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Sample records for activities including cell

  1. ΔPK oncolytic activity includes modulation of the tumour cell milieu.

    PubMed

    Bollino, Dominique; Colunga, Aric; Li, Baiquan; Aurelian, Laure

    2016-02-01

    Oncolytic virotherapy is a unique cancer therapeutic that encompasses tumour cell lysis through both virus replication and programmed cell death (PCD) pathways. Nonetheless, clinical efficacy is relatively modest, likely related to the immunosuppressive tumour milieu. Our studies use the herpes simplex virus type 2 (HSV-2)-based oncolytic virus ΔPK that has documented anti-tumour activity associated with virus replication, PCD and cancer stem cell lysis. They are designed to examine whether ΔPK-mediated oncolysis includes the ability to reverse the immunosuppressive tumour microenvironment by altering the balance of cytokines directly secreted by the melanoma cells and to define its mechanism. Here, we show that melanoma cells secreted the immunosuppressive cytokine IL-10, and that secretion was inhibited by ΔPK through virus replication and c-Jun N-terminal kinase/c-Jun activation. ΔPK-induced IL-10 inhibition upregulated surface expression of MHC class I chain-related protein A, the ligand for the activating NKG2D receptor expressed on NK- and cytotoxic T-cells. Concomitantly, ΔPK also upregulated the secretion of inflammatory cytokines TNF-α, granulocyte macrophage colony-stimulating factor and IL-1β through autophagy-mediated activation of Toll-like receptor 2 pathways and pyroptosis, and it inhibited the expression of the negative immune checkpoint regulator cytotoxic T-lymphocyte antigen 4. Pharmacologic inhibition of these processes significantly reduces the oncolytic activity of ΔPK. PMID:26602205

  2. Electrode including porous particles with embedded active material for use in a secondary electrochemical cell

    DOEpatents

    Vissers, Donald R.; Nelson, Paul A.; Kaun, Thomas D.; Tomczuk, Zygmunt

    1978-04-25

    Particles of carbonaceous matrices containing embedded electrode active material are prepared for vibratory loading within a porous electrically conductive substrate. In preparing the particles, active materials such as metal chalcogenides, solid alloys of alkali or alkaline earth metals along with other metals and their oxides in powdered or particulate form are blended with a thermosetting resin and particles of a volatile to form a paste mixture. The paste is heated to a temperature at which the volatile transforms into vapor to impart porosity at about the same time as the resin begins to cure into a rigid, solid structure. The solid structure is then comminuted into porous, carbonaceous particles with the embedded active material.

  3. Antiviral activity of human Vδ2 T-cells against WNV includes both cytolytic and non-cytolytic mechanisms.

    PubMed

    Agrati, Chiara; Castilletti, Concetta; Cimini, Eleonora; Romanelli, Antonella; Lapa, Daniele; Quartu, Serena; Martini, Federico; Capobianchi, Maria Rosaria

    2016-04-01

    West Nile virus (WNV) causes a severe central nervous system infection in humans, primarily in the elderly and immunocompromised subjects. Human γδ T-cells play a critical role in the immune response against viruses, and studies of WNV meningoencephalitis in laboratory mice described a role of γδ T-cells in the protective immune response. Aim of this study was to analyze the cytolytic and non-cytolytic antiviral activity of human Vδ2 T-cells against WNV replication. The anti-WNV activity of soluble factor released by zoledronic acid (ZA)-activated Vδ2 T-cell lines and the cytotoxic capability of Vδ2 T-cell lines against WNV-infected cells were tested in vitro. The activation of Vδ2 T-cell lines was able to inhibit WNV replication through the release of soluble factors. IFN-γ is massively released by activated Vδ2 T-cell lines and is involved in the anti-WNV activity. Moreover, the Vδ2 T-cell lines can efficiently kill WNV-infected cells possibly through perforin-mediated mechanism. Altogether, our results provide insight into the effector functions of human Vδ2 T-cells against WNV. The possibility to target these cells by ZA, a commercially available drug used in humans, could potentially offer a new immunotherapeutic strategy for WNV infection. PMID:27196553

  4. Selective Non-nucleoside Inhibitors of Human DNA Methyltransferases Active in Cancer Including in Cancer Stem Cells

    PubMed Central

    2015-01-01

    DNA methyltransferases (DNMTs) are important enzymes involved in epigenetic control of gene expression and represent valuable targets in cancer chemotherapy. A number of nucleoside DNMT inhibitors (DNMTi) have been studied in cancer, including in cancer stem cells, and two of them (azacytidine and decitabine) have been approved for treatment of myelodysplastic syndromes. However, only a few non-nucleoside DNMTi have been identified so far, and even fewer have been validated in cancer. Through a process of hit-to-lead optimization, we report here the discovery of compound 5 as a potent non-nucleoside DNMTi that is also selective toward other AdoMet-dependent protein methyltransferases. Compound 5 was potent at single-digit micromolar concentrations against a panel of cancer cells and was less toxic in peripheral blood mononuclear cells than two other compounds tested. In mouse medulloblastoma stem cells, 5 inhibited cell growth, whereas related compound 2 showed high cell differentiation. To the best of our knowledge, 2 and 5 are the first non-nucleoside DNMTi tested in a cancer stem cell line. PMID:24387159

  5. Photoelectrochemical cells including chalcogenophosphate photoelectrodes

    NASA Technical Reports Server (NTRS)

    Reichman, B.; Byvik, C. E. (Inventor)

    1984-01-01

    Photoelectrochemical cells employing chalcogenophosphate (MPX3) photoelectrodes are described where M is selected from the group of transition metal series of elements beginning with scandium (atomic number 21) through germanium (atomic number 32) yttrium (atomic number 39) through antimony (atomic number 51) and lanthanum (atomic number 57) through polonium (atomic number 84); P is phosphorus; and X is selected from the chalogenide series consisting of sulfur, selenium, and tellurium. These compounds have bandgaps in the desirable range from 2.0 eV to 2.2 eV for the photoelectrolysis of water and are stable when used as photoelectrodes for the same.

  6. Cutting edge: identification of a novel chemokine receptor that binds dendritic cell- and T cell-active chemokines including ELC, SLC, and TECK.

    PubMed

    Gosling, J; Dairaghi, D J; Wang, Y; Hanley, M; Talbot, D; Miao, Z; Schall, T J

    2000-03-15

    Searching for new receptors of dendritic cell- and T cell-active chemokines, we used a combination of techniques to interrogate orphan chemokine receptors. We report here on human CCX CKR, previously represented only by noncontiguous expressed sequence tags homologous to bovine PPR1, a putative gustatory receptor. We employed a two-tiered process of ligand assignment, where immobilized chemokines constructed on stalks (stalkokines) were used as bait for adhesion of cells expressing CCX CKR. These cells adhered to stalkokines representing ELC, a chemokine previously thought to bind only CCR7. Adhesion was abolished in the presence of soluble ELC, SLC (CCR7 ligands), and TECK (a CCR9 ligand). Complete ligand profiles were further determined by radiolabeled ligand binding and competition with >80 chemokines. ELC, SLC, and TECK comprised high affinity ligands (IC50 <15 nM); lower affinity ligands include BLC and vMIP-II (IC50 <150 nM). With its high affinity for CC chemokines and homology to CC receptors, we provisionally designate this new receptor CCR10. PMID:10706668

  7. PREFACE: 9th International Fröhlich's Symposium: Electrodynamic Activity of Living Cells (Including Microtubule Coherent Modes and Cancer Cell Physics)

    NASA Astrophysics Data System (ADS)

    Cifra, Michal; Pokorný, Jirí; Kucera, Ondrej

    2011-12-01

    This volume contains papers presented at the International Fröhlich's Symposium entitled 'Electrodynamic Activity of Living Cells' (1-3 July 2011, Prague, Czech Republic). The Symposium was the 9th meeting devoted to physical processes in living matter organized in Prague since 1987. The hypothesis of oscillation systems in living cells featured by non-linear interaction between elastic and electrical polarization fields, non-linear interactions between the system and the heat bath leading to energy downconversion along the frequency scale, energy condensation in the lowest frequency mode and creation of a coherent state was formulated by H Fröhlich, founder of the theory of dielectric materials. He assumed that biological activity is based not only on biochemical but also on biophysical mechanisms and that their disturbances form basic links along the cancer transformation pathway. Fröhlich outlined general ideas of non-linear physical processes in biological systems. The downconversion and the elastic-polarization interactions should be connected in a unified theory and the solution based on comprehensive non-linear characteristics. Biochemical and genetic research of biological systems are highly developed and have disclosed a variety of cellular and subcellular structures, chemical reactions, molecular information transfer, and genetic code sequences - including their pathological development. Nevertheless, the cancer problem is still a big challenge. Warburg's discovery of suppressed oxidative metabolism in mitochondria in cancer cells suggested the essential role of physical mechanisms (but his discovery has remained without impact on cancer research and on the study of physical properties of biological systems for a long time). Mitochondria, the power plants of the cell, have several areas of activity-oxidative energy production is connected with the formation of a strong static electric field around them, water ordering, and liberation of non

  8. CD161++CD8+ T cells, including the MAIT cell subset, are specifically activated by IL-12+IL-18 in a TCR-independent manner

    PubMed Central

    Ussher, James E; Bilton, Matthew; Attwod, Emma; Shadwell, Jonathan; Richardson, Rachel; de Lara, Catherine; Mettke, Elisabeth; Kurioka, Ayako; Hansen, Ted H; Klenerman, Paul; Willberg, Christian B

    2014-01-01

    CD161++CD8+ T cells represent a novel subset that is dominated in adult peripheral blood by mucosal-associated invariant T (MAIT) cells, as defined by the expression of a variable-α chain 7.2 (Vα7.2)-Jα33 TCR, and IL-18Rα. Stimulation with IL-18+IL-12 is known to induce IFN-γ by both NK cells and, to a more limited extent, T cells. Here, we show the CD161++ CD8+ T-cell population is the primary T-cell population triggered by this mechanism. Both CD161++Vα7.2+ and CD161++Vα7.2− T-cell subsets responded to IL-12+IL-18 stimulation, demonstrating this response was not restricted to the MAIT cells, but to the CD161++ phenotype. Bacteria and TLR agonists also indirectly triggered IFN-γ expression via IL-12 and IL-18. These data show that CD161++ T cells are the predominant T-cell population that responds directly to IL-12+IL-18 stimulation. Furthermore, our findings broaden the potential role of MAIT cells beyond bacterial responsiveness to potentially include viral infections and other inflammatory stimuli. PMID:24019201

  9. Origin of Rebound Plasma HIV Includes Cells with Identical Proviruses That Are Transcriptionally Active before Stopping of Antiretroviral Therapy

    PubMed Central

    Wiegand, Ann; Shao, Wei; Coffin, John M.; Mellors, John W.; Lederman, Michael; Gandhi, Rajesh T.; Keele, Brandon F.

    2015-01-01

    ABSTRACT Understanding the origin of HIV variants during viral rebound may provide insight into the composition of the HIV reservoir and has implications for the design of curative interventions. HIV single-genome sequences were obtained from 10 AIDS Clinical Trials Group participants who underwent analytic antiretroviral therapy (ART) interruption (ATI). Rebounding variants were compared with those in pre-ART plasma in all 10 participants and with on-ART peripheral blood mononuclear cell (PBMC)-associated DNA and RNA (CA-RNA) in 7/10 participants. The highest viral diversities were found in the DNA and CA-RNA populations. In 3 of 7 participants, we detected multiple, identical DNA and CA-RNA sequences during suppression on ART that exactly matched plasma HIV sequences. Hypermutated DNA and CA-RNA were detected in four participants, contributing to diversities in these compartments that were higher than in the pre-ART and post-ATI plasma. Shifts in the viral rebound populations could be detected in some participants over the 2- to 3-month observation period. These findings suggest that a source of initial rebound viremia could be populations of infected cells that clonally expanded prior to and/or during ART, some of which were already expressing HIV RNA before treatment was interrupted. These clonally expanding populations of HIV-infected cells may represent an important target for strategies aimed at achieving reservoir reduction and sustained virologic remission. IMPORTANCE Antiretroviral therapy alone cannot eradicate the HIV reservoir, and viral rebound is generally rapid after treatment interruption. It has been suggested that clonal expansion of HIV-infected cells is an important mechanism of HIV reservoir persistence, but the contribution of these clonally proliferating cells to the rebounding virus is unknown. We report a study of AIDS Clinical Trials Group participants who underwent treatment interruption and compared rebounding plasma virus with that

  10. The ethanol extract of Scutellaria baicalensis and the active compounds induce cell cycle arrest and apoptosis including upregulation of p53 and Bax in human lung cancer cells

    SciTech Connect

    Gao Jiayu; Morgan, Winston A.; Sanchez-Medina, Alberto; Corcoran, Olivia

    2011-08-01

    Despite a lack of scientific authentication, Scutellaria baicalensis is clinically used in Chinese medicine as a traditional adjuvant to chemotherapy of lung cancer. In this study, cytotoxicity assays demonstrated that crude ethanolic extracts of S. baicalensis were selectively toxic to human lung cancer cell lines A549, SK-LU-1 and SK-MES-1 compared with normal human lung fibroblasts. The active compounds baicalin, baicalein and wogonin did not exhibit such selectivity. Following exposure to the crude extracts, cellular protein expression in the cancer cell lines was assessed using 2D gel electrophoresis coupled with MALDI-TOF-MS/Protein Fingerprinting. The altered protein expression indicated that cell growth arrest and apoptosis were potential mechanisms of cytotoxicity. These observations were supported by PI staining cell cycle analysis using flow cytometry and Annexin-V apoptotic analysis by fluorescence microscopy of cancer cells treated with the crude extract and pure active compounds. Moreover, specific immunoblotting identification showed the decreased expression of cyclin A results in the S phase arrest of A549 whereas the G{sub 0}/G{sub 1} phase arrest in SK-MES-1 cells results from the decreased expression of cyclin D1. Following treatment, increased expression in the cancer cells of key proteins related to the enhancement of apoptosis was observed for p53 and Bax. These results provide further insight into the molecular mechanisms underlying the clinical use of this herb as an adjuvant to lung cancer therapy. - Research Highlights: > Scutellaria baicalensis is a clinical adjuvant to lung cancer chemotherapy in China. > Scutellaria ethanol extracts selectively toxic to A549, SK-LU-1 and SK-MES-1. > Baicalin, baicalein and wogonin were toxic to all lung cancer cell lines. > Proteomics identified increased p53 and BAX in response to Scutellaria extracts.

  11. Rhinacanthus nasutus Extracts Prevent Glutamate and Amyloid-β Neurotoxicity in HT-22 Mouse Hippocampal Cells: Possible Active Compounds Include Lupeol, Stigmasterol and β-Sitosterol

    PubMed Central

    Brimson, James M.; Brimson, Sirikalaya J.; Brimson, Christopher A.; Rakkhitawatthana, Varaporn; Tencomnao, Tewin

    2012-01-01

    The Herb Rhinacanthus nasutus (L.) Kurz, which is native to Thailand and Southeast Asia, has become known for its antioxidant properties. Neuronal loss in a number of diseases including Alzheimer’s disease is thought to result, in part, from oxidative stress. Glutamate causes cell death in the mouse hippocampal cell line, HT-22, by unbalancing redox homeostasis, brought about by a reduction in glutathione levels, and amyloid-β has been shown to induce reactive oxygen species (ROS) production. Here in, we show that ethanol extracts of R. nasutus leaf and root are capable of dose dependently attenuating the neuron cell death caused by both glutamate and amyloid-β treatment. We used free radical scavenging assays to measure the extracts antioxidant activities and as well as quantifying phenolic, flavonoid and sterol content. Molecules found in R. nasutus, lupeol, stigmasterol and β-sitosterol are protective against glutamate toxicity. PMID:22606031

  12. Chimeric Beta-Defensin Analogs, Including the Novel 3NI Analog, Display Salt-Resistant Antimicrobial Activity and Lack Toxicity in Human Epithelial Cell Lines

    PubMed Central

    Scudiero, Olga; Galdiero, Stefania; Nigro, Ersilia; Del Vecchio, Luigi; Di Noto, Rosa; Cantisani, Marco; Colavita, Irene; Galdiero, Massimiliano; Cassiman, Jean-Jacques; Daniele, Aurora; Pedone, Carlo

    2013-01-01

    Human beta-defensins (hBDs) are crucial peptides for the innate immune response and are thus prime candidates as therapeutic agents directed against infective diseases. Based on the properties of wild-type hBD1 and hBD3 and of previously synthesized analogs (1C, 3I, and 3N), we have designed a new analog, 3NI, and investigated its potential as an antimicrobial drug. Specifically, we evaluated the antimicrobial activities of 3NI versus those of hBD1, hBD3, 1C, 3I, and 3N. Our results show that 3NI exerted greater antibacterial activity against Pseudomonas aeruginosa, Escherichia coli, and Enterococcus faecalis than did hBD1 and hBD3, even with elevated salt concentrations. Moreover, its antiviral activity against herpes simplex virus 1 was greater than that of hBD1 and similar to that of hBD3. Subsequently, we investigated the cytotoxic effects of all peptides in three human epithelial carcinoma cell lines: A549 from lung, CaCo-2 from colon, and Capan-1 from pancreas. None of the analogs significantly reduced cell viability versus wild-type hBD1 and hBD3. They did not induce genotoxicity or cause an increase in the number of apoptotic cells. Using confocal microscopy, we also investigated the localization of the peptides during their incubation with epithelial cells and found that they were distributed on the cell surface, from which they were internalized. Finally, we show that hBD1 and hBD3 are characterized by high resistance to serum degradation. In conclusion, the new analog 3NI seems to be a promising anti-infective agent, particularly given its high salt resistance—a feature that is relevant in diseases such as cystic fibrosis. PMID:23357761

  13. Hematopoietic stem cell transplantation activity worldwide in 2012 and a SWOT analysis of the Worldwide Network for Blood and Marrow Transplantation Group including the global survey.

    PubMed

    Niederwieser, D; Baldomero, H; Szer, J; Gratwohl, M; Aljurf, M; Atsuta, Y; Bouzas, L F; Confer, D; Greinix, H; Horowitz, M; Iida, M; Lipton, J; Mohty, M; Novitzky, N; Nunez, J; Passweg, J; Pasquini, M C; Kodera, Y; Apperley, J; Seber, A; Gratwohl, A

    2016-06-01

    Data on 68 146 hematopoietic stem cell transplants (HSCTs) (53% autologous and 47% allogeneic) gathered by 1566 teams from 77 countries and reported through their regional transplant organizations were analyzed by main indication, donor type and stem cell source for the year 2012. With transplant rates ranging from 0.1 to 1001 per 10 million inhabitants, more HSCTs were registered from unrelated 16 433 donors than related 15 493 donors. Grafts were collected from peripheral blood (66%), bone marrow (24%; mainly non-malignant disorders) and cord blood (10%). Compared with 2006, an increase of 46% total (57% allogeneic and 38% autologous) was observed. Growth was due to an increase in reporting teams (18%) and median transplant activity/team (from 38 to 48 HSCTs/team). An increase of 167% was noted in mismatched/haploidentical family HSCT. A Strengths, Weaknesses, Opportunities, Threats (SWOT) analysis revealed the global perspective of WBMT to be its major strength and identified potential to be the key professional body for patients and authorities. The limited data collection remains its major weakness and threat. In conclusion, global HSCT grows over the years without plateauing (allogeneic>autologous) and at different rates in the four World Health Organization regions. Major increases were observed in allogeneic, haploidentical HSCT and, to a lesser extent, in cord blood transplantation. PMID:26901703

  14. Distinct speed dependence of entorhinal island and ocean cells, including respective grid cells.

    PubMed

    Sun, Chen; Kitamura, Takashi; Yamamoto, Jun; Martin, Jared; Pignatelli, Michele; Kitch, Lacey J; Schnitzer, Mark J; Tonegawa, Susumu

    2015-07-28

    Entorhinal-hippocampal circuits in the mammalian brain are crucial for an animal's spatial and episodic experience, but the neural basis for different spatial computations remain unknown. Medial entorhinal cortex layer II contains pyramidal island and stellate ocean cells. Here, we performed cell type-specific Ca(2+) imaging in freely exploring mice using cellular markers and a miniature head-mounted fluorescence microscope. We found that both oceans and islands contain grid cells in similar proportions, but island cell activity, including activity in a proportion of grid cells, is significantly more speed modulated than ocean cell activity. We speculate that this differential property reflects island cells' and ocean cells' contribution to different downstream functions: island cells may contribute more to spatial path integration, whereas ocean cells may facilitate contextual representation in downstream circuits. PMID:26170279

  15. Distinct speed dependence of entorhinal island and ocean cells, including respective grid cells

    PubMed Central

    Sun, Chen; Kitamura, Takashi; Yamamoto, Jun; Martin, Jared; Pignatelli, Michele; Kitch, Lacey J.; Schnitzer, Mark J.; Tonegawa, Susumu

    2015-01-01

    Entorhinal–hippocampal circuits in the mammalian brain are crucial for an animal’s spatial and episodic experience, but the neural basis for different spatial computations remain unknown. Medial entorhinal cortex layer II contains pyramidal island and stellate ocean cells. Here, we performed cell type-specific Ca2+ imaging in freely exploring mice using cellular markers and a miniature head-mounted fluorescence microscope. We found that both oceans and islands contain grid cells in similar proportions, but island cell activity, including activity in a proportion of grid cells, is significantly more speed modulated than ocean cell activity. We speculate that this differential property reflects island cells’ and ocean cells’ contribution to different downstream functions: island cells may contribute more to spatial path integration, whereas ocean cells may facilitate contextual representation in downstream circuits. PMID:26170279

  16. Hypocholesterolaemic Activity of Lupin Peptides: Investigation on the Crosstalk between Human Enterocytes and Hepatocytes Using a Co-Culture System Including Caco-2 and HepG2 Cells

    PubMed Central

    Lammi, Carmen; Zanoni, Chiara; Ferruzza, Simonetta; Ranaldi, Giulia; Sambuy, Yula; Arnoldi, Anna

    2016-01-01

    Literature indicates that peptic and tryptic peptides derived from the enzymatic hydrolysis of lupin protein are able to modulate cholesterol metabolism in human hepatic HepG2 cells and that part of these peptides are absorbed in a small intestine model based on differentiated human Caco-2 cells. In this paper, a co-culture system, including Caco-2 and HepG2 cells, was investigated with two objectives: (a) to verify whether cholesterol metabolism in HepG2 cells was modified by the peptides absorption through Caco-2 cells; (b) to investigate how lupin peptides influence cholesterol metabolism in Caco-2 cells. The experiments showed that the absorbed peptides, not only maintained their bioactivity on HepG2 cells, but that this activity was improved by the crosstalk of the two cells systems in co-culture. In addition, lupin peptides showed a positive influence on cholesterol metabolism in Caco-2 cells, decreasing the proprotein convertase subtilisin/kexin type 9 (PCSK9) secretion. PMID:27455315

  17. Hypocholesterolaemic Activity of Lupin Peptides: Investigation on the Crosstalk between Human Enterocytes and Hepatocytes Using a Co-Culture System Including Caco-2 and HepG2 Cells.

    PubMed

    Lammi, Carmen; Zanoni, Chiara; Ferruzza, Simonetta; Ranaldi, Giulia; Sambuy, Yula; Arnoldi, Anna

    2016-01-01

    Literature indicates that peptic and tryptic peptides derived from the enzymatic hydrolysis of lupin protein are able to modulate cholesterol metabolism in human hepatic HepG2 cells and that part of these peptides are absorbed in a small intestine model based on differentiated human Caco-2 cells. In this paper, a co-culture system, including Caco-2 and HepG2 cells, was investigated with two objectives: (a) to verify whether cholesterol metabolism in HepG2 cells was modified by the peptides absorption through Caco-2 cells; (b) to investigate how lupin peptides influence cholesterol metabolism in Caco-2 cells. The experiments showed that the absorbed peptides, not only maintained their bioactivity on HepG2 cells, but that this activity was improved by the crosstalk of the two cells systems in co-culture. In addition, lupin peptides showed a positive influence on cholesterol metabolism in Caco-2 cells, decreasing the proprotein convertase subtilisin/kexin type 9 (PCSK9) secretion. PMID:27455315

  18. Thymoquinone causes multiple effects, including cell death, on dividing plant cells.

    PubMed

    Hassanien, Sameh E; Ramadan, Ahmed M; Azeiz, Ahmed Z Abdel; Mohammed, Rasha A; Hassan, Sabah M; Shokry, Ahmed M; Atef, Ahmed; Kamal, Khalid B H; Rabah, Samar; Sabir, Jamal S M; Abuzinadah, Osama A; El-Domyati, Fotouh M; Martin, Gregory B; Bahieldin, Ahmed

    2013-01-01

    Thymoquinone (TQ) is a major constituent of Nigella sativa oil with reported anti-oxidative activity and anti-inflammatory activity in animal cells. It also inhibits proliferation and induces programmed cell death (apoptosis) in human skin cancer cells. The present study sought to detect the influence of TQ on dividing cells of three plant systems and on expression of Bcl2-associated athanogene-like (BAG-like) genes that might be involved during the process of cell death. BAG genes are known for the regulation of diverse physiological processes in animals, including apoptosis, tumorigenesis, stress responses, and cell division. Synthetic TQ at 0.1mg/mL greatly reduced wheat seed germination rate, whereas 0.2mg/mL completely inhibited germination. An Evans blue assay revealed moderate cell death in the meristematic zone of Glycine max roots after 1h of TQ treatment (0.2mg/mL), with severe cell death occurring in this zone after 2h of treatment. Light microscopy of TQ-treated (0.2mg/mL) onion hairy root tips for 1h revealed anti-mitotic activity and also cell death-associated changes, including nuclear membrane disruption and nuclear fragmentation. Transmission electron microscopy of TQ-treated cells (0.2mg/mL) for 1h revealed shrinkage of the plasma membrane, leakage of cell lysate, degradation of cell walls, enlargement of vacuoles and condensation of nuclei. Expression of one BAG-like gene, previously associated with cell death, was induced 20 min after TQ treatment in Glycine max root tip cells. Thus, TQ has multiple effects, including cell death, on dividing plant cells and plants may serve as a useful system to further investigate the mechanisms underlying the response of eukaryotic cells to TQ. PMID:24296078

  19. General Information about Plasma Cell Neoplasms (Including Multiple Myeloma)

    MedlinePlus

    ... Including Multiple Myeloma) Treatment (PDQ®)–Patient Version General Information About Plasma Cell Neoplasms Go to Health Professional ... the PDQ Adult Treatment Editorial Board . Clinical Trial Information A clinical trial is a study to answer ...

  20. Everglades National Park Including Biscayne National Park. Activity Book.

    ERIC Educational Resources Information Center

    Ruehrwein, Dick

    Intended to help elementary school children learn about the resources of the Everglades and Biscayne National Parks, this activity book includes information, puzzles, games, and quizzes. The booklet deals with concepts related to: (1) the seasons; (2) fire ecology; (3) water; (4) fish; (5) mammals; (6) mosquitos; (7) birds; (8) venomous snakes;…

  1. Fuel cell repeater unit including frame and separator plate

    DOEpatents

    Yamanis, Jean; Hawkes, Justin R; Chiapetta, Jr., Louis; Bird, Connie E; Sun, Ellen Y; Croteau, Paul F

    2013-11-05

    An example fuel cell repeater includes a separator plate and a frame establishing at least a portion of a flow path that is operative to communicate fuel to or from at least one fuel cell held by the frame relative to the separator plate. The flow path has a perimeter and any fuel within the perimeter flow across the at least one fuel cell in a first direction. The separator plate, the frame, or both establish at least one conduit positioned outside the flow path perimeter. The conduit is outside of the flow path perimeter and is configured to direct flow in a second, different direction. The conduit is fluidly coupled with the flow path.

  2. Electrolytes including fluorinated solvents for use in electrochemical cells

    SciTech Connect

    Tikhonov, Konstantin; Yip, Ka Ki; Lin, Tzu-Yuan

    2015-07-07

    Provided are electrochemical cells and electrolytes used to build such cells. The electrolytes include ion-supplying salts and fluorinated solvents capable of maintaining single phase solutions with the salts at between about -30.degree. C. to about 80.degree. C. The fluorinated solvents, such as fluorinated carbonates, fluorinated esters, and fluorinated esters, are less flammable than their non-fluorinated counterparts and increase safety characteristics of cells containing these solvents. The amount of fluorinated solvents in electrolytes may be between about 30% and 80% by weight not accounting weight of the salts. Fluorinated salts, such as fluoroalkyl-substituted LiPF.sub.6, fluoroalkyl-substituted LiBF.sub.4 salts, linear and cyclic imide salts as well as methide salts including fluorinated alkyl groups, may be used due to their solubility in the fluorinated solvents. In some embodiments, the electrolyte may also include a flame retardant, such as a phosphazene or, more specifically, a cyclic phosphazene and/or one or more ionic liquids.

  3. Stages of Plasma Cell Neoplasms (Including Multiple Myeloma)

    MedlinePlus

    ... Neoplasms for more information. High-dose chemotherapy with stem cell transplant This treatment is a way of giving ... blood -forming cells destroyed by the cancer treatment. Stem cells (immature blood cells) are removed from the blood ...

  4. Treatment Options for Plasma Cell Neoplasms (Including Multiple Myeloma)

    MedlinePlus

    ... Neoplasms for more information. High-dose chemotherapy with stem cell transplant This treatment is a way of giving ... blood -forming cells destroyed by the cancer treatment. Stem cells (immature blood cells) are removed from the blood ...

  5. Treatment Option Overview (Plasma Cell Neoplasms Including Multiple Myeloma)

    MedlinePlus

    ... Neoplasms for more information. High-dose chemotherapy with stem cell transplant This treatment is a way of giving ... blood -forming cells destroyed by the cancer treatment. Stem cells (immature blood cells) are removed from the blood ...

  6. Multiple cell photoresponsive amorphous photo voltaic devices including graded ban gaps

    SciTech Connect

    Ovshinsky, S.R.; Adler, D.

    1990-09-04

    This patent describes an improved photoresponsive tandem multiple cell device. It comprises: at least first and second superimposed solar cells; the first cell being formed of an amorphous silicon alloy material; the second amorphous silicon alloy cell having an active photoresponsive region in which radiation can impinge to produce charge carriers. The amorphous silicon alloy cell body including at least one element for reducing the density of defect states to about 10{sup 16} defects per cubic centimeter and a band gap adjusting element graded through at least a portion of the photoresponsive region thereof to enhance the radiation absorption; the adjusting element being germanium, and the band gap of the cell being adjusted for a specified photoresponse wavelength threshold function different from the first cell; the second cell being a multi-layer body having deposited silicon alloy layers of opposite (p and n) conductivity type; and the first cell being formed with the second cell in substantially direct junction contact therebetween.

  7. Solar cell activation system

    SciTech Connect

    Apelian, L.

    1983-07-05

    A system for activating solar cells involves the use of phosphorescent paint, the light from which is amplified by a thin magnifying lens and used to activate solar cells. In a typical system, a member painted with phosphorescent paint is mounted adjacent a thin magnifying lens which focuses the light on a predetermined array of sensitive cells such as selenium, cadmium or silicon, mounted on a plastic board. A one-sided mirror is mounted adjacent the cells to reflect the light back onto said cells for purposes of further intensification. The cells may be coupled to rechargeable batteries or used to directly power a small radio or watch.

  8. STEREOLOGICAL ESTIMATES OF THE BASAL FOREBRAIN CELL POPULATION IN THE RAT, INCLUDING NEURONS CONTAINING CHOLINE ACETYLTRANSFERASE (ChAT), GLUTAMIC ACID DECARBOXYLASE (GAD) OR PHOSPHATE-ACTIVATED GLUTAMINASE (PAG) AND COLOCALIZING VESICULAR GLUTAMATE TRANSPORTERS (VGluTs)

    PubMed Central

    GRITTI, I.; HENNY, P.; GALLONI, F.; MAINVILLE, L.; MARIOTTI, M.; JONES, B. E.

    2006-01-01

    The basal forebrain (BF) plays an important role in modulating cortical activity and influencing attention, learning and memory. These activities are fulfilled importantly yet not entirely by cholinergic neurons. Noncholinergic neurons also contribute and are comprised by GABAergic neurons and other possibly glutamatergic neurons. The aim of the present study was to estimate the total number of cells in the BF of the rat and the proportions of that total represented by cholinergic, GABAergic and glutamatergic neurons. For this purpose, cells were counted using unbiased stereological methods within the medial septum, diagonal band, magnocellular preoptic nucleus, substantia innominata and globus pallidus in sections stained for Nissl substance and/or the neurotransmitter enzymes, choline acetyltransferase (ChAT), glutamic acid decarboxylase (GAD) or phosphate-activated glutaminase (PAG). In Nissl-stained sections, the total number of neurons in the BF was estimated as ~355,000 and the numbers of ChAT-immuno-positive (+) as ~22,000, GAD+ ~119,000 and PAG+ ~316,000, corresponding to ~5%, ~35% and ~90% of the total. Thus, of the large population of BF neurons, only a small proportion has the capacity to synthesize acetylcholine (ACh), one third to synthesize GABA and the vast majority to synthesize glutamate (Glu). Moreover, through the presence of PAG, a proportion of ACh- and GABA-synthesizing neurons also have the capacity to synthesize Glu. In sections dual fluorescent immunostained for vesicular transporters, VGluT3 and not VGluT2 was present in the cell bodies of most PAG+ and ChAT+ and half the GAD+ cells. Given previous results showing that VGluT2 and not VGluT3 was present in BF axon terminals and not colocalized with VAChT or VGAT, we conclude that the BF cell population influences cortical and subcortical regions through neurons which release ACh, GABA or Glu from their terminals but which in part can also synthesize and release Glu from their soma or

  9. The Histone Demethylase Jumonji Coordinates Cellular Senescence Including Secretion of Neural Stem Cell-attracting Cytokines

    PubMed Central

    Perrigue, Patrick M.; Silva, Michael E.; Warden, Charles D.; Feng, Nathan L.; Reid, Michael A.; Mota, Daniel J.; Joseph, Lauren P.; Tian, Yangzi Isabel; Glackin, Carlotta A.; Gutova, Margarita; Najbauer, Joseph; Aboody, Karen S.; Barish, Michael E.

