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Sample records for beta1 integrins differentially

  1. Beta1 integrins differentially control extravasation of inflammatory cell subsets into the CNS during autoimmunity

    DEFF Research Database (Denmark)

    Bauer, Martina; Brakebusch, Cord; Coisne, Caroline;

    2009-01-01

    )-integrin gene either in all hematopoietic cells or selectively in T lymphocytes. Our results show that T cells critically rely on beta(1) integrins to accumulate in the central nervous system (CNS) during EAE, whereas CNS infiltration of beta(1)-deficient myeloid cells remains unaffected, suggesting that T...

  2. ADAM12 and alpha9beta1 integrin are instrumental in human myogenic cell differentiation

    DEFF Research Database (Denmark)

    Lafuste, Peggy; Sonnet, Corinne; Chazaud, Bénédicte; Dreyfus, Patrick A; Gherardi, Romain K; Wewer, Ulla M; Authier, François-Jérôme

    2005-01-01

    of alpha9 parallels that of ADAM12 and culminates at time of fusion. alpha9 and ADAM12 coimmunoprecipitate and participate to mpc adhesion. Inhibition of ADAM12/alpha9beta1 integrin interplay, by either ADAM12 antisense oligonucleotides or blocking antibody to alpha9beta1, inhibited overall mpc...

  3. Up-regulation of the integrin alpha 1/beta 1 in human neuroblastoma cells differentiated by retinoic acid: correlation with increased neurite outgrowth response to laminin.

    OpenAIRE

    Rossino, P; P. Defilippi; Silengo, L; Tarone, G.

    1991-01-01

    Retinoic acid (RA) is known to induce differentiation of neuroblastoma cells in vitro. Here we show that treatment of two human neuroblastoma cell lines, SY5Y and IMR32, with RA resulted in a fivefold increase of the integrin alpha 1/beta 1 expression. The effect was selective because expression of the alpha 3/beta 1 integrin, also present in these cells, was not increased. The up-regulation of the alpha 1/beta 1 differentiated SY5Y cells correlated with increased neurite response to laminin....

  4. Genetic analysis of beta1 integrin "activation motifs" in mice

    DEFF Research Database (Denmark)

    Czuchra, Aleksandra; Meyer, Hannelore; Legate, Kyle R;

    2006-01-01

    /beta tails, leading to tail separation and integrin activation. We analyzed mice in which we mutated the tyrosines of the beta1 tail and the membrane-proximal aspartic acid required for the salt bridge. Tyrosine-to-alanine substitutions abolished beta1 integrin functions and led to a beta1 integrin...

  5. Staged stromal extracellular 3D matrices differentially regulate breast cancer cell responses through PI3K and beta1-integrins

    International Nuclear Information System (INIS)

    Interactions between cancer cells and stroma are critical for growth and invasiveness of epithelial tumors. The biochemical mechanisms behind tumor-stromal interactions leading to increased invasiveness and metastasis are mostly unknown. The goal of this study was to analyze the direct effects of staged stroma-derived extracellular matrices on breast cancer cell behavior. Early and late three-dimensional matrices were produced by NIH-3T3 and tumor-associated murine fibroblasts, respectively. After removing fibroblasts, extracted matrices were re-cultured with breast epithelial cells of assorted characteristics: MCF-10A (non-tumorigenic), MCF-7 (tumorigenic, non-invasive), and MDA-MB-231 (tumorigenic, invasive). Effects prompted by staged matrices on epithelial cell's growth, morphology and invasion were determined. Also, matrix-induced velocity, directionality and relative track orientation of invasive cells were assessed in the presence or absence of inhibitors of phosphoinositide-3 kinase (PI3K) and/or beta-1 integrin. We observed that assorted breast epithelial cells reacted differently to two-dimensional vs. staged, control (early) and tumor-associated (late), three-dimensional matrices. MCF-10A had a proliferative advantage on two-dimensional substrates while MCF-7 and MDA-MB-231 showed no difference. MCF-10A and MCF-7 formed morphologically distinguishable aggregates within three-dimensional matrices, while MDA-MB-231 exhibited increased spindle-shape morphologies and directional movements within three-dimensional matrices. Furthermore, MDA-MB-231 acquired a pattern of parallel oriented organization within tumor-associated, but not control matrices. Moreover, tumor-associated matrices induced PI3K and beta1-integrin dependent Akt/PKB activity in MDA-MB-231 cells. Interestingly, beta1-integrin (but not PI3K) regulated tumor-associated matrix-induced mesenchymal invasion which, when inhibited, resulted in a change of invasive strategy rather than impeding

  6. Beta 1 integrin is essential for teratoma growth and angiogenesis

    DEFF Research Database (Denmark)

    Bloch, W; Forsberg, E; Lentini, S;

    1997-01-01

    Teratomas are benign tumors that form after ectopic injection of embryonic stem (ES) cells into mice and contain derivatives of all primitive germ layers. To study the role of beta 1 integrin during teratoma formation, we compared teratomas induced by normal and beta1-null ES cells. Injection of...... normal ES cells gave rise to large teratomas. In contrast, beta 1-null ES cells either did not grow or formed small teratomas with an average weight of <5% of that of normal teratomas. Histological analysis of beta 1-null teratomas revealed the presence of various differentiated cells, however, a much...... lower number of host-derived stromal cells than in normal teratomas. Fibronectin, collagen I, and nidogen were expressed but, in contrast to normal teratomas, diffusely deposited in beta1-null teratomas. Basement membranes were present but with irregular shape and detached from the cell surface. Normal...

  7. Conditional beta1-integrin gene deletion in neural crest cells causes severe developmental alterations of the peripheral nervous system

    DEFF Research Database (Denmark)

    Pietri, Thomas; Eder, Olivier; Breau, Marie Anne;

    2004-01-01

    Integrins are transmembrane receptors that are known to interact with the extracellular matrix and to be required for migration, proliferation, differentiation and apoptosis. We have generated mice with a neural crest cell-specific deletion of the beta1-integrin gene to analyse the role of beta1-...... almost complete absence of Schwann cells and sensory axon segregation and defective maturation in neuromuscular synaptogenesis. Thus, beta1-integrins are important for the control of embryonic and postnatal peripheral nervous system development....

  8. ADAM12 induces actin cytoskeleton and extracellular matrix reorganization during early adipocyte differentiation by regulating beta1 integrin function

    DEFF Research Database (Denmark)

    Kawaguchi, Nobuko; Sundberg, Christina; Kveiborg, Marie;

    2003-01-01

    Changes in cell shape are a morphological hallmark of differentiation. In this study we report that the expression of ADAM12, a disintegrin and metalloprotease, dramatically affects cell morphology in preadipocytes, changing them from a flattened, fibroblastic appearance to a more rounded shape. We...... early adipocyte differentiation....

  9. Discoidin domain receptor 1 is activated independently of beta(1) integrin

    DEFF Research Database (Denmark)

    Vogel, W; Brakebusch, C; Fässler, R;

    2000-01-01

    Various types of collagen have been identified as potential ligands for the two mammalian discoidin domain receptor (DDR) tyrosine kinases, DDR1 and DDR2. It is presently unclear whether collagen-induced DDR receptor activation, which occurs with very slow kinetics, involves additional proteins...... blocking antibodies for alpha(2)beta(1) integrin or in cells with a targeted deletion of the beta(1) integrin gene. Finally, we show that overexpression of dominant negative DDR1 in the myoblast cell line C2C12 blocks cellular differentiation and the formation of myofibers....

  10. Fetal and adult hematopoietic stem cells require beta1 integrin function for colonizing fetal liver, spleen, and bone marrow

    DEFF Research Database (Denmark)

    Potocnik, A J; Brakebusch, C; Fässler, R

    2000-01-01

    Homing of hematopoietic stem cells (HSCs) into hematopoietic organs is a prerequisite for the establishment of hematopoiesis during embryogenesis and after bone marrow transplantation. We show that beta1 integrin-deficient HSCs from the para-aortic splanchnopleura and the fetal blood had hematoly......Homing of hematopoietic stem cells (HSCs) into hematopoietic organs is a prerequisite for the establishment of hematopoiesis during embryogenesis and after bone marrow transplantation. We show that beta1 integrin-deficient HSCs from the para-aortic splanchnopleura and the fetal blood had...... hematolymphoid differentiation potential in vitro and in fetal organ cultures but were unable to seed fetal and adult hematopoietic tissues. Adult beta1 integrin null HSCs isolated from mice carrying loxP-tagged beta1 integrin alleles and ablated for beta1 integrin expression by retroviral cre transduction...... failed to engraft irradiated recipient mice. Moreover, absence of beta1 integrin resulted in sequestration of HSCs in the circulation and their reduced adhesion to endothelioma cells. These findings define beta1 integrin as an essential adhesion receptor for the homing of HSCs....

  11. Recombinant laminin-8 (alpha(4)beta(1)gamma(1)). Production, purification,and interactions with integrins.

    Science.gov (United States)

    Kortesmaa, J; Yurchenco, P; Tryggvason, K

    2000-05-19

    Laminins are a large family of heterotrimeric extracellular matrix glycoproteins that, in addition to having structural roles, take part in the regulation of processes such as cell migration, differentiation, and proliferation. The laminin alpha(4) chain is widely distributed both in adults and during development in tissues such as cardiac, skeletal and smooth muscle fibers, vascular endothelia, lungs, and in peripheral nerves. It can associate with laminin beta(1)/gamma(1) chains to form laminin-8 and with the beta(2)/gamma(1) chains to form laminin-9. Functional studies on these laminins have been hampered by poor availability of the protein in pure and soluble forms. To facilitate studies on laminin-8, recombinant laminin-8 was produced in a mammalian expression system, purified and shown to form native Y-shaped molecules in rotary shadowing electron microscopy. Integrins mediating cell adhesion to laminin-8 were identified using function-blocking mAbs. The integrin specificities were found to differ somewhat from that of laminin-1. Integrin alpha(6)beta(1) was found to be a major mediator of adhesion of HT-1080 and cultured capillary endothelial cells to laminin-8. Considering the expression patterns of laminin-8 and integrin alpha(6)beta(1) it is likely that the former is a ligand for the latter in vivo as well. PMID:10809728

  12. Dynamic expression of alpha 1 beta 1 and alpha 2 beta 1 integrin receptors by human vascular smooth muscle cells. Alpha 2 beta 1 integrin is required for chemotaxis across type I collagen-coated membranes.

    OpenAIRE

    Skinner, M P; Raines, E W; Ross, R.

    1994-01-01

    Vascular smooth muscle cells (SMCs) in the media of normal arteries express alpha 1 beta 1 integrin with no detectable alpha 2 beta 1 as determined by immunocytochemistry. In contrast, immunoprecipitation of integrins expressed by human SMCs cultured from medial explants shows strong expression of alpha 2 beta 1 and no expression of alpha 1 beta 1. The apparent reciprocal expression of these two collagen and laminin receptors was confirmed by flow cytometric analysis of fluorescent labeled ce...

  13. Simultaneous changes in the function and expression of beta 1 integrins during the growth arrest of poorly differentiated colorectal cells (LISP-1

    Directory of Open Access Journals (Sweden)

    R.A. Roela

    2003-08-01

    Full Text Available Cells usually lose adhesion and increase proliferation and migration during malignant transformation. Here, we studied how proliferation can affect the other two characteristics, which ultimately lead to invasion and metastasis. We determined the expression of ß1 integrins, as well as adhesion and migration towards laminin-1, fibronectin, collagens type I and type IV presented by LISP-1 colorectal cancer cells exposed to 2.5% dimethyl sulfoxide (DMSO, an agent capable of decreasing proliferation in this poorly differentiated colorectal cell line. Untreated cells (control, as shown by flow cytometry and monoclonal antibodies, expressed alpha2 (63.8 ± 11.3% positive cells, alpha3 (93.3 ± 7.0%, alpha5 (50.4 ± 12.0% and alpha6 (34.1 ± 4.9% integrins but not alpha1, alpha4, alphav or ß4. Cells adhered well to laminin-1 (73.4 ± 6.0% and fibronectin (40.0 ± 2.0% substrates but very little to collagens. By using blocking monoclonal antibodies, we showed that alpha2, alpha3 and alpha6 mediated laminin-1 adhesion, but neither alpha3 nor alpha5 contributed to fibronectin adherence. DMSO arrested cells at G0/G1 (control: 55.0 ± 2.4% vs DMSO: 70.7 ± 2.5% while simultaneously reducing alpha5 (24.2 ± 19% and alpha6 (14.3 ± 10.8% expression as well as c-myc mRNA (7-fold, the latter shown by Northern blotting. Although the adhesion rate did not change after exposure to DMSO, alpha3 and alpha5 played a major role in laminin-1 and fibronectin adhesion, respectively. Migration towards laminin-1, which was clearly increased upon exposure to DMSO (control: 6 ± 2 cells vs DMSO: 64 ± 6 cells, was blocked by an antibody against alpha6. We conclude that the effects of DMSO on LISP-1 proliferation were accompanied by concurrent changes in the expression and function of integrins, consequently modulating adhesion/migration, and revealing a complex interplay between function/expression and the proliferative state of cells.

  14. beta1-integrin-mediated signaling essentially contributes to cell survival after radiation-induced genotoxic injury

    DEFF Research Database (Denmark)

    Cordes, N; Seidler, J; Durzok, R; Geinitz, H; Brakebusch, C

    2006-01-01

    Integrin-mediated adhesion to extracellular matrix proteins confers resistance to radiation- or drug-induced genotoxic injury. To analyse the underlying mechanisms specific for beta1-integrins, wild-type beta1A-integrin-expressing GD25beta1A cells were compared to GD25beta1B cells, which express...... findings in tumor cells, human A-172 glioma cells were examined under the same conditions after siRNA-mediated silencing of beta1-integrins. We found that beta1A-integrin-mediated adhesion to fibronectin, collagen-III or beta1-IgG was essential for cell survival after radiation-induced genotoxic injury...... central role of beta1-integrins in Akt- and p130Cas/paxillin-mediated prosurvival signaling. These findings suggest beta1-integrins as critical regulators of cell survival after radiation-induced genotoxic injury. Elucidation of the molecular circuitry of prosurvival beta1-integrin-mediated signaling in...

  15. The fibronectin-binding integrins alpha5beta1 and alphavbeta3 differentially modulate RhoA-GTP loading, organization of cell matrix adhesions, and fibronectin fibrillogenesis

    DEFF Research Database (Denmark)

    Danen, Erik H J; Sonneveld, Petra; Brakebusch, Cord; Fassler, Reinhard; Sonnenberg, Arnoud

    2002-01-01

    We have studied the formation of different types of cell matrix adhesions in cells that bind to fibronectin via either alpha5beta1 or alphavbeta3. In both cases, cell adhesion to fibronectin leads to a rapid decrease in RhoA activity. However, alpha5beta1 but not alphavbeta3 supports high levels of...... RhoA activity at later stages of cell spreading, which are associated with a translocation of focal contacts to peripheral cell protrusions, recruitment of tensin into fibrillar adhesions, and fibronectin fibrillogenesis. Expression of an activated mutant of RhoA stimulates alphavbeta3-mediated...... fibrillogenesis. Despite the fact that alpha5beta1-mediated adhesion to the central cell-binding domain of fibronectin supports activation of RhoA, other regions of fibronectin are required for the development of alpha5beta1-mediated but not alphavbeta3-mediated focal contacts. Using chimeras of beta1 and beta3...

  16. Mycophenolate mofetil modulates adhesion receptors of the beta1 integrin family on tumor cells: impact on tumor recurrence and malignancy

    Directory of Open Access Journals (Sweden)

    Beecken Wolf-Dietrich

    2005-01-01

    Full Text Available Abstract Background Tumor development remains one of the major obstacles following organ transplantation. Immunosuppressive drugs such as cyclosporine and tacrolimus directly contribute to enhanced malignancy, whereas the influence of the novel compound mycophenolate mofetil (MMF on tumor cell dissemination has not been explored. We therefore investigated the adhesion capacity of colon, pancreas, prostate and kidney carcinoma cell lines to endothelium, as well as their beta1 integrin expression profile before and after MMF treatment. Methods Tumor cell adhesion to endothelial cell monolayers was evaluated in the presence of 0.1 and 1 μM MMF and compared to unstimulated controls. beta1 integrin analysis included alpha1beta1 (CD49a, alpha2beta1 (CD49b, alpha3beta1 (CD49c, alpha4beta1 (CD49d, alpha5beta1 (CD49e, and alpha6beta1 (CD49f receptors, and was carried out by reverse transcriptase-polymerase chain reaction, confocal microscopy and flow cytometry. Results Adhesion of the colon carcinoma cell line HT-29 was strongly reduced in the presence of 0.1 μM MMF. This effect was accompanied by down-regulation of alpha3beta1 and alpha6beta1 surface expression and of alpha3beta1 and alpha6beta1 coding mRNA. Adhesion of the prostate tumor cell line DU-145 was blocked dose-dependently by MMF. In contrast to MMF's effects on HT-29 cells, MMF dose-dependently up-regulated alpha1beta1, alpha2beta1, alpha3beta1, and alpha5beta1 on DU-145 tumor cell membranes. Conclusion We conclude that MMF possesses distinct anti-tumoral properties, particularly in colon and prostate carcinoma cells. Adhesion blockage of HT-29 cells was due to the loss of alpha3beta1 and alpha6beta1 surface expression, which might contribute to a reduced invasive behaviour of this tumor entity. The enhancement of integrin beta1 subtypes observed in DU-145 cells possibly causes re-differentiation towards a low-invasive phenotype.

  17. Mycophenolate mofetil modulates adhesion receptors of the beta1 integrin family on tumor cells: impact on tumor recurrence and malignancy

    International Nuclear Information System (INIS)

    Tumor development remains one of the major obstacles following organ transplantation. Immunosuppressive drugs such as cyclosporine and tacrolimus directly contribute to enhanced malignancy, whereas the influence of the novel compound mycophenolate mofetil (MMF) on tumor cell dissemination has not been explored. We therefore investigated the adhesion capacity of colon, pancreas, prostate and kidney carcinoma cell lines to endothelium, as well as their beta1 integrin expression profile before and after MMF treatment. Tumor cell adhesion to endothelial cell monolayers was evaluated in the presence of 0.1 and 1 μM MMF and compared to unstimulated controls. beta1 integrin analysis included alpha1beta1 (CD49a), alpha2beta1 (CD49b), alpha3beta1 (CD49c), alpha4beta1 (CD49d), alpha5beta1 (CD49e), and alpha6beta1 (CD49f) receptors, and was carried out by reverse transcriptase-polymerase chain reaction, confocal microscopy and flow cytometry. Adhesion of the colon carcinoma cell line HT-29 was strongly reduced in the presence of 0.1 μM MMF. This effect was accompanied by down-regulation of alpha3beta1 and alpha6beta1 surface expression and of alpha3beta1 and alpha6beta1 coding mRNA. Adhesion of the prostate tumor cell line DU-145 was blocked dose-dependently by MMF. In contrast to MMF's effects on HT-29 cells, MMF dose-dependently up-regulated alpha1beta1, alpha2beta1, alpha3beta1, and alpha5beta1 on DU-145 tumor cell membranes. We conclude that MMF possesses distinct anti-tumoral properties, particularly in colon and prostate carcinoma cells. Adhesion blockage of HT-29 cells was due to the loss of alpha3beta1 and alpha6beta1 surface expression, which might contribute to a reduced invasive behaviour of this tumor entity. The enhancement of integrin beta1 subtypes observed in DU-145 cells possibly causes re-differentiation towards a low-invasive phenotype

  18. Alpha9beta1 integrin in melanoma cells can signal different adhesion states for migration and anchorage

    DEFF Research Database (Denmark)

    Lydolph, Magnus C; Morgan-Fisher, Marie; Høye, Anette M;

    2009-01-01

    Cell surface integrins are the primary receptors for cell migration on extracellular matrix, and exist in several activation states regulated in part by ectodomain conformation. The alpha9 integrin subunit, which pairs only with beta1, has specific roles in the immune system and may regulate cell......beta1 integrin- and Rho kinase-dependent focal adhesion and stress fibre formation, suggesting that the activation status of alpha9beta1 integrin was altered. The effect of manganese ions in promoting focal adhesion formation was reproduced by beta1 integrin activating antibody. The alpha9beta1...

  19. alpha 11beta 1 integrin recognizes the GFOGER sequence in interstitial collagens.

    Science.gov (United States)

    Zhang, Wan-Ming; Kapyla, Jarmo; Puranen, J Santeri; Knight, C Graham; Tiger, Carl-Fredrik; Pentikainen, Olli T; Johnson, Mark S; Farndale, Richard W; Heino, Jyrki; Gullberg, Donald

    2003-02-28

    The integrins alpha(1)beta(1), alpha(2)beta(1), alpha(10)beta(1), and alpha(11)beta(1) are referred to as a collagen receptor subgroup of the integrin family. Recently, both alpha(1)beta(1) and alpha(2)beta(1) integrins have been shown to recognize triple-helical GFOGER (where single letter amino acid nomenclature is used, O = hydroxyproline) or GFOGER-like motifs found in collagens, despite their distinct binding specificity for various collagen subtypes. In the present study we have investigated the mechanism whereby the latest member in the integrin family, alpha(11)beta(1), recognizes collagens using C2C12 cells transfected with alpha(11) cDNA and the bacterially expressed recombinant alpha(11) I domain. The ligand binding properties of alpha(11)beta(1) were compared with those of alpha(2)beta(1). Mg(2+)-dependent alpha(11)beta(1) binding to type I collagen required micromolar Ca(2+) but was inhibited by 1 mm Ca(2+), whereas alpha(2)beta(1)-mediated binding was refractory to millimolar concentrations of Ca(2+). The bacterially expressed recombinant alpha(11) I domain preference for fibrillar collagens over collagens IV and VI was the same as the alpha(2) I domain. Despite the difference in Ca(2+) sensitivity, alpha(11)beta(1)-expressing cells and the alpha(11) I domain bound to helical GFOGER sequences in a manner similar to alpha(2)beta(1)-expressing cells and the alpha(2) I domain. Modeling of the alpha I domain-collagen peptide complexes could partially explain the observed preference of different I domains for certain GFOGER sequence variations. In summary, our data indicate that the GFOGER sequence in fibrillar collagens is a common recognition motif used by alpha(1)beta(1), alpha(2)beta(1), and also alpha(11)beta(1) integrins. Although alpha(10) and alpha(11) chains show the highest sequence identity, alpha(2) and alpha(11) are more similar with regard to collagen specificity. Future studies will reveal whether alpha(2)beta(1) and alpha(11)beta(1

  20. Beta1 integrin promotes but is not essential for metastasis of ras-myc transformed fibroblasts

    DEFF Research Database (Denmark)

    Brakebusch, C; Wennerberg, K; Krell, H W; Weidle, U H; Sallmyr, A; Johansson, S; Fässler, R

    1999-01-01

    integrin-expressing sublines. Tumors were induced by subcutaneous injection of GERM 11 cells and 3 independent beta1 integrin expressing sublines (GERM 116, 1A10, 2F2) into syngeneic mice. After 10 days tumors were surgically removed. While average weights of GERM 11 and GERM 116 tumors were similar...

  1. Beta1 integrin expression by podocytes is required to maintain glomerular structural integrity.

    Science.gov (United States)

    Pozzi, Ambra; Jarad, George; Moeckel, Gilbert W; Coffa, Sergio; Zhang, Xi; Gewin, Leslie; Eremina, Vera; Hudson, Billy G; Borza, Dorin-Bogdan; Harris, Raymond C; Holzman, Lawrence B; Phillips, Carrie L; Fassler, Reinhard; Quaggin, Susan E; Miner, Jeffrey H; Zent, Roy

    2008-04-15

    Integrins are transmembrane heteromeric receptors that mediate interactions between cells and extracellular matrix (ECM). beta1, the most abundantly expressed integrin subunit, binds at least 12 alpha subunits. beta1 containing integrins are highly expressed in the glomerulus of the kidney; however their role in glomerular morphogenesis and maintenance of glomerular filtration barrier integrity is poorly understood. To study these questions we selectively deleted beta1 integrin in the podocyte by crossing beta1(flox/flox) mice with podocyte specific podocin-cre mice (pod-Cre), which express cre at the time of glomerular capillary formation. We demonstrate that podocyte abnormalities are visualized during glomerulogenesis of the pod-Cre;beta1(flox/flox) mice and proteinuria is present at birth, despite a grossly normal glomerular basement membrane. Following the advent of glomerular filtration there is progressive podocyte loss and the mice develop capillary loop and mesangium degeneration with little evidence of glomerulosclerosis. By 3 weeks of age the mice develop severe end stage renal failure characterized by both tubulointerstitial and glomerular pathology. Thus, expression of beta1 containing integrins by the podocyte is critical for maintaining the structural integrity of the glomerulus. PMID:18328474

  2. Signaling through urokinase and urokinase receptor in lung cancer cells requires interactions with beta1 integrins.

    Science.gov (United States)

    Tang, Chi-Hui; Hill, Marla L; Brumwell, Alexis N; Chapman, Harold A; Wei, Ying

    2008-11-15

    The urokinase receptor (uPAR) is upregulated upon tumor cell invasion and correlates with poor lung cancer survival. Although a cis-interaction with integrins has been ascribed to uPAR, whether this interaction alone is critical to urokinase (uPA)- and uPAR-dependent signaling and tumor promotion is unclear. Here we report the functional consequences of point mutations of uPAR (H249A-D262A) that eliminate beta1 integrin interactions but maintain uPA binding, vitronectin attachment and association with alphaV integrins, caveolin and epidermal growth factor receptor. Disruption of uPAR interactions with beta1 integrins recapitulated previously reported findings with beta1-integrin-derived peptides that attenuated matrix-dependent ERK activation, MMP expression and in vitro migration by human lung adenocarcinoma cell lines. The uPAR mutant cells acquired enhanced capacity to adhere to vitronectin via uPAR-alphaVbeta5-integrin, rather than through the uPAR-alpha3beta1-integrin complex and they were unable to initiate uPA signaling to activate ERK, Akt or Stat1. In an orthotopic lung cancer model, uPAR mutant cells exhibited reduced tumor size compared with cells expressing wild-type uPAR. Taken together, the results indicate that uPAR-beta1-integrin interactions are essential to signals induced by integrin matrix ligands or uPA that support lung cancer cell invasion in vitro and progression in vivo. PMID:18940913

  3. Mouse egg integrin alpha 6 beta 1 functions as a sperm receptor.

    Science.gov (United States)

    Almeida, E A; Huovila, A P; Sutherland, A E; Stephens, L E; Calarco, P G; Shaw, L M; Mercurio, A M; Sonnenberg, A; Primakoff, P; Myles, D G; White, J M

    1995-06-30

    Binding between sperm and egg plasma membranes is an essential step in fertilization. Whereas fertilin, a mammalian sperm surface protein, is involved in this crucial interaction, sperm receptors on the egg plasma membrane have not been identified. Because fertilin contains a predicted integrin ligand domain, we investigated the expression and function of integrin subunits in unfertilized mouse eggs. Polymerase chain reactions detected mRNAs for alpha 5, alpha 6, alpha v, beta 1, beta 3, and beta 5. Immunofluorescence revealed alpha 6 beta 1 and alpha v beta 3 on the plasma membrane. GoH3, a function-blocking anti-alpha 6 monoclonal antibody, abolished sperm binding, but a nonfunction-blocking anti-alpha 6 monoclonal antibody, a function-blocking anti-alpha v beta 3 polyclonal antibody, and an RGD peptide had no effect. Somatic cells bound sperm avidly, but only if they expressed alpha 6 beta 1. A peptide analog of the fertilin integrin ligand domain inhibited sperm binding to eggs and alpha 6 beta 1+ cells and diminished GoH3 staining of eggs. Our results indicate a novel role for the integrin alpha 6 beta 1 as a cell-cell adhesion receptor that mediates sperm-egg binding. PMID:7600577

  4. Integrin alpha3beta1, a novel receptor for alpha3(IV) noncollagenous domain and a trans-dominant Inhibitor for integrin alphavbeta3.

    Science.gov (United States)

    Borza, Corina M; Pozzi, Ambra; Borza, Dorin-Bogdan; Pedchenko, Vadim; Hellmark, Thomas; Hudson, Billy G; Zent, Roy

    2006-07-28

    Exogenous soluble human alpha3 noncollagenous (NC1) domain of collagen IV inhibits angiogenesis and tumor growth. These biological functions are attributed to the binding of alpha3NC1 to integrin alphavbeta3. However, in some tumor cells that express integrin alphavbeta3, the alpha3NC1 domain does not inhibit proliferation, suggesting that integrin alphavbeta3 expression is not sufficient to mediate the anti-tumorigenic activity of this domain. Therefore, in the present study, we searched for novel binding receptors for the soluble alpha3NC1 domain in cells lacking alphavbeta3 integrin. In these cells, soluble alpha3NC1 bound integrin alpha3beta1; however, unlike alphavbeta3, alpha3beta1 integrin did not mediate cell adhesion to immobilized alpha3NC1 domain. Interestingly, in cells lacking integrin alpha3beta1, adhesion to the alpha3NC1 domain was enhanced due to activation of integrin alphavbeta3. These findings indicate that integrin alpha3beta1 is a receptor for the alpha3NC1 domain and transdominantly inhibits integrin alphavbeta3 activation. Thus integrin alpha3beta1, in conjunction with integrin alphavbeta3, modulates cellular responses to the alpha3NC1 domain, which may be pivotal in the mechanism underpinning its anti-angiogenic and anti-tumorigenic activities. PMID:16731529

  5. ADAM12-mediated focal adhesion formation is differently regulated by beta1 and beta3 integrins

    DEFF Research Database (Denmark)

    Thodeti, Charles Kumar; Frohlich, Camilla; Nielsen, Christian Kamp;

    2005-01-01

    ADAM12, adisintegrin and metalloprotease, has been demonstrated to be upregulated in human malignant tumors and to accelerate the malignant phenotype in a mouse model for breast cancer. ADAM12 is a substrate for beta1 integrins and may affect tumor and stromal cell behavior through its binding to...

  6. CD46 and beta1 integrin relocation in the sperm head during capacitation and acrosome reaction

    Czech Academy of Sciences Publication Activity Database

    Šebková, N.; Frolíková, M.; Dvořáková-Hortová, Kateřina

    Madison: Society for the Study of Reproduction, 2015. s. 212. [48th Annual Meeting of the Society for the Study of reproduction. 18.06.2015-22.06.2015, San Juan] R&D Projects: GA ČR(CZ) GA14-05547S Institutional support: RVO:86652036 Keywords : CD46 * beta1 integrin * Proximity Ligation Assay * acrosome reaction Subject RIV: CE - Biochemistry

  7. Expression and functional importance of collagen-binding integrins, alpha 1 beta 1 and alpha 2 beta 1, on virus-activated T cells

    DEFF Research Database (Denmark)

    Andreasen, Susanne Ø; Thomsen, Allan R; Koteliansky, Victor E; Novobrantseva, Tatiana I; Sprague, Andrew G; de Fougerolles, Antonin R; Christensen, Jan P

    2003-01-01

    Adhesive interactions are crucial to cell migration into inflammatory sites. Using murine lymphocytic choriomeningitis virus as an Ag model system, we have investigated expression and function of collagen-binding integrins, alpha(1)beta(1) and alpha(2)beta(1), on activated and memory T cells. Using...... this system and MHC tetramers to define Ag-specific T cells, we demonstrate that contrary to being VLAs, expression of alpha(1)beta(1) and alpha(2)beta(1) can be rapidly induced on acutely activated T cells, that expression of alpha(1)beta(1) remains elevated on memory T cells, and that expression of...... alpha(1)beta(1) parallels that of viral-specific effector CD8(+) T cells (defined by tetramer and IFN-gamma staining). In an adoptive transfer model, mAb-mediated blockade of these integrins on activated effector and memory T cells inhibited Ag-specific delayed-type hypersensitivity responses; similar...

  8. Cellular growth and survival are mediated by beta 1 integrins in normal human breast epithelium but not in breast carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Howlett, Anthony R; Bailey, Nina; Damsky, Caroline; Petersen, Ole W; Bissell, Mina J

    1994-11-28

    We previously established a rapid three-dimensional assay for discrimination of normal and malignant human breast epithelial cells using a laminin-rich reconstituted basement membrane. In this assay, normal epithelial cells differentiate into well-organized acinar structures whereas tumor cells fail to recapitulate this process and produce large, disordered colonies. The data suggest that breast acinar morphogenesis and differentiation is regulated by cell-extracellular matrix (ECM) interactions and that these interactions are altered in malignancy. Here, we investigated the role of ECM receptors (integrins) in these processes and report on the expression and function of potential laminin receptors in normal and tumorigenic breast epithelial cells. Immmunocytochemical analysis showed that normal and carcinoma cells in a three-dimensional substratum express profiles of integrins similar to normal and malignant breast tissues in situ. Normal cells express {alpha}1, {alpha}2, {alpha}3, {alpha}6, {beta}1 and {beta}4 integrin subunits, whereas breast carcinoma cells show variable losses, disordered expression, or down regulation of these subunits. Function-blocking experiments using inhibitory antiintegrin subunit antibodies showed a >5-fold inhibition of the formation of acinar structures by normal cells in the presence of either anti-{beta}1 or anti-{alpha}3 antibodies, whereas anti-{alpha}2 or -{alpha}6 had little or no effect. In experiments where collagen type I gels were used instead of basement membrane, acinar morphogenesis was blocked by anti-{beta}1 and -{alpha}2 antibodies but not by anti-{alpha}3. These data suggest a specificity of integrin utilization dependent on the ECM ligands encountered by the cell. The interruption of normal acinar morphogenesis by anti-integrin antibodies was associated with an inhibition of cell growth and induction of apoptosis. Function-blocking antibodies had no inhibitory effect on the rate of tumor cell growth, survival or

  9. Skin and hair follicle integrity is crucially dependent on beta 1 integrin expression on keratinocytes

    DEFF Research Database (Denmark)

    Brakebusch, C; Grose, R; Quondamatteo, F;

    2000-01-01

    developed severe hair loss due to a reduced proliferation of hair matrix cells and severe hair follicle abnormalities. Eventually, the malformed hair follicles were removed by infiltrating macrophages. The epidermis of the back skin became hyperthickened, the basal keratinocytes showed reduced expression of......, the integrity of the basement membrane surrounding the beta 1-deficient hair follicle was not affected. Finally, the dermis became fibrotic. These results demonstrate an important role of beta 1 integrins in hair follicle morphogenesis, in the processing of basement membrane components, in the...

  10. Lack of beta1 integrins in enteric neural crest cells leads to a Hirschsprung-like phenotype

    DEFF Research Database (Denmark)

    Breau, Marie A; Pietri, Thomas; Eder, Olivier;

    2006-01-01

    The enteric nervous system arises mainly from vagal and sacral neural crest cells that colonise the gut between 9.5 and 14 days of development in mice. Using the Cre-LoxP system, we removed beta1 integrins in the neural crest cells when they emerge from the neural tube. beta1-null enteric neural...... integrins are required for the villi innervation in the small intestine. Our findings highlight the crucial roles played by beta1 integrins at various steps of enteric nervous system development....

  11. beta 1 integrin inhibition dramatically enhances radiotherapy efficacy in human breast cancer xenografts

    Energy Technology Data Exchange (ETDEWEB)

    Park, Catherine C.; Park, Catherine C.; Zhang, Hui J.; Yao, Evelyn S.; Park, Chong J.; Bissell, Mina J.

    2008-06-02

    {beta}1 integrin signaling has been shown to mediate cellular resistance to apoptosis after exposure to ionizing radiation (IR). Other signaling molecules that increase resistance include Akt, which promotes cell survival downstream of {beta}1 integrin signaling. We showed previously that {beta}1 integrin inhibitory antibodies, AIIB2, enhance apoptosis and decrease growth in human breast cancer cells in 3 dimensional laminin-rich extracellular matrix (3D lrECM) cultures and in vivo. Here we asked whether AIIB2 could synergize with IR to modify Akt-mediated IR resistance. We used 3D lrECM cultures to test the optimal combination of AIIB2 with IR treatment of two breast cancer cell lines, MCF-7 and HMT3522-T4-2, as well as T4-2 myr-Akt breast cancer colonies or HMT3522-S-1, which form normal organotypic structures in 3D lrECM. Colonies were assayed for apoptosis and {beta}1 integrin/Akt signaling pathways were evaluated using western blot. In addition, mice bearing MCF-7 xenografts were used to validate the findings in 3D lrECM. We report that AIIB2 increased apoptosis optimally post-IR by down regulating Akt in breast cancer colonies in 3D lrECM. In vivo, addition of AIIB2 after IR significantly enhanced tumor growth inhibition and apoptosis compared to either treatment alone. Remarkably, the degree of tumor growth inhibition using AIIB2 plus 2 Gy radiation was similar to that of 8 Gy alone. We showed previously that AIIB2 had no discernible toxicity in mice; here, its addition allowed for a significant reduction in the IR dose that was necessary to achieve comparable growth inhibition and apoptosis in breast cancer xenografts in vivo.

  12. A preliminary optical and electron microscopic study of the beta(1) integrin distribution pattern of human osteosarcoma-derived cells.

    Science.gov (United States)

    Banai, Kiarash; Brady, Ken; McDonald, Fraser

    2004-07-01

    Immunogold labelling was used to study the organisation of the beta(1) integrins on osteosarcoma-derived osteoblasts (Saos-2 and MG-63). Monolayers of cells were prepared in multiwell culture plates on both uncovered and collagen-covered coverslips, and beta(1) integrins were primarily labelled using mouse monoclonal antibodies to beta(1) integrins. Indirect immunofluorescence labels using an anti-mouse fluorescein-conjugated goat antibody showed an even distribution of the beta(1) integrins on the cell membranes of all cell types used. A concentration of 2 microg/ml of the primary antibodies and a 1:100 dilution of the secondary antibodies were determined as the optimal concentration for labelling to use with indirect localisation of the primary antibodies gold conjugated to goat anti-mouse antibodies and viewed under an electron microscope. Ten nanometre gold particles were used for transmission electron microscopy (TEM) and 40 nm gold particles for scanning electron microscopy. TEM showed that beta(1) integrins were mainly clustered on the cell membrane processes with less labelling on the cell membranes themselves. The distribution of beta(1) integrins on osteosarcoma cells supports the concept that integrins may function by forming focal adhesions at the site of the cytoplasmic membrane processes. PMID:15241608

  13. Prolyl hydroxylation of collagen type I is required for efficient binding to integrin alpha 1 beta 1 and platelet glycoprotein VI but not to alpha 2 beta 1.

    Science.gov (United States)

    Perret, Stéephanie; Eble, Johannes A; Siljander, Pia R-M; Merle, Christine; Farndale, Richard W; Theisen, Manfred; Ruggiero, Florence

    2003-08-01

    Collagen is a potent adhesive substrate for cells, an event essentially mediated by the integrins alpha 1 beta 1 and alpha 2 beta 1. Collagen fibrils also bind to the integrin alpha 2 beta 1 and the platelet receptor glycoprotein VI to activate and aggregate platelets. The distinct triple helical recognition motifs for these receptors, GXOGER and (GPO)n, respectively, all contain hydroxyproline. Using unhydroxylated collagen I produced in transgenic plants, we investigated the role of hydroxyproline in the receptor-binding properties of collagen. We show that alpha 2 beta 1 but not alpha 1 beta 1 mediates cell adhesion to unhydroxylated collagen. Soluble recombinant alpha 1 beta 1 binding to unhydroxylated collagen is considerably reduced compared with bovine collagens, but binding can be restored by prolyl hydroxylation of recombinant collagen. We also show that platelets use alpha 2 beta 1 to adhere to the unhydroxylated recombinant molecules, but the adhesion is weaker than on fully hydroxylated collagen, and the unhydroxylated collagen fibrils fail to aggregate platelets. Prolyl hydroxylation is thus required for binding of collagen to platelet glycoprotein VI and to cells by alpha 1 beta 1. These observations give new insights into the molecular basis of collagen-receptor interactions and offer new selective applications for the recombinant unhydroxylated collagen I. PMID:12771137

  14. Loss of integrin alpha1beta1 ameliorates Kras-induced lung cancer.

    Science.gov (United States)

    Macias-Perez, Ines; Borza, Corina; Chen, Xiwu; Yan, Xuexian; Ibanez, Raquel; Mernaugh, Glenda; Matrisian, Lynn M; Zent, Roy; Pozzi, Ambra

    2008-08-01

    The collagen IV binding receptor integrin alpha1beta1 has been shown to regulate lung cancer due to its proangiogenic properties; however, it is unclear whether this receptor also plays a direct role in promoting primary lung tumors. To investigate this possibility, integrin alpha1-null mice were crossed with KrasLA2 mice that carry an oncogenic mutation of the Kras gene (G12D) and develop spontaneous primary tumors with features of non-small cell lung cancer. We provide evidence that KrasLA2/alpha1-null mice have a decreased incidence of primary lung tumors and longer survival compared with KrasLA2/alpha1 wild-type controls. Tumors from KrasLA2/alpha1-null mice were also smaller, less vascularized, and exhibited reduced cell proliferation and increased apoptosis, as determined by proliferating cell nuclear antigen and terminal deoxynucleotidyl-transferase-mediated dUTP nick-end staining, respectively. Moreover, tumors from the KrasLA2/alpha1-null mice showed diminished extracellular signal-regulated kinase (ERK) but enhanced p38 mitogen-activated protein kinase activation. Primary lung tumor epithelial cells isolated from KrasLA2/alpha1-null mice showed a significant decrease in anchorage-independent colony formation, collagen-mediated cell proliferation, ERK activation, and, most importantly, tumorigenicity when injected into nude mice compared with KrasLA2/alpha1 wild-type tumor cells. These results indicate that loss of the integrin alpha1 subunit decreases the incidence and growth of lung epithelial tumors initiated by oncogenic Kras, suggesting that both Kras and integrin alpha1beta1 cooperate to drive the growth of non-small cell lung cancer in vivo. PMID:18676835

  15. Blood platelets contain and secrete laminin-8 (alpha4beta1gamma1) and adhere to laminin-8 via alpha6beta1 integrin.

    Science.gov (United States)

    Geberhiwot, T; Ingerpuu, S; Pedraza, C; Neira, M; Lehto, U; Virtanen, I; Kortesmaa, J; Tryggvason, K; Engvall, E; Patarroyo, M

    1999-12-15

    Laminins, a family of heterotrimeric proteins with cell adhesive/signaling properties, are characteristic components of basement membranes of vasculature and tissues. In the present study, permeabilized platelets were found to react with a monoclonal antibody to laminin gamma1 chain by immunofluorescence. In Western blot analysis of platelet lysates, several monoclonal antibodies to gamma1 and beta1 laminin chains recognized 220- to 230-kDa polypeptides, under reducing conditions, and a structure with much slower electrophoretic mobility under nonreducing conditions. Immunoaffinity purification on a laminin beta1 antibody-Sepharose column yielded polypeptides of 230, 220, 200, and 180 kDa from platelet lysates. In the purified material, mAbs to beta1 and gamma1 reacted with the two larger polypeptides, while affinity-purified rabbit antibodies to laminin alpha4 chain recognized the smallest polypeptide. Identity of the polypeptides was confirmed by microsequencing. One million platelets contained on average 1 ng of laminin (approximately 700 molecules per cell), of which 20-35% was secreted within minutes after stimulation with either thrombin or phorbol ester. Platelets adhered to plastic surfaces coated with the purified platelet laminin, and this process was largely inhibited by antibodies to beta1 and alpha6 integrin chains. We conclude that platelets contain and, following activation, secrete laminin-8 (alpha4beta1gamma1) and that the cells adhere to the protein by using alpha6beta1 integrin. PMID:10585296

  16. Tumor cell adhesion to endothelial cells is increased by endotoxin via an upregulation of beta-1 integrin expression.

    LENUS (Irish Health Repository)

    Andrews, E J

    2012-02-03

    BACKGROUND: Recent studies have demonstrated that metastatic disease develops from tumor cells that adhere to endothelial cells and proliferate intravascularly. The beta-1 integrin family and its ligand laminin have been shown to be important in tumor-to-endothelial cell adhesion. Lipopolysaccharide (LPS) has been implicated in the increased metastatic tumor growth that is seen postoperatively. We postulated that LPS increases tumor cell expression of beta-1 integrins and that this leads to increased adhesion. METHODS: The human metastatic colon cancer cell line LS174T was labeled with an enhanced green fluorescent protein (eGFP) using retroviral transfection. Cell cultures were treated with LPS for 1, 2, and 4 h (n = 6 each) and were subsequently cocultured for 30 or 120 min with confluent human umbilical vein endothelial cells (HUVECs), to allow adherence. Adherent tumor cells were counted using fluorescence microscopy. These experiments were carried out in the presence or absence of a functional blocking beta-1 integrin monoclonal antibody (4B4). Expression of beta-1 integrin and laminin on tumor and HUVECs was assessed using flow cytometric analysis. Tumor cell NF-kappaB activation after incubation with LPS was measured. RESULTS: Tumor cell and HUVEC beta-1 integrin expression and HUVEC expression of laminin were significantly (P < 0.05) enhanced after incubation with LPS. Tumor cell adhesion to HUVECs was significantly increased. Addition of the beta-1 integrin blocking antibody reduced tumor cell adhesion to control levels. LPS increased tumor cell NF-kappaB activation. CONCLUSIONS: Exposure to LPS increases tumor cell adhesion to the endothelium through a beta-1 integrin-mediated pathway that is NF-kappaB dependent. This may provide a target for immunotherapy directed at reducing postoperative metastatic tumor growth.

  17. Interactions of the integrin subunit beta1A with protein kinase B/Akt, p130Cas and paxillin contribute to regulation of radiation survival

    DEFF Research Database (Denmark)

    Seidler, Julia; Durzok, Rita; Brakebusch, Cord; Cordes, Nils

    2005-01-01

    BACKGROUND AND PURPOSE: Cell adhesion-mediated radioresistance is a common phenomenon particularly relevant in tumor cells, which might hamper anticancer therapies. To analyze the role of adhesion-mediating beta1-integrins, stably transfected functional beta1A-integrin-expressing GD25beta1A and G...

  18. Beta 1 integrin predicts survival in breast cancer: a clinicopathological and immunohistochemical study

    Directory of Open Access Journals (Sweden)

    dos Santos Petra

    2012-08-01

    Full Text Available Abstract Background The main focus of several studies concerned with cancer progression and metastasis is to analyze the mechanisms that allow cancer cells to interact and quickly adapt with their environment. Integrins, a family of transmembrane glycoproteins, play a major role in invasive and metastatic processes. Integrins are involved in cell adhesion in both cell-extracellular matrix and cell-cell interactions, and particularly, β1 integrin is involved in proliferation and differentiation of cells in the development of epithelial tissues. This work aimed to investigate the putative role of β1 integrin expression on survival and metastasis in patients with breast invasive ductal carcinoma (IDC. In addition, we compared the expression of β1 integrin in patients with ductal carcinoma in situ (DCIS. Methods Through tissue microarray (TMA slides containing 225 samples of IDC and 67 samples of DCIS, β1 integrin expression was related with several immunohistochemical markers and clinicopathologic features of prognostic significance. Results β1 integrin was overexpressed in 32.8% of IDC. In IDC, β1 integrin was related with HER-2 (p = 0.019 and VEGF (p = 0.011 expression and it had a significant relationship with metastasis and death (p = 0.001 and p = 0.05, respectively. Kaplan-Meier survival analysis showed that the overexpression of this protein is very significant (p = 0.002 in specific survival (number of months between diagnosis and death caused by the disease. There were no correlation between IDC and DCIS (p = 0.559 regarding β1 integrin expression. Conclusions Considering that the expression of β1 integrin in breast cancer remains controversial, specially its relation with survival of patients, our findings provide further evidence that β1 integrin can be a marker of poor prognosis in breast cancer. Virtual slides The virtual slide(s for this article can be found here: http

  19. Differential expression of integrins and laminin-5 in normal oral epithelia

    DEFF Research Database (Denmark)

    Thorup, A K; Dabelsteen, Erik; Schou, S;

    1997-01-01

    /parabasal cells in keratinized epithelia, whereas there was increased suprabasal expression in nonkeratinized mucosa. These results indicate inhomogeneity in the basal cell population of oral squamous epithelia and differential expression of integrins, which may reflect differences in the underlying stroma......beta 1 and beta 4 integrins are receptors on epithelial cells mediating cell-extracellular matrix adhesion. Furthermore, alpha 2 beta 1 and alpha 3 beta 1 contribute to cell-cell adhesion. Laminin-5 in epithelial basement membranes (BMs) is a ligand for alpha 6 beta 4 and alpha 3 beta 1. Expression...... of different integrins and laminin-5 was studied in oral epithelium to characterize regional variations in these adhesion molecules. Monoclonal antibodies directed against alpha 2-alpha 6 beta 1/alpha 6 beta 4 and laminin-5 were examined in cryopreserved biopsies of normal mucosa by...

  20. Role of the beta1-integrin cytoplasmic tail in mediating invasin-promoted internalization of Yersinia

    DEFF Research Database (Denmark)

    Gustavsson, Anna; Armulik, Annika; Brakebusch, Cord;

    2002-01-01

    1B or different mutants of the beta1A subunit was used. Both beta1A and beta1B bound to invasin-expressing bacteria, but only beta1A was able to mediate internalization of the bacteria. The cytoplasmic region of beta1A, differing from beta1B, contains two NPXY motifs surrounding a double threonine...

  1. Increased expression of integrin alpha2 and abnormal response to TGF-beta1 in hereditary gingival fibromatosis.

    NARCIS (Netherlands)

    Zhou, J.; Meng, L.; Ye, X.Q.; Hoff, J.W. von den; Bian, Z.

    2009-01-01

    OBJECTIVE: To investigate the possible correlation between integrin alpha1, alpha2, and beta1 expression and excessive collagen synthesis in fibroblasts from 3 unrelated Chinese families with hereditary gingival fibromatosis (HGF). DESIGN: Gingival fibroblasts from three Chinese HGF patients and thr

  2. Integrin {beta}1-dependent invasive migration of irradiation-tolerant human lung adenocarcinoma cells in 3D collagen matrix

    Energy Technology Data Exchange (ETDEWEB)

    Ishihara, Seiichiro [Transdisciplinary Life Science Course, Faculty of Advanced Life Science, Hokkaido University, N10-W8, Kita-ku, Sapporo 060-0810 (Japan); Haga, Hisashi, E-mail: haga@sci.hokudai.ac.jp [Transdisciplinary Life Science Course, Faculty of Advanced Life Science, Hokkaido University, N10-W8, Kita-ku, Sapporo 060-0810 (Japan); Yasuda, Motoaki [Department of Oral Pathobiological Science, Graduate School of Dental Medicine, Hokkaido University, N13-W7, Kita-ku, Sapporo 060-8586 (Japan); Mizutani, Takeomi; Kawabata, Kazushige [Transdisciplinary Life Science Course, Faculty of Advanced Life Science, Hokkaido University, N10-W8, Kita-ku, Sapporo 060-0810 (Japan); Shirato, Hiroki [Department of Radiology, Hokkaido University Graduate School of Medicine, N15-W7, Kita-ku, Sapporo 060-8638 (Japan); Nishioka, Takeshi [Department of Biomedical Sciences and Engineering, Faculty of Health Sciences, Hokkaido University, N12-W5, Kita-ku, Sapporo 060-0812 (Japan)

    2010-06-04

    Radiotherapy is one of the effective therapies used for treating various malignant tumors. However, the emergence of tolerant cells after irradiation remains problematic due to their high metastatic ability, sometimes indicative of poor prognosis. In this study, we showed that subcloned human lung adenocarcinoma cells (A549P-3) that are irradiation-tolerant indicate high invasive activity in vitro, and exhibit an integrin {beta}1 activity-dependent migratory pattern. In collagen gel overlay assay, majority of the A549P-3 cells displayed round morphology and low migration activity, whereas a considerable number of A549P-3IR cells surviving irradiation displayed a spindle morphology and high migration rate. Blocking integrin {beta}1 activity reduced the migration rate of A549P-3IR cells and altered the cell morphology allowing them to assume a round shape. These results suggest that the A549P-3 cells surviving irradiation acquire a highly invasive integrin {beta}1-dependent phenotype, and integrin {beta}1 might be a potentially effective therapeutic target in combination with radiotherapy.

  3. Integrins (alpha7beta1) in muscle function and survival. Disrupted expression in merosin-deficient congenital muscular dystrophy

    DEFF Research Database (Denmark)

    Vachon, P H; Xu, H; Liu, L;

    1997-01-01

    Mutations in genes coding for dystrophin, for alpha, beta, gamma, and delta-sarcoglycans, or for the alpha2 chain of the basement membrane component merosin (laminin-2/4) cause various forms of muscular dystrophy. Analyses of integrins showed an abnormal expression and localization of alpha7beta1...

  4. Use of synthetic peptides to locate novel integrin alpha2beta1-binding motifs in human collagen III.

    Science.gov (United States)

    Raynal, Nicolas; Hamaia, Samir W; Siljander, Pia R-M; Maddox, Ben; Peachey, Anthony R; Fernandez, Rafael; Foley, Loraine J; Slatter, David A; Jarvis, Gavin E; Farndale, Richard W

    2006-02-17

    A set of 57 synthetic peptides encompassing the entire triplehelical domain of human collagen III was used to locate binding sites for the collagen-binding integrin alpha(2)beta(1). The capacity of the peptides to support Mg(2+)-dependent binding of several integrin preparations was examined. Wild-type integrins (recombinant alpha(2) I-domain, alpha(2)beta(1) purified from platelet membranes, and recombinant soluble alpha(2)beta(1) expressed as an alpha(2)-Fos/beta(1)-Jun heterodimer) bound well to only three peptides, two containing GXX'GER motifs (GROGER and GMOGER, where O is hydroxyproline) and one containing two adjacent GXX'GEN motifs (GLKGEN and GLOGEN). Two mutant alpha(2) I-domains were tested: the inactive T221A mutant, which recognized no peptides, and the constitutively active E318W mutant, which bound a larger subset of peptides. Adhesion of activated human platelets to GER-containing peptides was greater than that of resting platelets, and HT1080 cells bound well to more of the peptides compared with platelets. Binding of cells and recombinant proteins was abolished by anti-alpha(2) monoclonal antibody 6F1 and by chelation of Mg(2+). We describe two novel high affinity integrin-binding motifs in human collagen III (GROGER and GLOGEN) and a third motif (GLKGEN) that displays intermediate activity. Each motif was verified using shorter synthetic peptides. PMID:16326707

  5. Mechanotransduction molecules in the plant gravisensory response: amyloplast/statolith membranes contain a beta 1 integrin-like protein

    Science.gov (United States)

    Lynch, T. M.; Lintilhac, P. M.; Domozych, D.

    1998-01-01

    It has been hypothesized that the sedimentation of amyloplasts within root cap cells is the primary event in the plant gravisensory-signal transduction cascade. Statolith sedimentation, with its ability to generate weighty mechanical signals, is a legitimate means for organisms to discriminate the direction of the gravity vector. However, it has been demonstrated that starchless mutants with reduced statolith densities maintain some ability to sense gravity, calling into question the statolith sedimentation hypothesis. Here we report on the presence of a beta 1 integrin-like protein localized inside amyloplasts of tobacco NT-1 suspension culture, callus cells, and whole-root caps. Two different antibodies to the beta 1 integrin, one to the cytoplasmic domain and one to the extracellular domain, localize in the vicinity of the starch grains within amyloplasts of NT-1. Biochemical data reveals a 110-kDa protein immunoprecipitated from membrane fractions of NT-1 suspension culture indicating size homology to known beta 1 integrin in animals. This study provides the first direct evidence for the possibility of integrin-mediated signal transduction in the perception of gravity by higher plants. An integrin-mediated pathway, initiated by starch grain sedimentation within the amyloplast, may provide the signal amplification necessary to explain the gravitropic response in starch-depleted cultivars.

  6. The cysteine-rich domain of human ADAM 12 supports cell adhesion through syndecans and triggers signaling events that lead to beta1 integrin-dependent cell spreading

    DEFF Research Database (Denmark)

    Iba, K; Albrechtsen, R; Gilpin, B;

    2000-01-01

    spread on ADAM 12. However, spreading could be efficiently induced by the addition of either 1 mM Mn(2+) or the beta1 integrin-activating monoclonal antibody 12G10, suggesting that in these carcinoma cells, the ADAM 12-syndecan complex fails to modulate the function of beta1 integrin.......-dependent manner attach to ADAM 12 via members of the syndecan family. After binding to syndecans, mesenchymal cells spread and form focal adhesions and actin stress fibers. Integrin beta1 was responsible for cell spreading because function-blocking monoclonal antibodies completely inhibited cell spreading, and...... chondroblasts lacking beta1 integrin attached but did not spread. These data suggest that mesenchymal cells use syndecans as the initial receptor for the ADAM 12 cysteine-rich domain-mediated cell adhesion, and then the beta1 integrin to induce cell spreading. Interestingly, carcinoma cells attached but did not...

  7. Rhodocytin (aggretin) activates platelets lacking alpha(2)beta(1) integrin, glycoprotein VI, and the ligand-binding domain of glycoprotein Ibalpha

    DEFF Research Database (Denmark)

    Bergmeier, W; Bouvard, D; Eble, J A;

    2001-01-01

    collagen may activate platelets by a similar mechanism. In contrast to these findings, we provided evidence that rhodocytin does not bind to alpha(2)beta(1) integrin. Here we show that the Cre/loxP-mediated loss of beta(1) integrin on mouse platelets has no effect on rhodocytin-induced platelet activation......Although alpha(2)beta(1) integrin (glycoprotein Ia/IIa) has been established as a platelet collagen receptor, its role in collagen-induced platelet activation has been controversial. Recently, it has been demonstrated that rhodocytin (also termed aggretin), a snake venom toxin purified from the......, excluding an essential role of alpha(2)beta(1) integrin in this process. Furthermore, proteolytic cleavage of the 45-kDa N-terminal domain of glycoprotein (GP) Ibalpha either on normal or on beta(1)-null platelets had no significant effect on rhodocytin-induced platelet activation. Moreover, mouse platelets...

  8. Function of glycoprotein VI and integrin alpha2beta1 in the procoagulant response of single, collagen-adherent platelets.

    Science.gov (United States)

    Heemskerk, J W; Siljander, P; Vuist, W M; Breikers, G; Reutelingsperger, C P; Barnes, M J; Knight, C G; Lassila, R; Farndale, R W

    1999-05-01

    Various collagen-based materials were used to assess the structural requirements of collagen for inducing the procoagulant response of adhering platelets, as well as the collagen receptors involved. Cross-linked or monomeric collagen-related peptide (CRP), Gly-Cys-Hyp-(Gly-Pro-Hyp)10-Gly-Cys-Hyp-Gly was highly adhesive for platelets in a glycoprotein VI-(GpVI-)dependent manner. Adhesion was followed by a prolonged increase in cytosolic [Ca2+]i, formation of membrane blebs, exposure of phosphatidylserine (PS) and generation of prothrombinase-stimulating activity. Fibrils of type-I collagen were less adhesive but, once adhered, many of the platelets presented a full procoagulant response. Monomeric type-I collagen was unable to support adhesion, unless Mg(2+)-dependent integrin alpha2beta1 interactions were facilitated by omission of Ca2+ ions. With all surfaces, however, post-addition of CaCl2 to adhering platelets resulted in a potent Ca(2+)-influx signal, followed by PS exposure and bleb formation. The procoagulant response elicited by binding to CRP was inhibited by anti-GpVI Fab fragments, but not by impeding integrin alpha2beta1-mediated events. With fibrillar collagen, it was inhibited by blocking either the GpVI- or integrin alpha2beta1-mediated interactions. This suggests that the triple-helical Gly-Pro-Hyp repeat in CRP and analogous sequences in fibrillar collagen stimulate the procoagulant response of adherent platelets by acting as ligands for GpVI. Influx of Ca2+ is required for this response, and adhesion via integrin alpha2beta1 serves to potentiate the signaling effects of GpVI. PMID:10365754

  9. Antimigratory effect of TK1-2 is mediated in part by interfering with integrin alpha2beta1.

    Science.gov (United States)

    Kim, Hyun-Kyung; Oh, Dae-Shik; Lee, Sang-Bae; Ha, Jung-Min; Joe, Young Ae

    2008-07-01

    The recombinant two kringle domain of human tissue-type plasminogen activator (TK1-2) has been shown to inhibit endothelial cell proliferation, angiogenesis, and tumor cell growth despite of sharing a low amino acid sequence homology with angiostatin. Here, we explored a possible inhibitory mechanism of action of TK1-2 by focusing on antimigratory effect. TK1-2 effectively inhibited endothelial cell migration induced by basic fibroblast growth factor or vascular endothelial growth factor in a dose-dependent manner and tube formation on Matrigel. It blocked basic fibroblast growth factor-induced or vascular endothelial growth factor-induced phosphorylation of extracellular signal-regulated kinase 1/2 and formation of actin stress fibers and focal adhesions. Interestingly, TK1-2 alone induced the weak phosphorylation of focal adhesion kinase, whereas it inhibited focal adhesion kinase phosphorylation induced by growth factors. When immobilized, TK1-2 promoted adhesion and spreading of endothelial cells compared with bovine serum albumin. However, treatment with anti-alpha(2)beta(1) blocking antibody markedly diminished endothelial cell adhesion to immobilized TK1-2 compared with anti-alpha(v)beta(3) or anti-alpha(5)beta(1) antibody. Pretreatment of soluble TK1-2 also altered the binding level of anti-alpha(2)beta(1) antibody to endothelial cells in fluorescence-activated cell sorting analysis. Indeed, a blocking antibody against integrin alpha(2)beta(1) or knocking down of integrin alpha(2) expression prevented the inhibitory effect of TK1-2 in cell migration. Therefore, these results suggest that TK1-2 inhibits endothelial cell migration through inhibition of signaling and cytoskeleton rearrangement in part by interfering with integrin alpha(2)beta(1). PMID:18645023

  10. The Effect of dcEFs on migration behavior of A549 cells and Integrin beta1 expression

    Directory of Open Access Journals (Sweden)

    Yunjie WANG

    2008-04-01

    Full Text Available Background and objective The effect of direct-current electric fields (dcEFs on cells attracted extensive attention. Moreover the metastasis and its potential are considered to be related to dcEFs. The aim is to study the effect of dcEFs on migration behavior of A549 cells, Integrin ?1 and its signal pathways. Methods According to exposure to 5 V/cm dcEFs or not and the time of exposure, the A549 cells were divided into 4 groups. Images were taken per 5 min within 2 h to recode the migration of the cells. The data of results were analyzed statistically. Results Most of A549cells exposed to the dcEFs aligned and elongated perpendicularly to the electric field lines and migrated to the cathode continually during 2 h. On the contrary, cells unexposed to dcEFs showed slightly random movements. Immunofluorescence showed that Integrin ?1 on plasma membrane polarized to the cathode of the dcEFs. Western blot showed that Integrin beta1 downstream signal pathways p-FAK and p-ERK were overexpressed in the dcEFs. Conclusion A549 cells have a galvanotatic feature of cathodal directed migration while exposed to the dcEFs. The polarization of Integrin beta1 and the promotion of its downstream signal pathways may play an important roles in the galvanotaxis of A549 cells.

  11. Inhibiting Vimentin or beta 1-integrin Reverts Prostate Tumor Cells in IrECM and Reduces Tumor Growth

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Xueping; Fournier, Marcia V.; Ware, Joy L.; Bissell, Mina J.; Zehner, Zendra E.

    2009-07-27

    Prostate epithelial cells grown embedded in laminin-rich extracellular matrix (lrECM) undergo morphological changes that closely resemble their architecture in vivo. In this study, growth characteristics of three human prostate epithelial sublines derived from the same cellular lineage, but displaying different tumorigenic and metastatic properties in vivo, were assessed in three-dimensional (3D) lrECM gels. M12, a highly tumorigenic and metastatic subline, was derived from the parental prostate epithelial P69 cell line by selection in nude mice and found to contain a deletion of 19p-q13.1. The stable reintroduction of an intact human chromosome 19 into M12 resulted in a poorly tumorigenic subline, designated F6. When embedded in lrECM gels, the nontumorigenic P69 line produced acini with clearly defined lumena. Immunostaining with antibodies to {beta}-catenin, E-cadherin or {alpha}6-, {beta}4- and {beta}1-integrins showed polarization typical of glandular epithelium. In contrast, the metastatic M12 subline produced highly disorganized cells with no evidence of polarization. The F6 subline reverted to acini-like structures exhibiting basal polarity marked with integrins. Reducing either vimentin levels via siRNA interference or {beta}1-integrin expression by the addition of the blocking antibody, AIIB2, reorganized the M12 subline into forming polarized acini. The loss of vimentin significantly reduced M12-Vim tumor growth when assessed by subcutaneous injection in athymic mice. Thus, tumorigenicity in vivo correlated with disorganized growth in 3D lrECM gels. These studies suggest that the levels of vimentin and {beta}1-integrin play a key role in the homeostasis of the normal acini in prostate and that their dysregulation may lead to tumorigenesis.

  12. Plasmin-induced migration requires signaling through protease-activated receptor 1 and integrin alpha(9)beta(1).

    Science.gov (United States)

    Majumdar, Mousumi; Tarui, Takehiko; Shi, Biao; Akakura, Nobuaki; Ruf, Wolfram; Takada, Yoshikazu

    2004-09-01

    Plasmin is a major extracellular protease that elicits intracellular signals to mediate platelet aggregation, chemotaxis of peripheral blood monocytes, and release of arachidonate and leukotriene from several cell types in a G protein-dependent manner. Angiostatin, a fragment of plasmin(ogen), is a ligand and an antagonist for integrin alpha(9)beta(1). Here we report that plasmin specifically interacts with alpha(9)beta(1) and that plasmin induces of cells expressing migration recombinant alpha(9)beta(1) (alpha(9)-Chinese hamster ovary (CHO) cells). Migration was dependent on an interaction of the kringle domains of plasmin with alpha(9)beta(1) as well as the catalytic activity of plasmin. Angiostatin, representing the kringle domains of plasmin, alone did not induce the migration of alpha(9)-CHO cells, but simultaneous activation of the G protein-coupled protease-activated receptor (PAR)-1 with an agonist peptide induced the migration on angiostatin, whereas PAR-2 or PAR-4 agonist peptides were without effect. Furthermore, a small chemical inhibitor of PAR-1 (RWJ 58259) and a palmitoylated PAR-1-blocking peptide inhibited plasmin-induced migration of alpha(9)-CHO cells. These results suggest that plasmin induces migration by kringle-mediated binding to alpha(9)beta(1) and simultaneous proteolytic activation of PAR-1. PMID:15247268

  13. Cellular partitioning of beta-1 integrins and their phosphorylated forms is altered after transformation by Rous sarcoma virus or treatment with cytochalasin D.

    OpenAIRE

    Haimovich, B; Aneskievich, B J; Boettiger, D

    1991-01-01

    A sequential extraction procedure of 3-[(3-cholamidopropyl)-dimethylammonio]-1-propane sulfonate (CHAPS) buffer followed by RIPA or Laemmli sample buffer was developed to define two distinct subpopulations of beta-1 integrins in primary chicken embryo fibroblasts. Extraction of cells in culture revealed an association of adhesion plaque-localized integrin with the CHAPS-insoluble fraction. Phosphorylated integrins were found in both fractions, but the specific phosphorylation was 12-fold high...

  14. Integrin alpha1beta1 regulates epidermal growth factor receptor activation by controlling peroxisome proliferator-activated receptor gamma-dependent caveolin-1 expression.

    Science.gov (United States)

    Chen, Xiwu; Whiting, Carrie; Borza, Corina; Hu, Wen; Mont, Stacey; Bulus, Nada; Zhang, Ming-Zhi; Harris, Raymond C; Zent, Roy; Pozzi, Ambra

    2010-06-01

    Integrin alpha1beta1 negatively regulates the generation of profibrotic reactive oxygen species (ROS) by inhibiting epidermal growth factor receptor (EGFR) activation; however, the mechanism by which it does this is unknown. In this study, we show that caveolin-1 (Cav-1), a scaffolding protein that binds integrins and controls growth factor receptor signaling, participates in integrin alpha1beta1-mediated EGFR activation. Integrin alpha1-null mesangial cells (MCs) have reduced Cav-1 levels, and reexpression of the integrin alpha1 subunit increases Cav-1 levels, decreases EGFR activation, and reduces ROS production. Downregulation of Cav-1 in wild-type MCs increases EGFR phosphorylation and ROS synthesis, while overexpression of Cav-1 in the integrin alpha1-null MCs decreases EGFR-mediated ROS production. We further show that integrin alpha1-null MCs have increased levels of activated extracellular signal-regulated kinase (ERK), which leads to reduced activation of peroxisome proliferator-activated receptor gamma (PPARgamma), a transcription factor that positively regulates Cav-1 expression. Moreover, activation of PPARgamma or inhibition of ERK increases Cav-1 levels in the integrin alpha1-null MCs. Finally, we show that glomeruli of integrin alpha1-null mice have reduced levels of Cav-1 and activated PPARgamma but increased levels of phosphorylated EGFR both at baseline and following injury. Thus, integrin alpha1beta1 negatively regulates EGFR activation by positively controlling Cav-1 levels, and the ERK/PPARgamma axis plays a key role in regulating integrin alpha1beta1-dependent Cav-1 expression and consequent EGFR-mediated ROS production. PMID:20368353

  15. Endotoxin/lipopolysaccharide activates NF-kappa B and enhances tumor cell adhesion and invasion through a beta 1 integrin-dependent mechanism.

    LENUS (Irish Health Repository)

    Wang, Jiang Huai

    2012-02-03

    Beta(1) integrins play a crucial role in supporting tumor cell attachment to and invasion into the extracellular matrix. Endotoxin\\/LPS introduced by surgery has been shown to enhance tumor metastasis in a murine model. Here we show the direct effect of LPS on tumor cell adhesion and invasion in extracellular matrix proteins through a beta(1) integrin-dependent pathway. The human colorectal tumor cell lines SW480 and SW620 constitutively expressed high levels of the beta(1) subunit, whereas various low levels of alpha(1), alpha(2), alpha(4), and alpha(6) expression were detected. SW480 and SW620 did not express membrane-bound CD14; however, LPS in the presence of soluble CD14 (sCD14) significantly up-regulated beta(1) integrin expression; enhanced tumor cell attachment to fibronectin, collagen I, and laminin; and strongly promoted tumor cell invasion through the Matrigel. Anti-beta(1) blocking mAbs (4B4 and 6S6) abrogated LPS- plus sCD14-induced tumor cell adhesion and invasion. Furthermore, LPS, when combined with sCD14, resulted in NF-kappaB activation in both SW480 and SW620 cells. Inhibition of the NF-kappaB pathway significantly attenuated LPS-induced up-regulation of beta(1) integrin expression and prevented tumor cell adhesion and invasion. These results provide direct evidence that although SW480 and SW620 cells do not express membrane-bound CD14, LPS in the presence of sCD14 can activate NF-kappaB, up-regulate beta(1) integrin expression, and subsequently promote tumor cell adhesion and invasion. Moreover, LPS-induced tumor cell attachment to and invasion through extracellular matrix proteins is beta(1) subunit-dependent.

  16. Induction of cell scattering by expression of beta1 integrins in beta1-deficient epithelial cells requires activation of members of the rho family of GTPases and downregulation of cadherin and catenin function

    DEFF Research Database (Denmark)

    Gimond, C; van Der Flier, A; van Delft, S;

    1999-01-01

    was required for a complete morphological transition towards the spindle-shaped fibroblast-like phenotype. The expression of an interleukin-2 receptor (IL2R)-beta1A chimera and its incorporation into focal adhesions also induced the disruption of cadherin-based adhesions and the reorganization of ECM......Adhesion receptors, which connect cells to each other and to the surrounding extracellular matrix (ECM), play a crucial role in the control of tissue structure and of morphogenesis. In this work, we have studied how intercellular adhesion molecules and beta1 integrins influence each other using two......-catenin protein levels accompanied by their redistribution from the cytoskeleton-associated fraction to the detergent-soluble fraction. Regulation of alpha-catenin protein levels by beta1 integrins is likely to play a role in the morphological transition, since overexpression of alpha-catenin in GE11 cells before...

  17. Highly Potent, Water Soluble Benzimidazole Antagonist for Activated (alpha)4(beta)1 Integrin

    Energy Technology Data Exchange (ETDEWEB)

    Carpenter, R D; Andrei, M; Lau, E Y; Lightstone, F C; Liu, R; Lam, K S; Kurth, M J

    2007-08-29

    The cell surface receptor {alpha}{sub 4}{beta}{sub 1} integrin, activated constitutively in lymphoma, can be targeted with the bisaryl urea peptidomimetic antagonist 1 (LLP2A). However, concerns on its preliminary pharmacokinetic (PK) profile provided an impetus to change the pharmacophore from a bisaryl urea to a 2-arylaminobenzimidazole moiety resulting in improved solubility while maintaining picomolar potency [5 (KLCA4); IC{sub 50} = 305 pM]. With exceptional solubility, this finding has potential for improving PK to help diagnose and treat lymphomas.

  18. Modeled Microgravity Disrupts Collagen I/Integrin Signaling During Osteoblastic Differentiation of Human Mesenchymal Stem Cells

    Science.gov (United States)

    Meyers, Valerie E.; Zayzafoon, Majd; Gonda, Steven R.; Gathings, William E.; McDonald, Jay M.

    2004-01-01

    Spaceflight leads to reduced bone mineral density in weight bearing bones that is primarily attributed to a reduction in bone formation. We have previously demonstrated severely reduced osteoblastogenesis of human mesenchymal stem cells (hMSC) following seven days culture in modeled microgravity. One potential mechanism for reduced osteoblastic differentiation is disruption of type I collagen-integrin interactions and reduced integrin signaling. Integrins are heterodimeric transmembrane receptors that bind extracellular matrix proteins and produce signals essential for proper cellular function, survival, and differentiation. Therefore, we investigated the effects of modeled microgravity on integrin expression and function in hMSC. We demonstrate that seven days of culture in modeled microgravity leads to reduced expression of the extracellular matrix protein, type I collagen (Col I). Conversely, modeled microgravity consistently increases Col I-specific alpha2 and beta1 integrin protein expression. Despite this increase in integrin sub-unit expression, autophosphorylation of adhesion-dependent kinases, focal adhesion kinase (FAK) and proline-rich tyrosine kinase 2 (PYK2), is significantly reduced. Activation of Akt is unaffected by the reduction in FAK activation. However, reduced downstream signaling via the Ras-MAPK pathway is evidenced by a reduction in Ras and ERK activation. Taken together, our findings indicate that modeled microgravity decreases integrin/MAPK signaling, which likely contributes to the observed reduction in osteoblastogenesis.

  19. Integrin {alpha}{beta}1, {alpha}{sub v}{beta}, {alpha}{sub 6}{beta} effectors p130Cas, Src and talin regulate carcinoma invasion and chemoresistance

    Energy Technology Data Exchange (ETDEWEB)

    Sansing, Hope A. [Department of Oral and Craniofacial Biology, Louisiana State University Health Sciences Center-New Orleans, School of Dentistry, New Orleans, LA (United States); Sarkeshik, Ali; Yates, John R. [Department of Chemical Physiology, Scripps Research Institute, La Jolla, CA (United States); Patel, Vyomesh; Gutkind, J. Silvio [Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD (United States); Yamada, Kenneth M. [Laboratory of Cell and Developmental Biology, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD (United States); Berrier, Allison L., E-mail: allison.berrier@gmail.com [Department of Oral and Craniofacial Biology, Louisiana State University Health Sciences Center-New Orleans, School of Dentistry, New Orleans, LA (United States)

    2011-03-11

    Research highlights: {yields} Proteomics of clustered integrin {alpha}{beta}1, {alpha}{sub v}{beta}, {alpha}{sub 6}{beta} receptors in oral carcinoma. {yields} p130Cas, Dek, Src and talin regulate oral carcinoma invasion. {yields} p130Cas, talin, Src and zyxin regulate oral carcinoma resistance to cisplatin. -- Abstract: Ligand engagement by integrins induces receptor clustering and formation of complexes at the integrin cytoplasmic face that controls cell signaling and cytoskeletal dynamics critical for adhesion-dependent processes. This study searches for a subset of integrin effectors that coordinates both tumor cell invasion and resistance to the chemotherapeutic drug cisplatin in oral carcinomas. Candidate integrin effectors were identified in a proteomics screen of proteins recruited to clustered integrin {alpha}{beta}1, {alpha}{sub v}{beta} or {alpha}{sub 6}{beta} receptors in oral carcinomas. Proteins with diverse functions including microtubule and actin binding proteins, and factors involved in trafficking, transcription and translation were identified in oral carcinoma integrin complexes. Knockdown of effectors in the oral carcinoma HN12 cells revealed that p130Cas, Dek, Src and talin were required for invasion through Matrigel. Disruption of talin or p130Cas by RNA interference increased resistance to cisplatin, whereas targeting Dek, Src or zyxin reduced HN12 resistance to cisplatin. Analysis of the spreading of HN12 cells on collagen I and laminin I revealed that a decrease in p130Cas or talin expression inhibited spreading on both matrices. Interestingly, a reduction in zyxin expression enhanced spreading on laminin I and inhibited spreading on collagen I. Reduction of Dek, Src, talin or zyxin expression reduced HN12 proliferation by 30%. Proliferation was not affected by a reduction in p130Cas expression. We conclude that p130Cas, Src and talin function in both oral carcinoma invasion and resistance to cisplatin.

  20. Complementary roles of glycoprotein VI and alpha2beta1 integrin in collagen-induced thrombus formation in flowing whole blood ex vivo

    DEFF Research Database (Denmark)

    Kuijpers, Marijke J E; Schulte, Valerie; Bergmeier, Wolfgang; Lindhout, Theo; Brakebusch, Cord; Offermanns, Stefan; Fässler, Reinhard; Heemskerk, Johan W M; Nieswandt, Bernhard

    2003-01-01

    function in ex vivo thrombus formation during perfusion of whole blood over collagen. With mice deficient in GPVI or blocking antibodies, we found that GPVI was indispensable for collagen-dependent Ca2+ mobilization, exposure of PS, and aggregation of platelets. Deficiency of integrin beta1 reduces the......Platelets interact vigorously with subendothelial collagens that are exposed by injury or pathological damage of a vessel wall. The collagen-bound platelets trap other platelets to form aggregates, and they expose phosphatidylserine (PS) required for coagulation. Both processes are implicated in...... the formation of vaso-occlusive thrombi. We previously demonstrated that the immunoglobulin receptor glycoprotein VI (GPVI), but not integrin alpha2beta1, is essential in priming platelet-collagen interaction and subsequent aggregation. Here, we report that these receptors have yet a complementary...

  1. Metastatic dissemination of human ovarian epithelial carcinoma is promoted by alpha2beta1-integrin-mediated interaction with type I collagen.

