing peptide GRGDSPK alone did not inhibit nodule initiation, although it did block nodule maturation. Thus, in addition to the RGD sequence, other features of the large cell-binding fragments contribute to the full osteogenic effects of fibronectin. Nodule formation and osteoblast differentiation resumed after anti-fibronectin antibodies or GRGDSPK peptides were omitted from the media, showing that the inhibition was reversible and the treatments were not cytotoxic. Outside the central cell-binding domain, peptides from the IIICS region and antibodies to the N terminus did not

  3. Fibronectin regulates calvarial osteoblast differentiation

    NASA Technical Reports Server (NTRS)

    Moursi, A. M.; Damsky, C. H.; Lull, J.; Zimmerman, D.; Doty, S. B.; Aota, S.; Globus, R. K.

    1996-01-01

    The secretion of fibronectin by differentiating osteoblasts and its accumulation at sites of osteogenesis suggest that fibronectin participates in bone formation. To test this directly, we determined whether fibronectin-cell interactions regulate progressive differentiation of cultured fetal rat calvarial osteoblasts. Spatial distributions of alpha 5 integrin subunit, fibronectin, osteopontin (bone sialoprotein I) and osteocalcin (bone Gla-protein) were similar in fetal rat calvaria and mineralized, bone-like nodules formed by cultured osteoblasts. Addition of anti-fibronectin antibodies to cultures at confluence reduced subsequent formation of nodules to less than 10% of control values, showing that fibronectin is required for normal nodule morphogenesis. Anti-fibronectin antibodies selectively inhibited steady-state expression of mRNA for genes associated with osteoblast differentiation; mRNA levels for alkaline phosphatase and osteocalcin were suppressed, whereas fibronectin, type I collagen and osteopontin were unaffected. To identify functionally relevant domains of fibronectin, we treated cells with soluble fibronectin fragments and peptides. Cell-binding fibronectin fragments (type III repeats 6-10) containing the Arg-Gly-Asp (RGD) sequence blocked both nodule initiation and maturation, whether or not they contained a functional synergy site. In contrast, addition of the RGD-containing peptide GRGDSPK alone did not inhibit nodule initiation, although it did block nodule maturation. Thus, in addition to the RGD sequence, other features of the large cell-binding fragments contribute to the full osteogenic effects of fibronectin. Nodule formation and osteoblast differentiation resumed after anti-fibronectin antibodies or GRGDSPK peptides were omitted from the media, showing that the inhibition was reversible and the treatments were not cytotoxic. Outside the central cell-binding domain, peptides from the IIICS region and antibodies to the N terminus did not

  4. SSX2 regulates focal adhesion but does not drive the epithelial to mesenchymal transition in prostate cancer

    PubMed Central

    Bloom, Jordan E.; McNeel, Douglas G.

    2016-01-01

    Prostate cancer is the most commonly diagnosed malignancy for men in the United States. Metastatic prostate cancer, the lethal form of the disease, has a life expectancy of approximately five years. Identification of factors associated with this transition to metastatic disease is crucial for future therapies. One such factor is the SSX gene family, a family of cancer/testis antigens (CTA) transcription factors which have been shown to be aberrantly expressed in other cancers and associated with the epithelial to mesenchymal transition (EMT). We have previously shown that SSX expression in prostate cancers was restricted to metastatic tissue and not primary tumors. In this study, we have identified SSX2 as the predominant SSX family member expressed in prostate cancer, and found its expression in the peripheral blood of 19 of 54 (35%) prostate cancer patients, with expression restricted to circulating tumor cells, and in 7 of 15 (47%) metastatic cDNA samples. Further, we examined SSX2 function in prostate cancer through knockdown and overexpression in prostate cancer cell lines. While overexpression had little effect on morphology or gene transcript changes, knockdown of SSX2 resulted in an epithelial morphology, increased cell proliferation, increased expression of genes involved in focal adhesion, decreased anchorage independent growth, increased invasion, and increased tumorigenicity in vivo. We conclude from these findings that SSX2 expression in prostate cancer is not a driver of EMT, but is involved in processes associated with EMT including loss of focal adhesion that may be related to tumor cell dissemination. PMID:27276714

  5. Role of β-Pix in corneal epithelial cell migration on fibronectin.

    PubMed

    Kimura, Kazuhiro; Teranishi, Shinichiro; Orita, Tomoko; Zhou, Hongyan; Nishida, Teruo

    2011-05-16

    Corneal epithelial migration during wound healing is important for maintenance of corneal transparency, and fibronectin plays a key role in regulation of the adhesion and migration of corneal epithelial cells. The role of β-Pix in intracellular signaling that underlies the stimulatory effects of fibronectin on the adhesion and migration of corneal epithelial cells was examined. Simian virus 40-transformed human corneal epithelial (HCE) cells were cultured on fibronectin or on bovine serum albumin as a control. The localization and tyrosine phosphorylation of β-Pix were examined by immunofluorescence and immunoprecipitation analyses, respectively. The actin cytoskeleton and focal adhesions were detected by staining of cells with rhodamine-phalloidin and antibodies to phosphotyrosine, respectively. The effects of depletion of β-Pix on HCE cell adhesion and migration on fibronectin were investigated by cell transfection with a small interfering RNA specific for β-Pix mRNA. Fibronectin induced the tyrosine phosphorylation of β-Pix as well as its apparent accumulation at focal adhesions in HCE cells. Depletion of β-Pix inhibited the effects of fibronectin on remodeling of the actin cytoskeleton and the formation of focal adhesions. It also inhibited the migration of HCE cells on fibronectin in an in vitro model of wound healing, but it did not affect cell adhesion to fibronectin. β-Pix contributes to the regulation of the formation of focal adhesions as well as that of cell migration by fibronectin in HCE cells. This protein therefore likely plays an important role in signal transduction underlying corneal epithelial wound healing.

  6. Fibronectin in multiple sclerosis lesions.

    PubMed Central

    Sobel, R. A.; Mitchell, M. E.

    1989-01-01

    Cryostat sections of central nervous system (CNS) tissues of patients with multiple sclerosis (MS) and other CNS diseases were stained with antibodies to fibronectin, a macrophage fibronectin receptor component, fibrin/fibrinogen, and albumin using immunoperoxidase. In active, but not inactive, MS plaques vessel fibronectin was increased (to approximately 57% of Factor VIII+ vessels) over uninvolved MS and normal control white matter (P less than 0.001 for both). Fibronectin was primarily localized to vessel walls and amount of staining correlated with degree of inflammation. Active plaques and necrotic lesions also had extracellular fibronectin and fibrin/ogen. These molecules and the fibronectin receptor were found on macrophages. Albumin was more widely and diffusely distributed in lesions than fibronectin. Thus, in addition to extravasation from damaged vessels, fibronectin may be deposited on or synthesized by endothelial cells and macrophages in the CNS. Fibronectin could facilitate monocyte adhesion to endothelial cell luminal surfaces, promote migration of mononuclear cells, and enhance myelin phagocytosis in MS lesions. Images Figure 1 Figure 2 PMID:2528301

  7. Dynamic Structure of Plasma Fibronectin

    PubMed Central

    Maurer, Lisa M.; Ma, Wenjiang; Mosher, Deane F.

    2016-01-01

    Fibronectin is a large vertebrate glycoprotein that is found in soluble and insoluble forms and involved in diverse processes. Protomeric fibronectin is a dimer of subunits, each of which comprises 29 to 31 modules—12 type I, two type II, and 15-17 type III. Plasma fibronectin is secreted by hepatocytes and circulates in a compact conformation before it binds to cell surfaces, converts to an extended conformation, and is assembled into fibronectin fibrils. Here we review biophysical and structural studies that have shed light on how plasma fibronectin transitions from the compact to the extended conformation. The three types of modules each have a well-organized secondary and tertiary structure as defined by NMR and crystallography and have been likened to “beads on a string”. There are flexible sequences in the N-terminal tail, between the fifth and sixth type I modules, between the first two and last two of the type III modules, and at the C-terminus. Several specific module-module interactions have been identified that likely maintain the compact quaternary structure of circulating fibronectin. The quaternary structure is perturbed in response to binding events, including binding of fibronectin to the surface of vertebrate cells for fibril assembly and to bacterial adhesins. PMID:27185500

  8. THE α4β1 INTEGRIN AND THE EDA DOMAIN OF FIBRONECTIN REGULATE A PROFIBROTIC PHENOTYPE IN DERMAL FIBROBLASTS*

    PubMed Central

    Shinde, Arti V.; Kelsh, Rhiannon; Peters, John H.; Sekiguchi, Kiyotoshi; Van De Water, Livingston; McKeown-Longo, Paula J.

    2015-01-01

    Prompt deposition of fibronectin-rich extracellular matrix is a critical feature of normal development and the host-response to injury. Fibronectin isoforms that include the EDA and EDB domains are prominent in these fibronectin matrices. We now report using human dermal fibroblast cultures that the EDA domain of fibronectin or EDA-derived peptides modeled after the C-C’ loop promote stress fiber formation and myosin-light chain phosphorylation. These changes are accompanied by an increase in fibronectin synthesis and fibrillogenesis. These effects are blocked by pretreating cells with either siRNA or blocking antibody to the α4 integrin. Our data indicate that the interaction between the α4β1 integrin and the EDA domain of fibronectin helps to drive tissue fibrosis by promoting a contractile phenotype and an increase in fibronectin synthesis and deposition. PMID:25433338

  9. Variants of BALB/c 3T3 cells lacking complex gangliosides retain a fibronectin matrix and spread normally on fibronectin-coated substrates

    PubMed Central

    1986-01-01

    Evidence has accumulated that di- and trisialogangliosides are involved in the interaction of cells with fibronectin. We have therefore tested the ability of variants of BALB/c 3T3 deficient in such gangliosides to organize a fibronectin matrix and to spread on fibronectin-coated substrates. Whereas BALB/c 3T3 cells contained gangliosides GM3, GM1, and GD1a, direct chemical analysis showed that five out of six variants isolated contained no detectable GD1a. By the overlaying of thin layer chromatograms of cellular gangliosides with 125I-cholera toxin, these variants were also found to lack ganglioside GM1. In contrast, the sialogalactoprotein profile of these cells, analyzed using an 125I- ricin/SDS polyacrylamide gel overlay technique, was similar to that of the parent cell line. All variants organized an extensive fibronectin matrix comparable to that of BALB/c 3T3, as shown using either immunofluorescence or lactoperoxidase-catalyzed iodination. The variants could also spread on fibronectin-coated substrates and adopt a morphology similar to that of BALB/c 3T3 cells, with little or no difference in the concentration of fibronectin required for 50% cell spreading. Cell spreading of the variants was accompanied by the formation of focal contacts and microfilament bundles, in a manner closely resembling that seen with BALB/c 3T3 cells. Treatment of BALB/c 3T3 cells with neuraminidase, which converts much of the cellular GD1a to GM1, did not affect cell spreading on fibronectin. The results clearly demonstrate that complex gangliosides are not essential for retention of a fibronectin matrix or for spreading on fibronectin- coated substrates. PMID:2935542

  10. Fibronectin Aggregation and Assembly

    PubMed Central

    Ohashi, Tomoo; Erickson, Harold P.

    2011-01-01

    The mechanism of fibronectin (FN) assembly and the self-association sites are still unclear and contradictory, although the N-terminal 70-kDa region (I1–9) is commonly accepted as one of the assembly sites. We previously found that I1–9 binds to superfibronectin, which is an artificial FN aggregate induced by anastellin. In the present study, we found that I1–9 bound to the aggregate formed by anastellin and a small FN fragment, III1–2. An engineered disulfide bond in III2, which stabilizes folding, inhibited aggregation, but a disulfide bond in III1 did not. A gelatin precipitation assay showed that I1–9 did not interact with anastellin, III1, III2, III1–2, or several III1–2 mutants including III1–2KADA. (In contrast to previous studies, we found that the III1–2KADA mutant was identical in conformation to wild-type III1–2.) Because I1–9 only bound to the aggregate and the unfolding of III2 played a role in aggregation, we generated a III2 domain that was destabilized by deletion of the G strand. This mutant bound I1–9 as shown by the gelatin precipitation assay and fluorescence resonance energy transfer analysis, and it inhibited FN matrix assembly when added to cell culture. Next, we introduced disulfide mutations into full-length FN. Three disulfide locks in III2, III3, and III11 were required to dramatically reduce anastellin-induced aggregation. When we tested the disulfide mutants in cell culture, only the disulfide bond in III2 reduced the FN matrix. These results suggest that the unfolding of III2 is one of the key factors for FN aggregation and assembly. PMID:21949131

  11. Circulating Fibronectin Controls Tumor Growth12

    PubMed Central

    von Au, Anja; Vasel, Matthaeus; Kraft, Sabrina; Sens, Carla; Hackl, Norman; Marx, Alexander; Stroebel, Philipp; Hennenlotter, Jörg; Todenhöfer, Tilman; Stenzl, Arnulf; Schott, Sarah; Sinn, Hans-Peter; Wetterwald, Antoinette; Bermejo, Justo Lorenzo; Cecchini, Marco G; Nakchbandi, Inaam A

    2013-01-01

    Fibronectin is ubiquitously expressed in the extracellular matrix, and experimental evidence has shown that it modulates blood vessel formation. The relative contribution of local and circulating fibronectin to blood vessel formation in vivo remains unknown despite evidence for unexpected roles of circulating fibronectin in various diseases. Using transgenic mouse models, we established that circulating fibronectin facilitates the growth of bone metastases by enhancing blood vessel formation and maturation. This effect is more relevant than that of fibronectin produced by endothelial cells and pericytes, which only exert a small additive effect on vessel maturation. Circulating fibronectin enhances its local production in tumors through a positive feedback loop and increases the amount of vascular endothelial growth factor (VEGF) retained in the matrix. Both fibronectin and VEGF then cooperate to stimulate blood vessel formation. Fibronectin content in the tumor correlates with the number of blood vessels and tumor growth in the mouse models. Consistent with these results, examination of three separate arrays from patients with breast and prostate cancers revealed that a high staining intensity for fibronectin in tumors is associated with increased mortality. These results establish that circulating fibronectin modulates blood vessel formation and tumor growth by modifying the amount of and the response to VEGF. Furthermore, determination of the fibronectin content can serve as a prognostic biomarker for breast and prostate cancers and possibly other cancers. PMID:23908593

  12. Localization of fibronectin matrix assembly sites on fibroblasts and endothelial cells.

    PubMed

    Christopher, R A; Kowalczyk, A P; McKeown-Longo, P J

    1997-03-01

    Polymerization of soluble fibronectin into extracellular matrix fibers occurs through the interaction between the amino terminus of fibronectin contained within a 70 kDa fragment and 'matrix assembly sites' on the cell surface. The present studies were performed to localize the 'matrix assembly sites' (defined by 70 kDa binding sites) on newly adherent cells and on cells containing preformed fibronectin matrix. Matrix nucleation sites on newly spread cells were visualized using Texas Red conjugated 70 kDa fragment and were found to colocalize with vinculin and substrate fibronectin fibrils. Cells plated onto vitronectin coated coverslips did not exhibit any 70 kDa binding sites although these cells were well-spread with fully developed focal adhesions. Time course studies indicated that 70 kDa binding sites could be detected on newly adherent cells within 30-40 minutes following cell plating onto fibronectin coated coverslips, prior to the reorganization of substrate fibronectin into fibrils. Similarly, exogenous fibronectin conjugated with Texas Red was also colocalized with vinculin when added to newly adherent cells. The disruption of actin filaments with cytochalasin D both prevented the expression of 70 kDa binding sites and also resulted in the loss of established 70 kDa binding sites on newly spread cells. After 3 days in culture, cells organized an extensive fibronectin matrix and 70 kDa was colocalized with two distinct types of matrix fibronectin fibers: fine linear cell-associated fibers which co-stained with the beta1 integrin and coarse extracellular fibers which did not stain for the beta1 integrin. There was also a third type of fibronectin fiber which was organized into a meshwork structure. There was no localization of either beta1 or 70 kDa to these structures. Treatment of 3-day cells with cytochalasin D resulted in the disruption of cell-matrix fibers and cell-associated 70 kDa binding sites. In contrast, the coarse extracellular matrix fibers

  13. Integrin alpha 8 beta 1 promotes attachment, cell spreading, and neurite outgrowth on fibronectin.

    PubMed Central

    Müller, U; Bossy, B; Venstrom, K; Reichardt, L F

    1995-01-01

    The integrin alpha 8 subunit, isolated by low stringency hybridization, is a novel integrin subunit that associates with beta 1. To identify ligands, we have prepared a function-blocking antiserum to the extracellular domain of alpha 8, and we have established by transfection K562 cell lines that stably express alpha 8 beta 1 heterodimers on the cell surface. We demonstrate here by cell adhesion and neurite outgrowth assays that alpha 8 beta 1 is a fibronectin receptor. Studies on fibronectin fragments using RGD peptides as inhibitors show that alpha 8 beta 1 binds to the RGD site of fibronectin. In contrast to the endogenous alpha 5 beta 1 fibronectin receptor in K562 cells, alpha 8 beta 1 not only promotes cell attachment but also extensive cell spreading, suggesting functional differences between the two receptors. In chick embryo fibroblasts, alpha 8 beta 1 is localized to focal adhesions. We conclude that alpha 8 beta 1 is a receptor for fibronectin and can promote attachment, cell spreading, and neurite outgrowth on fibronectin. Images PMID:7626807

  14. Rho-mediated Contractility Exposes a Cryptic Site in Fibronectin and Induces Fibronectin Matrix Assembly

    PubMed Central

    Zhong, Cuiling; Chrzanowska-Wodnicka, Magdalena; Brown, James; Shaub, Amy; Belkin, Alexey M.; Burridge, Keith

    1998-01-01

    Many factors influence the assembly of fibronectin into an insoluble fibrillar extracellular matrix. Previous work demonstrated that one component in serum that promotes the assembly of fibronectin is lysophosphatidic acid (Zhang, Q., W.J. Checovich, D.M. Peters, R.M. Albrecht, and D.F. Mosher. 1994. J. Cell Biol. 127:1447–1459). Here we show that C3 transferase, an inhibitor of the low molecular weight GTP-binding protein Rho, blocks the binding of fibronectin and the 70-kD NH2-terminal fibronectin fragment to cells and blocks the assembly of fibronectin into matrix induced by serum or lysophosphatidic acid. Microinjection of recombinant, constitutively active Rho into quiescent Swiss 3T3 cells promotes fibronectin matrix assembly by the injected cells. Investigating the mechanism by which Rho promotes fibronectin polymerization, we have used C3 to determine whether integrin activation is involved. Under conditions where C3 decreases fibronectin assembly we have only detected small changes in the state of integrin activation. However, several inhibitors of cellular contractility, that differ in their mode of action, inhibit cell binding of fibronectin and the 70-kD NH2-terminal fibronectin fragment, decrease fibronectin incorporation into the deoxycholate insoluble matrix, and prevent fibronectin's assembly into fibrils on the cell surface. Because Rho stimulates contractility, these results suggest that Rho-mediated contractility promotes assembly of fibronectin into a fibrillar matrix. One mechanism by which contractility could enhance fibronectin assembly is by tension exposing cryptic self-assembly sites within fibronectin that is being stretched. Exploring this possibility, we have found a monoclonal antibody, L8, that stains fibronectin matrices differentially depending on the state of cell contractility. L8 was previously shown to inhibit fibronectin matrix assembly (Chernousov, M.A., A.I. Faerman, M.G. Frid, O.Y. Printseva, and V.E. Koteliansky. 1987

  15. The fibronectin receptor is organized by extracellular matrix fibronectin: implications for oncogenic transformation and for cell recognition of fibronectin matrices.

    PubMed

    Roman, J; LaChance, R M; Broekelmann, T J; Kennedy, C J; Wayner, E A; Carter, W G; McDonald, J A

    1989-06-01

    Cells interact with extracellular fibronectin (FN) via adhesive fibronectin receptors (FNRs) that are members of the very late antigens (VLAs) subgroup of the integrin family. In stationary fibroblasts, the FNR is highly organized and distributed identically to extracellular FN fibrils. However, in highly migratory neural crest cells and embryonic somatic fibroblasts, this organization is lost and the FNR appears diffuse. Similarly, oncogenic transformation typically leads to disorganization of the FN receptor and loss of matrix FN. Two models can account for these observations. First, the FN matrix may organize the FN receptor at extracellular matrix contacts on the cell surface. Motile cells not depositing FN matrices thus lack organized receptors. Alternatively, as the FNR is required for optimal FN matrix assembly, (McDonald, J. A., B. J. Quade, T. J. Broekelmann, R. LaChance, K. Forseman, K. Hasegawa, and S. Akiyama. 1987. J. Biol. Chem. 272:2957-2967; Roman, J. R. M. LaChance, T. J. Broekelmann, C. J. R. Kennedy, E. A. Wayner, W. G. Carter, J. A. McDonald. 1989. J. Cell Biol. 108:2529-2543) and has putative cytoskeletal links, it could be organized from within the cell helping to position newly forming FN fibrils. To study this question, we developed peptide antibodies specifically recognizing the alpha 5 subunit of the FNR. Using these antibodies, we examined the organization of FN and of the FNR in normal, matrix assembly inhibited, and SV40-transformed human fibroblasts. On FN-coated substrates, the FNR is found in focal contacts rather than diffusely on the basal cell surface, suggesting FNR interaction with intracellular components. However, when FN fibrils are deposited, the FNR is co-distributed with these fibrils. Preventing FN matrix assembly prevents organization of the FNR. Moreover, when fibroblasts with well established FN matrices and co-distributed FNR are incubated briefly with monoclonal antibodies that block FNR binding to FN, the FNR is no

  16. Fibronectin modulates osteoblast behavior on Nitinol.

    PubMed

    Muhonen, V; Fauveaux, C; Olivera, G; Vigneron, P; Danilov, A; Nagel, M-D; Tuukkanen, J

    2009-03-01

    We have previously demonstrated that primary rat osteoclasts behave differently when cultured on austenite and martensite Nitinol. In this study, we coated the two phases of Nitinol with plasma fibronectin and studied if this modifies the proliferation and cell cycle of MC3T3-E1 osteoblasts. The influence of the crystalline structure of Nitinol on the remodeling and conformation of fibronectin was also studied. The results on austenite demonstrated that fibronectin was more strongly remodeled and the cells spread better compared with the martensite phase. Interestingly, the conformation of the protein showed no differences between austenite and martensite. In addition, fibronectin improved cell proliferation in both phases, but the effect of fibronectin coating was stronger on the austenite surface. In addition, in both Nitinol phases, the proportion of cells in the G(1) phase was observed to grow in the presence of fibronectin. This could indicate cell differentiation on Nitinol.

  17. Identification of CD9 extracellular domains important in regulation of CHO cell adhesion to fibronectin and fibronectin pericellular matrix assembly.

    PubMed

    Cook, George A; Longhurst, Celia M; Grgurevich, Svetozar; Cholera, Shila; Crossno, Joseph T; Jennings, Lisa K

    2002-12-15

    CD9, a 24-kDa member of the tetraspanin family, influences cellular growth and development, activation, adhesion, and motility. Our investigation focuses on the hypothesis that the CD9 second extracellular loop (EC2) is important in modulating cell adhesive events. Using a Chinese hamster ovary (CHO) cell expression system, we previously reported that CD9 expression inhibited cell adhesion to fibronectin and fibronectin matrix assembly. For the first time, a functional epitope on CD9 EC2 that regulates these processes is described. Binding of mAb7, an EC2-specific anti-CD9 monoclonal antibody, reversed the CD9 inhibitory activity on CHO cell adhesion and fibronectin matrix assembly. This reversal of cell phenotype also was observed in CHO cells expressing CD9 EC2 truncations. Furthermore, our data showed that the EC2 sequence (173)LETFTVKSCPDAIKEVFDNK(192) was largely responsible for the CD9-mediated CHO cell phenotype. Two peptides, (135)K-V(172) (peptide 5b) and (168)P-I(185) (peptide 6a), selectively blocked mAb7 binding to soluble CD9 and to CD9 on intact cells. These active peptides reversed the influence of CD9 expression on CHO cell adhesion to fibronectin. In addition, confocal microscopy revealed that CD9 colocalized with the integrin alpha(5)beta(1) and cytoskeletal F-actin in punctate clusters on the cell surface, particularly at the cell margins. Immunoprecipitation studies confirmed CD9 association with beta(1) integrin. The cellular distribution and colocalization of focal adhesion kinase and alpha-actinin with cytoskeletal actin was also influenced by CD9 expression. Thus, CD9 may exhibit its effect by modulating the composition of adhesive complexes important in facilitating cell adhesion and matrix assembly.

  18. Hemagglutinin activity of human plasma fibronectin.

    PubMed

    Vuento, M

    1979-09-01

    Purified human plasma fibronectin at concentrations of about 30 microgram/ml was found to agglutinate trypsin-treated erythrocytes from certain species. The hemagglutination reaction was inhibited by specific antibodies to fibronectin, by relatively low concentrations of polyamines and by higher concentrations of basic amino acids and nonacetylated amino sugars. The divalent cations Ca2+ and Mg2+ and the chelating agent ethylenediaminetetraacetate did not affect the reaction. None of the neutral amino acids, neutral sugars or polyanions tested was inhibitory. The results imply that plasma fibronectin is capable of interacting with cell surfaces and support the idea of a similarity between cellular and plasma fibronectins.

  19. Fibronectin Matrix Assembly after Spinal Cord Injury

    PubMed Central

    Zhu, Yunjiao; Soderblom, Cynthia; Trojanowsky, Michelle; Lee, Do-Hun

    2015-01-01

    Abstract After spinal cord injury (SCI), a fibrotic scar forms at the injury site that is best characterized by the accumulation of perivascular fibroblasts and deposition of the extracellular matrix protein fibronectin. While fibronectin is a growth-permissive substrate for axons, the fibrotic scar is inhibitory to axon regeneration. The mechanism behind how fibronectin contributes to the inhibitory environment and how the fibronectin matrix is assembled in the fibrotic scar is unknown. By deleting fibronectin in myeloid cells, we demonstrate that fibroblasts are most likely the major source of fibronectin in the fibrotic scar. In addition, we demonstrate that fibronectin is initially present in a soluble form and is assembled into a matrix at 7 d post-SCI. Assembly of the fibronectin matrix may be mediated by the canonical fibronectin receptor, integrin α5β1, which is primarily expressed by activated macrophages/microglia in the fibrotic scar. Despite the pronounced cavitation after rat SCI, fibrotic scar also is observed in a rat SCI model, which is considered to be more similar to human pathology. Taken together, our study provides insight into the mechanism of fibrotic scar formation after spinal cord injury. PMID:25492623

  20. Modulation of fibronectin gene expression in human mononuclear phagocytes.

    PubMed Central

    Yamauchi, K; Martinet, Y; Crystal, R G

    1987-01-01

    Under some conditions, mononuclear phagocytes spontaneously synthesize and release fibronectin, an extracellular matrix glycoprotein with versatile effects on cell-matrix interactions. To gain insight into the processes that modulate the level of fibronectin secretion by these cells, we used monocytes, in vitro matured monocytes and alveolar macrophages as models to compare fibronectin mRNA levels and fibronectin secretion in a variety of circumstances. Using Northern analysis and dot-blot analysis with a 32P-labeled human fibronectin cDNA probe, we evaluated steady-state mRNA levels and a human fibronectin-specific ELISA was used to evaluate fibronectin secretion. In all cases the amounts of fibronectin secreted paralleled fibronectin mRNA levels. Specifically (a) when fibronectin mRNA was undetectable, as in the case of normal blood monocytes, no fibronectin was secreted, but whenever fibronectin mRNA was present, as in normal alveolar macrophages, fibronectin was secreted by the cells; (b) as monocytes matured into macrophages in vitro, the cells began to express fibronectin mRNA and the cells secreted fibronectin; (c) when alveolar macrophages were activated with surface stimuli such as lipopolysaccharide (LPS) or immune complexes, fibronectin mRNA levels decreased and in parallel, the cells secreted less fibronectin; (d) in idiopathic pulmonary fibrosis (IPF), alveolar macrophages contained severalfold more fibronectin mRNA transcripts that normal and the cells spontaneously secreted severalfold more fibronectin than normal; and (e) when IPF alveolar macrophages were placed in culture the fibronectin mRNA levels in the cells decreased with time, and concurrently the amounts of fibronectin produced per unit time continually decreased. The observation of a strict concordance of fibronectin mRNA levels and fibronectin release by mononuclear phagocytes suggests that, at least in many circumstances, fibronectin secretion by mononuclear phagocytes is controlled by

  1. Matrix Gla Protein Binds to Fibronectin and Enhances Cell Attachment and Spreading on Fibronectin

    PubMed Central

    Nishimoto, Satoru Ken; Nishimoto, Miyako

    2014-01-01

    Background. Matrix Gla protein (MGP) is a vitamin K-dependent, extracellular matrix protein. MGP is a calcification inhibitor of arteries and cartilage. However MGP is synthesized in many tissues and is especially enriched in embryonic tissues and in cancer cells. The presence of MGP in those instances does not correlate well with the calcification inhibitory role. This study explores a potential mechanism for MGP to bind to matrix proteins and alter cell matrix interactions. Methods. To determine whether MGP influences cell behavior through interaction with fibronectin, we studied MGP binding to fibronectin, the effect of MGP on fibronectin mediated cell attachment and spreading and immunolocalized MGP and fibronectin. Results. First, MGP binds to fibronectin. The binding site for MGP is in a specific fibronectin fragment, called III1-C or anastellin. The binding site for fibronectin is in a MGP C-terminal peptide comprising amino acids 61–77. Second, MGP enhances cell attachment and cell spreading on fibronectin. MGP alone does not promote cell adhesion. Third, MGP is present in fibronectin-rich regions of tissue sections. Conclusions. MGP binds to fibronectin. The presence of MGP increased cell-fibronectin interactions. PMID:25210519

  2. mRNA expression and protein distribution of fibronectin splice variants and high-molecular weight tenascin-C in different phases of human fracture healing.

    PubMed

    Kilian, O; Dahse, R; Alt, V; Zardi, L; Hentschel, J; Schnettler, R; Kosmehl, H

    2008-08-01

    Fracture healing is a reparative physiological process, which proceeds in stages, each characterized by the predominant tissue in the fracture gap. The tissue matrix is continuously reorganized by cell migration, proliferation, and differentiation. Adhesive proteins such as fibronectin and tenascin transmit information between matrix and cells. As a result of alternative splicing of pre-RNA, EDA + fibronectin, EDB + fibronectin, and high-molecular weight (hm) tenascin-C are generated. By definition, EDB + fibronectin is an oncofetal protein because it is extremely rare in normal adult tissue and plasma, whereas it is expressed in fetal and tumor tissues and during wound healing. In this study, we for the first time describe EDA + fibronectin, EDB + fibronectin, and hm tenascin-C expression in human fracture gap tissue during various stages of differentiation. We demonstrate mRNA expression of all three splice variants in the initial fibrin matrix with upregulation in the enchondral ossification/osteoid and woven bone stages. Of all variants, EDA + fibronectin mRNA has the highest concentration in all stages. For the analysis, we used LightCycler-based relative mRNA quantification and immunohistochemistry. Our data demonstrate that EDA + fibronectin and hm tenascin-C show a diffuse distribution pattern in fracture gap connective tissue, while EDB + fibronectin is focally concentrated in osteoblastic cells at the margins of woven bone. EDA + fibronectin and hm tenascin represent markers for active granulation processes, whereas EDB + fibronectin is specific for cells forming the enchondral and osteoid matrix. The possibility of stimulating fracture healing by EDB + fibronectin-cytokine complexes should be tested in further investigations.

  3. Improved scaffold biocompatibility through anti-Fibronectin aptamer functionalization.

    PubMed

    Galli, C; Parisi, L; Piergianni, M; Smerieri, A; Passeri, G; Guizzardi, S; Costa, F; Lumetti, S; Manfredi, E; Macaluso, G M

    2016-09-15

    Protein adsorption is the first and decisive step to define cell-biomaterial interaction. Guiding the adsorption of desired protein species may represent a viable approach to promote cell activities conducive to tissue regeneration. The aim of the present study was to investigate whether immobilized anti-Fibronectin aptamers could promote the attachment and growth of osteoblastic cells. Polyethyleneglycole diacrylate/thiolated Hyaluronic Acid hydrogels (PEGDA/tHA) were coated with anti-Fibronectin aptamers. Hydrogel loading and Fibronectin bonding were investigated, through spectrophotometry and Bradford assay. Subsequently, human osteoblasts (hOBs) were cultured on hydrogels for 10days in 2D and 3D cultures. Cells were monitored through microscopy and stained for focal adhesions, microfilaments and nuclei using fluorescence microscopy. Samples were also included in paraffin and stained with Hematoxylin-Eosin. Cell number on hydrogels was quantitated over time. Cell migration into the hydrogels was also studied through Calcein AM staining. Aptamers increased the number of adherent hOBs and their cytoplasm appeared more spread and richer in adhesion complexes than on control hydrogels. Viability assays confirmed that significantly more cells were present on hydrogels in the presence of aptamers, already after 48h of culture. When hOBs were encapsulated into hydrogels, cells were more numerous on aptamer-containing PEGDA-tHA. Cells migrated deeper in the gel in the presence of DNA aptamers, appearing on different focus planes. Our data demonstrate that anti-Fibronectin aptamers promote scaffold enrichment for this protein, thus improving cell adhesion and scaffold colonization. We believe aptamer coating of biomaterials is a useful and viable approach to improve the performance of scaffold materials for both research and possibly clinical purposes, because different medical devices could be envisaged able to capture bioactive mediators from the patients' blood and

  4. Syndecan-1 Regulates Cell Migration and Fibronectin Fibril Assembly

    PubMed Central

    Stepp, Mary Ann; Daley, William P.; Bernstein, Audrey M.; Pal-Ghosh, Sonali; Tadvalkar, Gauri; Shashurin, Alexey; Palsen, Sarah; Jurjus, Rosalyn A.; Larsen, Melinda

    2011-01-01

    Corneal scarring is a major cause of blindness worldwide and can result from the deposition of abnormal amounts of collagen fibers lacking the correct size and spacing required to produce a clear cornea. Collagen fiber formation requires a preformed fibronectin (FN) matrix. We demonstrate that the loss of syndecan1 (sdc1) in corneal stromal cells (CSC) impacts cell migration rates, the sizes and composition of focal and fibrillar adhesions, the activation of integrins, and the assembly of fibronectin into fibrils. Integrin and fibronectin expression are not altered on sdc1 null CSCs. Cell adhesion, spreading, and migration studies using low compared to high concentrations of FN and collagen I (CNI) or vitronectin (VN) with and without activation of integrins by manganese chloride show that the impact of sdc1 depletion on integrin activation varies depending on the integrin-mediated activity evaluated. Differences in FN-fibrillogenesis and migration in sdc1 null CSCs are reversed by addition of manganese chloride but cell spreading differences remain. To determine if our findings on sdc1 were specific to the cornea, we compared the phenotypes of sdc1 null dermal fibroblasts with those of CSCs. We found that without sdc1, both cell types migrate faster; however, cell-type specific differences in FN expression and its assembly into fibrils exist between these two cell types. Together, our data demonstrate that sdc1 functions to regulate integrin activity in multiple cell types. Loss of sdc1-mediated integrin function results in cell-type specific differences in matrix assembly. A better understanding of how different cell types regulate FN fibril formation via syndecans and integrins will lead to better treatments for scarring and fibrosis. PMID:20580707

  5. Guidance of Mesenchymal Stem Cells on Fibronectin Structured Hydrogel Films

    PubMed Central

    Kasten, Annika; Naser, Tamara; Brüllhoff, Kristina; Fiedler, Jörg; Müller, Petra; Möller, Martin; Rychly, Joachim; Groll, Jürgen; Brenner, Rolf E.

    2014-01-01

    Designing of implant surfaces using a suitable ligand for cell adhesion to stimulate specific biological responses of stem cells will boost the application of regenerative implants. For example, materials that facilitate rapid and guided migration of stem cells would promote tissue regeneration. When seeded on fibronectin (FN) that was homogeneously immmobilized to NCO-sP(EO-stat-PO), which otherwise prevents protein binding and cell adhesion, human mesenchymal stem cells (MSC) revealed a faster migration, increased spreading and a more rapid organization of different cellular components for cell adhesion on fibronectin than on a glass surface. To further explore, how a structural organization of FN controls the behavior of MSC, adhesive lines of FN with varying width between 10 µm and 80 µm and spacings between 5 µm and 20 µm that did not allow cell adhesion were generated. In dependance on both line width and gaps, cells formed adjacent cell contacts, were individually organized in lines, or bridged the lines. With decreasing sizes of FN lines, speed and directionality of cell migration increased, which correlated with organization of the actin cytoskeleton, size and shape of the nuclei as well as of focal adhesions. Together, defined FN lines and gaps enabled a fine tuning of the structural organization of cellular components and migration. Microstructured adhesive substrates can mimic the extracellular matrix in vivo and stimulate cellular mechanisms which play a role in tissue regeneration. PMID:25329487

  6. Adhesion to fibronectin promotes the activation of the p125FAK/Zap‐70 complex in human T cells

    PubMed Central

    Bearz, A; Tell, G; Formisano, S; Merluzzi, S; Colombatti, A; Pucillo, C

    1999-01-01

    The β1 integrins are a family of heterodimeric adhesion receptors involved in cell‐to‐cell contacts and cell‐to‐extracellular matrix interactions. Through their adhesive role, integrins participate in transduction of outside/inside signals and contribute to trigger a multitude of cellular events such as differentiation, cell activation, and motility. The fibronectin integrin receptors, α4β1 and α5β1, can function as costimulatory molecules in T‐cell receptor (TCR)‐dependent T‐cell activation. In the current study the Jurkat T‐cell line was used as a model system to investigate the TCR‐independent role of cell adhesion to fibronectin in the activation of Zap‐70, a central molecule in the signalling events in T cells. Upon adhesion to plastic immobilized fibronectin but not to bovine serum albumin (BSA) the phosphorylation of p125FAK, a protein kinase that localizes to focal adhesion sites, was induced. Moreover, clustering of fibronectin receptors led to the detection of a p125FAK/Zap‐70 complex. Finally, while the complex between fak‐B, another protein kinase localized to focal adhesion sites, and Zap‐70 was detected in cells plated either on BSA or on fibronectin, the formation of the p125FAK/Zap‐70 complex appeared specifically induced following fibronectin‐mediated integrin clustering. These data suggest the existence of a high degree of specificity when the members of the β1 integrin family mediate signalling pathways in T cells. PMID:10594689

  7. Regulation of fibronectin receptor distribution by transformation, exogenous fibronectin, and synthetic peptides

    PubMed Central

    1986-01-01

    Recent studies have shown that fibronectin and its 140K membrane receptor complex are spatially associated with microfilaments to form cell surface linkage complexes which are thought to mediate adhesive interactions between fibroblasts and their substrata. We examined the regulation of the organization of these cell surface structures in transformed and fibronectin-reconstituted cells as well as in cells treated with a competitive synthetic peptide inhibitor of fibronectin binding to its receptor. Correlative localization experiments with interference reflection microscopy and double-label or triple-label immunofluorescence revealed a concomitant loss of fibronectin, 140K receptor, and alpha-actinin colocalization at cell substratum extracellular matrix contact sites after transformation of chick fibroblasts by wild-type or temperature-sensitive Rous sarcoma viruses (RSV). Western and dot immunoblot analyses established that although similar total quantities of intact 140K molecules were present in the transformed cell cultures, significantly more was released into the culture medium of transformed cells. The 140K molecules on transformed cells were available for interaction with exogenously added fibronectin, which could reconstitute fibronectin-140K linkage complexes. In such fibronectin reconstitution experiments, many cells expressed both fibronectin-140K-actin linkage complexes and RSV pp60src, indicating that the morphological reversion could occur even in the continued presence of RSV transformation. The synthetic peptide Gly-Arg-Gly-Asp-Ser derived from the sequence of the cell-binding region of fibronectin could also prevent the organization of fibronectin-140K linkage complexes. Our results suggest that fibronectin interaction with cells regulates the organization of fibronectin receptor complexes and cytoskeletal components at the cell surface. PMID:3023389

  8. α5β1-integrinpromotes tension-dependent mammary epithelial cell invasion by engaging the fibronectin synergy site.

    PubMed

    Miroshnikova, Y A; Rozenberg, G I; Cassereau, L; Pickup, M; Mouw, J K; Ou, G; Templeman, K L; Elloumi-Hannachi, I; Gooch, K J; Sarang-Sieminski, A L; García, A J; Weaver, V M

    2017-09-06

    Tumors are fibrotic and characterized by abundant, remodeled and cross-linked collagen that stiffens the extracellular matrix stroma. The stiffened collagenous stroma fosters malignant transformation of the tissue by increasing tumor cell tension to promote focal adhesion formation and potentiate growth factor receptor signaling through ERK and PI3 kinase. Importantly, collagen cross-linking requires fibronectin. Fibrotic tumors contain abundant fibronectin and tumor cells frequently up regulate the fibronectin receptor α5β1 integrin.. Using transgenic and xenograft models, and tunable two and three-dimensional substrates we show that fibronectin bound α5β1 integrin promotes tension-dependent malignant transformation through engagement of the synergy site that enhances integrin adhesion force. We determined that ligation of the synergy site of fibronectin permits tumor cells to engage a zyxin-stabilized, vinculin-linked scaffold that facilitates nucleation of PIP3 at the plasma membrane to enhance PI3 kinase-dependent tumor cell invasion. The data explain why rigid collagen fibrils potentiate PI3 kinase activation to promote malignancy and offer a perspective regarding the consistent up regulation of α5β1 integrin and fibronectin in many tumors and their correlation with cancer aggression. © 2017 by The American Society for Cell Biology.

  9. The evolution of tenascins and fibronectin.

    PubMed

    Adams, Josephine C; Chiquet-Ehrismann, Ruth; Tucker, Richard P

    2015-01-01

    Tenascins are extracellular matrix glycoproteins that act both as integrin ligands and as modifiers of fibronectin-integrin interactions to regulate cell adhesion, migration, proliferation and differentiation. In tetrapods, both tenascins and fibronectin bind to integrins via RGD and LDV-type tripeptide motifs found in exposed loops in their fibronectin-type III domains. We previously showed that tenascins appeared early in the chordate lineage and are represented by single genes in extant cephalochordates and tunicates. Here we have examined the genomes of the coelacanth Latimeria chalumnae, the elephant shark Callorhinchus milii as well as the lampreys Petromyzon marinus and Lethenteron japonicum to learn more about the evolution of the tenascin gene family as well as the timing of the appearance of fibronectin during chordate evolution. The coelacanth has 4 tenascins that are more similar to tetrapod tenascins than are tenascins from ray-finned fishes. In contrast, only 2 tenascins were identified in the elephant shark and the Japanese lamprey L. japonicum. An RGD motif exposed to integrin binding is observed in tenascins from many, but not all, classes of chordates. Tetrapods that lack this RGD motif in tenascin-C have a similar motif in the paralog tenascin-W, suggesting the potential for some overlapping function. A predicted fibronectin with the same domain organization as the fibronectin from tetrapods is found in the sea lamprey P. marinus but not in tunicates, leading us to infer that fibronectin first appeared in vertebrates. The motifs that recognize LDV-type integrin receptors are conserved in fibronectins from a broad spectrum of vertebrates, but the RGD integrin-binding motif may have evolved in gnathostomes.

  10. The evolution of tenascins and fibronectin

    PubMed Central

    Adams, Josephine C; Chiquet-Ehrismann, Ruth; Tucker, Richard P

    2015-01-01

    Tenascins are extracellular matrix glycoproteins that act both as integrin ligands and as modifiers of fibronectin-integrin interactions to regulate cell adhesion, migration, proliferation and differentiation. In tetrapods, both tenascins and fibronectin bind to integrins via RGD and LDV-type tripeptide motifs found in exposed loops in their fibronectin-type III domains. We previously showed that tenascins appeared early in the chordate lineage and are represented by single genes in extant cephalochordates and tunicates. Here we have examined the genomes of the coelacanth Latimeria chalumnae, the elephant shark Callorhinchus milii as well as the lampreys Petromyzon marinus and Lethenteron japonicum to learn more about the evolution of the tenascin gene family as well as the timing of the appearance of fibronectin during chordate evolution. The coelacanth has 4 tenascins that are more similar to tetrapod tenascins than are tenascins from ray-finned fishes. In contrast, only 2 tenascins were identified in the elephant shark and the Japanese lamprey L. japonicum. An RGD motif exposed to integrin binding is observed in tenascins from many, but not all, classes of chordates. Tetrapods that lack this RGD motif in tenascin-C have a similar motif in the paralog tenascin-W, suggesting the potential for some overlapping function. A predicted fibronectin with the same domain organization as the fibronectin from tetrapods is found in the sea lamprey P. marinus but not in tunicates, leading us to infer that fibronectin first appeared in vertebrates. The motifs that recognize LDV-type integrin receptors are conserved in fibronectins from a broad spectrum of vertebrates, but the RGD integrin-binding motif may have evolved in gnathostomes. PMID:25482621

  11. Distribution of fibronectin in the rectal mucosa.

    PubMed

    Scott, D L; Morris, C J; Blake, A E; Low-Beer, T S; Walton, K W

    1981-07-01

    Fibronectin is a glycoprotein of high molecular weight present in tissues, plasma, and tissue fluids. Its distribution in the rectal mucosa was studied by immunofluorescent and immunoperoxidase techniques using a monospecific antiserum. Immunofluorescent reactivity for fibronectin was present in the normal rectal mucosa of control subjects in epithelial cells, on basement membranes, and as a loose cribriform network of extracellular reactivity in the lamina propria that codistributed with histochemically demonstrable reticulin. Fibronectin was demonstrated immunoelectromicroscopically on collagen fibres, on smooth muscle cells and within and between columnar epithelial cells. In the rectal mucosa of patients with colitis with marked inflammatory changes, fibronectin appeared thickened and more prominent when present on basement membranes and as sparse strands between inflammatory cells infiltrating the lamina propria. In patients with longstanding colitis and less inflammatory cell infiltration there was a diffuse increase in fibronectin which was densely and uniformly present throughout the lamina propria. Fibronectin is a structural component of the rectal mucosa and changes in its distribution may form an important part of the local reaction to inflammatory bowel disease.

  12. Acetylcholinesterase modulates neurite outgrowth on fibronectin.

    PubMed

    Giordano, C; Poiana, G; Augusti-Tocco, G; Biagioni, S

    2007-05-04

    Acetylcholinesterase (AChE) has been reported to be involved in the modulation of neurite outgrowth. To understand the role played by different domains, we transfected neuroblastoma cells with three constructs containing the invariant region of AChE, differing in the exon encoding the C-terminus and therefore in AChE cellular fate and localization. All isoforms increased neurite extension, suggesting the involvement of the invariant domain [A. De Jaco, G. Augusti-Tocco, S. Biagioni, Alternative AChE molecular forms exhibit similar ability to induce neurite outgrowth, J. Neurosci. Res. 70 (2002) 756-765]. The peripheral anionic site (PAS) is encoded by invariant exons and represents the domain involved in non-cholinergic functions of AChE. Masking of PAS with fasciculin results in a significant decrease of neurite outgrowth in all clones overexpressing AChE. A strong reduction was also observed when clones were cultured on fibronectin. Treatment of clones with fasciculin, therefore masking PAS, abolished the fibronectin-induced reduction. The inhibition of the catalytic site cannot revert the fibronectin effect. Finally, when clones were cultured on fibronectin in the presence of heparin, a ligand of fibronectin, the inhibitory effect was completely reversed. Our results indicate that PAS could directly or indirectly mediate AChE/fibronectin interactions.

  13. Small GTPase Rab21 mediates fibronectin induced actin reorganization in Entamoeba histolytica: implications in pathogen invasion.

    PubMed

    Emmanuel, Merlyn; Nakano, Yumiko Saito; Nozaki, Tomoyoshi; Datta, Sunando

    2015-03-01

    The protozoan parasite Entamoeba histolytica causes a wide spectrum of intestinal infections. In severe cases, the trophozoites can breach the mucosal barrier, invade the intestinal epithelium and travel via the portal circulation to the liver, where they cause hepatic abscesses, which can prove fatal if left untreated. The host Extra Cellular Matrix (ECM) plays a crucial role in amoebic invasion by triggering an array of cellular responses in the parasite, including induction of actin rich adhesion structures. Similar actin rich protrusive structures, known as 'invadosomes', promote chemotactic migration of the metastatic cancer cells and non-transformed cells by remodeling the ECM. Recent studies showed a central role for Rab GTPases, the master regulators of vesicular trafficking, in biogenesis of invadosomes. Here, we showed that fibronectin, a major host ECM component induced actin remodeling in the parasite in a Rab21 dependent manner. The focalized actin structures formed were reminiscent of the mammalian invadosomes. By using various approaches, such as immunofluorescence confocal microscopy and scanning electron microscopy, along with in vitro invasion assay and matrix degradation assay, we show that the fibronectin induced formation of amoebic actin dots depend on the nucleotide status of the GTPase. The ECM components, fibronectin and collagen type I, displayed differential control over the formation of actin dots, with fibronectin positively and collagen type I negatively modulating it. The cell surface adhesion molecule Gal/GalNAc complex was also found to impose additional regulation on this process, which might have implication in collagen type I mediated suppression of actin dots.

  14. High-affinity binding of fibronectin to cultured Kupffer cells

    SciTech Connect

    Cardarelli, P.M.; Blumenstock, F.A.; McKeown-Longo, P.J.; Saba, T.M.; Mazurkiewicz, J.E.; Dias, J.A. )

    1990-11-01

    Hepatic Kupffer cells are a major component of the reticuloendothelial or macrophage system. They were the first phagocytic cell type whose phagocytosis was shown to be influenced by plasma fibronectin, a dimeric opsonic glycoprotein. In the current study, the binding of soluble radioiodinated fibronectin purified from rat serum to isolated rat hepatic Kupffer cells was investigated using a cultured Kupffer cell monolayer technique. Binding was specific, since unlabeled purified fibronectin competed in a dose-dependent manner with the 125I-fibronectin for binding to the Kupffer cells. Addition of gelatin enhanced the binding of 125I-fibronectin to Kupffer cells. The phagocytosis of gelatinized-coated red cells by Kupffer cells was increased either by preopsonizing the target particles with purified fibronectin or by the addition of purified fibronectin to the culture medium. In contrast, exposure of the Kupffer cells to medium containing purified fibronectin followed by wash-removal of the fibronectin did not increase the uptake of gelatin-coated red blood cells, even though fibronectin was detected on the surface of the Kupffer cells by immunofluorescence. Trypsinized monolayers expressed decreased capacity to bind 125I-fibronectin as well as fibronectin-coated sheep erythrocytes. The binding of 125I-fibronectin-gelatin complexes was inhibited by excess unlabeled fibronectin. We calculated that specific high-affinity (Kd = 7.46 x 10(-9) M) binding sites for fibronectin exist on Kupffer cells. There are approximately 2,800-3,500 binding sites or putative fibronectin receptors per Kupffer cell. These sites appear to mediate the enhanced phagocytosis of gelatin-coated particles opsonized by fibronectin.

  15. Fibronectin is a survival factor for differentiated osteoblasts

    NASA Technical Reports Server (NTRS)

    Globus, R. K.; Doty, S. B.; Lull, J. C.; Holmuhamedov, E.; Humphries, M. J.; Damsky, C. H.

    1998-01-01

    The skeletal extracellular matrix produced by osteoblasts contains the glycoprotein fibronectin, which regulates the adhesion, differentiation and function of various adherent cells. Interactions with fibronectin are required for osteoblast differentiation in vitro, since fibronectin antagonists added to cultures of immature fetal calvarial osteoblasts inhibit their progressive differentiation. To determine if fibronectin plays a unique role in fully differentiated osteoblasts, cultures that had already formed mineralized nodules in vitro were treated with fibronectin antagonists. Fibronectin antibodies caused >95% of the cells in the mature cultures to display characteristic features of apoptosis (nuclear condensation, apoptotic body formation, DNA laddering) within 24 hours. Cells appeared to acquire sensitivity to fibronectin antibody-induced apoptosis as a consequence of differentiation, since antibodies failed to kill immature cells and the first cells killed were those associated with mature nodules. Intact plasma fibronectin, as well as fragments corresponding to the amino-terminal, cell-binding, and carboxy-terminal domains of fibronectin, independently induced apoptosis of mature (day-13), but not immature (day-4), osteoblasts. Finally, transforming growth factor-beta1 partially protected cells from the apoptotic effects of fibronectin antagonists. Thus, in the course of maturation cultured osteoblasts switch from depending on fibronectin for differentiation to depending on fibronectin for survival. These data suggest that fibronectin, together with transforming growth factor-beta1, may affect bone formation, in part by regulating the survival of osteoblasts.

  16. Fibronectin Protects from Excessive Liver Fibrosis by Modulating the Availability of and Responsiveness of Stellate Cells to Active TGF-β

    PubMed Central

    Kawelke, Nina; Vasel, Matthaeus; Sens, Carla; von Au, Anja; Dooley, Steven; Nakchbandi, Inaam A.

    2011-01-01

    Fibrotic tissue in the liver is mainly composed of collagen. Fibronectin, which is also present in fibrotic matrices, is required for collagen matrix assembly in vitro. It also modulates the amount of growth factors and their release from the matrix. We therefore examined the effects of the absence of fibronectin on the development of fibrosis in mice. Conditional deletion of fibronectin in the liver using the Mx promoter to drive cre expression resulted in increased collagen production and hence a more pronounced fibrosis in response to dimethylnitrosamine in mice. Exclusive deletion of fibronectin in hepatocytes or normalization of circulating fibronectin in Mx-cKO mice did not affect the development of fibrosis suggesting a role for fibronectin production by other liver cell types. The boosted fibrosis in fibronectin-deficient mice was associated with enhanced stellate cell activation and proliferation, elevated concentrations of active TGF-β, and increased TGF-β-mediated signaling. In vitro experiments revealed that collagen-type-I production by fibronectin-deficient hepatic stellate cells stimulated with TGF-β was more pronounced, and was associated with augmented Smad3-mediated signaling. Interfering with TGF-β signaling using SB431542 normalized collagen-type-I production in fibronectin-deficient hepatic stellate cells. Furthermore, precoating culture plates with fibronectin, but not collagen, or providing fibronectin fibrils unable to interact with RGD binding integrins via the RGD domain significantly diminished the amount of active TGF-β in fibronectin-deficient stellate cells and normalized collagen-type-I production in response to TGF-β stimulation. Thus, excessive stellate cell activation and production of collagen results from increased active TGF-β and TGF-β signaling in the absence of fibronectin. In conclusion, our data indicate that fibronectin controls the availability of active TGF-β in the injured liver, which impacts the severity of

  17. Identification of a fibronectin-binding protein from Staphylococcus epidermidis.

    PubMed

    Williams, Rachel J; Henderson, Brian; Sharp, Lindsay J; Nair, Sean P

    2002-12-01

    Staphylococcus epidermidis has been reported to bind to a number of host cell extracellular matrix proteins, including fibronectin. Here we report the identification of a fibronectin-binding protein from S. epidermidis. A phage display library of S. epidermidis genomic DNA was constructed and panned against immobilized fibronectin. A number of phagemid clones containing overlapping inserts were identified, and one of these clones, pSE109FN, contained a 1.4-kb insert. Phage pSE109FN was found to bind to fibronectin but not to collagen, fibrinogen, laminin, or vitronectin. However, pSE109FN also bound to heparin, hyaluronate, and plasminogen, although to a lesser extent than it bound to fibronectin. Analysis of The Institute for Genomic Research S. epidermidis genome sequence database revealed a 1.85-kb region within a putative 30.5-kb open reading frame, to which the overlapping DNA inserts contained within the fibronectin-binding phagemids mapped. We have designated the gene encoding the fibronectin-binding domain embp. A recombinant protein, Embp32, which encompassed the fibronectin-binding domain of Embp, blocked the binding of S. epidermidis, but not the binding of Staphylococcus aureus, to fibronectin. In contrast, a recombinant protein, FnBPB[D1-D4], spanning the fibronectin-binding domain of the S. aureus fibronectin-binding protein FnBPB, blocked binding of S. aureus to fibronectin but had a negligible effect on the binding of S. epidermidis.

  18. Fibronectin fibrillogenesis on sulfonated polystyrene surfaces.

    PubMed

    Pernodet, Nadine; Rafailovich, Miriam; Sokolov, Jonathan; Xu, D; Yang, Nan-Loh; McLeod, Kenneth

    2003-03-15

    Extracellular matrix (ECM) protein adsorption and organization serves as a critical first step in the development and organization of tissues. Advances in tissue engineering, therefore, will depend on the ability to control the rate and pattern of ECM formation. Fibronectin is a prominent component of the ECM, which undergoes fibrillogenesis in the presence of cells. Using sulfonated polysyrene surfaces, we showed that fibronectin undergoes a transition from monolayer to multilayer adsorption at calculated surface charge densities above 0.03 Coulombs (C)/m(2). At charge densities above approximately 0.08 C/m(2), distinct fibronectin fibrillar networks are observed to form with a fibril morphology similar to those observed to form in situ on cell surfaces. This self-organization process is time dependent, with the fibrils achieving dimensions of 30-40 microm in length and 1 microm in height after 72 h of incubation. We suggest that the polarization of charge domains on the polyampholytic fibronectin molecules near high charge density surfaces is sufficient to initiate the multilayer adsorption and the organization of these fibrillar structures. These results suggest that the nonlinear dependence of adsorption on surface charge density may play an important role in the self-organization of many matrix components.

  19. Binding of fibronectin to Staphylococcus strains.

    PubMed Central

    Switalski, L M; Rydén, C; Rubin, K; Ljungh, A; Höök, M; Wadström, T

    1983-01-01

    Fibronectin, a major protein component of plasma and loose connective tissue has previously been shown to bind to several strains of Staphylococcus aureus. We examined a large number of strains of different species of Staphylococcus with respect to their ability to bind fibronectin. The relative numbers of strains defined as fibronectin-binders among the different species were as follows: S. aureus (22 of 23), S. haemolyticus (5 of 5), S. warneri (8 of 11), S. hyicus (5 of 6), S. hominis (13 of 17), S. saprophyticus (11 of 20), S. epidermidis (4 of 7), and S. simulans (8 of 10). Only three species showed a predominance of nonbinders over binders: S. capitis (4 of 14), S. xylosus (0 of 4), and S. cohnii (3 of 11). These data indicate that staphylococcal species isolated from soft tissue infections frequently have the ability to bind fibronectin and suggest that the ability to bind to this protein may contribute to the virulence of coagulase-positive and coagulase-negative staphylococci. PMID:6315582

  20. The binding of calcium to human fibronectin.

    PubMed

    Amphlett, G W; Hrinda, M E

    1983-03-29

    The binding of calcium to human plasma fibronectin has been measured by equilibrium dialysis at 25 degrees in 0.1 M NaCl 50mM Tris HCL, pH 7.4. Curve fitting of the binding data indicates that fibronectin has two strong calcium binding sites per chain (Mr 220,000), KD = 1.3 mM and approximately 12 weak sites, KD = 2.3 mM. No significant displacement of bound calcium by magnesium was observed at magnesium concentrations up to 1 mM. Calcium binding to a pair of tryptic fragments of fibronectin (Mr congruent to 160,000 and 180,000) that bind to gelatin has also been investigated. These fragments have a single class of calcium binding sites, with 2.2 sites per chain, KD = 1.1 mM. Negligible calcium binding to tryptic fragments derived from other regions of the fibronectin molecule was observed.

  1. Particle Size Influences Fibronectin Internalization and Degradation by Fibroblasts

    NASA Astrophysics Data System (ADS)

    Bozavikov, Peter

    Particle size is a crucial factor that influences the fate and biological impact of particles and their surface proteins upon internalization. Here, using fibronectin-coated polystyrene nanoparticles and microparticles we examined the effect of particle size on degradation of fibronectin. Microparticle uptake depended primarily on beta1 integrins and actin filaments, while nanoparticle uptake relied mainly on lipid rafts and specifically on clathrin-mediated endocytosis. Further, biotinylated fibronectin when coated on microparticles underwent more intracellular processing than fibronectin coated on to nanoparticles. Thus, particle size affects actin and clathrin- dependent internalization, which in turn regulates intracellular fibronectin degradation.

  2. Fibronectin from chicken embryo fibroblasts contains covalently bound phosphate

    PubMed Central

    1979-01-01

    Fibronectin isolated from cultures of chicken embryo fibroblasts (CEF) contains phosphorus linked to serine and threonine by monoester bonds. Normal and Rous sarcoma virus (RSV)-transformed cells were incubated with [32P]orthophosphate, and fibronectin was isolated from the cell surfaces and conditioned media. 32P was stably associated with fibronectin during immunoprecipitation, SDS-polyacrylamide gel electrophoresis, phospholipid solvent extraction, and hot acid but not alkaline treatment. After a limited acid hydrolysis of fibronectin, both phosphoserine and phosphothreonine were found. The specific radioactivity of the 32P-labeled fibronectin from the conditioned medium of normal CEF was higher than that from the cultures of transformed CEF. PMID:457771

  3. Hyaluronan Controls the Deposition of Fibronectin and Collagen and Modulates TGF-β1 Induction of Lung Myofibroblasts

    PubMed Central

    Evanko, Stephen P.; Potter-Perigo, Susan; Petty, Loreen J.; Workman, Gail A.; Wight, Thomas N.

    2015-01-01

    The contribution of hyaluronan-dependent pericellular matrix to TGF-β1-driven induction and maintenance of myofibroblasts is not understood. Hyaluronan is an extracellular matrix (ECM) glycosaminoglycan important in cell adhesion, proliferation and migration, and is implicated in myofibroblast formation and maintenance. Reduced turnover of hyaluronan has been linked to differentiation of myofibroblasts and potentiation of lung fibrosis. Fibronectin is a fibril forming adhesive glycoprotein that is also upregulated following induction with TGF-β1. Although they are known to bind each other, the interplay between hyaluronan and fibronectin in the pericellular matrix during myofibroblast induction and matrix assembly is not clear. This study addresses the role of hyaluronan and its interaction with fibrillar matrix components during myofibroblast formation. Hyaluronan and fibronectin were increased and co-localized in the ECM following myofibroblast induction by TGF-β1. Inhibition of hyaluronan synthesis in TGF-β1-induced lung myofibroblasts over a 4 day period with 4-methyl umbelliferone (4-MU) further enhanced myofibroblast morphology, caused increased deposition of fibronectin and type I collagen in the ECM, and increased expression of alpha-smooth muscle actin and hyaluronan synthase 2 (HAS2) mRNA. Hyaluronan oligosaccarides or hyaluronidase treatment, which more effectively disrupted the pericellular matrix, had similar effects. CD44 and β1 integrins co-localized in the cell membrane and along some stress fibers. However, CD44 and hyaluronan were specifically excluded from focal adhesions, and associated primarily with cortical actin. Time-lapse imaging of the immediate effects of hyaluronidase digestion showed that hyaluronan matrix primarily mediates attachment of membrane and cortical actin between focal contacts, suggesting that surface adhesion through hyaluronan and CD44 is distinct from focal adhesion through β1 integrins and fibronectin. Fluorescein

  4. Upregulation of Fibronectin and the α5β1 and αvβ3 Integrins on Blood Vessels within the Cerebral Ischemic Penumbra

    PubMed Central

    Li, Longxuan; Liu, Fudong; Welser-Alves, Jennifer V.; McCullough, Louise D.; Milner, Richard

    2012-01-01

    Following focal cerebral ischemia, blood vessels in the ischemic border, or penumbra, launch an angiogenic response. In light of the critical role for fibronectin in angiogenesis, and the observation that fibronectin and its integrin receptors are strongly upregulated on angiogenic vessels in the hypoxic CNS, the aim of this study was to establish whether angiogenic vessels in the ischemic CNS also show this response. Focal cerebral ischemia was established in C57/Bl6 mice by middle cerebral artery occlusion (MCA:O), and brain tissue analyzed seven days following re-perfusion, a time at which angiogenesis is ongoing. Within the ischemic core, immunofluorescent (IF) studies demonstrated vascular expression of MECA-32, a marker of leaky cerebral vessels, and vascular breakdown, defined by loss of staining for the endothelial marker, CD31, and the vascular adhesion molecules, laminin, dystroglycan and α6 integrin. Within the ischemic penumbra, dual-IF with CD31 and Ki67 revealed the presence of proliferating endothelial cells, indicating ongoing angiogenesis. Significantly, vessels in the ischemic penumbra showed strong upregulation of fibronectin and the fibronectin receptors, α5β1 and αvβ3 integrins. Taken together with our recent finding that the α5β1 integrin plays an important role in promoting cerebral angiogenesis in response to hypoxia, these results suggest that stimulation of the fibronectin-α5β1 integrin signalling pathway may provide a novel approach to amplifying the intrinsic angiogenic response to cerebral ischemia. PMID:22056225

  5. Regulation of fibronectin gene expression in cardiac fibroblasts by scleraxis.

    PubMed

    Bagchi, Rushita A; Lin, Justin; Wang, Ryan; Czubryt, Michael P

    2016-11-01

    The glycoprotein fibronectin is a key component of the extracellular matrix. By interacting with numerous matrix and cell surface proteins, fibronectin plays important roles in cell adhesion, migration and intracellular signaling. Up-regulation of fibronectin occurs in tissue fibrosis, and previous studies have identified the pro-fibrotic factor TGFβ as an inducer of fibronectin expression, although the mechanism responsible remains unknown. We have previously shown that a key downstream effector of TGFβ signaling in cardiac fibroblasts is the transcription factor scleraxis, which in turn regulates the expression of a wide variety of extracellular matrix genes. We noted that fibronectin expression tracked closely with scleraxis expression, but it was unclear whether scleraxis directly regulated the fibronectin gene. Here, we report that scleraxis acts via two E-box binding sites in the proximal human fibronectin promoter to govern fibronectin expression, with the second E-box being both sufficient and necessary for scleraxis-mediated fibronectin expression to occur. A combination of electrophoretic mobility shift and chromatin immunoprecipitation assays indicated that scleraxis interacted to a greater degree with the second E-box. Over-expression or knockdown of scleraxis resulted in increased or decreased fibronectin expression, respectively, and scleraxis null mice presented with dramatically decreased immunolabeling for fibronectin in cardiac tissue sections compared to wild-type controls. Furthermore, scleraxis was required for TGFβ-induced fibronectin expression: TGFβ lost its ability to induce fibronectin expression following scleraxis knockdown. Together, these results demonstrate a novel and required role for scleraxis in the regulation of cardiac fibroblast fibronectin gene expression basally or in response to TGFβ.

  6. Material-driven fibronectin assembly for high-efficiency presentation of growth factors

    PubMed Central

    Llopis-Hernández, Virginia; Cantini, Marco; González-García, Cristina; Cheng, Zhe A.; Yang, Jingli; Tsimbouri, Penelope M; García, Andrés J.; Dalby, Matthew J.; Salmerón-Sánchez, Manuel

    2016-01-01

    Growth factors (GFs) are powerful signaling molecules with the potential to drive regenerative strategies, including bone repair and vascularization. However, GFs are typically delivered in soluble format at supraphysiological doses because of rapid clearance and limited therapeutic impact. These high doses have serious side effects and are expensive. Although it is well established that GF interactions with extracellular matrix proteins such as fibronectin control GF presentation and activity, a translation-ready approach to unlocking GF potential has not been realized. We demonstrate a simple, robust, and controlled material-based approach to enhance the activity of GFs during tissue healing. The underlying mechanism is based on spontaneous fibrillar organization of fibronectin driven by adsorption onto the polymer poly(ethyl acrylate). Fibrillar fibronectin on this polymer, but not a globular conformation obtained on control polymers, promotes synergistic presentation of integrin-binding sites and bound bone morphogenetic protein 2 (BMP-2), which enhances mesenchymal stem cell osteogenesis in vitro and drives full regeneration of a nonhealing bone defect in vivo at low GF concentrations. This simple and translatable technology could unlock the full regenerative potential of GF therapies while improving safety and cost-effectiveness. PMID:27574702

  7. Small GTPase Rab21 Mediates Fibronectin Induced Actin Reorganization in Entamoeba histolytica: Implications in Pathogen Invasion

    PubMed Central

    Emmanuel, Merlyn; Nakano, Yumiko Saito; Nozaki, Tomoyoshi; Datta, Sunando

    2015-01-01

    The protozoan parasite Entamoeba histolytica causes a wide spectrum of intestinal infections. In severe cases, the trophozoites can breach the mucosal barrier, invade the intestinal epithelium and travel via the portal circulation to the liver, where they cause hepatic abscesses, which can prove fatal if left untreated. The host Extra Cellular Matrix (ECM) plays a crucial role in amoebic invasion by triggering an array of cellular responses in the parasite, including induction of actin rich adhesion structures. Similar actin rich protrusive structures, known as ‘invadosomes’, promote chemotactic migration of the metastatic cancer cells and non-transformed cells by remodeling the ECM. Recent studies showed a central role for Rab GTPases, the master regulators of vesicular trafficking, in biogenesis of invadosomes. Here, we showed that fibronectin, a major host ECM component induced actin remodeling in the parasite in a Rab21 dependent manner. The focalized actin structures formed were reminiscent of the mammalian invadosomes. By using various approaches, such as immunofluorescence confocal microscopy and scanning electron microscopy, along with in vitro invasion assay and matrix degradation assay, we show that the fibronectin induced formation of amoebic actin dots depend on the nucleotide status of the GTPase. The ECM components, fibronectin and collagen type I, displayed differential control over the formation of actin dots, with fibronectin positively and collagen type I negatively modulating it. The cell surface adhesion molecule Gal/GalNAc complex was also found to impose additional regulation on this process, which might have implication in collagen type I mediated suppression of actin dots. PMID:25730114

  8. αV-class integrins exert dual roles on α5β1 integrins to strengthen adhesion to fibronectin

    PubMed Central

    Bharadwaj, Mitasha; Strohmeyer, Nico; Colo, Georgina P.; Helenius, Jonne; Beerenwinkel, Niko; Schiller, Herbert B.; Fässler, Reinhard; Müller, Daniel J.

    2017-01-01

    Upon binding to the extracellular matrix protein, fibronectin, αV-class and α5β1 integrins trigger the recruitment of large protein assemblies and strengthen cell adhesion. Both integrin classes have been functionally specified, however their specific roles in immediate phases of cell attachment remain uncharacterized. Here, we quantify the adhesion of αV-class and/or α5β1 integrins expressing fibroblasts initiating attachment to fibronectin (≤120 s) by single-cell force spectroscopy. Our data reveals that αV-class integrins outcompete α5β1 integrins. Once engaged, αV-class integrins signal to α5β1 integrins to establish additional adhesion sites to fibronectin, away from those formed by αV-class integrins. This crosstalk, which strengthens cell adhesion, induces α5β1 integrin clustering by RhoA/ROCK/myosin-II and Arp2/3-mediated signalling, whereas overall cell adhesion depends on formins. The dual role of both fibronectin-binding integrin classes commencing with an initial competition followed by a cooperative crosstalk appears to be a basic cellular mechanism in assembling focal adhesions to the extracellular matrix. PMID:28128308

  9. Fibronectin Deposition Participates in Extracellular Matrix Assembly and Vascular Morphogenesis.

    PubMed

    Hielscher, Abigail; Ellis, Kim; Qiu, Connie; Porterfield, Josh; Gerecht, Sharon

    2016-01-01

    The extracellular matrix (ECM) has been demonstrated to facilitate angiogenesis. In particular, fibronectin has been documented to activate endothelial cells, resulting in their transition from a quiescent state to an active state in which the cells exhibit enhanced migration and proliferation. The goal of this study is to examine the role of polymerized fibronectin during vascular tubulogenesis using a 3 dimensional (3D) cell-derived de-cellularized matrix. A fibronectin-rich 3D de-cellularized ECM was used as a scaffold to study vascular morphogenesis of endothelial cells (ECs). Confocal analyses of several matrix proteins reveal high intra- and extra-cellular deposition of fibronectin in formed vascular structures. Using a small peptide inhibitor of fibronectin polymerization, we demonstrate that inhibition of fibronectin fibrillogenesis in ECs cultured atop de-cellularized ECM resulted in decreased vascular morphogenesis. Further, immunofluorescence and ultrastructural analyses reveal decreased expression of stromal matrix proteins in the absence of polymerized fibronectin with high co-localization of matrix proteins found in association with polymerized fibronectin. Evaluating vascular kinetics, live cell imaging showed that migration, migration velocity, and mean square displacement, are disrupted in structures grown in the absence of polymerized fibronectin. Additionally, vascular organization failed to occur in the absence of a polymerized fibronectin matrix. Consistent with these observations, we tested vascular morphogenesis following the disruption of EC adhesion to polymerized fibronectin, demonstrating that block of integrins α5β1 and αvβ3, abrogated vascular morphogenesis. Overall, fibronectin deposition in a 3D cell-derived de-cellularized ECM appears to be imperative for matrix assembly and vascular morphogenesis.

  10. Interaction of Fibronectin With Semen Amyloids Synergistically Enhances HIV Infection

    PubMed Central

    Roan, Nadia R.; Chu, Simon; Liu, Haichuan; Neidleman, Jason; Witkowska, H. Ewa; Greene, Warner C.

    2014-01-01

    Semen harbors amyloids that enhance human immunodeficiency virus type 1 (HIV-1) infection. We set out to identify factors that bind these amyloids and to determine whether these factors modulate amyloid-mediated HIV-enhancing activity. Using biochemical and mass spectrometric approaches, we identified fibronectin as a consistent interaction partner. Although monomeric fibronectin did not enhance HIV infection, it synergistically increased the infectivity enhancement activity of the amyloids. Depletion of fibronectin decreased the enhancing activity of semen, suggesting that interfering with the binding interface between fibronectin and the amyloids could be an approach to developing a novel class of microbicides targeting the viral-enhancing activity of semen. PMID:24719472

  11. Sorption of fibronectin to human root surfaces in vitro

    SciTech Connect

    Mendieta, C.; Caravana, C.; Fine, D.H. )

    1990-05-01

    The purpose of this study was to determine the conditions that favor the sorption and retention of human plasma fibronectin to cementum. Rectangular root segments prepared from teeth extracted for orthodontic reasons were mounted on a capillary pipette and immersed in solutions of {sup 125}I fibronectin for assay of cementum sorption under various conditions. Kinetic studies showed sorption to be rapid, with 77% of the maximum fibronectin sorption occurring within 1 minute. Fibronectin sorption was reduced when added in conjunction with serum and was inhibited by monovalent ions (such as sodium), but enhanced in the presence of divalent cations (such as calcium). Exposure of cementum to serum partially blocked subsequent sorption of fibronectin, while cementum bound fibronectin was eluted by subsequent exposure to serum. Treatment of cementum with citric acid pH 1.1 (4 minutes) followed by 5% sodium hypochlorite (5 minutes) caused a significant increase in fibronectin sorption with maximum retention upon subsequent exposure to serum (P less than 0.05). Fibronectin sorption to cementum was: rapid, electrostatic in nature, competitive, reversible, Ca+(+)-facilitated, and maximized by prior treatment of the root with citric acid and sodium hypochlorite. It is concluded that sorption of fibronectin to cementum can be achieved for clinical gain; however, conditions of application can significantly influence both accumulation and subsequent release of root sorbed material.