    2016-01-01

    Jumonji domain-containing protein 3 (JMJD3/KDM6B) demethylates lysine 27 on histone H3 (H3K27me3), a repressive epigenetic mark controlling chromatin organization and cellular senescence. To better understand the functional consequences of JMJD3 its expression was investigated in brain tumor cells. Querying patient expression profile databases confirmed JMJD3 over-expression in high-grade glioma. Immunochemical staining of two glioma cell lines, U251 and U87, indicated intrinsic differences in JMJD3 expression levels that were reflected in changes in cell phenotype and variations associated with cellular senescence, including senescence-associated β-galactosidase (SA-β-gal) activity and the senescence associated secretory phenotype (SASP). Over-expressing wild type JMJD3 (JMJD3wt) activated SASP-associated genes, enhanced SA-βgal activity, and induced nuclear blebbing. Conversely, over-expression of a catalytically inactive dominant negative mutant JMJD3 (JMJD3mut) increased proliferation. In addition, a large number of transcripts were identified by RNA-seq as altered in JMJD3 over-expressing cells, including cancer- and inflammation-related transcripts as defined by IPA analysis. These results suggest that expression of the SASP in the context of cancer undermines normal tissue homeostasis and contributes to tumorigenesis and tumor progression. These studies are therapeutically relevant because inflammatory cytokines have been linked to homing of neural stem cells and other stem cells to tumor loci. PMID:25652587

  10. Antiviral activity of 1-docosanol, an inhibitor of lipid-enveloped viruses including herpes simplex.

    PubMed Central

    Katz, D H; Marcelletti, J F; Khalil, M H; Pope, L E; Katz, L R

    1991-01-01

    This article reports that 1-docosanol, a 22-carbon-long saturated alcohol, exerts a substantial inhibitory effect on replication of certain viruses (e.g., herpes simplex virus and respiratory syncytial virus) within primary target cells in vitro. To study the basis for its viral inhibitory activity, a suspension of 1-docosanol was formulated in an inert and nontoxic surfactant, Pluronic F-68; this suspension exerted potent inhibitory activity on the ability of susceptible viruses to infect cultured target cells. Susceptible viruses included wild-type herpes simplex viruses 1 and 2 as well as acyclovir-resistant herpes simplex virus 2 and also respiratory syncytial virus--all of which are lipid-enveloped. In contrast, nonenveloped poliovirus was not susceptible to the inhibitory action of 1-docosanol. Although the precise mechanism has yet to be defined, current evidence suggests that 1-docosanol inhibits viral replication by interfering with the early intracellular events surrounding viral entry into target cells. It is possible that interaction between the highly lipophilic compound and components of target cell membranes renders such target cells less susceptible to viral fusion and/or entry. If this mechanism proves to be correct, 1-docosanol may provide a broad spectrum activity against many different viruses, especially those with lipid-containing envelopes. Images PMID:1660151

  11. A POROELASTIC MODEL FOR CELL CRAWLING INCLUDING MECHANICAL COUPLING BETWEEN CYTOSKELETAL CONTRACTION AND ACTIN POLYMERIZATION.

    PubMed

    Taber, L A; Shi, Y; Yang, L; Bayly, P V

    2011-01-01

    Much is known about the biophysical mechanisms involved in cell crawling, but how these processes are coordinated to produce directed motion is not well understood. Here, we propose a new hypothesis whereby local cytoskeletal contraction generates fluid flow through the lamellipodium, with the pressure at the front of the cell facilitating actin polymerization which pushes the leading edge forward. The contraction, in turn, is regulated by stress in the cytoskeleton. To test this hypothesis, finite element models for a crawling cell are presented. These models are based on nonlinear poroelasticity theory, modified to include the effects of active contraction and growth, which are regulated by mechanical feedback laws. Results from the models agree reasonably well with published experimental data for cell speed, actin flow, and cytoskeletal deformation in migrating fish epidermal keratocytes. The models also suggest that oscillations can occur for certain ranges of parameter values. PMID:21765817

  12. Measuring energy metabolism in cultured cells, including human pluripotent stem cells and differentiated cells

    PubMed Central

    Zhang, Jin; Nuebel, Esther; Wisidagama, Dona R R; Setoguchi, Kiyoko; Hong, Jason S; Van Horn, Christine M; Imam, Sarah S; Vergnes, Laurent; Malone, Cindy S; Koehler, Carla M; Teitell, Michael A

    2013-01-01

    Measurements of glycolysis and mitochondrial function are required to quantify energy metabolism in a wide variety of cellular contexts. In human pluripotent stem cells (hPSCs) and their differentiated progeny, this analysis can be challenging because of the unique cell properties, growth conditions and expense required to maintain these cell types. Here we provide protocols for analyzing energy metabolism in hPSCs and their early differentiated progenies that are generally applicable to mature cell types as well. Our approach has revealed distinct energy metabolism profiles used by hPSCs, differentiated cells, a variety of cancer cells and Rho-null cells. The protocols measure or estimate glycolysis on the basis of the extracellular acidification rate, and they measure or estimate oxidative phosphorylation on the basis of the oxygen consumption rate. Assays typically require 3 h after overnight sample preparation. Companion methods are also discussed and provided to aid researchers in developing more sophisticated experimental regimens for extended analyses of cellular bioenergetics. PMID:22576106

  13. Optimal design of active and semi-active suspensions including time delays and preview

    NASA Astrophysics Data System (ADS)

    Hac', A.; Youn, I.

    1993-10-01

    Several control laws for active and semi-active suspension based on a linear half car model are derived and investigated. The strategies proposed take full advantage of the fact that the road input to the rear wheels is a delayed version of that to the front wheels, which in turn can be obtained either from the measurements of the front wheels and body motions or by direct preview of road irregularities if preview sensors are available. The suspension systems are optimized with respect to ride comfort, road holding and suspension rattle space as expressed by the mean-square-values of body acceleration (including effects of heave and pitch), tire deflections and front and rear suspension travels. The optimal control laws that minimize the given performance index and include passivity constraints in the semi-active case are derived using calculus of variation. The optimal semi-active suspension becomes piecewise linear, varying between passive and fully active systems and combinations of them. The performances of active and semi-active systems with and without preview were evaluated by numerical simulation in the time and frequency domains. The results show that incorporation of time delay between the front and rear axles in controller design improves the dynamic behavior of the rear axle and control of body pitch motion, while additional preview improves front wheel dynamics and body heave.

  14. Measuring and Reducing Off-Target Activities of Programmable Nucleases Including CRISPR-Cas9.

    PubMed

    Koo, Taeyoung; Lee, Jungjoon; Kim, Jin-Soo

    2015-06-01

    Programmable nucleases, which include zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and RNA-guided engineered nucleases (RGENs) repurposed from the type II clustered, regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9) system are now widely used for genome editing in higher eukaryotic cells and whole organisms, revolutionising almost every discipline in biological research, medicine, and biotechnology. All of these nucleases, however, induce off-target mutations at sites homologous in sequence with on-target sites, limiting their utility in many applications including gene or cell therapy. In this review, we compare methods for detecting nuclease off-target mutations. We also review methods for profiling genome-wide off-target effects and discuss how to reduce or avoid off-target mutations. PMID:25985872

  15. A full model for simulation of electrochemical cells including complex behavior

    NASA Astrophysics Data System (ADS)

    Esperilla, J. J.; Félez, J.; Romero, G.; Carretero, A.

    This communication presents a model of electrochemical cells developed in order to simulate their electrical, chemical and thermal behavior showing the differences when thermal effects are or not considered in the charge-discharge process. The work presented here has been applied to the particular case of the Pb,PbSO 4|H 2SO 4 (aq)|PbO 2,Pb cell, which forms the basis of the lead-acid batteries so widely used in the automotive industry and as traction batteries in electric or hybrid vehicles. Each half-cell is considered independently in the model. For each half-cell, in addition to the main electrode reaction, a secondary reaction is considered: the hydrogen evolution reaction in the negative electrode and the oxygen evolution reaction in the positive. The equilibrium potential is calculated with the Nernst equation, in which the activity coefficients are fitted to an exponential function using experimental data. On the other hand, the two main mechanisms that produce the overpotential are considered, that is the activation or charge transfer and the diffusion mechanisms. First, an isothermal model has been studied in order to show the behavior of the main phenomena. A more complex model has also been studied including thermal behavior. This model is very useful in the case of traction batteries in electric and hybrid vehicles where high current intensities appear. Some simulation results are also presented in order to show the accuracy of the proposed models.

  16. Mitochondrial ROS fire up T cell activation.

    PubMed

    Murphy, Michael P; Siegel, Richard M

    2013-02-21

    Metabolic reprogramming has emerged as an important feature of immune cell activation. Two new studies, including Sena et al. (2013) in this issue of Immunity, identify mitochondrial reactive oxygen species (ROS) arising from metabolic reprogramming as signaling molecules in T cell activation. PMID:23438817

  17. Myosin II Activity Softens Cells in Suspension

    PubMed Central

    Chan, Chii J.; Ekpenyong, Andrew E.; Golfier, Stefan; Li, Wenhong; Chalut, Kevin J.; Otto, Oliver; Elgeti, Jens; Guck, Jochen; Lautenschläger, Franziska

    2015-01-01

    The cellular cytoskeleton is crucial for many cellular functions such as cell motility and wound healing, as well as other processes that require shape change or force generation. Actin is one cytoskeleton component that regulates cell mechanics. Important properties driving this regulation include the amount of actin, its level of cross-linking, and its coordination with the activity of specific molecular motors like myosin. While studies investigating the contribution of myosin activity to cell mechanics have been performed on cells attached to a substrate, we investigated mechanical properties of cells in suspension. To do this, we used multiple probes for cell mechanics including a microfluidic optical stretcher, a microfluidic microcirculation mimetic, and real-time deformability cytometry. We found that nonadherent blood cells, cells arrested in mitosis, and naturally adherent cells brought into suspension, stiffen and become more solidlike upon myosin inhibition across multiple timescales (milliseconds to minutes). Our results hold across several pharmacological and genetic perturbations targeting myosin. Our findings suggest that myosin II activity contributes to increased whole-cell compliance and fluidity. This finding is contrary to what has been reported for cells attached to a substrate, which stiffen via active myosin driven prestress. Our results establish the importance of myosin II as an active component in modulating suspended cell mechanics, with a functional role distinctly different from that for substrate-adhered cells. PMID:25902426

  18. Myosin II Activity Softens Cells in Suspension.

    PubMed

    Chan, Chii J; Ekpenyong, Andrew E; Golfier, Stefan; Li, Wenhong; Chalut, Kevin J; Otto, Oliver; Elgeti, Jens; Guck, Jochen; Lautenschläger, Franziska

    2015-04-21

    The cellular cytoskeleton is crucial for many cellular functions such as cell motility and wound healing, as well as other processes that require shape change or force generation. Actin is one cytoskeleton component that regulates cell mechanics. Important properties driving this regulation include the amount of actin, its level of cross-linking, and its coordination with the activity of specific molecular motors like myosin. While studies investigating the contribution of myosin activity to cell mechanics have been performed on cells attached to a substrate, we investigated mechanical properties of cells in suspension. To do this, we used multiple probes for cell mechanics including a microfluidic optical stretcher, a microfluidic microcirculation mimetic, and real-time deformability cytometry. We found that nonadherent blood cells, cells arrested in mitosis, and naturally adherent cells brought into suspension, stiffen and become more solidlike upon myosin inhibition across multiple timescales (milliseconds to minutes). Our results hold across several pharmacological and genetic perturbations targeting myosin. Our findings suggest that myosin II activity contributes to increased whole-cell compliance and fluidity. This finding is contrary to what has been reported for cells attached to a substrate, which stiffen via active myosin driven prestress. Our results establish the importance of myosin II as an active component in modulating suspended cell mechanics, with a functional role distinctly different from that for substrate-adhered cells. PMID:25902426

  19. Fluorescence activated cell sorting.

    NASA Technical Reports Server (NTRS)

    Bonner, W. A.; Hulett, H. R.; Sweet, R. G.; Herzenberg, L. A.

    1972-01-01

    An instrument has been developed for sorting biological cells. The cells are rendered differentially fluorescent and incorporated into a small liquid stream illuminated by a laser beam. The cells pass sequentially through the beam, and fluorescent light from the cells gives rise to electrical signals. The stream is broken into a series of uniform size drops downstream of the laser. The cell signals are used to give appropriate electrostatic charges to drops containing the cells. The drops then pass between two charged plates and are deflected to appropriate containers. The system has proved capable of providing fractions containing large numbers of viable cells highly enriched in a particular functional type.

  20. Stem cell tracking with optically active nanoparticles

    PubMed Central

    Gao, Yu; Cui, Yan; Chan, Jerry KY; Xu, Chenjie

    2013-01-01

    Stem-cell-based therapies hold promise and potential to address many unmet clinical needs. Cell tracking with modern imaging modalities offers insight into the underlying biological process of the stem-cell-based therapies, with the goal to reveal cell survival, migration, homing, engraftment, differentiation, and functions. Adaptability, sensitivity, resolution, and non-invasiveness have contributed to the longstanding use of optical imaging for stem cell tracking and analysis. To identify transplanted stem cells from the host tissue, optically active probes are usually used to label stem cells before the administration. In comparison to the traditional fluorescent probes like fluorescent proteins and dyes, nanoparticle-based probes are advantageous in terms of the photo-stabilities and minimal changes to the cell phenotype. The main focus here is to overview the recent development of optically active nanoparticles for stem cells tracking. The related optical imaging modalities include fluorescence imaging, photoacoustic imaging, Raman and surface enhanced Raman spectroscopy imaging. PMID:23638335

  1. A simple method for in situ freezing of anchorage-dependent cells including rat liver parenchymal cells.

    PubMed

    Ohno, T; Saijo-Kurita, K; Miyamoto-Eimori, N; Kurose, T; Aoki, Y; Yosimura, S

    1991-03-01

    We developed a simple method for freezing anchorage-dependent cells, including primary cultured rat liver parenchymal cells, without detaching the cells from the culture dish. The method consists of preculture of the cells to confluence, changing the growth medium to a conventional freezing medium, packaging in a container, and storage at -80 degrees C. After thawing and changing the freezing medium to regular growth medium, cell growth was nearly identical to that of cells freshly seeded into a new dish. PMID:1367380

  2. Advances in the theory and application of BSF cells. [including electrical resistivity and photovoltaic cells

    NASA Technical Reports Server (NTRS)

    Mandelkorn, J.; Lamneck, J. H.

    1975-01-01

    The characteristics and behavior of p(+), p solar cells were investigated. The p(+), p cells were made by the removal of the n(+) surface layers from n(+), p p(+), BSF cells followed by application of a suitable contact to the resultant p(+), p structures. The open circuit voltage of p(+), p cells was found to increase with increasing 'p' bulk resistivity. The measured open circuit velocity-temperature coefficients were positive and increased with increasing resistivity. An outline of prior limitations in solar cell design is presented, and the removal of these limitations through use of BSF effects is pointed out. The study of BSF effects made feasible production of very thin high efficiency silicon cells as well as high resistivity-high efficiency cells, two desirable types of silicon cells which were previously impossible to make.

  3. Colon-targeted delivery of live bacterial cell biotherapeutics including microencapsulated live bacterial cells

    PubMed Central

    Prakash, Satya; Malgorzata Urbanska, Aleksandra

    2008-01-01

    There has been an ample interest in delivery of therapeutic molecules using live cells. Oral delivery has been stipulated as best way to deliver live cells to humans for therapy. Colon, in particular, is a part of gastrointestinal (GI) tract that has been proposed to be an oral targeted site. The main objective of these oral therapy procedures is to deliver live cells not only to treat diseases like colorectal cancer, inflammatory bowel disease, and other GI tract diseases like intestinal obstruction and gastritis, but also to deliver therapeutic molecules for overall therapy in various diseases such as renal failure, coronary heart disease, hypertension, and others. This review provides a comprehensive summary of recent advancement in colon targeted live bacterial cell biotherapeutics. Current status of bacterial cell therapy, principles of artificial cells and its potentials in oral delivery of live bacterial cell biotherapeutics for clinical applications as well as biotherapeutic future perspectives are also discussed in our review. PMID:19707368

  4. 7 CFR 981.441 - Credit for market promotion activities, including paid advertising.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... promotion activities, including paid advertising. (a) In order for a handler to receive credit for his/her own promotional activities from his/her pro rata portion of advertising assessment payments, pursuant... professional practices and rates for the type of activity conducted. In the case of claims for...

  5. 7 CFR 981.441 - Credit for market promotion activities, including paid advertising.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... promotion activities, including paid advertising. (a) In order for a handler to receive credit for his/her own promotional activities from his/her pro rata portion of advertising assessment payments, pursuant... professional practices and rates for the type of activity conducted. In the case of claims for...

  6. 7 CFR 981.441 - Credit for market promotion activities, including paid advertising.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... promotion activities, including paid advertising. (a) In order for a handler to receive credit for his/her own promotional activities from his/her pro rata portion of advertising assessment payments, pursuant... professional practices and rates for the type of activity conducted. In the case of claims for...

  7. 7 CFR 981.441 - Credit for market promotion activities, including paid advertising.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... promotion activities, including paid advertising. (a) In order for a handler to receive credit for his/her own promotional activities from his/her pro rata portion of advertising assessment payments, pursuant... professional practices and rates for the type of activity conducted. In the case of claims for...

  8. Methods for the measurement of cell and tissue compatibility including tissue regeneration processes

    PubMed Central

    Wiegand, Cornelia; Hipler, Uta-Christina

    2008-01-01

    Biocompatibility is one of the main requirements for the safe use of medical devices. Determination of cytotoxicity is part of the initial evaluation stipulated by ISO standards for the biological evaluation of medical devices. The use of cell cultures to test the biocompatibility of drugs, biomaterials or treatment techniques used in various disciplines is gaining in importance. A wide variety of self-initiated and commercially available cell lines has been evaluated and used: cultured fibroblasts from human skin, buccal mucosa, periodontal membrane, embryonic lung, epithelial and HeLa cells; cultures of human keratinocytes and HaCaT cells; different murine cell lines (C3H-L, Balb/c 3T3, L929 and others) as well as murine cells cultured from liver and spleen; T-lymphocytes from lymph nodes and macrophages obtained by lavage. All of the above cells are suitable for use in biocompatibility tests. Nevertheless, the general opinion is that toxicity tests in vitro will be more convincing when performed with cells that are homologous with the human tissue concerned. In accordance, appropriate cell lines for use in cytotoxicity and tolerance tests concerning the skin would be human dermal fibroblasts and human epidermal keratinocytes, as they take an active part in the immune response, inflammatory processes, and wound healing. The evaluation of the in vitro cytotoxicity of a biomaterial is often a qualitative analysis based on the morphological examination of cell damage and growth after direct or indirect contact with the material. Different commercial assays based on the determination of nucleic acids, metabolic activity, protein content or membrane integrity are available to measure cell proliferation and cell viability. A small selection – Pico Green® DNA Cell Proliferation Assay, ATPLite™ Luminescence ATP Detection Assay, BC Assay: protein quantitation kit, AlamarBlue™ Proliferation Assay and Live/Dead Staining with SYTO-13 and EthD-2 – are discussed

  9. Improvements to the FATOLA computer program including added actively controlled landing gear subroutines

    NASA Technical Reports Server (NTRS)

    Mall, G. H.

    1983-01-01

    Modifications to a multi-degree-of-freedom flexible aircraft take-off and landing analysis (FATOLA) computer program, including a provision for actively controlled landing gears to expand the programs simulation capabilities, are presented. Supplemental instructions for preparation of data and for use of the modified program are included.

  10. Extension to PV OPTICS to include front electrode design in solar cells

    NASA Astrophysics Data System (ADS)

    Guhabiswas, Debraj

    Proper optical designing of solar cells and modules is of paramount importance towards achieving high photovoltaic conversion efficiencies. Modeling softwares such as PV OPTICS, BIRANDY and SUNRAYS have been created to aid such optical designing of cells and modules; but none of these modeling packages take the front metal electrode architecture of a solar cell into account. A new model, has been developed to include the front metal electrode architecture to finished solar cells for optical calculations. This has been implemented in C++ in order to add a new module to PV OPTICS (NREL's photovoltaic modeling tool) to include front metallization patterns for optical design and simulation of solar cells. This new addition also calculates the contribution of light that diffuses out of the illuminated (non-metallized) regions to the solar cell current. It also determines the optical loss caused by the absorption in the front metal and separates metallic losses due to front and back contacts. This added capability also performs the following functions: • calculates the total current that can be generated in a solar cell due to optical absorption in each region, including the region beneath the front metal electrodes for the radiation spectrum of AM 1.5, • calculates various losses in the solar cell due to front electrode shading, metal absorption, and reflectance, • makes a plot of how light is absorbed in the metal as well as silicon under the shaded region in the solar cell. Although Finite Difference Time Domain (FDTD) is the numerical technique of choice to solve Maxwell's equations for a propagating electromagnetic wave, it is both time consuming and very demanding on the computer processors. Furthermore, for complicated geometric structures, FDTD poses various limitations. Hence, ray tracing has been chosen as the means of implementing this new model. This new software has been used to carry out a detailed investigation on the effect of various parameters of

  11. Intracellular activity of clinical concentrations of phenothiazines including thioridiazine against phagocytosed Staphylococcus aureus.

    PubMed

    Ordway, Diane; Viveiros, Miguel; Leandro, Clara; Arroz, Maria Jorge; Amaral, Leonard

    2002-07-01

    The effect of thioridazine (TZ) was studied on the killing activity of human peripheral blood monocyte derived macrophages (HPBMDM) and of human macrophage cell line THP-1 at extracellular concentrations below those achievable clinically. These macrophages have nominal killing activity against bacteria and therefore, would not influence any activity that the compounds may have against intracellular localised Staphylococcus aureus. The results indicated that whereas TZ has an in vitro minimum inhibitory concentration (MIC) against the strains of S. aureus of 18, 0.1 mg/l of TZ in the medium completely inhibits the growth of S. aureus that has been phagocytosed by macrophages. The latter concentration was non-toxic to macrophages, did not cause cellular expression of activation marker CD69 nor induction of CD3+ T cell production of IFN-gamma, but blocked cellular proliferation and down-regulated the production of T cell-derived cytokines (IFN-gamma, IL-5). These results suggest that TZ induces intracellular bactericidal activities independent of the capacity to generate Type 1 responses against S. aureus. PMID:12127709

  12. Fabrication of contacts for silicon solar cells including printing burn through layers

    SciTech Connect

    Ginley, David S; Kaydanova, Tatiana; Miedaner, Alexander; Curtis, Calvin J; Van Hest, Marinus Franciscus Antonius Maria

    2014-06-24

    A method for fabricating a contact (240) for a solar cell (200). The method includes providing a solar cell substrate (210) with a surface that is covered or includes an antireflective coating (220). For example, the substrate (210) may be positioned adjacent or proximate to an outlet of an inkjet printer (712) or other deposition device. The method continues with forming a burn through layer (230) on the coating (220) by depositing a metal oxide precursor (e.g., using an inkjet or other non-contact printing method to print or apply a volume of liquid or solution containing the precursor). The method includes forming a contact layer (240) comprising silver over or on the burn through layer (230), and then annealing is performed to electrically connect the contact layer (240) to the surface of the solar cell substrate (210) through a portion of the burn through layer (230) and the coating (220).

  13. Liquid-Liquid Displacement Flows in a Hele-Shaw Cell including Viscoplastic Effects

    NASA Astrophysics Data System (ADS)

    de Souza Mendes, Paulo R.; Varges, Priscilla R.

    2008-07-01

    Viscous fingering in non-Newtonian fluids in a rectangular Hele-Shaw cell is investigated. This cell is filled with aqueous solutions of carbopol in two different concentrations. A Newtonian mineral oil is then injected into the cell and the displacement is observed. A digital camera is used to capture images of the interface between the fluids during the flow. Applications include displacement of heavy crude oil in reservoirs. The main parameters that govern this flow are the viscosity ratio, the rheological capillary number, and the (dimensionless) flow rate. The interface shape is given for two different values of flow rate and viscosity ratio.

  14. Activated mast cells promote differentiation of B cells into effector cells

    PubMed Central

    Palm, Anna-Karin E.; Garcia-Faroldi, Gianni; Lundberg, Marcus; Pejler, Gunnar; Kleinau, Sandra

    2016-01-01

    Based on the known accumulation of mast cells (MCs) in B cell-dependent inflammatory diseases, including rheumatoid arthritis, we hypothesized that MCs directly modulate B cells. We show here that degranulated, and to a lesser extent naïve or IgE-sensitized, MCs activate both naïve and B cell receptor-activated B cells. This was shown by increased proliferation, blast formation, and expression of CD19, MHC class II and CD86 in the B cells. Further, MCs stimulated the secretion of IgM and IgG in IgM+ B cells, indicating that MCs can induce class-switch recombination in B cells. We also show that coculture of MCs with B cells promotes surface expression of L-selectin, a homing receptor, on the B cells. The effects of MCs on B cells were partly dependent on cell-cell contact and both follicular and marginal zone B cells could be activated by MCs. Our findings suggest that degranulated MCs support optimal activation of B cells, a finding that is in line with in vivo studies showing that MCs frequently degranulate in the context of B-cell driven pathologies such as arthritis. Together, our findings show that MCs have the capacity to differentiate B cells to effector cells. PMID:26847186

  15. Universal cell frame for high-pressure water electrolyzer and electrolyzer including the same

    DOEpatents

    Schmitt, Edwin W.; Norman, Timothy J.

    2013-01-08

    Universal cell frame generic for use as an anode frame and as a cathode frame in a water electrolyzer. According to one embodiment, the universal cell frame includes a unitary annular member having a central opening. Four trios of transverse openings are provided in the annular member, each trio being spaced apart by about 90 degrees. A plurality of internal radial passageways fluidly interconnect the central opening and each of the transverse openings of two diametrically-opposed trios of openings, the other two trios of openings lacking corresponding radial passageways. Sealing ribs are provided on the top and bottom surfaces of the annular member. The present invention is also directed at a water electrolyzer that includes two such cell frames, one being used as the anode frame and the other being used as the cathode frame, the cathode frame being rotated 90 degrees relative to the anode frame.

  16. A&M. TAN607. Special service cubicle (hot cell). Details include Zpipe ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    A&M. TAN-607. Special service cubicle (hot cell). Details include Z-pipe and stepped plug penetrations through shielding wall. Ralph M. Parsons 902-3-ANP-607-A116. Date: December 1952. Approved by INEEL Classification Office for public release. INEEL index code no. 034-0607-693-106767 - Idaho National Engineering Laboratory, Test Area North, Scoville, Butte County, ID

  17. The spindle-shaped cells in cutaneous Kaposi's sarcoma. Histologic simulators include factor XIIIa dermal dendrocytes.

    PubMed Central

    Nickoloff, B. J.; Griffiths, C. E.

    1989-01-01

    Kaposi's sarcoma is a neoplasm that develops as multifocal lesions, often involving the skin, characterized by a complex histologic picture including numerous vascular spaces, perivascular and interstitial spindle-shaped cells, and extravasated erythrocytes, lymphocytes, and plasma cells. Using an antibody against factor XIIIa, which identifies dermal dendrocytes, numerous factor XIIIa-positive dermal dendrocytes were detected among the spindle-shaped cells in 12 acquired immune deficiency syndrome (AIDS)-associated, and five non-AIDS-associated Kaposi's sarcoma lesions. The factor XIIIa-positive dermal dendrocytes were also increased in histologic simulators of Kaposi's sarcoma such as dermatofibroma, angiomatoid malignant fibrous histiocytoma, granuloma annulare, and early wound healing, but were absent in keloids. The increased number of dermal dendrocytes, which are often in an angiocentric configuration and which also express CD4, lymphocyte function associated antigen-1 (LFA-1), and Leu M3 in Kaposi's sarcoma, may be important to the angioproliferative response. The results suggested that the spindle-shaped cells that are present in a variety of cutaneous lesions are dermal dendrocytes and belong to the reticuloendothelial system, unlike other mesenchymal cell types such as the endothelial cell. Apparently a diverse array of stimuli, including human immunodeficiency virus type-1 (HIV-1) infection and trauma, can stimulate the accumulation of factor XIIIa expressing dermal dendrocytes in the skin. These cells can then participate in different stages of a variety of cutaneous alterations including Kaposi's sarcoma, dermatofibroma, granuloma annulare, and early wound healing. Thus, the factor XIIIa-positive dermal dendrocyte is a common cellular denominator among diverse clinical entities that share some histologic features. Images Figure 1 Figure 2 Figure 3 p797-a PMID:2573283

  18. The spindle-shaped cells in cutaneous Kaposi's sarcoma. Histologic simulators include factor XIIIa dermal dendrocytes.

    PubMed

    Nickoloff, B J; Griffiths, C E

    1989-11-01

    Kaposi's sarcoma is a neoplasm that develops as multifocal lesions, often involving the skin, characterized by a complex histologic picture including numerous vascular spaces, perivascular and interstitial spindle-shaped cells, and extravasated erythrocytes, lymphocytes, and plasma cells. Using an antibody against factor XIIIa, which identifies dermal dendrocytes, numerous factor XIIIa-positive dermal dendrocytes were detected among the spindle-shaped cells in 12 acquired immune deficiency syndrome (AIDS)-associated, and five non-AIDS-associated Kaposi's sarcoma lesions. The factor XIIIa-positive dermal dendrocytes were also increased in histologic simulators of Kaposi's sarcoma such as dermatofibroma, angiomatoid malignant fibrous histiocytoma, granuloma annulare, and early wound healing, but were absent in keloids. The increased number of dermal dendrocytes, which are often in an angiocentric configuration and which also express CD4, lymphocyte function associated antigen-1 (LFA-1), and Leu M3 in Kaposi's sarcoma, may be important to the angioproliferative response. The results suggested that the spindle-shaped cells that are present in a variety of cutaneous lesions are dermal dendrocytes and belong to the reticuloendothelial system, unlike other mesenchymal cell types such as the endothelial cell. Apparently a diverse array of stimuli, including human immunodeficiency virus type-1 (HIV-1) infection and trauma, can stimulate the accumulation of factor XIIIa expressing dermal dendrocytes in the skin. These cells can then participate in different stages of a variety of cutaneous alterations including Kaposi's sarcoma, dermatofibroma, granuloma annulare, and early wound healing. Thus, the factor XIIIa-positive dermal dendrocyte is a common cellular denominator among diverse clinical entities that share some histologic features. PMID:2573283

  19. A multiscale model for glioma spread including cell-tissue interactions and proliferation.

    PubMed

    Engwer, Christian; Knappitsch, Markus; Surulescu, Christina

    2016-04-01

    Glioma is a broad class of brain and spinal cord tumors arising from glia cells, which are the main brain cells that can develop into neoplasms. They are highly invasive and lead to irregular tumor margins which are not precisely identifiable by medical imaging, thus rendering a precise enough resection very difficult. The understanding of glioma spread patterns is hence essential for both radiological therapy as well as surgical treatment. In this paper we propose a multiscale model for glioma growth including interactions of the cells with the underlying tissue network, along with proliferative effects. Our current accounting for two subpopulations of cells to accomodate proliferation according to the go-or-grow dichtomoty is an extension of the setting in [16]. As in that paper, we assume that cancer cells use neuronal fiber tracts as invasive pathways. Hence, the individual structure of brain tissue seems to be decisive for the tumor spread. Diffusion tensor imaging (DTI) is able to provide such information, thus opening the way for patient specific modeling of glioma invasion. Starting from a multiscale model involving subcellular (microscopic) and individual (mesoscale) cell dynamics, we perform a parabolic scaling to obtain an approximating reaction-diffusion-transport equation on the macroscale of the tumor cell population. Numerical simulations based on DTI data are carried out in order to assess the performance of our modeling approach. PMID:27105989

  20. Solar sail attitude control including active nutation damping in a fixed-momentum wheel satellite

    NASA Technical Reports Server (NTRS)

    Azor, Ruth

    1992-01-01

    In geostationary cruise of a momentum biased satellite, it is necessary to stabilize the roll/yaw attitude due to disturbances, caused mainly by solar radiation pressure. This work presents a roll/yaw control which is obtained by the use of solar arrays and fixed flaps as actuators, with a horizon sensor for roll measurement. The design also includes an active nutation damping.

  1. Population and Human Development: A Course Curriculum Including Lesson Plans, Activities, and Bibliography. Revised.

    ERIC Educational Resources Information Center

    Murphy, Elaine M.; Long, Alison T.

    This course outline suggests materials and learning activities on the interrelated causes and consequences of population growth and other population matters. The document describes 15 class sessions which integrate information for sociology, anthropology, psychology, biology, animal behavior, and education. Topics include the history of human…

  2. Dye laser amplifier including a dye cell contained within a support vessel

    DOEpatents

    Davin, James

    1992-01-01

    A large (high flow rate) dye laser amplifier in which a continous replenished supply of dye is excited by a first light beam, specifically a copper vapor laser beam, in order to amplify the intensity of a second different light beam, specifically a dye beam, passing through the dye is disclosed herein. This amplifier includes a dye cell defining a dye chamber through which a continuous stream of dye is caused to pass at a flow rate of greater than 30 gallons/minute at a static pressure greater than 150 pounds/square inch and a specifically designed support vessel for containing the dye cell.

  3. Dye laser amplifier including a dye cell contained within a support vessel

    DOEpatents

    Davin, J.

    1992-12-01

    A large (high flow rate) dye laser amplifier in which a continuous replenished supply of dye is excited by a first light beam, specifically a copper vapor laser beam, in order to amplify the intensity of a second different light beam, specifically a dye beam, passing through the dye is disclosed herein. This amplifier includes a dye cell defining a dye chamber through which a continuous stream of dye is caused to pass at a flow rate of greater than 30 gallons/minute at a static pressure greater than 150 pounds/square inch and a specifically designed support vessel for containing the dye cell. 6 figs.

  4. Simulation of E. coli Gene Regulation including Overlapping Cell Cycles, Growth, Division, Time Delays and Noise

    PubMed Central

    Luo, Ruoyu; Ye, Lin; Tao, Chenyang; Wang, Kankan

    2013-01-01

    Due to the complexity of biological systems, simulation of biological networks is necessary but sometimes complicated. The classic stochastic simulation algorithm (SSA) by Gillespie and its modified versions are widely used to simulate the stochastic dynamics of biochemical reaction systems. However, it has remained a challenge to implement accurate and efficient simulation algorithms for general reaction schemes in growing cells. Here, we present a modeling and simulation tool, called ‘GeneCircuits’, which is specifically developed to simulate gene-regulation in exponentially growing bacterial cells (such as E. coli) with overlapping cell cycles. Our tool integrates three specific features of these cells that are not generally included in SSA tools: 1) the time delay between the regulation and synthesis of proteins that is due to transcription and translation processes; 2) cell cycle-dependent periodic changes of gene dosage; and 3) variations in the propensities of chemical reactions that have time-dependent reaction rates as a consequence of volume expansion and cell division. We give three biologically relevant examples to illustrate the use of our simulation tool in quantitative studies of systems biology and synthetic biology. PMID:23638057

  5. Mast cells inhibit intramacrophage Francisella tularensis replication via contact and secreted products including IL-4

    PubMed Central

    Ketavarapu, Jyothi M.; Rodriguez, Annette R.; Yu, Jieh-Juen; Cong, Yu; Murthy, Ashlesh K.; Forsthuber, Thomas G.; Guentzel, M. Neal; Klose, Karl E.; Berton, Michael T.; Arulanandam, Bernard P.