    Science.gov (United States)

    Fishman, D A; Kearns, A; Chilukuri, K; Bafetti, L M; O'Toole, E A; Georgacopoulos, J; Ravosa, M J; Stack, M S

    1998-01-01

    Metastatic dissemination of epithelial ovarian carcinoma is thought to be mediated via tumor cell exfoliation into the peritoneal cavity, followed by adhesion to and invasion through the mesothelium which overlies the contents of the peritoneal cavity. In this study, we have utilized short-term primary cultures to analyze the effect of specific extracellular matrix proteins on properties of human ovarian epithelial carcinoma cells which contribute to the invasive phenotype. Analysis of cell:matrix adhesive profiles indicated that ovarian carcinoma cells adhere preferentially to type I collagen. Immunoprecipitation analyses demonstrated the presence of the collagen-binding alpha2beta1 integrin in biotin-labeled ovarian carcinoma cell membranes, and cellular adhesion was inhibited by blocking antibodies directed against the alpha2 and beta1 integrin subunits. The alpha2beta1-binding peptide Asp-Gly-Glu-Ala (DGEA) was also moderately effective at blocking adhesion to collagen relative to the control peptide Ala-Gly-Glu-Ala (AGEA). Analysis of cell motility on protein-coated colloidal gold coverslips demonstrated that ovarian carcinoma cells migrate preferentially on type I collagen coated surfaces. Type I collagen promoted migration in a concentration-dependent, saturable manner, with maximal migration observed at a collagen-coating concentration of 50 microg/ml. Migration on collagen was inhibited by antibodies directed against the alpha2 and beta1 integrin subunits and by DGEA peptide, providing evidence for the role of the alpha2beta1 integrin in ovarian carcinoma cell motility. Culturing ovarian carcinoma cells on type I collagen gels led to a significant increase in conversion of the matrix metalloproteinase 2 zymogen to the 66-kD form, suggesting that adhesion to collagen also influences matrix-degrading proteinases. These data suggest that alpha2beta1-integrin-mediated interaction of ovarian carcinoma cells with type I collagen, a protein prevalent both in the

  2. Treatment of marrow stroma with interferon-alpha restores normal beta 1 integrin-dependent adhesion of chronic myelogenous leukemia hematopoietic progenitors. Role of MIP-1 alpha.

    OpenAIRE

    R Bhatia; McGlave, P B; Verfaillie, C M

    1995-01-01

    The mechanisms by which interferon-alpha (IFN-alpha) restores normal hematopoiesis in chronic myelogenous leukemia (CML) are not well understood. We have recently demonstrated that IFN-alpha acts directly on CML hematopoietic progenitors to restore their adhesion to marrow stroma by modulating beta 1 integrin receptor function. In the present study we examined the effect of IFN-alpha treatment of marrow stroma on subsequent adhesion of CML progenitors. Stromal layers were preincubated with IF...

  3. Integrin {alpha}1{beta}1 promotes caveolin-1 dephosphorylation by activating T cell protein-tyrosine phosphatase.

    Science.gov (United States)

    Borza, Corina M; Chen, Xiwu; Mathew, Sijo; Mont, Stacey; Sanders, Charles R; Zent, Roy; Pozzi, Ambra

    2010-12-17

    Integrin α1β1 is a collagen receptor that down-regulates collagen and reactive oxygen species (ROS) production, and mice lacking this receptor show increased ROS levels and exacerbated glomerular sclerosis following injury. Caveolin-1 (Cav-1) is a multifunctional protein that is tyrosine-phosphorylated in response to injury and has been implicated in ROS-mediated injury. Cav-1 interacts with integrins, and integrin α1β1 binds/activates T cell protein-tyrosine phosphatase (TCPTP), which is homologous to the tyrosine phosphatase PTP1B known to dephosphorylate Cav-1. In this study, we analyzed whether phosphorylated Cav-1 (pCav-1) is a substrate of TCPTP and if integrin α1β1 is essential for promoting TCPTP-mediated Cav-1 dephosphorylation. We found that Cav-1 phosphorylation is significantly higher in cells lacking integrin α1β1 at base line and following oxidative stress. Overexpression of TCPTP leads to reduced pCav-1 levels only in cells expressing integrin α1β1. Using solid phase binding assays, we demonstrated that 1) purified Cav-1 directly interacts with TCPTP and the integrin α1 subunit, 2) pCav-1 is a substrate of TCPTP, and 3) TCPTP-mediated Cav-1 dephosphorylation is highly increased by the addition of purified integrin α1β1 or an integrin α1 cytoplasmic peptide to which TCPTP has been shown to bind. Thus, our results demonstrate that pCav-1 is a new substrate of TCPTP and that integrin α1β1 acts as a negative regulator of Cav-1 phosphorylation by activating TCPTP. This could explain the protective function of integrin α1β1 in oxidative stress-mediated damage and why integrin α1-null mice are more susceptible to fibrosis following injury. PMID:20940300

  4. The alpha3 laminin subunit, alpha6beta4 and alpha3beta1 integrin coordinately regulate wound healing in cultured epithelial cells and in the skin

    DEFF Research Database (Denmark)

    Goldfinger, L E; Hopkinson, S B; deHart, G W;

    1999-01-01

    function-inhibiting antibodies, we provide evidence that LN5 and its two integrin receptors (alpha6beta4 and alpha3beta1) appear necessary for wound healing to occur in MCF-10A cell culture wounds. We propose a model for healing of wounded epithelial tissues based on these results....... epithelial cells. We have prepared a monoclonal antibody (12C4) whose epitope is located toward the carboxy terminus of the globular domain of the alpha3 laminin subunit. This epitope is lost from the alpha3 subunit as a consequence of proteolytic processing. Antibody 12C4 stains throughout the matrix of...... cover the wound site. A similar phenomenon is observed in human skin wounds, since we also detect expression of the unprocessed alpha3 laminin subunit at the leading tip of the sheet of epidermal cells that epithelializes skin wounds in vivo. In addition, using alpha3 laminin subunit and integrin...

  5. Molecular cloning and phylogenetic analysis of integrins alpha v beta 1 and alpha v beta 6 of one-humped camel (Camelus dromedarius)

    DEFF Research Database (Denmark)

    Du, Junzheng; Larska, Magdalena Larska; Chang, Huiyun;

    2010-01-01

    Bactrian camels can relatively easily be infected with FMDV, but dromedary camels remain resistant even to high doses of the virus. To understand the different susceptibility between the two camel species from the standpoint of viral receptors, this work reports the sequences of the dromedary camel...... into the Artiodactyla group, together with those of Bactrian camel, pig, sheep, and cattle that are susceptible to FMDV infection. Compared with the Bactrian camel integrins, 4, 10, and 8 amino acid changes were found in the dromedary camel alpha v, beta 1, and beta 6 subunits, respectively. This study...... will be of importance in understanding the differences of integrins as FMDV receptors among dromedary camel and other species. Crown...

  6. Dystrophin Dp71f associates with the beta1-integrin adhesion complex to modulate PC12 cell adhesion

    Science.gov (United States)

    Cerna, Joel; Cerecedo, Doris; Ortega, Arturo; García-Sierra, Francisco; Centeno, Federico; Garrido, Efrain; Mornet, Dominique; Cisneros, Bulmaro

    2006-01-01

    Dystrophin Dp71 is the main product of the Duchenne muscular dystrophy gene in the brain; however, its function is unknown. To study the role of Dp71 in neuronal cells, we previously generated by antisense treatment PC12 neuronal cell clones with decreased Dp71 expression (antisense-Dp71 cells). PC12 cells express two different splicing isoforms of Dp71, a cytoplasmic variant called Dp71f and a nuclear isoform called Dp71d. We previously reported that antisense-Dp71 cells display deficient adhesion to substrate and reduced immunostaining of β1-integrin in the cell area contacting the substrate. In this study, we isolated additional antisenseDp71 clones to analyze in detail the potential involvement of Dp71f isoform with the β1-integrin adhesion system of PC12 cells. Immunofluorescence analyses as well as immunoprecipitation assays demonstrated that the PC12 cell β1-integrin adhesion complex is composed of β1-integrin, talin, paxillin, α-actinin, FAK and actin. In addition, our results showed that Dp71f associates with most of the β1-integrin complex components (β1-integrin, FAK, α-actinin, talin and actin). In the antisense-Dp71 cells, the deficiency of Dp71 provokes a significant reduction of the β1-integrin adhesion complex and, consequently, the deficient adhesion of these cells to laminin. In vitro binding experiments confirmed the interaction of Dp71f with FAK and β1-integrin. Our data indicate that Dp71f is a structural component of the β1-integrin adhesion complex of PC12 cells that modulates PC12 cell adhesion by conferring proper complex assembly and/or maintenance. PMID:16935300

  7. Production of IL1-beta by ovarian cancer cells induces mesothelial cell beta1-integrin expression facilitating peritoneal dissemination

    Directory of Open Access Journals (Sweden)

    Watanabe Takafumi

    2012-02-01

    Full Text Available Abstract Background A crucial step in the metastatic spread of ovarian cancer (OC is the adhesion and implantation of tumor cells to the peritoneal mesothelium. In order to study this step in the cascade, we derived a pro-metastatic human ovarian carcinoma cell line (MFOC3 from the non-metastatic FOC3 line. Methods Molecular profiling of the isogeneic lines identified differentially expressed genes, and investigation for a role in dissemination for specific factors was achieved by development of a co-culture adhesion assay utilizing monolayers of human mesothelial cells. Results After murine intraperitoneal inoculation, the FOC3 cell line formed no metastases, but the MFOC3 subline formed metastases in > 80% of SCID mice. MFOC3 cells also adhered 2-3 times more avidly to mesothelial monolayers. This adhesion was inhibited by neutralizing antibodies to IL-1β and enhanced by recombinant IL-1β (p in vitro and significantly reduced metastases in vivo. Immunohistochemical analysis of a cohort of 96 ovarian cancer cases showed that negative IL-1β expression was significantly associated with an improved overall survival rate. Conclusions These results suggest that a IL-1β/β1-integrin axis plays a role in ovarian tumor cell adhesion to mesothelia, a crucial step in ovarian cancer dissemination.

  8. Distinct roles of beta1 metal ion-dependent adhesion site (MIDAS), adjacent to MIDAS (ADMIDAS), and ligand-associated metal-binding site (LIMBS) cation-binding sites in ligand recognition by integrin alpha2beta1.

    Science.gov (United States)

    Valdramidou, Dimitra; Humphries, Martin J; Mould, A Paul

    2008-11-21

    Integrin-ligand interactions are regulated in a complex manner by divalent cations, and previous studies have identified ligand-competent, stimulatory, and inhibitory cation-binding sites. In collagen-binding integrins, such as alpha2beta1, ligand recognition takes place exclusively at the alpha subunit I domain. However, activation of the alphaI domain depends on its interaction with a structurally similar domain in the beta subunit known as the I-like or betaI domain. The top face of the betaI domain contains three cation-binding sites: the metal-ion dependent adhesion site (MIDAS), the ADMIDAS (adjacent to MIDAS), and LIMBS (ligand-associated metal-binding site). The role of these sites in controlling ligand binding to the alphaI domain has yet to be elucidated. Mutation of the MIDAS or LIMBS completely blocked collagen binding to alpha2beta1; in contrast mutation of the ADMIDAS reduced ligand recognition but this effect could be overcome by the activating monoclonal antibody TS2/16. Hence, the MIDAS and LIMBS appear to be essential for the interaction between alphaI and betaI, whereas occupancy of the ADMIDAS has an allosteric effect on the conformation of betaI. An activating mutation in the alpha2 I domain partially restored ligand binding to the MIDAS and LIMBS mutants. Analysis of the effects of Ca(2+), Mg(2+), and Mn(2+) on ligand binding to these mutants showed that the MIDAS is a ligand-competent site through which Mn(2+) stimulates ligand binding, whereas the LIMBS is a stimulatory Ca(2+)-binding site, occupancy of which increases the affinity of Mg(2+) for the MIDAS. PMID:18820259

  9. The signaling pathway of Campylobacter jejuni-induced Cdc42 activation: Role of fibronectin, integrin beta1, tyrosine kinases and guanine exchange factor Vav2

    LENUS (Irish Health Repository)

    Krause-Gruszczynska, Malgorzata

    2011-12-28

    Abstract Background Host cell invasion by the foodborne pathogen Campylobacter jejuni is considered as one of the primary reasons of gut tissue damage, however, mechanisms and key factors involved in this process are widely unclear. It was reported that small Rho GTPases, including Cdc42, are activated and play a role during invasion, but the involved signaling cascades remained unknown. Here we utilised knockout cell lines derived from fibronectin-\\/-, integrin-beta1-\\/-, focal adhesion kinase (FAK)-\\/- and Src\\/Yes\\/Fyn-\\/- deficient mice, and wild-type control cells, to investigate C. jejuni-induced mechanisms leading to Cdc42 activation and bacterial uptake. Results Using high-resolution scanning electron microscopy, GTPase pulldowns, G-Lisa and gentamicin protection assays we found that each studied host factor is necessary for induction of Cdc42-GTP and efficient invasion. Interestingly, filopodia formation and associated membrane dynamics linked to invasion were only seen during infection of wild-type but not in knockout cells. Infection of cells stably expressing integrin-beta1 variants with well-known defects in fibronectin fibril formation or FAK signaling also exhibited severe deficiencies in Cdc42 activation and bacterial invasion. We further demonstrated that infection of wild-type cells induces increasing amounts of phosphorylated FAK and growth factor receptors (EGFR and PDGFR) during the course of infection, correlating with accumulating Cdc42-GTP levels and C. jejuni invasion over time. In studies using pharmacological inhibitors, silencing RNA (siRNA) and dominant-negative expression constructs, EGFR, PDGFR and PI3-kinase appeared to represent other crucial components upstream of Cdc42 and invasion. siRNA and the use of Vav1\\/2-\\/- knockout cells further showed that the guanine exchange factor Vav2 is required for Cdc42 activation and maximal bacterial invasion. Overexpression of certain mutant constructs indicated that Vav2 is a linker

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  11. Function of the integrin alpha 6 beta 1 in metastatic breast carcinoma cells assessed by expression of a dominant-negative receptor

    DEFF Research Database (Denmark)

    Shaw, L M; Chao, C; Wewer, U M;

    1996-01-01

    : alpha 2 beta 1, alpha 3 beta 1, and alpha 6 beta 1, but uses only alpha 6 beta 1 to mediate adhesion and migration on laminin matrices. To investigate the contribution of alpha 6 beta 1 to the aggressive behavior of these cells, we developed a dominant-negative strategy for knocking out alpha 6 beta 1...

  12. Venom gland EST analysis of the saw-scaled viper, Echis ocellatus, reveals novel alpha9beta1 integrin-binding motifs in venom metalloproteinases and a new group of putative toxins, renin-like aspartic proteases.

    Science.gov (United States)

    Wagstaff, Simon C; Harrison, Robert A

    2006-08-01

    Echis ocellatus is the most medically important snake in West Africa. However, the composition of its venom and the differential contribution of these venom components to the severe haemorrhagic and coagulopathic pathology of envenoming are poorly understood. To address this situation we assembled a toxin transcriptome based upon 1000 expressed sequence tags (EST) from a cDNA library constructed from pooled venom glands of 10 individual E. ocellatus. We used a variety of bioinformatic tools to construct a fully annotated venom-toxin transcriptome that was interrogated with a combination of BLAST annotation, gene ontology cataloguing and disintegrin-motif searching. The results of these analyses revealed an unusually abundant and diverse expression of snake venom metalloproteinases (SVMP) and a broad toxin-expression profile including several distinct isoforms of bradykinin-potentiating peptides, phospholipase A(2), C-type lectins, serine proteinases and l-amino oxidases. Most significantly, we identified for the first time a conserved alpha(9)beta(1) integrin-binding motif in several SVMPs, and a new group of putative venom toxins, renin-like aspartic proteases. PMID:16713134

  13. Integrin beta 1 enhances the epithelial-mesenchymal transition in association with gefitinib resistance of non-small cell lung cancer.

    Science.gov (United States)

    Ju, Lixia; Zhou, Caicun

    2013-01-01

    We have previously shown that integrinβ1 associates with gefitinib resistance. As epithelial-mesenchymal transition (EMT) also induces gefitinib resistance in vitro, we wished to determine the relation of them in gefitinib resistance. In this study, we show that integrinβ1 induced epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) resistance in xenograft tumors and gefitinib-resistant NSCLC tumors acquired EMT phenotype. Furthermore, inhibition of integrinβ1 reverses EMT, meanwhile overexpression and activation of integrinβ1 aggravates EMT. Lastly, we further identified that integrinβ1 enhanced EMT via FAK-AKT signaling pathway. These findings highlight a novel relation of integrinβ1 and EMT in EGFR TKI resistant NSCLC. PMID:24440972

  14. Investigating the role of Integrin Linked Kinase in mammary epithelial cell differentiation

    OpenAIRE

    Rooney, Nicholas

    2014-01-01

    Epithelial cell adhesion to the surrounding extracellular matrix (ECM) is necessary for their proper behaviour and function. During pregnancy and lactation mammary epithelial cells (MECs) require signals imparted by specific β1 integrin-laminin interactions for their functional differentiation in response to Prolactin (Prl) and for the correct formation of polarised secretory acini. Downstream of β1 integrin (β1Itg), the scaffold protein Integrin Linked Kinase (ILK) has been identified as the...

  15. Integrin alpha 5 beta 1 Plays a Critical Role in Resistance to Temozolomide by Interfering with the p53 Pathway in High-Grade Glioma

    Czech Academy of Sciences Publication Activity Database

    Janoušková, Hana; Maglott, A.; Leger, D.Y.; Bossert, C.; Noulet, F.; Guerin, E.; Guenot, D.; Pinel, S.; Chastagner, P.; Plenat, F.; Entz-Werle, N.; Lehmann-Che, J.; Godet, J.; Martin, S.; Teisinger, Jan; Dontenwill, M.

    2012-01-01

    Roč. 72, č. 14 (2012), s. 3463-3470. ISSN 0008-5472 Institutional support: RVO:67985823 Keywords : cancer * integrins * high-grade glioma Subject RIV: CE - Biochemistry Impact factor: 8.650, year: 2012

  16. Measurement of the Interaction Between Recombinant I-domain from Integrin alpha 2 beta 1 and a Triple Helical Collagen Peptide with the GFOGER Binding Motif Using Molecular Force Spectroscopy

    Directory of Open Access Journals (Sweden)

    Mark E. Welland

    2013-01-01

    Full Text Available The role of the collagen-platelet interaction is of crucial importance to the haemostatic response during both injury and pathogenesis of the blood vessel wall. Of particular interest is the high affinity interaction of the platelet transmembrane receptor, alpha 2 beta 1, responsible for firm attachment of platelets to collagen at and around injury sites. We employ single molecule force spectroscopy (SMFS using the atomic force microscope (AFM to study the interaction of the I-domain from integrin alpha 2 beta 1 with a synthetic collagen related triple-helical peptide containing the high-affinity integrin-binding GFOGER motif, and a control peptide lacking this sequence, referred to as GPP. By utilising synthetic peptides in this manner we are able to study at the molecular level subtleties that would otherwise be lost when considering cell-to-collagen matrix interactions using ensemble techniques. We demonstrate for the first time the complexity of this interaction as illustrated by the complex multi-peaked force spectra and confirm specificity using control blocking experiments. In addition we observe specific interaction of the GPP peptide sequence with the I-domain. We propose a model to explain these observations.

  17. Discovery, SAR, and Radiolabeling of Halogenated Benzimidazole Carboxamide Antagonists as Useful Tools for (alpha)4(beta)1 Integrin Expressed on T- and B-cell Lymphomas

    Energy Technology Data Exchange (ETDEWEB)

    Carpenter, R D; Natarajan, A; Lau, E Y; Andrei, M; Solano, D M; Lightstone, F C; DeNardo, S J; Lam, K S; Kurth, M J

    2010-02-08

    The cell surface receptor {alpha}{sub 4}{beta}{sub 1} integrin is an attractive yet poorly understood target for selective diagnosis and treatment of T- and B-cell lymphomas. This report focuses on the rapid microwave preparation of medicinally pertinent benzimidazole heterocycles, structure-activity relationships (SAR) of novel halobenzimidazole carboxamide antagonists 3-6, and preliminary biological evaluation of radioiodinated agents 7, 8, and 18. The I-125 derivative 18 had good tumor uptake (12 {+-} 1% ID/g at 24 h; 4.5 {+-} 1% ID/g at 48 h) and tumor:kidney ratio ({approx}4:1 at 24 h; 2.5:1 at 48 h) in xenograft murine models of B-cell lymphoma. Molecular homology models of {alpha}{sub 4}{beta}{sub 1} integrin have predicted that docked halobenzimidazole carboxamides have the halogen atom in a suitable orientation for halogen-hydrogen bonding. These high affinity ({approx} pM binding) halogenated ligands are attractive tools for medicinal and biological use; the fluoro and iodo derivatives are potential radiodiagnostic ({sup 18}F) or radiotherapeutic ({sup 131}I) agents, whereas the chloro and bromo analogues could provide structural insight into integrin-ligand interactions through photoaffinity cross-linking/mass spectroscopy experiments, as well as co-crystallization X-ray studies.

  18. Signaling from beta1- and beta2-adrenergic receptors is defined by differential interactions with PDE4

    DEFF Research Database (Denmark)

    Richter, Wito; Day, Peter; Agrawal, Rani; Bruss, Matthew D; Granier, Sébastien; Wang, Yvonne L; Rasmussen, Søren Gøgsig Faarup; Horner, Kathleen; Wang, Ping; Lei, Tao; Patterson, Andrew J; Kobilka, Brian; Conti, Marco

    2008-01-01

    Beta1- and beta2-adrenergic receptors (betaARs) are highly homologous, yet they play clearly distinct roles in cardiac physiology and pathology. Myocyte contraction, for instance, is readily stimulated by beta1AR but not beta2AR signaling, and chronic stimulation of the two receptors has opposing...

  19. Osteogenic differentiation of mesenchymal stem cells from dental bud: Role of integrins and cadherins

    Directory of Open Access Journals (Sweden)

    Adriana Di Benedetto

    2015-11-01

    Full Text Available Several studies have reported the beneficial effects of mesenchymal stem cells (MSCs in tissue repair and regeneration. New sources of stem cells in adult organisms are continuously emerging; dental tissues have been identified as a source of postnatal MSCs. Dental bud is the immature precursor of the tooth, is easy to access and we show in this study that it can yield a high number of cells with ≥95% expression of mesenchymal stemness makers and osteogenic capacity. Thus, these cells can be defined as Dental Bud Stem Cells (DBSCs representing a promising source for bone regeneration of stomatognathic as well as other systems. Cell interactions with the extracellular matrix (ECM and neighboring cells are critical for tissue morphogenesis and architecture; such interactions are mediated by integrins and cadherins respectively. We characterized DBSCs for the expression of these adhesion receptors and examined their pattern during osteogenic differentiation. Our data indicate that N-cadherin and cadherin-11 were expressed in undifferentiated DBSCs and their expression underwent changes during the osteogenic process (decreasing and increasing respectively, while expression of E-cadherin and P-cadherin was very low in DBSCs and did not change during the differentiation steps. Such expression pattern reflected the mesenchymal origin of DBSCs and confirmed their osteoblast-like features. On the other hand, osteogenic stimulation induced the upregulation of single subunits, αV, β3, α5, and the formation of integrin receptors α5β1 and αVβ3. DBSCs differentiation toward osteoblastic lineage was enhanced when cells were grown on fibronectin (FN, vitronectin (VTN, and osteopontin (OPN, ECM glycoproteins which contain an integrin-binding sequence, the RGD motif. In addition we established that integrin αVβ3 plays a crucial role during the commitment of MSCs to osteoblast lineage, whereas integrin α5β1 seems to be dispensable. These data suggest

  20. Hierarchy of ADAM12 binding to integrins in tumor cells

    DEFF Research Database (Denmark)

    Thodeti, Charles Kumar; Fröhlich, Camilla; Nielsen, Christian Kamp;

    2005-01-01

    12. However, when alpha9beta1 integrin is not expressed--as in many carcinoma cells--other members of the beta1 integrin family can replace its ligand binding activity. In attachment assays, the recombinant disintegrin domain of ADAM12 only supported alpha9 integrin-dependent tumor cell attachment...... with a rounded morphology; attachment of cells with a spread morphology required further activation of the alpha9beta1 integrin. We demonstrated that phosphoinositide-3-kinase appears to be central in regulating alpha9beta1 integrin cell spreading activity in response to ADAM12....

  1. Evaluation of β1-integrin expression on chondrogenically differentiating human adipose-derived stem cells using atomic force microscopy.

    Science.gov (United States)

    Quisenberry, Chrystal R; Nazempour, Arshan; Van Wie, Bernard J; Abu-Lail, Nehal I

    2016-06-01

    The expression of β1-integrin on human adipose-derived stem cells, differentiating toward a chondrogenic lineage, is hypothesized to decrease when cells are grown under in vivo-like environments due to sufficient extracellular matrix (ECM) buildup in the engineered tissues. The opposite is true when cells are grown in static cultures such as in pellet or micromass. To probe β1-integrin distribution on cellular surfaces, atomic force microscopy cantilevers modified with anti-β1-integrin antibodies were used. Specific antibody-antigen adhesion forces were identified and indicated the locations of β1-integrins on cells. ECM properties were assessed by estimating the Young's modulus of the matrix. Specific single antibody-antigen interactions averaged 78 ± 10 pN with multiple bindings occurring at approximate multiples of 78 pN. The author's results show that upregulated β1-integrin expression coincided with a less robust ECM as assessed by mechanical properties of tissues. In micromass and pellet cultures, transforming growth factor β3(TGF-β3) elicited a decrease in Young's modulus by 3.7- and 4.4-fold while eliciting an increase in β1-integrin count by 1.1- and 1.3-fold, respectively. β1-integrin counts on cells grown in the presence of TGF-β3 with oscillating hydrostatic pressure decreased by a 1.1-fold while the Young's modulus increased by a 1.9-fold. Collectively, our results suggest that cells in insufficiently robust ECM express more integrin perhaps to facilitate cell-ECM adhesion and compensate for a looser less robust ECM. PMID:27106564

  2. α2 Integrin, extracellular matrix metalloproteinase inducer, and matrix metalloproteinase-3 act sequentially to induce differentiation of mouse embryonic stem cells into odontoblast-like cells

    Energy Technology Data Exchange (ETDEWEB)

    Ozeki, Nobuaki; Kawai, Rie; Hase, Naoko; Hiyama, Taiki; Yamaguchi, Hideyuki [Department of Endodontics, School of Dentistry, Aichi Gakuin University, Nagoya, Aichi 464-8651 (Japan); Kondo, Ayami [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, Nagoya 464-8650 (Japan); Nakata, Kazuhiko [Department of Endodontics, School of Dentistry, Aichi Gakuin University, Nagoya, Aichi 464-8651 (Japan); Mogi, Makio, E-mail: makio@dpc.agu.ac.jp [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, Nagoya 464-8650 (Japan)

    2015-02-01

    We previously reported that interleukin 1β acts via matrix metalloproteinase (MMP)-3 to regulate cell proliferation and suppress apoptosis in α2 integrin-positive odontoblast-like cells differentiated from mouse embryonic stem (ES) cells. Here we characterize the signal cascade underpinning odontoblastic differentiation in mouse ES cells. The expression of α2 integrin, extracellular matrix metalloproteinase inducer (Emmprin), and MMP-3 mRNA and protein were all potently increased during odontoblastic differentiation. Small interfering RNA (siRNA) disruption of the expression of these effectors potently suppressed the expression of the odontoblastic biomarkers dentin sialophosphoprotein, dentin matrix protein-1 and alkaline phosphatase, and blocked odontoblast calcification. Our siRNA, western blot and blocking antibody analyses revealed a unique sequential cascade involving α2 integrin, Emmprin and MMP-3 that drives ES cell differentiation into odontoblasts. This cascade requires the interaction between α2 integrin and Emmprin and is potentiated by exogenous MMP-3. Finally, although odontoblast-like cells potently express α2, α6, αV, β1, and β3, integrins, we confirmed that β1 integrin acts as the trigger for ES cell differentiation, apparently in complex with α2 integrin. These results demonstrate a unique and unanticipated role for an α2 integrin-, Emmprin-, and MMP-3-mediated signaling cascade in driving mouse ES cell differentiation into odontoblast-like cells. - Highlights: • Odontoblast differentiation requires activation of α2 integrin, Emmprin and MMP-3. • α2 integrin, Emmprin and MMP-3 form a sequential signaling cascade. • β1 integrin acts a specific trigger for odontoblast differentiation. • The role of these effectors is highly novel and unanticipated.

  3. α2 Integrin, extracellular matrix metalloproteinase inducer, and matrix metalloproteinase-3 act sequentially to induce differentiation of mouse embryonic stem cells into odontoblast-like cells

    International Nuclear Information System (INIS)

    We previously reported that interleukin 1β acts via matrix metalloproteinase (MMP)-3 to regulate cell proliferation and suppress apoptosis in α2 integrin-positive odontoblast-like cells differentiated from mouse embryonic stem (ES) cells. Here we characterize the signal cascade underpinning odontoblastic differentiation in mouse ES cells. The expression of α2 integrin, extracellular matrix metalloproteinase inducer (Emmprin), and MMP-3 mRNA and protein were all potently increased during odontoblastic differentiation. Small interfering RNA (siRNA) disruption of the expression of these effectors potently suppressed the expression of the odontoblastic biomarkers dentin sialophosphoprotein, dentin matrix protein-1 and alkaline phosphatase, and blocked odontoblast calcification. Our siRNA, western blot and blocking antibody analyses revealed a unique sequential cascade involving α2 integrin, Emmprin and MMP-3 that drives ES cell differentiation into odontoblasts. This cascade requires the interaction between α2 integrin and Emmprin and is potentiated by exogenous MMP-3. Finally, although odontoblast-like cells potently express α2, α6, αV, β1, and β3, integrins, we confirmed that β1 integrin acts as the trigger for ES cell differentiation, apparently in complex with α2 integrin. These results demonstrate a unique and unanticipated role for an α2 integrin-, Emmprin-, and MMP-3-mediated signaling cascade in driving mouse ES cell differentiation into odontoblast-like cells. - Highlights: • Odontoblast differentiation requires activation of α2 integrin, Emmprin and MMP-3. • α2 integrin, Emmprin and MMP-3 form a sequential signaling cascade. • β1 integrin acts a specific trigger for odontoblast differentiation. • The role of these effectors is highly novel and unanticipated

  4. Quantitative Phosphoproteomic Study Reveals that Protein Kinase A Regulates Neural Stem Cell Differentiation Through Phosphorylation of Catenin Beta-1 and Glycogen Synthase Kinase 3β.

    Science.gov (United States)

    Wang, Shuxin; Li, Zheyi; Shen, Hongyan; Zhang, Zhong; Yin, Yuxin; Wang, Qingsong; Zhao, Xuyang; Ji, Jianguo

    2016-08-01

    Protein phosphorylation is central to the understanding of multiple cellular signaling pathways responsible for regulating the self-renewal and differentiation of neural stem cells (NSCs). Here we performed a large-scale phosphoproteomic analysis of rat fetal NSCs using strong cation exchange chromatography prefractionation and citric acid-assisted two-step enrichment with TiO2 strategy followed by nanoLC-MS/MS analysis. Totally we identified 32,546 phosphosites on 5,091 phosphoproteins, among which 23,945 were class I phosphosites, and quantified 16,000 sites during NSC differentiation. More than 65% of class I phosphosites were novel when compared with PhosphoSitePlus database. Quantification results showed that the early and late stage of NSC differentiation differ greatly. We mapped 69 changed phosphosites on 20 proteins involved in Wnt signaling pathway, including S552 on catenin beta-1 (Ctnnb1) and S9 on glycogen synthase kinase 3β (Gsk3β). Western blotting and real-time PCR results proved that Wnt signaling pathway plays critical roles in NSC fate determination. Furthermore, inhibition and activation of PKA dramatically affected the phosphorylation state of Ctnnb1 and Gsk3β, which regulates the differentiation of NSCs. Our data provides a valuable resource for studying the self-renewal and differentiation of NSCs. Stem Cells 2016;34:2090-2101. PMID:27097102

  5. Differential effects of bone morphogenetic protein-2 and transforming growth factor-beta1 on gene expression of collagen-modifying enzymes in human adipose tissue-derived mesenchymal stem cells.

    Science.gov (United States)

    Knippenberg, Marlene; Helder, Marco N; Doulabi, Behrouz Zandieh; Bank, Ruud A; Wuisman, Paul I J M; Klein-Nulend, Jenneke

    2009-08-01

    Adipose tissue-derived mesenchymal stem cells (AT-MSCs) in combination with bone morphogenetic protein-2 (BMP-2) or transforming growth factor-beta1 (TGF-beta1) are under evaluation for bone tissue engineering. Posttranslational modification of type I collagen is essential for functional bone tissue with adequate physical and mechanical properties. We investigated whether BMP-2 (10-100 ng/mL) and/or TGF-beta1 (1-10 ng/mL) affect gene expression of alpha2(I) procollagen and collagen-modifying enzymes, that is, lysyl oxidase and lysyl hydroxylases 1, 2, and 3 (encoded by PLOD1, 2, and 3), by human AT-MSCs. BMP-2, but not TGF-beta1, increased alkaline phosphatase activity after 28 days, indicating osteogenic differentiation of AT-MSCs. At day 4, both BMP-2 and TGF-beta1 upregulated alpha2(I) procollagen and PLOD1, which was downregulated at day 28. TGF-beta1, but not BMP-2, downregulated PLOD3 at day 28. Lysyl oxidase was upregulated by TGF-beta1 at day 4 and by BMP-2 at day 7. Neither BMP-2 nor TGF-beta1 affected PLOD2. In conclusion, these results suggest that AT-MSCs differentially respond to BMP-2 and TGF-beta1 with changes in gene expression of collagen-modifying enzymes. AT-MSCs may thus be able to appropriately modify type I collagen to form a functional bone extracellular matrix for tissue engineering, dependent on the growth factor added. PMID:19231972

  6. β3-integrin is required for differentiation in OC-2 cells derived from mammalian embryonic inner ear

    Directory of Open Access Journals (Sweden)

    Brunetta Ivan

    2012-03-01

    Full Text Available Abstract Background The mammalian inner ear contains the organ of Corti which is responsible for the conversion of sound into neuronal signals. This specialised epithelial tissue is the product of a complex developmental process where a common precursor cell type differentiates into the sound transducing hair cells and the non-innervated supporting cells. We hypothesised that integrin proteins, which are involved in cell attachment to extracellular matrix proteins and cellular signalling, play a role in the differentiation process of the precursor inner ear epithelial cells. To test our hypothesis we have utilised a cell line (OC-2 derived from E13 embryonic immortomouse inner ears. In vitro, by switching the incubation temperature from 33°C to 39°C, the OC-2 cells can be induced to differentiate and express hair cells markers, such as Myosin VIIa. The OC-2 cells are thus a useful model system for testing mechanism of hair cells differentiation. Results We have identified 4 integrin subunits which are expressed in OC-2 cells: α6, αv, β1 and β3. Among these, the relative level of expression of the αv, β1 and β3 subunits increased in a time dependent manner when the cells were exposed to the differentiating temperature of 39°C, most notably so for β3 which was not detectable at 33°C. Treatment of fully differentiated OC-2 cells with siRNA against the four integrin subunits reduced the expression of not only the respective integrin proteins but also of the hair cell marker Myosin VIIa. Conversely over-expression of β3 was sufficient to induce the expression of Myosin VIIa at 33°C. Conclusions Our data demonstrate that modulation of integrin expression is associated with the differentiation process of the OC-2 cells. This suggests that the maturation of the organ of Corti, from where OC-2 cells are derived, may also depend on changes of gene expression associated with integrin expression.

  7. Measurement of the Interaction Between Recombinant I-domain from Integrin alpha 2 beta 1 and a Triple Helical Collagen Peptide with the GFOGER Binding Motif Using Molecular Force Spectroscopy

    OpenAIRE

    Welland, Mark E.; Farndale, Richard W.; Dominique Bihan; Debdulal Roy; Simon J Attwood; Simpson, Anna M. C.; Hamaia, Samir W.

    2013-01-01

    The role of the collagen-platelet interaction is of crucial importance to the haemostatic response during both injury and pathogenesis of the blood vessel wall. Of particular interest is the high affinity interaction of the platelet transmembrane receptor, alpha 2 beta 1, responsible for firm attachment of platelets to collagen at and around injury sites. We employ single molecule force spectroscopy (SMFS) using the atomic force microscope (AFM) to study the interaction of the I-domain from i...

  8. Natalizumab plus interferon beta-1a for relapsing multiple sclerosis.

    NARCIS (Netherlands)

    Rudick, R.A.; Stuart, W.H.; Calabresi, P.A.; Confavreux, C.; Galetta, S.L.; Radue, E.W.; Lublin, F.D.; Weinstock-Guttman, B.; Wynn, D.R.; Lynn, F.; Panzara, M.A.; Sandrock, A.W.

    2006-01-01

    BACKGROUND: Interferon beta is used to modify the course of relapsing multiple sclerosis. Despite interferon beta therapy, many patients have relapses. Natalizumab, an alpha4 integrin antagonist, appeared to be safe and effective alone and when added to interferon beta-1a in preliminary studies. MET

  9. The Integrin-Mediated ILK-Parvin-αPix Signaling Axis Controls Differentiation in Mammary Epithelial Cells.

    Science.gov (United States)

    Rooney, Nicholas; Wang, Pengbo; Brennan, Keith; Gilmore, Andrew P; Streuli, Charles H

    2016-11-01

    Epithelial cell adhesion to the surrounding extracellular matrix is necessary for their proper behavior and function. During pregnancy and lactation, mammary epithelial cells (MECs) receive signals from their interaction with laminin via β1-integrin (β1-itg) to establish apico-basal polarity and to differentiate in response to prolactin. Downstream of β1-itg, the scaffold protein Integrin Linked Kinase (ILK) has been identified as the key signal transducer that is required for both lactational differentiation and the establishment of apico-basal polarity. ILK is an adaptor protein that forms the IPP complex with PINCH and Parvins, which are central to its adaptor functions. However, it is not known how ILK and its interacting partners control tissue-specific gene expression. Expression of ILK mutants, which weaken the interaction between ILK and Parvin, revealed that Parvins have a role in mammary epithelial differentiation. This conclusion was supported by shRNA-mediated knockdown of the Parvins. In addition, shRNA knockdown of the Parvin-binding guanine nucleotide exchange factor αPix prevented prolactin-induced differentiation. αPix depletion did not disrupt focal adhesions, MEC proliferation, or polarity. This suggests that αPix represents a differentiation-specific bifurcation point in β1-itg-ILK adhesive signaling. In summary, this study has identified a new role for Parvin and αPix downstream of the integrin-ILK signaling axis for MEC differentiation. J. Cell. Physiol. 231: 2408-2417, 2016. © 2016 Wiley Periodicals, Inc. PMID:27019299

  10. Why cellular stress suppresses adipogenesis in skeletal tissue, but is ineffective in adipose tissue: Control of mesenchymal cell differentiation via integrin binding sites in extracellular matrices

    OpenAIRE

    Volloch, Vladimir; Olsen, Bjorn R.

    2013-01-01

    This Perspective addresses one of the major puzzles of adipogenesis in adipose tissue, namely its resistance to cellular stress. It introduces a concept of “density” of integrin binding sites in extracellular matrix, proposes a cellular signaling explanation for the observed effects of matrix elasticity and of cell shape on mesenchymal stem cell differentiation, and discusses how specialized integrin binding sites in collagen IV - containing matrices guard two pivotal physiological and evolut...

  11. Biomimetic integrin-specific surfaces to direct osteoblastic function and tissue healing

    Science.gov (United States)

    Petrie, Timothy Andrew

    Current orthopedic implant technologies used suffer from slow rates of osseointegration, short lifetime, and lack of mechanical integrity as a result of poorly controlled cell-surface interactions. Recent biologically-inspired surface strategies (biomimetic) have focused on mimicking the biofunctionality of the extracellular matrix (ECM) by using short, adhesive oligopeptides, such as arginine-glycine-aspartic acid (RGD) present in numerous ECM components. However, these strategies have yielded mixed results in vivo and marginal bone healing responses. The central goal of this dissertation project was to engineer bioactive surfaces that specifically target integrin receptors important for osteogenic functions in order to improve bone tissue repair. In order to create integrin-specific interfaces, integrin-specific ligands reconstituting the fibronectin (FN) secondary/tertiary structure were first engineered and functionalized on material surfaces using several robust presentation schemes. We demonstrated that FN-mimetic-functionalized surfaces that directed alpha 5beta1 binding enhanced osteoblast and stromal cell integrin binding and adhesion, osteogenic signaling, and osteoblastic differentiation compared to various other RGD-based ligand-functionalized surfaces. Next, we investigated the effect of integrin-specific biointerfaces to modulate bone healing in a rat tibia implant bone model. We demonstrated, using a robust polymer brush system, that bioactive coatings on titanium implants that conferred high alpha5beta1 integrin specificity in vitro enhanced bone formation and implant integration in vivo. Moreover, we showed that integrin specificity can be engineered using different immobilization schemes, including clinically-relevant ligand dip-coating, and promote the same robust in vivo effect. Furthermore, we investigate the synergistic roles of integrin specificity and ligand clustering on cell response by engineering biointerfaces presenting trimeric and

  12. Low-Shear Modelled Microgravity Environment Maintains Morphology and Differentiated Functionality of Primary Porcine Hepatocyte Cultures

    OpenAIRE

    Nelson, Leonard J; Walker, Simon W.; Peter C. Hayes; Plevris, John N

    2010-01-01

    Hepatocytes cultured in conventional static culture rapidly lose polarity and differentiated function. This could be explained by gravity-induced sedimentation, which prevents formation of complete three-dimensional (3D) cell-cell/cellmatrix interactions and disrupts integrin-mediated signals (including the most abundant hepatic integrin alpha(5)beta(1)), important for cellular polarity and differentiation. Cell culture in a low fluid shear modelled microgravity (about 10(-2) g) environment p...

  13. Interferon Beta-1b Injection

    Science.gov (United States)

    Interferon beta-1b injection is used to reduce episodes of symptoms in patients with relapsing-remitting (course ... and problems with vision, speech, and bladder control). Interferon beta-1b is in a class of medications ...

  14. Chondroitin sulfate proteoglycans regulate the growth, differentiation and migration of multipotent neural precursor cells through the integrin signaling pathway

    Directory of Open Access Journals (Sweden)

    Lü He-Zuo

    2009-10-01

    Full Text Available Abstract Background Neural precursor cells (NPCs are defined by their ability to proliferate, self-renew, and retain the potential to differentiate into neurons and glia. Deciphering the factors that regulate their behaviors will greatly aid in their use as potential therapeutic agents or targets. Chondroitin sulfate proteoglycans (CSPGs are prominent components of the extracellular matrix (ECM in the central nervous system (CNS and are assumed to play important roles in controlling neuronal differentiation and development. Results In the present study, we demonstrated that CSPGs were constitutively expressed on the NPCs isolated from the E16 rat embryonic brain. When chondroitinase ABC was used to abolish the function of endogenous CSPGs on NPCs, it induced a series of biological responses including the proliferation, differentiation and migration of NPCs, indicating that CSPGs may play a critical role in NPC development and differentiation. Finally, we provided evidence suggesting that integrin signaling pathway may be involved in the effects of CSPGs on NPCs. Conclusion The present study investigating the influence and mechanisms of CSPGs on the differentiation and migration of NPCs should help us to understand the basic biology of NPCs during CNS development and provide new insights into developing new strategies for the treatment of the neurological disorders in the CNS.

  15. Differential gene expression by integrin β7+ and β7- memory T helper cells

    Directory of Open Access Journals (Sweden)

    Yang Yee

    2004-07-01

    Full Text Available Abstract Background The cell adhesion molecule integrin α4β7 helps direct the migration of blood lymphocytes to the intestine and associated lymphoid tissues. We hypothesized that β7+ and β7- blood memory T helper cells differ in their expression of genes that play a role in the adhesion or migration of T cells. Results RNA was prepared from β7+ and β7- CD4+ CD45RA- blood T cells from nine normal human subjects and analyzed using oligonucleotide microarrays. Of 21357 genes represented on the arrays, 16 were more highly expressed in β7+ cells and 18 were more highly expressed in β7- cells (≥1.5 fold difference and adjusted P + memory/effector T cells than on β7- cells. Conclusions Memory/effector T cells that express integrin β7 have a distinct pattern of expression of a set of gene transcripts. Several of these molecules can affect cell adhesion or chemotaxis and are therefore likely to modulate the complex multistep process that regulates trafficking of CD4+ memory T cell subsets with different homing behaviors.

  16. Can alterations in integrin and laminin-5 expression be used as markers of malignancy?

    DEFF Research Database (Denmark)

    Thorup, A K; Reibel, J; Schiødt, M;

    1998-01-01

    Development of squamous cell carcinomas (SCCs) involves alterations in the adhesive interactions in the epithelium and invasion through the basement membrane. Therefore, changes in the expression of receptors and ligands involved in cell-cell and cell-matrix adhesion may be essential for the...... transformation of a premalignant into a malignant lesion. The aim of this study was to examine if expression of specific cell adhesion molecules can be used as markers of malignant development. By immunohistochemistry, we examined the expression pattern of integrins alpha2beta1, alpha3beta1, alpha6beta4 and...... laminin-5 in biopsies from SCCs (n=18), premalignant lesions (leukoplakias, n=21) and non-premalignant tissue with chronic inflammation (n=11). In poorly differentiated SCCs, patchy loss of alpha3beta1, alpha6beta4 and laminin-5 expression was pronounced at the invasion front, whereas there was a tendency...

  17. Increased Osteogenic Differentiation of Periodontal Ligament Stem Cells on Polydopamine Film Occurs via Activation of Integrin and PI3K Signaling Pathways

    Directory of Open Access Journals (Sweden)

    Jeong Seok Lee

    2014-11-01

    Full Text Available Background/Aims: Mussel-inspired polydopamine (PDA is known to be an effective bioadhesive and bioactive material for controlling stem cell fate, which is important in stem cell-based regenerative medicine; however, the effect of PDA on osteogenic differentiation of periodontal ligament stem cells (PDLSCs is not fully understood. In this study, we investigated the osteoinductive effect of PDA on PDLSCs and examined how this phenomenon is encouraged. Methods: Osteogenic induction of PDLSCs was established by culturing cells on PDA film or on an uncoated polystyrene surface as a control. Osteogenic differentiation of PDLSCs was assessed by measurement of intracellular calcium levels and alkaline phosphatase (ALP activity as well as by evaluation of protein expression of osteocalcin (OCN, osterix (OSX, and runt-related transcription factor 2 (RUNX2. Results: The PDLSCs cultured on PDA film showed higher osteogenic activity than those on the control surface. Moreover, PDLSCs on PDA film expressed increased levels of the integrin adhesion receptors integrin α5 and β1 compared to control cells. Expression of one isoform of the intracellular signaling protein phosphatidylinositol-3-kinase (PI3K, p110γ, was increased in PDLSCs on PDA film in a PDA dose-dependent manner. This signaling protein was found to interact with integrin β1, demonstrating integrin-linked PI3K activation in response to PDA. Finally, the blockage of PI3K reduced the PDA-induced osteogenic activity of PDLSCs. Conclusion: our findings suggest that the bioadhesive PDA stimulates osteogenic differentiation of PDLSCs via activation of the integrin α5/β1 and PI3K signaling pathways.

  18. Targeted disruption of the LAMA3 gene in mice reveals abnormalities in survival and late stage differentiation of epithelial cells.

    Science.gov (United States)

    Ryan, M C; Lee, K; Miyashita, Y; Carter, W G

    1999-06-14

    Laminin 5 regulates anchorage and motility of epithelial cells through integrins alpha6beta4 and alpha3beta1, respectively. We used targeted disruption of the LAMA3 gene, which encodes the alpha3 subunit of laminin 5 and other isoforms, to examine developmental functions that are regulated by adhesion to the basement membrane (BM). In homozygous null animals, profound epithelial abnormalities were detected that resulted in neonatal lethality, consistent with removal of all alpha3-laminin isoforms from epithelial BMs. Alterations in three different cellular functions were identified. First, using a novel tissue adhesion assay, we found that the mutant BM could not induce stable adhesion by integrin alpha6beta4, consistent with the presence of junctional blisters and abnormal hemidesmosomes. In the absence of laminin 5 function, we were able to detect a new ligand for integrin alpha3beta1 in the epidermal BM, suggesting that basal keratinocytes can utilize integrin alpha3beta1 to interact with an alternative ligand. Second, we identified a survival defect in mutant epithelial cells that could be rescued by exogenous laminin 5, collagen, or an antibody against integrin alpha6beta4, suggesting that signaling through beta1 or beta4 integrins is sufficient for survival. Third, we detected abnormalities in ameloblast differentiation in developing mutant incisors indicating that events downstream of adhesion are affected in mutant animals. These results indicate that laminin 5 has an important role in regulating tissue organization, gene expression, and survival of epithelium. PMID:10366601

  19. Muscle-specific integrins in masseter muscle fibers of chimpanzees: an immunohistochemical study.

    Directory of Open Access Journals (Sweden)

    Gianluigi Vaccarino

    2010-05-01

    differentiation, and cell-extracellular matrix interactions. Our results demonstrated a different quantitative composition of integrins, in alpha male in respect to human and non-alpha male, hypothesizing that the MYH16 gene could modify the expression of integrins, influencing, in turn, the phenotype of muscle. In this way, alpha 7A-and beta 1A-integrin could determine the presence of type II fibers and then they could play a key role in the determination of contraction force. Then, MYH16 gene could be a common interactor of signalling between sarcoglycans and integrins in chimpanzee muscles.

  20. Differential Expression Levels of Integrin α6 Enable the Selective Identification and Isolation of Atrial and Ventricular Cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Anne Maria Wiencierz

    Full Text Available Central questions such as cardiomyocyte subtype emergence during cardiogenesis or the availability of cardiomyocyte subtypes for cell replacement therapy require selective identification and purification of atrial and ventricular cardiomyocytes. However, current methodologies do not allow for a transgene-free selective isolation of atrial or ventricular cardiomyocytes due to the lack of subtype specific cell surface markers.In order to develop cell surface marker-based isolation procedures for cardiomyocyte subtypes, we performed an antibody-based screening on embryonic mouse hearts. Our data indicate that atrial and ventricular cardiomyocytes are characterized by differential expression of integrin α6 (ITGA6 throughout development and in the adult heart. We discovered that the expression level of this surface marker correlates with the intracellular subtype-specific expression of MLC-2a and MLC-2v on the single cell level and thereby enables the discrimination of cardiomyocyte subtypes by flow cytometry. Based on the differential expression of ITGA6 in atria and ventricles during cardiogenesis, we developed purification protocols for atrial and ventricular cardiomyocytes from mouse hearts. Atrial and ventricular identities of sorted cells were confirmed by expression profiling and patch clamp analysis.Here, we introduce a non-genetic, antibody-based approach to specifically isolate highly pure and viable atrial and ventricular cardiomyocytes from mouse hearts of various developmental stages. This will facilitate in-depth characterization of the individual cellular subsets and support translational research applications.

  1. Differential β3 and β1 Integrin Expression in Bone Marrow and Cortical Bone of Estrogen Deficient Rats.

    Science.gov (United States)

    Voisin, Muriel; McNamara, Laoise M

    2015-09-01

    Integrin-based (β3 ) attachments to the extracellular matrix (ECM) on osteocyte cell processes have recently been proposed to play an important role in facilitating osteocyte mechanosensation. However, it is not yet known whether integrin expression is altered in the mechanoregulatory osteocytes during osteoporosis. The objective of this study was to test the hypothesis that the expression of integrin-based mechanosensory complexes (β1 and β3 integrins) is altered as a direct response to estrogen deficiency, in an estrogen deficient animal model of osteoporosis. Four weeks post-operatively, immunohistochemistry was used to detect for β1 and β3 integrin subunits in bone tissue and marrow of ovariectomized (OVX; N = 4) and SHAM (N = 4) operated animals. A tartrate resistant acid phosphatase (TRAP) control stain was performed to quantify the presence of osteoclasts in the bone marrow and bone surfaces. Image analysis was performed to quantify expression patterns in different biological compartments, that is, bone marrow, endosteum, and cortical bone. Our results showed that β1 integrins were ubiquitously expressed throughout the bone and marrow, for both OVX and SHAM groups. β3 integrin subunit expression was lower in bone cells from osteoporotic animals compared to controls, whereas β3 expression in marrow cells did not differ significantly between groups. At the endosteum no difference was observed in β3 integrin subunit expression. As expected, the number of osteoclasts was higher in the OVX group validating an imbalance in bone remodeling. We propose that a reduction in β3 integrin expression in osteocytes might impair mechanosensation by bone cells during estrogen deficiency. PMID:25974241

  2. Laminin isoforms differentially regulate adhesion, spreading, proliferation, and ERK activation of β1 integrin-null cells

    International Nuclear Information System (INIS)

    The presence of many laminin receptors of the β1 integrin family on most cells makes it difficult to define the biological functions of other major laminin receptors such as integrin α6β4 and dystroglycan. We therefore tested the binding of a β1 integrin-null cell line GD25 to four different laminin variants. The cells were shown to produce dystroglycan, which based on affinity chromatography bound to laminin-1, -2/4, and -10/11, but not to laminin-5. The cells also expressed the integrin α6Aβ4A variant. GD25 β1 integrin-null cells are known to bind poorly to laminin-1, but we demonstrate here that these cells bind avidly to laminin-2/4, -5, and -10/11. The initial binding at 20 min to each of these laminins could be inhibited by an integrin α6 antibody, but not by a dystroglycan antibody. Hence, integrin α6Aβ4A of GD25 cells was identified as a major receptor for initial GD25 cell adhesion to three out of four tested laminin isoforms. Remarkably, cell adhesion to laminin-5 failed to promote cell spreading, proliferation, and extracellular signal-regulated kinase (ERK) activation, whereas all these responses occurred in response to adhesion to laminin-2/4 or -10/11. The data establish GD25 cells as useful tools to define the role integrin α6Aβ4A and suggest that laminin isoforms have distinctly different capacities to promote cell adhesion and signaling via integrin α6Aβ4A

  3. Adipose Tissue-Derived Stromal Cells Inhibit TGF-beta 1-Induced Differentiation of Human Dermal Fibroblasts and Keloid Scar-Derived Fibroblasts in a Paracrine Fashion

    NARCIS (Netherlands)

    Spiekman, Maroesjka; Przybyt, Ewa; Plantinga, Josee A.; Gibbs, Susan; van der Lei, Berend; Harmsen, Martin C.

    2014-01-01

    Background: Adipose tissue-derived stromal cells augment wound healing and skin regeneration. It is unknown whether and how they can also influence dermal scarring. The authors hypothesized that adipose tissue-derived stromal cells inhibit adverse differentiation of dermal fibroblasts induced by the

  4. Integrin binding specificity regulates biomaterial surface chemistry effects on cell differentiation

    OpenAIRE

    Keselowsky, Benjamin G.; Collard, David M.; Andrés J. García

    2005-01-01

    Biomaterial surface chemistry has profound consequences on cellular and host responses, but the underlying molecular mechanisms remain poorly understood. Using self-assembled monolayers as model biomaterial surfaces presenting well defined chemistries, we demonstrate that surface chemistry modulates osteoblastic differentiation and matrix mineralization independently from alterations in cell proliferation. Surfaces were precoated with equal densities of fibronectin (FN), and surface chemistry...

  5. Mouse-induced pluripotent stem cells differentiate into odontoblast-like cells with induction of altered adhesive and migratory phenotype of integrin.

    Directory of Open Access Journals (Sweden)

    Nobuaki Ozeki

    Full Text Available Methods for differentiating induced pluripotent stem (iPS cells into odontoblasts generally require epithelial-mesenchymal interactions. Here, we sought to characterize the cells produced by a 'hanging drop' technique for differentiating mouse iPS cells into odontoblast-like cells that requires no such interaction. Cells were cultured by the hanging drop method on a collagen type-I (Col-I scaffold (CS combined with bone morphogenetic protein (BMP-4 (CS/BMP-4 without an epithelial-mesenchymal interaction. We evaluated the expression of odontoblast-related mRNA and protein, and the proliferation rate of these cells using reverse-transcription polymerase chain reaction, immunofluorescence staining, and BrdU cell proliferation enzyme-linked immunosorbent assay, respectively. The differentiated cells strongly expressed the mRNA for dentin sialophosphoprotein (DSPP and dentin matrix protein-1 (Dmp-1, which are markers of mature odontoblasts. Osteopontin and osteocalcin were not expressed in the differentiated cells, demonstrating that the differentiated iPS cells bore little resemblance to osteoblasts. Instead, they acquired odontoblast-specific properties, including the adoption of an odontoblastic phenotype, typified by high alkaline phosphatase (ALP activity and calcification capacity. The cell-surface expression of proteins such as integrins α2, α6, αV and αVβ3 was rapidly up-regulated. Interestingly, antibodies and siRNAs against integrin α2 suppressed the expression of DSPP and Dmp-1, reduced the activity of ALP and blocked calcification, suggesting that integrin α2 in iPS cells mediates their differentiation into odontoblast-like cells. The adhesion of these cells to fibronectin and Col-I, and their migration on these substrata, was significantly increased following differentiation into odontoblast-like cells. Thus, we have demonstrated that integrin α2 is involved in the differentiation of mouse iPS cells into odontoblast-like cells

  6. Differential Dynamics of α5 Integrin, Paxillin, and α-Actinin during Formation and Disassembly of Adhesions in Migrating Cells

    OpenAIRE

    Laukaitis, Christina M; Webb, Donna J.; Donais, Karen; Horwitz, Alan F.

    2001-01-01

    To investigate the mechanisms by which adhesions form and disperse in migrating cells, we expressed α5 integrin, α-actinin, and paxillin as green fluorescent protein (GFP) fusions. All localized with their endogenous counterparts and did not perturb migration when expressed at moderate levels. α5-GFP also rescued the adhesive defects in CHO B2 cells, which are α5 integrin deficient. In ruffling cells, α5-GFP and α-actinin–GFP localized prominently at the leading edge in membrane protrusions. ...

  7. Spermidine/spermine N-1-acetyltransferase specifically binds to the integrin alpha 9 subunit cytoplasmic domain and enhances cell migration

    OpenAIRE

    Chen, C.; Young, B A; Coleman, C S; Pegg, A E; Sheppard, D

    2004-01-01

    T he integrin alpha9beta1 is expressed on migrating cells, such as leukocytes, and binds to multiple ligands that are present at sites of tissue injury and inflammation. alpha9beta1, like the structurally related integrin alpha4beta1, mediates accelerated cell migration, an effect that depends on the beta cytoplasmic domain. alpha4beta1 enhances migration through reversible binding to the adapter protein, paxillin, but alpha9beta1-dependent migration is paxillin independent. Using yeast two-h...

  8. Absence of alphavbeta6 integrin is linked to initiation and progression of periodontal disease.

    Science.gov (United States)

    Ghannad, Farzin; Nica, Daniela; Fulle, Maria I Garcia; Grenier, Daniel; Putnins, Edward E; Johnston, Sarah; Eslami, Ameneh; Koivisto, Leeni; Jiang, Guoqiao; McKee, Marc D; Häkkinen, Lari; Larjava, Hannu

    2008-05-01

    Integrin alphavbeta6 is generally not expressed in adult epithelia but is induced in wound healing, cancer, and certain fibrotic disorders. Despite this generalized absence, we observed that alphavbeta6 integrin is constitutively expressed in the healthy junctional epithelium linking the gingiva to tooth enamel. Moreover, expression of alphavbeta6 integrin was down-regulated in human periodontal disease, a common medical condition causing tooth loss and also contributing to the development of cardiovascular diseases by increasing the total systemic inflammatory burden. Remarkably, integrin beta6 knockout mice developed classic signs of spontaneous, chronic periodontal disease with characteristic inflammation, epithelial down-growth, pocket formation, and bone loss around the teeth. Integrin alphavbeta6 acts as a major activator of transforming growth factor-beta1 (TGF-beta1), a key anti-inflammatory regulator in the immune system. Co-expression of TGF-beta1 and alphavbeta6 integrin was observed in the healthy junctional epithelium. Moreover, an antibody that blocks alphavbeta6 integrin-mediated activation of TGF-beta1 initiated inflammatory periodontal disease in a rat model of gingival inflammation. Thus, alphavbeta6 integrin is constitutively expressed in the epithelium sealing the gingiva to the tooth and plays a central role in protection against inflammatory periodontal disease through activation of TGF-beta1. PMID:18385522

  9. EDA-containing cellular fibronectin induces fibroblast differentiation through binding to alpha4beta7 integrin receptor and MAPK/Erk 1/2-dependent signaling.

    Science.gov (United States)

    Kohan, Martin; Muro, Andres F; White, Eric S; Berkman, Neville

    2010-11-01

    Fibroblast differentiation is an essential step during wound healing and fibrosis. Fibronectin (FN) is a major component of the extracellular matrix and occurs in two main forms: plasma and cellular FN. The latter includes the alternatively spliced domain A (EDA). Although EDA-containing cellular fibronectin (EDA-FN) is associated with fibroblast differentiation, how EDA-FN promotes differentiation is incompletely understood. In this study, we investigate the mechanism by which EDA-FN contributes to fibroblast differentiation with emphasis on the characterization of the EDA-FN receptor. We show that EDA-FN increases α-SMA expression (immunofluorescence), collagen deposition, cell contractility, and focal adhesion kinase (FAK) activation (immunoblotting); whereas plasma FN, a form lacking EDA, shows no effect. Primary lung fibroblasts constitutively express α(4)β(7) integrin receptor (FACS and RT-PCR). Blocking of α(4)β(7) reduces fibroblast adhesion to EDA-FN and inhibits α-SMA expression, collagen deposition, and FAK activation induced by EDA-FN. Using recombinant EDA-containing peptides, we demonstrate that the EDA segment is sufficient to induce fibroblast differentiation via binding to α(4)β(7). EDA-FN induces MAPK-Erk1/2 activation and inhibition of MEK1/2 attenuates EDA-FN-induced α-SMA expression. Our findings demonstrate that EDA-FN induces fibroblast differentiation by a mechanism that involves binding of EDA to α(4)β(7) integrin followed by activation of FAK and MAPK-associated signaling pathways. PMID:20643910

  10. Interferon Beta-1a Intramuscular Injection

    Science.gov (United States)

    Interferon beta-1a intramuscular injection is used to reduce the number of episodes of symptoms and slow ... and problems with vision, speech, and bladder control). Interferon beta-1a is in a class of medications ...

  11. Interferon Beta-1a Subcutaneous Injection

    Science.gov (United States)

    Interferon beta-1a subcutaneous injection is used to reduce episodes of symptoms and slow the development of ... and problems with vision, speech, and bladder control). Interferon beta-1a is in a class of medications ...

  12. Interferon Beta-1a Intramuscular Injection

    Science.gov (United States)

    Interferon beta-1a intramuscular injection is used to reduce the number of episodes of symptoms and slow the development of disability in patients ... Interferon beta-1a intramuscular injection comes as a powder in vials to be mixed into a solution for injection. Interferon beta-1a intramuscular ...

  13. Recombinant human laminin-10 (alpha5beta1gamma1). Production, purification, and migration-promoting activity on vascular endothelial cells.

    Science.gov (United States)

    Doi, Masayuki; Thyboll, Jill; Kortesmaa, Jarkko; Jansson, Katarina; Iivanainen, Antti; Parvardeh, Masomeh; Timpl, Rupert; Hedin, Ulf; Swedenborg, Jesper; Tryggvason, Karl

    2002-04-12

    The laminin (LN) family of large heterotrimeric extracellular matrix glycoproteins has multiple functions: LNs take part in the regulation of processes such as cell migration, differentiation, and proliferation, in addition to contributing to the structure of basement membranes. LN-10, composed of alpha5, beta1, and gamma1 chains, is widely distributed in most basement membranes of both epithelia and endothelia. We determined the complete human cDNA sequence for the LN alpha5 chain and produced recombinant human LN-10 (rLN-10) in HEK293 cells by triple transfection of full-length cDNAs encoding the human LN alpha5, beta1, and gamma1 chains. The rLN-10 was purified using affinity chromatography and had an apparent molecular mass of approximately 800 kDa in SDS-PAGE and a native domain structure in rotary shadowing electron microscopy. By using function-blocking monoclonal antibodies, integrin alpha(3)beta(1) was found to be a major mediator of adhesion of HT-1080 and human saphenous vein endothelial cells. Human saphenous vein endothelial cells adhered more strongly to rLN-10 than to LN-1 and LN-8 and showed better migration on rLN-10, compared with several other matrices. Considering the cell adhesive and migration-promoting properties of rLN-10 on endothelial cells, this molecule could be useful in improving the biocompatibility and endothelialization of vascular grafts. PMID:11821406

  14. Disintegrins: integrin selective ligands which activate integrin-coupled signaling and modulate leukocyte functions

    Directory of Open Access Journals (Sweden)

    Barja-Fidalgo C.

    2005-01-01

    Full Text Available Extracellular matrix proteins and cell adhesion receptors (integrins play essential roles in the regulation of cell adhesion and migration. Interactions of integrins with the extracellular matrix proteins lead to phosphorylation of several intracellular proteins such as focal adhesion kinase, activating different signaling pathways responsible for the regulation of a variety of cell functions, including cytoskeleton mobilization. Once leukocytes are guided to sites of infection, inflammation, or antigen presentation, integrins can participate in the initiation, maintenance, or termination of the immune and inflammatory responses. The modulation of neutrophil activation through integrin-mediated pathways is important in the homeostatic control of the resolution of inflammatory states. In addition, during recirculation, T lymphocyte movement through distinct microenvironments is mediated by integrins, which are critical for cell cycle, differentiation and gene expression. Disintegrins are a family of low-molecular weight, cysteine-rich peptides first identified in snake venom, usually containing an RGD (Arg-Gly-Asp motif, which confers the ability to selectively bind to integrins, inhibiting integrin-related functions in different cell systems. In this review we show that, depending on the cell type and the microenvironment, disintegrins are able to antagonize the effects of integrins or to act agonistically by activating integrin-mediated signaling. Disintegrins have proven useful as tools to improve the understanding of the molecular events regulated by integrin signaling in leukocytes and prototypes in order to design therapies able to interfere with integrin-mediated effects.

  15. Alpha1 and Alpha2 Integrins Mediate Invasive Activity of Mouse Mammary Carcinoma Cells through Regulation of Stromelysin-1 Expression

    Energy Technology Data Exchange (ETDEWEB)

    Lochter, Andre; Navre, Marc; Werb, Zena; Bissell, Mina J

    1998-06-29

    Tumor cell invasion relies on cell migration and extracellular matrix proteolysis. We investigated the contribution of different integrins to the invasive activity of mouse mammary carcinoma cells. Antibodies against integrin subunits {alpha}6 and {beta}1, but not against {alpha}1 and {alpha}2, inhibited cell locomotion on a reconstituted basement membrane in two-dimensional cell migration assays, whereas antibodies against {beta}1, but not against a6 or {alpha}2, interfered with cell adhesion to basement membrane constituents. Blocking antibodies against {alpha}1 integrins impaired only cell adhesion to type IV collagen. Antibodies against {alpha}1, {alpha}2, {alpha}6, and {beta}1, but not {alpha}5, integrin subunits reduced invasion of a reconstituted basement membrane. Integrins {alpha}1 and {alpha}2, which contributed only marginally to motility and adhesion, regulated proteinase production. Antibodies against {alpha}1 and {alpha}2, but not {alpha}6 and {beta}1, integrin subunits inhibited both transcription and protein expression of the matrix metalloproteinase stromelysin-1. Inhibition of tumor cell invasion by antibodies against {alpha}1 and {alpha}2 was reversed by addition of recombinant stromelysin-1. In contrast, stromelysin-1 could not rescue invasion inhibited by anti-{alpha}6 antibodies. Our data indicate that {alpha}1 and {alpha}2 integrins confer invasive behavior by regulating stromelysin-1 expression, whereas {alpha}6 integrins regulate cell motility. These results provide new insights into the specific functions of integrins during tumor cell invasion.

  16. [Transforming growth factor beta-1: structure, function, and regulation mechanisms in cancer].

    Science.gov (United States)

    Peralta-Zaragoza, O; Lagunas-Martínez, A; Madrid-Marina, V

    2001-01-01

    Transforming growth factor beta-1 (TGF-beta 1) is produced by several cell lineages such as lymphocytes, macrophages, and dendritic cells, and its expression serves in both autocrine and paracrine modes to control the differentiation, proliferation, and state of activation of these and other cells. In general, TGF-beta 1 has pleiotropic properties on the immune response during the development of infection diseases and cancer; however, the mechanisms of action and regulation of gene expression of this cytokine are poorly understood, in this review, the biological properties and the molecular mechanisms that regulate TGF-beta 1 gene expression are described, to understand the role of this cytokine in growth and cell differentiation. The knowledge of molecular mechanisms of gene expression of TGF-beta 1 may serve to develop new cancer therapies. The English version of this paper is available at: http://www.insp.mx/salud/index.html PMID:11547595

  17. Intestinal epithelial restitution. Involvement of specific laminin isoforms and integrin laminin receptors in wound closure of a transformed model epithelium

    DEFF Research Database (Denmark)

    Lotz, M M; Nusrat, A; Madara, J L;

    1997-01-01

    cells adjoining wounds. Because T84 cells stained faintly with MAbs 4C7 (laminin alpha 1 subunit) and with MAbs 4F11 and 1B4 (laminin alpha 2 subunit), we suggest that expression of laminins 6 and 7 is enhanced in response to wounding. The alpha 3 beta 1 integrin and the alpha 6-containing integrins...

  18. INDUCCIÓN DIFERENCIAL DE POLIFENOLOXIDASA Y beta-1,3-GLUCANASA EN CLAVEL (Dianthus caryophyllus DURANTE LA INFECCIÓN POR Fusarium oxysporum f. sp. dianthi RAZA 2 Polyphenoloxidase and beta-1,3-Glucanase Differential Induction in Carnation (Dianthus caryophyllus Infectedby Fusarium oxysporum f. sp. dianthi race 2

    Directory of Open Access Journals (Sweden)

    BLANCA LIGIA HIGUERA

    Full Text Available Se evaluó el cambio en el comportamiento de las enzimas polifenoloxidasa (PFO y beta-1,3-glucanasa (Glu en tallos de plantas de clavel inoculadas con el patógeno Fusarium oxysporum f. sp. dianthi (Fod raza 2, con el fin de determinar su posible participación en la respuesta defensiva y en la resistencia de la planta al marchitamiento vascular. Se evaluaron parámetros para la extracción y determinación de actividad de dichas enzimas. Las condiciones que proporcionaron los mejores resultados de extracción fueron: obtención de polvos de acetona previa al tratamiento con buffer de fosfatos pH 6,5 con 3% de PVPP para PFO y con buffer de fosfatos pH 6,5 para Glu. La cuantificación de PFO se llevó a cabo usando catecol a pH 7,0 y 37 ºC y midiendo los productos de reacción a 420 nm, y la de Glu usando laminarina digitata a 37 ºC y pH 5,5. Una vez establecidos los métodos, esquejes de clavel de una variedad altamente tolerante (Carolina y de una susceptible (Uconn fueron inoculados con el patógeno y sometidos al análisis de las enzimas a diferentes tiempos post-inoculación. Mientras que en la variedad susceptible la actividad PFO no se vio afectada, en la tolerante se presentó una importante y significativa inducción de esta enzima a las 12 h y 24 h, indicando que puede desempeñar un papel clave en la defensa de la planta, en fenómenos metabólicos probablemente relacionados con lignificación y síntesis de fenólicos. La enzima Glu presentó inducción en ambas variedades, aunque a diferentes tiempos, lo cual hace parte de una respuesta metabólica inespecífica, no relacionada con mecanismos de defensa activa del clavel contra el patógeno causal del marchitamiento vascular.We evaluated the changes on the dynamics of polyphenoloxidase (PFO and beta-1,3-glucanase (Glu enzymes in carnation steems, which had been previously inoculated with the pathogen Fusarium oxysporum f. sp. dianthi race 2 (Fod. We established the experimental

  19. Monocytic cells synthesize, adhere to, and migrate on laminin-8 (alpha 4 beta 1 gamma 1).

    Science.gov (United States)

    Pedraza, C; Geberhiwot, T; Ingerpuu, S; Assefa, D; Wondimu, Z; Kortesmaa, J; Tryggvason, K; Virtanen, I; Patarroyo, M

    2000-11-15

    Laminins, a growing family of large heterotrimeric proteins with cell adhesive and signaling properties, are major components of vascular and other basement membranes. Expression, recognition, and use of laminin isoforms by leukocytes are poorly understood. In monoblastic THP-1 cells, transcripts for laminin gamma(1)-, beta(1)-, and alpha(4)-chains were detected by RT-PCR. Following immunoaffinity purification on a laminin beta(1) Ab-Sepharose column, laminin beta(1)- (220 kDa), gamma(1)- (200 kDa), and alpha(4)- (180/200 kDa) chains were detected by Western blotting in THP-1 cells and in two other monoblastic cell lines, U-937 and Mono Mac 6. After cell permeabilization, a mAb to laminin gamma(1)-chain reacted with practically all blood monocytes by immunofluorescence flow cytometry, and laminin-8 (alpha(4)beta(1)gamma(1)) could be isolated also from these cells. Monoblastic JOSK-I cells adhered constitutively to immobilized recombinant laminin-8, less than to laminin-10/11 (alpha(5)beta(1)gamma(1)/alpha(5)beta(2)gamma(1)) but to a higher level than to laminin-1 (alpha(1)beta(1)gamma(1)). Compared with the other laminin isoforms, adhesion to laminin-8 was preferentially mediated by alpha(6)beta(1) and beta(2) integrins. Laminin-8 and, to a lower extent, laminin-1 promoted spontaneous and chemokine-induced migration of blood monocytes, whereas laminin-10/11 was inhibitory. Altogether, the results indicate that leukocytes, as other cell types, are able to synthesize complete laminin molecules. Expression, recognition, and use of laminin-8 by leukocytes suggest a major role of this laminin isoform in leukocyte physiology. PMID:11067943

  20. Alterations in integrin expression modulates invasion of pancreatic cancer cells.

    LENUS (Irish Health Repository)

    Walsh, Naomi

    2009-01-01

    BACKGROUND: Factors mediating the invasion of pancreatic cancer cells through the extracellular matrix (ECM) are not fully understood. METHODS: In this study, sub-populations of the human pancreatic cancer cell line, MiaPaCa-2 were established which displayed differences in invasion, adhesion, anoikis, anchorage-independent growth and integrin expression. RESULTS: Clone #3 displayed higher invasion with less adhesion, while Clone #8 was less invasive with increased adhesion to ECM proteins compared to MiaPaCa-2. Clone #8 was more sensitive to anoikis than Clone #3 and MiaPaCa-2, and displayed low colony-forming efficiency in an anchorage-independent growth assay. Integrins beta 1, alpha 5 and alpha 6 were over-expressed in Clone #8. Using small interfering RNA (siRNA), integrin beta1 knockdown in Clone #8 cells increased invasion through matrigel and fibronectin, increased motility, decreased adhesion and anoikis. Integrin alpha 5 and alpha 6 knockdown also resulted in increased motility, invasion through matrigel and decreased adhesion. CONCLUSION: Our results suggest that altered expression of integrins interacting with different extracellular matrixes may play a significant role in suppressing the aggressive invasive phenotype. Analysis of these clonal populations of MiaPaCa-2 provides a model for investigations into the invasive properties of pancreatic carcinoma.

  1. Human liver alcohol dehydrogenase. 1. The primary structure of the beta 1 beta 1 isoenzyme.

    Science.gov (United States)

    Hempel, J; Bühler, R; Kaiser, R; Holmquist, B; de Zalenski, C; von Wartburg, J P; Vallee, B; Jörnvall, H

    1984-12-17

    Determination of the amino acid sequence of the beta 1 subunit from the class I (pyrazole-sensitive) human liver alcohol dehydrogenase isoenzyme beta 1 beta 1 revealed a 373-residue structure differing at 48 positions (including a gap) from that of the subunit of the well studied horse liver alcohol dehydrogenase EE isoenzyme. The structure deduced is compatible with known differences in composition, ultraviolet absorbance, electrophoretic mobility and catalytic properties between the horse and human enzymes. All zinc-liganding residues of the horse E subunit are strictly conserved in the human beta 1 subunit, despite an earlier report of a mutation involving Cys-46. This residue therefore remains conserved in all known alcohol dehydrogenase structures. However, the total cysteine content of the beta 1 structure is raised from 14 in the subunit of the horse enzyme to 15 by a Tyr----Cys exchange. Most exchanges are on the surface of the molecule and of a well conserved nature. Substitutions close to the catalytic centre are of interest to explain the altered substrate specificity and different catalytic activity of the beta 1 homodimer. Functionally, a Ser----Thr exchange at position 48 appears to be of special importance, since Thr-48 in beta 1 instead of Ser-48 in the horse enzyme can restrict available space. Four other substitutions also line the active-site pocket, and appear to constitute partly compensated exchanges. PMID:6391920

  2. IGF-I and TGF-beta1 have distinct effects on phenotype and proliferation of intestinal fibroblasts.

    Science.gov (United States)

    Simmons, James G; Pucilowska, Jolanta B; Keku, Temitope O; Lund, P Kay

    2002-09-01

    Insulin-like growth factor I (IGF-I) and transforming growth factor-beta1 (TGF-beta1) are upregulated in myofibroblasts at sites of fibrosis in experimental enterocolitis and in Crohn's disease (CD). We compared the sites of expression of IGF-I and TGF-beta1 in a rat peptidoglycan-polysaccharide (PG-PS) model of chronic granulomatous enterocolitis and fibrosis. We used the human colonic CCD-18Co fibroblast/myofibroblast cell line to test the hypothesis that TGF-beta1 and IGF-I interact to regulate proliferation, collagen synthesis, and activated phenotype typified by expression of alpha-smooth muscle actin and organization into stress fibers. IGF-I potently stimulated while TGF-beta1 inhibited basal DNA synthesis. TGF-beta1 and IGF-I each had similar but not additive effects to induce type I collagen. TGF-beta1 but not IGF-I potently stimulated expression of alpha-smooth muscle actin and stress fiber formation. IGF-I in combination with TGF-beta1 attenuated stress fiber formation without reducing alpha-smooth muscle actin expression. Stress fibers were not a prerequisite for increased collagen synthesis. TGF-beta1 upregulated IGF-I mRNA, which led us to examine the effects of IGF-I in cells previously activated by TGF-beta1 pretreatment. IGF-I potently stimulated proliferation of TGF-beta1-activated myofibroblasts without reversing activated fibrogenic phenotype. We conclude that TGF-beta1 and IGF-I both stimulate type I collagen synthesis but have differential effects on activated phenotype and proliferation. We propose that during intestinal inflammation, regulation of activated phenotype and proliferation may require sequential actions of TGF-beta1 and IGF-I, but they may act in concert to increase collagen deposition. PMID:12181198

  3. TGF-beta1 release from biodegradable polymer microparticles: its effects on marrow stromal osteoblast function

    Science.gov (United States)

    Lu, L.; Yaszemski, M. J.; Mikos, A. G.; McIntire, L. V. (Principal Investigator)

    2001-01-01

    BACKGROUND: Controlled release of transforming growth factor-beta1 (TGF-beta1) to a bone defect may be beneficial for the induction of a bone regeneration cascade. The objectives of this work were to assess the feasibility of using biodegradable polymer microparticles as carriers for controlled TGF-beta1 delivery and the effects of released TGF-beta1 on the proliferation and differentiation of marrow stromal cells in vitro. METHODS: Recombinant human TGF-beta1 was incorporated into microparticles of blends of poly(DL-lactic-co-glycolic acid) (PLGA) and poly(ethylene glycol) (PEG). Fluorescein isothiocynate-labeled bovine serum albumin (FITC-BSA) was co-encapsulated as a porogen. The effects of PEG content (0, 1, or 5% by weight [wt%]) and buffer pH (3, 5, or 7.4) on the protein release kinetics and the degradation of PLGA were determined in vitro for as long as 28 days. Rat marrow stromal cells were seeded on a biodegradable poly(propylene fumarate) (PPF) substrate. The dose response and biological activity of released TGF-beta1 was determined after 3 days in culture. The effects of TGF-beta1 released from PLGA/PEG microparticles on marrow stromal cell proliferation and osteoblastic differentiation were assessed during a 21-day period. RESULTS: TGF-beta1 was encapsulated along with FITC-BSA into PLGA/PEG blend microparticles and released in a multiphasic fashion including an initial burst for as long as 28 days in vitro. Increasing the initial PEG content resulted in a decreased cumulative mass of released proteins. Aggregation of FITC-BSA occurred at lower buffer pH, which led to decreased release rates of both proteins. The degradation of PLGA was increased at higher PEG content and significantly accelerated at acidic pH conditions. Rat marrow stromal cells cultured on PPF substrates showed a dose response to TGF-beta1 released from the microparticles similar to that of added TGF-beta1, indicating that the activity of TGF-beta1 was retained during microparticle

  4. Integrin Trafficking and Tumor Progression

    Directory of Open Access Journals (Sweden)

    Sejeong Shin

    2012-01-01

    Full Text Available Integrins are major mediators of cancer cell adhesion to extracellular matrix. Through this interaction, integrins play critical roles in cell migration, invasion, metastasis, and resistance to apoptosis during tumor progression. Recent studies highlight the importance of integrin trafficking, endocytosis and recycling, for the functions of integrins in cancer cells. Understanding the molecular mechanisms of integrin trafficking is pivotal for understanding tumor progression and for the development of anticancer drugs.