  12. Focal Choroidal Excavation

    PubMed Central

    Cebeci, Zafer; Bayraktar, Şerife; Oray, Merih; Kır, Nur

    2016-01-01

    Focal choroidal excavation is a choroidal pit that can be detected by optical coherence tomography. Central serous chorioretinopathy, choroidal neovascularization and polypoidal choroidal vasculopathy are pathologies associated with focal choroidal excavation. In this article, we present the follow-up and treatment outcomes of three eyes of two patients with focal choroidal excavation. PMID:28050329

  13. Focal Choroidal Excavation.

    PubMed

    Cebeci, Zafer; Bayraktar, Şerife; Oray, Merih; Kır, Nur

    2016-12-01

    Focal choroidal excavation is a choroidal pit that can be detected by optical coherence tomography. Central serous chorioretinopathy, choroidal neovascularization and polypoidal choroidal vasculopathy are pathologies associated with focal choroidal excavation. In this article, we present the follow-up and treatment outcomes of three eyes of two patients with focal choroidal excavation.

  14. Cartilage proteoglycans inhibit fibronectin-mediated adhesion

    NASA Astrophysics Data System (ADS)

    Rich, A. M.; Pearlstein, E.; Weissmann, G.; Hoffstein, S. T.

    1981-09-01

    Normal tissues and organs show, on histological examination, a pattern of cellular and acellular zones that is characteristic and unique for each organ or tissue. This pattern is maintained in health but is sometimes destroyed by disease. For example, in mobile joints, the articular surfaces consist of relatively acellular hyaline cartilage, and the joint space is enclosed by a capsule of loose connective tissue with a lining of fibroblasts and macrophages. In the normal joint these cells are confined to the synovial lining and the articular surface remains acellular. In in vitro culture, macrophages and their precursor monocytes are very adhesive, and fibroblasts can migrate and overgrow surfaces such as collagen or plastic used for tissue culture. The fibroblasts adhere to collagen by means of fibronectin, which they synthesize and secrete1. Because the collagen of cartilage is capable of binding serum fibronectin2 and fibronectin is present in cartilage during its development3, these cells should, in theory, slowly migrate from the synovial lining to the articular surface. It is their absence from the articular cartilage in normal circumstances, and then presence in such pathological states as rheumatoid arthritis, that is striking. We therefore set out to determine whether a component of cartilage could prevent fibroblast adherence in a defined adhesion assay. As normal cartilage is composed of 50% proteoglycans and 50% collagen by dry weight4, we tested the possibility that the proteoglycans in cartilage inhibit fibroblast adhesion to collagen. We present here evidence that fibroblast spreading and adhesion to collagenous substrates is inhibited by cartilage proteoglycans.

  15. Dualistic nature of adhesive protein function: fibronectin and its biologically active peptide fragments can autoinhibit fibronectin function

    PubMed Central

    1984-01-01

    Fibronectin and certain polypeptide regions of this adhesive glycoprotein mediate cell attachment and spreading on various substrates. We explored the theoretical prediction that this adhesive protein could become a competitive inhibitor of fibronectin-mediated processes if present in solution at appropriately high concentrations. Fibronectin function was inhibited by purified plasma fibronectin at 5- 10 mg/ml, by a 75,000-dalton cell-interaction fragment of the protein at 0.5-1 mg/ml, and even by two synthetic peptides containing a conserved, hydrophilic amino acid sequence at 0.1-0.5 mg/ml. Inhibition of fibronectin-dependent cell spreading was dose dependent, noncytotoxic, and reversible. It was competitive in nature, since increased quantities of substrate-adsorbed fibronectin or longer incubation periods decreased the inhibition. A peptide inhibitory for fibronectin-mediated cell spreading also inhibited fibronectin-mediated attachment of cells to type I collagen, but it did not affect concanavalin A-mediated spreading. These results demonstrate the potential of a cell adhesion molecule and its biologically active peptide fragments to act as competitive inhibitors, and they suggest that fibronectin may act by binding to a saturable cell surface receptor. PMID:6736130

  16. A Microaffinity Assay for Plasma Fibronectin.

    DTIC Science & Technology

    1983-03-25

    employed as the source of fibronectin. The purification procedure was a modification of the method of Vuento and Vaheri (15) which involves affinity...Coated Beads. The gelatin-coated Sepharose 4B beads were prepared by a minor modification of the method of Vuento and Vaheri (15). Cyanogen bromide...Soc. 23, 119-134. 8. Ruoslahti, E., Vuento , M., and Engvall, E. (1978) Biochim. Biophys. Acta 534, 210-218. 9. Todd-Kulikowski, H.D. and Parsons, R.G

  17. Molecular architecture of native fibronectin fibrils

    PubMed Central

    Früh, Susanna Maria; Schoen, Ingmar; Ries, Jonas; Vogel, Viola

    2015-01-01

    Fibronectin fibrils within the extracellular matrix play central roles in physiological and pathological processes, yet many structural details about their hierarchical and molecular assembly remain unknown. Here we combine site-specific protein labelling with single-molecule localization by stepwise photobleaching or direct stochastic optical reconstruction microscopy (dSTORM), and determine the relative positions of various labelled sites within native matrix fibrils. Single end-labelled fibronectin molecules in fibrils display an average end-to-end distance of ∼133 nm. Sampling of site-specific antibody epitopes along the thinnest fibrils (protofibrils) shows periodic punctate label patterns with ∼95 nm repeats and alternating N- and C-terminal regions. These measurements suggest an antiparallel 30–40 nm overlap between N-termini, suggesting that the first five type I modules bind type III modules of the adjacent molecule. Thicker fibres show random bundling of protofibrils without a well-defined line-up. This super-resolution microscopy approach can be applied to other fibrillar protein assemblies of unknown structure. PMID:26041410

  18. Molecular architecture of native fibronectin fibrils

    NASA Astrophysics Data System (ADS)

    Früh, Susanna Maria; Schoen, Ingmar; Ries, Jonas; Vogel, Viola

    2015-06-01

    Fibronectin fibrils within the extracellular matrix play central roles in physiological and pathological processes, yet many structural details about their hierarchical and molecular assembly remain unknown. Here we combine site-specific protein labelling with single-molecule localization by stepwise photobleaching or direct stochastic optical reconstruction microscopy (dSTORM), and determine the relative positions of various labelled sites within native matrix fibrils. Single end-labelled fibronectin molecules in fibrils display an average end-to-end distance of ~133 nm. Sampling of site-specific antibody epitopes along the thinnest fibrils (protofibrils) shows periodic punctate label patterns with ~95 nm repeats and alternating N- and C-terminal regions. These measurements suggest an antiparallel 30-40 nm overlap between N-termini, suggesting that the first five type I modules bind type III modules of the adjacent molecule. Thicker fibres show random bundling of protofibrils without a well-defined line-up. This super-resolution microscopy approach can be applied to other fibrillar protein assemblies of unknown structure.

  19. Treponema pallidum receptor binding proteins interact with fibronectin

    PubMed Central

    1983-01-01

    Analysis of plasma proteins avidly bound to T. pallidum surfaces revealed the ability of T. pallidum to acquire numerous host macromolecules. No acquisition was evident by the avirulent spirochete, T. phagedenis biotype Reiter. Western blotting technology using hyperimmune antifibronectin serum as a probe revealed the ability of virulent treponemes to avidly bind fibronectin from a complex medium such as plasma. The specificity of the tiplike adherence of motile T. pallidum to fibronectin-coated glass surfaces and to fibronectin on HEp- 2 cells was reinforced by the observation that pretreatment of coverslips or cell monolayers with monospecific antiserum against fibronectin substantially reduced T. pallidum attachment. The stoichiometric binding of T. pallidum to fibronectin-coated coverslips and the inability of unlabeled or 35S-radiolabeled treponemes to interact with glass surfaces treated with other plasma proteins further established the specific nature of the interaction between virulent T. pallidum and fibronectin. The avid association between three outer envelope proteins of T. pallidum and fibronectin was also demonstrated. These treponemal surface proteins have been previously identified as putative receptor-binding proteins responsible for T. pallidum parasitism of host cells. The data suggest that surface fibronectin mediates tip-oriented attachment of T. pallidum to host cells via a receptor-ligand mechanism of recognition. PMID:6304227

  20. Enhanced endotoxin effects in plasma fibronectin-deficient rats.

    PubMed Central

    Yoder, M C; Kilpatrick-Smith, L; Arbittier, D; Douglas, S D; Polin, R A

    1987-01-01

    Immunoreactive plasma fibronectin depletion has been associated with the presence of collagen-fibronectin complexes in patients after trauma and in animal models of traumatic and burn injuries. However, the role of plasma fibronectin in the development of sepsis after traumatic and burn injuries in patients is unknown. Treatment of patients and animals with purified human plasma fibronectin ameliorates some of the clinical and metabolic effects of systemic endotoxemia. We report that the induction of immunoreactive plasma fibronectin deficiency by gelatin infusion is associated with enhanced effects of intraperitoneal Escherichia coli endotoxin injection. We have observed a significant increase in the concentrations of ammonia in plasma of treated rats compared with those in control rats administered the same dose of endotoxin. PMID:3298066

  1. Cadherin-11 localizes to focal adhesions and promotes cell–substrate adhesion

    PubMed Central

    Langhe, Rahul P.; Gudzenko, Tetyana; Bachmann, Michael; Becker, Sarah F.; Gonnermann, Carina; Winter, Claudia; Abbruzzese, Genevieve; Alfandari, Dominique; Kratzer, Marie-Claire; Franz, Clemens M.; Kashef, Jubin

    2016-01-01

    Cadherin receptors have a well-established role in cell–cell adhesion, cell polarization and differentiation. However, some cadherins also promote cell and tissue movement during embryonic development and tumour progression. In particular, cadherin-11 is upregulated during tumour and inflammatory cell invasion, but the mechanisms underlying cadherin-11 stimulated cell migration are still incompletely understood. Here, we show that cadherin-11 localizes to focal adhesions and promotes adhesion to fibronectin in Xenopus neural crest, a highly migratory embryonic cell population. Transfected cadherin-11 also localizes to focal adhesions in different mammalian cell lines, while endogenous cadherin-11 shows focal adhesion localization in primary human fibroblasts. In focal adhesions, cadherin-11 co-localizes with β1-integrin and paxillin and physically interacts with the fibronectin-binding proteoglycan syndecan-4. Adhesion to fibronectin mediated by cadherin-11/syndecan-4 complexes requires both the extracellular domain of syndecan-4, and the transmembrane and cytoplasmic domains of cadherin-11. These results reveal an unexpected role of a classical cadherin in cell–matrix adhesion during cell migration. PMID:26952325

  2. Haemodynamics Regulate Fibronectin Assembly via PECAM

    PubMed Central

    Chen, Zhongming; Givens, Chris; Reader, John S.; Tzima, Ellie

    2017-01-01

    Fibronectin (FN) assembly and fibrillogenesis are critically important in both development and the adult organism, but their importance in vascular functions is not fully understood. Here we identify a novel pathway by which haemodynamic forces regulate FN assembly and fibrillogenesis during vascular remodelling. Induction of disturbed shear stress in vivo and in vitro resulted in complex FN fibril assembly that was dependent on the mechanosensor PECAM. Loss of PECAM also inhibited the cell-intrinsic ability to remodel FN. Gain- and loss-of-function experiments revealed that PECAM-dependent RhoA activation is required for FN assembly. Furthermore, PECAM−/− mice exhibited reduced levels of active β1 integrin that were responsible for reduced RhoA activation and downstream FN assembly. These data identify a new pathway by which endothelial mechanotransduction regulates FN assembly and flow-mediated vascular remodelling. PMID:28120882

  3. Fibronectin synthesized by a human hepatoma cell line

    SciTech Connect

    Glasgow, J.E.; Colman, R.W.

    1984-07-01

    Fibronectin is a family of immunologically similar glycoproteins which mediate a variety of cell-cell and cell-substratum interactions. It is a constituent of the extracellular matrix of connective tissue and circulates in plasma. When suspension and adherent cultures of a human hepatoma cell line (SK-HEP-1) were incubated in serum-free medium, the resulting conditioned medium contained material which was specifically immunoprecipitated by antisera to human plasma fibronectin. By double immunodiffusion, a component in the conditioned culture medium was shown to form a line of identity with fibronectin in human plasma and to migrate as an alpha 2- to beta-globulin during immunoelectrophoresis. Human fibronectin was quantified in conditioned medium by electroimmunodiffusion, and was found to increase for at least three days at about 0.1 micrograms/10(6) cells/day. Adherent cultures of SK-HEP-1 cells were incubated with L-(/sup 35/S)methionine to label newly synthesized proteins. Labeled fibronectin in conditioned medium or in cell extracts comigrated with fibronectin in human plasma as shown by autoradiography following crossed-immunoelectrophoresis. Fibronectin was demonstrated in the extra-cellular matrix of adherent SK-HEP-1 cultures by immunofluorescence. It was shown previously that SK-HEP-1 cells synthesize alpha 1-protease inhibitor, one of the products of normal hepatocytes. The finding that these hepatoma cells also synthesize fibronectin supports the concept that the hepatocyte may be one source of circulating fibronectin, a possibility consistent with the established role of this cell type in blood plasma protein synthesis.

  4. MT1-MMP regulates the turnover and endocytosis of extracellular matrix fibronectin.

    PubMed

    Shi, Feng; Sottile, Jane

    2011-12-01

    The extracellular matrix (ECM) is dynamically remodeled by cells during development, normal tissue homeostasis and in a variety of disease processes. We previously showed that fibronectin is an important regulator of ECM remodeling. The deposition and/or polymerization of fibronectin into the ECM controls the deposition and stability of other ECM molecules. In addition, agents that inhibit fibronectin polymerization promote the turnover of fibronectin fibrils and enhance ECM fibronectin endocytosis and intracellular degradation. Endocytosis of ECM fibronectin is regulated by β1 integrins, including α5β1 integrin. We have examined the role of extracellular proteases in regulating ECM fibronectin turnover. Our data show that membrane type matrix metalloproteinase 1 (MT1-MMP; also known as MMP14) is a crucial regulator of fibronectin turnover. Cells lacking MT1-MMP show reduced turnover and endocytosis of ECM fibronectin. MT1-MMP regulates ECM fibronectin remodeling by promoting extracellular cleavage of fibronectin and by regulating α5β1-integrin endocytosis. Our data also show that fibronectin polymerization stabilizes fibronectin fibrils and inhibits ECM fibronectin endocytosis by inhibiting α5β1-integrin endocytosis. These data are the first to show that an ECM protein and its modifying enzyme can regulate integrin endocytosis. These data also show that integrin trafficking plays a major role in modulating ECM fibronectin remodeling. The dual dependence of ECM fibronectin turnover on extracellular proteolysis and endocytosis highlights the complex regulatory mechanisms that control ECM remodeling to ensure maintenance of proper tissue function.

  5. A fibronectin mimetic motif improves integrin mediated cell biding to recombinant spider silk matrices.

    PubMed

    Widhe, Mona; Shalaly, Nancy Dekki; Hedhammar, My

    2016-01-01

    The cell binding motif RGD is the most widely used peptide to improve cell binding properties of various biomaterials, including recombinant spider silk. In this paper we use genetic engineering to further enhance the cell supportive capacity of spider silk by presenting the RGD motif as a turn loop, similar to the one found in fibronectin (FN), but in the silk stabilized by cysteines, and therefore denoted FNCC. Human primary cells cultured on FNCC-silk showed increased attachment, spreading, stress fiber formation and focal adhesions, not only compared to RGD-silk, but also to silk fused with linear controls of the RGD containing motif from fibronectin. Cell binding to FNCC-silk was shown to involve the α5β1 integrin, and to support proliferation and migration of keratinocytes. The FNCC-silk protein allowed efficient assembly, and could even be transformed into free standing films, on which keratinocytes could readily form a monolayer culture. The results hold promise for future applications within tissue engineering. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. The relationship between force and focal complex development

    PubMed Central

    Galbraith, Catherine G.; Yamada, Kenneth M.; Sheetz, Michael P.

    2002-01-01

    To adhere and migrate, cells must be capable of applying cytoskeletal force to the extracellular matrix (ECM) through integrin receptors. However, it is unclear if connections between integrins and the ECM are immediately capable of transducing cytoskeletal contraction into migration force, or whether engagement of force transmission requires maturation of the adhesion. Here, we show that initial integrin–ECM adhesions become capable of exerting migration force with the recruitment of vinculin, a marker for focal complexes, which are precursors of focal adhesions. We are able to induce the development of focal complexes by the application of mechanical force to fibronectin receptors from inside or outside the cell, and we are able to extend focal complex formation to vitronectin receptors by the removal of c-Src. These results indicate that cells use mechanical force as a signal to strengthen initial integrin–ECM adhesions into focal complexes and regulate the amount of migration force applied to individual adhesions at localized regions of the advancing lamella. PMID:12446745

  7. Small Molecules Antagonise the MIA-Fibronectin Interaction in Malignant Melanoma

    PubMed Central

    Yip, King Tuo; Zhong, Xue Yin; Seibel, Nadia; Pütz, Stefanie; Autzen, Jasmin; Gasper, Raphael; Hofmann, Eckhard; Scherkenbeck, Jürgen; Stoll, Raphael

    2016-01-01

    Melanoma inhibitory activity (MIA), an extracellular protein highly expressed by malignant melanoma cells, plays an important functional role in melanoma development, progression, and metastasis. After its secretion, MIA directly interacts with extracellular matrix proteins, such as fibronectin (FN). By this mechanism, MIA actively facilitates focal cell detachment from surrounding structures and strongly promotes tumour cell invasion and migration. Hence, the molecular understanding of MIA’s function provides a promising target for the development of new strategies in malignant melanoma therapy. Here, we describe for the first time the discovery of small molecules that are able to disrupt the MIA-FN complex by selectively binding to a new druggable pocket, which we could identify on MIA by structural analysis and fragment-based screening. Our findings may inspire novel drug discovery efforts aiming at a therapeutically effective treatment of melanoma by targeting MIA. PMID:27151361

  8. Avian pathogenic Escherichia coli bind fibronectin and laminin.

    PubMed

    Ramírez, Rosa María; Almanza, Yolanda; González, Rafael; García, Santos; Heredia, Norma

    2009-04-01

    Avian colisepticemia frequently occurs after respiratory tract damage, the primary site for infection allows bacteria to encounter an exposed basement membrane, where laminin and fibronectin are important components. We investigated the ability of an isolate of avian pathogenic Escherichia coli to bind fibronectin and laminin. Using Far-western dot blot analysis, we demonstrated the ability of this microorganism to bind basement membrane proteins fibronectin and laminin. Results from an ELISA-based approach indicate that the binding to these membrane proteins was bacterial-dose dependent. Furthermore, two specific E. coli polypeptides, of 32 kDa and 130 kDa, reacted with laminin and fibronectin, respectively. Further evaluation of these potential bacterial adhesins may provide insights into the pathogenesis of colibacillosis.

  9. The fiber diameter of synthetic bioresorbable extracellular matrix influences human fibroblast morphology and fibronectin matrix assembly.

    PubMed

    Hsia, Henry C; Nair, Mohan R; Mintz, R Candida; Corbett, Siobhan A

    2011-06-01

    The ideal scaffold material should provide immediate capacity to bear mechanical loads and also permit eventual resorption and replacement with native tissue of similar mechanical integrity. Scaffold characteristics such as fiber diameter provide environmental cues that can influence cell function and differentiation. In this study, the impact of fiber diameter of scaffolds constructed from a tyrosine-based bioresorbable polymer on cellular response was investigated. Electrospun bioresorbable poly(desamino tyrosyl-tyrosine ethyl ester carbonate) scaffolds composed of microfibers or nanofibers were constructed and seeded with human dermal fibroblasts. The impact of fiber diameter on actin cytoskeletal morphology, focal adhesion size, fibronectin matrix assembly, and cell proliferation was evaluated using immunofluorescent microscopy and computer-assisted image analysis. Actin stress fibers were more easily observed in cells on microfiber scaffolds compared with those on nanofiber scaffolds. Cells on nanofiber scaffolds developed smaller focal adhesion complexes compared with those on microfiber scaffolds (p < 0.0001). The temporal patterns of fibronectin matrix assembly were affected by scaffold fiber diameter, with cells on microfiber scaffolds showing a delayed response in dense fibril formation compared with nanofiber scaffolds. Cells on nanofiber scaffolds showed higher proliferation compared with microfiber scaffolds at time points under 1 week (p < 0.01), but by 2 weeks significantly higher cell proliferation was observed on microfiber scaffolds (p < 0.01). The fiber diameter of bioresorbable scaffolds can significantly influence cell response and suggests that the ability of scaffolds to elicit consistent biological responses depends on factors beyond scaffold composition. Such findings have important implications for the design of clinically useful engineered constructs.

  10. Lung vascular injury with protease infusion. Relationship to plasma fibronectin.

    PubMed Central

    Cohler, L F; Saba, T M; Lewis, E P

    1985-01-01

    Fibronectin exists in a soluble form in plasma and in an insoluble form in tissues. Plasma fibronectin can modulate phagocytic function as well as incorporate into the tissue matrix where it is believed to influence microvascular integrity and tissue repair. The temporal alterations in plasma and lung lymph fibronectin were studied in relation to increased pulmonary vascular permeability induced by protease infusion. The acute sheep lung lymph fistula model was used. A 39% decrease in plasma fibronectin (control = 421 +/- 67 micrograms/ml) was observed 2.5 hours (255 +/- 43 micrograms/ml) after protease infusion. There was an elevation of lymph fibronectin early after protease infusion, followed by a progressive decline. Concomitant with the decrease in plasma fibronectin, an increase in lymph flow (QL) of greater than 200% (from a control of 6.7 +/- 1.0 ml/hr to 13.9 +/- 1.4 ml/hr) was observed within 2.5 hours. Also, there was a sustained elevation in the total protein lymph/plasma concentration (L/P) ratio, which was maximal at 2.5 hours. The transvascular protein clearance (TVPC = QL X L/P) was 4.5 +/- 0.7 ml/hr at the control period and 13.1 +/- 2.0 ml/hr by 2.5 hours. This was indicative of increased flux of protein-rich fluid across the pulmonary endothelial barrier. Lung vascular permeability stabilized after 2.5 hours as manifested by a slowly declining L/P ratio. Thus, plasma fibronectin deficiency may contribute to the etiology of increased lung vascular permeability with protease infusion. Since the progressive decline in plasma fibronectin was not reflected in a proportional increase in lymph fibronectin, plasma fibronectin may have sequestered in tissues such as the lung, or perhaps in reticuloendothelial cells during the injury phase. Whether the progressive decrease in plasma fibronectin reflects its incorporation into the endothelial barrier matrix where it may mediate stabilization of the pulmonary microvascular barrier remains to be determined

  11. Generation of fibronectin receptors on macrophages by wheat germ lectin.

    PubMed

    Hörmann, H; Jelinić, V; Richter, H

    1983-08-01

    A chymotrypsin-derived and 125I-labelled 125-kDa fragment of human plasma fibronectin which contained the cell binding site, was only weakly bound by peritoneal macrophages of guinea pigs and binding was not saturable. In presence of wheat germ lectin binding increased proportionally to the logarithm of the lectin concentration. Association of 125I-fragment with cells was partially prevented by non-labelled fragment indicating a saturable receptor-ligand interaction. An apparent affinity constant of about 2--4 x 10(-5) M was evaluated. A considerable fraction of the cell-bound 125I-fragment resisted removal by proteases suggesting that it was internalized. In order to investigate an influence of wheat germ lectin on the binding of 125I-fibronectin by the cells the macrophages were preincubated with the lectin followed by washing and evaluation of 125I-fibronectin binding. A simultaneous incubation of the cells with 125I-fibronectin and lectin was impractical due to partial interaction of the two proteins giving rise to some unspecific precipitates. Preincubation with wheat germ lectin considerably improved the capacity of the macrophages for binding of 125I-fibronectin. Again the binding of 125I-labelled protein could be restricted by unlabelled one. N-acetyl-glucosamine inhibited the binding of 125I-fibronectin by wheat germ lectin-treated cells if applied in the preincubation phase and more effectively, if applied in the final 125I-fibronectin binding assay. N-Acetylneuraminic acid also inhibited this step. In addition to wheat germ lectin concanavalin A was capable of generating fibronectin receptors on the cell surface. Soy bean lectin, however, was ineffective.

  12. Fibronectin Assembly in the Crypts of Cytokinesis-Blocked Multilobular Cells Promotes Anchorage-Independent Growth

    PubMed Central

    Gupta, Rajesh Kumar; Johansson, Staffan

    2013-01-01

    Anchorage-independent growth is a characteristic feature of cancer cells. However, it is unclear whether it represents a cause or a consequence of tumorigenesis. For normal cells, integrin-mediated adhesion is required for completion of the G1 and cytokinesis stages of the cell cycle. This study identified a mechanism that can drive anchorage-independent growth if the G1 checkpoint is suppressed. Cells with defective G1 checkpoint progressed through several rounds of the cell cycle in suspension in spite of uncompleted cytokinesis, thereby forming bi- and multilobular cells. Aurora B and CEP55 were localized to midbodies between the lobes, suggesting that the cytokinesis process reached close to abscission. Integrin-mediated re-attachment of such cells induced cytokinesis completion uncoupled from karyokinesis in most cells. However, a portion of the cells instead lost the constriction and became binucleated. Also, long-term suspension culture in soft agar produced colonies where the cytokinesis block was overcome. This process was fibronectin-dependent since fibronectin-deficient cells did not form colonies unless fibronectin was expressed or exogenously added. While fibronectin normally is not deposited on non-adherent single cells, bi/multilobular cells accumulated fibronectin in the intussusceptions. Based on our data we conclude: 1) Suppression of the G1 checkpoint allows multiple rounds of the cell cycle in detached cells and thereby enables matrix formation on their surface. 2) Uncompleted cytokinesis due to cell detachment resumes if integrin interactions are re-formed, allowing colony formation in soft agar 3) Such delayed cell division can generate binucleated cells, a feature known to cause chromosomal instability. PMID:23951336

  13. Bimaxillary Oral Focal Mucinosis.

    PubMed

    Yadav, Sunil; Malik, Sunita; Mittal, Hitesh Chander; Singh, Gurdarshan; Kamra, Hemlata

    2016-10-01

    Oral focal mucinosis is considered as oral counterpart of cutaneous focal mucinosis. The preoperative diagnosis of mucinosis is almost impossible because of its rarity and clinical similarity to other lesions of various etiologies. The histological diagnosis of oral mucinosis is important to better understand the etiopathogenesis, treatment modalities, and any recurrence of the lesion besides differentiating from the other soft tissue lesions.The purpose of this paper is to report the first case of bimaxillary involvement with dome-shaped elevated, rounded, asymptomatic, normally colored swelling in left posterior palatal mucosa and left mandibular posterior region in a 25-year old woman who was diagnosed as oral focal mucinosis histopathologically.

  14. Involvement of rho-gtpases in fibroblast adhesion and fibronectine fibrillogenesis under stretch

    NASA Astrophysics Data System (ADS)

    Guignandon, A.; Lambert, C.; Rattner, A.; Servotte, S.; Lapiere, C.; Nusgens, B.; Vico, L.

    The Rho family small GTPases play a crucial role in mediating cellular adaptation to mechanical stimulation (MS), and possibly to microgravity (μg), through effects on the cytoskeleton and cell adhesion which is, in turn, mainly regulated by fibronectin fibrillogenesis (FnF). It remains unclear how mechanical stimulation is transduced to the Rho signaling pathways and how it impacts on fibronectin (fbn) fibrillogenesis (FnF). μg (2 days, mission STS-095) led to de-adhesion of fibroblasts and modification of the underlying extracellular matrix. To determine whether GTPases modulated FnF, we generated stable cell lines expressing high level of activated RhoA and Rac1 (QL) as compared to wild type (WI26-WT). After MS application [8% deformation, 1Hz, 15 min., 3 times/day for 1-2 days], we quantified focal adhesion (vinculin, paxillin, FAKY397), f-actin stress fibers (Sf) and FnF with home-developed softwares. We reported that after MS, Sf are more rapidly (30min) formed under the nucleus in Wi26-WT (+100%) and Rac1 (+200%) than in RhoA (+20%). Vinculin & paxillin were only restricted to the cell edge in static conditions and homogeneously distributed after MS in WT and Rac1. The relative area of contacts (vinculin & paxillin) was more dramatically enhanced by MS in Rac1 (+80%) than in WT (+40%) and RhoA (+25%) indicating that new focal contacts are formed under MS and supported the presence of Sf. MS Activation of FAK (FAKY397) was clear in WT and Rac1 and reduced in RhoA. FnF was restricted to cell-cell contacts zone without any change in the relative area of fbn after a 2-days MS. However we found more numerous spots of fbn at the cell center in Rac1 as compared with RhoA & WT suggesting that these fibrillar contacts will grow upon maturation and modulate FnF. The results indicate that MS induces formation of Sf and focal adhesions and enhances FF. RhoA has been shown to induce the formation of Sf and focal adhesions, and Rac1 activation decreases Rho activity in

  15. In Vitro Analysis of Fibronectin-Modified Titanium Surfaces

    PubMed Central

    Chang, Yu-Chi; Lee, Wei-Fang; Feng, Sheng-Wei; Huang, Haw-Ming; Lin, Che-Tong; Teng, Nai-Chia; Chang, Wei Jen

    2016-01-01

    Background Glow discharge plasma (GDP) procedure is an effective method for grafting various proteins, including albumin, type I collagen, and fibronectin, onto a titanium surface. However, the behavior and impact of titanium (Ti) surface modification is yet to be unraveled. Purpose The purpose of this study is to evaluate and analyze the biological properties of fibronectin-grafted Ti surfaces treated by GDP. Materials and Methods Grade II Ti discs were initially cleaned and autoclaved to obtain original specimens. Subsequently, the specimens were GDP treated and grafted with fibronectin to form Ar-GDP (Argon GDP treatment only) and GDP-fib (fibronectin coating following GDP treatment) groups. Blood coagulation test and MG-63 cell culture were performed to evaluate the biological effects on the specimen. Results There was no significant difference between Ar-GDP and GDP-fib groups in blood compatibility analysis. While in the MTT test, cellular proliferation was benefited from the presence of fibronectin coating. The numbers of cells on Ar-GDP and GDP-fib specimens were greater than those in the original specimens after 24 h of culturing. Conclusions GDP treatment combined with fibronectin grafting favored MG-63 cell adhesion, migration, and proliferation on titanium surfaces, which could be attributed to the improved surface properties. PMID:26731536

  16. Tendon synovial cells secrete fibronectin in vivo and in vitro

    SciTech Connect

    Banes, A.J.; Link, G.W.; Bevin, A.G.; Peterson, H.D.; Gillespie, Y.; Bynum, D.; Watts, S.; Dahners, L.

    1988-01-01

    The chemistry and cell biology of the tendon have been largely overlooked due to the emphasis on collagen, the principle structural component of the tendon. The tendon must not only transmit the force of muscle contraction to bone to effect movement, but it must also glide simultaneously over extratendonous tissues. Fibronectin is classified as a cell attachment molecule that induces cell spreading and adhesion to substratum. The external surface of intact avian flexor tendon stained positively with antibody to cellular fibronectin. However, if the surface synovial cells were first removed with collagenase, no positive reaction with antifibronectin antibody was detected. Analysis of immunologically stained frozen sections of tendon also revealed fibronectin at the tendon synovium, but little was associated with cells internal in tendon. The staining pattern with isolated, cultured synovial cells and fibroblasts from the tendon interior substantiated the histological observations. Analysis of polyacrylamide gel profiles of /sup 35/S-methionine-labeled proteins synthesized by synovial cells and internal fibroblasts indicated that fibronectin was synthesized principally by synovial cells. Fibronectin at the tendon surface may play a role in cell attachment to prevent cell removal by the friction of gliding. Alternatively, fibronectin, with its binding sites for hyaluronic acid and collagen, may act as a complex for boundary lubrication.

  17. Fibronectin inhibits chronic pain development after spinal cord injury.

    PubMed

    Lin, Ching-Yi; Lee, Yu-Shang; Lin, Vernon W; Silver, Jerry

    2012-02-10

    Chronic pain following spinal cord injury (SCI) is a highly prevalent clinical condition that is difficult to treat. Using both von Frey filaments and radiant infrared heat to assess mechanical allodynia and thermal hyperalgesia, respectively, we have demonstrated that a one-time injection of fibronectin (50 μg/mL) into the spinal dorsal column (1 μL/min each injection for a total of 5 μL) immediately after SCI inhibits the development of mechanical allodynia (but not thermal hyperalgesia) over an 8-month observation period following spinal cord dorsal column crush (DCC). DCC will only induce mechanical Allodynia, but not thermal hyperalgesia or overt motor deficits. By applying various fibronectin fragments as well as competitive inhibitors, these effects were shown to be dependent on the connecting segment-1 (CS-1) motif of fibronectin. Furthermore, we found that acute fibronectin treatment diminished inflammation and blood-spinal cord barrier permeability, which in turn leads to enhanced fiber sparing and sprouting. In particular, the reduction of serotonin (5-HT) in the superficial dorsal horn, an important descending brainstem system in the modulation of pain, was blocked with fibronectin treatment. We conclude that treatment of SCI with fibronectin preserves sensory regulation and prevents the development of chronic allodynia, providing a potential therapeutic intervention to treat chronic pain following SCI.

  18. Fibronectin Inhibits Chronic Pain Development after Spinal Cord Injury

    PubMed Central

    Lee, Yu-Shang; Lin, Vernon W.; Silver, Jerry

    2012-01-01

    Abstract Chronic pain following spinal cord injury (SCI) is a highly prevalent clinical condition that is difficult to treat. Using both von Frey filaments and radiant infrared heat to assess mechanical allodynia and thermal hyperalgesia, respectively, we have demonstrated that a one-time injection of fibronectin (50 μg/mL) into the spinal dorsal column (1 μL/min each injection for a total of 5 μL) immediately after SCI inhibits the development of mechanical allodynia (but not thermal hyperalgesia) over an 8-month observation period following spinal cord dorsal column crush (DCC). DCC will only induce mechanical Allodynia, but not thermal hyperalgesia or overt motor deficits. By applying various fibronectin fragments as well as competitive inhibitors, these effects were shown to be dependent on the connecting segment-1 (CS-1) motif of fibronectin. Furthermore, we found that acute fibronectin treatment diminished inflammation and blood–spinal cord barrier permeability, which in turn leads to enhanced fiber sparing and sprouting. In particular, the reduction of serotonin (5-HT) in the superficial dorsal horn, an important descending brainstem system in the modulation of pain, was blocked with fibronectin treatment. We conclude that treatment of SCI with fibronectin preserves sensory regulation and prevents the development of chronic allodynia, providing a potential therapeutic intervention to treat chronic pain following SCI. PMID:22022865

  19. Fibronectin receptors of mononuclear phagocytes: Binding characteristics and biochemical isolation

    SciTech Connect

    Garcia-Pardo, A.; Ferreira, O.C.; Valinsky, J.; Bianco, C. )

    1989-04-01

    Fibronectin receptors on mononuclear phagocytes are involved in the localization of monocytes at inflammatory sites and in the subsequent expression of macrophage-like phenotypes. In this study, the authors have investigated the hypothesis that preteolytically derived fragments of fibronectin may interfere with binding of fibronectin to monocytes in the extracellular matrix. They report on the reactivity of U937 cells with an 80-kDa tryptic fragment of fibronectin which contains the cell-binding domain but lacks the gelatin/collagen-binding domain. U937 cells attached to surfaces coated with the 80-kDa fragment as well as with intact fibronectin. Preincubation of the cells with the 80-kDa fragment inhibited attachment to both surfaces while intact fibronectin had little or no inhibitory effect. This complex resolved into a single diffuse band of 144 kDa upon reduction. Binding of the protein complex to the affinity column required divalent cations. The complex bound to wheat germ agglutinin and could be specifically eluted by N-acetylglucosamine. Similar cell-surface proteins were isolated from peripheral blood monocytes.

  20. Fibronectin matrix assembly is essential for cell condensation during chondrogenesis

    PubMed Central

    Singh, Purva; Schwarzbauer, Jean E.

    2014-01-01

    ABSTRACT Mesenchymal cell condensation is the initiating event in endochondral bone formation. Cell condensation is followed by differentiation into chondrocytes, which is accompanied by induction of chondrogenic gene expression. Gene mutations involved in chondrogenesis cause chondrodysplasias and other skeletal defects. Using mesenchymal stem cells (MSCs) in an in vitro chondrogenesis assay, we found that knockdown of the diastrophic dysplasia (DTD) sulfate transporter (DTDST, also known as SLC26A2), which is required for normal cartilage development, blocked cell condensation and caused a significant reduction in fibronectin matrix. Knockdown of fibronectin with small interfering RNAs (siRNAs) also blocked condensation. Fibrillar fibronectin matrix was detected prior to cell condensation, and its levels increased during and after condensation. Inhibition of fibronectin matrix assembly by use of the functional upstream domain (FUD) of adhesin F1 from Streptococcus pyogenes prevented cell condensation by MSCs and also by the chondrogenic cell line ATDC5. Our data show that cell condensation and induction of chondrogenesis depend on fibronectin matrix assembly and DTDST, and indicate that this transporter is required earlier in chondrogenesis than previously appreciated. They also raise the possibility that certain of the skeletal defects in DTD patients might derive from the link between DTDST, fibronectin matrix and condensation. PMID:25146392

  1. Fibronectin matrix assembly is essential for cell condensation during chondrogenesis.

    PubMed

    Singh, Purva; Schwarzbauer, Jean E

    2014-10-15

    Mesenchymal cell condensation is the initiating event in endochondral bone formation. Cell condensation is followed by differentiation into chondrocytes, which is accompanied by induction of chondrogenic gene expression. Gene mutations involved in chondrogenesis cause chondrodysplasias and other skeletal defects. Using mesenchymal stem cells (MSCs) in an in vitro chondrogenesis assay, we found that knockdown of the diastrophic dysplasia (DTD) sulfate transporter (DTDST, also known as SLC26A2), which is required for normal cartilage development, blocked cell condensation and caused a significant reduction in fibronectin matrix. Knockdown of fibronectin with small interfering RNAs (siRNAs) also blocked condensation. Fibrillar fibronectin matrix was detected prior to cell condensation, and its levels increased during and after condensation. Inhibition of fibronectin matrix assembly by use of the functional upstream domain (FUD) of adhesin F1 from Streptococcus pyogenes prevented cell condensation by MSCs and also by the chondrogenic cell line ATDC5. Our data show that cell condensation and induction of chondrogenesis depend on fibronectin matrix assembly and DTDST, and indicate that this transporter is required earlier in chondrogenesis than previously appreciated. They also raise the possibility that certain of the skeletal defects in DTD patients might derive from the link between DTDST, fibronectin matrix and condensation.

  2. Fibronectin-guided migration of carcinoma collectives

    PubMed Central

    Gopal, Sandeep; Veracini, Laurence; Grall, Dominique; Butori, Catherine; Schaub, Sébastien; Audebert, Stéphane; Camoin, Luc; Baudelet, Emilie; Radwanska, Agata; Beghelli-de la Forest Divonne, Stéphanie; Violette, Shelia M.; Weinreb, Paul H.; Rekima, Samah; Ilie, Marius; Sudaka, Anne; Hofman, Paul; Van Obberghen-Schilling, Ellen

    2017-01-01

    Functional interplay between tumour cells and their neoplastic extracellular matrix plays a decisive role in malignant progression of carcinomas. Here we provide a comprehensive data set of the human HNSCC-associated fibroblast matrisome. Although much attention has been paid to the deposit of collagen, we identify oncofetal fibronectin (FN) as a major and obligate component of the matrix assembled by stromal fibroblasts from head and neck squamous cell carcinomas (HNSCC). FN overexpression in tumours from 435 patients corresponds to an independent unfavourable prognostic indicator. We show that migration of carcinoma collectives on fibrillar FN-rich matrices is achieved through αvβ6 and α9β1 engagement, rather than α5β1. Moreover, αvβ6-driven migration occurs independently of latent TGF-β activation and Smad-dependent signalling in tumour epithelial cells. These results provide insights into the adhesion-dependent events at the tumour–stroma interface that govern the collective mode of migration adopted by carcinoma cells to invade surrounding stroma in HNSCC. PMID:28102238

  3. Focal segmental glomerulosclerosis

    MedlinePlus

    ... is called the glomerulus. The glomeruli serve as filters that help the body get rid of harmful substances. Each kidney has thousands of glomeruli. "Focal" means that some of the glomeruli become scarred. Others ...

  4. Partial (focal) seizure

    MedlinePlus

    ... Jacksonian seizure; Seizure - partial (focal); Temporal lobe seizure; Epilepsy - partial seizures ... Abou-Khalil BW, Gallagher MJ, Macdonald RL. Epilepsies. In: Daroff ... Practice . 7th ed. Philadelphia, PA: Elsevier; 2016:chap 101. ...

  5. Integrin activation and focal complex formation in cardiac hypertrophy

    NASA Technical Reports Server (NTRS)

    Laser, M.; Willey, C. D.; Jiang, W.; Cooper, G. 4th; Menick, D. R.; Zile, M. R.; Kuppuswamy, D.

    2000-01-01

    Cardiac hypertrophy is characterized by both remodeling of the extracellular matrix (ECM) and hypertrophic growth of the cardiocytes. Here we show increased expression and cytoskeletal association of the ECM proteins fibronectin and vitronectin in pressure-overloaded feline myocardium. These changes are accompanied by cytoskeletal binding and phosphorylation of focal adhesion kinase (FAK) at Tyr-397 and Tyr-925, c-Src at Tyr-416, recruitment of the adapter proteins p130(Cas), Shc, and Nck, and activation of the extracellular-regulated kinases ERK1/2. A synthetic peptide containing the Arg-Gly-Asp (RGD) motif of fibronectin and vitronectin was used to stimulate adult feline cardiomyocytes cultured on laminin or within a type-I collagen matrix. Whereas cardiocytes under both conditions showed RGD-stimulated ERK1/2 activation, only collagen-embedded cells exhibited cytoskeletal assembly of FAK, c-Src, Nck, and Shc. In RGD-stimulated collagen-embedded cells, FAK was phosphorylated only at Tyr-397 and c-Src association occurred without Tyr-416 phosphorylation and p130(Cas) association. Therefore, c-Src activation is not required for its cytoskeletal binding but may be important for additional phosphorylation of FAK. Overall, our study suggests that multiple signaling pathways originate in pressure-overloaded heart following integrin engagement with ECM proteins, including focal complex formation and ERK1/2 activation, and many of these pathways can be activated in cardiomyocytes via RGD-stimulated integrin activation.

  6. Integrin activation and focal complex formation in cardiac hypertrophy

    NASA Technical Reports Server (NTRS)

    Laser, M.; Willey, C. D.; Jiang, W.; Cooper, G. 4th; Menick, D. R.; Zile, M. R.; Kuppuswamy, D.

    2000-01-01

    Cardiac hypertrophy is characterized by both remodeling of the extracellular matrix (ECM) and hypertrophic growth of the cardiocytes. Here we show increased expression and cytoskeletal association of the ECM proteins fibronectin and vitronectin in pressure-overloaded feline myocardium. These changes are accompanied by cytoskeletal binding and phosphorylation of focal adhesion kinase (FAK) at Tyr-397 and Tyr-925, c-Src at Tyr-416, recruitment of the adapter proteins p130(Cas), Shc, and Nck, and activation of the extracellular-regulated kinases ERK1/2. A synthetic peptide containing the Arg-Gly-Asp (RGD) motif of fibronectin and vitronectin was used to stimulate adult feline cardiomyocytes cultured on laminin or within a type-I collagen matrix. Whereas cardiocytes under both conditions showed RGD-stimulated ERK1/2 activation, only collagen-embedded cells exhibited cytoskeletal assembly of FAK, c-Src, Nck, and Shc. In RGD-stimulated collagen-embedded cells, FAK was phosphorylated only at Tyr-397 and c-Src association occurred without Tyr-416 phosphorylation and p130(Cas) association. Therefore, c-Src activation is not required for its cytoskeletal binding but may be important for additional phosphorylation of FAK. Overall, our study suggests that multiple signaling pathways originate in pressure-overloaded heart following integrin engagement with ECM proteins, including focal complex formation and ERK1/2 activation, and many of these pathways can be activated in cardiomyocytes via RGD-stimulated integrin activation.

  7. Temporal relationships of F-actin bundle formation, collagen and fibronectin matrix assembly, and fibronectin receptor expression to wound contraction

    PubMed Central

    1990-01-01

    Wound contraction can substantially reduce the amount of new tissue needed to reestablish organ integrity after tissue loss. Fibroblasts, rich in F-actin bundles, generate the force of wound contraction. Fibronectin-containing microfibrils link fibroblasts to each other and to collagen bundles and thereby provide transduction cables across the wound for contraction. The temporal relationships of F-actin bundle formation, collagen and fibronectin matrix assembly, and fibronectin receptor expression to wound contraction have not been determined. To establish these relationships, we used a cutaneous gaping wound model in outbred Yorkshire pigs. Granulation tissue filled approximately 80% of the wound space by day 5 after injury while wound contraction was first apparent at day 10. Neither actin bundles nor fibronectin receptors were observed in 5-d wound fibroblasts. Although fibronectin fibrils were assembled on the surfaces of 5-d fibroblasts, few fibrils coursed between cells. Day-7 fibroblasts stained strongly for nonmuscle- type F-actin bundles consistent with a contractile fibroblast phenotype. These cells expressed fibronectin receptors, were embedded in a fibronectin matrix that appeared to connect fibroblasts to the matrix and to each other, and were coaligned across the wound. Transmission EM confirmed the presence of microfilament bundles, cell- cell and cell-matrix linkages at day 7. Fibroblast coalignment, matrix interconnections, and actin bundles became more pronounced at days 10 and 14 coinciding with tissue contraction. These findings demonstrate that granulation tissue formation, F-actin bundle and fibronectin receptor expression in wound fibroblasts, and fibroblast-matrix linkage precede wound contraction. PMID:2136860

  8. The dispersion-focalization theory of sound systems

    NASA Astrophysics Data System (ADS)

    Schwartz, Jean-Luc; Abry, Christian; Boë, Louis-Jean; Vallée, Nathalie; Ménard, Lucie

    2005-04-01

    The Dispersion-Focalization Theory states that sound systems in human languages are shaped by two major perceptual constraints: dispersion driving auditory contrast towards maximal or sufficient values [B. Lindblom, J. Phonetics 18, 135-152 (1990)] and focalization driving auditory spectra towards patterns with close neighboring formants. Dispersion is computed from the sum of the inverse squared inter-spectra distances in the (F1, F2, F3, F4) space, using a non-linear process based on the 3.5 Bark critical distance to estimate F2'. Focalization is based on the idea that close neighboring formants produce vowel spectra with marked peaks, easier to process and memorize in the auditory system. Evidence for increased stability of focal vowels in short-term memory was provided in a discrimination experiment on adult French subjects [J. L. Schwartz and P. Escudier, Speech Comm. 8, 235-259 (1989)]. A reanalysis of infant discrimination data shows that focalization could well be the responsible for recurrent discrimination asymmetries [J. L. Schwartz et al., Speech Comm. (in press)]. Recent data about children vowel production indicate that focalization seems to be part of the perceptual templates driving speech development. The Dispersion-Focalization Theory produces valid predictions for both vowel and consonant systems, in relation with available databases of human languages inventories.

  9. Analysis of the myosin-II-responsive focal adhesion proteome reveals a role for β-Pix in negative regulation of focal adhesion maturation.

    PubMed

    Kuo, Jean-Cheng; Han, Xuemei; Hsiao, Cheng-Te; Yates, John R; Waterman, Clare M

    2011-04-01

    Focal adhesions undergo myosin-II-mediated maturation wherein they grow and change composition to modulate integrin signalling for cell migration, growth and differentiation. To determine how focal adhesion composition is affected by myosin II activity, we performed proteomic analysis of isolated focal adhesions and compared protein abundance in focal adhesions from cells with and without myosin II inhibition. We identified 905 focal adhesion proteins, 459 of which changed in abundance with myosin II inhibition, defining the myosin-II-responsive focal adhesion proteome. The abundance of 73% of the proteins in the myosin-II-responsive focal adhesion proteome was enhanced by contractility, including proteins involved in Rho-mediated focal adhesion maturation and endocytosis- and calpain-dependent focal adhesion disassembly. During myosin II inhibition, 27% of proteins in the myosin-II-responsive focal adhesion proteome, including proteins involved in Rac-mediated lamellipodial protrusion, were enriched in focal adhesions, establishing that focal adhesion protein recruitment is also negatively regulated by contractility. We focused on the Rac guanine nucleotide exchange factor β-Pix, documenting its role in the negative regulation of focal adhesion maturation and the promotion of lamellipodial protrusion and focal adhesion turnover to drive cell migration. © 2011 Macmillan Publishers Limited. All rights reserved

  10. Treatment of focal dystonia.

    PubMed

    Batla, Amit; Stamelou, Maria; Bhatia, Kailash P

    2012-06-01

    Dystonia is characterized by repetitive twisting movements or abnormal postures due to involuntary muscle activity. When limited to a single body region it is called focal dystonia. Examples of focal dystonia include cervical dystonia (neck), blepharospasm (eyes), oromandibular dystonia, focal limb dystonia, and spasmodic dysphonia, which are discussed here. Once the diagnosis is established, the therapeutic plan is discussed with the patients. They are informed that there is no cure for dystonia and treatment is symptomatic. The main therapeutic option for treating focal dystonias is botulinum toxin (BoNT). There have been several attempts to characterize the procedure, the type of toxin, dosage, techniques, and combination with physical measures in each of the focal dystonia forms. The general treatment principles are similar. The affected muscles are injected at muscle sites based on evidence and experience using standard dosages based on the type of toxin used. The injections are repeated after 3 to 6 months based on the individual response duration. In the uncommon event of nonresponse with BoNT, the dose and site are reassessed. Oral drug treatment could be considered as an additional option. Once the condition is thought to be medically refractory, the opinion from the deep brain stimulation (DBS) team for the suitability of the patient for DBS is taken. The successful use of DBS in cervical dystonia has led to increased acceptance for trial in other forms of focal dystonias. DBS surgery in focal dystonias other than cervical is, however, still experimental. The patients may be offered the surgery with adequate explanation of the risks and benefits. Patient education and directing the patients towards dystonia support groups and relevant websites that provide scientific information may be useful for long-term compliance and benefit.

  11. Dexamethasone-Mediated Activation of Fibronectin Matrix Assembly Reduces Dispersal of Primary Human Glioblastoma Cells.

    PubMed

    Shannon, Stephen; Vaca, Connan; Jia, Dongxuan; Entersz, Ildiko; Schaer, Andrew; Carcione, Jonathan; Weaver, Michael; Avidar, Yoav; Pettit, Ryan; Nair, Mohan; Khan, Atif; Foty, Ramsey A

    2015-01-01

    Despite resection and adjuvant therapy, the 5-year survival for patients with Glioblastoma multiforme (GBM) is less than 10%. This poor outcome is largely attributed to rapid tumor growth and early dispersal of cells, factors that contribute to a high recurrence rate and poor prognosis. An understanding of the cellular and molecular machinery that drive growth and dispersal is essential if we are to impact long-term survival. Our previous studies utilizing a series of immortalized GBM cell lines established a functional causation between activation of fibronectin matrix assembly (FNMA), increased tumor cohesion, and decreased dispersal. Activation of FNMA was accomplished by treatment with Dexamethasone (Dex), a drug routinely used to treat brain tumor related edema. Here, we utilize a broad range of qualitative and quantitative assays and the use of a human GBM tissue microarray and freshly-isolated primary human GBM cells grown both as conventional 2D cultures and as 3D spheroids to explore the role of Dex and FNMA in modulating various parameters that can significantly influence tumor cell dispersal. We show that the expression and processing of fibronectin in a human GBM tissue-microarray is variable, with 90% of tumors displaying some abnormality or lack in capacity to secrete fibronectin or assemble it into a matrix. We also show that low-passage primary GBM cells vary in their capacity for FNMA and that Dex treatment reactivates this process. Activation of FNMA effectively "glues" cells together and prevents cells from detaching from the primary mass. Dex treatment also significantly increases the strength of cell-ECM adhesion and decreases motility. The combination of increased cohesion and decreased motility discourages in vitro and ex vivo dispersal. By increasing cell-cell cohesion, Dex also decreases growth rate of 3D spheroids. These effects could all be reversed by an inhibitor of FNMA and by the glucocorticoid receptor antagonist, RU-486. Our results

  12. Brain fibronectin expression in prenatally irradiated mice

    SciTech Connect

    Meznarich, H.K.; McCoy, L.S.; Bale, T.L.; Stiegler, G.L.; Sikov, M.R. )

    1993-01-01

    Activation of gene transcription by radiation has been recently demonstrated in vivo. However, little is known on the specificity of these alterations on gene transcription. Prenatal irradiation is a known teratogen that affects the developing mammalian central nervous system (CNS). Altered neuronal migration has been suggested as a mechanism for abnormal development of prenatally irradiated brains. Fibronectin (FN), an extracellular glycoprotein, is essential for neural crest cell migration and neural cell growth. In addition, elevated levels of FN have been found in the extracellular matrix of irradiated lung. To test whether brain FN is affected by radiation, either FN level in insoluble matrix fraction or expression of FN mRNA was examined pre- and postnatally after irradiation. Mice (CD1), at 13 d of gestation (DG), served either as controls or were irradiated with 14 DG, 17 DG, or 5,6, or 14 d postnatal. Brain and liver were collected from offspring and analyzed for either total FN protein levels or relative mRNAs for FN and tubulin. Results of prenatal irradiation on reduction of postnatal brain weight relative to whole are comparable to that reported by others. Insoluble matrix fraction (IMF) per gram of brain, liver, lung, and heart weight was not significantly different either between control and irradiated groups or between postnatal stages, suggesting that radiation did not affect the IMF. However, total amounts of FN in brain IMF at 17 DG were significantly different (p < .02) between normal (1.66 [+-] 0.80 [mu]g) and irradiated brains (0.58 [+-] 0.22 [mu]g). FN mRNA was detectable at 13, 14, and 17 DG, but was not detectable at 6 and 14 d postnatal, indicating that FN mRNA is developmentally regulated. 41 refs., 4 figs., 3 tabs.

  13. Mesothelial cells promote early ovarian cancer metastasis through fibronectin secretion.

    PubMed

    Kenny, Hilary A; Chiang, Chun-Yi; White, Erin A; Schryver, Elizabeth M; Habis, Mohammed; Romero, Iris L; Ladanyi, Andras; Penicka, Carla V; George, Joshy; Matlin, Karl; Montag, Anthony; Wroblewski, Kristen; Yamada, S Diane; Mazar, Andrew P; Bowtell, David; Lengyel, Ernst

    2014-10-01

    Ovarian cancer (OvCa) metastasizes to organs in the abdominal cavity, such as the omentum, which are covered by a single layer of mesothelial cells. Mesothelial cells are generally thought to be "bystanders" to the metastatic process and simply displaced by OvCa cells to access the submesothelial extracellular matrix. Here, using organotypic 3D cultures, we found that primary human mesothelial cells secrete fibronectin in the presence of OvCa cells. Moreover, we evaluated the tumor stroma of 108 human omental metastases and determined that fibronectin was consistently overexpressed in these patients. Blocking fibronectin production in primary mesothelial cells in vitro or in murine models, either genetically (fibronectin 1 floxed mouse model) or via siRNA, decreased adhesion, invasion, proliferation, and metastasis of OvCa cells. Using a coculture model, we determined that OvCa cells secrete TGF-β1, which in turn activates a TGF-β receptor/RAC1/SMAD-dependent signaling pathway in the mesothelial cells that promotes a mesenchymal phenotype and transcriptional upregulation of fibronectin. Additionally, blocking α5 or β1 integrin function with antibodies reduced metastasis in an orthotopic preclinical model of OvCa metastasis. These findings indicate that cancer-associated mesothelial cells promote colonization during the initial steps of OvCa metastasis and suggest that mesothelial cells actively contribute to metastasis.

  14. Mesothelial cells promote early ovarian cancer metastasis through fibronectin secretion

    PubMed Central

    Kenny, Hilary A.; Chiang, Chun-Yi; White, Erin A.; Schryver, Elizabeth M.; Habis, Mohammed; Romero, Iris L.; Ladanyi, Andras; Penicka, Carla V.; George, Joshy; Matlin, Karl; Montag, Anthony; Wroblewski, Kristen; Yamada, S. Diane; Mazar, Andrew P.; Bowtell, David; Lengyel, Ernst

    2014-01-01

    Ovarian cancer (OvCa) metastasizes to organs in the abdominal cavity, such as the omentum, which are covered by a single layer of mesothelial cells. Mesothelial cells are generally thought to be “bystanders” to the metastatic process and simply displaced by OvCa cells to access the submesothelial extracellular matrix. Here, using organotypic 3D cultures, we found that primary human mesothelial cells secrete fibronectin in the presence of OvCa cells. Moreover, we evaluated the tumor stroma of 108 human omental metastases and determined that fibronectin was consistently overexpressed in these patients. Blocking fibronectin production in primary mesothelial cells in vitro or in murine models, either genetically (fibronectin 1 floxed mouse model) or via siRNA, decreased adhesion, invasion, proliferation, and metastasis of OvCa cells. Using a coculture model, we determined that OvCa cells secrete TGF-β1, which in turn activates a TGF-β receptor/RAC1/SMAD-dependent signaling pathway in the mesothelial cells that promotes a mesenchymal phenotype and transcriptional upregulation of fibronectin. Additionally, blocking α5 or β1 integrin function with antibodies reduced metastasis in an orthotopic preclinical model of OvCa metastasis. These findings indicate that cancer-associated mesothelial cells promote colonization during the initial steps of OvCa metastasis and suggest that mesothelial cells actively contribute to metastasis. PMID:25202979

  15. Fibronectin coating of oxygenator membranes enhances endothelial cell attachment

    PubMed Central

    2013-01-01

    Background Extracorporeal membrane oxygenation (ECMO) can replace the lungs’ gas exchange capacity in refractory lung failure. However, its limited hemocompatibility, the activation of the coagulation and complement system as well as plasma leakage and protein deposition hamper mid- to long-term use and have constrained the development of an implantable lung assist device. In a tissue engineering approach, lining the blood contact surfaces of the ECMO device with endothelial cells might overcome these limitations. As a first step towards this aim, we hypothesized that coating the oxygenator’s gas exchange membrane with proteins might positively influence the attachment and proliferation of arterial endothelial cells. Methods Sheets of polypropylene (PP), polyoxymethylpentene (TPX) and polydimethylsiloxane (PDMS), typical material used for oxygenator gas exchange membranes, were coated with collagen, fibrinogen, gelatin or fibronectin. Tissue culture treated well plates served as controls. Endothelial cell attachment and proliferation were analyzed for a period of 4 days by microscopic examination and computer assisted cell counting. Results Endothelial cell seeding efficiency is within range of tissue culture treated controls for fibronectin treated surfaces only. Uncoated membranes as well as all other coatings lead to lower cell attachment. A confluent endothelial cell layer develops on fibronectin coated PDMS and the control surface only. Conclusions Fibronectin increases endothelial cells’ seeding efficiency on different oxygenator membrane material. PDMS coated with fibronectin shows sustained cell attachment for a period of four days in static culture conditions. PMID:23356939

  16. ADAMTS-10 and -6 differentially regulate cell-cell junctions and focal adhesions.

    PubMed

    Cain, Stuart A; Mularczyk, Ewa J; Singh, Mukti; Massam-Wu, Teresa; Kielty, Cay M

    2016-10-25

    ADAMTS10 and ADAMTS6 are homologous metalloproteinases with ill-defined roles. ADAMTS10 mutations cause Weill-Marchesani syndrome (WMS), implicating it in fibrillin microfibril biology since some fibrillin-1 mutations also cause WMS. However little is known about ADAMTS6 function. ADAMTS10 is resistant to furin cleavage, however we show that ADAMTS6 is effectively processed and active. Using siRNA, over-expression and mutagenesis, it was found ADAMTS6 inhibits and ADAMTS10 is required for focal adhesions, epithelial cell-cell junction formation, and microfibril deposition. Either knockdown of ADAMTS6, or disruption of its furin processing or catalytic sites restores focal adhesions, implicating its enzyme activity acts on targets in the focal adhesion complex. In ADAMTS10-depleted cultures, expression of syndecan-4 rescues focal adhesions and cell-cell junctions. Recombinant C-termini of ADAMTS10 and ADAMTS6, both of which induce focal adhesions, bind heparin and syndecan-4. However, cells overexpressing full-length ADAMTS6 lack heparan sulphate and focal adhesions, whilst depletion of ADAMTS6 induces a prominent glycocalyx. Thus ADAMTS10 and ADAMTS6 oppositely affect heparan sulphate-rich interfaces including focal adhesions. We previously showed that microfibril deposition requires fibronectin-induced focal adhesions, and cell-cell junctions in epithelial cultures. Here we reveal that ADAMTS6 causes a reduction in heparan sulphate-rich interfaces, and its expression is regulated by ADAMTS10.

  17. ADAMTS-10 and -6 differentially regulate cell-cell junctions and focal adhesions

    PubMed Central

    Cain, Stuart A.; Mularczyk, Ewa J.; Singh, Mukti; Massam-Wu, Teresa; Kielty, Cay M.

    2016-01-01

    ADAMTS10 and ADAMTS6 are homologous metalloproteinases with ill-defined roles. ADAMTS10 mutations cause Weill-Marchesani syndrome (WMS), implicating it in fibrillin microfibril biology since some fibrillin-1 mutations also cause WMS. However little is known about ADAMTS6 function. ADAMTS10 is resistant to furin cleavage, however we show that ADAMTS6 is effectively processed and active. Using siRNA, over-expression and mutagenesis, it was found ADAMTS6 inhibits and ADAMTS10 is required for focal adhesions, epithelial cell-cell junction formation, and microfibril deposition. Either knockdown of ADAMTS6, or disruption of its furin processing or catalytic sites restores focal adhesions, implicating its enzyme activity acts on targets in the focal adhesion complex. In ADAMTS10-depleted cultures, expression of syndecan-4 rescues focal adhesions and cell-cell junctions. Recombinant C-termini of ADAMTS10 and ADAMTS6, both of which induce focal adhesions, bind heparin and syndecan-4. However, cells overexpressing full-length ADAMTS6 lack heparan sulphate and focal adhesions, whilst depletion of ADAMTS6 induces a prominent glycocalyx. Thus ADAMTS10 and ADAMTS6 oppositely affect heparan sulphate-rich interfaces including focal adhesions. We previously showed that microfibril deposition requires fibronectin-induced focal adhesions, and cell-cell junctions in epithelial cultures. Here we reveal that ADAMTS6 causes a reduction in heparan sulphate-rich interfaces, and its expression is regulated by ADAMTS10. PMID:27779234

  18. SNAP focal plane

    SciTech Connect

    Lampton, Michael L.; Kim, A.; Akerlof, C.W.; Aldering, G.; Amanullah, R.; Astier, P.; Barrelet, E.; Bebek, C.; Bergstrom, L.; Berkovitz, J.; Bernstein, G.; Bester, M.; Bonissent, A.; Bower, C.; Carithers Jr., W.C.; Commins, E.D.; Day, C.; Deustua, S.E.; DiGennaro,R.; Ealet, A.; Ellis, R.S.; Eriksson, M.; Fruchter, A.; Genat, J.-F.; Goldhaber, G.; Goobar, A.; Groom, D.; Harris, S.E.; Harvey, P.R.; Heetderks, H.D.; Holland, S.E.; Huterer, D.; Karcher, A.; Kolbe, W.; Krieger, B.; Lafever, R.; Lamoureux, J.; Levi, M.E.; Levin, D.S.; Linder,E.V.; Loken, S.C.; Malina, R.; Massey, R.; McKay, T.; McKee, S.P.; Miquel, R.; Mortsell, E.; Mostek, N.; Mufson, S.; Musser, J.; Nugent, P.; Oluseyi, H.; Pain, R.; Palaio, N.; Pankow, D.; Perlmutter, S.; Pratt, R.; Prieto, E.; Refregier, A.; Rhodes, J.; Robinson, K.; Roe, N.; Sholl, M.; Schubnell, M.; Smadja, G.; Smoot, G.; Spadafora, A.; Tarle, G.; Tomasch,A.; von der Lippe, H.; Vincent, R.; Walder, J.-P.; Wang, G.

    2002-07-29

    The proposed SuperNova/Acceleration Probe (SNAP) mission will have a two-meter class telescope delivering diffraction-limited images to an instrumented 0.7 square-degree field sensitive in the visible and near-infrared wavelength regime. We describe the requirements for the instrument suite and the evolution of the focal plane design to the present concept in which all the instrumentation--visible and near-infrared imagers, spectrograph, and star guiders--share one common focal plane.

  19. Vitelliform focal choroidal excavation.

    PubMed

    Or, Chris; Forooghian, Farzin

    2014-05-30

    Focal choroidal excavations (FCE) are characterized by foveal or perifoveal choroid excavations seen on optical coherence tomography (OCT). The authors report a case of FCE associated with a vitelliform lesion within the excavation. A case of FCE associated with a small vitelliform lesion has been described previously, but the larger extent of the vitelliform lesion observed in the current case has not been previously reported. This may represent a novel category of FCE, vitelliform focal choroidal excavation, in which deposition of vitelliform material is associated with its development.

  20. [Focal epithelial hyperplasia].

    PubMed

    Vera-Iglesias, E; García-Arpa, M; Sánchez-Caminero, P; Romero-Aguilera, G; Cortina de la Calle, P

    2007-11-01

    Focal epithelial hyperplasia is a rare disease of the oral mucosa caused by the human papilloma virus (HPV). It appears as a benign epithelial growth, usually in the mucosa of the lower lip. It is mainly associated with HPV serotypes 13 and 32 and there is a clear racial predilection for the disease in Native Americans and Eskimos. We describe the case of a 17-year-old girl from Ecuador with multiple papular lesions in both lips that were clinically and histologically consistent with focal epithelial hyperplasia. Analysis by polymerase chain reaction detected HPV serotype 13.

  1. Variable focal length microlenses

    NASA Astrophysics Data System (ADS)

    L. G., Commander; Day, S. E.; Selviah, D. R.

    2000-04-01

    Refractive surface relief microlenses (150 μm diameter) are immersed in nematic liquid crystal in a cell. Application of a variable voltage across the cell effectively varies the refractive index of the liquid crystal and results in a change of the focal length by the lensmakers formula (E. Hecht, Optics, 2nd edn., Addison-Wesley, Reading, Massachusetts, 1987, p. 138). We describe the cell design and construction and demonstrate a range of focal lengths from +490 to +1000 μm for 2 to 12 V applied. A diverging lens results when the voltage is lower. Theoretical models are developed to account for some of the observed aberrations.

  2. Focal brainstem gliomas

    PubMed Central

    Sabbagh, Abdulrahman J.; Alaqeel, Ahmed M.

    2015-01-01

    Improved neuronavigation guidance as well as intraoperative imaging and neurophysiologic monitoring technologies have enhanced the ability of neurosurgeons to resect focal brainstem gliomas. In contrast, diffuse brainstem gliomas are considered to be inoperable lesions. This article is a continuation of an article that discussed brainstem glioma diagnostics, imaging, and classification. Here, we address open surgical treatment of and approaches to focal, dorsally exophytic, and cervicomedullary brainstem gliomas. Intraoperative neuronavigation, intraoperative neurophysiologic monitoring, as well as intraoperative imaging are discussed as adjunctive measures to help render these procedures safer, more acute, and closer to achieving surgical goals. PMID:25864061

  3. Predicting preterm birth: Cervical length and fetal fibronectin.

    PubMed

    Son, Moeun; Miller, Emily S

    2017-09-18

    Spontaneous preterm birth remains the leading cause of neonatal morbidity and mortality worldwide, and accounts for a significant global health burden. Several obstetric strategies to screen for spontaneous preterm delivery, such as cervical length and fetal fibronectin measurement, have emerged. However, the effectiveness of these strategies relies on their ability to accurately predict those pregnancies at increased risk for spontaneous preterm birth (SPTB). Transvaginal cervical shortening is predictive of preterm birth and when coupled with appropriate preterm birth prevention strategies, has been associated with reductions in SPTB in asymptomatic women with a singleton gestation. The use of qualitative fetal fibronectin may be useful in conjunction with cervical length assessment in women with acute preterm labor symptoms, but data supporting its clinical utility remain limited. As both cervical length and qualitative fetal fibronectin have limited capacity to predict preterm birth, further studies are needed to investigate other potential screening modalities. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Glomerulopathy associated with predominant fibronectin deposits: Exclusion of the genes for fibronectin, villin and desmin as causative genes

    SciTech Connect

    Hildebrandt, F.; Strahm, B.; Prochoroff, A.; Cybulla, M.; Brandis, M.

    1996-05-03

    Glomerulopathy with predominant fibronectin deposits (GFD) is a newly recognized autosomal dominant renal disease that leads to albuminuria, microscopic hematuria, hypertension, renal tubular acidosis type IV, and end-stage renal disease in the second to fourth decade of life. Light microscopy documents extensive deposits in the subendothelial space, which on electron microscopy consist of non-oriented 12 x 125 nm fibers. Deposits are strongly immunoreactive for antibodies to fibronectin. We examined the hypothesis that a genetic defect in the gene for fibronectin is responsible for the disease. In a 197 member pedigree, 13 relatives developed end-stage renal failure from the disease. In 99 individuals haplotype analysis was performed using 6 microsatellite markers spanning a >56 cM interval in chromosome region 2q34, where fibronectin, villin, and desmin map in close proximity. Haplotype analysis resulted in exclusion of the whole range of 78 cM covered by the markers examined. This result excludes fibronectin, villin, and desmin from being the causative genes for GFD in this large kindred. 13 refs., 2 figs.

  5. Oral focal epithelial hyperplasia.

    PubMed

    Bassioukas, K; Danielides, V; Georgiou, I; Photos, E; Zagorianakou, P; Skevas, A

    2000-01-01

    Focal epithelial hyperplasia (FEH) or Heck disease, is a rare viral infection of the oral mucosa caused by HPV 13 or HPV 32. In Caucasians there have been only a few cases reported. We present the first case in Greece in a young Caucasian girl in which HPV 13 was detected with PCR analysis. The patient was successfully treated with CO2 laser.

  6. Immunolocalization of integrins and fibronectin in tubal pregnancy.

    PubMed

    Inan, Sevinc; Giray, Gulsen; Vatansever, H Seda; Ozbilgin, Kemal; Kuscu, N Kemal; Sayhan, Sevil

    2004-01-01

    Integrins are a large family of cell adhesion molecules that serve as receptors involved in cell-to-cell and cell-to-matrix interactions during implantation. We studied immunohistochemical staining of integrins (alpha 3, alpha V, beta 1, and alpha 2 beta 1) and fibronectin in ectopic tubal pregnancy. Thirty fallopian tube samples with ectopic pregnancies and five normal tubal segments were obtained during ligation operations; the latter specimens served as controls in the study. Formalin-fixed paraffin-embedded tissue sections were stained with hematoxylin-eosin or primary antibodies against alpha 3, beta 1, alpha V, and alpha 2 beta 1 integrins and fibronectin, using the avidin-biotin-peroxidase method. A semi-quantitative grading system was used to compare staining intensities. In the control samples, immunostaining of all integrins was found in a single layer of tall columnar epithelial cells, the lamina propria (Lp) and the muscular layer. Fibronectin staining was detected in the Lp and the muscular layer. Staining intensities of alpha 3 and beta 1 integrins and fibronectin were increased in the normal part of fallopian tubes with ectopic pregnancies. Staining of beta 1 integrin was more intense than staining of alpha 3 and fibronectin, whereas there was no difference in alpha V and alpha 2 beta 1 integrin expression between normal tubal tissue in the ectopic pregnancy group and control tubal tissue. In the tubal pregnancy group at the site of implantation, staining intensity of alpha 3 and beta 1 integrins and fibronectin was strong in decidual cells, supporting tissue and placental villi, whereas alpha V and alpha 2 beta 1 staining was mild. We concluded that integrins, especially beta 1 and alpha 3, and fibronectin may play a role in progression of tubal implantation. Although the role of integrins has not yet been clearly defined, these molecules may function as markers of normal and abnormal states of receptivity. We like to suggest that integrins and

  7. Nanopore Diameters Tune Strain in Extruded Fibronectin Fibers.

    PubMed

    Raoufi, Mohammad; Das, Tamal; Schoen, Ingmar; Vogel, Viola; Brüggemann, Dorothea; Spatz, Joachim P

    2015-10-14

    Fibronectin is present in the extracellular matrix and can be assembled into nanofibers in vivo by undergoing conformational changes. Here, we present a novel approach to prepare fibronectin nanofibers under physiological conditions using an extrusion approach through nanoporous aluminum oxide membranes. This one-step process can prepare nanofiber bundles up to a millimeter in length and with uniform fiber diameters in the nanometer range. Most importantly, by using different pore diameters and protein concentrations in the extrusion process, we could induce varying lasting structural changes in the fibers, which were monitored by Förster resonance energy transfer and should impose different physiological functions.

  8. Endothelium-derived fibronectin regulates neonatal vascular morphogenesis in an autocrine fashion.

    PubMed

    Turner, Christopher J; Badu-Nkansah, Kwabena; Hynes, Richard O

    2017-06-30

    Fibronectin containing alternatively spliced EIIIA and EIIIB domains is largely absent from mature quiescent vessels in adults, but is highly expressed around blood vessels during developmental and pathological angiogenesis. The precise functions of fibronectin and its splice variants during developmental angiogenesis however remain unclear due to the presence of cardiac, somitic, mesodermal and neural defects in existing global fibronectin KO mouse models. Using a rare family of surviving EIIIA EIIIB double KO mice, as well as inducible endothelial-specific fibronectin-deficient mutant mice, we show that vascular development in the neonatal retina is regulated in an autocrine manner by endothelium-derived fibronectin, and requires both EIIIA and EIIIB domains and the RGD-binding α5 and αv integrins for its function. Exogenous sources of fibronectin do not fully substitute for the autocrine function of endothelial fibronectin, demonstrating that fibronectins from different sources contribute differentially to specific aspects of angiogenesis.