    2008-01-01

    Francisella tularensis is an intracellular, Gram-negative bacterium that is the causative agent of pulmonary tularemia. The pathogenesis and mechanisms related to innate resistance against F. tularensis are not completely understood. Mast cells are strategically positioned within mucosal tissues, the major interface with the external environment, to initiate innate responses at the site of infection. Mast cell numbers in the cervical lymph nodes and the lungs progressively increased as early as 48 h after intranasal F. tularensis live vaccine strain (LVS) challenge. We established a primary bone marrow-derived mast cell–macrophage coculture system and found that mast cells significantly inhibit F. tularensis LVS uptake and growth within macrophages. Importantly, mice deficient in either mast cells or IL-4 receptor displayed greater susceptibility to the infection when compared with corresponding wild-type animals. Contact-dependent events and secreted products including IL-4 from mast cells, and IL-4 production from other cellular sources, appear to mediate the observed protective effects. These results demonstrate a previously unrecognized role for mast cells and IL-4 and provide a new dimension to our understanding of the innate immune mechanisms involved in controlling intramacrophage Francisella replication. PMID:18591675

  6. Twisted nematic liquid crystal cell characterization using rotating polarizers including full-field cell gap thickness measurement

    NASA Astrophysics Data System (ADS)

    Dev, Kapil; Prakarsa, Andy; Jiang, Yin Xi; Lee, Hooi Leng; Asundi, Anand

    2009-12-01

    Liquid crystal cells have always been an important part of commercially available displays, modulators and projectors. In this paper, Jones matrix representation including four independent parameters for twisted nematic liquid crystal (TN-LC) cell has been demonstrated. The physical parameters of the TN-LC cell such as twist angle, birefringence and director orientation at the input face of cell has been calculated using intensity transmittance of an experimental set-up that includes circularly polarized light and TN-LC cell sandwiched between two polarizers. The physical parameters have been calculated without any ambiguity using three different wavelengths. The knowledge from above measurement gives the general information about the birefringence and hence liquid crystal cell gap thickness. To acquire full-field liquid crystal cell gap thickness measurement phase shift polariscope has been adopted. The four phase shifted images through the rotating analyzer gives the full-field cell gap thickness measurement. The experimental results have been compared with the commercial point wise measurement and are in good agreement.

  7. Twisted nematic liquid crystal cell characterization using rotating polarizers including full-field cell gap thickness measurement

    NASA Astrophysics Data System (ADS)

    Dev, Kapil; Prakarsa, Andy; Jiang, Yin Xi; Lee, Hooi Leng; Asundi, Anand

    2010-03-01

    Liquid crystal cells have always been an important part of commercially available displays, modulators and projectors. In this paper, Jones matrix representation including four independent parameters for twisted nematic liquid crystal (TN-LC) cell has been demonstrated. The physical parameters of the TN-LC cell such as twist angle, birefringence and director orientation at the input face of cell has been calculated using intensity transmittance of an experimental set-up that includes circularly polarized light and TN-LC cell sandwiched between two polarizers. The physical parameters have been calculated without any ambiguity using three different wavelengths. The knowledge from above measurement gives the general information about the birefringence and hence liquid crystal cell gap thickness. To acquire full-field liquid crystal cell gap thickness measurement phase shift polariscope has been adopted. The four phase shifted images through the rotating analyzer gives the full-field cell gap thickness measurement. The experimental results have been compared with the commercial point wise measurement and are in good agreement.

  8. Multitarget magnetic activated cell sorter

    PubMed Central

    Adams, Jonathan D.; Kim, Unyoung; Soh, H. Tom

    2008-01-01

    Magnetic selection allows high-throughput sorting of target cells based on surface markers, and it is extensively used in biotechnology for a wide range of applications from in vitro diagnostics to cell-based therapies. However, existing methods can only perform separation based on a single parameter (i.e., the presence or absence of magnetization), and therefore, the simultaneous sorting of multiple targets at high levels of purity, recovery, and throughput remains a challenge. In this work, we present an alternative system, the multitarget magnetic activated cell sorter (MT-MACS), which makes use of microfluidics technology to achieve simultaneous spatially-addressable sorting of multiple target cell types in a continuous-flow manner. We used the MT-MACS device to purify 2 types of target cells, which had been labeled via target-specific affinity reagents with 2 different magnetic tags with distinct saturation magnetization and size. The device was engineered so that the combined effects of the hydrodynamic force produced from the laminar flow and the magnetophoretic force produced from patterned ferromagnetic structures within the microchannel result in the selective purification of the differentially labeled target cells into multiple independent outlets. We demonstrate here the capability to simultaneously sort multiple magnetic tags with >90% purity and >5,000-fold enrichment and multiple bacterial cell types with >90% purity and >500-fold enrichment at a throughput of 109 cells per hour. PMID:19015523

  9. EGFR activating mutations correlate with a Fanconi anemia-like cellular phenotype that includes PARP inhibitor sensitivity

    PubMed Central

    Pfäffle, Heike N.; Wang, Meng; Gheorghiu, Liliana; Ferraiolo, Natalie; Greninger, Patricia; Borgmann, Kerstin; Settleman, Jeffrey; Benes, Cyril H.; Sequist, Lecia V.; Zou, Lee; Willers, Henning

    2013-01-01

    In lung cancer patients whose tumors harbor activating mutations in the epidermal growth factor receptor (EGFR), increased responses to platinum-based chemotherapies are seen compared to wild-type cancers. However, the mechanisms underlying this association have remained elusive. Here, we describe a cellular phenotype of crosslinker sensitivity in a subset of EGFR-mutant lung cancer cell lines that is reminiscent of the defects seen in cells impaired in the Fanconi Anemia pathway, including a pronounced G2/M cell-cycle arrest and chromosomal radial formation. We identified a defect downstream of FANCD2 at the level of recruitment of FAN1 nuclease and DNA interstrand crosslink (ICL) unhooking. The effect of EGFR mutation was epistatic with FANCD2. Consistent with the known role of FANCD2 in promoting RAD51 foci formation and homologous recombination repair (HRR), EGFR-mutant cells also exhibited an impaired RAD51 foci response to ICLs, but not to DNA double-strand breaks. EGFR kinase inhibition affected RAD51 foci formation neither in EGFR mutant nor wild-type cells. In contrast, EGFR depletion or overexpression of mutant EGFR in wild-type cells suppressed RAD51 foci, suggesting an EGFR kinase-independent regulation of DNA repair. Interestingly, EGFR-mutant cells treated with the PARP inhibitor olaparib also displayed decreased FAN1 foci induction, coupled with a putative block in a late HRR step. As a result, EGFR-mutant lung cancer cells exhibited olaparib sensitivity in-vitro and in-vivo. Our findings provide insight into the mechanisms of cisplatin and PARP inhibitor sensitivity of EGFR-mutant cells, yielding potential therapeutic opportunities for further treatment individualization in this genetically defined subset of lung cancer. PMID:23966292

  10. Method of preparing a negative electrode including lithium alloy for use within a secondary electrochemical cell

    DOEpatents

    Tomczuk, Zygmunt; Olszanski, Theodore W.; Battles, James E.

    1977-03-08

    A negative electrode that includes a lithium alloy as active material is prepared by briefly submerging a porous, electrically conductive substrate within a melt of the alloy. Prior to solidification, excess melt can be removed by vibrating or otherwise manipulating the filled substrate to expose interstitial surfaces. Electrodes of such as solid lithium-aluminum filled within a substrate of metal foam are provided.

  11. Observing a fictitious stressful event: haematological changes, including circulating leukocyte activation.

    PubMed

    Mian, Rubina; Shelton-Rayner, Graham; Harkin, Brendan; Williams, Paul

    2003-03-01

    The aim of this study was to assess the effect of watching a psychological stressful event on the activation of leukocytes in healthy human volunteers. Blood samples were obtained from 32 healthy male and female subjects aged between 20 and 26 years before, during and after either watching an 83-minute horror film that none of the subjects had previously seen (The Texas Chainsaw Massacre, 1974) or by sitting quietly in a room (control group). Total differential cell counts, leukocyte activation as measured by the nitroblue tetrazolium (NBT) test, heart rate and blood pressure (BP) measurements were taken at defined time points. There were significant increases in peripheral circulating leukocytes, the number of activated circulating leukocytes, haemoglobin (Hb) concentration and haematocrit (Hct) in response to the stressor. These were accompanied by significant increases in heart rate, systolic and diastolic BP (P<0.05 from baseline). This is the first reported study on the effects of observing a psychologically stressful, albeit fictitious event on circulating leukocyte numbers and the state of leukocyte activation as determined by the nitrotetrazolium test. PMID:12637206

  12. Proteasome inhibitors, including curcumin, improve pancreatic β-cell function and insulin sensitivity in diabetic mice

    PubMed Central

    Weisberg, S; Leibel, R; Tortoriello, D V

    2016-01-01

    Background: Type 2 diabetes stems from obesity-associated insulin resistance, and in the genetically susceptible, concomitant pancreatic β-cell failure can occur, which further exacerbates hyperglycemia. Recent work by our group and others has shown that the natural polyphenol curcumin attenuates the development of insulin resistance and hyperglycemia in mouse models of hyperinsulinemic or compensated type 2 diabetes. Although several potential downstream molecular targets of curcumin exist, it is now recognized to be a direct inhibitor of proteasome activity. We now show that curcumin also prevents β-cell failure in a mouse model of uncompensated obesity-related insulin resistance (Leprdb/db on the Kaliss background). Results: In this instance, dietary supplementation with curcumin prevented hyperglycemia, increased insulin production and lean body mass, and prolonged lifespan. In addition, we show that short-term in vivo treatment with low dosages of two molecularly distinct proteasome inhibitors celastrol and epoxomicin reverse hyperglycemia in mice with β-cell failure by increasing insulin production and insulin sensitivity. Conclusions: These studies suggest that proteasome inhibitors may prove useful for patients with diabetes by improving both β-cell function and relieving insulin resistance. PMID:27110686

  13. T helper cell activation and human retroviral pathogenesis.

    PubMed Central

    Copeland, K F; Heeney, J L

    1996-01-01

    T helper (Th) cells are of central importance in regulating many critical immune effector mechanisms. The profile of cytokines produced by Th cells correlates with the type of effector cells induced during the immune response to foreign antigen. Th1 cells induce the cell-mediated immune response, while Th2 cells drive antibody production. Th cells are the preferential targets of human retroviruses. Infections with human T-cell leukemia virus (HTLV) or human immunodeficiency virus (HIV) result in the expansion of Th cells by the action of HTLV (adult T-cell leukemia) or the progressive loss of T cells by the action of HIV (AIDS). Both retrovirus infections impart a high-level activation state in the host immune cells as well as systemically. However, diverging responses to this activation state have contrasting effects on the Th-cell population. In HIV infection, Th-cell loss has been attributed to several mechanisms, including a selective elimination of cells by apoptosis. The induction of apoptosis in HIV infection is complex, with many different pathways able to induce cell death. In contrast, infection of Th cells with HTLV-1 affords the cell a protective advantage against apoptosis. This advantage may allow the cell to escape immune surveillance, providing the opportunity for the development of Th-cell cancer. In this review, we will discuss the impact of Th-cell activation and general immune activation on human retrovirus expression with a focus upon Th-cell function and the progression to disease. PMID:8987361

  14. Active seat suspension for a small vehicle: considerations for control system including observer

    NASA Astrophysics Data System (ADS)

    Katsumata, Hiroyuki; Shiino, Hiroshi; Oshinoya, Yasuo; Ishibashi, Kazuhisa; Ozaki, Koichi; Ogino, Hirohiko

    2007-12-01

    We have examined the improvement of ride quality and the reduction of riding fatigue brought about by the active control of the seat suspension of small vehicles such as one-seater electric automobiles. A small active seat suspension, which is easy to install, was designed and manufactured for one-seater electric automobiles. For the actuator, a maintenance-free voice coil motor used as a direct drive was adopted. For fundamental considerations, we designed a one-degree-of-freedom model for the active seat suspension system. Then, we designed a disturbance cancellation control system that includes the observer for a two-degree-of-freedom model. In an actual driving test, a test road, in which the concavity and convexity of an actual road surface were simulated using hard rubber, was prepared and the control performance of vertical vibrations of the seat surface during driving was examined. As a result, in comparison with the one-degree-of-freedom control system, it was confirmed that the control performance was improved by the two-degree-of-freedom control system that includes the observer.

  15. Breast Implant Informed Consent Should Include the Risk of Anaplastic Large Cell Lymphoma.

    PubMed

    Clemens, Mark W; Miranda, Roberto N; Butler, Charles E

    2016-04-01

    Breast implant-associated anaplastic large cell lymphoma (ALCL) is a rare T-cell lymphoma arising around breast implants. Public awareness has increased following a safety communication warning of the association of breast implant-associated ALCL by the U.S. Food and Drug Administration in 2011. Difficulty with determining an accurate assessment of risk, including diagnosis, or standardized treatment regimen has led surgeons to commonly omit preoperative discussion of this rare and frequently misunderstood cancer. Risk disclosure is a form of respect for patient autonomy, and informed consent has positive practical and moral consequences for the practice of plastic surgery. A model of breast implant-associated ALCL informed consent implementation and health care provider education are reviewed with 1-year process follow-up at a tertiary cancer center. Breast implant-associated ALCL should be included during preoperative counseling on the risks of breast implantation when obtaining informed consent. Pertinent aspects of decision-making include disease awareness, presenting symptoms, and resources for concerned patients. Education of health care professionals and provision of patient-focused materials ensures effectiveness of the informed consent process. PMID:27018666

  16. Are language-based activities in science effective for all students, including low achievers?

    NASA Astrophysics Data System (ADS)

    Rivard, Léonard P.

    2004-05-01

    The study investigated achievement status as a factor determining the use of language-based activities for learning science. A total of 154 eighth-grade students were randomly assigned to four groups, all stratified for gender and achievement level. The treatments involved various combinations of talk and writing, and descriptive and explanatory tasks. The dependent measures included scores on multiple choice tests obtained at three times during the study. Records of student talk and writing were also analyzed to identify patterns of differences between groups of achievers. The findings suggested that low achievers complete more problems, and develop better understanding and comprehension of ecology concepts when they have engaged in peer discussions of explanatory tasks. In comparison, high achievers benefit more from writing than talking, and writing explanations enhances comprehension more than restricted writing activities.

  17. Nuclear Rocket Test Facility Decommissioning Including Controlled Explosive Demolition of a Neutron-Activated Shield Wall

    SciTech Connect

    Michael Kruzic

    2007-09-01

    Located in Area 25 of the Nevada Test Site, the Test Cell A Facility was used in the 1960s for the testing of nuclear rocket engines, as part of the Nuclear Rocket Development Program. The facility was decontaminated and decommissioned (D&D) in 2005 using the Streamlined Approach For Environmental Restoration (SAFER) process, under the Federal Facilities Agreement and Consent Order (FFACO). Utilities and process piping were verified void of contents, hazardous materials were removed, concrete with removable contamination decontaminated, large sections mechanically demolished, and the remaining five-foot, five-inch thick radiologically-activated reinforced concrete shield wall demolished using open-air controlled explosive demolition (CED). CED of the shield wall was closely monitored and resulted in no radiological exposure or atmospheric release.

  18. Method of preparing a negative electrode including lithium alloy for use within a secondary electrochemical cell

    DOEpatents

    Tomczuk, Z.; Olszanski, W.; Battles, J.E.

    1975-12-09

    A negative electrode that includes a lithium alloy as active material is prepared by briefly submerging a porous, electrically conductive substrate within a melt of the alloy. Prior to solidification, excess melt can be removed by vibrating or otherwise manipulating the filled substrate to expose interstitial surfaces. Electrodes of such a solid lithium--aluminum filled within a substrate of metal foam are provided. 1 figure, 1 table.

  19. Fatty acid-releasing activities in Sinorhizobium meliloti include unusual diacylglycerol lipase.

    PubMed

    Sahonero-Canavesi, Diana X; Sohlenkamp, Christian; Sandoval-Calderón, Mario; Lamsa, Anne; Pogliano, Kit; López-Lara, Isabel M; Geiger, Otto

    2015-09-01

    Phospholipids are well known for their membrane-forming properties and thereby delimit any cell from the exterior world. In addition, membrane phospholipids can act as precursors for signals and other biomolecules during their turnover. Little is known about phospholipid signalling, turnover and remodelling in bacteria. Recently, we showed that a FadD-deficient mutant of Sinorhizobium meliloti, unable to convert free fatty acids to their coenzyme A derivatives, accumulates free fatty acids during the stationary phase of growth. Enzymatic activities responsible for the generation of these free fatty acids were unknown in rhizobia. Searching the genome of S. meliloti, we identified a potential lysophospholipase (SMc04041) and two predicted patatin-like phospholipases A (SMc00930, SMc01003). Although SMc00930 as well as SMc01003 contribute to the release of free fatty acids in S. meliloti, neither one can use phospholipids as substrates. Here we show that SMc01003 converts diacylglycerol to monoacylglycerol and a fatty acid, and that monoacylglycerol can be further degraded by SMc01003 to another fatty acid and glycerol. A SMc01003-deficient mutant of S. meliloti transiently accumulates diacylglycerol, suggesting that SMc01003 also acts as diacylglycerol lipase (DglA) in its native background. Expression of the DglA lipase in Escherichia coli causes lysis of cells in stationary phase of growth. PMID:25711932

  20. Expression of Human Endogenous Retrovirus-W Including Syncytin-1 in Cutaneous T-Cell Lymphoma

    PubMed Central

    Maliniemi, Pilvi; Vincendeau, Michelle; Mayer, Jens; Frank, Oliver; Hahtola, Sonja; Karenko, Leena; Carlsson, Emilia; Mallet, Francois; Seifarth, Wolfgang; Leib-Mösch, Christine; Ranki, Annamari

    2013-01-01

    The pathomechanism of mycosis fungoides (MF), the most common type of primary cutaneous T-cell lymphomas (CTCLs) and a malignancy of non-recirculating, skin-resident T-cells, is unknown albeit underlying viral infections have been sought for. Human endogenous retroviruses (HERVs) are ancient retroviral sequences in the human genome and their transcription is often deregulated in cancers. We explored the transcriptional activity of HERV sequences in a total of 34 samples comprising MF and psoriasis skin lesions, as well as corresponding non-malignant skin using a retrovirus-specific microarray and quantitative RT-PCR. To identify active HERV-W loci, we cloned the HERV-W specific RT-PCR products, sequenced the cDNA clones and assigned the sequences to HERV-W loci. Finally, we used immunohistochemistry on MF patient and non-malignant inflammatory skin samples to confirm specific HERV-encoded protein expression. Firstly, a distinct, skin-specific transcription profile consisting of five constitutively active HERV groups was established. Although individual variability was common, HERV-W showed significantly increased transcription in MF lesions compared to clinically intact skin from the same patient. Predominantly transcribed HERV-W loci were found to be located in chromosomes 6q21 and 7q21.2, chromosomal regions typically altered in CTCL. Surprisingly, we also found the expression of 7q21.2/ERVWE1-encoded Syncytin-1 (Env) protein in MF biopsies and expression of Syncytin-1 was seen in malignant lymphocytes, especially in the epidermotropic ones, in 15 of 30 cases studied. Most importantly, no Syncytin-1 expression was detected in inflammatory dermatosis (Lichen ruber planus) with skin-homing, non-malignant T lymphocytes. The expression of ERVWE1 mRNA was further confirmed in 3/7 MF lesions analyzed. Our observations strengthen the association between activated HERVs and cancer. The study offers a new perspective into the pathogenesis of CTCL since we demonstrate

  1. Should Cost-Effectiveness Analysis Include the Cost of Consumption Activities? AN Empirical Investigation.

    PubMed

    Adarkwah, Charles Christian; Sadoghi, Amirhossein; Gandjour, Afschin

    2016-02-01

    There has been a debate on whether cost-effectiveness analysis should consider the cost of consumption and leisure time activities when using the quality-adjusted life year as a measure of health outcome under a societal perspective. The purpose of this study was to investigate whether the effects of ill health on consumptive activities are spontaneously considered in a health state valuation exercise and how much this matters. The survey enrolled patients with inflammatory bowel disease in Germany (n = 104). Patients were randomized to explicit and no explicit instruction for the consideration of consumption and leisure effects in a time trade-off (TTO) exercise. Explicit instruction to consider non-health-related utility in TTO exercises did not influence TTO scores. However, spontaneous consideration of non-health-related utility in patients without explicit instruction (60% of respondents) led to significantly lower TTO scores. Results suggest an inclusion of consumption costs in the numerator of the cost-effectiveness ratio, at least for those respondents who spontaneously consider non-health-related utility from treatment. Results also suggest that exercises eliciting health valuations from the general public may include a description of the impact of disease on consumptive activities. PMID:25684073

  2. Kinetic modeling of rhamnolipid production by Pseudomonas aeruginosa PAO1 including cell density-dependent regulation.

    PubMed

    Henkel, Marius; Schmidberger, Anke; Vogelbacher, Markus; Kühnert, Christian; Beuker, Janina; Bernard, Thomas; Schwartz, Thomas; Syldatk, Christoph; Hausmann, Rudolf

    2014-08-01

    The production of rhamnolipid biosurfactants by Pseudomonas aeruginosa is under complex control of a quorum sensing-dependent regulatory network. Due to a lack of understanding of the kinetics applicable to the process and relevant interrelations of variables, current processes for rhamnolipid production are based on heuristic approaches. To systematically establish a knowledge-based process for rhamnolipid production, a deeper understanding of the time-course and coupling of process variables is required. By combining reaction kinetics, stoichiometry, and experimental data, a process model for rhamnolipid production with P. aeruginosa PAO1 on sunflower oil was developed as a system of coupled ordinary differential equations (ODEs). In addition, cell density-based quorum sensing dynamics were included in the model. The model comprises a total of 36 parameters, 14 of which are yield coefficients and 7 of which are substrate affinity and inhibition constants. Of all 36 parameters, 30 were derived from dedicated experimental results, literature, and databases and 6 of them were used as fitting parameters. The model is able to describe data on biomass growth, substrates, and products obtained from a reference batch process and other validation scenarios. The model presented describes the time-course and interrelation of biomass, relevant substrates, and products on a process level while including a kinetic representation of cell density-dependent regulatory mechanisms. PMID:24770383

  3. 25 CFR 170.137 - What types of activities can a recreation, tourism, and trails program include?

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... interpretative signs; (4) Provision for non-motorized trail activities including pedestrians and bicycles; (5) Provision for motorized trail activities including all terrain vehicles, motorcycles, snowmobiles, etc.; (6...; (8) Maintenance and restoration of existing recreational trails; (9) Development and...

  4. 25 CFR 170.137 - What types of activities can a recreation, tourism, and trails program include?

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... interpretative signs; (4) Provision for non-motorized trail activities including pedestrians and bicycles; (5) Provision for motorized trail activities including all terrain vehicles, motorcycles, snowmobiles, etc.; (6...; (8) Maintenance and restoration of existing recreational trails; (9) Development and...

  5. 25 CFR 170.137 - What types of activities can a recreation, tourism, and trails program include?

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... interpretative signs; (4) Provision for non-motorized trail activities including pedestrians and bicycles; (5) Provision for motorized trail activities including all terrain vehicles, motorcycles, snowmobiles, etc.; (6...; (8) Maintenance and restoration of existing recreational trails; (9) Development and...

  6. 25 CFR 170.137 - What types of activities can a recreation, tourism, and trails program include?

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... interpretative signs; (4) Provision for non-motorized trail activities including pedestrians and bicycles; (5) Provision for motorized trail activities including all terrain vehicles, motorcycles, snowmobiles, etc.; (6...; (8) Maintenance and restoration of existing recreational trails; (9) Development and...

  7. An Antimicrobial Metabolite from Bacillus sp.: Significant Activity Against Pathogenic Bacteria Including Multidrug-Resistant Clinical Strains

    PubMed Central

    Chalasani, Ajay G.; Dhanarajan, Gunaseelan; Nema, Sushma; Sen, Ramkrishna; Roy, Utpal

    2015-01-01

    In this study, the cell free modified tryptone soya broth (pH 7.4 ± 0.2) of Bacillus subtilis URID 12.1 showed significant antimicrobial activity against multidrug-resistant strains of Staphylococcus aureus, S. epidermidis, Streptococcus pyogenes and Enterococcus faecalis. The partially purified antimicrobial molecule was found to be resistant to extremes of pH and temperatures and also to higher concentrations of trypsin and proteinase K. The antimicrobial molecule was purified by a three-step method that included reversed-phase high performance liquid chromatography (RP-HPLC). The minimum inhibitory concentration (MIC) values were determined for 14 species of bacteria using a microbroth dilution technique. The HPLC-purified fraction showed the MICs ranging from 0.5 to 16 μg/ml for methicillin and vancomycin-resistant Staphylococcus aureus (MVRSA) and methicillin-resistant Staphylococcus epidermidis (MRSE) strains. The molecular mass of the antimicrobial compound was determined to be 842.37 Da. The same antimicrobial fraction showed negligible haemolytic activity against human red blood cells even at a concentration as high as 100 μg/ml. Because of its significant antimicrobial activity at low MIC values coupled with its non-haemolytic property, it may prove to be a novel antimicrobial lead molecule. PMID:26696963

  8. Electrochemical cell including a solid electrolyte made from a cation conductive vitreous compound

    SciTech Connect

    Duchange, J.P.; Gabano, J.P.

    1984-04-24

    An electrochemical cell having a lithium based negative active material, an electrolyte in the form of a vitreous cation conductive compound having a formula: aP/sub 2/S/sub 5/, bLi/sub 2/S, cLiX, where X stands for chlorine, bromine or iodine, and a, b, and c are numbers chosen in such a manner that b/(a+b) lies in the range 0.61 to 0.70 and c/(a+b+c) is less than or equal to the maximum solubility in the vitreous phase of LiX in the compound aP/sub 2/S/sub 5/, bLi/sub 2/S, and a positive active material in the form of a compressed powder with particles of the electrolyte dispersed throughout said positive active material. The positive active material is chosen from the group constituted by: (CH /SUB x/ ; Cu /SUB d/ O(PO/sub 4/)/sub 2/; V/sub 6/O/sub 13/; V/sub 2/S/sub 5/; MoS/sub 3/; CuS; S; CuO; Cu/sub 3/B/sub 2/O/sub 6/; FeS/sub 2/; Pb/sub 3/O/sub 4/; Bi/sub 2/O/sub 3/; PbO; BiO(CrO/sub 4/)/sub 2/; AgBi(CrO/sub 4/)/sub 2/; I/sub 2/; MoO/sub 3/; WO/sub 3/; TiS/sub 2/; NiPS/sub 3/; copper bismuthate; and lead bismuthate.

  9. Tracking and treating activated T cells

    PubMed Central

    Kim, N.H.; Nadithe, V.; Elsayed, M.; Merkel, O.M.

    2014-01-01

    Upon activation, T cells of various subsets are the most important mediators in cell-mediated immune responses. Activated T cells play an important role in immune system related diseases such as chronic inflammatory diseases, viral infections, autoimmune disease, transplant rejection, Crohn disease, diabetes, and many more. Therefore, efforts have been made to both visualize and treat activated T cells specifically. This review summarizes imaging approaches and selective therapeutics for activated T cells and gives an outlook on how tracking and treating can be combined into theragnositc agents for activated T cells. PMID:24660025

  10. Single cell multiplexed assay for proteolytic activity using droplet microfluidics.

    PubMed

    Ng, Ee Xien; Miller, Miles A; Jing, Tengyang; Chen, Chia-Hung

    2016-07-15

    Cellular enzymes interact in a post-translationally regulated fashion to govern individual cell behaviors, yet current platform technologies are limited in their ability to measure multiple enzyme activities simultaneously in single cells. Here, we developed multi-color Förster resonance energy transfer (FRET)-based enzymatic substrates and use them in a microfluidics platform to simultaneously measure multiple specific protease activities from water-in-oil droplets that contain single cells. By integrating the microfluidic platform with a computational analytical method, Proteolytic Activity Matrix Analysis (PrAMA), we are able to infer six different protease activity signals from individual cells in a high throughput manner (~100 cells/experimental run). We characterized protease activity profiles at single cell resolution for several cancer cell lines including breast cancer cell line MDA-MB-231, lung cancer cell line PC-9, and leukemia cell line K-562 using both live-cell and in-situ cell lysis assay formats, with special focus on metalloproteinases important in metastasis. The ability to measure multiple proteases secreted from or expressed in individual cells allows us to characterize cell heterogeneity and has potential applications including systems biology, pharmacology, cancer diagnosis and stem cell biology. PMID:26995287

  11. NIAM-Deficient Mice Are Predisposed to the Development of Proliferative Lesions including B-Cell Lymphomas

    PubMed Central

    Reed, Sara M.; Hagen, Jussara; Muniz, Viviane P.; Rosean, Timothy R.; Borcherding, Nick; Sciegienka, Sebastian; Goeken, J. Adam; Naumann, Paul W.; Zhang, Weizhou; Tompkins, Van S.; Janz, Siegfried; Meyerholz, David K.; Quelle, Dawn E.

    2014-01-01

    Nuclear Interactor of ARF and Mdm2 (NIAM, gene designation Tbrg1) is a largely unstudied inhibitor of cell proliferation that helps maintain chromosomal stability. It is a novel activator of the ARF-Mdm2-Tip60-p53 tumor suppressor pathway as well as other undefined pathways important for genome maintenance. To examine its predicted role as a tumor suppressor, we generated NIAM mutant (NIAMm/m) mice homozygous for a β-galactosidase expressing gene-trap cassette in the endogenous gene. The mutant mice expressed significantly lower levels of NIAM protein in tissues compared to wild-type animals. Fifty percent of aged NIAM deficient mice (14 to 21 months) developed proliferative lesions, including a uterine hemangioma, pulmonary papillary adenoma, and a Harderian gland adenoma. No age-matched wild-type or NIAM+/m heterozygous animals developed lesions. In the spleen, NIAMm/m mice had prominent white pulp expansion which correlated with enhanced increased reactive lymphoid hyperplasia and evidence of systemic inflammation. Notably, 17% of NIAM mutant mice had splenic white pulp features indicating early B-cell lymphoma. This correlated with selective expansion of marginal zone B cells in the spleens of younger, tumor-free NIAM-deficient mice. Unexpectedly, basal p53 expression and activity was largely unaffected by NIAM loss in isolated splenic B cells. In sum, NIAM down-regulation in vivo results in a significant predisposition to developing benign tumors or early stage cancers. These mice represent an outstanding platform for dissecting NIAM's role in tumorigenesis and various anti-cancer pathways, including p53 signaling. PMID:25393878

  12. Monitoring active volcanoes and mitigating volcanic hazards: the case for including simple approaches

    NASA Astrophysics Data System (ADS)

    Stoiber, Richard E.; Williams, Stanley N.

    1990-07-01

    Simple approaches to problems brought about eruptions and their ensuing hazardous effects should be advocated and used by volcanologists while awaiting more sophisticated remedies. The expedients we advocate have all or many of the following attributes: only locally available materials are required; no extensive training of operators or installation is necessary; they are affordable and do not require foreign aid or exports; they are often labor intensive and are sustainable without outside assistance. Where appropriate, the involvement of local residents is advocated. Examples of simple expedients which can be used in forecasting or mitigating the effects of crises emphasize the relative ease and the less elaborate requirements with which simple approaches can be activated. Emphasis is on visual observations often by untrained observers, simple meteorogical measurements, observations of water level in lakes, temperature and chemistry of springs and fumaroles, new springs and collapse areas and observations of volcanic plumes. Simple methods are suggested which can be applied to mitigating damage from mudflows, nuées ardentes, tephra falls and gas discharge. A review in hindsight at Ruiz includes the use of both chemical indicators and simple mudflow alarms. Simple expedients are sufficiently effective that any expert volcanologist called to aid in a crisis must include them in the package of advice offered. Simple approaches are a critical and logical complement to highly technical solutions to hazardous situations.

  13. Immune therapy including dendritic cell based therapy in chronic hepatitis B virus infection.

    PubMed

    Akbar, Sk Md Fazle; Horiike, Norio; Onji, Morikazu

    2006-05-14

    Hepatitis B virus (HBV) infection is a global public health problem. Of the approximately 2 billion people who have been infected worldwide, more than 400 million are chronic carriers of HBV. Considerable numbers of chronic HBV carriers suffer from progressive liver diseases. In addition, all HBV carriers are permanent source of this virus. There is no curative therapy for chronic HBV carriers. Antiviral drugs are recommended for about 10% patients, however, these drugs are costly, have limited efficacy, and possess considerable side effects. Recent studies have shown that immune responses of the host to the HBV are critically involved at every stage of chronic HBV infection: (1) These influence acquisition of chronic HBV carrier state, (2) They are important in the context of liver damages, (3) Recovery from chronic HBV-related liver diseases is dependent on nature and extent of HBV-specific immune responses. However, induction of adequate levels of HBV-specific immune responses in chronic HBV carriers is difficult. During the last one decade, hepatitis B vaccine has been administered to chronic HBV carriers as a therapeutic approach (vaccine therapy). The present regimen of vaccine therapy is safe and cheap, but not so effective. A dendritic cell-based therapeutic vaccine has recently been developed for treating chronic HBV infection. In this review, we will discuss about the concept, scientific logics, strategies and techniques of development of HBV-specific immune therapies including vaccine therapy and dendritic cell-based vaccine therapy for treating chronic HBV infection. PMID:16718812

  14. Particle-in-cell simulations for virtual cathode oscillator including foil ablation effects

    NASA Astrophysics Data System (ADS)

    Singh, Gursharn; Chaturvedi, S.

    2011-06-01

    We have performed two- and three-dimensional, relativistic, electromagnetic, particle-in-cell simulations of an axially extracted virtual cathode oscillator (vircator). The simulations include, for the first time, self-consistent dynamics of the anode foil under the influence of the intense electron beam. This yields the variation of microwave output power as a function of time, including the role of anode ablation and anode-cathode gap closure. These simulations have been done using locally developed particle-in-cell (PIC) codes. The codes have been validated using two vircator designs available from the literature. The simulations reported in the present paper take account of foil ablation due to the intense electron flux, the resulting plasma expansion and shorting of the anode-cathode gap. The variation in anode transparency due to plasma formation is automatically taken into account. We find that damage is generally higher near the axis. Also, at all radial positions, there is little damage in the early stages, followed by a period of rapid erosion, followed in turn by low damage rates. A physical explanation has been given for these trends. As a result of gap closure due to plasma formation from the foil, the output microwave power initially increases, reaches a near-flat-top and then decreases steadily, reaching a minimum around 230 ns. This is consistent with a typical plasma expansion velocity of ˜2 cm/μs reported in the literature. We also find a significant variation in the dominant output frequency, from 6.3 to 7.6 GHz. This variation is small as long as the plasma density is small, up to ˜40 ns. As the AK gap starts filling with plasma, there is a steady increase in this frequency.

  15. Decidual Cell Polyploidization Necessitates Mitochondrial Activity

    PubMed Central

    Ma, Xinghong; Gao, Fei; Rusie, Allison; Hemingway, Jennifer; Ostmann, Alicia B.; Sroga, Julie M.; Jegga, Anil G.; Das, Sanjoy K.