  5. Breast Cancer Cells in Three-dimensional Culture Display an Enhanced Radioresponse after Coordinate Targeting of Integrin ?5?1 and Fibronectin

    Energy Technology Data Exchange (ETDEWEB)

    Nam, Jin-Min; Onodera, Yasuhito; Bissell, Mina J; Park, Catherine C

    2010-04-07

    Tactics to selectively enhance cancer radioresponse are of great interest. Cancer cells actively elaborate and remodel their extracellular matrix (ECM) to aid in survival and progression. Previous work has shown that {beta}1-integrin inhibitory antibodies can enhance the growth-inhibitory and apoptotic responses of human breast cancer cell lines to ionizing radiation, either when cells are cultured in three-dimensional laminin-rich ECM (3D lrECM) or grown as xenografts in mice. Here, we show that a specific {alpha} heterodimer of {beta}1-integrin preferentially mediates a prosurvival signal in human breast cancer cells that can be specifically targeted for therapy. 3D lrECM culture conditions were used to compare {alpha}-integrin heterodimer expression in malignant and nonmalignant cell lines. Under these conditions, we found that expression of {alpha}5{beta}1-integrin was upregulated in malignant cells compared with nonmalignant breast cells. Similarly, we found that normal and oncofetal splice variants of fibronectin, the primary ECM ligand of {alpha}5{beta}1-integrin, were also strikingly upregulated in malignant cell lines compared with nonmalignant acini. Cell treatment with a peptide that disrupts the interactions of {alpha}5{beta}1-integrin with fibronectin promoted apoptosis in malignant cells and further heightened the apoptotic effects of radiation. In support of these results, an analysis of gene expression array data from breast cancer patients revealed an association of high levels of {alpha}5-integrin expression with decreased survival. Our findings offer preclinical validation of fibronectin and {alpha}5{beta}1-integrin as targets for breast cancer therapy.

  6. Covisualization by computational optical-sectioning microscopy of integrin and associated proteins at the cell membrane of living onion protoplasts

    Science.gov (United States)

    Gens, J. S.; Reuzeau, C.; Doolittle, K. W.; McNally, J. G.; Pickard, B. G.; Evans, M. L. (Principal Investigator)

    1996-01-01

    Using higher-resolution wide-field computational optical-sectioning fluorescence microscopy, the distribution of antigens recognized by antibodies against animal beta 1 integrin, fibronectin, and vitronectin has been visualized at the outer surface of enzymatically protoplasted onion epidermis cells and in depectinated cell wall fragments. On the protoplast all three antigens are colocalized in an array of small spots, as seen in raw images, in Gaussian filtered images, and in images restored by two different algorithms. Fibronectin and vitronectin but not beta 1 integrin antigenicities colocalize as puncta in comparably prepared and processed images of the wall fragments. Several control visualizations suggest considerable specifity of antibody recognition. Affinity purification of onion cell extract with the same anti-integrin used for visualization has yielded protein that separates in SDS-PAGE into two bands of about 105-110 and 115-125 kDa. These bands are again recognized by the visualization antibody, which was raised against the extracellular domain of chicken beta 1 integrin, and are also recognized by an antibody against the intracellular domain of chicken beta 1 integrin. Because beta 1 integrin is a key protein in numerous animal adhesion sites, it appears that the punctate distribution of this protein in the cell membranes of onion epidermis represents the adhesion sites long known to occur in cells of this tissue. Because vitronectin and fibronection are matrix proteins that bind to integrin in animals, the punctate occurrence of antigenically similar proteins both in the wall (matrix) and on enzymatically prepared protoplasts reinforces the concept that onion cells have adhesion sites with some similarity to certain kinds of adhesion sites in animals.

  7. Is Serum Transforming Growth Factor beta-1 Superior to Serum Creatinine for assessing Renal Failure and Renal Transplant Rejection

    Directory of Open Access Journals (Sweden)

    Gyanendra Kumar Sonkar, Usha

    2009-04-01

    Full Text Available A sustained overexpression of Transforming Growth Factor beta1 (TGF beta1, a cytokine has beenimplicated in the pathogenesis of fibrosis of kidney leading to end stage . The main aim of present studywas to find the utility of TGF beta1 and serum creatinine in differentiating chronic renal failure (CRFfrom acute renal failure (ARF, renal transplant rejection (Tx Rej and stable renal transplant (Tx Stband to study has attempted histopathological correlation of rejection cases with TGF beta1 and serumcreatinine. TGF beta1 was determined by using ELISA and serum creatinine was done by autoanalyser.In normal healthy controls (NHC, in majority of cases (80.0% TGF beta1 was below 25 ng/ml while in6.0% cases it was upto 34 ng/ml. Rise of TGF beta1 was significant in CRF patients as compared to ARFand NHC (p<0.05 .In rejection cases, TGF beta1 level was significantly raised as compared to NHCand stable graft cases (p<0.05. In rejection cases, it was raised above 40 ng/ml in only 50% cases. In twocases inspite of more than 70% glomerular fibrosis, the patient had TGF beta1 level of only 5 ng/ml and inother three cases of acute cellular rejection the level was 70, 35 and 28 ng/ml respectively.Contrary to itserum creatinine was raised above 2 mg/dl in all cases of transplant rejection but in stable transplant casesin majority (70.6% it was below 1.5 mg/dl and in 5 cases it was between 1.5 – 1.9 mg/dl.Thus the studysuggests that TGF beta1 may not be a good marker for chronic transplant rejection, as it does notcorrelate well with glomerular fibrosis, probably it is more associated with interstitial inflammation but itcan differentiate CRF from ARF if cut off of 40 ng/ml is taken.

  8. Integrin-like proteins are localized to plasma membrane fractions, not plastids, in Arabidopsis

    Science.gov (United States)

    Swatzell, L. J.; Edelmann, R. E.; Makaroff, C. A.; Kiss, J. Z.

    1999-01-01

    Integrins are a large family of integral membrane proteins that function in signal transduction in animal systems. These proteins are conserved in vertebrates, invertebrates, and fungi. Evidence from previous research suggests that integrin-like proteins may be present in plants as well, and that these proteins may function in signal transduction during gravitropism. In past studies, researchers have used monoclonal and polyclonal antibodies to localize beta 1 integrin-like proteins in plants. However, there is a disparity between data collected from these studies, especially since molecular weights obtained from these investigations range from 55-120 kDa for integrin-like proteins. To date, a complete investigation which employs all three basic immunolabeling procedures, immunoblotting, immunofluorescence microscopy, and immunogold labeling, in addition to extensive fractionation and exhaustive controls, has been lacking. In this paper, we demonstrate that use of a polyclonal antibody against the cytoplasmic domain of avian beta 1-integrin can produce potential artifacts in immunolocalization studies. However, these problems can be eliminated through use of starchless mutants or proper specimen preparation prior to electrophoresis. We also show that this antibody, when applied within the described parameters and with careful controls, identifies a large (100 kDa) integrin-like protein that is localized to plasma membrane fractions in Arabidopsis.

  9. Chondroitin sulphate modification in the alpha4 chain of human recombinant laminin-8 (alpha4beta1gamma1).

    Science.gov (United States)

    Kortesmaa, Jarkko; Doi, Masayuki; Patarroyo, Manuel; Tryggvason, Karl

    2002-10-01

    We have produced human laminin-8 (alpha4beta1gamma1) using recombinant technology. Approximately half of the recombinant laminin-8 (rLN-8) molecules were found to have a chondroitin sulphate modification in the alpha4 chain. The substituted and non-substituted forms were separated and tested for cell adhesion activity. Lower cell adhesion promoting activity was seen for the substituted form, but the integrin receptor utilization was similar. We also found the human rLN-8 to behave identically in cell adhesion assays compared to a human/mouse hybrid variant of rLN-8. PMID:12392759

  10. An endothelial laminin isoform, laminin 8 (alpha4beta1gamma1), is secreted by blood neutrophils, promotes neutrophil migration and extravasation, and protects neutrophils from apoptosis.

    Science.gov (United States)

    Wondimu, Zenebech; Geberhiwot, Tarekegn; Ingerpuu, Sulev; Juronen, Erkki; Xie, Xun; Lindbom, Lennart; Doi, Masayuki; Kortesmaa, Jarkko; Thyboll, Jill; Tryggvason, Karl; Fadeel, Bengt; Patarroyo, Manuel

    2004-09-15

    During extravasation, neutrophils migrate through the perivascular basement membrane (BM), a specialized extracellular matrix rich in laminins. Laminins 8 (LN-8) (alpha4beta1gamma1) and 10 (LN-10) (alpha5beta1gamma1) are major components of the endothelial BM, but expression, recognition, and use of these laminin isoforms by neutrophils are poorly understood. In the present study, we provide evidence, using a panel of novel monoclonal antibodies against human laminin alpha4 (LNalpha4) chain, that neutrophils contain and secrete LN-8, and that this endogenous laminin contributes to chemoattractant-induced, alphaMbeta2-integrin-dependent neutrophil migration through albumin-coated filters. Phorbol ester-stimulated neutrophils adhered to recombinant human (rh) LN-8, rhLN-10, and mouse LN-1 (mLN-1) (alpha1beta1gamma1) via alphaMbeta2-integrin, and these laminin isoforms strongly promoted chemoattractant-induced neutrophil migration via the same integrin. However, only rhLN-8 enhanced the spontaneous migration. In addition, recruitment of neutrophils into the peritoneum following an inflammatory stimulus was impaired in LNalpha4-deficient mice. rhLN-8 also protected isolated neutrophils from spontaneous apoptosis. This study is the first to identify a specific laminin isoform in neutrophils and provides evidence for the role of LN-8 in the adhesion, migration, extravasation, and survival of these cells. PMID:15172971

  11. Pulmonary administration of integrin-nanoparticles regenerates collapsed alveoli.

    Science.gov (United States)

    Horiguchi, Michiko; Kojima, Hisako; Sakai, Hitomi; Kubo, Hiroshi; Yamashita, Chikamasa

    2014-08-10

    Chronic obstructive pulmonary disease (COPD) is an intractable pulmonary disease, causes widespread and irreversible alveoli collapse. In search of a treatment target molecule, which is able to regenerate collapsed alveoli, we sought to identify a factor that induces differentiation in human alveolar epithelial stem cells using all-trans retinoic acid (ATRA), whose alveolar repair capacity has been reported in animal experiments. When human alveolar epithelial stem cells were exposed to ATRA at a concentration of 10μM for over seven days, approximately 20% of the cells differentiated into each of the type-I and type-II alveolar epithelial cells that constitute the alveoli. In a microarray analysis, integrin-α1 and integrin-β3 showed the largest variation in the ATRA-treated group compared with the controls. Furthermore, the effect of the induction of differentiation in human alveolar epithelial stem cells using ATRA was suppressed by approximately one-fourth by siRNA treatments with integrin α1 and integrin β3. These results suggested that integrin α1 and β3 are factors responsible for the induction of differentiation in human alveolar epithelial stem cells. We accordingly investigated whether integrin nanoparticles also had a regenerative effect in vivo. Elastase-induced COPD model mouse was produced, and the alveolar repair effect of pulmonary administration using nanoparticles of integrin protein was evaluated by X-ray CT scanning. Improvement in the CT value in comparison with an untreated group indicated that there was an alveolar repair effect. In this study, it was shown that the differentiation-inducing effect on human alveolar epithelial stem cells by ATRA was induced by increased expression of integrin, and that the induced integrin enhanced phosphorylation signaling of AKT, resulting in inducing differentiations. Furthermore, the study demonstrated that lung administration of nanoparticles with increased solubility and stability of integrin

  12. Functional analysis of the cytoplasmic domain of the integrin {alpha}1 subunit in endothelial cells.

    Science.gov (United States)

    Abair, Tristin D; Bulus, Nada; Borza, Corina; Sundaramoorthy, Munirathinam; Zent, Roy; Pozzi, Ambra

    2008-10-15

    Integrin alpha1beta1, the major collagen type IV receptor, is expressed by endothelial cells and plays a role in both physiologic and pathologic angiogenesis. Because the molecular mechanisms whereby this collagen IV receptor mediates endothelial cell functions are poorly understood, truncation and point mutants of the integrin alpha1 subunit cytoplasmic tail (amino acids 1137-1151) were generated and expressed into alpha1-null endothelial cells. We show that alpha1-null endothelial cells expressing the alpha1 subunit, which lacks the entire cytoplasmic tail (mutant alpha1-1136) or expresses all the amino acids up to the highly conserved GFFKR motif (mutant alpha1-1143), have a similar phenotype to parental alpha1-null cells. Pro(1144) and Leu(1145) were shown to be necessary for alpha1beta1-mediated endothelial cell proliferation; Lys(1146) for adhesion, migration, and tubulogenesis and Lys(1147) for tubulogenesis. Integrin alpha1beta1-dependent endothelial cell proliferation is primarily mediated by ERK activation, whereas migration and tubulogenesis require both p38 MAPK and PI3K/Akt activation. Thus, distinct amino acids distal to the GFFKR motif of the alpha1 integrin cytoplasmic tail mediate activation of selective downstream signaling pathways and specific endothelial cell functions. PMID:18647959

  13. Integrin activation state determines selectivity for novel recognition sites in fibrillar collagens.

    Science.gov (United States)

    Siljander, Pia R-M; Hamaia, Samir; Peachey, Anthony R; Slatter, David A; Smethurst, Peter A; Ouwehand, Willem H; Knight, C Graham; Farndale, Richard W

    2004-11-12

    Only three recognition motifs, GFOGER, GLOGER, and GASGER, all present in type I collagen, have been identified to date for collagen-binding integrins, such as alpha(2)beta(1). Sequence alignment was used to investigate the occurrence of related motifs in other human fibrillar collagens, and located a conserved array of novel GER motifs within their triple helical domains. We compared the integrin binding properties of synthetic triple helical peptides containing examples of such sequences (GLSGER, GMOGER, GAOGER, and GQRGER) or the previously identified motifs. Recombinant inserted (I) domains of integrin subunits alpha(1), alpha(2) and alpha(11) all bound poorly to all motifs other than GFOGER and GLOGER. Similarly, alpha(2)beta(1) -containing resting platelets adhered well only to GFOGER and GLOGER, while ADP-activated platelets, HT1080 cells and two active alpha(2)I domain mutants (E318W, locked open) bound all motifs well, indicating that affinity modulation determines the sequence selectivity of integrins. GxO/SGER peptides inhibited platelet adhesion to collagen monomers with order of potency F >/= L >/= M > A. These results establish GFOGER as a high affinity sequence, which can interact with the alpha(2)I domain in the absence of activation and suggest that integrin reactivity of collagens may be predicted from their GER content. PMID:15345717

  14. Multiple integrin-ligand interactions synergize in shear-resistant platelet adhesion at sites of arterial injury in vivo

    DEFF Research Database (Denmark)

    Grüner, Sabine; Prostredna, Miroslava; Schulte, Valerie;

    2003-01-01

    intravital fluorescence microscopy that platelet adhesion and thrombus growth on the exposed ECM of the injured carotid artery is not significantly altered in alpha2-null mice and even in mice with a Cre/loxP-mediated loss of all beta1 integrins on their platelets. In contrast, inhibition of alphaIIbbeta3...

  15. Low-shear modelled microgravity environment maintains morphology and differentiated functionality of primary porcine hepatocyte cultures.

    Science.gov (United States)

    Nelson, Leonard J; Walker, Simon W; Hayes, Peter C; Plevris, John N

    2010-01-01

    Hepatocytes cultured in conventional static culture rapidly lose polarity and differentiated function. This could be explained by gravity-induced sedimentation, which prevents formation of complete three-dimensional (3D) cell-cell/cell-matrix interactions and disrupts integrin-mediated signals (including the most abundant hepatic integrin alpha(5)beta(1)), important for cellular polarity and differentiation. Cell culture in a low fluid shear modelled microgravity (about 10(-2) g) environment promotes spatial colocation/self-aggregation of dissociated cells and induction of 3D differentiated liver morphology. Previously, we demonstrated the utility of a NASA rotary bioreactor in maintaining key metabolic functions and 3D aggregate formation of high-density primary porcine hepatocyte cultures over 21 days. Using serum-free chemically defined medium, without confounding interactions of exogenous bioscaffolding or bioenhancing surface materials, we investigated features of hepatic cellular polarity and differentiated functionality, including expression of hepatic integrin alpha(5), as markers of functional morphology. We report here that in the absence of exogenous biomatrix scaffolding, hepatocytes cultured in serum-free chemically defined medium in a microgravity environment rapidly (microgravity environment may promote tissue-like self-organization of dissociated cells, and offer advantages over spheroids cultured in conventional formats to delineate optimal conditions for enhanced directed tissue self-assembly. PMID:20395654

  16. Elicitor and resistance-inducing activities of beta-1,4 cellodextrins in grapevine, comparison with beta-1,3 glucans and alpha-1,4 oligogalacturonides.

    Science.gov (United States)

    Aziz, Aziz; Gauthier, Adrien; Bézier, Annie; Poinssot, Benoît; Joubert, Jean-Marie; Pugin, Alain; Heyraud, Alain; Baillieul, Fabienne

    2007-01-01

    Cellodextrins (CD), water-soluble derivatives of cellulose composed of beta-1,4 glucoside residues, have been shown to induce a variety of defence responses in grapevine (Vitis vinifera L.) cells. The larger oligomers of CD rapidly induced transient generation of H2O2 and elevation in free cytosolic calcium, followed by a differential expression of genes encoding key enzymes of the phenylpropanoid pathway and pathogenesis-related (PR) proteins as well as stimulation of chitinase and beta-1,3 glucanase activities. Most of these defence reactions were also induced by linear beta-1,3 glucans (betaGlu) and alpha-1,4 oligogalacturonides (OGA) of different degree of polymerization (DP), but the intensity of some reactions induced by CD was different when compared with betaGlu and OGA effects. Moreover, desensitization assays using H2O2 production showed that cells treated with CD remained fully responsive to a second application of OGA, suggesting a different mode of perception of these oligosaccharides by grape cells. None of CD, betaGlu, or OGA induced HSR gene expression nor did they induce cell death. In accordance with elicitor activity in grapevine cells, CD-incubated leaves challenged with Botrytis cinerea also resulted in a significant reduction of the disease. Data suggest that CD could operate via other distinct reaction pathways than betaGlu and OGA. They also highlight the requirement of a specific DP for each oligosaccharide to induce the defence response. PMID:17322548

  17. Identification of novel inhibitors of the transforming growth factor beta1 (TGF-beta1) type 1 receptor (ALK5).

    Science.gov (United States)

    Callahan, James F; Burgess, Joelle L; Fornwald, James A; Gaster, Laramie M; Harling, John D; Harrington, Frank P; Heer, Jag; Kwon, Chet; Lehr, Ruth; Mathur, A; Olson, Barbara A; Weinstock, Joseph; Laping, Nicholas J

    2002-02-28

    Screening of our internal compound collection for inhibitors of the transforming growth factor beta1 (TGF-beta1) type I receptor (ALK5) identified several hits. Optimization of the dihydropyrroloimidazole hit 2 by introduction of a 2-pyridine and 3,4-methylenedioxyphenyl group gave 7, a selective ALK5 inhibitor. With this information, optimization of the triarylimidazole hit 8 gave the selective inhibitor 14, which inhibits TGF-beta1-induced fibronectin mRNA formation while displaying no measurable cytotoxicity in the 48 h XTT assay. PMID:11855979

  18. Laminin-8 (alpha4beta1gamma1) is synthesized by lymphoid cells, promotes lymphocyte migration and costimulates T cell proliferation.

    Science.gov (United States)

    Geberhiwot, T; Assefa, D; Kortesmaa, J; Ingerpuu, S; Pedraza, C; Wondimu, Z; Charo, J; Kiessling, R; Virtanen, I; Tryggvason, K; Patarroyo, M

    2001-01-01

    Laminins are a growing family of large heterotrimeric proteins with cell adhesive and signalling functions. They are major components of basement membranes and are found in many organs, including the vasculature and other compartments of bone marrow, thymus, lymph nodes and spleen. However, expression, recognition and use of laminin isoforms by lymphoid cells are poorly understood. In the present study, lymphoid T cells (Jurkat) were found to synthesize laminin alpha4, beta1 and gamma1 mRNAs and polypeptides and to assemble the chains into laminin-8. Lymphoblastoid B (NAD-20) cells, lymphoid NK (NKL) cells and blood lymphocytes also contained laminin-8 and, after cell permeabilization, practically all blood lymphocytes reacted with mAbs to laminin beta1 and gamma1 chains. Following stimulation, blood lymphocytes secreted laminin-8, and this laminin isoform, but not laminin-10/11(alpha5beta1gamma1/alpha5beta2gamma1), promoted chemokine-induced migration of the cells. In an activation-dependent manner, purified blood CD4 T cells adhered to immobilized laminin-8 and laminin-10/11 by using alpha6beta1 integrin, but minimally to laminin-1 (alpha1beta1gamma1). Accordingly, laminin-8 and laminin-10/11, but not laminin-1, strongly costimulated proliferation of the T cells via the same integrin. Thus, lymphoid cells are able to synthesize and secrete complete laminin molecules. In addition, synthesis of laminin-8 and recognition of laminin-8 and -10/11 by lymphocytes indicate relevance of these laminin isoforms in lymphocyte physiology. PMID:11148143

  19. Factor de crecimiento transformante beta-1: estructura, función y mecanismos de regulación en cáncer Transforming growth factor beta-1: structure, function and regulation mechanisms in cancer

    Directory of Open Access Journals (Sweden)

    Oscar Peralta-Zaragoza

    2001-08-01

    Full Text Available El factor de crecimiento transformante beta-1 (TGF-beta1 es sintetizado por muchas estirpes celulares como linfocitos, macrófagos y células dendríticas, y su expresión regula de manera autócrina o parácrina la diferenciación, proliferación y el estado de activación de éstas y muchas otras células. En general, el TGF-beta1 tiene propiedades pleiotrópicas en el contexto de la respuesta inmune durante el desarrollo de infecciones y procesos neoplásicos; sin embargo, los mecanismos de acción y regulación de la expresión de esta citocina aún no se comprenden del todo. En la presente revisión se describen las propiedades biológicas y los procesos moleculares que regulan la expresión del TGF-beta1, para entender los efectos de esta citocina durante la proliferación y la diferenciación celular. El conocimiento de los mecanismos moleculares de la regulación del TGF-beta1 puede representar una importante estrategia de tratamiento del cáncer. El texto completo en inglés de este artículo está disponible en: http://www.insp.mx/salud/index.htmlTransforming growth factor beta-1 (TGF-beta1 is produced by several cell lineages such as lymphocytes, macrophages, and dendritic cells, and its expression serves in both autocrine and paracrine modes to control the differentiation, proliferation, and state of activation of these and other cells. In general, TGF-beta1 has pleiotropic properties on the immune response during the development of infection diseases and cancer; however, the mechanisms of action and regulation of gene expression of this cytokine are poorly understood, In this review, the biological properties and the molecular mechanisms that regulate TGF-beta1 gene expression are described, to understand the role of this cytokine in growth and cell differentiation. The knowledge of molecular mechanisms of gene expression of TGF-beta1 may serve to develop new cancer therapies. The English version of this paper is available at: http://www.insp.mx/salud/index.html

  20. Integrin Activation and Viral Infection

    Institute of Scientific and Technical Information of China (English)

    Shan-dian GAO; Jun-zheng DU; Jian-hua ZHOU; Hui-yun CHANG; Qing-ge XIE

    2008-01-01

    Integrins are members of a ubiquitous membrane receptor family which includes 18 different α subunits and 8 β subunits forming more than 20 α/β heterodimers. Integrins play key functions in vascular endothelial cell and tumour cell adhesion, lymphocyte trafficking, tumor growth and viral infection. Current understanding of the molecular basis of integrins as viral receptors has been achieved through many decades of study into the biology of transmembrane glycoproteins and their interactions with several viruses. This review provides a summary of the current knowledge on the molecular bases of interactions between viruses and integrins, which are of potential practical significance. Inhibition of virus-integrin interactions at the points of virus attachment or entry will provide a novel approach for the therapeutic treatment of viral diseases.

  1. Mouse-Induced Pluripotent Stem Cells Differentiate into Odontoblast-Like Cells with Induction of Altered Adhesive and Migratory Phenotype of Integrin

    OpenAIRE

    Ozeki, Nobuaki; Mogi, Makio; Kawai, Rie; Yamaguchi, Hideyuki; Hiyama, Taiki; Nakata, Kazuhiko; Nakamura, Hiroshi

    2013-01-01

    Methods for differentiating induced pluripotent stem (iPS) cells into odontoblasts generally require epithelial–mesenchymal interactions. Here, we sought to characterize the cells produced by a ‘hanging drop’ technique for differentiating mouse iPS cells into odontoblast-like cells that requires no such interaction. Cells were cultured by the hanging drop method on a collagen type-I (Col-I) scaffold (CS) combined with bone morphogenetic protein (BMP)-4 (CS/BMP-4) without an epithelial–mesench...

  2. Functional display of an alpha2 integrin-specific motif (RKK) on the surface of baculovirus particles.

    Science.gov (United States)

    Riikonen, Reetta; Matilainen, Heli; Rajala, Nina; Pentikainen, Olli; Johnson, Mark; Heino, Jyrki; Oker-Blom, Christian

    2005-08-01

    The use of baculovirus vectors shows promise as a tool for gene delivery into mammalian cells. These insect viruses have been shown to transduce a variety of mammalian cell lines, and gene transfer has also been demonstrated in vivo. In this study, we generated two recombinant baculovirus vectors displaying an integrin-specific motif, RKK, as a part of two different loops of the green fluorescent protein (GFP) fused with the major envelope protein gp64 of Autographa californica M nucleopolyhedrovirus. By enzyme linked immunosorbent assays, these viruses were shown to bind a peptide representing the receptor binding site of an alpha2 integrin, the alpha2I-domain. However, the interaction was not strong enough to overcome binding of wild type gp64 to the unknown cellular receptor(s) on the surface of alpha2 integrin-expressing cells (CHO-alpha2beta1) or enhance the viral uptake. After treatment of these cells with phospholipase C, internalization of all viruses was blocked or decreased significantly. However, one of the RKK displaying viruses, AcGFP(K)gp64, was still able to internalize into CHO-alpha2beta1 cells, although at a lower level as compared to non-treated cells. This may indicate the possible utilization of a PLC independent alternative route via, in this case, the alpha2beta1 integrin. PMID:16029062

  3. Evaluation of in vivo partial beta 1/beta 2-agonist activity: a dose-ranging study with carteolol.

    OpenAIRE

    Wheeldon, N M; McDevitt, D G; Lipworth, B J

    1992-01-01

    1. The aims of this study were to investigate the partial agonist profile of carteolol and evaluate methodology for differentiating relative beta 1 and beta 2 partial agonist activity (PAA) in vivo. 2. Eight normal subjects received single oral doses of carteolol 10 mg, 30 mg and 60 mg; nadolol 40 mg; pindolol 30 mg and placebo, given in a single-blind, randomised crossover design. 3. beta 1-PAA was demonstrated with carteolol by dose-related increases in resting heart rate and systolic blood...

  4. Activation of p53 pathway by Nutlin-3a inhibits the expression of the therapeutic target alpha 5 integrin in colon cancer cells

    Czech Academy of Sciences Publication Activity Database

    Janoušková, Hana; Ray, A.M.; Noulet, F.; Lelong-Rebel, I.; Choulier, L.; Schaffner, F.; Lehmann, M.; Martin, S.; Teisinger, Jan; Dontenwill, M.

    2013-01-01

    Roč. 336, č. 2 (2013), s. 307-318. ISSN 0304-3835 Institutional support: RVO:67985823 Keywords : colon cancer * integrin alpha 5 beta 1 * p53 * Nutlin-3a Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.016, year: 2013

  5. Ubuntu 6.10 Edgy Beta 1发布

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    近日,Ubuntu项目发布Ubuntu 6.10 Edgy Eft Beta 1版本,Edgy Eft Beta 1包含新的2.6.17版本内核、Xorg 7.1、GNOME 2.16正式版、新的Firefox浏览器Firefox2.0 Beta2、OpenOffice.org2.0.4RC2版本、聊天软件Gaim的新版本2.0 Beta3.1和新的init系统。

  6. The change of transforming growth factor {beta} 1 (TGF- {beta} 1) expression by melatonin in irradiated lung

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Seong Soon; Choi, Ihl Bohng [College of Medicine, The Catholic University of Korea, Seoul (Korea, Republic of)

    2005-09-15

    The changed expressions of TGF- {beta} 1, as a key cytokine in the fibrotic process, due to melatonin with potent antioxidative effects, were investigated in the irradiated lung using fibrosis-sensitive C57BL/6 mice. Female C57BL/6 mice were divided into control irradiation-only, and melatonin (300 mg/kg i.p. 1 hr before irradiation) pretreatment groups. The thoraces of the mice were irradiated with a single dose of 12 Gy. The mRNA expressions of TGF-{beta} 1 in the lung tissue 2 and 4 weeks after irradiation were quantified using semiquantitive RT-PCR, and the cellular origin and expression levels of TGF- {beta} 1 protein were identified using immunohistochemical staining. The relative mRNA expression levels in the irradiation-only and melatonin pretreatment group 2 and 4 weeks after irradiation were 1.92- and 1.80-fold ({rho} = 0.064) and 2.38- and 1.94-fold ({rho} = 0.004) increased, respectively compared to those in the control group. Increased expressions of TGF- {beta} 1 protein were prominently detected in regions of histopathological radiation injury, with alveolar macrophages and septal epithelial cells serving as important sources of TGF- {beta} 1 expression. At 2 and 4 weeks after irradiation, the expression levels of protein were 15.8% vs. 16.9% ({rho} = 0.565) and 36.1% vs. 25.7% ({rho} = 0.009), respectively. The mRNA and protein expressions of TGF- {beta} 1 in the lung tissue following thoracic irradiation with 12 Gy were significantly decreased by melatonin pretreatment at 4 weeks. These results indicate that melatonin may have a possible application as an antifibrotic agent in radiation-induced lung injury.

  7. Different Phenotypes in Human Prostate Cancer: α6 or α3 Integrin in Cell-extracellular Adhesion Sites

    Directory of Open Access Journals (Sweden)

    Monika Schmelz

    2002-01-01

    Full Text Available The distribution of α6/α3 integrin in adhesion complexes at the basal membrane in human normal and cancer prostate glands was analyzed in 135 biopsies from 61 patients. The levels of the polarized α6/α3 integrin expression at the basal membrane of prostate tumor glands were determined by quantitative immunohistochemistry. The α6/α3 integrin expression was compared with Gleason sum score, pathological stage, and preoperative serum prostate-specific antigen (PSA. The associations were assessed by statistical methods. Eighty percent of the tumors expressed the α6 or α3 integrin and 20% was integrin-negative. Gleason sum score, but not serum PSA, was associated with the integrin expression. Low Gleason sum score correlated with increased integrin expression, high Gleason sum score with low and negative integrin expression. Three prostate tumor phenotypes were distinguished based on differential integrin expression. Type I coexpressed both α6 and α3 subunits, type II exclusively expressed a6 integrin, and type III expressed α3 integrin only. Fifteen cases were further examined for the codistribution of vinculin, paxillin, and CD 151 on frozen serial sections using confocal laser scanning microscopy. The α6/α3 integrins, CD151, paxillin, and vinculin were present within normal glands. In prostate carcinoma, α6 integrin was colocalized with CD 151, but not with vinculin or paxillin. In tumor phenotype I, the α6 subunit did not colocalize with the α3 subunit indicating the existence of two different adhesion complexes. Human prostate tumors display on their cell surface the α6β1 and/or α3β1 integrins. Three tumor phenotypes associated with two different adhesion complexes were identified, suggesting a reorganization of cell adhesion structures in prostate cancer.

  8. Integrin Targeted Delivery of Radiotherapeutics

    Directory of Open Access Journals (Sweden)

    Zhaofei Liu, Fan Wang, Xiaoyuan Chen

    2011-01-01

    Full Text Available Targeted radionuclide therapy, which is based on the selective delivery of a sufficient radiation dose to tumors without significantly affecting normal tissues, is a promising therapeutic approach for the treatment of a wide variety of malignancies. Integrins, a family of cell adhesion molecules, play key roles during tumor angiogenesis and metastasis. Among all the integrins, αvβ3 seems to be the most important in the process of tumor angiogenesis. Integrin αvβ3 is highly expressed on activated endothelial cells, new-born vessels as well as some tumor cells, but is not present in resting endothelial cells and most normal organ systems, making it a suitable target for anti-tumor therapy. In this review, we summarize the current development and applications of antibody-, peptide-, and other ligand-based integrin targeted radiotherapeutics for tumor radiation therapy.

  9. The role of integrins in the development and homeostasis of the epidermis and skin appendages.

    Science.gov (United States)

    Rippa, A L; Vorotelyak, E A; Vasiliev, A V; Terskikh, V V

    2013-10-01

    Integrins play a critical role in the regulation of adhesion, migration, proliferation, and differentiation of cells. Because of the variety of the functions they play in the cell, they are necessary for the formation and maintenance of tissue structure integrity. The trove of data accumulated by researchers suggests that integrins participate in the morphogenesis of the epidermis and its appendages. The development of mice with tissue-specific integrin genes knockout and determination of the genetic basis for a number of skin diseases in humans showed the significance of integrins in the biology, physiology, and morphogenesis of the epidermis and hair follicles. This review discusses the data on the role of different classes of integrin receptors in the biology of epidermal cells, as well as the development of the epidermis and hair follicles. PMID:24455180

  10. Transforming growth factor-beta1 stimulates the production of insulin-like growth factor-I and insulin-like growth factor-binding protein-3 in human bone marrow stromal osteoblast progenitors

    DEFF Research Database (Denmark)

    Kveiborg, Marie; Flyvbjerg, Allan; Eriksen, E F;

    2001-01-01

    While transforming growth factor-beta1 (TGF-beta1) regulates proliferation and differentiation of human osteoblast precursor cells, the mechanisms underlying these effects are not known. Several hormones and locally acting growth factors regulate osteoblast functions through changes in the insulin......-like growth factors (IGFs) and IGF-binding proteins (IGFBPs). Thus, we studied the effects of TGF-beta1 on IGFs and IGFBPs in human marrow stromal (hMS) osteoblast precursor cells. TGF-beta1 increased the steady-state mRNA level of IGF-I up to 8.5+/-0.6-fold (P...

  11. New insights into structure-function relationship of the DHPR beta1a subunit in skeletal muscle excitation-contraction coupling using zebrafish 'relaxed' as an expression system

    International Nuclear Information System (INIS)

    The paralyzed zebrafish strain relaxed carries a null mutation for the skeletal muscle dihydropyridine receptor (DHPR) [beta]1a subunit. The lack of [beta]1a not only impedes functional [alpha]1S membrane expression but also precludes the skeletal muscle-specific ultrastructural arrangement of DHPRs into tetrads opposite ryanodine receptor (RyR1), coherent with the absence of skeletal muscle excitation-contraction (EC) coupling. With the plethora of experimental approaches feasible with zebrafish model organism and importantly with the [beta]1-null mutation having a monogenetic inheritance and because of the survival of the relaxed larvae for some days, we were able to establish the zebrafish relaxed as an expression system. Linking in vitro to in vivo observations, a clear differentiation between the major functional roles of [beta] subunits in EC coupling was feasible. The skeletal muscle [beta]1a subunit was able to restore all parameters of EC coupling upon expression in relaxed myotubes and larvae. Expression of the phylogenetically closest isoform to [beta]1a, the cardiac/neuronal [beta]2a subunit or the most distant neuronal [beta]M from the housefly in relaxed myotubes and larvae was likewise able to fully restore [alpha]1S triad targeting and facilitate charge movement. However, efficient tetrad formation and thus intact DHPR-RyR1 coupling was exclusively promoted by the [beta]1a isoform. Consequently, we postulated a model according to which [beta]1a acts as a unique allosteric modifier of [alpha]1S conformation crucial for skeletal muscle EC coupling. Therefore, unique structural elements in [beta]1a must be present which endow it with this exclusive property. Earlier, a unique hydrophobic heptad repeat motif (LVV) in the [beta]1a C-terminus was postulated by others to be essential for skeletal muscle EC coupling. We wanted to address the question if the proposed [beta]1a heptad repeat motif could be an active element of the DHPR-RyR1 signal transduction

  12. The involvement of Gab1 and PI 3-kinase in β1 integrin signaling in keratinocytes

    International Nuclear Information System (INIS)

    The control of the stem cell compartment in epidermis is closely linked to the regulation of keratinocyte proliferation and differentiation. β1 integrins are expressed 2-fold higher by stem cells than transit-amplifying cells. Signaling from these β1 integrins is critical for the regulation of the epidermal stem cell compartment. To clarify the functional relevance of this differential expression of β1 integrins, we established HaCaT cells with high β1integrin expression by repeated flow cytometric sorting of this population from the parental cell line. In these obtained cells expressing β1 integrins by 5-fold, MAPK activation was markedly increased. Regarding the upstream of MAPK, Gab1 phosphorylation was also higher with high β1 integrin expression, while Shc phosphorylation was not altered. In addition, enhanced phosphatidylinositol 3-kinase activation was also observed. These observations suggest that Gab1 and phosphatidylinositol 3-kinase play pivotal roles in the β1 integrin-mediated regulation of the epidermal stem cell compartment

  13. The newcomer in the integrin family: Integrin a9 in biology and cancer

    DEFF Research Database (Denmark)

    Høye, Anette Melissa; Couchman, John Robert; Wewer, Ulla M.;

    2012-01-01

    Integrins are heterodimeric transmembrane receptors regulating cell-cell and cell-extracellular matrix interactions. Of the 24 integrin heterodimers identified in humans, a9ß1 integrin is one of the least studied. a9, together with a4, comprise a more recent evolutionary sub-family of integrins t...

  14. Fibronectin-binding protein acts as Staphylococcus aureus invasin via fibronectin bridging to integrin alpha5beta1

    NARCIS (Netherlands)

    Sinha, B; François, P P; Nüsse, O; Foti, M; Hartford, O M; Vaudaux, P; Foster, T J; Lew, D P; Herrmann, M; Krause, K H

    1999-01-01

    The ability of Staphylococcus aureus to invade mammalian cells may explain its capacity to colonize mucosa and to persist in tissues after bacteraemia. To date, the underlying molecular mechanisms of cellular invasion by S. aureus are unknown, despite its high prevalence and difficulties in treatmen

  15. HLA Dr beta 1 alleles in Pakistani patients with rheumatoid arthritis

    International Nuclear Information System (INIS)

    Objective: To determine frequencies of HLA DR beta 1 alleles in rheumatoid arthritis in Pakistani patients. Study Design: Cross sectional / analytical study. Place and Duration of Study: Department of Immunology, Armed Forces Institute of Pathology, Rawalpindi in collaboration with Rheumatology departments of Military Hospital, Rawalpindi and Fauji Foundation Hospital, Rawalpindi, from January 2009 to January 2010. Methodology: HLA DR beta 1 genotyping of one hundred Pakistani patients, diagnosed as having RA as per American College of Rheumatology revised criteria 1987, was done. HLA DR beta 1 genotyping was carried out at allele group level (DR beta 1*01-DR beta 1*16) by sequence specific primers in RA patients. Comparison of HLA DR beta 1 allele frequencies between patients and control groups was made using Pearson's chi-square test to find possible association of HLA DR?1 alleles with RA in Pakistani rheumatoid patients. Results: HLA DR beta 1*04 was expressed with significantly increased frequency in patients with rheumatoid arthritis (p <0.05). HLA DR?1*11 was expressed statistically significantly more in control group as compared to rheumatoid patients indicating a possible protective effect. There was no statistically significant difference observed in frequencies of HLA DR beta 1 allele *01, DR beta 1 allele *03, DR beta 1 allele *07, DR beta 1 allele *08, DR beta 1 allele *09, DR beta 1 allele *10, DR beta 1 allele *12, DR beta 1 allele *13, DR beta 1 allele *14, DR?1 allele *15 and DR beta 1 allele *16 between patients and control groups. Conclusion: The identification of susceptible HLA DR beta 1 alleles in Pakistani RA patients may help physicians to make early decisions regarding initiation of early intensive therapy with disease modifying anti rheumatic medicines and biological agents decreasing disability in RA patients. (author)

  16. Compound list: transforming growth factor beta 1 [Open TG-GATEs

    Lifescience Database Archive (English)

    Full Text Available transforming growth factor beta 1 TGFB1 00182 ftp://ftp.biosciencedbc.jp/archive/op...en-tggates/LATEST/Human/in_vitro/transforming_growth_factor_beta_1.Human.in_vitro.Liver.zip ...

  17. Arginine-Glycine-Aspartate-Binding Integrins as Therapeutic and Diagnostic Targets.

    Science.gov (United States)

    Sun, Cui-Cui; Qu, Xian-Jun; Gao, Zu-Hua

    2016-01-01

    Arginine-glycine-aspartate (RGD)-binding integrins, including αvβ1, αvβ3, αvβ5, αvβ6, αvβ8, α5β1, αIIbβ3, and α8β1, recognize the tripeptide motif RGD in their ligands. RGD-binding integrins are involved in various cell functions, including cell proliferation, survival, differentiation, and motility that are critically important to both health and disease. The diagnostic and therapeutic value of some RGD-binding integrin inhibitors are either clinically proven or at different stages of development. In this review, we first summarized the structure and signaling characteristics of RGD-binding integrins. We then discussed the functions of RGD-binding integrins and their association with human disease. Finally, we recapitulated the research efforts and clinical trials of targeting RGD-binding integrins for the diagnosis and treatment of human disease. This comprehensive review of the current advances in RGD-binding integrins could assist scientists and clinicians in gaining a complete understanding of this group of molecules. It can also contribute to the design of new projects to further advance this field of research and to better apply the research results to benefit patients in clinical practice. PMID:24621642

  18. Tumor targeting via integrin ligands

    Directory of Open Access Journals (Sweden)

    HorstKessler

    2013-08-01

    Full Text Available Selective and targeted delivery of drugs to tumors is a major challenge for an effective cancer therapy and also to overcome the side effects associated with current treatments. Overexpression of various receptors on tumor cells is a characteristic structural and biochemical aspect of tumors and distinguishes them from physiologically normal cells. This abnormal feature is therefore suitable for selectively directing anticancer molecules to tumors by using ligands that can preferentially recognize such receptors. Several subtypes of integrin receptors that are crucial for cell adhesion, cell signaling, cell viability and motility have been shown to have an upregulated expression on cancer cells. Thus, ligands that recognize specific integrin subtypes represent excellent candidates to be conjugated to drugs or drug carrier systems and be targeted to tumors. In this regard, integrins recognizing the RGD cell adhesive sequence have been extensively targeted for tumor specific drug delivery. Here we review key recent examples on the presentation of RGD-based integrin ligands by means of distinct drug delivery systems, and discuss the prospects of such therapies to specifically target tumor cells.

  19. Glomerular injury is exacerbated in diabetic integrin alpha1-null mice.

    Science.gov (United States)

    Zent, R; Yan, X; Su, Y; Hudson, B G; Borza, D-B; Moeckel, G W; Qi, Z; Sado, Y; Breyer, M D; Voziyan, P; Pozzi, A

    2006-08-01

    Excessive glomerular collagen IV and reactive oxygen species (ROS) production are key factors in the development of diabetic nephropathy. Integrin alpha1beta1, the major collagen IV receptor, dowregulates collagen IV and ROS production, suggesting this integrin might determine the severity of diabetic nephropathy. To test this possibility, wild-type and integrin alpha1-null mice were rendered diabetic with streptozotocin (STZ) (100 mg/kg single intraperitoneal injection), after which glomerular filtration rate (GFR), glomerular collagen deposition, and glomerular basement membrane (GBM) thickening were evaluated. In addition, ROS and collagen IV production by mesangial cells as well as their proliferation was measured in vitro. Diabetic alpha1-null mice developed worse renal disease than diabetic wild-type mice. A significant increase in GFR was evident in the alpha1-null mice at 6 weeks after the STZ injection; it started to decrease by week 24 and reached levels of non-diabetic mice by week 36. In contrast, GFR only increased in wild-type mice at week 12 and its elevation persisted throughout the study. Diabetic mutant mice also showed increased glomerular deposition of collagen IV and GBM thickening compared to diabetic wild-type mice. Primary alpha1-null mesangial cells exposed to high glucose produced more ROS than wild-type cells, which led to decreased proliferation and increased collagen IV synthesis, thus mimicking the in vivo finding. In conclusion, this study suggests that lack of integrin alpha1beta1 exacerbates the glomerular injury in a mouse model of diabetes by modulating GFR, ROS production, cell proliferation, and collagen deposition. PMID:16775606

  20. Interaction between the insulin-like growth factor family and the integrin receptor family in tissue repair processes. Evidence in a rabbit ear dermal ulcer model.

    OpenAIRE

    Galiano, R D; Zhao, L L; Clemmons, D R; Roth, S. I.; Lin, X.; Mustoe, T. A.

    1996-01-01

    We have determined previously that IGF-I is dependent on the presence of IGF binding protein-1 (IGFBP-1) to act as a wound healing agent. We sought to determine the mechanism whereby IGFBP-1 is able to enhance IGF-I bioactivity. As IGFBP-1 binds both the alpha5beta1 integrin as well as IGF-I in vitro, we asked which of the following interactions were important: (a) the ability of IGFBP-1 to interact with an integrin receptor, and/or (b) the binding of IGF-I by IGFBP-1. We used an IGF-1 analog...

  1. Current Perspectives on Interferon Beta-1b for the Treatment of Multiple Sclerosis

    OpenAIRE

    Marziniak, Martin; Meuth, Sven

    2014-01-01

    Interferon (IFN) beta-1b was the first disease-modifying therapy to be approved for the treatment of multiple sclerosis (MS), and over 21 years of follow-up data demonstrate its efficacy and long-term safety profile. Following recent regulatory approvals in the USA and European Union, IFN beta-1b is now one of the seven disease-modifying therapies [intramuscular IFN beta-1a; subcutaneous (SC) IFN beta-1a; IFN beta-1b SC; glatiramer acetate SC; oral dimethyl fumarate; oral teriflunomide; and i...

  2. Integrin Expression Regulates Neuroblastoma Attachment and Migration

    Directory of Open Access Journals (Sweden)

    Amy Meyer

    2004-07-01

    Full Text Available Neuroblastoma (NBL is the most common malignant disease of infancy, and children with bone metastasis have a mortality rate greater than 90%. Two major classes of proteins, integrins and growth factors, regulate the metastatic process. We have previously shown that tumorigenic NBL cells express higher levels of the type I insulin-like growth factor receptor (IGF-IR and that β1 integrin expression is inversely proportional to tumorigenic potential in NBL. In the current study, we analyze the effect of β1 integrin and IGF-IR on NBL cell attachment and migration. Nontumorigenic S-cells express high levels of β1 integrin, whereas tumorigenic N-cells express little β1 integrin. Alterations in (3, integrin are due to regulation at the protein level, as translation is decreased in N-type cells. Moreover, inhibition of protein synthesis shows that β1 integrin is degraded more slowly in S-type cells (SHEP than in N-type cells (SH-SY5Y and IMR32. Inhibition of α5β1 integrin prevents SHEP (but not SH-SY5Y or IMR32 cell attachment to fibronectin and increases SHEP cell migration. Increases in IGF-IR decrease β1 integrin expression, and enhance SHEP cell migration, potentially through increased expression of αvβ3. These data suggest that specific classes of integrins in concert with IGF-IR regulate NBL attachment and migration.

  3. Anti-Integrin Therapy for Multiple Sclerosis

    Directory of Open Access Journals (Sweden)

    Eiji Kawamoto

    2012-01-01

    Full Text Available Integrins are the foremost family of cell adhesion molecules that regulate immune cell trafficking in health and diseases. Integrin alpha4 mediates organ-specific migration of immune cells to the inflamed brain, thereby playing the critical role in the pathogenesis of multiple sclerosis. Anti-alpha4 integrin therapy aiming to block infiltration of autoreactive lymphocytes to the inflamed brain has been validated in several clinical trials for the treatment of multiple sclerosis. This paper provides readers with an overview of the molecular and structural bases of integrin activation as well as rationale for using anti-alpha4 integrin therapy for multiple sclerosis and then chronicles the rise and fall of this treatment strategy using natalizumab, a humanized anti-alpha4 integrin.

  4. Anti-Integrin Therapy for Multiple Sclerosis

    OpenAIRE

    Motomu Shimaoka; Hiroshi Imai; Takayuki Okamoto; Susumu Nakahashi; Eiji Kawamoto

    2012-01-01

    Integrins are the foremost family of cell adhesion molecules that regulate immune cell trafficking in health and diseases. Integrin alpha4 mediates organ-specific migration of immune cells to the inflamed brain, thereby playing the critical role in the pathogenesis of multiple sclerosis. Anti-alpha4 integrin therapy aiming to block infiltration of autoreactive lymphocytes to the inflamed brain has been validated in several clinical trials for the treatment of multiple sclerosis. This paper pr...

  5. Transforming growth factor beta 1 (TGF-beta 1) induced neutrophil recruitment to synovial tissues: implications for TGF-beta-driven synovial inflammation and hyperplasia

    International Nuclear Information System (INIS)

    We have studied the consequences of introducing human recombinant transforming growth factor beta 1 (hrTGF-beta 1) into synovial tissue of the rat, to begin to better understand the significance of the fact that biologically active TGF-beta is found in human arthritic synovial effusions. Within 4-6 h after the intra-articular injection of 1 microgram of hrTGF-beta 1 into rat knee joints, extensive recruitment of polymorphonuclear leukocytes (PMNs) was observed. Cytochemistry and high resolution histological techniques were used to quantitate the influx of PMNs, which peaked 6 h post-injection. In a Boyden chamber assay, hrTGF-beta 1 at 1-10 fg/ml elicited a chemotactic response from PMNs greater in magnitude than that evoked by FMLP, establishing that TGF-beta 1 is an effective chemotactic agent for PMNs in vitro as well as in vivo. That PMNs may represent an important source of TGF-beta in inflammatory infiltrates was strongly suggested by a demonstration that stored TGF-beta 1 was secreted during phorbol myristate acetate-stimulated degranulation in vitro. Acid/ethanol extracts of human PMNs assayed by ELISA contained an average of 355 ng of TGF/beta 1 per 10(9) cells potentially available for secretion during degranulation of PMNs. [3H]Thymidine incorporation in vivo and autoradiography of tissue sections revealed that widespread cell proliferation was triggered by TGF-beta 1 injection. Synovial lining cells and cells located deep within the subsynovial connective tissue were identified as sources of at least some of the new cells that contribute to TGF-beta 1-induced hyperplasia. Our results demonstrate that TGF-beta is capable of exerting pathogenic effects on synovial tissue and that PMNs may represent a significant source of the TGF-beta present in synovial effusions

  6. The Effect of Stromal Integrin β3-Deficiency on Two Different Tumors in Mice

    Directory of Open Access Journals (Sweden)

    Inga Reigstad

    2016-01-01

    Full Text Available There is an increasing focus on the tumor microenvironment in carcinogenesis. Integrins are important receptors and adhesion molecules in this environment and have been shown to be involved in cell adhesion, proliferation, differentiation and migration. The present study aimed to evaluate the effect of stromal integrin β3-deficiency on tumor growth, angiogenesis, interstitial fluid pressure (PIF, fibrosis and metastasis in a murine breast cancer (4T1 and a prostate tumor (RM11 model. We showed that stromal integrin β3-deficiency led to an elevation in PIF that correlated to a shift towards thicker collagen fibrils in the 4T1 mammary tumor. In the RM11 prostate carcinoma model there was no effect of integrin β3-deficiency on PIF and collagen fibril thickness. These findings support the notion that changes in the collagen scaffold influence PIF, and also indicate that there must be important crosstalk between the stroma and tumor cells, in a tumor cell line specific manner. Furthermore, stromal integrin β3-deficiency had no effect on tumor growth or angiogenesis in both tumor models and no effect on lung metastasis in the 4T1 mammary tumor model. In conclusion, the stromal β3 integrin influence PIF, possibly via its effect on the structure of the collagen network, in a tumor cell line dependent manner.

  7. Bilinexin, a snake C-type lectin from Agkistrodon bilineatus venom agglutinates platelets via GPIb and alpha2beta1.

    Science.gov (United States)

    Du, X Y; Navdaev, A; Clemetson, J M; Magnenat, E; Wells, T N; Clemetson, K J

    2001-11-01

    A new snake protein, named bilinexin, has been purified from Agkistrodon bilineatus venom by ion-exchange chromatography and gel filtration chromatography. Under non-reducing conditions it has a mass of 110 kDa protein on SDS-PAGE. On reduction, it can be separated into five subunits with masses in the range 13-25 kDa. The N-terminal sequences of these subunits are very similar to those of convulxin or the alboaggregins, identifying bilinexin as a new member of the snake C-type lectin family, unusual in having multiple subunits. Bilinexin agglutinates fixed platelets. washed platelets and platelet rich plasma (PRP) without obvious activation (shape change) as confirmed by light microscope examination. Both inhibitory and binding studies indicate that antibodies against alpha2beta1 inhibit not only platelet agglutination induced by bilinexin, but also bilinexin binding to platelets. VM16d, a monoclonal anti-GPIbalpha antibody, completely inhibits platelet agglutination induced by bilinexin, and polyclonal antibodies against GPIbalpha prevent its binding to platelets. However, neither convulxin, polyclonal anti-GPVI antibodies, nor GPIIb/IIIa inhibitors affect its binding to and agglutination of platelets. Bilinexin neither activates GPIIb/IIIa integrin on platelets nor induces tyrosine phosphorylation of platelet proteins, nor increases intracellular Ca2+ in platelets. Like alboaggregin B, bilinexin agglutinates platelets, which makes it a good tool to investigate the differences in mechanism between snake C-type lectins causing platelet agglutination and those that induce full activation. PMID:11816718

  8. In situ validation of VEGFR-2 and α v ß 3 integrin as targets for breast lesion characterization.

    Science.gov (United States)

    Ehling, Josef; Misiewicz, Matthias; von Stillfried, Saskia; Möckel, Diana; Bzyl, Jessica; Pochon, Sibylle; Lederle, Wiltrud; Knuechel, Ruth; Lammers, Twan; Palmowski, Moritz; Kiessling, Fabian

    2016-04-01

    Vascular endothelial growth factor receptor 2 (VEGFR-2) and α v ß 3 integrin are the most frequently addressed targets in molecular imaging of tumor angiogenesis. In preclinical studies, molecular imaging of angiogenesis has shown potential to detect and differentiate benign and malignant lesions of the breast. Thus, in this retrospective clinical study employing patient tissues, the diagnostic value of VEGFR-2, α v ß 3 integrin and vascular area fraction for the diagnosis and differentiation of breast neoplasia was evaluated. To this end, tissue sections of breast cancer (n = 40), pre-invasive ductal carcinoma in situ (DCIS; n = 8), fibroadenoma (n = 40), radial scar (n = 6) and normal breast tissue (n = 40) were used to quantify (1) endothelial VEGFR-2, (2) endothelial α v ß 3 integrin and (3) total α v ß 3 integrin expression, as well as (4) the vascular area fraction. Sensitivity and specificity to differentiate benign from malignant lesions were calculated for each marker by receiver operating characteristics (ROC) analyses. Whereas vessel density, as commonly used, did not significantly differ between benign and malignant lesions (AUROC: 0.54), VEGFR-2 and α v ß 3 integrin levels were gradually up-regulated in carcinoma versus fibroadenoma versus healthy tissue. The highest diagnostic accuracy for differentiating carcinoma from fibroadenoma was found for total α v ß 3 integrin expression (AUROC: 0.76), followed by VEGFR-2 (AUROC: 0.71) and endothelial α v ß 3 integrin expression (AUROC: 0.68). In conclusion, total α v ß 3 integrin expression is the best discriminator between breast cancer, fibroadenoma and normal breast tissue. With respect to vascular targeting and molecular imaging of angiogenesis, endothelial VEGFR-2 appeared to be slightly superior to endothelial α v ß 3 for differentiating benign from cancerous lesions. PMID:26902100

  9. Localization of insulin-like growth factor (IGFBP)-3 in cultured porcine embryonic myogenic cells before and after TGF-beta1 treatment.

    Science.gov (United States)

    Xi, G; Hathaway, M R; White, M E; Dayton, W R

    2007-11-01

    Insulin-like growth factor binding protein (IGFBP)-3 binds IGFs with high affinity and affects their biological activity. IGFBP-3 that is not bound to IGF also affects cells via mechanisms involving binding to specific cell surface receptors and/or transport into the cell. IGFBP-3 is produced by porcine embryonic myogenic cell (PEMC) cultures. Additionally, IGFBP-3 facilitates the proliferation-suppressing actions of TGF-beta(1) and myostatin in PEMC cultures via mechanisms that do not involve IGF binding. Moreover, these mechanisms do not involve preventing myostatin or TGF-beta(1)-induced increases in phosphosmad2 or phosphosmad3 level. Consequently, the mechanism(s) by which IGFBP-3 facilitates the proliferation-suppressing actions of TGF-beta(1) and myostatin in PEMC is unclear. Since IGFBP-3 reportedly interacts with nuclear proteins that regulate transcription, TGF-beta(1) or myostatin-induced translocation of IGFBP-3 into the nucleus may facilitate the proliferation-suppressing actions of these cytokines. Here, we show that IGFBP-3 is localized in cells containing the muscle specific protein desmin, thus establishing the presence of this IGFBP in myogenic cells. IGFBP-3 is present in the cytoplasm of all myogenic cells and approximately 50% of the nuclei of proliferating PEMC. IGFBP-3 is also detectable in fused myotubes. IGFBP-3 suppresses IGF-I-stimulated differentiation of PEMC but has no affect on Long-R3-IGF-I-stimulated differentiation of PEMC. Treatment of PEMC for 24h with TGF-beta(1) (20 ng/ml) results in a 78% (p<0.01) increase in the number of nuclei that contain detectable IGFBP-3. These results suggest that translocation of IGFBP-3 into the nucleus of PEMC could play a role in mediating the proliferation-suppressing action of TGF-beta(1). PMID:17049199

  10. Extracellular matrix stiffness dictates Wnt expression through integrin pathway.

    Science.gov (United States)

    Du, Jing; Zu, Yan; Li, Jing; Du, Shuyuan; Xu, Yipu; Zhang, Lang; Jiang, Li; Wang, Zhao; Chien, Shu; Yang, Chun

    2016-01-01

    It is well established that extracellular matrix (ECM) stiffness plays a significant role in regulating the phenotypes and behaviors of many cell types. However, the mechanism underlying the sensing of mechanical cues and subsequent elasticity-triggered pathways remains largely unknown. We observed that stiff ECM significantly enhanced the expression level of several members of the Wnt/β-catenin pathway in both bone marrow mesenchymal stem cells and primary chondrocytes. The activation of β-catenin by stiff ECM is not dependent on Wnt signals but is elevated by the activation of integrin/ focal adhesion kinase (FAK) pathway. The accumulated β-catenin then bound to the wnt1 promoter region to up-regulate the gene transcription, thus constituting a positive feedback of the Wnt/β-catenin pathway. With the amplifying effect of positive feedback, this integrin-activated β-catenin/Wnt pathway plays significant roles in mediating the enhancement of Wnt signal on stiff ECM and contributes to the regulation of mesenchymal stem cell differentiation and primary chondrocyte phenotype maintenance. The present integrin-regulated Wnt1 expression and signaling contributes to the understanding of the molecular mechanisms underlying the regulation of cell behaviors by ECM elasticity. PMID:26854061

  11. Determination of transforming growth factor-beta 1 (TGF-beta 1) and insulin-like growth factor (IGF-1) in bovine colostrum samples.

    Science.gov (United States)

    Ginjala, V; Pakkanen, R

    1998-01-01

    The major growth factors in bovine colostrum are transforming growth factor-beta s (TGF-beta 1 and TGF-beta 2) and insulin-like growth factors (IGF-1 and IGF-2). Recently, TGF-beta 2 content of bovine colostrum was measured using a TGF-beta 2 specific ELISA (1) and now we have validated ELISAs for for bovine TGF-beta 1 and IGF-1. The concentrations of IGF-1 and TGF-beta 1 in the first milking after calving were 248-1850 ng/ml and 12.4-42.6 ng/ml, respectively, and they declined in correlation with total protein concentration to 27.0-101 ng/ml (IGF-1) and 0.80-3.49 ng/ml(TGF-beta 1) by the fifth milkings. The amount of TGF-beta 1 was on average 5.3 +/- 1.4% of that of TGF-beta 2 and there is a high correlation (r = 0.966) between the concentrations of these growth factors in the same samples. No free TGF-beta 1 form of could be detected. PMID:9682131

  12. Molecular cloning and characterisation of a pattern recognition molecule, lipopolysaccharide- and beta-1,3-glucan binding protein (LGBP) from the white shrimp Litopenaeus vannamei.

    Science.gov (United States)

    Cheng, Winton; Liu, Chun-Hung; Tsai, Chiung-Hui; Chen, Jiann-Chu

    2005-04-01

    A lipopolysaccharide- and beta-1,3-glucan binding protein (LGBP) cDNA was cloned from the haemocyte and hepatopancreas of white shrimp Litopenaeus vannamei using oligonucleotide primers and RT-PCR. Both 3'- and 5'-regions were isolated by rapid amplification of cDNA end RACE method. Analysis of nucleotide sequence revealed that the cDNA clone has an open reading frame of 1101 bp encoding a protein of 367 amino acids including a 17 amino acid signal peptide. The calculated molecular mass of the mature proteins (350 amino acids) is 39.92 kDa with an estimated pI of 4.37. Two putative integrin binding motifs (cell adhesion site), RGD (Arg-Gly-Asp) and a potential recognition motif for beta- (1-->3) linkage of polysaccharides were observed in the LGBP. Sequence comparison showed that LGBP deduced amino acid of L. vannamei has an overall similarity of 95%, 92% and 61% to that of blue shrimp Litopenaeus stylirostris LGBP, tiger shrimp Penaeus monodon BGBP and crayfish Pacifastacus leniusculus LGBP, respectively. Quantitative real-time RT-PCR analysis showed that LGBP transcript in haemocyte of L. vannamei increased in 3- and 6-h post Vibrio alginolyticus injection. PMID:15561560

  13. Molecular cloning and characterisation of a pattern recognition protein, lipopolysaccharide and beta-1,3-glucan binding protein (LGBP) from Chinese shrimp Fenneropenaeus chinensis.

    Science.gov (United States)

    Liu, Fengsong; Li, Fuhua; Dong, Bo; Wang, Xiaomei; Xiang, Jianhai

    2009-03-01

    A pattern recognition protein (PRP), lipopolysaccharide and beta-1,3-glucan binding protein (LGBP) cDNA was cloned from the haemocyte of Chinese shrimp Fenneropenaeus chinensis by the techniques of homology cloning and RACE. Analysis of nucleotide sequence revealed that the full-length cDNA of 1,275 bp has an open reading frame of 1,098 bp encoding a protein of 366 amino acids including a 17 amino acid signal peptide. Sequence comparison of the deduced amino acid sequence of F. chinensis LGBP showed a high identity of 94%, 90%, 87%, 72% and 63% with Penaeus monodon BGBP, Litopenaeus stylirostris LGBP, Marsupenaeu japonicus BGBP, Homarus gammarus BGBP and Pacifastacus leniusculus LGBP, respectively. The calculated molecular mass of the mature protein is 39,857 Da with a deduced pI of 4.39. Two putative integrin binding motifs, RGD (Arg-Gly-Asp) and a potential recognition motif for beta-1,3-linkage of polysaccharides were observed in LGBP sequence. RT-PCR analysis showed that LGBP gene expresses in haemocyte and hepatopancreas only, but not in other tissues. Capillary electrophoresis RT-PCR method was used to quantify the variation of mRNA transcription level during artificial infection with heat-killed Vibrio anguillarum and Staphylococcus aureusin. A significant enhancement of LGBP transcription was appeared at 6 h post-injection in response to bacterial infection. These results have provided useful information to understand the function of LGBP in shrimp. PMID:18163220

  14. Integrin αvβ3 expression in tongue squamous carcinoma cells Cal27 confers anticancer drug resistance through loss of pSrc(Y418).

    Science.gov (United States)

    Stojanović, Nikolina; Brozovic, Anamaria; Majhen, Dragomira; Bosnar, Maja Herak; Fritz, Gerhard; Osmak, Maja; Ambriović-Ristov, Andreja

    2016-08-01

    Integrins play key roles in the regulation of tumor cell adhesion, migration, invasion and sensitivity to anticancer drugs. In the present study we investigate the mechanism of resistance of tongue squamous carcinoma cells Cal27 with de novo integrin αvβ3 expression to anticancer drugs. Cal27-derived cell clones, obtained by transfection of plasmid containing integrin subunit β3 cDNA, as compared to control cells demonstrate: expression of integrin αvβ3; increased expression of integrin αvβ5; increased adhesion to fibronectin and vitronectin; resistance to cisplatin, mitomycin C, doxorubicin and 5-fluorouracil; increased migration and invasion, increased amount of integrin-linked kinase (ILK) and decreased amounts of non-receptor tyrosine kinase (Src) and pSrc(Y418). Knockdown of ILK and integrin β5 in cells expressing integrin αvβ3 ruled out their involvement in drug resistance. Opposite, Src knockdown in Cal27 cells which led to a reduction in pSrc(Y418), as well as treatment with the pSrc(Y418) inhibitors dasatinib and PP2, conferred resistance to all four anticancer drugs, indicating that the loss of pSrc(Y418) is responsible for the observed effect. We identified differential integrin signaling between Cal27 and integrin αvβ3-expressing cells. In Cal27 cells integrin αv heterodimers signal through pSrc(Y418) while this is not the case in integrin αvβ3-expressing cells. Finally, we show that dasatinib counteracts the effect of cisplatin in two additional head and neck squamous cell carcinoma (HNSCC) cell lines Cal33 and Detroit562. Our results suggest that pSrc(Y418) inhibitors, potential drugs for cancer therapy, may reduce therapeutic efficacy if combined with chemotherapeutics, and might not be recommended for HNSCC treatment. PMID:27108184

  15. In vivo evaluation of expression of TGF{beta}1 in the irradiated heart; Expressao da proteina TGF{beta}1 em coracao irradiado in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Affonso Junior, Renato Jose [Universidade Federal de Sao Paulo (UNIFESP), SP (Brazil). Escola Paulista de Medicina. Dept. Diagnostico por Imagem; Oshima, Celina Tizuko Fujiyama; Silva, Maria Regina Regis [Universidade Federal de Sao Paulo (UNIFESP), SP (Brazil). Escola Paulista de Medicina. Setor de Anatomia Patologica; Kimura, Edna Teruko [Sao Paulo Univ., SP (Brazil). Inst. de Ciencias Biomedicas. Dept. de Histologia; Egami, Mizue Imoto [Universidade Federal de Sao Paulo (UNIFESP), SP (Brazil). Escola Paulista de Medicina. Dept. de Morfologia; Segreto, Roberto Araujo; Segreto, Helena Regina Comodo [Universidade Federal de Sao Paulo (UNIFESP), SP (Brazil). Escola Paulista de Medicina. Setor de Radioterapia]. E-mail: hrcs.dmed@unifesp.epm.br

    2004-04-01

    The objectives of this study were to assess the latent and active TGF{beta}1 localization in the heart, to evaluate whether or not radiation induces latent TGF{beta}1 activation, and to study the distribution of collagen fibers in the irradiated heart. Thirty-two C 57 BL mice were randomly assigned in two groups: GI (non irradiated animals) and GII (irradiated animals). The mice from G II received a single whole-body radiation dose of 7 Gy, using a {sup 60}Co source at a dose rate of 0.97 Gy/min. The animals were sacrificed by cervical dislocation at 1, 14, 30 and 90 days after irradiation. The irradiated hearts showed: nuclear changes and muscle cells with decreased striations; significant increase in the collagen deposition 90 days after irradiation; latent TGF{beta}1 activation in the cardio myocytes and connective tissue cells after irradiation. Our results show the importance of TGF{beta}1 protein in the process of radiation-induced heart fibrosis and suggest that cardio myocytes and connective cells may play a role in this mechanism acting as cellular sources of active TGF{beta}1. (author)

  16. Potential targets of transforming growth factor-beta1 during inhibition of oocyte maturation in zebrafish

    Directory of Open Access Journals (Sweden)

    Clelland Eric

    2005-09-01

    Full Text Available Abstract Background TGF-beta is a multifunctional growth factor involved in regulating a variety of cellular activities. Unlike mammals, the function of TGF-beta in the reproduction of lower vertebrates, such as fish, is not clear. Recently, we showed that TGF-beta1 inhibits gonadotropin- and 17alpha, 20beta-dihydroxyprogesterone (DHP-induced maturation in zebrafish. The aim of the present study was to investigate the mechanisms underlying this action. Method To determine if the effect of TGF-beta1 on oocyte maturation involves transcription and/or translation, ovarian follicles were pre-treated with actinomycin D, a blocker of transcription, and cyclohexamide, an inhibitor of translation, and incubated with hCG or DHP, either alone or in combination with TGF-beta1 and oocyte maturation scored. To determine the effect of TGF-beta1 on mRNA levels of several key effectors of oocyte maturation, three sets of experiments were performed. First, follicles were treated with control medium or TGF-beta1 for 2, 6, 12, and 24 h. Second, follicles were treated with different concentrations of TGF-beta1 (0 to 10 ng/ml for 18 h. Third, follicles were incubated with hCG in the absence or presence of TGF-beta1 for 18 h. At the end of each experiment, total RNA was extracted and reverse transcribed. PCR using primers specific for 20beta-hydroxysteroid dehydrogenase (20beta-HSD which is involved in DHP production, follicle stimulating hormone receptor (FSHR, luteinizing hormone receptor (LHR, the two forms of membrane progestin receptor: mPR-alpha and mPR-beta, as well as GAPDH (control, were performed. Results Treatment with actinomycin D, a blocker of transcription, reduced the inhibitory effect of TGF-beta1 on DHP-induced oocyte maturation, indicating that the inhibitory action of TGF-beta1 is in part due to regulation of gene transcription. Treatment with TGF-beta1 caused a dose and time-dependent decrease in mRNA levels of 20beta-HSD, LHR and mPR-beta in

  17. Curcumin inhibits cellular condensation and alters microfilament organization during chondrogenic differentiation of limb bud mesenchymal cells.

    Science.gov (United States)

    Kim, Dong Kyun; Kim, Song Ja; Kang, Shin Sung; Jin, Eun Jung

    2009-09-30

    Curcumin is a well known natural polyphenol product isolated from the rhizome of the plant Curcuma longa, anti-inflammatory agent for arthritis by inhibiting synthesis of inflammatory prostaglandins. However, the mechanisms by which curcumin regulates the functions of chondroprogenitor, such as proliferation, precartilage condensation, cytoskeletal organization or overall chondrogenic behavior, are largely unknown. In the present report, we investigated the effects and signaling mechanism of curcumin on the regulation of chondrogenesis. Treating chick limb bud mesenchymal cells with curcumin suppressed chondrogenesis by stimulating apoptotic cell death. It also inhibited reorganization of the actin cytoskeleton into a cortical pattern concomitant with rounding of chondrogenic competent cells and down-regulation of integrin beta1 and focal adhesion kinase (FAK) phosphorylation. Curcumin suppressed the phosphorylation of Akt leading to Akt inactivation. Activation of Akt by introducing a myristoylated, constitutively active form of Akt reversed the inhibitory actions of curcumin during chondrogenesis. In summary, for the first time, we describe biological properties of curcumin during chondrogenic differentiation of chick limb bud mesenchymal cells. Curcumin suppressed chondrogenesis by stimulating apoptotic cell death and down-regulating integrin-mediated reorganization of actin cytoskeleton via modulation of Akt signaling. PMID:19478554

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

    Directory of Open Access Journals (Sweden)

    Kan Saito

    Full Text Available 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.

  19. Inhibition of vimentin or B1 integrin reverts morphology of prostate tumor cells grown in laminin-rich extracellular matrix gels and reduces tumor growth in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Xueping; Fournier, Marcia V; Ware, Joy L; Bissell, Mina J; Yacoub, Adly; Zehner, Zendra E

    2008-06-12

    Prostate epithelial cells grown embedded in laminin-rich extracellular matrix (lrECM) undergo morphologic changes that closely resemble their architecture in vivo. In this study, growth characteristics of three human prostate epithelial sublines derived from the same cellular lineage, but displaying different tumorigenic and metastatic properties in vivo, were assessed in three-dimensional lrECM gels. M12, a highly tumorigenic and metastatic subline, was derived from the immortalized, prostate epithelial P69 cell line by selection in athymic, nude mice and found to contain a deletion of 19p-q13.1. The stable reintroduction of an intact human chromosome 19 into M12 resulted in a poorly tumorigenic subline, designated F6. When embedded in lrECM gels, the parental, nontumorigenic P69 line produced acini with clearly defined lumena. Immunostaining with antibodies to {beta}-catenin, E-cadherin, or {alpha}6 and {beta}1 integrins showed polarization typical of glandular epithelium. In contrast, the metastatic M12 subline produced highly disorganized cells with no evidence of polarization. The F6 subline reverted to acini-like structures exhibiting basal polarity marked with integrins. Reducing either vimentin levels via small interfering RNA interference or the expression of {alpha}6 and {beta}1 integrins by the addition of blocking antibodies, reorganized the M12 subline into forming polarized acini. The loss of vimentin significantly reduced M12-Vim tumor growth when assessed by s.c. injection in athymic mice. Thus, tumorigenicity in vivo correlated with disorganized growth in three-dimensional lrECM gels. These studies suggest that the levels of vimentin and {beta}1 integrin play a key role in the homeostasis of the normal acinus in prostate and that their dysregulation may lead to tumorigenesis. [Mol Cancer Ther 2009;8(3):499-508].

  20. Endoglin negatively regulates transforming growth factor beta1-induced profibrotic responses in intestinal fibroblasts.

    LENUS (Irish Health Repository)

    Burke, J P

    2012-02-01

    BACKGROUND: Fibroblasts isolated from strictures in Crohn\\'s disease (CD) exhibit reduced responsiveness to stimulation with transforming growth factor (TGF) beta1. TGF-beta1, acting through the smad pathway, is critical to fibroblast-mediated intestinal fibrosis. The membrane glycoprotein, endoglin, is a negative regulator of TGF-beta1. METHODS: Intestinal fibroblasts were cultured from seromuscular biopsies of patients undergoing intestinal resection for CD strictures or from control patients. Endoglin expression was assessed using confocal microscopy, flow cytometry and western blot. The effect of small interfering (si) RNA-mediated knockdown and plasmid-mediated overexpression of endoglin on fibroblast responsiveness to TGF-beta1 was assessed by examining smad phosphorylation, smad binding element (SBE) promoter activity, connective tissue growth factor (CTGF) expression and ability to contract collagen. RESULTS: Crohn\\'s stricture fibroblasts expressed increased constitutive cell-surface and whole-cell endoglin relative to control cells. Endoglin co-localized with filamentous actin. Fibroblasts treated with siRNA directed against endoglin exhibited enhanced TGF-beta1-mediated smad-3 phosphorylation, and collagen contraction. Cells transfected with an endoglin plasmid did not respond to TGF-beta1 by exhibiting SBE promoter activity or producing CTGF. CONCLUSION: Fibroblasts from strictures in CD express increased constitutive endoglin. Endoglin is a negative regulator of TGF-beta1 signalling in the intestinal fibroblast, modulating smad-3 phosphorylation, SBE promoter activity, CTGF production and collagen contraction.

  1. Two pine endo-beta-1,4-glucanases are associated with rapidly growing reproductive structures.

    Science.gov (United States)

    Loopstra, C A; Mouradov, A; Vivian-Smith, A; Glassick, T V; Gale, B V; Southerton, S G; Marshall, H; Teasdale, R D

    1998-03-01

    Two cDNA clones encoding endo-beta-1,4-glucanases (EGases) were isolated from a radiata pine (Pinus radiata) cDNA library prepared from immature female strobili. The cDNAs PrCel1 (inus adiata cellulase ) and PrCel2 encode proteins 509 and 515 amino acids in length, respectively, including putative signal peptides. Both proteins contain domains conserved in plant and bacterial EGases. The proteins PRCEL1 and PRCEL2 showed strong similarity to each other (76% amino acid identity), and higher similarity to TPP18 (73 and 67%, respectively), an EGase cloned from tomato (Lycopersicon esculentum) pistils, than to any other reported EGases. Northern-blot analyses indicated that both genes displayed a similar pattern of expression. The only significant difference was in the level of expression. In situ hybridizations were used to demonstrate that, within differentiating pine reproductive structures, PrCel1 expression was greatest in microsporangia in pollen strobili and near the developing ovule in the seed strobili. Expression was also found in vegetative tissues, especially in regions experiencing cell elongation, such as the elongating region of root tips. Both proteins have an ability to degrade carboxymethylcellulose in vitro. Genomic-blot analysis indicated the presence of a family of EGase genes in the radiata pine genome, and that PrCel1 and PrCel2 are transcribed from distinct one-copy genes. PMID:9501128

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

    International Nuclear Information System (INIS)

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

  3. Isolation of extracellular 28- and 42-kilodalton beta-1,3-glucanases and comparison of three beta-1,3-glucanases produced by Bacillus circulans IAM1165.

    OpenAIRE

    Aono, R; Hammura, M; Yamamoto, M.; Asano, T.

    1995-01-01

    Bacillus circulans IAM1165 produces three major extracellular beta-1,3-glucanases (molecular masses, 28, 42, and 91 kDa) during the stationary phase of growth. The 28- and 42-kDa enzymes were purified to homogeneity from the culture supernatant in this study. The properties of these two enzymes were examined, together with those of the 91-kDa enzyme previously isolated. The enzymatic properties of the 28- and 42-kDa beta-1,3-glucanases closely resemble each other. The enzymes belong to a cate...