  9. Fibronectin: How Its Aberrant Expression in Tumors May Improve Therapeutic Targeting

    PubMed Central

    Wang, Jennifer Peyling; Hielscher, Abigail

    2017-01-01

    Fibronectin is a matrix glycoprotein which has not only been found to be over-expressed in several cancers, but has been shown to participate in several steps of tumorigenesis. The purpose of this review is to illustrate how aberrant fibronectin expression influences tumor growth, invasion, metastasis and therapy resistance. In particular, this review will focus on the interactions between cell receptor ligands and fibronectin and how this interaction influences downstream signaling events that aid tumor progression. This review will further discuss the possible implications of therapeutic drugs directed against fibronectin and/or cellular interactions with fibronectin and will additionally discuss novel approaches by which to limit intra- and extra-tumoral fibronectin expression and the cellular events which lead to aberrant fibronectin expression. It is anticipated that these studies will set a basis for future research that will not only aid understanding of fibronectin and its prognostic significance, but will further elucidate novel targets for therapeutics. PMID:28367247

  10. Selective fibronectin adsorption against albumin and enhanced stem cell attachment on helium atmospheric pressure glow discharge treated titanium

    NASA Astrophysics Data System (ADS)

    Han, Inho; Vagaska, Barbora; Joo Park, Bong; Lee, Mi Hee; Jin Lee, Seung; Park, Jong-Chul

    2011-06-01

    Successful tissue integration of implanted medical devices depends on appropriate initial cellular response. In this study, the effect of helium atmospheric pressure glow discharge (He-APGD) treatment of titanium on selective protein adsorption and the initial attachment processes and focal adhesion formation of osteoprogenitor cells and stem cells were examined. Titanium disks were treated in a self-designed He-APGD system. Initial attachment of MC3T3-E1 mouse pre-osteoblasts and human mesenchymal stem cells (MSCs) was evaluated by MTT assay and plasma membrane staining followed by morphometric analysis. Fibronectin adsorption was investigated by Enzyme-Linked ImmunoSorbant Assay. MSCs cell attachment to treated and non-treated titanium disks coated with different proteins was verified also in serum-free culture. Organization of actin cytoskeleton and focal adhesions was evaluated microscopically. He-APGD treatment effectively modified the titanium surfaces by creating a super-hydrophilic surface, which promoted selectively higher adsorption of fibronectin, a protein of critical importance for cell/biomaterial interaction. In two different types of cells, the He-APGD treatment enhanced the number of attaching cells as well as their attachment area. Moreover, cells had higher organization of actin cytoskeleton and focal adhesions. Faster acceptance of the material by the progenitor cells in the early phases of tissue integration after the implantation may significantly reduce the overall healing time; therefore, titanium treatment with He-APGD seems to be an effective method of surface modification of titanium for improving its tissue inductive properties.

  11. Selective Cell Growth on Fibronectin-Carbon Nanotube Hybrid Nanostructures

    NASA Astrophysics Data System (ADS)

    Namgung, Seon; Park, Sung Young; Lee, Byung Yang; Lee, Minbaek; Nam, Jwa-Min; Hong, Seunghun

    2008-03-01

    Carbon nanotubes (CNT) have been considered a promising material for biological applications including biosensors, therapeutic application, and nano-structured scaffolds. However, there are still controversies associated with toxicity and biocompatibility of CNTs on live cells. Here, we report general strategy to functionalize CNTs with cell adhesion molecules (fibronectins) for selective and stable adhesion of cells on CNTs. Interestingly, more fibronectins were adsorbed and activated on CNTs rather than on hydrophobic self assembled monolayers (SAMs) or bare substrates (SiO2). We demonstrate the functionality of fibronectins on CNTs with immunofluorescence and molecule-level force measurement study using atomic force microscopy (AFM). These fibronectin-CNT hybrid nanostructures were successfully applied to attract cells selectively onto predefined regions on the substrate. Our strategy was generally available on various cell types including mesenchymal stem cells, KB cells, and NIH3T3 fibroblast cells (Advanced Materials 19, 2530-2534 (2007)). We will also discuss about its impacts on cell biology combined with CNTs.

  12. Modulation of Human Plasma Fibronectin Levels Following Exercise,

    DTIC Science & Technology

    1988-01-01

    increased under conditions of pathology, such as in obesity (6). cancer (3). proteinuria (4). diabetic retinopathy (5). and preeclampsia (27). in the absence...Res. 1977: 22:709-716. 27. Stubbs. T.M.. Lazarchick. J.. and Horger. E.O. Plasma fibronectin levels in preeclampsia : A possible biochemical marker

  13. Modulated fibronectin anchorage at polymer substrates controls angiogenesis.

    PubMed

    Pompe, Tilo; Markowski, Manuela; Werner, Carsten

    2004-01-01

    A set of maleic anhydride copolymer thin films exhibiting well-defined differences in hydrophobicity and reactivity was compared with respect to the capability of supporting angiogenesis of human endothelial cells grown in contact. The physicochemical surface characteristics of the polymer substrates were found to modulate the anchorage of immobilized fibronectin. This was demonstrated to determine whether endothelial cells grow as a monolayer or form capillary networks. Enhanced reorganization of predeposited fibronectin into cell-matrix adhesions and slightly elevated levels of membrane-type matrix metalloproteinase 14 (MMP-14) occurred with weakly bound fibronectin layers where angiogenesis was most obvious. The key role of fibronectin-substrate binding for angiogenesis-under otherwise constant conditions-was further confirmed by the absence of variations in the expression of angiogenesis-related integrins (alpha(v)beta(3)) and in the secretion of the metalloproteinase MMP-2. Altogether, the results of this study point at the relevance of physicochemical surface characteristics of polymer materials for the stimulation of angiogenesis.

  14. Nicotine enhances colon cancer cell migration by induction of fibronectin.

    PubMed

    Wei, Po-Li; Kuo, Li-Jen; Huang, Ming-Te; Ting, Wen-Chien; Ho, Yuan-Soon; Wang, Weu; An, Jane; Chang, Yu-Jia

    2011-06-01

    Long-term cigarette smoking increases the risk of colorectal cancer mortality. Tobacco's addictive toxin, nicotine, was reported to increase DNA synthesis of colon cancer cells. Because metastasis is the major cause of cancer death, the influence of nicotine on the migration of colon cancer cells remains to be determined. The influence of nicotine on the migration of colon cancer cells was evaluated using transwell assay. Nicotine receptor-mediated migration was studied by using both inhibitors and small interfering RNA (siRNA). The role of COX-2 signal was studied using pharmacological inhibitors. The expression of epithelial mesenchymal transition (EMT) marker and COX-2 signal was evaluated using real-time polymerase chain reaction (PCR). Nicotine enhanced DLD-1 and SW480 cell migration in a dose-dependent manner. We used inhibitors and siRNA to demonstrate that α7-nAChR mediates nicotine-enhanced colon cancer cell migration and upregulates fibronectin expression, which is involved in nicotine-enhanced migration. Furthermore, COX-2 signal was induced by nicotine treatment and is involved in nicotine-enhanced fibronectin expression. Nicotine, tobacco's additive toxin, enhances colon cancer metastasis through α7-nAChR and fibronectin--a mesenchymal marker for epithelial mesenchymal transition. Furthermore, COX-2 signal was involved in the induction of fibronectin. Therefore, smoking may play role in the progression of colon cancer.

  15. Specific affinity between fibronectin and the epidermolysis bullosa acquisita antigen.

    PubMed Central

    Woodley, D T; O'Keefe, E J; McDonald, J A; Reese, M J; Briggaman, R A; Gammon, W R

    1987-01-01

    Autoantibodies in the skin and sera of patients with epidermolysis bullosa acquisita bind to a large matrix molecule within the lamina densa region of skin basement membrane. At the site of these immune complexes, the epidermis separates from the dermis, which creates a subepidermal blister just below the lamina densa. The target molecule for the autoantibodies is in close apposition to fibronectin, a major extracellular matrix molecule that is abundant in the upper dermis of skin. In this report, we show specific affinity between fibronectin and the 290,000-D chain of the epidermolysis bullosa acquisita antigen, and that this affinity is mediated by the gelatin/collagen-binding domain of fibronectin (Mr = 60,000). Since blistering in epidermolysis bullosa acquisita often occurs in the absence of clinical and histological inflammation, a direct interruption in the fibronectin-epidermolysis bullosa acquisita antigen bond may be involved in the pathogenesis of epidermal-dermal disadherence that occurs in this bullous disease. Images PMID:3584471

  16. Focal dystonia in musicians.

    PubMed

    Aránguiz, R; Chana-Cuevas, P; Alburquerque, D; León, M

    2011-01-01

    A special group of focal dystonia is that known as occupational, which include dystonic disorders triggered by repetitive motor activity, closely associated with the professional activity of a specific task that the affected person performs. In this sense, musicians are a population particularly vulnerable to this disorder, which is presented during the execution of highly trained movements. This article reviews the pathophysiology of focal dystonia and its therapeutic implications. The pathophysiological basis of focal dystonia in the musician is still not well established. However, due to the contribution of neurophysiological studies and functional neuroimaging, there is growing evidence of anomalies in the processing of sensory information, sensory-motor integration, cortical and subcortical inhibitory processes, which underline this disease. Clinically, it is characterised by the appearance of involuntary muscle contractions, and is associated with loss of motor control while practicing music. It is a gradual appearance and sometimes there may be a history of musculoskeletal injuries or non-physiological postures preceding the appearance of the symptoms. The neurological examination is usually normal, although subtle dystonic postures can develop spontaneously or with movements that involve the affected segments. The dystonia remains focal and is not generalised. Treatment is based on using multiple strategies for the management of the dystonia, with variable results. Although a specific therapy has not been defined, there are general principles that are combined in each situation looking for results. This includes, among others, pharmacological interventions, management with botulinum toxin, and sensory re-training techniques. Copyright © 2010 Sociedad Española de Neurología. Published by Elsevier Espana. All rights reserved.

  17. Oral focal epithelial hyperplasia.

    PubMed

    López-Jornet, Pía; Camacho-Alonso, Fabio; Berdugo, Lucero

    2010-01-01

    Focal epithelial hyperplasia (FEH) is a benign, asymptomatic disease. It appears as papules, principally on the lower lip, although it can also be found on the retro-commissural mucosa and tongue and, less frequently, on the upper lip, gingiva and palate. FEH is caused by human papillomavirus subtype 13 or 32. The condition occurs in many populations and ethnic groups. We present the clinical case of a 31-year-old male with lesions that clinically and histologically corresponded to FEH.

  18. TES is a novel focal adhesion protein with a role in cell spreading.

    PubMed

    Coutts, Amanda S; MacKenzie, Elaine; Griffith, Elen; Black, Donald M

    2003-03-01

    Previously, we identified TES as a novel candidate tumour suppressor gene that mapped to human chromosome 7q31.1. In this report we demonstrate that the TES protein is localised at focal adhesions, actin stress fibres and areas of cell-cell contact. TES has three C-terminal LIM domains that appear to be important for focal adhesion targeting. Additionally, the N-terminal region is important for targeting TES to actin stress fibres. Yeast two-hybrid and biochemical analyses yielded interactions with several focal adhesion and/or cytoskeletal proteins including mena, zyxin and talin. The fact that TES localises to regions of cell adhesion suggests that it functions in events related to cell motility and adhesion. In support of this, we demonstrate that fibroblasts stably overexpressing TES have an increased ability to spread on fibronectin.

  19. Identification of a fibronectin receptor specific for rat liver endothelial cells

    SciTech Connect

    Johansson, S.; Gustafson, S.; Pertoft, H. )

    1987-10-01

    Antibodies raised against the fibronectin receptor of rat hepatocytes recognized one protein in immunoblotting of solubilized rat liver endothelial cells (LEC). The antibodies specifically precipitated a 200-kDa protein together with the 135-kDa component from {sup 125}I-labeled LEC. Spreading of LEC on fibronectin, but not on laminin or collagen, was inhibited by monovalent Fab fragments of the antibodies, implicating that the 135/200-kDa complex is a specific fibronectin receptor. The results indicate that LEC, hepatocytes, and fibroblasts of rat carry different fibronectin receptors, suggesting that the interaction of fibronectin with these cells may have different functional roles.

  20. Inhibition of fibronectin binding and fibronectin-mediated cell adhesion to collagen by a peptide from the second type I repeat of thrombospondin

    PubMed Central

    1993-01-01

    The platelet and extracellular matrix glycoprotein thrombospondin interacts with various types of cells as both a positive and negative modulator of cell adhesion, motility, and proliferation. These effects may be mediated by binding of thrombospondin to cell surface receptors or indirectly by binding to other extracellular matrix components. The role of peptide sequences from the type I repeats of thrombospondin in its interaction with fibronectin were investigated. Fibronectin bound specifically to the peptide Gly-Gly-Trp-Ser-His-Trp from the second type I repeat of thrombospondin but not to the corresponding peptides from the first or third repeats or flanking sequences from the second repeat. The two Trp residues and the His residue were essential for binding, and the two Gly residues enhanced the affinity of binding. Binding of the peptide and intact thrombospondin to fibronectin were inhibited by the gelatin-binding domain of fibronectin. The peptide specifically inhibited binding of fibronectin to gelatin or type I collagen and inhibited fibronectin-mediated adhesion of breast carcinoma and melanoma cells to gelatin or type I collagen substrates but not direct adhesion of the cells to fibronectin, which was inhibited by the peptide Gly-Arg-Gly-Asp-Ser. Thus, the fibronectin- binding thrombospondin peptide Gly-Gly-Trp-Ser-His-Trp is a selective inhibitor of fibronectin-mediated interactions of cells with collagen in the extracellular matrix. PMID:8468356

  1. Fibronectin in tissue regeneration: timely disassembly of the scaffold is necessary to complete the build.

    PubMed

    Stoffels, Josephine M J; Zhao, Chao; Baron, Wia

    2013-11-01

    Tissue injury initiates extracellular matrix molecule expression, including fibronectin production by local cells and fibronectin leakage from plasma. To benefit tissue regeneration, fibronectin promotes opsonization of tissue debris, migration, proliferation, and contraction of cells involved in the healing process, as well as angiogenesis. When regeneration proceeds, the fibronectin matrix is fully degraded. However, in a diseased environment, fibronectin clearance is often disturbed, allowing structural variants to persist and contribute to disease progression and failure of regeneration. Here, we discuss first how fibronectin helps tissue regeneration, with a focus on normal cutaneous wound healing as an example of complete tissue recovery. Then, we continue to argue that, although the fibronectin matrix generated following cartilage and central nervous system white matter (myelin) injury initially benefits regeneration, fibronectin clearance is incomplete in chronic wounds (skin), osteoarthritis (cartilage), and multiple sclerosis (myelin). Fibronectin fragments or aggregates persist, which impair tissue regeneration. The similarities in fibronectin-mediated mechanisms of frustrated regeneration indicate that complete fibronectin clearance is a prerequisite for recovery in any tissue. Also, they provide common targets for developing therapeutic strategies in regenerative medicine.

  2. Extracellular Matrix Fibronectin Stimulates the Self-Assembly of Microtissues on Native Collagen Gels

    PubMed Central

    Sevilla, Carlos A.; Dalecki, Diane

    2010-01-01

    Fibronectin is an adhesive glycoprotein that is polymerized into extracellular matrices via a tightly regulated, cell-dependent process. Here, we demonstrate that fibronectin matrix polymerization induces the self-assembly of multicellular structures in vitro, termed tissue bodies. Fibronectin-null mouse embryonic fibroblasts adherent to compliant gels of polymerized type I collagen failed to spread or proliferate. In contrast, addition of fibronectin to collagen-adherent fibronectin-null mouse embryonic fibroblasts resulted in a dose-dependent increase in cell number, and induced the formation of three-dimensional (3D) multicellular structures that remained adherent and well-spread on the native collagen substrate. An extensive fibrillar fibronectin matrix formed throughout the microtissue. Blocking fibronectin matrix polymerization inhibited both cell proliferation and microtissue formation, demonstrating the importance of fibronectin fibrillogenesis in triggering cellular self-organization. Cell proliferation, tissue body formation, and tissue body shape were dependent on both fibronectin and collagen concentrations, suggesting that the relative proportion of collagen and fibronectin fibrils polymerized into the extracellular matrix influences the extent of cell proliferation and the final shape of microtissues. These data demonstrate a novel role for cell-mediated fibronectin fibrillogenesis in the formation and vertical assembly of microtissues, and provide a novel approach for engineering complex tissue architecture. PMID:20673131

  3. Co-assembly of plasma and cellular fibronectins into fibrils in human fibroblast cultures

    PubMed Central

    1990-01-01

    Exogenous plasma and endogenous cellular fibronectins on the surface of cultured fibroblasts and in extracellular matrix fibrils were colocalized by fluorescent and high voltage immunoelectron microscopy. Fibroblast cultures grown in the presence or absence of cycloheximide were incubated with exogenous plasma fibronectin labeled with fluorescein isothiocyanate. A monoclonal antibody specific for the EIIIA sequence of cellular fibronectin was used to detect cellular fibronectin. A rabbit antifluorescein antibody identified fluoresceinated plasma fibronectin. In cultures incubated in the presence of cycloheximide, plasma fibronectin was bound to the cell surface and was assembled into extracellular fibrils. In cultures grown in the absence of cycloheximide, plasma and cellular fibronectins were observed in the same matrix fibrils and in the same locations on the cell surface. There was not, however, random admixture of the two proteins. PMID:2114411

  4. Fibroblast-fibronectin patterning and network formation in 3D fibrin matrices.

    PubMed

    Miron-Mendoza, Miguel; Graham, Eric; Manohar, Sujal; Petroll, W Matthew

    2017-06-07

    We previously reported that fibroblasts migrating within 3-D collagen matrices move independently, whereas fibroblasts within 3-D fibrin matrices form an interconnected network. Similar networks have been identified previously during in vivo corneal wound healing. In this study, we investigate the role of fibronectin in mediating this mechanism of collective cell spreading, migration and patterning. To assess cell spreading, corneal fibroblasts were plated within fibrillar collagen or fibrin matrices. To assess migration, compacted cell-populated collagen matrices were nested inside cell-free fibrin matrices. Constructs were cultured in serum-free media containing PDGF, with or without RGD peptide, anti-α5 or anti-fibronectin blocking antibodies. In some experiments, LifeAct and fluorescent fibronectin were used to allow dynamic assessment of cell-induced fibronectin reorganization. 3-D and 4-D imaging were used to assess cell mechanical behavior, connectivity, F-actin, α5 integrin and fibronectin organization. Corneal fibroblasts within 3-D fibrin matrices formed an interconnected network that was lined with cell-secreted fibronectin. Live cell imaging demonstrated that fibronectin tracks were formed at the leading edge of spreading and migrating cells. Furthermore, fibroblasts preferentially migrated through fibronectin tracks laid down by other cells. Interfering with cell-fibronectin binding with RGD, anti α5 integrin or anti fibronectin antibodies inhibited cell spreading and migration through fibrin, but did not affect cell behavior in collagen. In this study, a novel mode of cell patterning was identified in which corneal fibroblasts secrete and attach to fibronectin via α5β1 integrin to facilitate spreading and migration within 3-D fibrin matrices, resulting in the formation of localized fibronectin tracks. Other cells use these fibronectin tracks as conduits, resulting in an interconnected cell-fibronectin network. Copyright © 2017 International

  5. Focal adhesions in osteoneogenesis

    PubMed Central

    Biggs, M.J.P; Dalby, M.J

    2010-01-01

    As materials technology and the field of tissue engineering advances, the role of cellular adhesive mechanisms, in particular the interactions with implantable devices, becomes more relevant in both research and clinical practice. A key tenet of medical device technology is to use the exquisite ability of biological systems to respond to the material surface or chemical stimuli in order to help develop next-generation biomaterials. The focus of this review is on recent studies and developments concerning focal adhesion formation in osteoneogenesis, with an emphasis on the influence of synthetic constructs on integrin mediated cellular adhesion and function. PMID:21287830

  6. [Focal epithelial hyperplasia].

    PubMed

    Delgado, Yolanda; Torrelo, Antonio; Colmenero, Isabel; Zambrano, Antonio

    2005-12-01

    Focal epithelial hyperplasia (FEH) is a benign proliferation of the oral mucosa with well defined clinical and histological characteristics. It has been associated with infection of the oral mucosa by types 13 and 32 of the human papillomavirus (HPV), and to a lesser extent, with other types. Its clinical course is variable, although it usually persists for months or years; cases with spontaneous resolution have been described, as have others with prolonged persistence. We present the case of an Ecuadorian boy whose visit was motivated by lesions in the oral mucosa consistent with a diagnosis of FEH, which were confirmed in the histological study, and in which HPV type 13 DNA was identified.

  7. iDriving (Intelligent Driving)

    SciTech Connect

    Malikopoulos, Andreas

    2012-09-17

    iDriving identifies the driving style factors that have a major impact on fuel economy. An optimization framework is used with the aim of optimizing a driving style with respect to these driving factors. A set of polynomial metamodels is constructed to reflect the responses produced in fuel economy by changing the driving factors. The optimization framework is used to develop a real-time feedback system, including visual instructions, to enable drivers to alter their driving styles in responses to actual driving conditions to improve fuel efficiency.

  8. Allosteric Regulation of Fibronectin/α5β1 Interaction by Fibronectin-Binding MSCRAMMs

    PubMed Central

    Liang, Xiaowen; Garcia, Brandon L.; Visai, Livia; Prabhakaran, Sabitha; Meenan, Nicola A. G.; Potts, Jennifer R.; Humphries, Martin J.; Höök, Magnus

    2016-01-01

    Adherence of microbes to host tissues is a hallmark of infectious disease and is often mediated by a class of adhesins termed MSCRAMMs (Microbial Surface Components Recognizing Adhesive Matrix Molecules). Numerous pathogens express MSCRAMMs that specifically bind the heterodimeric human glycoprotein fibronectin (Fn). In addition to roles in adhesion, Fn-binding MSCRAMMs exploit physiological Fn functions. For example, several pathogens can invade host cells by a mechanism whereby MSCRAMM-bound Fn bridges interaction with α5β1 integrin. Here, we investigate two Fn-binding MSCRAMMs, FnBPA (Staphylococcus aureus) and BBK32 (Borrelia burgdorferi) to probe structure-activity relationships of MSCRAMM-induced Fn/α5β1integrin activation. Circular dichroism, fluorescence resonance energy transfer, and dynamic light scattering techniques uncover a conformational rearrangement of Fn involving domains distant from the MSCRAMM binding site. Surface plasmon resonance experiments demonstrate a significant enhancement of Fn/α5β1 integrin affinity in the presence of FnBPA or BBK32. Detailed kinetic analysis of these interactions reveal that this change in affinity can be attributed solely to an increase in the initial Fn/α5β1 on-rate and that this rate-enhancement is dependent on high-affinity Fn-binding by MSCRAMMs. These data implicate MSCRAMM-induced perturbation of specific intramolecular contacts within the Fn heterodimer resulting in activation by exposing previously cryptic α5β1 interaction motifs. By correlating structural changes in Fn to a direct measurement of increased Fn/α5β1 affinity, this work significantly advances our understanding of the structural basis for the modulation of integrin function by Fn-binding MSCRAMMs. PMID:27434228

  9. Nuclear transport of paxillin depends on focal adhesion dynamics and FAT domains

    PubMed Central

    Sathe, Aneesh R.; Shivashankar, G. V.; Sheetz, Michael P.

    2016-01-01

    ABSTRACT The nuclear transport of paxillin appears to be crucial for paxillin function but the mechanism of transport remains unclear. Here, we show that the nuclear transport of paxillin is regulated by focal adhesion turnover and the presence of FAT domains. Focal adhesion turnover was controlled using triangular or circular fibronectin islands. Circular islands caused higher focal adhesion turnover and increased the nuclear transport of paxillin relative to triangular islands. Mutating several residues of paxillin had no effect on its nuclear transport, suggesting that the process is controlled by multiple domains. Knocking out FAK (also known as PTK2) and vinculin caused an increase in nuclear paxillin. This could be reversed by rescue with wild-type FAK but not by FAK with a mutated FAT domain, which inhibits paxillin binding. Expressing just the FAT domain of FAK not only brought down nuclear levels of paxillin but also caused a large immobile fraction of paxillin to be present at focal adhesions, as demonstrated by fluorescence recovery after photobleaching (FRAP) studies. Taken together, focal adhesion turnover and FAT domains regulate the nuclear localization of paxillin, suggesting a possible role for transcriptional control, through paxillin, by focal adhesions. PMID:27068537

  10. Dexamethasone-Mediated Activation of Fibronectin Matrix Assembly Reduces Dispersal of Primary Human Glioblastoma Cells

    PubMed Central

    Shannon, Stephen; Vaca, Connan; Jia, Dongxuan; Entersz, Ildiko; Schaer, Andrew; Carcione, Jonathan; Weaver, Michael; Avidar, Yoav; Pettit, Ryan; Nair, Mohan; Khan, Atif; Foty, Ramsey A.

    2015-01-01

    Despite resection and adjuvant therapy, the 5-year survival for patients with Glioblastoma multiforme (GBM) is less than 10%. This poor outcome is largely attributed to rapid tumor growth and early dispersal of cells, factors that contribute to a high recurrence rate and poor prognosis. An understanding of the cellular and molecular machinery that drive growth and dispersal is essential if we are to impact long-term survival. Our previous studies utilizing a series of immortalized GBM cell lines established a functional causation between activation of fibronectin matrix assembly (FNMA), increased tumor cohesion, and decreased dispersal. Activation of FNMA was accomplished by treatment with Dexamethasone (Dex), a drug routinely used to treat brain tumor related edema. Here, we utilize a broad range of qualitative and quantitative assays and the use of a human GBM tissue microarray and freshly-isolated primary human GBM cells grown both as conventional 2D cultures and as 3D spheroids to explore the role of Dex and FNMA in modulating various parameters that can significantly influence tumor cell dispersal. We show that the expression and processing of fibronectin in a human GBM tissue-microarray is variable, with 90% of tumors displaying some abnormality or lack in capacity to secrete fibronectin or assemble it into a matrix. We also show that low-passage primary GBM cells vary in their capacity for FNMA and that Dex treatment reactivates this process. Activation of FNMA effectively “glues” cells together and prevents cells from detaching from the primary mass. Dex treatment also significantly increases the strength of cell-ECM adhesion and decreases motility. The combination of increased cohesion and decreased motility discourages in vitro and ex vivo dispersal. By increasing cell-cell cohesion, Dex also decreases growth rate of 3D spheroids. These effects could all be reversed by an inhibitor of FNMA and by the glucocorticoid receptor antagonist, RU-486. Our

  11. Curli mediate bacterial adhesion to fibronectin via tensile multiple bonds

    NASA Astrophysics Data System (ADS)

    Oh, Yoo Jin; Hubauer-Brenner, Michael; Gruber, Hermann J.; Cui, Yidan; Traxler, Lukas; Siligan, Christine; Park, Sungsu; Hinterdorfer, Peter

    2016-09-01

    Many enteric bacteria including pathogenic Escherichia coli and Salmonella strains produce curli fibers that bind to host surfaces, leading to bacterial internalization into host cells. By using a nanomechanical force-sensing approach, we obtained real-time information about the distribution of molecular bonds involved in the adhesion of curliated bacteria to fibronectin. We found that curliated E. coli and fibronectin formed dense quantized and multiple specific bonds with high tensile strength, resulting in tight bacterial binding. Nanomechanical recognition measurements revealed that approximately 10 bonds were disrupted either sequentially or simultaneously under force load. Thus the curli formation of bacterial surfaces leads to multi-bond structural components of fibrous nature, which may explain the strong mechanical binding of curliated bacteria to host cells and unveil the functions of these proteins in bacterial internalization and invasion.

  12. Curli mediate bacterial adhesion to fibronectin via tensile multiple bonds

    PubMed Central

    Oh, Yoo Jin; Hubauer-Brenner, Michael; Gruber, Hermann J.; Cui, Yidan; Traxler, Lukas; Siligan, Christine; Park, Sungsu; Hinterdorfer, Peter

    2016-01-01

    Many enteric bacteria including pathogenic Escherichia coli and Salmonella strains produce curli fibers that bind to host surfaces, leading to bacterial internalization into host cells. By using a nanomechanical force-sensing approach, we obtained real-time information about the distribution of molecular bonds involved in the adhesion of curliated bacteria to fibronectin. We found that curliated E. coli and fibronectin formed dense quantized and multiple specific bonds with high tensile strength, resulting in tight bacterial binding. Nanomechanical recognition measurements revealed that approximately 10 bonds were disrupted either sequentially or simultaneously under force load. Thus the curli formation of bacterial surfaces leads to multi-bond structural components of fibrous nature, which may explain the strong mechanical binding of curliated bacteria to host cells and unveil the functions of these proteins in bacterial internalization and invasion. PMID:27652888

  13. Suberoylanilide hydroxamic acid (SAHA) at subtoxic concentrations increases the adhesivity of human leukemic cells to fibronectin.

    PubMed

    Kuzelová, Katerina; Pluskalová, Michaela; Brodská, Barbora; Otevrelová, Petra; Elknerová, Klára; Grebenová, Dana; Hrkal, Zbynek

    2010-01-01

    Suberoylanilide hydroxamic acid (SAHA) is an inhibitor of histone deacetylases (HDACs) which is being introduced into clinic for the treatment of hematological diseases. We studied the effect of this compound on six human hematopoietic cell lines (JURL-MK1, K562, CML-T1, Karpas-299, HL-60, and ML-2) as well as on normal human lymphocytes and on leukemic primary cells. SAHA induced dose-dependent and cell type-dependent cell death which displayed apoptotic features (caspase-3 activation and apoptotic DNA fragmentation) in most cell types including the normal lymphocytes. At subtoxic concentrations (0.5-1 microM), SAHA increased the cell adhesivity to fibronectin (FN) in all leukemia/lymphoma-derived cell lines but not in normal lymphocytes. This increase was accompanied by an enhanced expression of integrin beta1 and paxillin, an essential constituent of focal adhesion complexes, both at the protein and mRNA level. On the other hand, the inhibition of ROCK protein, an important regulator of cytoskeleton structure, had no consistent effect on SAHA-induced increase in the cell adhesivity. The promotion of cell adhesivity to FN seems to be specific for SAHA as we observed no such effects with other HDAC inhibitors (trichostatin A and sodium butyrate).

  14. Fibronectin alters the rate of formation and structure of the fibrin matrix.

    PubMed

    Ramanathan, Anand; Karuri, Nancy

    2014-01-10

    Plasma fibronectin is a vital component of the fibrin clot; however its role on clot structure is not clearly understood. The goal of this study was to examine the influence of fibronectin on the kinetics of formation, structural characteristics and composition of reconstituted fibrin clots or fibrin matrices. Fibrin matrices were formed by adding thrombin to 1, 2 or 4 mg/ml fibrinogen supplemented with 0-0.4 mg/ml fibronectin. The rate of fibrin matrix formation was then monitored by measuring light absorbance properties at different time points. Confocal microscopy of fluorescein conjugated fibrinogen was used to visualize the structural characteristics of fibrin matrices. The amount of fibronectin in fibrin matrices was determined through electrophoresis and immunoblotting of solubilized matrices. Fibronectin concentration positively correlated with the initial rate of fibrin matrix formation and with steady state light absorbance values of fibrin matrices. An increase in fibronectin concentration resulted in thinner and denser fibers in the fibrin matrices. Electrophoresis and immunoblotting showed that fibronectin was covalently and non-covalently bound to fibrin matrices and in the form of high molecular weight multimers. The formation of fibronectin multimers was attributed to cross-linking of fibronectin by trace amounts Factor XIIIa. These findings are novel because they link results from light absorbance studies to microcopy analyses and demonstrate an influence of fibronectin on fibrin matrix structural characteristics. This data is important in developing therapies that destabilize fibrin clots.

  15. Definition of fibronectin-mediated uptake of gelatinized latex by liver slices and macrophages.

    PubMed

    Molnar, J; Galles, M; Beezhold, D; Lai, M Z; Ku, C S; Van Alten, P J

    1987-10-01

    These studies show that both liver slices and macrophages carried out fibronectin concentration-dependent uptake of 125I-labeled gelatin-coated latex (test latex). Lack of phagocytosis of test latex by liver slices was shown directly by electron microscopy and indirectly by trypsin treatment, which caused the release of all test latex taken up in response to fibronectin. Inhibitors of phagocytosis did not alter this uptake. On the other hand, trypsin released only a portion of test latex from macrophages. Inhibitors of phagocytosis did not effect the released radioactive particles from macrophages but greatly reduced the trypsin-resistant radioactivity, taken as representing phagocytized particles. Opsonization of test latex with fibronectin did not require heparin but its association with liver slices occurred only in the presence of heparin. Macrophages, however, readily bound and internalized the opsonized test latex and heparin only potentiated these reactions. Gelatin competed with test latex for fibronectin for opsonization, but did not inhibit binding and phagocytosis of fibronectin-test latex complexes. Finally, soluble fibronectin-gelatin complexes did not compete for binding and phagocytosis of fibronectin-test latex complexes. Thus, fibronectin concentrated on the surface of latex is preferred for interaction with the fibronectin receptor of macrophages. Gelatin, however, was not essential for this reaction, because fibronectin directly coupled to latex was also readily taken up.

  16. Fibronectin deposition in delayed-type hypersensitivity. Reactions of normals and a patient with afibrinogenemia.

    PubMed Central

    Clark, R A; Horsburgh, C R; Hoffman, A A; Dvorak, H F; Mosesson, M W; Colvin, R B

    1984-01-01

    During development of delayed hypersensitivity (DH) skin reactions, fibronectin accumulates in two distinct sites: (a) the dermal interstitium in a pattern similar to fibrin and with a time course similar to that of fibrin deposition and mononuclear cell infiltration, and (b) blood vessel walls in a pattern suggestive of basement membrane staining and with a time course similar to that of endothelial cell proliferation. In vitro fibronectin can bind to monocytes or endothelial cells and simultaneously bind to fibrin or collagen matrices; by such interaction in vivo it may affect cell migration or proliferation. Thus, fibronectin deposition in DH reactions may facilitate cell-matrix interactions; however, the possibility exists that extravascular fibronectin accumulation may be only secondary to interstitial fibrin clot formation, and that blood vessel-associated fibronectin may be only a function of adsorption onto basement membrane (type IV) collagen. To address these possibilities, we investigated the association of fibronectin with fibrin, type IV collagen, and mononuclear cell infiltrates in DH reactions. Skin sites of DH reactions in normal volunteers were biopsied at 24, 48, and 72 h after intradermal challenge and examined by immunofluorescence technique. At all time points most of the interstitial fibronectin coincided with fibrin; however, some interstitial fibronectin was coincident with mononuclear cells positive for HLA-DR or monocyte-specific antigen. The coincidence of fibronectin with mononuclear cells was more apparent in a 48-h DH reaction from a patient with congenital afibrinogenemia. Vessel wall fibronectin was increased by 48 h after challenge and appeared as a fine linear band on the luminal side of a much thicker band of type IV collagen. Thus, the coincidence of extravascular fibronectin with mononuclear cells, its appearance without fibrin in the site from a patient with afibrinogenemia, and incomplete correspondence of vessel wall

  17. Fish Rhabdovirus Cell Entry Is Mediated by Fibronectin

    PubMed Central

    Bearzotti, Monique; Delmas, Bernard; Lamoureux, Annie; Loustau, Anne-Marie; Chilmonczyk, Stefan; Bremont, Michel

    1999-01-01

    Three monoclonal antibodies (MAbs) generated against rainbow trout gonad cells (RTG-2) have been selected for their ability to protect cells from the viral hemorrhagic septicemia virus (VHSV) infection, a salmonid rhabdovirus. Protection from infection was restricted to the salmonid-derived cell lines indicating species specificity of the blocking MAbs. Surprisingly, the blocking activity of these MAbs was also effective against other nonantigenically related fish rhabdoviruses. Indirect immunofluorescence and immunoelectron microscopy observations demonstrated that the three MAbs were all directed against an abundant cell plasma membrane component, and immunoprecipitation studies indicated that the target consisted of a heterodimeric complex with molecular masses of 200 and 44 kDa. Biochemical data provided the following evidence that fibronectin is part of this complex and that it could represent the main receptor for fish rhabdoviruses. (i) An antiserum generated against the 200-kDa protein reacted against the recombinant rainbow trout fibronectin expressed in Escherichia coli. (ii) The purified rainbow trout fibronectin was able to bind specifically to VHSV. To our knowledge, this is the first identification of a cellular component acting as a primary receptor for a virus replicating in lower vertebrates and, more interestingly, for viruses belonging to the Rhabdoviridae family. PMID:10438860

  18. Fibronectin Extra Domain A Promotes Liver Sinusoid Repair following Hepatectomy

    PubMed Central

    Sackey-Aboagye, Bridget; Olsen, Abby L.; Mukherjee, Sarmistha M.; Ventriglia, Alexander; Yokosaki, Yasuyuki; Greenbaum, Linda E.; Lee, Gi Yun; Naga, Hani

    2016-01-01

    Liver sinusoidal endothelial cells (LSECs) are the main endothelial cells in the liver and are important for maintaining liver homeostasis as well as responding to injury. LSECs express cellular fibronectin containing the alternatively spliced extra domain A (EIIIA-cFN) and increase expression of this isoform after liver injury, although its function is not well understood. Here, we examined the role of EIIIA-cFN in liver regeneration following partial hepatectomy. We carried out two-thirds partial hepatectomies in mice lacking EIIIA-cFN and in their wild type littermates, studied liver endothelial cell adhesion on decellularized, EIIIA-cFN-containing matrices and investigated the role of cellular fibronectins in liver endothelial cell tubulogenesis. We found that liver weight recovery following hepatectomy was significantly delayed and that sinusoidal repair was impaired in EIIIA-cFN null mice, especially females, as was the lipid accumulation typical of the post-hepatectomy liver. In vitro, we found that liver endothelial cells were more adhesive to cell-deposited matrices containing the EIIIA domain and that cellular fibronectin enhanced tubulogenesis and vascular cord formation. The integrin α9β1, which specifically binds EIIIA-cFN, promoted tubulogenesis and adhesion of liver endothelial cells to EIIIA-cFN. Our findings identify a role for EIIIA-cFN in liver regeneration and tubulogenesis. We suggest that sinusoidal repair is enhanced by increased LSEC adhesion, which is mediated by EIIIA-cFN. PMID:27741254

  19. Molecular simulation of fibronectin adsorption onto polyurethane surfaces.

    PubMed

    Panos, Melisa; Sen, Taner Z; Ahunbay, M Göktuğ

    2012-08-28

    Poly(ethylene glycol)-based polyurethanes have been widely used in biomedical applications; however, they are prone to swelling. A natural polyol, castor oil, can be incorporated into these polyurethanes to control the degree of the swelling, which alters mechanical properties and protein adsorption characteristic of the polymers. In this work, we modeled poly(ethylene glycol) and castor oil copolymers of hexamethylene diisocyanate-based polyurethanes (PEG-HDI and CO-HDI, respectively) and compared their mechanisms for fibronectin adsorption using molecular mechanics and molecular dynamics simulations. Results showed that the interplay between the hydrophobic residues concentrated at the N-terminal end of the protein, the surface roughness, and the hydrophilicity of the polymer surface determine the overall protein adsorption affinity. Incorporating explicit water molecules in the simulations results in higher affinity for fibronectin adsorption to more hydrophobic surface of CO-HDI surfaces, emphasizing the role that water molecules play during adsorption. We also observed that the strain energies that are indicative of flexibility and consequently entropy are significantly affected by the changes in the patterns of β-sheet formation/breaking. Our study lends supports to the view that while castor oil controls the degree of swelling, it increases the adsorption of fibronectin to a limited extent due to the interplay between its hydrophobicity and its surface roughness, which needs to be taken into account during the design of polyurethane-based biomaterials.

  20. TENASCIN AND FIBRONECTIN IN PLEOMORPHIC ADENOMA OF THE SALIVARY GLAND

    PubMed Central

    Bento, Patrícia Meira; Freitas, Roseana de Almeida; Pinto, Leão Pereira; de Souza, Lélia Batista

    2006-01-01

    Objectives: To analyze the expression and distribution pattern of extracellular matrix components in pleomorphic adenomas of the major and minor salivary glands and to compare the morphological findings of these tumors with the immunohistochemical expression, considering the different types of stroma predominating in each case. Methods and Results: The expression of tenascin (TN) and fibronectin (FN) was analyzed in 23 cases of pleomorphic adenomas, 11 major and 12 minor salivary gland tumors, by the streptavidin-biotin method using anti-tenascin and anti-fibronectin antibodies. In addition, the immunohistochemical results were correlated with the morphological findings of the lesions. All cases analyzed were immunoreactive for the antibodies used. Fibronectin showed strong labeling in fibrous and chondroid stroma, while labeling was weak in hyaline and myxoid stroma. Tenascin expression was more intense in fibrous and chondroid stroma and moderate in hyaline and myxoid stroma. Conclusions: No difference in the expression of these proteins was observed between major and minor salivary gland tumors. PMID:19089073

  1. Semaphorin 3A Increases FAK Phosphorylation at Focal Adhesions to Modulate MDA-MB-231 Cell Migration and Spreading on Different Substratum Concentrations

    PubMed Central

    Compere, Frances V.; Miller, Alex M.

    2017-01-01

    Interactions between integrin-mediated adhesions and the extracellular matrix (ECM) are important regulators of cell migration and spreading. However, mechanisms by which extracellular ligands regulate cell migration and spreading in response to changes in substratum concentration are not well understood. Semaphorin 3A (Sema3A) has been shown to inhibit cell motility and alter integrin signaling in various cell types. We propose that Sema3A alters focal adhesions to modulate breast carcinoma cell migration and spreading on substrata coated with different concentrations of ECM. We demonstrate that Sema3A inhibits MDA-MB-231 cell migration and spreading on substrata coated with high concentrations of collagen and fibronectin but enhances migration and spreading at lower concentrations of collagen and fibronectin. Sema3A increases focal adhesion kinase phosphorylation at tyrosine 397 (pFAK397) at focal adhesions on all substratum concentrations of collagen and fibronectin but decreased pFAK397 levels on laminin. Rho-associated protein kinase (ROCK) inhibition blocks the Sema3A-mediated effects on cell migration, spreading, and pFAK397 at focal adhesions when cultured on all concentrations of collagen. These results suggest that Sema3A shifts the optimal level of cell-matrix adhesions to a nonoptimal ECM coating concentration, in particular collagen, to yield maximal cell migration and spreading that may be mediated through a ROCK-dependent mechanism. PMID:28182100

  2. Impaired Driving

    MedlinePlus

    Impaired driving is dangerous. It's the cause of more than half of all car crashes. It means operating a ... texting Having a medical condition which affects your driving For your safety and the safety of others, ...

  3. Focal plane polarimeter design

    SciTech Connect

    McClelland, J.B.

    1983-10-12

    Measurement of polarization transfer or so-called triple-scattering parameters have been made recently for proton-nucleon scattering at TRIUMF, SIN, and LAMPF using carbon polarimeters and have been essential in determining the proton-nucleon amplitudes up to 800 MeV. An extension to the case is described where the scattered proton polarization is analyzed after passage through some type of spectrometer. Most of the experience with this type of focal plane polarimeter (FPP) has been gained in the field of proton-nucleus scattering at intermediate energies but is certainly not confined to such specific cases. The salient features of an FPP are emphasized by describing a minimal system which includes all the necessary components then go on to a more complete system. 10 references. (WHK)

  4. Extensive focal epithelial hyperplasia.

    PubMed

    Hashemipour, Maryam Alsadat; Shoryabi, Ali; Adhami, Shahrzad; Mehrabizadeh Honarmand, Hoda

    2010-01-01

    Heck's disease or focal epithelial hyperplasia is a benign contagious disease caused by human papillomavirus types 13 or 32. It occurs with low frequency in the Iranian population. This condition is characterized by the occurrence of multiple, small papules or nodules in the oral cavity, especially on the labial and buccal mucosa and tongue. In some populations, up to 39% of children are affected. Conservative surgical excision of lesions may be performed for diagnostic or aesthetic purposes. The risk of recurrence after this therapy is minimal, and there seems to be no malignant transformation potential. In the present work, we presented the clinical case of a 12-year-old Iranian girl with oral lesions that clinically and histologically correspond to Heck's disease.

  5. Fibronectin changes in eosinophilic meningitis with blood-CSF barrier disruption.

    PubMed

    Shyu, Ling-Yuh; Hu, Ming-E; Chou, Chun-Hui; Chen, Ke-Min; Chiu, Ping-Sung; Lai, Shih-Chan

    2015-01-01

    Fibronectin, which is present at relatively low levels in healthy central nervous systems (CNS), shows increased levels in meningitis. In this study, fibronectin processing was correlated with the increased permeability of the blood-cerebrospinal fluid (CSF) barrier as well as with the formation of eosinophil infiltrates in angiostrongyliasis meningitis. The immunohistochemistry results show matrix metalloproteinase-9 (MMP-9) is localized in the choroid plexus epithelium. Coimmunoprecipitation demonstrated fibronectin strongly binds MMP-9. Furthermore, treatment with the MMP-9 inhibitor GM6001 significantly inhibited fibronectin processing, reduced the blood-CSF barrier permeability, and decreased the eosinophil counts. The decreased fibronectin processing in CSF implies decreased cellular invasion of the subarachnoid space across the blood-CSF barrier. Therefore, increased fibronectin processing may be associated with barrier disruption and participate in the extravasation and migration of eosinophils into the CNS during experimental parasitic infection. Copyright © 2015 Elsevier Inc. All rights reserved.

  6. Mechanism of acute depletion of plasma fibronectin following thermal injury in rats. Appearance of a gelatinlike ligand in plasma

    SciTech Connect

    Deno, D.C.; McCafferty, M.H.; Saba, T.M.; Blumenstock, F.A.

    1984-01-01

    Plasma fibronectin was depleted within 15 min following sublethal burn, followed by partial recovery at 8 h and complete restoration by 24 h in anesthetized rats. Radiolabeled /sup 75/Se-plasma fibronectin, injected intravenously before burn, was rapidly sequestered in burn skin as well as the liver. Fibronectin levels at 2 h postburn as detected by immunoassay vs. /sup 75/Se-plasma fibronectin indicated that more fibronectin was in the plasma than detected by electroimmunoassay. Crossed immunoelectrophoretic analysis of fibronectin in early postburn plasma demonstrated a reduced electrophoretic mobility of the fibronectin antigen. Addition of heparin or fibrin, both of which have affinity for fibronectin, to normal plasma was unable to reproduce this altered fibronectin electrophoretic pattern. In contrast, addition of gelatin or native collagen to normal plasma reproduced the abnormal electrophoretic pattern of fibronectin seen in burn plasma. Extracts of burned skin, but not extracts of normal skin, when added to normal plasma, elicited a similar altered electrophoretic pattern for fibronectin. By gel filtration, fibronectin in burn plasma had an apparent molecular weight approximately 40% greater than that observed in normal plasma. These data suggest the release into the blood of a gelatinlike ligand from burned skin, which complexes with plasma fibronectin. Thus, fibronectin deficiency acutely postburn appears mediated by (a) its accumulation at the site of burn injury; (b) its removal from the circulation by the liver; and (c) its presence in the plasma in a form that is less detectable by immunoassay.

  7. Integrins in point contacts mediate cell spreading: factors that regulate integrin accumulation in point contacts vs. focal contacts

    PubMed Central

    1993-01-01

    We have studied the function and distribution of the alpha 1 beta 1, alpha 5 beta 1 and alpha 6 beta 1 heterodimers on type-1 astrocytes with antibodies specific for integrin subunits (alpha 1, alpha 5, alpha 6, and beta 1). The alpha 1 beta 1 heterodimer mediates adhesion to laminin and collagen, the alpha 5 beta 1 to fibronectin in an RGD- dependent manner. The alpha 5 beta 1 integrin is found in focal contacts in long-term cultures of well-spread astrocytes colocalizing with vinculin and the termini of actin stress fibers. alpha 1 beta 1 heterodimers can occasionally be found as small aggregates within focal contacts but they do not accumulate there. Instead, alpha 1 beta 1 integrins are found in punctate deposits called point contacts which are distributed over the upper and the lower cell surfaces whether laminin, collagen, fibronectin or polylysine is used as a substratum. Unlike focal contacts, point contacts contain clathrin but rarely codistribute with actin or vinculin. Two observations indicate that these point contacts are functional. First, mAb 3A3, directed against the rat alpha 1 subunit, inhibits the attachment of astrocytes to laminin and collagen. Second, during the spreading of astrocytes, a band of point contacts forms around the cell perimeter at a time when no focal contacts are visible. While alpha 1 beta 1 integrins are found only in point contacts in astrocytes, the alpha 6 beta 1 integrin, another laminin receptor, is localized within focal contacts. Moreover, alpha 1 beta 1 heterodimers accumulate in focal contacts in fibroblasts. Thus, the alpha subunit contributes, independent of its ligand, to functional integrin heterodimer accumulation in focal contacts or in point contacts. This accumulation varies among different cell types with apparently identical heterodimers as well as with the motile state (spreading vs. flattened) of the same cells. PMID:8416993

  8. Electromyogram-evoked focal myositis

    PubMed Central

    Snipes, George; Quan, Carolyn

    2017-01-01

    Focal myositis is a rarely reported inflammatory disease of skeletal muscle, particularly of an extremity. It is often misinterpreted as an infectious syndrome, leading to prolonged antibiotic use and a delay in immunosuppressive therapy. Without a confirmed etiology to date, we present a case of recurrent focal myositis following an electromyogram. PMID:28127151

  9. Staphylococcus aureus fibronectin binding proteins A and B possess a second fibronectin binding region that may have biological relevance to bone tissues.

    PubMed

    Williams, R J; Henderson, B; Nair, S P

    2002-05-01

    Staphylococcus aureus is a major human pathogen that has a propensity for targeting to bone tissues and thereby causing bone disease. A plausible hypothesis is that S. aureus targets to bone using the MSCRAMM family of surface proteins possessed by this organism. Two such proteins that have recently been shown to be important in bone infections are the S. aureus fibronectin binding proteins (FnBP) A and B. To identify fibronectin-binding domains from S. aureus that have biological relevance to bone, a phage display library of S. aureus genomic DNA was constructed and panned sequentially against immobilized fibronectin and cultured osteoblasts. Using this system, phage displaying a second fibronectin-binding region within the N-terminal part of FnBPA and FnBPB, which is distinct from the primary fibronectin-binding domain located within the D repeat region of these proteins, was isolated. Phage displaying this second region bound to both immobilized fibronectin and to osteoblasts and/or the extracellular matrix synthesized by these cells, thereby suggesting a biological relevance for these regions in S. aureus binding to bone tissues. Analysis of these binding regions for their ability to bind to other extracellular matrix proteins revealed a preference for fibronectin, with slight binding to fibrinogen and no binding to collagen or laminin.

  10. Fibronectin and alpha5 integrin regulate keratinocyte cell cycling. A mechanism for increased fibronectin potentiation of T cell lymphokine-driven keratinocyte hyperproliferation in psoriasis.

    PubMed Central

    Bata-Csorgo, Z; Cooper, K D; Ting, K M; Voorhees, J J; Hammerberg, C

    1998-01-01

    In addition to being T lymphocyte-driven, psoriasis may be due in part to abnormal integrin expression. Normal-appearing (uninvolved) skin from psoriatic patients was examined to determine whether altered fibronectin or its receptor expression is detectable before development of psoriatic lesions. In contrast to skin from normal subjects, we detect by immunofluorescence the abnormal presence of plasma fibronectin in the basal cell layer of the epidermis of psoriatic uninvolved skin. Furthermore, increased fibronectin exposure superinduces the in vitro cell cycle induction and expansion of psoriatic nonlesional keratinocytes in response to a cocktail of T cell lymphokines. Fibronectin alone also appeared to increase cell cycle entry among uninvolved but not normal keratinocytes. Concordantly, the alpha5 integrin fibronectin receptor, but not alpha2 or alpha3, is overexpressed in the in vivo nonlesional psoriatic epidermis. The involvement of alpha5beta1 in the early outgrowth of clonogenic keratinocytes in the ex vivo culture was demonstrated by the ability of anti-alpha5 mAb to inhibit keratinocyte growth on fibronectin. Thus, the fibronectin receptor appears to be one of the components required for the development of the hyperresponsiveness of psoriatic keratinocytes to signals for proliferation provided by lymphokines produced by intralesional T lymphocytes in psoriasis. PMID:9525994

  11. Self-assembly of fibronectin mimetic peptide-amphiphile nanofibers

    NASA Astrophysics Data System (ADS)

    Rexeisen, Emilie Lynn

    Many therapeutic strategies incorporate peptides into their designs to mimic the natural protein ligands found in vivo. A few examples are the short peptide sequences RGD and PHSRN that mimic the primary and synergy-binding domains of the extracellular matrix protein, fibronectin, which is recognized by the cell surface receptor, alpha5beta 1 integrin. Even though scaffold modification with biomimetic peptides remains one of the most promising approaches for tissue engineering, the use of these peptides in therapeutic tissue-engineered products and drug delivery systems available on the commercial market is limited because the peptides are not easily able to mimic the natural protein. The design of a peptide that can effectively target the alpha5beta1 integrin would greatly increase biomimetic scaffold therapeutic potential. A novel peptide containing both the RGD primary binding domain and PHSRN synergy-binding domain for fibronectin joined with the appropriate linker should bind alpha 5beta1 integrin more efficiently and lead to greater cell adhesion over RGD alone. Several fibronectin mimetic peptides were designed and coupled to dialkyl hydrocarbon tails to make peptide-amphiphiles. The peptides contained different linkers connecting the two binding domains and different spacers separating the hydrophobic tails from the hydrophilic headgroups. The peptide-amphiphiles were deposited on mica substrates using the Langmuir-Blodgett technique. Langmuir isotherms indicated that the peptide-amphiphiles that contained higher numbers of serine residues formed a more tightly packed monolayer, but the increased number of serines also made transferring the amphiphiles to the mica substrate more difficult. Atomic force microscopy (AFM) images of the bilayers showed that the headgroups might be bent, forming small divots in the surface. These divots may help expose the PHSRN synergy-binding domain. Parallel studies undertaken by fellow group members showed that human

  12. Pile Driving

    NASA Technical Reports Server (NTRS)

    1987-01-01

    Machine-oriented structural engineering firm TERA, Inc. is engaged in a project to evaluate the reliability of offshore pile driving prediction methods to eventually predict the best pile driving technique for each new offshore oil platform. Phase I Pile driving records of 48 offshore platforms including such information as blow counts, soil composition and pertinent construction details were digitized. In Phase II, pile driving records were statistically compared with current methods of prediction. Result was development of modular software, the CRIPS80 Software Design Analyzer System, that companies can use to evaluate other prediction procedures or other data bases.

  13. A Small Chimeric Fibronectin Fragment Accelerates Dermal Wound Repair in Diabetic Mice

    PubMed Central

    Hocking, Denise C.; Brennan, James R.; Raeman, Carol H.

    2016-01-01

    Objective: During wound repair, soluble fibronectin is converted into biologically active, insoluble fibrils via a cell-mediated process. This fibrillar, extracellular matrix (ECM) form of fibronectin stimulates cell processes critical to tissue repair. Nonhealing wounds show reduced levels of ECM fibronectin fibrils. The objective of this study was to produce a small, recombinant wound supplement with the biological activity of insoluble fibronectin fibrils. Approach: A chimeric fibronectin fragment was produced by inserting the integrin-binding Arg-Gly-Asp (RGD) loop from the tenth type III repeat of fibronectin (FNIII10) into the analogous site within the heparin-binding, bioactive fragment of the first type III repeat (FNIII1H). FNIII1HRGD was tested for its ability to support cell functions necessary for wound healing, and then evaluated for its capacity to accelerate healing of full-thickness dermal wounds in diabetic mice. Results: In vitro, FNIII1HRGD supported cell adhesion, proliferation, and ECM fibronectin deposition. Application of FNIII1HRGD to dermal wounds of diabetic mice significantly enhanced wound closure compared with controls (73.9% ±4.1% vs. 58.1% ±4.7% closure on day 9, respectively), and significantly increased granulation tissue thickness (2.88 ± 0.75-fold increase over controls on day 14). Innovation: Recombinant proteins designed to functionally mimic the ECM form of fibronectin provide a novel therapeutic approach to circumvent diminished fibronectin fibril formation by delivering ECM fibronectin signals in a soluble form to chronic wounds. Conclusion: A small, chimeric fibronectin protein was developed. FNIII1HRGD demonstrated enhanced bioactivity in vitro and stimulated wound repair in a murine model of chronic wounds. PMID:27867754

  14. Fibronectin in Immune Responses in Organ Transplant Recipients

    PubMed Central

    De Sousa, Maria; Kupiec-Weglinski, Jerzy W.

    2000-01-01

    The immune response to an organ allograft involves perpetuation of T cell infiltration and activation. Advances in understanding the mechanisms of T cell activation have placed particular emphasis on the interactions between the T-cell receptor and antigen presenting cells, with little reference to the fact that in vivo activation occurs in the physical context of extracellular matrix proteins (ECM). Indeed, the possibility that ECM proteins may have a determining role in lymphocyte adhesion and tissue localization and function is now becoming more appreciated in view of growing evidence indicating that integrins and other T cell antigens bind ECM components, with some of these components exerting synergistic effects on T- cell activation. This review focuses on the importance of interactions between lymphocytes and fibronectin, a prominent ECM component, for cell migration and function in organ allograft recipients. It explores novel therapeutic approaches based on the assumption that fibronectin represents an active element in the process of T cell activation in the immune cascade triggered by organ transplantation. PMID:11097215

  15. Stretching Fibroblasts Remodels Fibronectin and Alters Cancer Cell Migration

    NASA Astrophysics Data System (ADS)

    Ao, Mingfang; Brewer, Bryson M.; Yang, Lijie; Franco Coronel, Omar E.; Hayward, Simon W.; Webb, Donna J.; Li, Deyu

    2015-02-01

    Most investigations of cancer-stroma interactions have focused on biochemical signaling effects, with much less attention being paid to biophysical factors. In this study, we investigated the role of mechanical stimuli on human prostatic fibroblasts using a microfluidic platform that was adapted for our experiments and further developed for both repeatable performance among multiple assays and for compatibility with high-resolution confocal microscopy. Results show that mechanical stretching of normal tissue-associated fibroblasts (NAFs) alters the structure of secreted fibronectin. Specifically, unstretched NAFs deposit and assemble fibronectin in a random, mesh-like arrangement, while stretched NAFs produce matrix with a more organized, linearly aligned structure. Moreover, the stretched NAFs exhibited an enhanced capability for directing co-cultured cancer cell migration in a persistent manner. Furthermore, we show that stretching NAFs triggers complex biochemical signaling events through the observation of increased expression of platelet derived growth factor receptor α (PDGFRα). A comparison of these behaviors with those of cancer-associated fibroblasts (CAFs) indicates that the observed phenotypes of stretched NAFs are similar to those associated with CAFs, suggesting that mechanical stress is a critical factor in NAF activation and CAF genesis.

  16. Fibronectin and stem cell differentiation – lessons from chondrogenesis

    PubMed Central

    Singh, Purva; Schwarzbauer, Jean E.

    2012-01-01

    Summary The extracellular matrix (ECM) is an intricate network of proteins that surrounds cells and has a central role in establishing an environment that is conducive to tissue-specific cell functions. In the case of stem cells, this environment is the stem cell niche, where ECM signals participate in cell fate decisions. In this Commentary, we describe how changes in ECM composition and mechanical properties can affect cell shape and stem cell differentiation. Using chondrogenic differentiation as a model, we examine the changes in the ECM that occur before and during mesenchymal stem cell differentiation. In particular, we focus on the main ECM protein fibronectin, its temporal expression pattern during chondrogenic differentiation, its potential effects on functions of differentiating chondrocytes, and how its interactions with other ECM components might affect cartilage development. Finally, we discuss data that support the possibility that the fibronectin matrix has an instructive role in directing cells through the condensation, proliferation and/or differentiation stages of cartilage formation. PMID:22976308

  17. Enhanced bacterial adhesion on surfaces pretreated with fibrinogen and fibronectin

    SciTech Connect

    Mohammad, S.F.; Topham, N.S.; Burns, G.L.; Olsen, D.B.

    1988-07-01

    The effect of certain plasma proteins on the adhesion of Pseudomonas aeruginosa and Staphylococcus epidermidis on polyurethane, polyvinylchloride, or glass was investigated. Test surfaces were treated with serum, plasma, albumin, immunoglobulin G, fibrinogen, or fibronectin. Using a specially designed test chamber, surfaces previously treated with test proteins were incubated with bacterial suspension. During the experiment, the test chamber was placed on a rotator to prevent settling of bacteria. At the end of the experiment, each test well was rinsed repeatedly to remove non-adherent bacteria. The number of bacteria adherent to the test surfaces was quantitated by a combination of methods including microscopic counting of cells, scintillation counting and autoradiography. It was noted that a greater number of bacteria adhered to surfaces coated with fibrinogen or fibronectin whereas surfaces treated with serum showed reduced bacterial adhesion. The inhibitory effect of serum appeared more pronounced with S. epidermidis when compared with P. aeruginosa under identical experimental conditions. Scanning electron microscopy revealed that adherent bacteria were randomly distributed on the test surfaces and appeared to replicate while still adherent. These observations suggested that bacterial adhesion to biomaterials can be significantly influenced by the composition of the adsorbed proteins at the interface.

  18. ED-B fibronectin expression is a marker of epithelial-mesenchymal transition in translational oncology.

    PubMed

    Petrini, Iacopo; Barachini, Serena; Carnicelli, Vittoria; Galimberti, Sara; Modeo, Letizia; Boni, Roberto; Sollini, Martina; Erba, Paola Anna

    2017-01-17

    Fibronectin is a component of the extracellular matrix that links collagen fibers to integrins on the cell's surface. The splicing isoforms, containing the ED-B domain, are not expressed in adult tissues but only in tumor stroma or during embryonic development. Fibroblasts and endothelial cells express ED-B fibronectin during angiogenesis. Also cancer cells can synthetize ED-B fibronectin, but its function in tumor growth needs to be further elucidated.We evaluated the expression of ED-B fibronectin in prostate cancer cell lines: PC3 and DU145. Using TGF-β, we induced epithelial to mesenchymal transition in culture and observed an increase of ED-B fibronectin expression. Thereafter, we evaluated the expression of ED-B fibronectin in multipotent mesangiogenic progenitor cells, and in mesenchymal stromal cells. The expression of ED-B fibronectin was much higher in mesenchymal than prostate cancer cells even after the epithelial to mesenchymal transition.Epithelial to mesenchymal transition is a key step for tumor progression contributing to the metastatic spread. Therefore, circulating cancer cells could seed into the metastatic niche taking advantage from the ED-B fibronectin that secrete their own.

  19. Augmentation of human monocyte opsonin-independent phagocytosis by fragments of human plasma fibronectin.

    PubMed

    Czop, J K; Kadish, J L; Austen, K F

    1981-06-01

    Human plasma fibronectin isolated by gelatin-affinity chromatography increases in a dose-dependent fashion the number of human monocytes that ingest particulate activators of the human alternative complement pathway in a fully synthetic medium. The fibronectin effect is selective for these particulate activators, does not extend to particles whose ingestion is dependent upon opsonization with IgG, and is not observed with pretreatment of the monocytes. Affinity chromatography with monoclonal antibody to plasma fibronectin of 440,000 daltons reveals that only 12-53% of the protein in a phagocytically active gelatin-affinity-purified fibronectin preparations is bound to the antibody. The protein eluted after affinity chromatography with monoclonal antibody of active preparations, which represented 10-43% of the protein applied, exhibits a 2- to 10-fold increment of activity per microgram of protein above the starting gelatin-affinity-purified material. Thus, the activity that augments the percent of human monocytes ingesting particulate activators of the alternative pathway is antigenically defined as plasma fibronectin. Preparations containing only intact 440,000-dalton fibronectin are also bound to and eluted from the monoclonal antibody, but they fail to augment phagocytosis. When inactive 440,000-dalton plasma fibronectin is subjected to limited trypsin cleavage, phagocytosis-enhancing activity develops that is bound to and elutes from the affinity column prepared with monoclonal antibody, thereby indicating that the enhancing activity of plasma fibronectin resides in cleavage fragments.

  20. Statistical Earthquake Focal Mechanism Forecasts

    NASA Astrophysics Data System (ADS)

    Kagan, Y. Y.; Jackson, D. D.

    2013-12-01

    The new whole Earth focal mechanism forecast, based on the GCMT catalog, has been created. In the present forecast, the sum of normalized seismic moment tensors within 1000 km radius is calculated and the P- and T-axes for the focal mechanism are evaluated on the basis of the sum. Simultaneously we calculate an average rotation angle between the forecasted mechanism and all the surrounding mechanisms. This average angle shows tectonic complexity of a region and indicates the accuracy of the prediction. The method was originally proposed by Kagan and Jackson (1994, JGR). Recent interest by CSEP and GEM has motivated some improvements, particularly to extend the previous forecast to polar and near-polar regions. The major problem in extending the forecast is the focal mechanism calculation on a spherical surface. In the previous forecast as our average focal mechanism was computed, it was assumed that longitude lines are approximately parallel within 1000 km radius. This is largely accurate in the equatorial and near-equatorial areas. However, when one approaches the 75 degree latitude, the longitude lines are no longer parallel: the bearing (azimuthal) difference at points separated by 1000 km reach about 35 degrees. In most situations a forecast point where we calculate an average focal mechanism is surrounded by earthquakes, so a bias should not be strong due to the difference effect cancellation. But if we move into polar regions, the bearing difference could approach 180 degrees. In a modified program focal mechanisms have been projected on a plane tangent to a sphere at a forecast point. New longitude axes which are parallel in the tangent plane are corrected for the bearing difference. A comparison with the old 75S-75N forecast shows that in equatorial regions the forecasted focal mechanisms are almost the same, and the difference in the forecasted focal mechanisms rotation angle is close to zero. However, though the forecasted focal mechanisms are similar

  1. Design of large aperture focal plane shutter

    NASA Astrophysics Data System (ADS)

    Hu, Jia-wen; Ma, Wen-li; Huang, Jin-long

    2012-09-01

    To satisfy the requirement of large telescope, a large aperture focal plane shutter with aperture size of φ200mm was researched and designed to realize, which could be started and stopped in a relative short time with precise position, and also the blades could open and close at the same time at any orientation. Timing-belts and stepper motors were adopted as the drive mechanism. Velocity and position of the stepper motors were controlled by the PWM pulse generated by DSP. Exponential curve is applied to control the velocity of the stepper motors to make the shutter start and stop in a short time. The closing/open time of shutter is 0.2s, which meets the performance requirements of large telescope properly.

  2. Different Organization of Type I Collagen Immobilized on Silanized and Nonsilanized Titanium Surfaces Affects Fibroblast Adhesion and Fibronectin Secretion.

    PubMed

    Marín-Pareja, Nathalia; Cantini, Marco; González-García, Cristina; Salvagni, Emiliano; Salmerón-Sánchez, Manuel; Ginebra, Maria-Pau

    2015-09-23

    Silanization has emerged in recent years as a way to obtain a stronger and more stable attachment of biomolecules to metallic substrates. However, its impact on protein conformation, a key aspect that influences cell response, has hardly been studied. In this work, we analyzed by atomic force microscopy (AFM) the distribution and conformation of type I collagen on plasma-treated surfaces before and after silanization. Subsequently, we investigated the effect of the different collagen conformations on fibroblasts adhesion and fibronectin secretion by immunofluorescence analyses. Two different organosilanes were used on plasma-treated titanium surfaces, either 3-chloropropyl-triethoxy-silane (CPTES) or 3-glycidyloxypropyl-triethoxy-silane (GPTES). The properties and amount of the adsorbed collagen were assessed by contact angle, X-ray photoelectron spectroscopy, optical waveguide lightmode spectroscopy, and AFM. AFM studies revealed different conformations of type I collagen depending on the silane employed. Collagen was organized in fibrillar networks over very hydrophilic (plasma treated titanium) or hydrophobic (silanized with CPTES) surfaces, the latter forming little globules with a beads-on-a-string appearance, whereas over surfaces presenting an intermediate hydrophobic character (silanized with GPTES), collagen was organized into clusters with a size increasing at higher protein concentration in solution. Cell response was strongly affected by collagen conformation, especially at low collagen density. The samples exhibiting collagen organized in globular clusters (GPTES-functionalized samples) favored a faster and better fibroblast adhesion as well as better cell spreading, focal adhesions formation, and more pronounced fibronectin fibrillogenesis. In contrast, when a certain protein concentration was reached at the material surface, the effect of collagen conformation was masked, and similar fibroblast response was observed in all samples.

  3. Fibronectin induces macrophage migration through a SFK-FAK/CSF-1R pathway.

    PubMed

    Digiacomo, Graziana; Tusa, Ignazia; Bacci, Marina; Cipolleschi, Maria Grazia; Dello Sbarba, Persio; Rovida, Elisabetta

    2017-07-04

    Integrins, following binding to proteins of the extracellular matrix (ECM) including collagen, laminin and fibronectin (FN), are able to transduce molecular signals inside the cells and to regulate several biological functions such as migration, proliferation and differentiation. Besides activation of adaptor molecules and kinases, integrins transactivate Receptor Tyrosine Kinases (RTK). In particular, adhesion to the ECM may promote RTK activation in the absence of growth factors. The Colony-Stimulating Factor-1 Receptor (CSF-1R) is a RTK that supports the survival, proliferation, and motility of monocytes/macrophages, which are essential components of innate immunity and cancer development. Macrophage interaction with FN is recognized as an important aspect of host defense and wound repair. The aim of the present study was to investigate on a possible cross-talk between FN-elicited signals and CSF-1R in macrophages. FN induced migration in BAC1.2F5 and J774 murine macrophage cell lines and in human primary macrophages. Adhesion to FN determined phosphorylation of the Focal Adhesion Kinase (FAK) and Src Family Kinases (SFK) and activation of the SFK/FAK complex, as witnessed by paxillin phosphorylation. SFK activity was necessary for FAK activation and macrophage migration. Moreover, FN-induced migration was dependent on FAK in either murine macrophage cell lines or human primary macrophages. FN also induced FAK-dependent/ligand-independent CSF-1R phosphorylation, as well as the interaction between CSF-1R and β1. CSF-1R activity was necessary for FN-induced macrophage migration. Indeed, genetic or pharmacological inhibition of CSF-1R prevented FN-induced macrophage migration. Our results identified a new SFK-FAK/CSF-1R signaling pathway that mediates FN-induced migration of macrophages.

  4. Relationship between fibronectin expression during gastrulation and heart formation in the rat embryo.

    PubMed

    Suzuki, H R; Solursh, M; Baldwin, H S

    1995-11-01

    By utilizing myosin immunostaining, we were able to identify early rat myocardium as a thin epithelial sheet and realized that its cohesive movement toward the midline leads to the straight heart tube formation. Localization study of fibronectin mRNA and protein was, therefore, carried out to investigate its tissue origin and possible roles in facilitating mesoderm migration and heart formation. Fibronectin mRNAs were first detected throughout the mesoderm during the early primitive streak stage, suggesting that the mesoderm is the source of fibronectin. By pre-head fold (pre-somite) and head fold (early somite) stages, the mesoderm became largely down-regulated for fibronectin mRNAs, while it was also at these stages when myosin-positive myocardium formed itself into the epithelium and was subsequently folding toward the midline. Thus, there appears to be little fibronectin synthesis during and directly relevant to early heart tube formation. Later, during the early straight heart tube stage (5 somite and older), endocardium became highly positive for fibronectin mRNAs, suggesting that the endocardium is the major source of fibronectin for the cardiac jelly. Based on the results, we present a map for the early mammalian heart in which the heart is a single crescentic band lying in front of the prechordal plate. We also suggest a process for heart tube formation based on the cohesive movement of the myocardial epithelium. During heart tube formation, fibronectin protein had been deposited previously by the mesoderm and was found uniformly in the ECM and not newly produced by any adjacent tissue. The data contradict the endodermal guidance of heart migration by fibronectin gradient and suggest, instead, a permissive role for the fibronectin substrate.

  5. Amino acid polymorphisms in the fibronectin-binding repeats of fibronectin-binding protein A affect bond strength and fibronectin conformation.

    PubMed

    Casillas-Ituarte, Nadia N; Cruz, Carlos H B; Lins, Roberto D; DiBartola, Alex C; Howard, Jessica; Liang, Xiaowen; Höök, Magnus; Viana, Isabelle F T; Sierra-Hernández, M Roxana; Lower, Steven K

    2017-04-11

    The Staphylococcus aureus cell surface contains cell wall-anchored proteins such as fibronectin-binding protein A (FnBPA) that bind to host ligands (e.g. fibronectin; Fn) present in the extracellular matrix of tissue or coatings on cardiac implants. Recent clinical studies have found a correlation between cardiovascular infections caused by S. aureus and nonsynonymous single nucleotide polymorphisms (SNPs) in FnBPA. Atomic force microscopy (AFM), surface plasmon resonance (SPR), and molecular simulations were used to investigate interactions between Fn and each of eight, 20-mer peptide variants containing amino acids A, H, I, K, N, and Q at positions equivalent to 782 and/or 786 in Fn-binding repeat-9 (FnBR-9) of FnBPA. Experimentally measured bond lifetimes (1/koff ) and dissociation constants (Kd = koff / kon ), determined by mechanically dissociating the Fn-peptide complex at loading rates relevant to the cardiovascular system varied from the lowest-affinity H782A+K786A peptide (0.011 sec, 747 µM) to the highest-affinity H782Q+K786N peptide (0.192 sec, 15.7 µM). These AFM results tracked remarkably well to metadynamics simulations in which peptide detachment was defined solely by the free-energy landscape. Simulations and SPR experiments suggested that an Fn conformational change may enhance the stability of the binding complex for peptides with K786I or H782Q+K786I (Kd(app) = 0.2 to 0.5 µM. as determined by SPR) compared with the lowest-affinity double alanine peptide (Kd(app) = 3.8 µM). Together, these findings demonstrate that amino acid substitutions in FnBR-9 can significantly affect bond strength and influence the conformation of Fn upon binding. They provide a mechanistic explanation for the observation of nonsynonymous SNPs in fnbA) among clinical isolates of S. aureus that cause endovascular infections.

  6. Cooperative Effects of Fibronectin Matrix Assembly and Initial Cell-Substrate Adhesion Strength in Cellular Self-Assembly

    PubMed Central

    Brennan, James R.; Hocking, Denise C.