    2011-01-01

    Cellular polyploidy has been widely reported in nature, yet its developmental mechanism and function remain poorly understood. In the present study, to better define the aspects of decidual cell polyploidy, we isolated pure polyploid and non-polyploid decidual cell populations from the in vivo decidual bed. Three independent RNA pools prepared for each population were then subjected to the Affymetrix gene chip analysis for the whole mouse genome transcripts. Our data revealed up-regulation of 1015 genes and down-regulation of 1207 genes in the polyploid populations, as compared to the non-polyploid group. Comparative RT-PCR and in situ hybridization results indeed confirmed differential expressional regulation of several genes between the two populations. Based on functional enrichment analyses, up-regulated polyploidy genes appeared to implicate several functions, which primarily include cell/nuclear division, ATP binding, metabolic process, and mitochondrial activity, whereas that of down-regulated genes primarily included apoptosis and immune processes. Further analyses of genes that are related to mitochondria and bi-nucleation showed differential and regional expression within the decidual bed, consistent with the pattern of polyploidy. Consistently, studies revealed a marked induction of mitochondrial mass and ATP production in polyploid cells. The inhibition of mitochondrial activity by various pharmacological inhibitors, as well as by gene-specific targeting using siRNA-mediated technology showed a dramatic attenuation of polyploidy and bi-nucleation development during in vitro stromal cell decidualization, suggesting mitochondria play a major role in positive regulation of decidual cell polyploidization. Collectively, analyses of unique polyploidy markers and molecular signaling networks may be useful to further characterize functional aspects of decidual cell polyploidy at the site of implantation. PMID:22046353

  16. Viral Evasion of Natural Killer Cell Activation

    PubMed Central

    Ma, Yi; Li, Xiaojuan; Kuang, Ersheng

    2016-01-01

    Natural killer (NK) cells play a key role in antiviral innate defenses because of their abilities to kill infected cells and secrete regulatory cytokines. Additionally, NK cells exhibit adaptive memory-like antigen-specific responses, which represent a novel antiviral NK cell defense mechanism. Viruses have evolved various strategies to evade the recognition and destruction by NK cells through the downregulation of the NK cell activating receptors. Here, we review the recent findings on viral evasion of NK cells via the impairment of NK cell-activating receptors and ligands, which provide new insights on the relationship between NK cells and viral actions during persistent viral infections. PMID:27077876

  17. Mechanisms of Cell Propulsion by Active Stresses.

    PubMed

    Carlsson, A E

    2011-07-01

    The mechanisms by which cytoskeletal flows and cell-substrate interactions interact to generate cell motion are explored using a simplified model of the cytoskeleton as a viscous gel containing active stresses. This model yields explicit general results relating cell speed and traction forces to the distributions of active stress and cell-substrate friction. It is found that 1) the cell velocity is given by a function that quantifies the asymmetry of the active-stress distribution, 2) gradients in cell-substrate friction can induce motion even when the active stresses are symmetrically distributed, 3) the traction-force dipole is enhanced by protrusive stresses near the cell edges or contractile stresses near the center of the cell, and 4) the cell velocity depends biphasically on the cell-substrate adhesion strength if active stress is enhanced by adhesion. Specific experimental tests of the calculated dependences are proposed. PMID:21804763

  18. Mechanisms of Cell Propulsion by Active Stresses

    PubMed Central

    Carlsson, A. E.

    2011-01-01

    The mechanisms by which cytoskeletal flows and cell-substrate interactions interact to generate cell motion are explored using a simplified model of the cytoskeleton as a viscous gel containing active stresses. This model yields explicit general results relating cell speed and traction forces to the distributions of active stress and cell-substrate friction. It is found that 1) the cell velocity is given by a function that quantifies the asymmetry of the active-stress distribution, 2) gradients in cell-substrate friction can induce motion even when the active stresses are symmetrically distributed, 3) the traction-force dipole is enhanced by protrusive stresses near the cell edges or contractile stresses near the center of the cell, and 4) the cell velocity depends biphasically on the cell-substrate adhesion strength if active stress is enhanced by adhesion. Specific experimental tests of the calculated dependences are proposed. PMID:21804763

  19. Electrolytic/fuel cell bundles and systems including a current collector in communication with an electrode thereof

    DOEpatents

    Hawkes, Grant L.; Herring, James S.; Stoots, Carl M.; O& #x27; Brien, James E.

    2013-03-05

    Electrolytic/fuel cell bundles and systems including such bundles include an electrically conductive current collector in communication with an anode or a cathode of each of a plurality of cells. A cross-sectional area of the current collector may vary in a direction generally parallel to a general direction of current flow through the current collector. The current collector may include a porous monolithic structure. At least one cell of the plurality of cells may include a current collector that surrounds an outer electrode of the cell and has at least six substantially planar exterior surfaces. The planar surfaces may extend along a length of the cell, and may abut against a substantially planar surface of a current collector of an adjacent cell. Methods for generating electricity and for performing electrolysis include flowing current through a conductive current collector having a varying cross-sectional area.

  20. Chronic variable stress activates hematopoietic stem cells

    PubMed Central

    Courties, Gabriel; Dutta, Partha; Iwamoto, Yoshiko; Zaltsman, Alex; von zur Muhlen, Constantin; Bode, Christoph; Fricchione, Gregory L.; Denninger, John; Lin, Charles P.; Vinegoni, Claudio; Libby, Peter; Swirski, Filip K.; Weissleder, Ralph; Nahrendorf, Matthias

    2014-01-01

    Exposure to psychosocial stress is a risk factor for many diseases, including atherosclerosis1,2. While incompletely understood, interaction between the psyche and the immune system provides one potential mechanism linking stress and disease inception and progression. Known crosstalk between the brain and immune system includes the hypothalamic–pituitary–adrenal axis, which centrally drives glucocorticoid production in the adrenal cortex, and the sympathetic–adrenal–medullary axis, which controls stress–induced catecholamine release in support of the fight–or–flight reflex3,4. It remains unknown however if chronic stress changes hematopoietic stem cell activity. Here we show that stress increases proliferation of these most primitive progenitors, giving rise to higher levels of disease–promoting inflammatory leukocytes. We found that chronic stress induced monocytosis and neutrophilia in humans. While investigating the source of leukocytosis in mice, we discovered that stress activates upstream hematopoietic stem cells. Sympathetic nerve fibers release surplus noradrenaline, which uses the β3 adrenergic receptor to signal bone marrow niche cells to decrease CXCL12 levels. Consequently, elevated hematopoietic stem cell proliferation increases output of neutrophils and inflammatory monocytes. When atherosclerosis–prone ApoE−/− mice encounter chronic stress, accelerated hematopoiesis promotes plaque features associated with vulnerable lesions that cause myocardial infarction and stroke in humans. PMID:24952646

  1. Tryptophan protects hepatocytes against reactive oxygen species-dependent cell death via multiple pathways including Nrf2-dependent gene induction.

    PubMed

    Kimura, Takuya; Watanabe, Yoshifumi

    2016-05-01

    Hepatocyte apoptosis plays a key role in the pathogenesis of immune-mediated hepatitis. However, the detailed mechanisms of apoptosis signaling are still unclear and effective therapeutic drugs for hepatitis have been explored. Here, we show that tryptophan (Trp) suppressed IFN-γ-mediated hepatic apoptosis in vitro. Trp inhibited the downstream apoptotic events of mitochondria disruption, such as cell death and caspase-3 activation, while it did not influence upstream signaling including STAT1 activation and IRF1 expression. Trp suppressed reactive oxygen species (ROS) generation at the mitochondria. IFN-γ induced ROS in mitochondria by inhibiting complex I and III, but not II. This ROS generation by IFN-γ required de novo protein synthesis. Trp showed relatively weak direct scavenging activity but antagonized IFN-γ against the suppression of complex I. In addition, Trp increased the expression of the Nrf2-dependent antioxidant genes NQO1, HO-1 and GCS in hepatocytes both in vitro and in vivo. Finally, the administration of Trp in an acetaminophen-induced ROS-dependent hepatitis model suppressed the liver injury in vivo. Thus, Trp protects hepatocytes from ROS-dependent cell injury via multiple pathways. This study suggests Trp as a therapeutic antioxidant drug for hepatitis and a regulator for Nrf2-dependent genes. PMID:26795536

  2. Be BOLD: Encouraging Girls to Include Unstructured Bouts of Physical Activity into Daily Routines

    ERIC Educational Resources Information Center

    Hill, Kory; Williams, Gwynne M.

    2014-01-01

    Adolescent girls are less active than their male counterparts and physical activity levels tend to decline as one ages. One of the goals of concerned physical educators is to promote a physically active lifestyle and to teach skills and promote behaviors that will allow students to be active both in and out of school. This article presents a…

  3. Transition metals activate TFEB in overexpressing cells

    PubMed Central

    Peña, Karina A.; Kiselyov, Kirill

    2015-01-01

    Transition metal toxicity is an important factor in the pathogenesis of numerous human disorders, including neurodegenerative diseases. Lysosomes have emerged as important factors in transition metal toxicity because they handle transition metals via endocytosis, autophagy, absorption from the cytoplasm and exocytosis. Transcription factor EB (TFEB) regulates lysosomal biogenesis and the expression of lysosomal proteins in response to lysosomal and/or metabolic stresses. Since transition metals cause lysosomal dysfunction, we proposed that TFEB may be activated to drive gene expression in response to transition metal exposure and that such activation may influence transition metal toxicity. We found that transition metals copper (Cu) and iron (Fe) activate recombinant TFEB and stimulate the expression of TFEB-dependent genes in TFEB-overexpressing cells. In cells that show robust lysosomal exocytosis, TFEB was cytoprotective at moderate levels of Cu exposure, decreasing oxidative stress as reported by the expression of heme oxygenase-1 (HMOX1) gene. However, at high levels of Cu exposure, particularly in cells with low levels of lysosomal exocytosis, activation of overexpressed TFEB was toxic, increasing oxidative stress and mitochondrial damage. Based on these data, we conclude that TFEB-driven gene network is a component of the cellular response to transition metals. These data suggest limitations and disadvantages of TFEB overexpression as a therapeutic approach. PMID:26251447

  4. Inactivation of Ricin Toxin by Nanosecond Pulsed Electric Fields Including Evidences from Cell and Animal Toxicity

    NASA Astrophysics Data System (ADS)

    Wei, Kai; Li, Wei; Gao, Shan; Ji, Bin; Zang, Yating; Su, Bo; Wang, Kaile; Yao, Maosheng; Zhang, Jue; Wang, Jinglin

    2016-01-01

    Ricin is one of the most toxic and easily produced plant protein toxin extracted from the castor oil plant, and it has been classified as a chemical warfare agent. Here, nanosecond pulsed electric fields (nsPEFs) at 30 kV/cm (pulse durations: 10 ns, 100 ns, and 300 ns) were applied to inactivating ricin up to 4.2 μg/mL. To investigate the efficacy, cells and mice were tested against the ricin treated by the nsPEFs via direct intraperitoneal injection and inhalation exposure. Results showed that nsPEFs treatments can effectively reduce the toxicity of the ricin. Without the nsPEFs treatment, 100% of mice were killed upon the 4 μg ricin injection on the first day, however 40% of the mice survived the ricin treated by the nsPEFs. Compared to injection, inhalation exposure even with higher ricin dose required longer time to observe mice fatality. Pathological observations revealed damages to heart, lung, kidney, and stomach after the ricin exposure, more pronounced for lung and kidney including severe bleeding. Sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE) and circular dichroism (CD) analyses revealed that although the primary structure of ricin was not altered, its secondary structures (beta-sheet and beta-turn) underwent transition upon the nsPEFs treatment.

  5. Inactivation of Ricin Toxin by Nanosecond Pulsed Electric Fields Including Evidences from Cell and Animal Toxicity

    PubMed Central

    Wei, Kai; Li, Wei; Gao, Shan; Ji, Bin; Zang, Yating; Su, Bo; Wang, Kaile; Yao, Maosheng; Zhang, Jue; Wang, Jinglin

    2016-01-01

    Ricin is one of the most toxic and easily produced plant protein toxin extracted from the castor oil plant, and it has been classified as a chemical warfare agent. Here, nanosecond pulsed electric fields (nsPEFs) at 30 kV/cm (pulse durations: 10 ns, 100 ns, and 300 ns) were applied to inactivating ricin up to 4.2 μg/mL. To investigate the efficacy, cells and mice were tested against the ricin treated by the nsPEFs via direct intraperitoneal injection and inhalation exposure. Results showed that nsPEFs treatments can effectively reduce the toxicity of the ricin. Without the nsPEFs treatment, 100% of mice were killed upon the 4 μg ricin injection on the first day, however 40% of the mice survived the ricin treated by the nsPEFs. Compared to injection, inhalation exposure even with higher ricin dose required longer time to observe mice fatality. Pathological observations revealed damages to heart, lung, kidney, and stomach after the ricin exposure, more pronounced for lung and kidney including severe bleeding. Sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE) and circular dichroism (CD) analyses revealed that although the primary structure of ricin was not altered, its secondary structures (beta-sheet and beta-turn) underwent transition upon the nsPEFs treatment. PMID:26728251

  6. Inactivation of Ricin Toxin by Nanosecond Pulsed Electric Fields Including Evidences from Cell and Animal Toxicity.

    PubMed

    Wei, Kai; Li, Wei; Gao, Shan; Ji, Bin; Zang, Yating; Su, Bo; Wang, Kaile; Yao, Maosheng; Zhang, Jue; Wang, Jinglin

    2016-01-01

    Ricin is one of the most toxic and easily produced plant protein toxin extracted from the castor oil plant, and it has been classified as a chemical warfare agent. Here, nanosecond pulsed electric fields (nsPEFs) at 30 kV/cm (pulse durations: 10 ns, 100 ns, and 300 ns) were applied to inactivating ricin up to 4.2 μg/mL. To investigate the efficacy, cells and mice were tested against the ricin treated by the nsPEFs via direct intraperitoneal injection and inhalation exposure. Results showed that nsPEFs treatments can effectively reduce the toxicity of the ricin. Without the nsPEFs treatment, 100% of mice were killed upon the 4 μg ricin injection on the first day, however 40% of the mice survived the ricin treated by the nsPEFs. Compared to injection, inhalation exposure even with higher ricin dose required longer time to observe mice fatality. Pathological observations revealed damages to heart, lung, kidney, and stomach after the ricin exposure, more pronounced for lung and kidney including severe bleeding. Sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE) and circular dichroism (CD) analyses revealed that although the primary structure of ricin was not altered, its secondary structures (beta-sheet and beta-turn) underwent transition upon the nsPEFs treatment. PMID:26728251

  7. Antiviral Regulation in Porcine Monocytic Cells at Different Activation States

    PubMed Central

    Rowland, Raymond R. R.

    2014-01-01

    ABSTRACT Monocytic cells, including macrophages and dendritic cells, exist in different activation states that are critical to the regulation of antimicrobial immunity. Many pandemic viruses are monocytotropic, including porcine reproductive and respiratory syndrome virus (PRRSV), which directly infects subsets of monocytic cells and interferes with antiviral responses. To study antiviral responses in PRRSV-infected monocytic cells, we characterized inflammatory cytokine responses and genome-wide profiled signature genes to investigate response pathways in uninfected and PRRSV-infected monocytic cells at different activation states. Our findings showed suppressed interferon (IFN) production in macrophages in non-antiviral states and an arrest of lipid metabolic pathways in macrophages at antiviral states. Importantly, porcine monocytic cells at different activation states were susceptible to PRRSV and responded differently to viral infection. Based on Gene Ontology (GO) analysis, two approaches were used to potentiate antiviral activity: (i) pharmaceutical modulation of cellular lipid metabolism and (ii) in situ PRRSV replication-competent expression of interferon alpha (IFN-α). Both approaches significantly suppressed exogenous viral infection in monocytic cells. In particular, the engineered IFN-expressing PRRSV strain eliminated exogenous virus infection and sustained cell viability at 4 days postinfection in macrophages. These findings suggest an intricate interaction of viral infection with the activation status of porcine monocytic cells. An understanding and integration of antiviral infection with activation status of monocytic cells may provide a means of potentiating antiviral immunity. IMPORTANCE Activation statuses of monocytic cells, including monocytes, macrophages (Mϕs), and dendritic cells (DCs), are critically important for antiviral immunity. Unfortunately, the activation status of porcine monocytic cells or how cell activation status

  8. Thiamine induced resistance to Plasmopara viticola in grapevine and elicited host-defense responses, including HR like-cell death.

    PubMed

    Boubakri, Hatem; Wahab, Mohamed Ali; Chong, Julie; Bertsch, Christophe; Mliki, Ahmed; Soustre-Gacougnolle, Isabelle

    2012-08-01

    Recently, thiamine (VitaminB1) has been shown to induce resistance against Pseudomonas syringae in Arabidopsis plants through priming of defense responses. In this paper, we have demonstrated the efficiency of thiamine to induce resistance against downy mildew caused by the oomycete Plasmopara viticola in a susceptible Vitis vinifera cultivar "Chardonnay" under glasshouse controlled conditions by providing a dual mode of action involving direct antifungal activity and elicitation of host-defense responses. Thiamine-induced defense responses included the generation of hydrogen peroxide (H(2)O(2)) in both grapevine suspension cultured cells (SCC) and plant leaves, upregulation of an array of defense-related genes and the induction of other defense responses at subcellular level such as callose deposition in stomata cells, phenolic compounds accumulation and hypersensitive response (HR) like-cell death. Epifluorescence microscopy studies revealed dramatic changes in P. viticola individual developmental stages during its colonization of the intercellular space of the leaf mesophyll in thiamine-treated plants. Collectively, our report evidenced the efficiency of thiamine in the control of downy mildew in grapevine by direct and indirect effects, suggesting that thiamine could be an attractive alternative to chemical fungicides in disease management in vineyards. PMID:22698755

  9. Calcium alloy as active material in secondary electrochemical cell

    DOEpatents

    Roche, Michael F.; Preto, Sandra K.; Martin, Allan E.

    1976-01-01

    Calcium alloys such as calcium-aluminum and calcium-silicon, are employed as active material within a rechargeable negative electrode of an electrochemical cell. Such cells can use a molten salt electrolyte including calcium ions and a positive electrode having sulfur, sulfides, or oxides as active material. The calcium alloy is selected to prevent formation of molten calcium alloys resulting from reaction with the selected molten electrolytic salt at the cell operating temperatures.

  10. Mast cells enhance T cell activation: Importance of mast cell-derived TNF

    NASA Astrophysics Data System (ADS)

    Nakae, Susumu; Suto, Hajime; Kakurai, Maki; Sedgwick, Jonathon D.; Tsai, Mindy; Galli, Stephen J.

    2005-05-01

    Mast cells are not only important effector cells in immediate hypersensitivity reactions and immune responses to pathogens but also can contribute to T cell-mediated disorders. However, the mechanisms by which mast cells might influence T cells in such settings are not fully understood. We find that mast cells can enhance proliferation and cytokine production in multiple T cell subsets. Mast cell-dependent enhancement of T cell activation can be promoted by FcRI-dependent mast cell activation, TNF production by both mast cells and T cells, and mast cell-T cell contact. However, at high concentrations of cells, mast cells can promote T cell activation independent of IgE or TNF. Finally, mast cells also can promote T cell activation by means of soluble factors. These findings identify multiple mechanisms by which mast cells can influence T cell proliferation and cytokine production. allergy | asthma | autoimmunity | cytokines | immune response

  11. Using Assistive Technology Adaptations To Include Students with Learning Disabilities in Cooperative Learning Activities.

    ERIC Educational Resources Information Center

    Bryant, Diane Pedrotty; Bryant, Brian R.

    1998-01-01

    Discusses a process for integrating technology adaptations for students with learning disabilities into cooperative-learning activities in terms of three components: (1) selecting adaptations; (2) monitoring use of adaptations during cooperative-learning activities; and (3) evaluating the adaptations' effectiveness. Barriers to and support systems…

  12. Interest Inventory. [Includes Academic Interest Measure, Pupil Activity Inventory, and Semantic Differential].

    ERIC Educational Resources Information Center

    Harvard Univ., Cambridge, MA. Harvard Project Physics.

    This Interest Inventory contains three inventories: Academic Interest Measure (AIM), Pupil Activity Inventory (PAI), and Semantic Differential test (SD). The AIM measures six subscales of academic interests; the PAI measures non-school activities in science; and the SD measures attitudes toward science and physics. The inventories are designed for…

  13. Persistent neural activity in head direction cells

    NASA Technical Reports Server (NTRS)

    Taube, Jeffrey S.; Bassett, Joshua P.; Oman, C. M. (Principal Investigator)

    2003-01-01

    Many neurons throughout the rat limbic system discharge in relation to the animal's directional heading with respect to its environment. These so-called head direction (HD) cells exhibit characteristics of persistent neural activity. This article summarizes where HD cells are found, their major properties, and some of the important experiments that have been conducted to elucidate how this signal is generated. The number of HD and angular head velocity cells was estimated for several brain areas involved in the generation of the HD signal, including the postsubiculum, anterior dorsal thalamus, lateral mammillary nuclei and dorsal tegmental nucleus. The HD cell signal has many features in common with what is known about how neural integration is accomplished in the oculomotor system. The nature of the HD cell signal makes it an attractive candidate for using neural network models to elucidate the signal's underlying mechanisms. The conditions that any network model must satisfy in order to accurately represent how the nervous system generates this signal are highlighted and areas where key information is missing are discussed.

  14. Fabrication method for cores of structural sandwich materials including star shaped core cells

    DOEpatents

    Christensen, Richard M.

    1997-01-01

    A method for fabricating structural sandwich materials having a core pattern which utilizes star and non-star shaped cells. The sheets of material are bonded together or a single folded sheet is used, and bonded or welded at specific locations, into a flat configuration, and are then mechanically pulled or expanded normal to the plane of the sheets which expand to form the cells. This method can be utilized to fabricate other geometric cell arrangements than the star/non-star shaped cells. Four sheets of material (either a pair of bonded sheets or a single folded sheet) are bonded so as to define an area therebetween, which forms the star shaped cell when expanded.

  15. Fabrication method for cores of structural sandwich materials including star shaped core cells

    DOEpatents

    Christensen, R.M.

    1997-07-15

    A method for fabricating structural sandwich materials having a core pattern which utilizes star and non-star shaped cells is disclosed. The sheets of material are bonded together or a single folded sheet is used, and bonded or welded at specific locations, into a flat configuration, and are then mechanically pulled or expanded normal to the plane of the sheets which expand to form the cells. This method can be utilized to fabricate other geometric cell arrangements than the star/non-star shaped cells. Four sheets of material (either a pair of bonded sheets or a single folded sheet) are bonded so as to define an area therebetween, which forms the star shaped cell when expanded. 3 figs.

  16. The DNA methylation profile of activated human natural killer cells.

    PubMed

    Wiencke, John K; Butler, Rondi; Hsuang, George; Eliot, Melissa; Kim, Stephanie; Sepulveda, Manuel A; Siegel, Derick; Houseman, E Andres; Kelsey, Karl T

    2016-05-01

    Natural killer (NK) cells are now recognized to exhibit characteristics akin to cells of the adaptive immune system. The generation of adaptive memory is linked to epigenetic reprogramming including alterations in DNA methylation. The study herein found reproducible genome wide DNA methylation changes associated with human NK cell activation. Activation led predominately to CpG hypomethylation (81% of significant loci). Bioinformatics analysis confirmed that non-coding and gene-associated differentially methylated sites (DMS) are enriched for immune related functions (i.e., immune cell activation). Known DNA methylation-regulated immune loci were also identified in activated NK cells (e.g., TNFA, LTA, IL13, CSF2). Twenty-one loci were designated high priority and further investigated as potential markers of NK activation. BHLHE40 was identified as a viable candidate for which a droplet digital PCR assay for demethylation was developed. The assay revealed high demethylation in activated NK cells and low demethylation in naïve NK, T- and B-cells. We conclude the NK cell methylome is plastic with potential for remodeling. The differentially methylated region signature of activated NKs revealed similarities with T cell activation, but also provided unique biomarker candidates of NK activation, which could be useful in epigenome-wide association studies to interrogate the role of NK subtypes in global methylation changes associated with exposures and/or disease states. PMID:26967308

  17. Methods of improving the efficiency of photovoltaic cells. [including X ray analysis

    NASA Technical Reports Server (NTRS)

    Loferski, J. J.; Roessler, B.; Crisman, E. E.; Chen, L. Y.; Kaul, R.

    1974-01-01

    Work on aluminum-alloyed silicon grating cells is continued. Optimization of the geometry (grating line width and spacing) confirms the analysis of such cells. A 1 sq cm grating cell was fabricated and its i-V characteristic was measured under an AMO solar simulator. It is found that the efficiency of this cell would be about 7.9%, if it were covered by the usual antireflection coating. The surface of the cell is not covered by a diffused junction. The response is blue shifted; the current is somewhat higher than that produced by a commercial Si cell. However, the open circuit voltage is low, and attempts to optimize the open circuit voltage of the aluminum-alloy junctions are described. A preliminary X-ray topographic examination of GaAs specimens of the type commonly used to make solar cells is studied. The X-ray study shows that the wafers are filled with regions having strain gradients, possibly caused by precipitates. It is possible that a correlation exists between the presence of low mechanical perfection and minority carrier diffusion lengths of GaAs crystals.

  18. Mobilized peripheral blood grafts include more than hematopoietic stem cells: the immunological perspective.

    PubMed

    Saraceni, F; Shem-Tov, N; Olivieri, A; Nagler, A

    2015-07-01

    Although stem cell mobilization has been performed for more than 20 years, little is known about the effects of mobilizing agents on apheresis composition and the impact of graft cell subsets on patients' outcome. With the increasing use of plerixafor and the inclusion of poor mobilizers in autologous transplant procedures, new parameters other than CD34(+) stem cell dose are emerging; plerixafor seems to mobilize more primitive CD34(+)/CD38(-) stem cells compared with G-CSF, but their correlation with stable hematopoietic engraftment is still obscure. Immune recovery is as crucial as hematopoietic reconstitution, and higher T and natural killer cells infused within the graft have been correlated with better outcome in autologous transplant; recent studies showed increased mobilization of immune effectors with plerixafor compared with G-CSF, but further data are needed to clarify the clinical impact of these findings. In the allogeneic setting, much evidence suggests that mobilized T-cell alloreactivity is tempered by G-CSF, probably with the mediation of dendritic cells, even though no clear correlation with GVL and GVHD has been found. Plerixafor is not approved in healthy donors yet; early data suggest it might mobilize a GVHD protective balance of immune effectors, but further studies are needed to define its role in allogeneic transplant. PMID:25665044

  19. Remote System Technologies for Deactivating Hanford Hot Cells (for WM'03 - abstract included)

    SciTech Connect

    BERLIN, G.T.

    2003-01-28

    Remote system technologies are being deployed by Fluor Hanford to help accelerate the deactivation of highly-radioactive hot cell facilities. This paper highlights the application of several remotely deployed technologies enabling the deactivation tasks.

  20. Diffractive laser beam homogenizer including a photo-active material and method of fabricating the same

    DOEpatents

    Bayramian, Andy J; Ebbers, Christopher A; Chen, Diana C

    2014-05-20

    A method of manufacturing a plurality of diffractive optical elements includes providing a partially transmissive slide, providing a first piece of PTR glass, and directing first UV radiation through the partially transmissive slide to impinge on the first piece of PTR glass. The method also includes exposing predetermined portions of the first piece of PTR glass to the first UV radiation and thermally treating the exposed first piece of PTR glass. The method further includes providing a second piece of PTR glass and directing second UV radiation through the thermally treated first piece of PTR glass to impinge on the second piece of PTR glass. The method additionally includes exposing predetermined portions of the second piece of PTR glass to the second UV radiation, thermally treating the exposed second piece of PTR glass, and repeating providing and processing of the second piece of PTR glass using additional pieces of PTR glass.

  1. Activation-induced necroptosis contributes to B-cell lymphopenia in active systemic lupus erythematosus

    PubMed Central

    Fan, H; Liu, F; Dong, G; Ren, D; Xu, Y; Dou, J; Wang, T; Sun, L; Hou, Y

    2014-01-01

    B-cell abnormality including excessive activation and lymphopenia is a central feature of systemic lupus erythematosus (SLE). Although activation threshold, auto-reaction and death of B cells can be affected by intrinsical and/or external signaling, the underlying mechanisms are unclear. Herein, we demonstrate that co-activation of Toll-like receptor 7 (TLR7) and B-cell receptor (BCR) pathways is a core event for the survival/dead states of B cells in SLE. We found that the mortalities of CD19+CD27- and CD19+IgM+ B-cell subsets were increased in the peripheral blood mononuclear cells (PBMCs) of SLE patients. The gene microarray analysis of CD19+ B cells from active SLE patients showed that the differentially expressed genes were closely correlated to TLR7, BCR, apoptosis, necroptosis and immune pathways. We also found that co-activation of TLR7 and BCR could trigger normal B cells to take on SLE-like B-cell characters including the elevated viability, activation and proliferation in the first 3 days and necroptosis in the later days. Moreover, the necroptotic B cells exhibited mitochondrial dysfunction and hypoxia, along with the elevated expression of necroptosis-related genes, consistent with that in both SLE B-cell microarray and real-time PCR verification. Expectedly, pretreatment with the receptor-interacting protein kinase 1 (RIPK1) inhibitor Necrostatin-1, and not the apoptosis inhibitor zVAD, suppressed B-cell death. Importantly, B cells from additional SLE patients also significantly displayed high expression levels of necroptosis-related genes compared with those from healthy donors. These data indicate that co-activation of TLR7 and BCR pathways can promote B cells to hyperactivation and ultimately necroptosis. Our finding provides a new explanation on B-cell lymphopenia in active SLE patients. These data suggest that extrinsic factors may increase the intrinsical abnormality of B cells in SLE patients. PMID:25210799

  2. Natural killer cell activity during measles.

    PubMed Central

    Griffin, D E; Ward, B J; Jauregui, E; Johnson, R T; Vaisberg, A

    1990-01-01

    Natural killer cells are postulated to play an important role in host anti-viral defences. We measured natural killer cell activity in 30 individuals with acute measles (73 +/- 21 lytic units (LU)/10(7) cells) and 16 individuals with other infectious diseases (149 +/- 95 LU) and found it reduced compared with values for adults (375 +/- 70 LU; P less than 0.001) or children (300 +/- 73 LU, P less than 0.01) without infection. Reduced natural killer cell activity was found in measles patients with (84 +/- 30 LU) and without (55 +/- 18 LU) complications and was present for at least 3 weeks after the onset of the rash. Activity was increased by in vitro exposure of cells to interleukin-2. Depressed natural killer cell activity parallels in time the suppression of other parameters of cell-mediated immunity that occurs during measles. PMID:1696863

  3. Modeling Active Mechanosensing in Cell-Matrix Interactions.

    PubMed

    Chen, Bin; Ji, Baohua; Gao, Huajian

    2015-01-01

    Cells actively sense the mechanical properties of the extracellular matrix, such as its rigidity, morphology, and deformation. The cell-matrix interaction influences a range of cellular processes, including cell adhesion, migration, and differentiation, among others. This article aims to review some of the recent progress that has been made in modeling mechanosensing in cell-matrix interactions at different length scales. The issues discussed include specific interactions between proteins, the structure and mechanosensitivity of focal adhesions, the cluster effects of the specific binding, the structure and behavior of stress fibers, cells' sensing of substrate stiffness, and cell reorientation on cyclically stretched substrates. The review concludes by looking toward future opportunities in the field and at the challenges to understanding active cell-matrix interactions. PMID:26098510

  4. Sixty Minutes of Physical Activity per Day Included within Preschool Academic Lessons Improves Early Literacy

    ERIC Educational Resources Information Center

    Kirk, Stacie M.; Kirk, Erik P.

    2016-01-01

    Background: The effects of increases in physical activity (PA) on early literacy skills in preschool children are not known. Methods: Fifty-four African-American preschool children from a low socioeconomic urban Head Start participated over 8 months. A 2-group, quasi-experimental design was used with one preschool site participating in the PA…

  5. 7 CFR 981.441 - Credit for market promotion activities, including paid advertising.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... AGRICULTURE ALMONDS GROWN IN CALIFORNIA Administrative Rules and Regulations § 981.441 Credit for market... each activity shall be to promote the sale, consumption or use of California almonds, and nothing... in California almond growing counties with more than 1,000 bearing acres: Provided, That...

  6. Backyards and Butterflies: Ways to Include Children with Disabilities in Outdoor Activities.

    ERIC Educational Resources Information Center

    Greenstein, Doreen; And Others

    This sourcebook is designed for children, parents, and families, detailing ideas for outdoor play and learning activities, with emphasis on involving children with disabilities in outdoor play. A rural perspective permeates the guide, although each chapter contains ideas for making outdoor environments more accessible and safer for all children,…

  7. Beyond Right or Wrong: Challenges of Including Creative Design Activities in the Classroom

    ERIC Educational Resources Information Center

    Brennan, Karen

    2015-01-01

    In this article, we explore challenges encountered by K-12 educators in establishing classroom cultures that support creative learning activities with the Scratch programming language. Providing opportunities for students to understand and to build capacities for creative work was described by many of the teachers that we interviewed as a central…

  8. Physical Activity Programs in Higher Education: Modifying Net/Wall Games to Include Individuals with Disabilities

    ERIC Educational Resources Information Center

    Braga, Luciana; Tracy, Julia F.; Taliaferro, Andrea R.

    2015-01-01

    The growing number of students with disabilities in higher education settings has presented challenges for instructors with regards to appropriate inclusion. Concerning physical activity courses in higher education, instructors may not have the knowledge or resources to make modifications and accommodations that will ultimately result in…

  9. Population and Human Development: A Course Curriculum Including Lesson Plans, Activities and Bibliography.

    ERIC Educational Resources Information Center

    Murphy, Elaine M.

    This course outline suggests materials and learning activities on the interrelated causes and consequences of population growth and other population concerns. Designed to educate general college audiences, it is also intended for use as a preservice course for teachers. In addition, the course can be modified for high school students. The course…

  10. Estimation of Cell-Type Composition Including T and B Cell Subtypes for Whole Blood Methylation Microarray Data

    PubMed Central

    Waite, Lindsay L.; Weaver, Benjamin; Day, Kenneth; Li, Xinrui; Roberts, Kevin; Gibson, Andrew W.; Edberg, Jeffrey C.; Kimberly, Robert P.; Absher, Devin M.; Tiwari, Hemant K.

    2016-01-01

    DNA methylation levels vary markedly by cell-type makeup of a sample. Understanding these differences and estimating the cell-type makeup of a sample is an important aspect of studying DNA methylation. DNA from leukocytes in whole blood is simple to obtain and pervasive in research. However, leukocytes contain many distinct cell types and subtypes. We propose a two-stage model that estimates the proportions of six main cell types in whole blood (CD4+ T cells, CD8+ T cells, monocytes, B cells, granulocytes, and natural killer cells) as well as subtypes of T and B cells. Unlike previous methods that only estimate overall proportions of CD4+ T cell, CD8+ T cells, and B cells, our model is able to estimate proportions of naïve, memory, and regulatory CD4+ T cells as well as naïve and memory CD8+ T cells and naïve and memory B cells. Using real and simulated data, we are able to demonstrate that our model is able to reliably estimate proportions of these cell types and subtypes. In studies with DNA methylation data from Illumina's HumanMethylation450k arrays, our estimates will be useful both for testing for associations of cell type and subtype composition with phenotypes of interest as well as for adjustment purposes to prevent confounding in epigenetic association studies. Additionally, our method can be easily adapted for use with whole genome bisulfite sequencing (WGBS) data or any other genome-wide methylation data platform. PMID:26925097

  11. A novel peptide inhibitor of classical and lectin complement activation including ABO incompatibility

    PubMed Central

    Mauriello, Clifford T.; Pallera, Haree K.; Sharp, Julia A.; Woltmann, Jon L.; Qian, Shizhi; Hair, Pamela S.; van der Pol, Pieter; van Kooten, Cees; Thielens, Nicole M.; Lattanzio, Frank A.; Cunnion, Kenji M.; Krishna, Neel K.