  4. Effects of beta-1,3-glucan from Septoria tritici on structural defence responses in wheat

    DEFF Research Database (Denmark)

    Shetty, N.P.; Jensen, J.D.; Knudsen, A.;

    2009-01-01

    -1,3-glucanase and chitinase transcripts followed by a subsequent reduction in level. Resistance was also associated with high activity of beta-1,3-glucanase, especially in the apoplastic fluid, in accordance with the biotrophic/endophytic lifestyle of the pathogen in the apoplastic spaces, thus...... of callose. Collectively, these data indicate that resistance is dependent on a fast, initial recognition of the pathogen, probably due to beta-1,3-glucan in the fungal cell walls, and this results in the accumulation of beta-1,3-glucanase and structural defence responses, which may directly inhibit...... the pathogen and protect the host against fungal enzymes and toxins....

  5. Factor de crecimiento transformante beta-1: estructura, función y mecanismos de regulación en cáncer

    OpenAIRE

    Oscar Peralta Zaragoza; A. Lagunas Martínez; Vicente Madrid Marina

    2001-01-01

    Transforming growth factor beta-1 (TGF-b1) is produced by several cell lineages such as lymphocytes, macrophages, and dendritic cells, and its expression serves in both autocrine and paracrine modes to control the differentiation, proliferation, and state of activation of these and other cells. In general, TGF-b1 has pleiotropic properties on the immune response during the development of infection diseases and cancer; however, the mechanisms of action and regulation of gene expression of this...

  6. The antifibrotic effects of TGF-{beta}1 siRNA on hepatic fibrosis in rats

    Energy Technology Data Exchange (ETDEWEB)

    Lang, Qing; Liu, Qi [Key Laboratory of Molecular Biology for Infectious Diseases, Ministry of Education, Instituted for Virus Hepatitis and Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing (China); Xu, Ning [The Second Hospital of YuLin, Shanxi Province (China); Qian, Ke-Li; Qi, Jing-Hu; Sun, Yin-Chun; Xiao, Lang [Key Laboratory of Molecular Biology for Infectious Diseases, Ministry of Education, Instituted for Virus Hepatitis and Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing (China); Shi, Xiao-Feng, E-mail: sxff2003@yahoo.com.cn [Key Laboratory of Molecular Biology for Infectious Diseases, Ministry of Education, Instituted for Virus Hepatitis and Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing (China)

    2011-06-10

    Highlights: {yields} We constructed CCL4 induced liver fibrosis model successfully. {yields} We proofed that the TGF-{beta}1 siRNA had a definite therapy effect to CCL4 induced liver fibrosis. {yields} The therapy effect of TGF-{beta}1 siRNA had dose-dependent. -- Abstract: Background/aims: Hepatic fibrosis results from the excessive secretion of matrix proteins by hepatic stellate cells (HSCs), which proliferate during fibrotic liver injury. Transforming growth factor (TGF)-{beta}1 is the dominant stimulus for extracellular matrix (ECM) production by stellate cells. Our study was designed to investigate the antifibrotic effects of using short interference RNA (siRNA) to target TGF-{beta}1 in hepatic fibrosis and its mechanism in rats exposed to a high-fat diet and carbon tetrachloride (CCL4). Methods: A total of 40 healthy, male SD (Sprague-Dawley) rats were randomly divided into five even groups containing of eight rats each: normal group, model group, TGF-{beta}1 siRNA 0.125 mg/kg treatment group, TGF-{beta}1 siRNA 0.25 mg/kg treatment group and TGF-{beta}1 siRNA negative control group (0.25 mg/kg). CCL4 and a high-fat diet were used for 8 weeks to induce hepatic fibrosis. All the rats were then sacrificed to collect liver tissue samples. A portion of the liver samples were soaked in formalin for Hematoxylin-Eosin staining, classifying the degree of liver fibrosis, and detecting the expression of type I and III collagen and TGF-{beta}1; the remaining liver samples were stored in liquid nitrogen to be used for detecting TGF-{beta}1 by Western blotting and for measuring the mRNA expression of type I and III collagen and TGF-{beta}1 by quantitative real-time polymerase chain reaction. Results: Comparing the TGF-{beta}1 siRNA 0.25 mg/kg treatment group to the model group, the TGF-{beta}1 siRNA negative control group and the TGF-{beta}1 siRNA 0.125 mg/kg treatment group showed significantly reduced levels of pathological changes, protein expression and the m

  7. Integrin αIIb (CD41 plays a role in the maintenance of hematopoietic stem cell activity in the mouse embryonic aorta

    Directory of Open Access Journals (Sweden)

    Jean-Charles Boisset

    2013-04-01

    Integrins are transmembrane receptors that play important roles as modulators of cell behaviour through their adhesion properties and the initiation of signaling cascades. The αIIb integrin subunit (CD41 is one of the first cell surface markers indicative of hematopoietic commitment. αIIb pairs exclusively with β3 to form the αIIbβ3 integrin. β3 (CD61 also pairs with αv (CD51 to form the αvβ3 integrin. The expression and putative role of these integrins during mouse hematopoietic development is as yet unknown. We show here that hematopoietic stem cells (HSCs differentially express αIIbβ3 and αvβ3 integrins throughout development. Whereas the first HSCs generated in the aorta at mid-gestation express both integrins, HSCs from the placenta only express αvβ3, and most fetal liver HSCs do not express either integrin. By using αIIb deficient embryos, we show that αIIb is not only a reliable HSC marker but it also plays an important and specific function in maintaining the HSC activity in the mouse embryonic aorta.

  8. Symbiont-derived beta-1,3-glucanases in a social insect: mutualism beyond nutrition

    OpenAIRE

    Rosengaus, Rebeca B.; Schultheis, Kelley F.; Alla eYalonetskaya; Bulmer, Mark S.; William S DuComb; Benson, Ryan W.; John Paul eThottam; Veronica eGodoy-Carter

    2014-01-01

    Termites have had a long co-evolutionary history with prokaryotic and eukaryotic gut microbes. Historically, the role of these anaerobic obligate symbionts has been attributed to the nutritional welfare of the host. We provide evidence that protozoa (and/or their associated bacteria) colonizing the hindgut of the dampwood termite Zootermopsis angusticollis, synthesize multiple functional beta-1,3-glucanases, enzymes known for breaking down beta-1,3-glucans, the main component of fungal cell w...

  9. TM4SF5 suppression disturbs integrin α5-related signalling and muscle development in zebrafish.

    Science.gov (United States)

    Choi, Yoon-Ju; Kim, Hyun Ho; Kim, Jeong-Gyun; Kim, Hye-Jin; Kang, Minkyung; Lee, Mi-Sook; Ryu, Jihye; Song, Haeng Eun; Nam, Seo Hee; Lee, Doohyung; Kim, Kyu-Won; Lee, Jung Weon

    2014-08-15

    TM4SF5 (transmembrane 4 L six family member 5) is involved in EMT (epithelial-mesenchymal transition) for liver fibrosis and cancer metastasis; however, the function(s) of TM4SF5 during embryogenesis remains unknown. In the present study the effects of TM4SF5 on embryogenesis of zebrafish were investigated. tm4sf5 mRNA was expressed in the posterior somites during somitogenesis and in whole myotome 1 dpf (day post-fertilization). tm4sf5 suppression impaired development of the trunk with aberrant morphology of muscle fibres and altered expression of integrin α5. The arrangement and adhesion of muscle cells were abnormally disorganized in tm4sf5 morphants with reduced muscle fibre masses, where integrin α5-related signalling molecules, including fibronectin, FAK (focal adhesion kinase), vinculin and actin were aberrantly localized, compared with those in control fish. Aberrant muscle developments in tm4sf5 morphants were recovered by additional tm4sf5 or integrin α5 mRNA injection. Such a role for TM4SF5 was observed in the differentiation of C2C12 mouse myoblast cells to multinuclear muscle cells. Taken together, the results show that TM4SF5 controls muscle differentiation via co-operation with integrin α5-related signalling. PMID:24897542

  10. Human fallopian tube epithelium constitutively expresses integrin endometrial receptivity markers: no evidence for a tubal implantation window.

    Science.gov (United States)

    Brown, J K; Shaw, J L V; Critchley, H O D; Horne, A W

    2012-03-01

    Understanding of ectopic implantation within the Fallopian tube (FT) is limited. In the human uterus, the putative 'window of implantation' in the mid-luteal phase of the menstrual cycle is accompanied by increased endometrial epithelial expression of the integrins α(1)β(1), α(4)β(1) and α(v)β(3) and its ligand osteopontin. Similar cyclical changes in FT integrin expression have been proposed to contribute to ectopic implantation, but supporting data are limited. In the current study, we present quantitative data on human FT transcription and translation of the integrin subunits α(1), α(4), α(V), β(1) and β(3) during the follicular and mid-luteal phases of the menstrual cycle, together with a supporting immuocytochemical analysis of their spatial distribution within the FT, and that of osteopontin. In contrast to previous studies, our data indicate that all five integrin receptivity markers are constitutively transcribed and translated in the FT, with no evidence for changes in their expression or distribution during the window of implantation in the mid-luteal phase of the cycle. Furthermore, we could find no evidence for cyclic redistribution of the integrin α(v)β(3) ligand osteopontin within the FT. Although we do not rule out the involvement of integrin endometrial receptivity markers in the establishment of ectopic pregnancy, our findings do not support their differential expression during a tubal implantation window. PMID:22002573

  11. Discovery of platyhelminth-specific α/β-integrin families and evidence for their role in reproduction in Schistosoma mansoni.

    Directory of Open Access Journals (Sweden)

    Svenja Beckmann

    Full Text Available In all metazoa, the response of cells to molecular stimuli from their environment represents a fundamental principle of regulatory processes controlling cell growth and differentiation. Among the membrane-linked receptors mediating extracellular communication processes are integrin receptors. Besides managing adhesion to the extracellular matrix or to other cells, they arrange information flow into the cells by activating intracellular signaling pathways often acting synergistically through cooperation with growth factor receptors. Although a wealth of information exists on integrins in different model organisms, there is a big gap of knowledge for platyhelminths. Here we report on the in silico detection and reconstruction of α and β integrins from free-living and parasitic platyhelminths, which according to structural and phylogenetic analyses form specific clades separate from each other and from further metazoan integrins. As representative orthologs of parasitic platyhelminths we have cloned one beta-integrin (Smβ-Int1 and four alpha-integrins (Smα-Int1 - Smα-Int4 from Schistosoma mansoni; they were characterized by molecular and biochemical analyses. Evidence is provided that Smβ-Int1 interacts and co-localizes in the reproductive organs with known schistosome cellular tyrosine kinases (CTKs, of which the Syk kinase SmTK4 appeared to be the strongest interaction partner as shown by yeast two-hybrid analyses and coimmunoprecipitation experiments. By a novel RNAi approach with adult schistosomes in vitro we demonstrate for the first time multinucleated oocytes in treated females, indicating a decisive role Smβ-Int1 during oogenesis as phenotypically analyzed by confocal laser scanning microscopy (CLSM. Our findings provide a first comprehensive overview about platyhelminth integrins, of which the parasite group exhibits unique features allowing a clear distinction from the free-living groups. Furthermore, we shed first lights on the

  12. Endocytosis of Integrin-Binding Human Picornaviruses

    Directory of Open Access Journals (Sweden)

    Pirjo Merilahti

    2012-01-01

    Full Text Available Picornaviruses that infect humans form one of the largest virus groups with almost three hundred virus types. They include significant enteroviral pathogens such as rhino-, polio-, echo-, and coxsackieviruses and human parechoviruses that cause wide range of disease symptoms. Despite the economic importance of picornaviruses, there are no antivirals. More than ten cellular receptors are known to participate in picornavirus infection, but experimental evidence of their role in cellular infection has been shown for only about twenty picornavirus types. Three enterovirus types and one parechovirus have experimentally been shown to bind and use integrin receptors in cellular infection. These include coxsackievirus A9 (CV-A9, echovirus 9, and human parechovirus 1 that are among the most common and epidemic human picornaviruses and bind to αV-integrins via RGD motif that resides on virus capsid. In contrast, echovirus 1 (E-1 has no RGD and uses integrin α2β1 as cellular receptor. Endocytosis of CV-A9 has recently been shown to occur via a novel Arf6- and dynamin-dependent pathways, while, contrary to collagen binding, E-1 binds inactive β1 integrin and enters via macropinocytosis. In this paper, we review what is known about receptors and endocytosis of integrin-binding human picornaviruses.

  13. Integrins, muscle agrin and sarcoglycans during muscular inactivity conditions: an immunohistochemical study

    Directory of Open Access Journals (Sweden)

    G Anastasi

    2009-06-01

    Full Text Available Sarcoglycans are transmembrane proteins that seem to be functionally and pathologically as important as dystrophin. Sarcoglycans cluster together to form a complex, which is localized in the cell membrane of skeletal, cardiac, and smooth muscle. It has been proposed that the dystrophin-glycoprotein complex (DGC links the actin cytoskeleton with the extracellular matrix and the proper maintenance of this connection is thought to be crucial to the mechanical stability of the sarcolemma. The integrins are a family of heterodimeric cell surface receptors which play a crucial role in cell adhesion including cell-matrix and intracellular interactions and therefore are involved in various biological phenomena, including cell migration, and differentiation tissue repair. Sarcoglycans and integrins play a mechanical and signaling role stabilizing the systems during cycles of contraction and relaxation.Several studies suggested the possibility that integrins might play a role in muscle agrin signalling. On these basis, we performed an immunohistochemical analyzing sarcoglycans, integrins and agrin, on human skeletal muscle affected by sensitive-motor polyneuropathy, in order to better define the correlation between these proteins and neurogenic atrophy due to peripheral neuropathy. Our results showed the existence of a cascade mechanism which provoke a loss of regulatory effects of muscle activity on costameres, due to loss of muscle and neural agrin.This cascade mechanism could determine a quantitative modification of transmembrane receptors and loss of ?7B could be replaced and reinforced by enhanced expression of the ?7A integrin to restore muscle fiber viability. Second, it is possible that the reduced cycles of contraction and relaxation of muscle fibers, during muscular atrophy, provoke a loss of mechanical stresses transmitted over cell surface receptors that physically couple the cytoskeleton to extracellular matrix. Consequently, these mechanical

  14. Dietary Beta-1,3/1,6-Glucans Reduce Clinical Signs of Canine Atopy

    Directory of Open Access Journals (Sweden)

    A. C. Beynen

    2011-01-01

    Full Text Available Problem statement: There was evidence that beta-1,3/1,6-glucans modulate inflammatory activity. In an open, non-controlled trial, purified beta-1,3/1,6-glucans were found to improve the clinical signs of dogs with undefined chronic skin disorders. Given the design of that study, further work was required on the efficacy of beta-1,3/1,6-glucans in the treatment of canine atopy. Approach: The influence of a purified preparation of beta-1,3/1,6-glucans (MacroGard® on canine atopy was assessed in a double-blind, placebo-controlled trial. Privately owned dogs were used and the clinical signs of atopic dermatitis were evaluated by the owners. For a period of 8 weeks, the dogs daily received a complete dry food without (n = 16 or with 800 ppm beta-1,3/1,6-glucans (n = 15. During the trial, all dogs were treated three times with the use of a flea remedy in order to exclude any influence of flea-bite allergy. To assess the severity of atopic dermatitis, the clinical signs scored were itching, redness, scaling, thickening and stripping of skin. Results: For all five clinical signs, the group-mean improvement, expressed as change of severity score over time, was greater in the test group than in the controls. Within each group, the changes for the five clinical signs were added up to arrive at an overall index of improvement of atopic dermatitis. The extra improvement caused by the ingestion of beta-1,3/1,6-glucans was 63%. The difference between the pooled group-mean changes of the scores for the control and test dogs was statistically significant (PConclusion: Beta-1,3/1,6-glucans can be considered safe and it is put forward that a dose of 800 ppm in a dry food is beneficial for dogs with atopic dermatitis.

  15. IL-4 and TGF-beta 1 counterbalance one another while regulating mast cell homeostasis.

    Science.gov (United States)

    Macey, Matthew R; Sturgill, Jamie L; Morales, Johanna K; Falanga, Yves T; Morales, Joshua; Norton, Sarah K; Yerram, Nitin; Shim, Hoon; Fernando, Josephine; Gifillan, Alasdair M; Gomez, Gregorio; Schwartz, Lawrence; Oskeritzian, Carole; Spiegel, Sarah; Conrad, Daniel; Ryan, John J

    2010-05-01

    Mast cell responses can be altered by cytokines, including those secreted by Th2 and regulatory T cells (Treg). Given the important role of mast cells in Th2-mediated inflammation and recent demonstrations of Treg-mast cell interactions, we examined the ability of IL-4 and TGF-beta1 to regulate mast cell homeostasis. Using in vitro and in vivo studies of mouse and human mast cells, we demonstrate that IL-4 suppresses TGF-beta1 receptor expression and signaling, and vice versa. In vitro studies demonstrated that IL-4 and TGF-beta1 had balancing effects on mast cell survival, migration, and FcepsilonRI expression, with each cytokine cancelling the effects of the other. However, in vivo analysis of peritoneal inflammation during Nippostrongylus brasiliensis infection in mice revealed a dominant suppressive function for TGF-beta1. These data support the existence of a cytokine network involving the Th2 cytokine IL-4 and the Treg cytokine TGF-beta1 that can regulate mast cell homeostasis. Dysregulation of this balance may impact allergic disease and be amenable to targeted therapy. PMID:20304823

  16. Synergistic effects of 1,25-Dihydroxyvitamin D3 and TGF-beta1 on the production of insulin-like growth factor binding protein 3 in human bone marrow stromal cell cultures

    DEFF Research Database (Denmark)

    Kveiborg, Marie; Flyvbjerg, Allan; Kassem, M

    2002-01-01

    1,25-Dihydroxyvitamin D3 (calcitriol), transforming growth factor-beta (TGF-beta), and insulin-like growth factors (IGFs) are all important bone regulatory factors known to affect proliferation and differentiation of human bone-forming cells (osteoblasts). We have previously shown that TGF-beta1 ...

  17. HGF Accelerates Wound Healing by Promoting the Dedifferentiation of Epidermal Cells through β1-Integrin/ILK Pathway

    Directory of Open Access Journals (Sweden)

    Jin-Feng Li

    2013-01-01

    Full Text Available Skin wound healing is a critical and complex biological process after trauma. This process is activated by signaling pathways of both epithelial and nonepithelial cells, which release a myriad of different cytokines and growth factors. Hepatocyte growth factor (HGF is a cytokine known to play multiple roles during the various stages of wound healing. This study evaluated the benefits of HGF on reepithelialization during wound healing and investigated its mechanisms of action. Gross and histological results showed that HGF significantly accelerated reepithelialization in diabetic (DB rats. HGF increased the expressions of the cell adhesion molecules β1-integrin and the cytoskeleton remodeling protein integrin-linked kinase (ILK in epidermal cells in vivo and in vitro. Silencing of ILK gene expression by RNA interference reduced expression of β1-integrin, ILK, and c-met in epidermal cells, concomitantly decreasing the proliferation and migration ability of epidermal cells. β1-Integrin can be an important maker of poorly differentiated epidermal cells. Therefore, these data demonstrate that epidermal cells become poorly differentiated state and regained some characteristics of epidermal stem cells under the role of HGF after wound. Taken together, the results provide evidence that HGF can accelerate reepithelialization in skin wound healing by dedifferentiation of epidermal cells in a manner related to the β1-integrin/ILK pathway.

  18. Exercise-induced changes in circulating levels of transforming growth factor-beta-1 in humans

    DEFF Research Database (Denmark)

    Heinemeier, Katja; Langberg, Henning; Kjaer, Michael

    resting subjects (reported values range from 500 to 18,300 pg ml(-1)) and also the extent of intra-individual variation is unknown. As a basis for detecting exercise-induced changes in transforming growth factor-beta-1, we measured its concentration, by enzyme-linked immunosorbent assay, in plasma from...... eight healthy resting subjects. Plasma was sampled from each subject on five successive days according to a procedure designed to minimize activation of platelets, as platelet alpha-granules contain large amounts of transforming growth factor-beta-1. The mean plasma level was relatively low [1155 (30......Mechanical loading of cells induces the expression of transforming growth factor-beta-1, and acute exercise, which involves mechanical loading of several tissues, could thus increase its circulating level in humans. However, no consensus exists regarding the plasma concentration of this cytokine in...

  19. Integrin α6Bβ4 inhibits colon cancer cell proliferation and c-Myc activity

    International Nuclear Information System (INIS)

    Integrins are known to be important contributors to cancer progression. We have previously shown that the integrin β4 subunit is up-regulated in primary colon cancer. Its partner, the integrin α6 subunit, exists as two different mRNA splice variants, α6A and α6B, that differ in their cytoplasmic domains but evidence for distinct biological functions of these α6 splice variants is still lacking. In this work, we first analyzed the expression of integrin α6A and α6B at the protein and transcript levels in normal human colonic cells as well as colorectal adenocarcinoma cells from both primary tumors and established cell lines. Then, using forced expression experiments, we investigated the effect of α6A and α6B on the regulation of cell proliferation in a colon cancer cell line. Using variant-specific antibodies, we observed that α6A and α6B are differentially expressed both within the normal adult colonic epithelium and between normal and diseased colonic tissues. Proliferative cells located in the lower half of the glands were found to predominantly express α6A, while the differentiated and quiescent colonocytes in the upper half of the glands and surface epithelium expressed α6B. A relative decrease of α6B expression was also identified in primary colon tumors and adenocarcinoma cell lines suggesting that the α6A/α6B ratios may be linked to the proliferative status of colonic cells. Additional studies in colon cancer cells showed that experimentally restoring the α6A/α6B balance in favor of α6B caused a decrease in cellular S-phase entry and repressed the activity of c-Myc. The findings that the α6Bβ4 integrin is expressed in quiescent normal colonic cells and is significantly down-regulated in colon cancer cells relative to its α6Aβ4 counterpart are consistent with the anti-proliferative influence and inhibitory effect on c-Myc activity identified for this α6Bβ4 integrin. Taken together, these findings point out the importance of integrin

  20. Alterated integrin expression in lichen planopilaris

    Directory of Open Access Journals (Sweden)

    Erriquez Roberta

    2007-02-01

    Full Text Available Abstract Background Lichen planopilaris (LPP is an inflammatory disease characterized by a lymphomononuclear infiltrate surrounding the isthmus and infundibulum of the hair follicle of the scalp, that evolves into atrophic/scarring alopecia. In the active phase of the disease hairs are easily plucked with anagen-like hair-roots. In this study we focused on the expression of integrins and basement membrane components of the hair follicle in active LPP lesions. Methods Scalp biopsies were taken in 10 patients with LPP and in 5 normal controls. Using monoclonal antibodies against α3β1 and α6β4 integrins we showed the expression of these integrins and of the basement membrane components of the hair follicle in active LPP lesions and in healthy scalp skin. Results In the LPP involved areas, α3β1 was distributed in a pericellular pattern, the α6 subunit was present with a basolateral distribution while the β4 subunit showed discontinuous expression at the basal pole and occasionally, basolateral staining of the hair follicle. Conclusion: An altered distribution of the integrins in active LPP lesions can explain the phenomenon of easy pulling-out of the hair with a "gelatinous" root-sheath.

  1. Brucella beta 1,2 cyclic glucan is an activator of human and mouse dendritic cells

    OpenAIRE

    A Martirosyan; Perez-Gutierrez, C. (Camino); Banchereau, R; Dutartre, H; Lecine, P.; Dullaers, M. (Melissa); Mello, M.; Pinto, S; Muller, A; Leserman, L; Levy, Y.; Zurawski, G; Zurawski, S; Moreno, E; Moriyon, I

    2012-01-01

    Bacterial cyclic glucans are glucose polymers that concentrate within the periplasm of alpha-proteobacteria. These molecules are necessary to maintain the homeostasis of the cell envelope by contributing to the osmolarity of Gram negative bacteria. Here, we demonstrate that Brucella beta 1,2 cyclic glucans are potent activators of human and mouse dendritic cells. Dendritic cells activation by Brucella beta 1,2 cyclic glucans requires TLR4, MyD88 and TRIF, but not CD14. The Brucella cyclic glu...

  2. Integrin-mediated function of Rab GTPases in cancer progression

    Directory of Open Access Journals (Sweden)

    Alahari Suresh K

    2010-12-01

    Full Text Available Abstract The RAS (rat sarcoma superfamily of small GTPases is broadly subdivided into five groups: Ras, Rho, Rab, Ran, and Arf. Rab family proteins are important in regulating signal transduction and cellular processes such as differentiation, proliferation, vesicle transport, nuclear assembly, and cytoskeleton formation. However, some Rab proteins have been reported to be necessary for the adhesion and migration of cancer cells. Although Ras and Rho family members have been strongly implicated in cancer progression, knowledge of Rabs action in this regard is limited. Some reports have also linked Rab GTPases with cancer cell migration and invasiveness. This review discusses the implications of the involvement of Rabs in malignant transformation and cancer therapy through integrin-mediated signaling events, with particular emphasis on breast cancer.

  3. Effects of dietary beta-1,3-glucan on the growth, survival, physiological and immune response of marron, Cherax tenuimanus (smith, 1912).

    Science.gov (United States)

    Sang, Huynh Minh; Fotedar, Ravi

    2010-01-01

    Six isonitrogenous and isocalorific diets supplemented with five different levels of beta-1,3-glucan (0.08%, 0.1%, 0.2%, 0.4% and 0.8%) were formulated and tested for marron (Cherax tenuimanus) growth, survival, organosomatic indices, osmoregulatory capacity and immunological parameters (total and differential haemocyte counts, haemolymph clotting time and bacteraemia). The sixth diet without any beta-1,3-glucan was used as a control. Each diet was provided to 18 marron (0.47 +/- 0.02 g initial weight) replicated 3 times in individual 250 L fiberglass cylindrical tanks. Each tank was provided with a biological filtration recirculating water system. After 84 days of culture, the survival and yield were higher in the marron fed 0.1% beta glucan supplemented diet. The different levels of beta glucan did not alter any of the physiological parameters of marron. However, dietary supplementation with beta glucan resulted in significantly higher (P < 0.05) total haemocyte count (THC) and granular cells. The bacteraemia rank was lower in all diets having beta glucan supplemented with more than or equal to 0.1% compared to the control and 0.08% beta glucan supplemented diets. Results suggest that dietary beta-1,3-glucan at a minimum concentration of 0.1-0.2% can improve the immune system of marron. PMID:20139008

  4. ADAM12/syndecan-4 signaling promotes beta 1 integrin-dependent cell spreading through protein kinase Calpha and RhoA

    DEFF Research Database (Denmark)

    Thodeti, Charles Kumar; Albrechtsen, Reidar; Grauslund, Morten;

    2002-01-01

    The ADAMs (a disintegrin and metalloprotease) comprise a large family of multidomain proteins with cell-binding and metalloprotease activities. The ADAM12 cysteine-rich domain (rADAM12-cys) supports cell attachment using syndecan-4 as a primary cell surface receptor that subsequently triggers bet...

  5. Proinsulin C-peptide antagonizes the profibrotic effects of TGF-beta1 via up-regulation of retinoic acid and HGF-related signaling pathways.

    Science.gov (United States)

    Hills, Claire E; Willars, Gary B; Brunskill, Nigel J

    2010-04-01

    Novel signaling roles for C-peptide have recently been discovered with evidence that it can ameliorate complications of type 1 diabetes. Here we sought to identify new pathways regulated by C-peptide of relevance to the pathophysiology of diabetic nephropathy. Microarray analysis was performed to identify genes regulated by either C-peptide and/or TGF-beta1 in a human proximal tubular cell line, HK-2. Expression of retinoic acid receptor beta (RARbeta), hepatocyte growth factor (HGF), cellular retinoic acid-binding protein II (CRABPII), vimentin, E-cadherin, Snail, and beta-catenin was assessed by immunoblotting. The cellular localization of vimentin and beta-catenin was determined by immunocytochemistry. Changes in cell morphology were assessed by phase contrast microscopy. Gene expression profiling demonstrated differential expression of 953 and 1458 genes after C-peptide exposure for 18 h or 48 h, respectively. From these, members of the antifibrotic retinoic acid (RA)- and HGF-signaling pathways were selected. Immunoblotting demonstrated that C-peptide increased RARbeta, CRABPII, and HGF. We confirmed a role for RA in reversal of TGF-beta1-induced changes associated with epithelial-mesenchymal transition, including expression changes in Snail, E-cadherin, vimetin, and redistribution of beta-catenin. Importantly, these TGF-beta1-induced changes were inhibited by C-peptide. Further, effects of TGF-beta1 on Snail and E-cadherin expression were blocked by HGF, and inhibitory effects of C-peptide were removed by blockade of HGF activity. This study identifies a novel role for HGF as an effector of C-peptide, possibly via an RA-signaling pathway, highlighting C-peptide as a potential therapy for diabetic nephropathy. PMID:20197308

  6. The function of the human interferon-beta 1a glycan determined in vivo

    DEFF Research Database (Denmark)

    Dissing-Olesen, Lasse; Thaysen-Andersen, Morten; Meldgaard, Michael; Højrup, Peter; Finsen, Bente

    2008-01-01

    counterpart, due to a protein stabilization/solubilization effect of the glycan. It is interesting to note that the terminating sialic acids were essential for these effects. Conclusively, the structure/bioactivity relationship of rhIFN-beta1a was determined in vivo, and it provided a novel insight into the...

  7. Natalizumab plus interferon beta-1a reduces lesion formation in relapsing multiple sclerosis

    DEFF Research Database (Denmark)

    Radue, Ernst-Wilhelm; Stuart, William H; Calabresi, Peter A;

    2010-01-01

    The SENTINEL study showed that the addition of natalizumab improved outcomes for patients with relapsing multiple sclerosis (MS) who had experienced disease activity while receiving interferon beta-1a (IFNbeta-1a) alone. Previously unreported secondary and tertiary magnetic resonance imaging (MRI...

  8. Long-term impact of interferon beta-1b in patients with CIS

    DEFF Research Database (Denmark)

    Edan, G; Kappos, L; Montalbán, X;

    2014-01-01

    OBJECTIVE: To examine the long-term impact of early treatment initiation of interferon beta-1b (IFNB1b, Betaferon/Betaseron) in patients with a first event suggestive of multiple sclerosis (MS). METHODS: In the original placebo-controlled phase of BENEFIT, patients were randomised to IFNB1b 250 μg...

  9. NMR characterization of a 264-residue hyperthermostable endo-beta-1,3-glucanase.

    NARCIS (Netherlands)

    Ippel, J.H.; Koutsopoulos, S.; Nabuurs, S.M.; Berkel, W.J. van; Oost, J. van der; Mierlo, C.P. van

    2010-01-01

    Insight into the hyperthermostable endo-beta-1,3-glucanase pfLamA from Pyrococcus furiosus is obtained by using NMR spectroscopy. pfLamA functions optimally at 104 degrees C and recently the X-ray structure of pfLamA has been obtained at 20 degrees C, a temperature at which the enzyme is inactive. I

  10. Transgenic mice overexpressing the beta 1-adrenergic receptor in adipose tissue are resistant to obesity.

    Science.gov (United States)

    Soloveva, V; Graves, R A; Rasenick, M M; Spiegelman, B M; Ross, S R

    1997-01-01

    The ratio of alpha- to beta-receptors is thought to regulate the lipolytic index of adipose depots. To determine whether increasing the activity of the beta 1-adrenergic receptor (AR) in adipose tissue would affect the lipolytic rate or the development of this tissue, we used the enhancer-promoter region of the adipocyte lipid-binding protein (aP2) gene to direct expression of the human beta 1 AR cDNA to adipose tissue. Expression of the transgene was seen only in brown and white adipose tissue. Adipocytes from transgenic mice were more responsive to beta AR agonists than were adipocytes from nontransgenic mice, both in terms of cAMP production and lipolytic rates. Transgenic animals were partially resistant to diet-induced obesity. They had smaller adipose tissue depots than their nontransgenic littermates, reflecting decreased lipid accumulation in their adipocytes. In addition to increasing the lipolytic rate, overexpression of the beta 1 AR induced the abundant appearance of brown fat cells in subcutaneous white adipose tissue. These results demonstrate that the beta 1 AR is involved in both stimulation of lipolysis and the proliferation of brown fat cells in the context of the whole organism. Moreover, it appears that it is the overall beta AR activity, rather than the particular subtype, that controls these phenomena. PMID:8994185

  11. The journey of integrins and partners in a complex interactions landscape studied by super-resolution microscopy and single protein tracking.

    Science.gov (United States)

    Rossier, Olivier; Giannone, Grégory

    2016-04-10

    Cells adjust their adhesive and cytoskeletal organizations according to changes in the biochemical and physical nature of their surroundings. In return, by adhering and generating forces on the extracellular matrix (ECM) cells organize their microenvironment. Integrin-dependent focal adhesions (FAs) are the converging zones integrating biochemical and biomechanical signals arising from the ECM and the actin cytoskeleton. Thus, integrin-mediated adhesion and mechanotransduction, the conversion of mechanical forces into biochemical signals, are involved in critical cellular functions such as migration, proliferation and differentiation, and their deregulation contributes to pathologies including cancer. A challenging problem is to decipher how stochastic protein movements and interactions lead to formation of dynamic architecture such as integrin-dependent adhesive structures. In this review, we will describe recent advances made possible by super-resolution microscopies and single molecule tracking approaches that provided new understanding on the organization and the dynamics of integrins and intracellular regulators at the nanoscale in living cells. PMID:26571074

  12. Transforming growth factor beta 1-induced changes in cell migration, proliferation, and angiogenesis in the chicken chorioallantoic membrane

    OpenAIRE

    1990-01-01

    Application of TGF beta 1 (10-100 ng) to the chicken chorioallantoic membrane (CAM) for 72 h resulted in a dose-dependent, gross angiogenic response. The vascular effects induced by TGF beta 1 were qualitatively different than those induced by maximal doses of basic FGF (bFGF) (500 ng). While TGF beta 1 induced the formation of large blood vessels by 72 h, bFGF induced primarily small blood vessels. Histologic analysis revealed that TGF beta 1 stimulated pleiotropic cellular responses in the ...

  13. Tyrosine dephosphorylation of nuclear proteins mimics transforming growth factor beta 1 stimulation of alpha 2(I) collagen gene expression.

    OpenAIRE

    Greenwel, P; Hu, W.; Kohanski, R A; Ramirez, F.

    1995-01-01

    Transforming growth factor beta 1 (TGF-beta 1) exerts a positive effect on the transcription of genes coding for several extracellular matrix-related products, including collagen I. We have previously identified a strong TGF-beta 1-responsive element (TbRE) in the upstream promoter sequence of the alpha 2(I) collagen (COL1A2) gene. Our experiments have shown that TGF-beta 1 stimulates COL1A2 transcription by increasing binding of an Sp1-containing complex (TbRC) to the TbRE. They have also su...

  14. Patient satisfaction with the BETACONNECT™ autoinjector for interferon beta-1b

    Directory of Open Access Journals (Sweden)

    Weller I

    2015-07-01

    Full Text Available Ivonne Weller,1 Anna Saake,1 Thomas Schreiner,1 Julika Vogelreuter,1 Nicolas Petroff2 1Bayer Vital GmbH, Leverkusen, Germany; 2Vitartis Medizin-Service GmbH, Göttingen, Germany Purpose: Multiple sclerosis (MS is a chronic demyelinating, degenerative disease requiring long-term treatment. Patient adherence to treatment may be challenging in such scenarios, especially since treatment often involves self-injection, for example, with interferon beta-1b during therapy. BETACONNECT™ is a novel electronic autoinjector for patient support in interferon beta-1b administration. The purpose of this survey was to assess patient satisfaction with the BETACONNECT™ device and its features.Patients and methods: A total of 2,299 MS patients using the BETACONNECT™ device were asked to participate in a survey in October 2014. All of these candidates participated in the BETAPLUS® program and had provided written informed consent. The participants were asked to answer 13 device-related questions.Results: Of these candidates, 1,365 replied to the questionnaire, with more than 60% of the participants being 40–59 years of age. Among them, 69% were women and 21% were men (10% not specified. Approximately half of the participants received treatment with interferon beta-1b for more than 5 years. Most participants (85% had used self-injection devices before, with 59% previously using BETACOMFORT®, 23% using BETAJECT® Comfort, and 3% using BETAJECT® Lite, while less than 4% manually injected interferon beta-1b. The majority of the participants had received the BETACONNECT™ device from a BETAPLUS® nurse (87% and 48% had already used the device for more than 2 months (49% for 2 months or less. Among the participants, more than 90% evaluated the BETACONNECT™ device as “very helpful” or “helpful” in supporting their interferon beta-1b therapy with only marginal sex differences. Features that were rated “very important” by more than half of the

  15. α-Spectrin and integrins act together to regulate actomyosin and columnarization, and to maintain a monolayered follicular epithelium.

    Science.gov (United States)

    Ng, Bing Fu; Selvaraj, Gokul Kannan; Santa-Cruz Mateos, Carmen; Grosheva, Inna; Alvarez-Garcia, Ines; Martín-Bermudo, María Dolores; Palacios, Isabel M

    2016-04-15

    The spectrin cytoskeleton crosslinks actin to the membrane, and although it has been greatly studied in erythrocytes, much is unknown about its function in epithelia. We have studied the role of spectrins during epithelia morphogenesis using theDrosophilafollicular epithelium (FE). As previously described, we show that α-Spectrin and β-Spectrin are essential to maintain a monolayered FE, but, contrary to previous work, spectrins are not required to control proliferation. Furthermore, spectrin mutant cells show differentiation and polarity defects only in the ectopic layers of stratified epithelia, similar to integrin mutants. Our results identify α-Spectrin and integrins as novel regulators of apical constriction-independent cell elongation, asα-Spectrinand integrin mutant cells fail to columnarize. Finally, we show that increasing and reducing the activity of the Rho1-Myosin II pathway enhances and decreases multilayering ofα-Spectrincells, respectively. Similarly, higher Myosin II activity enhances the integrin multilayering phenotype. This work identifies a primary role for α-Spectrin in controlling cell shape, perhaps by modulating actomyosin. In summary, we suggest that a functional spectrin-integrin complex is essential to balance adequate forces, in order to maintain a monolayered epithelium. PMID:26952981

  16. Force via integrins but not E-cadherin decreases Oct3/4 expression in embryonic stem cells

    International Nuclear Information System (INIS)

    Highlights: ► Force via integrins or cadherins induces similar cell stiffening responses. ► Force via integrins but not cadherins induces cell spreading. ► Force via integrins but not cadherins induces differentiation of embryonic stem cells. -- Abstract: Increasing evidence suggests that mechanical factors play a critical role in fate decisions of stem cells. Recently we have demonstrated that a local force applied via Arg-Gly-Asp (RGD) peptides coated magnetic beads to mouse embryonic stem (ES) cells increases cell spreading and cell stiffness and decreases Oct3/4 (Pou5f1) gene expression. However, it is not clear whether the effects of the applied stress on these functions of ES cells can be extended to natural extracellular matrix proteins or cell–cell adhesion molecules. Here we show that a local cyclic shear force applied via fibronectin or laminin to integrin receptors increased cell spreading and stiffness, downregulated Oct3/4 gene expression, and decreased cell proliferation rate. In contrast, the same cyclic force applied via cell–cell adhesion molecule E-cadherin (Cdh1) had no effects on cell spreading, Oct3/4 gene expression, and the self-renewal of mouse ES cells, but induced significant cell stiffening. Our findings demonstrate that biological responses of ES cells to force applied via integrins are different from those to force via E-cadherin, suggesting that mechanical forces might play different roles in different force transduction pathways to shape early embryogenesis.

  17. Direct interaction of the kringle domain of urokinase-type plasminogen activator (uPA) and integrin alpha v beta 3 induces signal transduction and enhances plasminogen activation.

    Science.gov (United States)

    Tarui, Takehiko; Akakura, Nobuaki; Majumdar, Mousumi; Andronicos, Nicholas; Takagi, Junichi; Mazar, Andrew P; Bdeir, Khalil; Kuo, Alice; Yarovoi, Serge V; Cines, Douglas B; Takada, Yoshikazu

    2006-03-01

    It has been questioned whether there are receptors for urokinase-type plasminogen activator (uPA) that facilitate plasminogen activation other than the high affinity uPA receptor (uPAR/CD87) since studies of uPAR knockout mice did not support a major role of uPAR in plasminogen activation. uPA also promotes cell adhesion, chemotaxis, and proliferation besides plasminogen activation. These uPA-induced signaling events are not mediated by uPAR, but mediated by unidentified, lower-affinity receptors for the uPA kringle. We found that uPA binds specifically to integrin alpha v beta 3 on CHO cells depleted of uPAR. The binding of uPA to alpha v beta 3 required the uPA kringle domain. The isolated uPA kringle domain binds specifically to purified, recombinant soluble, and cell surface alpha v beta 3, and other integrins (alpha 4 beta 1 and alpha 9 beta 1), and induced migration of CHO cells in an alpha v beta 3-dependent manner. The binding of the uPA kringle to alpha v beta 3 and uPA kringle-induced alpha v beta 3-dependent cell migration were blocked by homologous plasminogen kringles 1-3 or 1-4 (angiostatin), a known integrin antagonist. We studied whether the binding of uPA to integrin alpha v beta 3 through the kringle domain plays a role in plasminogen activation. On CHO cell depleted of uPAR, uPA enhanced plasminogen activation in a kringle and alpha v beta 3-dependent manner. Endothelial cells bound to and migrated on uPA and uPA kringle in an alpha v beta 3-dependent manner. These results suggest that uPA binding to integrins through the kringle domain plays an important role in both plasminogen activation and uPA-induced intracellular signaling. The uPA kringle-integrin interaction may represent a novel therapeutic target for cancer, inflammation, and vascular remodeling. PMID:16525582

  18. β1 Integrins Mediate Mechanosensitive Signaling Pathways in Osteocytes

    OpenAIRE

    Litzenberger, Julie B.; Tummala, Padmaja; Kim, Jae-Beom; Jacobs, Christopher R.

    2010-01-01

    Integrins are cell-substrate adhesion proteins that initiate intracellular signaling and may serve as mechanosensors in bone. MLO-Y4 cells were stably transfected with a dominant negative form of the β1 integrin subunit (β1DN) containing the transmembrane domain and cytoplasmic tail of β1 integrin. Cells expressing β1DN had reduced vinculin localization to focal contacts but no change in intracellular actin organization. When exposed to oscillatory fluid flow, β1DN cells exhibited a significa...

  19. Traction forces mediated by integrin signaling are necessary for definitive endoderm specification.

    Science.gov (United States)

    Taylor-Weiner, Hermes; Ravi, Neeraja; Engler, Adam J

    2015-05-15

    Pluripotent embryonic stem cells (ESCs) exert low-traction forces on their niche in vitro whereas specification to definitive endoderm in vivo coincides with force-mediated motility, suggesting a differentiation-mediated switch. However, the onset of contractility and extent to which force-mediated integrin signaling regulates fate choices is not understood. To address the requirement of tractions forces for differentiation, we examined mouse embryonic stem cell (ESC) specification towards definitive endoderm on fibrillar fibronectin containing a deformation-sensitive FRET probe. Inhibiting contractility resulted in an increase in the observed fibronectin FRET intensity ratio but also decreased the amount of phosphorylated nuclear SMAD2, leading to reduced expression of the definitive endoderm marker SOX17. By contrast ESCs maintained in pluripotency medium did not exert significant tractions against the fibronectin matrix. When laminin-111 was added to fibrillar matrices to improve the efficiency of definitive endoderm induction, ESCs decreased their fibronectin traction forces in a laminin-dependent manner; blocking the laminin-binding α3-integrin restored fibronectin matrix deformation and reduced SOX17 expression and SMAD2 phosphorylation, probably because of compensation of inhibitory signaling from SMAD7 after 5 days in culture. These data imply that traction forces and integrin signaling are important regulators of early fate decisions in ESCs. PMID:25908864

  20. Expression of VLA-integrins and their related basement membrane ligands in gingiva from patients of various periodontitis categories

    DEFF Research Database (Denmark)

    Gürses, N.; Thorup, Alis Karabulut; Reibel, J.; Carter, G.W.; Holmstrup, Palle

    integrins, basement membrane, gingiva, periodontitis, periodontal disease activity immunofluorescence......integrins, basement membrane, gingiva, periodontitis, periodontal disease activity immunofluorescence...

  1. Inhibition of integrin α2β1 ameliorates glomerular injury.

    Science.gov (United States)

    Borza, Corina M; Su, Yan; Chen, Xiwu; Yu, Ling; Mont, Stacey; Chetyrkin, Sergei; Voziyan, Paul; Hudson, Billy G; Billings, Paul C; Jo, Hyunil; Bennett, Joel S; Degrado, William F; Eckes, Beate; Zent, Roy; Pozzi, Ambra

    2012-06-01

    Mesangial cells and podocytes express integrins α1β1 and α2β1, which are the two major collagen receptors that regulate multiple cellular functions, including extracellular matrix homeostasis. Integrin α1β1 protects from glomerular injury by negatively regulating collagen production, but the role of integrin α2β1 in renal injury is unclear. Here, we subjected wild-type and integrin α2-null mice to injury with adriamycin or partial renal ablation. In both of these models, integrin α2-null mice developed significantly less proteinuria and glomerulosclerosis. In addition, selective pharmacological inhibition of integrin α2β1 significantly reduced adriamycin-induced proteinuria, glomerular injury, and collagen deposition in wild-type mice. This inhibitor significantly reduced collagen synthesis in wild-type, but not integrin α2-null, mesangial cells in vitro, demonstrating that its effects are integrin α2β1-dependent. Taken together, these results indicate that integrin α2β1 contributes to glomerular injury by positively regulating collagen synthesis and suggest that its inhibition may be a promising strategy to reduce glomerular injury and proteinuria. PMID:22440900

  2. Flavobacterium sp. strain 4221 and Pedobacter sp. strain 4236 beta-1,3-glucanases that are active at low temperatures

    DEFF Research Database (Denmark)

    Rasmussen, Mikkel A.; Madsen, Søren; Stougaard, Peter; Johnsen, Mads Grønvald

    2008-01-01

    Secretion of beta-1,3-glucanases by the arctic bacterial isolates 4221 and 4236, related to the genera Flavobacterium and Pedobacter, was discovered. Escherichia coli and Lactococcus lactis expression of beta-1,3-glucanases Glc4221-1 and Glc4236-1 from the respective isolates was achieved. The en...

  3. Leukocyte arrest: Biomechanics and molecular mechanisms of β2 integrin activation

    Science.gov (United States)

    Fan, Zhichao; Ley, Klaus

    2016-01-01

    Integrins are a group of heterodimeric transmembrane receptors that play essential roles in cell–cell and cell–matrix interaction. Integrins are important in many physiological processes and diseases. Integrins acquire affinity to their ligand by undergoing molecular conformational changes called activation. Here we review the molecular biomechanics during conformational changes of integrins, integrin functions in leukocyte biorheology (adhesive functions during rolling and arrest) and molecules involved in integrin activation. PMID:26684674

  4. Role of TGF-beta1 in relation to exercise-induced type I collagen synthesis in human tendinous tissue

    DEFF Research Database (Denmark)

    Heinemeier, Katja; Langberg, Henning; Olesen, Jens L;

    2003-01-01

    Mechanical loading of tissue is known to influence local collagen synthesis, and microdialysis studies indicate that mechanical loading of human tendon during exercise elevates tendinous type I collagen production. Transforming growth factor-beta1 (TGF-beta1), a potent stimulator of type I collagen...... synthesis, is released from cultured tendon fibroblasts in response to mechanical loading. Thus TGF-beta1 could link mechanical loading and collagen synthesis in tendon tissue in vivo. Tissue levels of TGF-beta1 and type I collagen metabolism markers [procollagen I COOH-terminal propeptide (PICP) and COOH......-terminal telopeptide of type I collagen (ICTP)] were measured by microdialysis in peritendinous tissue of the Achilles' tendon in six male volunteers before and after treadmill running (1 h, 12 km/h, 3% uphill). In addition, blood levels of TGF-beta1, PICP, and ICTP were obtained. PICP levels increased 68 h after...

  5. Symbiont-derived beta-1,3-glucanases in a social insect: mutualism beyond nutrition

    Directory of Open Access Journals (Sweden)

    Rebeca B Rosengaus

    2014-11-01

    Full Text Available Termites have had a long co-evolutionary history with prokaryotic and eukaryotic gut microbes. Historically, the role of these anaerobic obligate symbionts has been attributed to the nutritional welfare of the host. We provide evidence that protozoa (and/or their associated bacteria colonizing the hindgut of the dampwood termite Zootermopsis angusticollis, synthesize multiple functional beta-1,3-glucanases, enzymes known for breaking down beta-1,3-glucans, the main component of fungal cell walls. These enzymes, we propose, may help in both digestion of ingested fungal hyphae and protection against invasion by fungal pathogens. This research points to an additional novel role for the mutualistic hindgut microbial consortia of termites, an association that may extend beyond ligno-cellulolytic activity and nitrogen fixation to include a reduction in the risks of mycosis at both the individual- and colony-levels while nesting in and feeding on microbial-rich decayed wood.

  6. Can alterations in integrin and laminin-5 expression be used as markers of malignancy?

    DEFF Research Database (Denmark)

    Thorup, Alis Karabulut; Reibel, J.; Schjødt, Morten;

    1998-01-01

    Integrins, laminin-5, cell adhesion molecules, oral, leukoplakia, premalignant, squamous cell carcinomas......Integrins, laminin-5, cell adhesion molecules, oral, leukoplakia, premalignant, squamous cell carcinomas...

  7. Interaction of interleukin-6 and transforming growth factor-beta1 pathways in prostate epithelial cells

    Czech Academy of Sciences Publication Activity Database

    Staršíchová, Andrea; Lincová, Eva; Pernicová, Zuzana; Kozubík, Alois; Souček, Karel

    2009-01-01

    Roč. 276, č. 1 (2009), s. 253. ISSN 1742-464X. [34th FEBS Congress. 04.07.2009-09.07.2009, Prague] R&D Projects: GA ČR(CZ) GA204/07/0834 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : transforming growth factor-beta1 * interleukin -6 * prostate Subject RIV: BO - Biophysics

  8. Endothelin-1 activates phospholipase D and thymidine incorporation in fibroblasts overexpressing protein kinase C beta 1.

    OpenAIRE

    Pai, J K; Dobek, E A; Bishop, W R

    1991-01-01

    Endothelins (ETs) are a family of extremely potent vasoconstrictor peptides. In addition, ET-1 acts as a potent mitogen and activates phospholipase C in smooth muscle cells and fibroblasts. We examined the effects of ET-1 on phosphatidylcholine (PC) metabolism and thymidine incorporation in control Rat-6 fibroblasts and in cells that overexpress protein kinase C beta 1 (PKC). PC pools were labeled with [3H]myristic acid, and formation of phosphatidylethanol (PEt), an unambiguous marker of pho...

  9. Renal thrombotic microangiopathy caused by interferon beta-1a treatment for multiple sclerosis

    Directory of Open Access Journals (Sweden)

    Mahe J

    2013-08-01

    Full Text Available Julien Mahe,1 Aurélie Meurette,2 Anne Moreau,3 Caroline Vercel,2 Pascale Jolliet1,4 1Clinical Pharmacology Department, Institute of Biology, University Hospital, Nantes, France; 2Clinical Nephrology and Immunology Department, University Hospital, Nantes, France; 3Laboratory of Pathology, University Hospital, Nantes, France; 4EA 4275 Biostatistics, Pharmacoepidemiology and Subjective Measures in Health Sciences, University of Nantes, Nantes, France Abstract: Interferon beta-1a is available as an immunomodulating agent for relapsing forms of multiple sclerosis. Common side effects include flu-like symptoms, asthenia, anorexia, and administration site reaction. Kidney disorders are rarely reported. In this study we describe the case of a woman who has been undergoing treatment with interferon beta-1a for multiple sclerosis for 5 years. She developed a hemolytic-uremic syndrome with intravascular hemolysis in a context of severe hypertension. A kidney biopsy showed a thrombotic microangiopathy. This observation highlights an uncommon side effect of long-term interferon beta-1a therapy. Pathophysiological mechanisms leading to this complication might be explained by the antiangiogenic activity of interferon. Keywords: thrombotic microangiopathy, interferon beta, hemolytic-uremic syndrome, antiangiogenic activity

  10. Crosstalk between Fibroblast Growth Factor (FGF Receptor and Integrin through Direct Integrin Binding to FGF and Resulting Integrin-FGF-FGFR Ternary Complex Formation

    Directory of Open Access Journals (Sweden)

    Seiji Mori

    2013-08-01

    Full Text Available Fibroblast growth factors (FGFs play a critical role in diverse physiological processes and the pathogenesis of diseases. Integrins are involved in FGF signaling, since integrin antagonists suppress FGF signaling. This is called integrin-FGF crosstalk, while the specifics of the crosstalk are unclear. This review highlights recent findings that FGF1 directly interacts with integrin αvβ3, and the resulting integrin-FGF-fibroblast growth factor receptor (FGFR ternary complex formation is essential for FGF1-induced cell proliferation, migration and angiogenesis. An integrin-binding defective FGF1 mutant (Arg-50 to Glu, R50E is defective in ternary complex formation and in inducing cell proliferation, migration and angiogenesis, while R50E still binds to the FGF receptor and heparin. In addition, R50E suppressed tumorigenesis in vivo, while wild-type (WT FGF1 enhanced it. Thus, the direct interaction between FGF1 and integrin αvβ3 is a potential therapeutic target, and R50E is a potential therapeutic agent.

  11. Functional consequences of integrin gene mutations in mice

    DEFF Research Database (Denmark)

    Bouvard, D; Brakebusch, C; Gustafsson, E; Aszódi, A; Bengtsson, T; Berna, A; Fässler, R

    2001-01-01

    Integrins are cell-surface receptors responsible for cell attachment to extracellular matrices and to other cells. The application of mouse genetics has significantly increased our understanding of integrin function in vivo. In this review, we summarize the phenotypes of mice carrying mutant inte...

  12. TGF-beta 1 attenuates myocardial ischemia-reperfusion injury via inhibition of upregulation of MMP-1.

    Science.gov (United States)

    Chen, Hongjiang; Li, Dayuan; Saldeen, Tom; Mehta, Jawahar L

    2003-05-01

    Ischemia-reperfusion (I/R) is thought to upregulate the expression and activity of matrix metalloproteinases (MMPs), which regulate myocardial and vascular remodeling. Previous studies have shown that transforming growth factor-beta(1) (TGF-beta(1)) can attenuate myocardial injury induced by I/R. TGF-beta(1) is also reported to suppress the release of MMPs. To study the modulation of MMP-1 by TGF-beta(1) in I/R myocardium, Sprague-Dawley rats were given saline and subjected to 1 h of myocardial ischemia [total left coronary artery (LCA) ligation] followed by 1 h of reperfusion (n = 9). Parallel groups of rats were pretreated with recombinant TGF-beta(1) (rTGF-beta(1), 1 mg/rat, n = 9) before reperfusion or exposure to sham I/R (control group). I/R caused myocardial necrosis and dysfunction, indicated by decreased first derivative of left ventricular pressure, mean arterial blood pressure, and heart rate (all P injury and death of cultured myocytes, measured as lactate dehydrogenase release and trypan blue staining, in a dose- and time-dependent manner (P injury and death induced by active MMP-1. The present study for the first time shows that MMP-1 can directly cause myocyte injury or death and that attenuation of myocardial I/R injury by TGF-beta(1) may, at least partly, be mediated by the inhibition of upregulation of MMP-1. PMID:12679326

  13. On gaps in Rényi $\\beta$-expansions of unity for $\\beta > 1$ an algebraic number.

    OpenAIRE

    Verger-Gaugry, Jean-Louis

    2006-01-01

    Let $\\beta> 1$ be an algebraic number. We study the strings of zeros (“gaps”) in the Rényi $\\beta$ -expansion $d_{\\beta}(1)$ of unity which controls the set $\\mathbb{Z}_{\\beta}$ of $\\beta$-integers. Using a version of Liouville's inequality which extends Mahler's and Güting's approximation theorems, the strings of zeros in $d_{\\beta}(1)$ are shown to exhibit a “gappiness” asymptotically bounded above by $log(M(\\beta ))/ log(\\beta)$, where $M(\\beta)$ is the Mahler measure of $\\beta$ . The proo...

  14. Chain specificity assignment of monoclonal antibodies to human laminins by using recombinant laminin beta1 and gamma1 chains.

    Science.gov (United States)

    Geberhiwot, T; Wondimu, Z; Salo, S; Pikkarainen, T; Kortesmaa, J; Tryggvason, K; Virtanen, I; Patarroyo, M

    2000-05-01

    In the present study, the chain specificity of 16 commonly used monoclonal antibodies to human laminin(s) was analysed by using recombinant laminin beta1 and gamma1 chains. By ELISA, all antibodies reacted with purified placenta laminin, and most antibodies recognised either recombinant beta1 or gamma1 chains. Reactivity and chain specificity was confirmed against the recombinant chains in Western blotting under non-reducing conditions, and only a few antibodies were reactive under reducing conditions. Most antibodies were able to immunoprecipitate associated laminin beta1/gamma1 chains from platelet lysates. Based on these results and data from the literature, a tentative epitope map is presented. PMID:10842099

  15. Lamellipodial tension, not integrin/ligand binding, is the crucial factor to realise integrin activation and cell migration.

    Science.gov (United States)

    Schulte, Carsten; Ferraris, Gian Maria Sarra; Oldani, Amanda; Galluzzi, Massimiliano; Podestà, Alessandro; Puricelli, Luca; de Lorenzi, Valentina; Lenardi, Cristina; Milani, Paolo; Sidenius, Nicolai

    2016-01-01

    The molecular clutch (MC) model proposes that actomyosin-driven force transmission permits integrin-dependent cell migration. To investigate the MC, we introduced diverse talin (TLN) and integrin variants into Flp-In™ T-Rex™ HEK293 cells stably expressing uPAR. Vitronectin variants served as substrate providing uPAR-mediated cell adhesion and optionally integrin binding. This particular system allowed us to selectively analyse key MC proteins and interactions, effectively from the extracellular matrix substrate to intracellular f-actin, and to therewith study mechanobiological aspects of MC engagement also uncoupled from integrin/ligand binding. With this experimental approach, we found that for the initial PIP2-dependent membrane/TLN/f-actin linkage and persistent lamellipodia formation the C-terminal TLN actin binding site (ABS) is dispensable. The establishment of an adequate MC-mediated lamellipodial tension instead depends predominantly on the coupling of this C-terminal TLN ABS to the actomyosin-driven retrograde actin flow force. This lamellipodial tension is crucial for full integrin activation eventually determining integrin-dependent cell migration. In the integrin/ligand-independent condition the frictional membrane resistance participates to these processes. Integrin/ligand binding can also contribute but is not necessarily required. PMID:26616200

  16. IFN beta 1a as Glucocorticoids-Sparing Therapy in a Patient with CLIPPERS

    Science.gov (United States)

    Rico, María; Villafani, Javier; Tuñón, Alberto; Mateos, Valentín; Oliva-Nacarino, Pedro

    2016-01-01

    Patient: Male, 31 Final Diagnosis: CLIPPERS Symptoms: Ataxia • diplopia Medication: IFNbeta 1a Clinical Procedure: — Specialty: Neurology Objective: Rare disease Background: Chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids (CLIPPERS) is a recently described inflammatory disease of the central nervous system, distinguished by brainstem- and spinal cord-centered lesions with a characteristic contrast enhancement on MRI, a lymphocytic perivascular infiltrate on pathological exam, and a dramatic response to and dependence on steroids therapy. Since its initial description in 2010, different glucocorticoid-sparing agents, mostly immunosuppressant drugs, have been used to minimize the dosage, but these therapies also carry the risk of important secondary effects. We present the first reported case of CLIPPERS treated with interferon beta 1a as add-on therapy. Case Report: A previously healthy 31-year-old man presented with gait ataxia and dysarthria. MRI showed pons-centered hyperintense patchy lesions on T2-weighted images. Additional tests ruled out other possible diagnoses and symptoms reversed with intravenous methylprednisolone. Over the years the patient presented with several episodes of deterioration each year, which were partly reversed with glucocorticoid therapy, but leaving him with growing sequelae. Four years after the initial event, treatment with interferon-beta-1a was initiated, achieving reduced frequency of the relapses to 1 every 4 years, which were no longer associated to increasing disability. This allowed reducing glucocorticoids to 30 mg of Deflazacort every other day. Conclusions: Interferon beta-1a could be an alternative to corticosteroid-combined therapy in CLIPPERS and its more benign profile of secondary effects compared to immunosuppressants could make it an attractive choice. PMID:26813773

  17. Exposure to particulates, microorganisms, beta(1-3)-glucans, and endotoxins during soybean harvesting.

    Science.gov (United States)

    Roy, Chad J; Thorne, Peter S

    2003-01-01

    Sclerotinia sclerotiorum is an emerging fungal pathogen affecting soybeans in the United States. In response to its emergence, exposures to particulates, bioaerosols, endotoxins, S. sclerotiorum, and beta(1-3)-glucans were characterized during soybean harvesting. Air sampling was performed on soybean harvesters (combines) and on the farmers in closed cabs as personal samples during harvesting at 17 farms in 1997 and repeated at 15 in 1998. S. sclerotiorum infestation was evident in the fields at 8 of the sites (44%). The geometric mean concentrations (and geometric standard deviations) measured on the combines in 1998 were as follows: total dust, 11.9 (2.8) mg/m(3); inhalable dust 11.7 (6.4) mg/m(3); and beta(1-3)-glucans, 5027 (7) ng/m(3). Values for the personal samples in 1998 were as follows: total dust, 1.2 (6.7) mg/m(3); inhalable dust, 1.1 (5.3); and beta(1-3)-glucans, 674 (9) ng/m(3). These concentrations were two- to threefold higher than in the previous year. Ambient endotoxin concentrations were 4438 EU/m(3) in Year I and 459 EU/m(3) in Year II. Particle size distribution measurements on the combines yielded mass median aerodynamic diameters of 6.6 microm on the combine and 4.0 microm inside the combine cab. Closed combine cabs provided an average workplace protection factor of 11.7 for total dust. Nevertheless, personal exposures to organisms inside combine cabins ranged from 3.6 x 10(4) to 4.0 x 10(8) organisms/m(3). These data indicate the potential exists for high exposures to organic dust and bioaerosols during soybean harvesting. PMID:12908864

  18. Research of TGF-beta1 Inducing Lung Adencarcinoma PC9 Cells to Mesenchymal Cells Transition

    OpenAIRE

    Chen, Xiaofeng; Wang, Heyong; Zhang, Lei; Zhang, Huijun

    2010-01-01

    Background and objective It has been proven that epithelial-mesenchymal transition (EMT) not only correlated with embryonic development but also could promote tumor invasion and metastasis. Transforming growth factor beta-1 (TGF-β1) has been identified as the main inducer of tumor EMT. The aim of this study was to investigate the effects of TGF-β1 on EMT and PI3K/AKT signaling pathway in lung adencarcinoma PC9 cells. Methods Cultured PC9 cells were treated with different concentrations of TGF...

  19. VLA-4 integrin concentrates at the peripheral supramolecular activation complex of the immune synapse and drives T helper 1 responses

    Science.gov (United States)

    Mittelbrunn, María; Molina, Ana; Escribese, María M.; Yáñez-Mó, María; Escudero, Ester; Ursa, Ángeles; Tejedor, Reyes; Mampaso, Francisco; Sánchez-Madrid, Francisco

    2004-07-01

    The integrin 41 (VLA-4) not only mediates the adhesion and transendothelial migration of leukocytes, but also provides costimulatory signals that contribute to the activation of T lymphocytes. However, the behavior of 41 during the formation of the immune synapse is currently unknown. Here, we show that 41 is recruited to both human and murine antigen-dependent immune synapses, when the antigen-presenting cell is a B lymphocyte or a dendritic cell, colocalizing with LFA-1 at the peripheral supramolecular activation complex. However, when conjugates are formed in the presence of anti-4 antibodies, VLA-4 colocalizes with the CD3- chain at the center of the synapse. In addition, antibody engagement of 4 integrin promotes polarization toward a T helper 1 (Th1) response in human in vitro models of CD4+ T cell differentiation and naïve T cell priming by dendritic cells. The in vivo administration of anti-4 integrin antibodies also induces an immune deviation to Th1 response that dampens a Th2-driven autoimmune nephritis in Brown Norway rats. These data reveal a regulatory role of 4 integrins on T lymphocyte-antigen presenting cell cognate immune interactions.

  20. FAK, talin and PIPKIγ regulate endocytosed integrin activation to polarize focal adhesion assembly.

    Science.gov (United States)

    Nader, Guilherme P F; Ezratty, Ellen J; Gundersen, Gregg G

    2016-05-01

    Integrin endocytic recycling is critical for cell migration, yet how recycled integrins assemble into new adhesions is unclear. By synchronizing endocytic disassembly of focal adhesions (FAs), we find that recycled integrins reassemble FAs coincident with their return to the cell surface and dependent on Rab5 and Rab11. Unexpectedly, endocytosed integrins remained in an active but unliganded state in endosomes. FAK and Src kinases co-localized with endocytosed integrin and were critical for FA reassembly by regulating integrin activation and recycling, respectively. FAK sustained the active integrin conformation by maintaining talin association with Rab11 endosomes in a type I phosphatidylinositol phosphate kinase (PIPKIγ)-dependent manner. In migrating cells, endocytosed integrins reassembled FAs polarized towards the leading edge, and this polarization required FAK. These studies identify unanticipated roles for FA proteins in maintaining endocytosed integrin in an active conformation. We propose that the conformational memory of endocytosed integrin enhances polarized reassembly of FAs to enable directional cell migration. PMID:27043085

  1. Human Pregnancy Specific Beta-1-Glycoprotein 1 (PSG1) Has a Potential Role in Placental Vascular Morphogenesis1

    OpenAIRE

    Ha, Cam T.; Wu, Julie A.; Irmak, Ster; Lisboa, Felipe A.; Dizon, Anne M.; Warren, James W; Ergun, Suleyman; Dveksler, Gabriela S.

    2010-01-01

    Previous studies suggest that human pregnancy specific beta-1-glycoproteins (PSGs) play immunomodulatory roles during pregnancy; however, other possible functions of PSGs have yet to be explored. We have observed that PSGs induce transforming growth factor beta 1 (TGFB1), which among its other diverse functions inhibits T-cell function and has proangiogenic properties. The present study investigates a potential role for PSG1, the most abundant PSG in maternal serum, as a possible inducer of p...

  2. Downregulation of IL-6-induced STAT3 tyrosine phosphorylation by TGF-beta 1 is mediated by caspase-dependent and -independent processes

    NARCIS (Netherlands)

    Wierenga, ATJ; Schuringa, JJ; Eggen, BJL; Kruijer, W; Vellenga, E

    2002-01-01

    To explore the possible cross-talk between the IL-6 and TGF-beta1 pathways in AML blast cells, the effect of TGF-beta1 pretreatment on IL-6-induced STAT3 tyrosine phosphorylation was studied. A reduction of STAT3 tyrosine phosphorylation after TGF-beta1 pretreatment was observed in four out of 40 AM

  3. Bone response and mechanical strength of rabbit femoral defects filled with injectable CaP cements containing TGF-beta 1 loaded gelatin microparticles.

    NARCIS (Netherlands)

    Link, D.P.; Dolder, J. van den; Beucken, J.J.J.P van den; Wolke, J.G.C.; Mikos, A.G.; Jansen, J.A.

    2008-01-01

    This study focused at the potential of transforming growth factor beta 1 (TGF-beta 1) loaded gelatin microparticles to enhance the bone response and mechanical strength of rabbit femoral defects filled with injectable calcium phosphate (CaP)/gelatin microparticle composites. Therefore, TGF-beta1 loa

  4. TGF beta-1 dependent fast stimulation of ATM and p53 phosphorylation following exposure to ionizing radiation does not involve TGF beta-receptor I signalling

    NARCIS (Netherlands)

    Wiegman, Erwin M.; Blaese, Marcet A.; Loeffler, Heidi; Coppes, Rob P.; Rodemann, H. Peter

    2007-01-01

    Background and purpose: It has been proposed that radiation induced stimulation of ATM and downstream components involves activation of TGF beta-1 and that this may be due to TGF beta-1-receptor I-Smad signalling. Therefore, the aim of this study was to clarify the distinct role of TGF beta-1-recept

  5. Suppression of integrin activation by the membrane-distal sequence of the integrin alphaIIb cytoplasmic tail.

    Science.gov (United States)

    Yamanouchi, Jun; Hato, Takaaki; Tamura, Tatsushiro; Fujita, Shigeru

    2004-01-01

    Integrin cytoplasmic tails regulate integrin activation including an increase in integrin affinity for ligands. Although there is ample evidence that the membrane-proximal regions of the alpha and beta tails interact with each other to maintain integrins in a low-affinity state, little is known about the role of the membrane-distal region of the alpha tail in regulation of integrin activation. We report a critical sequence for regulation of integrin activation in the membrane-distal region of the alphaIIb tail. Alanine substitution of the RPP residues in the alphaIIb tail rendered alphaIIbbeta3 constitutively active in a metabolic energy-dependent manner. Although an alphaIIb/alpha6Abeta3 chimaeric integrin, in which the alphaIIb tail was replaced by the alpha6A tail, was in an energy-dependent active state to bind soluble ligands, introduction of the RPP sequence into the alpha6A tail inhibited binding of an activation-dependent antibody PAC1. In alphaIIb/alpha6Abeta3, deleting the TSDA sequence from the alpha6A tail or single amino acid substitutions of the TSDA residues inhibited alphaIIb/alpha6Abeta3 activation and replacing the membrane-distal region of the alphaIIb tail with TSDA rendered alphaIIbbeta3 active, suggesting a stimulatory role of TSDA in energy-dependent integrin activation. However, adding TSDA to the alphaIIb tail containing the RPP sequence of the membrane-distal region failed to activate alphaIIbbeta3. These results suggest that the RPP sequence after the GFFKR motif of the alphaIIb tail suppresses energy-dependent alphaIIbbeta3 activation. These findings provide a molecular basis for the regulation of energy-dependent integrin activation by alpha subunit tails. PMID:14723599

  6. Angiotensin II increases CTGF expression via MAPKs/TGF-{beta}1/TRAF6 pathway in atrial fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Gu, Jun [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University School of medicine, Shanghai (China); Liu, Xu, E-mail: xkliuxu@yahoo.cn [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University School of medicine, Shanghai (China); Wang, Quan-xing, E-mail: shmywqx@126.com [National Key Laboratory of Medical Immunology, Second Military Medical University, Shanghai (China); Tan, Hong-wei [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University School of medicine, Shanghai (China); Guo, Meng [National Key Laboratory of Medical Immunology, Second Military Medical University, Shanghai (China); Jiang, Wei-feng; Zhou, Li [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University School of medicine, Shanghai (China)

    2012-10-01

    The activation of transforming growth factor-{beta}1(TGF-{beta}1)/Smad signaling pathway and increased expression of connective tissue growth factor (CTGF) induced by angiotensin II (AngII) have been proposed as a mechanism for atrial fibrosis. However, whether TGF{beta}1/non-Smad signaling pathways involved in AngII-induced fibrogenetic factor expression remained unknown. Recently tumor necrosis factor receptor associated factor 6 (TRAF6)/TGF{beta}-associated kinase 1 (TAK1) has been shown to be crucial for the activation of TGF-{beta}1/non-Smad signaling pathways. In the present study, we explored the role of TGF-{beta}1/TRAF6 pathway in AngII-induced CTGF expression in cultured adult atrial fibroblasts. AngII (1 {mu}M) provoked the activation of P38 mitogen activated protein kinase (P38 MAPK), extracellular signal-regulated kinase 1/2(ERK1/2) and c-Jun NH(2)-terminal kinase (JNK). AngII (1 {mu}M) also promoted TGF{beta}1, TRAF6, CTGF expression and TAK1 phosphorylation, which were suppressed by angiotensin type I receptor antagonist (Losartan) as well as p38 MAPK inhibitor (SB202190), ERK1/2 inhibitor (PD98059) and JNK inhibitor (SP600125). Meanwhile, both TGF{beta}1 antibody and TRAF6 siRNA decreased the stimulatory effect of AngII on TRAF6, CTGF expression and TAK1 phosphorylation, which also attenuated AngII-induced atrial fibroblasts proliferation. In summary, the MAPKs/TGF{beta}1/TRAF6 pathway is an important signaling pathway in AngII-induced CTGF expression, and inhibition of TRAF6 may therefore represent a new target for reversing Ang II-induced atrial fibrosis. -- Highlights: Black-Right-Pointing-Pointer MAPKs/TGF{beta}1/TRAF6 participates in AngII-induced CTGF expression in atrial fibroblasts. Black-Right-Pointing-Pointer TGF{beta}1/TRAF6 participates in AngII-induced atrial fibroblasts proliferation. Black-Right-Pointing-Pointer TRAF6 may represent a new target for reversing Ang II-induced atrial fibrosis.

  7. Cost-effectiveness analysis of peginterferon beta-1a compared with interferon beta-1a and glatiramer acetate in the treatment of relapsing-remitting multiple sclerosis in the United States.

    Science.gov (United States)

    Hernandez, Luis; Guo, Shien; Kinter, Elizabeth; Fay, Monica

    2016-07-01

    Objective Peginterferon beta-1a 125 mcg, administered subcutaneously (SC) every 2 weeks, a new disease-modifying therapy (DMT) for relapsing-remitting multiple sclerosis (RRMS), was approved by the US Food and Drug Administration in 2014. This study assesses the cost-effectiveness of peginterferon beta-1a vs interferon beta-1a (44 mcg SC 3 times per week) and glatiramer acetate (20 mg SC once-daily) in the treatment of RRMS from the perspective of a US payer over 10 years. Methods A Markov cohort economic model was developed for this analysis. The model predicts disability progression, occurrence of relapses and other adverse events and translates them into quality-adjusted life years (QALYs) and costs. Natural history data were obtained from the placebo arm of the ADVANCE trial of peginterferon beta-1a, the London Ontario (Canada) database and a large population-based MS survey. Comparative efficacy of each DMT vs placebo was obtained from a network meta-analysis. Costs (in 2014 US dollars) were sourced from public databases and literature. Clinical and economic outcomes were discounted at 3% per year. Results Over 10 years, peginterferon beta-1a was dominant (i.e., more effective and less costly), with cost-savings of $22,070 and additional 0.06 QALYs when compared with interferon beta-1a 44 mcg and with cost-savings of $19,163 and 0.07 QALYs gained when compared with glatiramer acetate 20 mg. Results were most sensitive to variations in the treatment effect of each DMT, treatment acquisition costs of each DMT and the time horizon. Probabilistic sensitivity analyses indicated that peginterferon beta-1a remains dominant in >90% of 5,000 replications compared with either DMTs. Conclusion This analysis suggests that long-term treatment with peginterferon beta-1a improves clinical outcomes at reduced costs compared with interferon beta-1a 44 mcg and glatiramer acetate 20 mg and should be a valuable addition to managed care formularies for treating

  8. Quantitative analysis of transforming growth factor beta 1 mRNA in patients with alcoholic liver disease

    Institute of Scientific and Technical Information of China (English)

    Wei-Xing Chen; You-Ming Li; Chao-Hui Yu; Wei-Min Cai; Min Zheng; Feng Chen

    2002-01-01

    AIM: To investigate the expression of the transforminggrowth factor beta 1 (TGF- beta 1 ) mRNA in different stagesof alcoholic liver disease (ALD) and its clinical value.METHODS: One hundred and seven male alcoholics weregrouped by clinical findings into four groups: alcoholabusers without liver impairment (n=22 ), alcoholicsteatosis ( n = 30 ); alcoholic hepatitis ( n = 31 ); andalcoholic cirrhosis ( n = 24 ) Using peripheral bloodmononuclear cells(PBMC) as samples the gene expressionof TGF-beta 1 was examined quantitatively by reversetranscription polymerase chain reaction (RT-PCR) and dotblot. There are 34 healthy subjects served as control.RESULTS: The expression of TGF-beta 1 from all ALDpatients was significantly greater than that in controls ( 1. 320± 1.162 vs 0.808±0.276, P<0.001). The differences of theexpressions were significant between the patients from eachgroups ( alcoholic steatosis, alcoholic hepatitis andalcoholic cirrhosis) and the controls ( 1. 168 ± 0.852, 1.462 ±1.657, 1.329± 0.610 vs 0.808 ± 0.276, P< 0.050). Nosignificant differences of TGF -beta 1 mRNA expression wereobserved between alcohol abusers without liver impairmentand controls. The expressions in patients with alcoholichepatitis and alcoholic cirrhosis were significantly greaterthan that in alcohol abusers respectively (1.462 ± 1. 657, 1.329 ± 0. 610 vs 0. 841 ± 0. 706, P < 0. 050). No significantdifferences of TGF -beta 1 mRNA expression were observedbetween alcoholic fatty liver men and alcohol abusers.CONCLUSION: TGF-beta 1 expression level can be a riskfactor for alcoholic liver disease and might be related to theinflammatory activity and fibrosis of the liver in patients .

  9. Skin Necrosis Following a Recombinant Interferon-beta-1b Injection

    Directory of Open Access Journals (Sweden)

    Chih-Hsun Yang

    2002-11-01

    Full Text Available Recombinant interferon beta-1b (INF-β-1b has been proven to be an effective means oftreating relapsing-remitting multiple sclerosis (MS. Adverse reactions to interferon therapyhave been well documented. The most common side effects are transient influenza-likesymptoms, including fever, fatigue, nausea, and myalgia. Cutaneous necrosis has occasionallybeen reported, mostly involving small and limited lesions. This article describes an MSpatient who developed multiple large, deep cutaneous ulcers on INF-β-1b injection sites,which subsequently required surgical treatment. Vessel thrombosis in the subcutaneous fattylayer and the clinical appearance of livedoid erythema beside the ulcers indicated that INF-β-1b may have caused skin necrosis through its vascular effects.

  10. IGF-binding proteins mediate TGF-beta 1-induced apoptosis in bovine mammary epithelial BME-UV1 cells.

    Science.gov (United States)

    Gajewska, Małgorzata; Motyl, Tomasz

    2004-10-01

    TGF-beta 1 is an antiproliferative and apoptogenic factor for mammary epithelial cells (MEC) acting in an auto/paracrine manner and thus considered an important local regulator of mammary tissue involution. However, the apoptogenic signaling pathway induced by this cytokine in bovine MEC remains obscure. The present study was focused on identification of molecules involved in apoptogenic signaling of transforming growth factor-beta 1 (TGF-beta 1) in the model of bovine mammary epithelial cell line (BME-UV1). Laser scanning cytometry (LSC), Western blot and electrophoretic mobility shift assay (EMSA) were used for analysis of expression and activity of TGF-beta 1-related signaling molecules. The earliest response occurring within 1-2 h after TGF-beta 1 administration was an induction and activation of R-Smads (Smad2 and Smad3) and Co-Smad (Smad4). An evident formation of Smad-DNA complexes began from 2nd hour after MEC exposure to TGF-beta 1. Similarly to Smads, proteins of AP1 complex: phosphorylated c-Jun and JunD appeared to be early reactive molecules; however, an increase in their expression was detected only in cytosolic fraction. In the next step, an increase of IGF binding protein-3 (IGFBP-3) and IGFBP-4 expression was observed from 6th hour followed by a decrease in the activity of protein kinase B (PKB/Akt), which occurred after 24 h of MEC exposure to TGF-beta 1. The decrease in PKB/Akt activity coincided in time with the decline of phosphorylated Bad expression (inactive form). Present study supported additional evidence that stimulation of insulin-like growth factor I (IGF-I) was associated with complete abrogation of TGF-beta 1-induced activation of Bad and Bax and in the consequence protection against apoptosis. In conclusion, apoptotic effect of TGF-beta 1 in bovine MEC is mediated by IGFBPs and occurs through IGF-I sequestration, resulting in inhibition of PKB/Akt-dependent survival pathway. PMID:15556067

  11. Loss of bone marrow adrenergic beta 1 and 2 receptors modifies transcriptional networks, reduces circulating inflammatory factors, and regulates blood pressure.