    2015-01-01

    The cell-dependent polymerization of intercellular fibronectin fibrils can stimulate cells to self-assemble into multicellular structures. The local physical cues that support fibronectin-mediated cellular self-assembly are largely unknown. Here, fibronectin matrix analogs were used as synthetic adhesive substrates to model cell-matrix fibronectin fibrils having different integrin-binding specificity, affinity, and/or density. We utilized this model to quantitatively assess the relationship between adhesive forces derived from cell-substrate interactions and the ability of fibronectin fibril assembly to induce cellular self-assembly. Results indicate that the strength of initial, rather than mature, cell-substrate attachments correlates with the ability of substrates to support fibronectin-mediated cellular self-assembly. The cellular response to soluble fibronectin was bimodal and independent of the integrin-binding specificity of the substrate; increasing soluble fibronectin levels above a critical threshold increased aggregate cohesion on permissive substrates. Once aggregates formed, continuous fibronectin polymerization was necessary to maintain cohesion. During self-assembly, soluble fibronectin decreased cell-substrate adhesion strength and induced aggregate cohesion via a Rho-dependent mechanism, suggesting that the balance of contractile forces derived from fibronectin fibrils within cell-cell versus cell-substrate adhesions controls self-assembly and aggregate cohesion. Thus, initial cell-substrate attachment strength may provide a quantitative basis with which to build predictive models of fibronectin-mediated microtissue fabrication on a variety of substrates. PMID:26712598

  7. The Focal Surface of the JEM-EUSO Instrument

    SciTech Connect

    Kawasaki, Y.; Casolino, M.; Gorodetzky, P.; Santangelo, A.; Ricci, M.; Kajino, F.; Ebisuzaki, T.

    2011-09-22

    The Extreme Universe Space Observatory on JEM/EF (JEM-EUSO) is a space mission to study extremely high-energy cosmic rays. The JEM-EUSO instrument is a wide-angle refractive telescope in the near-ultraviolet wavelength region which will be mounted to the International Space Station. Its goal is to measure time-resolved fluorescence images of extensive air showers in the atmosphere. In this paper we describe in detail the main features and technological aspects of the focal surface of the instrument. The JEM-EUSO focal surface is a spherically curved surface, with an area of about 4.5m{sup 2}. The focal surface detector is made of more than 5,000 multi-anode photomultipliers (MAPMTs). Current baseline is Hamamatsu R11265-03-M64. The approach to the focal surface detector is highly modular. Photo-Detector-Modules (PDM) are the basic units that drive the mechanical structure and data acquisition. Each PDM consists of 9 Elementary Cells (ECs). The EC, which is the basic unit of the MAPMT support structure and of the front-end electronics, contains 4 units of MAPMTs. In total, about 1,200 ECs or about 150 PDMs are arranged on the whole of the focal surface of JEM-EUSO.

  8. Impaired Driving

    MedlinePlus

    ... people were killed in alcohol-impaired driving crashes, accounting for nearly one-third (31%) of all traffic- ... promotion efforts into practice that influence economic, organizational, policy, and school/community action. 13,14 Using community- ...

  9. Distracted Driving

    MedlinePlus

    ... other distractions. 3 At 55 mph, the average text takes your eyes off the road long enough ... risk behaviors among high school students, including sending texts while driving. 6,7 In 2013, more than ...

  10. Pile driving

    SciTech Connect

    Merjan, S.

    1988-02-16

    Process for producing in the ground a driven composite pile is described having (a) a lower pipe stem having an upper part having a top, the lower pipe stem being capable of withstanding pile driving blows applied to the top and (b) an upper corrugated shell stem incapable of withstanding pile driving blows, the corrugated shell stem having a lower end, which process comprises driving the lower pipe stem into the ground fitting to the top of the lower pipe stem a splicer. The splicer comprises a plate having a top face and a bottom face, an integral body portion depending from the plate and surrounding the upper part of the pipe stem and, welded to the top face of the plate, an upwardly extending corrugated shell stub up to about three feet long, screwing the lower end of the upper corrugated shell stem to the shell stub after driving the lower pipe stem into the ground, placing a non-expanding pipe mandrel into the shell stem with the bottom of the mandrel resting on the plate, striking pile-driving blows on the top of the mandrel to drive the composite pile into the ground, and filling the shell stem and pipe stem with concrete from above.

  11. NEDD9 stabilizes focal adhesions, increases binding to the extra-cellular matrix and differentially effects 2D versus 3D cell migration.

    PubMed

    Zhong, Jessie; Baquiran, Jaime B; Bonakdar, Navid; Lees, Justin; Ching, Yu Wooi; Pugacheva, Elena; Fabry, Ben; O'Neill, Geraldine M

    2012-01-01

    The speed of cell migration on 2-dimensional (2D) surfaces is determined by the rate of assembly and disassembly of clustered integrin receptors known as focal adhesions. Different modes of cell migration that have been described in 3D environments are distinguished by their dependence on integrin-mediated interactions with the extra-cellular matrix. In particular, the mesenchymal invasion mode is the most dependent on focal adhesion dynamics. The focal adhesion protein NEDD9 is a key signalling intermediary in mesenchymal cell migration, however whether NEDD9 plays a role in regulating focal adhesion dynamics has not previously been reported. As NEDD9 effects on 2D migration speed appear to depend on the cell type examined, in the present study we have used mouse embryo fibroblasts (MEFs) from mice in which the NEDD9 gene has been depleted (NEDD9 -/- MEFs). This allows comparison with effects of other focal adhesion proteins that have previously been demonstrated using MEFs. We show that focal adhesion disassembly rates are increased in the absence of NEDD9 expression and this is correlated with increased paxillin phosphorylation at focal adhesions. NEDD9-/- MEFs have increased rates of migration on 2D surfaces, but conversely, migration of these cells is significantly reduced in 3D collagen gels. Importantly we show that myosin light chain kinase is activated in 3D in the absence of NEDD9 and is conversely inhibited in 2D cultures. Measurement of adhesion strength reveals that NEDD9-/- MEFs have decreased adhesion to fibronectin, despite upregulated α5β1 fibronectin receptor expression. We find that β1 integrin activation is significantly suppressed in the NEDD9-/-, suggesting that in the absence of NEDD9 there is decreased integrin receptor activation. Collectively our data suggest that NEDD9 may promote 3D cell migration by slowing focal adhesion disassembly, promoting integrin receptor activation and increasing adhesion force to the ECM.

  12. Precise Measurement of Effective Focal Length

    NASA Technical Reports Server (NTRS)

    Wise, T. D.; Young, J. B.

    1983-01-01

    Computerized instrument measures effective focal lengths to 0.01 percent accuracy. Laser interferometers measure mirror angle and stage coordinate y in instrument for accurate measurment of focal properties of optical systems. Operates under computer control to measure effective focal length, focal surface shape, modulation transfer function, and astigmatism.

  13. Nanotopographical manipulation of focal adhesion formation for enhanced differentiation of human neural stem cells.

    PubMed

    Yang, Kisuk; Jung, Kyuhwan; Ko, Eunkyung; Kim, Jin; Park, Kook In; Kim, Jinseok; Cho, Seung-Woo

    2013-11-13

    Manipulating neural stem cell (NSC) fate is of great importance for improving the therapeutic efficacy of NSCs to treat neurodegenerative disorders. Biophysical cues, in addition to biochemical factors, regulate NSC phenotype and function. In this study, we assessed the extent to which surface nanotopography of culture substrates modulates human NSC (hNSC) differentiation. Fibronectin-coated polymer substrates with diverse nanoscale shapes (groove and pillar) and dimensions (ranging from 300 to 1500 nm groove width and pillar gap) were used to investigate the effects of topographical cues on hNSC morphology, alignment, focal adhesion, and differentiation. The majority of nanopatterned substrates induced substantial changes in cellular morphology and alignment along the patterned shapes, leading to alterations in focal adhesion and F-actin reorganization. Certain types of nanopatterned substrates, in particular the ones with small nanostructures (e.g., 300-300 nm groove ridges and 300-300 nm pillar diameter gaps), were found to effectively enhance focal adhesion complex development. Consequently, these substrates enhanced hNSC differentiation toward neurons and astrocytes. Nanotopographical-induced formation of focal adhesions in hNSCs activates integrin-mediated mechanotransduction and intracellular signaling pathways such as MEK-ERK, which may ultimately promote gene expression related to NSC differentiation. This strategy of manipulating matrix surface topography could be applied to develop culture substrates and tissue engineered scaffolds that improve the efficacy of NSC therapeutics.

  14. Fibronectin tetrapeptide is target for syphilis spirochete cytadherence

    SciTech Connect

    Thomas, D.D.; Baseman, J.B.; Alderete, J.F.

    1985-11-01

    The syphilis bacterium, Treponema pallidum, parasitizes host cells through recognition of fibronectin (Fn) on cell surfaces. The active site of the Fn molecule has been identified as a four-amino acid sequence, arg-gly-asp-ser (RGDS), located on each monomer of the cell-binding domain. The synthetic heptapeptide gly-arg-gly-asp-ser-pro-cys (GRGDSPC), with the active site sequence RGDS, specifically competed with SVI-labeled cell-binding domain acquisition by T. pallidum. Additionally, the same heptapeptide with the RGDS sequence diminished treponemal attachment to HEp-2 and HT1080 cell monolayers. Related heptapeptides altered in one key amino acid within the RGDS sequence failed to inhibit Fn cell-binding domain acquisition or parasitism of host cells by T. pallidum. The data support the view that T. pallidum cytadherence of host cells is through recognition of the RGDS sequence also important for eukaryotic cell-Fn binding.

  15. Structural Insights into Fibronectin Type III Domain Mediated Signaling

    PubMed Central

    Bencharit, Sompop; Cui, Cai Bin; Siddiqui, Adnan; Howard-Williams, Escher L.; Sondek, John; Zuobi-Hasona, Kheir; Aukhil, Ikramuddin

    2007-01-01

    The alternatively spliced type-III extradomain B (EIIIB) of Fibronectin (FN) is only expressed during embryogenesis, wound healing and tumorigenesis. The biological function of this domain remains unclear. We describe here the first crystal structure of the interface between alternatively-spliced domain EIIIB and its adjacent FN type-III domain 8 (FN B-8). The opened CC′ loop of EIIIB and the rotation and tilt of EIIIB domain allows good access to the FG loop of FN-8 which is normally hindered by the CC′ loop of FN-7. In addition, the AGEGIP sequence of the CC′ loop of EIIIB replaces the NGQQGN sequence of the CC′ loop of FN-7. Finally, the CC” loop of EIIIB forms an acidic groove with FN-8. These structural findings warrant future studies directed at identifying potential binding partners for FN B-8 interface, linking EIIIB to skeletal and cartilagenous development, wound healing, and tumorigenesis, respectively. PMID:17261313

  16. A fibronectin receptor on Candida albicans mediates adherence of the fungus to extracellular matrix

    SciTech Connect

    Klotz, S.A.; Smith, R.L. )

    1991-03-01

    Binding of fibronectin, an extracellular matrix (ECM) protein, to Candida albicans was measured, and adherence of the fungus to immobilized ECM proteins, fibronectin, laminin, types I and IV collagen, and subendothelial ECM was studied. 125I-labeled fibronectin was inhibited from binding to the fungus by unlabeled human plasma fibronectin and by Arg-Gly-Asp (RGD), Gly-Arg-Gly-Glu-Ser-Pro (GRGESP), and Gly-Arg-Gly-Asp-Thr-Pro (GRGDTP), but binding was not inhibited by Gly-Arg-Gly-Asp-Ser-Pro. Soluble fibronectin, RGD, GRGESP, and GRGDTP also inhibited fungal adherence to the individual immobilized ECM proteins in a complex pattern, but only soluble fibronectin (10(-7) M) inhibited fungal adherence to subendothelial ECM. Thus, C. albicans possesses at least one type of cell surface receptor for binding soluble fibronectin that can be inhibited with peptides. This receptor apparently is used to bind the fungus to immobilized ECM proteins and to subendothelial ECM and may play a role in the initiation of disseminated disease by bloodborne fungi by providing for adherence of the microorganisms to ECM proteins.

  17. Identification and characterization of a fibronectin-binding protein from Granulicatella adiacens.

    PubMed

    Yamaguchi, T; Soutome, S; Oho, T

    2011-12-01

    The interaction of microorganisms with fibronectin plays an important role in infective endocarditis. Granulicatella adiacens is a member of the oral microbiota, formerly known as nutritionally variant streptococci, and is often isolated from endocarditis patients. In the present study we identified a surface protein, designated Cha, which binds to fibronectin, by a plaque hybridization procedure using the cshA sequence as probe, which encodes a fibronectin-binding molecule of Streptococcus gordonii DL1. The cha sequence was highly homologous to cshA and encoded a product of 2351 amino acid residues. The protein comprised a unique sequence in the N-terminal half region. The C-terminal region contained nine complete, and one incomplete, 115-amino acid residue repeat blocks. Among eight strains of nutritionally variant streptococci, three G. adiacens strains and one Abiotrophia defectiva strain carried the cha gene. Heterologous expression studies suggested that Cha adhered to immobilized fibronectin, and that this function was located in the unique region. Recombinant Cha protein also adhered to immobilize fibronectin and partially inhibited adherence of G. adiacens to fibronectin in a dose-dependent manner. These results suggest that Cha is a cell surface protein that mediates adherence of G. adiacens to fibronectin.

  18. Role of Fibronectin in the Adhesion of Acinetobacter baumannii to Host Cells

    PubMed Central

    Smani, Younes; McConnell, Michael J.; Pachón, Jerónimo

    2012-01-01

    Adhesion to host cells is an initial and important step in Acinetobacter baumannii pathogenesis. However, there is relatively little information on the mechanisms by which A. baumannii binds to and interacts with host cells. Adherence to extracellular matrix proteins, such as fibronectin, affords pathogens with a mechanism to invade epithelial cells. Here, we found that A. baumannii adheres more avidly to immobilized fibronectin than to control protein. Free fibronectin used as a competitor resulted in dose-dependent decreased binding of A. baumannii to fibronectin. Three outer membrane preparations (OMPs) were identified as fibronectin binding proteins (FBPs): OMPA, TonB-dependent copper receptor, and 34 kDa OMP. Moreover, we demonstrated that fibronectin inhibition and neutralization by specific antibody prevented significantly the adhesion of A. baumannii to human lung epithelial cells (A549 cells). Similarly, A. baumannii OMPA neutralization by specific antibody decreased significantly the adhesion of A. baumannii to A549 cells. These data indicate that FBPs are key adhesins that mediate binding of A. baumannii to human lung epithelial cells through interaction with fibronectin on the surface of these host cells. PMID:22514602

  19. Characterization of a fibronectin-binding protein from Lactobacillus casei BL23.

    PubMed

    Muñoz-Provencio, D; Pérez-Martínez, G; Monedero, V

    2010-03-01

    To characterize the functionality of the Lactobacillus casei BL23 fbpA gene encoding a putative fibronectin-binding protein. Adhesion tests showed that L. casei BL23 binds immobilized and soluble fibronectin in a protease-sensitive manner. A mutant with inactivated fbpA showed a decrease in binding to immobilized fibronectin and a strong reduction in the surface hydrophobicity as reflected by microbial adhesion to solvents test. However, minor effects were seen on adhesion to the human Caco-2 or HT-29 cell lines. Purified 6X(His)FbpA bound to immobilized fibronectin in a dose-dependent manner. Western blot experiments with FbpA-specific antibodies showed that FbpA could be extracted from the cell surface by LiCl treatment and that protease digestion of the cells reduced the amount of extracted FbpA. Furthermore, surface exposition of FbpA was detected in other L. casei strains by LiCl extraction and whole-cell ELISA. FbpA can be found at the L. casei BL23 surface and participates in cell attachment to immobilized fibronectin. We showed that FbpA is an important, but not the only, factor contributing to fibronectin binding in BL23 strain. This is the first report showing the involvement of FbpA in fibronectin binding in L. casei BL23 and represents a new contribution to the study of attachment factors in probiotic bacteria.

  20. External fibronectin of cultured human fibroblasts is predominantly a matrix protein

    PubMed Central

    1978-01-01

    The distribution of a major glycoprotein (fibronectin) of human fibroblast cultures was studied in immunoelectron microscopy with peroxidase- or ferritin-labeled antibodies. External fibronectin was visualized in pericellular structures, in some areas on the growth substratum, and to a lesser degree in close association with the upper and lower surface membranes of the cell. The pericellular fibronectin- containing structures consisted of amorphous or vaguely fibrillar material forming strands or patches, 50-500 nm in diameter; the structures appeared to mediate distant cell-to-cell and cell-to- substrate contacts. When in close association with the plasma membrane, fibronectin markers were seen as discrete patches. The exact relationship between this form of fibronectin and the plasma membrane, however, remained open. Filamentous material was commonly seen in the cortical cytoplasm under patches of membrane-associated fibronectin. The distribution that we observed is consistent with the proposed roles of fibronectin in cell interactions with neighboring structures and with its presence in vivo as an extracellular glycoprotein in connective tissue matrix and basal laminae. PMID:344328

  1. Fibronectin-Containing Extracellular Vesicles Protect Melanocytes against Ultraviolet Radiation-Induced Cytotoxicity.

    PubMed

    Bin, Bum-Ho; Kim, Dae-Kyum; Kim, Nan-Hyung; Choi, Eun-Jeong; Bhin, Jinhyuk; Kim, Sung Tae; Gho, Yong Song; Lee, Ai-Young; Lee, Tae Ryong; Cho, Eun-Gyung

    2016-05-01

    Skin melanocytes are activated by exposure to UV radiation to secrete melanin-containing melanosomes to protect the skin from UV-induced damage. Despite the continuous renewal of the epidermis, the turnover rate of melanocytes is very slow, and they survive for long periods. However, the mechanisms underlying the survival of melanocytes exposed to UV radiation are not known. Here, we investigated the role of melanocyte-derived extracellular vesicles in melanocyte survival. Network analysis of the melanocyte extracellular vesicle proteome identified the extracellular matrix component fibronectin at a central node, and the release of fibronectin-containing extracellular vesicles was increased after exposure of melanocytes to UVB radiation. Using an anti-fibronectin neutralizing antibody and specific inhibitors of extracellular vesicle secretion, we demonstrated that extracellular vesicles enriched in fibronectin were involved in melanocyte survival after UVB radiation. Furthermore, we observed that in the hyperpigmented lesions of patients with melasma, the extracellular space around melanocytes contained more fibronectin compared with normal skin, suggesting that fibronectin is involved in maintaining melanocytes in pathological conditions. Collectively, our findings suggest that melanocytes secrete fibronectin-containing extracellular vesicles to increase their survival after UVB radiation. These data provide important insight into how constantly stimulated melanocytes can be maintained in pathological conditions such as melasma. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  2. Genetic models of focal epilepsies.

    PubMed

    Boillot, Morgane; Baulac, Stéphanie

    2016-02-15

    Focal epilepsies were for a long time thought to be acquired disorders secondary to cerebral lesions. However, the important role of genetic factors in focal epilepsies is now well established. Several focal epilepsy syndromes are now proven to be monogenic disorders. While earlier genetic studies suggested a strong contribution of ion channel and neurotransmitter receptor genes, later work has revealed alternative pathways, among which the mammalian target of rapamycin (mTOR) signal transduction pathway with DEPDC5. In this article, we provide an update on the mutational spectrum of neuronal nicotinic acetylcholine receptor genes (CHRNA4, CHRNB2, CHRNA2) and KCNT1 causing autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE), and of LGI1 in autosomal dominant epilepsy with auditory features (ADEAF). We also emphasize, through a review of the current literature, the contribution of in vitro and in vivo models developed to unveil the pathogenic mechanisms underlying these two epileptic syndromes.

  3. Continuously variable focal length lens

    DOEpatents

    Adams, Bernhard W; Chollet, Matthieu C

    2013-12-17

    A material preferably in crystal form having a low atomic number such as beryllium (Z=4) provides for the focusing of x-rays in a continuously variable manner. The material is provided with plural spaced curvilinear, optically matched slots and/or recesses through which an x-ray beam is directed. The focal length of the material may be decreased or increased by increasing or decreasing, respectively, the number of slots (or recesses) through which the x-ray beam is directed, while fine tuning of the focal length is accomplished by rotation of the material so as to change the path length of the x-ray beam through the aligned cylindrical slows. X-ray analysis of a fixed point in a solid material may be performed by scanning the energy of the x-ray beam while rotating the material to maintain the beam's focal point at a fixed point in the specimen undergoing analysis.

  4. Fibronectin Matrix Mimetics Promote Full-Thickness Wound Repair in Diabetic Mice

    PubMed Central

    Roy, Daniel C.; Mooney, Nancie A.; Raeman, Carol H.; Dalecki, Diane

    2013-01-01

    During tissue repair, fibronectin is converted from a soluble, inactive form into biologically active extracellular matrix (ECM) fibrils through a cell-dependent process. ECM fibronectin promotes numerous cell processes that are critical to tissue repair and regulates the assembly of other proteins into the matrix. Nonhealing wounds show reduced levels of ECM fibronectin. To functionally mimic ECM fibronectin, a series of fibronectin matrix mimetics was developed by directly coupling the matricryptic, heparin-binding fragment of the first type III repeat of fibronectin (FNIII1H) to various sequences from the integrin-binding domain (FNIII8–10). The recombinant proteins were produced as glutathione-S-transferase (GST)-tagged fusion proteins for ease of production and purification. Full-thickness, excisional wounds were produced in genetically diabetic mice, and fibronectin matrix mimetics were applied directly to the wounds. A significant enhancement of wound closure was observed by day 9 in response to GST/III1H,8–10 versus GST-treated controls (73.9%±4.1% vs. 58.1%±4.7% closure, respectively). Two weeks after injury, fibronectin matrix mimetic-treated wounds had developed a multi-layered epithelium that completely covered the wound space. Furthermore, significant increases in granulation tissue thickness were observed in response to treatment with GST/III1H,8–10 (4.05±0.93-fold), GST/III1H,8,10 (2.91±0.49-fold), or GST/III1H,8RGD (3.55±0.59-fold) compared with GST controls, and was accompanied by dense collagen deposition, the presence of myofibroblasts, and functional vasculature. Thus, the recombinant fibronectin matrix analogs normalized the impairment in repair observed in this chronic wound model and may provide a new approach to accelerate the healing of diabetic wounds. PMID:23808793

  5. Fibronectin matrix polymerization regulates smooth muscle cell phenotype through a Rac1 dependent mechanism.

    PubMed

    Shi, Feng; Long, Xiaochun; Hendershot, Allison; Miano, Joseph M; Sottile, Jane

    2014-01-01

    Smooth muscle cells are maintained in a differentiated state in the vessel wall, but can be modulated to a synthetic phenotype following injury. Smooth muscle phenotypic modulation is thought to play an important role in the pathology of vascular occlusive diseases. Phenotypically modulated smooth muscle cells exhibit increased proliferative and migratory properties that accompany the downregulation of smooth muscle cell marker proteins. Extracellular matrix proteins, including fibronectin, can regulate the smooth muscle phenotype when used as adhesive substrates. However, cells produce and organize a 3-dimensional fibrillar extracellular matrix, which can affect cell behavior in distinct ways from the protomeric 2-dimensional matrix proteins that are used as adhesive substrates. We previously showed that the deposition/polymerization of fibronectin into the extracellular matrix can regulate the deposition and organization of other extracellular matrix molecules in vitro. Further, our published data show that the presence of a fibronectin polymerization inhibitor results in increased expression of smooth muscle cell differentiation proteins and inhibits vascular remodeling in vivo. In this manuscript, we used an in vitro cell culture system to determine the mechanism by which fibronectin polymerization affects smooth muscle phenotypic modulation. Our data show that fibronectin polymerization decreases the mRNA levels of multiple smooth muscle differentiation genes, and downregulates the levels of smooth muscle α-actin and calponin proteins by a Rac1-dependent mechanism. The expression of smooth muscle genes is transcriptionally regulated by fibronectin polymerization, as evidenced by the increased activity of luciferase reporter constructs in the presence of a fibronectin polymerization inhibitor. Fibronectin polymerization also promotes smooth muscle cell growth, and decreases the levels of actin stress fibers. These data define a Rac1-dependent pathway wherein

  6. Fibronectin on the Surface of Myeloma Cell-derived Exosomes Mediates Exosome-Cell Interactions.

    PubMed

    Purushothaman, Anurag; Bandari, Shyam Kumar; Liu, Jian; Mobley, James A; Brown, Elizabeth E; Sanderson, Ralph D

    2016-01-22

    Exosomes regulate cell behavior by binding to and delivering their cargo to target cells; however, the mechanisms mediating exosome-cell interactions are poorly understood. Heparan sulfates on target cell surfaces can act as receptors for exosome uptake, but the ligand for heparan sulfate on exosomes has not been identified. Using exosomes isolated from myeloma cell lines and from myeloma patients, we identify exosomal fibronectin as a key heparan sulfate-binding ligand and mediator of exosome-cell interactions. We discovered that heparan sulfate plays a dual role in exosome-cell interaction; heparan sulfate on exosomes captures fibronectin, and on target cells it acts as a receptor for fibronectin. Removal of heparan sulfate from the exosome surface releases fibronectin and dramatically inhibits exosome-target cell interaction. Antibody specific for the Hep-II heparin-binding domain of fibronectin blocks exosome interaction with tumor cells or with marrow stromal cells. Regarding exosome function, fibronectin-mediated binding of exosomes to myeloma cells activated p38 and pERK signaling and expression of downstream target genes DKK1 and MMP-9, two molecules that promote myeloma progression. Antibody against fibronectin inhibited the ability of myeloma-derived exosomes to stimulate endothelial cell invasion. Heparin or heparin mimetics including Roneparstat, a modified heparin in phase I trials in myeloma patients, significantly inhibited exosome-cell interactions. These studies provide the first evidence that fibronectin binding to heparan sulfate mediates exosome-cell interactions, revealing a fundamental mechanism important for exosome-mediated cross-talk within tumor microenvironments. Moreover, these results imply that therapeutic disruption of fibronectin-heparan sulfate interactions will negatively impact myeloma tumor growth and progression.

  7. Fibronectin on the Surface of Myeloma Cell-derived Exosomes Mediates Exosome-Cell Interactions*

    PubMed Central

    Purushothaman, Anurag; Bandari, Shyam Kumar; Liu, Jian; Mobley, James A.; Brown, Elizabeth E.; Sanderson, Ralph D.

    2016-01-01

    Exosomes regulate cell behavior by binding to and delivering their cargo to target cells; however, the mechanisms mediating exosome-cell interactions are poorly understood. Heparan sulfates on target cell surfaces can act as receptors for exosome uptake, but the ligand for heparan sulfate on exosomes has not been identified. Using exosomes isolated from myeloma cell lines and from myeloma patients, we identify exosomal fibronectin as a key heparan sulfate-binding ligand and mediator of exosome-cell interactions. We discovered that heparan sulfate plays a dual role in exosome-cell interaction; heparan sulfate on exosomes captures fibronectin, and on target cells it acts as a receptor for fibronectin. Removal of heparan sulfate from the exosome surface releases fibronectin and dramatically inhibits exosome-target cell interaction. Antibody specific for the Hep-II heparin-binding domain of fibronectin blocks exosome interaction with tumor cells or with marrow stromal cells. Regarding exosome function, fibronectin-mediated binding of exosomes to myeloma cells activated p38 and pERK signaling and expression of downstream target genes DKK1 and MMP-9, two molecules that promote myeloma progression. Antibody against fibronectin inhibited the ability of myeloma-derived exosomes to stimulate endothelial cell invasion. Heparin or heparin mimetics including Roneparstat, a modified heparin in phase I trials in myeloma patients, significantly inhibited exosome-cell interactions. These studies provide the first evidence that fibronectin binding to heparan sulfate mediates exosome-cell interactions, revealing a fundamental mechanism important for exosome-mediated cross-talk within tumor microenvironments. Moreover, these results imply that therapeutic disruption of fibronectin-heparan sulfate interactions will negatively impact myeloma tumor growth and progression. PMID:26601950

  8. The Non-Equilibrium Thermodynamics and Kinetics of Focal Adhesion Dynamics

    PubMed Central

    Olberding, Joseph E.; Thouless, Michael D.; Arruda, Ellen M.; Garikipati, Krishna

    2010-01-01

    Background We consider a focal adhesion to be made up of molecular complexes, each consisting of a ligand, an integrin molecule, and associated plaque proteins. Free energy changes drive the binding and unbinding of these complexes and thereby controls the focal adhesion's dynamic modes of growth, treadmilling and resorption. Principal Findings We have identified a competition among four thermodynamic driving forces for focal adhesion dynamics: (i) the work done during the addition of a single molecular complex of a certain size, (ii) the chemical free energy change associated with the addition of a molecular complex, (iii) the elastic free energy change associated with deformation of focal adhesions and the cell membrane, and (iv) the work done on a molecular conformational change. We have developed a theoretical treatment of focal adhesion dynamics as a nonlinear rate process governed by a classical kinetic model. We also express the rates as being driven by out-of-equilibrium thermodynamic driving forces, and modulated by kinetics. The mechanisms governed by the above four effects allow focal adhesions to exhibit a rich variety of behavior without the need to introduce special constitutive assumptions for their response. For the reaction-limited case growth, treadmilling and resorption are all predicted by a very simple chemo-mechanical model. Treadmilling requires symmetry breaking between the ends of the focal adhesion, and is achieved by driving force (i) above. In contrast, depending on its numerical value (ii) causes symmetric growth, resorption or is neutral, (iii) causes symmetric resorption, and (iv) causes symmetric growth. These findings hold for a range of conditions: temporally-constant force or stress, and for spatially-uniform and non-uniform stress distribution over the FA. The symmetric growth mode dominates for temporally-constant stress, with a reduced treadmilling regime. Significance In addition to explaining focal adhesion dynamics, this

  9. Focal epithelial hyperplasia: Case report.

    PubMed

    Puriene, Alina; Rimkevicius, Arunas; Gaigalas, Mindaugas

    2011-01-01

    The purpose of the present article is to present a 15 year-old patient with focal epithelial hyperplasia and to review the references on the subject-related etiological, pathological, diagnostic and treatment aspects. Focal epithelial hyperplasia is a rare human papilloma virus (HPV) related to oral lesion with very low frequency within our population. Surgical treatment with a biopsy was performed, acanthosis and parakeratosis are consistent histopathological features, since the patient had no history of sexual contact and HIV infection, the virus was probably acquired from environmental sources.

  10. Localization of fibronectin within the renal glomerulus and its production by cultured glomerular cells.

    PubMed Central

    Oberley, T. D.; Mosher, D. F.; Mills, M. D.

    1979-01-01

    Fibronectin was shown in the kidney glomerulus by the use of the peroxidase-labeled antibody technique. At the ultrastructural level, fibronectin was present in greatest quantities along those cell surfaces that abut the capillary basement membrane, especially along the capillary epithelial cell foot process. Intracellular staining was also seen in the glomerulus, most notably in the mesangial cell. Fibronectin was present extracellularly in large amounts in glomerular cell culture and was also demonstrated on the cell membrane and intracellularly. Images Figure 5 Figure 2 Figure 3 Figure 4 Figure 1 PMID:382867

  11. Effect of sterilization methods on cell binding activity of surface-immobilized fibronectin.

    PubMed

    Xing, Hai Bo; Pan, Qin; Wang, Kai Feng; Lou, Fang; Li, Da

    2014-05-01

    The effect of sterilization methods on biological activity of fibronectin on the surface of biomaterials was elaborated in the present study. Sterile protein- modified biomaterials were fabricated by microfilter filtration and UV irradiation, respectively. UV irradiation altered the conformation of surface- adsorbed fibronectin and further affected the attachment, morphology and biological function of endothelial cells. However, microfilter filtration did not to change the normal conformation of fibronectin, or the proliferation and biological function of endothelial cells, indicating that microfilter filtration sterilization is the most suitable method for protein-substrate. Copyright © 2014 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

  12. Fibronectin is a hypoxia-independent target of the tumor suppressor VHL.

    PubMed

    Bluyssen, Hans A R; Lolkema, Martijn P J K; van Beest, Moniek; Boone, Michelle; Snijckers, Cristel M J T; Los, Maartje; Gebbink, Martijn F B G; Braam, Branko; Holstege, Frank C P; Giles, Rachel H; Voest, Emile E

    2004-01-02

    The von Hippel-Lindau (VHL) tumor suppressor gene regulates the extracellular matrix by controlling fibronectin deposition. To identify novel VHL target genes, we subjected mRNA from VHL-deficient RCC cells (786-0-pRC) and a transfectant re-expressing wildtype VHL (786-0-VHL) to differential expression profiling. Among the differentially expressed genes, we detected that fibronectin is upregulated in the presence of VHL, while it is not affected by hypoxia. Thus regulation of fibronectin deposition by VHL occurs at the transcriptional level, irrespective of oxygen levels.

  13. The distribution of tissue fibronectin and sialic acid in human breast cancer.

    PubMed

    Süer, S; Baloğlu, H; Güngör, Z; Sönmez, H; Kökoğlu, E

    1998-06-01

    Our findings indicate that sialic acid and fibronectin levels in breast tumors are higher than those in normal tissues. The mean tissue fibronectin and sialic acid concentrations for patients with breast cancer were 30.90 +/- 9.68 microg/mg protein and 21.60 +/- 9.35 microg/mg protein, respectively, and for normal controls were 12.47 +/- 5.69 microg/mg protein, respectively. Tissue fibronectin and sialic acid can be important markers for human breast cancer.

  14. Clinical studies on plasma fibronectin and factor XIII; with special reference to hyperlipoproteinemia.

    PubMed

    Cucuianu, M; Rus, H G; Cristea, A; Niculescu, F; Bedeleanu, D; Poruţiu, D; Roman, S

    1985-04-30

    When compared to age-matched normal weight normolipidemic control subjects, plasma factor XIII, plasma fibronectin and serum cholinesterase levels were found to be markedly decreased in patients with decompensated cirrhosis of the liver, not significantly changed in hyperlipoproteinemia type IIa (heterozygous subjects) and increased in hypertriglyceridemic subjects (type IIb and IV) as well as in hyperlipidemic nephrotic patients. A possible accelerated hepatic synthesis of certain plasma proteins including factor XIII and fibronectin in patients with the nephrotic syndrome as well as in endogenous hypertriglyceridemia is envisaged. It is also considered that mural thrombi, richer in factor XIII and fibronectin, would be more resistant to fibrinolysis and more readily attached to subendothelial structures.

  15. Covalent grafting of fibronectin onto plasma-treated PTFE: influence of the conjugation strategy on fibronectin biological activity.

    PubMed

    Vallières, Karine; Petitclerc, Eric; Laroche, Gaétan

    2007-05-10

    Surface coating of synthetic materials is often considered to improve biomedical devices biocompatibility. In this study, we covalently bound fibronectin (FN) onto ammonia plasma-treated PTFE via two crosslinkers, namely glutaric anhydride (GA) and sulfosuccinimidyl-4-(p-maleimidophenyl)butyrate (sulfo-SMPB). With respect to clean PTFE, cell adhesion increased markedly on both FN grafted surfaces, although it was twice higher on PTFE-GA-FN than on PTFE-SMPB-FN. ELISA experiments performed with a polyclonal antibody revealed that the amount of FN is identical on both surfaces while monoclonal antibody specific to the RGD binding site clearly demonstrated a greater availability when FN is surface grafted through GA. These results provide evidence of a variation in protein conformation correlated with the surface conjugation strategy.

  16. Dissociation Rate Constants of Human Fibronectin Binding to Fibronectin-binding Proteins on Living Staphylococcus aureus Isolated from Clinical Patients*

    PubMed Central

    Casillas-Ituarte, Nadia N.; Lower, Brian H.; Lamlertthon, Supaporn; Fowler, Vance G.; Lower, Steven K.

    2012-01-01

    Staphylococcus aureus is part of the indigenous microbiota of humans. Sometimes, S. aureus bacteria enter the bloodstream, where they form infections on implanted cardiovascular devices. A critical, first step in such infections is a bond that forms between fibronectin-binding protein (FnBP) on S. aureus and host proteins, such as fibronectin (Fn), that coat the surface of implants in vivo. In this study, native FnBPs on living S. aureus were shown to form a mechanically strong conformational structure with Fn by atomic force microscopy. The tensile acuity of this bond was probed for 46 bloodstream isolates, each from a patient with a cardiovascular implant. By analyzing the force spectra with the worm-like chain model, we determined that the binding events were consistent with a multivalent, cluster bond consisting of ∼10 or ∼80 proteins in parallel. The dissociation rate constant (koff, s−1) of each multibond complex was determined by measuring strength as a function of the loading rate, normalized by the number of bonds. The bond lifetime (1/koff) was two times longer for bloodstream isolates from patients with an infected device (1.79 or 69.47 s for the 10- or 80-bond clusters, respectively; n = 26 isolates) relative to those from patients with an uninfected device (0.96 or 34.02 s; n = 20 isolates). This distinction could not be explained by different amounts of FnBP, as confirmed by Western blots. Rather, amino acid polymorphisms within the Fn-binding repeats of FnBPA explain, at least partially, the statistically (p < 0.05) longer bond lifetime for isolates associated with an infected cardiovascular device. PMID:22219202

  17. Disk Drives

    NASA Technical Reports Server (NTRS)

    1994-01-01

    A new material known as AlBeMet, developed by Brush Wellman for research applications in the National Aero-Space Plane (NASP) program, is now used for high performance disk drives. AlBeMet is a compression of aluminum, beryllium metal matrix composite. It reduces system weight and its high thermal conductivity can effectively remove heat and increase an electrical system's lifetime. The lighter, stiffer AlBeMet (AlBeMet 160) used in the disk drive means heads can be moved faster, improving disk performance.

  18. Focal fits during chlorambucil therapy

    PubMed Central

    Naysmith, A.; Robson, R. H.

    1979-01-01

    An elderly man receiving chlorambucil for chronic lymphatic leukaemia developed focal fits. The onset and frequency were dose related. There was no evidence of metabolic disturbance or of meningeal leukaemia. Although reported in children and well recognized in animals, chlorambucil-induced fits in an adult have not been previously recorded. PMID:118440

  19. Vermont management in focal areas

    Treesearch

    Judy Rosovsky; Bruce L. Parker; Luke Curtis

    1991-01-01

    Following the 1979 outbreak of gypsy moths Lymantria dispar L. in Vermont, state personnel began monitoring a number of focal areas for signs of increase in gypsy moth populations. In 1986 data from this early warning system indicated an incipient outbreak. We took advantage of this increase to test an experimental management technique. Would...

  20. Statistical earthquake focal mechanism forecasts

    NASA Astrophysics Data System (ADS)

    Kagan, Yan Y.; Jackson, David D.

    2014-04-01

    Forecasts of the focal mechanisms of future shallow (depth 0-70 km) earthquakes are important for seismic hazard estimates and Coulomb stress, and other models of earthquake occurrence. Here we report on a high-resolution global forecast of earthquake rate density as a function of location, magnitude and focal mechanism. In previous publications we reported forecasts of 0.5° spatial resolution, covering the latitude range from -75° to +75°, based on the Global Central Moment Tensor earthquake catalogue. In the new forecasts we have improved the spatial resolution to 0.1° and the latitude range from pole to pole. Our focal mechanism estimates require distance-weighted combinations of observed focal mechanisms within 1000 km of each gridpoint. Simultaneously, we calculate an average rotation angle between the forecasted mechanism and all the surrounding mechanisms, using the method of Kagan & Jackson proposed in 1994. This average angle reveals the level of tectonic complexity of a region and indicates the accuracy of the prediction. The procedure becomes problematical where longitude lines are not approximately parallel, and where shallow earthquakes are so sparse that an adequate sample spans very large distances. North or south of 75°, the azimuths of points 1000 km away may vary by about 35°. We solved this problem by calculating focal mechanisms on a plane tangent to the Earth's surface at each forecast point, correcting for the rotation of the longitude lines at the locations of earthquakes included in the averaging. The corrections are negligible between -30° and +30° latitude, but outside that band uncorrected rotations can be significantly off. Improved forecasts at 0.5° and 0.1° resolution are posted at http://eq.ess.ucla.edu/kagan/glob_gcmt_index.html.

  1. Immunohistochemical expression of fibronectin in the lungs of fire victims proves intravital reaction in fatal burns.

    PubMed

    Bohnert, Michael; Anderson, Jürina; Rothschild, Markus A; Böhm, Joachim

    2010-11-01

    Immunohistochemical studies about the presence of fibronectin in the lungs were performed in a group of 73 fire victims (63 cases of intravital and 10 cases of postmortem burn) as well as in an unselected control group of 55 individuals not exposed to fire before death. The cases of intravital burn showed a significantly stronger fibronectin expression than the control cases and the cases of postmortem burn. Fibronectin was mainly present in macrophages of the peribronchial lung parenchyma and, not associated with cells, in the matrix of peribronchial tissue. Our findings suggest that higher levels of fibronectin expression in the lung tissue of burn victims compared to fire-unrelated deaths may serve as an indicator of an early intravital inflammatory response to fire damage.

  2. Fibrillar organization of fibronectin is expressed coordinately with cell surface gangliosides in a variant murine fibroblast

    PubMed Central

    1986-01-01

    NCTC 2071A cells, a line of transformed murine fibroblasts, grow in serum-free medium, are deficient in gangliosides, synthesize fibronectin, but do not retain and organize it on the cell surface. When the cells are exposed to exogenous gangliosides, fibrillar strands of fibronectin become attached to the cell surface. A morphologically distinct variant of NCTC 2071A cells was observed to both retain cell surface fibronectin and organize it into a fibrillar network when the cells were stained with anti-fibronectin antibodies and a fluorescent second antibody. A revertant cell type appeared to resemble the parental NCTC 2071A cells in terms of morphology and fibronectin organization. All three cell types were subjected to mild NaIO4 oxidation and reduction with KB3H4 of very high specific radioactivity in order to label the sialic acid residues of surface gangliosides. The variant had much more surface gangliosides than the parental, particularly more complex gangliosides corresponding to GM1 and GD1a. The surface gangliosides of the revertant were intermediate between the parental and the variant. By using sialidase, which hydrolyzes GD1a to GM1, and 125I-labeled cholera toxin, which binds specifically to GM1, the identity and levels of these gangliosides were confirmed in the three cell types. When variant cells were exposed to sialidase for 2 d, there appeared to be little change in fibronectin organization. Concomitant treatment of the cells with the B subunit of cholera toxin, which bound to all the surface GM1 including that generated by the sialidase, however, eliminated the fibrillar network of fibronectin. In addition, exposure of the variant cells to a 70,000-mol-wt fragment of fibronectin, which lacks the cell attachment domain but contains a matrix assembly domain, inhibited the formation of fibers. Finally, all three cell types were assayed for their ability to attach to and spread on fibronectin-coated surfaces; no significant differences were found

  3. Focal Adhesion Kinase Regulates Fibroblast Migration via Integrin beta-1 and Plays a Central Role in Fibrosis.

    PubMed

    Zhao, Xue-Ke; Cheng, Yiju; Liang Cheng, Ming; Yu, Lei; Mu, Mao; Li, Hong; Liu, Yang; Zhang, Baofang; Yao, Yumei; Guo, Hui; Wang, Rong; Zhang, Quan

    2016-01-14

    Lung fibrosis is a major medical problem for the aging population worldwide. Fibroblast migration plays an important role in fibrosis. Focal Adhesion Kinase (FAK) senses the extracellular stimuli and initiates signaling cascades that promote cell migration. This study first examined the dose and time responses of FAK activation in human lung fibroblasts treated with platelet derived growth factor BB (PDGF-BB). The data indicate that FAK is directly recruited by integrin β1 and the subsequent FAK activation is required for fibroblast migration on fibronectin. In addition, the study has identified that α5β1 and α4β1 are the major integrins for FAK-mediated fibroblast migration on fibronect. In contrast, integrins αvβ3, αvβ6, and αvβ8 play a minor but distinct role in fibroblast migration on fibronectin. FAK inhibitor significantly reduces PDGF-BB stimulated fibroblast migration. Importantly, FAK inhibitor protects bleomycin-induced lung fibrosis in mice. FAK inhibitor blocks FAK activation and significantly reduces signaling cascade of fibroblast migration in bleomycin-challenged mice. Furthermore, FAK inhibitor decreases lung fibrotic score, collagen accumulation, fibronectin production, and myofibroblast differentiation in in bleomycin-challenged mice. These data demonstrate that FAK mediates fibroblast migration mainly via integrin β1. Furthermore, the findings suggest that targeting FAK signaling is an effective therapeutic strategy against fibrosis.

  4. Adhesion, unfolding forces, and molecular elasticity of fibronectin coatings: An atomic force microscopy study.

    PubMed

    Sumarokova, Maria; Iturri, Jagoba; Toca-Herrera, José L

    2017-10-07

    Fibronectin is an extracellular matrix protein that is involved in cell adhesion, growth, migration, differentiation, and wound healing. Fibronectin coatings are currently used in many laboratories for biomedical and biotechnology purposes. In this study we have investigated the adhesion and mechanical properties of fibronectin coatings. The coatings were also used to study the role of the residence time and the influence of the loading rate in nonspecific interactions. The results showed that the adhesion force between silica and fibronectin increased with loading rate delivering similar values for residence times of 1 and 2 s. Further analysis indicated that the distance to the transition state was about 0.5 nm. Moreover, the adhesion force did not vary with the loading rate for contact time of 0 s. The unfolding of fibronectin domains also depended of the Dwell time (no unfolding events were observed for zero residence time). Applied loads of 2 nN were able to stretch the fibronectin layer up to 200 nm and to unfold the three fibronectin domains, which were similar for a Dwell time of 1 and 2 s. However, the unfolding length increased with loading rate: below 2.5 µm s(-1) the obtained lengths matched the value of FN I (13.5 nm), while for higher speeds the measured values corresponded to the lengths of FN II (18 nm) and FN III (27 nm). This investigation has answered and opened new questions about the mechanical stability and function of fibronectin coatings. The results have also raised theoretical questions about the difference between specific and nonspecific interactions to be addressed in future work. © 2017 Wiley Periodicals, Inc.

  5. Interaction between Fibronectin and β1 Integrin Is Essential for Tooth Development

    PubMed Central

    Yamada, Aya; Yuasa, Kenji; Yoshizaki, Keigo; Iwamoto, Tsutomu; Saito, Masahiro; Nakamura, Takashi; Fukumoto, Satoshi

    2015-01-01

    The dental epithelium and extracellular matrix interact to ensure that cell growth and differentiation lead to the formation of teeth of appropriate size and quality. To determine the role of fibronectin in differentiation of the dental epithelium and tooth formation, we analyzed its expression in developing incisors. Fibronectin mRNA was expressed during the presecretory stage in developing dental epithelium, decreased in the secretory and early maturation stages, and then reappeared during the late maturation stage. The binding of dental epithelial cells derived from postnatal day-1 molars to a fibronectin-coated dish was inhibited by the RGD but not RAD peptide, and by a β1 integrin-neutralizing antibody, suggesting that fibronectin-β1 integrin interactions contribute to dental epithelial-cell binding. Because fibronectin and β1 integrin are highly expressed in the dental mesenchyme, it is difficult to determine precisely how their interactions influence dental epithelial differentiation in vivo. Therefore, we analyzed β1 integrin conditional knockout mice (Intβ1lox-/lox-/K14-Cre) and found that they exhibited partial enamel hypoplasia, and delayed eruption of molars and differentiation of ameloblasts, but not of odontoblasts. Furthermore, a cyst-like structure was observed during late ameloblast maturation. Dental epithelial cells from knockout mice did not bind to fibronectin, and induction of ameloblastin expression in these cells by neurotrophic factor-4 was inhibited by treatment with RGD peptide or a fibronectin siRNA, suggesting that the epithelial interaction between fibronectin and β1 integrin is important for ameloblast differentiation and enamel formation. PMID:25830530

  6. Interaction between fibronectin and β1 integrin is essential for tooth development.

    PubMed

    Saito, Kan; Fukumoto, Emiko; Yamada, Aya; Yuasa, Kenji; Yoshizaki, Keigo; Iwamoto, Tsutomu; Saito, Masahiro; Nakamura, Takashi; Fukumoto, Satoshi

    2015-01-01

    The dental epithelium and extracellular matrix interact to ensure that cell growth and differentiation lead to the formation of teeth of appropriate size and quality. To determine the role of fibronectin in differentiation of the dental epithelium and tooth formation, we analyzed its expression in developing incisors. Fibronectin mRNA was expressed during the presecretory stage in developing dental epithelium, decreased in the secretory and early maturation stages, and then reappeared during the late maturation stage. The binding of dental epithelial cells derived from postnatal day-1 molars to a fibronectin-coated dish was inhibited by the RGD but not RAD peptide, and by a β1 integrin-neutralizing antibody, suggesting that fibronectin-β1 integrin interactions contribute to dental epithelial-cell binding. Because fibronectin and β1 integrin are highly expressed in the dental mesenchyme, it is difficult to determine precisely how their interactions influence dental epithelial differentiation in vivo. Therefore, we analyzed β1 integrin conditional knockout mice (Intβ1lox-/lox-/K14-Cre) and found that they exhibited partial enamel hypoplasia, and delayed eruption of molars and differentiation of ameloblasts, but not of odontoblasts. Furthermore, a cyst-like structure was observed during late ameloblast maturation. Dental epithelial cells from knockout mice did not bind to fibronectin, and induction of ameloblastin expression in these cells by neurotrophic factor-4 was inhibited by treatment with RGD peptide or a fibronectin siRNA, suggesting that the epithelial interaction between fibronectin and β1 integrin is important for ameloblast differentiation and enamel formation.

  7. Quantitative Studies of Endothelial Cell Fibronectin and Filamentous Actin (F-Actin) Coalignment in Response to Shear Stress.

    PubMed

    Gong, Xianghui; Zhao, Xixi; Li, Bin; Sun, Yan; Liu, Meili; Huang, Yan; Jia, Xiaoling; Ji, Jing; Fan, Yubo

    2017-09-12

    Both fibronectin (FN) and filamentous actin (F-actin) fibers play a critical role for endothelial cells (ECs) in responding to shear stress and modulating cell alignment and functions. FN is dynamically coupled to the F-actin cytoskeleton via focal adhesions. However, it is unclear how ECs cooperatively remodel their subcellular FN matrix and intracellular F-actin cytoskeleton in response to shear stress. Current studies are hampered by the lack of a reliable and sensitive quantification method of FN orientation. In this study, we developed a MATLAB-based feature enhancement method to quantify FN and F-actin orientation. The role of F-actin in FN remodeling was also studied by treating ECs with cytochalasin D. We have demonstrated that FN and F-actin codistributed and coaligned parallel to the flow direction, and that F-actin alignment played an essential role in regulating FN alignment in response to shear stress. Our findings offer insight into how ECs cooperatively remodel their subcellular ECM and intracellular F-actin cytoskeleton in response to mechanical stimuli, and are valuable for vascular tissue engineering.

  8. Fibronectin gene expression in human adipose tissue and its associations with obesity-related genes and metabolic parameters.

    PubMed

    Lee, Seoung Hee; Park, Hye Soon; Lee, Jung Ah; Song, Young Sook; Jang, Yeon Jin; Kim, Jong-Hyeok; Lee, Yeon Ji; Heo, Yoonseok

    2013-04-01

    Limited data are available on the in vivo expression of fibronectin, one of the main extracellular matrix components. We investigated the expression of fibronectin in abdominal visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) and the associations of leptin, adiponectin, and vaspin gene expression with metabolic parameters in obese women. We recruited female subjects undergoing bariatric surgery for obesity (n = 24) and patients undergoing benign gynecological surgery as the control group (n = 23). We measured anthropometric variables, abdominal fat distribution, metabolic parameters, serum concentrations of leptin, adiponectin, and vaspin, and depot-specific mRNA expression of fibronectin, leptin, adiponectin, and vaspin. Fibronectin expression in both VAT and SAT was significantly lower in the obese group than in the control group. Fibronectin expression in both VAT and SAT were negatively correlated with body mass index or waist circumference, with higher prominence in VAT. In multiple regression analysis, fibronectin expression in both VAT and SAT was negatively correlated with serum leptin concentration. Fibronectin expression in VAT was negatively correlated with leptin expression in VAT. Additionally, fibronectin expression in SAT was negatively correlated with leptin expression in SAT and positively correlated with adiponectin expression in VAT and SAT. We found significant negative associations between depot-specific fibronectin expression in human adipose tissue and obesity indices and obesity-related biomarkers. Our results suggest that fibronectin expression may contribute to obesity and metabolic dysregulation in humans.

  9. Neuropilin-1 stimulates tumor growth by increasing fibronectin fibril assembly in the tumor microenvironment.

    PubMed

    Yaqoob, Usman; Cao, Sheng; Shergill, Uday; Jagavelu, Kumaravelu; Geng, Zhimin; Yin, Meng; de Assuncao, Thiago M; Cao, Ying; Szabolcs, Anna; Thorgeirsson, Snorri; Schwartz, Martin; Yang, Ju Dong; Ehman, Richard; Roberts, Lewis; Mukhopadhyay, Debabrata; Shah, Vijay H

    2012-08-15

    The tumor microenvironment, including stromal myofibroblasts and associated matrix proteins, regulates cancer cell invasion and proliferation. Here, we report that neuropilin-1 (NRP-1) orchestrates communications between myofibroblasts and soluble fibronectin that promote α5β1 integrin-dependent fibronectin fibril assembly, matrix stiffness, and tumor growth. Tumor growth and fibronectin fibril assembly were reduced by genetic depletion or antibody neutralization of NRP-1 from stromal myofibroblasts in vivo. Mechanistically, the increase in fibronectin fibril assembly required glycosylation of serine 612 of the extracellular domain of NRP-1, an intact intracellular NRP-1 SEA domain, and intracellular associations between NRP-1, the scaffold protein GIPC, and the nonreceptor tyrosine kinase c-Abl that augmented α5β1 fibronectin fibril assembly activity. Analysis of human cancer specimens established an association between tumoral NRP-1 levels and clinical outcome. Our findings indicate that NRP-1 activates the tumor microenvironment, thereby promoting tumor growth. These results not only identify new molecular mechanisms of fibronectin fibril assembly but also have important implications for therapeutic targeting of the myofibroblast in the tumor microenvironment. ©2012 AACR.

  10. Women’s perspectives of the fetal fibronectin testing process: a qualitative descriptive study

    PubMed Central

    2014-01-01

    Background In 2009 the Ontario Ministry of Health and Long Term Care funded the implementation of province-wide fetal fibronectin testing in Ontario hospitals. This paper reports results from the provincial evaluation that sought to describe the experience of fetal fibronectin testing from the perspective of women with symptoms of preterm labour. Methods A descriptive qualitative design was used, employing semi-structured telephone and face-to-face interviews with women who had fetal fibronectin testing. Results Five hospitals participated in recruiting women for the study and 17 women were interviewed. Women described their experiences of fetal fibronectin testing as an emotional process that moves from expecting, to feeling, to hoping for reassurance; and then to re-defining what is required to feel reassured. Women described feeling anxious while waiting for fetal fibronectin results. When test results were negative, women described feeling a sense of relief that their symptoms would not likely lead to an imminent preterm birth. Women with positive results expressed feeling reassured by the care decisions and quick action taken by the health care team. Conclusion Fetal fibronectin testing was acceptable and beneficial to these women with symptoms of preterm labour. Implications for practice and future research are suggested. PMID:24894630

  11. Women's perspectives of the fetal fibronectin testing process: a qualitative descriptive study.

    PubMed

    Peterson, Wendy E; Sprague, Ann E; Reszel, Jessica; Walker, Mark; Fell, Deshayne B; Perkins, Sherry L; Dunn, Sandra I; Johnson, Moya

    2014-06-04

    In 2009 the Ontario Ministry of Health and Long Term Care funded the implementation of province-wide fetal fibronectin testing in Ontario hospitals. This paper reports results from the provincial evaluation that sought to describe the experience of fetal fibronectin testing from the perspective of women with symptoms of preterm labour. A descriptive qualitative design was used, employing semi-structured telephone and face-to-face interviews with women who had fetal fibronectin testing. Five hospitals participated in recruiting women for the study and 17 women were interviewed. Women described their experiences of fetal fibronectin testing as an emotional process that moves from expecting, to feeling, to hoping for reassurance; and then to re-defining what is required to feel reassured. Women described feeling anxious while waiting for fetal fibronectin results. When test results were negative, women described feeling a sense of relief that their symptoms would not likely lead to an imminent preterm birth. Women with positive results expressed feeling reassured by the care decisions and quick action taken by the health care team. Fetal fibronectin testing was acceptable and beneficial to these women with symptoms of preterm labour. Implications for practice and future research are suggested.

  12. Galectin-3-induced cell spreading and motility relies on distinct signaling mechanisms compared to fibronectin.

    PubMed

    More, Shyam K; Chiplunkar, Shubhada V; Kalraiya, Rajiv D

    2016-05-01

    Secreted galectin-3 often gets incorporated into extracellular matrix and is utilized by cancer cells for spreading, movement, and metastatic dissemination. Here we investigate molecular mechanisms by which galectin-3 brings about these effects and compare it with fibronectin. Imaging of cells spread on fibronectin showed stress fibers throughout cell body, however, galectin-3-induced formation of parallel actin bundles in the lamellipodial region resulting in unique morphological features. FRAP analysis showed that the actin turnover in the lamellipodial region was much higher in cells spread on galectin-3 as compared to that on fibronectin. Rac1 activation is correlated with lamellipodial organization on both the substrates. Activation of Akt and Rac1, the regulators of actin dynamics, show inverse correlation with each other on both galectin-3 and fibronectin. Activation of Erk however, remained similar. Further, inhibition of activation of Akt and Erk inhibited spreading and motility of cells on galectin-3 but not on fibronectin. The results very comprehensively demonstrate distinct signaling pathways that regulate microfilament organization, lamellipodial structures, spreading, and movement of cells plated on galectin-3 as opposed to fibronectin.

  13. Fibronectin-like protein in Porifera: its role in cell aggregation.

    PubMed Central

    Labat-Robert, J; Robert, L; Auger, C; Lethias, C; Garrone, R

    1981-01-01

    Experiments were carried out on a freshwater sponge (Ephydatia mulleri) in order to demonstrate the presence of fibronectin in Porifera. By using antibodies to highly purified human plasma fibronectin, the presence of a similar or identical protein could be demonstrated in the membranes of E. mulleri cells such as epithelial cells, fibroblast-like cells, and choanocytes. The reaction was specific, could be abolished by the addition of excess fibronectin, and was not observed with nonimmune rabbit serum. The immune fluorescent reaction became stronger when the sponge cells were pretreated with acetone and could also be observed, although with a less intense staining, on the intercellular matrix. This shows the predominant presence of a sponge fibronectin-like protein in the cell membranes and also its presence to a lesser extent in the intercellular matrix. When dissociated sponge cells were led to reassociate under the microscope, reassociation could be completely inhibited by anti-human fibronectin antiserum up to a dilution of 1:120 and partially inhibited up to a dilution of 1:240. The reassociation of dissociated sponge cells could also be inhibited by the addition of purified gelatin but not with serum albumin or with a normal, nonimmune rabbit serum. These results clearly indicate that a sponge cell fibronectin-like protein may play an important role as the (or one of the) recognition site(s) of the aggregation factor(s) and can therefore be directly involved in cell association, morphogenesis, and differentiation. Images PMID:7031644

  14. Binding of human fibronectin to group A, C, and G streptococci.

    PubMed Central

    Myhre, E B; Kuusela, P

    1983-01-01

    A total of 387 bacterial strains belonging to 35 species were tested in direct binding experiments for the uptake of purified radiolabeled human fibronectin. Positive binding was found in group A, C, and G streptococci and in Staphylococcus aureus. The group C streptococcal species Streptococcus equisimilis, Streptococcus zooepidemicus, Streptococcus equi, and Streptococcus dysgalactiae were uniformly reactive with fibronectin. Beta-hemolytic bovine group G streptococci showed the same degree of reactivity as human group G strains. In contrast, only 4 out of 15 alpha-hemolytic bovine group G strains were able to bind fibronectin. The uptake of fibronectin measured at room temperature with a human group G streptococcus was a slow, time-dependent process with maximum binding after approximately 1 h. Human polyclonal immunoglobulin G and serum albumin tested in inhibition experiments did not affect the fibronectin binding. Fibronectin seems, therefore, to interact with a surface component that is different from the specific binding sites previously described for human immunoglobulin G and serum albumin. PMID:6299959

  15. Akt1 Signaling Regulates Integrin Activation, Matrix Recognition, and Fibronectin Assembly*S

    PubMed Central

    Somanath, Payaningal R.; Kandel, Eugene S.; Hay, Nissim; Byzova, Tatiana V.

    2009-01-01

    Akt, a serine-threonine kinase, regulates multiple cellular processes in vascular cells. We have previously documented that Akt activates integrins and Akt1 deficiency results in matrix abnormalities in skin and blood vessels in vivo. Based on these observations, we hypothesized that Akt1 is necessary for integrin activation and matrix assembly by fibroblasts. In this study, using various cell systems, we show that Akt1 is essential for the inside-out activation of integrins in endothelial cells and fibroblasts, which in turn, mediates matrix assembly. Fibronectin is a major extracellular matrix component of the skin and the vascular basement membrane, which possesses binding sites for many integrins and extracellular matrix proteins. Akt1−/− fibroblasts and NIH fibroblasts expressing dominant negative Akt1 (K179M-Akt1) showed impaired fibronectin assembly compared with control fibroblasts. In contrast, expression of constitutively active Akt1 (myrAkt1) resulted in enhanced fibronectin assembly. Although increased fibronectin assembly by myrAkt1-expressing human foreskin fibroblasts was abolished by treatment with anti-integrin β1 blocking antibodies, treatment with β1-stimulating antibodies rescued the impaired fibronectin assembly that was due to lack of Akt activity. Finally, expression of myrAkt1 corrected the phenotype of Akt1−/− fibroblasts thus showing that Akt1 regulates fibronectin assembly through activation of integrin α5β1. PMID:17562714

  16. Predictive value of cervical length measurement and fibronectin testing in threatened preterm labor.

    PubMed

    van Baaren, Gert-Jan; Vis, Jolande Y; Wilms, Femke F; Oudijk, Martijn A; Kwee, Anneke; Porath, Martina M; Oei, Guid; Scheepers, Hubertina C J; Spaanderman, Marc E A; Bloemenkamp, Kitty W M; Haak, Monique C; Bolte, Antoinette C; Bax, Caroline J; Cornette, Jérôme M J; Duvekot, Johannes J; Nij Bijvanck, Bas W A; van Eyck, Jim; Franssen, Maureen T M; Sollie, Krystyna M; Vandenbussche, Frank P H A; Woiski, Mallory; Grobman, William A; van der Post, Joris A M; Bossuyt, Patrick M M; Opmeer, Brent C; Mol, Ben W J

    2014-06-01

    To estimate the performance of combining cervical length measurement with fetal fibronectin testing in predicting delivery in women with symptoms of preterm labor. We conducted a prospective nationwide cohort study in all 10 perinatal centers in The Netherlands. Women with symptoms of preterm labor between 24 and 34 weeks of gestation with intact membranes were included. In all women, qualitative fibronectin testing (0.050-microgram/mL cutoff) and cervical length measurement were performed. Logistic regression was used to predict spontaneous preterm delivery within 7 days after testing. A risk less than 5%, corresponding to the risk for women with a cervical length of at least 25 mm, was considered as low risk. Between December 2009 and August 2012, 714 women were enrolled. Fibronectin results and cervical length were available for 665 women, of whom 80 (12%) delivered within 7 days. Women with a cervical length of at least 30 mm or with a cervical length between 15 and 30 mm with a negative fibronectin result were at low risk (less than 5%) of spontaneous delivery within 7 days. Fibronectin testing in case of a cervical length between 15 and 30 mm additionally classified 103 women (15% of the cohort) as low risk and 36 women (5% of the cohort) as high risk. Cervical length measurement, combined with fetal fibronectin testing in case of a cervical length between 15 and 30 mm, improves identification of women with a low risk to deliver spontaneously within 7 days. II.

  17. Fibronectin biosynthesis and cell-surface expression by cardiac and non-cardiac endothelial cells.

    PubMed Central

    Johnson, C. M.; Helgeson, S. C.

    1993-01-01

    We examined the biosynthesis and surface expression of fibronectin, an adhesive glycoprotein, in several types of cultured porcine endothelial cells: pulmonary artery, thoracic aorta, coronary artery, aortic valve, and mitral valve. We used immunocytochemical staining to compare the levels of fibronectin present in these same tissues in vivo. Using endogenous radiolabeling, we found that all cell types except aortic valve endothelial cells synthesized and released into the culture media substantial quantities of fibronectin. Using radioiodination of intact cells, we found that, whereas both thoracic aorta and pulmonary artery cells had measurable fibronectin on the surface, aortic valve, mitral valve, and coronary artery cells had little cell-surface fibronectin present. Immunocytochemical staining showed that all endothelial regions except aortic valve had substantial quantities of immunoreactive fibronectin in vivo. These data suggest that the aortic valve endothelium may be distinct from other endothelia. Such differences could be important for the pathogenesis of valvular disease. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:8494044

  18. Conformational changes of plasma fibronectin detected upon adsorption to solid substrates: A spin-label study

    SciTech Connect

    Narasimhan, C.; Lai, Chingsan )

    1989-06-13

    Changes in local environment of the free sulfhydryl groups in plasma fibronectin upon adsorption of the protein to polystyrene beads have been examined by electron spin resonance (ESR) spin-label spectroscopy. The two free sulfhydryl groups per subunit of plasma fibronectin were modified chemically with an ({sup 15}N,{sup 2}H)maleimide spin-label. For soluble fibronectin, both free sulfhydryl groups shown to be in confined environments as evidenced from the labeled protein exhibiting a strongly immobilized ESR spectrum as described previously using ({sup 14}N,{sup 1}H)maleimide spin-labels. When the labeled protein was adsorbed to the beads, half of the strongly immobilized component was found to convert into a weakly immobilized component, a result indicating that one of the two labeled sites becomes exposed and exhibits a fast tumbling motion. Experiments conducted using various spin-labeled fibronectin fragments suggest that the newly exposed labeled site is located between the DNA-binding and the cell-binding regions of the molecule. The data obtained indicate that, upon adsorption to polystyrene beads, plasma fibronectin undergoes a conformational change through which the buried free sulfhydryl group near the cell-binding region of the molecule is exposed. This observation may have important implications regarding the expression of cell adhesive properties of the fibronectin molecule.

  19. Decreased interaction of fibronectin, type IV collagen, and heparin due to nonenzymatic glycation. Implications for diabetes mellitus

    SciTech Connect

    Tarsio, J.F.; Reger, L.A.; Furcht, L.T.

    1987-02-24

    The nonenzymatic glycation of basement membrane proteins, such as fibronectin and type IV collagen, occurs in diabetes mellitus. These proteins are nonenzymatically glycated in vivo and can also be nonenzymatically glycated in vitro. After 12 days of incubation at 37 /sup 0/C with 500 mM glucose, purified samples of human plasma fibronectin and native type IV collagen showed a 13.0- and 4.2-fold increase, respectively, in glycated amino acid levels in comparison to control samples incubated in the absence of glucose. Gelatin (denatured calfskin collagen) was glycated 22.3-fold under the same conditions. Scatchard analyses were performed on the binding of radiolabeled fibronectin to gelatin or type IV collagen. It was found that there is a 3-fold reduction in the affinity of fibronectin to type IV collagen due to the nonenzymatic glycation of fibronectin. The molecular association of control fibronectin or its glycated counterpart with (/sup 3/H)heparin was also determined. Scatchard analyses of this interaction showed no difference between control fibronectin and glycated fibronectin in (/sup 3/H)heparin binding. When fibronectin and (/sup 3/H)heparin were incubated together with type IV collagen at fixed concentrations, the binding of (/sup 3/H)heparin to fibronectin was significantly enhanced by the addition of type IV collagen. This positive cooperative binding of heparin with fibronectin and type IV collagen was reduced approximately 36% when fibronectin was nonenzymatically glycated, 64% when type IV collagen was nonenzymatically glycated, and 70% when both fibronectin and type IV collagen were nonenzymatically glycated. These observations with native type IV collagen are discussed relevant to changes in the molecular composition of basement membranes that occur in diabetes mellitus.

  20. Small Molecule Inhibitors Target the Tissue Transglutaminase and Fibronectin Interaction

    PubMed Central

    Yakubov, Bakhtiyor; Chen, Lan; Belkin, Alexey M.; Zhang, Sheng; Chelladurai, Bhadrani; Zhang, Zhong-Yin; Matei, Daniela

    2014-01-01

    Tissue transglutaminase (TG2) mediates protein crosslinking through generation of ε−(γ-glutamyl) lysine isopeptide bonds and promotes cell adhesion through interaction with fibronectin (FN) and integrins. Cell adhesion to the peritoneal matrix regulated by TG2 facilitates ovarian cancer dissemination. Therefore, disruption of the TG2-FN complex by small molecules may inhibit cell adhesion and metastasis. A novel high throughput screening (HTS) assay based on AlphaLISA™ technology was developed to measure the formation of a complex between His-TG2 and the biotinylated FN fragment that binds TG2 and to discover small molecules that inhibit this protein-protein interaction. Several hits were identified from 10,000 compounds screened. The top candidates selected based on >70% inhibition of the TG2/FN complex formation were confirmed by using ELISA and bioassays measuring cell adhesion, migration, invasion, and proliferation. In conclusion, the AlphaLISA bead format assay measuring the TG2-FN interaction is robust and suitable for HTS of small molecules. One compound identified from the screen (TG53) potently inhibited ovarian cancer cell adhesion to FN, cell migration, and invasion and could be further developed as a potential inhibitor for ovarian cancer dissemination. PMID:24586660

  1. Fibronectin-Grafted Titanium Dental Implants: An In Vivo Study.

    PubMed

    Chang, Yu-Chi; Ho, Kuo-Ning; Feng, Sheng-Wei; Huang, Haw-Ming; Chang, Chia-Hsun; Lin, Che-Tong; Teng, Nai-Chia; Pan, Yu Hwa; Chang, Wei-Jen

    2016-01-01

    Modification of the physiochemical properties of titanium surfaces using glow discharge plasma (GDP) and fibronectin coating has been shown to enhance the surface hydrophilicity, surface roughness, cell adhesion, migration, and proliferation. This in vivo study aimed to evaluate the bone integration efficacy of a biologically modified implant surface. Two different surface-modified implants (Ar-GDP and GDP-fib) were placed in the mandibular premolar area of six beagle dogs for 2-8 weeks. Three techniques [histologic evaluation, resonance frequency analysis (RFA), and microcomputed tomography (micro-CT) evaluation] were used to detect the implant stability and bone-implant contact. The implant stability quotient values of GDP-fib implants were significantly greater than the Ar-GDP implants at 2 and 4 weeks (P < 0.01). The bone volume/total volume ratio of GDP-fib implants was greater than the Ar-GDP implants in micro-CT evaluation. A high positive correlation was observed between RFA and micro-CT measurements. At 2 weeks, osteoblasts were seen to line the implant surface, and multinuclear osteoclasts could be seen on the surface of old parent bone. After 8 weeks, a majority of the space in the wound chamber appeared to be replaced by bone. Enhancement of the stability of biologically modified implants was proved by the results of RFA, micro-CT, and histological analysis. This enhanced stability may help fasten treatment and be clinically beneficial.

  2. Dissecting the interaction between transglutaminase 2 and fibronectin.

    PubMed

    Cardoso, Inês; Østerlund, Eva Christina; Stamnaes, Jorunn; Iversen, Rasmus; Andersen, Jan Terje; Jørgensen, Thomas J D; Sollid, Ludvig M

    2017-03-01

    In the extracellular environment, the enzyme transglutaminase 2 (TG2) is involved in cell-matrix interactions through association with the extracellular matrix protein, fibronectin (FN). The 45 kDa gelatin-binding domain of FN (45FN) is responsible for the binding to TG2. Previous studies have demonstrated that the FN-binding site of TG2 is located in the N-terminal domain of the enzyme although with conflicting results regarding the specific residues involved. Here we have mapped the FN interaction site of human TG2 by use of hydrogen/deuterium exchange coupled with mass spectrometry, and we confirm that the FN-binding site is located in the N-terminal domain of TG2. Furthermore, by combination of site-directed mutagenesis and surface plasmon resonance analysis we have identified the TG2 residues K30, R116 and H134 as crucial to maintain the high affinity interaction with FN. Mutation of all three residues simultaneously reduced binding to 45FN by more than 2000-fold. We also identified residues in the catalytic core domain of TG2 that contributed to FN binding, hence extending the binding interface between TG2 and FN. This study provides new insights into the high affinity interaction between TG2 and FN.

  3. Exposure to biomass smoke extract enhances fibronectin release from fibroblasts.

    PubMed

    Krimmer, David; Ichimaru, Yukikazu; Burgess, Janette; Black, Judith; Oliver, Brian

    2013-01-01

    COPD induced following biomass smoke exposure has been reported to be associated with a more fibrotic phenotype than cigarette smoke induced COPD. This study aimed to investigate if biomass smoke induced extracellular matrix (ECM) protein production from primary human lung fibroblasts in vitro. Primary human lung fibroblasts (n=5-10) were stimulated in vitro for up to 72 hours with increasing concentrations of biomass smoke extract (BME) or cigarette smoke extract (CSE) prior to being assessed for deposition of ECM proteins, cytokine release, and activation of intracellular signalling molecules. Deposition of the ECM proteins perlecan and fibronectin was upregulated by both CSE (p<0.05) and BME (p<0.05). The release of the neutrophilic chemokine IL-8 was also enhanced by BME. ERK1/2 phosphorylation was significantly upregulated by BME (p<0.05). Chemical inhibition of ERK signalling molecules partially attenuated these effects (p<0.05). Stimulation with endotoxin had no effect. This study demonstrated that BME had similar effects to CSE in vitro and had the capacity to directly induce fibrosis by upregulating production of ECM proteins. The mechanisms by which both biomass and cigarette smoke exposure cause lung damage may be similar.

  4. Human macrophage differentiation involves an interaction between integrins and fibronectin

    SciTech Connect

    Laouar, A.; Chubb, C.B.H.; Collart, F.; Huberman, E.

    1997-03-14

    The authors have examined the role of integrins and extracellular matrix (ECM) proteins in macrophage differentiation of (1) human HL-60 myeloid leukemia cells induced by phorbol 12-myristate 13-acetate (PMA) and (2) human peripheral blood monocytes induced by either PMA or macrophage-colony stimulating factor (M-CSF). Increased {beta}{sub 1} integrin and fibronectin (FN) gene expression was observed in PMA-treated HL-60 cells and PMA- or M-CSF-treated monocytes, even at a time preceding the manifestation of macrophage markers. Treated HL-60 cells and monocytes also released and deposited FN on the culture dishes. An HL-60 cell variant, HL-525, which is deficient in protein kinase C {beta} (PKC{beta}) and resistant to PMA-induced differentiation, failed to express FN after PMA treatment. Restoration of PKC{beta} resulted in PMA-induced FN gene expression and macrophage differentiation. The macrophage phenotype induced in HL-60 cells or monocytes was attenuated by anti-{beta}{sub 1} integrin or anti-FN MAbs. The authors suggest that macrophage differentiation involves activation of PKC and expression of specific integrins and ECM proteins. The stimulated cells, through their integrins, attach and spread on these substrates by binding to the deposited ECM proteins. This attachment and spreading in turn, through integrin signaling, leads to the macrophage phenotype.