    2012-01-01

    Previous experiments from our laboratories have identified peptides derived from the human astrovirus coat protein (CP) that bind C1q and mannose binding lectin (MBL) inhibiting activation of the classical and lectin pathways of complement, respectively. The purpose of this study was to evaluate the function of these coat protein peptides (CPPs) in an in vitro model of complement-mediated disease (ABO incompatibility), preliminarily assess their in vivo complement suppression profile and develop more highly potent derivatives of these molecules. E23A, a 30 amino acid CPP derivative previously demonstrated to inhibit classical pathway activation was able to dose-dependently inhibit lysis of AB erythrocytes treated with mismatched human O serum. Additionally, when injected into rats, E23A inhibited the animals’ serum from lysing antibody-sensitized erythrocytes, providing preliminary in vivo functional evidence that this CPP can cross the species barrier to inhibit serum complement activity in rodents. A rational drug design approach was implemented to identify more potent CPP derivatives, resulting in the identification and characterization of a 15 residue peptide (Polar Assortant (PA)), which demonstrated both superior inhibition of classical complement pathway activation and robust binding to C1q collagen-like tails. PA also inhibited ABO incompatibility in vitro and demonstrated in vivo complement suppression up to 24 hours post-injection. CPP’s ability to inhibit ABO incompatibility in vitro, proof of concept in vivo inhibitory activity in rats and the development of the highly potent PA derivative set the stage for preclinical testing of this molecule in small animal models of complement-mediated disease. PMID:22906481

  12. An Updated Review of Interventions that Include Promotion of Physical Activity for Adult Men.

    PubMed

    Bottorff, Joan L; Seaton, Cherisse L; Johnson, Steve T; Caperchione, Cristina M; Oliffe, John L; More, Kimberly; Jaffer-Hirji, Haleema; Tillotson, Sherri M

    2015-06-01

    The marked disparity in life expectancy between men and women suggests men are a vulnerable group requiring targeted health promotion programs. As such, there is an increasing need for health promotion strategies that effectively engage men with their health and/or illness management. Programs that promote physical activity could significantly improve the health of men. Although George et al. (Sports Med 42(3):281, 30) reviewed physical activity programs involving adult males published between 1990 and 2010, developments in men's health have prompted the emergence of new sex- and gender-specific approaches targeting men. The purpose of this review was to: (1) extend and update the review undertaken by George et al. (Sports Med 42(3):281, 30) concerning the effectiveness of physical activity programs in males, and (2) evaluate the integration of gender-specific influences in the content, design, and delivery of men's health promotion programs. A search of MEDLINE, CINAHL, ScienceDirect, Web of Science, PsycINFO, the Cochrane Library, and the SPORTDiscus databases for articles published between January 2010 and August 2014 was conducted. In total, 35 studies, involving evaluations of 31 programs, were identified. Findings revealed that a variety of techniques and modes of delivery could effectively promote physical activity among men. Though the majority of programs were offered exclusively to men, 12 programs explicitly integrated gender-related influences in male-specific programs in ways that recognized men's interests and preferences. Innovations in male-only programs that focus on masculine ideals and gender influences to engage men in increasing their physical activity hold potential for informing strategies to promote other areas of men's health. PMID:25430599

  13. Cell death sensitization of leukemia cells by opioid receptor activation

    PubMed Central

    Friesen, Claudia; Roscher, Mareike; Hormann, Inis; Fichtner, Iduna; Alt, Andreas; Hilger, Ralf A.; Debatin, Klaus-Michael; Miltner, Erich

    2013-01-01

    Cyclic AMP (cAMP) regulates a number of cellular processes and modulates cell death induction. cAMP levels are altered upon stimulation of specific G-protein-coupled receptors inhibiting or activating adenylyl cyclases. Opioid receptor stimulation can activate inhibitory Gi-proteins which in turn block adenylyl cyclase activity reducing cAMP. Opioids such as D,L-methadone induce cell death in leukemia cells. However, the mechanism how opioids trigger apoptosis and activate caspases in leukemia cells is not understood. In this study, we demonstrate that downregulation of cAMP induced by opioid receptor activation using the opioid D,L-methadone kills and sensitizes leukemia cells for doxorubicin treatment. Enhancing cAMP levels by blocking opioid-receptor signaling strongly reduced D,L-methadone-induced apoptosis, caspase activation and doxorubicin-sensitivity. Induction of cell death in leukemia cells by activation of opioid receptors using the opioid D,L-methadone depends on critical levels of opioid receptor expression on the cell surface. Doxorubicin increased opioid receptor expression in leukemia cells. In addition, the opioid D,L-methadone increased doxorubicin uptake and decreased doxorubicin efflux in leukemia cells, suggesting that the opioid D,L-methadone as well as doxorubicin mutually increase their cytotoxic potential. Furthermore, we found that opioid receptor activation using D,L-methadone alone or in addition to doxorubicin inhibits tumor growth significantly in vivo. These results demonstrate that opioid receptor activation via triggering the downregulation of cAMP induces apoptosis, activates caspases and sensitizes leukemia cells for doxorubicin treatment. Hence, opioid receptor activation seems to be a promising strategy to improve anticancer therapies. PMID:23633472

  14. Including MIR of a primary bone leiomyosarcoma that radiologically mimics a giant cell tumor.

    PubMed

    Sirikulchayanonta, Vorachai; Jaovisidh, Suphaneewan

    2008-02-01

    The authors present a case of a 42-year-old female who developed a leiomyosarcoma of the right proximal tibia that appeared radiologically similar to a giant cell tumor Histology revealed spindle cells running in whorl-like fashion with focal atypia and low mitotic figures. The immuno-stains revealed positive reactivity for alpha-smooth muscle (SMA), muscle actin and cytokeratin (AE1/AE3). The authors rendered a diagnosis of low-grade leiomyosarcoma of bone. The lesion was considered a primary lesion since the patient did not have other leiomyomatous tumors. The MRI showed hypo- to iso- signal intensity on T1-weighted imaging and heterogeneous intensity on T2-weighted imaging. This was likely due to admixed fibrotic tissue in the lesion. The tumor cells were not positive for Ebstein-Barr virus by in-situ hybridization as seen in leiomyomatous tumors in immunodeficiency patients. PMID:18389991

  15. Space Resources for Teachers: Biology, Including Suggestions for Classroom Activities and Laboratory Experiments.

    ERIC Educational Resources Information Center

    Lee, Tom E.; And Others

    This compilation of resource units concerns the latest developments in space biology. Some of the topics included are oxygen consumption, temperature, radiation, rhythms, weightlessness, acceleration and vibration stress, toxicity, and sensory and perceptual problems. Many of the topics are interdisciplinary and relate biology, physiology,…

  16. Organ Preference of Cancer Metastasis and Metastasis-Related Cell Adhesion Molecules Including Carbohydrates.

    PubMed

    Kawaguchi, Takanori

    2016-01-01

    This review starts on one of our special interests, the organ preference of metastasis. We examined data on 1,117 autopsy cases and found that the organ distribution of metastasis of cancers of the lung, pancreas, stomach, colon, rectum, uterine cervix, liver, bile duct, and esophagus involved the lung, liver, adrenal gland, bone/bone marrow, lymph node, and pleura/peritoneum. Cancers of the kidney, thyroid, ovary, choriocarcinoma, and breast, however, manifested different metastatic patterns. The distribution of leukemia and lymphoma metastases was quite different from that of epithelial cancers. On the basis of experimental studies, we believe that the anatomical-mechanical hypothesis should be replaced by the microinjury hypothesis, which suggests that tissue microinjury induced by temporal tumor cell embolization is crucial for successful metastasis. This hypothesis may actually reflect the so-called inflammatory oncotaxis concept. To clarify the mechanisms underlying metastasis, we developed an experimental model system of a rat hepatoma AH7974 that embraced substrate adhesiveness. This model did not prove a relationship between substrate-adhesion potential and metastatic lung-colonizing potential of tumor cells, but metastatic potential was correlated with the expression of the laminin carbohydrate that was recognized by Griffonia (Bandeiraea) simplicifolia isolectin G4. Therefore, we investigated the relationship between carbohydrate expression profiles and metastasis and prognosis. We indeed found an intimate relationship between the carbohydrate expression of cancer cells and the progression of malignant tumors, organ preference of metastasis, metastatic potential of tumor cells, and prognosis of patients. PMID:26521885

  17. Activity of faropenem tested against Neisseria gonorrhoeae isolates including fluoroquinolone-resistant strains.

    PubMed

    Jones, Ronald N; Critchley, Ian A; Whittington, William L H; Janjic, Nebojsa; Pottumarthy, Sudha

    2005-12-01

    We evaluated the anti-gonococcal potency of faropenem along with 7 comparator reference antimicrobials against a preselected collection of clinical isolates. The 265 isolates were inclusive of 2 subsets: 1) 76 well-characterized resistant phenotypes of gonococcal strains (53 quinolone-resistant strains--31 with documented quinolone resistance-determining region changes from Japan, 15 strains resistant to penicillin and tetracycline, and 8 strains with intermediate susceptibility to penicillin) and 2) 189 recent isolates from clinical specimens in 2004 from 6 states across the United States where quinolone resistance is prevalent. Activity of faropenem was adversely affected by l-cysteine hydrochloride in IsoVitaleX (4-fold increase in [minimal inhibitory concentration] MIC50; 0.06 versus 0.25 microg/mL). The rank order of potency of the antimicrobials for the entire collection was ceftriaxone (MIC90, 0.06 microg/mL) > faropenem (0.25 microg/mL) > azithromycin (0.5 microg/mL) > cefuroxime (1 microg/mL) > tetracycline (2 microg/mL) > penicillin = ciprofloxacin = levofloxacin (4 microg/mL). Using MIC90 for comparison, faropenem was 4-fold more potent than cefuroxime (0.25 versus 1 microg/mL), but was 4-fold less active than ceftriaxone (0.25 versus 0.06 microg/mL). Although the activity of faropenem was not affected by either penicillinase production (MIC90, 0.12 microg/mL, penicillinase-positive) or increasing ciprofloxacin MIC (0.25 microg/mL, ciprofloxacin-resistant), increasing penicillin MIC was associated with an increase in MIC90 values (0.016 microg/mL for penicillin-susceptible to 0.25 microg/mL for penicillin-resistant strains). Among the recent (2004) clinical gonococcal isolates tested, reduced susceptibility to penicillins, tetracycline, and fluoroquinolones was high (28.0-94.2%). Geographic distribution of the endemic resistance rates of gonococci varied considerably, with 16.7-66.7% of the gonococcal isolates being ciprofloxacin-resistant in Oregon

  18. Phytophthora infestans Has a Plethora of Phospholipase D Enzymes Including a Subclass That Has Extracellular Activity

    PubMed Central

    Meijer, Harold J. G.; Hassen, Hussen Harrun; Govers, Francine

    2011-01-01

    In eukaryotes phospholipase D (PLD) is involved in many cellular processes. Currently little is known about PLDs in oomycetes. Here we report that the oomycete plant pathogen Phytophthora infestans has a large repertoire of PLDs divided over six subfamilies: PXPH-PLD, PXTM-PLD, TM-PLD, PLD-likes, and type A and B sPLD-likes. Since the latter have signal peptides we developed a method using metabolically labelled phospholipids to monitor if P. infestans secretes PLD. In extracellular medium of ten P. infestans strains PLD activity was detected as demonstrated by the production of phosphatidic acid and the PLD specific marker phosphatidylalcohol. PMID:21423760

  19. Thermodynamic Modeling and Dispatch of Distributed Energy Technologies including Fuel Cell -- Gas Turbine Hybrids

    NASA Astrophysics Data System (ADS)

    McLarty, Dustin Fogle

    Distributed energy systems are a promising means by which to reduce both emissions and costs. Continuous generators must be responsive and highly efficiency to support building dynamics and intermittent on-site renewable power. Fuel cell -- gas turbine hybrids (FC/GT) are fuel-flexible generators capable of ultra-high efficiency, ultra-low emissions, and rapid power response. This work undertakes a detailed study of the electrochemistry, chemistry and mechanical dynamics governing the complex interaction between the individual systems in such a highly coupled hybrid arrangement. The mechanisms leading to the compressor stall/surge phenomena are studied for the increased risk posed to particular hybrid configurations. A novel fuel cell modeling method introduced captures various spatial resolutions, flow geometries, stack configurations and novel heat transfer pathways. Several promising hybrid configurations are analyzed throughout the work and a sensitivity analysis of seven design parameters is conducted. A simple estimating method is introduced for the combined system efficiency of a fuel cell and a turbine using component performance specifications. Existing solid oxide fuel cell technology is capable of hybrid efficiencies greater than 75% (LHV) operating on natural gas, and existing molten carbonate systems greater than 70% (LHV). A dynamic model is calibrated to accurately capture the physical coupling of a FC/GT demonstrator tested at UC Irvine. The 2900 hour experiment highlighted the sensitivity to small perturbations and a need for additional control development. Further sensitivity studies outlined the responsiveness and limits of different control approaches. The capability for substantial turn-down and load following through speed control and flow bypass with minimal impact on internal fuel cell thermal distribution is particularly promising to meet local demands or provide dispatchable support for renewable power. Advanced control and dispatch

  20. Fabrication of Mediatorless/Membraneless Glucose/Oxygen Based Biofuel Cell using Biocatalysts Including Glucose Oxidase and Laccase Enzymes

    NASA Astrophysics Data System (ADS)

    Christwardana, Marcelinus; Kim, Ki Jae; Kwon, Yongchai

    2016-07-01

    Mediatorless and membraneless enzymatic biofuel cells (EBCs) employing new catalytic structure are fabricated. Regarding anodic catalyst, structure consisting of glucose oxidase (GOx), poly(ethylenimine) (PEI) and carbon nanotube (CNT) is considered, while three cathodic catalysts consist of glutaraldehyde (GA), laccase (Lac), PEI and CNT that are stacked together in different ways. Catalytic activities of the catalysts for glucose oxidation and oxygen reduction reactions (GOR and ORR) are evaluated. As a result, it is confirmed that the catalysts work well for promotion of GOR and ORR. In EBC tests, performances of EBCs including 150 μm-thick membrane are measured as references, while those of membraneless EBCs are measured depending on parameters like glucose flow rate, glucose concentration, distance between two electrodes and electrolyte pH. With the measurements, how the parameters affect EBC performance and their optimal conditions are determined. Based on that, best maximum power density (MPD) of membraneless EBC is 102 ± 5.1 μW · cm‑2 with values of 0.5 cc · min‑1 (glucose flow rate), 40 mM (glucose concentration), 1 mm (distance between electrodes) and pH 3. When membrane and membraneless EBCs are compared, MPD of the membraneless EBC that is run at the similar operating condition to EBC including membrane is speculated as about 134 μW · cm‑2.

  1. Fabrication of Mediatorless/Membraneless Glucose/Oxygen Based Biofuel Cell using Biocatalysts Including Glucose Oxidase and Laccase Enzymes

    PubMed Central

    Christwardana, Marcelinus; Kim, Ki Jae; Kwon, Yongchai

    2016-01-01

    Mediatorless and membraneless enzymatic biofuel cells (EBCs) employing new catalytic structure are fabricated. Regarding anodic catalyst, structure consisting of glucose oxidase (GOx), poly(ethylenimine) (PEI) and carbon nanotube (CNT) is considered, while three cathodic catalysts consist of glutaraldehyde (GA), laccase (Lac), PEI and CNT that are stacked together in different ways. Catalytic activities of the catalysts for glucose oxidation and oxygen reduction reactions (GOR and ORR) are evaluated. As a result, it is confirmed that the catalysts work well for promotion of GOR and ORR. In EBC tests, performances of EBCs including 150 μm-thick membrane are measured as references, while those of membraneless EBCs are measured depending on parameters like glucose flow rate, glucose concentration, distance between two electrodes and electrolyte pH. With the measurements, how the parameters affect EBC performance and their optimal conditions are determined. Based on that, best maximum power density (MPD) of membraneless EBC is 102 ± 5.1 μW · cm−2 with values of 0.5 cc · min−1 (glucose flow rate), 40 mM (glucose concentration), 1 mm (distance between electrodes) and pH 3. When membrane and membraneless EBCs are compared, MPD of the membraneless EBC that is run at the similar operating condition to EBC including membrane is speculated as about 134 μW · cm−2. PMID:27426264

  2. Fabrication of Mediatorless/Membraneless Glucose/Oxygen Based Biofuel Cell using Biocatalysts Including Glucose Oxidase and Laccase Enzymes.

    PubMed

    Christwardana, Marcelinus; Kim, Ki Jae; Kwon, Yongchai

    2016-01-01

    Mediatorless and membraneless enzymatic biofuel cells (EBCs) employing new catalytic structure are fabricated. Regarding anodic catalyst, structure consisting of glucose oxidase (GOx), poly(ethylenimine) (PEI) and carbon nanotube (CNT) is considered, while three cathodic catalysts consist of glutaraldehyde (GA), laccase (Lac), PEI and CNT that are stacked together in different ways. Catalytic activities of the catalysts for glucose oxidation and oxygen reduction reactions (GOR and ORR) are evaluated. As a result, it is confirmed that the catalysts work well for promotion of GOR and ORR. In EBC tests, performances of EBCs including 150 μm-thick membrane are measured as references, while those of membraneless EBCs are measured depending on parameters like glucose flow rate, glucose concentration, distance between two electrodes and electrolyte pH. With the measurements, how the parameters affect EBC performance and their optimal conditions are determined. Based on that, best maximum power density (MPD) of membraneless EBC is 102 ± 5.1 μW · cm(-2) with values of 0.5 cc · min(-1) (glucose flow rate), 40 mM (glucose concentration), 1 mm (distance between electrodes) and pH 3. When membrane and membraneless EBCs are compared, MPD of the membraneless EBC that is run at the similar operating condition to EBC including membrane is speculated as about 134 μW · cm(-2). PMID:27426264

  3. Pluripotent embryonic stem cells and multipotent adult germline stem cells reveal similar transcriptomes including pluripotency-related genes.

    PubMed

    Meyer, S; Nolte, J; Opitz, L; Salinas-Riester, G; Engel, W

    2010-11-01

    DNA microarray analysis was performed with mouse multipotent adult germline stem cells (maGSCs) and embryonic stem cells (ESCs) from different genetic backgrounds cultured under standard ESC-culture conditions and under differentiation-promoting conditions by the withdrawal of the leukemia inhibitory factor (LIF) and treatment with retinoic acid (RA). The analyzed undifferentiated cell lines are very similar based on their global gene expression pattern and show 97-99% identity dependent on the analyzed background. Only 621 genes are differentially expressed in cells derived from mouse 129SV-background and 72 genes show differences in expression in cells generated from transgenic Stra8-EGFP/Rosa26-LacZ-background. Both maGSCs and ESCs express the same genes involved in the regulation of pluripotency and even show no differences in the expression level of these genes. When comparing maGSCs with previously published signature genes of other pluripotent cell lines, we found that maGSCs shared a very similar gene expression pattern with embryonic germ cells (EGCs). Also after differentiation of maGSCs and ESCs the transcriptomes of the cell lines are nearly identical which suggests that both cell types differentiate spontaneously in a very similar way. This is the first study, at transcriptome level, to compare ESCs and a pluripotent cell line derived from an adult organism (maGSCs). PMID:20624824

  4. Evaluation of school-based dental health activities including fluoride mouth-rinsing in Hiraizumi, Japan.

    PubMed

    Ohara, S; Kawaguchi, Y; Shinada, K; Sasaki, Y

    2000-06-01

    School-based dental health activities conducted in Hiraizumi over the past 20 years have remarkably improved the dental health status of schoolchildren. For example, DMFT index of 12-year-old children decreased to 1.5 in 1998, one-half that of the national average. School dental health activities, which were focused on dental health education, resulted in an increase of filled teeth rates, a decrease in the number of missing teeth, and a decline in incisor caries (1979-1986). In addition, the introduction of a school-based fluoride mouth-rinsing program (1986 - ) showed a positive effect on the prevention of dental caries; a significant decrease was observed in the overall prevalence of dental caries, particularly in the molars. In Japan it seems advantageous to promote the dental health of schoolchildren by school-based programs that combine dental health examination, dental health education and fluoride mouth-rinsing program. Especially, to prevent dental caries in the mandibular first molars more effectively, it is recommended to start fluoride mouth-rinsing at age 5. PMID:12160185

  5. New Pyrrole Derivatives with Potent Tubulin Polymerization Inhibiting Activity As Anticancer Agents Including Hedgehog-Dependent Cancer

    PubMed Central

    La Regina, Giuseppe; Bai, Ruoli; Coluccia, Antonio; Famiglini, Valeria; Pelliccia, Sveva; Passacantilli, Sara; Mazzoccoli, Carmela; Ruggieri, Vitalba; Sisinni, Lorenza; Bolognesi, Alessio; Rensen, Whilelmina Maria; Miele, Andrea; Nalli, Marianna; Alfonsi, Romina; Di Marcotullio, Lucia; Gulino, Alberto; Brancale, Andrea; Novellino, Ettore; Dondio, Giulio; Vultaggio, Stefania; Varasi, Mario; Mercurio, Ciro; Hamel, Ernest; Lavia, Patrizia; Silvestri, Romano

    2014-01-01

    We synthesized 3-aroyl-1-arylpyrrole (ARAP) derivatives as potential anticancer agents having different substituents at the pendant 1-phenyl ring. Both the 1-phenyl ring and 3-(3,4,5-trimethoxyphenyl)carbonyl moieties were mandatory to achieve potent inhibition of tubulin polymerization, binding of colchicine to tubulin, and cancer cell growth. ARAP 22 showed strong inhibition of the P-glycoprotein-overexpressing NCI-ADR-RES and Messa/Dx5MDR cell lines. Compounds 22 and 27 suppressed in vitro the Hedgehog signaling pathway, strongly reducing luciferase activity in SAG treated NIH3T3 Shh-Light II cells, and inhibited the growth of medulloblastoma D283 cells at nanomolar concentrations. ARAPs 22 and 27 represent a new potent class of tubulin polymerization and cancer cell growth inhibitors with the potential to inhibit the Hedgehog signaling pathway. PMID:25025991

  6. Polarization birefringence measurements for characterizing the myocardium, including healthy, infarcted, and stem-cell-regenerated tissues

    NASA Astrophysics Data System (ADS)

    Wood, Michael F. G.; Ghosh, Nirmalya; Wallenburg, Marika A.; Li, Shu-Hong; Weisel, Richard D.; Wilson, Brian C.; Li, Ren-Ke; Vitkin, I. Alex

    2010-07-01

    Myocardial infarction leads to structural remodeling of the myocardium, in particular to the loss of cardiomyocytes due to necrosis and an increase in collagen with scar formation. Stem cell regenerative treatments have been shown to alter this remodeling process, resulting in improved cardiac function. As healthy myocardial tissue is highly fibrous and anisotropic, it exhibits optical linear birefringence due to the different refractive indices parallel and perpendicular to the fibers. Accordingly, changes in myocardial structure associated with infarction and treatment-induced remodeling will alter the anisotropy exhibited by the tissue. Polarization-based linear birefringence is measured on the myocardium of adult rat hearts after myocardial infarction and compared with hearts that had received mesenchymal stem cell treatment. Both point measurement and imaging data show a decrease in birefringence in the region of infarction, with a partial rebound back toward the healthy values following regenerative treatment with stem cells. These results demonstrate the ability of optical polarimetry to characterize the micro-organizational state of the myocardium via its measured anisotropy, and the potential of this approach for monitoring regenerative treatments of myocardial infarction.

  7. A Methodology for Post Operational Clean Out of a Highly Active Facility Including Solids Behaviour - 12386

    SciTech Connect

    Edmondson, Michael J.; Ward, Tracy R.; Maxwell, Lisa J.

    2012-07-01

    The Highly Active Liquor Evaporation and Storage (HALES) plant at Sellafield handles acidic fission product containing liquor with typical activities of the order of 18x10{sup 9} Bq/ml. A strategy experimental feedback approach has been used to establish a wash regime for the Post Operational Clean Out (POCO) of the oldest storage tanks for this liquor. Two different wash reagents have been identified as being potentially suitable for removal of acid insoluble fission product precipitates. Ammonium carbamate and sodium carbonate yield similar products during the proposed wash cycle. The proposed wash reagents provide dissolution of caesium phosphomolybdate (CPM) and zirconium molybdate (ZM) solid phases but yields a fine, mobile precipitate of metal carbonates from the Highly Active Liquor (HAL) supernate. Addition of nitric acid to the wash effluent can cause CPM to precipitate where there is sufficient caesium and phosphorous available. Where they are not present (from ZM dissolution) the nitric acid addition initially produces a nitrate precipitate which then re-dissolves, along with the metal carbonates, to give a solid-free solution. The different behaviour of the two solids during the wash cycle has led to the proposal for an amended flowsheet. Additional studies on the potential to change the morphology of crystallising ZM have presented opportunities for changing the rheology of ZM sediments through doping with tellurium or particular organic acids. Two different wash reagents have been identified as being potentially suitable for the POCO of HALES Oldside HASTs. AC and SC both yield similar products during the proposed wash cycle. However, the different behaviour of the two principle HAL solids, CPM and ZM, during the wash cycle has led to the proposal for an amended flowsheet. Additional studies on the potential to change the morphology of crystallising ZM have presented opportunities for changing its rheology through doping with tellurium or certain

  8. Reduced Toxicity Fuel Satellite Propulsion System Including Fuel Cell Reformer with Alcohols Such as Methanol

    NASA Technical Reports Server (NTRS)

    Schneider, Steven J. (Inventor)

    2001-01-01

    A reduced toxicity fuel satellite propulsion system including a reduced toxicity propellant supply for consumption in an axial class thruster and an ACS class thruster. The system includes suitable valves and conduits for supplying the reduced toxicity propellant to the ACS decomposing element of an ACS thruster. The ACS decomposing element is operative to decompose the reduced toxicity propellant into hot propulsive gases. In addition the system includes suitable valves and conduits for supplying the reduced toxicity propellant to an axial decomposing element of the axial thruster. The axial decomposing element is operative to decompose the reduced toxicity propellant into hot gases. The system further includes suitable valves and conduits for supplying a second propellant to a combustion chamber of the axial thruster, whereby the hot gases and the second propellant auto-ignite and begin the combustion process for producing thrust.

  9. Steady-state analysis of activated sludge processes with a settler model including sludge compression.

    PubMed

    Diehl, S; Zambrano, J; Carlsson, B

    2016-01-01

    A reduced model of a completely stirred-tank bioreactor coupled to a settling tank with recycle is analyzed in its steady states. In the reactor, the concentrations of one dominant particulate biomass and one soluble substrate component are modelled. While the biomass decay rate is assumed to be constant, growth kinetics can depend on both substrate and biomass concentrations, and optionally model substrate inhibition. Compressive and hindered settling phenomena are included using the Bürger-Diehl settler model, which consists of a partial differential equation. Steady-state solutions of this partial differential equation are obtained from an ordinary differential equation, making steady-state analysis of the entire plant difficult. A key result showing that the ordinary differential equation can be replaced with an approximate algebraic equation simplifies model analysis. This algebraic equation takes the location of the sludge-blanket during normal operation into account, allowing for the limiting flux capacity caused by compressive settling to easily be included in the steady-state mass balance equations for the entire plant system. This novel approach grants the possibility of more realistic solutions than other previously published reduced models, comprised of yet simpler settler assumptions. The steady-state concentrations, solids residence time, and the wastage flow ratio are functions of the recycle ratio. Solutions are shown for various growth kinetics; with different values of biomass decay rate, influent volumetric flow, and substrate concentration. PMID:26476681

  10. Quasielastic neutron scattering experiments including activation energies and mathematical modeling of methyl halide dynamics

    NASA Astrophysics Data System (ADS)

    Kirstein, O.; Prager, M.; Grimm, H.; Buchsteiner, A.; Wischnewski, A.

    2007-09-01

    Quasielastic neutron scattering experiments were carried out using the multichopper time-of-flight spectrometer V3 at the Hahn-Meitner Institut, Germany and the backscattering spectrometer at Forschungszentrum Jülich, Germany. Activation energies for CH3X, X =F, Cl, Br, and I, were obtained. In combination with results from previous inelastic neutron scattering experiments the data were taken to describe the dynamics of the halides in terms of two different models, the single particle model and the coupling model. Coupled motions of methyl groups seem to explain the dynamics of the methyl fluoride and chloride; however, the coupling vanishes with the increase of the mass of the halide atom in CH3Br and CH3I.

  11. LIPID PEROXIDATION GENERATES BIOLOGICALLY ACTIVE PHOSPHOLIPIDS INCLUDING OXIDATIVELY N-MODIFIED PHOSPHOLIPIDS

    PubMed Central

    Davies, Sean S.; Guo, Lilu

    2014-01-01

    Peroxidation of membranes and lipoproteins converts “inert” phospholipids into a plethora of oxidatively modified phospholipids (oxPL) that can act as signaling molecules. In this review, we will discuss four major classes of oxPL: mildly oxygenated phospholipids, phospholipids with oxidatively truncated acyl chains, phospholipids with cyclized acyl chains, and phospholipids that have been oxidatively N-modified on their headgroups by reactive lipid species. For each class of oxPL we will review the chemical mechanisms of their formation, the evidence for their formation in biological samples, the biological activities and signaling pathways associated with them, and the catabolic pathways for their elimination. We will end by briefly highlighting some of the critical questions that remain about the role of oxPL in physiology and disease. PMID:24704586

  12. Multilayer-omics analysis of renal cell carcinoma, including the whole exome, methylome and transcriptome

    PubMed Central

    Arai, Eri; Sakamoto, Hiromi; Ichikawa, Hitoshi; Totsuka, Hirohiko; Chiku, Suenori; Gotoh, Masahiro; Mori, Taisuke; Nakatani, Tamao; Ohnami, Sumiko; Nakagawa, Tohru; Fujimoto, Hiroyuki; Wang, Linghua; Aburatani, Hiroyuki; Yoshida, Teruhiko; Kanai, Yae

    2014-01-01

    The aim of this study was to identify pathways that have a significant impact during renal carcinogenesis. Sixty-seven paired samples of both noncancerous renal cortex tissue and cancerous tissue from patients with clear cell renal cell carcinomas (RCCs) were subjected to whole-exome, methylome and transcriptome analyses using Agilent SureSelect All Exon capture followed by sequencing on an Illumina HiSeq 2000 platform, Illumina Infinium HumanMethylation27 BeadArray and Agilent SurePrint Human Gene Expression microarray, respectively. Sanger sequencing and quantitative reverse transcription-PCR were performed for technical verification. MetaCore software was used for pathway analysis. Somatic nonsynonymous single-nucleotide mutations, insertions/deletions and intragenic breaks of 2,153, 359 and 8 genes were detected, respectively. Mutations of GCN1L1, MED12 and CCNC, which are members of CDK8 mediator complex directly regulating β-catenin-driven transcription, were identified in 16% of the RCCs. Mutations of MACF1, which functions in the Wnt/β-catenin signaling pathway, were identified in 4% of the RCCs. A combination of methylome and transcriptome analyses further highlighted the significant role of the Wnt/β-catenin signaling pathway in renal carcinogenesis. Genetic aberrations and reduced expression of ERC2 and ABCA13 were frequent in RCCs, and MTOR mutations were identified as one of the major disrupters of cell signaling during renal carcinogenesis. Our results confirm that multilayer-omics analysis can be a powerful tool for revealing pathways that play a significant role in carcinogenesis. PMID:24504440

  13. A feedback model for leukemia including cell competition and the action of the immune system

    NASA Astrophysics Data System (ADS)

    Balea, S.; Halanay, A.; Neamtu, M.

    2014-12-01

    A mathematical model, coupling the dynamics of short-term stem-like cells and mature leukocytes in leukemia with that of the immune system, is investigated. The model is described by a system of nine delay differential equations with nine delays. Three equilibrium points E0, E1, E2 are highlighted. The stability and the existence of the Hopf bifurcation for the equilibrium points are investigated. In the analysis of the model, the rate of asymmetric division and the rate of symmetric division are very important.

  14. Ozone control of biological activity during Earth's history, including the KT catastrophe

    NASA Technical Reports Server (NTRS)

    Sheldon, W. R.

    1994-01-01

    There have been brief periods since the beginning of the Cambrian some 600 m.y. ago when mass extinctions destroyed a significant fraction of living species. The most widely studied of these events is the catastrophe at the KT boundary that ended the long dominance of the dinosaurs. In addition to mass extinctions, there is another profound discontinuity in the history of Earth's biota, the explosion of life at the end of the Precambrian era which is an episode that is not explained well at all. For some 3 b.y. before the Cambrian, life had been present on Earth, but maintained a low level of activity which is an aspect of the biota that is puzzling, especially during the last two-thirds of that period. During the last 2 b.y. before the Cambrian, conditions at the Earth's surface were suitable for a burgeoning of the biota, according to most criteria: the oceans neither boiled nor were fozen solid during this time, and the atmosphere contained sufficient O for the development of animals. The purpose of this paper is to suggest that mass extinctions and the lackluster behavior of the Precambrian biota share a common cause: an inadequate amount of ozone in the atmosphere.

  15. Design of a high-lift experiment in water including active flow control

    NASA Astrophysics Data System (ADS)

    Beutel, T.; Sattler, S.; El Sayed, Y.; Schwerter, M.; Zander, M.; Büttgenbach, S.; Leester-Schädel, M.; Radespiel, R.; Sinapius, M.; Wierach, P.

    2014-07-01

    This paper describes the structural design of an active flow-control experiment. The aim of the experiment is to investigate the increase in efficiency of an internally blown Coanda flap using unsteady blowing. The system uses tailor-made microelectromechanical (MEMS) pressure sensors to determine the state of the oncoming flow and an actuated lip to regulate the mass flow and velocity of a stream near a wall over the internally blown flap. Sensors and actuators are integrated into a highly loaded system that is extremely compact. The sensors are connected to a bus system that feeds the data into a real-time control system. The piezoelectric actuators using the d 33 effect at a comparable low voltage of 120 V are integrated into a lip that controls the blowout slot height. The system is designed for closed-loop control that efficiently avoids flow separation on the Coanda flap. The setup is designed for water-tunnel experiments in order to reduce the free-stream velocity and the system’s control frequency by a factor of 10 compared with that in air. This paper outlines the function and verification of the system’s main components and their development.

  16. Accurate expressions for solar cell fill factors including series and shunt resistances

    NASA Astrophysics Data System (ADS)

    Green, Martin A.

    2016-02-01

    Together with open-circuit voltage and short-circuit current, fill factor is a key solar cell parameter. In their classic paper on limiting efficiency, Shockley and Queisser first investigated this factor's analytical properties showing, for ideal cells, it could be expressed implicitly in terms of the maximum power point voltage. Subsequently, fill factors usually have been calculated iteratively from such implicit expressions or from analytical approximations. In the absence of detrimental series and shunt resistances, analytical fill factor expressions have recently been published in terms of the Lambert W function available in most mathematical computing software. Using a recently identified perturbative relationship, exact expressions in terms of this function are derived in technically interesting cases when both series and shunt resistances are present but have limited impact, allowing a better understanding of their effect individually and in combination. Approximate expressions for arbitrary shunt and series resistances are then deduced, which are significantly more accurate than any previously published. A method based on the insights developed is also reported for deducing one-diode fits to experimental data.