    Science.gov (United States)

    Ahmari, Niousha; Schmidt, Jordan T; Krane, Gregory A; Malphurs, Wendi; Cunningham, Bruce E; Owen, Jennifer L; Martyniuk, Christopher J; Zubcevic, Jasenka

    2016-07-01

    Hypertension (HTN) is a prevalent condition with complex etiology and pathophysiology. Evidence exists of significant communication between the nervous system and the immune system (IS), and there appears to be a direct role for inflammatory bone marrow (BM) cells in the pathophysiology of hypertension. However, the molecular and neural mechanisms underlying this interaction have not been characterized. Here, we transplanted whole BM cells from the beta 1 and 2 adrenergic receptor (AdrB1(tm1Bkk)AdrB2(tm1Bkk)/J) knockout (KO) mice into near lethally irradiated C57BL/6J mice to generate a BM AdrB1.B2 KO chimera. This allowed us to evaluate the role of the BM beta 1 and beta 2 adrenergic receptors in mediating BM IS homeostasis and regulating blood pressure (BP) in an otherwise intact physiological setting. Fluorescence-activated cell sorting demonstrated that a decrease in systolic and mean BP in the AdrB1.B2 KO chimera is associated with a decrease in circulating inflammatory T cells, macrophage/monocytes, and neutrophils. Transcriptomics in the BM identified 7,419 differentially expressed transcripts between the C57 and AdrB1.B2 KO chimera. Pathway analysis revealed differentially expressed transcripts related to several cell processes in the BM of C57 compared with AdrB1.B2 KO chimera, including processes related to immunity (e.g., T-cell activation, T-cell recruitment, cytokine production, leukocyte migration and function), the cardiovascular system (e.g., blood vessel development, peripheral nerve blood flow), and the brain (e.g., central nervous system development, neurite development) among others. This study generates new insight into the molecular events that underlie the interaction between the sympathetic drive and IS in modulation of BP. PMID:27235450

  12. Factor de crecimiento transformante beta-1: estructura, función y mecanismos de regulación en cáncer Transforming growth factor beta-1: structure, function and regulation mechanisms in cancer

    OpenAIRE

    Oscar Peralta-Zaragoza; A. Lagunas-Martínez; Vicente Madrid-Marina

    2001-01-01

    El factor de crecimiento transformante beta-1 (TGF-beta1) es sintetizado por muchas estirpes celulares como linfocitos, macrófagos y células dendríticas, y su expresión regula de manera autócrina o parácrina la diferenciación, proliferación y el estado de activación de éstas y muchas otras células. En general, el TGF-beta1 tiene propiedades pleiotrópicas en el contexto de la respuesta inmune durante el desarrollo de infecciones y procesos neoplásicos; sin embargo, los mecanismos de acción y r...

  13. Transforming growth factor-beta 1 stimulates synthesis of proteoglycan aggregates in calf articular cartilage organ cultures

    Energy Technology Data Exchange (ETDEWEB)

    Morales, T.I. (Bone Research Branch, National Institute of Dental Research, National Institutes of Health, Bethesda, MD (United States))

    1991-04-01

    Previous work showed that transforming growth factor-beta 1 (TGF-beta 1), added alone to bovine cartilage organ cultures, stimulated (35S)sulfate incorporation into macromolecular material but did not investigate the fidelity of the stimulated system to maintain synthesis of cartilage-type proteoglycans. This paper provides evidence that chondrocytes synthesize the appropriate proteoglycan matrix under TGF-beta 1 stimulation: (1) there is a coordinated increase in hyaluronic acid and proteoglycan monomer synthesis, (2) link-stable proteoglycan aggregates are assembled, (3) the hybrid chondroitin sulfate/keratan sulfate monomeric species is synthesized, and (4) there is an increase in protein core synthesis. Some variation in glycosylation patterns was observed when proteoglycans synthesized under TGF-beta 1 stimulation were compared to those synthesized under basal conditions. Thus comparing TGF-beta 1 to basal samples respectively, the monomers were larger (Kav on Sepharose CL-2B = 0.29 vs 0.41), the chondroitin sulfate chains were longer by approximately 3.5 kDa, the percentage of total glycosaminoglycan in keratan sulfate increased slightly from approximately 4% (basal) to approximately 6%, and the unsulfated disaccharide decreased from 28% (basal) to 12%. All of these variations are in the direction of a more anionic proteoglycan. Since the ability of proteoglycans to confer resiliency to the cartilage matrix is directly related to their anionic nature, these changes would presumably have a beneficial effect on tissue function.

  14. Tumor-host interactions in the gallbladder suppress distal angiogenesis and tumor growth: involvement of transforming growth factor beta1.

    Science.gov (United States)

    Gohongi, T; Fukumura, D; Boucher, Y; Yun, C O; Soff, G A; Compton, C; Todoroki, T; Jain, R K

    1999-10-01

    Angiogenesis inhibitors produced by a primary tumor can create a systemic anti-angiogenic environment and maintain metastatic tumor cells in a state of dormancy. We show here that the gallbladder microenvironment modulates the production of transforming growth factor (TGF)-beta1, a multifunctional cytokine that functions as an endogenous anti-angiogenic and anti-tumor factor in a cranial window preparation. We found that a wide variety of human gallbladder tumors express TGF-beta1 irrespective of histologic type. We implanted a gel impregnated with basic fibroblast growth factor or Mz-ChA-2 tumor in the cranial windows of mice without tumors or mice with subcutaneous or gallbladder tumors to study angiogenesis and tumor growth at a secondary site. Angiogenesis, leukocyte-endothelial interaction in vessels and tumor growth in the cranial window were substantially inhibited in mice with gallbladder tumors. The concentration of TGF-beta1 in the plasma of mice with gallbladder tumors was 300% higher than that in the plasma of mice without tumors or with subcutaneous tumors. In contrast, there was no difference in the plasma levels of other anti- and pro-angiogenic factors. Treatment with neutralizing antibody against TGF-beta1 reversed both angiogenesis suppression and inhibition of leukocyte rolling induced by gallbladder tumors. TGF-beta1 also inhibited Mz-ChA-2 tumor cell proliferation. Our results indicate that the production of anti-angiogenesis/proliferation factors is regulated by tumor-host interactions. PMID:10502827

  15. Regulation of proliferation of embryonic heart mesenchyme: Role of transforming growth factor-beta 1 and the interstitial matrix

    International Nuclear Information System (INIS)

    Proliferation of atrioventricular cushion mesenchyme of the embryonic avian heart maintained in three-dimensional aggregate culture is stimulated by interaction with the interstitial matrix. Chicken serum or transforming growth factor-beta 1, which stimulates proliferation, induces matrix deposition in regions of the aggregate showing high labeling indices with tritiated thymidine. Dispersed heart mesenchyme interstitial matrix introduced into serum-free culture is incorporated into the aggregate and stimulates cellular proliferation similar to serum or transforming growth factor-beta 1. Proliferation is reversibly inhibited by the peptide Gly-Arg-Gly-Asp-Ser-Pro. It is suggested that transforming growth factor-beta 1 stimulates the production of interstitial matrix and that a sufficient stimulus for proliferation in this system is the presence of the matrix, which acts as the adhesive support for cellular anchorage

  16. GSK3 inactivation is involved in mitochondrial complex IV defect in transforming growth factor (TGF) {beta}1-induced senescence

    Energy Technology Data Exchange (ETDEWEB)

    Byun, Hae-Ok; Jung, Hyun-Jung; Seo, Yong-Hak; Lee, Young-Kyoung [Department of Biochemistry and Molecular Biology, Ajou University School of Medicine, Suwon 443-721 (Korea, Republic of); Department of Molecular Science and Technology, The Graduate School, Ajou University, Suwon 443-721 (Korea, Republic of); Hwang, Sung-Chul [Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine, Suwon 443-721 (Korea, Republic of); Seong Hwang, Eun [Department of Life Science, University of Seoul, Seoul 130-743 (Korea, Republic of); Yoon, Gyesoon, E-mail: ypeace@ajou.ac.kr [Department of Biochemistry and Molecular Biology, Ajou University School of Medicine, Suwon 443-721 (Korea, Republic of); Department of Molecular Science and Technology, The Graduate School, Ajou University, Suwon 443-721 (Korea, Republic of)

    2012-09-10

    Transforming growth factor {beta}1 (TGF {beta}1) induces Mv1Lu cell senescence by persistently producing mitochondrial reactive oxygen species (ROS) through decreased complex IV activity. Here, we investigated the molecular mechanism underlying the effect of TGF {beta}1 on mitochondrial complex IV activity. TGF {beta}1 progressively phosphorylated the negative regulatory sites of both glycogen synthase kinase 3 (GSK3) {alpha} and {beta}, corresponding well to the intracellular ROS generation profile. Pre-treatment of N-acetyl cysteine, an antioxidant, did not alter this GSK3 phosphorylation (inactivation), whereas pharmacological inhibition of GSK3 by SB415286 significantly increased mitochondrial ROS, implying that GSK3 phosphorylation is an upstream event of the ROS generation. GSK3 inhibition by SB415286 decreased complex IV activity and cellular O{sub 2} consumption rate and eventually induced senescence of Mv1Lu cell. Similar results were obtained with siRNA-mediated knockdown of GSK3. Moreover, we found that GSK3 not only exists in cytosol but also in mitochondria of Mv1Lu cell and the mitochondrial GSK3 binds complex IV subunit 6b which has no electron carrier and is topologically located in the mitochondrial intermembrane space. Involvement of subunit 6b in controlling complex IV activity and overall respiration rate was proved with siRNA-mediated knockdown of subunit 6b. Finally, TGF {beta}1 treatment decreased the binding of the subunit 6b to GSK3 and subunit 6b phosphorylation. Taken together, our results suggest that GSK3 inactivation is importantly involved in TGF {beta}1-induced complex IV defects through decreasing phosphorylation of the subunit 6b, thereby contributing to senescence-associated mitochondrial ROS generation.

  17. Toxin A from Clostridium difficile binds to rabbit erythrocyte glycolipids with terminal Gal alpha 1-3Gal beta 1-4GlcNAc sequences

    International Nuclear Information System (INIS)

    The binding of Toxin A isolated from Clostridium difficile to rabbit erythrocyte glycolipids has been studied. Total lipid extracts from rabbit erythrocytes were subjected to thin-layer chromatography and toxin-binding glycolipids detected by using 125I-labeled Toxin A in a direct binding overlay technique. Two major and several minor toxin-binding glycolipids were detected in rabbit erythrocytes by this method. The results of structural analyses of the major toxin-binding glycolipids were consistent with a pentasaccharide-ceramide (Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc-Cer) and a branched decasaccharide-ceramide (Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-3[Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-6]Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc-Cer) previously identified as the two most abundant glycolipids in rabbit erythrocytes. 125I-Toxin A binding to these glycolipids could be inhibited by bovine thyroglobulin, monospecific antiserum to the toxin, or by treatment of the glycolipids with alpha-galactosidase. The absence of toxin interaction with isoglobotriaosylceramide (Gal alpha 1-3Gal beta 1-4Glc-Cer) isolated from canine intestine suggested that the GlcNAc residue present in the terminal Gal alpha 1-3Gal beta 1-4GLcNAc sequence common to all known toxin binding glycoconjugates is required for carbohydrate-specific recognition by Toxin A. These observations are consistent with the proposed carbohydrate binding specificity of Toxin A for the nonreducing terminal sequence, Gal alpha 1-3Gal beta 1-4GlcNAc

  18. Toxin A from Clostridium difficile binds to rabbit erythrocyte glycolipids with terminal Gal alpha 1-3Gal beta 1-4GlcNAc sequences

    Energy Technology Data Exchange (ETDEWEB)

    Clark, G.F.; Krivan, H.C.; Wilkins, T.D.; Smith, D.F.

    1987-08-15

    The binding of Toxin A isolated from Clostridium difficile to rabbit erythrocyte glycolipids has been studied. Total lipid extracts from rabbit erythrocytes were subjected to thin-layer chromatography and toxin-binding glycolipids detected by using /sup 125/I-labeled Toxin A in a direct binding overlay technique. Two major and several minor toxin-binding glycolipids were detected in rabbit erythrocytes by this method. The results of structural analyses of the major toxin-binding glycolipids were consistent with a pentasaccharide-ceramide (Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc-Cer) and a branched decasaccharide-ceramide (Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-3(Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-6)Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc-Cer) previously identified as the two most abundant glycolipids in rabbit erythrocytes. /sup 125/I-Toxin A binding to these glycolipids could be inhibited by bovine thyroglobulin, monospecific antiserum to the toxin, or by treatment of the glycolipids with alpha-galactosidase. The absence of toxin interaction with isoglobotriaosylceramide (Gal alpha 1-3Gal beta 1-4Glc-Cer) isolated from canine intestine suggested that the GlcNAc residue present in the terminal Gal alpha 1-3Gal beta 1-4GLcNAc sequence common to all known toxin binding glycoconjugates is required for carbohydrate-specific recognition by Toxin A. These observations are consistent with the proposed carbohydrate binding specificity of Toxin A for the nonreducing terminal sequence, Gal alpha 1-3Gal beta 1-4GlcNAc.

  19. Molecular cloning of the first metazoan beta-1,3 glucanase from eggs of the sea urchin Strongylocentrotus purpuratus.

    OpenAIRE

    Bachman, E S; McClay, D R

    1996-01-01

    We report the molecular cloning of the first beta-1,3 glucanase from animal tissue. Three peptide sequences were obtained from beta-1,3 glucanase that had been purified from eggs of the sea urchin Strongylocentrotus purpuratus and the gene was cloned by PCR using oligonucleotides deduced from the peptide sequences. The full-length cDNA shows a predicted enzyme structure of 499 aa with a hydrophobic signal sequence. A 3.2-kb message is present in eggs, during early embryogenesis, and in adult ...

  20. Expression of β2-integrin on leukocytes in liver cirrhosis

    Institute of Scientific and Technical Information of China (English)

    Anatol Panasiuk; Janusz Zak; Elzbieta Maciorkowska; Bozena Panasiuk; Danuta Prokopowicz

    2006-01-01

    AIM: To analyze β2-integrin expression on blood leukocytes in liver cirrhosis.METHODS: In 40 patients with liver cirrhosis and 20healthy individuals, the evaluation of expression of CD11a (LFA-1α), CD11b (Mac-1α), CD11c (αX) and CD49d (VLA-4α) on peripheral blood leukocytes was performed using flow cytometry. The analysis was carried out in groups of patients divided into B and C according to Child-Pugh's classification.RESULTS: An increased CD11a, CD11b, CD11c and CD49d integrin expression was observed on peripheral blood leukocytes in liver cirrhosis. The integrin levels were elevated as the advancement of liver failure progressed. The highest expression of integrins occurred predominantly on monocytes. A slight expression of VLA-4 was found on lymphocytes and granulocytes and it increased together with liver failure. A positive correlation was noted between median intensity of fluorescence (MIF) expression on polymorphonuclear cells of CD11a and CD11c and CD49d (r = 0.42, P < 0.01; r = 053, P < 0.01, respectively) in liver cirrhosis stage C. However,no correlation was observed between integrin expression on leukocytes. The concentrations of sICAM-1, sVCAM-1,and TNFα, were significantly elevated in liver cirrhosis.CONCLUSION: β2-integrin expression on leukocytes increases in liver cirrhosis decompensated as the stage of liver failure increases, which is a result of permanent activation of leukocytes circulating through the inflamed liver environment. β2-integrin expression on circulating leukocytes can intensify liver cirrhosis.

  1. Injectable interferon beta-1b for the treatment of relapsing forms of multiple sclerosis

    Directory of Open Access Journals (Sweden)

    Slobodan M Jankovic

    2010-03-01

    Full Text Available Slobodan M JankovicPharmacology Department, Medical Faculty, University of Kragujevac, Kragujevac, SerbiaAbstract: Multiple sclerosis (MS is chronic inflammatory and demyelinating disease with either a progressive (10%–15% or relapsing-remitting (85%–90% course. The pathological hallmarks of MS are lesions of both white and grey matter in the central nervous system. The onset of the disease is usually around 30 years of age. The patients experience an acute focal neurologic dysfunction which is not characteristic, followed by partial or complete recovery. Acute episodes of neurologic dysfunction with diverse signs and symptoms will then recur throughout the life of a patient, with periods of partial or complete remission and clinical stability in between. Currently, there are several therapeutic options for MS with disease-modifying properties. Immunomodulatory therapy with interferon beta-1b (IFN-β1b or -1a, glatiramer and natalizumab shows similar efficacy; in a resistant or intolerant patient, the most recently approved therapeutic option is mitoxantrone. IFN-β1b in patients with MS binds to specific receptors on surface of immune cells, changing the expression of several genes and leading to a decrease in quantity of cell-associated adhesion molecules, inhibition of major histocompatibility complex class II expression and reduction in inflammatory cells migration into the central nervous system. After 2 years of treatment, IFN-β1b reduces the risk of development of clinically defined MS from 45% (with placebo to 28% (with IFN-β1b. It also reduces relapses for 34% (1.31 exacerbations annually with placebo and 0.9 with higher dose of IFN-β1b and makes 31% more patients relapse-free. In secondary-progressive disease annual rate of progression is 3% lower with IFN-β1b. In recommended doses IFN-β1b causes the following frequent adverse effects: injection site reactions (redness, discoloration, inflammation, pain, necrosis and non

  2. Photon-induced cell migration and integrin expression promoted by DNA integration of HPV16 genome

    International Nuclear Information System (INIS)

    Persistent human papilloma virus 16 (HPV16) infections are a major cause of cervical cancer. The integration of the viral DNA into the host genome causes E2 gene disruption which prevents apoptosis and increases host cell motility. In cervical cancer patients, survival is limited by local infiltration and systemic dissemination. Surgical control rates are poor in cases of parametrial infiltration. In these patients, radiotherapy (RT) is administered to enhance local control. However, photon irradiation itself has been reported to increase cell motility. In cases of E2-disrupted cervical cancers, this phenomenon would impose an additional risk of enhanced tumor cell motility. Here, we analyze mechanisms underlying photon-increased migration in keratinocytes with differential E2 gene status. Isogenic W12 (intact E2 gene status) and S12 (disrupted E2 gene status) keratinocytes were analyzed in fibronectin-based and serum-stimulated migration experiments following single photon doses of 0, 2, and 10 Gy. Quantitative FACS analyses of integrin expression were performed. Migration and adhesion are increased in E2 gene-disrupted keratinocytes. E2 gene disruption promotes attractability by serum components, therefore, effectuating the risk of local infiltration and systemic dissemination. In S12 cells, migration is further increased by photon RT which leads to enhanced expression of fibronectin receptor integrins. HPV16-associated E2 gene disruption is a main predictor of treatment-refractory cancer virulence. E2 gene disruption promotes cell motility. Following photon RT, E2-disrupted tumors bear the risk of integrin-related infiltration and dissemination. (orig.)

  3. Interferon beta 1-a na esclerose múltipla: experiência de um ano em 62 pacientes Interferon beta 1-a in multiple sclerosis: experience of one year in 62 patients

    Directory of Open Access Journals (Sweden)

    CHARLES P. TILBERY

    2000-06-01

    Full Text Available Relatamos os resultados de estudo com o interferon beta 1-a em 62 pacientes ambulatoriais com a forma remitente-recorrente da esclerose múltipla durante um ano. Os critérios de inclusão para este tratamento foram de escore do EDSS entre 0 e 5,5 e de pelo menos relato de dois surtos nos dois últimos anos. Administramos 3 milhões de unidades internacionais de interferon beta 1-a três vezes por semana. Os objetivos deste estudo foram verificar o efeito da medicação no número de surtos e avaliar a eficácia da droga na progressão da doença. O índice anual de surtos nos pacientes que não tomaram a medicação foi 1,32 e naqueles medicados 0,63. O escore do EDSS em pacientes não medicados foi 4,7 e naqueles com medicamento 2,0. O interferon beta 1-a foi bem tolerado e 85% dos pacientes completaram um ano de tratamento.We report the results of a trial of interferon beta 1-a in 62 ambulatory patients with relapsing-remitting multiple sclerosis. Entry criteria included EDSS of 0 to 5.5 and at least two exacerbations in the previous 2 years. The patients received 3 million international units by subcutaneous injections three times a week. The end points were differences in exacerbation rate and treatment effect on disease progression. The annual exacerbation rate for patients that did not take the interferon beta 1-a was 1.32 and for the patients under medication 0.63. The EDSS score in patients that did not take the mediaction was 4.7 and 2.0 for the patients with interferon beta 1-a. Interferon beta 1-a was well tolerated and 85% of patients completed 1 year treatment.

  4. Integrin activation by a cold atmospheric plasma jet

    International Nuclear Information System (INIS)

    Current breakthrough research on cold atmospheric plasma (CAP) demonstrates that CAP has great potential in various areas, including medicine and biology, thus providing a new tool for living tissue treatment. In this paper, we explore potential mechanisms by which CAP alters cell migration and influences cell adhesion. We focus on the study of CAP interaction with fibroblasts and corneal epithelial cells. The data show that fibroblasts and corneal epithelial cells have different thresholds (treatment times) required to achieve maximum inhibition of cell migration. Both cell types reduced their migration rates by ∼30-40% after CAP compared to control cells. Also, the impact of CAP treatment on cell migration and persistence of fibroblasts after integrin activation by MnCl2, serum starvation or replating cells onto surfaces coated with integrin ligands is assessed; the results show that activation by MnCl2 or starvation attenuates cells’ responses to plasma. Studies carried out to assess the impact of CAP treatment on the activation state of β1 integrin and focal adhesion size by using immunofluorescence show that fibroblasts have more active β1 integrin on their surface and large focal adhesions after CAP treatment. Based on these data, a thermodynamic model is presented to explain how CAP leads to integrin activation and focal adhesion assembly. (paper)

  5. Primary structure determination of five sialylated oligosaccharides derived from bronchial mucus glycoproteins of patients suffering from cystic fibrosis. The occurrence of the NeuAc alpha(2----3)Gal beta(1----4)[Fuc alpha(1----3)] GlcNAc beta(1----.) structural element revealed by 500-MHz 1H NMR spectroscopy.

    Science.gov (United States)

    Lamblin, G; Boersma, A; Klein, A; Roussel, P; van Halbeek, H; Vliegenthart, J F

    1984-07-25

    The structure of sialylated carbohydrate units of bronchial mucins obtained from cystic fibrosis patients was investigated by 500-MHz 1H NMR spectroscopy in conjunction with sugar analysis. After subjecting the mucins to alkaline borohydride degradation, sialylated oligosaccharide-alditols were isolated by anion-exchange chromatography and fractionated by high performance liquid chromatography. Five compounds could be obtained in a rather pure state; their structures were established as the following: A-1, NeuAc alpha(2----3)Gal beta(1----4) [Fuc alpha(1----3)]GlcNAc beta(1----3)Gal-NAc-ol; A-2, NeuAc alpha(2----3)Gal beta(1----4)GlcNAc beta(1----6)-[GlcNAc beta (1----3)]GalNAc-o1; A-3, NeuAc alpha(2----3)Gal beta-(1----4)[Fuc alpha(1----3)]GlcNAc beta(1----3)Gal beta(1----3) GalNAc-o1; A-4, NeuAc alpha(2----3)Gal beta(1----4)[Fuc alpha(1----3)]Glc-NAc NAc beta(1----6)[GlcNAc beta(1----3)]GalNAc-o1; A-6,NeuAc alpha-(2----3) Gal beta(1----4)[Fuc alpha(1----3)]GlcNAc beta(1----6)[Gal beta-(1----4) GlcNAc beta(1----3)]GalNAc-o1. The simultaneous presence of sialic acid in alpha(2----3)-linkage to Gal and fucose in alpha(1----3)-linkage to GlcNAc of the same N-acetyllactosamine unit could be adequately proved by high resolution 1H NMR spectroscopy. This sequence constitutes a novel structural element for mucins. PMID:6746638

  6. Generalized epilepsy with febrile seizures plus-associated sodium channel beta1 subunit mutations severely reduce beta subunit-mediated modulation of sodium channel function.

    Science.gov (United States)

    Xu, R; Thomas, E A; Gazina, E V; Richards, K L; Quick, M; Wallace, R H; Harkin, L A; Heron, S E; Berkovic, S F; Scheffer, I E; Mulley, J C; Petrou, S

    2007-08-10

    Two novel mutations (R85C and R85H) on the extracellular immunoglobulin-like domain of the sodium channel beta1 subunit have been identified in individuals from two families with generalized epilepsy with febrile seizures plus (GEFS+). The functional consequences of these two mutations were determined by co-expression of the human brain NaV1.2 alpha subunit with wild type or mutant beta1 subunits in human embryonic kidney (HEK)-293T cells. Patch clamp studies confirmed the regulatory role of beta1 in that relative to NaV1.2 alone the NaV1.2+beta1 currents had right-shifted voltage dependence of activation, fast and slow inactivation and reduced use dependence. In addition, the NaV1.2+beta1 current entered fast inactivation slightly faster than NaV1.2 channels alone. The beta1(R85C) subunit appears to be a complete loss of function in that none of the modulating effects of the wild type beta1 were observed when it was co-expressed with NaV1.2. Interestingly, the beta1(R85H) subunit also failed to modulate fast kinetics, however, it shifted the voltage dependence of steady state slow inactivation in the same way as the wild type beta1 subunit. Immunohistochemical studies revealed cell surface expression of the wild type beta1 subunit and undetectable levels of cell surface expression for both mutants. The functional studies suggest association of the beta1(R85H) subunit with the alpha subunit where its influence is limited to modulating steady state slow inactivation. In summary, the mutant beta1 subunits essentially fail to modulate alpha subunits which could increase neuronal excitability and underlie GEFS+ pathogenesis. PMID:17629415

  7. Research of TGF-beta1 Inducing Lung Adencarcinoma PC9 Cells to Mesenchymal Cells Transition

    Directory of Open Access Journals (Sweden)

    Xiaofeng CHEN

    2010-01-01

    Full Text Available Background and objective It has been proven that epithelial-mesenchymal transition (EMT not only correlated with embryonic development but also could promote tumor invasion and metastasis. Transforming growth factor beta-1 (TGF-β1 has been identified as the main inducer of tumor EMT. The aim of this study was to investigate the effects of TGF-β1 on EMT and PI3K/AKT signaling pathway in lung adencarcinoma PC9 cells. Methods Cultured PC9 cells were treated with different concentrations of TGF-β1 for 48 h. The morphological changes were observed under phase-contrast microscopy; EMT relative marker protein changes were assessed by Western blot and immunoflurescence staining. In addition, the expression of AKT and P-AKT were also measured by Western blot. Results The data showed that TGF-β1 could induce PC9 morphological alteration from epithelial to mesenchymal and upregulate the expression of mesenchymal maker protein Fibronectin. Obviously, the expression of P-AKT was downregulated by TGF-β1 treatment for 48 h. Conclusion TGF-β1 might induce EMT of PC9 cells , accompanied by the changes of PI3K/AKT signaling pathway.

  8. Structural Requirements for Activation in αIIbβ3 Integrin*

    OpenAIRE

    Kamata, Tetsuji; Handa, Makoto; Ito, Sonomi; Sato, Yukiko; Ohtani, Toshimitsu; Kawai, Yohko; Ikeda, Yasuo; Aiso, Sadakazu

    2010-01-01

    Integrins are postulated to undergo structural rearrangement from a low affinity bent conformer to a high affinity extended conformer upon activation. However, some reports have shown that a bent conformer is capable of binding a ligand, whereas another report has shown that integrin extension does not absolutely lead to activation. To clarify whether integrin affinity is indeed regulated by the so-called switchblade-like movement, we have engineered a series of mutant αIIbβ3 integrins that a...

  9. Characterization and partial purification of beta-1,3-D-glucan (callose) synthase from barley (Hordeum vulgare) leaves

    DEFF Research Database (Denmark)

    Pedersen, L.H.; Jacobsen, S.; Hejgaard, J.;

    1993-01-01

    was inhibited by UDP and uridine 5' triphosphate (UTP). Glucanase digestion of the in vitro product showed that it was a beta-1,3-linked polysaccharide. Two different procedures were used for further enrichment of polypeptides involved in callose synthase activity. Sucrose gradient centrifugation with...

  10. beta-1,3-Glucan recognition by an insect pathogen recognition domain causes self-association of protein: carbohydrate complex

    Science.gov (United States)

    In response to invading microorganisms, insect beta-1,3-glucan recognition protein (beta-GRP), a soluble receptor in the hemolymph, binds to the surfaces of bacteria and fungi and activates serine protease cascades associated with the prophenoloxidase (proPO) and Toll pathways; it also agglutinates ...

  11. Ozone and ultraviolet B effects on the defense-related proteins beta-1,3-glucanase and chitinase in tobacco

    International Nuclear Information System (INIS)

    The air pollutant ozone is a potent abiotic inducer of defense-related enzymes such as pathogenesis-related proteins. Here we report on the accumulation of beta-1,3-glucanase and chitinase in Nicotiana tabacumL. treated with ozone and ultraviolet B radiation, singly and in combination, under a simulated sunlight spectrum. Ozone (0.16 mu L . L-1, 2 x 5 h) induced the basic isoforms of beta-1,3-glucanase in both, ozone-sensitive (Eel W3) and -tolerant (Bel B) cultivars, while chitinase was only affected in cv. Bel W3. Ultraviolet B radiation (7.5 MED) alone did not lead to beta-1,3-glucanase or chitinase induction. In combined treatments ultraviolet B increased the ozone-dependent lesion formation and reduced chitinase accumulation in the sensitive cv. Bel W3. Analysis of the intercellular washing fluid of ozone-treated plants revealed the accumulation of a major ozone-related protein (O(3)R-1) of 28 kDa within 32 h. Microsequence analysis of two tryptic peptides showed 100 % homology to acidic chitinase PR-3b. These results indicate that basic beta-1,3-glucanase and chitinase are distinctly regulated in ozone and ultraviolet B treated tobacco, and that ultraviolet B radiation with a similar UV edge as the solar spectrum does not lead to an accumulation of basic pathogenesis-related proteins

  12. Microglia and macrophages are major sources of locally produced transforming growth factor-beta1 after transient middle cerebral artery occlusion in rats

    DEFF Research Database (Denmark)

    Lehrmann, E; Kiefer, R; Christensen, Thomas;

    1998-01-01

    The potentially neurotrophic cytokine transforming growth factor-beta1 (TGF-beta1) is locally expressed following human stroke and experimental ischemic lesions, but the cellular source(s) and profile of induction have so far not been established in experimental focal cerebral ischemia. This study...... middle cerebral artery. Six hours after ischemia, an early and transient neuronal and microglial expression of TGF-beta1 mRNA was observed in the extraischemic cingulate and frontal cortices. Both early and protracted expression of TGF-beta1 mRNA in the caudate-putamen and neocortical infarcts and in the...

  13. Beta(1-->3)-glucan in house dust of German homes: housing characteristics, occupant behavior, and relations with endotoxins, allergens, and molds.

    OpenAIRE

    Gehring, U; Douwes, J; Doekes, G.; Koch, A.; Bischof, W; B. Fahlbusch; Richter, K.; Wichmann, H. E.; Heinrich, J.; ,

    2001-01-01

    beta(1-->3)-Glucans are potent proinflammatory agents that have been suggested to play a role in indoor-related respiratory health effects. The aim of this study was to assess whether beta(1-->3)-glucan concentrations in house dust are correlated with levels of endotoxins, allergens, and culturable mold spore counts in house dust. Further, the associations of beta(1-->3)-glucan with housing characteristics and occupant behavior were assessed. beta(1-->3)-Glucan was measured in settled house d...

  14. Synthesis and biological evaluation of potent alphavbeta3-integrin receptor antagonists.

    NARCIS (Netherlands)

    Dijkgraaf, I.; Kruijtzer, J.A.; Frielink, C.; Soede, A.C.; Hilbers, H.W.; Oyen, W.J.G.; Corstens, F.H.M.; Liskamp, R.M.; Boerman, O.C.

    2006-01-01

    INTRODUCTION: alpha(v)beta(3) Integrin is expressed in sprouting endothelial cells in growing tumors, whereas it is absent in quiescent blood vessels. In addition, various tumor cell types express alpha(v)beta(3) integrin. alpha(v)beta(3) Integrin, a transmembrane heterodimeric protein, binds to the

  15. The talin head domain reinforces integrin-mediated adhesion by promoting adhesion complex stability and clustering.

    Directory of Open Access Journals (Sweden)

    Stephanie J Ellis

    2014-11-01

    Full Text Available Talin serves an essential function during integrin-mediated adhesion in linking integrins to actin via the intracellular adhesion complex. In addition, the N-terminal head domain of talin regulates the affinity of integrins for their ECM-ligands, a process known as inside-out activation. We previously showed that in Drosophila, mutating the integrin binding site in the talin head domain resulted in weakened adhesion to the ECM. Intriguingly, subsequent studies showed that canonical inside-out activation of integrin might not take place in flies. Consistent with this, a mutation in talin that specifically blocks its ability to activate mammalian integrins does not significantly impinge on talin function during fly development. Here, we describe results suggesting that the talin head domain reinforces and stabilizes the integrin adhesion complex by promoting integrin clustering distinct from its ability to support inside-out activation. Specifically, we show that an allele of talin containing a mutation that disrupts intramolecular interactions within the talin head attenuates the assembly and reinforcement of the integrin adhesion complex. Importantly, we provide evidence that this mutation blocks integrin clustering in vivo. We propose that the talin head domain is essential for regulating integrin avidity in Drosophila and that this is crucial for integrin-mediated adhesion during animal development.

  16. The Brucella abortus cyclic beta-1,2-glucan virulence factor is substituted with O-ester-linked succinyl residues.

    Science.gov (United States)

    Roset, Mara S; Ciocchini, Andrés E; Ugalde, Rodolfo A; Iñón de Iannino, Nora

    2006-07-01

    Brucella periplasmic cyclic beta-1,2-glucan plays an important role during bacterium-host interaction. Nuclear magnetic resonance spectrometry analysis, thin-layer chromatography, and DEAE-Sephadex chromatography were used to characterize Brucella abortus cyclic glucan. In the present study, we report that a fraction of B. abortus cyclic beta-1,2-glucan is substituted with succinyl residues, which confer anionic character on the cyclic beta-1,2-glucan. The oligosaccharide backbone is substituted at C-6 positions with an average of two succinyl residues per glucan molecule. This O-ester-linked succinyl residue is the only substituent of Brucella cyclic glucan. A B. abortus open reading frame (BAB1_1718) homologous to Rhodobacter sphaeroides glucan succinyltransferase (OpgC) was identified as the gene encoding the enzyme responsible for cyclic glucan modification. This gene was named cgm for cyclic glucan modifier and is highly conserved in Brucella melitensis and Brucella suis. Nucleotide sequencing revealed that B. abortus cgm consists of a 1,182-bp open reading frame coding for a predicted membrane protein of 393 amino acid residues (42.7 kDa) 39% identical to Rhodobacter sphaeroides succinyltransferase. cgm null mutants in B. abortus strains 2308 and S19 produced neutral glucans without succinyl residues, confirming the identity of this protein as the cyclic-glucan succinyltransferase enzyme. In this study, we demonstrate that succinyl substituents of cyclic beta-1,2-glucan of B. abortus are necessary for hypo-osmotic adaptation. On the other hand, intracellular multiplication and mouse spleen colonization are not affected in cgm mutants, indicating that cyclic-beta-1,2-glucan succinylation is not required for virulence and suggesting that no low-osmotic stress conditions must be overcome during infection. PMID:16816173

  17. Adhesive and migratory effects of phosphophoryn are modulated by flanking peptides of the integrin binding motif.

    Directory of Open Access Journals (Sweden)

    Shigeki Suzuki

    Full Text Available Phosphophoryn (PP is generated from the proteolytic cleavage of dentin sialophosphoprotein (DSPP. Gene duplications in the ancestor dentin matrix protein-1 (DMP-1 genomic sequence created the DSPP gene in toothed animals. PP and DMP-1 are phosphorylated extracellular matrix proteins that belong to the family of small integrin-binding ligand N-linked glycoproteins (SIBLINGs. Many SIBLING members have been shown to evoke various cell responses through the integrin-binding Arg-Gly-Asp (RGD domain; however, the RGD-dependent function of PP is not yet fully understood. We demonstrated that recombinant PP did not exhibit any obvious cell adhesion ability, whereas the simultaneously purified recombinant DMP-1 did. A cell adhesion inhibitory analysis was performed by pre-incubating human osteosarcoma MG63 cells with various PP peptides before seeding onto vitronectin. The results obtained revealed that the incorporation of more than one amino acid on both sides of the PP-RGD domain was unable to inhibit the adhesion of MG63 cells onto vitronectin. Furthermore, the inhibitory activity of a peptide containing the PP-RGD domain with an open carboxyl-terminal side (H-463SDESDTNSESANESGSRGDA482-OH was more potent than that of a peptide containing the RGD domain with an open amino-terminal side (H-478SRGDASYTSDESSDDDNDSDSH499-OH. This phenomenon was supported by the potent cell adhesion and migration abilities of the recombinant truncated PP, which terminated with Ala482. Furthermore, various point mutations in Ala482 and/or Ser483 converted recombinant PP into cell-adhesive proteins. Therefore, we concluded that the Ala482-Ser483 flanking sequence, which was detected in primates and mice, was the key peptide bond that allowed the PP-RGD domain to be sequestered. The differential abilities of PP and DMP-1 to act on integrin imply that DSPP was duplicated from DMP-1 to serve as a crucial extracellular protein for tooth development rather than as an integrin

  18. α2 integrin as regulator of metastatic potential

    Institute of Scientific and Technical Information of China (English)

    Miroslav BARANCIK; Albert BREIER

    2011-01-01

    @@ Regulation of cellular events is a complex process that involves several factors in their specific interactions and interplays.However, this complexity signs also exist specificity and changes in single protein, and could significantly influence the properties and responses of cells. Recently, Ramirez and colleagues[1]provided innovative results that emphasize the important and selective role of one specific protein, α2 integrin, in the complex process of tumor metastasis.This protein, as a part of heterodimeric 2β1 integrin, was identified as a metastasis suppressor in both breast and prostate cancer.

  19. Vicrostatin - an anti-invasive multi-integrin targeting chimeric disintegrin with tumor anti-angiogenic and pro-apoptotic activities.

    Directory of Open Access Journals (Sweden)

    Radu O Minea

    Full Text Available Similar to other integrin-targeting strategies, disintegrins have previously shown good efficacy in animal cancer models with favorable pharmacological attributes and translational potential. Nonetheless, these polypeptides are notoriously difficult to produce recombinantly due to their particular structure requiring the correct pairing of multiple disulfide bonds for biological activity. Here, we show that a sequence-engineered disintegrin (called vicrostatin or VCN can be reliably produced in large scale amounts directly in the oxidative cytoplasm of Origami B E. coli. Through multiple integrin ligation (i.e., alphavbeta3, alphavbeta5, and alpha5beta1, VCN targets both endothelial and cancer cells significantly inhibiting their motility through a reconstituted basement membrane. Interestingly, in a manner distinct from other integrin ligands but reminiscent of some ECM-derived endogenous anti-angiogenic fragments previously described in the literature, VCN profoundly disrupts the actin cytoskeleton of endothelial cells (EC inducing a rapid disassembly of stress fibers and actin reorganization, ultimately interfering with EC's ability to invade and form tubes (tubulogenesis. Moreover, here we show for the first time that the addition of a disintegrin to tubulogenic EC sandwiched in vitro between two Matrigel layers negatively impacts their survival despite the presence of abundant haptotactic cues. A liposomal formulation of VCN (LVCN was further evaluated in vivo in two animal cancer models with different growth characteristics. Our data demonstrate that LVCN is well tolerated while exerting a significant delay in tumor growth and an increase in the survival of treated animals. These results can be partially explained by potent tumor anti-angiogenic and pro-apoptotic effects induced by LVCN.

  20. Essential function for PDLIM2 in cell polarization in three-dimensional cultures by feedback regulation of the β1-integrin-RhoA signaling axis.

    Science.gov (United States)

    Deevi, Ravi Kiran; Cox, Orla T; O'Connor, Rosemary

    2014-05-01

    PDLIM2 is a cytoskeletal and nuclear PDZ-LIM domain protein that regulates the stability of Nuclear Factor kappa-B (NFκB) and other transcription factors, and is required for polarized cell migration. PDLIM2 expression is suppressed by methylation in different cancers, but is strongly expressed in invasive breast cancer cells that have undergone an Epithelial Mesenchymal Transition (EMT). PDLIM2 is also expressed in non-transformed breast myoepithelial MCF10A cells and here we asked whether it is important for maintaining the polarized, epithelial phenotype of these cells. Suppression of PDLIM2 in MCF10A cells was sufficient to disrupt cell polarization and acini formation with increased proliferation and reduced apoptosis in the luminal space compared to control acini with hollow lumina. Spheroids with suppressed PDLIM2 exhibited increased expression of cell-cell and cell-matrix adhesion proteins including beta 1 (β1) integrin. Interestingly, levels of the Insulin-like growth factor 1 receptor (IGF-1 R) and Receptor of activated protein kinase C 1 (RACK1), which scaffolds IGF-1R to β1 integrin, were also increased, indicating a transformed phenotype. Focal Adhesion Kinase (FAK) and cofilin phosphorylation, and RhoA Guanosine Triphosphatase (GTPase) activity were all enhanced in these spheroids compared to control acini. Importantly, inhibition of either FAK or Rho Kinase (ROCK) was sufficient to rescue the polarity defect. We conclude that PDLIM2 expression is essential for feedback regulation of the β1-integrin-RhoA signalling axis and integration of cellular microenvironment signals with gene expression to control the polarity of breast epithelial acini structures. This is a mechanism by which PDLIM2 could mediate tumour suppression in breast epithelium. PMID:24863845

  1. Serum Transforming Growth Factor Beta-1 as an Index of Chemical Hepato carcinogenesis in Rats

    International Nuclear Information System (INIS)

    Transforming growth factor beta-1 (TGF β1) is an important mediator which controls liver cell proliferation and replication. The relation between TGF β1, Alpha-fetoprotein (AFP) and clinically thought hepatocellular carcinoma (HCC) in rats were investigated to clarify the clinical value of measuring peripheral serum TGF β1 and AFP in evaluation of HCC. Peripheral serum TGF β1 and AFP were measured during chemically induced hepato carcinogenesis. Male rats were given a genotoxic compound diethylnitrosamine (DEN) in drinking water for 149 days with control receiving drinking water only. Animals were killed at different times intervals 54, 86 and 149 days, serum TGF β1 levels were measured by, Enzyme Linked Immunosorbent Assay (ELISA) and AFP levels were assayed by immunoradiometric assay (IRMA). In DEN treated rats 54 days, there was mild portal tract inflammatory cellular infiltrate, serum TGF β1 and AFP levels were both significantly elevated above control (P>0.05 and P<0.001). At 86 days there were moderate inflammation (portal and peri portal), serum TGF β1 and AFP levels were significantly increased, (P<0.001). At 149 days typical HCC were present in ten of ten rats and serum TGF β1 and AFP were both significantly elevated compared with controls, (P<0.001). It can be concluded that serum TGF β1 and AFP levels are elevated during chemically induced HCC and have roles during the stages of process (initiation, promotion and progression); both serum TGF β1 and AFP levels can be used in parallel as a non invasive tumor markers for early diagnosis and prognosis of HCC

  2. Crystal Structure of the Catalytic Domain of Drosophila [beta]1,4-Galactosyltransferase-7

    Energy Technology Data Exchange (ETDEWEB)

    Ramakrishnan, Boopathy; Qasba, Pradman K. (NIH)

    2010-11-03

    The {beta}1,4-galactosyltransferase-7 ({beta}4Gal-T7) enzyme, one of seven members of the {beta}4Gal-T family, transfers in the presence of manganese Gal from UDP-Gal to an acceptor sugar (xylose) that is attached to a side chain hydroxyl group of Ser/Thr residues of proteoglycan proteins. It exhibits the least protein sequence similarity with the other family members, including the well studied family member {beta}4Gal-T1, which, in the presence of manganese, transfers Gal from UDP-Gal to GlcNAc. We report here the crystal structure of the catalytic domain of {beta}4Gal-T7 from Drosophila in the presence of manganese and UDP at 1.81 {angstrom} resolution. In the crystal structure, a new manganese ion-binding motif (HXH) has been observed. Superposition of the crystal structures of {beta}4Gal-T7 and {beta}4Gal-T1 shows that the catalytic pocket and the substrate-binding sites in these proteins are similar. Compared with GlcNAc, xylose has a hydroxyl group (instead of an N-acetyl group) at C2 and lacks the CH{sub 2}OH group at C5; thus, these protein structures show significant differences in their acceptor-binding site. Modeling of xylose in the acceptor-binding site of the {beta}4Gal-T7 crystal structure shows that the aromatic side chain of Tyr{sup 177} interacts strongly with the C5 atom of xylose, causing steric hindrance to any additional group at C5. Because Drosophila Cd7 has a 73% protein sequence similarity to human Cd7, the present crystal structure offers a structure-based explanation for the mutations in human Cd7 that have been linked to Ehlers-Danlos syndrome.

  3. Ganglioside GM2 mediates migration of tumor cells by interacting with integrin and modulating the downstream signaling pathway.

    Science.gov (United States)

    Kundu, Manjari; Mahata, Barun; Banerjee, Avisek; Chakraborty, Sohini; Debnath, Shibjyoti; Ray, Sougata Sinha; Ghosh, Zhumur; Biswas, Kaushik

    2016-07-01

    The definitive role of ganglioside GM2 in mediating tumor-induced growth and progression is still unknown. Here we report a novel role of ganglioside GM2 in mediating tumor cell migration and uncovered its mechanism. Data shows differential expression levels of GM2-synthase as well as GM2 in different human cancer cells. siRNA mediated knockdown of GM2-synthase in CCF52, A549 and SK-RC-26B cells resulted in significant inhibition of tumor cell migration as well as invasion in vitro without affecting cellular proliferation. Over-expression of GM2-synthase in low-GM2 expressing SK-RC-45 cells resulted in a consequent increase in migration thus confirming the potential role GM2 and its downstream partners play in tumor cell migration and motility. Further, treatment of SK-RC-45 cells with exogenous GM2 resulted in a dramatic increase in migratory and invasive capacity with no change in proliferative capacity, thereby confirming the role of GM2 in tumorigenesis specifically by mediating tumor migration and invasion. Gene expression profiling of GM2-synthase silenced cells revealed altered expression of several genes involved in cell migration primarily those controlling the integrin mediated signaling. GM2-synthase knockdown resulted in decreased phosphorylation of FAK, Src as well as Erk, while over-expression and/or exogenous GM2 treatment caused increased FAK and Erk phosphorylation respectively. Again, GM2 mediated invasion and Erk phosphorylation is blocked in integrin knockdown SK-RC-45 cells, thus confirming that GM2 mediated migration and phosphorylation of Erk is integrin dependent. Finally, confocal microscopy suggested co-localization while co-immunoprecipitation and surface plasmon resonance (SPR) confirmed direct interaction of membrane bound ganglioside, GM2 with the integrin receptor. PMID:27066976

  4. MicroRNA-26a modulates transforming growth factor beta-1-induced proliferation in human fetal lung fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xiaoou [Division of Pulmonary Diseases, State Key Laboratory of Biotherapy of China, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Department of Respiratory Medicine, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Liu, Lian [Division of Pulmonary Diseases, State Key Laboratory of Biotherapy of China, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Shen, Yongchun; Wang, Tao; Chen, Lei; Xu, Dan [Division of Pulmonary Diseases, State Key Laboratory of Biotherapy of China, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Department of Respiratory Medicine, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Wen, Fuqiang, E-mail: wenfuqiang.scu@gmail.com [Division of Pulmonary Diseases, State Key Laboratory of Biotherapy of China, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Department of Respiratory Medicine, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China)

    2014-11-28

    Highlights: • Endogenous miR-26a inhibits TGF-beta 1 induced proliferation of lung fibroblasts. • miR-26a induces G1 arrest through directly targeting 3′-UTR of CCND2. • TGF indispensable receptor, TGF-beta R I, is regulated by miR-26a. • miR-26a acts through inhibiting TGF-beta 2 feedback loop to reduce TGF-beta 1. • Collagen type I and connective tissue growth factor are suppressed by miR-26a. - Abstract: MicroRNA-26a is a newly discovered microRNA that has a strong anti-tumorigenic capacity and is capable of suppressing cell proliferation and activating tumor-specific apoptosis. However, whether miR-26a can inhibit the over-growth of lung fibroblasts remains unclear. The relationship between miR-26a and lung fibrosis was explored in the current study. We first investigated the effect of miR-26a on the proliferative activity of human lung fibroblasts with or without TGF-beta1 treatment. We found that the inhibition of endogenous miR-26a promoted proliferation and restoration of mature miR-26a inhibited the proliferation of human lung fibroblasts. We also examined that miR-26a can block the G1/S phase transition via directly targeting 3′-UTR of CCND2, degrading mRNA and decreasing protein expression of Cyclin D2. Furthermore, we showed that miR-26a mediated a TGF-beta 2-TGF-beta 1 feedback loop and inhibited TGF-beta R I activation. In addition, the overexpression of miR-26a also significantly suppressed the TGF-beta 1-interacting-CTGF–collagen fibrotic pathway. In summary, our studies indicated an essential role of miR-26a in the anti-fibrotic mechanism in TGF-beta1-induced proliferation in human lung fibroblasts, by directly targeting Cyclin D2, regulating TGF-beta R I as well as TGF-beta 2, and suggested the therapeutic potential of miR-26a in ameliorating lung fibrosis.

  5. MicroRNA-26a modulates transforming growth factor beta-1-induced proliferation in human fetal lung fibroblasts

    International Nuclear Information System (INIS)

    Highlights: • Endogenous miR-26a inhibits TGF-beta 1 induced proliferation of lung fibroblasts. • miR-26a induces G1 arrest through directly targeting 3′-UTR of CCND2. • TGF indispensable receptor, TGF-beta R I, is regulated by miR-26a. • miR-26a acts through inhibiting TGF-beta 2 feedback loop to reduce TGF-beta 1. • Collagen type I and connective tissue growth factor are suppressed by miR-26a. - Abstract: MicroRNA-26a is a newly discovered microRNA that has a strong anti-tumorigenic capacity and is capable of suppressing cell proliferation and activating tumor-specific apoptosis. However, whether miR-26a can inhibit the over-growth of lung fibroblasts remains unclear. The relationship between miR-26a and lung fibrosis was explored in the current study. We first investigated the effect of miR-26a on the proliferative activity of human lung fibroblasts with or without TGF-beta1 treatment. We found that the inhibition of endogenous miR-26a promoted proliferation and restoration of mature miR-26a inhibited the proliferation of human lung fibroblasts. We also examined that miR-26a can block the G1/S phase transition via directly targeting 3′-UTR of CCND2, degrading mRNA and decreasing protein expression of Cyclin D2. Furthermore, we showed that miR-26a mediated a TGF-beta 2-TGF-beta 1 feedback loop and inhibited TGF-beta R I activation. In addition, the overexpression of miR-26a also significantly suppressed the TGF-beta 1-interacting-CTGF–collagen fibrotic pathway. In summary, our studies indicated an essential role of miR-26a in the anti-fibrotic mechanism in TGF-beta1-induced proliferation in human lung fibroblasts, by directly targeting Cyclin D2, regulating TGF-beta R I as well as TGF-beta 2, and suggested the therapeutic potential of miR-26a in ameliorating lung fibrosis

  6. The Effect of the Arg389Gly Beta-1 Adrenoceptor Polymorphism on Plasma Renin Activity and Heart Rate and the Genotype-Dependent Response to Metoprolol Treatment

    DEFF Research Database (Denmark)

    Petersen, Morten; Andersen, Jon T; Jimenez-Solem, Espen;

    2012-01-01

    A gene-drug interaction has been indicated between beta-1 selective beta-blockers and the Arg389Gly polymorphism (rs1801253) in the adrenergic beta-1 receptor gene (ADRB1). We studied the effect of the ADRB1 Arg389Gly polymorphism on plasma renin activity (PRA) and heart rate (HR) and the genotype...

  7. Antithrombotic effects of beta2-adrenergic receptor blockade on top of beta1-receptor blockade in patients with acute coronary syndrome or heart failure : a systematic review

    NARCIS (Netherlands)

    De Peuter, O.R.; Lussana, F.; Peters, R.J.; Büller, H.R.; Kamphuisen, P.W.

    2009-01-01

    Background: Non-selective beta1 + 2 blockers may have specific antithrombotic effects not present in eta1-blockers, due to a eta2-specific effect on sympathetic activity. Our aim was to assess the influence of eta2-receptor suppression on top of selective beta1-receptor blockade on the occurrence of

  8. Role of TGF-beta1-independent changes in protein neosynthesis, p38alphaMAPK, and cdc42 in hydrogen peroxide-induced senescence-like morphogenesis

    DEFF Research Database (Denmark)

    Chrétien, Aline; Dierick, Jean-François; Delaive, Edouard; Larsen, Martin Røssel; Dieu, Marc; Raes, Martine; Deroanne, Christophe F; Roepstorff, Peter; Toussaint, Olivier

    2008-01-01

    The role of TGF-beta1 in hydrogen peroxide-induced senescence-like morphogenesis has been described. The aim of this work was to investigate whether TGF-beta1-independent changes in protein synthesis are involved in this morphogenesis and to study possible mechanisms occurring earlier than TGF-be...

  9. Immunogenicity of Recombinant Human Interferon Beta-1b in Immune-Tolerant Transgenic Mice Corresponds with the Biophysical Characteristics of Aggregates

    NARCIS (Netherlands)

    Haji Abdolvahab, Mohadeseh; Fazeli, Ahmad; Halim, Andhyk; Sediq, Ahmad S; Fazeli, Mohammad Reza; Schellekens, Huub

    2016-01-01

    Determining to what extent biophysical characteristics of aggregates affect immunogenicity of therapeutic interferon beta-1b. Three recombinant human interferon beta-1b (rhIFNβ-1b) samples with different levels of aggregates generated by copper oxidation, thermal stress, or left untreated, as well a

  10. Silencing of VAMP3 inhibits cell migration and integrin-mediated adhesion

    International Nuclear Information System (INIS)

    Integrins are transmembrane receptors for cell adhesion to the extracellular matrix. In cell migration, integrins are endocytosed from the plasma membrane or the cell surface, transported in vesicles and exocytosed actively at the cell front. In the present study, we examined the roles of VAMP3, a SNARE protein that mediates exocytosis, in cell migration and integrin trafficking. Small interfering RNA (siRNA)-induced silencing of VAMP3 inhibited chemotactic cell migration by more than 60% without affecting cell proliferation. VAMP3 silencing reduced the levels of β1 integrin at the cell surface but had no effect on total cellular β1 integrin, indicating that VAMP3 is required for trafficking of β1 integrin to the plasma membrane. Furthermore, VAMP3 silencing diminished cell adhesion to laminin but not to fibronectin or collagen. Taken together, these data suggest that VAMP3-dependent integrin trafficking is crucial in cell migration and cell adhesion to laminin.

  11. PET Imaging of Integrin αVβ3 Expression

    Directory of Open Access Journals (Sweden)

    Ambros J. Beer, Horst Kessler, Hans-Jürgen Wester, Markus Schwaiger

    2011-01-01

    Full Text Available PET imaging of integrin αvβ3 expression has been studied intensely by the academia and recently also by the industry. Imaging of integrin αvβ3 expression is of great potential value, as the integrin αvβ3 is a key player in tumor metastasis and angiogenesis. Therefore PET imaging of this target might be a suitable in-vivo biomarker of angiogenesis and metastatic potential of tumors. In this manuscript, the various strategies for PET imaging of the integrin αvβ3 will be summarized, including monomeric and multimeric radiolabelled RGD peptides and nanoparticles. While most experiments have been performed using preclinical tumor models, more and more clinical results on PET imaging of αvβ3 expression are available and will be discussed in detail. However, while a multitude of radiotracer strategies have been successfully evaluated for PET imaging of αvβ3, the ultimate clinical value of this new imaging biomarker still has to be evaluated in large clinical trials.

  12. INTERFERON BETA-1A TREATMENT IN HTLV-1-ASSOCIATED MYELOPATHY/TROPICAL SPASTIC PARAPARESIS: A CASE REPORT

    Directory of Open Access Journals (Sweden)

    Graça Maria de Castro Viana

    2014-09-01

    Full Text Available Here a young patient (< 21 years of age with a history of infective dermatitis is described. The patient was diagnosed with myelopathy associated with HTLV-1/tropical spastic paraparesis and treated with interferon beta-1a. The disease was clinically established as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP, and laboratory tests confirmed the presence of antibodies to HTLV-1 in the cerebrospinal fluid (CSF. Mumps, cytomegalovirus, Epstein-Barr virus, schistosomiasis, herpes virus 1 and 2, rubella, measles, varicella-zoster toxoplasmosis, hepatitis, HIV, and syphilis were excluded by serology. The patient was diagnosed with neurogenic bladder and presented with nocturia, urinary urgency, paresthesia of the lower left limb, a marked reduction of muscle strength in the lower limbs, and a slight reduction in upper limb strength. During the fourth week of treatment with interferon beta-1a, urinary urgency and paresthesia disappeared and clinical motor skills improved.

  13. Role of interleukin-10 and transforming growth factor beta 1 in otitis media with effusion

    Institute of Scientific and Technical Information of China (English)

    ZHAO Shou-qin; LI Jie; LIU Hua; ZHANG Quan-geng; WANG Yang; HAN De-min

    2009-01-01

    Background Otitis media with effusion (OME) is a disease with complicated pathogeneses which are not clearly known. Increasing interest has been focused on immunological cells, cytokines and their roles in chronic inflammatory states. This study was designed to disclose the existence and roles of interleukin-10 (IL-10) and transforming growth factor beta1 (TGF-β1) in the cause of OME in adults, and to investigate the probable role of Foxp3+CD4+CD25+ T cells in OME.Methods The concentrations of IL-10 and TGF-β1 in the middle ear effusions (MEEs) and plasmas of 36 adults (45 ears) with OME were measured by means of enzyme linked immunosorbent assay (ELISA). As contrast, the concentrations of IL-10 and TGF-β1 in the plasma of 30 normal volunteers were measured using the same method. Furthermore, the proportion of Foxp3+CD4+CD25+ T cells in CD4+ T cells of blood was tested by flow cytometry. Results (1) The concentrations of IL-10 in all MEEs and plasmas of the chronic OME patients were higher than those in patients with acute OME (both P 0.05). The concentration of IL-10 in MEEs had a strong correlation with the duration of the illness (r=0.547, P<0.01). The same correlation was also found between the concentration of TGF-β1 in MEEs and the times patients being treated (r=0.579, P <0.01). (3) The proportion of Foxp3+CD4+CD25+T/CD4+ T cells in the blood of chronic OME was not only significantly higher than that in the acute OME (P<0.01), but also higher than that in normal volunteers (P <0.01). In chronic OME, there was a correlation between the proportion of Foxp3+CD4+CD25+ T/CD4+ T cells in the blood and the concentration of IL-10 in the plasmas (r=0.602, P <0.05). Conclusions IL-10 and TGF-β1, as two important immunoregulatory mediators, participate in middle ear inflammatory response, especially in chronic course of OME in adults. Foxp3+CD4+CD25+ T cells may play some immunoregulatory roles in the course of this disease.

  14. Translation of myelin basic protein mRNA in oligodendrocytes is regulated by integrin activation and hnRNP-K

    DEFF Research Database (Denmark)

    Laursen, Lisbeth Schmidt; Chan, Colin W; ffrench-Constant, Charles

    2011-01-01

    Myelination in the central nervous system provides a unique example of how cells establish asymmetry. The myelinating cell, the oligodendrocyte, extends processes to and wraps multiple axons of different diameter, keeping the number of wraps proportional to the axon diameter. Local regulation of...... translation of a key sheath protein, myelin basic protein (MBP), by reversing the inhibitory effect of the mRNA 3′UTR. During oligodendrocyte differentiation and myelination α6β1-integrin interacts with hnRNP-K, an mRNA-binding protein, which binds to MBP mRNA and translocates from the nucleus to the myelin...

  15. C-peptide reverses TGF-beta1-induced changes in renal proximal tubular cells: implications for treatment of diabetic nephropathy.

    Science.gov (United States)

    Hills, Claire E; Al-Rasheed, Nawal; Al-Rasheed, Nouf; Willars, Gary B; Brunskill, Nigel J

    2009-03-01

    The crucial pathology underlying progressive chronic kidney disease in diabetes is tubulointerstitial fibrosis. Central to this process is epithelial-mesenchymal transformation (EMT) of proximal tubular epithelial cells driven by maladaptive transforming growth factor-beta1 (TGF-beta1) signaling. Novel signaling roles for C-peptide have recently been discovered with evidence emerging that C-peptide may mitigate microvascular complications of diabetes. We studied the potential for C-peptide to interrupt injurious TGF-beta1 signaling pathways and thus block development of EMT in HK2 human kidney proximal tubular cells. Cells were incubated with TGF-beta1 either alone or with C-peptide in low or high glucose. Changes in cell morphology, TGF-beta1 receptor expression, vimentin, E-cadherin, and phosphorylated Smads were assessed. Luciferase reporters were used to assess Smad activity. The cytoskeleton was visualized by TRITC-phalloidin staining. The typical TGF-beta1-stimulated, EMT-associated morphological alterations of proximal tubular cells, including increased vimentin expression, decreased E-cadherin expression, and cytoskeletal rearrangements, were prevented by C-peptide treatment. C-peptide also blocked TGF-beta1-induced upregulation of expression of both type I and type II TGF-beta1 receptors and attenuated TGF-beta1-mediated Smad phosphorylation and Smad transcriptional activity. These effects of C-peptide were inhibited by pertussis toxin. The results demonstrate that C-peptide almost completely reversed the morphological changes in PT cells induced by TGF-beta1 and suggest a role or C-peptide as a renoprotective agent in diabetic nephropathy. PMID:19091788

  16. Insulin-like growth factors require phosphatidylinositol 3-kinase to signal myogenesis: dominant negative p85 expression blocks differentiation of L6E9 muscle cells.

    Science.gov (United States)

    Kaliman, P; Canicio, J; Shepherd, P R; Beeton, C A; Testar, X; Palacín, M; Zorzano, A

    1998-01-01

    Phosphatidylinositol 3 (PI 3)-kinases are potently inhibited by two structurally unrelated membrane-permeant reagents: wortmannin and LY294002. By using these two inhibitors we first suggested the involvement of a PI 3-kinase activity in muscle cell differentiation. However, several reports have described that these compounds are not as selective for PI 3-kinase activity as assumed. Here we show that LY294002 blocks the myogenic pathway elicited by insulin-like growth factors (IGFs), and we confirm the specific involvement of PI 3-kinase in IGF-induced myogenesis by overexpressing in L6E9 myoblasts a dominant negative p85 PI 3-kinase-regulatory subunit (L6E9-delta p85). IGF-I, des(1-3)IGF-I, or IGF-II induced L6E9 skeletal muscle cell differentiation as measured by myotube formation, myogenin gene expression, and GLUT4 glucose carrier induction. The addition of LY294002 to the differentiation medium totally inhibited these IGF-induced myogenic events without altering the expression of a non-muscle-specific protein, beta1-integrin. Independent clones of L6E9 myoblasts expressing a dominant negative mutant of the p85-regulatory subunit (delta p85) showed markedly impaired glucose transport activity and formation of p85/p110 complexes in response to insulin, consistent with the inhibition of PI 3-kinase activity. IGF-induced myogenic parameters in L6E9-delta p85 cells, ie. cell fusion and myogenin gene and GLUT4 expression, were severely impaired compared with parental cells or L6E9 cells expressing wild-type p85. In all, data presented here indicate that PI 3-kinase is essential for IGF-induced muscle differentiation and that the specific PI 3-kinase subclass involved in myogenesis is the heterodimeric p85-p110 enzyme. PMID:9440811

  17. Clinical efficacy of beta1 selective adrenergic blockers in the treatment of neurocardiogenic syncope – a meta-analysis

    OpenAIRE

    Vallurupalli, Srikanth

    2010-01-01

    Srikanth Vallurupalli1, Smita Das21Division of General Internal Medicine, Southern Illinois University School of Medicine, Springfield, IL, USA; 2Department of Kinesiology and Community Health, University of Illinois College of Medicine, Champaign, IL, USABackground: Beta1 (B1) selective blockers have been widely used for the treatment of neurocardiogenic syncope though clinical trials have shown conflicting degrees of efficacy.Objective: To study the clinical efficacy of B1 selective blocker...

  18. Cloning and in-silico analysis of beta-1,3-xylanase from psychrophilic yeast, Glaciozyma antarctica PI12

    Science.gov (United States)

    Nor, Nooraisyah Mohamad; Bakar, Farah Diba Abu; Mahadi, Nor Muhammad; Murad, Abdul Munir Abdul

    2015-09-01

    A beta-1,3-xylanase (EC 3.2.1.32) gene from psychrophilic yeast, Glaciozyma antarctica has been identified via genome data mining. The enzyme was grouped into GH26 family based on Carbohydrate Active Enzyme (CaZY) database. The molecular weight of this protein was predicted to be 42 kDa and is expected to be soluble for expression. The presence of signal peptide suggested that this enzyme may be released extracellularly into the marine environment of the host's habitat. This supports the theory that such enzymatic activity is required for degradation of nutrients of polysaccharide origins into simpler carbohydrates outside the environment before it could be taken up inside the cell. The sequence for this protein showed very little conservation (< 30%) with other beta-1,3-xylanases from available databases. Based on the phylogenetic analysis, this protein also showed distant relationship to other xylanases from eukaryotic origin. The protein may have undergone major substitution in its gene sequence order to adapt to the cold climate. This is the first report of beta-1,3-xylanase gene isolated from a psychrophilic yeast.

  19. Transforming growth factor beta 1 effects on cartilage tissue metabolism%转化生长因子β1对软骨组织代谢影响的研究进展*★

    Institute of Scientific and Technical Information of China (English)

    郭铁峰; 周明旺; 李盛华; 孙凤岐; 穆欢喜

    2013-01-01

      背景:转化生长因子β1可以介导软骨合成、抑制胶原和蛋白多糖分解,在诱导软骨分化和维持软骨表型上起着重要作用,实现软骨缺损的功能性修复。  目的:从生物学特性、在生物工程中的应用、基因多态性、信号通路及微小 RNA等方面综述转化生长因子β1对软骨组织代谢影响的研究进展。  方法:以“transforming growth factor-β1,Cartilage Differentiation,cartilage matrix”为英文检索词,以“转化生长因子β1,软骨分化,软骨基质”为中文检索词。经第一作者检索2007/2012CNKI数据库及SPRINGERLINK数据库有关转化生长因子β1对软骨组织代谢影响的研究进展方面的文献130篇,根据纳入排除标准保留54篇进行总结。  结果与结论:转化生长因子β1可诱导间充质细胞向软骨细胞分化,促进软骨特异性基质的合成,保护软骨基质不被各种蛋白酶水解破坏,能够增强软骨组织自身再生能力,实现使软骨的损伤逆转,在软骨修复领域展现了巨大的潜在应用价值。%BACKGROUND:Transforming growth factor beta 1 can mediate cartilage synthesis and inhibit decomposition of col agen and protein polysaccharide, which has a most important effect on induction of cartilage differentiation in vitro and maintenance of cartilage phenotype, realizing the functional repair of cartilage defects. OBJECTIVE:Based on the biological characteristics, applications in biotechnology, gene polymorphism, signaling pathways and microRNA, to introduce research progress of transforming growth factor beta 1 influence on cartilage tissue metabolism. METHODS:The first author searched CNKI and SPRINGERLINK databases (2007/2012) to retrieve articles related to transforming growth factor beta 1 influence on cartilage tissue metabolism using the key words of“transforming growth factor beta 1, cartilage differentiation, cartilage matrix

  20. Identification of retinoic acid in a high content screen for agents that overcome the anti-myogenic effect of TGF-beta-1.

    Directory of Open Access Journals (Sweden)

    Chateen Krueger

    Full Text Available BACKGROUND: Transforming growth factor beta 1 (TGF-β1 is an inhibitor of muscle cell differentiation that is associated with fibrosis, poor regeneration and poor function in some diseases of muscle. When neutralizing antibodies to TGF-β1 or the angiotensin II inhibitor losartan were used to reduce TGF-β1 signaling, muscle morphology and function were restored in mouse models of Marfan Syndrome and muscular dystrophy. The goal of our studies was to identify additional agents that overcome the anti-myogenic effect of TGF-β1. METHODOLOGY/PRINCIPAL FINDINGS: A high-content cell-based assay was developed in a 96-well plate format that detects the expression of myosin heavy chain (MHC in C2C12 cells. The assay was used to quantify the dose-dependent responses of C2C12 cell differentiation to TGF-β1 and to the TGF-β1 Type 1 receptor kinase inhibitor, SB431542. Thirteen agents previously described as promoting C2C12 differentiation in the absence of TGF-β1 were screened in the presence of TGF-β1. Only all-trans retinoic acid and 9-cis retinoic acid allowed a maximal level of C2C12 cell differentiation in the presence of TGF-β1; the angiotensin-converting enzyme inhibitor captopril and 10 nM estrogen provided partial rescue. Vitamin D was a potent inhibitor of retinoic acid-induced myogenesis in the presence of TGF-β1. TGF-β1 inhibits myoblast differentiation through activation of Smad3; however, retinoic acid did not inhibit TGF-β1-induced activation of a Smad3-dependent reporter gene in C2C12 cells. CONCLUSIONS/SIGNIFICANCE: Retinoic acid alleviated the anti-myogenic effect of TGF-β1 by a Smad3-independent mechanism. With regard to the goal of improving muscle regeneration and function in individuals with muscle disease, the identification of retinoic acid is intriguing in that some retinoids are already approved for human therapy. However, retinoids also have well-described adverse effects. The quantitative, high-content assay will be

  1. LFA-1 and Mac-1 integrins bind to the serine/threonine-rich domain of thrombomodulin.

    Science.gov (United States)

    Kawamoto, Eiji; Okamoto, Takayuki; Takagi, Yoshimi; Honda, Goichi; Suzuki, Koji; Imai, Hiroshi; Shimaoka, Motomu

    2016-05-13

    LFA-1 (αLβ2) and Mac-1 (αMβ2) integrins regulate leukocyte trafficking in health and disease by binding primarily to IgSF ligand ICAM-1 and ICAM-2 on endothelial cells. Here we have shown that the anti-coagulant molecule thrombomodulin (TM), found on the surface of endothelial cells, functions as a potentially new ligand for leukocyte integrins. We generated a recombinant extracellular domain of human TM and Fc fusion protein (TM-domains 123-Fc), and showed that pheripheral blood mononuclear cells (PBMCs) bind to TM-domains 123-Fc dependent upon integrin activation. We then demonstrated that αL integrin-blocking mAb, αM integrin-blocking mAb, and β2 integrin-blocking mAb inhibited the binding of PBMCs to TM-domains 123-Fc. Furthermore, we show that the serine/threonine-rich domain (domain 3) of TM is required for the interaction with the LFA-1 (αLβ2) and Mac-1 (αMβ2) integrins to occur on PBMCs. These results demonstrate that the LFA-1 and Mac-1 integrins on leukocytes bind to TM, thereby establishing the molecular and structural basis underlying LFA-1 and Mac-1 integrin interaction with TM on endothelial cells. In fact, integrin-TM interactions might be involved in the dynamic regulation of leukocyte adhesion with endothelial cells. PMID:27055590

  2. Mutually Exclusive Roles of SHARPIN in Integrin Inactivation and NF-κB Signaling

    Science.gov (United States)

    De Franceschi, Nicola; Peuhu, Emilia; Parsons, Maddy; Rissanen, Sami; Vattulainen, Ilpo; Salmi, Marko

    2015-01-01

    SHANK-associated RH domain interactor (SHARPIN) inhibits integrins through interaction with the integrin α-subunit. In addition, SHARPIN enhances nuclear factor-kappaB (NF-κB) activity as a component of the linear ubiquitin chain assembly complex (LUBAC). However, it is currently unclear how regulation of these seemingly different roles is coordinated. Here, we show that SHARPIN binds integrin and LUBAC in a mutually exclusive manner. We map the integrin binding site on SHARPIN to the ubiquitin-like (UBL) domain, the same domain implicated in SHARPIN interaction with LUBAC component RNF31 (ring finger protein 31), and identify two SHARPIN residues (V267, L276) required for both integrin and RNF31 regulation. Accordingly, the integrin α-tail is capable of competing with RNF31 for SHARPIN binding in vitro. Importantly, the full SHARPIN RNF31-binding site contains residues (F263A/I272A) that are dispensable for SHARPIN-integrin interaction. Importantly, disrupting SHARPIN interaction with integrin or RNF31 abolishes SHARPIN-mediated regulation of integrin or NF-κB activity, respectively. Altogether these data suggest that the roles of SHARPIN in inhibiting integrin activity and supporting linear ubiquitination are (molecularly) distinct. PMID:26600301

  3. Neutralization of pathogenic beta1-receptor autoantibodies by aptamers in vivo: the first successful proof of principle in spontaneously hypertensive rats.

    Science.gov (United States)

    Haberland, Annekathrin; Wallukat, Gerd; Berg, Sabine; Schulz, Angela-Martina; Freyse, Ernst-Joachim; Vetter, Roland; Salzsieder, Eckhard; Müller, Johannes; Kreutz, Reinhold; Schimke, Ingolf

    2014-08-01

    Autoantibodies (AABs) against the second extracellular loop of the beta1-receptor (beta1(II)-AABs) are found as a pathogenic driver in patients with idiopathic dilated cardiomyopathy, Chagas cardiomyopathy, peripartum cardiomyopathy, and myocarditis, and have been increasingly seen as a treatment target. We recently identified an aptamer (single short DNA strand) that specifically binds and neutralizes beta1(II)-AABs. Via application of this aptamer, a new treatment strategy for diseases associated with the cardio-pathogenic beta1(II)-AABs could be developed. Spontaneously hypertensive rats (SHR) positive for beta1(II)-AABs were treated five times at weekly intervals (bolus application of 2 mg/kg body weight followed by an infusion of the same amount over 20 min). SHR responded to aptamer treatment with a strong reduction in the cardio-pathogenic beta1(II)-AABs. The AABs did not substantially return within the study period. No signs for aptamer toxicity were observed by visual examination of the heart, liver, and kidney, or by measurement of plasma CK, ALT, and creatinine. The aptamer's potential for beta1(II)-AAB neutralization and consequently for cardiomyopathy treatment has been shown for the first time in vivo. PMID:24744110

  4. Effect of beta 1,3 glucan in stress responses of the pencilfish (Nannostomus trifasciatus during transport within the rio Negro basin

    Directory of Open Access Journals (Sweden)

    Janessa S. Abreu

    2014-09-01

    Full Text Available We investigated the use of beta 1,3 glucan as an imunostimulant during a transport experiment to determine the effects upon the stress response of the pencilfish (Nannostomus trifasciatus. Pencilfish were fed for seven days with different concentrations of beta 1,3 glucan: 0.0% (control; 0.01%; 0.1% and 0.5% of beta 1,3 glucan per kg of feed-1. Fish were then transported for 24 hours by boat from Barcelos to Manaus. The highest dose of beta 1,3 glucan in the food increased Na+influx after 12 hours of transport and 0.1 and 0.5% beta 1,3 glucan maintained the flux of both ions close to zero at 24 hours. All doses of beta 1,3 glucan reduced K+ loss significantly in the beginning of the transport, but after 12 to 24 hours did not. No significant differences in whole body cortisol or survival were observed. Our results indicate that pencilfish had ionic alterations during transport from Barcelos to Manaus. The lack of significant differences in whole body cortisol and survival rate in addition to the maintenance of Na+ and K+ balance during transport reinforce the positive effects of beta 1,3 glucan immunostimulant on fish homeostasis. Therefore, we recommend its addition to food prior to transport.

  5. E6 and E7 oncoproteins from human papillomavirus type 16 induce activation of human transforming growth factor beta1 promoter throughout Sp1 recognition sequence.

    Science.gov (United States)

    Peralta-Zaragoza, Oscar; Bermúdez-Morales, Víctor; Gutiérrez-Xicotencatl, Lourdes; Alcocer-González, Juan; Recillas-Targa, Félix; Madrid-Marina, Vicente

    2006-01-01

    Human Papillomavirus (HPV) infection is the main etiologic agent of cervical cancer and HPV E6 and E7 oncogenes trans-regulate many cellular genes. An association between TGF-beta1 gene expression and cervical cancer development has been suggested; however, the mechanisms by which HPV influences TGF-beta1 expression remain unclear. In the present study we analyzed the mechanism through which HPV-16 E6 and E7 oncoproteins regulate the TGF-beta1 promoter in cervical tumor cells. Our results showed that E6 and E7 increased TGF-beta1 promoter activity. Furthermore, we identified a specific DNA sequence motif in the TGF-beta1 core promoter that is responsible for trans-activation and that corresponds to the Sp1e-binding site associated with HPV-16 E6 and E7 oncoproteins. Mutational analysis showed that the Sp1e recognition site abolished the trans-activation caused by E6 and E7. These results suggest a physical interaction and functional cooperation between viral oncoproteins and cellular regulatory elements of the TGF-beta1 promoter, and may explain the contribution of HPV-16 to TGF-beta1 gene expression in cervical cancer. PMID:16987065

  6. Osteopontin binding to the alpha 4 integrin requires highest affinity integrin conformation, but is independent of post-translational modifications of osteopontin

    DEFF Research Database (Denmark)

    Hui, Tommy; Sørensen, Esben Skipper; Rittling, Susan R.

    2015-01-01

    Osteopontin (OPN) is a ligand for the α4 integrin, but the physiological importance of this binding is not well understood. Here, we have assessed the effect of posttranslational modifications on OPN binding to the α4 integrin on cultured human leukocyte cell lines, and compared OPN interaction...

  7. Thalidomide and the immune system. 3. Simultaneous up- and down-regulation of different integrin receptors on human white blood cells.

    Science.gov (United States)

    Nogueira, A C; Neubert, R; Helge, H; Neubert, D

    1994-01-01

    Time-dependent changes in the surface receptor expression of various maturational and integrin receptors on peripheral blood cells were studied in two healthy human volunteers following oral applications of thalidomide (Thd). In each measurement the receptor density was quantified by prior calibration of the flow cytometer with latex beads bearing a determined number of fluorescence molecules. The effects observed in the course of the Thd-treatment were practically identical or at least very similar in both the volunteers during four different trials, and were in accord with previous results obtained in large-scale studies (68 treated animals) with non-human primates. It should be stressed that no clear-cut changes were observed in the percentage or absolute numbers of primary lymphocyte subsets such as CD3, CD4 and CD20. After the first two doses of 7 mg Thd/kg body wt the CD18 (the common beta-chain of the beta 2-integrins) marker already decreased in surface density or was no longer detectable on granulocytes, monocytes and lymphocytes. This effect persisted throughout the treatment period and slowly subsided after discontinuation of treatment. With a few days lag phase, the surface density of CD54 (ICAM-1) on granulocytes increased and many cells previously not bearing this receptor newly acquired such surface markers. On monocytes however, the CD54 receptor was lost on many cells. Within the lymphocyte fraction a loss of the CD54 marker could be noted on CD4 cells but not on CD8 cells, where an increase of the receptor expression could be observed. Other markers, such as the alpha chains of the beta 1 integrins CD49b (VLA alpha 2) and CD49d (VLA alpha 4) showed contrasting reactions to the Thd-treatment. Whereas a pronounced loss of the receptor density of CD49d was observed and only few cells with high epitope density were left in the blood at the end of the complete dosing schedule, no such effect was observable on cells bearing the CD49b epitope. A distinct

  8. Stimulation of Odontogenesis and Angiogenesis via Bioactive Nanocomposite Calcium Phosphate Cements Through Integrin and VEGF Signaling Pathways.