  5. Fibronectin regulates Wnt7a signaling and satellite cell expansion

    PubMed Central

    Bentzinger, C. Florian; Wang, Yu Xin; von Maltzahn, Julia; Soleimani, Vahab D.; Yin, Hang; Rudnicki, Michael A.

    2012-01-01

    SUMMARY The influence of the extracellular matrix (ECM) within the stem cell niche remains poorly understood. We found that Syndecan-4 (Sdc4) and Frizzled-7 (Fzd7) form a co-receptor complex in satellite cells and that binding of the ECM glycoprotein Fibronectin (FN) to Sdc4 stimulates the ability of Wnt7a to induce the symmetric expansion of satellite stem cells. Newly activated satellite cells dynamically remodel their niche by transient high-level expression of FN. Knockdown of FN in prospectively isolated satellite cells severely impaired their ability to repopulate the satellite cell niche. Conversely, in vivo over-expression of FN with Wnt7a dramatically stimulated the expansion of satellite stem cells in regenerating muscle. Therefore, activating satellite cells remodel their niche through autologous expression of FN that provides feedback to stimulate Wnt7a signaling through the Fzd7/Sdc4 co-receptor complex. Thus, FN and Wnt7a together regulate the homeostatic levels of satellite stem cells and satellite myogenic cells during regenerative myogenesis. PMID:23290138

  6. Pleiotropic virulence factor - Streptococcus pyogenes fibronectin-binding proteins.

    PubMed

    Yamaguchi, Masaya; Terao, Yutaka; Kawabata, Shigetada

    2013-04-01

    Streptococcus pyogenes causes a broad spectrum of infectious diseases, including pharyngitis, skin infections and invasive necrotizing fasciitis. The initial phase of infection involves colonization, followed by intimate contact with the host cells, thus promoting bacterial uptake by them. S. pyogenes recognizes fibronectin (Fn) through its own Fn-binding proteins to obtain access to epithelial and endothelial cells in host tissue. Fn-binding proteins bind to Fn to form a bridge to α5 β1 -integrins, which leads to rearrangement of cytoskeletal actin in host cells and uptake of invading S. pyogenes. Recently, several structural analyses of the invasion mechanism showed molecular interactions by which Fn converts from a compact plasma protein to a fibrillar component of the extracellular matrix. After colonization, S. pyogenes must evade the host innate immune system to spread into blood vessels and deeper organs. Some Fn-binding proteins contribute to evasion of host innate immunity, such as the complement system and phagocytosis. In addition, Fn-binding proteins have received focus as non-M protein vaccine candidates, because of their localization and conservation among different M serotypes.Here, we review the roles of Fn-binding proteins in the pathogenesis and speculate regarding possible vaccine antigen candidates. © 2012 Blackwell Publishing Ltd.

  7. Plasma fibronectin promotes thrombus growth and stability in injured arterioles

    PubMed Central

    Ni, Heyu; Yuen, Peter S. T.; Papalia, Jessie M.; Trevithick, Jane E.; Sakai, Takao; Fässler, Reinhard; Hynes, Richard O.; Wagner, Denisa D.

    2003-01-01

    Mice lacking both of the best-known platelet ligands, von Willebrand factor and fibrinogen, can still form occlusive thrombi in injured arterioles. The platelets of these animals accumulate excessive amounts of fibronectin (FN). These observations led us to examine the contribution of plasma FN (pFN) to thrombus formation. Inactivation of the FN gene in FN conditional knockout mice reduced pFN levels to <2% and platelet FN to ≈20% of the levels in similarly treated control mice. The mice were then observed in a model of arterial injury to evaluate their capacity to form thrombi. The deficiency of pFN did not affect the initial platelet adhesion, but a delay of several minutes in thrombus formation was observed in the arterioles of pFN-deficient mice as compared with control mice. The thrombi that formed in the absence of pFN were stably anchored to the vessel wall but continuously shed platelets or small platelet clumps, thus slowing their growth significantly; the platelet/platelet cohesion was apparently diminished. Consequently the occlusion of pFN-deficient vessels was delayed, with the majority of vessels remaining patent at the end of the 40-min observation period. We conclude that, in addition to von Willebrand factor and fibrinogen, FN plays a significant role in thrombus initiation, growth, and stability at arterial shear rates and that deficiency in each of the three platelet ligands has its own specific impact on platelet plug formation. PMID:12606706

  8. Thrombin Increases Expression of Fibronectin Antigen on the Platelet Surface

    NASA Astrophysics Data System (ADS)

    Ginsberg, Mark H.; Painter, Richard G.; Forsyth, Jane; Birdwell, Charles; Plow, Edward F.

    1980-02-01

    Fibronectins (fn) are adhesive glycoproteins which bind to collagen and to fibrin and appear to be important in cellular adhesion to other cells or surfaces. Fn-related antigen is present in human platelets, suggesting a possible role for fn in the adhesive properties of platelets. We have studied the localization of fn in resting and thrombin-stimulated platelets by immunofluorescence and quantitative binding of radiolabeled antibody. In resting fixed platelets, variable light surface staining for fn was observed. When these cells were made permeable to antibody with detergent, staining for fn was markedly enhanced and was present in a punctate distribution, suggesting intracellular localization. Stimulation with thrombin, which is associated with increased platelet adhesiveness, resulted in increased staining for fn antigen on intact platelets. These stimulated cells did not leak 51Cr nor did they stain for F-actin, thus documenting that the increased fn staining was not due to loss of plasma membrane integrity. The thrombin-induced increase in accessible platelet fn antigen was confirmed by quantitative antibody binding studies in which thrombin-stimulated platelets specifically bound 15 times as much radiolabeled F(ab')2 anti-fn as did resting cells. Thus, thrombin stimulation results in increased expression of fn antigen on the platelet surface. Here it may participate in interactions with fibrin, connective tissue, or other cells.

  9. Fibronectin-Grafted Titanium Dental Implants: An In Vivo Study

    PubMed Central

    Chang, Yu-Chi; Ho, Kuo-Ning; Feng, Sheng-Wei; Huang, Haw-Ming; Chang, Chia-Hsun; Lin, Che-Tong; Teng, Nai-Chia; Pan, Yu Hwa; Chang, Wei-Jen

    2016-01-01

    Modification of the physiochemical properties of titanium surfaces using glow discharge plasma (GDP) and fibronectin coating has been shown to enhance the surface hydrophilicity, surface roughness, cell adhesion, migration, and proliferation. This in vivo study aimed to evaluate the bone integration efficacy of a biologically modified implant surface. Two different surface-modified implants (Ar-GDP and GDP-fib) were placed in the mandibular premolar area of six beagle dogs for 2–8 weeks. Three techniques [histologic evaluation, resonance frequency analysis (RFA), and microcomputed tomography (micro-CT) evaluation] were used to detect the implant stability and bone-implant contact. The implant stability quotient values of GDP-fib implants were significantly greater than the Ar-GDP implants at 2 and 4 weeks (P < 0.01). The bone volume/total volume ratio of GDP-fib implants was greater than the Ar-GDP implants in micro-CT evaluation. A high positive correlation was observed between RFA and micro-CT measurements. At 2 weeks, osteoblasts were seen to line the implant surface, and multinuclear osteoclasts could be seen on the surface of old parent bone. After 8 weeks, a majority of the space in the wound chamber appeared to be replaced by bone. Enhancement of the stability of biologically modified implants was proved by the results of RFA, micro-CT, and histological analysis. This enhanced stability may help fasten treatment and be clinically beneficial. PMID:27366739

  10. EDA-containing fibronectin increases proliferation of embryonic stem cells.

    PubMed

    Losino, Noelia; Waisman, Ariel; Solari, Claudia; Luzzani, Carlos; Espinosa, Darío Fernández; Sassone, Alina; Muro, Andrés F; Miriuka, Santiago; Sevlever, Gustavo; Barañao, Lino; Guberman, Alejandra

    2013-01-01

    Embryonic stem cells (ESC) need a set of specific factors to be propagated. They can also grow in conditioned medium (CM) derived from a bovine granulosa cell line BGC (BGC-CM), a medium that not only preserves their main features but also increases ESC´s proliferation rate. The mitogenic properties of this medium were previously reported, ascribing this effect to an alternative spliced generated fibronectin isoform that contains the extra domain A (FN EDA(+)). Here, we investigated if the FN EDA(+) isoform increased proliferation of mouse and human ES cells. We analyzed cell proliferation using conditioned media produced by different mouse embryonic fibroblast (MEF) lines genetically engineered to express FN constitutively including or excluding the EDA domain (FN EDA(-)), and in media supplemented with recombinant peptides containing or not the EDA. We found that the presence of EDA in the medium increased mouse and human ESC's proliferation rate. Here we showed for the first time that this FN isoform enhances ESC's proliferation. These findings suggest a possible conserved behavior for regulation of ES cells proliferation by this FN isoform and could contribute to improve their culturing conditions both for research and cell therapy.

  11. EDA-Containing Fibronectin Increases Proliferation of Embryonic Stem Cells

    PubMed Central

    Losino, Noelia; Waisman, Ariel; Solari, Claudia; Luzzani, Carlos; Espinosa, Darío Fernández; Sassone, Alina; Muro, Andrés F.; Miriuka, Santiago; Sevlever, Gustavo; Barañao, Lino; Guberman, Alejandra

    2013-01-01

    Embryonic stem cells (ESC) need a set of specific factors to be propagated. They can also grow in conditioned medium (CM) derived from a bovine granulosa cell line BGC (BGC-CM), a medium that not only preserves their main features but also increases ESC´s proliferation rate. The mitogenic properties of this medium were previously reported, ascribing this effect to an alternative spliced generated fibronectin isoform that contains the extra domain A (FN EDA+). Here, we investigated if the FN EDA+ isoform increased proliferation of mouse and human ES cells. We analyzed cell proliferation using conditioned media produced by different mouse embryonic fibroblast (MEF) lines genetically engineered to express FN constitutively including or excluding the EDA domain (FN EDA-), and in media supplemented with recombinant peptides containing or not the EDA. We found that the presence of EDA in the medium increased mouse and human ESC’s proliferation rate. Here we showed for the first time that this FN isoform enhances ESC’s proliferation. These findings suggest a possible conserved behavior for regulation of ES cells proliferation by this FN isoform and could contribute to improve their culturing conditions both for research and cell therapy. PMID:24244705

  12. Focal hyperhidrosis: diagnosis and management

    PubMed Central

    Haider, Aamir; Solish, Nowell

    2005-01-01

    HYPERHIDROSIS, A CONDITION CHARACTERIZED by excessive sweating, can be generalized or focal. Generalized hyperhidrosis involves the entire body and is usually part of an underlying condition, most often an infectious, endocrine or neurologic disorder. Focal hyperhidrosis is idiopathic, occurring in otherwise healthy people. It affects 1 or more body areas, most often the palms, armpits, soles or face. Almost 3% of the general population, largely people aged between 25 and 64 years, experience hyperhidrosis. The condition carries a substantial psychological and social burden, since it interferes with daily activities. However, patients rarely seek a physician's help because many are unaware that they have a treatable medical disorder. Early detection and management of hyperhidrosis can significantly improve a patient's quality of life. There are various topical, systemic, surgical and nonsurgical treatments available with efficacy rates greater than 90%–95%. PMID:15632408

  13. Variable focal length deformable mirror

    DOEpatents

    Headley, Daniel; Ramsey, Marc; Schwarz, Jens

    2007-06-12

    A variable focal length deformable mirror has an inner ring and an outer ring that simply support and push axially on opposite sides of a mirror plate. The resulting variable clamping force deforms the mirror plate to provide a parabolic mirror shape. The rings are parallel planar sections of a single paraboloid and can provide an on-axis focus, if the rings are circular, or an off-axis focus, if the rings are elliptical. The focal length of the deformable mirror can be varied by changing the variable clamping force. The deformable mirror can generally be used in any application requiring the focusing or defocusing of light, including with both coherent and incoherent light sources.

  14. Stigmatic flat focal field spectrograph

    NASA Astrophysics Data System (ADS)

    Niemczyk, T. M.; Gobeli, G. W.

    Advances in two dimensional detectors have created a need for a spectrograph that can produce high fidelity images. Aberrations that detract from image quality in conventional spectrographs are discussed. Ray trace and experimental results obtained for a unique spectrograph designed for use with array detectors, i.e., designed to produce high fidelity images, are presented. The spectrograph employs toroidal mirrors to achieve stigmatic imaging in a flat focal field.

  15. Persistent Focal Behavior and Physical Activity Performance

    ERIC Educational Resources Information Center

    Erfle, Stephen E.

    2014-01-01

    This article examines the proclivity and performance attributes of focal students across time and activities using data from 9,345 students. Three systematic focal behavior partitions are examined: Across activities, across time, and across activities and time. A student's performance is focal if it ends in 0 or 5 for push-ups and 0 for curl-ups.…

  16. Persistent Focal Behavior and Physical Activity Performance

    ERIC Educational Resources Information Center

    Erfle, Stephen E.

    2014-01-01

    This article examines the proclivity and performance attributes of focal students across time and activities using data from 9,345 students. Three systematic focal behavior partitions are examined: Across activities, across time, and across activities and time. A student's performance is focal if it ends in 0 or 5 for push-ups and 0 for curl-ups.…

  17. F-actin-anchored focal adhesions distinguish endothelial phenotypes of human arteries and veins.

    PubMed

    van Geemen, Daphne; Smeets, Michel W J; van Stalborch, Anne-Marieke D; Woerdeman, Leonie A E; Daemen, Mat J A P; Hordijk, Peter L; Huveneers, Stephan

    2014-09-01

    Vascular endothelial-cadherin- and integrin-based cell adhesions are crucial for endothelial barrier function. Formation and disassembly of these adhesions controls endothelial remodeling during vascular repair, angiogenesis, and inflammation. In vitro studies indicate that vascular cytokines control adhesion through regulation of the actin cytoskeleton, but it remains unknown whether such regulation occurs in human vessels. We aimed to investigate regulation of the actin cytoskeleton and cell adhesions within the endothelium of human arteries and veins. We used an ex vivo protocol for immunofluorescence in human vessels, allowing detailed en face microscopy of endothelial monolayers. We compared arteries and veins of the umbilical cord and mesenteric, epigastric, and breast tissues and find that the presence of central F-actin fibers distinguishes the endothelial phenotype of adult arteries from veins. F-actin in endothelium of adult veins as well as in umbilical vasculature predominantly localizes cortically at the cell boundaries. By contrast, prominent endothelial F-actin fibers in adult arteries anchor mostly to focal adhesions containing integrin-binding proteins paxillin and focal adhesion kinase and follow the orientation of the extracellular matrix protein fibronectin. Other arterial F-actin fibers end in vascular endothelial-cadherin-based endothelial focal adherens junctions. In vitro adhesion experiments on compliant substrates demonstrate that formation of focal adhesions is strongly induced by extracellular matrix rigidity, irrespective of arterial or venous origin of endothelial cells. Our data show that F-actin-anchored focal adhesions distinguish endothelial phenotypes of human arteries from veins. We conclude that the biomechanical properties of the vascular extracellular matrix determine this endothelial characteristic. © 2014 American Heart Association, Inc.

  18. Extracellular matrix-specific focal adhesions in vascular smooth muscle produce mechanically active adhesion sites

    PubMed Central

    Sun, Zhe; Martinez-Lemus, Luis A.; Hill, Michael A.; Meininger, Gerald A.

    2008-01-01

    Integrin-mediated mechanotransduction in vascular smooth muscle cells (VSMCs) plays an important role in the physiological control of tissue blood flow and vascular resistance. To test whether force applied to specific extracellular matrix (ECM)-integrin interactions could induce myogenic-like mechanical activity at focal adhesion sites, we used atomic force microscopy (AFM) to apply controlled forces to specific ECM adhesion sites on arteriolar VSMCs. The tip of AFM probes were fused with a borosilicate bead (2∼5 μm) coated with fibronectin (FN), collagen type I (CNI), laminin (LN), or vitronectin (VN). ECM-coated beads induced clustering of α5- and β3-integrins and actin filaments at sites of bead-cell contact indicative of focal adhesion formation. Step increases of an upward (z-axis) pulling force (800∼1,600 pN) applied to the bead-cell contact site for FN-specific focal adhesions induced a myogenic-like, force-generating response from the VSMC, resulting in a counteracting downward pull by the cell. This micromechanical event was blocked by cytochalasin D but was enhanced by jasplakinolide. Function-blocking antibodies to α5β1- and αvβ3-integrins also blocked the micromechanical cell event in a concentration-dependent manner. Similar pulling experiments with CNI, VN, or LN failed to induce myogenic-like micromechanical events. Collectively, these results demonstrate that mechanical force applied to integrin-FN adhesion sites induces an actin-dependent, myogenic-like, micromechanical event. Focal adhesions formed by different ECM proteins exhibit different mechanical characteristics, and FN appears of particular relevance in its ability to strongly attach to VSMCs and to induce myogenic-like, force-generating reactions from sites of focal adhesion in response to externally applied forces. PMID:18495809

  19. Almost Unilateral Focal Dermal Hypoplasia

    PubMed Central

    Lee, Solam; Choe, Sung Jay

    2017-01-01

    Focal dermal hypoplasia, caused by mutations in PORCN, is an X-linked ectodermal dysplasia, also known as Goltz syndrome. Only seven cases of unilateral or almost unilateral focal dermal hypoplasia have been reported in the English literature and there have been no previously reported cases in the Republic of Korea. A 19-year-old female presented with scalp defects, skin lesions on the right leg and the right trunk, and syndactyly of the right fourth and fifth toes. Cutaneous examination revealed multiple atrophic plaques and a brown and yellow mass with fat herniation and telangiectasia that was mostly located on the lower right leg. She had syndactyly on the right foot and the scalp lesion appeared to be an atrophic, membranous, fibrotic alopecic scar. A biopsy of the calf revealed upper dermal extension of fat cells, dermal atrophy, and loss of dermal collagen. A diagnosis of almost unilateral focal dermal hypoplasia was made on the basis of physical and histologic findings. Henceforth, the patient was referred to a plastic surgeon and an orthopedics department to repair her syndactyly. PMID:28223754

  20. Plasma fibronectin synthesis in normal and injured humans as determined by stable isotope incorporation.

    PubMed Central

    Thompson, C; Blumenstock, F A; Saba, T M; Feustel, P J; Kaplan, J E; Fortune, J B; Hough, L; Gray, V

    1989-01-01

    In humans, plasma fibronectin decreases early after operative injury, burn, or trauma, followed by a rapid restoration with a secondary decline typically observed if such patients become septic. We determined the rate of plasma fibronectin and plasma fibrinogen synthesis in normal subjects and injured patients using a stable isotope incorporation technique with [15N]glycine. During a constant 14-h infusion of [15N]glycine, the enrichment of [15N]glycine in both the free plasma glycine precursor pool as well as the urinary hippurate pool was determined; the latter used as an estimate of intracellular hepatic precursor enrichment. [15N]Glycine enrichment in both plasma fibronectin and fibrinogen was also quantified. The synthesis rate (Js/V) expressed in micrograms per milliliter of plasma per hour and the fractional synthesis rate (FSR) expressed as percentage of the plasma pool produced per day were determined. In normal subjects, the FSR for plasma fibronectin using 15N enrichment into urinary hippurate was 35.35 +/- 1.46%/d, whereas the Js/V was 4.45 +/- 0.19 micrograms/ml plasma per h. In normal subjects, the FSR for plasma fibronectin using 15N enrichment into free plasma glycine was 14.73 +/- 0.63%/d, whereas the Js/V was 1.98 +/- 0.09 micrograms/ml plasma per h. Early (2-3 d) after burn injury, fibronectin synthesis was increased (Js/V = 5.74 +/- 0.36; P less than 0.05), whereas later after injury, fibronectin synthesis began to decline (Js/V = 3.52 +/- 0.24; P less than 0.05) based on 15N enrichment of urinary hippurate. In contrast, the Js/V and FSR of plasma fibrinogen, a well-documented acute-phase plasma protein, revealed a sustained elevation (P less than 0.05) after injury in both the trauma and burn patients. Thus, plasma fibronectin synthesis is elevated early postinjury, which may contribute to the rapid restoration of its blood level. However, once fibronectin levels have normalized, the synthesis of plasma fibronectin appears to decline. PMID

  1. Quantification of fibronectin as a method to assess ex vivo extracellular matrix remodeling.

    PubMed

    Bager, C L; Gudmann, N; Willumsen, N; Leeming, D J; Karsdal, M A; Bay-Jensen, A C; Høgdall, E; Balslev, I; He, Y

    2016-09-16

    Altered architecture, composition and quality of the extracellular matrix (ECM) are pathological hallmarks of several inflammatory and fibro-proliferative pathological processes such as osteoarthritis (OA), rheumatoid arthritis (RA), fibrosis and cancer. One of the most important components of the ECM is fibronectin. Fibronectin serves as an adhesion molecule anchoring cells to the underlying basement membrane through direct interaction with integrin receptors. Fibronectin hereby modulates the properties of the ECM and affects cellular processes. Quantification of fibronectin remodeling could therefore be used to assess the changes in the ECM that occur during progression of fibro-proliferative pathologies. Ex vivo models are becoming state-of-the-art tools to study ECM remodeling as the cellular composition and the organization of the ECM are preserved. Ex vivo models may therefore be a valuable tool to study the ECM remodeling that occurs during progression of fibro-proliferative pathologies. The aim of this study was to quantify fibronectin remodeling in ex vivo models of cartilage and cancer. A competitive The enzyme-linked immunosorbent assay (ELISA) against the C-terminus of fibronectin was developed (FBN-C). The assay was evaluated in relation to specificity, technical performance and as a marker for quantification of fibronectin in cartilage and cancer ex vivo models. The ELISA was specific and technically stable. Cleavage of tumor tissue with MMP-2 released significantly higher levels of FBN-C compared to tissue with buffer only and western blot analysis revealed that FBN-C recognizes both full length and degraded fibronectin. When ex vivo cartilage cultures were stimulated with the anabolic factor TGFβ and catabolic factors TNF-α and OSM, significantly higher levels of FBN-C were found in the conditioned media. Lastly, FBN-C was released from a cancer ex vivo model. In conclusion, we were able to quantify fibronectin remodeling in ex vivo models

  2. Interactions between integrin receptors and fibronectin are required for calvarial osteoblast differentiation in vitro

    NASA Technical Reports Server (NTRS)

    Moursi, A. M.; Globus, R. K.; Damsky, C. H.

    1997-01-01

    We previously showed that anti-fibronectin antibodies or soluble fibronectin fragments containing the central cell-binding domain inhibit formation of mineralized nodules by fetal calvarial osteoblasts in vitro. These findings suggest a critical role for fibronectin in osteoblast differentiation and morphogenesis. In this study we tested the hypothesis that fibronectin's effects on osteogenesis are mediated via direct interactions with integrin receptors for fibronectin on osteoblasts. Immunocytochemical analysis identified the integrin fibronectin receptor alpha5ss1 in fetal rat calvarial tissue and in cultured osteoblasts at all stages of differentiation. Three other integrins, alpha3ss1, alpha8ss1 and alphavss3, which can bind fibronectin, as well as other matrix components, were also identified in tissue and at all stages of cell culture. Immunoprecipitation data showed that alpha5ss1 levels are constant throughout osteoblast differentiation whereas levels of alpha3ss1 and alpha8ss1 decline in mature mineralized cultures. To determine whether integrin fibronectin receptors are required for osteoblast formation of mineralized nodules, we examined the extent of nodule formation in the presence and absence of function-perturbing anti-integrin antibodies. The antibodies were present continuously in cultures beginning at confluence (day 3), and nodule formation was measured at days 10 and 20. An anti-alpha5 integrin subunit antibody reduced nodule formation to less than 5% of control values at both time points. Inhibition of nodule formation was reversible and did not affect cell attachment and viability. Function-perturbing antibodies against alpha3ss1 and alpha8ss1 also reduced nodule formation, to less than 20% of control values. In contrast, function-perturbing antibodies to alphavss3 and alphavss5 did not affect nodule formation, indicating that the inhibitions noted were indeed specific. To determine the effect of antibody treatment on gene expression, steady

  3. Interactions between integrin receptors and fibronectin are required for calvarial osteoblast differentiation in vitro

    NASA Technical Reports Server (NTRS)

    Moursi, A. M.; Globus, R. K.; Damsky, C. H.

    1997-01-01

    We previously showed that anti-fibronectin antibodies or soluble fibronectin fragments containing the central cell-binding domain inhibit formation of mineralized nodules by fetal calvarial osteoblasts in vitro. These findings suggest a critical role for fibronectin in osteoblast differentiation and morphogenesis. In this study we tested the hypothesis that fibronectin's effects on osteogenesis are mediated via direct interactions with integrin receptors for fibronectin on osteoblasts. Immunocytochemical analysis identified the integrin fibronectin receptor alpha5ss1 in fetal rat calvarial tissue and in cultured osteoblasts at all stages of differentiation. Three other integrins, alpha3ss1, alpha8ss1 and alphavss3, which can bind fibronectin, as well as other matrix components, were also identified in tissue and at all stages of cell culture. Immunoprecipitation data showed that alpha5ss1 levels are constant throughout osteoblast differentiation whereas levels of alpha3ss1 and alpha8ss1 decline in mature mineralized cultures. To determine whether integrin fibronectin receptors are required for osteoblast formation of mineralized nodules, we examined the extent of nodule formation in the presence and absence of function-perturbing anti-integrin antibodies. The antibodies were present continuously in cultures beginning at confluence (day 3), and nodule formation was measured at days 10 and 20. An anti-alpha5 integrin subunit antibody reduced nodule formation to less than 5% of control values at both time points. Inhibition of nodule formation was reversible and did not affect cell attachment and viability. Function-perturbing antibodies against alpha3ss1 and alpha8ss1 also reduced nodule formation, to less than 20% of control values. In contrast, function-perturbing antibodies to alphavss3 and alphavss5 did not affect nodule formation, indicating that the inhibitions noted were indeed specific. To determine the effect of antibody treatment on gene expression, steady

  4. Trop-2 is up-regulated in invasive prostate cancer and displaces FAK from focal contacts

    PubMed Central

    Trerotola, Marco; Ganguly, Kirat K.; Fazli, Ladan; Fedele, Carmine; Lu, Huimin; Dutta, Anindita; Liu, Qin; De Angelis, Tiziana; Riddell, Luke W.; Riobo, Natalia A.; Gleave, Martin E.; Zoubeidi, Amina; Pestell, Richard G.; Altieri, Dario C.; Languino, Lucia R.

    2015-01-01

    In this study, we show that the transmembrane glycoprotein Trop-2 is up-regulated in human prostate cancer (PCa) with extracapsular extension (stages pT3/pT4) as compared to organ-confined (stage pT2) PCa. Consistent with this evidence, Trop-2 expression is found to be increased in metastatic prostate tumors of Transgenic Adenocarcinoma of Mouse Prostate mice and to strongly correlate with α5β1 integrin levels. Using PCa cells, we show that Trop-2 specifically associates with the α5 integrin subunit, as binding to α3 is not observed, and that Trop-2 displaces focal adhesion kinase from focal contacts. In support of the role of Trop-2 as a promoter of PCa metastatic phenotype, we observe high expression of this molecule in exosomes purified from Trop-2-positive PCa cells. These vesicles are then found to promote migration of Trop-2-negative PCa cells on fibronectin, an α5β1 integrin/focal adhesion kinase substrate, thus suggesting that the biological function of Trop-2 may be propagated to recipient cells. In summary, our findings show that Trop-2 promotes an α5β1 integrin-dependent pro-metastatic signaling pathway in PCa cells and that the altered expression of Trop-2 may be utilized for early identification of capsule-invading PCa. PMID:26015409

  5. SdrI of Staphylococcus saprophyticus is a multifunctional protein: localization of the fibronectin-binding site.

    PubMed

    Sakinç, Türkân; Kleine, Britta; Michalski, Nadine; Kaase, Martin; Gatermann, Sören G

    2009-11-01

    Staphylococcus saprophyticus, an important cause of urinary tract infections in young women, expresses the surface protein SdrI, a member of the serine-aspartate repeat (SD) protein family. Here we analyse the fibronectin-binding ability of SdrI, as S. saprophyticus is known to bind fibronectin and there is no known SD protein with this function. This protein does not contain the binding motif typical for fibronectin-binding proteins. Using recombinant fragments of SdrI, we localized the binding domain in the A region and show that SdrI bound to the N-terminal 30-kDa fragment of fibronectin. The fibronectin-binding function was shown in the natural host using an SdrI knockout mutant that showed decreased binding to fibronectin compared with wild-type strain 7108.

  6. Crystal structures of fibronectin-binding sites from Staphylococcus aureus FnBPA in complex with fibronectin domains

    PubMed Central

    Bingham, Richard J.; Rudiño-Piñera, Enrique; Meenan, Nicola A. G.; Schwarz-Linek, Ulrich; Turkenburg, Johan P.; Höök, Magnus; Garman, Elspeth F.; Potts, Jennifer R.

    2008-01-01

    Staphylococcus aureus can adhere to and invade endothelial cells by binding to the human protein fibronectin (Fn). FnBPA and FnBPB, cell wall-attached proteins from S. aureus, have multiple, intrinsically disordered, high-affinity binding repeats (FnBRs) for Fn. Here, 30 years after the first report of S. aureus/Fn interactions, we present four crystal structures that together comprise the structures of two complete FnBRs, each in complex with four of the N-terminal modules of Fn. Each ≈40-residue FnBR forms antiparallel strands along the triple-stranded β-sheets of four sequential F1 modules (2–5F1) with each FnBR/2–5F1 interface burying a total surface area of ≈4,300 Å2. The structures reveal the roles of residues conserved between S. aureus and Streptococcus pyogenes FnBRs and show that there are few linker residues between FnBRs. The ability to form large intermolecular interfaces with relatively few residues has been proposed to be a feature of disordered proteins, and S. aureus/Fn interactions provide an unusual illustration of this efficiency. PMID:18713862

  7. Large format focal plane array integration with precision alignment, metrology and accuracy capabilities

    NASA Astrophysics Data System (ADS)

    Neumann, Jay; Parlato, Russell; Tracy, Gregory; Randolph, Max

    2015-09-01

    Focal plane alignment for large format arrays and faster optical systems require enhanced precision methodology and stability over temperature. The increase in focal plane array size continues to drive the alignment capability. Depending on the optical system, the focal plane flatness of less than 25μm (.001") is required over transition temperatures from ambient to cooled operating temperatures. The focal plane flatness requirement must also be maintained in airborne or launch vibration environments. This paper addresses the challenge of the detector integration into the focal plane module and housing assemblies, the methodology to reduce error terms during integration and the evaluation of thermal effects. The driving factors influencing the alignment accuracy include: datum transfers, material effects over temperature, alignment stability over test, adjustment precision and traceability to NIST standard. The FPA module design and alignment methodology reduces the error terms by minimizing the measurement transfers to the housing. In the design, the proper material selection requires matched coefficient of expansion materials minimizes both the physical shift over temperature as well as lowering the stress induced into the detector. When required, the co-registration of focal planes and filters can achieve submicron relative positioning by applying precision equipment, interferometry and piezoelectric positioning stages. All measurements and characterizations maintain traceability to NIST standards. The metrology characterizes the equipment's accuracy, repeatability and precision of the measurements.

  8. Glycosylated fibronectin as a first-trimester biomarker for prediction of gestational diabetes.

    PubMed

    Rasanen, Juha P; Snyder, Caryn K; Rao, Paturi V; Mihalache, Raluca; Heinonen, Seppo; Gravett, Michael G; Roberts, Charles T; Nagalla, Srinivasa R

    2013-09-01

    To evaluate the potential clinical utility of serum biomarkers for first-trimester prediction of gestational diabetes mellitus (GDM). Maternal serum concentrations of glycosylated (Sambucus nigra lectin-reactive) fibronectin, adiponectin, sex hormone-binding globulin, placental lactogen, and high-sensitivity C-reactive protein (CRP) were measured at 5-13 weeks of gestation in a case-control study of 90 pregnant women with subsequent development of GDM and in 92 control group participants. Ability to detect GDM was assessed using logistic regression modeling and receiver operating characteristic (ROC) curves. Classification performance and positive and negative predictive values were reported at specific thresholds. Glycosylated fibronectin variation across trimesters was evaluated using a serial-measures analysis of 35 nondiabetic control group participants. First-trimester serum concentrations of glycosylated fibronectin, adiponectin, high-sensitivity CRP, and placental lactogen were significantly associated (P<.001) with GDM. After adjustment for maternal factors and other biomarkers, glycosylated fibronectin demonstrated an independent association with GDM (P<.001). Adiponectin, high-sensitivity CRP, and placental lactogen demonstrated modest classification performance compared with glycosylated fibronectin (respectively: area under the curve [AUC] 0.63; 95% confidence interval [CI] 0.53-0.71; AUC 0.68; 95% CI 0.60-0.76; and AUC 0.67, 95% CI 0.59-0.75; compared with AUC 0.91; 95% CI 0.87-0.96). Glycosylated fibronectin levels above a threshold of 120 mg/L correctly identified 57 GDM case group participants with a positive predictive value of 63% (95% CI 53-72%) and a negative predictive value of 95% (95% CI 94-95%) at a population prevalence of 12%. There was no association between sex hormone-binding globulin and GDM. First-trimester glycosylated fibronectin is a potential pregnancy-specific biomarker for early identification of women at risk for GDM. II.

  9. Intracellular signaling via ERK/MAPK completes the pathway for tubulogenic fibronectin in MDCK cells.

    PubMed

    Liu, Zhao; Greco, Andres J; Hellman, Nathan E; Spector, June; Robinson, Jonathan; Tang, Oliver T; Lipschutz, Joshua H

    2007-02-16

    A classic in vitro model of branching morphogenesis utilizes the Madin-Darby canine kidney (MDCK) cell line. MDCK Strain II cells form hollow monoclonal cysts in a three-dimensional collagen matrix over the course of 10 days and tubulate in response to hepatocyte growth factor (HGF). We and our colleagues previously showed that activation of the extracellular-signal regulated kinase (ERK, aka MAPK) pathway is necessary and sufficient to induce tubulogenesis in MDCK cells. We also showed in a microarray study that one of the genes upregulated by HGF was the known tubulogene fibronectin. Given that HGF activates a multitude of signaling pathways, including ERK/MAPK, to test the intracellular regulatory pathway, we used two distinct inhibitors of ERK activation (U0126 and PD098059). Following induction of MDCK Type II cells with HGF, tubulogenic fibronectin mRNA was upregulated fourfold by real-time PCR, and minimal or no change in fibronectin expression was seen when HGF was added with either U0126 or PD098059. We confirmed these results using an MDCK cell line inducible for Raf, which is upstream of ERK. Following activation of Raf, fibronectin mRNA and protein expression were increased to a similar degree as was seen following HGF induction. Furthermore, MDCK Strain I cells, which originate from collecting ducts and have constitutively active ERK, spontaneously initiate tubulogenesis. We show here that MDCK Strain I cells have high levels of fibronectin mRNA and protein compared to MDCK Strain II cells. When U0126 and PD098059 were added to MDCK Strain I cells, fibronectin mRNA, and protein levels were decreased to levels seen in MDCK Strain II cells. These data allow us to complete what we believe is the first description of a tubulogenic pathway from receptor/ligand (HGF/CMET), through an intracellular signaling pathway (ERK/MAPK), to transcription and, finally, secretion of a critical tubuloprotein (fibronectin).

  10. Intracellular Signaling via ERK/MAPK Completes the Pathway for Tubulogenic Fibronectin in MDCK Cells

    PubMed Central

    Liu, Zhao; Greco, Andres J.; Hellman, Nathan E.; Spector, June; Robinson, Jonathan; Tang, Oliver T.; Lipschutz, Joshua H.

    2007-01-01

    A classic in vitro model of branching morphogenesis utilizes the Madin-Darby canine kidney (MDCK) cell line. MDCK Strain II cells form hollow monoclonal cysts in a three-dimensional collagen matrix over the course of ten days and tubulate in response to hepatocyte growth factor (HGF). We and our colleagues previously showed that activation of the extracellular-signal regulated kinase (ERK, aka MAPK) pathway is necessary and sufficient to induce tubulogenesis in MDCK cells. We also showed in a microarray study that one of the genes upregulated by HGF was the known tubulogene fibronectin. Given that HGF activates a multitude of signaling pathways, including ERK/MAPK, to test the intracellular regulatory pathway, we used two distinct inhibitors of ERK activation (U0126 and PD098059). Following induction of MDCK Type II cells with HGF, tubulogenic fibronectin mRNA was upregulated 4-fold by real time PCR, and minimal or no change in fibronectin expression was seen when HGF was added with either U0126 or PD098059. We confirmed these results using an MDCK cell line inducible for Raf, which is upstream of ERK. Following activation of Raf, fibronectin mRNA and protein expression were increased to a similar degree as was seen following HGF induction. Furthermore, MDCK Strain I cells, which originate from collecting ducts and have constitutively active ERK, spontaneously initiate tubulogenesis. We show here that MDCK Strain I cells have high levels of fibronectin mRNA and protein compared to MDCK Strain II cells. When U0126 and PD098059 were added to MDCK Strain I cells, fibronectin mRNA and protein levels were decreased to levels seen in MDCK Strain II cells. These data allow us to complete what we believe is the first description of a tubulogenic pathway from receptor/ligand (HGF/CMET), through an intracellular signaling pathway (ERK/MAPK), to transcription and, finally, secretion of a critical tubuloprotein (fibronectin). PMID:17196167

  11. Anti-Idiotypic Monobodies Derived from a Fibronectin Scaffold

    PubMed Central

    Sullivan, Mark A.; Brooks, Lauren R.; Weidenborner, Philip; Domm, William; Mattiacio, Jonelle; Xu, Qingfu; Tiberio, Michael; Wentworth, Timothy; Kobie, James; Bryk, Peter; Zheng, Bo; Murphy, Mary; Sanz, Ignacio; Dewhurst, Stephen

    2014-01-01

    Mimetics of conformational protein epitopes have broad applications, but have been difficult to identify using conventional peptide phage display. The tenth type III domain of human fibronectin (FNfn10) has two extended, randomizable surface-exposed loops and might be more amenable to the identification of such mimetics. We therefore selected a library of FNfn10 clones, randomized in both loops (15 residues in all), for binding to monoclonal antibodies (MAb) that recognize the HIV-1 envelope glycoprotein. Anti-idiotypic monobodies (αIMs) mimicking both “linear” epitopes (2F5 and 4E10 MAbs) and conformational epitopes (b12 and VRC01 MAbs) were generated. αIMs selected against 2F5 and 4E10 frequently displayed sequence homology to the corresponding linear native epitopes. In the case of b12 and VRC01, we expected that the two constrained loop domains of FNfn10 would both contribute to complex conformational interactions with target antibodies. However, mutagenesis studies revealed differences from this simple model An αIM selected against b12 was found to bind its cognate antibody via only a few residues within the BC loop of FNfn10, with minimal contribution from the FG loop. Unexpectedly, this was sufficient to generate a protein that engaged its cognate antibody in a manner very similar to HIV-1 Env, and with a strong KD (43 nM). In contrast, an αIM selected against VRC01 engaged its cognate antibody in a manner that was dependent on both BC and FG loop sequences. Overall, these data suggest that the FNfn10 scaffold can be used to identify complex structures that mimic conformational protein epitopes. PMID:23394681

  12. Stromelysin generates a fibronectin fragment that inhibits Schwann cell proliferation

    PubMed Central

    1992-01-01

    Our previous report (Muir, D., S. Varon, and M. Manthorpe. 1990. J. Cell Biol. 109:2663-2672) described the isolation and partial characterization of a 55-kD antiproliferative protein found in Schwann cell (SC) and schwannoma cell line-conditioned media and we concluded that SC proliferation is under negative autocrine control. In the present study the 55-kD protein was found to possess metalloprotease activity and stromelysin immunoreactivity. The SC-derived metalloprotease shares many properties with stromelysin isolated from other sources including the ability to cleave fibronectin (FN). Furthermore, limited proteolysis of FN by the SC-derived protease generated a FN fragment which itself expresses a potent antiproliferative activity for SCs. The active FN fragment corresponds to the 29-kD amino-terminal region of the FN molecule which was also identified as an active component in SC CM. Additional evidence that a proteolytic fragment of FN can possess antiproliferative activity for SCs was provided by the finding that plasmin can generate an amino- terminal FN fragment which mimicked the activity of the SC metalloprotease-generated antiproliferative FN fragment. Both the 55-kD SC metalloprotease and the 29-kD FN fragment could completely and reversibly inhibit proliferation of SCs treated with various mitogens and both were largely ineffective at inhibiting proliferation by immortalized or transformed SC lines. Normal and transformed SC types do secrete the proform of stromelysin, however, transformed cultures do not produce activated stromelysin and thus cannot generate the antiproliferative fragment of FN. These results suggest that, once activated, a SC-derived protease similar to stromelysin cleaves FN and generates an antiproliferative activity which can maintain normal SC quiescence in vitro. PMID:1730742

  13. Heparin-dependent regulation of fibronectin matrix conformation

    PubMed Central

    Hubbard, Brant; Buczek-Thomas, Jo Ann; Nugent, Matthew A.; Smith, Michael L.

    2013-01-01

    Extracellular matrix (ECM) conformation is regulated by a variety of stimuli in vivo, including mechanical forces and allosteric binding partners, and these conformational changes contribute to the regulation of cell behavior. Heparin and heparan sulfate, for example, have been shown to regulate the sequestration and presentation of numerous growth factors, including vascular endothelial growth factor, on the heparin 2 binding domain in fibronectin (Fn). However, mechanical force also alters Fn conformation, indicating that the growth factor binding region may be co-regulated by both heparin and mechanical force. Herein, we describe a simple antibody-based method for evaluating the conformation of the heparin 2 binding domain in Fn, and use it to determine the relative contributions of heparin and mechanical strain to the regulation of Fn conformation. We achieved specificity in quantifying conformational changes in this region of Fn by measuring the ratio of two fluorescent monoclonal antibodies, one that is insensitive to Fn conformational changes and a second whose binding is reduced or enhanced by non-equilibrium conformational changes. Importantly, this technique is shown to work on Fn adsorbed on surfaces, single Fn fibers, and Fn matrix fibers in cell culture. Using our dual antibody approach, we show that heparin and mechanical strain co-regulate Fn conformation in matrix fibrils, which is the first demonstration of heparin-dependent regulation of Fn in its physiologically-relevant fibrillar state. Furthermore, the dual antibody approach utilizes commercially available antibodies and simple immunohistochemistry, thus making it accessible to a wide range of scientists interested in Fn mechanobiology. PMID:24148804

  14. Mutations in FN1 cause glomerulopathy with fibronectin deposits.

    PubMed

    Castelletti, Federica; Donadelli, Roberta; Banterla, Federica; Hildebrandt, Friedhelm; Zipfel, Peter F; Bresin, Elena; Otto, Edgar; Skerka, Christine; Renieri, Alessandra; Todeschini, Marta; Caprioli, Jessica; Caruso, Rosa Maria; Artuso, Rosangela; Remuzzi, Giuseppe; Noris, Marina

    2008-02-19

    Glomerulopathy with fibronectin (FN) deposits (GFND) is an autosomal dominant disease with age-related penetrance, characterized by proteinuria, microscopic hematuria, hypertension, and massive glomerular deposits of FN that lead to end-stage renal failure. The genetic abnormality underlying GFND was still unknown. We hypothesized that mutations in FN1, which encodes FN, were the cause of GFND. In a large Italian pedigree with eight affected subjects, we found linkage with GFND at the FN1 locus at 2q32. We sequenced the FN1 in 15 unrelated pedigrees and found three heterozygous missense mutations, the W1925R, L1974R, and Y973C, that cosegregated with the disease in six pedigrees. The mutations affected two domains of FN (Hep-II domain for the W1925R and the L1974R, and Hep-III domain for the Y973C) that play key roles in FN-cell interaction and in FN fibrillogenesis. Mutant recombinant Hep-II fragments were expressed, and functional studies revealed a lower binding to heparin and to endothelial cells and podocytes compared with wild-type Hep-II and an impaired capability to induce endothelial cell spreading and cytoskeletal reorganization. Overall dominant mutations in FN1 accounted for 40% of cases of GFND in our study group. These findings may help understanding the pathogenesis of proteinuria and glomerular FN deposits in GFND and possibly in more common renal diseases such as diabetic nephropathy, IgA nephropathy, and lupus nephritis. To our knowledge no FN1 mutation causing a human disease was previously reported.

  15. In vitro selection of fibronectin gain-of-function mutations.

    PubMed Central

    Tani, Patricia H; Loftus, Joseph C; Bowditch, Ron D

    2002-01-01

    Directed protein evolution, which employs a combination of random mutagenesis, phage display, and in vitro selection, was used to identify second-site suppressors of the fibronectin (Fn) cell binding domain mutation Asp1495Ala (RGA). The mutations in the Fn 9th (3fn9) and 10th (3fn10) type III repeats obtained after selection on purified integrins alphaIIbbeta3(D119Y) and alpha5beta1 are reported. The 3fn9-10(D1495A) phage with substitution mutations at Asp1418, which is located within the linker region between 3fn9 and 3fn10, enhanced binding to the integrins alphaIIbbeta3 and alpha5beta1, but not alphavbeta3. The substitution mutations identified at residue Asp1418 were introduced into the native recombinant 3fn9-10 sequence and found to augment binding to alphaIIbbeta3, demonstrating that the observed gain-of-function phenotype was independent of the multivalent character of the phage. These results support the following conclusions. First, regions of Fn in addition to the RGD loop are in close proximity to alphaIIbbeta3 and alpha5beta1 and are capable of participating in the binding to these integrins. Secondly, the conformational relationship between the 3fn9 and 3fn10 modules may be an important factor in the binding of Fn to these two integrins. Thirdly, other altered properties of Fn-integrin interactions, such as integrin specificity, may also be selected. This is the first description of Fn mutations that augment binding to integrins. The ability to select for particular phenotypes in vitro and the subsequent characterization of these mutations should further our understanding of the molecular details involved in the association of integrins and their ligands. Additionally, these higher-affinity 3fn9-10 ligands provide a starting point for further in vitro evolution and engineering of integrin-specific modules. PMID:12071856

  16. Anti-idiotypic monobodies derived from a fibronectin scaffold.

    PubMed

    Sullivan, Mark A; Brooks, Lauren R; Weidenborner, Philip; Domm, William; Mattiacio, Jonelle; Xu, Qingfu; Tiberio, Michael; Wentworth, Timothy; Kobie, James; Bryk, Peter; Zheng, Bo; Murphy, Mary; Sanz, Ignacio; Dewhurst, Stephen

    2013-03-12

    Mimetics of conformational protein epitopes have broad applications but have been difficult to identify using conventional peptide phage display. The 10th type III domain of human fibronectin (FNfn10) has two extended, randomizable surface-exposed loops and might be more amenable to the identification of such mimetics. We therefore selected a library of FNfn10 clones, randomized in both loops (15 residues in all), for binding to monoclonal antibodies (mAbs) that recognize the HIV-1 envelope glycoprotein. Anti-idiotypic monobodies (αIMs) mimicking both "linear" epitopes (2F5 and 4E10 mAbs) and conformational epitopes (b12 and VRC01 mAbs) were generated. αIMs selected against 2F5 and 4E10 frequently displayed sequence homology to the corresponding linear native epitopes. In the case of b12 and VRC01, we expected that the two constrained loop domains of FNfn10 would both contribute to complex conformational interactions with target antibodies. However, mutagenesis studies revealed differences from this simple model. An αIM selected against b12 was found to bind its cognate antibody via only a few residues within the BC loop of FNfn10, with minimal contribution from the FG loop. Unexpectedly, this was sufficient to generate a protein that engaged its cognate antibody in a manner very similar to that of HIV-1 Env, and with a strong KD (43 nM). In contrast, an αIM selected against VRC01 engaged its cognate antibody in a manner that was dependent on both BC and FG loop sequences. Overall, these data suggest that the FNfn10 scaffold can be used to identify complex structures that mimic conformational protein epitopes.

  17. Human macrophage differentiation involves an interaction between integrins and fibronectin

    SciTech Connect

    Laouar, A.; Chubb, C.B.H.; Collart, F.; Huberman, E.

    1996-11-15

    The authors have examined the role of the {beta}{sub 1} integrin family of adhesion receptors (VLA) and the extracellular matrix protein fibronectin (FN) in macrophage differentiation of (1) human HL-60 myeloid leukemia cells induced by phorbol 12-myristate 13-acetate (PMA) and (2) human peripheral blood monocytes induced by either PMA or macrophage-colony stimulating factor (M=CSF). Increased VLA and FN gene expression was observed as early as 4 h after PMA treatment of HL-60 cells and PMA- or M-CSF-treatment of monocytes, and it preceded the manifestation of macrophage markers. Treated HL-60 cells and monocytes also released and deposited FN on the surface of the tissue culture dishes. An HL-60 cell variant, HL-525, which is deficient in protein kinase C {beta} and resistant to PMA-induced differentiation, exhibited elevated levels of the VLA antigen but failed to express the FN gene. Incubation of HL-525 cells on dishes precoated with exogenous FN resulted in a macrophage differentiation. The macrophage phenotype induced in HL-60 cells, HL-525 cells, or monocytes was attenuated to various degrees by anti-VLA or anti-FN MAbs or by exogenous RGDS, a VLA-binding motif on FN. The authors suggest that macrophage differentiation is initiated by the activation of protein kinase C, which leads to the expression of the integrin, FN and related genes. The integrins mediate cell attachment and spreading on appropriate substrates by binding to deposited extracellular proteins such as FN. This attachment and spreading, in turn, leads to the expression of genes that code for the macrophage functions.

  18. Origin of tumor-promoter released fibronectin in fibroblasts

    SciTech Connect

    Burrous, B.A.; Wolf, G.

    1986-05-01

    Previous work from the laboratory showed that the chemical tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) stimulated release of the cell surface glycoprotein, fibronectin (FN) from human lung fibroblasts (HLF), leading to depletion of cell surface FN, while FN synthesis is not altered by TPA. To further investigate the mechanism(s) by which TPA stimulates FN release, two types of experiments were performed. In the first, HLF were pulsed with /sup 35/S-methionine-labeled medium with or without TPA. In the second, cell-surface proteins were labeled by iodination (/sup 125/I) and then incubated in unlabeled medium with or without TPA. In both cases, the fate of labeled FN was followed over 12 hr. The /sup 35/S-meth-labeled HLF showed a rapid loss of labeled FN, first into a small, highly-labeled pool of cell surface FN (1 hr), later into the medium (4 hr or longer). Specific activities showed that this small pool in the cell surface turned over rapidly. TPA treatment resulted in more rapid movement of /sup 35/S-meth pulse-labeled FN to the cell surface and into the medium than in control cells. TPA thus affected the fate of intracellular FN. TPA treatment of HLF also resulted in more rapid removal of /sup 125/I-cell surface-labeled FN into the medium than in control cells. Thus, TPA affects the fate of preexisting cell surface FN in HLF. From these results, they hypothesize that TPA has two separate effects: it stimulates depletion of preexisting intracellular FN during the first hr of treatment, and it stimulates release of preexisting cell surface FN over all treatment times.

  19. Cooperative effects of fibronectin matrix assembly and initial cell-substrate adhesion strength in cellular self-assembly.

    PubMed

    Brennan, James R; Hocking, Denise C

    2016-03-01

    The cell-dependent polymerization of intercellular fibronectin fibrils can stimulate cells to self-assemble into multicellular structures. The local physical cues that support fibronectin-mediated cellular self-assembly are largely unknown. Here, fibronectin matrix analogs were used as synthetic adhesive substrates to model cell-matrix fibronectin fibrils having different integrin-binding specificity, affinity, and/or density. We utilized this model to quantitatively assess the relationship between adhesive forces derived from cell-substrate interactions and the ability of fibronectin fibril assembly to induce cellular self-assembly. Results indicate that the strength of initial, rather than mature, cell-substrate attachments correlates with the ability of substrates to support fibronectin-mediated cellular self-assembly. The cellular response to soluble fibronectin was bimodal and independent of the integrin-binding specificity of the substrate; increasing soluble fibronectin levels above a critical threshold increased aggregate cohesion on permissive substrates. Once aggregates formed, continuous fibronectin polymerization was necessary to maintain cohesion. During self-assembly, soluble fibronectin decreased cell-substrate adhesion strength and induced aggregate cohesion via a Rho-dependent mechanism, suggesting that the balance of contractile forces derived from fibronectin fibrils within cell-cell versus cell-substrate adhesions controls self-assembly and aggregate cohesion. Thus, initial cell-substrate attachment strength may provide a quantitative basis with which to build predictive models of fibronectin-mediated microtissue fabrication on a variety of substrates. Cellular self-assembly is a process by which cells and extracellular matrix (ECM) proteins spontaneously organize into three-dimensional (3D) tissues in the absence of external forces. Cellular self-assembly can be initiated in vitro, and represents a potential tool for tissue engineers to

  20. The predictive value of quantitative fibronectin testing in combination with cervical length measurement in symptomatic women.

    PubMed

    Bruijn, Merel M C; Kamphuis, Esme I; Hoesli, Irene M; Martinez de Tejada, Begoña; Loccufier, Anne R; Kühnert, Maritta; Helmer, Hanns; Franz, Marie; Porath, Martina M; Oudijk, Martijn A; Jacquemyn, Yves; Schulzke, Sven M; Vetter, Grit; Hoste, Griet; Vis, Jolande Y; Kok, Marjolein; Mol, Ben W J; van Baaren, Gert-Jan

    2016-12-01

    The combination of the qualitative fetal fibronectin test and cervical length measurement has a high negative predictive value for preterm birth within 7 days; however, positive prediction is poor. A new bedside quantitative fetal fibronectin test showed potential additional value over the conventional qualitative test, but there is limited evidence on the combination with cervical length measurement. The purpose of this study was to compare quantitative fetal fibronectin and qualitative fetal fibronectin testing in the prediction of spontaneous preterm birth within 7 days in symptomatic women who undergo cervical length measurement. We performed a European multicenter cohort study in 10 perinatal centers in 5 countries. Women between 24 and 34 weeks of gestation with signs of active labor and intact membranes underwent quantitative fibronectin testing and cervical length measurement. We assessed the risk of preterm birth within 7 days in predefined strata based on fibronectin concentration and cervical length. Of 455 women who were included in the study, 48 women (11%) delivered within 7 days. A combination of cervical length and qualitative fibronectin resulted in the identification of 246 women who were at low risk: 164 women with a cervix between 15 and 30 mm and a negative fibronectin test (<50 ng/mL; preterm birth rate, 2%) and 82 women with a cervix at >30 mm (preterm birth rate, 2%). Use of quantitative fibronectin alone resulted in a predicted risk of preterm birth within 7 days that ranged from 2% in the group with the lowest fibronectin level (<10 ng/mL) to 38% in the group with the highest fibronectin level (>500 ng/mL), with similar accuracy as that of the combination of cervical length and qualitative fibronectin. Combining cervical length and quantitative fibronectin resulted in the identification of an additional 19 women at low risk (preterm birth rate, 5%), using a threshold of 10 ng/mL in women with a cervix at <15 mm, and 6 women at high risk

  1. Effect of fibronectin- and collagen I-coated titanium fiber mesh on proliferation and differentiation of osteogenic cells.

    PubMed

    van den Dolder, Juliette; Bancroft, Gregory N; Sikavitsas, Vassilios I; Spauwen, Paul H M; Mikos, Antonios G; Jansen, John A

    2003-06-01

    The objective of this study was to evaluate the effects of fibronectin and collagen I coatings on titanium fiber mesh on the proliferation and osteogenic differentiation of rat bone marrow cells. Three main treatment groups were investigated in addition to uncoated titanium fiber meshes: meshes coated with fibronectin, meshes coated with collagen I, and meshes coated first with collagen I and then subsequently with fibronectin. Rat bone marrow cells were cultured for 1, 4, 8, and 16 days in plain and coated titanium fiber meshes. In addition, a portion of each of these coating treatment groups was cultured in the presence of antibodies against fibronectin and collagen I integrins. To evaluate cellular proliferation and differentiation, constructs were examined for DNA, osteocalcin, and calcium content and alkaline phosphatase activity. There were no significant effects of the coatings on cellular proliferation as indicated by the DNA quantification analysis. When antibodies against fibronectin and collagen I integrins were used, a significant reduction (p < 0.05) in cell proliferation was observed for the uncoated titanium meshes, meshes coated with collagen, and meshes coated with collagen and fibronectin. The different coatings also did not affect the alkaline phosphatase activity of the cells seeded on the coated meshes. However, the presence of antibodies against fibronectin or collagen I integrins resulted in significantly delayed expression of alkaline phosphatase activity for uncoated titanium meshes, meshes coated with collagen, and meshes coated with collagen and fibronectin. Calcium measurements did not reveal a significant effect of fibronectin or collagen I coating on calcium deposition in the meshes. Also, no difference in calcium content was observed in the uncoated titanium meshes and meshes coated with fibronectin when antibodies against fibronectin or collagen I integrins were present. Meshes coated with both collagen I and fibronectin showed

  2. Definition of the native and denatured type II collagen binding site for fibronectin using a recombinant collagen system.

    PubMed

    An, Bo; Abbonante, Vittorio; Yigit, Sezin; Balduini, Alessandra; Kaplan, David L; Brodsky, Barbara

    2014-02-21

    Interaction of collagen with fibronectin is important for extracellular matrix assembly and regulation of cellular processes. A fibronectin-binding region in collagen was identified using unfolded fragments, but it is not clear if the native protein binds fibronectin with the same primary sequence. A recombinant bacterial collagen is utilized to characterize the sequence requirement for fibronectin binding. Chimeric collagens were generated by inserting the putative fibronectin-binding region from human collagen into the bacterial collagen sequence. Insertion of a sufficient length of human sequence conferred fibronectin affinity. The minimum sequence requirement was identified as a 6-triplet sequence near the unique collagenase cleavage site and was the same in both triple-helix and denatured states. Denaturation of the chimeric collagen increased its affinity for fibronectin, as seen for mammalian collagens. The fibronectin binding recombinant collagen did not contain hydroxyproline, indicating hydroxyproline is not essential for binding. However, its absence may account, in part, for the higher affinity of the native chimeric protein and the lower affinity of the denatured protein compared with type II collagen. Megakaryocytes cultured on chimeric collagen with fibronectin affinity showed improved adhesion and differentiation, suggesting a strategy for generating bioactive materials in biomedical applications.

  3. Modulation of endothelial cell adhesion to synthetic vascular grafts using biotinylated fibronectin in a dual ligand protein system

    NASA Astrophysics Data System (ADS)

    Anamelechi, Charles Chibuzor

    Over half a million coronary artery bypass operations are performed annually in the US yielding an annual health care cost of over 16 billion dollars. Only five percent of bypasses are repeat operations in spite of the procedures prevalence. Patients facing repeat coronary artery bypass operations often lack transplantable autologous arteries or veins, necessitating the use of substitutes. Unfortunately, synthetic small diameter vascular grafts have unacceptable patency rates, primarily due to lumenal thrombus formation and intimal thickening. Endothelial cells (EC) mediate the anti-thrombotic activity in healthy blood vessels, and due to the scarcity of suitable autologous vascular replacement, EC-seeded small diameter synthetic vascular grafts represent a clear, immediate, and practical solution. The fundamental goal of this project was to optimize the dual ligand (DL) system on synthetic vascular graft (SVG) surrogates to show enhanced cell adhesion, retention, and native functionality compared to fibronectin alone. Initially, two SVG surrogates were identified through characterization by x-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM), and 125I radiolabeling. The first modification to the DL system involved direct biotinylation of fibronectin (bFN) as a replacement for co-adsorption of FN with biotinylated bovine serum albumin (bBSA). This was analyzed with a Langmuir model using surface plasmon resonance (SPR) spectroscopy to verify the binding affinity of bFN and ELISA to detect the availability of the RGD binding motif post biotinylation. The second major change in this project examined cell binding and formation of focal adhesion after shifting from direct incubation of HUVECs with RGD-SA to sequentially adsorbing bFN(9) and RGD-SA prior to introducing unmodified HUVECs. These experiments were conducted under static seeding conditions. Next, dynamic cell seeding onto the sequentially adsorbed protein surface was examined as a function

  4. Research of aerial camera focal pane micro-displacement measurement system based on Michelson interferometer

    NASA Astrophysics Data System (ADS)

    Wang, Shu-juan; Zhao, Yu-liang; Li, Shu-jun

    2014-09-01

    The aerial camera focal plane in the correct position is critical to the imaging quality. In order to adjust the aerial camera focal plane displacement caused in the process of maintenance, a new micro-displacement measuring system of aerial camera focal plane in view of the Michelson interferometer has been designed in this paper, which is based on the phase modulation principle, and uses the interference effect to realize the focal plane of the micro-displacement measurement. The system takes He-Ne laser as the light source, uses the Michelson interference mechanism to produce interference fringes, changes with the motion of the aerial camera focal plane interference fringes periodically, and records the periodicity of the change of the interference fringes to obtain the aerial camera plane displacement; Taking linear CCD and its driving system as the interference fringes picking up tool, relying on the frequency conversion and differentiating system, the system determines the moving direction of the focal plane. After data collecting, filtering, amplifying, threshold comparing, counting, CCD video signals of the interference fringes are sent into the computer processed automatically, and output the focal plane micro displacement results. As a result, the focal plane micro displacement can be measured automatically by this system. This system uses linear CCD as the interference fringes picking up tool, greatly improving the counting accuracy and eliminated the artificial counting error almost, improving the measurement accuracy of the system. The results of the experiments demonstrate that: the aerial camera focal plane displacement measurement accuracy is 0.2nm. While tests in the laboratory and flight show that aerial camera focal plane positioning is accurate and can satisfy the requirement of the aerial camera imaging.

  5. Interictal spikes in focal epileptogenesis.

    PubMed

    de Curtis, M; Avanzini, G

    2001-04-01

    Interictal electroencephalography (EEG) potentials in focal epilepsies are sustained by synchronous paroxysmal membrane depolarization generated by assemblies of hyperexcitable neurons. It is currently believed that interictal spiking sets a condition that preludes to the onset of an ictal discharge. Such an assumption is based on little experimental evidence. Human pre-surgical studies and recordings in chronic and acute models of focal epilepsy showed that: (i) interictal spikes (IS) and ictal discharges are generated by different populations of neuron through different cellular and network mechanisms; (ii) the cortical region that generates IS (irritative area) does not coincide with the ictal-onset area; (iii) IS frequency does not increase before a seizure and is enhanced just after an ictal event; (iv) spike suppression is found to herald ictal discharges; and (v) enhancement of interictal spiking suppresses ictal events. Several experimental evidences indicate that the highly synchronous cellular discharge associated with an IS is generated by a multitude of mechanisms involving synaptic and non-synaptic communication between neurons. The synchronized neuronal discharge associated with a single IS induces and is followed by a profound and prolonged refractory period sustained by inhibitory potentials and by activity-dependent changes in the ionic composition of the extracellular space. Post-spike depression may be responsible for pacing interictal spiking periodicity commonly observed in both animal models and human focal epilepsies. It is proposed that the strong after-inhibition produced by IS protects against the occurrence of ictal discharges by maintaining a low level of excitation in a general condition of hyperexcitability determined by the primary epileptogenic dysfunction.

  6. Streptococcus mutans and Streptococcus intermedius adhesion to fibronectin films are oppositely influenced by ionic strength.

    PubMed

    Busscher, Henk J; van de Belt-Gritter, Betsy; Dijkstra, Rene J B; Norde, Willem; van der Mei, Henny C

    2008-10-07

    Bacterial adhesion to protein-coated surfaces is mediated by an interplay of specific and nonspecific interactions. Although nonspecific interactions are ubiquitously present, little is known about the physicochemical mechanisms of specific interactions. The aim of this paper is to determine the influence of ionic strength on the adhesion of two streptococcal strains to fibronectin films. Streptococcus mutans LT11 and Streptococcus intermedius NCTC11324 both possess antigen I/II with the ability to bind fibronectin from solution, but S. intermedius binds approximately 20x less fibronectin than does the S. mutans strain under identical conditions. Both strains as well as fibronectin films are negatively charged in low ionic strength phosphate buffered saline (PBS, 10x diluted), but bacteria appear uncharged in high ionic strength PBS. Physicochemical modeling on the basis of overall cell surface properties (cell surface hydrophobicity and zeta potentials) demonstrates that both strains should favor adhesion to fibronectin films in a high ionic strength environment as compared to in a low ionic strength environment, where electrostatic repulsion between equally charged surfaces is dominant. Adhesion of S. intermedius to fibronectin films in a parallel plate flow chamber was completely in line with this modeling, while in addition atomic force microscopy (AFM) indicated stronger adhesion forces upon retraction between fibronectin-coated tips and the cell surfaces in high ionic strength PBS than in low ionic strength PBS. Thus, the dependence of the interaction on ionic strength is dominated by the overall negative charge on the interacting surfaces. Adhesion of S. mutans to fibronectin films, however, was completely at odds with theoretical modeling, and the strain adhered best in low ionic strength PBS. Moreover, AFM indicated weaker repulsive forces upon approach between fibronectin-coated tips and the cell surfaces in low ionic strength PBS than in high ionic

  7. Fetal fibronectin as a predictor of preterm birth in patients with symptoms: a multicenter trial.

    PubMed

    Peaceman, A M; Andrews, W W; Thorp, J M; Cliver, S P; Lukes, A; Iams, J D; Coultrip, L; Eriksen, N; Holbrook, R H; Elliott, J; Ingardia, C; Pietrantoni, M

    1997-07-01

    Our aim was to determine whether the presence of fetal fibronectin in vaginal secretions of patients with symptoms suggestive of preterm labor predicts preterm delivery. Patients who were examined at the hospital between 24 weeks' and 34 weeks 6 days' gestation with intact membranes, no prior tocolysis, symptoms suggestive of preterm labor, and cervical dilation < 3 cm were recruited at 10 sites. Swabs of the posterior fornix were assayed for the presence of fetal fibronectin by monoclonal antibody assay, with a positive result defined as > or = 50 ng/ml. Results were not available to the managing physicians. Tocolysis was used when clinically indicated after specimen collection. A total of 763 patients had fetal fibronectin results and pregnancy outcome data available for analysis. Fetal fibronectin was detected in specimens from 150 (20%) patients. Compared with patients who had negative results, patients who had positive results for fetal fibronectin were more likely to be delivered within 7 days (relative risk 25.9 [95% confidence interval 7.8 to 86]), within 14 days (relative risk 20.4 [95% confidence interval 8.0 to 53]), and before 37 completed weeks (relative risk 2.9 [95% confidence interval 2.2 to 3.7]). The negative predictive values for delivery within 7 days, within 14 days, and at < 37 weeks were 99.5%, 99.2%, and 84.5%, respectively. When we used multiple logistic regression analysis to control for potential confounding variables among singleton pregnancies, only the presence of fetal fibronectin (odds ratio 48.8, 95% confidence interval 7.4 to 320), prior preterm birth (odds ratio 8.3, 95% confidence interval 1.5 to 46.6), and tocolysis (odds ratio 4.1, 95% confidence interval 1.0 to 16.0) were associated with birth within 7 days; fetal fibronectin (odds ratio 3.6, 95% confidence interval 2.2 to 5.9), prior preterm birth (odds ratio 2.5, 95% confidence interval 1.4 to 4.4), cervical dilatation > 1 cm (odds ratio 2.9, 95% confidence interval 1.6 to 5

  8. Supporting prostate cancer focal therapy: a multidisciplinary International Consensus of Experts ("ICE").

    PubMed

    Reis, Leonardo O; Billis, Athanase; Zequi, Stenio C; Tobias-Machado, Marcos; Viana, Publio; Cerqueira, Michael; Ward, John F

    2014-06-01

    Prostate cancer is a common malignancy among men, and the current screening, imaging and sampling approaches aim to detect early-stage, organ-confined disease. In such scenario, focal prostate cancer therapy currently relies on the index lesion concept as the dominant lesion that drives the disease natural history. Focal therapy demands the essential imaging and sampling techniques to strategically locate and qualify the disease, but, despite advances in technology, prostate imaging and biopsy have several limitations that need to be overcome if focal therapy is to be developed further. The I Prostate Cancer Focal Treatment International Symposium was convened to foster discussion on this topic that sits at the crossroads of multiple disciplines (Urology, Pathology, Radiology, Radiation Oncology and Medical Oncology) all of which were represented for this comprehensive multidisciplinary review of the current literature.

  9. SNAP Satellite Focal Plane Development

    SciTech Connect

    Bebek, C.; Akerlof, C.; Aldering, G.; Amanullah, R.; Astier, P.; Baltay, C.; Barrelet, E.; Basa, S.; Bercovitz, J.; Bergstrom, L.; Berstein, G.P.; Bester, M.; Bohlin, R.; Bonissent, A.; Bower, C.; Campbell, M.; Carithers, W.; Commins, E.; Day, C.; Deustua, S.; DiGennaro, R.; Ealet, A.; Ellis, R.; Emmett, W.; Eriksson, M.; Fouchez,D.; Fruchter, A.; Genat, J-F.; Goldhaber, G.; Goobar, A.; Groom, D.; Heetderks, H.; Holland, S.; Huterer, D.; Johnson, W.; Kadel, R.; Karcher,A.; Kim, A.; Kolbe, W.; Lafever, R.; Lamoureaux, J.; Lampton, M.; Lefevre, O.; Levi, M.; Levin, D.; Linder, E.; Loken, S.; Malina, R.; Mazure, A.; McKay, T.; McKee, S.; Miquel, R.; Morgan, N.; Mortsell, E.; Mostek, N.; Mufson, S.; Musser, J.; Roe, N.; Nugent, P.; Oluseyi, H.; Pain, R.; Palaio, N.; Pankow, D.; Perlmutter, S.; Prieto, E.; Rabinowitz,D.; Refregier, A.; Rhodes, J.; Schubnell, M.; Sholl, M.; Smadja, G.; Smith, R.; Smoot, G.; Snyder, J.; Spadafora, A.; Szymkowiak, A.; Tarle,G.; Taylor, K.; Tilquin, A.; Tomasch, A.; Vincent, D.; von der Lippe, H.; Walder, J-P.; Wang, G.

    2003-07-07

    The proposed SuperNova/Acceleration Probe (SNAP) mission will have a two-meter class telescope delivering diffraction-limited images to an instrumented 0.7 square degree field in the visible and near-infrared wavelength regime. The requirements for the instrument suite and the present configuration of the focal plane concept are presented. A two year R&D phase, largely supported by the Department of Energy, is just beginning. We describe the development activities that are taking place to advance our preparedness for mission proposal in the areas of detectors and electronics.

  10. Interferon Induced Focal Segmental Glomerulosclerosis

    PubMed Central

    Bayram Kayar, Nuket; Alpay, Nadir; Hamdard, Jamshid; Emegil, Sebnem; Bag Soydas, Rabia; Baysal, Birol

    2016-01-01

    Behçet's disease is an inflammatory disease of unknown etiology which involves recurring oral and genital aphthous ulcers and ocular lesions as well as articular, vascular, and nervous system involvement. Focal segmental glomerulosclerosis (FSGS) is usually seen in viral infections, immune deficiency syndrome, sickle cell anemia, and hyperfiltration and secondary to interferon therapy. Here, we present a case of FSGS identified with kidney biopsy in a patient who had been diagnosed with Behçet's disease and received interferon-alpha treatment for uveitis and presented with acute renal failure and nephrotic syndrome associated with interferon. PMID:27847659

  11. Focal epithelial hyperplasia: Heck disease.

    PubMed

    Cohen, P R; Hebert, A A; Adler-Storthz, K

    1993-09-01

    Two sisters of Mexican ancestry had focal epithelial hyperplasia (FEH). The lesions on the oral mucosa of the older child were initially misinterpreted as representing sexual abuse. Microscopic evaluation of a hematoxylin and eosin-stained section from a lower lip papule demonstrated the histologic features of FEH. Although human papillomavirus (HPV) type 13 and HPV32 have been most consistently present in FEH lesions, types 6, 11, 13, and 32 were not detected in the paraffin-embedded tissue specimen of our patient using an in situ hybridization technique. The lesions persisted or recurred during management using destructive modalities; subsequently, they completely resolved spontaneously.

  12. A TonB-dependent outer membrane protein as a Bacteroides fragilis fibronectin-binding molecule.

    PubMed

    Pauer, Heidi; Ferreira, Eliane de Oliveira; dos Santos-Filho, Joaquim; Portela, Maristela Barbosa; Zingali, Russolina Benedeta; Soares, Rosangela Maria Araújo; Domingues, Regina Maria Cavalcanti Pilotto

    2009-04-01

    The binding of Bacteroides fragilis to plasmatic fibronectin was investigated using strains isolated from healthy subjects and from patients with bacteremia. They were cultivated in a synthetic media in which variations in cysteine concentrations determined alterations in the oxidation-reduction potential (Eh). All the strains assayed were capable of adhering to plasmatic fibronectin when cultivated under oxidizing and reducing conditions. Bacteroides fragilis 1405 showed the greatest difference when the results under these conditions were compared and it was selected for further investigations. Chemical treatments suggested the involvement of a protein in the interaction between B. fragilis and plasmatic fibronectin. Sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis of outer membrane proteins (OMPs) revealed differences between the extracts obtained from cultures grown under the two conditions. Protein bands of c. 102, 100, 77, 73, 50 and 40 kDa were more highly expressed under oxidizing than reducing conditions. Dot blot analysis showed a stronger recognition of plasmatic fibronectin by OMPs obtained from cultures grown under higher Eh, and Western blot assays confirmed a band of c. 102 kDa as fibronectin-binding protein. This protein was sequenced and revealed to be a putative TonB-dependent OMPs. PCR analysis confirmed the presence of this gene in all the studied strains.

  13. Matrix fibronectin disruption and altered endothelial cell adhesion induced by activated leukocytes

    SciTech Connect

    Vincent, P.; Richards, P.; Saba, T.; DelVecchio, P.

    1986-03-01

    Sequestration of activated leukocytes (PMN) within the lung may contribute to pulmonary vascular injury following trauma, sepsis, or intravascular coagulation. Monolayers of cultured rat endothelial cells were utilized to evaluate the effect of activated PMNs on endothelial cell attachment and the extracellular fibronectin matrix over a 4 hr incubation interval. Rat endothelial cells were identified by immunofluorescent staining of Factor VIII R:Ag. Endothelial cells were labeled with /sup 51/Cr in order to establish a cell injury assay in which the release of pelletable (cell associated) or non-pelletable activity was measured in the media. PMN activation was verified by chemiluminescence activity. Following phorbol myristate acetate (PMA) the leukocytes aggregated, chemiluminesced, and caused detachment of /sup 51/Cr endothelial cells. Endothelial detachment increased as a function of time with a plateau by 3 hrs. Immunofluorescent analysis of extracellular fibronectin in endothelial cell cultures revealed disruption of the fibrillar matrix fibronectin in association with endothelial cell disadhesion. Matrix fibronectin disruption was not seen with PMNs or PMA alone. Thus, disruption of the fibronectin matrix by released proteases may contribute to endothelial cell detachment.