  17. Differentially expressed epigenome modifiers, including Aurora kinase A and B, in immune cells of rheumatoid arthritis

    PubMed Central

    Glant, Tibor T.; Besenyei, Timea; Kádár, András; Kurkó, Júlia; Tryniszewska, Beata; Gál, János; Soós, Györgyi; Szekanecz, Zoltán; Hoffmann, Gyula; Block, Joel A.; Katz, Robert S.; Mikecz, Katalin; Rauch, Tibor A.

    2014-01-01

    Objective The aim of this study was to identify epigenetic factors that are implicated in the pathogenesis of rheumatoid arthritis (RA) and to explore the therapeutic potential of the targeted inhibition of these factors. Methods PCR arrays were utilized to investigate the expression profile of genes that encod key epigenetic regulator enzymes. Mononuclear cells from RA patients and mice were monitored for gene expression changes, in association with arthritis development in murine models of RA. Selected genes were further characterized by quantitative real-time PCR, Western blot and flow cytometry methods. The targeted inhibition of the upregulated enzymes was studied in arthritic mice. Results A set of genes with arthritis-specific expression was identified by the PCR arrays. Aurora kinase A and B, both of which were highly expressed in arthritic mice and treatment naïve RA patients, were selected for detailed analysis. Elevated Aurora kinase expression was accompanied with an increased phosphorylation of histone H3, which promotes proliferation of T lymphocytes. Treatment with VX-680, a pan-Aurora kinase inhibitor, promoted B cell apoptosis, provided significant protection against the onset, and attenuated the inflammatory reactions in arthritic mice. Conclusions Arthritis development is accompanied the changes in the expression of a number of epigenome-modifying enzymes. Drug-induced downregulation of the Aurora kinases, among other targets, seems to be sufficient to treat experimental arthritis. Development of new therapeutics that target the Aurora kinases can potentially improve RA management. PMID:23653330

  18. Differences in Lifestyles Including Physical Activity According to Sexual Orientation among Korean Adolescents

    PubMed Central

    YOON, Jin-Ho; SO, Wi-Young

    2013-01-01

    Abstract Background The purpose of the present study was to examine differences in lifestyle factors such as physical activity among homosexual (gay or lesbian), bisexual, and heterosexual Korean adolescents. Methods The sample consisted of 74,186 adolescents from grades 7—12 (ages 12—18) who participated in the 8th annual Korea Youth Risk Behavior Web-based Survey in 2012. Of this sample, only 11,829 provided enough information regarding their romantic and sexual experiences to define them as gay, lesbian, bisexual, or heterosexual. From this information, males were divided into gay (n = 323), bisexual (n = 243), and heterosexual (n = 6,501) groups, and females were divided into lesbian (n = 208), bisexual (n = 113), and heterosexual (n = 4,441) groups. Differences in lifestyle factors according to sexual orientation were analyzed using one-way analysis of variance. Results Males showed significant differences by sexual orientation group in terms of frequency of smoking (P = 0.029), alcohol consumption (P < 0.001), muscular strength exercises (P = 0.020), and walking for at least 10 minutes per week (P < 0.001). Females showed significant differences by sexual orientation group in terms of frequency of smoking (P < 0.001), alcohol consumption (P < 0.001), vigorous physical exercise (P < 0.001), moderate physical exercise (P < 0.001), and muscular strength exercises (P < 0.001), as well as for self-reported mental stress (P < 0.001). Conclusion We concluded those gay and bisexual males and lesbian and bisexual females had significant lifestyle differences as compared with heterosexual adolescents. This effect was stronger for females than for males. PMID:26060636

  19. RADIO PROPERTIES OF LOW-REDSHIFT BROAD-LINE ACTIVE GALACTIC NUCLEI INCLUDING EXTENDED RADIO SOURCES

    SciTech Connect

    Rafter, Stephen E.; Crenshaw, D. Michael; Wiita, Paul J.

    2011-03-15

    We present a study of the extended radio emission in a sample of 8434 low-redshift (z < 0.35) broad-line active galactic nuclei (AGNs) from the Sloan Digital Sky Survey. To calculate the jet and lobe contributions to the total radio luminosity, we have taken the 846 radio core sources detected in our previous study of this sample and performed a systematic search in the FIRST database for extended radio emission that is likely associated with the optical counterparts. We found that 51 out of 846 radio core sources have extended emission (>4'' from the optical AGN) that is positively associated with the AGN, and we have identified an additional 12 AGNs with extended radio emission but no detectable radio core emission. Among these 63 AGNs, we found 6 giant radio galaxies, with projected emission exceeding 750 kpc in length, and several other AGNs with unusual radio morphologies also seen in higher redshift surveys. The optical spectra of many of the extended sources are similar to those of typical broad-line radio galaxy spectra, having broad H{alpha} emission lines with boxy profiles and large M{sub BH}. With extended emission taken into account, we find strong evidence for a bimodal distribution in the radio-loudness parameter R ({identical_to}{nu}{sub radio} L{sub radio}/{nu}{sub opt} L{sub opt}), where the lower radio luminosity core-only sources appear as a population separate from the extended sources, with a dividing line at log(R) {approx}1.75. This dividing line ensures that these are indeed the most radio-loud AGNs, which may have different or extreme physical conditions in their central engines when compared to the more numerous radio-quiet AGNs.

  20. Mast cell activation syndrome masquerading as agranulocytosis.

    PubMed

    Afrin, Lawrence B

    2012-01-01

    Acquired agranulocytosis is a rare, life-threatening disorder. The few known causes/associations usually are readily identifiable (e.g., drug reaction, Felty syndrome, megaloblastosis, large granular lymphocytic leukemia, etc.). We report a novel association with mast cell disease. A 61-year-old morbidly obese man developed rheumatoid arthritis unresponsive to several medications. Agranulocytosis developed shortly after sulfasalazine was started but did not improve when the drug was soon stopped. Other symptoms across many systems developed including hives and presyncope. Marrow aspiration and biopsy showed only neutropenia. Serum tryptase was mildly elevated; urinary prostaglandin D2 was markedly elevated. Other causes were not found. Mast cell activation syndrome (MCAS) was diagnosed. Oral antihistamines, montelukast, and cromolyn were unhelpful; aspirin was initially felt contraindicated. Imatinib immediately increased neutrophils from 0% to 25% but did not help symptoms; subsequent addition of aspirin increased neutrophils further and abated symptoms. Different presentations of different MCAS patients reflect elaboration of different mediators likely consequent to different Kit mutations. Mast cells (MCs) help regulate adipocytes, and adipocytes can inhibit granulopoiesis; thus, a Kit-mutated MC clone may have directly and/or indirectly driven agranulocytosis. MCAS should be considered in otherwise idiopathic agranulocytosis presenting with comorbidities best explained by MC mediator release. PMID:22338992

  1. [Inhibitory interactions in neuronal networks including cells of the auditory cortex and the medial geniculate body].

    PubMed

    Sil'kis, I G

    1994-01-01

    Cross-correlation method was used for revealing effective inhibitory interactions in neural networks containing simultaneously recorded neurons from different loci of auditory cortex (A1) and medial geniculate body (MGB). It was shown that (i) inhibitory connections were "divergent", i. e., one neuron in A1 (MGB) depressed activity of neurons in different loci of A1 and MGB simultaneously; (ii) inputs to inhibitory neuron were "convergent", i.e., one neuron in A1 (MGB) was excited by neurons from different loci of A1 and MGB simultaneously. There were inhibitory neurons which selectively depressed activity of only one neighbouring neuron. The results allow to suggest that the same inhibitory neuron may be involved in afferent and feedback inhibition. We supposed that the principles of organization of inhibitory connections in thalamo-cortical networks underlie the observed exceptions to mapping (tonotopic) principle of organization of receptive fields of A1 and MGB. PMID:7879428

  2. 45 CFR 287.130 - Can NEW Program activities include job market assessments, job creation and economic development...

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... assessments, job creation and economic development activities? 287.130 Section 287.130 Public Welfare... creation and economic development activities? (a) A Tribe may conduct job market assessments within its NEW Program. These might include the following: (1) Consultation with the Tribe's economic development...

  3. 45 CFR 287.130 - Can NEW Program activities include job market assessments, job creation and economic development...

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... assessments, job creation and economic development activities? 287.130 Section 287.130 Public Welfare... creation and economic development activities? (a) A Tribe may conduct job market assessments within its NEW Program. These might include the following: (1) Consultation with the Tribe's economic development...

  4. 45 CFR 287.130 - Can NEW Program activities include job market assessments, job creation and economic development...

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... assessments, job creation and economic development activities? 287.130 Section 287.130 Public Welfare... creation and economic development activities? (a) A Tribe may conduct job market assessments within its NEW Program. These might include the following: (1) Consultation with the Tribe's economic development...

  5. 45 CFR 287.130 - Can NEW Program activities include job market assessments, job creation and economic development...

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... assessments, job creation and economic development activities? 287.130 Section 287.130 Public Welfare... creation and economic development activities? (a) A Tribe may conduct job market assessments within its NEW Program. These might include the following: (1) Consultation with the Tribe's economic development...

  6. 45 CFR 287.130 - Can NEW Program activities include job market assessments, job creation and economic development...

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... assessments, job creation and economic development activities? 287.130 Section 287.130 Public Welfare... creation and economic development activities? (a) A Tribe may conduct job market assessments within its NEW Program. These might include the following: (1) Consultation with the Tribe's economic development...

  7. Activity of tigecycline tested against a global collection of Enterobacteriaceae, including tetracycline-resistant isolates.

    PubMed

    Fritsche, Thomas R; Strabala, Patty A; Sader, Helio S; Dowzicky, Michael J; Jones, Ronald N

    2005-07-01

    Steadily increasing resistance among the Enterobacteriaceae to beta-lactams, fluoroquinolones, aminoglycosides, tetracyclines, and trimethoprim/sulfamethoxazole has compromised the utility of these commonly used antimicrobial classes for many community- or hospital-acquired infections. The development of tigecycline, the sentinel representative of a novel class of broad-spectrum agents (the glycylcyclines), represents an important milestone in addressing this critical need. Resistance to tigecycline might be expected to occur via the same mechanisms that produce tetracycline resistance; however, tigecycline remains stable and largely unaffected by the commonly occurring efflux and ribosomal protection resistance mechanisms. In this study, an international collection of Enterobacteriaceae (11327 isolates; 32.8% tetracycline-resistant) from global surveillance studies (2000-2004) were evaluated against tigecycline and other comparator antimicrobials. Although the most active agents were the carbapenems and aminoglycosides (97.5-99.7% susceptible), tigecycline displayed high potency (MIC50 and MIC90, 0.25 and 1 microg/mL) with 95.7% of all strains being inhibited at < or =2 microg/mL. Despite higher MIC values observed with Serratia spp. and Proteae, between 90.5% and 97.5% of isolates were inhibited by < or =4 microg/mL of tigecycline. Tetracycline-resistant populations demonstrated only modest decreases in potency to tigecycline, which appeared to be species-dependent (up to 2-fold only for Escherichia coli, Salmonella spp., Shigella spp., and Panteoa agglomerans; and up to 4-fold for Klebsiella spp., Enterobacter spp., and Citrobacter spp.). Among E. coli (263 isolates) and Klebsiella spp. (356) that meet recognized screening definitions for extended-spectrum beta-lactamase production, 100.0% and 94.4% were inhibited by tigecycline at 2 microg/mL, respectively. These findings confirm that tigecycline exhibits potency, breadth of spectrum, and stability to the

  8. Electrodes including a polyphosphazene cyclomatrix, methods of forming the electrodes, and related electrochemical cells

    SciTech Connect

    Gering, Kevin L; Stewart, Frederick F; Wilson, Aaron D; Stone, Mark L

    2014-10-28

    An electrode comprising a polyphosphazene cyclomatrix and particles within pores of the polyphosphazene cyclomatrix. The polyphosphazene cyclomatrix comprises a plurality of phosphazene compounds and a plurality of cross-linkages. Each phosphazene compound of the plurality of phosphazene compounds comprises a plurality of phosphorus-nitrogen units, and at least one pendant group bonded to each phosphorus atom of the plurality of phosphorus-nitrogen units. Each phosphorus-nitrogen unit is bonded to an adjacent phosphorus-nitrogen unit. Each cross-linkage of the plurality of cross-linkages bonds at least one pendant group of one phosphazene compound of the plurality of phosphazene compounds with the at least one pendant group of another phosphazene compound of the plurality of phosphazene compounds. A method of forming a negative electrode and an electrochemical cell are also described.

  9. Characterization of tissue plasminogen activator binding proteins isolated from endothelial cells and other cell types

    SciTech Connect

    Beebe, D.P.; Wood, L.L.; Moos, M. )

    1990-07-15

    Human tissue plasminogen activator (t-PA) was shown to bind specifically to human osteosarcoma cells (HOS), and human epidermoid carcinoma cells (A-431 cells). Crosslinking studies with DTSSP demonstrated high molecular weight complexes (130,000) between {sup 125}I-t-PA and cell membrane protein on human umbilical vein endothelial cells (HUVEC), HOS, and A-431 cells. A 48-65,000 molecular weight complex was demonstrated after crosslinking t-PA peptide (res. 7-20) to cells. Ligand blotting of cell lysates which had been passed over a t-PA affinity column revealed binding of t-PA to 54,000 and 95,000 molecular weight proteins. Several t-PA binding proteins were identified in immunopurified cell lysates, including tubulin beta chain, plasminogen activator inhibitor type 1 and single chain urokinase.

  10. Novel Treatment for Mantle Cell Lymphoma including Therapy-resistant Tumor by NF-κB and mTOR Dual Targeting Approach

    PubMed Central

    Chaturvedi, Nagendra K.; Rajule, Rajkumar N.; Shukla, Ashima; Radhakrishnan, Prakash; Todd, Gordon L.; Natarajan, Amarnath; Vose, Julie M.; Joshi, Shantaram S.

    2014-01-01

    Mantle cell lymphoma (MCL) is one of the most aggressive B cell non-Hodgkin lymphomas with a median survival of about five years. Currently, there is no curative therapy available for refractory MCL because of relapse from therapy-resistant tumor cells. The NF-κB and mTOR pathways are constitutively active in refractory MCL leading to increased proliferation and survival. Targeting these pathways is an ideal strategy to improve therapy for refractory MCL. Therefore, we investigated the in vitro and in vivo antilymphoma activity and associated molecular mechanism of action of a novel compound 13-197, a quinoxaline analog that specifically perturbs IκB kinase (IKK) β, a key regulator of the NF-κB pathway. 13-197 decreased the proliferation and induced apoptosis in MCL cells including therapy-resistant cells compared to control cells. Furthermore, we observed down-regulation of IκBα phosphorylation and inhibition of NF-κB nuclear translocation by 13-197 in MCL cells. In addition, NF-κB regulated genes such as cyclin D1, Bcl-XL and Mcl-1 were down-regulated in 13-197-treated cells. 13-197 also inhibited the phosphorylation of S6K and 4E-BP1, the downstream molecules of mTOR pathway that are also activated in refractory MCL. Further, 13-197 reduced the tumor burden in vivo in the kidney, liver, and lungs of therapy-resistant MCL bearing NOD-SCID mice compared to vehicle treated mice; indeed, 13-197 significantly increased the survival of MCL transplanted mice. Together, results suggest that 13-197 as a single agent disrupts the NF-κB and mTOR pathways leading suppression of proliferation and increased apoptosis in malignant MCL cells including reduction in tumor burden in mice. PMID:23963361

  11. Should Physical Activity Be Included in Nutrition Education? A Comparison of Nutrition Outcomes with and without In-Class Activities

    ERIC Educational Resources Information Center

    Palmer-Keenan, Debra M.; Corda, Kirsten

    2014-01-01

    Limited-resource adults' dietary intakes and nutrition behaviors improve as a result of Expanded Food and Nutrition Education Program (EFNEP)/Supplemental Nutrition Assistance Program Education (SNAP-Ed) participation; however, physical activity education is needed for improved health. The experimental study reported here assessed if spending…

  12. Upstream stimulatory factor activates the vasopressin promoter via multiple motifs, including a non-canonical E-box.

    PubMed Central

    Coulson, Judy M; Edgson, Jodie L; Marshall-Jones, Zoe V; Mulgrew, Robert; Quinn, John P; Woll, Penella J

    2003-01-01

    We have described previously a complex E-box enhancer (-147) of the vasopressin promoter in small-cell lung cancer (SCLC) extracts [Coulson, Fiskerstrand, Woll and Quinn, (1999) Biochem. J. 344, 961-970]. Upstream stimulatory factor (USF) heterodimers were one of the complexes binding to this site in vitro. We now report that USF overexpression in non-SCLC (NSCLC) cells can functionally activate vasopressin promoter-driven reporters that are otherwise inactive in this type of lung cancer cell. Site-directed mutagenesis and electrophoretic mobility-shift analysis demonstrate that although the -147 E-box contributes, none of the previously predicted E-boxes (-147, -135, -34) wholly account for this USF-mediated activation in NSCLC. 5' Deletion showed the key promoter region as -52 to +42; however, USF-2 binding was not reliant on the -34 E-box, but on a novel adjacent CACGGG non-canonical E-box at -42 (motif E). This mediated USF binding in both SCLC and USF-2-transfected NSCLC cells. Mutation of motif E or the non-canonical TATA box abolished activity, implying both are required for transcriptional initiation on overexpression of USF-2. Co-transfected dominant negative USF confirmed that binding was required through motif E for function, but that the classical activation domain of USF was not essential. USF-2 bound motif E with 10-fold lower affinity than the -147 E-box. In NSCLC, endogenous USF-2 expression is low, and this basal level appears to be insufficient to activate transcription of arginine vasopressin (AVP). In summary, we have demonstrated a novel mechanism for USF activation, which contributes to differential vasopressin expression in lung cancer. PMID:12403649

  13. Rapidly rendering cells phagocytic through a cell surface display technique and concurrent Rac activation.

    PubMed

    Onuma, Hiroki; Komatsu, Toru; Arita, Makoto; Hanaoka, Kenjiro; Ueno, Tasuku; Terai, Takuya; Nagano, Tetsuo; Inoue, Takanari

    2014-07-15

    Cell surfaces represent a platform through which extracellular signals that determine diverse cellular processes, including migration, division, adhesion, and phagocytosis, are transduced. Techniques to rapidly reconfigure the surface properties of living cells should thus offer the ability to harness these cellular functions. Although the molecular mechanism of phagocytosis is well characterized, the minimal molecular players that are sufficient to activate this elaborate process remain elusive. We developed and implemented a technique to present a molecule of interest at the cell surface in an inducible manner on a time scale of minutes. We simultaneously induced the cell surface display of the C2 domain of milk fat globule epidermal growth factor factor 8 (MFG-E8) and activated the intracellular small guanosine triphosphatase Rac, which stimulates actin polymerization at the cell periphery. The C2 domain binds to phosphatidylserine, a lipid exposed on the surface of apoptotic cells. By integrating the stimulation of these two processes, we converted HeLa cells into a phagocytic cell line that bound to and engulfed apoptotic human Jurkat cells. Inducing either the cell surface display of the C2 domain or activating Rac alone was not sufficient to stimulate phagocytosis, which suggests that attachment to the target cell and actin reorganization together constitute the minimal molecular events that are needed to induce phagocytosis. This cell surface display technique might be useful as part of a targeted, cell-based therapy in which unwanted cells with characteristic surface molecules could be rapidly consumed by engineered cells. PMID:25028719

  14. Rapidly rendering cells phagocytic through a cell-surface display technique and concurrent Rac activation

    PubMed Central

    Onuma, Hiroki; Arita, Makoto; Hanaoka, Kenjiro; Ueno, Tasuku; Terai, Takuya; Nagano, Tetsuo

    2014-01-01

    Cell surfaces represent a platform through which extracellular signals that determine diverse cellular processes, including migration, division, adhesion, and phagocytosis, are transduced. Techniques to rapidly reconfigure the surface properties of living cells should thus offer the ability to harness these cellular functions. Although the molecular mechanism of phagocytosis is well-characterized, the minimal molecular players that are sufficient to activate this elaborate process remain elusive. We developed and implemented a technique to present a molecule of interest at the cell surface in an inducible manner on a timescale of minutes. We simultaneously induced the cell-surface display of the C2 domain of milk fat globule-EGF factor 8 (MFG-E8) and activated the intracellular small guanosine triphosphatase Rac, which stimulates actin polymerization at the cell periphery. The C2 domain binds to phosphatidylserine, a lipid exposed on the surface of apoptotic cells. By integrating the stimulation of these two processes, we converted HeLa cells into a phagocytic cell line that bound to and engulfed apoptotic human Jurkat cells. Inducing either the cell-surface display of the C2 domain or activating Rac alone was not sufficient to stimulate phagocytosis, which suggests that attachment to the target cell and actin reorganization together constitute the minimal molecular events that are needed to induce phagocytosis. This cell-surface display technique might be useful as part of a targeted, cell-based therapy in which unwanted cells with characteristic surface molecules could be rapidly consumed by engineered cells. PMID:25028719

  15. Activity-driven fluctuations in living cells

    NASA Astrophysics Data System (ADS)

    Fodor, É.; Guo, M.; Gov, N. S.; Visco, P.; Weitz, D. A.; van Wijland, F.

    2015-05-01

    We propose a model for the dynamics of a probe embedded in a living cell, where both thermal fluctuations and nonequilibrium activity coexist. The model is based on a confining harmonic potential describing the elastic cytoskeletal matrix, which undergoes random active hops as a result of the nonequilibrium rearrangements within the cell. We describe the probe's statistics and we bring forth quantities affected by the nonequilibrium activity. We find an excellent agreement between the predictions of our model and experimental results for tracers inside living cells. Finally, we exploit our model to arrive at quantitative predictions for the parameters characterizing nonequilibrium activity, such as the typical time scale of the activity and the amplitude of the active fluctuations.

  16. HIV-1 Nef and Vpu Are Functionally Redundant Broad-Spectrum Modulators of Cell Surface Receptors, Including Tetraspanins

    PubMed Central

    Haller, Claudia; Müller, Birthe; Fritz, Joëlle V.; Lamas-Murua, Miguel; Stolp, Bettina; Pujol, François M.; Keppler, Oliver T.

    2014-01-01

    ABSTRACT HIV-1 Nef and Vpu are thought to optimize virus replication in the infected host, at least in part via their ability to interfere with vesicular host cell trafficking. Despite the use of distinct molecular mechanisms, Nef and Vpu share specificity for some molecules such as CD4 and major histocompatibility complex class I (MHC-I), while disruption of intracellular transport of the host cell restriction factor CD317/tetherin represents a specialized activity of Vpu not exerted by HIV-1 Nef. To establish a profile of host cell receptors whose intracellular transport is affected by Nef, Vpu, or both, we comprehensively analyzed the effect of these accessory viral proteins on cell surface receptor levels on A3.01 T lymphocytes. Thirty-six out of 105 detectable receptors were significantly downregulated by HIV-1 Nef, revealing a previously unappreciated scope with which HIV-1 Nef remodels the cell surface of infected cells. Remarkably, the effects of HIV-1 Vpu on host cell receptor exposure largely matched those of HIV-1 Nef in breadth and specificity (32 of 105, all also targeted by Nef), even though the magnitude was generally less pronounced. Of particular note, cell surface exposure of all members of the tetraspanin (TSPAN) protein family analyzed was reduced by both Nef and Vpu, and the viral proteins triggered the enrichment of TSPANs in a perinuclear area of the cell. While Vpu displayed significant colocalization and physical association with TSPANs, interactions of Nef with TSPANs were less robust. TSPANs thus emerge as a major target of deregulation in host cell vesicular transport by HIV-1 Nef and Vpu. The conservation of this activity in two independent accessory proteins suggests its importance for the spread of HIV-1 in the infected host. IMPORTANCE In this paper, we define that HIV-1 Nef and Vpu display a surprising functional overlap and affect the cell surface exposure of a previously unexpected breadth of cellular receptors. Our analyses

  17. Active unjamming of confluent cell layers

    NASA Astrophysics Data System (ADS)

    Marchetti, M. Cristina

    Cell motion inside dense tissues governs many biological processes, including embryonic development and cancer metastasis, and recent experiments suggest that these tissues exhibit collective glassy behavior. Motivated by these observations, we have studied a model of dense tissues that combines self-propelled particle models and vertex models of confluent cell layers. In this model, referred to as self-propelled Voronoi (SPV), cells are described as polygons in a Voronoi tessellation with directed noisy cell motility and interactions governed by a shape energy that incorporates the effects of cell volume incompressibility, contractility and cell-cell adhesion. Using this model, we have demonstrated a new density-independent solid-liquid transition in confluent tissues controlled by cell motility and a cell-shape parameter measuring the interplay of cortical tension and cell-cell adhesion. An important insight of this work is that the rigidity and dynamics of cell layers depends sensitively on cell shape. We have also used the SPV model to test a new method developed by our group to determine cellular forces and tissue stresses from experimentally accessible cell shapes and traction forces, hence providing the spatio-temporal distribution of stresses in motile dense tissues. This work was done with Dapeng Bi, Lisa Manning and Xingbo Yang. MCM was supported by NSF-DMR-1305184 and by the Simons Foundation.

  18. Cathode material comparison of thermal runaway behavior of Li-ion cells at different state of charges including over charge

    NASA Astrophysics Data System (ADS)

    Mendoza-Hernandez, Omar Samuel; Ishikawa, Hiroaki; Nishikawa, Yuuki; Maruyama, Yuki; Umeda, Minoru

    2015-04-01

    The analysis of Li-ion secondary cells under outstanding conditions, as overcharge and high temperatures, is important to determine thermal abuse characteristics of electroactive materials and precise risk assessments on Li-ion cells. In this work, the thermal runaway behavior of LiCoO2 and LiMn2O4 cathode materials were compared at different state of charges (SOCs), including overcharge, by carrying out accelerating rate calorimetry (ARC) measurements using 18650 Li-ion cells. Onset temperatures of self-heating reactions and thermal runaway behavior were identified, and by using these onset points thermal mapping plots were made. We were able to identify non-self-heating, self-heating and thermal runaway regions as a function of state of charge and temperature. The cell using LiMn2O4 cathode material was found to be more thermally stable than the cell using LiCoO2. In parallel with the ARC measurements, the electrochemical behavior of the cells was monitored by measuring the OCV and internal resistance of the cells. The electrochemical behavior of the cells showed a slightly dependency on SOC.

  19. Kinetic discrimination in T-cell activation.

    PubMed Central

    Rabinowitz, J D; Beeson, C; Lyons, D S; Davis, M M; McConnell, H M

    1996-01-01

    We propose a quantitative model for T-cell activation in which the rate of dissociation of ligand from T-cell receptors determines the agonist and antagonist properties of the ligand. The ligands are molecular complexes between antigenic peptides and proteins of the major histocompatibility complex on the surfaces of antigen-presenting cells. Binding of ligand to receptor triggers a series of biochemical reactions in the T cell. If the ligand dissociates after these reactions are complete, the T cell receives a positive activation signal. However, dissociation of ligand after completion of the first reaction but prior to generation of the final products results in partial T-cell activation, which acts to suppress a positive response. Such a negative signal is brought about by T-cell ligands containing the variants of antigenic peptides referred to as T-cell receptor antagonists. Results of recent experiments with altered peptide ligands compare favorably with T-cell responses predicted by this model. PMID:8643643

  20. The VOLMAX Transient Electromagnetic Modeling System, Including Sub-Cell Slots and Wires on Random Non-Orthogonal Cells

    SciTech Connect

    Riley, D.J.; Turner, C.D.

    1997-12-31

    VOLMAX is a three-dimensional transient volumetric Maxwell equation solver that operates on standard rectilinear finite-difference time-domain (FDTD) grids, non-orthogonal unstructured grids, or a combination of both types (hybrid grids). The algorithm is fully explicit. Open geometries are typically solved by embedding multiple unstructured regions into a simple rectilinear FDTD mesh. The grid types are fully connected at the mesh interfaces without the need for complex spatial interpolation. The approach permits detailed modeling of complex geometry while mitigating the large cell count typical of non-orthogonal cells such as tetrahedral elements. To further improve efficiency, the unstructured region carries a separate time step that sub-cycles relative to the time-step used in the FDTD mesh.

  1. Lipolytic activity in adipocyte cell fractions.

    PubMed

    Oschry, Y; Shapiro, B

    1980-05-28

    Adipocytes release only negligible amounts of free fatty acids unless stimulated, but reveal considerable lipolytic activity when homogenized. Epinephrine treatment of the cells caused only a 20-40% increase in the activity of infranatants of homogenates while raising the activity associated with the fat layer up to 10-fold. Full activity (i.e. that of intact-activated cells) could be revealed by epinephrine treatment of the homogenate as well as by sonication of the fat layer in buffer. The combination of both treatments did not yield higher activities. The fat cake contains the bulk of the potential activities which are only realized when dispersed in the aqueous phase by sonication, or upon hormone activation of the whole homogenate. Increase in activity could also be obtained by removal of most of the lipid from the fat layer by extraction with petroleum ether. Re-introduction of extracted lipid inhibited lipolysis. The active enzyme could be separated by flotation at 1.12 specific gravity. The data suggest that the lack of activity in the intact non-stimulated cell may be due to the lack of availability of the aqueous phase to the enzyme. PMID:7378439

  2. Short-Range Temporal Interactions in Sleep; Hippocampal Spike Avalanches Support a Large Milieu of Sequential Activity Including Replay.

    PubMed

    Mahoney, J Matthew; Titiz, Ali S; Hernan, Amanda E; Scott, Rod C

    2016-01-01

    Hippocampal neural systems consolidate multiple complex behaviors into memory. However, the temporal structure of neural firing supporting complex memory consolidation is unknown. Replay of hippocampal place cells during sleep supports the view that a simple repetitive behavior modifies sleep firing dynamics, but does not explain how multiple episodes could be integrated into associative networks for recollection during future cognition. Here we decode sequential firing structure within spike avalanches of all pyramidal cells recorded in sleeping rats after running in a circular track. We find that short sequences that combine into multiple long sequences capture the majority of the sequential structure during sleep, including replay of hippocampal place cells. The ensemble, however, is not optimized for maximally producing the behavior-enriched episode. Thus behavioral programming of sequential correlations occurs at the level of short-range interactions, not whole behavioral sequences and these short sequences are assembled into a large and complex milieu that could support complex memory consolidation. PMID:26866597

  3. Short-Range Temporal Interactions in Sleep; Hippocampal Spike Avalanches Support a Large Milieu of Sequential Activity Including Replay

    PubMed Central

    Mahoney, J. Matthew; Titiz, Ali S.; Hernan, Amanda E.; Scott, Rod C.

    2016-01-01

    Hippocampal neural systems consolidate multiple complex behaviors into memory. However, the temporal structure of neural firing supporting complex memory consolidation is unknown. Replay of hippocampal place cells during sleep supports the view that a simple repetitive behavior modifies sleep firing dynamics, but does not explain how multiple episodes could be integrated into associative networks for recollection during future cognition. Here we decode sequential firing structure within spike avalanches of all pyramidal cells recorded in sleeping rats after running in a circular track. We find that short sequences that combine into multiple long sequences capture the majority of the sequential structure during sleep, including replay of hippocampal place cells. The ensemble, however, is not optimized for maximally producing the behavior-enriched episode. Thus behavioral programming of sequential correlations occurs at the level of short-range interactions, not whole behavioral sequences and these short sequences are assembled into a large and complex milieu that could support complex memory consolidation. PMID:26866597

  4. Laser activated nanothermolysis of leukemia cells monitored by photothermal microscopy

    NASA Astrophysics Data System (ADS)

    Lapotko, Dmitri; Lukianova, Ekaterina; Shnip, Alexander; Zheltov, George; Potapnev, Michail; Savitsky, Valeriy; Klimovich, Olga; Oraevsky, Alexander

    2005-04-01

    We are developing new diagnostic and therapeutic technologies for leukemia based on selective targeting of leukemia cells with gold nanoparticles and thermomechanical destruction of the tumor cells with laser-induced microbubbles. Clusters of spherical gold nanoparticles that have strong optical absorption of laser pulses at 532 nm served as nucleation sites of vapor microbubbles. The nanoparticles were targeted selectively to leukemia cells using leukemia-specific surface receptors and a set of two monoclonal antibodies. Application of a primary myeloid-specific antibody to tumor cells followed by targeting the cells with 30-nm nanoparticles conjugated with a secondary antibody (IgG) resulted in formation of nanoparticulate clusters due to aggregation of IgGs. Formation of clusters resulted in substantial decrease of the damage threshold for target cells. The results encourage development of Laser Activated Nanothermolysis as a Cell Elimination Therapy (LANCET) for leukemia. The proposed technology can be applied separately or in combination with chemotherapy for killing leukemia cells without damage to other blood cells. Potential applications include initial reduction of concentration of leukemia cells in blood prior to chemotherapy and treatment of residual tumor cells after the chemotherapy. Laser-induced bubbles in individual cells and cell damage were monitored by analyzing profile of photothermal response signals over the entire cell after irradiation with a single 10-ns long laser pulse. Photothermal microscopy was utilized for imaging formation of microbubbles around nanoparticulate clusters.

  5. 25 CFR 170.623 - How are IRR Program projects and activities included in a self-governance agreement?

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... other information required under 25 CFR 1000 subpart K. ... self-governance agreement? 170.623 Section 170.623 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE... self-governance agreement? To include an IRR Program project or activity in a self-governance...

  6. Effects of Leisure Education Programme Including Sportive Activities on Perceived Freedom in Leisure of Adolescents with Intellectual Disabilities

    ERIC Educational Resources Information Center

    Ertuzun, Ezgi

    2015-01-01

    The objective of this experimental study is to determine the effect of leisure education programme including sportive activities on the perceived freedom in leisure of adolescents with mild intellectual disabilities. The research was designed with an experimental group (n = 37) and a control group (n = 34), and was conducted among a total of 71…

  7. 25 CFR 170.137 - What types of activities can a recreation, tourism, and trails program include?

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 25 Indians 1 2014-04-01 2014-04-01 false What types of activities can a recreation, tourism, and trails program include? 170.137 Section 170.137 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR LAND AND WATER INDIAN RESERVATION ROADS PROGRAM Indian Reservation Roads Program Policy and Eligibility Recreation, Tourism and Trails...

  8. 25 CFR 170.623 - How are IRR Program projects and activities included in a self-governance agreement?

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... other information required under 25 CFR 1000 subpart K. ... self-governance agreement? 170.623 Section 170.623 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE... self-governance agreement? To include an IRR Program project or activity in a self-governance...

  9. 25 CFR 170.623 - How are IRR Program projects and activities included in a self-governance agreement?

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... other information required under 25 CFR 1000 subpart K. ... self-governance agreement? 170.623 Section 170.623 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE... self-governance agreement? To include an IRR Program project or activity in a self-governance...

  10. 25 CFR 170.623 - How are IRR Program projects and activities included in a self-governance agreement?

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... other information required under 25 CFR 1000 subpart K. ... self-governance agreement? 170.623 Section 170.623 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE... self-governance agreement? To include an IRR Program project or activity in a self-governance...

  11. 25 CFR 170.623 - How are IRR Program projects and activities included in a self-governance agreement?