    Science.gov (United States)

    Lee, Sang-Im; Lee, Eui-Suk; El-Fiqi, Ahmed; Lee, So-Youn; Eun-Cheol Kim; Kim, Hae-Won

    2016-05-01

    Formulating self-setting calcium phosphate cements (CPCs) with secondary phases particularly in the nanoscale order holds great promise to improve biological properties. Here, we focus on the effect that bioactive glass nanoparticles (BGN) incorporated in CPC compositions can have on the proliferation, odontogenic differentiation, and angiogenic stimulation of stem cells derived from human dental pulp (HDPSCs). These odontogenic and angiogenic events are of special importance in the dentin-pulp regeneration processes. In comparison to pure CPCs, nanocomposite cements exhibit a significantly improved proliferation of HDPSCs, and the improvement is more significant as the BGN content increases. The nanocomposite cements substantially enhance the adhesion of cells, and significantly up-regulate odontogenic differentiation, including alkaline phosphatase (ALP) activity and the expressions of odontogenic genes (sialophosphoprotein, dentin matrix protein I, ALP, osteopontin and osteocalcin). Furthermore, the use of nanocomposite cements result in stimulation of angiogenic gene expression (VEGF, FGF-2, VEGFRs, PECAM-1, and VE-cadherin) and protein production (VEGF, VEGFR-1). The angiogenic stimulation by the HDPSCs significantly affects the endothelial cell behaviors, that is, the endothelial cell migration and the tubular network formation are substantially improved when treated with HDPSC-conditioned medium, particularly with the help of nanocomposite cements. The integrin and VEGF signaling pathways are reasoned for the stimulation of the odontogenesis and angiogenesis of cells, where the nanocomposite cements up-regulate the integrin subsets α1, α2, α3, and β1, and activate the integrin downstream signal pathways, such as p-FAK, p-Akt, p-paxillin, JNK, EK, and NF-κB, as well as other nuclear transcriptional factors, including CREB, STAT-3, and ELK-1. The current results indicate that the new formulation of the nanocomposite self-setting cements might provide some

  9. Isolation and overexpression of a gene encoding an extracellular beta-(1,3-1,4)-glucanase from Streptococcus bovis JB1.

    OpenAIRE

    Ekinci, M S; McCrae, S I; Flint, H.J. (Harry J.)

    1997-01-01

    Streptococcus bovis JB1 was found to produce a 25-kDa extracellular enzyme active against beta-(1,3-1,4)-glucans. A gene was isolated encoding a specific beta-(1,3-1,4)-glucanase that corresponds to this size and belongs to glycoside hydrolase family 16. A 4- to 10-fold increase in supernatant beta-glucanase activity was obtained when the cloned beta-glucanase gene was reintroduced into S. bovis JB1 by use of constructs based on the plasmid vector pTRW10 or pIL253. The beta-(1,3-1,4)-glucanas...

  10. Matrix crosslinking forces tumor progression by enhancing integrin signaling

    DEFF Research Database (Denmark)

    Levental, Kandice R; Yu, Hongmei; Kass, Laura; Lakins, Johnathon N; Egeblad, Mikala; Erler, Janine Terra; Fong, Sheri F T; Csiszar, Katalin; Giaccia, Amato; Weninger, Wolfgang; Yamauchi, Mitsuo; Gasser, David L; Weaver, Valerie M

    2009-01-01

    Tumors are characterized by extracellular matrix (ECM) remodeling and stiffening. The importance of ECM remodeling to cancer is appreciated; the relevance of stiffening is less clear. We found that breast tumorigenesis is accompanied by collagen crosslinking, ECM stiffening, and increased focal...... adhesions. Induction of collagen crosslinking stiffened the ECM, promoted focal adhesions, enhanced PI3 kinase (PI3K) activity, and induced the invasion of an oncogene-initiated epithelium. Inhibition of integrin signaling repressed the invasion of a premalignant epithelium into a stiffened, crosslinked ECM...... and forced integrin clustering promoted focal adhesions, enhanced PI3K signaling, and induced the invasion of a premalignant epithelium. Consistently, reduction of lysyl oxidase-mediated collagen crosslinking prevented MMTV-Neu-induced fibrosis, decreased focal adhesions and PI3K activity, impeded...

  11. Carbon Ion Irradiation Inhibits Glioma Cell Migration Through Downregulation of Integrin Expression

    International Nuclear Information System (INIS)

    Purpose: To investigate the effect of carbon ion irradiation on glioma cell migration. Methods and Materials: U87 and Ln229 glioma cells were irradiated with photons and carbon ions. Migration was analyzed 24 h after irradiation. Fluorescence-activated cell sorting analysis was performed in order to quantify surface expression of integrins. Results: Single photon doses of 2 Gy and 10 Gy enhanced ανβ3 and ανβ5 integrin expression and caused tumor cell hypermigration on both vitronectin (Vn) and fibronectin (Fn). Compared to integrin expression in unirradiated cells, carbon ion irradiation caused decreased integrin expression and inhibited cell migration on both Vn and Fn. Conclusion: Photon radiotherapy (RT) enhances the risk of tumor cell migration and subsequently promotes locoregional spread via photon induction of integrin expression. In contrast to photon RT, carbon ion RT causes decreased integrin expression and suppresses glioma cell migration on both Vn and Fn, thus promising improved local control.

  12. Augmentation of integrin-mediated mechanotransduction by hyaluronic acid

    OpenAIRE

    Chopra, Anant; Murray, Maria E.; Byfield, Fitzroy; Mendez, Melissa; Halleluyan, Ran; Restle, David; Aroush, Dikla Raz-Ben; Peter A. Galie; Pogoda, Katarzyna; Bucki, Robert; Marcinkiewicz, Cezary; Prestwich, Glenn D.; Zarembinski, Thomas; Chen, Christopher S.; Puré, Ellen

    2013-01-01

    Changes in tissue and organ stiffness occur during development and are frequently symptoms of disease. Many cell types respond to the stiffness of substrates and neighboring cells in vitro and most cell types increase adherent area on stiffer substrates that are coated with ligands for integrins or cadherins. In vivo cells engage their extracellular matrix (ECM) by multiple mechanosensitive adhesion complexes and other surface receptors that potentially modify the mechanical signals transduce...

  13. Integrin-dependent Neutrophil Migration in the Injured Mouse Cornea

    OpenAIRE

    Hanlon, Samuel D.; Smith, C. Wayne; Sauter, Marika N; Burns, Alan R.

    2014-01-01

    As an early responder to an inflammatory stimulus, neutrophils (PMNs) must exit the vasculature and migrate through the extravascular tissue to the site of insult, which is often remote from the point of extravasation. Following a central epithelial corneal abrasion, PMNs recruited from the peripheral limbal vasculature migrate into the avascular corneal stroma. In vitro studies suggest PMN locomotion over 2-D surfaces is dependent on integrin binding while migration within 3-D matrices can b...

  14. β1 Integrin Is Essential for Teratoma Growth and Angiogenesis

    OpenAIRE

    Bloch, Wilhelm; Forsberg, Erik; Lentini, Sylvia; Brakebusch, Cord; Martin, Karl; Krell, Hans W.; Weidle, Ulrich H.; Addicks, Klaus; Fässler, Reinhard

    1997-01-01

    Teratomas are benign tumors that form after ectopic injection of embryonic stem (ES) cells into mice and contain derivatives of all primitive germ layers. To study the role of β1 integrin during teratoma formation, we compared teratomas induced by normal and β1-null ES cells. Injection of normal ES cells gave rise to large teratomas. In contrast, β1-null ES cells either did not grow or formed small teratomas with an average weight of

  15. Roles of integrin activation in eosinophil function and the eosinophilic inflammation of asthma

    OpenAIRE

    Barthel, Steven R; Johansson, Mats W.; McNamee, Dawn M.; Deane F Mosher

    2007-01-01

    Eosinophilic inflammation is a characteristic feature of asthma. Integrins are highly versatile cellular receptors that regulate extravasation of eosinophils from the postcapillary segment of the bronchial circulation to the airway wall and airspace. Such movement into the asthmatic lung is described as a sequential, multistep paradigm, whereby integrins on circulating eosinophils become activated, eosinophils tether in flow and roll on bronchial endothelial cells, integrins on rolling eosino...

  16. CCN2: a mechanosignaling sensor modulating integrin-dependent connective tissue remodeling in fibroblasts?

    OpenAIRE

    Leask, Andrew

    2013-01-01

    Tensegrity (tensional integrity) is an emerging concept governing the structure of the body. Integrin-mediated mechanical tension is essential for connective tissue function in vivo. For example, in adult skin fibroblasts, the integrin β1 subunit mediates adhesion to collagen and fibronectin. Moreover, integrin β1, through its abilities to activate latent TGFβ1 and promote collagen production through focal adhesion kinase/rac1/nicotinamide adenine dinucleotide phosphate oxidase (NOX)/reactive...

  17. Inhibition of a novel specific neuroglial integrin signaling pathway increases STAT3-mediated CNTF expression

    OpenAIRE

    Keasey, Matthew P.; Kang, Seong Su; Lovins, Chiharu; Hagg, Theo

    2013-01-01

    Background Ciliary neurotrophic factor (CNTF) expression is repressed in astrocytes by neuronal contact in the CNS and is rapidly induced by injury. Here, we defined an inhibitory integrin signaling pathway. Results The integrin substrates laminin, fibronectin and vitronectin, but not collagen, thrombospondin or fibrinogen, reduced CNTF expression in C6 astroglioma cells. Antibodies against αv and β5, but not α6 or β1, integrin induced CNTF. Together, the ligand and antibody specificity sugge...

  18. Polarized Integrin Mediates Human Keratinocyte Adhesion to Basal Lamina

    Science.gov (United States)

    de Luca, Michele; Tamura, Richard N.; Kajiji, Shama; Bondanza, Sergio; Rossino, Paola; Cancedda, Ranieri; Carlo Marchisio, Pier; Quaranta, Vito

    1990-09-01

    Epithelial cell interactions with matrices are critical to tissue organization. Indirect immunofluorescence and immunoprecipitations of cell lysates prepared from stratified cultures of human epidermal cells showed that the major integrins expressed by keratinocytes are α_Eβ_4 (also called α_6β_4) and α_2β_1/α_3β_1. The α_Eβ_4 integrin is localized at the surface of basal cells in contact with the basement membrane, whereas α_2β_1/ α_3β_1 integrins are absent from the basal surface and are localized only on the lateral surface of basal and spinous keratinocytes. Anti-β_4 antibodies potently inhibited keratinocyte adhesion to matrigel or purified laminin, whereas anti-β_1 antibodies were ineffective. Only anti-β_4 antibodies were able to detach established keratinocyte colonies. These data suggest that α_Eβ_4 mediates keratinocyte adhesion to basal lamina, whereas the β_1 subfamily is involved in cell-cell adhesion of keratinocytes.

  19. Integrin αβ3-Targeted Imaging of Lung Cancer

    Directory of Open Access Journals (Sweden)

    Xiaoyuan Chen

    2005-03-01

    Full Text Available A series of radiolabeled cyclic arginine-glycineaspartic acid (RGD peptide ligands for cell adhesion molecule integrin αβ3-targeted tumor angiogenesis targeting are being developed in our laboratory. In this study, this effort continues by applying a positron emitter 64Cu-labeled PEGylated dimeric RGD peptide radiotracer 64Cu-DOTA-PEG-E[c(RGDyK]2 for lung cancer imaging. The PEGylated RGD peptide indicated integrin αβ3 avidity, but the PEGylation reduced the receptor binding affinity of this ligand compared to the unmodified RGD dimer. The radiotracer revealed rapid blood clearance and predominant renal clearance route. The minimum nonspecific activity accumulation in normal lung tissue and heart rendered high-quality orthotopic lung cancer tumor images, enabling clear demarcation of both the primary tumor at the upper lobe of the left lung, as well as metastases in the mediastinum, contralateral lung, diaphragm. As a comparison, fluorodeoxyglucose (FDG scans on the same mice were only able to identify the primary tumor, with the metastatic lesions masked by intense cardiac uptake and high lung background. 64Cu-DOTA-PEGE[c(RGDyK]2 is an excellent positron emission tomography (PET tracer for integrin-positive tumor imaging. Further studies to improve the receptor binding affinity of the tracer and subsequently to increase the magnitude of tumor uptake without comprising the favorable in vivo kinetics are currently in progress.

  20. Linkage of two human pregnancy-specific beta 1-glycoprotein genes: one is associated with hydatidiform mole.

    OpenAIRE

    Leslie, K. K.; Watanabe, S; Lei, K J; Chou, D Y; Plouzek, C A; Deng, H C; Torres, J.; Chou, J Y

    1990-01-01

    A genomic clone containing two linked human pregnancy-specific beta 1-glycoprotein (PS beta G) genes has been isolated and characterized. The two genes are arranged in the same 5'----3' orientation; the 3' region (including the A2 and B-C exons) of the upstream gene, PSGGA, is linked to the 5' region (including the 5'/L and L/N exons) of PSGGB, the downstream gene. Depending upon the domains compared, PSGGA and PSGGB share 92-98% nucleotide and 86-95% amino acid sequence identity with PSG93, ...

  1. Beta-1 adrenergic agonist treatment mitigates negative changes in cancellous bone microarchitecture and inhibits osteocyte apoptosis during disuse.

    Science.gov (United States)

    Swift, Joshua M; Swift, Sibyl N; Allen, Matthew R; Bloomfield, Susan A

    2014-01-01

    The sympathetic nervous system (SNS) plays an important role in mediating bone remodeling. However, the exact role that beta-1 adrenergic receptors (beta1AR) have in this process has not been elucidated. We have previously demonstrated the ability of dobutamine (DOB), primarily a beta1AR agonist, to inhibit reductions in cancellous bone formation and mitigate disuse-induced loss of bone mass. The purpose of this study was to characterize the independent and combined effects of DOB and hindlimb unloading (HU) on cancellous bone microarchitecture, tissue-level bone cell activity, and osteocyte apoptosis. Male Sprague-Dawley rats, aged 6-mos, were assigned to either normal cage activity (CC) or HU (n = 18/group) for 28 days. Animals were administered either daily DOB (4 mg/kg BW/d) or an equal volume of saline (VEH) (n = 9/gp). Unloading resulted in significantly lower distal femur cancellous BV/TV (-33%), Tb.Th (-11%), and Tb.N (-25%) compared to ambulatory controls (CC-VEH). DOB treatment during HU attenuated these changes in cancellous bone microarchitecture, resulting in greater BV/TV (+29%), Tb.Th (+7%), and Tb.N (+21%) vs. HU-VEH. Distal femur cancellous vBMD (+11%) and total BMC (+8%) were significantly greater in DOB- vs. VEH-treated unloaded rats. Administration of DOB during HU resulted in significantly greater osteoid surface (+158%) and osteoblast surface (+110%) vs. HU-VEH group. Furthermore, Oc.S/BS was significantly greater in HU-DOB (+55%) vs. CC-DOB group. DOB treatment during unloading fully restored bone formation, resulting in significantly greater bone formation rate (+200%) than in HU-VEH rats. HU resulted in an increased percentage of apoptotic cancellous osteocytes (+85%), reduced osteocyte number (-16%), lower percentage of occupied osteocytic lacunae (-30%) as compared to CC-VEH, these parameters were all normalized with DOB treatment. Altogether, these data indicate that beta1AR agonist treatment during disuse mitigates negative

  2. Beta-1 adrenergic agonist treatment mitigates negative changes in cancellous bone microarchitecture and inhibits osteocyte apoptosis during disuse.

    Directory of Open Access Journals (Sweden)

    Joshua M Swift

    Full Text Available The sympathetic nervous system (SNS plays an important role in mediating bone remodeling. However, the exact role that beta-1 adrenergic receptors (beta1AR have in this process has not been elucidated. We have previously demonstrated the ability of dobutamine (DOB, primarily a beta1AR agonist, to inhibit reductions in cancellous bone formation and mitigate disuse-induced loss of bone mass. The purpose of this study was to characterize the independent and combined effects of DOB and hindlimb unloading (HU on cancellous bone microarchitecture, tissue-level bone cell activity, and osteocyte apoptosis. Male Sprague-Dawley rats, aged 6-mos, were assigned to either normal cage activity (CC or HU (n = 18/group for 28 days. Animals were administered either daily DOB (4 mg/kg BW/d or an equal volume of saline (VEH (n = 9/gp. Unloading resulted in significantly lower distal femur cancellous BV/TV (-33%, Tb.Th (-11%, and Tb.N (-25% compared to ambulatory controls (CC-VEH. DOB treatment during HU attenuated these changes in cancellous bone microarchitecture, resulting in greater BV/TV (+29%, Tb.Th (+7%, and Tb.N (+21% vs. HU-VEH. Distal femur cancellous vBMD (+11% and total BMC (+8% were significantly greater in DOB- vs. VEH-treated unloaded rats. Administration of DOB during HU resulted in significantly greater osteoid surface (+158% and osteoblast surface (+110% vs. HU-VEH group. Furthermore, Oc.S/BS was significantly greater in HU-DOB (+55% vs. CC-DOB group. DOB treatment during unloading fully restored bone formation, resulting in significantly greater bone formation rate (+200% than in HU-VEH rats. HU resulted in an increased percentage of apoptotic cancellous osteocytes (+85%, reduced osteocyte number (-16%, lower percentage of occupied osteocytic lacunae (-30% as compared to CC-VEH, these parameters were all normalized with DOB treatment. Altogether, these data indicate that beta1AR agonist treatment during disuse mitigates negative changes

  3. Platelet-derived growth factor-BB and transforming growth factor beta 1 selectively modulate glycosaminoglycans, collagen, and myofibroblasts in excisional wounds.

    OpenAIRE

    Pierce, G F; Vande Berg, J.; Rudolph, R; Tarpley, J.; Mustoe, T A

    1991-01-01

    Recombinant platelet-derived growth factor (PDGF) and transforming growth factor beta 1 (TGF-beta 1) influence the rate of extracellular matrix formed in treated incisional wounds. Because incisional healing processes are difficult to quantify, a full-thickness excisional wound model in the rabbit ear was developed to permit detailed analyses of growth-factor-mediated tissue repair. In the present studies, quantitative and qualitative differences in acute inflammatory cell influx, glycosamino...

  4. Enhanced tumor formation in cyclin D1 x transforming growth factor beta1 double transgenic mice with characterization by magnetic resonance imaging.

    Science.gov (United States)

    Deane, Natasha G; Lee, Haakil; Hamaamen, Jalal; Ruley, Anna; Washington, M Kay; LaFleur, Bonnie; Thorgeirsson, Snorri S; Price, Ronald; Beauchamp, R Daniel

    2004-02-15

    Transgenic mice that overexpress cyclin D1 protein in the liver develop liver carcinomas with high penetrance. Transforming growth factor beta (TGF-beta) serves as either an epithelial cell growth inhibitor or a tumor promoter, depending on the cellular context. We interbred LFABP-cyclin D1 and Alb-TGF-beta1 transgenic mice to produce cyclin D1/TGF-beta1 double transgenic mice and followed the development of liver tumors over time, characterizing cellular and molecular changes, tumor incidence, tumor burden, and tumor physiology noninvasively by magnetic resonance imaging. Compared with age-matched LFABP-cyclin D1 single transgenic littermates, cyclin D1/TGF-beta1 mice exhibited a significant increase in tumor incidence. Tumor multiplicity, tumor burden, and tumor heterogeneity were higher in cyclin D1/TGF-beta1 mice compared with single transgenic littermates. Characteristics of cyclin D1/TGF-beta1 livers correlated with a marked induction of the peripheral periductal oval cell/stem cell compartment of the liver. A number of cancerous lesions from cyclin D1/TGF-beta1 mice exhibited unique features such as ductal plate malformations and hemorrhagic nodules. Some lesions were contiguous with the severely diseased background liver and, in some cases, replaced the normal architecture of the entire organ. Cyclin D1/TGF-beta1 lesions, in particular, were associated with malignant features such as areas of vascular invasion by hepatocytes and heterogeneous hyperintensity of signal on T2-weighted magnetic resonance imaging. These findings demonstrate that TGF-beta1 promotes stem cell activation and tumor progression in the context of cyclin D1 overexpression in the liver. PMID:14973059

  5. Regulation of extracellular matrix synthesis by TNF-alpha and TGF-beta1 in type II cells exposed to coal dust.

    Science.gov (United States)

    Lee, Y C; Rannels, D E

    1998-10-01

    Type II pulmonary epithelial cells respond to anthracite coal dust PSOC 867 with increased synthesis of extracellular matrix (ECM) components. Alveolar macrophages modulate this response by pathways that may involve soluble mediators, including tumor necrosis factor-alpha (TNF-alpha) or transforming growth factor-beta1 (TGF-beta1). The effects of TNF-alpha (10 ng/ml) and/or TGF-beta1 (2 ng/ml) were thus investigated in dust-exposed primary type II cell cultures. In control day 1 or day 3 cultures, TNF-alpha and/or TGF-beta1 had little or no effect on the synthesis of type II cellular proteins, independent of whether the cells were exposed to dust. With PSOC 867 exposure, where ECM protein synthesis is elevated, TNF-alpha and TGF-beta1 further increased both the absolute and relative rates of ECM synthesis on day 3 but had little effect on day 1. Each mediator increased expression of fibronectin mRNA, as well as of ECM fibronectin content, in a manner qualitatively similar to their effects on synthesis. Thus TNF-alpha and TGF-beta1 modulate both ECM synthesis and fibronectin content in coal dust-exposed type II cell cultures. PMID:9755095

  6. Coexistence of beta 1- and beta 2-adrenoceptors in the rabbit heart: quantitative analysis of the regional distribution by (-)-3H-dihydroalprenolol binding

    International Nuclear Information System (INIS)

    We determined the amount of beta 1- and beta 2-adrenoceptors in right and left atria and ventricles of rabbits. For this purpose inhibition of specific (-)-3H-dihydroalprenolol [(-)-3H-DHA] binding (5 nM) by beta 1-selective (practolol, metoprolol) and beta 2-selective (zinterol, IPS 339) adrenergic drugs was determined and analyzed by pseudo-Scatchard (Hofstee) plots. For both atria, inhibition of binding by the four selective beta-adrenergic drugs resulted in non-linear Hofstee plots, suggesting the coexistence of both beta-adrenoceptor subtypes. From these plots we calculated a beta 1:beta 2-adrenoceptor ratio of 72:28 for the right atrium and of 82:18 for the left. In contrast, only a very small amount of beta 2-adrenoceptors (approximately 5-7% of the total beta-adrenoceptor population) could be detected in the ventricles. For comparison we analyzed the inhibition of specific (-)-3H-DHA binding in tissues with homogeneous population of beta-adrenoceptors (beta 1:guinea pig left ventricle; beta 2: cerebellum of mature rats). For both tissues the four selective beta-adrenergic drugs showed linear Hofstee plots, demonstrating that in tissues with homogeneous beta-receptor population interaction of each drug with the receptor followed simple mass-action kinetics. We conclude that beta 1- and beta 2-adrenoceptors coexist in rabbit atria while the ventricles are predominantly endowed the beta 1-adrenoceptors

  7. Coexistence of beta 1- and beta 2-adrenoceptors in the rabbit heart: quantitative analysis of the regional distribution by (-)-/sup 3/H-dihydroalprenolol binding

    Energy Technology Data Exchange (ETDEWEB)

    Brodde, O.E.; Leifert, F.J.; Krehl, H.J.

    1982-01-01

    We determined the amount of beta 1- and beta 2-adrenoceptors in right and left atria and ventricles of rabbits. For this purpose inhibition of specific (-)-/sup 3/H-dihydroalprenolol ((-)-/sup 3/H-DHA) binding (5 nM) by beta 1-selective (practolol, metoprolol) and beta 2-selective (zinterol, IPS 339) adrenergic drugs was determined and analyzed by pseudo-Scatchard (Hofstee) plots. For both atria, inhibition of binding by the four selective beta-adrenergic drugs resulted in non-linear Hofstee plots, suggesting the coexistence of both beta-adrenoceptor subtypes. From these plots we calculated a beta 1:beta 2-adrenoceptor ratio of 72:28 for the right atrium and of 82:18 for the left. In contrast, only a very small amount of beta 2-adrenoceptors (approximately 5-7% of the total beta-adrenoceptor population) could be detected in the ventricles. For comparison we analyzed the inhibition of specific (-)-/sup 3/H-DHA binding in tissues with homogeneous population of beta-adrenoceptors (beta 1:guinea pig left ventricle; beta 2: cerebellum of mature rats). For both tissues the four selective beta-adrenergic drugs showed linear Hofstee plots, demonstrating that in tissues with homogeneous beta-receptor population interaction of each drug with the receptor followed simple mass-action kinetics. We conclude that beta 1- and beta 2-adrenoceptors coexist in rabbit atria while the ventricles are predominantly endowed the beta 1-adrenoceptors.

  8. Localization of the {alpha}7 integrin gene (ITGA7) on human chromosome 12q13: Clustering of integrin and Hox genes implies parallel evolution of these gene families

    Energy Technology Data Exchange (ETDEWEB)

    Wang, W.; Wu, W.; Kaufman, S.J. [Univ. of Illinois, Urbana, IL (United States)] [and others

    1995-04-10

    Expression of the {alpha}7 integrin gene (ITGA7) is developmentally regulated during the formation of skeletal muscle. Increased levels of expression and production of isoforms containing different cytoplasmic and extracellular domains accompany myogenesis. To determine whether a single or multiple {alpha}7 gene(s) underlie the structural diversity in this alpha chain that accompanies development, we have examined the rat and human genomes by Southern blotting and in situ hybridization. Our results demonstrate that there is only one {alpha}7 gene in both the rat and the human genomes. In the human, ITGA7 is present on chromosome 12q13. Phylogenetic analysis of the integrin alpha chain sequences suggests that the early integrin genes evolved in two pathways to form the I-integrins and the non-I-integrins. The I-integrin alpha chains contain an additional sequence of approximately 180 amino acids and arose as a result of an early insertion into the non-I-gene. The I-chain subfamily further evolved by duplications within the same chromosome. The non-I-integrin alpha chain genes are localized in clusters on chromosomes 2, 12, and 17, and this closely coincides with the localization of the human homeobox gene clusters. Non-I-integrin alpha chain genes appear to have evolved in parallel and in proximity to the Hox clusters. Thus, the Hox genes that underlie the design of body structure and the Integrin genes that underlie informed cell-cell and cell-matrix interactions appear to have evolved in parallel and coordinate fashions. 52 refs., 5 figs., 2 tabs.

  9. Cilengitide inhibits proliferation and differentiation of human endothelial progenitor cells in vitro

    International Nuclear Information System (INIS)

    Bone marrow derived hematopoietic stem cells can function as endothelial progenitor cells. They are recruited to malignant tumors and differentiate into endothelial cells. This mechanism of neovascularization termed vasculogenesis is distinct from proliferation of pre-existing vessels. To better understand vasculogenesis we developed a cell culture model with expansion and subsequent endothelial differentiation of human CD133+ progenitor cells in vitro. αvβ3-integrins are expressed by endothelial cells and play a role in the attachment of endothelial cells to the extracellular matrix. We investigated the effect of Cilengitide, a peptide-like, high affinity inhibitor of αvβ3- and αvβ5-integrins in our in vitro system. We could show expression of αvβ3-integrin on 60 ± 9% of non-adherent endothelial progenitors and on 91 ± 7% of differentiated endothelial cells. αvβ3-integrin was absent on CD133+ hematopoietic stem cells. Cilengitide inhibited proliferation of CD133+ cells in a dose-dependent manner. The development of adherent endothelial cells from expanded CD133+ cells was reduced even stronger by Cilengitide underlining its effect on integrin mediated cell adhesion. Expression of endothelial antigens CD144 and von Willebrand factor on differentiating endothelial precursors was decreased by Cilengitide. In summary, Cilengitide inhibits proliferation and differentiation of human endothelial precursor cells underlining its anti-angiogenic effects

  10. The dermatan sulfate proteoglycan decorin modulates α2β1 integrin and the vimentin intermediate filament system during collagen synthesis.

    Directory of Open Access Journals (Sweden)

    Oliver Jungmann

    Full Text Available Decorin, a small leucine-rich proteoglycan harboring a dermatan sulfate chain at its N-terminus, is involved in regulating matrix organization and cell signaling. Loss of the dermatan sulfate of decorin leads to an Ehlers-Danlos syndrome characterized by delayed wound healing. Decorin-null (Dcn(-/- mice display a phenotype similar to that of EDS patients. The fibrillar collagen phenotype of Dcn(-/- mice could be rescued in vitro by decorin but not with decorin lacking the glycosaminoglycan chain. We utilized a 3D cell culture model to investigate the impact of the altered extracellular matrix on Dcn(-/- fibroblasts. Using 2D gel electrophoresis followed by mass spectrometry, we identified vimentin as one of the proteins that was differentially upregulated by the presence of decorin. We discovered that a decorin-deficient matrix leads to abnormal nuclear morphology in the Dcn(-/- fibroblasts. This phenotype could be rescued by the decorin proteoglycan but less efficiently by the decorin protein core. Decorin treatment led to a significant reduction of the α2β1 integrin at day 6 in Dcn(-/- fibroblasts, whereas the protein core had no effect on β1. Interestingly, only the decorin core induced mRNA synthesis, phosphorylation and de novo synthesis of vimentin indicating that the proteoglycan decorin in the extracellular matrix stabilizes the vimentin intermediate filament system. We could support these results in vivo, because the dermis of wild-type mice have more vimentin and less β1 integrin compared to Dcn(-/-. Furthermore, the α2β1 null fibroblasts also showed a reduced amount of vimentin compared to wild-type. These data show for the first time that decorin has an impact on the biology of α2β1 integrin and the vimentin intermediate filament system. Moreover, our findings provide a mechanistic explanation for the reported defects in wound healing associated with the Dcn(-/- phenotype.

  11. Quantitative gene-expression of the tumor angiogenesis markers vascular endothelial growth factor, integrin alphaV and integrin beta3 in human neuroendocrine tumors

    DEFF Research Database (Denmark)

    Oxboel, Jytte; Binderup, Tina; Knigge, Ulrich; Kjaer, Andreas

    2009-01-01

    molecules VEGF and integrin beta3 were lower in neuroendocrine tumors than in colorectal liver metastases and were highly variable. Therefore, individual selection of patients may be necessary if anti-angiogenesis treatment is to be successful in patients with neuroendocrine tumors......., in neuroendocrine tumors. We used quantitative real-time PCR for measuring mRNA gene-expression of vascular endothelial growth factor (VEGF), integrin alphaV, and integrin beta3, and CD34 for a group of patients with neuroendocrine tumors (n=13). Tissue from patients with colorectal cancer liver...

  12. Targeting ILK and β4 integrin abrogates the invasive potential of ovarian cancer

    International Nuclear Information System (INIS)

    Highlights: ► The potential of targeting ILK and integrins for highly aggressive ovarian cancer. ► Unanticipated synergistic effect for the combination of ILK/β4 integrin. ► Combination of ILK/β4 integrin effectively inhibited the PI3K/Akt/Rac1 cascade. ► Targeting of β4 integrin/ILK had potent inhibitory effects in ovarian cancer. -- Abstract: Integrins and integrin-linked kinase (ILK) are essential to cancerous invasion because they mediate physical interactions with the extracellular matrix, and regulate oncogenic signaling pathways. The purpose of our study is to determine whether deletion of β1 and β4 integrin and ILK, alone or in combination, has antitumoral effects in ovarian cancer. Expression of β1 and β4 integrin and ILK was analyzed by immunohistochemistry in 196 ovarian cancer tissue samples. We assessed the effects of depleting these molecules with shRNAs in ovarian cancer cells by Western blot, conventional RT-PCR, cell proliferation, migration, invasion, and in vitro Rac1 activity assays, and in vivo xenograft formation assays. Overexpression of β4 integrin and ILK in human ovarian cancer specimens was found to correlate with tumor aggressiveness. Depletion of these targets efficiently suppresses ovarian cancer cell proliferation, migration, and invasion in vitro and xenograft tumor formation in vivo. We also demonstrated that single depletion of ILK or combination depletion of β4 integrin/ILK inhibits phosphorylation of downstream signaling targets, p-Ser 473 Akt and p-Thr202/Tyr204 Erk1/2, and activation of Rac1, as well as reduce expression of MMP-2 and MMP-9 and increase expression of caspase-3 in vitro. In conclusion, targeting β4 integrin combined with ILK can instigate the latent tumorigenic potential and abrogate the invasive potential in ovarian cancer.

  13. Conservation of the Human Integrin-Type Beta-Propeller Domain in Bacteria

    Science.gov (United States)

    Chouhan, Bhanupratap; Denesyuk, Alexander; Heino, Jyrki; Johnson, Mark S.; Denessiouk, Konstantin

    2011-01-01

    Integrins are heterodimeric cell-surface receptors with key functions in cell-cell and cell-matrix adhesion. Integrin α and β subunits are present throughout the metazoans, but it is unclear whether the subunits predate the origin of multicellular organisms. Several component domains have been detected in bacteria, one of which, a specific 7-bladed β-propeller domain, is a unique feature of the integrin α subunits. Here, we describe a structure-derived motif, which incorporates key features of each blade from the X-ray structures of human αIIbβ3 and αVβ3, includes elements of the FG-GAP/Cage and Ca2+-binding motifs, and is specific only for the metazoan integrin domains. Separately, we searched for the metazoan integrin type β-propeller domains among all available sequences from bacteria and unicellular eukaryotic organisms, which must incorporate seven repeats, corresponding to the seven blades of the β-propeller domain, and so that the newly found structure-derived motif would exist in every repeat. As the result, among 47 available genomes of unicellular eukaryotes we could not find a single instance of seven repeats with the motif. Several sequences contained three repeats, a predicted transmembrane segment, and a short cytoplasmic motif associated with some integrins, but otherwise differ from the metazoan integrin α subunits. Among the available bacterial sequences, we found five examples containing seven sequential metazoan integrin-specific motifs within the seven repeats. The motifs differ in having one Ca2+-binding site per repeat, whereas metazoan integrins have three or four sites. The bacterial sequences are more conserved in terms of motif conservation and loop length, suggesting that the structure is more regular and compact than those example structures from human integrins. Although the bacterial examples are not full-length integrins, the full-length metazoan-type 7-bladed β-propeller domains are present, and sometimes two tandem

  14. Conservation of the human integrin-type beta-propeller domain in bacteria.

    Directory of Open Access Journals (Sweden)

    Bhanupratap Chouhan

    Full Text Available Integrins are heterodimeric cell-surface receptors with key functions in cell-cell and cell-matrix adhesion. Integrin α and β subunits are present throughout the metazoans, but it is unclear whether the subunits predate the origin of multicellular organisms. Several component domains have been detected in bacteria, one of which, a specific 7-bladed β-propeller domain, is a unique feature of the integrin α subunits. Here, we describe a structure-derived motif, which incorporates key features of each blade from the X-ray structures of human αIIbβ3 and αVβ3, includes elements of the FG-GAP/Cage and Ca(2+-binding motifs, and is specific only for the metazoan integrin domains. Separately, we searched for the metazoan integrin type β-propeller domains among all available sequences from bacteria and unicellular eukaryotic organisms, which must incorporate seven repeats, corresponding to the seven blades of the β-propeller domain, and so that the newly found structure-derived motif would exist in every repeat. As the result, among 47 available genomes of unicellular eukaryotes we could not find a single instance of seven repeats with the motif. Several sequences contained three repeats, a predicted transmembrane segment, and a short cytoplasmic motif associated with some integrins, but otherwise differ from the metazoan integrin α subunits. Among the available bacterial sequences, we found five examples containing seven sequential metazoan integrin-specific motifs within the seven repeats. The motifs differ in having one Ca(2+-binding site per repeat, whereas metazoan integrins have three or four sites. The bacterial sequences are more conserved in terms of motif conservation and loop length, suggesting that the structure is more regular and compact than those example structures from human integrins. Although the bacterial examples are not full-length integrins, the full-length metazoan-type 7-bladed β-propeller domains are present, and

  15. Photon-induced cell migration and integrin expression promoted by DNA integration of HPV16 genome

    Energy Technology Data Exchange (ETDEWEB)

    Rieken, Stefan; Simon, Florian; Habermehl, Daniel; Dittmar, Jan Oliver; Combs, Stephanie E.; Weber, Klaus; Debus, Juergen; Lindel, Katja [University Hospital of Heidelberg, Department of Radiation Therapy and Radiation Oncology, Heidelberg (Germany)

    2014-10-15

    Persistent human papilloma virus 16 (HPV16) infections are a major cause of cervical cancer. The integration of the viral DNA into the host genome causes E2 gene disruption which prevents apoptosis and increases host cell motility. In cervical cancer patients, survival is limited by local infiltration and systemic dissemination. Surgical control rates are poor in cases of parametrial infiltration. In these patients, radiotherapy (RT) is administered to enhance local control. However, photon irradiation itself has been reported to increase cell motility. In cases of E2-disrupted cervical cancers, this phenomenon would impose an additional risk of enhanced tumor cell motility. Here, we analyze mechanisms underlying photon-increased migration in keratinocytes with differential E2 gene status. Isogenic W12 (intact E2 gene status) and S12 (disrupted E2 gene status) keratinocytes were analyzed in fibronectin-based and serum-stimulated migration experiments following single photon doses of 0, 2, and 10 Gy. Quantitative FACS analyses of integrin expression were performed. Migration and adhesion are increased in E2 gene-disrupted keratinocytes. E2 gene disruption promotes attractability by serum components, therefore, effectuating the risk of local infiltration and systemic dissemination. In S12 cells, migration is further increased by photon RT which leads to enhanced expression of fibronectin receptor integrins. HPV16-associated E2 gene disruption is a main predictor of treatment-refractory cancer virulence. E2 gene disruption promotes cell motility. Following photon RT, E2-disrupted tumors bear the risk of integrin-related infiltration and dissemination. (orig.) [German] Persistierende Infektionen mit humanen Papillomaviren 16 (HPV16) sind ein Hauptausloeser des Zervixkarzinoms. Die Integration der viralen DNS in das Wirtszellgenom fuehrt zum Integritaetsverlust des E2-Gens, wodurch in der Wirtszelle Apoptose verhindert und Motilitaet gesteigert werden. In

  16. Human Adipose-Derived Mesenchymal Stem Cells Cryopreservation and Thawing Decrease α4-Integrin Expression

    Science.gov (United States)

    Irioda, Ana Carolina; Cassilha, Rafael; Zocche, Larissa; Francisco, Julio Cesar; Cunha, Ricardo Correa; Ferreira, Priscila Elias; Guarita-Souza, Luiz Cesar; Ferreira, Reginaldo Justino; Mogharbel, Bassam Felipe; Garikipati, Venkata Naga Srikanth; Souza, Daiany; Beltrame, Mirian Perlingeiro; de Carvalho, Katherine Athayde Teixeira

    2016-01-01

    Aim. The effects of cryopreservation on adipose tissue-derived mesenchymal stem cells are not clearly documented, as there is a growing body of evidence about the importance of adipose-derived mesenchymal stem cells for regenerative therapies. The aim of this study was to analyze human adipose tissue-derived mesenchymal stem cells phenotypic expression (CD34, CD45, CD73, CD90, CD105, and CD49d), colony forming unit ability, viability, and differentiation potential before and after cryopreservation. Materials and Methods. 12 samples of the adipose tissue were collected from a healthy donor using the liposuction technique. The cell isolation was performed by enzymatic digestion and then the cells were cultured up to passage 2. Before and after cryopreservation the immunophenotype, cellular viability analysis by flow cytometer, colony forming units ability, differentiation potential into adipocytes and osteoblasts as demonstrated by Oil Red O and Alizarin Red staining, respectively. Results. The immunophenotypic markers expression was largely preserved, and their multipotency was maintained. However, after cryopreservation, the cells decreased α4-integrin expression (CD49d), cell viability, and number of colony forming units. Conclusions. These findings suggest that ADMSC transplanted after cryopreservation might compromise the retention of transplanted cells in the host tissue. Therefore, further studies are warranted to standardize protocols related to cryopreservation to attain full benefits of stem cell therapy. PMID:26981129

  17. Human Adipose-Derived Mesenchymal Stem Cells Cryopreservation and Thawing Decrease α4-Integrin Expression

    Directory of Open Access Journals (Sweden)

    Ana Carolina Irioda

    2016-01-01

    Full Text Available Aim. The effects of cryopreservation on adipose tissue-derived mesenchymal stem cells are not clearly documented, as there is a growing body of evidence about the importance of adipose-derived mesenchymal stem cells for regenerative therapies. The aim of this study was to analyze human adipose tissue-derived mesenchymal stem cells phenotypic expression (CD34, CD45, CD73, CD90, CD105, and CD49d, colony forming unit ability, viability, and differentiation potential before and after cryopreservation. Materials and Methods. 12 samples of the adipose tissue were collected from a healthy donor using the liposuction technique. The cell isolation was performed by enzymatic digestion and then the cells were cultured up to passage 2. Before and after cryopreservation the immunophenotype, cellular viability analysis by flow cytometer, colony forming units ability, differentiation potential into adipocytes and osteoblasts as demonstrated by Oil Red O and Alizarin Red staining, respectively. Results. The immunophenotypic markers expression was largely preserved, and their multipotency was maintained. However, after cryopreservation, the cells decreased α4-integrin expression (CD49d, cell viability, and number of colony forming units. Conclusions. These findings suggest that ADMSC transplanted after cryopreservation might compromise the retention of transplanted cells in the host tissue. Therefore, further studies are warranted to standardize protocols related to cryopreservation to attain full benefits of stem cell therapy.

  18. Deletion of integrin-linked kinase from neural crest cells in mice results in aortic aneurysms and embryonic lethality

    OpenAIRE

    Arnold, Thomas D.; Keling Zang; Ainara Vallejo-Illarramendi

    2013-01-01

    SUMMARY Neural crest cells (NCCs) participate in the remodeling of the cardiac outflow tract and pharyngeal arch arteries during cardiovascular development. Integrin-linked kinase (ILK) is a serine/threonine kinase and a major regulator of integrin signaling. It links integrins to the actin cytoskeleton and recruits other adaptor molecules into a large complex to regulate actin dynamics and integrin function. Using the Cre-lox system, we deleted Ilk from NCCs of mice to investigate its rol...

  19. A role for the actin-bundling protein l-plastin in the regulation of leukocyte integrin function

    OpenAIRE

    Jones, Samuel L.; Wang, Jun; Turck, Christoph W; Brown, Eric J.

    1998-01-01

    Regulation of leukocyte integrin avidity is a crucial aspect of inflammation and immunity. The actin cytoskeleton has an important role in the regulation of integrin function, but the cytoskeletal proteins involved are largely unknown. Because inflammatory stimuli that activate integrin-mediated adhesion in human polymorphonuclear neutrophils (PMN) and monocytes cause phosphorylation of the actin-bundling protein l-plastin, we tested whether l-plastin phosphorylation was involved in integrin ...

  20. Interactions between the discoidin domain receptor 1 and β1 integrin regulate attachment to collagen

    Directory of Open Access Journals (Sweden)

    Lisa A. Staudinger

    2013-09-01

    Collagen degradation by phagocytosis is essential for physiological collagen turnover and connective tissue homeostasis. The rate limiting step of phagocytosis is the binding of specific adhesion receptors, which include the integrins and discoidin domain receptors (DDR, to fibrillar collagen. While previous data suggest that these two receptors interact, the functional nature of these interactions is not defined. In mouse and human fibroblasts we examined the effects of DDR1 knockdown and over-expression on β1 integrin subunit function. DDR1 expression levels were positively associated with enhanced contraction of floating and attached collagen gels, increased collagen binding and increased collagen remodeling. In DDR1 over-expressing cells compared with control cells, there were increased numbers, area and length of focal adhesions immunostained for talin, paxillin, vinculin and activated β1 integrin. After treatment with the integrin-cleaving protease jararhagin, in comparison to controls, DDR1 over-expressing cells exhibited increased β1 integrin cleavage at the cell membrane, indicating that DDR1 over-expression affected the access and susceptibility of cell-surface β1 integrin to the protease. DDR1 over-expression was associated with increased glycosylation of the β1 integrin subunit, which when blocked by deoxymannojirimycin, reduced collagen binding. Collectively these data indicate that DDR1 regulates β1 integrin interactions with fibrillar collagen, which positively impacts the binding step of collagen phagocytosis and collagen remodeling.

  1. Interaction of the α2A domain of integrin with small collagen fragments

    NARCIS (Netherlands)

    Siebert, H.-C.; Burg-Roderfeld, M.; Eckert, T.; Stötzel, S.; Kirch, U.; Diercks, T.; Humphries, M.J.; Frank, M.; Wechselberger, R.W.; Tajkhorshid, E.; Oesser, S.

    2010-01-01

    We here present a detailed study of the ligand-receptor interactions between single and triple-helical strands of collagen and the α2A domain of integrin (α2A), providing valuable new insights into the mechanisms and dynamics of collagen-integrin binding at a sub-molecular level. The occurrence of s

  2. Ligand-Occupied Integrin Internalization Links Nutrient Signaling to Invasive Migration

    Directory of Open Access Journals (Sweden)

    Elena Rainero

    2015-01-01

    Full Text Available Integrin trafficking is key to cell migration, but little is known about the spatiotemporal organization of integrin endocytosis. Here, we show that α5β1 integrin undergoes tensin-dependent centripetal movement from the cell periphery to populate adhesions located under the nucleus. From here, ligand-engaged α5β1 integrins are internalized under control of the Arf subfamily GTPase, Arf4, and are trafficked to nearby late endosomes/lysosomes. Suppression of centripetal movement or Arf4-dependent endocytosis disrupts flow of ligand-bound integrins to late endosomes/lysosomes and their degradation within this compartment. Arf4-dependent integrin internalization is required for proper lysosome positioning and for recruitment and activation of mTOR at this cellular subcompartment. Furthermore, nutrient depletion promotes subnuclear accumulation and endocytosis of ligand-engaged α5β1 integrins via inhibition of mTORC1. This two-way regulatory interaction between mTORC1 and integrin trafficking in combination with data describing a role for tensin in invasive cell migration indicate interesting links between nutrient signaling and metastasis.

  3. Targeting of Alpha-V Integrins Reduces Malignancy of Bladder Carcinoma

    Science.gov (United States)

    van der Horst, Geertje; Bos, Lieke; van der Mark, Maaike; Cheung, Henry; Heckmann, Bertrand; Clément-Lacroix, Philippe; Lorenzon, Giocondo; Pelger, Rob C. M.; Bevers, Rob F. M.; van der Pluijm, Gabri

    2014-01-01

    Low survival rates of metastatic cancers emphasize the need for a drug that can prevent and/or treat metastatic cancer. αv integrins are involved in essential processes for tumor growth and metastasis and targeting of αv integrins has been shown to decrease angiogenesis, tumor growth and metastasis. In this study, the role of αv integrin and its potential as a drug target in bladder cancer was investigated. Treatment with an αv integrin antagonist as well as knockdown of αv integrin in the bladder carcinoma cell lines, resulted in reduced malignancy invitro, as illustrated by decreased proliferative, migratory and clonogenic capacity. The CDH1/CDH2 ratio increased, indicating a shift towards a more epithelial phenotype. This shift appeared to be associated with downregulation of EMT-inducing transcription factors including SNAI2. The expression levels of the self-renewal genes NANOG and BMI1 decreased as well as the number of cells with high Aldehyde Dehydrogenase activity. In addition, self-renewal ability decreased as measured with the urosphere assay. In line with these observations, knockdown or treatment of αv integrins resulted in decreased metastatic growth in preclinical invivo models as assessed by bioluminescence imaging. In conclusion, we show that αv integrins are involved in migration, EMT and maintenance of Aldehyde Dehydrogenase activity in bladder cancer cells. Targeting of αv integrins might be a promising approach for treatment and/or prevention of metastatic bladder cancer. PMID:25247809

  4. Role of aVb3 integrin in embryo implantation in the mouse

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Integrin, a heterodimeric adhesive molecule composed of a and b subunits, can regulate cell adhesion and trafficking. Recent data have documented that, at the "implantation window" stage, aVb3 integrin participates in the maternal-fetal interaction and becomes a potential marker of uterine receptivity. Furthermore, it can affect invasiveness of embryo. This work made a further study about its action mechanism. Results of indirect immunofluorescence and laser scanning confocal microscopy showed that aVb3 integrin was clearly expressed in the mouse blastocyst. Injection of aVb3 integrin antiserum into a uterine horn of a pregnant mouse on day 3 markedly decreased the number of embryos implanted (P < 0.001). In a co-culture model, aVb3 integrin antisera at 1:100 and 1:200 dilutions significantly depressed the attachment and outgrowth reactions of blastocysts on monolayer of uterine epithelial cells. Analysis of correlation manifested that the inhibitory effect of aVb3 integrin antiserum was dosage/dilution-dependent. Thus, aVb3 integrin is an essential factor in the uterine endometrium for embryo implantation in the mouse. This integrin distinctly expressed in the mouse blastocyst at "implantation" stage affected the process of embryo implantation by route of mediating both the attachment and the outgrowth processes of blastocyst on uterine epithelial cells.

  5. Integrin β3 is required in infection and proliferation of classical swine fever virus.

    Directory of Open Access Journals (Sweden)

    Weiwei Li

    Full Text Available Classical Swine Fever (CSF is a highly infectious fatal pig disease, resulting in huge economic loss to the swine industry. Integrins are membrane-bound signal mediators, expressed on a variety of cell surfaces and are known as receptors or co-receptors for many viruses. However, the role of integrin β3 in CSFV infection is unknown. Here, through quantitive PCR, immunofluorescence (IFC and immunocytohistochemistry (ICC, we revealed that ST (swine testicles epithelial cells have a prominent advantage in CSFV proliferation as compared to EC (swine umbilical vein endothelial cell, IEC (swine intestinal epithelial cell and PK (porcine kidney epithelial cells. Meanwhile, ST cells had remarkably more integrin β3 expression as compared to EC, IEC and PK cells, which was positively correlated with CSFV infection and proliferation. Integrin β3 was up-regulated post CSFV infection in all the four cell lines, while the CSFV proliferation rate was decreased in integrin β3 function-blocked cells. ShRNA1755 dramatically decreased integrin β3, with a deficiency of 96% at the mRNA level and 80% at the protein level. CSFV proliferation was dramatically reduced in integrin β3 constantly-defected cells (ICDC, with the deficiencies of 92.6%, 99% and 81.7% at 24 h, 48 h and 72 h post CSFV infection, respectively. These results demonstrate that integrin β3 is required in CSFV infection and proliferation, which provide a new insight into the mechanism of CSFV infection.

  6. Integrin αv in the mechanical response of osteoblast lineage cells

    Energy Technology Data Exchange (ETDEWEB)

    Kaneko, Keiko [Department of Bone and Joint Disease, National Center for Geriatrics and Gerontology, Obu, Aichi 474-8511 (Japan); Ito, Masako [Medical Work-Life-Balance Center, Nagasaki University Hospital, Nagasaki 852-8501 (Japan); Naoe, Yoshinori [Department of Mechanism of Aging, National Center for Geriatrics and Gerontology, Obu, Aichi 474-8511 (Japan); Lacy-Hulbert, Adam [Department of Pediatrics, Massachusetts General Hospital, Boston, MA 02114 (United States); Ikeda, Kyoji, E-mail: kikeda@ncgg.go.jp [Department of Bone and Joint Disease, National Center for Geriatrics and Gerontology, Obu, Aichi 474-8511 (Japan)

    2014-05-02

    Highlights: • Deletion of integrin αv in osteoblast lineage results in an impaired SOST response to loading in vivo. • c-Src–p130Cas–JNK–YAP/TAZ is activated via integrin αv on osteoblasts in response to FSS. • Deletion of integrin αv in osteoblasts results in impaired responses to mechanical stimulation. • Integrin αv is a key component of the mechanosensing machinery in bone. - Abstract: Although osteoblast lineage cells, especially osteocytes, are thought to be a primary mechanosensory cell in bone, the identity of the mechano-receptor and downstream mechano-signaling pathways remain largely unknown. Here we show using osteoblastic cell model of mechanical stimulation with fluid shear stress that in the absence of integrin αv, phosphorylation of the Src substrate p130Cas and JNK was impaired, culminating in an inhibition of nuclear translocation of YAP/TAZ and subsequent transcriptional activation of target genes. Targeted deletion of the integrin αv in osteoblast lineage cells results in an attenuated response to mechanical loading in terms of Sost gene expression, indicative of a role for integrin αv in mechanoreception in vivo. Thus, integrin αv may be integral to a mechanosensing machinery in osteoblastic cells and involved in activation of a Src–JNK–YAP/TAZ pathway in response to mechanical stimulation.

  7. Integrin β1 Participates in Atrial Remodeling in Rapid Atrial Pacing Induced Canine Atrial Fibrillation Model

    Institute of Scientific and Technical Information of China (English)

    Zhang wei; Yang guirong; Zheng zhaotong; Wang sujia; Zhang yun

    2004-01-01

    @@ Objective Integrin β1 regulates cell to cell and cell to extracellualr matrix interaction in heart. however, its pathop hysiological role in atrial fibrillation is unclear. The purpose of t his study was to determine whether atrial structural remodeling during atrial fibrillation is associated with altered integrinβ1.

  8. Transforming growth factor-beta1 promotes the migration and invasion of sphere-forming stem-like cell subpopulations in esophageal cancer

    International Nuclear Information System (INIS)

    Esophageal cancer is one of the most lethal solid malignancies. Mounting evidence demonstrates that cancer stem cells (CSCs) are able to cause tumor initiation, metastasis and responsible for chemotherapy and radiotherapy failures. As CSCs are thought to be the main reason of therapeutic failure, these cells must be effectively targeted to elicit long-lasting therapeutic responses. We aimed to enrich and identify the esophageal cancer cell subpopulation with stem-like properties and help to develop new target therapy strategies for CSCs. Here, we found esophageal cancer cells KYSE70 and TE1 could form spheres in ultra low attachment surface culture and be serially passaged. Sphere-forming cells could redifferentiate and acquire morphology comparable to parental cells, when return to adherent culture. The sphere-forming cells possessed the key criteria that define CSCs: persistent self-renewal, overexpression of stemness genes (SOX2, ALDH1A1 and KLF4), reduced expression of differentiation marker CK4, chemoresistance, strong invasion and enhanced tumorigenic potential. SB525334, transforming growth factor-beta 1(TGF-β1) inhibitor, significantly inhibited migration and invasion of sphere-forming stem-like cells and had no effect on sphere-forming ability. In conclusion, esophageal cancer sphere-forming cells from KYSE70 and TE1 cultured in ultra low attachment surface possess cancer stem cell properties, providing a model for CSCs targeted therapy. TGF-β1 promotes the migration and invasion of sphere-forming stem-like cells, which may guide future studies on therapeutic strategies targeting these cells. - Highlights: • Esophageal cancer sphere-forming cells possess cancer stem cell properties. • Sphere-forming cells enhance TGF-β1 pathway activity. • TGF-β 1 inhibitor suppresses the migration and invasion of sphere-forming cells

  9. Transforming growth factor beta-1 and interleukin-17 gene transcription in peripheral blood mononuclear cells and the human response to infection.

    LENUS (Irish Health Repository)

    White, Mary

    2012-02-01

    INTRODUCTION: The occurrence of severe sepsis may be associated with deficient pro-inflammatory cytokine production. Transforming growth factor beta-1 (TGFbeta-1) predominantly inhibits inflammation and may simultaneously promote IL-17 production. Interleukin-17 (IL-17) is a recently described pro-inflammatory cytokine, which may be important in auto-immunity and infection. We investigated the hypothesis that the onset of sepsis is related to differential TGFbeta-1 and IL-17 gene expression. METHODS: A prospective observational study in a mixed intensive care unit (ICU) and hospital wards in a university hospital. Patients (59) with severe sepsis; 15 patients with gram-negative bacteraemia but without critical illness and 10 healthy controls were assayed for TGFbeta-1, IL-17a, IL-17f, IL-6 and IL-1beta mRNA in peripheral blood mononuclear cells (PBMC) by quantitative real-time PCR and serum protein levels by ELISA. RESULTS: TGFbeta-1 mRNA levels are reduced in patients with bacteraemia and sepsis compared with controls (p=0.02). IL-6 mRNA levels were reduced in bacteraemic patients compared with septic patients and controls (p=0.008). IL-1beta mRNA levels were similar in all groups, IL-17a and IL-17f mRNA levels are not detectable in peripheral blood mononuclear cells. IL-6 protein levels were greater in patients with sepsis than bacteraemic and control patients (p<0.0001). Activated TGFbeta-1 and IL-17 protein levels were similar in all groups. IL-1beta protein was not detectable in the majority of patients. CONCLUSIONS: Down regulation of TGFbeta-1 gene transcription was related to the occurrence of infection but not the onset of sepsis. Interleukin-17 production in PBMC may not be significant in the human host response to infection.

  10. Transforming growth factor-beta1 promotes the migration and invasion of sphere-forming stem-like cell subpopulations in esophageal cancer

    Energy Technology Data Exchange (ETDEWEB)

    Yue, Dongli; Zhang, Zhen; Li, Jieyao; Chen, Xinfeng [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); Department of Oncology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052 (China); Ping, Yu; Liu, Shasha [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); School of Life Sciences, Zhengzhou University, Zhengzhou 450000 (China); Shi, Xiaojuan; Li, Lifeng [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); Department of Oncology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052 (China); Wang, Liping [Department of Oncology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052 (China); Huang, Lan [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); Zhang, Bin [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); Robert H. Lurie Comprehensive Cancer Center, Department of Medicine-Division of Hematology/Oncology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611 (United States); Sun, Yan [Department of Oncology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052 (China); Department of Medical Oncology, Cancer Hospital, Chinese Academy of Medical Sciences (China); and others

    2015-08-01

    Esophageal cancer is one of the most lethal solid malignancies. Mounting evidence demonstrates that cancer stem cells (CSCs) are able to cause tumor initiation, metastasis and responsible for chemotherapy and radiotherapy failures. As CSCs are thought to be the main reason of therapeutic failure, these cells must be effectively targeted to elicit long-lasting therapeutic responses. We aimed to enrich and identify the esophageal cancer cell subpopulation with stem-like properties and help to develop new target therapy strategies for CSCs. Here, we found esophageal cancer cells KYSE70 and TE1 could form spheres in ultra low attachment surface culture and be serially passaged. Sphere-forming cells could redifferentiate and acquire morphology comparable to parental cells, when return to adherent culture. The sphere-forming cells possessed the key criteria that define CSCs: persistent self-renewal, overexpression of stemness genes (SOX2, ALDH1A1 and KLF4), reduced expression of differentiation marker CK4, chemoresistance, strong invasion and enhanced tumorigenic potential. SB525334, transforming growth factor-beta 1(TGF-β1) inhibitor, significantly inhibited migration and invasion of sphere-forming stem-like cells and had no effect on sphere-forming ability. In conclusion, esophageal cancer sphere-forming cells from KYSE70 and TE1 cultured in ultra low attachment surface possess cancer stem cell properties, providing a model for CSCs targeted therapy. TGF-β1 promotes the migration and invasion of sphere-forming stem-like cells, which may guide future studies on therapeutic strategies targeting these cells. - Highlights: • Esophageal cancer sphere-forming cells possess cancer stem cell properties. • Sphere-forming cells enhance TGF-β1 pathway activity. • TGF-β 1 inhibitor suppresses the migration and invasion of sphere-forming cells.

  11. Effects of polymorphisms in beta1-adrenoceptor and alpha-subunit of G protein on heart rate and blood pressure during exercise test. The Finnish Cardiovascular Study.

    Science.gov (United States)

    Nieminen, Tuomo; Lehtimäki, Terho; Laiho, Jarno; Rontu, Riikka; Niemelä, Kari; Kööbi, Tiit; Lehtinen, Rami; Viik, Jari; Turjanmaa, Väinö; Kähönen, Mika

    2006-02-01

    We tested whether the Arg389Gly and Ser49Gly polymorphisms of the beta1-adrenergic receptor gene ADRB1 and the T393C polymorphism of the G protein alpha-subunit gene GNAS1 modulate heart rate (HR) and blood pressure responses during an exercise stress test. The study population comprised 890 participants (563 men and 327 women, mean age 58.1 +/- 12.6 yr) of the Finnish Cardiovascular Study. Their HR, systolic (SAP), and diastolic arterial pressures (DAP) at rest, during exercise, and 4 min after the test were measured and analyzed by repeated-measurement ANOVA (RANOVA). Genotypes were detected by TaqMan 5' nuclease assay. In all subjects, and in men and women separately, the T393C of GNAS1 was the only polymorphism with genotype x time interaction in HR over the three study phases (P = 0.04, RANOVA). None of the polymorphisms presented genotype x time interaction in SAP or DAP responses (P > 0.10, RANOVA). In all subjects at rest, the Ser49Gly polymorphism of ADRB1 tended (P = 0.06, ANOVA) to differentiate HR. Arg389Gly polymorphism of ADRB1 affected maximal SAP during exercise (P = 0.04, ANOVA) and the change in SAP from rest to maximal (P = 0.03, ANOVA). Arg389 homozygotes, particularly men, were less likely to have ventricular extrasystoles during the exercise (odds ratio = 0.68, 95% confidence interval = 0.51-0.91, P = 0.009, and odds ratio = 0.60, 95% confidence interval = 0.42-0.86, P = 0.006, respectively) than did Gly389 carriers. In conclusion, polymorphisms examined appear to have modulatory effects on hemodynamics in a clinical exercise test setting. However, the effects in absolute numbers were minor and clinically possibly insignificant. PMID:16210433

  12. Vesicle-associated membrane protein 2 mediates trafficking of α5β1 integrin to the plasma membrane

    International Nuclear Information System (INIS)

    Integrins are major receptors for cell adhesion to the extracellular matrix (ECM). As transmembrane proteins, the levels of integrins at the plasma membrane or the cell surface are ultimately determined by the balance between two vesicle trafficking events: endocytosis of integrins at the plasma membrane and exocytosis of the vesicles that transport integrins. Here, we report that vesicle-associated membrane protein 2 (VAMP2), a SNARE protein that mediates vesicle fusion with the plasma membrane, is involved in the trafficking of α5β1 integrin. VAMP2 was present on vesicles containing endocytosed β1 integrin. Small interfering RNA (siRNA) silencing of VAMP2 markedly reduced cell surface α5β1 and inhibited cell adhesion and chemotactic migration to fibronectin, the ECM ligand of α5β1, without altering cell surface expression of α2β1 integrin or α3β1 integrin. By contrast, silencing of VAMP8, another SNARE protein, had no effect on cell surface expression of the integrins or cell adhesion to fibronectin. In addition, VAMP2-mediated trafficking is involved in cell adhesion to collagen but not to laminin. Consistent with disruption of integrin functions in cell proliferation and survival, VAMP2 silencing diminished proliferation and triggered apoptosis. Collectively, these data indicate that VAMP2 mediates the trafficking of α5β1 integrin to the plasma membrane and VAMP2-dependent integrin trafficking is critical in cell adhesion, migration and survival.

  13. Alpha V integrin prolongs collagenase production through Jun activation binding protein 1.

    Science.gov (United States)

    Levinson, Howard; Sil, Alok K; Conwell, John E; Hopper, James E; Ehrlich, H Paul

    2004-08-01

    Robust expression of alphav integrin and matrix metalloproteinase 1 (MMP1) plays an important role in cancer metastasis and wound healing. A patient with an abnormal scar that appeared stretched and thinned out was found to have fibroblasts that overexpressed alphav integrin; therefore, a relationship between alphav integrin expression and MMP1 production was sought. A yeast 2 hybrid screen revealed alphav integrin interacts with jun activation binding domain-1 (JAB1). Mesenchymal-derived cells were transfected with the alphav integrin gene and incorporated into collagen lattices. Transfected cells maximally contracted collagen lattices beginning on day 5, whereas control transfected cells did not contract lattices. Late-phase collagen lattice contraction was inhibited by a pan-MMP inhibitor, BB4. Overexpression of alphav correlated with enhanced MMP1 transcription, as determined by a luciferase assay (P production and that this signaling pathway in fibroblasts may lead to abnormal scarring. PMID:15269586

  14. Identification of integrin-like in guard cells of Vicia faba

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    We addressed the existence and localization of integrin-like in guard cells of Vicia faba by using a probe of polyclonal antibody against the human integrin (avb3/b5). Western blot results showed that three integrins-like of about 47.3, 43.7 and 41.1 ku were detected from the preparation of membrane fragments of purified guard cell protoplasts. Further research with immunofluorescent scanning micro-scopy indicated that those integrins-like were localized on plasma membrane of guard cells, most nearing the dorsal wall, which is consistent with the reception of signals from epidermal cells to guard cells. Thus our results indicate, for the first time, that integrins-like are present at guard cell plasma membrane of Vicia faba.

  15. Laminin and integrin expression in the ventral ectodermal ridge of the mouse embryo: implications for regulation of BMP signalling

    Science.gov (United States)

    Lopez-Escobar, Beatriz; de Felipe, Beatriz; Sanchez-Alcazar, Jose Antonio; Sasaki, Takako; Copp, Andrew J.; Ybot-Gonzalez, Patricia

    2013-01-01

    Background The ventral ectodermal ridge (VER) is an important signalling centre in the mouse tail-bud following completion of gastrulation. BMP regulation is essential for VER function, but how these signals are transmitted between adjacent tissues is unclear. Results We investigated the idea that extracellular matrix components might be involved, using immunohistochemistry and in situ hybridisation to detect all known α, β and γ laminin chains and their mRNAs in the early tail bud. We identified an apparently novel laminin variant, comprising α5, β3 and γ2 chains, as a major component of the VER basement membrane at E9.5. Strikingly, only the mRNAs for these chains were co-expressed in VER cells, suggesting that lamin532 may be the sole basement membrane laminin at this stage. Since α6 integrin was also expressed in VER cells, this raises the possibility of cell-matrix interactions regulating BMP signalling at this site of caudal morphogenesis. Conclusions Laminin532 could interact with α6-containing integrin to direct differentiation of the specialised VER cells from surface ectoderm. PMID:22911573

  16. Purification and characterization of pathogenesis-related antifungal beta 1,3 glucanase from basrai banana fruit

    International Nuclear Information System (INIS)

    Pathogenesis-related proteins have been described as proteins that are encoded by the plant genome and that are induced specifically in response to infections by pathogens. These represent a collection of unrelated protein families which function as part of the plant defense system. Pathogenesis-related antifungal protein has been isolated from the pulp of ripe Basrai bananas and purified through ammonium sulphate precipitation, Sephadex G- 75 gel filtration chromatography and electro-elution. The purified protein with acidic character (pI 6.81). has molecular weight of 34.5kDa, as determined by MALOI- TOF mass spectrometry. Mascot score obtained was 473 greater than 82, indicate extensive homology at a significant level (p.0.05) and the protein was identified as beta 1,3-glucanase with antifungal activity. It inhibited the growth of Fusarium oxysporum demonstrating the potential role of Basrai banana antifungal protein to control fungal diseases in plants, animals and human. (author)

  17. Variation in the levels of pregnancy-specific beta-1-glycoprotein in maternal serum from chromosomally abnormal pregnancies.

    Science.gov (United States)

    Graham, G W; Crossley, J A; Aitken, D A; Connor, J M

    1992-06-01

    Human pregnancy-specific beta-1-glycoprotein (SP1) was assayed retrospectively in stored maternal serum (MS) samples from 82 chromosomally abnormal pregnancies and 377 matched controls. The median MSSP1 concentration in 48 Down's syndrome pregnancies was significantly elevated at 1.17 multiples of the control median (MOM), and significantly reduced (0.5 MOM) in a group of eight cases of unbalanced translocations. There was no significant difference in median SP1 concentrations in cases of trisomy 18, trisomy 13, balanced translocations, or sex chromosome abnormalities. A comparison with human chorionic gonadotrophin results in the same series of samples indicates that SP1 is a less sensitive predictor of Down's syndrome pregnancies. PMID:1387478

  18. Co-translational formation and pharmacological characterization of beta1-adrenergic receptor/nanodisc complexes with different lipid environments.

    Science.gov (United States)

    Rues, Ralf-Bernhardt; Dötsch, Volker; Bernhard, Frank

    2016-06-01

    G protein-coupled receptors are of key significance for biomedical research. Streamlined approaches for their efficient recombinant production are of pivotal interest in order to explore their intrinsic conformational dynamics and complex ligand binding behavior. We have systematically optimized the co-translational association and folding of G protein-coupled receptors with defined membranes of nanodiscs by cell-free expression approaches. Each optimization step was quantified and the ligand binding active fraction of the receptor samples could drastically be improved. The strategy was exemplified with a stabilized and a non-stabilized derivative of the turkey beta1-adrenergic receptor. Systematic lipid screens with preformed nanodiscs revealed that generation of ligand binding active conformations of the analyzed beta1-adrenergic receptors strongly depends on lipid charge, flexibility and chain length. The lipid composition of the nanodisc membranes modulates the affinities to a variety of ligands of both receptor derivatives. In addition, the thermostabilization procedure had a significant impact on specific ligand affinities of the receptor and abolished or reduced the binding of certain antagonists. Both receptors were highly stable after purification with optimized nanodisc membranes. The procedure avoids any detergent contact of the receptors and sample production takes less than two days. Moreover, even non-stabilized receptors can be analyzed and their prior purification is not necessary for the formation of nanodisc complexes. The established process appears therefore to be suitable as a new platform for the functional or even structural characterization of recombinant G protein-coupled receptors associated with defined lipid environments. PMID:26922884

  19. Wisp2/CCN5 up-regulated in the central nervous system of GM3-only mice facilitates neurite formation in Neuro2a cells via integrin-Akt signaling

    Energy Technology Data Exchange (ETDEWEB)

    Ohkawa, Yuki, E-mail: tomilbio@med.nagoya-u.ac.jp [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Ohmi, Yuhsuke, E-mail: ooumi82@med.nagoya-u.ac.jp [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Tajima, Orie, E-mail: oriet@isc.chubu.ac.jp [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Department of Biomedical Sciences, Chubu University College of Life and Health Sciences, 1200 Matsumoto-cho, Kasugai 487-8501 (Japan); Yamauchi, Yoshio, E-mail: yyoshio@med.nagoya-u.ac.jp [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Furukawa, Keiko, E-mail: keikofu@isc.chubu.ac.jp [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Department of Biomedical Sciences, Chubu University College of Life and Health Sciences, 1200 Matsumoto-cho, Kasugai 487-8501 (Japan); Furukawa, Koichi, E-mail: koichi@med.nagoya-u.ac.jp [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan)

    2011-08-05

    Highlights: {yields} Wisp2/CCN5 was up-regulated in nervous tissues of GM3-only mutant mice. {yields} Wisp2/CCN5 was found in neurons more strongly in the mutant mice. {yields} Wisp2/CCN5 induces Akt phosphorylation via integrins and facilitates neurite formation. {yields} Wisp2/CCN5 conferred resistance to H{sub 2}O{sub 2}-induced apoptosis. {yields} Up-regulation of Wisp2/CCN5 in GM3-only mice seemed for protection of brains from neurodegeneration. -- Abstract: Wisp2/CCN5 belongs to CCN family proteins which are involved in cell proliferation, angiogenesis, tumorigenesis and wound healing. Although a number of studies on the roles of Wisp2/CCN5 in cancers have been reported, no study on the expression and function of Wisp2/CCN5 in the central nervous system has been reported. In this study, we focused on Wisp2/CCN5 that was up-regulated in nervous tissues in GM3-only mice. Over-expression of Wisp2/CCN5 enhanced neurite outgrowth potently after serum withdrawal with increased phosphorylation levels of Akt and ERKs. When cells were cultured with recombinant Wisp2/CCN5 proteins, more and longer neurites were formed than in the controls. Thus, we demonstrated for the first time that Wisp2/CCN5 facilitates neurite formation in a mouse neuroblastoma cell line, Neuro2a. Akt phosphorylation induced by recombinant Wisp2/CCN5 was suppressed after knockdown of integrin {beta}1. Moreover, Wisp2/CCN5-over-expressing cells were resistant to apoptosis induced by H{sub 2}O{sub 2}. These results suggested that secreted Wisp2/CCN5 induces Akt and ERK phosphorylation via integrins, and consequently facilitates neurite formation and conferred resistance to apoptosis. Up-regulation of Wisp2/CCN5 in GM3-only mice should be, therefore, a reaction to protect nervous tissues from neurodegeneration caused by ganglioside deficiency.

  20. The Mycoparasite Ampelomyces quisqualis Expresses exgA Encoding an exo-beta-1,3-Glucanase in Culture and During Mycoparasitism.

    Science.gov (United States)

    Rotem, Y; Yarden, O; Sztejnberg, A

    1999-08-01

    ABSTRACT Ampelomyces quisqualis, a mycoparasite of fungi causing powdery mildews, exhibited high levels of extracellular exo-beta-1,3-glucanase activity in culture compared with Neurospora crassa and Gliocladium roseum. A. quisqualis culture filtrates affected powdery mildew caused by Sphaerotheca fusca in a manner indicative of cell wall degradation, as determined by microscopic examination. A gene encoding an exo-beta-1,3-glucanase in A. quisqualis, designated exgA, was isolated and sequenced. The predicted polypeptide deduced from exgA had 46, 42, and 30% identity with amino acid sequences of Trichoderma harzianum exo-beta-1,3-glucanase and Cochliobolus carbonum EXG1 (both encoding exo-beta-1,3-glucanase) and T. harzianum bng13.1 (encoding an endo-beta-1,3-glucanase), respectively. The exgA gene had a predicted molecular mass of 84 kDa and a pI of 4.79. The gene was expressed during the late stages of growth in culture, and transcription was induced by fungal cell wall components. Transcript levels for exgA were present during late stages of hyperpar-asitism and were abundant along A. quisqualis mycelium and were slightly less abundant in A. quisqualis pycnidia. PMID:18944674

  1. Factor de crecimiento transformante beta-1: estructura, función y mecanismos de regulación en cáncer

    Directory of Open Access Journals (Sweden)

    Peralta-Zaragoza Oscar

    2001-01-01

    Full Text Available El factor de crecimiento transformante beta-1 (TGF-beta1 es sintetizado por muchas estirpes celulares como linfocitos, macrófagos y células dendríticas, y su expresión regula de manera autócrina o parácrina la diferenciación, proliferación y el estado de activación de éstas y muchas otras células. En general, el TGF-beta1 tiene propiedades pleiotrópicas en el contexto de la respuesta inmune durante el desarrollo de infecciones y procesos neoplásicos; sin embargo, los mecanismos de acción y regulación de la expresión de esta citocina aún no se comprenden del todo. En la presente revisión se describen las propiedades biológicas y los procesos moleculares que regulan la expresión del TGF-beta1, para entender los efectos de esta citocina durante la proliferación y la diferenciación celular. El conocimiento de los mecanismos moleculares de la regulación del TGF-beta1 puede representar una importante estrategia de tratamiento del cáncer. El texto completo en inglés de este artículo está disponible en: http://www.insp.mx/salud/index.html

  2. IL-1 beta and TGF-beta 1 modulate the sulphation grade of chondro-disaccharides in porcine articular cartilage: a capillary electrophoresis study.

    Science.gov (United States)

    Zanni, M; Tamburro, A; Rotilio, D

    1995-07-01

    This report describes the effect of interleukin-1 beta (IL-1 beta) and transforming growth factor-beta 1 (TGF-beta 1) on proteoglycan release from cartilage explants and modification at the sulphation level. Matrix proteoglycans purified by ion-exchange chromatography were composed of two distinct peaks (1 and 2) each showing a different Kav value when they were subjected to size-exclusion chromatography on a Sepharose CL-2B column. Glycosaminoglycans (GAGs) of conditioned medium and extracellular matrix proteoglycans were digested by chondroitin ABC and AC lyase, suggesting that chondroitin sulphate (CS) is the major GAG present (80-90%). Structural analysis of disaccharides, by capillary zone electrophoresis, revealed a different pattern of sulphated glycosaminoglycans when cartilage was treated with either IL-1 beta or TGF-beta 1. Analysis of GAGs released into the medium from TGF-beta 1 treated cartilage showed a reduction in the level of 4-S-disaccharide (delta Di4S) and an increase in non-sulphated disaccharides (delta Di0S), while no significant changes were found in IL-1 beta treated cartilage. In the extracellular matrix, IL-1 beta and TGF-beta 1 induced a more complex rearrangement of the GAGs. The level of non-sulphated disaccharides was increased whereas that of total sulphated disaccharides was reduced. Taken together, these results suggest that both cytokines modify the structure of GAGs, probably by interfering with the activity or the synthesis of sulphotransferases involved in GAG turnover. PMID:7551687

  3. Toxin a from Clostridium difficile binds to rabbit erythrocyte glycolipids with therminal Gal. cap alpha. 1-3Gal. beta. 1-4GlcNaC sequences

    Energy Technology Data Exchange (ETDEWEB)

    Clark, G.F.; Krivan, H.; Wilkins, T.; Smith, D.F.

    1987-05-01

    Toxin A is one of two clostridial toxins implicated as the causative agent of pseudomembranous colitis in patients undergoing postoperative antibiotic therapy. Evidence that the carbohydrate binding determinant for this toxin is a glycoconjugate(s) with non-reducing Gal..cap alpha..1-3Gal..beta..1-4GlcNAc has recently been reported. Specific agglutination of rabbit erythrocytes by Toxin A is inhibited by bovine thyroglobulin and prevented by pretreatment of cells with ..cap alpha..-galactosidase. Total lipid extracts from rabbit erythrocytes were subjected to thin layer chromatography and the chromatogram overlaid with purified /sup 125/I-labeled Toxin A. Two major and several minor toxin-binding glycolipids were detected following autoradiography. The major toxin-binding glycolipids were identified as pentasaccharide- and decasaccharide-ceramides expressing terminal Gal..cap alpha..1-3Gal..beta..1-4GlcNAc sequences. Treatment of the toxin-binding glycolipids with ..cap alpha..-galactosidase abolished binding. Forsmann glycolipid, globoside, Gal..cap alpha..1-4 Gal..beta..1-4Glc-cer, and Gal..cap alpha..1-3Gal..beta..1-4Glc-cer did not bind the toxin. These observations are consistent with the proposed carbohydrate specificity of the toxin for the non-reducing terminal sequence, Gal..cap alpha..1-3Gal..beta..1-4GlcNAc.