  14. Fibronectin adsorption on functionalized electrospun polycaprolactone scaffolds: experimental and molecular dynamics studies.

    PubMed

    Regis, Shawn; Youssefian, Sina; Jassal, Manisha; Phaneuf, Matthew D; Rahbar, Nima; Bhowmick, Sankha

    2014-06-01

    Designing scaffolds to modulate protein adsorption is a key to building advanced scaffolds for tissue regeneration. Protein adsorption to tissue engineering scaffolds is critical in early cell attachment, survival, and eventual proliferation. The goal of this study is to examine the effect of functionalization on fibronectin adsorption to electrospun polycaprolactone (PCL) scaffolds through experimentation using fluorescently labeled fibronectin and to couple this experimental data with analysis of interaction energies obtained through molecular dynamics (MD) simulations to develop a better understanding of the adsorption process. This study is the first to analyze and compare experimental and MD simulation results of fibronectin adsorption on functionalized electrospun PCL scaffolds. Electrospun nanofiber PCL scaffolds were treated with either 1 N NaOH (hydrolyzed) or 46% hexamethylenediamine (HMD) (aminated) solution to be compared with untreated (control) scaffolds. We found that aminated PCL scaffolds experimentally adsorbed more fibronectin than control scaffolds, whereas hydrolyzed scaffolds showed decreased adsorption. MD simulations carried out with NVT ensemble at a temperature of 310 K indicated a higher work of adhesion for both functionalized scaffolds over control. Also, the simulations revealed different conformations of fibronectin on each scaffold type after adsorption, with the arginine-glycine-aspartic acid sequence appearing most accessible on the aminated scaffolds. This suggests that functionalization affects not only the quantity of protein that will adsorb on a scaffold but how it attaches as well, which could affect subsequent cell attachment.

  15. Chemically grafted fibronectin for use in QCM-D cell studies

    PubMed Central

    Sobolewski, Peter; Tomczyk, Nancy; Composto, Russell J.; Eckmann, David M.

    2014-01-01

    Traditionally, fibronectin has been used as a physisorbed surface coating (physFN) in cell culture experiments due to its critical role in cell adhesion. However, because the resulting layer is thick, unstable, and of unpredictable uniformity, this method of fibronectin deposition is unsuitable for some types of research, including quartz crystal microbalance (QCM) experiments involving cells. Here, we present a new method for chemical immobilization of fibronectin onto silicon oxide surfaces, including QCM crystals pre-coated with silicon oxide. We characterize these chemically coated fibronectin surfaces (chemFN) as well as physFN ones using surface ellipsometry (SE), Fourier transform infrared spectroscopy (FTIR), atomic force microscopy (AFM), and contact angle measurements. A cell culture model demonstrates that cells on chemFN and physFN surfaces exhibit similar viability, structure, adhesion and metabolism. Finally, we perform QCM experiments using cells on both surfaces which demonstrate the superior suitability of chemFN coatings for QCM research, and provide real-time QCM-D data from cells subjected to an actin depolymerizing agent. Overall, our method of chemical immobilization of fibronectin yields great potential for furthering cellular experiments in which thin, stable and uniform coatings are desirable. As QCM research with cells has been rather limited in success thus far, we anticipate that this new technique will particularly benefit this experimental system by availing it to the much broader field of cell mechanics. PMID:24657645

  16. Fibronectin and Cyclic Strain Improve Cardiac Progenitor Cell Regenerative Potential In Vitro

    PubMed Central

    Ghosh-Choudhary, Shohini; Fierro, Marcos J.; Christman, Karen L.; Taylor, W. Robert

    2016-01-01

    Cardiac progenitor cells (CPCs) have rapidly advanced to clinical trials, yet little is known regarding their interaction with the microenvironment. Signaling cues present in the microenvironment change with development and disease. This work aims to assess the influence of two distinct signaling moieties on CPCs: cyclic biaxial strain and extracellular matrix. We evaluate four endpoints for improving CPC therapy: paracrine signaling, proliferation, connexin43 expression, and alignment. Vascular endothelial growth factor A (about 900 pg/mL) was secreted by CPCs cultured on fibronectin and collagen I. The application of mechanical strain increased vascular endothelial growth factor A secretion 2–4-fold for CPCs cultured on poly-L-lysine, laminin, or a naturally derived cardiac extracellular matrix. CPC proliferation was at least 25% higher on fibronectin than that on other matrices, especially for lower strain magnitudes. At 5% strain, connexin43 expression was highest on fibronectin. With increasing strain magnitude, connexin43 expression decreased by as much as 60% in CPCs cultured on collagen I and a naturally derived cardiac extracellular matrix. Cyclic mechanical strain induced the strongest CPC alignment when cultured on fibronectin or collagen I. This study demonstrates that culturing CPCs on fibronectin with 5% strain magnitude is optimal for their vascular endothelial growth factor A secretion, proliferation, connexin43 expression, and alignment. PMID:27610140

  17. Chemically grafted fibronectin for use in QCM-D cell studies.

    PubMed

    Kandel, Judith; Lee, Hyun-Su; Sobolewski, Peter; Tomczyk, Nancy; Composto, Russell J; Eckmann, David M

    2014-08-15

    Traditionally, fibronectin has been used as a physisorbed surface coating (physFN) in cell culture experiments due to its critical role in cell adhesion. However, because the resulting layer is thick, unstable, and of unpredictable uniformity, this method of fibronectin deposition is unsuitable for some types of research, including quartz crystal microbalance (QCM) experiments involving cells. Here, we present a new method for chemical immobilization of fibronectin onto silicon oxide surfaces, including QCM crystals pre-coated with silicon oxide. We characterize these chemically coated fibronectin surfaces (chemFN) as well as physFN ones using spectroscopic ellipsometry (SE), Fourier transform infrared spectroscopy (FTIR), atomic force microscopy (AFM), and contact angle measurements. A cell culture model demonstrates that cells on chemFN and physFN surfaces exhibit similar viability, structure, adhesion and metabolism. Finally, we perform QCM experiments using cells on both surfaces which demonstrate the superior suitability of chemFN coatings for QCM research, and provide real-time QCM-D data from cells subjected to an actin depolymerizing agent. Overall, our method of chemical immobilization of fibronectin yields great potential for furthering cellular experiments in which thin, stable and uniform coatings are desirable. As QCM research with cells has been rather limited in success thus far, we anticipate that this new technique will particularly benefit this experimental system by availing it to the much broader field of cell mechanics.

  18. Stuck in the Middle: Fibronectin-Binding Proteins in Gram-Positive Bacteria

    PubMed Central

    Hymes, Jeffrey P.; Klaenhammer, Todd R.

    2016-01-01

    Fibronectin is a multidomain glycoprotein found ubiquitously in human body fluids and extracellular matrices of a variety of cell types from all human tissues and organs, including intestinal epithelial cells. Fibronectin plays a major role in the regulation of cell migration, tissue repair, and cell adhesion. Importantly, fibronectin also serves as a common target for bacterial adhesins in the gastrointestinal tract. Fibronectin-binding proteins (FnBPs) have been identified and characterized in a wide variety of host-associated bacteria. Single bacterial species can contain multiple, diverse FnBPs. In pathogens, some FnBPs contribute to virulence via host cell attachment, invasion, and interference with signaling pathways. Although FnBPs in commensal and probiotic strains are not sufficient to confer virulence, they are essential for attachment to their ecological niches. Here we describe the interaction between human fibronectin and bacterial adhesins by highlighting the FnBPs of Gram-positive pathogens and commensals. We provide an overview of the occurrence and diversity of FnBPs with a focus on the model pathogenic organisms in which FnBPs are most characterized. Continued investigation of FnBPs is needed to fully understand their divergence and specificity in both pathogens and commensals. PMID:27713740

  19. Stuck in the Middle: Fibronectin-Binding Proteins in Gram-Positive Bacteria.

    PubMed

    Hymes, Jeffrey P; Klaenhammer, Todd R

    2016-01-01

    Fibronectin is a multidomain glycoprotein found ubiquitously in human body fluids and extracellular matrices of a variety of cell types from all human tissues and organs, including intestinal epithelial cells. Fibronectin plays a major role in the regulation of cell migration, tissue repair, and cell adhesion. Importantly, fibronectin also serves as a common target for bacterial adhesins in the gastrointestinal tract. Fibronectin-binding proteins (FnBPs) have been identified and characterized in a wide variety of host-associated bacteria. Single bacterial species can contain multiple, diverse FnBPs. In pathogens, some FnBPs contribute to virulence via host cell attachment, invasion, and interference with signaling pathways. Although FnBPs in commensal and probiotic strains are not sufficient to confer virulence, they are essential for attachment to their ecological niches. Here we describe the interaction between human fibronectin and bacterial adhesins by highlighting the FnBPs of Gram-positive pathogens and commensals. We provide an overview of the occurrence and diversity of FnBPs with a focus on the model pathogenic organisms in which FnBPs are most characterized. Continued investigation of FnBPs is needed to fully understand their divergence and specificity in both pathogens and commensals.

  20. Astrocyte-derived tissue Transglutaminase affects fibronectin deposition, but not aggregation, during cuprizone-induced demyelination

    PubMed Central

    Espitia Pinzon, Nathaly; Sanz-Morello, Berta; Brevé, John J. P.; Bol, John G. J. M.; Drukarch, Benjamin; Bauer, Jan; Baron, Wia; van Dam, Anne-Marie

    2017-01-01

    Astrogliosis as seen in Multiple Sclerosis (MS) develops into astroglial scarring, which is beneficial because it seals off the site of central nervous system (CNS) damage. However, astroglial scarring also forms an obstacle that inhibits axon outgrowth and (re)myelination in brain lesions. This is possibly an important cause for incomplete remyelination in the CNS of early stage MS patients and for failure in remyelination when the disease progresses. In this study we address whether under demyelinating conditions in vivo, tissue Transglutaminase (TG2), a Ca2+ -dependent enzyme that catalyses posttranslational modification of proteins, contributes to extracellular matrix (ECM) deposition and/or aggregation. We used the cuprizone model for de- and remyelination. TG2 immunoreactivity and enzymatic activity time-dependently appeared in astrocytes and ECM, respectively, in the corpus callosum of cuprizone-treated mice. Enhanced presence of soluble monomeric and multimeric fibronectin was detected during demyelination, and fibronectin immunoreactivity was slightly decreased in cuprizone-treated TG2−/− mice. In vitro TG2 overexpression in astrocytes coincided with more, while knock-down of TG2 with less fibronectin production. TG2 contributes, at least partly, to fibronectin production, and may play a role in fibronectin deposition during cuprizone-induced demyelination. Our observations are of interest in understanding the functional implications of TG2 during astrogliosis. PMID:28128219

  1. Polymorphisms in Fibronectin Binding Protein A of Staphylococcus Aureus are Associated with Infection of Cardiovascular Devices

    SciTech Connect

    Lower, Steven; Lamlertthon, Supaporn; Casillas-Ituarte, Nadia; Lins, Roberto D.; Yongsunthon, Ruchirej; Taylor, Eric S.; DiBartola, Alex; Edmondson, Catherine; McIntyre, Lauren M.; Reller, L. Barth; Que, Yok-Ai; Ros, Robert; Lower, Brian; Fowler, Vance

    2011-11-08

    Medical implants, like cardiovascular devices, improve the quality of life for countless individuals but may become infected with bacteria like Staphylococcus aureus. Such infections take the form of a bio-film, a structured community of bacterial cells adherent to the surface of a solid substrate. Every bio-film begins with an attractive force or bond between bacterium and substratum. We used atomic force microscopy to probe experimentally forces between a fibronectin-coated surface (i.e., proxy for an implanted cardiac device) and fibronectin-binding receptors on the surface of individual living bacteria from each of 80 clinical isolates of S. aureus. These isolates originated from humans with infected cardiac devices (CDI; n = 26), uninfected cardiac devices (n = 20), and the anterior nares of asymptomatic subjects (n = 34). CDI isolates exhibited a distinct bindingforce signature and had speci!c single amino acid polymorphisms in fibronectin-binding protein A corresponding to E652D, H782Q, and K786N. In silico molecular dynamics simulations demonstrate that residues D652, Q782, and N786 in fibronectin-binding protein A form extra hydrogen bonds with fibronectin, complementing the higher binding force and energy measured by atomic force microscopy for the CDI isolates. This study is significant, because it links pathogenic bacteria biofilms from the length scale of bonds acting across a nanometer-scale space to the clinical presentation of disease at the human dimension.

  2. Proapoptotic fibronectin fragment induces the degradation of ubiquitinated p53 via proteasomes in periodontal ligament cells

    PubMed Central

    Ghosh, Abhijit; Joo, Nam Eok; Chen, Tina Chunyuan; Kapila, Yvonne L.

    2009-01-01

    Background and Objective The extracellular matrix (ECM) plays a key role in signaling necessary for tissue remodeling and cell survival. However, signals from disease-altered ECMs, as that present in inflammatory diseases like periodontitis and arthritis, may lead to apoptosis or programmed cell death of resident cells. Previously, we found that a disease-associated fibronectin fragment triggers apoptosis of primary human periodontal ligament (PDL) cells via a novel apoptotic pathway in which the tumor suppressor, p53, is transcriptionally downregulated. Materials and Methods We used immunofluorescence, transfection assays, western blotting and ELISAs to show that p53 is degraded by a proteasomal pathway in response to a proapoptotic disease-associated fibronectin fragment. Results We now show that under these same apoptotic conditions p53 is further downregulated by post-translational ubiquitination and subsequent targeting to the proteasome for degradation. Pretreatment of cells with the proteasomal inhibitors MG132 and lactacystin rescued the cells from apoptosis. p53 levels in cells transfected with ubiquitin siRNA were resistant to degradation induced by the proapoptotic fibronectin fragment, showing that ubiquitination is important for the proapoptotic fibronectin fragment-induced degradation of p53. Conclusions These data show that a proapoptotic fibronectin matrix induces ubiquitination and degradation of p53 in the proteasome as part of a novel mechanism of apoptosis associated with inflammatory diseases. PMID:20337881

  3. Implementation of focal zooming on the Nike KrF laser.

    PubMed

    Kehne, D M; Karasik, M; Aglitsky, Y; Smyth, Z; Terrell, S; Weaver, J L; Chan, Y; Lehmberg, R H; Obenschain, S P

    2013-01-01

    In direct drive inertial confinement laser fusion, a pellet containing D-T fuel is imploded by ablation arising from absorption of laser energy at its outer surface. For optimal coupling, the focal spot of the laser would continuously decrease to match the reduction in the pellet's diameter, thereby minimizing wasted energy. A krypton-fluoride laser (λ = 248 nm) that incorporates beam smoothing by induced spatial incoherence has the ability to produce a high quality focal profile whose diameter varies with time, a property known as focal zooming. A two-stage focal zoom has been demonstrated on the Nike laser at the Naval Research Laboratory. In the experiment, a 4.4 ns laser pulse was created in which the on-target focal spot diameter was 1.3 mm (full width at half maximum) for the first 2.4 ns and 0.28 mm for the final 2 ns. These two diameters appear in time-integrated focal plane equivalent images taken at several locations in the amplification chain. Eight of the zoomed output beams were overlapped on a 60 μm thick planar polystyrene target. Time resolved images of self-emission from the rear of the target show the separate shocks launched by the two corresponding laser focal diameters.

  4. Implementation of focal zooming on the Nike KrF laser

    SciTech Connect

    Kehne, D. M.; Karasik, M.; Weaver, J. L.; Chan, Y.; Obenschain, S. P.; Aglitsky, Y.; Smyth, Z.; Lehmberg, R. H.; Terrell, S.

    2013-01-15

    In direct drive inertial confinement laser fusion, a pellet containing D-T fuel is imploded by ablation arising from absorption of laser energy at its outer surface. For optimal coupling, the focal spot of the laser would continuously decrease to match the reduction in the pellet's diameter, thereby minimizing wasted energy. A krypton-fluoride laser ({lambda}= 248 nm) that incorporates beam smoothing by induced spatial incoherence has the ability to produce a high quality focal profile whose diameter varies with time, a property known as focal zooming. A two-stage focal zoom has been demonstrated on the Nike laser at the Naval Research Laboratory. In the experiment, a 4.4 ns laser pulse was created in which the on-target focal spot diameter was 1.3 mm (full width at half maximum) for the first 2.4 ns and 0.28 mm for the final 2 ns. These two diameters appear in time-integrated focal plane equivalent images taken at several locations in the amplification chain. Eight of the zoomed output beams were overlapped on a 60 {mu}m thick planar polystyrene target. Time resolved images of self-emission from the rear of the target show the separate shocks launched by the two corresponding laser focal diameters.

  5. Pathogenesis of Focal Segmental Glomerulosclerosis

    PubMed Central

    Lim, Beom Jin; Yang, Jae Won; Do, Woo Sung; Fogo, Agnes B.

    2016-01-01

    Focal segmental glomerulosclerosis (FSGS) is characterized by focal and segmental obliteration of glomerular capillary tufts with increased matrix. FSGS is classified as collapsing, tip, cellular, perihilar and not otherwise specified variants according to the location and character of the sclerotic lesion. Primary or idiopathic FSGS is considered to be related to podocyte injury, and the pathogenesis of podocyte injury has been actively investigated. Several circulating factors affecting podocyte permeability barrier have been proposed, but not proven to cause FSGS. FSGS may also be caused by genetic alterations. These genes are mainly those regulating slit diaphragm structure, actin cytoskeleton of podocytes, and foot process structure. The mode of inheritance and age of onset are different according to the gene involved. Recently, the role of parietal epithelial cells (PECs) has been highlighted. Podocytes and PECs have common mesenchymal progenitors, therefore, PECs could be a source of podocyte repopulation after podocyte injury. Activated PECs migrate along adhesion to the glomerular tuft and may also contribute to the progression of sclerosis. Markers of activated PECs, including CD44, could be used to distinguish FSGS from minimal change disease. The pathogenesis of FSGS is very complex; however, understanding basic mechanisms of podocyte injury is important not only for basic research, but also for daily diagnostic pathology practice. PMID:27744657

  6. Fibronectin From Oviductal Cells Fluctuates During the Estrous Cycle and Contributes to Sperm-Oviduct Interaction in Cattle.

    PubMed

    Osycka-Salut, Claudia E; Castellano, Luciana; Fornes, Daiana; Beltrame, Jimena S; Alonso, Carlos A I; Jawerbaum, Alicia; Franchi, Ana; Díaz, Emilce S; Perez Martinez, Silvina

    2017-11-01

    During the passage of sperm through the oviduct, spermatozoa bind to the oviductal epithelium and form the oviductal reservoir. This interaction keeps the fertilizing capacity of sperm until ovulation-associated signals induce sperm release from the oviductal epithelium, allowing the transit of spermatozoa to the fertilization site. Fibronectin is a glycoprotein from the extracellular matrix that binds to α5β1 receptors. Fibronectin has been found to be expressed in the oviduct, whereas α5β1 has been found to be expressed in the sperm of different species. Fibronectin is involved through α5β1 in sperm functions. The aim of this work was to study the participation of oviductal fibronectin in the regulation of the sperm-oviduct interaction in cattle. We found that oviductal epithelial cells differentially expressed all mRNA splice variants of fibronectin during the estrous cycle. Fibronectin was localized in the apical region of oviductal epithelial cells and fibronectin levels in the oviductal fluid fluctuated during the estrous cycle. Also, bovine spermatozoa expressed α5β1. Using in vitro sperm-oviduct co-cultures, we found that spermatozoa were attached to the oviductal epithelium through α5β1. The incubation of co-cultures with fibronectin induced sperm release from the oviductal cells through α5β1. The sperm population released from oviductal cells by fibronectin was enriched in motile and capacitated spermatozoa. Based on our in vitro culture system results, we propose that fibronectin and α5β1 are involved in the sperm-oviduct interaction. Also, an increase in fibronectin levels in the oviductal fluid during the pre-ovulatory period may promote sperm release from the oviductal epithelium in cattle. J. Cell. Biochem. 118: 4095-4108, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  7. Fluorescent gangliosides as probes for the retention and organization of fibronectin by ganglioside-deficient mouse cells

    PubMed Central

    1985-01-01

    Ganglioside-deficient transformed mouse fibroblasts (NCTC 2071A cells), which grow in serum-free medium, synthesize fibronectin but do not retain it on the cell surface. When fluorescent derivatives of gangliosides, containing either rhodamine or Lucifer yellow CH attached to the sialic acid residues, were added to the culture medium, the cells incorporated the derivatives and their surfaces became highly fluorescent. When the cells were stained with anti-fibronectin antibodies and a fluorescent second antibody, fibrillar strands of fibronectin were observed to be attached to the cell surface, with partial coincidence of the patterns of direct ganglioside fluorescence and indirect fibronectin immunofluorescence at the cell surface. When the cells were exposed to bacterial neuraminidase during the time of ganglioside insertion, similar patterns of fluorescence were observed. Because the fluorescent gangliosides are resistant to the enzyme, these results suggest that neuraminidase-sensitive endogenous glycoconjugates were not involved in the ganglioside-mediated retention and organization of endogenous fibronectin. After cells were exposed to exogenous chicken fibronectin, most of the fibronectin was attached to the substratum and only a few fibrils were attached to the cells. When exogenous gangliosides were included in the incubation, there was a striking increase in cell-associated exogenous fibronectin, which was highly organized into a fibrillar network. Conversely, cells incubated for 18 h with exogenous unmodified gangliosides exhibited a highly organized network of endogenously derived fibronectin. Upon further incubation of the cells for 2 h with fluorescent gangliosides, there was considerable co-distribution of the fluorescent gangliosides with the fibronectin network as revealed by immunofluorescence. Our results support the concept that gangliosides can mediate the attachment of fibronectin to the cell surface and its organization into a fibrillar

  8. Regulation of the L-type calcium channel by alpha 5beta 1 integrin requires signaling between focal adhesion proteins.

    PubMed

    Wu, X; Davis, G E; Meininger, G A; Wilson, E; Davis, M J

    2001-08-10

    The L-type calcium channel is the major calcium influx pathway in vascular smooth muscle and is regulated by integrin ligands, suggesting an important link between extracellular matrix and vascular tone regulation in tissue injury and remodeling. We examined the role of integrin-linked tyrosine kinases and focal adhesion proteins in regulation of L-type calcium current in single vascular myocytes. Soluble tyrosine kinase inhibitors blocked the increase in current produced by alpha(5) integrin antibody or fibronectin, whereas tyrosine phosphatase inhibition enhanced the effect. Cell dialysis with an antibody to focal adhesion kinase or with FRNK, the C-terminal noncatalytic domain of focal adhesion kinase, produced moderate (24 or 18%, respectively) inhibition of basal current but much greater inhibition (63 or 68%, respectively) of integrin-enhanced current. A c-Src antibody and peptide inhibitors of the Src homology-2 domain or a putative Src tyrosine phosphorylation site on the channel produced similar inhibition. Antibodies to the cytoskeletal proteins paxillin and vinculin, but not alpha-actinin, inhibited integrin-dependent current by 65-80%. Therefore, alpha(5)beta(1) integrin appears to regulate a tyrosine phosphorylation cascade involving Src and various focal adhesion proteins that control the function of the L-type calcium channel. This interaction may represent a novel mechanism for control of calcium influx in vascular smooth muscle and other cell types.

  9. Determination of the Clostridium perfringens-binding site on fibronectin.

    PubMed

    Katayama, Seiichi; Tagomori, Mika; Morita, Naomi; Yamasaki, Tsutomu; Nariya, Hirofumi; Okada, Mariko; Watanabe, Mariko; Hitsumoto, Yasuo

    2015-08-01

    The extracellular matrix protein fibronectin (Fn) is known to bind to the surface of Clostridium perfringens cells. Fn is a disulfide-linked homodimer protein, with each Fn polypeptide consisting of three types of repeating modules: 12 type I, 2 type II, and 15-17 type III modules. To determine the epitope on Fn recognized by C. perfringens cells, anti-Fn monoclonal antibodies (mAbs) and various Fn fragments (III2-10, rIII2-4, rIII5-7, rIII8, rIII9, rIII10) were employed. Although two C. perfringens-derived Fn-binding proteins, FbpA and FbpB, have been reported, they appear not to be the bacterium's surface Fn receptor. Moreover, both FbpA and FbpB were found to bind to C. perfringens cells. To avoid confusion, a mutant C. perfringens lacking both the fbpA and fbpB genes (MW5) was prepared using an in-frame deletion system. MW5 cells bound Fn on their surface, suggesting the presence of a putative Fn receptor(s) on C. perfringens cells. Of several anti-Fn mAbs, both HB39 and MO inhibited the binding of Fn to MW5 cells. HB39 reacted strongly with III2-10 and rIII9, and weakly with rIII2-4, rIII10 and rIII5-7 in Western blotting analysis. Binding of HB39 to Fn was inhibited in the presence of either rIII9 or rIII10, but not in the presence of rIII2-4, rIII5-7, or rIII8. Binding of Fn to MW5 cells was strongly inhibited by both III2-10 and rIII9, marginally inhibited by rIII2-4, but not affected by rIII5-7, rIII8, or rIII10. Significant binding of MW5 cells to immobilized rIII9 and rIII10 as well as immobilized III2-10 was observed. The region of Fn recognized by C. perfringens was thus mapped to the region encompassed by III9 and III10. Copyright © 2014 Elsevier Ltd. All rights reserved.

  10. A comparison of prevaginal and postvaginal manipulation fetal fibronectin.

    PubMed

    Turitz, Amy L; Ackerman, Christina M; Johnson, Denise L; Bank, Tracy C; Duong, Jimmy K; Lee, Shing M; Gyamfi-Bannerman, Cynthia

    2016-05-01

    Fetal fibronectin (fFN) is used as a biomarker for preterm delivery. Currently, its use is discouraged if there has been vaginal manipulation in the previous 24 hours. Our objective is to determine if there are differences between fFN results before and after vaginal manipulation in the form of sterile vaginal exam or transvaginal ultrasound. This was a prospective observational cohort study at a single center of women between 22-33 6/7 weeks at risk for preterm delivery due to: (1) a history of preterm delivery, short cervix, or multifetal gestation; or (2) symptoms of preterm labor. We excluded women with vaginal bleeding or infection, placenta previa, ruptured membranes, cervical dilation >3 cm, or any form of vaginal manipulation in the previous 24 hours. Specimen A was collected prior to planned vaginal exam or transvaginal ultrasound and specimen B was collected within 4 hours. The agreement between specimens A and B was assessed using descriptive statistics. Test characteristics of specimens A and B using the outcome of preterm delivery (<37 weeks) were calculated. In all, 310 specimen pairs from 237 women were collected. Specimen A was positive in 37 (12%) instances and negative in 273 (88%) while specimen B was positive in 39 (13%) and negative in 271 (87%). There were discordant results in 26 specimen pairs. Of these, 14 (5%) negative specimen A results subsequently became positive for specimen B, and 12 (32%) positive specimen A results became negative for specimen B. Overall, there was a 92% agreement between specimens A and B (confidence interval, 88-94%). The specificity of specimens A and B for preterm birth was 90% vs 89%, respectively, with a negative predictive value of 87% for both. The false-negative rate was 12.8% for specimen A and 13.3% for specimen B. There is a moderately high degree of agreement between prevaginal and postvaginal manipulation fFN results. Their test characteristics for evaluating preterm birth are similar with strong

  11. Potential regulation of cartilage metabolism in osteoarthritis by fibronectin fragments.

    PubMed

    Homandberg, G A

    1999-10-15

    There are few candidates for biochemical pathways that either initiate or amplify catabolic processes involved in osteoarthritis (OA). Perhaps, one of the most likely sources for such pathways may be within the extracellular matrix itself. This review focuses on an example of how specific degradation products of the extracellular matrix of cartilage, produced during proteolytic damage, have the potential to enhance OA-like processes. In this example, these products can induce or activate other factors, such as catabolic cytokines, that amplify the damage. The damage, in turn, enhances levels of the degradation products themselves, as in a positive feedback loop. Since these products are derived from the cartilage matrix, they could be considered barometers of the health of the cartilage that signal to the chondrocyte, through outside to inside signaling, the health or status of the surrounding matrix. The best example and most characterized system is that of fragments of the matrix protein, fibronectin (Fn), although as discussed later, other recently discovered fragment systems may also have the potential to regulate cartilage metabolism. In the case of Fn fragments (Fn-fs), the Fn-fs enhance levels of catabolic cytokines as in OA and, thus, are potentially earlier damage mediators than catabolic cytokines. The Fn-fs up-regulate matrix metalloproteinase (MMP) expression, significantly enhance degradation and loss of proteoglycan (PG) from cartilage and temporarily suppress PG synthesis, all events observed in OA. However, this Fn-f system may be involved in normal cartilage homeostasis as well. For example, low concentrations of Fn-fs enhance anabolic activities and could play a role in normal homeostasis. This system may also be involved in not only amplifying damage but also coupling damage to repair. For example, high concentrations of Fn-fs that might arise in OA temporarily offset the anabolic response of lower Fn-f concentrations and cause short

  12. Platelet-rich plasma and chronic wounds: remaining fibronectin may influence matrix remodeling and regeneration success.

    PubMed

    Moroz, Andrei; Deffune, Elenice

    2013-11-01

    Platelet-rich plasma has been largely used as a therapeutic option for the treatment of chronic wounds of different etiologies. The enhanced regeneration observed after the use of platelet-rich plasma has been systematically attributed to the growth factors that are present inside platelets' granules. We hypothesize that the remaining plasma and platelet-bound fibronectin may act as a further bioactive protein in platelet-rich plasma preparations. Recent reports were analyzed and presented as direct evidences of this hypotheses. Fibronectin may directly influence the extracellular matrix remodeling during wound repair. This effect is probably through matrix metalloproteinase expression, thus exerting an extra effect on chronic wound regeneration. Physicians should be well aware of the possible fibronectin-induced effects in their future endeavors with PRP in chronic wound treatment. Copyright © 2013 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  13. klf2a couples mechanotransduction and zebrafish valve morphogenesis through fibronectin synthesis

    PubMed Central

    Steed, Emily; Faggianelli, Nathalie; Roth, Stéphane; Ramspacher, Caroline; Concordet, Jean-Paul; Vermot, Julien

    2016-01-01

    The heartbeat and blood flow signal to endocardial cell progenitors through mechanosensitive proteins that modulate the genetic program controlling heart valve morphogenesis. To date, the mechanism by which mechanical forces coordinate tissue morphogenesis is poorly understood. Here we use high-resolution imaging to uncover the coordinated cell behaviours leading to heart valve formation. We find that heart valves originate from progenitors located in the ventricle and atrium that generate the valve leaflets through a coordinated set of endocardial tissue movements. Gene profiling analyses and live imaging reveal that this reorganization is dependent on extracellular matrix proteins, in particular on the expression of fibronectin1b. We show that blood flow and klf2a, a major endocardial flow-responsive gene, control these cell behaviours and fibronectin1b synthesis. Our results uncover a unique multicellular layering process leading to leaflet formation and demonstrate that endocardial mechanotransduction and valve morphogenesis are coupled via cellular rearrangements mediated by fibronectin synthesis. PMID:27221222

  14. Single-molecule dynamic force spectroscopy of the fibronectin-heparin interaction

    SciTech Connect

    Mitchell, Gabriel; Lamontagne, Charles-Antoine; Lebel, Rejean; Grandbois, Michel Malouin, Francois

    2007-12-21

    The integrity of cohesive tissues strongly depends on the presence of the extracellular matrix, which provides support and anchorage for cells. The fibronectin protein and the heparin-like glycosaminoglycans are key components of this dynamic structural network. In this report, atomic force spectroscopy was used to gain insight into the compliance and the resistance of the fibronectin-heparin interaction. We found that this interaction can be described by an energetic barrier width of 3.1 {+-} 0.2 A and an off-rate of 0.2 {+-} 0.1 s{sup -1}. These dissociation parameters are similar to those of other carbohydrate-protein interactions and to off-rate values reported for more complex interactions between cells and extracellular matrix components. Our results indicate that the function of the fibronectin-heparin interaction is supported by its capacity to sustain significant deformations and considerable external mechanical forces.

  15. Multiplicative and subtractive focal volume engineering in coherent Raman microscopy

    PubMed Central

    Raghunathan, Varun; Potma, Eric Olaf

    2012-01-01

    Rigorous calculations are performed to study the effective reduction of the nonlinear excitation volumes when using phase-only masks to condition the pump and Stokes driving fields. Focal volume reduction was achieved using both a multiplicative operation of the excitation fields as well as a subtractive operation. Using a tunable optical bottle beam for the Stokes field, an effective reduction of the width of the excitation volume by a factor of 1.5 can be achieved in the focal plane. Further reduction of the focal volume introduces a rapid growth of sidelobes, which renders such volumes unsuitable for imaging applications. In addition, phase sensitive detection was found to provide information from selective sub-divisions of the engineered coherent anti-Stokes Raman scattering excitation volume. In the case of isolated nanoparticles, an apparent resolution improvement by a factor of 3 is demonstrated, and it is shown that the size of sub-diffraction-limited particles can be accurately determined using phase sensitive detection. PMID:21045900

  16. Expression of fibronectin and integrins in cultured periodontal ligament epithelial cells.

    PubMed

    Uitto, V J; Larjava, H; Peltonen, J; Brunette, D M

    1992-05-01

    The process of attachment of epithelial cells obtained from the porcine periodontal ligament (cell rests of Malassez) to different extracellular matrix proteins and their expression of fibronectin and integrin receptors were studied by means of immunocytochemistry, in situ hybridization, and time-lapse cinemicrography techniques. The cell lines of periodontal ligament epithelial cells (PLE cells) attached to and spread rapidly on fibronectin, vitronectin, and type I collagen. One of the cell lines also attached to laminin, while the other cell line showed poor attachment to both laminin and Matrigel, a basement membrane material. By use of the in situ hybridization technique, some PLE cells were found to express the fibronectin gene strongly. Immunocytochemical staining localized fibronectin in extracellular fibrils and intracellular granules. Fibronectin was also found in the tracks left behind by the cells migrating on the substratum. Arg-gly-asp-ser peptide inhibited the attachment of the PLE cells to fibronectin, laminin, type I collagen, and vitronectin by 47%, 43%, 83%, and 94%, respectively, suggesting that the cell-matrix interactions were partly mediated by receptors related to the integrin family. Antibodies against the beta 1-integrin subunit stained the cell bodies and the plasma membrane projections of spreading cells. After 24 h or longer in culture, beta 1-integrins were localized to the regions of cell-cell contact. Cinemicrography of the arg-gly-asp-ser-peptide-treated cells demonstrated that the spreading and migration of isolated cells were prevented by the peptide. The peptide did not appear to dissociate the cell-cell contacts or interfere with migration of spread-cell colonies.(ABSTRACT TRUNCATED AT 250 WORDS)

  17. Isolation and characterization of fibronectin-alpha 1-microglobulin complex in rat plasma.

    PubMed Central

    Falkenberg, C; Enghild, J J; Thøgersen, I B; Salvesen, G; Akerström, B

    1994-01-01

    Molecules containing the 28 kDa immunoregulatory protein alpha 1-microglobulin (alpha 1-m), also known as protein HC, were isolated from rat plasma or serum by immunoaffinity chromatography. Three molecular species were distinguished on the basis of nondenaturing PAGE. Two of these have been described previously: uncomplexed alpha 1-m, and the complex of alpha 1-m with alpha 1-inhibitor-3. The third species was analysed by denaturing PAGE, immunoblotting, proteinase digestion and N-terminal-sequence analyses, and shown to consist of a complex between alpha 1-m and fibronectin. This complex, with a mass of about 560 kDa, was resistant to dissociation in the presence of denaturants, but not in the presence of reducing agents in combination with denaturants, and we conclude that the two components are linked by disulphide bonds. About 60% of the total detectable plasma alpha 1-m exists as high-molecular-mass complexes distributed approximately evenly between fibronectin and alpha 1-inhibitor-3. Immunochemical analyses were used to determine the proportion of the total plasma pools of fibronectin and alpha 1-inhibitor-3 that circulate in complex with alpha 1-m. About 3-7% of the total plasma fibronectin from three different rat strains contained alpha 1-m, whereas 0.3-0.8% of the total plasma alpha 1-inhibitor-3 contained alpha 1-m. Complexes were found at similar levels in plasma and serum, indicating that coagulation is not responsible for complex formation. Moreover, immunochemical analyses of human plasma revealed small amounts of alpha 1-m in complex with fibronectin and alpha 2-macroglobulin (an alpha 1-inhibitor-3 homologue). The existence of a complex between alpha 1-m and fibronectin in rats and humans suggests a mechanism for the incorporation of the immunoregulatory molecule alpha 1-m into the extracellular matrix. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:7519849

  18. Gene expression profiling identifies Fibronectin 1 and CXCL9 as candidate biomarkers for breast cancer screening

    PubMed Central

    Ruiz-Garcia, E; Scott, V; Machavoine, C; Bidart, J M; Lacroix, L; Delaloge, S; Andre, F

    2010-01-01

    Background: There is a need to develop blood-based bioassays for breast cancer (BC) screening. In this study, differential gene expression between BC samples and benign tumours was used to identify candidate biomarkers for blood-based screening. Methods: We identified two proteins (Fibronectin 1 and CXCL9) from a gene expression data set that included 120 BC samples and 45 benign lesions. These proteins fulfil the following criteria: differential gene expression between cancer and benign lesion, protein released in the extracellular medium and stable in the serum, commercially available ELISA kit, ELISA accuracy in a feasibility study. Protein concentrations were determined by ELISA. Blood samples were from normal volunteers (n=119) and early BC patients (n=133). Results: Seventy-three per cent of patients had cT1-T2 tumour. Patients had higher CXCL9 and Fibronectin 1 concentrations than volunteers. CXCL9 mean concentration was 851 and 635 pg ml−1 for patients and volunteers respectively (P=0.013). CXCL9 concentration was significantly higher in patients with estrogen receptor (ER)-negative compared with volunteers (P=0.003), data consistent with gene expression profile. Fibronectin 1 mean concentration was 190 μg ml−1 for patients and 125 μg ml−1 for volunteers (P<0.001). Areas under the curve for BC diagnosis were 0.78 and 0.62 for Fibronectin 1 and CXCL9 respectively. A combined score including Fibronectin 1 and CXCL9 dosages presented 53% of sensitivity and 98% of specificity. Similar performances were observed for ER-negative tumours. Conclusions: This study suggests that Fibronectin 1/CXCL9 dosage in serum could screen a significant rate of BC, including ER-negative, and that differential gene expression analysis is a good approach to select candidate biomarkers to set up blood assays cancer screening. PMID:20068563

  19. Hyaluronan and the hyaluronan receptor RHAMM promote focal adhesion turnover and transient tyrosine kinase activity

    PubMed Central

    1994-01-01

    The molecular mechanisms whereby hyaluronan (HA) stimulates cell motility was investigated in a C-H-ras transformed 10T 1/2 fibroblast cell line (C3). A significant (p < 0.001) stimulation of C3 cell motility with HA (10 ng/ml) was accompanied by an increase in protein tyrosine phosphorylation as detected by anti-phosphotyrosine antibodies using immunoblot analysis and immunofluorescence staining of cells. Tyrosine phosphorylation of several proteins was found to be both rapid and transient with phosphorylation occurring within 1 min of HA addition and dissipating below control levels 10-15 min later. These responses were also elicited by an antibody generated against a peptide sequence within the HA receptor RHAMM. Treatment of cells with tyrosine kinase inhibitors (genistein, 10 micrograms/ml or herbimycin A, 0.5 micrograms/ml) or microinjection of anti-phosphotyrosine antibodies inhibited the transient protein tyrosine phosphorylation in response to HA as well as prevented HA stimulation of cell motility. To determine a link between HA-stimulated tyrosine phosphorylation and the resulting cell locomotion, cytoskeletal reorganization was examined in C3 cells plated on fibronectin and treated with HA or anti-RHAMM antibody. These agents caused a rapid assembly and disassembly of focal adhesions as revealed by immunofluorescent localization of vinculin. The time course with which HA and antibody induced focal adhesion turnover exactly paralleled the induction of transient protein tyrosine phosphorylation. In addition, phosphotyrosine staining colocalized with vinculin within structures in the lamellapodia of these cells. Notably, the focal adhesion kinase, pp125FAK, was rapidly phosphorylated and dephosphorylated after HA stimulation. These results suggest that HA stimulates locomotion via a rapid and transient protein tyrosine kinase signaling event mediated by RHAMM. They also provide a possible molecular basis for focal adhesion turnover, a process that is

  20. Towards Dualband Megapixel QWIP Focal Plane Arrays

    DTIC Science & Technology

    2006-01-01

    Journal Article PREPRINT 3. DATES COVERED (From - To) 2006 4. TITLE AND SUBTITLE Towards dualband megapixel QWIP focal plane arrays (PREPRINT) 5a...pixel quantum well infrared photodetector ( QWIP ) focal planes have been demonstrated with excellent imaging performance. The MWIR QWIP detector array...registered simultaneously readable dualband QWIP focal plane arrays. In this paper, we will discuss the performance in terms of quantum efficiency

  1. RF/Optical Demonstration: Focal Plane Assembly

    NASA Astrophysics Data System (ADS)

    Hoppe, D. J.; Chung, S.; Kovalik, J.; Gama, E.; Fernandez, M. M.

    2016-11-01

    In this article, we describe the second-generation focal plane optical assembly employed in the RF/optical demonstration at DSS-13. This assembly receives reflected light from the two mirror segments mounted on the RF primary. The focal plane assembly contains a fast steering mirror (FSM) to stabilize the focal plane spot, a pupil camera to aid in aligning the two segments, and several additional cameras for receiving the optical signal prior to as well as after the FSM loop.

  2. Modulation of the Metastatic Activity of Melanoma Cells by Laminin and Fibronectin

    NASA Astrophysics Data System (ADS)

    Terranova, Victor P.; Williams, Jeannette E.; Liotta, Lance A.; Martin, George R.

    1984-11-01

    Metastatic mouse melanoma cells have a high affinity for the basement membrane and the ability to degrade it; these properties may allow tumor cells to invade the membrane and disseminate. In this study it was found that the metastatic potential of mouse melanoma cells varied when the cells were exposed in culture to fibronectin or laminin. After removal of fibronectin or exposure to laminin, the cells had an increased affinity for basement membrane collagen, were more invasive of basement membranes in vitro, and produced more lung colonies in vivo. These changes are correlated with and may be due to an increase in the laminin-binding capacity of the tumor cell surface.

  3. Activated platelets form protected zones of adhesion on fibrinogen and fibronectin-coated surfaces

    PubMed Central

    1993-01-01

    Leukocytes form zones of close apposition when they adhere to ligand- coated surfaces. Because plasma proteins are excluded from these contact zones, we have termed them protected zones of adhesion. To determine whether platelets form similar protected zones of adhesion, gel-filtered platelets stimulated with thrombin or ADP were allowed to adhere to fibrinogen- or fibronectin-coated surfaces. The protein- coated surfaces with platelets attached were stained with either fluorochrome-conjugated goat anti-human fibrinogen or anti-human fibronectin antibodies, or with rhodamine-conjugated polyethylene glycol polymers. Fluorescence microscopy revealed that F(ab')2 anti- fibrinogen (100 kD) did not penetrate into the contact zones between stimulated platelets and the underlying fibrinogen-coated surface, while Fab antifibrinogen (50 kD) and 10 kD polyethylene glycol readily penetrated and stained the substrate beneath the platelets. Thrombin- or ADP-stimulated platelets also formed protected zones of adhesion on fibronectin-coated surfaces. F(ab')2 anti-fibronectin and 10 kD polyethylene glycol were excluded from these adhesion zones, indicating that they are much less permeable than those formed by platelets on fibrinogen-coated surfaces. The permeability properties of protected zones of adhesion formed by stimulated platelets on surfaces coated with both fibrinogen and fibronectin were similar to the zones of adhesion formed on fibronectin alone. mAb 7E3, directed against the alpha IIb beta 3 integrin blocked the formation of protected adhesion zones between thrombin-stimulated platelets and fibrinogen or fibronectin coated surfaces. mAb C13 is directed against the alpha 5 beta 1 integrin on platelets. Stimulated platelets treated with this mAb formed protected zones of adhesion on surfaces coated with fibronectin. These protected zones were impermeable to F(ab')2 antifibronectin but were permeable to 10 kD polyethylene glycol. These results show that activated

  4. Attention and driving in traumatic brain injury: a question of coping with time-pressure.

    PubMed

    Brouwer, Wiebo H; Withaar, Frederiec K; Tant, Mark L M; van Zomeren, Adriaan H

    2002-02-01

    Diffuse and focal traumatic brain injury (TBI) can result in perceptual, cognitive, and motor dysfunction possibly leading to activity limitations in driving. Characteristic dysfunctions for severe diffuse TBI are confronted with function requirements derived from the hierarchical task analysis of driving skill. Specifically, we focus on slow information processing, divided attention, and the development of procedural knowledge. Also the effects of a combination of diffuse and focal dysfunctions, specifically homonymous hemianopia and the dysexecutive syndrome, are discussed. Finally, we turn to problems and challenges with regard to assessment and rehabilitation methods in the areas of driving and fitness to drive.

  5. Mislocalized scaffolding by the Na-H exchanger NHE1 dominantly inhibits fibronectin production and TGF-beta activation.

    PubMed

    Karydis, Anastasios; Jimenez-Vidal, Maite; Denker, Sheryl P; Barber, Diane L

    2009-04-01

    Secretion and assembly of the extracellular matrix protein fibronectin regulates a number of normal cell and tissue functions and is dysregulated in disease states such as fibrosis, diabetes, and cancer. We found that mislocalized scaffolding by the plasma membrane Na-H exchanger NHE1 suppresses fibronectin expression, secretion, and assembly. In fibroblasts, wild-type NHE1 localizes to the distal margin of membrane protrusions or lamellipodia but a mutant NHE1-KRA2 lacking binding sites for PI(4,5)P2 and the ERM proteins ezrin, radixin, and moesin is mislocalized and found uniformly along the plasma membrane. Although NHE1 regulates intracellular pH homeostasis, fibronectin production is not regulated by changes in intracellular pH, nor is it attenuated in NHE1-deficient cells, indicating fibronectin expression is independent of NHE1 activity. However, fibronectin production is nearly absent in cells expressing NHE1-KRA2 because scaffolding by NHE1 is mislocalized. Additionally, secretion of active but not latent TGF-beta is reduced and exogenous TGF-beta restores fibronectin secretion and assembly. Our data indicate that scaffolding by NHE1-KRA2 dominantly suppresses fibronectin synthesis and TGF-beta activation, and they suggest that NHE1-KRA2 can be used for obtaining a mechanistic understanding of how fibronectin production is regulated and speculatively for therapeutic control of dysregulated production in pathological conditions.

  6. Fibronectin 1 promotes migration and invasion of papillary thyroid cancer and predicts papillary thyroid cancer lymph node metastasis

    PubMed Central

    Xia, Shujun; Wang, Chuandong; Postma, Emily Louise; Yang, Yanhua; Ni, Xiaofeng; Zhan, Weiwei

    2017-01-01

    Lymph node metastasis (LNM) is common in papillary thyroid cancer (PTC), and is an indicator of recurrence. The detailed molecular mechanism of LNM in PTC has not been well described. This study aimed to investigate the role of fibronectin 1 in PTC LNM and its clinical relevance. The expression of fibronectin 1 was confirmed in PTC tissues and cell lines. A correlation analysis was conducted and a receiver-operating characteristic curve obtained. The effect of fibronectin 1 on the proliferation of PTC cell lines was performed using a colony-formation assay and Cell Counting Kit 8. Cell-cycle analysis was performed with a flow-cytometry assay. Migration and invasion ability were evaluated by transwell and wound-healing assays. Fibronectin 1 was overexpressed in metastasized PTC. Overexpressed fibronectin 1 was positively correlated with PTC LNM. Receiver-operating characteristic analysis showed that the diagnostic accuracy of fibronectin 1 was 81.1%, with sensitivity of 80% and specificity of 82%. Overexpression of fibronectin 1 promoted proliferation, migration, and invasion in PTC. Fibronectin 1 plays a critical role in PTC metastasis by modulating the proliferation, migration, and invasion ability of PTC cells, and it is a valuable diagnostic biomarker for predicting PTC LNM. PMID:28367057

  7. Fluorescence polarization assay and SDS-PAGE confirms matrilysin degrades fibronectin and collagen IV whereas gelatinase A degrades collagen IV but not fibronectin.

    PubMed

    Kraft, P J; Haynes-Johnson, D E; Patel, L; Lenhart, J A; Zivin, R A; Palmer, S S

    2001-10-01

    Matrilysin and gelatinase A are hypothesized to have significant roles in uterine and ovarian function. However, proteolytic activity assays for these enzymes are limited. We describe the development of simple and rapid assays for the proteolysis of fluorescein-labeled full-length substrates, collagen IV (Col-IV) and fibronectin (FN), and demonstrate the selectivity of matrilysin (MMP-7) compared to gelatinase A (MMP-2) for fibronectin. Changes in fluorescence intensity (FIU) and fluorescence polarization (mP) resulting from the protease activity of matrilysin and gelatinase A were measured. These studies show that the fluorescently labeled substrates, Col-IV and FN, are as reliable and amenable to rapid in vitro assay as peptide substrates. In addition, they are easier to use than previously described, non-fluorescent methods. The results demonstrate that assays using full-length, biological matrix proteins are more sensitive indicators of MMP-specific substrate activity than peptide based assays.

  8. Primary focal hyperhidrosis: diagnosis and management. .

    PubMed

    Wang, Rena; Solish, Nowell; Murray, Christian A

    2008-12-01

    Primary focal hyperhidrosis is a common and serious medical condition that causes considerable psychosocial morbidity. Diagnostic and effective management strategies can improve patients' quality of living dramatically.

  9. Digital scanner infrared focal plane technology

    NASA Astrophysics Data System (ADS)

    Ortiz, M. A.; Malone, N. R.; Harris, M.; Shin, J.; Byers, S.; Price, D.; Vampola, J.

    2011-09-01

    Advancements in finer geometry and technology advancements in circuit design now allow placement of digital architecture on cryogenic focal planes while using less power than heritage analog designs. These advances in technology reduce the size, weight, and power of modern focal planes. In addition, the interface to the focal plane is significantly simplified and is more immune to Electromagnetic Interference (EMI). The cost of the customer's instrument after integration with the digital scanning Focal Plane Array (FPA) has been significantly reduced by placing digital architecture such as Analog to digital convertors and Low Voltage Differential Signaling (LVDS) Inputs and Outputs (I/O) on the Read Out Integrated Circuit (ROIC).

  10. Focal properties of geodesic waveguide lenses

    NASA Technical Reports Server (NTRS)

    Verber, C. M.; Vahey, D. W.; Wood, V. E.

    1976-01-01

    The focal properties of uncorrected geodesic lenses in ion-exchanged glass waveguides are reported. A 13.8-mm-focal-length lens resolved beams with an angular separation of 27.6 mrad, while a 28-mm-focal-length lens resolved beams with an angular separation of only 3.3 mrad. Intensity profiles of the focal region of the former lens revealed a 40-micron spot size when the input aperture was 5 mm, and a spot size of 7.7 microns when the aperture was reduced to 1 mm. This value is close to the diffraction-limited spot size of 5.7 microns.

  11. Smov Baseline Focal Plane Check

    NASA Astrophysics Data System (ADS)

    Gilmozzi, Roberto

    1994-01-01

    This test will be executed during the period after the servicing mission and before the extension of the COSTAR assembly. Its purpose is to verify that the FOS, HRS, and FOC focal planes have not been altered by the activities performed by Story and the Astronauts during the servicing mission. A large unknown deviation in aperture position would severly impact subsequent COSTAR alignment activities. If this test reveals a deviation, we may be able to compensate for any offsets prior to the complex and delicate COSTAR alignment calibrations. This enhanced version of the Heptathlon is designed to verify course alignments and measure relative aperture positions to within a few arcsecs. SPECIAL REQUIREMENTS: ***This test uses pre-servicing parameters for HRS, FOS, and FOC and the Cycle 4 parameters for WFPC2.*** ***This test requires special alignment and special guide stars.** ***This test requires special commanding for telemetry setups.**

  12. Focal epithelial hyperplasia - an update.

    PubMed

    Said, Ahmed K; Leao, Jair C; Fedele, Stefano; Porter, Stephen R

    2013-07-01

    Focal epithelial hyperplasia (FEH) is an asymptomatic benign mucosal disease, which is mostly observed in specific groups in certain geographical regions. FEH is usually a disease of childhood and adolescence and is generally associated with people who live in poverty and of low socioeconomic status. Clinically, FEH is typically characterized by multiple, painless, soft, sessile papules, plaques or nodules, which may coalesce to give rise to larger lesions. Human papillomavirus (HPV), especially genotypes 13 and 32, have been associated and detected in the majority of FEH lesions. The clinical examination and social history often allow diagnosis, but histopathological examination of lesional tissue is usually required to confirm the exact diagnosis. FEH sometimes resolves spontaneously however, treatment is often indicated as a consequence of aesthetic effects or any interference with occlusion. There remains no specific therapy for FEH, although surgical removal, laser excision or possibly topical antiviral agents may be of benefit. There remains no evidence that FEH is potentially malignant.

  13. Enhanced focal adhesion assembly reflects increased mechanosensation and mechanotransduction at maternal-conceptus interface and uterine wall during ovine pregnancy.

    PubMed

    Burghardt, Robert C; Burghardt, James R; Taylor, James D; Reeder, Adele T; Nguen, Bar T; Spencer, Thomas E; Bayless, Kayla J; Johnson, Greg A

    2009-03-01

    The integrity of the fetal-maternal interface is critical for proper fetal nourishment during pregnancy. Integrins are important adhesion molecules present at the interface during implantation; however, in vivo evidence for integrin activation and focal adhesion formation at the maternal-conceptus interface is limited. We hypothesized that focal adhesion assembly in uterine luminal epithelium (LE) and conceptus trophectoderm (Tr) results from integrin binding of extracellular matrix (ECM) at this interface to provide increased tensile forces and signaling to coordinate utero-placental development. An ovine model of unilateral pregnancy was used to evaluate mechanotransduction events leading to focal adhesion assembly at the maternal-conceptus interface and within the uterine wall. Animals were hysterectomized on days 40, 80, or 120 of pregnancy, and uteri immunostained for integrins (ITGAV, ITGA4, ITGA5, ITGB1, ITGB3, and ITGB5), ECM proteins (SPP1, LGALS15, fibronectin (FN), and vitronectin (VTN)), cytoskeletal molecules (ACTN and TLN1), and a signal generator (PTK2). Focal adhesion assembly in myometrium and stroma was also studied to provide a frame of reference for mechanical stretch of the uterine wall. Large focal adhesions containing aggregates of ITGAV, ITGA4, ITGA5, ITGB1, ITGB5, ACTN, and PTK2 were detected in interplacentomal uterine LE and Tr of gravid but not non-gravid uterine horns and increased during pregnancy. SPP1 and LGALS15, but not FN or VTN, were present along LE and Tr interfaces in both uterine horns. These data support the idea that focal adhesion assembly at the maternal-conceptus interface reflects adaptation to increasing forces caused by the growing fetus. Cooperative binding of multiple integrins to SPP1 deposited at the maternal-conceptus interface forms an adhesive mosaic to maintain a tight connection between uterine and placental surfaces along regions of epitheliochorial placentation in sheep.

  14. High expression of fibronectin is associated with poor prognosis, cell proliferation and malignancy via the NF-κB/p53-apoptosis signaling pathway in colorectal cancer

    PubMed Central

    Yi, Wenzhong; Xiao, Enhua; Ding, Ru; Luo, Ping; Yang, Yi

    2016-01-01

    Fibronectin is a glycoprotein of the extracellular matrix, and regulates the processes of self-renewal and cell cycle progression. This study aimed to investigate fibronectin expression in colorectal cancer (CRC) and elucidate the effects of fibronectin on CRC by using a knockdown approach. Immunohistochemistry was used to evaluate the expression of fibronectin in 107 CRC patient tissues and gene expression was detected by real-time quantitative PCR (qPCR) and western blot analysis. Based on the above findings, the association among fibronectin expression, clinicopathological features and prognosis was analyzed. Next, fibronectin expression was silenced by small-interfering RNAs (siRNAs) and the effects of fibronectin siRNA transfection on CRC cells and tumor growth in nude mice were assessed. Expression of genes in the NF-κB/p53-apoptosis signaling pathway were analyzed after fibronectin siRNA transfection both in vitro and in vivo. Based on the results, high expression of fibronectin was observed both in the CRC tissues and CRC cell lines. The expression level was positively correlated with TNM stage (P=0.0025) and distant metastasis (P=0.0013). By Kaplan-Meier analysis, the patients with low fibronectin expression had a longer survival time comparing to those with relatively high expression. Knockdown of fibronectin suppressed SW480 cell proliferation, migration and invasion. In addition, knockdown of fibronectin led to S phase cell cycle arrest. The following study showed that the NF-κB/p53-apoptosis signaling pathway in CRC was affected by fibronectin knockdown. Tumor formation was also depressed by fibronectin siRNA transfection of CRC cells. These results showed the significant role of fibronectin in CRC tissues and cell lines. Therefore, fibronectin may be regarded as a potential target for CRC treatment. PMID:27748871

  15. Dynamics of fibronectin adsorption on TiO2 surfaces.

    PubMed

    Sousa, S R; Brás, M Manuela; Moradas-Ferreira, P; Barbosa, M A

    2007-06-19

    In the present work we analyze the dynamics of fibronectin (FN) adsorption on two different stable titanium oxides, with varied surface roughness, and chemically similar to those used in clinical practice. The two types of titanium oxide surfaces used were TiO2 sputtered on Si (TiO2 sp) and TiO2 formed on commercially pure titanium after immersion in H2O2 (TiO2 cp). Surface characterization was previously carried out using different techniques (Sousa, S. R.; Moradas-Ferreira, P.; Melo, L. V.; Saramago, B.; Barbosa, M. A. Langmuir 2004, 20 (22), 9745-9754). Imaging and roughness analysis before and after FN adsorption used atomic force microscopy (AFM) in tapping mode, in air, and in magnetic alternating current mode, in liquid (water). FN adsorption as a function of time was followed by X-ray photoelectron spectroscopy (XPS), by radiolabeling of FN with 125I (125I-FN), and by ellipsometry. Exchangeability studies were performed using FN and HSA. AFM roughness analysis revealed that, before FN adsorption, both TiO2 surfaces exhibited a lower root-mean-square (Rq) and maximum peak with the depth of the maximum valley (Rmax) roughness in air than in water, due to TiO2 hydration. After protein adsorption, the same behavior was observed for the TiO2 sp substrate, while Rq and Rmax roughness values in air and in water were similar in the case of the TiO2 cp substrate, for the higher FN concentration used. Surface roughness was always significantly higher on the TiO2 cp surfaces. AFM led to direct visualization of adsorbed FN on both surfaces tested, indicating that after 10 min of FN incubation the TiO2 sp surface was partially covered by FN. The adsorbed protein seems to form globular aggregates or ellipsoids, and FN aggregates coalesce, forming clusters as the time of adsorption and the concentration increase. Radiolabeling of FN revealed that a rapid adsorption occurs on both surfaces and the amount adsorbed increased with time, reaching a maximum after 60 min of

  16. Interactions of immunoglobulin G, fibrinogen and fibronectin with Staphylococcus hyicus and Staphylococcus intermedius.

    PubMed

    Lämmler, C; de Freitas, J C; Chhatwal, G S; Blobel, H

    1985-10-01

    Binding of immunoglobulin G, fibrinogen and fibronectin to 112 cultures of coagulase-positive staphylococci together with 7 of coagulase-negative S. hyicus subsp. chromogenes were investigated. Of the coagulase-positive staphylococcal cultures 45 were S. hyicus subsp. hyicus, 51 S. intermedius and 16 S. aureus. All 45 S. hyicus subsp. hyicus cultures coagulated plasma preparations from pigs and not always those from sheep, rabbits and dogs. Labelled IgG was bound by all cultures of S. hyicus subsp. hyicus and S. aureus, but only by 6 of 51 S. intermedius cultures. Fibrinogen interacted with 28 of the 45 S. hyicus subsp. hyicus cultures, with 17 of the 51 S. intermedius cultures and with S. aureus throughout. Fibronectin reacted with 19 cultures of S. hyicus subsp. hyicus, 11 of S. intermedius and all S. aureus. The binding activities for labelled IgG were more pronounced than those for fibrinogen and fibronectin. None of the 7 cultures of S. hyicus subsp. chromogenes bound any of these plasma proteins. Bindings of fibrinogen and fibronectin to S. hyicus subsp. hyicus and S. intermedius elicited only in part distinct clumping reactions of the staphylococci in the respective plasma proteins.

  17. Functional Analysis of an S-Layer-Associated Fibronectin-Binding Protein in Lactobacillus acidophilus NCFM.

    PubMed

    Hymes, Jeffrey P; Johnson, Brant R; Barrangou, Rodolphe; Klaenhammer, Todd R

    2016-05-01

    Bacterial surface layers (S-layers) are crystalline arrays of self-assembling proteinaceous subunits called S-layer proteins (Slps) that comprise the outermost layer of the cell envelope. Many additional proteins that are associated with or embedded within the S-layer have been identified in Lactobacillus acidophilus NCFM, an S-layer-forming bacterium that is widely used in fermented dairy products and probiotic supplements. One putative S-layer-associated protein (SLAP), LBA0191, was predicted to mediate adhesion to fibronectin based on the in silico detection of a fibronectin-binding domain. Fibronectin is a major component of the extracellular matrix (ECM) of intestinal epithelial cells. Adhesion to intestinal epithelial cells is considered an important trait for probiotic microorganisms during transit and potential association with the intestinal mucosa. To investigate the functional role of LBA0191 (designated FbpB) in L. acidophilus NCFM, an fbpB-deficient strain was constructed. The L. acidophilus mutant with a deletion off bpB lost the ability to adhere to mucin and fibronectin in vitro Homologues off bpB were identified in five additional putative S-layer-forming species, but no homologues were detected in species outside theL. acidophilus homology group.

  18. Antibody Response to Fibronectin-Binding Adhesin FnbpA in Patients with Staphylococcus aureus Infections

    PubMed Central

    Casolini, Fabrizia; Visai, Livia; Joh, Danny; Conaldi, Pier Giulio; Toniolo, Antonio; Höök, Magnus; Speziale, Pietro

    1998-01-01

    We have analyzed antibody reactivity to a fibronectin-binding microbial surface component that recognizes adhesive matrix molecules (MSCRAMM) in blood plasma collected from patients with staphylococcal infections. All patients had elevated levels of anti-MSCRAMM antibodies compared to those of young children who, presumably, had not been exposed to staphylococcal infections. The anti-MSCRAMM antibodies preferentially reacted with the ligand-binding repeat domain of the adhesin. However, these antibodies did not inhibit fibronectin binding. Essentially, all patients had antibodies which specifically recognized the fibronectin-MSCRAMM complex but not the isolated components. Epitopes recognized by these anti-ligand-induced binding sites antibodies were found in each repeat unit of the MSCRAMM. These results demonstrate that staphylococci have bound fibronectin some time during infection and that each repeat unit in the MSCRAMM can engage in ligand binding. Furthermore, our previously proposed model, suggesting that an unordered structure in the MSCRAMM undergoes a conformational change upon ligand binding (K. House-Pompeo, Y. Xu, D. Joh, P. Speziale, and M. Höök, J. Biol. Chem. 271:1379–1384, 1996), is presumably operational in patients during infections. PMID:9784554

  19. Functional Analysis of an S-Layer-Associated Fibronectin-Binding Protein in Lactobacillus acidophilus NCFM

    PubMed Central

    Hymes, Jeffrey P.; Johnson, Brant R.; Barrangou, Rodolphe

    2016-01-01

    Bacterial surface layers (S-layers) are crystalline arrays of self-assembling proteinaceous subunits called S-layer proteins (Slps) that comprise the outermost layer of the cell envelope. Many additional proteins that are associated with or embedded within the S-layer have been identified in Lactobacillus acidophilus NCFM, an S-layer-forming bacterium that is widely used in fermented dairy products and probiotic supplements. One putative S-layer-associated protein (SLAP), LBA0191, was predicted to mediate adhesion to fibronectin based on the in silico detection of a fibronectin-binding domain. Fibronectin is a major component of the extracellular matrix (ECM) of intestinal epithelial cells. Adhesion to intestinal epithelial cells is considered an important trait for probiotic microorganisms during transit and potential association with the intestinal mucosa. To investigate the functional role of LBA0191 (designated FbpB) in L. acidophilus NCFM, an fbpB-deficient strain was constructed. The L. acidophilus mutant with a deletion of fbpB lost the ability to adhere to mucin and fibronectin in vitro. Homologues of fbpB were identified in five additional putative S-layer-forming species, but no homologues were detected in species outside the L. acidophilus homology group. PMID:26921419

  20. Biochemical and immunological relationships among fibronectin-like proteins from different sea urchin species.

    PubMed

    Matranga, V; Zito, F; Tesoro, V; Yokota, Y; Nakano, E

    1995-08-01

    Fibronectin-like proteins were purified from ovaries of the sea urchin species, Paracentrotus lividus (PI), Sphaerechinus granularis (Sg), Arbacia lixula (Al), Pseudocentrotus depressus (Pd), and Anthocidaris crassispina (Ac), by gelatin-Sepharose affinity chromatography. The major component had a molecular mass of 180 kDa and was eluted by 1 M NaCl or 8 M urea, depending on the species used. By substrate adhesion assay, we tested the biological activity of the 180 kDa protein purified from Paracentrotus lividus (P1-180K) and showed that it promotes the adhesion of homologous embryonic cells to the substrate. An antiserum, developed against Temnopleurus hardwickii fibronectin-like protein (Th-180K), was used in Western blots of the proteins purified from the five species. The antibody cross-reacted with Pl-180K, Pd-180K and Ac-180K. A peptide map of P1-180K, obtained by V8 protease partial digestion, was compared with those obtained from the other four proteins and showed an homology between 40 and 56%. This report confirms that fibronectin-like proteins can be purified from sea urchins on the basis of their binding to gelatin-Sepharose; the proteins differ for their binding affinity to gelatin and share different epitopes, suggesting that they are members of a sea urchin fibronectin super family.

  1. Hormonal regulation of focal adhesions in bovine adrenocortical cells: induction of paxillin dephosphorylation by adrenocorticotropic hormone.

    PubMed Central

    Vilgrain, I; Chinn, A; Gaillard, I; Chambaz, E M; Feige, J J

    1998-01-01

    A study of bovine adrenocortical cell shape on adrenocorticotropic hormone (ACTH) challenge showed that the cells round up and develop arborized processes. This effect was found to be (1) specific for ACTH because angiotensin II and basic fibroblast growth factor have no effect; (2) mediated by a cAMP-dependent pathway because forskolin reproduces the effect of the hormone; (3) inhibited by sodium orthovanadate, a phosphotyrosine phosphatase inhibitor, but unchanged by okadaic acid, a serine/threonine phosphatase inhibitor; and (4) correlated with a complete loss of focal adhesions. Biochemical studies of the focal-adhesion-associated proteins showed that pp125fak, vinculin (110 kDa) and paxillin (70 kDa) were detected in the Triton X-100-insoluble fraction from adrenocortical cells. During cell adhesion on fibronectin as substratum, two major phosphotyrosine-containing proteins of molecular masses 125 and 68 kDa were immunodetected in the same fraction. A dramatic decrease in the extent of tyrosine phosphorylation of these proteins was observed within 60 min after treatment with ACTH. No change in pp125fak tyrosine phosphorylation nor in Src activity was detected. In contrast, paxillin was found to be tyrosine-dephosphorylated in a time-dependent manner in ACTH-treated cells. Sodium orthovanadate completely prevented the effect of ACTH. These observations suggest a possible role for phosphotyrosine phosphatases in hormone-dependent cellular regulatory processes. PMID:9601084

  2. Hormonal regulation of focal adhesions in bovine adrenocortical cells: induction of paxillin dephosphorylation by adrenocorticotropic hormone.

    PubMed

    Vilgrain, I; Chinn, A; Gaillard, I; Chambaz, E M; Feige, J J

    1998-06-01

    A study of bovine adrenocortical cell shape on adrenocorticotropic hormone (ACTH) challenge showed that the cells round up and develop arborized processes. This effect was found to be (1) specific for ACTH because angiotensin II and basic fibroblast growth factor have no effect; (2) mediated by a cAMP-dependent pathway because forskolin reproduces the effect of the hormone; (3) inhibited by sodium orthovanadate, a phosphotyrosine phosphatase inhibitor, but unchanged by okadaic acid, a serine/threonine phosphatase inhibitor; and (4) correlated with a complete loss of focal adhesions. Biochemical studies of the focal-adhesion-associated proteins showed that pp125fak, vinculin (110 kDa) and paxillin (70 kDa) were detected in the Triton X-100-insoluble fraction from adrenocortical cells. During cell adhesion on fibronectin as substratum, two major phosphotyrosine-containing proteins of molecular masses 125 and 68 kDa were immunodetected in the same fraction. A dramatic decrease in the extent of tyrosine phosphorylation of these proteins was observed within 60 min after treatment with ACTH. No change in pp125fak tyrosine phosphorylation nor in Src activity was detected. In contrast, paxillin was found to be tyrosine-dephosphorylated in a time-dependent manner in ACTH-treated cells. Sodium orthovanadate completely prevented the effect of ACTH. These observations suggest a possible role for phosphotyrosine phosphatases in hormone-dependent cellular regulatory processes.

  3. Measuring integrated cellular mechanical stress response at focal adhesions by optical tweezers

    NASA Astrophysics Data System (ADS)

    Bordeleau, François; Bessard, Judicael; Marceau, Normand; Sheng, Yunlong

    2011-09-01

    The ability of cells to sustain mechanical stress is largely modulated by the cytoskeleton. We present a new application of optical tweezers to study cell's mechanical properties. We trap a fibronectin-coated bead attached to an adherent H4II-EC3 rat hepatoma cell in order to apply the force to the cell surface membrane. The bead position corresponding to the cell's local mechanical response at focal adhesions is measured with a quadrant detector. We assessed the cell response by tracking the evolution of the equilibrium force for 40 cells selected at random and selected a temporal window to assess the cell initial force expression at focal adhesions. The mean value of the force within this time window over 40 randomly selected bead/cell bounds was 52.3 pN. Then, we assessed the responses of the cells with modulation of the cytoskeletons, namely the ubiquitous actin-microfilaments and microtubules, plus the differentiation-dependent keratin intermediate filaments. Notably, a destabilization of the first two networks led to around 50 and 30% reductions in the mean equilibrium forces, respectively, relative to untreated cells, whereas a loss of the third one yielded a 25% increase. The differences in the forces from untreated and treated cells are resolved by the optical tweezers experiment.

  4. GABAergic networks jump-start focal seizures

    PubMed Central

    de Curtis, Marco; Avoli, Massimo

    2016-01-01

    Summary Abnormally enhanced glutamatergic excitation is commonly believed to mark the onset of a focal seizure. This notion, however, is not supported by firm evidence, and it will be challenged here. A general reduction of unit firing has been indeed observed in association with low-voltage fast activity at the onset of seizures recorded during presurgical intracranial monitoring in patients with focal, drug-resistant epilepsies. Moreover, focal seizures in animal models start with increased γ-aminobutyric acid (GABA)ergic interneuronal activity that silences principal cells. In vitro studies have shown that synchronous activation of GABAA receptors occurs at seizure onset and causes sizeable elevations in extracellular potassium, thus facilitating neuronal recruitment and seizure progression. A paradoxical involvement of GABAergic networks is required for the initiation of focal seizures characterized by low-voltage fast activity, which represents the most common seizure-onset pattern in focal epilepsies. PMID:27061793

  5. Cell adhesion to fibronectin and tenascin: quantitative measurements of initial binding and subsequent strengthening response

    PubMed Central

    1989-01-01

    Cell-substratum adhesion strengths have been quantified using fibroblasts and glioma cells binding to two extracellular matrix proteins, fibronectin and tenascin. A centrifugal force-based adhesion assay was used for the adhesive strength measurements, and the corresponding morphology of the adhesions was visualized by interference reflection microscopy. The initial adhesions as measured at 4 degrees C were on the order of 10(-5)dynes/cell and did not involve the cytoskeleton. Adhesion to fibronectin after 15 min at 37 degrees C were more than an order of magnitude stronger; the strengthening response required cytoskeletal involvement. By contrast to the marked strengthening of adhesion to FN, adhesion to TN was unchanged or weakened after 15 min at 37 degrees C. The absolute strength of adhesion achieved varied according to protein and cell type. When a mixed substratum of fibronectin and tenascin was tested, the presence of tenascin was found to reduce the level of the strengthening of cell adhesion normally observed at 37 degrees C on a substratum of fibronectin alone. Parallel analysis of corresponding interference reflection micrographs showed that differences in the area of cell surface within 10-15 nm of the substratum correlated closely with each of the changes in adhesion observed: after incubation for 15 min on fibronectin at 37 degrees C, glioma cells increased their surface area within close contact to the substrate by integral to 125- fold. Cells on tenascin did not increase their surface area of contact. The increased surface area of contact and the inhibitory activity of cytochalasin b suggest that the adhesive "strengthening" in the 15 min after initial binding brings additional adhesion molecules into the adhesive site and couples the actin cytoskeleton to the adhesion complex. PMID:2477381

  6. Human fibronectin contains distinct adhesion- and motility-promoting domains for metastatic melanoma cells

    PubMed Central

    1986-01-01

    The active migration of tumor cells through extracellular matrices has been proposed to play a role in certain aspects of metastasis. Metastatic tumor cells migrate in vitro in response to substratum-bound adhesive glycoproteins such as fibronectin. The present studies use affinity-purified proteolytic fragments of fibronectin to determine the nature of adhesion- and/or motility-promoting domains within the protein. Two distinct fragments were identified with cell adhesion- promoting activities. By a number of criteria, the adhesive activity promoted by these two fragments was distinct. One fragment, a 75-kD tryptic fragment purified by monoclonal antibody chromatography, promoted the adhesion, spreading, and haptotactic motility of melanoma cells. Experiments using a synthetic cell attachment peptide in solution indicated that at least part of the attachment activity exhibited by the 75-kD fragment is mediated by the sequence arg-gly-asp- ser. It was not possible to demonstrate migration-stimulating activity using a small (11.5 kD) peptic fragment containing this sequence (Pierschbacher, M.D., E. G. Hayman, and E. Ruoslahti, 1981, Cell, 26:259-267) suggesting that another cell-binding activity within the 75 kD fragment distinct from arg-gly-asp-ser might be required for motility. The second fragment that stimulated melanoma adhesion was a 33-kD tryptic/catheptic carboxyl-terminal heparin-binding fragment, which is localized to the A chain of fibronectin. This fragment promotes adhesion and spreading but not the motility of these cells. Melanoma adhesion to this heparin-binding fragment was sensitive to the effects of cycloheximide, which contrasted adhesion to the haptotaxis- promoting fragment. Importantly, these studies illustrate that haptotaxis in response to fibronectin is not due to simple adhesion gradients of this protein. The results are discussed in light of a model for multiple distinct cell surface constituents mediating cell adhesion and motility on

  7. Focal brain inflammation and autism.

    PubMed

    Theoharides, Theoharis C; Asadi, Shahrzad; Patel, Arti B

    2013-04-09

    Increasing evidence indicates that brain inflammation is involved in the pathogenesis of neuropsychiatric diseases. Autism spectrum disorders (ASD) are characterized by social and learning disabilities that affect as many as 1/80 children in the USA. There is still no definitive pathogenesis or reliable biomarkers for ASD, thus significantly curtailing the development of effective therapies. Many children with ASD regress at about age 3 years, often after a specific event such as reaction to vaccination, infection, stress or trauma implying some epigenetic triggers, and may constitute a distinct phenotype. ASD children respond disproportionally to stress and are also affected by food and skin allergies. Corticotropin-releasing hormone (CRH) is secreted under stress and together with neurotensin (NT) stimulates mast cells and microglia resulting in focal brain inflammation and neurotoxicity. NT is significantly increased in serum of ASD children along with mitochondrial DNA (mtDNA). NT stimulates mast cell secretion of mtDNA that is misconstrued as an innate pathogen triggering an auto-inflammatory response. The phosphatase and tensin homolog (PTEN) gene mutation, associated with the higher risk of ASD, which leads to hyper-active mammalian target of rapamycin (mTOR) signalling that is crucial for cellular homeostasis. CRH, NT and environmental triggers could hyperstimulate the already activated mTOR, as well as stimulate mast cell and microglia activation and proliferation. The natural flavonoid luteolin inhibits mTOR, mast cells and microglia and could have a significant benefit in ASD.