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... other information required under 25 CFR 1000 subpart K. ... self-governance agreement? 170.623 Section 170.623 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE... self-governance agreement? To include an IRR Program project or activity in a self-governance...

  12. Lymphatic endothelial cells actively regulate prostate cancer cell invasion.

    PubMed

    Shah, Tariq; Wildes, Flonne; Kakkad, Samata; Artemov, Dmitri; Bhujwalla, Zaver M

    2016-07-01

    Lymphatic vessels serve as the primary route for metastatic spread to lymph nodes. However, it is not clear how interactions between cancer cells and lymphatic endothelial cells (LECs), especially within hypoxic microenvironments, affect the invasion of cancer cells. Here, using an MR compatible cell perfusion assay, we investigated the role of LEC-prostate cancer (PCa) cell interaction in the invasion and degradation of the extracellular matrix (ECM) by two human PCa cell lines, PC-3 and DU-145, under normoxia and hypoxia, and determined the metabolic changes that occurred under these conditions. We observed a significant increase in the invasion of ECM by invasive PC-3 cells, but not poorly invasive DU-145 cells when human dermal lymphatic microvascular endothelial cells (HMVEC-dlys) were present. Enhanced degradation of ECM by PC-3 cells in the presence of HMVEC-dlys identified interactions between HMVEC-dlys and PCa cells influencing cancer cell invasion. The enhanced ECM degradation was partly attributed to increased MMP-9 enzymatic activity in PC-3 cells when HMVEC-dlys were in close proximity. Significantly higher uPAR and MMP-9 expression levels observed in PC-3 cells compared to DU-145 cells may be one mechanism for increased invasion and degradation of matrigel by these cells irrespective of the presence of HMVEC-dlys. Hypoxia significantly decreased invasion by PC-3 cells, but this decrease was significantly attenuated when HMVEC-dlys were present. Significantly higher phosphocholine was observed in invasive PC-3 cells, while higher glycerophosphocholine was observed in DU-145 cells. These metabolites were not altered in the presence of HMVEC-dlys. Significantly increased lipid levels and lipid droplets were observed in PC-3 and DU-145 cells under hypoxia reflecting an adaptive survival response to oxidative stress. These results suggest that in vivo, invasive cells in or near lymphatic endothelial cells are likely to be more invasive and degrade the ECM

  13. Reserve, thin form-factor, hypochlorite-based cells for powering portable systems: Manufacture (including MEMS processes), performance and characterization

    NASA Astrophysics Data System (ADS)

    Cardenas-Valencia, Andres M.; Biver, Carl J.; Langebrake, Larry

    This work focuses on fabrication routes and performance evaluation of thin form-factors, reserve cells, as a powering alternative for expendable and/or remotely operated systems. The catalytic decomposition of sodium hypochlorite solutions is revisited herein with two cost-effective anodes: zinc and aluminum. Aluminum, even though the most expensive of the utilized anodes, constituted cells with double the energy content (up to 55 Wh kg -1) than those fabricated with zinc. Even though the hypochlorite concentration in the solution limits the cells' operational life, attractive performances (1.0 V with a current of 10 mA) for the manufactured cells are obtained. It is shown that micro fabrication processes, allowing for close electrodes interspacing, provided high faradic and columbic efficiencies of up to 70 and 100%, respectively. Obtained specific energies (50-120 Wh kg -1) are in the same order of magnitude than batteries currently used for powering deployable systems. Experimental results show that a simple model that linearly relates over potentials and the electrical load, adequately describe all the cell designs. A mathematical model based on a kinetic-mechanistic scheme that relates the current output as a function of time agrees fairly well with results obtained activating cells with various concentrations of NaOCl solutions.

  14. Enhancement of endothelial cell migration by constitutively active LPA{sub 1}-expressing tumor cells

    SciTech Connect

    Kitayoshi, Misaho; Kato, Kohei; Tanabe, Eriko; Yoshikawa, Kyohei; Fukui, Rie; Fukushima, Nobuyuki; Tsujiuchi, Toshifumi

    2012-06-01

    Highlights: Black-Right-Pointing-Pointer Mutated LPA{sub 1} stimulates cell migration of endothelial cells. Black-Right-Pointing-Pointer VEGF expressions are increased by mutated LPA{sub 1}. Black-Right-Pointing-Pointer LPA signaling via mutated LPA{sub 1} is involved in angiogenesis. Black-Right-Pointing-Pointer Mutated LPA{sub 1} promotes cancer cell progression. -- Abstract: Lysophosphatidic acid (LPA) receptors belong to G protein-coupled transmembrane receptors (LPA receptors; LPA{sub 1} to LPA{sub 6}). They indicate a variety of cellular response by the interaction with LPA, including cell proliferation, migration and differentiation. Recently, we have reported that constitutive active mutated LPA{sub 1} induced the strong biological effects of rat neuroblastoma B103 cells. In the present study, we examined the effects of mutated LPA{sub 1} on the interaction between B103 cells and endothelial F-2 cells. Each LPA receptor expressing B103 cells were maintained in serum-free DMEM and cell motility assay was performed with a Cell Culture Insert. When F-2 cells were cultured with conditioned medium from Lpar1 and Lpar3-expressing cells, the cell motility of F-2 cells was significantly higher than control cells. Interestingly, the motile activity of F-2 cells was strongly induced by mutated LPA{sub 1} than other cells, correlating with the expression levels of vascular endothelial growth factor (Vegf)-A and Vegf-C. Pretreatment of LPA signaling inhibitors inhibited F-2 cell motility stimulated by mutated LPA{sub 1}. These results suggest that activation of LPA signaling via mutated LPA{sub 1} may play an important role in the promotion of angiogenesis in rat neuroblastoma cells.

  15. Ceftaroline versus isolates from animal bite wounds: comparative in vitro activities against 243 isolates, including 156 Pasteurella species isolates.

    PubMed

    Goldstein, Ellie J C; Citron, Diane M; Merriam, C Vreni; Tyrrell, Kerin L

    2012-12-01

    More than 5 million Americans are bitten by animals, usually dogs, annually. Bite patients comprise ∼1% of all patients who visit emergency departments (300,000/year), and approximately 10,000 require hospitalization and intravenous antibiotics. Ceftaroline is the bioactive component of the prodrug ceftaroline fosamil, which is FDA approved for the treatment of acute bacterial skin and skin structure infections (ABSSSIs), including those containing methicillin-resistant Staphylococcus aureus (MRSA). There are no in vitro data about the activity of ceftaroline against Pasteurella multocida subsp. multocida and Pasteurella multocida subsp. septica, other Pasteurella spp., or other bite wound isolates. We therefore studied the in vitro activity of ceftaroline against 243 animal bite isolates. MICs were determined using the broth microdilution method according to CLSI guidelines. Comparator drugs included cefazolin, ceftriaxone, ertapenem, ampicillin-sulbactam, azithromycin, doxycycline, and sulfamethoxazole-trimethoprim (SMX-TMP). Ceftaroline was the most active agent against all 5 Pasteurella species, including P. multocida subsp. multocida and P. multocida subsp. septica, with a maximum MIC of ≤0.008 μg/ml; more active than ceftriaxone and ertapenem (MIC(90)s, ≤0.015 μg/ml); and more active than cefazolin (MIC(90), 0.5 μg/ml) doxycycline (MIC(90), 0.125 μg/ml), azithromycin (MIC(90), 0.5 μg/ml), ampicillin-sulbactam (MIC(90), 0.125 μg/ml), and SMX-TMP (MIC(90), 0.125 μg/ml). Ceftaroline was also very active against all S. aureus isolates (MIC(90), 0.125 μg/ml) and other Staphylococcus and Streptococcus species, with a maximum MIC of 0.125 μg/ml against all bite isolates tested. Ceftaroline has potential clinical utility against infections involving P. multocida, other Pasteurella species, and aerobic Gram-positive isolates, including S. aureus. PMID:23027193

  16. Ceftaroline versus Isolates from Animal Bite Wounds: Comparative In Vitro Activities against 243 Isolates, Including 156 Pasteurella Species Isolates

    PubMed Central

    Citron, Diane M.; Merriam, C. Vreni; Tyrrell, Kerin L.

    2012-01-01

    More than 5 million Americans are bitten by animals, usually dogs, annually. Bite patients comprise ∼1% of all patients who visit emergency departments (300,000/year), and approximately 10,000 require hospitalization and intravenous antibiotics. Ceftaroline is the bioactive component of the prodrug ceftaroline fosamil, which is FDA approved for the treatment of acute bacterial skin and skin structure infections (ABSSSIs), including those containing methicillin-resistant Staphylococcus aureus (MRSA). There are no in vitro data about the activity of ceftaroline against Pasteurella multocida subsp. multocida and Pasteurella multocida subsp. septica, other Pasteurella spp., or other bite wound isolates. We therefore studied the in vitro activity of ceftaroline against 243 animal bite isolates. MICs were determined using the broth microdilution method according to CLSI guidelines. Comparator drugs included cefazolin, ceftriaxone, ertapenem, ampicillin-sulbactam, azithromycin, doxycycline, and sulfamethoxazole-trimethoprim (SMX-TMP). Ceftaroline was the most active agent against all 5 Pasteurella species, including P. multocida subsp. multocida and P. multocida subsp. septica, with a maximum MIC of ≤0.008 μg/ml; more active than ceftriaxone and ertapenem (MIC90s, ≤0.015 μg/ml); and more active than cefazolin (MIC90, 0.5 μg/ml) doxycycline (MIC90, 0.125 μg/ml), azithromycin (MIC90, 0.5 μg/ml), ampicillin-sulbactam (MIC90, 0.125 μg/ml), and SMX-TMP (MIC90, 0.125 μg/ml). Ceftaroline was also very active against all S. aureus isolates (MIC90, 0.125 μg/ml) and other Staphylococcus and Streptococcus species, with a maximum MIC of 0.125 μg/ml against all bite isolates tested. Ceftaroline has potential clinical utility against infections involving P. multocida, other Pasteurella species, and aerobic Gram-positive isolates, including S. aureus. PMID:23027193

  17. Alkaline pH activates the transport activity of GLUT1in L929 fibroblast cells

    PubMed Central

    Gunnink, Stephen M.; Kerk, Samuel A.; Kuiper, Benjamin D.; Alabi, Ola D.; Kuipers, David P.; Praamsma, Riemer C.; Wrobel, Kathryn E.; Louters, Larry L.

    2016-01-01

    The widely expressed mammalian glucose transporter, GLUT1, can be acutely activated in L929 fibroblast cells by a variety of conditions, including glucose deprivation, or treatment with various respiration inhibitors. Known thiol reactive compounds including phenylarsine oxide and nitroxyl are the fastest acting stimulators of glucose uptake, implicating cysteine biochemistry as critical to the acute activation of GLUT1. In this study, we report that in L929 cells glucose uptake increases 6-fold as the pH of the uptake solution is increased from 6 to 9 with the half-maximal activation at pH 7.5; consistent with the pKa of cysteine residues. This pH effect is essentially blocked by the pretreatment of the cells with either iodoacetamide or cinnamaldehyde, compounds that form covalent adducts with reduced cysteine residues. In addition, the activation by alkaline pH is not additive at pH 8 with known thiol reactive activators such as phenylarsine oxide or hydroxylamine. Kinetic analysis in L929 cells at pH 7 and 8 indicate that alkaline conditions both increases the Vmax and decreases the Km of transport. This is consistent with the observation that pH activation is additive to methylene blue, which activates uptake by increasing the Vmax, as well as to berberine, which activates uptake by decreasing the Km. This suggests that cysteine biochemistry is utilized in both methylene blue and berberine activation of glucose uptake. In contrast a pH increase from 7 to 8 in HCLE cells does not further activate glucose uptake. HCLE cells have a 25-fold higher basal glucose uptake rate than L929 cells and the lack of a pH effect suggests that the cysteine biochemistry has already occurred in HCLE cells. The data are consistent with pH having a complex mechanism of action, but one likely mediated by cysteine biochemistry. PMID:24333987

  18. Alkaline pH activates the transport activity of GLUT1 in L929 fibroblast cells.

    PubMed

    Gunnink, Stephen M; Kerk, Samuel A; Kuiper, Benjamin D; Alabi, Ola D; Kuipers, David P; Praamsma, Riemer C; Wrobel, Kathryn E; Louters, Larry L

    2014-04-01

    The widely expressed mammalian glucose transporter, GLUT1, can be acutely activated in L929 fibroblast cells by a variety of conditions, including glucose deprivation, or treatment with various respiration inhibitors. Known thiol reactive compounds including phenylarsine oxide and nitroxyl are the fastest acting stimulators of glucose uptake, implicating cysteine biochemistry as critical to the acute activation of GLUT1. In this study, we report that in L929 cells glucose uptake increases 6-fold as the pH of the uptake solution is increased from 6 to 9 with the half-maximal activation at pH 7.5; consistent with the pKa of cysteine residues. This pH effect is essentially blocked by the pretreatment of the cells with either iodoacetamide or cinnamaldehyde, compounds that form covalent adducts with reduced cysteine residues. In addition, the activation by alkaline pH is not additive at pH 8 with known thiol reactive activators such as phenylarsine oxide or hydroxylamine. Kinetic analysis in L929 cells at pH 7 and 8 indicate that alkaline conditions both increases the Vmax and decreases the Km of transport. This is consistent with the observation that pH activation is additive to methylene blue, which activates uptake by increasing the Vmax, as well as to berberine, which activates uptake by decreasing the Km. This suggests that cysteine biochemistry is utilized in both methylene blue and berberine activation of glucose uptake. In contrast a pH increase from 7 to 8 in HCLE cells does not further activate glucose uptake. HCLE cells have a 25-fold higher basal glucose uptake rate than L929 cells and the lack of a pH effect suggests that the cysteine biochemistry has already occurred in HCLE cells. The data are consistent with pH having a complex mechanism of action, but one likely mediated by cysteine biochemistry. PMID:24333987

  19. Regulation of the proliferation of colon cancer cells by compounds that affect glycolysis, including 3-bromopyruvate, 2-deoxyglucose and biguanides.

    PubMed

    Lea, Michael A; Qureshi, Mehreen S; Buxhoeveden, Michael; Gengel, Nicolette; Kleinschmit, Jessica; Desbordes, Charles

    2013-02-01

    In previous studies performed by our group, we observed that 2-deoxyglucose blocked the acidification of the medium used for culture of colon cancer cells caused by incubation with biguanides and it had an additive inhibitory effect on growth. In the present work, we found that 3-bromopyruvate can also prevent the lowering of pH caused by biguanide treatment. 3-Bromopyruvate inhibited colonic cancer cell proliferation, but the effect was not always additive to that of biguanides and an additive effect was more notable in combined treatment with 3-bromopyruvate and 2-deoxyglucose. The induction of alkaline phosphatase activity by butyrate was not consistently affected by combination with other agents that modified glucose metabolism. The drug combinations that were examined inhibited proliferation of wild-type and p53-null cells and affected colonic cancer lines with different growth rates. PMID:23393330

  20. Activation of Human T-Helper/Inducer Cell, T-Cytotoxic Cell, B-Cell, and Natural Killer (NK)-Cells and induction of Natural Killer Cell Activity against K562 Chronic Myeloid Leukemia Cells with Modified Citrus Pectin

    PubMed Central

    2011-01-01

    Background Modified citrus pectin (MCP) is known for its anti-cancer effects and its ability to be absorbed and circulated in the human body. In this report we tested the ability of MCP to induce the activation of human blood lymphocyte subsets like T, B and NK-cells. Methods MCP treated human blood samples were incubated with specific antibody combinations and analyzed in a flow cytometer using a 3-color protocol. To test functionality of the activated NK-cells, isolated normal lymphocytes were treated with increasing concentrations of MCP. Log-phase PKH26-labeled K562 leukemic cells were added to the lymphocytes and incubated for 4 h. The mixture was stained with FITC-labeled active form of caspase 3 antibody and analyzed by a 2-color flow cytometry protocol. The percentage of K562 cells positive for PKH26 and FITC were calculated as the dead cells induced by NK-cells. Monosaccharide analysis of the MCP was performed by high-performance anion-exchange chromatography with pulse amperometric detection (HPAEC-PAD). Results MCP activated T-cytotoxic cells and B-cell in a dose-dependent manner, and induced significant dose-dependent activation of NK-cells. MCP-activated NK-cells demonstrated functionality in inducing cancer cell death. MCP consisted of oligogalacturonic acids with some containing 4,5-unsaturated non-reducing ends. Conclusions MCP has immunostimulatory properties in human blood samples, including the activation of functional NK cells against K562 leukemic cells in culture. Unsaturated oligogalacturonic acids appear to be the immunostimulatory carbohydrates in MCP. PMID:21816083

  1. Activation of intraislet lymphoid cells causes destruction of islet cells.

    PubMed Central

    Lacy, P. E.; Finke, E. H.

    1991-01-01

    In vitro culture of rat islets at 24 degrees C for 7 days in tissue culture medium CMRL 1066 almost completely eliminated lymphoid cells from the islets. Immunostaining of the islets with monoclonal antibody OX4 for demonstration of class II major histocompatibility complex (MHC)-expressing cells revealed a decrease from 13.1 +/- 0.6 positive cells per islet on day 0 to 0.7 +/- 0.1 cells per islet on day 7. A comparable decrease was found using OX1 for demonstration of all leukocytes. In contrast, culture of rat islets at 24 degrees C for 7 days with tissue culture Roswell Park Memorial Institute (RPMI) 1640 medium was not as effective in eliminating lymphoid cells as in medium CMRL 1066 (3.0 +/- 0.2 class II MHC positive cells per islet at 7 days). Effective elimination of intraislet lymphoid cells apparently is due to the combined effect of low temperature culture and the tissue culture medium CMRL-1066. The second goal of the study was to determine whether the destructive effect of interferon gamma (IFN-gamma) on rat islets in culture was due to intraislet lymphoid cells. In vitro culture of rat islets with IFN-gamma (1000 units/ml) at 37 degrees C caused almost complete destruction of the islets at 7 days. If intraislet lymphoid cells were eliminated from the islets by in vitro culture at 24 degrees C followed by exposure to IFN-gamma (1000 units/ml) for 7 days at 37 degrees C, then IFN-gamma did not cause destruction of the islets and transplants of the treated islets produced normoglycemia in diabetic recipient mice. These findings indicate that intraislet lymphoid cells are responsible for destruction of islet cells when these cells (presumably macrophages) are activated by IFN-gamma. Intraislet lymphoid cells may play a significant role in destroying islet cells in autoimmune diabetes. Images Figure 1 Figure 2 PMID:1902627

  2. Enteric Bacterial Metabolites Propionic and Butyric Acid Modulate Gene Expression, Including CREB-Dependent Catecholaminergic Neurotransmission, in PC12 Cells - Possible Relevance to Autism Spectrum Disorders

    PubMed Central

    Nankova, Bistra B.; Agarwal, Raj; MacFabe, Derrick F.; La Gamma, Edmund F.

    2014-01-01

    Alterations in gut microbiome composition have an emerging role in health and disease including brain function and behavior. Short chain fatty acids (SCFA) like propionic (PPA), and butyric acid (BA), which are present in diet and are fermentation products of many gastrointestinal bacteria, are showing increasing importance in host health, but also may be environmental contributors in neurodevelopmental disorders including autism spectrum disorders (ASD). Further to this we have shown SCFA administration to rodents over a variety of routes (intracerebroventricular, subcutaneous, intraperitoneal) or developmental time periods can elicit behavioral, electrophysiological, neuropathological and biochemical effects consistent with findings in ASD patients. SCFA are capable of altering host gene expression, partly due to their histone deacetylase inhibitor activity. We have previously shown BA can regulate tyrosine hydroxylase (TH) mRNA levels in a PC12 cell model. Since monoamine concentration is known to be elevated in the brain and blood of ASD patients and in many ASD animal models, we hypothesized that SCFA may directly influence brain monoaminergic pathways. When PC12 cells were transiently transfected with plasmids having a luciferase reporter gene under the control of the TH promoter, PPA was found to induce reporter gene activity over a wide concentration range. CREB transcription factor(s) was necessary for the transcriptional activation of TH gene by PPA. At lower concentrations PPA also caused accumulation of TH mRNA and protein, indicative of increased cell capacity to produce catecholamines. PPA and BA induced broad alterations in gene expression including neurotransmitter systems, neuronal cell adhesion molecules, inflammation, oxidative stress, lipid metabolism and mitochondrial function, all of which have been implicated in ASD. In conclusion, our data are consistent with a molecular mechanism through which gut related environmental signals such as

  3. Protrusive Activity Guides Changes in Cell-Cell Tension during Epithelial Cell Scattering

    PubMed Central

    Maruthamuthu, Venkat; Gardel, Margaret L.

    2014-01-01

    Knowing how epithelial cells regulate cell-matrix and cell-cell adhesions is essential to understand key events in morphogenesis as well as pathological events such as metastasis. During epithelial cell scattering, epithelial cell islands rupture their cell-cell contacts and migrate away as single cells on the extracellular matrix (ECM) within hours of growth factor stimulation, even as adhesion molecules such as E-cadherin are present at the cell-cell contact. How the stability of cell-cell contacts is modulated to effect such morphological transitions is still unclear. Here, we report that in the absence of ECM, E-cadherin adhesions continue to sustain substantial cell-generated forces upon hepatocyte growth factor (HGF) stimulation, consistent with undiminished adhesion strength. In the presence of focal adhesions, constraints that preclude the spreading and movement of cells at free island edges also prevent HGF-mediated contact rupture. To explore the role of cell motion and cell-cell contact rupture, we examine the biophysical changes that occur during the scattering of cell pairs. We show that the direction of cell movement with respect to the cell-cell contact is correlated with changes in the average intercellular force as well as the initial direction of cell-cell contact rupture. Our results suggest an important role for protrusive activity resulting in cell displacement and force redistribution in guiding cell-cell contact rupture during scattering. PMID:25099795

  4. Aberrant activation of nuclear factor of activated T cell 2 in lamina propria mononuclear cells in ulcerative colitis

    PubMed Central

    Shih, Tsung-Chieh; Hsieh, Sen-Yung; Hsieh, Yi-Yueh; Chen, Tse-Chin; Yeh, Chien-Yu; Lin, Chun-Jung; Lin, Deng-Yn; Chiu, Cheng-Tang

    2008-01-01

    AIM: To investigate the role of nuclear factor of activated T cell 2 (NFAT2), the major NFAT protein in peripheral T cells, in sustained T cell activation and intractable inflammation in human ulcerative colitis (UC). METHODS: We used two-dimensional gel-electrophoresis, immunohistochemistry, double immunohistochemical staining, and confocal microscopy to inspect the expression of NFAT2 in 107, 15, 48 and 5 cases of UC, Crohn’s disease (CD), non-specific colitis, and 5 healthy individuals, respectively. RESULTS: Up-regulation with profound nucleo-translocation/activation of NFAT2 of lamina propria mononuclear cells (LPMC) of colonic mucosa was found specifically in the affected colonic mucosa from patients with UC, as compared to CD or NC (P < 0.001, Kruskal-Wallis test). Nucleo-translocation/activation of NFAT2 primarily occurred in CD8+T, but was less prominent in CD4+ T cells or CD20+B cells. It was strongly associated with the disease activity, including endoscopic stage (τ = 0.2145, P = 0.0281) and histologic grade (τ = 0.4167, P < 0.001). CONCLUSION: We disclose for the first time the nucleo-translocation/activatin of NFAT2 in lamina propria mononuclear cells in ulcerative colitis. Activation of NFAT2 was specific for ulcerative colitis and highly associated with disease activity. Since activation of NFAT2 is implicated in an auto-regulatory positive feedback loop of sustained T-cell activation and NFAT proteins play key roles in the calcium/calcineurin signaling pathways, our results not only provide new insights into the mechanism for sustained intractable inflammation, but also suggest the calcium-calcineurin/NFAT pathway as a new therapeutic target for ulcerative colitis. PMID:18350607

  5. Human Mesenchymal Stem Cells Retain Multilineage Differentiation Capacity Including Neural Marker Expression after Extended In Vitro Expansion

    PubMed Central

    Okolicsanyi, Rachel K.; Camilleri, Emily T.; Oikari, Lotta E; Yu, Chieh; Cool, Simon M.; van Wijnen, Andre J.; Griffiths, Lyn R.; Haupt, Larisa M.

    2015-01-01

    The suitability of human mesenchymal stem cells (hMSCs) in regenerative medicine relies on retention of their proliferative expansion potential in conjunction with the ability to differentiate toward multiple lineages. Successful utilisation of these cells in clinical applications linked to tissue regeneration requires consideration of biomarker expression, time in culture and donor age, as well as their ability to differentiate towards mesenchymal (bone, cartilage, fat) or non-mesenchymal (e.g., neural) lineages. To identify potential therapeutic suitability we examined hMSCs after extended expansion including morphological changes, potency (stemness) and multilineage potential. Commercially available hMSC populations were expanded in vitro for > 20 passages, equating to > 60 days and > 50 population doublings. Distinct growth phases (A-C) were observed during serial passaging and cells were characterised for stemness and lineage markers at representative stages (Phase A: P+5, approximately 13 days in culture; Phase B: P+7, approximately 20 days in culture; and Phase C: P+13, approximately 43 days in culture). Cell surface markers, stem cell markers and lineage-specific markers were characterised by FACS, ICC and Q-PCR revealing MSCs maintained their multilineage potential, including neural lineages throughout expansion. Co-expression of multiple lineage markers along with continued CD45 expression in MSCs did not affect completion of osteogenic and adipogenic specification or the formation of neurospheres. Improved standardised isolation and characterisation of MSCs may facilitate the identification of biomarkers to improve therapeutic efficacy to ensure increased reproducibility and routine production of MSCs for therapeutic applications including neural repair. PMID:26356539

  6. Activation of radiosensitizers by hypoxic cells.

    PubMed Central

    Olive, P. L.; Durand, R. E.

    1978-01-01

    Hypoxic cells can metabolize nitroheterocyclic compounds to produce toxic intermediates capable of affecting the survival of neighbouring oxygenated cells. Mutagenesis experiments with E. coli WP-2 343 (deficient in nitroreductase) indicated that reduction of nitroheterocyclics outside bacteria causes killing and mutations within bacteria, presumably due to the transfer of the "active" specie (s). Using animal tissue slices to reduce nitrofurans, cultured L-929 cells incubated under aerobic conditions were far more sensitive to the toxic and DNA damaging effects of these drugs. Transfer of the active species also occurs in a tissue-like environment in multicell spheroids where the presence of a hypoxic central core served to convert the nitroheterocyclics to intermediates which also damaged the neighbouring oxygenated cells. PMID:354676

  7. Polyclonal B cell activation in ankylosing spondylitis.

    PubMed Central

    Barbieri, P; Olivieri, I; Benedettini, G; Marelli, P; Ciompi, M L; Pasero, G; Campa, M

    1990-01-01

    The peripheral blood lymphocyte response of patients with ankylosing spondylitis (AS) to several polyclonal B cell activators was investigated. No differences were found in the reactivity to pokeweed mitogen and protein A between patients and controls; in contrast, the peripheral blood lymphocyte response to Staphylococcus aureus strain Cowan I (SAC) was significantly higher in patients with AS than in controls. This responsiveness was not influenced either by the presence of the HLA-B27 antigen or by environmental factors or associated diseases, and it was higher in patients with active AS than in those with inactive disease. The percentage of circulating B cells was normal. The responses to T cell mitogens and the percentages of T cell subpopulations were similar in patients and in controls. The peripheral blood lymphocyte hyperactivity of patients with AS to SAC was associated with an increased in vitro production of immunoglobulins. PMID:2383063

  8. (+)-Catechin attenuates activation of hepatic stellate cells.

    PubMed

    Bragança de Moraes, Cristina Machado; Bitencourt, Shanna; de Mesquita, Fernanda Cristina; Mello, Denizar; de Oliveira, Leticia Paranhos; da Silva, Gabriela Viegas; Lorini, Vinicius; Caberlon, Eduardo; de Souza Basso, Bruno; Schmid, Julia; Ferreira, Gabriela Acevedo; de Oliveira, Jarbas Rodrigues

    2014-04-01

    (+)-Catechin is a type of catechin present in large amounts in açaí fruits and cocoa seeds. Besides its antioxidant and anti-inflammatory activities, little is known about its effects in the liver, especially during hepatic fibrosis. We report here the effects of (+)-catechin on hepatic stellate cells. (+)-Catechin induced quiescent phenotype in GRX cells, along with an increase in lipid droplets. Proliferator-activated receptor γ mRNA expression was upregulated, whereas type I collagen mRNA expression was downregulated. Pro-inflammatory cytokines were not influenced by (+)-catechin, whereas the levels of interleukin 10 were significantly increased. The data provide evidence that (+)-catechin can reduce hepatic stellate cell activation. PMID:24353036

  9. Studies of T-cell activation in chronic inflammation

    PubMed Central

    2002-01-01

    Chapter summary The strong association between specific alleles encoded within the MHC class II region and the development of rheumatoid arthritis (RA) has provided the best evidence to date that CD4+ T cells play a role in the pathogenesis of this chronic inflammatory disease. However, the unusual phenotype of synovial T cells, including their profound proliferative hyporesponsiveness to TCR ligation, has challenged the notion that T-cell effector responses are driven by cognate cartilage antigens in inflamed synovial joints. The hierarchy of T-cell dysfunction from peripheral blood to inflamed joint suggests that these defects are acquired through prolonged exposure to proinflammatory cytokines such as tumour necrosis factor (TNF)-α. Indeed, there are now compelling data to suggest that chronic cytokine activation may contribute substantially to the phenotype and effector function of synovial T cells. Studies reveal that chronic exposure of T cells to TNF uncouples TCR signal transduction pathways by impairing the assembly and stability of the TCR/CD3 complex at the cell surface. Despite this membrane-proximal effect, TNF selectively uncouples downstream signalling pathways, as is shown by the dramatic suppression of calcium signalling responses, while Ras/ERK activation is spared. On the basis of these data, it is proposed that T-cell survival and effector responses are driven by antigen-independent, cytokine-dependent mechanisms, and that therapeutic strategies that seek to restore T-cell homeostasis rather than further depress T-cell function should be explored in the future. PMID:12110140

  10. The Difference of Lymphocyte Subsets Including Regulatory T-Cells in Umbilical Cord Blood between AGA Neonates and SGA Neonates

    PubMed Central

    Yoon, Sang Hee; Hur, Mina; Hwang, Han Sung; Kwon, Han Sung

    2015-01-01

    Purpose This study aimed to compare the regulatory T cells in cord blood of appropriate for gestational age (AGA) neonates with those of small for gestational age (SGA) neonates. Materials and Methods Umbilical cord blood was collected upon labor in 108 healthy full-term (between 37 and 41 gestational weeks) neonates, who were born between November 2010 and April 2012. Among them, 77 samples were obtained from AGA neonates, and 31 samples were obtained from SGA neonates. Regulatory T cells and lymphocyte subsets were determined using a flow cytometer. Student's t-test for independent samples was used to compare differences between AGA and SGA neonates. Results Regulatory T cells in cord blood were increased in the SGA group compared with normal controls (p=0.041). However, cytotoxic T cells in cord blood were significantly decreased in the SGA group compared with normal controls (p=0.007). Conclusion This is the first study to compare the distribution of lymphocyte subsets including regulatory T cells in cord blood between AGA neonates and SGA neonates. PMID:25837188

  11. Oncogenic functions of IGF1R and INSR in prostate cancer include enhanced tumor growth, cell migration and angiogenesis.

    PubMed

    Heidegger, Isabel; Kern, Johann; Ofer, Philipp; Klocker, Helmut; Massoner, Petra

    2014-05-15

    We scrutinized the effect of insulin receptor (INSR) in addition to IGF1R in PCa using in vitro and in vivo models. In-vitro overexpression of IGF1R and INSRA, but not INSRB increased cell proliferation, colony formation, migration, invasion and resistance to apoptosis in prostate cancer cells (DU145, LNCaP, PC3). Opposite effects were induced by downregulation of IGF1R and total INSR, but not INSRB. In contrast to tumor cells, non-cancerous epithelial cells of the prostate (EP156T, RWPE-1) were inhibited on overexpression and stimulated by knockdown of receptors. In-vivo analyses using the chicken allantoic membrane assay confirmed the tumorigenic effects of IGF1R and INSR. Apart of promoting tumor growth, IGF1R and INSR overexpression also enhanced angiogenesis indicated by higher vessel density and increased number of desmin-immunoreactive pericytes. Our study underscores the oncogenic impact of IGF1R including significant effects on tumor growth, cell migration, sensitivity to apoptotic/chemotherapeutic agents and angiogenesis, and characterizes the INSR, in particular the isoform INSRA, as additional cancer-promoting receptor in prostate cancer. Both, the insulin-like growth factor receptor 1 and the insulin receptor exert oncogenic functions, thus proposing that both receptors need to be considered in therapeutic settings. PMID:24809298

  12. Oncogenic functions of IGF1R and INSR in prostate cancer include enhanced tumor growth, cell migration and angiogenesis

    PubMed Central

    Heidegger, Isabel; Kern, Johann; Ofer, Philipp

    2014-01-01

    We scrutinized the effect of insulin receptor (INSR) in addition to IGF1R in PCa using in vitro and in vivo models. In-vitro overexpression of IGF1R and INSRA, but not INSRB increased cell proliferation, colony formation, migration, invasion and resistance to apoptosis in prostate cancer cells (DU145, LNCaP, PC3). Opposite effects were induced by downregulation of IGF1R and total INSR, but not INSRB. In contrast to tumor cells, non-cancerous epithelial cells of the prostate (EP156T, RWPE-1) were inhibited on overexpression and stimulated by knockdown of receptors. In-vivo analyses using the chicken allantoic membrane assay confirmed the tumorigenic effects of IGF1R and INSR. Apart from promoting tumor growth, IGF1R and INSR overexpression also enhanced angiogenesis indicated by higher vessel density and increased number of desmin-immunoreactive pericytes. Our study underscores the oncogenic impact of IGF1R including significant effects on tumor growth, cell migration, sensitivity to apoptotic/chemotherapeutic agents and angiogenesis, and characterizes the INSR, in particular the isoform INSRA, as additional cancer-promoting receptor in prostate cancer. Both, the insulin-like growth factor receptor 1 and the insulin receptor exert oncogenic functions, thus proposing that both receptors need to be considered in therapeutic settings. PMID:24809298

  13. Evaluating cell-surface expression and measuring activation of mammalian odorant receptors in heterologous cells

    PubMed Central

    Zhuang, Hanyi; Matsunami, Hiroaki

    2009-01-01

    A fundamental question in olfaction is which odorant receptors (ORs) are activated by a given odorant. A major roadblock to investigate odorant-OR relationship in mammals has been an inability to express ORs in heterologous cells suitable for screening active ligands for ORs. The discovery of the receptor-transporting protein (RTP) family has facilitated the effective cell-surface expression of ORs in heterologous cells. The establishment of a robust heterologous expression system for mammalian ORs facilitates the high-throughput “deorphanization” of these receptors by matching them to their cognate ligands. This protocol details the method used for evaluating the cell-surface expression and measuring the functional activation of ORs of transiently-expressed mammalian odorant receptors in HEK293T cells. The stages of odorant receptor cell-surface expression include cell culture preparation, transfer of cells, transfection, and immunocytochemistry/flow cytometry, odorant stimulation, and luciferase assay. This protocol can be completed in a period of 3 days from transfer of cells to cell-surface expression detection and/or measurement of functional activation. PMID:18772867

  14. Cell Cycle-Regulated Protein Abundance Changes in Synchronously Proliferating HeLa Cells Include Regulation of Pre-mRNA Splicing Proteins

    PubMed Central

    Lane, Karen R.; Yu, Yanbao; Lackey, Patrick E.; Chen, Xian; Marzluff, William F.; Cook, Jeanette Gowen

    2013-01-01

    Cell proliferation involves dramatic changes in DNA metabolism and cell division, and control of DNA replication, mitosis, and cytokinesis have received the greatest attention in the cell cycle field. To catalogue a wider range of cell cycle-regulated processes, we employed quantitative proteomics of synchronized HeLa cells. We quantified changes in protein abundance as cells actively progress from G1 to S phase and from S to G2 phase. We also describe a cohort of proteins whose abundance changes in response to pharmacological inhibition of the proteasome. Our analysis reveals not only the expected changes in proteins required for DNA replication and mitosis but also cell cycle-associated changes in proteins required for biological processes not known to be cell-cycle regulated. For example, many pre-mRNA alternative splicing proteins are down-regulated in S phase. Comparison of this dataset to several other proteomic datasets sheds light on global mechanisms of cell cycle phase transitions and underscores the importance of both phosphorylation and ubiquitination in cell cycle changes. PMID:23520512

  15. Entangled active matter: From cells to ants

    NASA Astrophysics Data System (ADS)

    Hu, D. L.; Phonekeo, S.; Altshuler, E.; Brochard-Wyart, F.