  4. Mechanosensitive components of integrin adhesions: Role of vinculin.

    Science.gov (United States)

    Atherton, Paul; Stutchbury, Ben; Jethwa, Devina; Ballestrem, Christoph

    2016-04-10

    External forces play a key role in shaping development and normal physiology. Aberrant responses to forces, or changes in the nature of such forces, are implicated in a variety of diseases. Cells contain several types of adhesions, linking them to their external environment. It is through these adhesions that forces are both sensed (from the outside inwards) and applied (from inside to out). Furthermore, several adhesion-based proteins are sensitive to changes in intracellular forces, utilising them for activation and regulation. Here, we outline how vinculin, a key component of integrin-mediated adhesions linking the actin cytoskeleton to the extracellular matrix (ECM), is regulated by force and acts as force transducing protein. We discuss the role of vinculin in vivo and its place in health and disease; summarise the proposed mechanisms by which vinculin is recruited to and activated at integrin-ECM adhesions; and discuss recent findings that place vinculin as the major force sensing and transmitting component of cell-matrix adhesion complexes. Finally, we discuss the role of vinculin in regulating the cellular responses to both the physical properties of the external environment and to externally applied physical stimuli. PMID:26607713

  5. Integrin-linked kinase regulates interphase and mitotic microtubule dynamics.

    Directory of Open Access Journals (Sweden)

    Simin Lim

    Full Text Available Integrin-linked kinase (ILK localizes to both focal adhesions and centrosomes in distinct multiprotein complexes. Its dual function as a kinase and scaffolding protein has been well characterized at focal adhesions, where it regulates integrin-mediated cell adhesion, spreading, migration and signaling. At the centrosomes, ILK regulates mitotic spindle organization and centrosome clustering. Our previous study showed various spindle defects after ILK knockdown or inhibition that suggested alteration in microtubule dynamics. Since ILK expression is frequently elevated in many cancer types, we investigated the effects of ILK overexpression on microtubule dynamics. We show here that overexpressing ILK in HeLa cells was associated with a shorter duration of mitosis and decreased sensitivity to paclitaxel, a chemotherapeutic agent that suppresses microtubule dynamics. Measurement of interphase microtubule dynamics revealed that ILK overexpression favored microtubule depolymerization, suggesting that microtubule destabilization could be the mechanism behind the decreased sensitivity to paclitaxel, which is known to stabilize microtubules. Conversely, the use of a small molecule inhibitor selective against ILK, QLT-0267, resulted in suppressed microtubule dynamics, demonstrating a new mechanism of action for this compound. We further show that treatment of HeLa cells with QLT-0267 resulted in higher inter-centromere tension in aligned chromosomes during mitosis, slower microtubule regrowth after cold depolymerization and the presence of a more stable population of spindle microtubules. These results demonstrate that ILK regulates microtubule dynamics in both interphase and mitotic cells.

  6. Astroglial Integrins in the Development and Regulation of Neurovascular Units

    Directory of Open Access Journals (Sweden)

    Hironobu Tanigami

    2012-01-01

    Full Text Available In the neurovascular units of the central nervous system, astrocytes form extensive networks that physically and functionally connect the neuronal synapses and the cerebral vascular vessels. This astrocytic network is thought to be critically important for coupling neuronal signaling activity and energy demand with cerebral vascular tone and blood flow. To establish and maintain this elaborate network, astrocytes must precisely calibrate their perisynaptic and perivascular processes in order to sense and regulate neuronal and vascular activities, respectively. Integrins, a prominent family of cell-adhesion molecules that support astrocytic migration in the brain during developmental and normal adult stages, have been implicated in regulating the integrity of the blood brain barrier and the tripartite synapse to facilitate the formation of a functionally integrated neurovascular unit. This paper describes the significant roles that integrins and connexins play not only in regulating astrocyte migration during the developmental and adult stages of the neurovascular unit, but also in general health and in such diseases as hepatic encephalopathy.

  7. Integrin activation and focal complex formation in cardiac hypertrophy

    Science.gov (United States)

    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.

  8. Genetic analysis of the Drosophila alphaPS2 integrin subunit reveals discrete adhesive, morphogenetic and sarcomeric functions.

    OpenAIRE

    Bloor, J. W.; Brown, N H

    1998-01-01

    The integrin family of cell surface receptors mediates cell-substrate and cell-to-cell adhesion and transmits intracellular signals. In Drosophila there is good evidence for an adhesive role of integrins, but evidence for integrin signalling has remained elusive. Each integrin is an alphabeta heterodimer, and the Drosophila betaPS subunit forms at least two integrins by association with different alpha subunits: alphaPS1betaPS (PS1) and alphaPS2betaPS (PS2). The complex pattern of PS2 integri...

  9. Factor de crecimiento transformante beta-1: estructura, función y mecanismos de regulación en cáncer

    OpenAIRE

    Peralta-Zaragoza Oscar; Lagunas-Martínez A.; Madrid-Marina Vicente

    2001-01-01

    El factor de crecimiento transformante beta-1 (TGF-beta1) es sintetizado por muchas estirpes celulares como linfocitos, macrófagos y células dendríticas, y su expresión regula de manera autócrina o parácrina la diferenciación, proliferación y el estado de activación de éstas y muchas otras células. En general, el TGF-beta1 tiene propiedades pleiotrópicas en el contexto de la respuesta inmune durante el desarrollo de infecciones y procesos neoplásicos; sin embargo, los mecanismos de acción y r...

  10. Transforming growth factor-beta 1 downregulates dexamethasone-induced tetranectin gene expression during the in vitro mineralization of the human osteoblastic cell line SV-HFO

    DEFF Research Database (Denmark)

    Iba, K; Sawada, N; Chiba, H;

    1995-01-01

    In the present study, we examined the regulation of tetranectin gene expression using a human osteoblastic cell line, SV-HFO, that undergoes mineralization upon treatment with dexamethasone. We found that the expression of tetranectin and alkaline phosphatase mRNA was induced by dexamethasone...... treatment as evidenced by Northern blotting. When transforming growth factor-beta 1 (TGF-beta 1) was added together with dexamethasone to the SV-HFO cell cultures, the mineralization process was markedly suppressed and the expression of tetra nectin and alkaline phosphatase was downregulated in a dose......-dependent manner. These results demonstrate that the expression of tetranectin in these osteoblastic cells is regulated by dexamethasone and TGF-beta 1 and that tetranectin expression is tightly linked to the process of mineralization....

  11. Combined effects of moderately elevated blood glucose and locally produced TGF-beta1 on glomerular morphology and renal collagen production

    DEFF Research Database (Denmark)

    Krag, Søren; Nyengaard, Jens R; Wogensen, Lise

    2007-01-01

    BACKGROUND: There is a correlation between renal graft rejection and blood glucose (BG) levels. Furthermore, diabetic patients may develop non-diabetic renal diseases, which in some circumstances progress rapidly. Since transforming growth factor-beta1 (TGF-beta) levels are elevated in many renal...... glomerular volume, total mesangial volume and renal collagen content. Moreover, high BG exaggerated TGF-beta1-induced changes in the BMT, MMP-2 and TIMP-1 expression and pERK1/2 staining. CONCLUSION: Even moderate elevations in BG accelerate the progression of those kidney diseases in which TGF-beta1 is...... involved. This emphasizes the importance of strict BG control in renal transplant patients and diabetic patients with renal malfunctions unrelated to diabetes. Udgivelsesdato: 2007-Sep...

  12. The Modifying Effect of Beta 1,3-D Glucan on Gamma Radiation Induced Oxidative Stress and Genotoxicity in Male Mice

    International Nuclear Information System (INIS)

    Beta 1,3-D glucan is a natural polysacharide dericed from the cell walls of baker's yeast Saccharomyces Cerevisiae, with immunoenhancing capabilities. This study was performed to evaluate the efficacy of beta 1,3-D glucan on antioxidant status (superoxide dismutase, SOD and catalase activities and the contents of reduced glutathione (GSH) and lipid peroxides (TBARS) in liver and blood of male mice. In addition, cytogenetic end points (sister chromatid exchanges) were identified in bone marrow. Mice were exposed to whole body gamma radiation, delivered as 2 Gy every other day up to dose 8 Gy. Beta 1,3-D glucan was supplemented daily (65 mg/kg b wt/day) by gavage for 15 days before and during exposure to gamma irradiation. Experimental investigations were carried out 1, 7 and 14 days after exposure to the last radiation dose. The results obtained showed that beta 1,3-D glucan significantly minimized the radiation induced increase in the amount of TBARS in liver and blood of irradiated mice. Furthermore, significant amelioration in SOD and catalase activities was observed 7 and 14 days from the last irradiation fraction in liver and blood. The results obtained showed that the amelioration in the decrease of reduced glutatgione in liver was obvious 7 and 14 days after irradiation Supplementation of beta 1,3-D glucan was also effective in minimizing the radiation induced increase in sister chromatid exchanges throughout the experimental period. It could be concluded that intensifying antioxidant capability of irradiated mice by beta 1,3-D glucan administration, could ameliorate the genotoxicity signs that appears after radiation exposure

  13. The α11 integrin mediates fibroblast-extracellular matrix-cardiomyocyte interactions in health and disease.

    Science.gov (United States)

    Civitarese, Robert A; Talior-Volodarsky, Ilana; Desjardins, Jean-Francois; Kabir, Golam; Switzer, Jennifer; Mitchell, Melissa; Kapus, Andras; McCulloch, Christopher A; Gullberg, Donald; Connelly, Kim A

    2016-07-01

    Excessive cardiac interstitial fibrosis impairs normal cardiac function. We have shown that the α11β1 (α11) integrin mediates fibrotic responses to glycated collagen in rat myocardium by a pathway involving transforming growth factor-β. Little is known of the role of the α11 integrin in the developing mammalian heart. Therefore, we examined the impact of deletion of the α11 integrin in wild-type mice and in mice treated with streptozotocin (STZ) to elucidate the role of the α11 integrin in normal cardiac homeostasis and in the pathogenesis of diabetes-related fibrosis. As anticipated, cardiac fibrosis was reduced in α11 integrin knockout mice (α11(-/-); C57BL/6 background) treated with STZ compared with STZ-treated wild-type mice (P organization at intercalated disks and impaired gap-junction development. Overall, deletion of the α11 integrin attenuates cardiac fibrosis in the mammalian mouse heart and reduces ECM formation as a result of diabetes. Furthermore, α11 integrin deletion impairs cardiac function and alters cardiomyocyte morphology. These findings shed further light on the poorly understood interaction between the fibroblast-cardiomyocyte and the ECM. PMID:27199132

  14. Untersuchung der Sicherheit und Verträglichkeit von Interferon-beta-1b bei Kindern und Jugendlichen mit Multipler Sklerose

    OpenAIRE

    Schelensky, Lilien

    2012-01-01

    Multiple sclerosis (MS) is the most common neurological disease of non-traumatic origin, which leads in young adulthood to permanent disability. 3-5% of patients experience the clinical onset of their disease in childhood and adolescence. The treatment of MS currently consists mainly in an immunemodulatory therapy with interferon beta-1a (INFß-1a, i.m. / s.c.), interferon beta-1b (INFß-1b) or glatiramer acetate. For beta-interferons here the safety, tolerability and effectiveness, especially ...

  15. The formation of tertiary dentin after pulp capping with a calcium phosphate cement, loaded with PLGA microparticles containing TGF-beta1.

    OpenAIRE

    W. Zhang; Walboomers, X.F.; Jansen, J. A.

    2008-01-01

    The aim of the current study was to evaluate the effect of a calcium phosphate material equipped with poly (lactic-co-glycolic acid) microspheres for pulp capping, and to measure the dentin bridge formation, when using various concentrations of transforming growth factor (TGF) beta1. Preset samples were made (2 mm diameter; 2 mm height), containing 0 (controls), 20, or 400 ng TGF-beta1. These were placed in goat incisors. Incisors capped with glass-ionomer cement only were used as negative co...

  16. Loss of β1-integrin from urothelium results in overactive bladder and incontinence in mice: a mechanosensory rather than structural phenotype

    OpenAIRE

    Kanasaki, Keizo; Yu, Weiqun; von Bodungen, Maximilian; Larigakis, John D.; Kanasaki, Megumi; Ayala de la Pena, Francisco; Kalluri, Raghu; Hill, Warren G.

    2013-01-01

    Bladder urothelium senses and communicates information about bladder fullness. However, the mechanoreceptors that respond to tissue stretch are poorly defined. Integrins are mechanotransducers in other tissues. Therefore, we eliminated β1-integrin selectively in urothelium of mice using Cre-LoxP targeted gene deletion. β1-Integrin localized to basal/intermediate urothelial cells by confocal microscopy. β1-Integrin conditional-knockout (β1-cKO) mice lacking urothelial β1-integrin exhibited dow...

  17. The role of mutated amyloid beta 1-42 stimulating dendritic cells in a PDAPP transgenic mouse

    Directory of Open Access Journals (Sweden)

    LI Jia-lin

    2012-06-01

    Full Text Available Background Amyloid plaque is one of the pathological hallmarks of Alzheimer's disease (AD. Anti-beta-amyloid (Aβ immunotherapy is effective in removing brain Aβ, but has shown to be associated with detrimental effects. To avoid severe adverse effects such as meningoencephalitis induced by amyloid beta vaccine with adjuvant, and take advantage of amyloid beta antibody's therapeutic effect on Alzheimer's disease sufficiently, our group has developed a new Alzheimer vaccine with mutated amyloid beta 1-42 peptide stimulating dendritic cells (DC. Our previous work has confirmed that DC vaccine can induce adequate anti-amyloid beta antibody in PDAPP Tg mice safely and efficiently. The DC vaccine can improve impaired learning and memory in the Alzheimer's animal model, and did not cause microvasculitis, microhemorrhage or meningoencephalitis in the animal model. However, the exact mechanism of immunotherapy which reduces Aβ deposition remains unknown. In this report, we studied the mechanism of the vaccine, thinking that this may have implications for better understanding of the pathogenesis of Alzheimer's disease. Methods A new Alzheimer vaccine with mutated amyloid beta 1-42 peptide stimulating DC which were obtained from C57/B6 mouse bone marrow was developed. Amyloid beta with Freund's adjuvant was inoculated at the same time to act as positive control. After the treatment was done, the samples of brains were collected, fixed, cut. Immunohistochemical staining was performed to observe the expression of the nuclear hormone liver X receptor (LXR, membrane-bound protein tyrosine phosphatase (CD45, the ATP-binding cassette family of active transporters (ABCA1, receptor for advanced glycation end products (RAGE, β-site APP-cleaving enzyme (BACE and Aβ in mouse brain tissue. Semi-quantitative analysis was used to defect CA1, CA2, CA3, DG, Rad in hippocampus region and positive neuron in cortex region. Results Aβ was significantly reduced in the

  18. β1 integrins regulate mammary gland proliferation and maintain the integrity of mammary alveoli

    OpenAIRE

    Li, Na; Zhang, Yu; Naylor, Matthew J.; Schatzmann, Franziska; Maurer, Francisca; Wintermantel, Tim; Schuetz, Gunther; Mueller, Ulrich; Streuli, Charles H; Hynes, Nancy E.

    2005-01-01

    Integrin–extracellular matrix interactions play important roles in the coordinated integration of external and internal cues that are essential for proper development. To study the role of β1 integrin in the mammary gland, Itgβ1flox/flox mice were crossed with WAPiCre transgenic mice, which led to specific ablation of β1 integrin in luminal alveolar epithelial cells. In the β1 integrin mutant mammary gland, individual alveoli were disorganized resulting from alterations in cell–basement membr...

  19. Progresses in optimization strategy for radiolabeled molecular probes targeting integrin αvβ3

    International Nuclear Information System (INIS)

    Tumor angiogenesis is critical in the growth, invasion and metastasis of malignant tumors. The integrins, which express on many types of tumor cells and activated vascular endothelial cells, play an important role in regulation of the tumor angiogenesis. RGD peptide, which contains Arg-Gly-Asp sequence, binds specifically to integrin αvβ3. Therefore, the radiolabeled RGD peptides may have broad application prospects in radionuclide imaging and therapy. Major research interests include the selection of radionuclides, modification and improvement of RGD structures. In this article, we give a review on research progresses in optimization strategy for radiolabeled molecular probes targeting integrin αvβ3. (authors)

  20. Integrin β1, Osmosensing, and Chemoresistance in Mouse Ehrlich Carcinoma Cells

    DEFF Research Database (Denmark)

    Sørensen, Belinda Halling; Rasmussen, Line Jee Hartmann; Broberg, Bjørn Sindballe;

    2015-01-01

    RNA was used to silence integrin β1 expression. Regulatory volume decrease (RVD) after cell swelling was studied with calcein-fluorescence-self-quenching and Coulter counter analysis. Taurine efflux was estimated with tracer technique. Caspase assay was used to determine apoptosis. RESULTS: We show...... that adherent cells have stronger fibronectin binding and a significantly increased expression of integrin α5, αv, and β1 at mRNA and protein level, compared to non-adherent cells. Knockdown of integrin β1 reduced RVD of the adherent but not of the non-adherent cells. Efflux of taurine was unaffected...

  1. Isolation and sequencing of a putative promoter region of the murine G protein beta 1 subunit (GNB1) gene.

    Science.gov (United States)

    Kitanaka, Junichi; Kitanaka, Nobue; Takemura, Motohiko; Wang, Xiao-Bing; Hembree, Cambria M; Goodman, Nancy L; Uhl, George R

    2002-02-01

    The expression of the heterotrimeric GTP-binding protein beta 1 subunit gene (GNB1) is regulated by psychostimulants such as cocaine and amphetamines. Since the up-regulation appears to be one of the candidate processes of sensitization, it is necessary to elucidate the cellular and molecular mechanism of the GNB1 gene regulation for a better understanding the establishment of sensitization. In the present study, we describe the isolation and nucleotide sequence analysis of the GNB1 gene promoter region. We have isolated approximately 10 kb of the 5'-flanking region of the mouse of GNB1 gene and found potential elements involved in putative transcriptional control of the GNB1, such as AP1, AP2, Sp1, cyclic AMP response element, and nuclear factor kappa B recognition sites, within the sequences 0.3 kb upstream from the putative transcription start site. This region was highly rich in G + C content, but lacked TATA or CATT boxes. Comparing the nucleotide sequence of the cDNA clone with the human genome databases using the BLAST program a region containing putative exon 1 and promoter of the human GNB1 gene in chromosome 1 was found. The cloning and sequence analysis of an extensive portion of the 5'-flanking regulatory region of the GNB1 gene provides new insights into the factors involved in the regulation by psychostimulants of GNB1 expression. PMID:12180136

  2. CDNA CLONING, PURIFICATION, PROPERTIES, AND FUNCTION OF A BETA-1, E-GLUCAN RECOGNITION PROTEIN FROM A PYRALID MOTH, PLODIA INTERPUNCTELLA

    Science.gov (United States)

    Microorganisms possess distinctive biochemical or molecular patterns on their cell surfaces, such as those formed by the lipopolysaccharides, lipoteichoic acids, and/or peptidoglycans of bacteria and the beta-1,3-glucans of fungi. Pattern recognition proteins that bind to these surface moieties have...

  3. Molecular cloning and characterization of cgt, the Brucella abortus cyclic beta-1,2-glucan transporter gene, and its role in virulence.

    Science.gov (United States)

    Roset, Mara S; Ciocchini, Andrés E; Ugalde, Rodolfo A; Iñón de Iannino, Nora

    2004-04-01

    The animal pathogen Brucella abortus contains a gene cgt, which complemented Sinorhizobium meliloti nodule development (ndvA) and Agrobacterium tumefaciens chromosomal virulence (chvA) mutants. Complemented strains recovered the presence of anionic cyclic beta-1,2-glucan, motility, tumor induction in A. tumefaciens, and nodule occupancy in S. meliloti, all traits strictly associated with the presence of cyclic beta-1,2-glucan in the periplasm. Nucleotide sequencing revealed that B. abortus cgt contains a 1,797-bp open reading frame coding for a predicted membrane protein of 599 amino acids (65.9 kDa) that is 58.5 and 59.9% identical to S. meliloti NdvA and A. tumefaciens ChvA, respectively. Additionally, B. abortus cgt, like S. meliloti ndvA and A. tumefaciens chvA possesses ATP-binding motifs and the ABC signature domain features of a typical ABC transporter. Characterization of Cgt was carried out by the construction of null mutants in B. abortus 2308 and S19 backgrounds. Both mutants do not transport cyclic beta-1,2-glucan to the periplasm, as shown by the absence of anionic cyclic glucan, and they display reduced virulence in mice and defective intracellular multiplication in HeLa cells. These results suggest that cyclic beta-1,2-glucan must be transported into the periplasmatic space to exert its action as a virulence factor. PMID:15039351

  4. CD44 disruption prevents degeneration of the capillary network in obstructive nephropathy via reduction of TGF-beta 1-induced apoptosis

    NARCIS (Netherlands)

    K.M.A. Rouschop; N. Claessen; S.T. Pals; J.J. Weening; S. Florquin

    2006-01-01

    CD44 is a glycoprotein that is involved in inflammation and cell-cell/cell-matrix interactions, is upregulated in the kidney upon injury, and leads to fibrosis through enhancement of TGF-beta 1 signaling. Absence of CD44 prevents development of renal fibrosis in unilateral ureteral obstruction (UUO)

  5. Adverse events of interferon beta-1a: a prospective multi-centre international ICH-GCP-based CRO-supported external validation study in daily practice.

    NARCIS (Netherlands)

    Jongen, P.J.H.; Sindic, C.; Sanders, E.; Hawkins, S.; Linssen, W.; Munster, E. van; Frequin, S.T.F.M.; Borm, G.F.

    2011-01-01

    BACKGROUND: Due to methodological shortcomings the available post-registration data on the adverse events (AEs) occurring in interferon beta-1a (INFb-1a)-treated patients fail to adequately validate phase III data and only partially inform on safety in daily practice. We assessed AEs in relapsing re

  6. Role of beta 1-4 linked polymers in the biofilm structure of marine Pseudomonas sp. CE-2 on 304 stainless steel coupons

    Digital Repository Service at National Institute of Oceanography (India)

    Jain, A.; Bhosle, N.B.

    . 85%). Conversely, the lectins Con A or WGA did not influence the CE-2 biofilms on the SS coupons. Furthermore, treatment with cellulase, an enzyme specific for the degradation of beta 1-4 linked polymers, removed substantial amounts of CE-2 biofilm...

  7. Phosphatase of regenerating liver-3 directly interacts with integrin β1 and regulates its phosphorylation at tyrosine 783

    Directory of Open Access Journals (Sweden)

    Tian Wei

    2012-10-01

    Full Text Available Abstract Background Phosphatase of regenerating liver-3 (PRL-3 or PTP4A3 has been implicated in controlling cancer cell proliferation, motility, metastasis, and angiogenesis. Deregulated expression of PRL-3 is highly correlated with cancer progression and predicts poor survival. Although PRL-3 was categorized as a tyrosine phosphatase, its cellular substrates remain largely unknown. Results We demonstrated that PRL-3 interacts with integrin β1 in cancer cells. Recombinant PRL-3 associates with the intracellular domain of integrin β1 in vitro. Silencing of integrin α1 enhances PRL-3-integrin β1 interaction. Furthermore, PRL-3 diminishes tyrosine phosphorylation of integrin β1 in vitro and in vivo. With site-specific anti-phosphotyrosine antibodies against residues in the intracellular domain of integrin β1, tyrosine-783, but not tyrosine-795, is shown to be dephosphorylated by PRL-3 in a catalytic activity-dependant manner. Phosphorylation of Y783 is potentiated by ablation of PRL-3 or by treatment with a chemical inhibitor of PRL-3. Conversely, depletion of integrin α1 decreases the phosphorylation of this site. Conclusions Our results revealed a direct interaction between PRL-3 and integrin β1 and characterized Y783 of integrin β1 as a bona fide substrate of PRL-3, which is negatively regulated by integrin α1.

  8. CCN2: a mechanosignaling sensor modulating integrin-dependent connective tissue remodeling in fibroblasts?

    Science.gov (United States)

    Leask, Andrew

    2013-08-01

    Tensegrity (tensional integrity) is an emerging concept governing the structure of the body. Integrin-mediated mechanical tension is essential for connective tissue function in vivo. For example, in adult skin fibroblasts, the integrin β1 subunit mediates adhesion to collagen and fibronectin. Moreover, integrin β1, through its abilities to activate latent TGFβ1 and promote collagen production through focal adhesion kinase/rac1/nicotinamide adenine dinucleotide phosphate oxidase (NOX)/reactive oxygen species (ROS), is essential for dermal homeostasis, repair and fibrosis. The integrin β1-interacting protein CCN2, a member of the CCN family of proteins, is induced by TGFβ1; yet, CCN2 is not a simple downstream mediator of TGFβ1, but instead synergistically promote TGFβ1-induced adhesive signaling and fibrosis. Due to its selective ability to sense mechanical forces in the microenvironment, CCN2 may represent an exquisitely precise target for therapeutic intervention. PMID:23729366

  9. The Adenovirus Type 3 Dodecahedron's RGD Loop Comprises an HSPG Binding Site That Influences Integrin Binding

    Directory of Open Access Journals (Sweden)

    H. Lortat-Jacob

    2010-01-01

    Full Text Available Human type 3 adenovirus dodecahedron (a virus like particle made of twelve penton bases features the ability to enter cells through Heparan Sulphate Proteoglycans (HSPGs and integrins interaction and is used as a versatile vector to deliver DNA or proteins. Cryo-EM reconstruction of the pseudoviral particle with Heparan Sulphate (HS oligosaccharide shows an extradensity on the RGD loop. A set of mutants was designed to study the respective roles of the RGD sequence (RGE mutant and of a basic sequence located just downstream. Results showed that the RGE mutant binding to the HS deficient CHO-2241 cells was abolished and unexpectedly, mutation of the basic sequence (KQKR to AQAS dramatically decreased integrin recognition by the viral pseudoparticle. This basic sequence is thus involved in integrin docking, showing a close interplay between HSPGs and integrin receptors.

  10. Diabetes-induced upregulation of urotensin II and its receptor plays an important role in TGF-beta1-mediated renal fibrosis and dysfunction.

    Science.gov (United States)

    Tian, Lin; Li, Cai; Qi, Jiping; Fu, Peng; Yu, Xiaoyan; Li, Xiaokun; Cai, Lu

    2008-11-01

    Urotensin II (UII) was identified as the ligand for a novel G protein-coupled receptor, GPR14. UII was found not only to have a potent vasoconstrictive action but also to have profibrotic effects in the heart. The present study was to define whether UII and GPR14 also play important roles in diabetes-induced renal fibrosis and dysfunction. Diabetic rats were induced using streptozotocin, and the rat proximal tubular epithelial cells (NRK-52E) were used for the in vitro mechanism study. Results showed that expression of UII and GPR14 was significantly upregulated at both mRNA and protein levels in the diabetic kidneys compared with controls. The upregulated expressions of UII and GPR14 in the kidney were accompanied by significant increases in the renal profibrotic factor transforming growth factor (TGF)-beta1 expression, the renal extracellular matrix (fibronectin and collagen IV) accumulation, and the renal dysfunction (increases in urinal N-acetyl-beta-d-glucosaminidase content, 24-h urinary retinol-binding protein excretion rate, and decrease in creatinine clearance rate). Exposure of NRK-52E cells to 10(-8) mol/l UII for 48 h caused a significant increase of TGF-beta1, but not ANG II, production that was GPR14- and calcium-dependent, since GPR14 small-interfering RNA and calcium channel blocker nimodipine or calcium chelator EDTA all could abolish the induction of TGF- beta1 by UII. Furthermore, exposure of NRK-52E cells to TGF-beta1 or ANG II also increased UII and GPR14 mRNA expressions. These results suggested that diabetes-induced upregulation of UII and GPR14, most likely through autocrine and/or paracrine mechanisms, plays an important role in TGF-beta1-mediated renal fibrosis and dysfunction. PMID:18796544

  11. Characterization of a Full-Length Endogenous Beta-Retrovirus, EqERV-Beta1, in the Genome of the Horse (Equus caballus

    Directory of Open Access Journals (Sweden)

    Antoinette C. van der Kuyl

    2011-06-01

    Full Text Available Information on endogenous retroviruses fixed in the horse (Equus caballus genome is scarce. The recent availability of a draft sequence of the horse genome enables the detection of such integrated viruses by similarity search. Using translated nucleotide fragments from gamma-, beta-, and delta-retroviral genera for initial searches, a full-length beta-retrovirus genome was retrieved from a horse chromosome 5 contig. The provirus, tentatively named EqERV-beta1 (for the first equine endogenous beta-retrovirus, was 10434 nucleotide (nt in length with the usual retroviral genome structure of 5’LTR-gag-pro-pol-env-3’LTR. The LTRs were 1361 nt long, and differed approximately 1% from each other, suggestive of a relatively recent integration. Coding sequences for gag, pro and pol were present in three different reading-frames, as common for beta-retroviruses, and the reading frames were completely open, except that the env gene was interrupted by a single stopcodon. No reading frame was apparent downstream of the env gene, suggesting that EqERV-beta1 does not encode a superantigen like mouse mammary tumor virus (MMTV. A second proviral genome of EqERV-beta1, with no stopcodon in env, is additionally integrated on chromosome 5 downstream of the first virus. Single EqERV-beta1 LTRs were abundantly present on all chromosomes except chromosome 24. Phylogenetically, EqERV-beta1 most closely resembles an unclassified retroviral sequence from cattle (Bos taurus, and the murine beta-retrovirus MMTV.

  12. Delivery of dexamethasone from bioactive nanofiber matrices stimulates odontogenesis of human dental pulp cells through integrin/BMP/mTOR signaling pathways.

    Science.gov (United States)

    Lim, Hyun-Chang; Nam, Ok Hyung; Kim, Mi-Joo; El-Fiqi, Ahmed; Yun, Hyung-Mun; Lee, Yoo-Mi; Jin, Guang-Zhen; Lee, Hae-Hyoung; Kim, Hae-Won; Kim, Eun-Cheol

    2016-01-01

    Therapeutically relevant design of scaffolds is of special importance in the repair and regeneration of tissues including dentin and pulp. Here we exploit nanofiber matrices that incorporate bioactive glass nanoparticles (BGNs) and deliver the odontogenic drug dexamethasone (DEX) to stimulate the odontogenic differentiation of human dental pulp cells (HDPCs). DEX molecules were first loaded onto the BGN, and then the DEX-BGN complex was incorporated within the biopolymer nanofiber matrix through electrospinning. The release of DEX continued over a month, showing a slow releasing profile. HDPCs cultured on the DEX-releasing BGN matrices were viable, proliferating well up to 14 days. The odontogenic differentiation, as assessed by alkaline phosphatase activity, mRNA expression of genes, and mineralization, was significantly stimulated on the matrices incorporating BGN and further on those releasing DEX. The DEX-releasing BGN matrices highly upregulated the expression of the integrin subsets α1, α5, and β3 as well as integrin downstream signaling molecules, including focal adhesion kinase (FAK), Paxillin, and RhoA, and activated bone morphogenetic protein mRNA and phosphorylation of Smad1/5/8. Furthermore, the DEX-releasing BGN-matrices stimulated Akt and mammalian target of rapamycin (mTOR), which was proven by the inhibition study. Collectively, the designed therapeutic nanofiber matrices that incorporate BGN and deliver DEX were demonstrated to promote odontogenesis of HDPCs, and the integrins, bone morphogenetic protein, and mTOR signaling pathways are proposed to be the possible molecular mechanisms. While further in vivo studies are still needed, the DEX-releasing bioactive scaffolds are considered as a potential therapeutic nanomatrix for regenerative endodontics and tissue engineering. PMID:27354790

  13. Delivery of dexamethasone from bioactive nanofiber matrices stimulates odontogenesis of human dental pulp cells through integrin/BMP/mTOR signaling pathways

    Science.gov (United States)

    Lim, Hyun-Chang; Nam, Ok Hyung; Kim, Mi-joo; El-Fiqi, Ahmed; Yun, Hyung-Mun; Lee, Yoo-Mi; Jin, Guang-Zhen; Lee, Hae-Hyoung; Kim, Hae-Won; Kim, Eun-Cheol

    2016-01-01

    Therapeutically relevant design of scaffolds is of special importance in the repair and regeneration of tissues including dentin and pulp. Here we exploit nanofiber matrices that incorporate bioactive glass nanoparticles (BGNs) and deliver the odontogenic drug dexamethasone (DEX) to stimulate the odontogenic differentiation of human dental pulp cells (HDPCs). DEX molecules were first loaded onto the BGN, and then the DEX-BGN complex was incorporated within the biopolymer nanofiber matrix through electrospinning. The release of DEX continued over a month, showing a slow releasing profile. HDPCs cultured on the DEX-releasing BGN matrices were viable, proliferating well up to 14 days. The odontogenic differentiation, as assessed by alkaline phosphatase activity, mRNA expression of genes, and mineralization, was significantly stimulated on the matrices incorporating BGN and further on those releasing DEX. The DEX-releasing BGN matrices highly upregulated the expression of the integrin subsets α1, α5, and β3 as well as integrin downstream signaling molecules, including focal adhesion kinase (FAK), Paxillin, and RhoA, and activated bone morphogenetic protein mRNA and phosphorylation of Smad1/5/8. Furthermore, the DEX-releasing BGN-matrices stimulated Akt and mammalian target of rapamycin (mTOR), which was proven by the inhibition study. Collectively, the designed therapeutic nanofiber matrices that incorporate BGN and deliver DEX were demonstrated to promote odontogenesis of HDPCs, and the integrins, bone morphogenetic protein, and mTOR signaling pathways are proposed to be the possible molecular mechanisms. While further in vivo studies are still needed, the DEX-releasing bioactive scaffolds are considered as a potential therapeutic nanomatrix for regenerative endodontics and tissue engineering. PMID:27354790

  14. Transendothelial migration of neutrophils involves integrin-associated protein (CD47).

    OpenAIRE

    Cooper, D; Lindberg, F P; Gamble, J R; Brown, E J; Vadas, M A

    1995-01-01

    Inflammation is a primary pathological process. The development of an inflammatory reaction involves the movement of white blood cells through the endothelial lining of blood vessels into tissues. This process of transendothelial cell migration of neutrophils has been shown to involve neutrophil beta 2 integrins (CD18) and endothelial cell platelet-endothelium cell adhesion molecules (PECAM-1; CD31). We now show that F(ab')2 fragments of the monoclonal antibody B6H12 against integrin-associat...

  15. Targeting of alpha-v integrins reduces malignancy of bladder carcinoma.

    Directory of Open Access Journals (Sweden)

    Geertje van der Horst

    Full Text Available Low survival rates of metastatic cancers emphasize the need for a drug that can prevent and/or treat metastatic cancer. αv integrins are involved in essential processes for tumor growth and metastasis and targeting of αv integrins has been shown to decrease angiogenesis, tumor growth and metastasis. In this study, the role of αv integrin and its potential as a drug target in bladder cancer was investigated. Treatment with an αv integrin antagonist as well as knockdown of αv integrin in the bladder carcinoma cell lines, resulted in reduced malignancy in vitro, as illustrated by decreased proliferative, migratory and clonogenic capacity. The CDH1/CDH2 ratio increased, indicating a shift towards a more epithelial phenotype. This shift appeared to be associated with downregulation of EMT-inducing transcription factors including SNAI2. The expression levels of the self-renewal genes NANOG and BMI1 decreased as well as the number of cells with high Aldehyde Dehydrogenase activity. In addition, self-renewal ability decreased as measured with the urosphere assay. In line with these observations, knockdown or treatment of αv integrins resulted in decreased metastatic growth in preclinical in vivo models as assessed by bioluminescence imaging. In conclusion, we show that αv integrins are involved in migration, EMT and maintenance of Aldehyde Dehydrogenase activity in bladder cancer cells. Targeting of αv integrins might be a promising approach for treatment and/or prevention of metastatic bladder cancer.

  16. Fibronectin increases the force production of mouse papillary muscles via α5β1 integrin

    OpenAIRE

    Wu, Xin; Chakraborty, Sanjukta; Heaps, Cristine L.; Davis, Michael J.; Meininger, Gerald A.; Muthuchamy, Mariappan

    2010-01-01

    The extracellular matrix (ECM) protein-integrin-cytoskeleton axis plays a central role as a mechanotransducing protein assemblage in many cell types. However, how the process of mechanotransduction and the mechanical generated signals arising from this axis affect myofilament function in cardiac muscle are not completely understood. We hypothesize that ECM proteins can regulate cardiac function through integrin binding, and thereby alter the intracellular calcium concentration ([Ca2+]i) and/o...

  17. Structure and binding interface of the cytosolic tails of αXβ2 integrin.

    Directory of Open Access Journals (Sweden)

    Geok-Lin Chua

    Full Text Available BACKGROUND: Integrins are signal transducer proteins involved in a number of vital physiological processes including cell adhesion, proliferation and migration. Integrin molecules are hetero-dimers composed of two distinct subunits, α and β. In humans, 18 α and 8 β subunits are combined into 24 different integrin molecules. Each of the subunit comprises a large extracellular domain, a single pass transmembrane segment and a cytosolic tail (CT. The CTs of integrins are vital for bidirectional signal transduction and in maintaining the resting state of the receptors. A large number of intracellular proteins have been found to interact with the CTs of integrins linking integrins to the cytoskeleton. METHODOLOGY/PRINCIPAL FINDINGS: In this work, we have investigated structure and interactions of CTs of the leukocyte specific integrin αXβ2. We determined the atomic resolution structure of a myristoylated CT of αX in perdeuterated dodecylphosphocholine (DPC by NMR spectroscopy. Our results reveal that the 35-residue long CT of αX adopts an α-helical conformation for residues F4-N17 at the N-terminal region. The remaining residues located at the C-terminal segment of αX delineate a long loop of irregular conformations. A segment of the loop maintains packing interactions with the helical structure by an extended non-polar surface of the αX CT. Interactions between αX and β2 CTs are demonstrated by (15N-(1H HSQC NMR experiments. We find that residues constituting the polar face of the helical conformation of αX are involved in interactions with the N-terminal residues of β2 CT. A docked structure of the CT complex indicates that a network of polar and/or salt-bridge interactions may sustain the heteromeric interactions. CONCLUSIONS/SIGNIFICANCE: The current study provides important insights into the conservation of interactions and structures among different CTs of integrins.

  18. Therapeutic Targeting of Integrin αvβ6 in Breast Cancer

    OpenAIRE

    Moore, Kate M.; Thomas, Gareth J; Duffy, Stephen W; Warwick, Jane; Gabe, Rhian; Chou, Patrick; Ellis, Ian O.; Green, Andrew R.; Haider, Syed; Brouilette, Kellie; Saha, Antonio; Vallath, Sabari; Bowen, Rebecca; Chelala, Claude; Eccles, Diana

    2014-01-01

    Background Integrin αvβ6 promotes migration, invasion, and survival of cancer cells; however, the relevance and role of αvβ6 has yet to be elucidated in breast cancer. Methods Protein expression of integrin subunit beta6 (β6) was measured in breast cancers by immunohistochemistry (n > 2000) and ITGB6 mRNA expression measured in the Molecular Taxonomy of Breast Cancer International Consortium dataset. Overall survival was assessed using Kaplan Meier curves, and bioinformatics statistical analy...

  19. Integrin β3 Is Required in Infection and Proliferation of Classical Swine Fever Virus

    OpenAIRE

    Weiwei Li; Gang Wang; Wulong Liang; Kai Kang; Kangkang Guo; Yanming Zhang

    2014-01-01

    Classical Swine Fever (CSF) is a highly infectious fatal pig disease, resulting in huge economic loss to the swine industry. Integrins are membrane-bound signal mediators, expressed on a variety of cell surfaces and are known as receptors or co-receptors for many viruses. However, the role of integrin β3 in CSFV infection is unknown. Here, through quantitive PCR, immunofluorescence (IFC) and immunocytohistochemistry (ICC), we revealed that ST (swine testicles epithelial) cells have a prominen...

  20. Actin flow and talin dynamics govern rigidity sensing in actin–integrin linkage through talin extension

    OpenAIRE

    Hirata, Hiroaki; Chiam, Keng-Hwee; Lim, Chwee Teck; Sokabe, Masahiro

    2014-01-01

    At cell–substrate adhesion sites, the linkage between actin filaments and integrin is regulated by mechanical stiffness of the substrate. Of potential molecular regulators, the linker proteins talin and vinculin are of particular interest because mechanical extension of talin induces vinculin binding with talin, which reinforces the actin–integrin linkage. For understanding the molecular and biophysical mechanism of rigidity sensing at cell–substrate adhesion sites, we constructed a simple ph...

  1. Coalition of Oct4A and β1 integrins in facilitating metastasis in ovarian cancer

    OpenAIRE

    Samardzija, Chantel; Luwor, Rodney B.; Quinn, Michael A; Kannourakis, George; Jock K Findlay; Ahmed, Nuzhat

    2016-01-01

    Background Ovarian cancer is a metastatic disease and one of the leading causes of gynaecology malignancy-related deaths in women. Cancer stem cells (CSCs) are key contributors of cancer metastasis and relapse. Integrins are a family of cell surface receptors which allow interactions between cells and their surrounding microenvironment and play a fundamental role in promoting metastasis. This study investigates the molecular mechanism which associates CSCs and integrins in ovarian cancer meta...

  2. Single particle tracking with sterol modulation reveals the cholesterol-mediated diffusion properties of integrin receptors

    International Nuclear Information System (INIS)

    A combination of sterol modulation with cyclodextrins plus fluorescence microscopy revealed a biophysical mechanism behind cholesterol’s influence on the diffusion of a ubiquitous class of receptors called integrins. The heterogeneous diffusion of integrins bound to ligand-coated quantum dots was measured using single particle tracking (SPT), and the ensemble changes in integrin diffusion were measured by fluorescence recovery after photobleaching (FRAP). A 25 ± 1% reduction of membrane cholesterol resulted in three significant changes to the diffusion of ligand-bound αPS2CβPS integrins as measured by SPT. There was a 23% increase in ligand-bound mobile integrins; there was a statistically significant increase in the average diffusion coefficient inside zones of confined diffusion, and histograms of confined integrin trajectories showed an increased frequency in the range of 0.1–1 μm2 s−1 and a decreased frequency in the 0.001–0.1 μm2 s−1 range. No statistical change was measured in the duration of confinement nor the size of confined zones. Restoring the cholesterol-depleted cells with exogenous cholesterol or exogenous epicholesterol resulted in similar diffusion properties. Epicholesterol differs from cholesterol in the orientation of a single hydroxyl group. The ability of epicholesterol to substitute for cholesterol suggests a biophysical mechanism for cholesterol’s effect on integrin diffusion. Influences of bilayer thickness, viscosity and organization are discussed as possible explanations for the measured changes in integrin diffusion when the membrane cholesterol concentration is reduced. (paper)

  3. The integrin-collagen connection--a glue for tissue repair?

    Science.gov (United States)

    Zeltz, Cédric; Gullberg, Donald

    2016-02-15

    The α1β1, α2β1, α10β1 and α11β1 integrins constitute a subset of the integrin family with affinity for GFOGER-like sequences in collagens. Integrins α1β1 and α2β1 were originally identified on a subset of activated T-cells, and have since been found to be expressed on a number of cell types including platelets (α2β1), vascular cells (α1β1, α2β1), epithelial cells (α1β1, α2β1) and fibroblasts (α1β1, α2β1). Integrin α10β1 shows a distribution that is restricted to mesenchymal stem cells and chondrocytes, whereas integrin α11β1 appears restricted to mesenchymal stem cells and subsets of fibroblasts. The bulk of the current literature suggests that collagen-binding integrins only have a limited role in adult connective tissue homeostasis, partly due to a limited availability of cell-binding sites in the mature fibrillar collagen matrices. However, some recent data suggest that, instead, they are more crucial for dynamic connective tissue remodeling events--such as wound healing--where they might act specifically to remodel and restore the tissue architecture. This Commentary discusses the recent development in the field of collagen-binding integrins, their roles in physiological and pathological settings with special emphasis on wound healing, fibrosis and tumor-stroma interactions, and include a discussion of the most recently identified newcomers to this subfamily--integrins α10β1 and α11β1. PMID:26857815

  4. Integrins Regulate Apical Constriction via Microtubule Stabilization in the Drosophila Eye Disc Epithelium

    Directory of Open Access Journals (Sweden)

    Vilaiwan M. Fernandes

    2014-12-01

    Full Text Available During morphogenesis, extracellular signals trigger actomyosin contractility in subpopulations of cells to coordinate changes in cell shape. To illuminate the link between signaling-mediated tissue patterning and cytoskeletal remodeling, we study the progression of the morphogenetic furrow (MF, the wave of apical constriction that traverses the Drosophila eye imaginal disc preceding photoreceptor neurogenesis. Apical constriction depends on actomyosin contractility downstream of the Hedgehog (Hh and bone morphogenetic protein (BMP pathways. We identify a role for integrin adhesion receptors in MF progression. We show that Hh and BMP regulate integrin expression, the loss of which disrupts apical constriction and slows furrow progression; conversely, elevated integrins accelerate furrow progression. We present evidence that integrins regulate MF progression by promoting microtubule stabilization, since reducing microtubule stability rescues integrin-mediated furrow acceleration. Thus, integrins act as a genetic link between tissue-level signaling events and morphological change at the cellular level, leading to morphogenesis and neurogenesis in the eye.

  5. RGD-Binding Integrins in Prostate Cancer: Expression Patterns and Therapeutic Prospects against Bone Metastasis

    International Nuclear Information System (INIS)

    Prostate cancer is the third leading cause of male cancer deaths in the developed world. The current lack of highly specific detection methods and efficient therapeutic agents for advanced disease have been identified as problems requiring further research. The integrins play a vital role in the cross-talk between the cell and extracellular matrix, enhancing the growth, migration, invasion and metastasis of cancer cells. Progression and metastasis of prostate adenocarcinoma is strongly associated with changes in integrin expression, notably abnormal expression and activation of the β3 integrins in tumour cells, which promotes haematogenous spread and tumour growth in bone. As such, influencing integrin cell expression and function using targeted therapeutics represents a potential treatment for bone metastasis, the most common and debilitating complication of advanced prostate cancer. In this review, we highlight the multiple ways in which RGD-binding integrins contribute to prostate cancer progression and metastasis, and identify the rationale for development of multi-integrin antagonists targeting the RGD-binding subfamily as molecularly targeted agents for its treatment

  6. Multiple proteins of White spot syndrome virus involved in recognition of -integrin

    Indian Academy of Sciences (India)

    Jing-Yan Zhang; Qing-Hui Liu; Jie Huang

    2014-06-01

    The recognition and attachment of virus to its host cell surface is a critical step for viral infection. Recent research revealed that -integrin was involved in White spot syndrome virus (WSSV) infection. In this study, the interaction of -integrin with structure proteins of WSSV and motifs involved in WSSV infection was examined. The results showed that envelope proteins VP26, VP31, VP37, VP90 and nucleocapsid protein VP136 interacted with LvInt. RGD-, YGL- and LDV-related peptide functioned as motifs of WSSV proteins binding with -integrin. The -integrin ligand of RGDT had better blocking effect compared with that of YGL- and LDV-related peptides. In vivo assay indicated that RGD-, LDV- and YGL-related peptides could partially block WSSV infection. These data collectively indicate that multiple proteins were involved in recognition of -integrin. Identification of proteins in WSSV that are associated with -integrin will assist development of new agents for effective control of the white spot syndrome.

  7. Association of Transforming Growth Factor Beta-1-509C/T Gene Polymorphism with Ischemic Stroke: A Meta Analysis

    Science.gov (United States)

    Kumar, Pradeep; Kumar, Amit; Srivastava, Mukesh Kumar; Misra, Shubham; Pandit, Awadh Kishor; Prasad, Kameshwar

    2016-01-01

    Introduction: Transforming Growth Factor-Beta 1 (TGF-β1) is a pleiotropic cytokine with potent anti-inflammatory property, which has been considered as an essential risk factor in the inflammatory process of Ischemic Stroke (IS), by involving in the pathophysiological progression of hypertension, atherosclerosis, and lipid metabolisms. -509C/T TGF-β1 gene polymorphism has been found to be associated with the risk of IS. The aim of this meta-analysis was to provide a relatively comprehensive account of the relation between -509C/T gene polymorphisms of TGF-β1 and susceptibility to IS. Methods: A review of literature for eligible genetic association Studies published before October 20, 2014 was conducted in the PubMed, EMBASE, Google Scholar and Trip database. The strength of association was calculated by pooled odds ratios (ORs) with 95% confidence intervals using RevMan 5.3 software. Heterogeneity was examined using Higgins I-squared, Tau-squared, and Chi-squared tests. Results: A total of 2 studies involving 614 cases and 617 controls were found. The overall estimates did not show any significant relation between TGF-β1-509C/T polymorphism and risk of IS under dominant (CC+CT vs. TT: OR=1.01, 95%CI=0.31 to 3.26; P=0.99), recessive (CC vs. CT+TT: OR=0.94, 95%CI=0.47 to 1.90; P=0.87), and allelic models (T vs. C: OR=1.06, 95%CI=0.55 to 2.04; P=0.86). Conclusion: This meta-analysis showed that TGF-β1-509C/T gene polymorphism has no significant association with the susceptibility of IS. Further well-designed prospective studies with larger sample size are needed to confirm these findings. PMID:27303603

  8. Interferon Beta-1a (AVONEX® as a Treatment Option for Untreated Patients with Multiple Sclerosis (AXIOM: A Prospective, Observational Study

    Directory of Open Access Journals (Sweden)

    Christoph Kleinschnitz

    2015-07-01

    Full Text Available The efficacy and safety of first-line disease-modifying therapies (DMT for relapsing-remitting multiple sclerosis (RRMS has been demonstrated in pivotal, randomized trials, but these studies do not reflect the routine care setting where treatment gaps or switches are common. The Avonex as Treatment Option for Untreated MS Patients (AXIOM trial assessed the efficacy of newly-initiated intramuscular interferon beta-1a (IM IFNb-1a after a treatment-free interval, with particular consideration of the previous course of disease and therapy. The AXIOM trial was an open, 12-month, observational, non-interventional study with a retrospective and a prospective part conducted in Germany. RRMS patients with a treatment-free interval of at least three months were included and treated with IFNb-1a for up to 12 months. Relapse rate, disability progression, injection-related parameters and quality of life observed during the prospective part were compared with retrospectively-collected data. Two hundred and thirty five RRMS patients participated in AXIOM. The mean relapse rate decreased from 1.1 in the three months before baseline to 0.2 per quarter during the twelve-month observational period; the Multiple Sclerosis Functional Composite score improved during twelve months of IM IFNb-1a treatment, while the Expanded Disability Status Scale score did not change over the course of this study. Compared to previous DMTs (IM IFNb-1a, subcutaneous IFNb-1a (SC IFNb-1a, SC IFNb-1b, glatiramer acetate, the patients experienced less injection site reactions and flu-like symptoms, with a stated improved quality of life. IM IFNb-1a was effective and well accepted in RRMS patients with no or discontinued previous therapy. These results from the routine care setting may inform optimization of DMT treatment in RRMS, but need confirmation in further studies.

  9. Computational analysis of the R85C and R85H epilepsy mutations in Na+ channel beta1 subunits.

    Science.gov (United States)

    Thomas, E A; Xu, R; Petrou, S

    2007-07-29

    Mutations in Na+ channels cause a variety of epilepsy syndromes. Analysis of these mutations shows a range of simultaneous functional consequences, each of which may increase or decrease membrane excitability, making it difficult to predict the combined effect on neuron firing. This may be addressed by building mathematical models of Na+ channel gating and using them in neuron models to predict responses to natural stimuli. The R85C and R85H mutations of the beta1 subunit cause generalized epilepsy syndromes in humans, and an experimental study showed that these mutations shift steady-state activation in the negative direction, which predicts increased excitability, and shift fast inactivation in the negative direction, which predicts decreased excitability. In addition, the R85C also shifts slow inactivation in the negative direction. To predict changes in neuron excitability resulting from these contradictory effects we built Na+ channel models based on our earlier data and on new measurements of the rate of slow inactivation over a range of potentials. Use of these Na+ channel models in simple neuron models revealed that both mutations cause an increase in excitability but the R85H mutation was more excitable. This is due to differences in steady-state slow inactivation and to subtle differences in fast kinetics captured by the model fitting process. To understand the effect of changes in different gating processes and to provide a simple guide for interpreting changes caused by mutations, we performed a sensitivity analysis. Using the wild-type model we shifted each activation curve by +/-5 mV or altered gating rates up or down by 20%. Excitability was most sensitive to changes in voltage dependence of activation, followed by voltage dependence of inactivation and then slow inactivation. By contrast, excitability was relatively insensitive to gating rates. PMID:17604911

  10. Expression and clinical significance of dendritic cell and transforming growth factor-beta 1 in cervical cancer

    Institute of Scientific and Technical Information of China (English)

    Zhao; Shan; Rong; Fengnian

    2006-01-01

    Objective:To explore the density and mature status of Dendritic cell(DC) in cervical cancer and correlation with the expression of transforming growth factor-beta 1(TGF-β1).Methods:Streptavidin-peroxidase(SP) immunohistochemistry methods were used to detect S-100 DC and the expression of TGF-β1 in 20 normal cervical tissues and 53 cervical cancer tissues without any sort of chemotherapy or radiation therapy prior to resection.Medical records were reviewed,clinicopathological variables were retrieved and used for analysis.Results:Two types of DC were observed under the microscope.The expression of DC in cervical cancer was significantly higher than that in normal tissues(23.34 cells/mm2 vs 29.91 cells/mm2,P<0.05),and significantly higher in early stage than that in advanced stage(P<0.05).The expression of TGF-β1 was significantly higher in cervical cancer than that in normal tissues (P<0.025).However,there was no correaction between TGF-β1 and lymph nodes metastasis.The index of DC in cervical cancer was negatively correlated to the expression of TGF-β1 in tumor cells (r=-0.8875,P=0.0001).Conclusion:Maturation of DC in cervical cancer is inhibited.The decreased number of DC and the higher expression of TGF-β1 are due to the failure of the immunity,these may play an important role in the development of the cervical cancer.

  11. Management Strategies for Flu-Like Symptoms and Injection-Site Reactions Associated with Peginterferon Beta-1a

    Science.gov (United States)

    Centonze, Diego; Newsome, Scott D.; Huang, DeRen; Robertson, Christopher; You, Xiaojun; Sabatella, Guido; Evilevitch, Vladimir; Leahy, Leslie

    2016-01-01

    Background: Flu-like symptoms (FLSs) and injection-site reactions (ISRs) have been reported with interferon beta treatments for multiple sclerosis (MS). We sought to obtain consensus on the characteristics/management of FLSs/ISRs in patients with relapsing-remitting MS based on experiences from the randomized, placebo-controlled ADVANCE study of peginterferon beta-1a. Methods: ADVANCE investigators with a predefined number of enrolled patients were eligible to participate in a consensus-generating exercise using a modified Delphi method. An independent steering committee oversaw the development of two sequential Delphi questionnaires. An average rating (AR) of 2.7 or more was defined as consensus a priori. Results: Thirty and 29 investigators (ie, responders) completed questionnaires 1 and 2, respectively, representing 374 patients from ADVANCE. Responders reported that the incidence/duration of FLSs/ISRs in their typical patient generally declined after 3 months of treatment. Responders reached consensus that FLSs typically last up to 24 hours (AR = 3.17) and have mild/moderate effects on activities of daily living (AR = 3.34). Patients should initiate acetaminophen/nonsteroidal anti-inflammatory drug treatment on a scheduled basis (AR = 3.31) and change the timing of injection (AR = 3.28) to manage FLSs. Injection-site rotation/cooling and drug administration at room temperature (all AR ≥ 3.10) were recommended for managing ISRs. Patient education on FLSs/ISRs was advocated before treatment initiation. Conclusions: Delphi responders agreed on the management strategies for FLSs/ISRs and agreed that patient education is critical to set treatment expectations and promote adherence. PMID:27551246

  12. Development of a high-sensitivity immunoassay for amyloid-beta 1-42 using a silicon microarray platform.

    Science.gov (United States)

    Gagni, Paola; Sola, Laura; Cretich, Marina; Chiari, Marcella

    2013-09-15

    In this work, we present a highly sensitive immunoassay for the detection of the Alzheimer's disease (AD) biomarker amyloid-beta 1-42 (Aβ42) based on a label/label-free microarray platform that utilises silicon/silicon oxide (Si/SiO2) substrates. Due to constructive interference, Si/SiO2 layered slides allow enhancement of the fluorescence intensity on the surface with significant improvements in sensitivity of detection. The same substrate allows the label-free multiplexed detection of targets using the Interferometric Reflectance Imaging Sensor (IRIS), a platform amenable to high-throughput detection of mass changes on microarray substrates. Silicon chips are coated with copoly(DMA-NAS-MAPS), a ter-copolymer made from dimethylacrylamide (DMA), 3-(trimethoxysilyl)propyl methacrylate (MAPS) and N-Acryloyloxy succinimide (NAS). Aβ42 aggregation was studied by circular dichroism (CD), and an optimal antibody pair was selected based on specificity of recognition, binding yield and spot morphology of the capture antibody on the coated silicon surface as analysed by IRIS. Finally, incubation conditions were optimised, and an unprecedented Aβ42 detection sensitivity of 73pg/mL was achieved using an artificial cerebrospinal fluid (CSF) sample. Because of their multiplexing capability, low volume sample consumption and efficient sample-to-result time for population-wide screening, microarrays are ideal tools for the identification of individuals with preclinical AD who are still cognitively healthy. The high sensitivity of this assay format, potentially coupled to a pre-concentration step or signal-enhancing modifications, could lead to a non-invasive, inexpensive diagnostic tool for population-wide screening of AD biomarkers in biological fluids other than CSF, such as serum or plasma. PMID:23624018

  13. Rational design of a protein that binds integrin αvβ3 outside the ligand binding site

    Science.gov (United States)

    Turaga, Ravi Chakra; Yin, Lu; Yang, Jenny J.; Lee, Hsiauwei; Ivanov, Ivaylo; Yan, Chunli; Yang, Hua; Grossniklaus, Hans E.; Wang, Siming; Ma, Cheng; Sun, Li; Liu, Zhi-Ren

    2016-01-01

    Integrin αvβ3 expression is altered in various diseases and has been proposed as a drug target. Here we use a rational design approach to develop a therapeutic protein, which we call ProAgio, that binds to integrin αvβ3 outside the classical ligand-binding site. We show ProAgio induces apoptosis of integrin αvβ3-expressing cells by recruiting and activating caspase 8 to the cytoplasmic domain of integrin αvβ3. ProAgio also has anti-angiogenic activity and strongly inhibits growth of tumour xenografts, but does not affect the established vasculature. Toxicity analyses demonstrate that ProAgio is not toxic to mice. Our study reports a new integrin-targeting agent with a unique mechanism of action, and provides a template for the development of integrin-targeting therapeutics. PMID:27241473

  14. Correlation study of integrin genes and bronchial asthma%整合素基因与支气管哮喘相关性的研究

    Institute of Scientific and Technical Information of China (English)

    甄丽华; 邵玉霞

    2014-01-01

    their proliferation,apoptosis and differentiation.A variety of human integrin heterodimers are formed from 9 types of β subunits and 24 types of α subunits.They are heterodimers and function as transmembrane linkers between the extracellular matrix and the actin cytoskeleton nction as signal transducers,activating various intracellular signaling pathways when activated by matrix binding.Integrins and conventional signaling receptors often cooperate to promote cell growth,cell survival,and cell proliferation.In recent years different integrin associated with asthma research more and more,the paper elaborated following different integrin.

  15. Integrin α1β1 Promotes Caveolin-1 Dephosphorylation by Activating T Cell Protein-tyrosine Phosphatase*

    Science.gov (United States)

    Borza, Corina M.; Chen, Xiwu; Mathew, Sijo; Mont, Stacey; Sanders, Charles R.; Zent, Roy; Pozzi, Ambra

    2010-01-01

    Integrin α1β1 is a collagen receptor that down-regulates collagen and reactive oxygen species (ROS) production, and mice lacking this receptor show increased ROS levels and exacerbated glomerular sclerosis following injury. Caveolin-1 (Cav-1) is a multifunctional protein that is tyrosine-phosphorylated in response to injury and has been implicated in ROS-mediated injury. Cav-1 interacts with integrins, and integrin α1β1 binds/activates T cell protein-tyrosine phosphatase (TCPTP), which is homologous to the tyrosine phosphatase PTP1B known to dephosphorylate Cav-1. In this study, we analyzed whether phosphorylated Cav-1 (pCav-1) is a substrate of TCPTP and if integrin α1β1 is essential for promoting TCPTP-mediated Cav-1 dephosphorylation. We found that Cav-1 phosphorylation is significantly higher in cells lacking integrin α1β1 at base line and following oxidative stress. Overexpression of TCPTP leads to reduced pCav-1 levels only in cells expressing integrin α1β1. Using solid phase binding assays, we demonstrated that 1) purified Cav-1 directly interacts with TCPTP and the integrin α1 subunit, 2) pCav-1 is a substrate of TCPTP, and 3) TCPTP-mediated Cav-1 dephosphorylation is highly increased by the addition of purified integrin α1β1 or an integrin α1 cytoplasmic peptide to which TCPTP has been shown to bind. Thus, our results demonstrate that pCav-1 is a new substrate of TCPTP and that integrin α1β1 acts as a negative regulator of Cav-1 phosphorylation by activating TCPTP. This could explain the protective function of integrin α1β1 in oxidative stress-mediated damage and why integrin α1-null mice are more susceptible to fibrosis following injury. PMID:20940300

  16. Key Interactions in Integrin Ectodomain Responsible for Global Conformational Change Detected by Elastic Network Normal-Mode Analysis

    OpenAIRE

    Matsumoto, Atsushi; Kamata, Tetsuji; Takagi, Junichi; Iwasaki, Kenji; Yura, Kei

    2008-01-01

    Integrin, a membrane protein with a huge extracellular domain, participates in cell-cell and cell-extracellular-matrix interactions for metazoan. A group of integrins is known to perform a large-scale structural change when the protein is activated, but the activation mechanism and generality of the conformational change remain to be elucidated. We performed normal-mode analysis of the elastic network model on integrin α V β 3 ectodomain in the bent form and identified key residues that influ...

  17. Increasing α7β1-integrin promotes muscle cell proliferation, adhesion, and resistance to apoptosis without changing gene expression

    OpenAIRE

    LIU, Jianming; Burkin, Dean J.; Kaufman, Stephen J.

    2007-01-01

    The dystrophin-glycoprotein complex maintains the integrity of skeletal muscle by associating laminin in the extracellular matrix with the actin cytoskeleton. Several human muscular dystrophies arise from defects in the components of this complex. The α7β1-integrin also binds laminin and links the extracellular matrix with the cytoskeleton. Enhancement of α7-integrin levels alleviates pathology in mdx/utrn−/− mice, a model of Duchenne muscular dystrophy, and thus the integrin may functionally...

  18. Adrenergic activation of electrogenic K+ secretion in guinea pig distal colonic epithelium: involvement of beta1- and beta2-adrenergic receptors.

    Science.gov (United States)

    Zhang, Jin; Halm, Susan T; Halm, Dan R

    2009-08-01

    Adrenergic stimulation of electrogenic K+ secretion in isolated mucosa from guinea pig distal colon required activation of two beta-adrenergic receptor subtypes (beta-AdrR). Addition of epinephrine (epi) or norepinephrine (norepi) to the bathing solution of mucosae in Ussing chambers increased short-circuit current (Isc) and transepithelial conductance (Gt), consistent with this cation secretion. A beta-adrenergic classification was supported by propranolol antagonism of this secretory response and the lack of effect by the alpha-AdrR antagonists BE2254 (alpha1-AdrR) and yohimbine (alpha2-AdrR). Subtype-selective antagonists CGP20712A (beta1-AdrR), ICI-118551 (beta2-AdrR), and SR59320A (beta3-AdrR) were relatively ineffective at inhibiting the epi-stimulated Isc response. In combination, CGP20712A and ICI-118551 inhibited the response, which supported a synergistic action by beta1-AdrR and beta2-AdrR. Expression of mRNA for both beta1-AdrR and beta2-AdrR was indicated by RT-PCR of RNA from colonic epithelial cells. Protein expression was indicated by immunoblot showing bands at molecular weights consistent with monomers and oligomers. Immunoreactivity (ir) for beta1-AdrR and beta2-AdrR was prominent in basolateral membranes of columnar epithelial cells in the crypts of Lieberkühn as well as intercrypt surface epithelium. Cells in the pericryptal sheath also had beta1-AdrR(ir) but did not have discernable beta2-AdrR(ir). The adrenergic sensitivity of K+ secretion measured by Isc and Gt was relatively low as indicated by EC(50)s of 41 +/- 7 nM for epi and 50 +/- 14 nM for norepi. Adrenergic activation of electrogenic K+ secretion required the involvement of both beta1-AdrR and beta2-AdrR, occurring with an agonist sensitivity reduced compared with reported values for either receptor subtype. PMID:19460844

  19. Bidirectional remodeling of β1-integrin adhesions during chemotropic regulation of nerve growth

    Directory of Open Access Journals (Sweden)

    Carlstrom Lucas P

    2011-11-01

    Full Text Available Abstract Background Chemotropic factors in the extracellular microenvironment guide nerve growth by acting on the growth cone located at the tip of extending axons. Growth cone extension requires the coordination of cytoskeleton-dependent membrane protrusion and dynamic adhesion to the extracellular matrix, yet how chemotropic factors regulate these events remains an outstanding question. We demonstrated previously that the inhibitory factor myelin-associated glycoprotein (MAG triggers endocytic removal of the adhesion receptor β1-integrin from the growth cone surface membrane to negatively remodel substrate adhesions during chemorepulsion. Here, we tested how a neurotrophin might affect integrin adhesions. Results We report that brain-derived neurotropic factor (BDNF positively regulates the formation of substrate adhesions in axonal growth cones during stimulated outgrowth and prevents removal of β1-integrin adhesions by MAG. Treatment of Xenopus spinal neurons with BDNF rapidly triggered β1-integrin clustering and induced the dynamic formation of nascent vinculin-containing adhesion complexes in the growth cone periphery. Both the formation of nascent β1-integrin adhesions and the stimulation of axon extension by BDNF required cytoplasmic calcium ion signaling and integrin activation at the cell surface. Exposure to MAG decreased the number of β1-integrin adhesions in the growth cone during inhibition of axon extension. In contrast, the BDNF-induced adhesions were resistant to negative remodeling by MAG, correlating with the ability of BDNF pretreatment to counteract MAG-inhibition of axon extension. Pre-exposure to MAG prevented the BDNF-induced formation of β1-integrin adhesions and blocked the stimulation of axon extension by BDNF. Conclusions Altogether, these findings demonstrate the neurotrophin-dependent formation of integrin-based adhesions in the growth cone and reveal how a positive regulator of substrate adhesions can block

  20. Glioma cell dispersion is driven by α5 integrin-mediated cell-matrix and cell-cell interactions.

    Science.gov (United States)

    Blandin, Anne-Florence; Noulet, Fanny; Renner, Guillaume; Mercier, Marie-Cécile; Choulier, Laurence; Vauchelles, Romain; Ronde, Philippe; Carreiras, Franck; Etienne-Selloum, Nelly; Vereb, Gyorgy; Lelong-Rebel, Isabelle; Martin, Sophie; Dontenwill, Monique; Lehmann, Maxime

    2016-07-01

    Glioblastoma multiform (GBM) is the most common and most aggressive primary brain tumor. The fibronectin receptor, α5 integrin is a pertinent novel therapeutic target. Despite numerous data showing that α5 integrin support tumor cell migration and invasion, it has been reported that α5 integrin can also limit cell dispersion by increasing cell-cell interaction. In this study, we showed that α5 integrin was involved in cell-cell interaction and gliomasphere formation. α5-mediated cell-cell cohesion limited cell dispersion from spheroids in fibronectin-poor microenvironment. However, in fibronectin-rich microenvironment, α5 integrin promoted cell dispersion. Ligand-occupied α5 integrin and fibronectin were distributed in fibril-like pattern at cell-cell junction of evading cells, forming cell-cell fibrillar adhesions. Activated focal adhesion kinase was not present in these adhesions but was progressively relocalized with α5 integrin as cell migrates away from the spheroids. α5 integrin function in GBM appears to be more complex than previously suspected. As GBM overexpressed fibronectin, it is most likely that in vivo, α5-mediated dissemination from the tumor mass overrides α5-mediated tumor cell cohesion. In this respect, α5-integrin antagonists may be useful to limit GBM invasion in brain parenchyma. PMID:27063097

  1. TGF-beta receptor 2 downregulation in tumour-associated stroma worsens prognosis and high-grade tumours show more tumour-associated macrophages and lower TGF-beta1 expression in colon carcinoma: a retrospective study

    Directory of Open Access Journals (Sweden)

    Papadopoulos Thomas

    2007-08-01

    Full Text Available Abstract Background Histological phenotype and clinical behaviour of malignant tumours are not only dependent on alterations in the epithelial cell compartment, but are affected by their interaction with inflammatory cells and tumour-associated stroma. Studies in animal models have shown influence of tumour-associated macrophages (TAM on histological grade of differentiation in colon carcinoma. Disruption of transforming growth factor beta (TGF-beta signalling in tumour cells is related to more aggressive clinical behaviour. Expression data of components of this pathway in tumour-associated stroma is limited. Methods Tissue micro arrays of 310 colon carcinomas from curatively resected patients in UICC stage II and III were established. In a first step we quantified amount of CD68 positive TAMs and expression of components of TGF-beta signalling (TGF-beta1, TGF-beta receptors type 1 and 2, Smad 3 and 4 in tumour and associated stroma. Further we analyzed correlation to histological and clinical parameters (histological grade of differentiation (low-grade (i.e. grade 1 and 2 vs. high-grade (i.e. grade 3 and 4, lymph node metastasis, distant metastasis, 5 year cancer related survival using Chi-square or Fisher's exact test, when appropriate, to compare frequencies, Kaplan-Meier method to calculate 5-year rates of distant metastases and cancer-related survival and log rank test to compare the rates of distant metastases and survival. To identify independent prognostic factors Cox regression analysis including lymph node status and grading was performed. Results High-grade tumours and those with lymph node metastases showed higher rates of TAMs and lower expression of TGF-beta1. Loss of nuclear Smad4 expression in tumor was associated with presence of lymph node metastasis, but no influence on prognosis could be demonstrated. Decrease of both TGF-beta receptors in tumour-associated stroma was associated with increased lymph node metastasis and

  2. Fibrinogen interaction of CHO cells expressing chimeric αIIb/αvβ3 integrin

    Institute of Scientific and Technical Information of China (English)

    Juan-juan CHEN; Xiao-yu SU; Xiao-dong XI; Li-ping LIN; Jian DING; He LU

    2008-01-01

    Aim: The molecular mechanisms of the affinity regulation of αvβ3 integrin are important in tumor development, wound repairing, and angiogenesis. It has been established that the cytoplasmic domains of αvβ3 integrin play an important role in integrin-ligand affinity regulation. However, the relationship of structure-func-tion within these domains remains unclear. Methods: The extracellular and trans-membrane domain of αⅡb was fused to the αv integrin cytoplasmic domain, and the chimeric α subunit was coexpressed in Chinese hamster ovary (CHO) cells with the wild-type β3 subunit or with 3 mutant 133 sequences bearing truncations at the positions of T741, Y747, and F754, respectively. The CHO cells expressing these recombinant integrins were tested for soluble fibrinogen binding and the cell adhesion and spreading on immobilized fibrinogen. Results: All 4 types of integrins bound soluble fibrinogen in the absence of agonist stimulation, and only the cells expressing the chimeric α subunit with the wild-type β3 subunit, but not those with truncated β3, could adhere to and spread on immobilized fibrinogen. Conclusion: The substitution αⅡb at the cytoplasmic domain with the ctv cyto-plasmic sequence rendered the extracellular αⅡbβ3 a constitutively activated con-formation for ligands without the need of "inside-out" signals. Our results also indicated that the COOH-terminal sequence of β3 might play a key role in integrin αⅡb/αvβ3-mediated cell adhesion and spreading on immobilized fibrinogen. The cells expressing αⅡb/αvβ3 have enormous potential for facilitating drug screen-ing for antagonists either to αvβ3 intracellular interactions or to αⅡbβ3 receptor functions.

  3. The role of integrin α(V)β(3) in osteocyte mechanotransduction.

    Science.gov (United States)

    Haugh, Matthew G; Vaughan, Ted J; McNamara, Laoise M

    2015-02-01

    Recent in vivo studies have proposed that integrin αvβ3 attachments between osteocyte cell processes and the extracellular matrix may facilitate mechanosensation in bone. However the role of these attachments in osteocyte biochemical response to mechanical stimulus has yet to be investigated. With this in mind, the objective of this study was to determine the effect of blocking integrin αvβ3 function on the biochemical response of osteocytes to mechanical stimulus. Antagonists specific to integrin subunit β3 were used to block integrin αvβ3 on MLO-Y4 mouse osteocytes. After treatment, cells were subjected to laminar oscillatory fluid flow stimulus (1 Pa, 1 Hz) for one hour. Fluorescent staining was performed to visualise cell morphology. Prostaglandin E2 (PGE2) release was assayed using an enzyme immunoassay and qRT-PCR was used to analyse the relative expression of cyclooxygenase-2 (COX-2), receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG). Our results show that blocking integrin αvβ3 disrupts osteocyte morphology, causing a reduction in spread area and process retraction. Integrin αvβ3 blocking also disrupted COX-2 expression and PGE2 release in response to fluid shear stress. Taken together, the results of this study indicate that integrin αvβ3 is essential for the maintenance of osteocyte cell processes and also for mechanosensation and mechanotransduction by osteocytes. A better understanding of this process may lead to the development of novel treatments for bone pathologies where mechanosensitivity is thought to be compromised. PMID:25460927

  4. Integrin Receptors Play a Key Role in the Regulation of Hepatic CYP3A.

    Science.gov (United States)

    Jonsson-Schmunk, Kristina; Wonganan, Piynauch; Choi, Jin Huk; Callahan, Shellie M; Croyle, Maria A

    2016-05-01

    Landmark studies describing the effect of microbial infection on the expression and activity of hepatic CYP3A used bacterial lipopolysaccharide as a model antigen. Our efforts to determine whether these findings were translatable to viral infections led us to observations suggesting that engagement of integrin receptors is key in the initiation of processes responsible for changes in hepatic CYP3A4 during infection and inflammation. Studies outlined in this article were designed to evaluate whether engagement of integrins, receptors commonly used by a variety of microbes to enter cellular targets, is vital in the regulation of CYP3A in the presence and absence of virus infection. Mice infected with a recombinant adenovirus (AdlacZ) experienced a 70% reduction in hepatic CYP3A catalytic activity. Infection with a mutant virus with integrin-binding arginine-glycine-aspartic acid (RGD) sequences deleted from the penton base protein of the virus capsid (AdΔRGD) did not alter CYP3A activity. CYP3A mRNA and protein levels in AdlacZ-treated animals were also suppressed, whereas those of mice given AdΔRGD were not significantly different from uninfected control mice. Silencing of the integrinβ-subunit reverted adenovirus-mediated CYP3A4 suppression in vitro. Silencing of theα-subunit did not. Suppression of integrin subunits had a profound effect on nuclear receptors pregnane X receptor and constitutive androstane receptor, whereas retinoid X receptorαwas largely unaffected. To our knowledge, this is the first time that extracellular receptors, like integrins, have been indicated in the regulation of CYP3A. This finding has several implications owing to the important role of integrins in normal physiologic process and in many disease states. PMID:26868618

  5. Phosphorylated tau/amyloid beta 1-42 ratio in ventricular cerebrospinal fluid reflects outcome in idiopathic normal pressure hydrocephalus

    Directory of Open Access Journals (Sweden)

    Patel Sunil

    2012-03-01

    Full Text Available Abstract Background Idiopathic normal pressure hydrocephalus (iNPH is a potentially reversible cause of dementia and gait disturbance that is typically treated by operative placement of a ventriculoperitoneal shunt. The outcome from shunting is variable, and some evidence suggests that the presence of comorbid Alzheimer's disease (AD may impact shunt outcome. Evidence also suggests that AD biomarkers in cerebrospinal fluid (CSF may predict the presence of AD. The aim of this study was to investigate the relationship between the phosphorylated tau/amyloid beta 1-42 (ptau/Aβ1-42 ratio in ventricular CSF and shunt outcome in patients with iNPH. Methods We conducted a prospective trial with a cohort of 39 patients with suspected iNPH. Patients were clinically and psychometrically assessed prior to and approximately 4 months after ventriculoperitoneal shunting. Lumbar and ventricular CSF obtained intraoperatively, and tissue from intraoperative cortical biopsies were analyzed for AD biomarkers. Outcome measures included performance on clinical symptom scales, supplementary gait measures, and standard psychometric tests. We investigated relationships between the ptau/Aβ1-42 ratio in ventricular CSF and cortical AD pathology, initial clinical features, shunt outcome, and lumbar CSF ptau/Aβ1-42 ratios in the patients in our cohort. Results We found that high ptau/Aβ1-42 ratios in ventricular CSF correlated with the presence of cortical AD pathology. At baseline, iNPH patients with ratio values most suggestive of AD presented with better gait performance but poorer cognitive performance. Patients with high ptau/Aβ1-42 ratios also showed a less robust response to shunting on both gait and cognitive measures. Finally, in a subset of 18 patients who also underwent lumbar puncture, ventricular CSF ratios were significantly correlated with lumbar CSF ratios. Conclusions Levels of AD biomarkers in CSF correlate with the presence of cortical AD pathology

  6. ADAM2 interactions with mouse eggs and cell lines expressing α4/α9 (ITGA4/ITGA9 integrins: implications for integrin-based adhesion and fertilization.

    Directory of Open Access Journals (Sweden)

    Ulyana V Desiderio

    Full Text Available BACKGROUND: Integrins are heterodimeric cell adhesion molecules, with 18 α (ITGA and eight β (ITGB subunits forming 24 heterodimers classified into five families. Certain integrins, especially the α(4/α(9 (ITGA4/ITGA9 family, interact with members of the ADAM (a disintegrin and metalloprotease family. ADAM2 is among the better characterized and also of interest because of its role in sperm function. Having shown that ITGA9 on mouse eggs participates in mouse sperm-egg interactions, we sought to characterize ITGA4/ITGA9-ADAM2 interactions. METHODOLOGY/PRINCIPAL FINDINGS: An anti-β(1/ITGB1 function-blocking antibody that reduces sperm-egg binding significantly inhibited ADAM2 binding to mouse eggs. Analysis of integrin subunit expression indicates that mouse eggs could express at least ten different integrins, five in the RGD-binding family, two in the laminin-binding family, two in the collagen-binding family, and ITGA9-ITGB1. Adhesion assays to characterize ADAM2 interactions with ITGA4/ITGA9 family members produced the surprising result that RPMI 8866 cell adhesion to ADAM2 was inhibited by an anti-ITGA9 antibody, noteworthy because ITGA9 has only been reported to dimerize with ITGB1, and RPMI 8866 cells lack detectable ITGB1. Antibody and siRNA studies demonstrate that ITGB7 is the β subunit contributing to RPMI 8866 adhesion to ADAM2. CONCLUSIONS/SIGNIFICANCE: These data indicate that a novel integrin α-β combination, ITGA9-ITGB7 (α(9β(7, in RPMI 8866 cells functions as a binding partner for ADAM2. ITGA9 had previously only been reported to dimerize with ITGB1. Although ITGA9-ITGB7 is unlikely to be a widely expressed integrin and appears to be the result of "compensatory dimerization" occurring in the context of little/no ITGB1 expression, the data indicate that ITGA9-ITGB7 functions as an ADAM binding partner in certain cellular contexts, with implications for mammalian fertilization and integrin function.