  8. Focal liver lesions found incidentally

    PubMed Central

    Algarni, Abdullah A; Alshuhri, Abdullah H; Alonazi, Majed M; Mourad, Moustafa Mabrouk; Bramhall, Simon R

    2016-01-01

    Incidentally found focal liver lesions are a common finding and a reason for referral to hepatobiliary service. They are often discovered in patients with history of liver cirrhosis, colorectal cancer, incidentally during work up for abdominal pain or in a trauma setting. Specific points should considered during history taking such as risk factors of liver cirrhosis; hepatitis, alcohol consumption, substance exposure or use of oral contraceptive pills and metabolic syndromes. Full blood count, liver function test and tumor markers can act as a guide to minimize the differential diagnosis and to categorize the degree of liver disease. Imaging should start with B-mode ultrasound. If available, contrast enhanced ultrasound is a feasible, safe, cost effective option and increases the ability to reach a diagnosis. Contrast enhanced computed tomography should be considered next. It is more accurate in diagnosis and better to study anatomy for possible operation. Contrast enhanced magnetic resonance is the gold standard with the highest sensitivity. If doubt still remains, the options are biopsy or surgical excision. PMID:27028805

  9. Early vision and focal attention

    NASA Astrophysics Data System (ADS)

    Julesz, Bela

    1991-07-01

    At the thirty-year anniversary of the introduction of the technique of computer-generated random-dot stereograms and random-dot cinematograms into psychology, the impact of the technique on brain research and on the study of artificial intelligence is reviewed. The main finding-that stereoscopic depth perception (stereopsis), motion perception, and preattentive texture discrimination are basically bottom-up processes, which occur without the help of the top-down processes of cognition and semantic memory-greatly simplifies the study of these processes of early vision and permits the linking of human perception with monkey neurophysiology. Particularly interesting are the unexpected findings that stereopsis (assumed to be local) is a global process, while texture discrimination (assumed to be a global process, governed by statistics) is local, based on some conspicuous local features (textons). It is shown that the top-down process of "shape (depth) from shading" does not affect stereopsis, and some of the models of machine vision are evaluated. The asymmetry effect of human texture discrimination is discussed, together with recent nonlinear spatial filter models and a novel extension of the texton theory that can cope with the asymmetry problem. This didactic review attempts to introduce the physicist to the field of psychobiology and its problems-including metascientific problems of brain research, problems of scientific creativity, the state of artificial intelligence research (including connectionist neural networks) aimed at modeling brain activity, and the fundamental role of focal attention in mental events.

  10. Development and evaluation of a new kinetic assay for the quantitation of fibronectin gelatin-binding activity.

    PubMed

    Gelder, F B; Brown, S T; Moore, C A

    1985-10-01

    A new rapid and sensitive kinetic assay that measures the gelatin-binding activity of fibronectin has been developed. This assay is based on the rate of fibronectin-mediated aggregation of covalently coupled latex-gelatin particles. The addition of human plasma and serum resulted in aggregation rates proportional to the concentration of fibronectin in the test sample. This assay was inhibited by the addition of gelatin, demonstrating substrate specificity. This new assay requires 50 microliter of sample and can be performed within 5 minutes. Particle aggregation rates were affected by pH, heparin, and coupled gelatin concentration per milligram of latex. Maximum aggregation rates were observed at pH 8.0. Heparin was not an absolute requirement for particle aggregation but enhanced rates up to 1 U heparin/ml with little additive effect at greater concentrations. Heparin had a relatively greater effect on assays performed in acidic buffers. The concentration of gelatin per milligram of latex was rate limiting up to approximately 50 micrograms gelatin/mg latex with little change in aggregation rates at higher concentrations. Good correlation between total antigenic fibronectin (electroimmunoassay) and fibronectin gelatin-binding activity (latex-gelatin kinetic aggregation assay) was demonstrated in plasma from normal blood donors. This new assay will allow further definition of the relationship between fibronectin gelatin-binding activity and antigenic fibronectin in normal and pathophysiologic states.

  11. Release of transforming growth factor-beta (TGF-β) and fibronectin by alveolar macrophages in airway diseases

    PubMed Central

    VIGNOLA, A M; CHANEZ, P; CHIAPPARA, G; MERENDINO, A; ZINNANTI, E; BOUSQUET, J; BELLIA, V; BONSIGNORE, G

    1996-01-01

    Asthma and chronic bronchitis are associated with airway remodelling, and airway macrophages are present in bronchial inflammation. TGF-β and fibronectin released by alveolar macrophages possess a fibrogenic potency. The potential role of alveolar macrophages in airway remodelling was studied in asthma and chronic bronchitis by the release of TGF-β and fibronectin. Alveolar macrophages were isolated by bronchoalveolar lavage in 14 control subjects, 14 asthmatics and 14 chronic bronchitics. The spontaneous and lipopolysaccharide (LPS)- or concanavalin A (Con A)-induced release of TGF-β and fibronectin was measured by ELISA. Alveolar macrophages from chronic bronchitics spontaneously release greater amounts of TGF-β and fibronectin than those from asthmatic and control subjects. Alveolar macrophages from asthmatics release greater amounts of TGF-β and fibronectin than those from control subjects. The spontaneous release of TGF-β is significantly correlated with that of fibronectin. Fibronectin release was significantly reduced after LPS stimulation, and TGF-β release was significantly increased after LPS stimulation, except in chronic bronchitis patients. Con A increased the release of TGF-β in cells from normal subjects. This study suggests that activated macrophages play a role in airway remodelling in chronic bronchitis and to a lesser extent in asthma. PMID:8870708

  12. Human osteosarcoma cells resistant to detachment by an Arg-Gly-Asp- containing peptide overproduce the fibronectin receptor

    PubMed Central

    1987-01-01

    MG-63 human osteosarcoma cells were selected for attachment and growth in the presence of increasing concentrations of a synthetic peptide containing the cell attachment-promoting Arg-Gly-Asp sequence derived from the cell-binding region of fibronectin. Cells capable of attachment and growth in 5-mM concentrations of a peptide having the sequence Gly-Arg-Gly-Asp-Ser-Pro overproduce the cell surface receptor for fibronectin. In contrast, these cells show no differences in the numbers of vitronectin receptor they express as compared with the parental MG-63 cells. In agreement with the resistance of the selected cells to detachment by the peptide, 25-fold more Arg-Gly-Asp-containing peptide is required to prevent the attachment of these cells to fibronectin-coated surfaces than is needed to inhibit the attachment of MG-63 cells to the same substrate. However, similar concentrations of this peptide inhibit attachment of both cell lines to vitronectin- coated surfaces. The increase in fibronectin receptor is due to an increase in the levels of mRNA encoding the fibronectin receptor. Because of the nature of the selection process, we reasoned that this increase might be due to amplification of the fibronectin receptor gene, but no increase in gene copy number was detected by Southern blot analysis. The peptide-resistant cells display a very different morphology from that of the MG-63 cells, one that has a greater resemblance to that of osteocytes. The resistant cells also grow much more slowly than the MG-63 cells. The increased fibronectin receptor and altered morphology and growth properties were stable for at least 3 mo in the absence of peptide. The enhanced expression of the fibronectin receptor on the resistant cells indicates that cells are capable of altering the amount of fibronectin receptor on their surface in response to environmental factors and that this may in turn affect the phenotypic properties of the cell. PMID:2443508

  13. Cleavage of extracellular matrix in periodontitis: gingipains differentially affect cell adhesion activities of fibronectin and tenascin-C

    PubMed Central

    Ruggiero, Sabrina; Cosgarea, Raluca; Potempa, Jan; Potempa, Barbara; Eick, Sigrun; Chiquet, Matthias

    2014-01-01

    Gingipains are cysteine proteases that represent major virulence factors of the periodontopathogenic bacterium Porphyromonas gingivalis. Gingipains are reported to degrade extracellular matrix (ECM) of periodontal tissues, leading to tissue destruction and apoptosis. The exact mechanism is not known, however. Fibronectin and tenascin-C are pericellular ECM glycoproteins present in periodontal tissues. Whereas fibronectin mediates fibroblast adhesion, tenascin-C binds to fibronectin and inhibits its cell-spreading activity. Using purified proteins in vitro, we asked whether fibronectin and tenascin-C are cleaved by gingipains at clinically relevant concentrations, and how fragmentation by the bacterial proteases affects their biological activity in cell adhesion. Fibronectin was cleaved into distinct fragments by all three gingipains; however, only arginine-specific HRgpA and RgpB but not lysine-specific Kgp destroyed its cell-spreading activity. This result was confirmed with recombinant cell-binding domain of fibronectin. Of the two major tenascin-C splice variants, the large but not the small was a substrate for gingipains, indicating that cleavage occurred primarily in the alternatively spliced domain. Surprisingly, cleavage of large tenascin-C variant by all three gingipains generated fragments with increased anti-adhesive activity towards intact fibronectin. Fibronectin and tenascin-C fragments were detected in gingival crevicular fluid of a subset of periodontitis patients. We conclude that cleavage by gingipains directly affects the biological activity of both fibronectin and tenascin-C in a manner that might lead to increased cell detachment and loss during periodontal disease. PMID:23313574

  14. Production of Fibronectin by the Human Alveolar Macrophage: Mechanism for the Recruitment of Fibroblasts to Sites of Tissue Injury in Interstitial Lung Diseases

    NASA Astrophysics Data System (ADS)

    Rennard, Stephen I.; Hunninghake, Gary W.; Bitterman, Peter B.; Crystal, Ronald G.

    1981-11-01

    Because cells of the mononuclear phagocyte system are known to produce fibronectin and because alveolar macrophages are activated in many interstitial lung diseases, the present study was designed to evaluate a role for the alveolar macrophage as a source of the increased levels of fibronectin found in the lower respiratory tract in interstitial lung diseases and to determine if such fibronectin might contribute to the development of the fibrosis found in these disorders by being a chemoattractant for human lung fibroblasts. Production of fibronectin by human alveolar macrophages obtained by bronchoalveolar lavage and maintained in short-term culture in serum-free conditions was demonstrated; de novo synthesis was confirmed by the incorporation of [14C]proline. This fibronectin had a monomer molecular weight of 220,000 and was antigenically similar to plasma fibronectin. Macrophages from patients with idiopathic pulmonary fibrosis produced fibronectin at a rate 20 times higher than did normal macrophages; macrophages from patients with pulmonary sarcoidosis produced fibronectin at 10 times the normal rate. Macrophages from 6 of 10 patients with various other interstitial disorders produced fibronectin at rates greater than the rate of highest normal control. Human alveolar macrophage fibronectin was chemotactic for human lung fibroblasts, suggesting a functional role for this fibronectin in the derangement of the alveolar structures that is characteristic of these disorders.

  15. Genetics Home Reference: familial focal epilepsy with variable foci

    MedlinePlus

    ... familial focal epilepsy with variable foci familial focal epilepsy with variable foci Enable Javascript to view the ... PDF Open All Close All Description Familial focal epilepsy with variable foci (FFEVF) is an uncommon form ...

  16. Mosaic focal plane for star sensors

    NASA Astrophysics Data System (ADS)

    Chang, N. C.

    1981-02-01

    The basic principles of star sensors are reviewed with reference to the advantages of replacing photodiodes, image dissectors, and vidicons with mosaic charge transfer device (CTD) focal planes. The desirable characteristics of CTD focal planes include: high uniformity, high transfer effect, low dark current, low hot and cold spots, low dead space, low angular misalignment, high coplanarity, and high thermal stability. An implementation of a mosaic CTD array star sensor which achieves high angular position accuracy and frequency attitude update is presented. Two focal plane packaging concepts, the planar and vertical board packagings, are examined.

  17. Unilateral Isolated Proximal Femoral Focal Deficiency

    PubMed Central

    Doğer, Emek; Köpük, Şule Y.; Çakıroğlu, Yiğit; Çakır, Özgür; Yücesoy, Gülseren

    2013-01-01

    Objective. To discuss a patient with a prenatal diagnosis of unilateral isolated femoral focal deficiency. Case. Antenatal diagnosis of unilateral isolated femoral focal deficiency was made at 20 weeks of gestation. The length of left femur was shorter than the right, and fetal femur length was below the fifth percentile. Proximal femoral focal deficiency was diagnosed. After delivery, the diagnosis was confirmed with skeletal radiographs and magnetic resonance imaging. In prenatal ultrasonographic examination, the early recognition and exclusion of skeletal dysplasias is important; moreover, treatment plans should be initiated, and valuable information should be provided to the family. PMID:23984135

  18. A Review of Focal Reducer Interferometer Systems

    DTIC Science & Technology

    1986-06-01

    searches. G.Courtes has also proposed a design for a focal reducer for the E.S.O. 3.6m. Ritchey -Chretien telescope (Fig. 4b). He uses a Schmidt camera...spaced components, with a modified Wynne camera design (Fig. 7). The instrument proposed used folded optics and would operate at the F/8 Ritchey Cretien ...G.Courtes for the 3.6m Ritchey Chretien telescope . a, telescope focal plane and field lens; b, collimator; c, camera. Figure 5 The focal reducer of A.B

  19. Do focal colors look particularly "colorful"?

    PubMed

    Witzel, Christoph; Franklin, Anna

    2014-04-01

    If the most typical red, yellow, green, and blue were particularly colorful (i.e., saturated), they would "jump out to the eye." This would explain why even fundamentally different languages have distinct color terms for these focal colors, and why unique hues play a prominent role in subjective color appearance. In this study, the subjective saturation of 10 colors around each of these focal colors was measured through a pairwise matching task. Results show that subjective saturation changes systematically across hues in a way that is strongly correlated to the visual gamut, and exponentially related to sensitivity but not to focal colors.

  20. [The influence of immobilized fibronectin on karyotypic variability of two rat kangaroo kidney cell lines].

    PubMed

    Polianskaia, G G; Goriachaia, T S; Pinaev, G P

    2007-01-01

    The numerical and structural karyotypic variability has been investigated in "markerless" Rat kangaroo kidney cell lines NBL-3-17 and NBL-3-11 when cultivating on a fibronectin-coated surface. In cell line NBL-3-17, cultivated on the fibronectin-coated surface for 1, 2, 4 and 8 days, the character of cell distribution for the chromosome number has changed. These changes involve a significant decrease in frequency of cells with modal number of chromosomes, and an increase in frequency of cells with lower chromosomal number. Many new additional structural variants of the karyotype (SVK) appear. The observed alterations seem to be due preference adhesion of cells with lower chromosome number, disturbances of mitotic apparatus and selection of SVK, which are more adopted to changes in culture conditions. Detachment of cells from the fibronectin-coated surface, followed by 5 days cultivation on a hydrophilic surface restored control distribution. In cell line NBL-3-11, cultivated on the fibronectin-coated surface for 1, 2, 4 and 8 days, the character of numerical karyotypic variability did not change compared to control variants. In cell line NBL-3-17 the frequency of chromosomal aberrations under cultivation on the fibronectin-coated surface for 1, 2, 4 and 8 days did not change relative to control variants. In cell line NBL-3-11 the frequency of chromosomal aberrations under the same conditions significantly increases, mainly at the expence of chromosomal, chromatid breaks and dicentrics (telomeric association) relative to control variants. We discuss possible reasons of differences in the character of numerical and structural karyotypic variability between cell lines NBL-3-17 (hypotriploid) and NBL-3-11 (hypodiploid) under cultivation on fibronectin. The reasons of the observed interline karyotypic differences possibly consist in peculiarity of karyotypic structure of cell line NBL-3-11 and in the change of gene expression, namely in a dose of certain functioning

  1. Coaxial Redundant Drives

    NASA Technical Reports Server (NTRS)

    Brissette, R.

    1983-01-01

    Harmonic drives allow redundancy and high out put torque in small package. If main drive fails, standby drive takes over and produces torque along same axis as main drive. Uses include power units in robot for internal pipeline inspection, manipulators in deep submersible probes or other applications in which redundancy protects against costly failures.

  2. Focal myositis presenting with radial nerve palsy.

    PubMed

    Alzagatiti, B I; Bertorini, T E; Horner, L H; Maccarino, V S; O'Brien, T

    1999-07-01

    Focal myositis is a rare inflammatory pseudotumor of skeletal muscle which usually has a benign course. We report a 56-year-old woman with a painful mass in the left arm with a radial nerve palsy. Magnetic resonance imaging (MRI) of the left arm showed a mass in the triceps muscle that was suggestive of a soft-tissue sarcoma. Electromyography showed a severe radial neuropathy involving both motor and sensory axons. An open biopsy showed focal myositis. Treatment with corticosteroids resulted in complete disappearance of the mass clinically and by MRI, without recurrence for more than 2 years. Radial nerve function also recovered completely. As a treatable cause of focal neuropathy, focal myositis should be included in the differential diagnosis of a muscle mass. Copyright 1999 John Wiley & Sons, Inc.

  3. An uncommon focal epithelial hyperplasia manifestation.

    PubMed

    dos Santos-Pinto, Lourdes; Giro, Elisa Maria Aparecida; Pansani, Cyneu Aguiar; Ferrari, Junia; Massucato, Elaine Maria Sgavioli; Spolidório, Luis Carlos

    2009-01-01

    Focal epithelial hyperplasia is a rare, contagious disease associated with infection of the oral mucosa by human papillomavirus types 13 or 32, characterized by multiple soft papules of the same color as the adjacent normal mucosa. It mainly affects the lower lip, buccal mucosa, and tongue. The purpose of this case report was to describe a rare verrucal lesion located in the upper gingiva that was clinically and histologically consistent with focal epithelial hyperplasia.

  4. Finding the Focal Axes of Offset Antennas

    NASA Technical Reports Server (NTRS)

    Schmidt, R. F.

    1982-01-01

    Focal axis of offset paraboloidal reflector antennas determined by direct measurement instead of trial and error. Two feed horns transmit sum or difference pattern to antenna under test, which reflects energy to far-field detector. When axis of feed horns coincides with focal axis of antenna reflector, far-field detector records minimum in amplitude difference and maximum in absolute-magnitude phase difference between sum and difference signals.

  5. Fibronectin-synthesizing activity of free and membrane-bound polyribosomes from human embryonic fibroblasts and chick embryos

    SciTech Connect

    Belkin, V.M.; Volodarskaya, S.M.

    1986-06-20

    The fibronectin-synthesizing activity of membrane-bound and free polyribosomes in a cell-free system was studied using immunochemical methods. It was found that fibronectin biosynthesis on membrane-bound polyribosomes from human embryonic fibroblasts accounts for 4.9% and those from 10-day-old chick embryos for 1.1% of the total amount of newly synthesized proteins, whereas on free polyribosomes it is 1.0 and 0.3%, respectively. Fibronectin monomers with a molecular weight of 220,000 were found only in the material of the cell-free system containing heavy fractions of membrane-bound polyribosomes newly synthesized in the presence of spermidine. Thus, it was shown that fibronectin is synthesized primarily on membrane-bound polyribosomes.

  6. Cooperative signaling by alpha 5 beta 1 and alpha 4 beta 1 integrins regulates metalloproteinase gene expression in fibroblasts adhering to fibronectin

    PubMed Central

    1995-01-01

    Rabbit synovial fibroblasts (RSF) express basal levels of the metalloproteinases (MMP) collagenase, stromelysin-1 and 92-kD gelatinase when plated on intact fibronectin (FN), but elevated levels when plated on either the central RGD-containing cell-binding region of FN (120FN) or antibody against the alpha 5 beta 1 integrin, suggesting that domains outside 120FN may suppress the induction of MMP (Werb, Z., P. M. Tremble, O. Behrendtsen, E. Crowley, and C.H. Damsky. 1989. J. Cell Biol. 109:877-889). We therefore attempted to reconstitute the basal signaling of intact FN by plating RSF on 120FN together with domains of FN outside this region. Large COOH-terminal fragments containing both the heparin-binding and HICS domains suppressed MMP when combined with 120FN. To map the active sequences, peptides from this region and larger fragments that did, or did not, include the CS-1 portion of IIICS were tested. Only CS-1 peptide, or larger fragments containing CS-1, suppressed MMP expression induced by 120FN. In contrast, peptide V from the heparin-binding region, shown previously to stimulate focal contact formation, further enhanced MMP expression by RSF when present on the substrate with 120FN. RSF expressed alpha 4 beta 1 integrin, the receptor for CS-1, and the anti-alpha 4 mAb blocked the ability of CS-1 to suppress MMP induction by 120FN. These results show that signals modulating MMP expression and focal contact assembly are regulated independently, and that cooperative signaling by alpha 5 beta 1 and alpha 4 beta 1 integrins plays a dominant role in regulating expression of these extracellular matrix-remodeling genes in response to FN. This work demonstrates directly the modular way in which information in the extracellular matrix is detected and processed by cell surface receptors. PMID:7537277

  7. Folding of beta-sandwich proteins: three-state transition of a fibronectin type III module.

    PubMed Central

    Cota, E.; Clarke, J.

    2000-01-01

    An analysis of the folding of the 94 residue tenth fibronectin type III (fnIII) domain of human fibronectin (FNfn10) is presented. Use of guanidine isothiocyanate as a denaturant allows us to obtain equilibrium and kinetic data across a broad range of denaturant concentrations that are unavailable in guanidine hydrochloride. Equilibrium unfolding experiments show that FNfn10 is significantly more stable than has been reported previously. Comparison of equilibrium and kinetic parameters reveals the presence of an intermediate that accumulates at low denaturant concentrations. This is the first demonstration of three-state folding kinetics for a fnIII domain. We have previously shown that a homologous domain from human tenascin (TNfn3) folds by a two-state mechanism, but this does not necessarily indicate that the two proteins fold by different folding pathways. PMID:10739253

  8. Identification of Surface Proteins from Lactobacillus casei BL23 Able to Bind Fibronectin and Collagen.

    PubMed

    Muñoz-Provencio, Diego; Pérez-Martínez, Gaspar; Monedero, Vicente

    2011-03-01

    Strains of lactobacilli show the capacity to attach to extracellular matrix proteins. Cell-wall fractions of Lactobacillus casei BL23 enriched in fibronectin, and collagen-binding proteins were isolated. Mass spectrometry analysis of their protein content revealed the presence of stress-related proteins (GroEL, ClpL), translational elongation factors (EF-Tu, EF-G), oligopeptide solute-binding proteins, and the glycolytic enzymes enolase and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The latter two enzymes were expressed in Escherichia coli and purified as glutathione-S-transferase (GST) fusion proteins, and their in vitro binding activity to fibronectin and collagen was confirmed. These results reinforce the idea that lactobacilli display on their surfaces a variety of moonlighting proteins that can be important in their adaptation to survive at intestinal mucosal sites and in the interaction with host cells.

  9. Sighting optics including an optical element having a first focal length and a second focal length

    DOEpatents

    Crandall, David Lynn [Idaho Falls, ID

    2011-08-01

    One embodiment of sighting optics according to the teachings provided herein may include a front sight and a rear sight positioned in spaced-apart relation. The rear sight includes an optical element having a first focal length and a second focal length. The first focal length is selected so that it is about equal to a distance separating the optical element and the front sight and the second focal length is selected so that it is about equal to a target distance. The optical element thus brings into simultaneous focus, for a user, images of the front sight and the target.

  10. Surface oxide net charge of a titanium alloy: modulation of fibronectin-activated attachment and spreading of osteogenic cells.

    PubMed

    Rapuano, Bruce E; MacDonald, Daniel E

    2011-01-01

    In the current study, we have altered the surface oxide properties of a Ti6Al4V alloy using heat treatment or radiofrequency glow discharge (RFGD) in order to evaluate the relationship between the physico-chemical and biological properties of the alloy's surface oxide. The effects of surface pretreatments on the attachment of cells from two osteogenic cell lines (MG63 and MC3T3) and a mesenchymal stem cell line (C3H10T1/2) to fibronectin adsorbed to the alloy were measured. Both heat and RFGD pretreatments produced a several-fold increase in the number of cells that attached to fibronectin adsorbed to the alloy at a range of coating concentrations (0.001-10nM FN) for each cell line tested. An antibody (HFN7.1) directed against the central integrin binding domain of fibronectin produced a 65-70% inhibition of cell attachment to fibronectin-coated disks, indicating that cell attachment to the metal discs was dependent on fibronectin binding to cell integrin receptors. Both treatments also accelerated the cell spreading response manifested by extensive flattening and an increase in mean cellular area. The treatment-induced increases in the cell attachment activity of adsorbed fibronectin were correlated with previously demonstrated increases in Ti6Al4V oxide negative net surface charge at physiological pH produced by both heat and RFGD pretreatments. Since neither treatment increased the adsorption mass of fibronectin, these findings suggest that negatively charged surface oxide functional groups in Ti6Al4V can modulate fibronectin's integrin receptor activity by altering the adsorbed protein's conformation. Our results further suggest that negatively charged functional groups in the surface oxide can play a prominent role in the osseointegration of metallic implant materials.

  11. Anti-citrullinated fibronectin antibodies in rheumatoid arthritis are associated with human leukocyte antigen-DRB1 shared epitope alleles

    PubMed Central

    2012-01-01

    Introduction Fibronectin is one of the most abundant proteins present in the inflamed joint. Here, we characterized the citrullination of fibronectin in the joints of rheumatoid arthritis (RA) patients and studied the prevalence, epitope specificity and human leukocyte antigen (HLA) association of autoantibodies against citrullinated fibronectin in RA. Methods Citrullinated residues in fibronectin isolated from RA patient synovial fluid were identified by mass spectrometry. The corresponding citrullinated and non-citrullinated peptides were synthesized and used to analyze the presence of autoantibodies to these peptides in RA sera and sera from other diseases and healthy controls by ELISA. The data were compared with risk factors like shared epitope HLA alleles and smoking, and with clinical features. Results Five citrullinated residues were identified in fibronectin from RA synovial fluid. RA sera reacted in a citrulline-dependent manner with two out of four citrullinated fibronectin peptides, one of which contains two adjacent citrulline residues, in contrast to non-RA sera, which were not reactive. The most frequently recognized peptide (FN-Cit1035,1036, LTVGLTXXGQPRQY, in which × represents citrulline) was primarily targeted by anti-CCP (cyclic citrullinated peptide) 2-positive RA patients. Anti-FN-Cit1035,1036 autoantibodies were detected in 50% of established anti-CCP2-positive RA patients and in 45% of such patients from a early arthritis clinic. These antibodies appeared to be predominantly of the immunoglobulin G (IgG) isotype and to be associated with HLA shared epitope alleles (odds ratio = 2.11). Conclusions Fibronectin in the inflamed synovia of RA patients can be citrullinated at least at five positions. Together with the flanking amino acids, three of these citrullinated residues comprise two epitopes recognized by RA autoantibodies. Anti-citrullinated fibronectin peptide antibodies are associated with HLA shared epitope alleles. PMID:22339947

  12. Fibronectin-binding protein of Streptococcus pyogenes: sequence of the binding domain involved in adherence of streptococci to epithelial cells.

    PubMed Central

    Talay, S R; Valentin-Weigand, P; Jerlström, P G; Timmis, K N; Chhatwal, G S

    1992-01-01

    The sequence of the fibronectin-binding domain of the fibronectin-binding protein of Streptococcus pyogenes (Sfb protein) was determined, and its role in streptococcal adherence was investigated by use of an Sfb fusion protein in adherence studies. A 1-kb DNA fragment coding for the binding domain of Sfb protein was cloned into the expression vector pEX31 to produce an Sfb fusion protein consisting of the N-terminal part of MS2 polymerase and a C-terminal fragment of the streptococcal protein. Induction of the vector promoter resulted in hyperexpression of fibronectin-binding fusion protein in the cytoplasm of the recombinant Escherichia coli cells. Sequence determination of the cloned 1-kb fragment revealed an in-frame reading frame for a 268-amino-acid peptide composed of a 37-amino-acid sequence which is completely repeated three times and incompletely repeated a fourth time. Cloning of one repeat into pEX31 resulted in expression of small fusion peptides that show fibronectin-binding activity, indicating that one repeat contains at least one binding domain. Each repeat exhibits two charged domains and shows high homology with the 38-amino-acid D3 repeat of the fibronectin-binding protein of Staphylococcus aureus. Sequence comparison with other streptococcal ligand-binding surface proteins, including M protein, failed to reveal significant homology, which suggests that Sfb protein represents a novel type of functional protein in S. pyogenes. The Sfb fusion protein isolated from the cytoplasm of recombinant cells was purified by fast protein liquid chromatography. It showed a strong competitive inhibition of fibronectin binding to S. pyogenes and of the adherence of bacteria to cultured epithelial cells. In contrast, purified streptococcal lipoteichoic acid showed only a weak inhibition of fibronectin binding and streptococcal adherence. These results demonstrate that Sfb protein is directly involved in the fibronectin-mediated adherence of S. pyogenes to

  13. Fibronectin and vitronectin promote human fetal osteoblast cell attachment and proliferation on nanoporous titanium surfaces

    PubMed Central

    Rivera-Chacon, D. M.; Alvarado-Velez, M.; Acevedo-Morantes, C.Y.; Singh, S.P.; Gultepe, E.; Nagesha, D.; Sridhar, S.; Ramirez-Vick, J.E.

    2013-01-01

    Improvements in osteoconduction of implant biomaterials require focusing on the bone-implant interface, which is a complex multifactorial system. Surface topography of implants plays a crucial role at this interface. Nanostructured surfaces have been shown to promote serum protein adsorption and osteoblast adhesion when compared to microstructured surfaces for bone-implant materials. We studied the influence of the serum proteins fibronectin and vitronectin on the attachment and proliferation of osteoblasts onto nanostructured titania surfaces. Human fetal osteoblastic cells hFOB 1.19 were used as model osteoblasts and were grown on nanoporous TiO2 templates, using Ti6Al4V and commercially pure Ti substrates as controls. Results show a significant increase in cell proliferation on nanoporous TiO2 over flat substrates. Initial cell attachment data exhibited a significant effect by either fibronectin or vitronectin on cell adhesion at the surface of any of the tested materials. In addition, the extent of cell adhesion was significantly different between the nanoporous TiO2 and both Ti6Al4V and commercially pure Ti substrates, with the first showing the highest surface coverage. There was no significant difference on osteoblast attachment or proliferation between the presence of fibronectin or vitronectin using any of the material substrates. Taken together, these results suggest that the increase in osteoblast attachment and proliferation shown on the nanoporous TiO2 is due to an increase in the adsorption of fibronectin and vitronectin because of the higher surface area and to an enhanced protein unfolding, which allows access to osteoblast binding motifs within these proteins. PMID:23858975

  14. Pigment epithelium-derived factor (PEDF) inhibits breast cancer metastasis by down-regulating fibronectin.

    PubMed

    Hong, Honghai; Zhou, Ti; Fang, Shuhuan; Jia, Minghan; Xu, Zumin; Dai, Zhiyu; Li, Cen; Li, Shuai; Li, Lei; Zhang, Ting; Qi, Weiwei; Bardeesi, Adham Sameer A; Yang, Zhonghan; Cai, Weibin; Yang, Xia; Gao, Guoquan

    2014-11-01

    Pigment epithelium-derived factor (PEDF) plays an important role in the tumor growth and metastasis inhibition. It has been reported that PEDF expression is significantly reduced in breast cancer, and associated with disease progression and poor patient outcome. However, the exact mechanism of PEDF on breast cancer metastasis including liver and lung metastasis remains unclear. The present study aims to reveal the impact of PEDF on breast cancer. The orthotopic tumor mice model inoculated by MDA-MB-231 cells stably expressing PEDF or control cells was used to assess liver and lung metastasis of breast cancer. In vitro, migration and invasion experiments were used to detect the metastatic abilities of MDA-MB-231 and SKBR3 breast cancer cells with or without overexpression of PEDF. The metastatic-related molecules including EMT makers, fibronectin, and p-AKT and p-ERK were detected by qRT-PCR, Western blot, and Fluorescent immunocytochemistry. PEDF significantly inhibited breast cancer growth and metastasis in vivo and in vitro. Mechanically, PEDF inhibited breast cancer cell migration and invasion by down-regulating fibronectin and subsequent MMP2/MMP9 reduction via p-ERK and p-AKT signaling pathways. However, PEDF had no effect on EMT conversion in the breast cancer cells which was usually involved in cancer metastasis. Furthermore, the study showed that laminin receptor mediated the down-regulation of fibronectin by PEDF. These results reported for the first time that PEDF inhibited breast cancer metastasis by down-regulating fibronectin via laminin receptor/AKT/ERK pathway. Our findings demonstrated PEDF as a dual effector in limiting breast cancer growth and metastasis and highlighted a new avenue to block breast cancer progression.

  15. Synthetic immunomodulatory peptide IDR-1002 enhances monocyte migration and adhesion on fibronectin.

    PubMed

    Madera, Laurence; Hancock, Robert E W

    2012-01-01

    Regulation of the immune system by immunomodulatory agents, such as the synthetic innate defense regulator (IDR) peptides, has been proposed as a potential strategy to strengthen host immune responses against infection. IDR peptides confer protection in vivo against a range of bacterial infections and have been developed as components of single-dose vaccine adjuvants due to their ability to modulate innate immunity, correlating with an increased recruitment of monocytes to sites of infection or immunization. However, the mechanisms by which IDR peptides augment monocyte recruitment remain poorly defined. Anti-infective peptide IDR-1002 was demonstrated here to lack direct monocyte chemoattractive activity yet enhance, by up to 5-fold, the ability of human monocytes to migrate on fibronectin towards chemokines. This effect correlated with an increased adhesion of monocytes and THP-1 cells to fibronectin by IDR-1002 and other IDR peptides and the adhesion of THP-1 cells to fibronectin occurred in a β(1)-integrin-dependent manner, corresponding with an increased activation of β(1)-integrins and the phosphoinositide 3-kinase (PI3K)-Akt pathway. PI3K- and Akt-specific inhibitors abrogated IDR-1002-induced adhesion and activation of β(1)-integrins, whereas p38 and MEK1 inhibitors did not affect, or moderately inhibited, adhesion, respectively. Furthermore, IDR-1002 enhancement of monocyte migration towards chemokines and activation of β(1)-integrins was abrogated in the presence of PI3K- and Akt-specific inhibitors. In summary, IDR-1002 enhanced monocyte migration on fibronectin through promotion of β(1)-integrin-mediated interactions regulated by the PI3K-Akt pathway, revealing a mechanism by which IDR-1002 promotes monocyte recruitment. Copyright © 2012 S. Karger AG, Basel.

  16. Inhibition of Hyaluronan Synthesis Reduces Versican and Fibronectin Levels in Trabecular Meshwork Cells

    PubMed Central

    Keller, Kate E.; Sun, Ying Ying; Vranka, Janice A.; Hayashi, Lauren; Acott, Ted S.

    2012-01-01

    Hyaluronan (HA) is a major component of the extracellular matrix (ECM) and is synthesized by three HA synthases (HAS). Similarities between the HAS2 knockout mouse and the hdf mutant mouse, which has a mutation in the versican gene, suggest that HA and versican expression may be linked. In this study, the relationship between HA synthesis and levels of versican, fibronectin and several other ECM components in trabecular meshwork cells from the anterior segment of the eye was investigated. HA synthesis was inhibited using 4-methylumbelliferone (4MU), or reduced by RNAi silencing of each individual HAS gene. Quantitative RT-PCR and immunoblotting demonstrated a reduction in mRNA and protein levels of versican and fibronectin. Hyaluronidase treatment also reduced versican and fibronectin levels. These effects could not be reversed by addition of excess glucose or glucosamine or exogenous HA to the culture medium. CD44, tenascin C and fibrillin-1 mRNA levels were reduced by 4MU treatment, but SPARC and CSPG6 mRNA levels were unaffected. Immunostaining of trabecular meshwork tissue after exposure to 4MU showed an altered localization pattern of HA-binding protein, versican and fibronectin. Reduction of versican by RNAi silencing did not affect HA concentration as assessed by ELISA. Together, these data imply that HA concentration affects synthesis of certain ECM components. Since precise regulation of the trabecular meshwork ECM composition and organization is required to maintain the aqueous humor outflow resistance and intraocular pressure homeostasis in the eye, coordinated coupling of HA levels and several of its ECM binding partners should facilitate this process. PMID:23139787

  17. Ras modulation of superoxide activates ERK-dependent fibronectin expression in diabetes-induced renal injuries.

    PubMed

    Lin, C-L; Wang, F-S; Kuo, Y-R; Huang, Y-T; Huang, H-C; Sun, Y-C; Kuo, Y-H

    2006-05-01

    Although previous studies have demonstrated that diabetic nephropathy is attributable to early extracellular matrix accumulation in glomerular mesangial cells, the molecular mechanism by which high glucose induces matrix protein deposition remains not fully elucidated. Rat mesangial cells pretreated with or without inhibitors were cultured in high-glucose or advanced glycation end product (AGE) conditions. Streptozotocin-induced diabetic rats were given superoxide dismutase (SOD)-conjugated propylene glycol to scavenge superoxide. Transforming growth factor (TGF)-beta1, fibronectin expression, Ras, ERK, p38, and c-Jun activation of glomerular mesangial cells or urinary albumin secretion were assessed. Superoxide, not nitric oxide or hydrogen peroxide, mediated high glucose- and AGE-induced TGF-beta1 and fibronectin expression. Pretreatment with diphenyliodonium, not allopurinol or rotenone, reduced high-glucose and AGE augmentation of superoxide synthesis and fibronection expression. High glucose and AGEs rapidly enhanced Ras activation and progressively increased cytosolic ERK and nuclear c-Jun activation. Inhibiting Ras by manumycin A reduced the stimulatory effects of high glucose and AGEs on superoxide and fibronectin expression. SOD or PD98059 pretreatment reduced high-glucose and AGE promotion of ERK and c-Jun activation. Exogenous SOD treatment in diabetic rats significantly attenuated diabetes induction of superoxide, urinary albumin excretion, 8-hydroxy-2'-deoxyguanosine, TGF-beta1, and fibronectin immunoreactivities in renal glomerular mesangial cells. Ras induction of superoxide activated ERK-dependent fibrosis-stimulatory factor and extracellular matrix gene transcription of mesangial cells. Reduction of oxidative stress by scavenging superoxide may provide an alternative strategy for controlling diabetes-induced early renal injury.

  18. Structural and dynamical characterization of a biologically active unfolded fibronectin-binding protein from Staphylococcus aureus.

    PubMed

    Penkett, C J; Redfield, C; Jones, J A; Dodd, I; Hubbard, J; Smith, R A; Smith, L J; Dobson, C M

    1998-12-01

    A 130-residue fragment (D1-D4) taken from a fibronectin-binding protein of Staphylococcus aureus, which contains four fibronectin-binding repeats and is unfolded but biologically active at neutral pH, has been studied extensively by NMR spectroscopy. Using heteronuclear multidimensional techniques, the conformational properties of D1-D4 have been defined at both a global and a local level. Diffusion studies give an average effective radius of 26.2 +/- 0.1 A, approximately 75% larger than that expected for a globular protein of this size. Analysis of chemical shift, 3JHNalpha coupling constant, and NOE data show that the experimental parameters agree well overall with values measured in short model peptides and with predictions from a statistical model for a random coil. Sequences where specific features give deviations from these predictions for a random coil have however been identified. These arise from clustering of hydrophobic side chains and electrostatic interactions between charged groups. 15N relaxation studies demonstrate that local fluctuations of the chain are the dominant motional process that gives rise to relaxation of the 15N nuclei, with a persistence length of approximately 7-10 residues for the segmental motion. The consequences of the structural and dynamical properties of this unfolded protein for its biological role of binding to fibronectin have been considered. It is found that the regions of the sequence involved in binding have a high propensity for populating extended conformations, a feature that would allow a number of both charged and hydrophobic groups to be presented to fibronectin for highly specific binding.

  19. Workforce. Focal Point: Research, Policy, and Practice in Children's Mental Health. Volume 22, Number 1, Winter 2008

    ERIC Educational Resources Information Center

    Walker, Janet S., Ed.; Gowen, L. Kris, Ed.; Aue, Nicole, Ed.

    2008-01-01

    This issue of "Focal Point" explores how the increasing emphasis on using evidence-based practices and a "system of care" approach is driving changes in jobs and roles related to children's mental health. Articles in the issue describe how agencies and providers of services and supports have responded to these changes by creating new types of…

  20. Workforce. Focal Point: Research, Policy, and Practice in Children's Mental Health. Volume 22, Number 1, Winter 2008

    ERIC Educational Resources Information Center

    Walker, Janet S., Ed.; Gowen, L. Kris, Ed.; Aue, Nicole, Ed.

    2008-01-01

    This issue of "Focal Point" explores how the increasing emphasis on using evidence-based practices and a "system of care" approach is driving changes in jobs and roles related to children's mental health. Articles in the issue describe how agencies and providers of services and supports have responded to these changes by creating new types of…

  1. Fetal fibronectin, interleukin-6, and C-reactive protein are useful in establishing prognostic subcategories of idiopathic preterm labor.

    PubMed

    Burrus, D R; Ernest, J M; Veille, J C

    1995-10-01

    Our purpose was to evaluate fetal fibronectin, interleukin-6, and C-reactive protein from patients with preterm labor to establish prognostic subcategories. Thirty-seven patients with preterm labor had cervical fetal fibronectin and plasma C-reactive protein sampled. Eighteen of these patients had amniotic fluid interleukin-6 levels measured. Outcome variables were (1) delivery before 34 weeks and (2) delivery within 48 hours. Detectable cervical fetal fibronectin identified 89% of patients who were delivered before 34 weeks' gestation. Interleukin-6 > 1500 pg/ml identified 88% of patients who were delivered within 48 hours. C-reactive protein > 1.5 mg/dl correlated with elevated interleukin-6 levels (p < 0.001). Three subcategories of idiopathic preterm labor were evident: (1) fetal fibronectin nondetectable (37% likely to be delivered before 34 weeks), (2) fetal fibronectin detectable but interleukin-6 < 1500 pg/ml (79% likely to be delivered before 34 weeks but 85% with > 48 hours' latency), and (3) fetal fibronectin present and interleukin-6 > 1500 pg/ml (91% likely to be delivered with < 48 hours' latency).

  2. Fibronectin-Alginate microcapsules improve cell viability and protein secretion of encapsulated Factor IX-engineered human mesenchymal stromal cells.

    PubMed

    Sayyar, Bahareh; Dodd, Megan; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2015-01-01

    Continuous delivery of proteins by engineered cells encapsu-lated in biocompatible polymeric microcapsules is of considerable therapeutic potential. However, this technology has not lived up to expectations due to inadequate cell--matrix interactions and subsequent cell death. In this study we hypoth-esize that the presence of fibronectin in an alginate matrix may enhance the viability and functionality of encapsulated human cord blood-derived mesenchymal stromal cells (MSCs) expressing the human Factor IX (FIX) gene. MSCs were encapsulated in alginate-PLL microcapsules containing 10, 100, or 500 μg/ml fibronectin to ameliorate cell survival. MSCs in microcapsules with 100 and 500 μg/ml fibronectin demonstrated improved cell viability and proliferation and higher FIX secretion compared to MSCs in non-supplemented microcapsules. In contrast, 10 μg/ml fibronectin did not significantly affect the viability and protein secretion from the encapsulated cells. Differentiation studies demonstrated osteogenic (but not chondrogenic or adipogenic) differentiation capability and efficient FIX secretion of the enclosed MSCs in the fibronectin-alginate suspension culture. Thus, the use of recombinant MSCs encapsulated in fibronectin-alginate microcapsules in basal or osteogenic cultures may be of practical use in the treatment of hemophilia B.

  3. Identification and isolation of a 140 kd cell surface glycoprotein with properties expected of a fibronectin receptor

    SciTech Connect

    Pytela, R.; Pierschbacher, M.D.; Ruoslahti, E.

    1985-01-01

    Affinity chromatography was used to identify a putative cell surface receptor for fibronectin. A large cell-attachment-promoting fibronectin fragment was used as the affinity matrix, and specific elution was effected by using synthetic peptides containing the sequence Arg-Gly-Asp, which is derived from the cell recognition sequence in the fibronectin cell attachment site. A 140 kd protein was bound by the affinity matrix from octylglucoside extracts of MG-63 human osteosarcoma cells and specifically eluted with the synthetic peptide Gly-Arg-Gly-Asp-Ser-Pro. The 140 kd protein was labeled by cell surface specific radioiodination and became incorporated into liposomes at a high efficiency. Liposomes containing this protein showed specific affinity toward fibronectin-coated surfaces, and this binding could be selectively inhibited by the synthetic cell-attachment peptide but not by inactive peptides. Affinity chromatography on wheat germ agglutinin-Sepharose showed that the 140 kd protein is a glycoprotein and, in combination with the fibronectin fragment chromatography, gave highly enriched preparations of the 140 kd protein. These properties suggest that the 140 kd glycoprotein is a membrane-embedded cell surface protein directly involved in the initial step of cell adhesion to fibronectin substrates.

  4. Fibronectin induction abrogates the BRAF inhibitor response of BRAF V600E/PTEN-null melanoma cells

    PubMed Central

    Fedorenko, Inna V.; Abel, Ethan V.; Koomen, John M.; Fang, Bin; Wood, Elizabeth R.; Chen, Y. Ann; Fisher, Kate J.; Iyengar, Sanjana; Dahlman, Kimberly B.; Wargo, Jennifer A.; Flaherty, Keith T.; Sosman, Jeffrey A.; Sondak, Vernon K.; Messina, Jane L.; Gibney, Geoffrey T.; Smalley, Keiran S.M.

    2015-01-01

    The mechanisms by which some melanoma cells adapt to BRAF inhibitor therapy are incompletely understood. In the present study, we used mass spectrometry-based phosphoproteomics to determine how BRAF inhibition remodeled the signaling network of melanoma cell lines that were BRAF-mutant and PTEN-null. Short-term BRAF inhibition was associated with marked changes in fibronectin-based adhesion signaling that were PTEN-dependent. These effects were recapitulated through BRAF siRNA knockdown and following treatment with chemotherapeutic drugs. Increased fibronectin expression was also observed in mouse xenograft models as well as specimens from melanoma patients undergoing BRAF inhibitor treatment. Analysis of a melanoma TMA showed loss of PTEN expression to predict for a lower overall survival, with a trend for even lower survival being seen when loss of fibronectin was included in the analysis. Mechanistically, the induction of fibronectin limited the responses of these PTEN-null melanoma cell lines to vemurafenib, with enhanced cytotoxicity observed following the knockdown of either fibronectin or its receptor α5β1 integrin. This in turn abrogated the cytotoxic response to BRAF inhibition via increased AKT signaling, which prevented the induction of cell death by maintaining the expression of the pro-survival protein Mcl-1. The protection conveyed by the induction of fibronectin expression could be overcome through combined treatment with a BRAF and PI3K inhibitor. PMID:26073081

  5. Tuning Mesenchymal Stem Cell Response onto Titanium-Niobium-Hafnium Alloy by Recombinant Fibronectin Fragments.

    PubMed

    Herranz-Diez, C; Mas-Moruno, C; Neubauer, S; Kessler, H; Gil, F J; Pegueroles, M; Manero, J M; Guillem-Marti, J

    2016-02-03

    Since metallic biomaterials used for bone replacement possess low bioactivity, the use of cell adhesive moieties is a common strategy to improve cellular response onto these surfaces. In recent years, the use of recombinant proteins has emerged as an alternative to native proteins and short peptides owing to the fact that they retain the biological potency of native proteins, while improving their stability. In the present study, we investigated the biological effect of two different recombinant fragments of fibronectin, spanning the 8-10th and 12-14th type III repeats, covalently attached to a new TiNbHf alloy using APTES silanization. The fragments were studied separately and mixed at different concentrations and compared to a linear RGD, a cyclic RGD and the full-length fibronectin protein. Cell culture studies using rat mesenchymal stem cells demonstrated that low to medium concentrations (30% and 50%) of type III 8-10th fragment mixed with type III 12-14th fragment stimulated cell spreading and proliferation compared to RGD peptides and the fragments separately. On the other hand, type III 12-14th fragment alone or mixed at low volume percentages ≤50% with type III 8-10th fragment increased alkaline phosphatase levels compared to the other molecules. These results are significant for the understanding of the role of fibronectin recombinant fragments in cell responses and thus to design bioactive coatings for biomedical applications.

  6. Topical tretinoin increases the tropoelastin and fibronectin content of photoaged hairless mouse skin.

    PubMed

    Schwartz, E; Kligman, L H

    1995-04-01

    Topical tretinoin treatment of photoaged hairless mice has been shown in previous studies to stimulate formation of a subepidermal zone of new connective tissue characterized by enhanced collagen synthesis. The aims of this study were to localize and/or quantify elastin, fibronectin, and glycosaminoglycans in the same model. Hairless mice (Skh-1) were irradiated thrice weekly for 10 weeks with gradually increasing doses of ultraviolet (up to 4.5 minimal erythema doses per exposure) from Westinghouse FS-40 bulbs. Mice were then treated five times a week with either 0.05% tretinoin, the ethanol:propylene glycol vehicle, or nothing for another 10 weeks. Controls included mice sacrificed after 10 weeks of ultraviolet treatment and age-matched untreated animals. The distribution of elastin and fibronectin was examined by immunofluorescence microscopy, which revealed fine fibrils in the subepidermal zone in tretinoin-treated skin. A quantitative slot-blot immunobinding assay showed that tretinoin induced a threefold higher amount of tropoelastin compared with controls. Insoluble elastin content (desmosine levels) was similar in all groups. Although fibronectin content was increased by ultraviolet radiation, tretinoin treatment induced the largest increase. In contrast, the amount of glycosaminoglycans, although increased by UVB radiation, was reduced by tretinoin treatment.

  7. Effect of a collagen membrane enriched with fibronectin on guided tissue regeneration in dogs.

    PubMed

    Kurtis, Bülent; Balos, Köksal; Oygür, Tülin

    2002-01-01

    The present study was planned to assess the capacity of a resorbable collagen membrane enriched with fibronectin to prevent the apical migration of epithelium and to facilitate new attachment and new bone. Experimental osseous dehiscence defects were produced on the bilateral labial aspect of mandibular 2nd, 3rd and 4th premolar teeth in six mongrel dogs. Guided tissue regeneration therapy using collagen membranes, which were rehydrated with fibronectin solution, was performed on one quadrant (group A). In the contralateral quadrant, the same collagen membranes, but rehydrated only with saline (group B), were placed over the bony defects. The third premolar teeth, which were treated by open-flap debridement, served as control (group C). Flaps were positioned slightly coronally and sutured; sutures were removed 10 days later. The dogs were killed 30 days after reconstructive therapy. Tissue blocks containing the experimental and control teeth were excised, demineralized in EDTA, and embedded in paraffin. Histological and histometric evaluation revealed that all groups demonstrated similar effects on preventing the down-growth of epithelium and formation of new cementum and new bone. Collagen membranes were tolerated well within the tissues, and membrane remnants were identified at 30 days. In summary, this study indicated that in this dog model similar healing results could be achieved with a bovine type I collagen membrane with or without fibronectin solution and open-flap debridement.

  8. Periodontal healing following guided tissue regeneration with citric acid and fibronectin application.

    PubMed

    Caffesse, R G; Nasjleti, C E; Anderson, G B; Lopatin, D E; Smith, B A; Morrison, E C

    1991-01-01

    This study was undertaken to determine the effects of guided tissue regeneration (GTR) with and without citric acid conditioning and autologous fibronectin application. The study subjects were four female beagle dogs with spontaneous periodontitis. The dogs were given thorough root debridement and 4 weeks later, mucoperiosteal flaps were raised on both sides of the mandible involving the 2nd, 3rd, and 4th premolar and 1st molar teeth. After debridement, notches were placed on the roots at the level of supporting bone. Citric acid (pH 1) was topically applied for 3 minutes on the exposed root surfaces of one side (experimental). The roots were irrigated with normal saline solution. Both the root surfaces and the inner surface of the flap were then bathed in autologous fibronectin in saline. Following this, Gore-Tex periodontal material was adapted to the roots of each tooth and sutured. The contralateral side, serving as control, was treated by surgery and application of Gore-Tex periodontal material only. All membranes were removed 1 month after surgery, and the dogs sacrificed at 3 months. Both mesio-distal and bucco-lingual microscopic histological sections were evaluated by descriptive histology, and linear measurements and surface area determination of the furcal tissues were made. Periodontal healing following the use of GTR procedure resulted in an increase in connective tissue and alveolar bone regeneration. Adjunctive critic acid plus autologous fibronectin produced slightly better results, but these differences were not statistically significant for this sample.

  9. Mechanism of Fibronectin Binding to Human Trabecular Meshwork Exosomes and Its Modulation by Dexamethasone

    PubMed Central

    Klingeborn, Mikael; Stamer, W. Daniel

    2016-01-01

    Exosomes are emerging as important mediators of cell-matrix interactions by means of specific adhesion proteins. Changes in the tissue-specific exosomal protein expression may underlie pathological conditions whereby extracellular matrix turnover and homeostasis is disrupted. Ocular hypertension due to extracellular matrix accumulation in the trabecular meshwork is a hallmark of glucocorticoid-induced glaucoma. In the trabecular meshwork, exosomal fibronectin mediates cell matrix interactions at cellular structures called “invadosomes”. Trabecular meshwork cells use invadosomes to turn over their surrounding matrix and maintain passageways for flow of aqueous humor. In this study, we observed that human trabecular meshwork explants treated with dexamethasone released exosomes with significantly reduced amounts of fibronectin bound per exosome. Further, we found that exosome-fibronectin binding is heparan sulfate-dependent, consistent with our observation that trabecular meshwork exosomes are enriched in the heparin/heparan sulfate binding annexins A2 and A6. In this way, dexamethasone-treated explants released exosomes with a significant reduction in annexin A2 and A6 per exosome. Interestingly, we did not detect exosomal matrix metalloproteinases, but we identified abundant dipeptidyl peptidase 4, a serine protease whose activity was reduced on exosomes isolated from dexamethasone-treated explants. Together, our findings demonstrate mechanistically how corticosteroid-induced alterations in exosomal adhesion cargo and properties can account for the pathological matrix accumulation seen in many glaucoma patients. PMID:27783649

  10. Fibronectin is deposited by injury-activated epicardial cells and is necessary for zebrafish heart regeneration

    PubMed Central

    Wang, Jinhu; Karra, Ravi; Dickson, Amy L.; Poss, Kenneth D.

    2013-01-01

    Unlike adult mammals, adult zebrafish vigorously regenerate lost heart muscle in response to injury. The epicardium, a mesothelial cell layer enveloping the myocardium, is activated to proliferate after cardiac injury and can contribute vascular support cells or provide mitogens to regenerating muscle. Here, we applied proteomics to identify secreted proteins that are associated with heart regeneration. We found that Fibronectin, a main component of the extracellular matrix, is induced and deposited after cardiac damage. In situ hybridization and transgenic reporter analyses indicated that expression of two fibronectin paralogues, fn1 and fn1b, are induced by injury in epicardial cells, while the itgb3 receptor is induced in cardiomyocytes near the injury site. fn1, the more dynamic of these paralogs, is induced chamber-wide within one day of injury before localizing epicardial Fn1 synthesis to the injury site. fn1 loss-of-function mutations disrupted zebrafish heart regeneration, as did induced expression of a dominant-negative Fibronectin cassette, defects that were not attributable to direct inhibition of cardiomyocyte proliferation. These findings reveal a new role for the epicardium in establishing an extracellular environment that supports heart regeneration. PMID:23988577

  11. Osteoblast mineralization requires β1 integrin/ICAP-1–dependent fibronectin deposition

    PubMed Central

    Brunner, Molly; Millon-Frémillon, Angélique; Chevalier, Genevieve; Nakchbandi, Inaam A.; Mosher, Deane; Block, Marc R.

    2011-01-01

    The morphogenetic and differentiation events required for bone formation are orchestrated by diffusible and insoluble factors that are localized within the extracellular matrix. In mice, the deletion of ICAP-1, a modulator of β1 integrin activation, leads to severe defects in osteoblast proliferation, differentiation, and mineralization and to a delay in bone formation. Deposition of fibronectin and maturation of fibrillar adhesions, adhesive structures that accompany fibronectin deposition, are impaired upon ICAP-1 loss, as are type I collagen deposition and mineralization. Expression of β1 integrin with a mutated binding site for ICAP-1 recapitulates the ICAP-1–null phenotype. Follow-up experiments demonstrated that ICAP-1 negatively regulates kindlin-2 recruitment onto the β1 integrin cytoplasmic domain, whereas an excess of kindlin-2 binding has a deleterious effect on fibrillar adhesion formation. These results suggest that ICAP-1 works in concert with kindlin-2 to control the dynamics of β1 integrin–containing fibrillar adhesions and, thereby, regulates fibronectin deposition and osteoblast mineralization. PMID:21768292

  12. Structure of integrin α5β1 in complex with fibronectin

    PubMed Central

    Takagi, Junichi; Strokovich, Konstantin; Springer, Timothy A.; Walz, Thomas

    2003-01-01

    The membrane-distal headpiece of integrins has evolved to specifically bind large extracellular protein ligands, but the molecular architecture of the resulting complexes has not been determined. We used molecular electron microscopy to determine the three-dimensional structure of the ligand-binding headpiece of integrin α5β1 complexed with fragments of its physiological ligand fibronectin. The density map for the unliganded α5β1 headpiece shows a ‘closed’ conformation similar to that seen in the αVβ3 crystal structure. By contrast, binding to fibronectin induces an ‘open’ conformation with a dramatic, ∼80° change in the angle of the hybrid domain of the β subunit relative to its I-like domain. The fibronectin fragment binds to the interface between the β-propeller and I-like domains in the integrin headpiece through the RGD-containing module 10, but direct contact of the synergy-region-containing module 9 to integrin is not evident. This finding is corroborated by kinetic analysis of real-time binding data, which shows that the synergy site greatly enhances kon but has little effect on the stability or koff of the complex. PMID:12970173

  13. Focal Mechanism determination of local M

    NASA Astrophysics Data System (ADS)

    Vales, Dina; Custório, Susana; Carrilho, Fernando

    2015-04-01

    We determine the focal mechanisms of local small (ML<3.9) earthquakes that occurred between 2013 and 2014 in mainland Portugal. These low magnitude events were recorded by several stations that provide first-motion polarity solutions. However, only few stations are located near the epicenter and record a waveform with a signal-to-noise ratio (SNR) high enough to allow full waveform modelling. To overcome this limitation, we used a new approach called cyclic scanning of the polarity solutions (CSPS) (Fojtíková and Zahradnik, 2014), which performs a joint inversion of full waveform and first motion polarities to retrieve the focal mechanism. This methodology has the advantage of yielding reliable focal mechanism solutions, even when high SNR waveforms are available from only a few near field stations (or in the limiting case, only with one single station). To apply the CSPS method one needs to: i) run the the FOCal MEChanism (FOCMEC) code (Snoke, 2003) to obtain a suite of the DC solutions corresponding to the first motion polarities, and then ii) perform the waveform modelling in order to decrease the uncertainty. The ISOLated Asperities (ISOLA) software (Sokos and Zahradník, 2008, 2013) is used in this second step. We applied this method to weak events recorded by a network of 30 broadband seismic stations that transmit data in real-time to Instituto Português do Mar e da Atmosfera (IPMA), the institution responsible for seismic monitoring in Portugal. We interpret the obtained fault plane solutions in light of active faults and regional tectonics, and in comparison with focal mechanisms previously inferred for events in the region. The focal mechanisms obtained for small earthquakes allow us to significantly expand the database of available focal mechanisms in mainland Portugal, contributing to the understanding of active deformation in the region.

  14. NMDA receptor binding in focal epilepsies.

    PubMed

    McGinnity, C J; Koepp, M J; Hammers, A; Riaño Barros, D A; Pressler, R M; Luthra, S; Jones, P A; Trigg, W; Micallef, C; Symms, M R; Brooks, D J; Duncan, J S

    2015-10-01

    To demonstrate altered N-methyl-d-aspartate (NMDA) receptor availability in patients with focal epilepsies using positron emission tomography (PET) and [(18)F]GE-179, a ligand that selectively binds to the open NMDA receptor ion channel, which is thought to be overactive in epilepsy. Eleven patients (median age 33 years, 6 males) with known frequent interictal epileptiform discharges had an [(18)F]GE-179 PET scan, in a cross-sectional study. MRI showed a focal lesion but discordant EEG changes in two, was non-localising with multifocal EEG abnormalities in two, and was normal in the remaining seven patients who all had multifocal EEG changes. Individual patient [(18)F]GE-179 volume-of-distribution (VT) images were compared between individual patients and a group of 10 healthy controls (47 years, 7 males) using Statistical Parametric Mapping. Individual analyses revealed a single cluster of focal VT increase in four patients; one with a single and one with multifocal MRI lesions, and two with normal MRIs. Post hoc analysis revealed that, relative to controls, patients not taking antidepressants had globally increased [(18)F]GE-179 VT (+28%; p<0.002), and the three patients taking an antidepressant drug had globally reduced [(18)F]GE-179 VT (-29%; p<0.002). There were no focal abnormalities common to the epilepsy group. In patients with focal epilepsies, we detected primarily global increases of [(18)F]GE-179 VT consistent with increased NMDA channel activation, but reduced availability in those taking antidepressant drugs, consistent with a possible mode of action of this class of drugs. [(18)F]GE-179 PET showed focal accentuations of NMDA binding in 4 out of 11 patients, with difficult to localise and treat focal epilepsy. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

  15. Transparent Meta-Analysis: Does Aging Spare Prospective Memory with Focal vs. Non-Focal Cues?

    PubMed Central

    Uttl, Bob

    2011-01-01

    Background Prospective memory (ProM) is the ability to become aware of a previously-formed plan at the right time and place. For over twenty years, researchers have been debating whether prospective memory declines with aging or whether it is spared by aging and, most recently, whether aging spares prospective memory with focal vs. non-focal cues. Two recent meta-analyses examining these claims did not include all relevant studies and ignored prevalent ceiling effects, age confounds, and did not distinguish between prospective memory subdomains (e.g., ProM proper, vigilance, habitual ProM) (see Uttl, 2008, PLoS ONE). The present meta-analysis focuses on the following questions: Does prospective memory decline with aging? Does prospective memory with focal vs. non-focal cues decline with aging? Does the size of age-related declines with focal vs. non-focal cues vary across ProM subdomains? And are age-related declines in ProM smaller than age-related declines in retrospective memory? Methods and Findings A meta-analysis of event-cued ProM using data visualization and modeling, robust count methods, and conventional meta-analysis techniques revealed that first, the size of age-related declines in ProM with both focal and non-focal cues are large. Second, age-related declines in ProM with focal cues are larger in ProM proper and smaller in vigilance. Third, age-related declines in ProM proper with focal cues are as large as age-related declines in recall measures of retrospective memory. Conclusions The results are consistent with Craik's (1983) proposal that age-related declines on ProM tasks are generally large, support the distinction between ProM proper vs. vigilance, and directly contradict widespread claims that ProM, with or without focal cues, is spared by aging. PMID:21304905

  16. "Focal thyroid inferno" on color Doppler ultrasonography: a specific feature of focal Hashimoto's thyroiditis.

    PubMed

    Fu, Xianshui; Guo, Limei; Zhang, Huabin; Ran, Weiqiang; Fu, Peng; Li, Zhiqiang; Chen, Wen; Jiang, Ling; Wang, Jinrui; Jia, Jianwen

    2012-11-01

    To evaluate color-Doppler features predictive of focal Hashimoto's thyroiditis. A total of 521 patients with 561 thyroid nodules that underwent surgeries or gun biopsies were included in this study. These nodules were divided into three groups: focal Hashimoto's thyroiditis (104 nodules in 101 patients), benignity other than focal Hashimoto's thyroiditis (73 nodules in 70 patients), and malignancy (358 nodules in 350 patients). On color Doppler sonography, four vascularity types were determined as: hypovascularity, marked internal flow, marked peripheral flow and focal thyroid inferno. The χ2 test was performed to seek the potential vascularity type with the predictive ability of certain thyroid pathology. Furthermore, the gray-scale features of each nodule were also studied. The vascularity type I (hypovascularity) was more often seen in focal Hashimoto's thyroiditis than other benignity and malignancy (46% vs. 20.5% and 19%). While the type II (marked internal flow) showed the opposite tendency (26.9% [focal Hashimoto's thyroiditis] vs. 45.2% [other benignity] and 52.8% [malignancy]). However, type III (marked peripheral flow) was unable to predict any thyroid pathology. Importantly, type IV (focal thyroid inferno) was exclusive to focal Hashimoto's thyroiditis. All 8 type IV nodules appeared to be solid, hypoechoic, and well-defined. Using "focal thyroid inferno" as an indicator of FHT, the diagnostic sensitivity and specificity were 7.7% and 100% respectively. The vascularity type of "focal thyroid inferno" is specific for focal Hashimoto thyroiditis. Recognition of this particular feature may avoid unnecessary interventional procedures for some solid hypoechoic thyroid nodules suspicious of malignancy. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  17. Power semiconductor controlled drives

    NASA Astrophysics Data System (ADS)

    Dubey, Gopal K.

    This book presents power semiconductor controlled drives employing dc motors, induction motors, and synchronous motors. The dynamics of motor and load systems are covered. Open-loop and closed-loop drives are considered, and thyristor, power transistor, and GTO converters are discussed. In-depth coverage is given to ac drives, particularly those fed by voltage and current source inverters and cycloconverters. Full coverage is given to brushless and commutatorless dc drives, including load-commuted synchronous motor drives. Rectifier-controlled dc drives are presented in detail.

  18. Biosynthesis, surface expression and function of the fibronectin receptor after rat liver cell transformation to tumorigenicity.

    PubMed Central

    Decastel, M; Doyennette-Moyne, M A; Gouet, E; Aubery, M; Codogno, P

    1993-01-01

    Zajdela hepatoma cells are poorly-adherent cells derived from an undifferentiated tumour and transplanted into rat. We compared the biosynthesis, structure and function of the fibronectin receptor in normal rat hepatocytes with that in Zajdela hepatoma cells. The rat hepatocyte fibronectin receptor has been isolated. It is composed of two subunits: alpha 5 (molecular mass 155 kDa) and beta 1 (molecular mass 115 kDa). However, its biosynthesis has not yet been described. Using polyclonal antibodies raised against each of the subunits of the receptor, we observed that the alpha 5-subunit was synthesized as a 155-kDa polypeptide in normal rat hepatocytes and Zajdela hepatoma cells. In contrast, the molecular mass of the beta 1-subunit was 130 kDa in Zajdela hepatoma cells versus 115 kDa in normal rat hepatocytes. Pulse-chase experiments showed that the apparent transition time from the 100-kDa beta 1-precursor to the 130-kDa mature form was abnormally prolonged in Zajdela hepatoma cells since the latter was not detected until 24 h, while the transition from the 100-kDa precursor to the 115-kDa mature form began within 3 h in normal rat hepatocytes. Digestion of both the normal rat hepatocytes and Zajdela hepatoma cells 100-kDa beta 1-precursors with endo-beta-N-acetylglucosaminidase H and peptide N-glycosidase yielded products from 100 kDa to 84 kDa and 82 kDa, respectively, as judged by SDS/PAGE, suggesting that the same polypeptide chain is synthesized in normal rat hepatocytes and in Zajdela hepatoma cells. Incubation of the mature normal rat hepatocyte beta 1-subunit with peptide N-glycosidase reduced its molecular mass from 115 kDa to 82 kDa, as judged by SDS/PAGE, while the molecular mass of the abnormal mature Zajdela hepatoma cell beta 1-subunit decreased from 130 to 110 kDa. Thus, in addition to alterations in the Asn-linked oligosaccharide processing, 'ascitic growth' induced other post-translational modifications in the Zajdela hepatoma cell beta 1-subunit

  19. Focal Plane Metrology for the LSST Camera

    SciTech Connect

    A Rasmussen, Andrew P.; Hale, Layton; Kim, Peter; Lee, Eric; Perl, Martin; Schindler, Rafe; Takacs, Peter; Thurston, Timothy; /SLAC

    2007-01-10

    Meeting the science goals for the Large Synoptic Survey Telescope (LSST) translates into a demanding set of imaging performance requirements for the optical system over a wide (3.5{sup o}) field of view. In turn, meeting those imaging requirements necessitates maintaining precise control of the focal plane surface (10 {micro}m P-V) over the entire field of view (640 mm diameter) at the operating temperature (T {approx} -100 C) and over the operational elevation angle range. We briefly describe the hierarchical design approach for the LSST Camera focal plane and the baseline design for assembling the flat focal plane at room temperature. Preliminary results of gravity load and thermal distortion calculations are provided, and early metrological verification of candidate materials under cold thermal conditions are presented. A detailed, generalized method for stitching together sparse metrology data originating from differential, non-contact metrological data acquisition spanning multiple (non-continuous) sensor surfaces making up the focal plane, is described and demonstrated. Finally, we describe some in situ alignment verification alternatives, some of which may be integrated into the camera's focal plane.

  20. Neuroimaging of focal cortical dysplasia: neuropathological correlations.

    PubMed

    Colombo, Nadia; Citterio, Alberto; Galli, Carlo; Tassi, Laura; Lo Russo, Giorgio; Scialfa, G; Spreafico, Roberto

    2003-09-01

    Focal cortical dysplasia is a well-known cause of intractable epilepsy with early onset of seizures, and is potentially amenable to surgical therapy. It was first described by Taylor in 1971 as a peculiar malformative disorganisation of the neocortex characterised at histology by loss of cortical lamination and accompanied by giant, dysmorphic neurones and, most frequently, by "balloon cells" littered throughout the cortex and sub-cortical white matter. While in the past decades the term "cortical dysplasia" has referred to various malformations of cortical development, such as agyria, pachygyria, polymicrogyria, heterotopia and hemimegalencephaly, it is now widely accepted that the entity identified by Taylor should be considered separately, from both histological and neuroimaging standpoints. More recently, the recognition of various histological subtypes of focal cortical dysplasia characterised by different degrees of cortical disruption with or without cytological abnormalities has generated several classifications that are still unsatisfactory. With better magnetic resonance capability, subtle and very small focal cortical dysplasias may now be visualised and the differential magnetic resonance aspects of the histological subgroups can be established. We will discuss the problem of histopathological classification and magnetic resonance imaging differentiation of the various subtypes of focal cortical dysplasia in the light of personal data collected from a large series of epileptic patients who underwent surgery and had a histological diagnosis of focal cortical dysplasia. Copyright John Libbey Eurotext 2003.

  1. Dementia and driving

    MedlinePlus

    ... medlineplus.gov/ency/patientinstructions/000028.htm Dementia and driving To use the sharing features on this page, ... their independence is being taken away. Signs That Driving May No Longer be Safe People with signs ...

  2. Ocular disease and driving.

    PubMed

    Wood, Joanne M; Black, Alex A

    2016-09-01

    As the driving population ages, the number of drivers with visual impairment resulting from ocular disease will increase given the age-related prevalence of ocular disease. The increase in visual impairment in the driving population has a number of implications for driving outcomes. This review summarises current research regarding the impact of common ocular diseases on driving ability and safety, with particular focus on cataract, glaucoma, age-related macular degeneration, hemianopia and diabetic retinopathy. The evidence considered includes self-reported driving outcomes, driving performance (on-road and simulator-based) and various motor vehicle crash indices. Collectively, this review demonstrates that driving ability and safety are negatively affected by ocular disease; however, further research is needed in this area. Older drivers with ocular disease need to be aware of the negative consequences of their ocular condition and in the case where treatment options are available, encouraged to seek these earlier for optimum driving safety and quality of life benefits.

  3. Impaired Driving - Multiple Languages

    MedlinePlus

    ... Are Here: Home → Multiple Languages → All Health Topics → Impaired Driving URL of this page: https://medlineplus.gov/languages/ ... V W XYZ List of All Topics All Impaired Driving - Multiple Languages To use the sharing features on ...

  4. The Focal Adhesion Analysis Server: a web tool for analyzing focal adhesion dynamics.

    PubMed

    Berginski, Matthew E; Gomez, Shawn M

    2013-01-01

    The Focal Adhesion Analysis Server (FAAS) is a web-based implementation of a set of computer vision algorithms designed to quantify the behavior of focal adhesions in cells imaged in 2D cultures. The input consists of one or more images of a labeled focal adhesion protein. The outputs of the system include a range of static and dynamic measurements for the adhesions present in each image as well as how these properties change over time. The user is able to adjust several parameters important for proper focal adhesion identification. This system provides a straightforward tool for the global, unbiased assessment of focal adhesion behavior common in optical microscopy studies. The webserver is available at: http://faas.bme.unc.edu/.

  5. Gear bearing drive

    NASA Technical Reports Server (NTRS)

    Weinberg, Brian (Inventor); Mavroidis, Constantinos (Inventor); Vranish, John M. (Inventor)

    2011-01-01

    A gear bearing drive provides a compact mechanism that operates as an actuator providing torque and as a joint providing support. The drive includes a gear arrangement integrating an external rotor DC motor within a sun gear. Locking surfaces maintain the components of the drive in alignment and provide support for axial loads and moments. The gear bearing drive has a variety of applications, including as a joint in robotic arms and prosthetic limbs.

  6. Solid-state curved focal plane arrays

    NASA Technical Reports Server (NTRS)

    Nikzad, Shouleh (Inventor); Hoenk, Michael (Inventor); Jones, Todd (Inventor)

    2010-01-01

    The present invention relates to curved focal plane arrays. More specifically, the present invention relates to a system and method for making solid-state curved focal plane arrays from standard and high-purity devices that may be matched to a given optical system. There are two ways to make a curved focal plane arrays starting with the fully fabricated device. One way, is to thin the device and conform it to a curvature. A second way, is to back-illuminate a thick device without making a thinned membrane. The thick device is a special class of devices; for example devices fabricated with high purity silicon. One surface of the device (the non VLSI fabricated surface, also referred to as the back surface) can be polished to form a curved surface.

  7. Focal Adhesion-Independent Cell Migration.

    PubMed

    Paluch, Ewa K; Aspalter, Irene M; Sixt, Michael

    2016-10-06

    Cell migration is central to a multitude of physiological processes, including embryonic development, immune surveillance, and wound healing, and deregulated migration is key to cancer dissemination. Decades of investigations have uncovered many of the molecular and physical mechanisms underlying cell migration. Together with protrusion extension and cell body retraction, adhesion to the substrate via specific focal adhesion points has long been considered an essential step in cell migration. Although this is true for cells moving on two-dimensional substrates, recent studies have demonstrated that focal adhesions are not required for cells moving in three dimensions, in which confinement is sufficient to maintain a cell in contact with its substrate. Here, we review the investigations that have led to challenging the requirement of specific adhesions for migration, discuss the physical mechanisms proposed for cell body translocation during focal adhesion-independent migration, and highlight the remaining open questions for the future.

  8. Measuring microfocus focal spots using digital radiography

    SciTech Connect

    Fry, David A

    2009-01-01

    Measurement of microfocus spot size can be important for several reasons: (1) Quality assurance during manufacture of microfocus tubes; (2) Tracking performance and stability of microfocus tubes; (3) Determining magnification (especially important for digital radiography where the native spatial resolution of the digital system is not adequate for the application); (4) Knowledge of unsharpness from the focal spot alone. The European Standard EN 12543-5 is based on a simple geometrical method of calculating focal spot size from unsharpness of high magnification film radiographs. When determining microfocus focal spot dimensions using unsharpness measurements both signal-to-noise (SNR) and magnification can be important. There is a maximum accuracy that is a function of SNR and therefore an optimal magnification. Greater than optimal magnification can be used but it will not increase accuracy.

  9. Grieving while Driving

    ERIC Educational Resources Information Center

    Rosenblatt, Paul C.

    2004-01-01

    Secondary analysis of data from 84 people in 2 interview studies shows that some bereaved people grieve actively while driving. The grief can be intense, even years after a death. Grief while driving may erupt spontaneously or be set off by a wide range of reminders. Some bereaved people seem to save their grieving for times when they drive,…

  10. Sequential Dependencies in Driving

    ERIC Educational Resources Information Center

    Doshi, Anup; Tran, Cuong; Wilder, Matthew H.; Mozer, Michael C.; Trivedi, Mohan M.

    2012-01-01

    The effect of recent experience on current behavior has been studied extensively in simple laboratory tasks. We explore the nature of sequential effects in the more naturalistic setting of automobile driving. Driving is a safety-critical task in which delayed response times may have severe consequences. Using a realistic driving simulator, we find…

  11. Sequential Dependencies in Driving

    ERIC Educational Resources Information Center

    Doshi, Anup; Tran, Cuong; Wilder, Matthew H.; Mozer, Michael C.; Trivedi, Mohan M.

    2012-01-01

    The effect of recent experience on current behavior has been studied extensively in simple laboratory tasks. We explore the nature of sequential effects in the more naturalistic setting of automobile driving. Driving is a safety-critical task in which delayed response times may have severe consequences. Using a realistic driving simulator, we find…

  12. Magnetic drive coupling

    NASA Technical Reports Server (NTRS)

    Carter, Edward L. (Inventor)

    1987-01-01

    The driving and driven members of a magnetic drive are separated by en enlarged gap to provide clearance for a conduit or other member. Flux pins in the gap maintain the torque transmitting capability of the drive. The spacing between two of the flux pins is increased to provide space for the conduit.

  13. Expansive focal cemento-osseous dysplasia.

    PubMed

    Bulut, Emel Uzun; Acikgoz, Aydan; Ozan, Bora; Zengin, Ayse Zeynep; Gunhan, Omer

    2012-01-01

    To present a case of expansive focal cemento-osseous dysplasia and emphasize the importance of differential diagnosis. Cemento-osseous dysplasia is categorized into three subtypes on the basis of the clinical and radiographic features: Periapical, focal and florid. The focal type exhibits a single site of involvement in any tooth-bearing or edentulous area of the jaws. These lesions are usually asymptomatic; therefore, they are frequently diagnosed incidentally during routine radiographic examinations. Lesions are usually benign, show limited growth, and do not require further surgical intervention, but periodic follow-up is recommended because occasionally, this type of dysplasia progresses into florid osseous dysplasia and simple bone cysts are formed. A 24-year-old female patient was referred to our clinic for swelling in the left edentulous mandibular premolarmolar region and felt discomfort when she wore her prosthetics. She had no pain, tenderness or paresthesia. Clinical examination showed that the swelling in the posterior mandible that was firm, nonfluctuant and covered by normal mucosa. On panoramic radiography and computed tomography, a well defined lesion of approximately 1.5 cm in diameter of mixed density was observed. The swelling increased slightly in size over 2 years making it difficult to use prosthetics and, therefore, the lesion was totally excised under local anesthesia, and surgical specimens were submitted for histopathological examination. The histopathological diagnosis was focal cemento-osseous dysplasia. In the present case, because of the increasing size of the swelling making it difficult to use prosthetics, young age of the patient and localization of the lesion, in the initial examination, cemento-ossifying fibroma was suspected, and the lesion was excised surgically; the histopathological diagnosis confirmed it as focal cemento-osseous dysplasia. We present a case of expansive focal cemento-osseous dysplasia. Differential diagnosis

  14. Achromatic phase shifting focal plane masks

    NASA Astrophysics Data System (ADS)

    Newman, Kevin

    The search for life on other worlds is an exciting scientific endeavor that could change the way we perceive our place in the universe. Thousands of extrasolar planets have been discovered using indirect detection techniques. One of the most promising methods for discovering new exoplanets and searching for life is direct imaging with a coronagraph. Exoplanet coronagraphy of Earth-like planets is a challenging task, but we have developed many of the tools necessary to make it feasible. The Phase-Induced Amplitude Apodization (PIAA) Coronagraph is one of the highest-performing architectures for direct exoplanet imaging. With a complex phase-shifting focal plane mask, the PIAA Complex Mask Coronagraph (PIAACMC) can approach the theoretical performance limit for any direct detection technique. The architecture design is flexible enough to be applied to any arbitrary aperture shape, including segmented and obscured apertures. This is an important feature for compatibility with next-generation ground and space-based telescopes. PIAA and PIAACMC focal plane masks have been demonstrated in monochromatic light. An important next step for high-performance coronagraphy is the development of broadband phase-shifting focal plane masks. In this dissertation, we present an algorithm for designing the PIAA and PIAACMC focal plane masks to operate in broadband. We also demonstrate manufacturing of the focal plane masks, and show laboratory results. We use simulations to show the potential performance of the coronagraph system, and the use of wavefront control to correct for mask manufacturing errors. Given the laboratory results and simulations, we show new areas of exoplanet science that can potentially be explored using coronagraph technology. The main conclusion of this dissertation is that we now have the tools required to design and manufacture PIAA and PIAACMC achromatic focal plane masks. These tools can be applied to current and future telescope systems to enable new

  15. The Interaction between the Third Type III Domain from Fibronectin and Anastellin Involves β-Strand Exchange.

    PubMed

    Stine, Jessica M; Ahl, Gabriel J H; Schlenker, Casey; Rusnac, Domnita-Valeria; Briknarová, Klára

    2017-09-05

    Anastellin is a small recombinant fragment derived from the extracellular matrix protein fibronectin; it comprises the first type III (FN3) domain without the two N-terminal β-strands. It inhibits angiogenesis, tumor growth, and metastasis in mouse models and requires endogenous fibronectin for its in vivo anti-angiogenic activity. It binds to fibronectin in vitro and converts the soluble protein to insoluble fibrils that structurally and functionally resemble fibronectin fibrils deposited in the extracellular matrix by cells. Anastellin binds to several FN3 domains in fibronectin, but how it interacts with these domains and why the interactions lead to aggregation of fibronectin are not well understood. In this work, we investigated the interaction between anastellin and the third FN3 domain (3FN3) from fibronectin. We show that anastellin binds with high affinity to a peptide comprising the two N-terminal β-strands from 3FN3, and we present here the structure of the resulting complex. The peptide and anastellin form a composite FN3 domain, with the two N-terminal β-strands from 3FN3 bound in place of the two β-strands that are missing in anastellin. We also demonstrate using disulfide cross-linking that a similar interaction involving the two N-terminal β-strands of 3FN3 occurs when intact 3FN3 binds to anastellin. 3FN3 adopts a compact globular fold in solution, and to interact with anastellin in a manner consistent with our data, it has to open up and expose a β-strand edge that is not accessible in the context of the folded domain.

  16. Layer-by-layer construction of the heparin/fibronectin coatings on titanium surface:stability and functionality

    NASA Astrophysics Data System (ADS)

    Li, Guicai; Yang, Ping; Huang, Nan

    Layer-by-layer assembly as a versatile bottom-up nanofabrication technique has been widely used in the development of biomimetic materials with superior mechanical and biological properties. In this study, layer-by-layer assembled heparin/fibronectin biofunctional films were fabricated on titanium (Ti) surface to enhance the blood anticoagulation and accelerate the endothelialization simultaneously. The wettability and chemical changes of the assembled films were investigated by static water contact angle measurement and fourier transform infrared spectroscopy (FTIR). The morphology of modified Ti surfaces were observed using scanning electron microscopy (SEM). The real time assembly process was in-situ monitored by quartz crystal microbalance with dissipation (QCM-D). The stability of the films was evaluated by measuring the changes in wettability and the quantity of heparin and fibronectin on the surfaces. The anticoagulation properties of the films were quantitatively rated using Activated partial thromboplastin time (APTT) analysis. New peaks of hydroxyl and amino group were observed on the assembled Ti srufaces by FTIR. The contact angles varied among the films with different bilayer numbers, indicating the successful graft of the heparin and fibronectin layer-by-layer. QCM-D results showed that the frequency shift increased with the bilayer numbers, and the heparin and fibronectin could form multilayers. The assembly films were stable after incubation in PBS for 24 h based on the results of the contact angle measurement and the quantity of heparin and fibronectin analysis. APTT results suggested that the assembled films kept excellent antithrombotic properties. All these results revealed that the assembled heparin/fibronectin films with stabiltiy and anticoagulation property could be firmly formed on titanium surfaces. Our study further demonstrates that layer-by-layer assembly of heparin and fibronectin will provide a potential and effective tool for

  17. Identification of a novel glycoprotein (AGp110) involved in interactions of rat liver parenchymal cells with fibronectin

    PubMed Central

    1990-01-01

    We have identified an integral membrane glycoprotein in rat liver that mediates adhesion of cultured hepatocytes on fibronectin substrata. The protein was isolated by affinity chromatography of detergent extracts on wheat germ lectin-Agarose followed by chromatography of the WGA binding fraction on fibronectin-Sepharose. The glycoprotein (AGp110), eluted at high salt concentrations from the fibronectin column, has a molecular mass of 110 kD and a pI of 4.2. Binding of immobilized AGp110 to soluble rat plasma fibronectin required Ca2+ ions but was not inhibited by RGD peptides. Fab' fragments of immunoglobulins raised in rabbits against AGp110 reversed the spreading of primary hepatocytes attached onto fibronectin-coated substrata, but had no effect on cells spread on type IV collagen or laminin substrata. The effect of the antiserum on cell spreading was reversible. AGp110 was detected by immunofluorescence around the periphery of the ventral surface of substratum attached hepatocytes, and scattered on the dorsal surface. Immunohistochemical evidence and Western blotting of fractionated liver plasma membranes indicated a bile canalicular (apical) localization of AGp110 in the liver parenchyma. Expression of AGp110 is tissue specific: it was found mainly in liver, kidney, pancreas, and small intestine but was not detected in stomach, skeletal muscle, heart, and large intestine. AGp110 could be labeled by lactoperoxidase-catalyzed surface iodination of intact liver cells and, after phase partitioning of liver plasma membranes with the detergent Triton X-114, it was preferentially distributed in the hydrophobic phase. Treatment with glycosidases indicated extensive sialic acid substitution in at least 10 O-linked carbohydrate chains and 1-2 N-linked glycans. Immunological comparisons suggest that AGp110, the integrin fibronectin receptor and dipeptidyl peptidase IV, an enzyme involved in fibronectin-mediated adhesion of hepatocytes on collagen, are distinct proteins

  18. The muscle-specific laminin receptor alpha7 beta1 integrin negatively regulates alpha5 beta1 fibronectin receptor function.

    PubMed

    Tomatis, D; Echtermayer, F; Schöber, S; Balzac, F; Retta, S F; Silengo, L; Tarone, G

    1999-02-01

    alpha7 beta1 is the major integrin complex expressed in differentiated muscle cells where it functions as a laminin receptor. In this work we have expressed the alpha7 integrin subunit in CHO cells to investigate the functional properties of this receptor. After transfection with alpha7 CHO cells acquired the ability to adhere and spread on laminin 1 consistent with the laminin receptor activity of the alpha7 beta1. alpha7 transfectants, however, showed a 70% reduction in the ability to adhere to fibronectin and were unable to assemble a fibronectin matrix. The degree of reduction was inversely related to the level of alpha7 expression. To define the mechanisms underlying this adhesive defect we analyzed surface expression and functional properties of the alpha5 beta1 fibronectin receptor. Although cell surface expression of alpha5 beta1 was reduced by a factor of 20-25% in alpha7 transfectants compared to control untransfected cells, this slight reduction was not sufficient to explain the dramatic reduction in cell adhesion (70%) and matrix assembly (close to 100%). Binding studies showed that the affinity of 125I-fibronectin for its surface receptor was decreased by 50% in alpha7 transfectants, indicating that the alpha5 beta1 integrin is partially inactivated in these cells. Inactivation can be reversed by Mn2+, a cation known to increase integrin affinity for their ligands. In fact, incubation of cells with Mn2+ restored fibronectin binding affinity, adhesion to fibronectin, and assembly of fibronectin matrix in alpha7 transfectants. These data indicate that alpha7 expression leads to the functional down regulation of alpha5beta1 integrin by decreasing ligand binding affinity and surface expression. In conclusion, the data reported establish the existence of a negative cooperativity between alpha7 and alpha5 integrins that may be important in determining functional regulation of integrins during myogenic differentiation.

  19. Role of a bacillus Calmette-Guérin fibronectin attachment protein in BCG-induced antitumor activity.

    PubMed

    Zhao, W; Schorey, J S; Bong-Mastek, M; Ritchey, J; Brown, E J; Ratliff, T L

    2000-04-01

    Intravesical Mycobacterium bovis bacillus Calmette-Gu*erin (BCG) is the treatment of choice for superficial bladder cancer. Previous studies showed that attachment of BCG to fibronectin within the bladder was necessary for mediation of the antitumor response. Further studies identified a bacterial receptor, fibronectin attachment protein (FAP), as an important mediator of BCG attachment to fibronectin. In vitro studies showed that a stable BCG/fibronectin interaction was dependent on FAP binding to fibronectin; however, no role for FAP in the attachment of BCG in vivo has been characterized. We now report the cloning of the M. bovis BCG FAP (FAP-B) and demonstrate an important role for FAP in the in vivo attachment of BCG to the bladder wall and in the induction of BCG-mediated antitumor activity. The predicted amino acid sequence for FAP-B shows 61% and 71% homology, respectively, with Mycobacterium avium FAP (FAP-A) and Mycobacterium leprae FAP (FAP-L). Rabbit polyclonal antibodies against Mycobacterium vaccae FAP (FAP-V) reacted with all 3 recombinant FAP proteins on Western blots. Functional studies show FAP-B to bind fibronectin via the highly conserved attachment regions previously identified for FAP-A and FAP-L and also to competitively inhibit attachment of BCG to matrix fibronectin. In vivo studies show FAP to be a necessary protein for the stable attachment of BCG to the bladder wall. Moreover, stable binding of BCG via FAP was shown to be necessary for the expression of BCG-induced antitumor activity. Our results demonstrate a biological role for FAP in the mediation of BCG-induced antitumor activity.

  20. Crystal diffraction lens with variable focal length

    DOEpatents

    Smither, Robert K.

    1991-01-01

    A method and apparatus for altering the focal length of a focusing element o one of a plurality of pre-determined focal lengths by changing heat transfer within selected portions of the element by controlled quantities. Control over heat transfer is accomplished by manipulating one or more of a number of variables, including: the amount of heat or cold applied to surfaces; type of fluids pumped through channels for heating and cooling; temperatures, directions of flow and rates of flow of fluids; and placement of channels.

  1. Crystal diffraction lens with variable focal length

    DOEpatents

    Smither, R.K.

    1991-04-02

    A method and apparatus for altering the focal length of a focusing element of one of a plurality of pre-determined focal lengths by changing heat transfer within selected portions of the element by controlled quantities is disclosed. Control over heat transfer is accomplished by manipulating one or more of a number of variables, including: the amount of heat or cold applied to surfaces; type of fluids pumped through channels for heating and cooling; temperatures, directions of flow and rates of flow of fluids; and placement of channels. 19 figures.

  2. Staring Focal Plane Array System Modeling

    DTIC Science & Technology

    1989-12-01

    C’, DTIC SELECTENI ODEC271 D B mU STARING FOCAL PLANE ARRAY SYSTEM MODELING THESIS John Gerard Murphy Captain, USAF AFIT/GEO/ENP/89D- 3 DEPARTMENT OF...m mmmmmmmm .. \\FlIT/GEO/ENP/89D- 3 STARING FOCAL PLANE ARRAY SYSTEM MODELING THESIS Presented to the Faculty of the School of Engineering of the Air...Background. .. .. ... ... .... ... .... ..... 1-2 1.1.1 MRTD .. .. .. ... ... .... .... ..... 1-2 01.1.2 MIRTD in Modeling. .. .. .. ... .... .... 1- 3 113MRTD

  3. [Antiperspirants for the therapy of focal hyperhidrosis].

    PubMed

    Streker, M; Kerscher, M

    2012-06-01

    In Europe often no clear distinction is made between deodorant and antiperspirant. Particularly in Germany, the labeling "deo" is used for both. Only antiperspirants are capable of influencing the activity of eccrine sweat glands. In the treatment of focal hyperhidrosis, the use of aluminum chloride solutions represents the first choice. The efficacy is well documented in a variety of studies. Subjective side effects include pruritus and - less often - irritant dermatitis, which can be treated symptomatically and usually does not require discontinuation of the treatment. Rare variants of focal hyperhidrosis like auriculotemporal syndrome, Ross syndrome and nevus sudoriferus also are suitable for treatment with topical aluminum chloride hexahydrate solutions.

  4. [Liver ultrasound: focal lesions and diffuse diseases].

    PubMed

    Segura Grau, A; Valero López, I; Díaz Rodríguez, N; Segura Cabral, J M

    2016-01-01

    Liver ultrasound is frequently used as a first-line technique for the detection and characterization of the most common liver lesions, especially those incidentally found focal liver lesions, and for monitoring of chronic liver diseases. Ultrasound is not only used in the Bmode, but also with Doppler and, more recently, contrast-enhanced ultrasound. It is mainly used in the diagnosis of diffuse liver diseases, such as steatosis or cirrhosis. This article presents a practical approach for diagnosis workup, in which the different characteristics of the main focal liver lesions and diffuse liver diseases are reviewed.

  5. Natural course of symptomatic focal choroidal excavation.

    PubMed

    Pierro, Luisa; Casalino, Giuseppe; Introini, Ugo; Gagliardi, Marco; Sergenti, Jessica; Cascavilla, Maria Lucia; Bandello, Francesco

    2015-01-01

    A 32-year-old man was referred to the authors' department for nonspecified macular dystrophy with persistent metamorphopsia in the right eye diagnosed 10 years before and followed using optical coherence tomography. The patient underwent a comprehensive ocular examination, including multimodal imaging evaluation and electrofunctional testing. The diagnosis was consistent with nonconforming focal choroid excavation. Over 10 years, no complications occurred, visual acuity was stable, and optical coherence tomography showed no progression of the lesion during follow-up. In this case, nonconforming symptomatic focal choroid excavation was a nonprogressive condition with good long-term visual outcome.

  6. A simple method for focal length measurement

    NASA Astrophysics Data System (ADS)

    Ma, Hua; Ren, Huan; Zhang, Lin; Shi, Zhengdong; Yuan, Quan; Yang, Yi

    2016-09-01

    A simple method for focal length measurement based on image processing is demonstrated and discussed. The collimated beam, detector, motorized translation stage and computer make up of this test system. The two spots pass through the tested lens is accepted by detector, which is transferred twice by motorized translation stage. By acquired the difference of two spots by image processing, the focal length of the tested lens can be gained. The error sources in the measurement are analyzed. Then the results of experiment show that the relative error was 0.1%. This method can be used in workshop and labs for its convenience and low cost.

  7. Extensive Focal Epithelial Hyperplasia: A Case Report.

    PubMed

    Mansouri, Zahra; Bakhtiari, Sedigheh; Noormohamadi, Robab

    2015-01-01

    Focal epithelial hyperplasia (FEH) or Heck's disease is a rare viral infection of the oral mucosa caused by human papilloma virus especially subtypes 13 or 32. The frequency of this disease varies widely from one geographic region and ethnic groups to another. This paper reports an Iranian case of extensive focal epithelial hyperplasia. A 35-year-old man with FEH is described, in whom the lesions had persisted for more than 25 years. The lesion was diagnosed according to both clinical and histopathological features. Dental practitioner should be aware of these types of lesions and histopathological examination together and a careful clinical observation should be carried out for a definitive diagnosis.

  8. Actinic Granuloma with Focal Segmental Glomerulosclerosis

    PubMed Central

    Phasukthaworn, Ruedee; Chanprapaph, Kumutnart; Vachiramon, Vasanop

    2016-01-01

    Actinic granuloma is an uncommon granulomatous disease, characterized by annular erythematous plaque with central clearing predominately located on sun-damaged skin. The pathogenesis is not well understood, ultraviolet radiation is recognized as precipitating factor. We report a case of a 52-year-old woman who presented with asymptomatic annular erythematous plaques on the forehead and both cheeks persisting for 2 years. The clinical presentation and histopathologic findings support the diagnosis of actinic granuloma. During that period of time, she also developed focal segmental glomerulosclerosis. The association between actinic granuloma and focal segmental glomerulosclerosis needs to be clarified by further studies. PMID:27293392

  9. Syncope and Driving.

    PubMed

    Guzman, Juan C; Morillo, Carlos A

    2015-08-01

    The occurrence of syncope while driving has obvious implications for personal and public safety. Neurally mediated syncope is the most common type of syncope in general and, thereby, also while driving. The presence of structural heart disease (reduced ejection fraction, previous myocardial infarction, significant congenital heart disease) potentially leads to high risk and should determine driving restrictions pending clarification of underlying heart disease and etiology of syncope. The clinical approach to syncope evaluation and recommendations for driving should not differ, whether or not the syncopal spell occurred while driving.

  10. Reconfigurable Drive Current System

    NASA Technical Reports Server (NTRS)

    Alhorn, Dean C. (Inventor); Dutton, Kenneth R. (Inventor); Howard, David E. (Inventor); Smith, Dennis A. (Inventor)

    2017-01-01

    A reconfigurable drive current system includes drive stages, each of which includes a high-side transistor and a low-side transistor in a totem pole configuration. A current monitor is coupled to an output of each drive stage. Input channels are provided to receive input signals. A processor is coupled to the input channels and to each current monitor for generating at least one drive signal using at least one of the input signals and current measured by at least one of the current monitors. A pulse width modulation generator is coupled to the processor and each drive stage for varying the drive signals as a function of time prior to being supplied to at least one of the drive stages.

  11. Drill drive mechanism

    DOEpatents

    Dressel, Michael O.

    1979-01-01

    A drill drive mechanism is especially adapted to provide both rotational drive and axial feed for a drill of substantial diameter such as may be used for drilling holes for roof bolts in mine shafts. The drill shaft is made with a helical pattern of scroll-like projections on its surface for removal of cuttings. The drill drive mechanism includes a plurality of sprockets carrying two chains of drive links which are arranged to interlock around the drill shaft with each drive link having depressions which mate with the scroll-like projections. As the chain links move upwardly or downwardly the surfaces of the depressions in the links mate with the scroll projections to move the shaft axially. Tangs on the drive links mate with notch surfaces between scroll projections to provide a means for rotating the shaft. Projections on the drive links mate together at the center to hold the drive links tightly around the drill shaft. The entire chain drive mechanism is rotated around the drill shaft axis by means of a hydraulic motor and gear drive to cause rotation of the drill shaft. This gear drive also connects with a differential gearset which is interconnected with a second gear. A second motor is connected to the spider shaft of the differential gearset to produce differential movement (speeds) at the output gears of the differential gearset. This differential in speed is utilized to drive said second gear at a speed different from the speed of said gear drive, this speed differential being utilized to drive said sprockets for axial movement of said drill shaft.

  12. Placental inflammation and its relationship to cervicovaginal fetal fibronectin in preterm birth.

    PubMed

    van der Krogt, Laura; Ridout, Alexandra E; Seed, Paul T; Shennan, Andrew H

    2017-07-01

    Late miscarriage and preterm birth are frequently thought to be associated with inflammation and infection, although in most cases the underlying cause of early delivery remains unknown. The placenta is the organ that links mother and fetus during pregnancy, and postnatal examination may provide useful information about pathophysiology. The relationship between placental pathological lesions and predictive markers of early delivery has not been explored. We sought to characterize preterm deliveries according to placental pathology and relate these to the performance of reliable predictive markers, fetal fibronectin and cervical length. This is a retrospective subanalysis from a larger prospective cohort study on sonographic cervical length, quantitative fetal fibronectin and risk of spontaneous preterm birth. Our cohort was comprised of high-risk asymptomatic women attending the Prematurity Surveillance Clinic at St Thomas' Hospital between 2002 and 2015, who went on to have a late miscarriage or preterm delivery (16-36(+6) weeks') and who had available placental histology. The placental pathology of these preterm deliveries was characterized according to the lesions identified, and categorized (according to the Redman classification) into inflammatory (e.g. chorioamnionitis) or non-inflammatory (histologically normal or vascular lesions indicating e.g. malperfusion). We sought to relate placental findings to the performance of reliable predictive markers, in women who delivered early. Standard clinical cut offs for cervical length (<25mm) and fetal fibronectin (>50ng/mL) were used to identify the proportion of preterm births that were accurately predicted by the tests or who showed a false negative result, in relation to their placental histology findings. Binomial logistic regression was carried out to evaluate the relationship between placental inflammation, quantitative fFN and cervical length as continuous variables. 105 women who had a late miscarriage or

  13. MTI Focal Plane Assembly Design and Performance

    SciTech Connect

    Ballard, M.; Rienstra, J.L.

    1999-06-17

    The focal plane assembly for the Multispectral Thermal Imager (MTI) consists of sensor chip assemblies, optical filters, and a vacuum enclosure. Sensor chip assemblies, composed of linear detector arrays and readout integrated circuits, provide spatial resolution in the cross-track direction for the pushbroom imager. Optical filters define 15 spectral bands in a range from 0.45 {micro}m to 10.7 {micro}m. All the detector arrays are mounted on a single focal plane and are designed to operate at 75 K. Three pairs of sensor chip assemblies (SCAs) are required to provide cross-track coverage in all 15 spectral bands. Each pair of SCAs includes detector arrays made from silicon, iridium antimonide, and mercury cadmium telluride. Read out integrated circuits multiplex the signals from the detectors to 18 separate video channels. Optical filter assemblies defining the spectral bands are mounted over the linear detector arrays. Each filter assembly consists of several filter strips bonded together side-by-side. The MTI focal plane assembly has been integrated with the rest of the payload and has undergone detailed testing and calibration. This paper includes representative test data for the various spectral bands and the overall performance of the focal plane assembly.

  14. Towards Dualband Megapixel QWIP Focal Plane Arrays

    NASA Technical Reports Server (NTRS)

    Gunapala, S. D.; Bandara, S. V.; Liu, J. K.; Mumolo, J. M.; Hill, C. J.; Rafol, S. B.; Salazar, D.; Woolaway, J.; LeVan, P. D.; Tidrow, M. Z.

    2006-01-01

    Mid-wavelength infrared (MWIR) and long-wavelength infrared (LWIR) 1024 x 1024 pixel quantum well infrared photodetector (QWIP) focal planes have been demonstrated with excellent imaging performance. The MWIR QWIP detector array has demonstrated a noise equivalent differential temperature (NEDT) of 17 mK at a 95 K operating temperature with f/2.5 optics at 300 K background and the LWIR detector array has demonstrated a NEDT of 13 mK at a 70 K operating temperature with the same optical and background conditions as the MWIR detector array after the subtraction of system noise. Both MWIR and LWIR focal planes have shown background limited performance (BLIP) at 90 K and 70 K operating temperatures respectively, with similar optical and background conditions. In addition, we have demonstrated MWIR and LWIR pixel co-registered simultaneously readable dualband QWIP focal plane arrays. In this paper, we will discuss the performance in terms of quantum efficiency, NEDT, uniformity, operability, and modulation transfer functions of the 1024 x 1024 pixel arrays and the progress of dualband QWIP focal plane array development work.

  15. Sensory-motor integration in focal dystonia.

    PubMed

    Avanzino, Laura; Tinazzi, Michele; Ionta, Silvio; Fiorio, Mirta

    2015-12-01

    Traditional definitions of focal dystonia point to its motor component, mainly affecting planning and execution of voluntary movements. However, focal dystonia is tightly linked also to sensory dysfunction. Accurate motor control requires an optimal processing of afferent inputs from different sensory systems, in particular visual and somatosensory (e.g., touch and proprioception). Several experimental studies indicate that sensory-motor integration - the process through which sensory information is used to plan, execute, and monitor movements - is impaired in focal dystonia. The neural degenerations associated with these alterations affect not only the basal ganglia-thalamic-frontal cortex loop, but also the parietal cortex and cerebellum. The present review outlines the experimental studies describing impaired sensory-motor integration in focal dystonia, establishes their relationship with changes in specific neural mechanisms, and provides new insight towards the implementation of novel intervention protocols. Based on the reviewed state-of-the-art evidence, the theoretical framework summarized in the present article will not only result in a better understanding of the pathophysiology of dystonia, but it will also lead to the development of new rehabilitation strategies.

  16. Dual band QWIP focal plane array

    NASA Technical Reports Server (NTRS)

    Gunapala, Sarath D. (Inventor); Choi, Kwong Kit (Inventor); Bandara, Sumith V. (Inventor)

    2005-01-01

    A quantum well infrared photodetector (QWIP) that provides two-color image sensing. Two different quantum wells are configured to absorb two different wavelengths. The QWIPs are arrayed in a focal plane array (FPA). The two-color QWIPs are selected for readout by selective electrical contact with the two different QWIPs or by the use of two different wavelength sensitive gratings.

  17. Pure Gerstmann's syndrome from a focal lesion.

    PubMed

    Roeltgen, D P; Sevush, S; Heilman, K M

    1983-01-01

    It is controversial whether a focal lesion can specifically induce Gerstmann's syndrome (dyscalculia, left-right disorientation, finger agnosia, and agraphia). Also, Gerstmann's tetrad has been attributed to other cerebral symptoms, particularly aphasia. We examined a patient who had all four symptoms of Gerstmann's syndrome, without other symptoms or signs, and who had a discrete left parietal lesion.

  18. Universities: A Focal Point for Economic Development.

    ERIC Educational Resources Information Center

    Maidique, Modesto A.

    1988-01-01

    Higher education can act as a focal point of economic development. The most widely recognized type of economic development entails an association between a university, its research facilities, and private industry. An example of this partnership is the one between Stanford University and the industries in the "Silicon Valley." (MLW)

  19. Large Format Multicolor QWIP Focal Plane Arrays

    NASA Technical Reports Server (NTRS)

    Soibel, A.; Gunapala, S. D.; Bandara, S. V.; Liu, J. K.; Mumolo, J. M.; Ting, D. Z.; Hill, C. J.; Nguyen, J.

    2009-01-01

    Mid-wave infrared (MWIR) and long-wave infrared (LWIR) multicolor focal plane array (FPA) cameras are essential for many DoD and NASA applications including Earth and planetary remote sensing. In this paper we summarize our recent development of large format multicolor QWIP FPA that cover MWIR and LWIR bands.

  20. [Focal epithelial hyperplasia. An unusual clinical aspect].

    PubMed

    Bodokh, I; Lacour, J P; Rainero, C; Orth, G; Perrin, C; Hoffman, P; Santini, J; Ortonne, J P

    1993-01-01

    We report a case of focal epithelial hyperplasia in a child born in France of Algerian parents. The clinical appearance was unusual in that certain lesions were verrucous and pediculate. A virological study revealed the presence of papillomavirus 32, one of the two types of HPV specifically associated with this entity.

  1. Optical interconnections to focal plane arrays

    SciTech Connect

    Rienstra, J.L.; Hinckley, M.K.

    2000-11-01

    The authors have successfully demonstrated an optical data interconnection from the output of a focal plane array to the downstream data acquisition electronics. The demonstrated approach included a continuous wave laser beam directed at a multiple quantum well reflectance modulator connected to the focal plane array analog output. The output waveform from the optical interconnect was observed on an oscilloscope to be a replica of the input signal. They fed the output of the optical data link to the same data acquisition system used to characterize focal plane array performance. Measurements of the signal to noise ratio at the input and output of the optical interconnection showed that the signal to noise ratio was reduced by a factor of 10 or more. Analysis of the noise and link gain showed that the primary contributors to the additional noise were laser intensity noise and photodetector receiver noise. Subsequent efforts should be able to reduce these noise sources considerably and should result in substantially improved signal to noise performance. They also observed significant photocurrent generation in the reflectance modulator that imposes a current load on the focal plane array output amplifier. This current loading is an issue with the demonstrated approach because it tends to negate the power saving feature of the reflectance modulator interconnection concept.

  2. Genetics Home Reference: focal dermal hypoplasia

    MedlinePlus

    ... HYPOPLASIA Sources for This Page Clements SE, Mellerio JE, Holden ST, McCauley J, McGrath JA. PORCN gene ... on PubMed Clements SE, Wessagowit V, Lai-Cheong JE, Arita K, McGrath JA. Focal dermal hypoplasia resulting ...

  3. Sonographic features of focal Hashimoto's thyroiditis.

    PubMed

    Fu, Xian-shui; Li, Zhi-qiang; Zhang, Hua-bin; Wang, Jin-rui

    2014-06-01

    To evaluate the diagnostic values of gray-scale and color Doppler ultrasound in the diagnosis of focal Hashimoto's thyroiditis (FHT). The gray-scale and colour Doppler ultrasound data of 120 patients with histopathologically confirmed FHT were retrospectively studied. The background of thyroid parenchyma was subjectively evaluated as absence or presence of diffuse Hashimoto's thyroiditis. The vascular types of thyroid nodules were determined as follows: hypovascularity, marked internal flow, marked peripheral flow, and focal thyroid inferno. Among all 120 nodules, 71 (59.2%) were located in normal thyroid parenchyma, 49 (40.8%) in the background of diffuse Hashimoto's thyroiditis. In the normal thyroid background, hypoechoic nodules accounted for 85.9% (61/71), and only 14.1% (10/71)nodules were isoechoic or hyperechoic. However, in the background of diffuse Hashimoto's thyroiditis, 40.8% were hypoechoic and 59.2% were isoechoic or hyperechoic. The difference was statistically significant (P<0.001). In terms of the vascular types, the hypovascularity, marked internal flow, marked peripheral flow, and focal thyroid inferno accounted for 45 %, 25.9%, 20.8%, and 8.3%, respectively. The "focal inferno" is a relative specific color Doppler feature of FHT.

  4. Estimation of focal and extra-focal radiation profiles based on Gaussian modeling in medical linear accelerators.

    PubMed

    Anai, Shigeo; Arimura, Hidetaka; Nakamura, Katsumasa; Araki, Fujio; Matsuki, Takaomi; Yoshikawa, Hideki; Yoshidome, Satoshi; Shioyama, Yoshiyuki; Honda, Hiroshi; Ikeda, Nobuo

    2011-07-01

    The X-ray source or focal radiation is one of the factors that can degrade the conformal field edge in stereotactic body radiotherapy. For that reason, it is very important to estimate the total focal radiation profiles of linear accelerators, which consists of X-ray focal-spot radiation and extra-focal radiation profiles. Our purpose in this study was to propose an experimental method for estimating the focal-spot and extra-focal radiation profiles of linear accelerators based on triple Gaussian functions. We measured the total X-ray focal radiation profiles of the accelerators by moving a slit in conjunction with a photon field p-type silicon diode. The slit width was changed so that the extra-focal radiation could be optimally included in the total focal radiation. The total focal radiation profiles of an accelerator at 4-MV and 10-MV energies were approximated with a combination of triple Gaussian functions, which correspond to the focal-spot radiation, extra-focal radiation, and radiation transmitted through the slit assembly. As a result, the ratios of the Gaussian peak value of the extra-focal radiation to that of the focal spot for 4 and 10 MV were 0.077 and 0.159, respectively. The peak widths of the focal-spot and extra-focal radiation profiles were 0.57 and 25.0 mm for 4 MV, respectively, and 0.60 and 22.0 mm for 10 MV, respectively. We concluded that the proposed focal radiation profile model based on the triple Gaussian functions may be feasible for estimating the X-ray focal-spot and extra-focal radiation profiles.

  5. Chronic cardiac allograft rejection: critical role of ED-A(+) fibronectin and implications for targeted therapy strategies.

    PubMed

    Franz, Marcus; Neri, Dario; Berndt, Alexander

    2012-03-01

    Chronic cardiac allograft rejection is characterized by cardiac allograft vasculopathy (CAV) and cardiac interstitial fibrosis (CIF) causing severe long-term complications after heart transplantation and determining allograft function and patients' prognosis. Until now, there have been no sufficient preventive or therapeutic strategies. CAV and CIF are accompanied by changes in the extracellular matrix, including re-expression of the fetal fibronectin splice variant known as ED-A(+) fibronectin. This molecule has been shown to be crucial for the development of myofibroblasts (MyoFbs) as the main cell type in CIF and for the activation of vascular smooth muscle cells (VSMCs) as the main cell type in CAV. Relevant re-expression and protein deposition of ED-A(+) fibronectin has been demonstrated in animal models of chronic rejection, with spatial association to CAV and CIF, and a quantitative correlation to the rejection grade. The paper by Booth et al published in this issue of The Journal of Pathology could prove for the first time the functional importance of ED-A(+) fibronectin for the development of CIF as a main component of chronic cardiac rejection. Thus, promising conclusions for the development of new diagnostic, preventive, and therapeutic strategies for chronic cardiac rejection focusing on ED-A(+) fibronectin can be suggested.

  6. Fibronectin induction abrogates the BRAF inhibitor response of BRAF V600E/PTEN-null melanoma cells.

    PubMed

    Fedorenko, I V; Abel, E V; Koomen, J M; Fang, B; Wood, E R; Chen, Y A; Fisher, K J; Iyengar, S; Dahlman, K B; Wargo, J A; Flaherty, K T; Sosman, J A; Sondak, V K; Messina, J L; Gibney, G T; Smalley, K S M

    2016-03-10

    The mechanisms by which some melanoma cells adapt to Serine/threonine-protein kinase B-Raf (BRAF) inhibitor therapy are incompletely understood. In the present study, we used mass spectrometry-based phosphoproteomics to determine how BRAF inhibition remodeled the signaling network of melanoma cell lines that were BRAF mutant and PTEN null. Short-term BRAF inhibition was associated with marked changes in fibronectin-based adhesion signaling that were PTEN dependent. These effects were recapitulated through BRAF siRNA knockdown and following treatment with chemotherapeutic drugs. Increased fibronectin expression was also observed in mouse xenograft models as well as specimens from melanoma patients undergoing BRAF inhibitor treatment. Analysis of a melanoma tissue microarray showed loss of PTEN expression to predict for a lower overall survival, with a trend for even lower survival being seen when loss of fibronectin was included in the analysis. Mechanistically, the induction of fibronectin limited the responses of these PTEN-null melanoma cell lines to vemurafenib, with enhanced cytotoxicity observed following the knockdown of either fibronectin or its receptor α5β1 integrin. This in turn abrogated the cytotoxic response to BRAF inhibition via increased AKT signaling, which prevented the induction of cell death by maintaining the expression of the pro-survival protein Mcl-1. The protection conveyed by the induction of FN expression could be overcome through combined treatment with a BRAF and PI3K inhibitor.

  7. Novel Potential Interacting Partners of Fibronectin in Spontaneous Animal Model of Interstitial Cystitis

    PubMed Central

    Treutlein, Gudrun; Dorsch, Roswitha; Euler, Kerstin N.; Hauck, Stefanie M.; Amann, Barbara; Hartmann, Katrin; Deeg, Cornelia A.

    2012-01-01

    Feline idiopathic cystitis (FIC) is the only spontaneous animal model for human interstitial cystitis (IC), as both possess a distinctive chronical and relapsing character. Underlying pathomechanisms of both diseases are not clearly established yet. We recently detected increased urine fibronectin levels in FIC cases. The purpose of this study was to gain further insight into the pathogenesis by assessing interacting partners of fibronectin in urine of FIC affected cats. Several candidate proteins were identified via immunoprecipitation and mass spectrometry. Considerable changes in FIC conditions compared to physiological expression of co-purified proteins were detected by Western blot and immunohistochemistry. Compared to controls, complement C4a and thioredoxin were present in higher levels in urine of FIC patients whereas loss of signal intensity was detected in FIC affected tissue. Galectin-7 was exclusively detected in urine of FIC cats, pointing to an important role of this molecule in FIC pathogenesis. Moderate physiological signal intensity of galectin-7 in transitional epithelium shifted to distinct expression in transitional epithelium under pathophysiological conditions. I-FABP expression was reduced in urine and urinary bladder tissue of FIC cats. Additionally, transduction molecules of thioredoxin, NF-κB p65 and p38 MAPK, were examined. In FIC affected tissue, colocalization of thioredoxin and NF-κB p65 could be demonstrated compared to absent coexpression of thioredoxin and p38 MAPK. These considerable changes in expression level and pattern point to an important role for co-purified proteins of fibronectin and thioredoxin-regulated signal transduction pathways in FIC pathogenesis. These results could provide a promising starting point for novel therapeutic approaches in the future. PMID:23236492

  8. Leishmania infection modulates beta-1 integrin activation and alters the kinetics of monocyte spreading over fibronectin

    PubMed Central

    Figueira, Cláudio Pereira; Carvalhal, Djalma Gomes Ferrão; Almeida, Rafaela Andrade; Hermida, Micely d’ El-Rei; Touchard, Dominique; Robert, Phillipe; Pierres, Anne; Bongrand, Pierre; dos-Santos, Washington LC

    2015-01-01

    Contact with Leishmania leads to a decreases in mononuclear phagocyte adherence to connective tissue. In this work, we studied the early stages of bond formation between VLA4 and fibronectin, measured the kinetics of membrane alignment and the monocyte cytoplasm spreading area over a fibronectin-coated surface, and studied the expression of high affinity integrin epitope in uninfected and Leishmania-infected human monocytes. Our results show that the initial VLA4-mediated interaction of Leishmania-infected monocyte with a fibronectin-coated surface is preserved, however, the later stage, leukocyte spreading over the substrate is abrogated in Leishmania-infected cells. The median of spreading area was 72 [55–89] μm2 for uninfected and 41 [34–51] μm2 for Leishmania-infected monocyte. This cytoplasm spread was inhibited using an anti-VLA4 blocking antibody. After the initial contact with the fibronectrin-coated surface, uninfected monocyte quickly spread the cytoplasm at a 15 μm2 s−1 ratio whilst Leishmania-infected monocytes only made small contacts at a 5.5 μm2 s−1 ratio. The expression of high affinity epitope by VLA4 (from 39 ± 21% to 14 ± 3%); and LFA1 (from 37 ± 32% to 18 ± 16%) molecules was reduced in Leishmania-infected monocytes. These changes in phagocyte function may be important for parasite dissemination and distribution of lesions in leishmaniasis. PMID:26249106

  9. Leishmania infection modulates beta-1 integrin activation and alters the kinetics of monocyte spreading over fibronectin.

    PubMed

    Figueira, Cláudio Pereira; Carvalhal, Djalma Gomes Ferrão; Almeida, Rafaela Andrade; Hermida, Micely d' El-Rei; Touchard, Dominique; Robert, Phillipe; Pierres, Anne; Bongrand, Pierre; dos-Santos, Washington L C

    2015-08-07

    Contact with Leishmania leads to a decreases in mononuclear phagocyte adherence to connective tissue. In this work, we studied the early stages of bond formation between VLA4 and fibronectin, measured the kinetics of membrane alignment and the monocyte cytoplasm spreading area over a fibronectin-coated surface, and studied the expression of high affinity integrin epitope in uninfected and Leishmania-infected human monocytes. Our results show that the initial VLA4-mediated interaction of Leishmania-infected monocyte with a fibronectin-coated surface is preserved, however, the later stage, leukocyte spreading over the substrate is abrogated in Leishmania-infected cells. The median of spreading area was 72 [55-89] μm(2) for uninfected and 41 [34-51] μm(2) for Leishmania-infected monocyte. This cytoplasm spread was inhibited using an anti-VLA4 blocking antibody. After the initial contact with the fibronectrin-coated surface, uninfected monocyte quickly spread the cytoplasm at a 15 μm(2) s(-1) ratio whilst Leishmania-infected monocytes only made small contacts at a 5.5 μm(2) s(-1) ratio. The expression of high affinity epitope by VLA4 (from 39 ± 21% to 14 ± 3%); and LFA1 (from 37 ± 32% to 18 ± 16%) molecules was reduced in Leishmania-infected monocytes. These changes in phagocyte function may be important for parasite dissemination and distribution of lesions in leishmaniasis.

  10. A Processed Multidomain Mycoplasma hyopneumoniae Adhesin Binds Fibronectin, Plasminogen, and Swine Respiratory Cilia

    PubMed Central

    Seymour, Lisa M.; Deutscher, Ania T.; Jenkins, Cheryl; Kuit, Tracey A.; Falconer, Linda; Minion, F. Chris; Crossett, Ben; Padula, Matthew; Dixon, Nicholas E.; Djordjevic, Steven P.; Walker, Mark J.

    2010-01-01

    Porcine enzootic pneumonia is a chronic respiratory disease that affects swine. The etiological agent of the disease, Mycoplasma hyopneumoniae, is a bacterium that adheres to cilia of the swine respiratory tract, resulting in loss of cilia and epithelial cell damage. A M. hyopneumoniae protein P116, encoded by mhp108, was investigated as a potential adhesin. Examination of P116 expression using proteomic analyses observed P116 as a full-length protein and also as fragments, ranging from 17 to 70 kDa in size. A variety of pathogenic bacterial species have been shown to bind the extracellular matrix component fibronectin as an adherence mechanism. M. hyopneumoniae cells were found to bind fibronectin in a dose-dependent and saturable manner. Surface plasmon resonance was used to show that a recombinant C-terminal domain of P116 bound fibronectin at physiologically relevant concentrations (KD 24 ± 6 nm). Plasmin(ogen)-binding proteins are also expressed by many bacterial pathogens, facilitating extracellular matrix degradation. M. hyopneumoniae cells were found to also bind plasminogen in a dose-dependent and saturable manner; the C-terminal domain of P116 binds to plasminogen (KD 44 ± 5 nm). Plasminogen binding was abolished when the C-terminal lysine of P116 was deleted, implicating this residue as part of the plasminogen binding site. P116 fragments adhere to the PK15 porcine kidney epithelial-like cell line and swine respiratory cilia. Collectively these data suggest that P116 is an important adhesin and virulence factor of M. hyopneumoniae. PMID:20813843

  11. A processed multidomain mycoplasma hyopneumoniae adhesin binds fibronectin, plasminogen, and swine respiratory cilia.

    PubMed

    Seymour, Lisa M; Deutscher, Ania T; Jenkins, Cheryl; Kuit, Tracey A; Falconer, Linda; Minion, F Chris; Crossett, Ben; Padula, Matthew; Dixon, Nicholas E; Djordjevic, Steven P; Walker, Mark J

    2010-10-29

    Porcine enzootic pneumonia is a chronic respiratory disease that affects swine. The etiological agent of the disease, Mycoplasma hyopneumoniae, is a bacterium that adheres to cilia of the swine respiratory tract, resulting in loss of cilia and epithelial cell damage. A M. hyopneumoniae protein P116, encoded by mhp108, was investigated as a potential adhesin. Examination of P116 expression using proteomic analyses observed P116 as a full-length protein and also as fragments, ranging from 17 to 70 kDa in size. A variety of pathogenic bacterial species have been shown to bind the extracellular matrix component fibronectin as an adherence mechanism. M. hyopneumoniae cells were found to bind fibronectin in a dose-dependent and saturable manner. Surface plasmon resonance was used to show that a recombinant C-terminal domain of P116 bound fibronectin at physiologically relevant concentrations (K(D) 24 ± 6 nm). Plasmin(ogen)-binding proteins are also expressed by many bacterial pathogens, facilitating extracellular matrix degradation. M. hyopneumoniae cells were found to also bind plasminogen in a dose-dependent and saturable manner; the C-terminal domain of P116 binds to plasminogen (K(D) 44 ± 5 nm). Plasminogen binding was abolished when the C-terminal lysine of P116 was deleted, implicating this residue as part of the plasminogen binding site. P116 fragments adhere to the PK15 porcine kidney epithelial-like cell line and swine respiratory cilia. Collectively these data suggest that P116 is an important adhesin and virulence factor of M. hyopneumoniae.

  12. Decipher the dynamic coordination between enzymatic activity and structural modulation at focal adhesions in living cells

    NASA Astrophysics Data System (ADS)

    Lu, Shaoying; Seong, Jihye; Wang, Yi; Chang, Shiou-Chi; Eichorst, John Paul; Ouyang, Mingxing; Li, Julie Y.-S.; Chien, Shu; Wang, Yingxiao

    2014-07-01

    Focal adhesions (FAs) are dynamic subcellular structures crucial for cell adhesion, migration and differentiation. It remains an enigma how enzymatic activities in these local complexes regulate their structural remodeling in live cells. Utilizing biosensors based on fluorescence resonance energy transfer (FRET), we developed a correlative FRET imaging microscopy (CFIM) approach to quantitatively analyze the subcellular coordination between the enzymatic Src activation and the structural FA disassembly. CFIM reveals that the Src kinase activity only within the microdomain of lipid rafts at the plasma membrane is coupled with FA dynamics. FA disassembly at cell periphery was linearly dependent on this raft-localized Src activity, although cells displayed heterogeneous levels of response to stimulation. Within lipid rafts, the time delay between Src activation and FA disassembly was 1.2 min in cells seeded on low fibronectin concentration ([FN]) and 4.3 min in cells on high [FN]. CFIM further showed that the level of Src-FA coupling, as well as the time delay, was regulated by cell-matrix interactions, as a tight enzyme-structure coupling occurred in FA populations mediated by integrin αvβ3, but not in those by integrin α5β1. Therefore, different FA subpopulations have distinctive regulation mechanisms between their local kinase activity and structural FA dynamics.

  13. Imaging modalities in Focal Therapy: Multiparametric Ultrasound.

    PubMed

    Wildeboer, R R; Panfilova, A P; Mischi, M; Wijkstra, H

    2016-07-01

    Prostate cancer (PCa) is the most common form of cancer among men in the US and the second most common cause of death. It has been observed that an increasing number of newly diagnosed patients exhibit low-risk features and that over-treatment with radical prostatectomy is a growing problem. The feasibility of focal therapy as an organsparing alternative, however, depends on the reliability of imaging techniques to identify, localize and monitor clinically relevant PCa lesions. The aim of this review is to investigate the potential of multiparametric ultrasound (mpUS) for focal therapy. We briefly introduce the most common focal therapies and thoroughly discuss the ability of available ultrasound modalities to localize PCa and reflect tissue properties. The imaging requirements of the focal therapies are studied to put the performance of the US techniques into perspective. We found that transrectal greyscale echography, Doppler sonography, elastography, contrast-enhanced ultrasonography and computerized ultrasound have been studied for the purpose of prostate imaging. Several of these modalities are already frequently used in current clinical practice; to add to the diagnostic process of PCa, to guide and monitor the application of focal therapy or to perform follow-up after treatment. Despite their capability to detect a large fraction of the PCa lesions, none of these modalities is currently considered sufficiently accurate for stand-alone tumour detection and localization. However, although there are only few studies reporting on a combined use of different ultrasound modalities, the results of an mpUS approach seem promising. Several US modalities have been successfully applied as a viable alternative to monitor tissue destruction during and after treatment. In view of the advantages of US and the promising results of a multiparametric approach in PCa detection and localization, researchers are urged to further investigate mpUS for therapeutic purposes.

  14. Surface displaced alfa-enolase of Lactobacillus plantarum is a fibronectin binding protein

    PubMed Central

    Castaldo, Cristiana; Vastano, Valeria; Siciliano, Rosa Anna; Candela, Marco; Vici, Manuela; Muscariello, Lidia; Marasco, Rosangela; Sacco, Margherita

    2009-01-01

    Background Lactic acid bacteria of the genus Lactobacillus and Bifidobacterium are one of the most important health promoting groups of the human intestinal microbiota. Their protective role within the gut consists in out competing invading pathogens for ecological niches and metabolic substrates. Among the features necessary to provide health benefits, commensal microorganisms must have the ability to adhere to human intestinal cells and consequently to colonize the gut. Studies on mechanisms mediating adhesion of lactobacilli to human intestinal cells showed that factors involved in the interaction vary mostly among different species and strains, mainly regarding interaction between bacterial adhesins and extracellular matrix or mucus proteins. We have investigated the adhesive properties of Lactobacillus plantarum, a member of the human microbiota of healthy individuals. Results We show the identification of a Lactobacillus plantarum LM3 cell surface protein (48 kDa), which specifically binds to human fibronectin (Fn), an extracellular matrix protein. By means of mass spectrometric analysis this protein was identified as the product of the L. plantarum enoA1 gene, coding the EnoA1 alfa-enolase. Surface localization of EnoA1 was proved by immune electron microscopy. In the mutant strain LM3-CC1, carrying the enoA1 null mutation, the 48 kDa adhesin was not anymore detectable neither by anti-enolase Western blot nor by Fn-overlay immunoblotting assay. Moreover, by an adhesion assay we show that LM3-CC1 cells bind to fibronectin-coated surfaces less efficiently than wild type cells, thus demonstrating the significance of the surface displaced EnoA1 protein for the L. plantarum LM3 adhesion to fibronectin. Conclusion Adhesion to host tissues represents a crucial early step in the colonization process of either pathogens or commensal bacteria. We demonstrated the involvement of the L. plantarum Eno A1 alfa-enolase in Fn-binding, by studying LM3 and LM3-CC1 surface

  15. Accuracy of cervicovaginal fetal fibronectin test in predicting risk of spontaneous preterm birth: systematic review

    PubMed Central

    Honest, Honest; Bachmann, Lucas M; Gupta, Janesh K; Kleijnen, Jos; Khan, Khalid S

    2002-01-01

    Objective To determine the accuracy with which a cervicovaginal fetal fibronectin test predicts spontaneous preterm birth in women with or without symptoms of preterm labour. Design Systematic quantitative review of studies of test accuracy. Data sources Medline, Embase, PASCAL, Biosis, Cochrane Library, Medion, National Research Register, SCISEARCH, conference papers, manual searching of bibliographies of known primary and review articles, and contact with experts and manufacturer. Study selection Two reviewers independently selected and extracted data on study characteristics, quality, and accuracy. Data extraction Accuracy data were used to form 2×2 contingency tables with spontaneous preterm birth before 34 and 37 weeks' gestation and birth within 7-10 days of testing (for symptomatic pregnant women) as reference standards. Data were pooled to produce summary receiver operating characteristic curves and summary likelihood ratios for positive and negative test results. Data synthesis 64 primary articles were identified, consisting of 28 studies in asymptomatic women and 40 in symptomatic women, with a total of 26 876 women. Among asymptomatic women the best summary likelihood ratio for positive results was 4.01 (95% confidence interval 2.93 to 5.49) for predicting birth before 34 weeks' gestation, with corresponding summary likelihood ratio for negative results of 0.78 (0.72 to 0.84). Among symptomatic women the best summary likelihood ratio for positive results was 5.42 (4.36 to 6.74) for predicting birth within 7-10 days of testing, with corresponding ratio for negative results of 0.25 (0.20 to 0.31). Conclusion Cervicovaginal fetal fibronectin test is most accurate in predicting spontaneous preterm birth within 7-10 days of testing among women with symptoms of threatened preterm birth before advanced cervical dilatation. What is already known on this topicSpontaneous preterm birth is a major cause of neonatal morbidity and mortalityIf spontaneous preterm

  16. Nanofibrous glass tailored with apatite-fibronectin interface for bone cell stimulation.

    PubMed

    Kim, Hae-Won; Lee, Hae-Hyoung; Knowles, Jonathan C

    2008-06-01

    Exploring a material with smart and biomimetic interface has great potential in the biomaterials and tissue engineering field. This paper reports a novel nanofibrous bone matrix that was developed to retain a cell-stimulating and bone-mimetic biointerface. The bone mineral, apatite, and the cell adhesive protein, fibronectin (FN), were hybridized on the interface of a bioactive glass nanofibrous mesh, through the dissolution-and-reprecipitation process. The hybridized nanofibrous mesh showed significant improvement in the initial responses of the bone-derived cells. It is believed that this biomimetic and cell-stimulating nanofibrous mesh can be used as a potential bone regeneration matrix.

  17. Focal adhesion kinase activity is required for actomyosin contractility-based invasion of cells into dense 3D matrices

    PubMed Central

    Mierke, Claudia T.; Fischer, Tony; Puder, Stefanie; Kunschmann, Tom; Soetje, Birga; Ziegler, Wolfgang H.

    2017-01-01

    The focal adhesion kinase (FAK) regulates the dynamics of integrin-based cell adhesions important for motility. FAK’s activity regulation is involved in stress-sensing and focal-adhesion turnover. The effect of FAK on 3D migration and cellular mechanics is unclear. We analyzed FAK knock-out mouse embryonic fibroblasts and cells expressing a kinase-dead FAK mutant, R454-FAK, in comparison to FAK wild-type cells. FAK knock-out and FAKR454/R454 cells invade dense 3D matrices less efficiently. These results are supported by FAK knock-down in wild-type fibroblasts and MDA-MB-231 human breast cancer cells showing reduced invasiveness. Pharmacological interventions indicate that in 3D matrices, cells deficient in FAK or kinase-activity behave similarly to wild-type cells treated with inhibitors of Src-activity or actomyosin-contractility. Using magnetic tweezers experiments, FAKR454/R454 cells are shown to be softer and exhibit impaired adhesion to fibronectin and collagen, which is consistent with their reduced 3D invasiveness. In line with this, FAKR454/R454 cells cannot contract the matrix in contrast to FAK wild-type cells. Finally, our findings demonstrate that active FAK facilitates 3D matrix invasion through increased cellular stiffness and transmission of actomyosin-dependent contractile force in dense 3D extracellular matrices. PMID:28202937

  18. Nano-Stenciled RGD-Gold Patterns That Inhibit Focal Contact Maturation Induce Lamellipodia Formation in Fibroblasts

    PubMed Central

    Lutz, Roman; Pataky, Kristopher; Gadhari, Neha; Marelli, Mattia; Brugger, Juergen; Chiquet, Matthias

    2011-01-01

    Cultured fibroblasts adhere to extracellular substrates by means of cell-matrix adhesions that are assembled in a hierarchical way, thereby gaining in protein complexity and size. Here we asked how restricting the size of cell-matrix adhesions affects cell morphology and behavior. Using a nanostencil technique, culture substrates were patterned with gold squares of a width and spacing between 250 nm and 2 µm. The gold was functionalized with RGD peptide as ligand for cellular integrins, and mouse embryo fibroblasts were plated. Limiting the length of cell-matrix adhesions to 500 nm or less disturbed the maturation of vinculin-positive focal complexes into focal contacts and fibrillar adhesions, as indicated by poor recruitment of α5-integrin. We found that on sub-micrometer patterns, fibroblasts spread extensively, but did not polarize. Instead, they formed excessive numbers of lamellipodia and a fine actin meshwork without stress fibers. Moreover, these cells showed aberrant fibronectin fibrillogenesis, and their speed of directed migration was reduced significantly compared to fibroblasts on 2 µm square patterns. Interference with RhoA/ROCK signaling eliminated the pattern-dependent differences in cell morphology. Our results indicate that manipulating the maturation of cell-matrix adhesions by nanopatterned surfaces allows to influence morphology, actin dynamics, migration and ECM assembly of adhering fibroblasts. PMID:21980465

  19. Spatial focalization of pheromone/MAPK signaling triggers commitment to cell-cell fusion.

    PubMed

    Dudin, Omaya; Merlini, Laura; Martin, Sophie G

    2016-10-01

    Cell fusion is universal in eukaryotes for fertilization and development, but what signals this process is unknown. Here, we show in Schizosaccharomyces pombe that fusion does not require a dedicated signal but is triggered by spatial focalization of the same pheromone-GPCR (G-protein-coupled receptor)-MAPK signaling cascade that drives earlier mating events. Autocrine cells expressing the receptor for their own pheromone trigger fusion attempts independently of cell-cell contact by concentrating pheromone release at the fusion focus, a dynamic actin aster underlying the secretion of cell wall hydrolases. Pheromone receptor and MAPK cascade are similarly enriched at the fusion focus, concomitant with fusion commitment in wild-type mating pairs. This focalization promotes cell fusion by immobilizing the fusion focus, thus driving local cell wall dissolution. We propose that fusion commitment is imposed by a local increase in MAPK concentration at the fusion focus, driven by a positive feedback between fusion focus formation and focalization of pheromone release and perception.

  20. Spatial focalization of pheromone/MAPK signaling triggers commitment to cell–cell fusion

    PubMed Central

    Merlini, Laura

    2016-01-01

    Cell fusion is universal in eukaryotes for fertilization and development, but what signals this process is unknown. Here, we show in Schizosaccharomyces pombe that fusion does not require a dedicated signal but is triggered by spatial focalization of the same pheromone–GPCR (G-protein-coupled receptor)–MAPK signaling cascade that drives earlier mating events. Autocrine cells expressing the receptor for their own pheromone trigger fusion attempts independently of cell–cell contact by concentrating pheromone release at the fusion focus, a dynamic actin aster underlying the secretion of cell wall hydrolases. Pheromone receptor and MAPK cascade are similarly enriched at the fusion focus, concomitant with fusion commitment in wild-type mating pairs. This focalization promotes cell fusion by immobilizing the fusion focus, thus driving local cell wall dissolution. We propose that fusion commitment is imposed by a local increase in MAPK concentration at the fusion focus, driven by a positive feedback between fusion focus formation and focalization of pheromone release and perception. PMID:27798845

  1. X-ray sources of medical linear accelerators: focal and extra-focal radiation.

    PubMed

    Jaffray, D A; Battista, J J; Fenster, A; Munro, P

    1993-01-01

    A computerized tomography (CT) reconstruction technique has been used to make quantitative measurements of the size and shape of the focal spot in medical linear accelerators. Using this technique, we have measured the focal spots in a total of nine accelerators, including (i) two Varian Clinac 2100c's, (ii) two Atomic Energy of Canada Ltd. (AECL) Therac-25's, (iii) two AECL Therac 6's, (iv) a Siemens KD-2, (v) a Varian Clinac 600c (4 MV), and (vi) an AECL Therac-20. Some of these focal spots were monitored for changes over a 2-yr period. It has been found that (i) the size and shape of the source spot varies greatly between accelerators of different design ranging from 0.5 to 3.4 mm in full width at half maximum (FWHM); and (ii) for accelerators of the same design, the focal spots are very similar. In addition to the measurements of the focal spot, a new technique for measuring the magnitude and distribution of extra-focal radiation originating from the linear accelerator head (flattening filter, primary collimator) has also been developed. The extra-focal radiation produced by a Varian Clinac 2100c accelerator was measured using this technique and it was found that the extra-focal radiation accounts for as much as 8% of the total photon fluence reaching the isocenter. The majority (75%) of this extra-focal radiation originates from within a circle 6 cm in diameter at the target plane. The source MTFs for each of the measured focal spots have been calculated in order to assess their influence on the spatial resolution of verification images. The limiting spatial resolution (i.e., 10% modulation) for all the source MTFs is 1.8 mm-1 or greater when used for transmission radiography at a magnification of 1.2. The extra-focal radiation, which produces a low-frequency drop in the source MTFs of up to 8%, changes with field size. As a result, the source MTFs of linear accelerators depend not only on the design of individual accelerators and image magnification, but also

  2. Restoring Conscious Arousal During Focal Limbic Seizures with Deep Brain Stimulation.

    PubMed

    Kundishora, Adam J; Gummadavelli, Abhijeet; Ma, Chanthia; Liu, Mengran; McCafferty, Cian; Schiff, Nicholas D; Willie, Jon T; Gross, Robert E; Gerrard, Jason; Blumenfeld, Hal

    2016-03-03

    Impaired consciousness occurs suddenly and unpredictably in people with epilepsy, markedly worsening quality of life and increasing risk of mortality. Focal seizures with impaired consciousness are the most common form of epilepsy and are refractory to all current medical and surgical therapies in about one-sixth of cases. Restoring consciousness during and following seizures would be potentially transformative for these individuals. Here, we investigate deep brain stimulation to improve level of conscious arousal in a rat model of focal limbic seizures. We found that dual-site stimulation of the central lateral nucleus of the intralaminar thalamus (CL) and the pontine nucleus oralis (PnO) bilaterally during focal limbic seizures restored normal-appearing cortical electrophysiology and markedly improved behavioral arousal. In contrast, single-site bilateral stimulation of CL or PnO alone was insufficient to achieve the same result. These findings support the "network inhibition hypothesis" that focal limbic seizures impair consciousness through widespread inhibition of subcortical arousal. Driving subcortical arousal function would be a novel therapeutic approach to some forms of refractory epilepsy and may be compatible with devices already in use for responsive neurostimulation. Multisite deep brain stimulation of subcortical arousal structures may benefit not only patients with epilepsy but also those with other disorders of consciousness.

  3. Hybrid drive arrangement

    SciTech Connect

    Oetting, H.; Heidemeyer, P.

    1985-02-19

    The invention concerns a hybrid drive arrangement for vehicles, with an engine drive and with a flywheel storage drive, which includes a storage flywheel supported concentrically relative to the crankshaft for storing kinetic energy during such operations as braking operations of the vehicle. Both drives can be connected with the driving wheels of the vehicle by means of a common, preferably continuously variable, transmission. In order to obtain a faster response of the engine drive on suddenly occurring power demands and in order to achieve a more favorable design of the storage flywheel, there is to be provided in accordance with the invention, in addition to a storage flywheel, an engine flywheel associated with the reciprocating-piston internal combustion engine, which compensates for torque irregularities of the engine. The engine flywheel can be connected with a crankshaft by means of a first clutch and with the storage flywheel by means of at least one further clutch.

  4. Up-regulation of gastric cancer cell invasion by Twist is accompanied by N-cadherin and fibronectin expression.

    PubMed

    Yang, Zhou; Zhang, Xiaohong; Gang, Haiju; Li, Xiaojun; Li, Zumao; Wang, Tao; Han, Juan; Luo, Ting; Wen, Fuqiang; Wu, Xiaoting

    2007-07-06

    Twist, a newly found EMT-inducer, has been reported to be up-regulated in those of diffuse-type gastric carcinomas with high N-cadherin level. We show here MKN45, a cell line derived from undifferentiated carcinomas cells, expresses high levels of Twist. Down-regulation of Twist, using an antisense Twist vector in MKN45 cells, inhibits cell migration and invasion, companied with a morphologic changes associated with MET. Suppression of Twist also decreases the expressions of N-cadherin and fibronectin, but not of E-cadherin in MKN45. In contrast, overexpression of Twist in MKN28, a cell line derived from moderate differentiated carcinomas, results in up-regulation of N-cadherin and fibronectin, companied with down-regulation of E-cadherin. Taken together, our results suggest that Twist regulates cell motility and invasion in gastric cancer cell lines, probably through the N-cadherin and fibronectin production.

  5. Marihuana and driving.

    PubMed

    Moskowitz, H

    1985-08-01

    A review was performed of the marihuana and driving literature, both epidemiological and experimental. It was noted that epidemiological studies face considerable difficulties in obtaining estimates of risks involved for drivers utilizing marihuana due to the rapid decline in blood levels of tetrahydrocannabinol. On the other hand, experimental studies examining the relationship between administered marihuana dose and performance have identified many driving-related areas as exhibiting impairment. Areas impaired include coordination, tracking, perception, vigilance and performance in both driving simulators and on the road. Other behavioral areas of lesser importance for driving also exhibited evidence of impairment by marihuana. Areas for further research are suggested.

  6. Vehicle drive system

    SciTech Connect

    Kurata, N.

    1986-08-19

    A vehicle is described having driving wheels both on the left and right sides of the chassis frame thereof comprising: a power unit including an engine and a transmission system for transmitting the power from an output shaft of the engine to the driving wheels independently. The power unit has a casing constructed as a rigid member for supporting the driving wheels and pivotally connected through a pivot shaft to the chassis frame so as to permit vertical movement of the driving wheels, the transmission system including a differential gear means having a case connected through speed reduction gears to the output shaft. The differential gear means include left and right side output gears, the transmission system including left and right input drive shafts extending laterally from the left and right side output gears of the differential gear means. The transmission system includes left and right output sections to which the input drive shafts are drivingly connected and output shafts connected to the respective driving wheels, and the output section including V-belt type automatic transmissions connected between the input drive shafts and the output shafts.

  7. DESI focal plate mechanical integration and cooling

    NASA Astrophysics Data System (ADS)

    Lambert, A. R.; Besuner, R. W.; Claybaugh, T. M.; Silber, J. H.

    2016-08-01

    The Dark Energy Spectroscopic Instrument (DESI) is under construction to measure the expansion history of the Universe using the Baryon Acoustic Oscillation technique[1]. The spectra of 40 million galaxies over 14000 sq. deg will be measured during the life of the experiment. A new prime focus corrector for the KPNO Mayall telescope will deliver light to 5000 fiber optic positioners. The fibers in turn feed ten broad-band spectrographs. This paper describes the mechanical integration of the DESI focal plate and the thermal system design. The DESI focal plate is comprised of ten identical petal assemblies. Each petal contains 500 robotic fiber positioners. Each petal is a complete, self-contained unit, independent from the others, with integrated power supply, controllers, fiber routing, and cooling services. The major advantages of this scheme are: (1) supports installation and removal of complete petal assemblies in-situ, without disturbing the others, (2) component production, assembly stations, and test procedures are repeated and parallelizable, (3) a complete, full-scale prototype can be built and tested at an early date, (4) each production petal can be surveyed and tested as a complete unit, prior to integration, from the fiber tip at the focal surface to the fiber slit at the spectrograph. The ten petal assemblies will be installed in a single integration ring, which is mounted to the DESI corrector. The aluminum integration ring attaches to the steel corrector barrel via a flexured steel adapter, isolating the focal plate from differential thermal expansions. The plate scale will be kept stable by conductive cooling of the petal assembly. The guider and wavefront sensors (one per petal) will be convectively cooled by forced flow of air. Heat will be removed from the system at ten liquid-cooled cold plates, one per petal, operating at ambient temperature. The entire focal plate structure is enclosed in an insulating shroud, which serves as a thermal barrier

  8. Maternal soluble vascular cytoplasmic adhesion molecule-1 and fibronectin levels in early- and late-onset preeclamptic pregnancies.

    PubMed

    Dogan, E; Demir, S Cansun; Gulec, U Kucukgoz

    2014-01-01

    The purpose of this study was to investigate maternal plasma soluble vascular cytoplasmic adhesion molecule-1 (sVCAM-1) and fibronectin levels in the patients with early-onset preeclampsia (EOP) and late-onset preeclampsia (LOP) and also to determine whether different mechanisms are involved in these two forms of disorders. The authors performed a case control study consisting of randomly selected 80 healthy pregnant women (group 1 = control group) and 80 preeclamptic women (group 2 = defined study group). Study group consisted of43 patients with EOP and 37 patients with LOP. sVCAM-1 and fibronectin concentrations were measured by enzyme-linked immunosorbent assay (ELISA) and the findings were compared between the groups. The mean levels of sVCAM-1 and fibronectin were significantly higher in the LOP group than those in the normotensive group (p = 0.043 and 0.010 respectively). Markers were significantly different between the two hypertensive groups of pregnancy. The EOP group had a higher level of sVCAM-1 and fibronectin concentration than the LOP group (p = 0.01, for both markers). There was a positive correlation both between the values of plasma fibronectin and the systolic-diastolic blood pressure measurements (r:0.43 and 0.44, respectively), and between sVCAM-1 and the systolic/diastolic blood pressure measurements (r = 0.54 and 0.64, respectively). Increased plasma levels of fibronectin and sVCAM-1 were found in the preeclamptic patients, especially in those with early-onset preeclampsia. These markers might be related to the pathogenesis of different types of preeclampsia.

  9. [Histological study of cell attachment and proliferation of periodontal ligament cells on the tooth surface. Effect of the fibronectin application].

    PubMed

    Takigami, T

    1989-09-01

    The purpose of this study was to examine the effect of fibronectin on cell attachment and the growth of cultured cells on the surface of the dentin. In the first experiment, dentin specimens were divided into following three groups: dentin surface polished with carborundum-point, carborundum-point and #150 sandpaper and carborundum-point, #150 sandpaper and #240 sandpaper. The roughness of the surface was measured with a profilometer (Surfcom e-st-sa) for the purpose of comparing differences among the three groups. The foregoing treated samples were added to cultures of periodontal ligament (PDL) cells and following fixation, the number of cells was counted under a light microscope and examination under a scanning electron microscope (SEM) was also performed. 1. Greater numbers of cells attached to the smooth surface (#240) than the rough surface. 2. SEM observations showed increased numbers of attached and proliferated cells on the smooth surface. In the second experiment, the dentin surfaces were polished with carborundum point, #150 and #240 sand paper. The dentin tube side was treated with fibronectin. MEM-treated samples not immersed in fibronectin were employed as controls. The foregoing treated root samples were cultured with PDL cells. Following fixation, the cells attached to the root surface were examined under the light microscope and using a SEM. 1. A significant increase in the number of the attached cells as observed