  7. Crossroads of integrins and cadherins in epithelia and stroma remodeling.

    Science.gov (United States)

    Epifano, Carolina; Perez-Moreno, Mirna

    2012-01-01

    Adhesion events mediated by cadherin and integrin adhesion receptors have fundamental roles in the maintenance of the physiological balance of epithelial tissues, and it is well established that perturbations in their normal functional activity and/or changes in their expression are associated with tumorigenesis. Over the last decades, increasing evidence of a dynamic collaborative interaction between these complexes through their shared interactions with cytoskeletal proteins and common signaling pathways has emerged not only as an important regulator of several aspects of epithelial cell behavior, but also as a coordinated adhesion module that senses and transmits signals from and to the epithelia surrounding microenvironment. The tight regulation of their crosstalk is particularly important during epithelial remodeling events that normally take place during morphogenesis and tissue repair, and when defective it leads to cell transformation and aggravated responses of the tumor microenvironment that contribute to tumorigenesis. In this review we highlight some of the interactions that regulate their crosstalk and how this could be implicated in regulating signals across epithelial tissues to sustain homeostasis. PMID:22568988

  8. Molecular physiology of the tensin brotherhood of integrin adaptor proteins.

    Science.gov (United States)

    Haynie, Donald T

    2014-07-01

    Numerous proteins have been identified as constituents of the adhesome, the totality of molecular components in the supramolecular assemblies known as focal adhesions, fibrillar adhesions and other kinds of adhesive contact. The transmembrane receptor proteins called integrins are pivotal adhesome members, providing a physical link between the extracellular matrix (ECM) and the actin cytoskeleton. Tensins are ever more widely investigated intracellular adhesome constituents. Involved in cell attachment and migration, cytoskeleton reorganization, signal transduction and other processes relevant to cancer research, tensins have recently been linked to functional properties of deleted in liver cancer 1 (DLC1) and a mitogen-activated protein kinases (MAPK), to cell migration in breast cancer, and to metastasis suppression in the kidney. Tensins are close relatives of phosphatase homolog/tensin homolog (PTEN), an extensively studied tumor suppressor. Such findings are recasting the earlier vision of tensin (TNS) as an actin-filament (F-actin) capping protein in a different light. This critical review aims to summarize current knowledge on tensins and thus to highlight key points concerning the expression, structure, function, and evolution of the various members of the TNS brotherhood. Insight is sought by comparisons with homologous proteins. Some historical points are added for perspective. PMID:24634006

  9. Structural basis of substrate discrimination and integrin binding by autotaxin

    Energy Technology Data Exchange (ETDEWEB)

    Hausmann, Jens; Kamtekar, Satwik; Christodoulou, Evangelos; Day, Jacqueline E.; Wu, Tao; Fulkerson, Zachary; Albers, Harald M.H.G.; van Meeteren, Laurens A.; Houben, Anna J.S.; van Zeijl, Leonie; Jansen, Silvia; Andries, Maria; Hall, Troii; Pegg, Lyle E.; Benson, Timothy E.; Kasiem, Mobien; Harlos, Karl; Vander Kooi, Craig W.; Smyth, Susan S.; Ovaa, Huib; Bollen, Mathieu; Morris, Andrew J.; Moolenaar, Wouter H.; Perrakis, Anastassis (Pfizer); (Leuven); (Oxford); (NCI-Netherlands); (Kentucky)

    2013-09-25

    Autotaxin (ATX, also known as ectonucleotide pyrophosphatase/phosphodiesterase-2, ENPP2) is a secreted lysophospholipase D that generates the lipid mediator lysophosphatidic acid (LPA), a mitogen and chemoattractant for many cell types. ATX-LPA signaling is involved in various pathologies including tumor progression and inflammation. However, the molecular basis of substrate recognition and catalysis by ATX and the mechanism by which it interacts with target cells are unclear. Here, we present the crystal structure of ATX, alone and in complex with a small-molecule inhibitor. We have identified a hydrophobic lipid-binding pocket and mapped key residues for catalysis and selection between nucleotide and phospholipid substrates. We have shown that ATX interacts with cell-surface integrins through its N-terminal somatomedin B-like domains, using an atypical mechanism. Our results define determinants of substrate discrimination by the ENPP family, suggest how ATX promotes localized LPA signaling and suggest new approaches for targeting ATX with small-molecule therapeutic agents.

  10. Avβ3 integrin: Pathogenetic role in osteotropic tumors.

    Science.gov (United States)

    Stucci, Stefania; Tucci, Marco; Passarelli, Anna; Silvestris, Franco

    2015-10-01

    The interplay of cancer cells and accessory cells within the microenvironment drives signals regulating the proliferation, migration and skeleton colonization. Osteotropism of tumor cells depends on chemokine activation, production of soluble factors and defective gene expression that cooperate within the metastatic niche to the bone resorbing functions of osteoclasts. Adhesion of cancer cells to the extracellular matrix is regulated by integrins as αvβ3 that enhances their invasiveness, pro-tumor angiogenesis and skeleton invasion. Therefore, αvβ3 signaling is implicated in enhancing osteotropism of breast and prostate cancers as well as of multiple myeloma. Targeting of αvβ3 has been adopted to restrain the tumor progression in several cancer models leading to improvement of overall survival as effect of the reduction of both tumor burden and osteotropism by malignant cells. Here, we review both the role of αvβ3 in malignant osteoclastogenesis and its potential targeting to restrain the bone colonization by skeleton invading cancers. PMID:26126493

  11. Expression of transforming growth factor-beta 1 in dystrophic patient muscles correlates with fibrosis. Pathogenetic role of a fibrogenic cytokine.

    OpenAIRE

    Bernasconi, P.; Torchiana, E; Confalonieri, P; Brugnoni, R; Barresi, R.; Mora, M.; Cornelio, F; Morandi, L.; Mantegazza, R.

    1995-01-01

    Duchenne muscular dystrophy is a fatal disorder characterized by progressive muscular weakness, wasting, and severe muscle contractures in later disease stages. Muscle biopsy reveals conspicuous myofiber degeneration and fibrosis substituting muscle tissue. We quantitatively determined mRNA of the potent fibrogenic cytokine transforming growth factor-beta 1 by quantitative PCR in 15 Duchenne muscular dystrophy, 13 Becker muscular dystrophy, 11 spinal muscular atrophy patients, and 16 controls...

  12. Contribution of beta 1- and beta 2-adrenoceptors of human atrium and ventricle to the effects of noradrenaline and adrenaline as assessed with (-)-atenolol.

    OpenAIRE

    Lemoine, H.; Schönell, H.; Kaumann, A. J.

    1988-01-01

    1. (-)-Atenolol was used as a tool to assess the function of beta 1- and beta 2-adrenoceptors in human heart. Right atrial and left ventricular preparations from patients undergoing open heart surgery were set up to contract isometrically. Membrane particles were prepared for beta-adrenoceptor labelling with [3H]-(-)-bupranolol and adenylate cyclase assays. 2. The positive inotropic effects of (-)-noradrenaline were antagonized to a similar extent by (-)-atenolol in atrial and ventricular pre...

  13. Studies of associations between the Arg389Gly polymorphism of the beta1-adrenergic receptor gene (ADRB1) and hypertension and obesity in 7677 Danish white subjects

    DEFF Research Database (Denmark)

    Gjesing, A P; Andersen, G; Albrechtsen, A; Glümer, C; Borch-Johnsen, K; Jørgensen, T; Hansen, T; Pedersen, O

    2007-01-01

    Activation of the beta(1)-adrenergic receptor (ADRB1) causes increased lipolysis in adipose tissue and enhances cardiac output. Analysis of the association of the functional ADRB1 Arg389Gly variant with obesity and hypertension has given ambiguous results. To clarify the potential impact of this...... variant on obesity and hypertension in the general population, we examined the Arg389Gly variant in a relatively large-scale population-based study....

  14. A pharmacodynamic study on clenbuterol-induced toxicity: beta1- and beta2-adrenoceptors involvement in guinea-pig tachycardia in an in vitro model.

    Science.gov (United States)

    Mazzanti, Gabriela; Di Sotto, Antonella; Daniele, Claudia; Battinelli, Lucia; Brambilla, Gianfranco; Fiori, Maurizio; Loizzo, Stefano; Loizzo, Alberto

    2007-09-01

    Beta(2)-receptor adrenergic agonists as clenbuterol and analogues are illegally used as growth promoters in cattle, in Europe, as well as in other countries. Following consumption of meat or liver, intoxication cases were described, and cardiovascular toxic effects (tachycardia, hypertension) were of clinical relevance. Therefore, we investigated whether heart rate increase induced by clenbuterol could depend upon stimulation of beta(1)- and/or beta(2)-adrenergic receptors, and in which ratio. We used in vitro guinea-pig atria, a model in which beta(1)-/beta(2)-receptors ratio is similar to that found in men. In our experiments both beta(1)- and beta(2)-receptors contributed to clenbuterol-induced heart rate increase, but with a different potency. The selective beta(2)-antagonist ICI-118,551 competitively antagonized responses to clenbuterol with high affinity (pA(2) 9.47+/-0.28, SchildSlope 0.98+/-0.20 not significantly different from unity, K(B) 0.34 nM). The selective beta(1)-antagonist atenolol antagonized clenbuterol with a relatively lower affinity (pA(2)=7.59+/-0.14), the SchildSlope=1.97+/-0.33 was significantly different from unity (Pclenbuterol stimulates guinea-pig heart rate by acting chiefly on beta(2)-adrenoceptor, although responses to clenbuterol apparently are mediated by an inter-play between both beta-adrenoceptors. Further experiments are necessary to understand which beta-adrenergic antagonists are of effectiveness to counteract cardiovascular effects in case of intoxication following clenbuterol, or other beta-adrenergic stimulants. PMID:17449161

  15. Integrin-linked kinase (ILK) modulates wound healing through regulation of hepatocyte growth factor (HGF)

    Energy Technology Data Exchange (ETDEWEB)

    Serrano, Isabel; Diez-Marques, Maria L.; Rodriguez-Puyol, Manuel [Department of Physiology, University of Alcala, Alcala de Henares, Madrid (Spain); Red de Investigacion Renal Cooperativa (RedinRen) (Spain); Instituto Reina Sofia de Investigacion Nefrologica (Spain); Herrero-Fresneda, Inmaculada [Nephrology Unit, IDIBELL, Hospital de Bellvitge, Barcelona (Spain); Red de Investigacion Renal Cooperativa (RedinRen) (Spain); Garcia del Moral, Raimundo [Department of Pathology, University of Granada (Spain); Red de Investigacion Renal Cooperativa (RedinRen) (Spain); Dedhar, Shoukat [Department of Integrative Oncology, BC Cancer Research Center, Vancouver, BC (Canada); Ruiz-Torres, Maria P., E-mail: mpiedad.ruiz@uah.es [Department of Physiology, University of Alcala, Alcala de Henares, Madrid (Spain); Red de Investigacion Renal Cooperativa (RedinRen) (Spain); Instituto Reina Sofia de Investigacion Nefrologica (Spain); Rodriguez-Puyol, Diego [Nephrology Unit, Hospital Universitario Principe de Asturias, Alcala de Henares, Madrid (Spain); Red de Investigacion Renal Cooperativa (RedinRen) (Spain); Instituto Reina Sofia de Investigacion Nefrologica (Spain)

    2012-11-15

    Integrin-linked kinase (ILK) is an intracellular effector of cell-matrix interactions and regulates many cellular processes, including growth, proliferation, survival, differentiation, migration, invasion and angiogenesis. The present work analyzes the role of ILK in wound healing in adult animals using a conditional knock-out of the ILK gene generated with the tamoxifen-inducible Cre-lox system (CRE-LOX mice). Results show that ILK deficiency leads to retarded wound closure in skin. Intracellular mechanisms involved in this process were analyzed in cultured mouse embryonic fibroblast (MEF) isolated from CRE-LOX mice and revealed that wounding promotes rapid activation of phosphatidylinositol 3-kinase (PI3K) and ILK. Knockdown of ILK resulted in a retarded wound closure due to a decrease in cellular proliferation and loss of HGF protein expression during the healing process, in vitro and in vivo. Alterations in cell proliferation and wound closure in ILK-deficient MEF or mice could be rescued by exogenous administration of human HGF. These data demonstrate, for the first time, that the activation of PI3K and ILK after skin wounding are critical for HGF-dependent tissue repair and wound healing. -- Highlights: Black-Right-Pointing-Pointer ILK deletion results in decreased HGF expression and delayed scratch wound repair. Black-Right-Pointing-Pointer PI3K/ILK/AKT pathway signals through HGF to regulate wound healing. Black-Right-Pointing-Pointer An ILK-dependent increase in HGF expression is responsible for wound healing in vivo. Black-Right-Pointing-Pointer ILK-KO mice are used to confirm the requirement for ILK function in wound healing. Black-Right-Pointing-Pointer Human HGF treatment restores delayed wound closure in vitro and in vivo.

  16. Integrin-linked kinase (ILK) modulates wound healing through regulation of hepatocyte growth factor (HGF)

    International Nuclear Information System (INIS)

    Integrin-linked kinase (ILK) is an intracellular effector of cell–matrix interactions and regulates many cellular processes, including growth, proliferation, survival, differentiation, migration, invasion and angiogenesis. The present work analyzes the role of ILK in wound healing in adult animals using a conditional knock-out of the ILK gene generated with the tamoxifen-inducible Cre-lox system (CRE-LOX mice). Results show that ILK deficiency leads to retarded wound closure in skin. Intracellular mechanisms involved in this process were analyzed in cultured mouse embryonic fibroblast (MEF) isolated from CRE-LOX mice and revealed that wounding promotes rapid activation of phosphatidylinositol 3-kinase (PI3K) and ILK. Knockdown of ILK resulted in a retarded wound closure due to a decrease in cellular proliferation and loss of HGF protein expression during the healing process, in vitro and in vivo. Alterations in cell proliferation and wound closure in ILK-deficient MEF or mice could be rescued by exogenous administration of human HGF. These data demonstrate, for the first time, that the activation of PI3K and ILK after skin wounding are critical for HGF-dependent tissue repair and wound healing. -- Highlights: ► ILK deletion results in decreased HGF expression and delayed scratch wound repair. ► PI3K/ILK/AKT pathway signals through HGF to regulate wound healing. ► An ILK-dependent increase in HGF expression is responsible for wound healing in vivo. ► ILK-KO mice are used to confirm the requirement for ILK function in wound healing. ► Human HGF treatment restores delayed wound closure in vitro and in vivo.

  17. Preferential reduction of binding of 125I-iodopindolol to beta-1 adrenoceptors in the amygdala of rat after antidepressant treatments

    International Nuclear Information System (INIS)

    This study utilized quantitative receptor autoradiography to examine the effects of repeated administration of antidepressants to rats on the binding of the beta adrenoceptor antagonist, 125I-iodopindolol (125I-IPIN) to either beta-1 or beta-2 adrenoceptors in various regions of brain. Antidepressants were selected to represent various chemical and pharmacological classes including tricyclic compounds (desipramine and protriptyline), monoamine oxidase inhibitors (clorgyline, phenelzine and tranylcypromine), atypical antidepressants (mianserin and trazodone) and selective inhibitors of the uptake of serotonin (citalopram and sertraline). Additionally, rats were treated with various psychotropic drugs that lack antidepressant efficacy (cocaine, deprenyl, diazepam and haloperidol). Repeated treatment of rats with desipramine, protriptyline, clorgyline, phenelzine, tranylcypromine or mianserin reduced the binding of 125I-IPIN to beta-1 adrenoceptors in many brain areas. Only in the basolateral and lateral nuclei of the amygdala did all six of these antidepressants significantly reduce 125I-IPIN binding to beta-1 adrenoceptors. In these amygdaloid nuclei, the magnitude of the reduction in the binding of 125I-IPIN caused by each of these drugs was comparable to or greater than the reduction in binding produced in any other region of brain. Reductions of binding of 125I-IPIN after antidepressant treatments were not consistently observed in the cortex, the area of brain examined most often in homogenate binding studies. Only the monoamine oxidase inhibitors caused reductions in the binding of 125I-IPIN to beta-2 adrenoceptors, and this effect was generally localized to the amygdala and hypothalamus

  18. Characterization of a Thermobifida fusca beta-1,3-glucanase (Lam81A) with a potential role in plant biomass degradation.

    Science.gov (United States)

    McGrath, Colleen E; Wilson, David B

    2006-11-28

    Thermobifida fusca is a filamentous soil bacterium that plays a major role in the breakdown of plant biomass. In this paper, we report the cloning, expression, purification, and characterization of the T. fusca enzyme, Lam81A. The Carbohydrate Active Enzymes Database (http://afmb.cnrs-mrs.fr/CAZY/) indicates that Lam81A belongs to a relatively uncharacterized family of beta-1,3-glucanases, family GH-81 [Coutinho, P. M., and Henrissat, B. (1999) in Recent Advances in Carbohydrate Bioengineering (Gilbert, H. J., Davies, G., Henrissat, B., and Svensson, B., Eds.) pp 3-12, The Royal Society of Chemistry, Cambridge, U.K.]. Microarray analysis suggests that Lam81A plays a role in biomass degradation, where its natural substrate may be the plant cell wall polysaccharide, callose, which is a polymer of beta-1,3-linked glucose. Characterization of Lam81A has shown that the enzyme is specific for beta-1,3-linked glucose polysaccharides, is endohydrolytic, and utilizes an inverting mechanism for substrate hydrolysis. In addition, the enzyme has a broad pH optimum from 5.5 to 10, a temperature optimum of 50 degrees C, and demonstrates substrate inhibition, as well as showing a high basal level of expression. PMID:17115704

  19. Anti-acetylcholinesterase and Antioxidant Activities of Inhaled Juniper Oil on Amyloid Beta (1-42)-Induced Oxidative Stress in the Rat Hippocampus.

    Science.gov (United States)

    Cioanca, Oana; Hancianu, Monica; Mihasan, Marius; Hritcu, Lucian

    2015-05-01

    Juniper volatile oil is extracted from Juniperus communis L., of the Cupressaceae family, also known as common juniper. Also, in aromatherapy the juniper volatile oil is used against anxiety, nervous tension and stress-related conditions. In the present study, we identified the effects of the juniper volatile oil on amyloid beta (1-42)-induced oxidative stress in the rat hippocampus. Rats received a single intracerebroventricular injection of amyloid beta (1-42) (400 pmol/rat) and then were exposed to juniper volatile oil (200 μl, either 1 or 3 %) for controlled 60 min period, daily, for 21 continuous days. Also, the antioxidant activity in the hippocampus was assessed using superoxide dismutase, glutathione peroxidase and catalase specific activities, the total content of the reduced glutathione, protein carbonyl and malondialdehyde levels. Additionally, the acetylcholinesterase activity in the hippocampus was assessed. The amyloid beta (1-42)-treated rats exhibited the following: increase of the acetylcholinesterase, superoxide dismutase and catalase specific activities, decrease of glutathione peroxidase specific activity and the total content of the reduced glutathione along with an elevation of malondialdehyde and protein carbonyl levels. Inhalation of the juniper volatile oil significantly decreases the acetylcholinesterase activity and exhibited antioxidant potential. These findings suggest that the juniper volatile oil may be a potential candidate for the development of therapeutic agents to manage oxidative stress associated with Alzheimer's disease through decreasing the activity of acetylcholinesterase and anti-oxidative mechanism. PMID:25743585

  20. Integrin α4 Induces Lymphangiogenesis and Metastasis via Upregulation of VEGF-C in Human Colon Cancer.

    Science.gov (United States)

    Lv, Xiao-Hong; Liu, Bao-Quan; Li, Xue-Mei; Wang, Xiang-Chen; Li, Xin-Lei; Ahmed, Naila; Zhang, Ya-Fang

    2016-06-01

    Vascular endothelial growth factor-C (VEGF-C) is a key regulator in lymphangiogenesis, and is overexpressed in various malignancies. Integrin α4β1, a new member of the VEGF-C/VEGF receptor pathway, was found to be overexpressed in melanoma tumors. However, little is known regarding the potential role of integrin α4β1 in lymphangiogenesis and other solid tumors. The aim of this study was to investigate the expression patterns of integrin α4 and VEGF-C in relation to lymphangiogenesis and clinicopathological parameters in human colon cancer. The expression of integrin α4, VEGF-C, and VEGFR-3 was assessed in 71 human colon cancer tissues and 30 paracancerous normal tissues by immunohistochemical staining. Lymphatic microvessel density (LMVD) was measured after D2-40-labeling, and the correlations among different factors were statistically analyzed. The expression of integrin α4, VEGF-C, VEGFR-3, and LMVD was higher in colon cancer tissues compared with the normal paracancerous colon tissues. There was a positive correlation between the expression of integrin α4 and VEGF-C. Integrin α4 and VEGF-C were significantly associated with the clinicopathological parameters (LMVD, Duke's stage, and lymph node metastasis). Kaplan-Meier analyses indicated that patients with high integrin α4 or VEGF-C expression had significantly shorter overall survival and tumor-free survival time. Multivariate analyses suggested that integrin α4 and VEGF-C may serve as independent prognostic factors for human colon cancer. Both integrin α4 and VEGF-C are involved in lymphangiogenesis and lymphatic metastasis. Our results demonstrated that integrin α4 is a novel prognostic indicator for human colon cancer. Anat Rec, 299:741-747, 2016. © 2016 Wiley Periodicals, Inc. PMID:26917449

  1. IGF-IR promotes prostate cancer growth by stabilizing α5β1 integrin protein levels.

    Directory of Open Access Journals (Sweden)

    Aejaz Sayeed

    Full Text Available Dynamic crosstalk between growth factor receptors, cell adhesion molecules and extracellular matrix is essential for cancer cell migration and invasion. Integrins are transmembrane receptors that bind extracellular matrix proteins and enable cell adhesion and cytoskeletal organization. They also mediate signal transduction to regulate cell proliferation and survival. The type 1 insulin-like growth factor receptor (IGF-IR mediates tumor cell growth, adhesion and inhibition of apoptosis in several types of cancer. We have previously demonstrated that β1 integrins regulate anchorage-independent growth of prostate cancer (PrCa cells by regulating IGF-IR expression and androgen receptor-mediated transcriptional functions. Furthermore, we have recently reported that IGF-IR regulates the expression of β1 integrins in PrCa cells. We have dissected the mechanism through which IGF-IR regulates β1 integrin expression in PrCa. Here we report that IGF-IR is crucial for PrCa cell growth and that β1 integrins contribute to the regulation of proliferation by IGF-IR. We demonstrate that β1 integrin regulation by IGF-IR does not occur at the mRNA level. Exogenous expression of a CD4 - β1 integrin cytoplasmic domain chimera does not interfere with such regulation and fails to stabilize β1 integrin expression in the absence of IGF-IR. This appears to be due to the lack of interaction between the β1 cytoplasmic domain and IGF-IR. We demonstrate that IGF-IR stabilizes the β1 subunit by protecting it from proteasomal degradation. The α5 subunit, one of the binding partners of β1, is also downregulated along with β1 upon IGF-IR knockdown while no change is observed in the expression of the α2, α3, α4, α6 and α7 subunits. Our results reveal a crucial mechanistic role for the α5β1 integrin, downstream of IGF-IR, in regulating cancer growth.

  2. Changes in membrane sphingolipid composition modulate dynamics and adhesion of integrin nanoclusters.

    Science.gov (United States)

    Eich, Christina; Manzo, Carlo; de Keijzer, Sandra; Bakker, Gert-Jan; Reinieren-Beeren, Inge; García-Parajo, Maria F; Cambi, Alessandra

    2016-01-01

    Sphingolipids are essential constituents of the plasma membrane (PM) and play an important role in signal transduction by modulating clustering and dynamics of membrane receptors. Changes in lipid composition are therefore likely to influence receptor organisation and function, but how this precisely occurs is difficult to address given the intricacy of the PM lipid-network. Here, we combined biochemical assays and single molecule dynamic approaches to demonstrate that the local lipid environment regulates adhesion of integrin receptors by impacting on their lateral mobility. Induction of sphingomyelinase (SMase) activity reduced sphingomyelin (SM) levels by conversion to ceramide (Cer), resulting in impaired integrin adhesion and reduced integrin mobility. Dual-colour imaging of cortical actin in combination with single molecule tracking of integrins showed that this reduced mobility results from increased coupling to the actin cytoskeleton brought about by Cer formation. As such, our data emphasizes a critical role for the PM local lipid composition in regulating the lateral mobility of integrins and their ability to dynamically increase receptor density for efficient ligand binding in the process of cell adhesion. PMID:26869100

  3. Functional analysis of the putative integrin recognition motif on adeno-associated virus 9.

    Science.gov (United States)

    Shen, Shen; Berry, Garrett E; Castellanos Rivera, Ruth M; Cheung, Roland Y; Troupes, Andrew N; Brown, Sarah M; Kafri, Tal; Asokan, Aravind

    2015-01-16

    Adeno-associated viruses (AAVs) display a highly conserved NGR motif on the capsid surface. Earlier studies have established this tripeptide motif as being essential for integrin-mediated uptake of recombinant AAV serotype 2 (AAV2) in cultured cells. However, functional attributes of this putative integrin recognition motif in other recombinant AAV serotypes displaying systemic transduction in vivo remain unknown. In this study, we dissect the biology of an integrin domain capsid mutant derived from the human isolate AAV9 in mice. The AAV9/NGA mutant shows decreased systemic transduction in mice. This defective phenotype was accompanied by rapid clearance of mutant virions from the blood circulation and nonspecific sequestration by the spleen. Transient vascular hyperpermeability, induced by histamine coinjection, exacerbated AAV9/NGA uptake by the spleen but not the liver. However, such treatment did not affect AAV9 virions, suggesting a potential entry/post-entry defect for the mutant in different tissues. Further characterization revealed modestly decreased cell surface binding but a more pronounced defect in the cellular entry of mutant virions. These findings were corroborated by the observation that blocking multiple integrins adversely affected recombinant AAV9 transduction in different cell types, albeit with variable efficiencies. From a structural perspective, we observed that the integrin recognition motif is located in close proximity to the galactose binding footprint on AAV9 capsids and postulate that this feature could influence cell surface attachment, cellular uptake at the tissue level, and systemic clearance by the reticuloendothelial system. PMID:25404742

  4. Localized LoxL3-Dependent Fibronectin Oxidation Regulates Myofiber Stretch and Integrin-Mediated Adhesion.

    Science.gov (United States)

    Kraft-Sheleg, Ortal; Zaffryar-Eilot, Shelly; Genin, Olga; Yaseen, Wesal; Soueid-Baumgarten, Sharon; Kessler, Ofra; Smolkin, Tatyana; Akiri, Gal; Neufeld, Gera; Cinnamon, Yuval; Hasson, Peleg

    2016-03-01

    For muscles to function, myofibers have to stretch and anchor at the myotendinous junction (MTJ), a region rich in extracellular matrix (ECM). Integrin signaling is required for MTJ formation, and mutations affecting the cascade lead to muscular dystrophies in mice and humans. Underlying mechanisms for integrin activation at the MTJ and ECM modifications regulating its signaling are unclear. We show that lysyl oxidase-like 3 (LoxL3) is a key regulator of integrin signaling that ensures localized control of the cascade. In LoxL3 mutants, myofibers anchor prematurely or overshoot to adjacent somites, and are loose and lack tension. We find that LoxL3 complexes with and directly oxidizes Fibronectin (FN), an ECM scaffold protein and integrin ligand enriched at the MTJ. We identify a mechanism whereby localized LoxL3 secretion from myofiber termini oxidizes FN, enabling enhanced integrin activation at the tips of myofibers and ensuring correct positioning and anchoring of myofibers along the MTJ. PMID:26954549

  5. Modulation of radiation-induced oral mucositis (mouse) by selective inhibition of β1 integrin

    International Nuclear Information System (INIS)

    Introduction: Oral mucositis is a severe side effect of radio(chemo)therapy for head and neck tumors, for which β1 integrins have been proposed as potential therapeutic targets. The present study was initiated to determine the effect of selective inhibition of β1 integrin on the oral epithelial radiation response. Materials and methods: Daily fractionated irradiation was given with 5 × 3 Gy/week over 1 or 2 weeks with/without the β1 integrin-inhibiting monoclonal antibody AIIB2 or an IgG control. Each protocol was terminated by graded test doses to generate full dose–effect curves for mucosal ulceration. The same technique was used for single dose irradiation. Results: Combined single dose irradiation plus AIIB2 resulted in a significant decrease of the ED50 compared to irradiation alone or control IgG. No effect of AIIB2 was found with fractionated irradiation over 1 week. With 2 weeks of fractionation, AIIB2 induced a significant increase in the ED50 for the terminating test irradiation when administered in week 2. The time course of the response was largely unaffected by β1 integrin inhibition. Conclusions: A reduction of mucosal reactions by β1 integrin inhibition later in a course of fractionation was observed, i.e. when epithelial repopulation processes were active. Further mechanistic studies are required.

  6. The cytoplasmic extension of the integrin β6 subunit regulates epithelial-to-mesenchymal transition.

    Science.gov (United States)

    Lee, Carlin; Lee, Casey; Lee, Stacey; Siu, Amanda; Ramos, Daniel M

    2014-02-01

    Prognosis for oral cancer patients has not improved in over 60 years due to invasion and recurrence. To understand the invasive behavior of this tumor, we evaluated the role of the αvβ6 integrin. Invasive oral SCC cells express the αvβ6 integrin, which contains an 11-amino-acid extension on its β-subunit unique to the integrin family. We determined that this β6 cytoplasmic extension regulates the composition of the intermediate filament network and the organization of signaling structures called focal contacts. The auto-phosphorylation of FAK, which is localized to focal contacts, was also regulated by the β6-cytoplasmic tail, as were the transcription factors Notch and STAT3. Lastly, we also determined that activation of MAPK required the full-length β6 integrin. Together these results indicate that the signaling critical to epithelial-to-mesenchymal transition (EMT) is regulated by the β6 integrin cytoplasmic domain. PMID:24510996

  7. Antimicrobial actions of the human epididymis 2 (HE2 protein isoforms, HE2alpha, HE2beta1 and HE2beta2

    Directory of Open Access Journals (Sweden)

    French Frank S

    2004-08-01

    Full Text Available Abstract Background The HE2 gene encodes a group of isoforms with similarities to the antimicrobial beta-defensins. We demonstrated earlier that the antimicrobial activity of HE2 proteins and peptides is salt resistant and structure dependent and involves permeabilization of bacterial membranes. In this study, we further characterize the antimicrobial properties of HE2 peptides in terms of the structural changes induced in E. coli and the inhibition of macromolecular synthesis. Methods E. coli treated with 50 micro g/ml of HE2alpha, HE2beta1 or HE2beta2 peptides for 30 and 60 min were visualized using transmission and scanning electron microscopy to investigate the impact of these peptides on bacterial internal and external structure. The effects of HE2alpha, HE2beta1 and HE2beta2 on E. coli macromolecular synthesis was assayed by incubating the bacteria with 2, 10 and 25 micro g/ml of the individual peptides for 0–60 min and measuring the incorporation of the radioactive precursors [methyl-3H]thymidine, [5-3H]uridine and L-[4,5-3H(N]leucine into DNA, RNA and protein. Statistical analyses using Student's t-test were performed using Sigma Plot software. Values shown are Mean ± S.D. Results E. coli treated with HE2alpha, HE2beta1 and HE2beta2 peptides as visualized by transmission electron microscopy showed extensive damage characterized by membrane blebbing, thickening of the membrane, highly granulated cytoplasm and appearance of vacuoles in contrast to the smooth and continuous membrane structure of the untreated bacteria. Similarly, bacteria observed by scanning electron microscopy after treating with HE2alpha, HE2beta1 or HE2beta2 peptides exhibited membrane blebbing and wrinkling, leakage of cellular contents, especially at the dividing septa, and external accumulation of fibrous materials. In addition, HE2alpha, HE2beta1 and HE2beta2 peptides inhibited E. coli DNA, RNA and protein synthesis. Conclusions The morphological changes observed

  8. Regulation of the expression of Gal alpha 1-3Gal beta 1-4GlcNAc glycosphingolipids in kidney.

    Science.gov (United States)

    Hendricks, S P; He, P; Stults, C L; Macher, B A

    1990-10-15

    Previous studies (Galili, U., Clark, M. R., Shohet, S. B., Buehler, J., and Macher, B. A. (1987) Proc. Natl. Acad. Sci. U. S. A. 84, 1369-1373; Galili, U., Shohet, S. B., Korbrin, E., Stults, C. L. M., and Macher, B. A. (1988) J. Biol. Chem. 263, 17755-17762) have established that there is a unique evolutionary distribution of glycoconjugates carrying the Gal alpha 1-3Gal beta 1-4GlcNAc epitope. These glycoconjugates are expressed by cells from New World monkeys and non-primate mammals, but not by cells from humans, Old World monkeys, or apes. The lack of expression of this epitope in the latter species appears to result from the suppression of gene expression for the enzyme UDP-galactose:nLc4Cer alpha 1-3-galactosyltransferase (alpha 1-3GalT) (Joziasse, D. H., Shaper, J. H., Van den Eijnden, D. H., Van Tunen, A. J., and Shaper, N. L. (1989) J. Biol. Chem. 264, 14290-14297). Although many non-primate species are known to express this carbohydrate epitope, the nature (i.e. glycoprotein or glycosphingolipid) of the glycoconjugate carrying this epitope is only known for a few tissues in a few animal species. Furthermore, it is not known whether all animal species express this epitope in the same tissues. We have investigated these questions by analyzing the glycosphingolipids in kidney from several non-primate animal species. Immunostained thin layer chromatograms of glycosphingolipids from sheep, pig, rabbit, cow, and rat kidney with the Gal alpha 1-3Gal beta 1-4GlcNAc glycosphingolipid-specific monoclonal antibody, Gal-13, demonstrated that kidney from all of these species except rat contained Gal alpha 1-3Gal beta 1-4GlcNAc neutral glycosphingolipids. A lack of expression of Gal alpha 1-3Gal beta 1-4GlcNAc glycosphingolipids in rat may be due to the lack of expression of the enzyme (alpha 1-3GalT) which catalyzes the formation of the Gal alpha 1-3Gal nonreducing terminal sequence of these compounds or to the lack of expression of glycosyltransferases which are

  9. Irradiation and various cytotoxic drugs enhance tyrosine phosphorylation and {beta}{sub 1}-integrin clustering in human A549 lung cancer cells in a substratum-dependent manner in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Cordes, N.; Beinke, C.; Beuningen, D. van [Inst. of Radiobiology, German Armed Forces, Munich (Germany); Plasswilm, L. [Dept. of Radiation Oncology, Univ. Hospital Basel (Swaziland)

    2004-03-01

    Background and purpose: interactions of cells with a substratum, especially extracellular matrix proteins, initiate clustering of integrin receptors in the cell membrane. This process represents the initial step for the activation of signaling pathways regulating survival, proliferation, differentiation, adhesion, and migration, and could, furthermore, be important for cellular resistance-mediating mechanisms against radiation or cytotoxic drugs. The lack of data elucidating the impact of irradiation or cytotoxic drugs on this important phenomenon led to this study on human A549 lung cancer cells in vitro. Material and methods: the human lung carcinoma cell line A549 grown on polystyrene or fibronectin (FN) was irradiated with 0-8 Gy or treated with cisplatin (0.1-50 {mu}M), paclitaxel (0.1-50 nM), or mitomycin (0.1-50 {mu}M). Colony formation assays, immunofluorescence staining in combination with activation of integrin clustering using anti-{beta}{sub 1}-integrin antibodies (K20), and Western blotting for tyrosine phosphorylation under treatment of cells with the IC{sub 50} for irradiation (2 Gy; IC{sub 50} = 2.2 Gy), cisplatin (2 {mu}M), paclitaxel (5 nM), or mitomycin (7 {mu}M) were performed. Results: attachment of cells to FN resulted in a significantly reduced radio- and chemosensitivity compared to polystyrene. The clustering of {beta}{sub 1}-integrins examined by immunofluorescence staining was only stimulated by irradiation, cisplatin, paclitaxel, or mitomycin in case of cell attachment to FN. By contrast, tyrosine phosphorylation, as one of the major events following {beta}{sub 1}-integrin clustering, showed a 3.7-fold, FN-related enhancement, and treatment of cells with the IC{sub 50} of radiation, cisplatin, paclitaxel, or mitomycin showed a substratum-dependent induction. Conclusion: for the first time, a strong influence of irradiation and a variety of cytotoxic drugs on the clustering of {beta}{sub 1}-integrins could be shown. This event is a

  10. Integrin β3 and LKB1 are independently involved in the inhibition of proliferation by lovastatin in human intrahepatic cholangiocarcinoma.

    Science.gov (United States)

    Yang, Sheng-Huei; Lin, Hung-Yun; Changou, Chun A; Chen, Chun-Han; Liu, Yun-Ru; Wang, Jinghan; Jiang, Xiaoqing; Luh, Frank; Yen, Yun

    2016-01-01

    Human intrahepatic cholangiocarcinomas are one of the most difficult cancers to treat. In our study, Lovastatin, a 3-hydroxy-3-methylglutaryl-coenzyme-CoA (HMG-CoA) reductase inhibitor, demonstrated anticancer properties by inhibiting cancer cell proliferation, cell migration and cell adhesion. Lovastatin inhibited the expressions of transforming growth factor (TGF)-β1, cyclooxygenase (COX)-2, and intercellular adhesion molecule (ICAM)-1. Furthermore, lovastatin inhibited the expressions of integrin β1 and integrin β3 but not integrin αv or integrin β5. While Lovastatin's inhibitory effects on TGFβ1, COX2, and ICAM-1 expression were independently controlled by the tumor suppressor LKB1, integrin β3 expression was not affected. Lovastatin's inhibitory effect on cell adhesion was associated with the decreased expression of integrin β3 and cell surface heterodimer integrin αvβ3. Quantitative real time PCR, fluorescent microscopy, and cell migration assays all confirmed that Lovastatin inhibits integrin αvβ3 downstream signaling including FAK activation, and β-catenin, vimentin, ZO-1, and β-actin. Overall, Lovastatin reduced tumor cell proliferation and migration by modifying the expression of genes involved in cell adhesion and other critical cellular processes. Our study highlights novel anti-cancer properties of Lovastatin and supports further exploration of statins in the context of cholangiocarcinoma therapy. PMID:26517522

  11. RPTP-alpha acts as a transducer of mechanical force on alphav/beta3-integrin-cytoskeleton linkages

    DEFF Research Database (Denmark)

    von Wichert, Gotz; Jiang, Guoying; Kostic, Ana;

    2003-01-01

    -integrins at the leading edge during early spreading, and coimmunoprecipitates with alphav-integrins during spreading on fibronectin and vitronectin. RPTPalpha-dependent activation of Src family kinases, in particular activation of Fyn, is required for the force-dependent formation of focal complexes and...

  12. Conditional deletion of the Itgb4 integrin gene in Schwann cells leads to delayed peripheral nerve regeneration.

    NARCIS (Netherlands)

    Zee, C.E.E.M. van der; Kreft, M.; Beckers, G.; Kuipers, A.; Sonnenberg, A.

    2008-01-01

    Several different integrins participate in the complex interactions that promote repair of the peripheral nervous system. The role of the integrin alpha6beta4 in peripheral nerve regeneration was investigated in mice by cre-mediated deletion of the Itgb4 (beta4) gene in Schwann cells. After a crush

  13. Induction of Partial Protection against Foot and Mouth Disease Virus in Guinea Pigs by Neutralization with the Integrin β6-1 Subunit

    Directory of Open Access Journals (Sweden)

    Zhidong Zhang

    2013-04-01

    Full Text Available The mechanism by which the foot-and-mouth disease virus (FMDV initiates infection of cells is thought to involve the attachment of the viral capsid to host integrins on the surface of target cells. However, the role of integrins in FMDV infection still needs to be fully understood, although it has been demonstrated that integrin αvβ6 interferes with FMDV in vitro and results in neutralization of its infectivity. In the present study, we describe the cloning and sequencing of suckling mouse integrin β6 and the subsequent expression of two segments of integrin β6 extracellular domains: β6-1 (which contains the ligand-binding domain and β6-2. Sequencing of the mouse integrin β6 subunit revealed close homology (~90% with its human counterpart. When recombinant integrin extracellular domains β6-1 and β6-2 formulated with adjuvant were inoculated into guinea pigs, anti-integrin antibody expression was high before FMDV challenge. Interestingly, guinea pigs (50% inoculated with integrin β6-1 were protected from FMDV infection; in contrast, none of the animals inoculated with integrin β6-2 were protected. This result indicates that an integrin blockade may be able to interfere with FMDV infection in vivo, which raises the possibility that targeting integrin in vivo may be the basis for a new strategy to control FMDV infection.

  14. Integrin beta3 Leu33Pro polymorphism and risk of hip fracture: 25 years follow-up of 9233 adults from the general population

    DEFF Research Database (Denmark)

    Tofteng, Charlotte L; Bach-Mortensen, Pernille; Bojesen, Stig E;

    2007-01-01

    integrin beta3 Leu33Pro polymorphism have a two-fold risk of hip fracture, mainly confined to postmenopausal women. Integrin beta3 Leu33Pro homozygosity could prove a useful marker for risk of future hip fracture and may contribute to pharmacogenetic variation in effects of integrin alphavbeta3 antagonists....

  15. A ligand-independent integrin β1 mechanosensory complex guides spindle orientation.

    Science.gov (United States)

    Petridou, Nicoletta I; Skourides, Paris A

    2016-01-01

    Control of spindle orientation is a fundamental process for embryonic development, morphogenesis and tissue homeostasis, while defects are associated with tumorigenesis and other diseases. Force sensing is one of the mechanisms through which division orientation is determined. Here we show that integrin β1 plays a critical role in this process, becoming activated at the lateral regions of the cell cortex in a ligand-independent manner. This activation is force dependent and polar, correlating with the spindle capture sites. Inhibition of integrin β1 activation on the cortex and disruption of its asymmetric distribution leads to spindle misorientation, even when cell adhesion is β1 independent. Examining downstream targets reveals that a cortical mechanosensory complex forms on active β1, and regulates spindle orientation irrespective of cell context. We propose that ligand-independent integrin β1 activation is a conserved mechanism that allows cell responses to external stimuli. PMID:26952307

  16. Heterocyclic Scaffolds in the Design of Peptidomimetic Integrin Ligands: Synthetic Strategies, Structural Aspects, and Biological Activity.

    Science.gov (United States)

    De Marco, Rossella; Mazzotti, Giacomo; Greco, Arianna; Gentilucci, Luca

    2016-01-01

    The integrin receptors represent valuable targets for therapeutic interventions; being overexpressed in many pathological states, their inhibition can be effective to treat a number of severe diseases. Since integrin functions are mediated by interactions with ECM protein ligands, the inhibition can be achieved by interfering with such interactions using small mimetics of the integrin-ligand recognition motifs (e.g. RGD, LDV, etc.). In this review, we focus on the antagonists with peptideheterocycle hybrid structures. The introduction of well-designed scaffolds has met considerable success in the rational design of highly stable, bioavailable, and conformationally defined antagonists. Two main approaches are discussed herein. The first approach is the use of scaffolds external to the main recognition motifs, aimed at improving conformational definition. In the second approach, heterocyclic cores are introduced within the recognition motifs, giving access to libraries of 3D diverse candidate antagonists. PMID:26265351

  17. Syndecans promote integrin-mediated adhesion of mesenchymal cells in two distinct pathways

    DEFF Research Database (Denmark)

    Whiteford, James; Behrends, Volker; Kirby, Hishani; Kusche-Gullberg, Marion; Muramatsu, Takashi; Couchman, John R

    2007-01-01

    Syndecans are transmembrane proteoglycans that support integrin-mediated adhesion. Well documented is the contribution of syndecan-4 that interacts through its heparan sulphate chains to promote focal adhesion formation in response to fibronectin domains. This process has requirements for integrin...... and signaling through the cytoplasmic domain of syndecan-4. Here an alternate pathway mediated by the extracellular domains of syndecans-2 and -4 is characterized that is independent of both heparan sulphate and syndecan signaling. This pathway is restricted to mesenchymal cells and was not seen in...... any epithelial cell line tested, apart from vascular endothelia. The syndecan ectodomains coated as substrates promoted integrin-dependent attachment, spreading and focal adhesion formation. Syndecan-4 null cells were competent, as were fibroblasts compromised in heparan sulphate synthesis that were...

  18. NMR structure of integrin α4 cytosolic tail and its interactions with paxillin.

    Directory of Open Access Journals (Sweden)

    Geok-Lin Chua

    Full Text Available BACKGROUND: Integrins are a group of transmembrane signaling proteins that are important in biological processes such as cell adhesion, proliferation and migration. Integrins are α/β hetero-dimers and there are 24 different integrins formed by specific combinations of 18 α and 8 β subunits in humans. Generally, each of these subunits has a large extracellular domain, a single pass transmembrane segment and a cytosolic tail (CT. CTs of integrins are important in bidirectional signal transduction and they associate with a large number of intracellular proteins. PRINCIPAL FINDINGS: Using NMR spectroscopy, we determined the 3-D structure of the full-length α4 CT (Lys968-Asp999 and characterize its interactions with the adaptor protein paxillin. The α4 CT assumes an overall helical structure with a kink in its membrane proximal region. Residues Gln981-Asn997 formed a continuous helical conformation that may be sustained by potential ionic and/or hydrogen bond interactions and packing of aromatic-aliphatic side-chains. ¹⁵N-¹H HSQC NMR experiments reveal interactions of the α4 CT C-terminal region with a fragment of paxillin (residues G139-K277 that encompassed LD2-LD4 repeats. Residues of these LD repeats including their adjoining linkers showed α4 CT binding-induced chemical shift changes. Furthermore, NMR studies using LD-containing peptides showed predominant interactions between LD3 and LD4 of paxillin and α4 CT. Docked structures of the α4 CT with these LD repeats suggest possible polar and/or salt-bridge and non-polar packing interactions. SIGNIFICANCE: The current study provides molecular insights into the structural diversity of α CTs of integrins and interactions of integrin α4 CT with the adaptor protein paxillin.

  19. Integrin-mediated cell migration is blocked by inhibitors of human neuraminidase.

    Science.gov (United States)

    Jia, Feng; Howlader, Md Amran; Cairo, Christopher W

    2016-09-01

    Integrins are critical receptors in cell migration and adhesion. A number of mechanisms are known to regulate the function of integrins, including phosphorylation, conformational change, and cytoskeletal anchoring. We investigated whether native neuraminidase (Neu, or sialidase) enzymes which modify glycolipids could play a role in regulating integrin-mediated cell migration. Using a scratch assay, we found that exogenously added Neu3 and Neu4 activity altered rates of cell migration. We observed that Neu4 increased the rate of migration in two cell lines (HeLa, A549); while Neu3 only increased migration in HeLa cells. A bacterial neuraminidase was able to increase the rate of migration in HeLa, but not in A549 cells. Treatment of cells with complex gangliosides (GM1, GD1a, GD1b, and GT1b) resulted in decreased cell migration rates, while LacCer was able to increase rates of migration in both lines. Importantly, our results show that treatment of cells with inhibitors of native Neu enzymes had a dramatic effect on the rates of cell migration. The most potent compound tested targeted the human Neu4 isoenzyme, and was able to substantially reduce the rate of cell migration. We found that the lateral mobility of integrins was reduced by treatment of cells with Neu3, suggesting that Neu3 enzyme activity resulted in changes to integrin-co-receptor or integrin-cytoskeleton interactions. Finally, our results support the hypothesis that inhibitors of human Neu can be used to investigate mechanisms of cell migration and for the development of anti-adhesive therapies. PMID:27344026

  20. Quantitative Proteomics Reveals β2 Integrin-mediated Cytoskeletal Rearrangement in Vascular Endothelial Growth Factor (VEGF)-induced Retinal Vascular Hyperpermeability.

    Science.gov (United States)

    Jo, Dong Hyun; Bae, Jingi; Chae, Sehyun; Kim, Jin Hyoung; Han, Jong-Hee; Hwang, Daehee; Lee, Sang-Won; Kim, Jeong Hun

    2016-05-01

    Retinal vascular hyperpermeability causes macular edema, leading to visual deterioration in retinal diseases such as diabetic retinopathy and retinal vascular occlusion. Dysregulation of junction integrity between endothelial cells by vascular endothelial growth factor (VEGF) was shown to cause retinal vascular hyperpermeability. Accordingly, anti-VEGF agents have been used to treat retinal vascular hyperpermeability. However, they can confer potential toxicity through their deleterious effects on maintenance and survival of neuronal and endothelial cells in the retina. Thus, it is important to identify novel therapeutic targets for retinal vascular hyperpermeability other than VEGF. Here, we prepared murine retinas showing VEGF-induced vascular leakage from superficial retinal vascular plexus and prevention of VEGF-induced leakage by anti-VEGF antibody treatment. We then performed comprehensive proteome profiling of these samples and identified retinal proteins for which abundances were differentially expressed by VEGF, but such alterations were inhibited by anti-VEGF antibody. Functional enrichment and network analyses of these proteins revealed the β2 integrin pathway, which can prevent dysregulation of junction integrity between endothelial cells through cytoskeletal rearrangement, as a potential therapeutic target for retinal vascular hyperpermeability. Finally, we experimentally demonstrated that inhibition of the β2 integrin pathway salvaged VEGF-induced retinal vascular hyperpermeability, supporting its validity as an alternative therapeutic target to anti-VEGF agents. PMID:26969716

  1. Interaction of the α2A domain of integrin with small collagen fragments

    OpenAIRE

    Siebert, Hans-Christian; Burg-Roderfeld, Monika; Eckert, Thomas; Stötzel, Sabine; Kirch, Ulrike; Diercks, Tammo; Humphries, Martin J.; Frank, Martin; Wechselberger, Rainer; Tajkhorshid, Emad; Oesser, Steffen

    2010-01-01

    We here present a detailed study of the ligand-receptor interactions between single and triple-helical strands of collagen and the α2A domain of integrin (α2A), providing valuable new insights into the mechanisms and dynamics of collagen-integrin binding at a sub-molecular level. The occurrence of single and triple-helical strands of the collagen fragments was scrutinized with atom force microscopy (AFM) techniques. Strong interactions of the triple-stranded fragments comparable to those of c...

  2. Intracellular signals direct integrin localization to sites of function in embryonic muscles

    OpenAIRE

    1996-01-01

    In the Drosophila embryo, the alphaPS2betaPS integrin heterodimer is localized tightly at the termini of the multinucleate muscles where they attach to the alphaPS1betaPS-containing epidermal tendon cells. Here we examine the basis for alphaPS2betaPS integrin subcellular localization. We show that the betaPS cytoplasmic tail is sufficient to direct the localization of a heterologous transmembrane protein, CD2, to the muscle termini in vivo. This localization does not occur via an association ...

  3. Stable Peptides Instead of Stapled Peptides: Highly Potent αvβ6-Selective Integrin Ligands.

    Science.gov (United States)

    Maltsev, Oleg V; Marelli, Udaya Kiran; Kapp, Tobias G; Di Leva, Francesco Saverio; Di Maro, Salvatore; Nieberler, Markus; Reuning, Ute; Schwaiger, Markus; Novellino, Ettore; Marinelli, Luciana; Kessler, Horst

    2016-01-22

    The αvβ6 integrin binds the RGD-containing peptide of the foot and mouth disease virus with high selectivity. In this study, the long binding helix of this ligand was downsized to an enzymatically stable cyclic peptide endowed with sub-nanomolar binding affinity toward the αvβ6 receptor and remarkable selectivity against other integrins. Computational studies were performed to disclose the molecular bases underlying the high binding affinity and receptor subtype selectivity of this peptide. Finally, the utility of the ligand for use in biomedical studies was also demonstrated here. PMID:26663660

  4. α6β4 integrin and dystroglycan cooperate to stabilize the myelin sheath

    OpenAIRE

    Nodari, A.; Previtali, S.C.; Dati, G.; Occhi, S.; Court, FA.; Colombelli, C.; Zambroni, D.; Dina, G.; Del Carro, U.; Campbell, K. P.; Quattrini, A; Wrabetz, L.; Feltri, ML.

    2008-01-01

    Schwann cells integrate signals deriving from the axon and the basal lamina to myelinate peripheral nerves. Integrin α6β4 is a laminin receptor synthesized by Schwann cells and displayed apposed to the basal lamina. α6β4 integrin expression in Schwann cells is induced by axons at the onset of myelination, and rise in adulthood. The β4 chain has a uniquely long cytoplasmic domain that interacts with intermediate filaments such as dystonin, important in peripheral myelination. Furthermore, α6β4...

  5. PKCθ signaling is required for myoblast fusion by regulating the expression of caveolin-3 and β1D integrin upstream focal adhesion kinase

    Science.gov (United States)

    Madaro, Luca; Marrocco, Valeria; Fiore, Piera; Aulino, Paola; Smeriglio, Piera; Adamo, Sergio; Molinaro, Mario; Bouché, Marina

    2011-01-01

    Fusion of mononucleated myoblasts to form multinucleated myofibers is an essential phase of skeletal myogenesis, which occurs during muscle development as well as during postnatal life for muscle growth, turnover, and regeneration. Many cell adhesion proteins, including integrins, have been shown to be important for myoblast fusion in vertebrates, and recently focal adhesion kinase (FAK), has been proposed as a key mediator of myoblast fusion. Here we focused on the possible role of PKCθ, the PKC isoform predominantly expressed in skeletal muscle, in myoblast fusion. We found that the expression of PKCθ is strongly up-regulated following freeze injury–induced muscle regeneration, as well as during in vitro differentiation of satellite cells (SCs; the muscle stem cells). Using both PKCθ knockout and muscle-specific PKCθ dominant-negative mutant mouse models, we observed delayed body and muscle fiber growth during the first weeks of postnatal life, when compared with wild-type (WT) mice. We also found that myofiber formation, during muscle regeneration after freeze injury, was markedly impaired in PKCθ mutant mice, as compared with WT. This phenotype was associated with reduced expression of the myogenic differentiation program executor, myogenin, but not with that of the SC marker Pax7. Indeed in vitro differentiation of primary muscle-derived SCs from PKCθ mutants resulted in the formation of thinner myotubes with reduced numbers of myonuclei and reduced fusion rate, when compared with WT cells. These effects were associated to reduced expression of the profusion genes caveolin-3 and β1D integrin and to reduced activation/phosphorylation of their up-stream regulator FAK. Indeed the exogenous expression of a constitutively active mutant form of PKCθ in muscle cells induced FAK phosphorylation. Moreover pharmacologically mediated full inhibition of FAK activity led to similar fusion defects in both WT and PKCθ-null myoblasts. We thus propose that PKC

  6. Delivery of dexamethasone from bioactive nanofiber matrices stimulates odontogenesis of human dental pulp cells through integrin/BMP/mTOR signaling pathways

    Directory of Open Access Journals (Sweden)

    Lim HC

    2016-06-01

    Full Text Available Hyun-Chang Lim,1,* Ok Hyung Nam,2,* Mi-joo Kim,3 Ahmed El-Fiqi,4,5 Hyung-Mun Yun,3 Yoo-Mi Lee,3 Guang-Zhen Jin,4,5 Hae-Hyoung Lee,5,6 Hae-Won Kim,4–6 Eun-Cheol Kim3 1Department of Periodontology, 2Department of Pediatric Dentistry, 3Department of Oral and Maxillofacial Pathology, Research Center for Tooth and Periodontal Regeneration (MRC, School of Dentistry, Kyung Hee University, Seoul, 4Department of Nanobiomedical Science, BK21 PLUS NBM Global Research Center for Regenerative Medicine, 5Institute of Tissue Regeneration Engineering, 6Department of Biomaterials Science, College of Dentistry, Dankook University, Cheonan, Republic of Korea *These authors contributed equally to this work as first authors Abstract: Therapeutically relevant design of scaffolds is of special importance in the repair and regeneration of tissues including dentin and pulp. Here we exploit nanofiber matrices that incorporate bioactive glass nanoparticles (BGNs and deliver the odontogenic drug dexamethasone (DEX to stimulate the odontogenic differentiation of human dental pulp cells (HDPCs. DEX molecules were first loaded onto the BGN, and then the DEX-BGN complex was incorporated within the biopolymer nanofiber matrix through electrospinning. The release of DEX continued over a month, showing a slow releasing profile. HDPCs cultured on the DEX-releasing BGN matrices were viable, proliferating well up to 14 days. The odontogenic differentiation, as assessed by alkaline phosphatase activity, mRNA expression of genes, and mineralization, was significantly stimulated on the matrices incorporating BGN and further on those releasing DEX. The DEX-releasing BGN matrices highly upregulated the expression of the integrin subsets α1, α5, and β3 as well as integrin downstream signaling molecules, including focal adhesion kinase (FAK, Paxillin, and RhoA, and activated bone morphogenetic protein mRNA and phosphorylation of Smad1/5/8. Furthermore, the DEX-releasing BGN

  7. Modulation of TGFβ1-Dependent Myofibroblast Differentiation by Hyaluronan

    OpenAIRE

    Webber, Jason; Jenkins, Robert H.; Meran, Soma; Phillips, Aled; Steadman, Robert

    2009-01-01

    Myofibroblasts are contractile cells that are characterized by the expression of α-smooth muscle actin and mediate the closure of wounds and the formation of collagen-rich scars. Their presence in organs such as lungs, liver, and kidney has long been established as a marker of progressive fibrosis. The transforming growth factor beta1-driven differentiation of fibroblasts is a major source of myofibroblasts, and recent data have shown that hyaluronan is a major modulator of this process. This...

  8. Preferential reduction of binding of sup 125 I-iodopindolol to beta-1 adrenoceptors in the amygdala of rat after antidepressant treatments

    Energy Technology Data Exchange (ETDEWEB)

    Ordway, G.A.; Gambarana, C.; Tejani-Butt, S.M.; Areso, P.; Hauptmann, M.; Frazer, A. (Veterans Affairs Medical Center, Philadelphia, PA (USA))

    1991-05-01

    This study utilized quantitative receptor autoradiography to examine the effects of repeated administration of antidepressants to rats on the binding of the beta adrenoceptor antagonist, {sup 125}I-iodopindolol ({sup 125}I-IPIN) to either beta-1 or beta-2 adrenoceptors in various regions of brain. Antidepressants were selected to represent various chemical and pharmacological classes including tricyclic compounds (desipramine and protriptyline), monoamine oxidase inhibitors (clorgyline, phenelzine and tranylcypromine), atypical antidepressants (mianserin and trazodone) and selective inhibitors of the uptake of serotonin (citalopram and sertraline). Additionally, rats were treated with various psychotropic drugs that lack antidepressant efficacy (cocaine, deprenyl, diazepam and haloperidol). Repeated treatment of rats with desipramine, protriptyline, clorgyline, phenelzine, tranylcypromine or mianserin reduced the binding of {sup 125}I-IPIN to beta-1 adrenoceptors in many brain areas. Only in the basolateral and lateral nuclei of the amygdala did all six of these antidepressants significantly reduce {sup 125}I-IPIN binding to beta-1 adrenoceptors. In these amygdaloid nuclei, the magnitude of the reduction in the binding of {sup 125}I-IPIN caused by each of these drugs was comparable to or greater than the reduction in binding produced in any other region of brain. Reductions of binding of {sup 125}I-IPIN after antidepressant treatments were not consistently observed in the cortex, the area of brain examined most often in homogenate binding studies. Only the monoamine oxidase inhibitors caused reductions in the binding of {sup 125}I-IPIN to beta-2 adrenoceptors, and this effect was generally localized to the amygdala and hypothalamus.

  9. A novel needle for subcutaneous injection of interferon beta-1a: effect on pain in volunteers and satisfaction in patients with multiple sclerosis

    Directory of Open Access Journals (Sweden)

    Prais Wes A

    2008-10-01

    Full Text Available Abstract Background To reduce injection pain and improve satisfaction, a thinner (29-gauge [29G], sharper (5-bevel needle than the 27G/3-bevel needle used previously to inject interferon (IFN beta-1a, 44 or 22 mcg subcutaneously (sc three times weekly (tiw, was developed for use in multiple sclerosis (MS. Methods Two clinical trials in healthy volunteers and five surveys of patients with MS were conducted to assess whether the 29G/5-bevel needle with a Thermo Plastic Elastomer (TPE needle shield (a sleeve that houses the tip of the needle in a secure location is an improvement over the 27G/3-bevel needle with a rubber shield for injection of IFN beta-1a, 44 or 22 mcg sc tiw. Parameters assessed were: pain and ease of insertion (healthy volunteer and nurse responses on subjective pain measurement scales; and patient satisfaction (surveys of patients with MS. Results In healthy volunteers, the 29G/5-bevel needle with TPE shield was associated with the least perceived pain on the Visual Analog Scale (VAS and Verbal VAS (VB-VAS; mean VAS pain scores decreased by 40% and skin penetration improved by 69% compared with the 27G/3-bevel needle with standard rubber shield (p Conclusion Together these studies indicate that the 29G/5-bevel needle with the TPE shield is an improvement over the 27G/3-bevel needle with standard rubber shield in terms of pain, ease of insertion and patient satisfaction. These improvements are expected to result in improved compliance in patients with MS treated with IFN beta-1a, 44 or 22 mcg sc tiw.

  10. Integrin αIIb (CD41) plays a role in the maintenance of hematopoietic stem cell activity in the mouse embryonic aorta

    OpenAIRE

    Boisset, Jean-Charles; Clapes, Thomas; van der Linden, Reinier; Dzierzak, Elaine; Robin, Catherine

    2013-01-01

    Integrins are transmembrane receptors that play important roles as modulators of cell behaviour through their adhesion properties and the initiation of signaling cascades. The αIIb integrin subunit (CD41) is one of the first cell surface markers indicative of hematopoietic commitment. αIIb pairs exclusively with β3 to form the αIIbβ3 integrin. β3 (CD61) also pairs with αv (CD51) to form the αvβ3 integrin. The expression and putative role of these integrins during mouse hematopoietic developme...

  11. Release of transforming growth factor beta 1 and platelet derived growth factor type AB from canine platelet gels obtained by the tube method and activated with calcium salts

    OpenAIRE

    RF Silva; GC Santana; FOP Leme; JU Carmona; CMF Rezende

    2013-01-01

    The objectives of this study were: 1) to measure the concentrations of transforming growth factor beta 1 (TGF-β1) and platelet-derived growth factor type AB (PDGF-AB) in plasma and platelet gel (PG) activated with calcium salts (gluconate or chloride) in dogs, and 2) to determine correlations between cell results and growth factors (GF) concentrations. Blood samples were collected from fourteen Brazilian Fila dogs. EDTA was used to obtain whole blood and plasma while ACD-A solution was used t...

  12. Oral administration of beta-1,3/1,6-glucan to dogs temporally changes total and antigen-specific IgA and IgM

    OpenAIRE

    Stuyven, E; Verdonck, F.; Hoek, I.; Daminet, S.; Duchateau, Luc; Remon, J P; Goddeeris, Bruno; Cox, Eric

    2010-01-01

    The effect of oral administration of beta-1,3/1,6-glucans from Saccharomyces cerevisiae on humoral immunity in domestic dogs is not known. In this study, 15 beagle dogs were orally given MacroGard tablets, which contain 150 mg of this beta-glucan, daily for 4 weeks. At the end of this period, the total serum immunoglobulin A (IgA) level decreased significantly in the group treated with the glucan compared to that in the control group as well as compared to the concentrations before supplement...

  13. Diabetes mellitus affects the biomechanical function of the callus and the expression of TGF-beta1 and BMP2 in an early stage of fracture healing

    OpenAIRE

    Xu, M. T.; Sun, S.; Zhang, L.; Xu, F.; Du, S.L.; Zhang, X. D.; Wang, D. W.

    2015-01-01

    Transforming growth factor beta 1 (TGF-β1) and bone morphogenetic protein-2 (BMP-2) are important regulators of bone repair and regeneration. In this study, we examined whether TGF-β1 and BMP-2 expressions were delayed during bone healing in type 1 diabetes mellitus. Tibial fractures were created in 95 diabetic and 95 control adult male Wistar rats of 10 weeks of age. At 1, 2, 3, 4, and 5 weeks after fracture induction, five rats were sacrificed from each group. The expressions of TGF-β1 and ...

  14. TGF-beta1-induced EMT of non-transformed prostate hyperplasia cells is characterized by early induction of SNA12/slug

    Czech Academy of Sciences Publication Activity Database

    Slabáková, Eva; Pernicová, Zuzana; Slavíčková, Eva; Staršíchová, Andrea; Kozubík, Alois; Souček, Karel

    2011-01-01

    Roč. 71, č. 12 (2011), s. 1332-1343. ISSN 0270-4137 R&D Projects: GA ČR(CZ) GA310/07/0961; GA ČR(CZ) GD303/09/H048; GA MZd NS9600; GA MZd NS9956 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : epithelial-mesenchymal transition * SNAI2/Slug * transforming growth factor-beta1 Subject RIV: BO - Biophysics Impact factor: 3.485, year: 2011

  15. Long-term effect of early treatment with interferon beta-1b after a first clinical event suggestive of multiple sclerosis: 5-year active treatment extension of the phase 3 BENEFIT trial

    DEFF Research Database (Denmark)

    Kappos, Ludwig; Freedman, Mark S; Polman, Chris H;

    2009-01-01

    with interferon beta-1b on time to clinically definite multiple sclerosis (CDMS) and other disease outcomes, including disability progression. METHODS: Patients with a first event suggestive of multiple sclerosis and a minimum of two clinically silent lesions in MRI were randomly assigned to receive...... interferon beta-1b 250 microg (n=292; early treatment) or placebo (n=176; delayed treatment) subcutaneously every other day for 2 years, or until diagnosis of CDMS. All patients were then eligible to enter a prospectively planned follow-up phase with open-label interferon beta-1b up to a maximum of 5 years...... 125 in both groups. No significant differences in other disability related outcomes were recorded. Frequency and severity of adverse events remained within the established safety and tolerability profile of interferon beta-1b. INTERPRETATION: Effects on the rate of conversion to CDMS and the...

  16. Improved targeting of the alpha(v)beta (3) integrin by multimerisation of RGD peptides.

    NARCIS (Netherlands)

    Dijkgraaf, I.; Kruijtzer, J.A.; Liu, S.; Soede, A.C.; Oyen, W.J.G.; Corstens, F.H.M.; Liskamp, R.M.; Boerman, O.C.

    2007-01-01

    PURPOSE: The integrin alpha(v)beta(3) is expressed on sprouting endothelial cells and on various tumour cell types. Due to the restricted expression of alpha(v)beta(3) in tumours, alpha(v)beta(3) is considered a suitable receptor for tumour targeting. In this study the alpha(v)beta(3) binding charac

  17. Actin flow and talin dynamics govern rigidity sensing in actin–integrin linkage through talin extension

    Science.gov (United States)

    Hirata, Hiroaki; Chiam, Keng-Hwee; Lim, Chwee Teck; Sokabe, Masahiro

    2014-01-01

    At cell–substrate adhesion sites, the linkage between actin filaments and integrin is regulated by mechanical stiffness of the substrate. Of potential molecular regulators, the linker proteins talin and vinculin are of particular interest because mechanical extension of talin induces vinculin binding with talin, which reinforces the actin–integrin linkage. For understanding the molecular and biophysical mechanism of rigidity sensing at cell–substrate adhesion sites, we constructed a simple physical model to examine a role of talin extension in the stiffness-dependent regulation of actin–integrin linkage. We show that talin molecules linking between retrograding actin filaments and substrate-bound integrin are extended in a manner dependent on substrate stiffness. The model predicts that, in adhesion complexes containing ≈30 talin links, talin is extended enough for vinculin binding when the substrate is stiffer than 1 kPa. The lifetime of talin links needs to be 2–5 s to achieve an appropriate response of talin extension against substrate stiffness. Furthermore, changes in actin velocity drastically shift the range of substrate stiffness that induces talin–vinculin binding. Our results suggest that talin extension is a key step in sensing and responding to substrate stiffness at cell adhesion sites. PMID:25142525

  18. Actin flow and talin dynamics govern rigidity sensing in actin-integrin linkage through talin extension.

    Science.gov (United States)

    Hirata, Hiroaki; Chiam, Keng-Hwee; Lim, Chwee Teck; Sokabe, Masahiro

    2014-10-01

    At cell-substrate adhesion sites, the linkage between actin filaments and integrin is regulated by mechanical stiffness of the substrate. Of potential molecular regulators, the linker proteins talin and vinculin are of particular interest because mechanical extension of talin induces vinculin binding with talin, which reinforces the actin-integrin linkage. For understanding the molecular and biophysical mechanism of rigidity sensing at cell-substrate adhesion sites, we constructed a simple physical model to examine a role of talin extension in the stiffness-dependent regulation of actin-integrin linkage. We show that talin molecules linking between retrograding actin filaments and substrate-bound integrin are extended in a manner dependent on substrate stiffness. The model predicts that, in adhesion complexes containing ≈30 talin links, talin is extended enough for vinculin binding when the substrate is stiffer than 1 kPa. The lifetime of talin links needs to be 2-5 s to achieve an appropriate response of talin extension against substrate stiffness. Furthermore, changes in actin velocity drastically shift the range of substrate stiffness that induces talin-vinculin binding. Our results suggest that talin extension is a key step in sensing and responding to substrate stiffness at cell adhesion sites. PMID:25142525

  19. DNA-based digital tension probes reveal integrin forces during early cell adhesion

    Science.gov (United States)

    Zhang, Yun; Ge, Chenghao; Zhu, Cheng; Salaita, Khalid

    2014-10-01

    Mechanical stimuli profoundly alter cell fate, yet the mechanisms underlying mechanotransduction remain obscure because of a lack of methods for molecular force imaging. Here to address this need, we develop a new class of molecular tension probes that function as a switch to generate a 20- to 30-fold increase in fluorescence upon experiencing a threshold piconewton force. The probes employ immobilized DNA hairpins with tunable force response thresholds, ligands and fluorescence reporters. Quantitative imaging reveals that integrin tension is highly dynamic and increases with an increasing integrin density during adhesion formation. Mixtures of fluorophore-encoded probes show integrin mechanical preference for cyclized RGD over linear RGD peptides. Multiplexed probes with variable guanine-cytosine content within their hairpins reveal integrin preference for the more stable probes at the leading tip of growing adhesions near the cell edge. DNA-based tension probes are among the most sensitive optical force reporters to date, overcoming the force and spatial resolution limitations of traction force microscopy.

  20. A computational analysis of the dynamic roles of talin, Dok1, and PIPKI for integrin activation.

    Directory of Open Access Journals (Sweden)

    Florian Geier

    Full Text Available Integrin signaling regulates cell migration and plays a pivotal role in developmental processes and cancer metastasis. Integrin signaling has been studied extensively and much data is available on pathway components and interactions. Yet the data is fragmented and an integrated model is missing. We use a rule-based modeling approach to integrate available data and test biological hypotheses regarding the role of talin, Dok1 and PIPKI in integrin activation. The detailed biochemical characterization of integrin signaling provides us with measured values for most of the kinetics parameters. However, measurements are not fully accurate and the cellular concentrations of signaling proteins are largely unknown and expected to vary substantially across different cellular conditions. By sampling model behaviors over the physiologically realistic parameter range we find that the model exhibits only two different qualitative behaviors and these depend mainly on the relative protein concentrations, which offers a powerful point of control to the cell. Our study highlights the necessity to characterize model behavior not for a single parameter optimum, but to identify parameter sets that characterize different signaling modes.

  1. Examination of soluble integrin resistant mutants of foot-and-mouth disease virus (FMDV)

    Science.gov (United States)

    Foot-and-mouth disease virus (FMDV) initiates infection in vitro via recognition of at least four cell-surface integrin molecules avb1, avb3, avb6 or avb8 through the interaction of a highly conserved Arg-Gly-Asp (RGD) amino acid sequence motif located in the GH loop of VP1. In this work, soluble i...

  2. Cordycepin suppresses integrin/FAK signaling and epithelial-mesenchymal transition in hepatocellular carcinoma.

    Science.gov (United States)

    Yao, Wen-Ling; Ko, Bor-Sheng; Liu, Tzu-An; Liang, Shu-Man; Liu, Chia-Chia; Lu, Yi-Jhu; Tzean, Shean-Shong; Shen, Tang-Long; Liou, Jun-Yang

    2014-01-01

    Cordycepin, also known as 3-deoxyadenosine, is an analogue of adenosine extracted from the traditional Chinese medicine "Dong Chong Xia Cao". Cordycepin is an active small molecular weight compound and is implicated in modulating multiple physiological functions including immune activation, anti-aging and anti-tumor effects. Several studies have indicated that cordycepin suppresses tumor progression. However, the signaling pathways involved in cordycepin regulating cancer cell motility, invasiveness and epithelial-mesenchymal transition (EMT) remain unclear. In this study, we found that cordycepin inhibits hepatocellular carcinoma (HCC) cell proliferation and migration/invasion. Treatment of cordycepin results in the increasing expression of epithelial marker, Ecadherin while no significant effect was found on N-cadherin α-catenin and β-catenin. Furthermore, although the expression of focal adhesion kinase (FAK) was slightly reduced, the level of phosphorylated FAK was significantly reduced by the treatment of cordycepin. In addition, cordycepin significantly suppresses the expression of integrin α3, integrin α6 and integrin β1 which are crucial interacting partners of FAK in regulating the focal adhesion complex. These results suggest cordycepin may contribute to EMT, antimigration/ invasion and growth inhibitory effects of HCC by suppressing E-cadherin and integrin/FAK signaling. Thus, cordycepin is a potential therapeutic or supplementary agent for preventing HCC tumor progression. PMID:23855336

  3. Polyvalent integrin antagonist-decorated superparamagnetic iron oxide nanoparticles for triggering apoptosis in human leukemia cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Say, R Latin-Small-Letter-Dotless-I dvan, E-mail: rsay@anadolu.edu.tr [Anadolu Universitesi, Kimya Boeluemue, Fen Fakueltesi (Turkey); Yazar, Suzan [Sanovel Pharmaceutical Company (Turkey); Ugur, Alper; Huer, Deniz [Anadolu Universitesi, Kimya Boeluemue, Fen Fakueltesi (Turkey); Denizli, Adil [Hacettepe University, Department of Chemistry (Turkey); Ersoez, Arzu [Anadolu Universitesi, Kimya Boeluemue, Fen Fakueltesi (Turkey)

    2013-01-15

    Integrin family members are the main mediators of cell adhesion to the extracellular matrix and active as intra- and extracellular signaling molecules in a variety of processes. They bind to their ligands by interacting with short amino acid sequences, that is, RGD (arginine-glycine-aspartic acid) sequence. RGD sequences have been used to enhance cell binding to artificial surfaces, so RGD mimics have been used to block integrin binding to its ligand. Integrin-ligand interactions are dependent on divalent cations, and Mg{sup 2+} provide higher-affinity binding to ligand for many integrins. In this study, we have designed new integrin antagonists using methacryloyl amidoaspartic acid (MAASP) monomer-conjugated silanized super paramagnetic iron oxide nanoparticles (SPIONs, the size of the nanoparticles was verified with an average size of 32.6 nm) and poly(MAASP-co-EDMA) shell-decorated silanized SPIONs. Several mechanisms have been proposed to describe uptake of modified SPIONs into the cells, including receptor-mediated endocytosis. Our aim is to bind these modified SPIONs to the integrin-mediated aspartic acid ends of MAASP monomers and block integrin binding to their ligand.

  4. Signal regulatory protein alpha negatively regulates beta2 integrin-mediated monocyte adhesion, transendothelial migration and phagocytosis.

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    Dan-Qing Liu

    Full Text Available BACKGROUND: Signal regulate protein alpha (SIRPalpha is involved in many functional aspects of monocytes. Here we investigate the role of SIRPalpha in regulating beta(2 integrin-mediated monocyte adhesion, transendothelial migration (TEM and phagocytosis. METHODOLOGY/PRINCIPAL FINDINGS: THP-1 monocytes/macropahges treated with advanced glycation end products (AGEs resulted in a decrease of SIRPalpha expression but an increase of beta(2 integrin cell surface expression and beta(2 integrin-mediated adhesion to tumor necrosis factor-alpha (TNFalpha-stimulated human microvascular endothelial cell (HMEC-1 monolayers. In contrast, SIRPalpha overexpression in THP-1 cells showed a significant less monocyte chemotactic protein-1 (MCP-1-triggered cell surface expression of beta(2 integrins, in particular CD11b/CD18. SIRPalpha overexpression reduced beta(2 integrin-mediated firm adhesion of THP-1 cells to either TNFalpha-stimulated HMEC-1 monolayers or to immobilized intercellular adhesion molecule-1 (ICAM-1. SIRPalpha overexpression also reduced MCP-1-initiated migration of THP-1 cells across TNFalpha-stimulated HMEC-1 monolayers. Furthermore, beta(2 integrin-mediated THP-1 cell spreading and actin polymerization in response to MCP-1, and phagocytosis of bacteria were both inhibited by SIRPalpha overexpression. CONCLUSIONS/SIGNIFICANCE: SIRPalpha negatively regulates beta(2 integrin-mediated monocyte adhesion, transendothelial migration and phagocytosis, thus may serve as a critical molecule in preventing excessive activation and accumulation of monocytes in the arterial wall during early stage of atherosclerosis.

  5. Vipegitide: a folded peptidomimetic partial antagonist of α2β1 integrin with antiplatelet aggregation activity

    Directory of Open Access Journals (Sweden)

    Momic T

    2015-01-01

    Full Text Available Tatjana Momic,1 Jehoshua Katzhendler,1 Ela Shai,2 Efrat Noy,3 Hanoch Senderowitz,3 Johannes A Eble,4 Cezary Marcinkiewicz,5 David Varon,2 Philip Lazarovici11School of Pharmacy Institute for Drug Research, Faculty of Medicine, The Hebrew University of Jerusalem, 2Department of Hematology, Coagulation Unit, Hadassah-Hebrew University Medical Center, Jerusalem, Israel; 3Department of Chemistry, Bar-Ilan University, Ramat-Gan, Israel; 4Institute for Physiological Chemistry and Pathobiochemistry, University of Münster, Münster, Germany; 5Department of Bioengineering, College of Engineering, Temple University, Philadelphia, PA, USAAbstract: Linear peptides containing the sequence WKTSRTSHY were used as lead compounds to synthesize a novel peptidomimetic antagonist of α2β1 integrin, with platelet aggregation-inhibiting activity, named Vipegitide. Vipegitide is a 13-amino acid, folded peptidomimetic molecule, containing two α-aminoisobutyric acid residues at positions 6 and 8 and not stable in human serum. Substitution of glycine and tryptophan residues at positions 1 and 2, respectively, with a unit of two polyethylene glycol (PEG molecules yielded peptidomimetic Vipegitide-PEG2, stable in human serum for over 3 hours. Vipegitide and Vipegitide-PEG2 showed high potency (7×10-10 M and 1.5×10-10 M, respectively and intermediate efficacy (40% and 35%, respectively as well as selectivity toward α2 integrin in inhibition of adhesion of α1/α2 integrin overexpressing cells toward respective collagens. Interaction of both peptidomimetics with extracellular active domain of α2 integrin was confirmed in cell-free binding assay with recombinant α2 A-domain. Integrin α2β1 receptor is found on the platelet membrane and triggers collagen-induced platelet aggregation. Vipegitide and Vipegitide-PEG2 inhibited α2β1 integrin-mediated adhesion of human and murine platelets under the flow condition, by 50%. They efficiently blocked adenosine diphosphate

  6. Radioinduced intestinal fibrosis: from molecular mechanisms to therapy applications. Contribution of the TGF--{beta}1, of the CTGF and of the transduction pathway of the Rho/ROCK signal; La fibrose intestinale radio-induite: des mecanismes moleculaires aux applications therapeutiques. Roles du TGF-{