    2016-07-01

    Both cells and ants belong to the broad field of active matter, a novel class of non-equilibrium materials composed of many interacting units that individually consume energy and collectively generate motion or mechanical stresses. However cells and ants differ from fish and birds in that they can support static loads. This is because cells and ants can be entangled, so that individual units are bound by transient links. Entanglement gives cells and ants a set of remarkable properties usually not found together, such as the ability to flow like a fluid, spring back like an elastic solid, and self-heal. In this review, we present the biology, mechanics and dynamics of both entangled cells and ants. We apply concepts from soft matter physics and wetting to characterize these systems as well as to point out their differences, which arise from their differences in size. We hope that our viewpoints will spur further investigations into cells and ants as active materials, and inspire the fabrication of synthetic active matter.

  16. Modulation of Dendritic Cell Activation and Subsequent Th1 Cell Polarization by Lidocaine.

    PubMed

    Jeon, Young-Tae; Na, Hyeongjin; Ryu, Heeju; Chung, Yeonseok

    2015-01-01

    Dendritic cells play an essential role in bridging innate and adaptive immunity by recognizing cellular stress including pathogen- and damage-associated molecular patterns and by shaping the types of antigen-specific T cell immunity. Although lidocaine is widely used in clinical settings that trigger cellular stress, it remains unclear whether such treatment impacts the activation of innate immune cells and subsequent differentiation of T cells. Here we showed that lidocaine inhibited the production of IL-6, TNFα and IL-12 from dendritic cells in response to toll-like receptor ligands including lipopolysaccharide, poly(I:C) and R837 in a dose-dependent manner. Notably, the differentiation of Th1 cells was significantly suppressed by the addition of lidocaine while the same treatment had little effect on the differentiation of Th17, Th2 and regulatory T cells in vitro. Moreover, lidocaine suppressed the ovalbumin-specific Th1 cell responses in vivo induced by the adoptive transfer of ovalbumin-pulsed dendritic cells. These results demonstrate that lidocaine inhibits the activation of dendritic cells in response to toll-like receptor signals and subsequently suppresses the differentiation of Th1 cell responses. PMID:26445366

  17. Imaging CREB Activation in Living Cells*

    PubMed Central

    Friedrich, Michael W.; Aramuni, Gayane; Mank, Marco; Mackinnon, Jonathan A. G.; Griesbeck, Oliver

    2010-01-01

    The Ca2+- and cAMP-responsive element-binding protein (CREB) and the related ATF-1 and CREM are stimulus-inducible transcription factors that link certain forms of cellular activity to changes in gene expression. They are attributed to complex integrative activation characteristics, but current biochemical technology does not allow dynamic imaging of CREB activation in single cells. Using fluorescence resonance energy transfer between mutants of green fluorescent protein we here develop a signal-optimized genetically encoded indicator that enables imaging activation of CREB due to phosphorylation of the critical serine 133. The indicator of CREB activation due to phosphorylation (ICAP) was used to investigate the role of the scaffold and anchoring protein AKAP79/150 in regulating signal pathways converging on CREB. We show that disruption of AKAP79/150-mediated protein kinase A anchoring or knock-down of AKAP150 dramatically reduces the ability of protein kinase A to activate CREB. In contrast, AKAP79/150 regulation of CREB via L-type channels may only have minor importance. ICAP allows dynamic and reversible imaging in living cells and may become useful in studying molecular components and cell-type specificity of activity-dependent gene expression. PMID:20484048

  18. Critical telomerase activity for uncontrolled cell growth.

    PubMed

    Wesch, Neil L; Burlock, Laura J; Gooding, Robert J

    2016-01-01

    The lengths of the telomere regions of chromosomes in a population of cells are modelled using a chemical master equation formalism, from which the evolution of the average number of cells of each telomere length is extracted. In particular, the role of the telomere-elongating enzyme telomerase on these dynamics is investigated. We show that for biologically relevant rates of cell birth and death, one finds a critical rate, R crit, of telomerase activity such that the total number of cells diverges. Further, R crit is similar in magnitude to the rates of mitosis and cell death. The possible relationship of this result to replicative immortality and its associated hallmark of cancer is discussed. PMID:27500377

  19. Critical telomerase activity for uncontrolled cell growth

    NASA Astrophysics Data System (ADS)

    Wesch, Neil L.; Burlock, Laura J.; Gooding, Robert J.

    2016-08-01

    The lengths of the telomere regions of chromosomes in a population of cells are modelled using a chemical master equation formalism, from which the evolution of the average number of cells of each telomere length is extracted. In particular, the role of the telomere-elongating enzyme telomerase on these dynamics is investigated. We show that for biologically relevant rates of cell birth and death, one finds a critical rate, R crit, of telomerase activity such that the total number of cells diverges. Further, R crit is similar in magnitude to the rates of mitosis and cell death. The possible relationship of this result to replicative immortality and its associated hallmark of cancer is discussed.

  20. Steering Organoids Toward Discovery: Self-Driving Stem Cells Are Opening a World of Possibilities, Including Drug Testing and Tissue Sourcing.

    PubMed

    Solis, Michele

    2016-01-01

    Since the 1980s, stem cells' shape-shifting abilities have wowed scientists. With proper handling, a few growth factors, and some time, stem cells can be cooked up into specific cell types, including neurons, muscle, and skin. PMID:27414630

  1. Buoyancy-activated cell sorting using targeted biotinylated albumin microbubbles.

    PubMed

    Liou, Yu-Ren; Wang, Yu-Hsin; Lee, Chia-Ying; Li, Pai-Chi

    2015-01-01

    Cell analysis often requires the isolation of certain cell types. Various isolation methods have been applied to cell sorting, including fluorescence-activated cell sorting and magnetic-activated cell sorting. However, these conventional approaches involve exerting mechanical forces on the cells, thus risking cell damage. In this study we applied a novel isolation method called buoyancy-activated cell sorting, which involves using biotinylated albumin microbubbles (biotin-MBs) conjugated with antibodies (i.e., targeted biotin-MBs). Albumin MBs are widely used as contrast agents in ultrasound imaging due to their good biocompatibility and stability. For conjugating antibodies, biotin is conjugated onto the albumin MB shell via covalent bonds and the biotinylated antibodies are conjugated using an avidin-biotin system. The albumin microbubbles had a mean diameter of 2 μm with a polydispersity index of 0.16. For cell separation, the MDA-MB-231 cells are incubated with the targeted biotin-MBs conjugated with anti-CD44 for 10 min, centrifuged at 10 g for 1 min, and then allowed 1 hour at 4 °C for separation. The results indicate that targeted biotin-MBs conjugated with anti-CD44 antibodies can be used to separate MDA-MB-231 breast cancer cells; more than 90% of the cells were collected in the MB layer when the ratio of the MBs to cells was higher than 70:1. Furthermore, we found that the separating efficiency was higher for targeted biotin-MBs than for targeted avidin-incorporated albumin MBs (avidin-MBs), which is the most common way to make targeted albumin MBs. We also demonstrated that the recovery rate of targeted biotin-MBs was up to 88% and the sorting purity was higher than 84% for a a heterogenous cell population containing MDA-MB-231 cells (CD44(+)) and MDA-MB-453 cells (CD44-), which are classified as basal-like breast cancer cells and luminal breast cancer cells, respectively. Knowing that the CD44(+) is a commonly used cancer-stem-cell biomarker, our

  2. Buoyancy-Activated Cell Sorting Using Targeted Biotinylated Albumin Microbubbles

    PubMed Central

    Liou, Yu-Ren; Wang, Yu-Hsin; Lee, Chia-Ying; Li, Pai-Chi

    2015-01-01

    Cell analysis often requires the isolation of certain cell types. Various isolation methods have been applied to cell sorting, including florescence-activated cell sorting and magnetic-activated cell sorting. However, these conventional approaches involve exerting mechanical forces on the cells, thus risking cell damage. In this study we applied a novel isolation method called buoyancy-activated cell sorting, which involves using biotinylated albumin microbubbles (biotin-MBs) conjugated with antibodies (i.e., targeted biotin-MBs). Albumin MBs are widely used as contrast agents in ultrasound imaging due to their good biocompatibility and stability. For conjugating antibodies, biotin is conjugated onto the albumin MB shell via covalent bonds and the biotinylated antibodies are conjugated using an avidin-biotin system. The albumin microbubbles had a mean diameter of 2μm with a polydispersity index of 0.16. For cell separation, the MDA-MB-231 cells are incubated with the targeted biotin-MBs conjugated with anti-CD44 for 10 min, centrifuged at 10g for 1 min, and then allowed 1 hour at 4°C for separation. The results indicate that targeted biotin-MBs conjugated with anti-CD44 antibodies can be used to separate MDA-MB-231 breast cancer cells; more than 90% of the cells were collected in the MB layer when the ratio of the MBs to cells was higher than 70:1. Furthermore, we found that the separating efficiency was higher for targeted biotin-MBs than for targeted avidin-incorporated albumin MBs (avidin-MBs), which is the most common way to make targeted albumin MBs. We also demonstrated that the recovery rate of targeted biotin-MBs was up to 88% and the sorting purity was higher than 84% for a a heterogenous cell population containing MDA-MB-231 cells (CD44+) and MDA-MB-453 cells (CD44–), which are classified as basal-like breast cancer cells and luminal breast cancer cells, respectively. Knowing that the CD44+ is a commonly used cancer-stem-cell biomarker, our targeted

  3. Active mechanics and geometry of adherent cells and cell colonies

    NASA Astrophysics Data System (ADS)

    Banerjee, Shiladitya

    2014-03-01

    Measurements of traction stresses exerted by adherent cells or cell colonies on elastic substrates have yielded new insight on how the mechanical and geometrical properties of the substrate regulate cellular force distribution, mechanical energy, spreading, morphology or stress ber architecture. We have developed a generic mechanical model of adherent cells as an active contractile gel mechanically coupled to an elastic substrate and to neighboring cells in a tissue. The contractile gel model accurately predicts the distribution of cellular and traction stresses as observed in single cell experiments, and captures the dependence of cell shape, traction stresses and stress ber polarization on the substrate's mechanical and geometrical properties. The model further predicts that the total strain energy of an adherent cell is solely regulated by its spread area, in agreement with recent experiments on micropatterned substrates with controlled geometry. When used to describe the behavior of colonies of adherent epithelial cells, the model demonstrates the crucial role of the mechanical cross-talk between intercellular and extracellular adhesion in regulating traction force distribution. Strong intercellular mechanical coupling organizes traction forces to the colony periphery, whereas weaker intercellular coupling leads to the build up of traction stresses at intercellular junctions. Furthermore, in agreement with experiments on large cohesive keratinocyte colonies, the model predicts a linear scaling of traction forces with the colony size. This scaling suggests the emergence of an effective surface tension as a scale-free material property of the adherent tissue, originating from actomyosin contractility.

  4. Immunohistochemical distinction of metastases of renal cell carcinoma to the adrenal from primary adrenal nodules, including oncocytic tumor.

    PubMed

    Li, Hongmei; Hes, Ondrej; MacLennan, Gregory T; Eastwood, Daniel C; Iczkowski, Kenneth A

    2015-05-01

    Metastases of clear cell renal cell carcinoma to the adrenal can mimic primary adrenal cortical neoplasms or normal adrenal, especially in biopsy material. We compared 34 cases of clear cell renal cell carcinoma metastasis to the adrenal with 49 primary adrenal lesions (16 carcinoma, 22 adenoma, 9 oncocytic tumor, and 2 hyperplasia). Normal adrenal was available in 59 cases. Each entity was represented on tissue microarrays by duplicate-triplicate evaluable spots taken from spatially separate areas. Two pathologists evaluated all reactivity from 0 to 3+. A panel of 12 immunohistochemical stains was performed, including the first diagnostic uses of steroid receptor coactivator (SRC1) and equilibrative nucleoside transporter 1 (ENT1). The most sensitive and specific renal cell carcinoma markers were membranous reactivity for carbonic anhydrase IX (CAIX) and RCC marker and nuclear reactivity for PAX8. For adrenal cortical carcinomas, best markers were synaptophysin, SRC1, and MelanA; and for adrenal oncocytic tumor, synaptophysin and ENT1. Optimal markers for adrenal cortical adenoma and normal adrenal were ENT1 (more specific) and either MelanA or SRC1 (more sensitive). Calretinin, cytokeratin 34βE12 and CAM5.2, inhibin, and steroidogenic factor 1 (SF1) proved less valuable to the panel. Nonspecific cytoplasmic biotin reactivity was frequent for CAIX and PAX8. Tumors with high-grade cytology should be worked up with 2 of the 3 stains: CAIX, PAX8, or RCC marker; and either SRC1 or MelanA. Adrenal adenoma, or normal adrenal, versus low-grade renal cell carcinoma are distinguished by a panel of: CAIX, PAX8, or RCC Marker; ENT1 and either SRC1 or MelanA. PMID:25690138

  5. Hydrogen peroxide stimulates cell motile activity through LPA receptor-3 in liver epithelial WB-F344 cells

    SciTech Connect

    Shibata, Ayano; Tanabe, Eriko; Inoue, Serina; Kitayoshi, Misaho; Okimoto, Souta; Hirane, Miku; Araki, Mutsumi; Fukushima, Nobuyuki; Tsujiuchi, Toshifumi

    2013-04-12

    Highlights: •Hydrogen peroxide stimulates cell motility of WB-F344 cells. •LPA{sub 3} is induced by hydrogen peroxide in WB-F344 cells. •Cell motility by hydrogen peroxide is inhibited in LPA{sub 3} knockdown cells. •LPA signaling is involved in cell migration by hydrogen peroxide. -- Abstract: Hydrogen peroxide which is one of reactive oxygen species (ROS) mediates a variety of biological responses, including cell proliferation and migration. In the present study, we investigated whether lysophosphatidic acid (LPA) signaling is involved in cell motile activity stimulated by hydrogen peroxide. The rat liver epithelial WB-F344 cells were treated with hydrogen peroxide at 0.1 or 1 μM for 48 h. In cell motility assays, hydrogen peroxide treated cells showed significantly high cell motile activity, compared with untreated cells. To measure the expression levels of LPA receptor genes, quantitative real time RT-PCR analysis was performed. The expressions of LPA receptor-3 (Lpar3) in hydrogen peroxide treated cells were significantly higher than those in control cells, but not Lpar1 and Lpar2 genes. Next, to assess the effect of LPA{sub 3} on cell motile activity, the Lpar3 knockdown cells from WB-F344 cells were also treated with hydrogen peroxide. The cell motile activity of the knockdown cells was not stimulated by hydrogen peroxide. Moreover, in liver cancer cells, hydrogen peroxide significantly activated cell motility of Lpar3-expressing cells, but not Lpar3-unexpressing cells. These results suggest that LPA signaling via LPA{sub 3} may be mainly involved in cell motile activity of WB-F344 cells stimulated by hydrogen peroxide.

  6. NKG2D is a Key Receptor for Recognition of Bladder Cancer Cells by IL-2-Activated NK Cells and BCG Promotes NK Cell Activation

    PubMed Central

    García-Cuesta, Eva María; López-Cobo, Sheila; Álvarez-Maestro, Mario; Esteso, Gloria; Romera-Cárdenas, Gema; Rey, Mercedes; Cassady-Cain, Robin L.; Linares, Ana; Valés-Gómez, Alejandro; Reyburn, Hugh Thomson; Martínez-Piñeiro, Luis; Valés-Gómez, Mar

    2015-01-01

    Intravesical instillation of bacillus Calmette–Guérin (BCG) is used to treat superficial bladder cancer, either papillary tumors (after transurethral resection) or high-grade flat carcinomas (carcinoma in situ), reducing recurrence in about 70% of patients. Initially, BCG was proposed to work through an inflammatory response, mediated by phagocytic uptake of mycobacterial antigens and cytokine release. More recently, other immune effectors such as monocytes, natural killer (NK), and NKT cells have been suggested to play a role in this immune response. Here, we provide a comprehensive study of multiple bladder cancer cell lines as putative targets for immune cells and evaluated their recognition by NK cells in the presence and absence of BCG. We describe that different bladder cancer cells can express multiple activating and inhibitory ligands for NK cells. Recognition of bladder cancer cells depended mainly on NKG2D, with a contribution from NKp46. Surprisingly, exposure to BCG did not affect the immune phenotype of bladder cells nor increased NK cell recognition of purified IL-2-activated cell lines. However, NK cells were activated efficiently when BCG was included in mixed lymphocyte cultures, suggesting that NK activation after mycobacteria treatment requires the collaboration of various immune cells. We also analyzed the percentage of NK cells in peripheral blood of a cohort of bladder cancer patients treated with BCG. The total numbers of NK cells did not vary during treatment, indicating that a more detailed study of NK cell activation in the tumor site will be required to evaluate the response in each patient. PMID:26106390

  7. Mechanically activated artificial cell by using microfluidics.

    PubMed

    Ho, Kenneth K Y; Lee, Lap Man; Liu, Allen P

    2016-01-01

    All living organisms sense mechanical forces. Engineering mechanosensitive artificial cell through bottom-up in vitro reconstitution offers a way to understand how mixtures of macromolecules assemble and organize into a complex system that responds to forces. We use stable double emulsion droplets (aqueous/oil/aqueous) to prototype mechanosensitive artificial cells. In order to demonstrate mechanosensation in artificial cells, we develop a novel microfluidic device that is capable of trapping double emulsions into designated chambers, followed by compression and aspiration in a parallel manner. The microfluidic device is fabricated using multilayer soft lithography technology, and consists of a control layer and a deformable flow channel. Deflections of the PDMS membrane above the main microfluidic flow channels and trapping chamber array are independently regulated pneumatically by two sets of integrated microfluidic valves. We successfully compress and aspirate the double emulsions, which result in transient increase and permanent decrease in oil thickness, respectively. Finally, we demonstrate the influx of calcium ions as a response of our mechanically activated artificial cell through thinning of oil. The development of a microfluidic device to mechanically activate artificial cells creates new opportunities in force-activated synthetic biology. PMID:27610921

  8. Mechanically activated artificial cell by using microfluidics

    PubMed Central

    Ho, Kenneth K. Y.; Lee, Lap Man; Liu, Allen P.

    2016-01-01

    All living organisms sense mechanical forces. Engineering mechanosensitive artificial cell through bottom-up in vitro reconstitution offers a way to understand how mixtures of macromolecules assemble and organize into a complex system that responds to forces. We use stable double emulsion droplets (aqueous/oil/aqueous) to prototype mechanosensitive artificial cells. In order to demonstrate mechanosensation in artificial cells, we develop a novel microfluidic device that is capable of trapping double emulsions into designated chambers, followed by compression and aspiration in a parallel manner. The microfluidic device is fabricated using multilayer soft lithography technology, and consists of a control layer and a deformable flow channel. Deflections of the PDMS membrane above the main microfluidic flow channels and trapping chamber array are independently regulated pneumatically by two sets of integrated microfluidic valves. We successfully compress and aspirate the double emulsions, which result in transient increase and permanent decrease in oil thickness, respectively. Finally, we demonstrate the influx of calcium ions as a response of our mechanically activated artificial cell through thinning of oil. The development of a microfluidic device to mechanically activate artificial cells creates new opportunities in force-activated synthetic biology. PMID:27610921

  9. Evaluation and optimization of a micro-tubular solid oxide fuel cell stack model including an integrated cooling system

    NASA Astrophysics Data System (ADS)

    Hering, Martin; Brouwer, Jacob; Winkler, Wolfgang

    2016-01-01

    A micro-tubular solid oxide fuel cell stack model including an integrated cooling system was developed using a quasi three-dimensional, spatially resolved, transient thermodynamic, physical and electrochemical model that accounts for the complex geometrical relations between the cells and cooling-tubes. For the purpose of model evaluation, reference operating, geometrical and material properties are determined. The reference stack design is composed of 3294 cells, with a diameter of 2 mm, and 61 cooling-tubes. The stack is operated at a power density of 300 mW/cm2 and air is used as the cooling fluid inside the integrated cooling system. Regarding the performance, the reference design achieves an electrical stack efficiency of around 57% and a power output of 1.1 kW. The maximum occurring temperature of the positive electrode electrolyte negative electrode (PEN)-structure is 1369 K. As a result of a design of experiments, parameters of a best-case design are determined. The best-case design achieves a comparable power output of 1.1 kW with an electrical efficiency of 63% and a maximum occurring temperature of the PEN-structure of 1268 K. Nevertheless, the best-case design has an increased volume based on the higher diameter of 3 mm and increased spacing between the cells.

  10. Direct determination of phosphatase activity from physiological substrates in cells.

    PubMed

    Ren, Zhongyuan; Do, Le Duy; Bechkoff, Géraldine; Mebarek, Saida; Keloglu, Nermin; Ahamada, Saandia; Meena, Saurabh; Magne, David; Pikula, Slawomir; Wu, Yuqing; Buchet, René

    2015-01-01

    A direct and continuous approach to determine simultaneously protein and phosphate concentrations in cells and kinetics of phosphate release from physiological substrates by cells without any labeling has been developed. Among the enzymes having a phosphatase activity, tissue non-specific alkaline phosphatase (TNAP) performs indispensable, multiple functions in humans. It is expressed in numerous tissues with high levels detected in bones, liver and neurons. It is absolutely required for bone mineralization and also necessary for neurotransmitter synthesis. We provided the proof of concept that infrared spectroscopy is a reliable assay to determine a phosphatase activity in the osteoblasts. For the first time, an overall specific phosphatase activity in cells was determined in a single step by measuring simultaneously protein and substrate concentrations. We found specific activities in osteoblast like cells amounting to 116 ± 13 nmol min(-1) mg(-1) for PPi, to 56 ± 11 nmol min(-1) mg(-1) for AMP, to 79 ± 23 nmol min(-1) mg(-1) for beta-glycerophosphate and to 73 ± 15 nmol min(-1) mg(-1) for 1-alpha-D glucose phosphate. The assay was also effective to monitor phosphatase activity in primary osteoblasts and in matrix vesicles. The use of levamisole--a TNAP inhibitor--served to demonstrate that a part of the phosphatase activity originated from this enzyme. An IC50 value of 1.16 ± 0.03 mM was obtained for the inhibition of phosphatase activity of levamisole in osteoblast like cells. The infrared assay could be extended to determine any type of phosphatase activity in other cells. It may serve as a metabolomic tool to monitor an overall phosphatase activity including acid phosphatases or other related enzymes. PMID:25785438

  11. Use of an Activated Beta-Catenin to Identify Wnt Pathway Target Genes in Caenorhabditis elegans, Including a Subset of Collagen Genes Expressed in Late Larval Development

    PubMed Central

    Jackson, Belinda M.; Abete-Luzi, Patricia; Krause, Michael W.; Eisenmann, David M.

    2014-01-01

    The Wnt signaling pathway plays a fundamental role during metazoan development, where it regulates diverse processes, including cell fate specification, cell migration, and stem cell renewal. Activation of the beta-catenin−dependent/canonical Wnt pathway up-regulates expression of Wnt target genes to mediate a cellular response. In the nematode Caenorhabditis elegans, a canonical Wnt signaling pathway regulates several processes during larval development; however, few target genes of this pathway have been identified. To address this deficit, we used a novel approach of conditionally activated Wnt signaling during a defined stage of larval life by overexpressing an activated beta-catenin protein, then used microarray analysis to identify genes showing altered expression compared with control animals. We identified 166 differentially expressed genes, of which 104 were up-regulated. A subset of the up-regulated genes was shown to have altered expression in mutants with decreased or increased Wnt signaling; we consider these genes to be bona fide C. elegans Wnt pathway targets. Among these was a group of six genes, including the cuticular collagen genes, bli-1col-38, col-49, and col-71. These genes show a peak of expression in the mid L4 stage during normal development, suggesting a role in adult cuticle formation. Consistent with this finding, reduction of function for several of the genes causes phenotypes suggestive of defects in cuticle function or integrity. Therefore, this work has identified a large number of putative Wnt pathway target genes during larval life, including a small subset of Wnt-regulated collagen genes that may function in synthesis of the adult cuticle. PMID:24569038

  12. The IKAROS Interaction with a Complex Including Chromatin Remodeling and Transcription Elongation Activities Is Required for Hematopoiesis

    PubMed Central

    Bottardi, Stefania; Mavoungou, Lionel; Pak, Helen; Daou, Salima; Bourgoin, Vincent; Lakehal, Yahia A.; Affar, El Bachir; Milot, Eric

    2014-01-01

    IKAROS is a critical regulator of hematopoietic cell fate and its dynamic expression pattern is required for proper hematopoiesis. In collaboration with the Nucleosome Remodeling and Deacetylase (NuRD) complex, it promotes gene repression and activation. It remains to be clarified how IKAROS can support transcription activation while being associated with the HDAC-containing complex NuRD. IKAROS also binds to the Positive-Transcription Elongation Factor b (P-TEFb) at gene promoters. Here, we demonstrate that NuRD and P-TEFb are assembled in a complex that can be recruited to specific genes by IKAROS. The expression level of IKAROS influences the recruitment of the NuRD-P-TEFb complex to gene regulatory regions and facilitates transcription elongation by transferring the Protein Phosphatase 1α (PP1α), an IKAROS-binding protein and P-TEFb activator, to CDK9. We show that an IKAROS mutant that is unable to bind PP1α cannot sustain gene expression and impedes normal differentiation of IkNULL hematopoietic progenitors. Finally, the knock-down of the NuRD subunit Mi2 reveals that the occupancy of the NuRD complex at transcribed regions of genes favors the relief of POL II promoter-proximal pausing and thereby, promotes transcription elongation. PMID:25474253

  13. Photochemical approaches to T-cell activation

    PubMed Central

    Huse, Morgan

    2010-01-01

    Despite decades of intensive research, T-cell activation has remained mysterious because of both the dizzying diversity of antigen recognition and the speed and comprehensiveness of the T-cell-receptor signalling network. Further progress will require new approaches and reagents that provide added levels of control. Photochemistry allows specific biochemical processes to be controlled with light and is well suited to mechanistic studies in complex cellular environments. In recent years, several laboratories have adopted approaches based on photoreactive peptide-major histocompatibility complex reagents in order to study T-cell activation and function with high precision. Here, I review these efforts and outline future directions for this exciting area of research. PMID:20406301

  14. Stability enhancement of an atomic force microscope for long-term force measurement including cantilever modification for whole cell deformation

    NASA Astrophysics Data System (ADS)

    Weafer, P. P.; McGarry, J. P.; van Es, M. H.; Kilpatrick, J. I.; Ronan, W.; Nolan, D. R.; Jarvis, S. P.

    2012-09-01

    Atomic force microscopy (AFM) is widely used in the study of both morphology and mechanical properties of living cells under physiologically relevant conditions. However, quantitative experiments on timescales of minutes to hours are generally limited by thermal drift in the instrument, particularly in the vertical (z) direction. In addition, we demonstrate the necessity to remove all air-liquid interfaces within the system for measurements in liquid environments, which may otherwise result in perturbations in the measured deflection. These effects severely limit the use of AFM as a practical tool for the study of long-term cell behavior, where precise knowledge of the tip-sample distance is a crucial requirement. Here we present a readily implementable, cost effective method of minimizing z-drift and liquid instabilities by utilizing active temperature control combined with a customized fluid cell system. Long-term whole cell mechanical measurements were performed using this stabilized AFM by attaching a large sphere to a cantilever in order to approximate a parallel plate system. An extensive examination of the effects of sphere attachment on AFM data is presented. Profiling of cantilever bending during substrate indentation revealed that the optical lever assumption of free ended cantilevering is inappropriate when sphere constraining occurs, which applies an additional torque to the cantilevers "free" end. Here we present the steps required to accurately determine force-indentation measurements for such a scenario. Combining these readily implementable modifications, we demonstrate the ability to investigate long-term whole cell mechanics by performing strain controlled cyclic deformation of single osteoblasts.

  15. Femtosecond laser fabricated microfluorescence-activated cell sorter for single cell recovery

    NASA Astrophysics Data System (ADS)

    Bragheri, F.; Paiè, P.; Nava, G.; Yang, T.; Minzioni, P.; Martinez Vazquez, R.; Bellini, N.; Ramponi, R.; Cristiani, I.; Osellame, R.

    2014-03-01

    Manipulation, sorting and recovering of specific live cells from samples containing less than a few thousand cells is becoming a major hurdle in rare cell exploration such as stem cell research or cell based diagnostics. Moreover the possibility of recovering single specific cells for culturing and further analysis would be of great impact in many biological fields ranging from regenerative medicine to cancer therapy. In recent years considerable effort has been devoted to the development of integrated and low-cost optofluidic devices able to handle single cells, which usually rely on microfluidic circuits that guarantee a controlled flow of the cells. Among the different microfabrication technologies, femtosecond laser micromachining (FLM) is ideally suited for this purpose as it provides the integration of both microfluidic and optical functions on the same glass chip leading to monolithic, robust and portable devices. Here a new optofluidic device is presented, which is capable of sorting and recovering of single cells, through optical forces, on the basis of their fluorescence and. Both fluorescence detection and single cell sorting functions are integrated in the microfluidic chip by FLM. The device, which is specifically designed to operate with a limited amount of cells but with a very high selectivity, is fabricated by a two-step process that includes femtosecond laser irradiation followed by chemical etching. The capability of the device to act as a micro fluorescence-activated cell sorter has been tested on polystyrene beads and on tumor cells and the results on the single live cell recovery are reported.

  16. [An electrochemical method for measuring metabolic activity and counting cells].

    PubMed

    Kuznetsov, B a; Khlupova, M e; Shleev, S V; Kaprel'iants, A S; Iaropolov, A I

    2006-01-01

    An express electrochemical method for determining the metabolic activity of live cells based on the possibility of an electron exchange between an electrode and elements of the biological electron transfer chain in the presence of a mediator is proposed. This method is useful for studying any live cells (animal, plant, and microbial), including anaerobic, dormant, and spore cells. The sample preparation and measurement itself does not take more than 30 min. The detection limit in a volume of 15 ml amounts to 10-5 cells/ml. The applicability of the assessment method of the metabolic activity level during the transition of the bacteria Mycobacterium smegmatis into an uncultivable dormant state was demonstrated. This method is of special value for medicine and environmental control, detecting latent forms of pathogens. An optimal combination of the methods for the express analysis of latent pathogens is proposed. PMID:17066962

  17. Epigenetic Changes during Hepatic Stellate Cell Activation

    PubMed Central

    Götze, Silke; Schumacher, Eva C.; Kordes, Claus; Häussinger, Dieter

    2015-01-01

    Background and Aims Hepatic stellate cells (HSC), which can participate in liver regeneration and fibrogenesis, have recently been identified as liver-resident mesenchymal stem cells. During their activation HSC adopt a myofibroblast-like phenotype accompanied by profound changes in the gene expression profile. DNA methylation changes at single genes have been reported during HSC activation and may participate in the regulation of this process, but comprehensive DNA methylation analyses are still missing. The aim of the present study was to elucidate the role of DNA methylation during in vitro activation of HSC. Methods and Results The analysis of DNA methylation changes by antibody-based assays revealed a strong decrease in the global DNA methylation level during culture-induced activation of HSC. To identify genes which may be regulated by DNA methylation, we performed a genome-wide Methyl-MiniSeq EpiQuest sequencing comparing quiescent and early culture-activated HSC. Approximately 400 differentially methylated regions with a methylation change of at least 20% were identified, showing either hypo- or hypermethylation during activation. Further analysis of selected genes for DNA methylation and expression were performed revealing a good correlation between DNA methylation changes and gene expression. Furthermore, global DNA demethylation during HSC activation was investigated by 5-bromo-2-deoxyuridine assay and L-mimosine treatment showing that demethylation was independent of DNA synthesis and thereby excluding a passive DNA demethylation mechanism. Conclusions In summary, in vitro activation of HSC initiated strong DNA methylation changes, which were associated with gene regulation. These results indicate that epigenetic mechanisms are important for the control of early HSC activation. Furthermore, the data show that global DNA demethylation during activation is based on an active DNA demethylation mechanism. PMID:26065684

  18. Activity of Eravacycline against Enterobacteriaceae and Acinetobacter baumannii, Including Multidrug-Resistant Isolates, from New York City

    PubMed Central

    Abdallah, Marie; Olafisoye, Olawole; Cortes, Christopher; Urban, Carl; Landman, David

    2014-01-01

    Eravacycline demonstrated in vitro activity against a contemporary collection of more than 4,000 Gram-negative pathogens from New York City hospitals, with MIC50/MIC90 values, respectively, for Escherichia coli of 0.12/0.5 μg/ml, Klebsiella pneumoniae of 0.25/1 μg/ml, Enterobacter aerogenes of 0.25/1 μg/ml, Enterobacter cloacae 0.5/1 μg/ml, and Acinetobacter baumannii of 0.5/1 μg/ml. Activity was retained against multidrug-resistant isolates, including those expressing KPC and OXA carbapenemases. For A. baumannii, eravacycline MICs correlated with increased expression of the adeB gene. PMID:25534744

  19. Contribution of myosin II activity to cell spreading dynamics.

    PubMed

    Nisenholz, Noam; Paknikar, Aishwarya; Köster, Sarah; Zemel, Assaf

    2016-01-14

    Myosin II activity and actin polymerization at the leading edge of the cell are known to be essential sources of cellular stress. However, a quantitative account of their separate contributions is still lacking; so is the influence of the coupling between the two phenomena on cell spreading dynamics. We present a simple analytic elastic theory of cell spreading dynamics that quantitatively demonstrates how actin polymerization and myosin activity cooperate in the generation of cellular stress during spreading. Consistent with experiments, myosin activity is assumed to polarize in response to the stresses generated during spreading. The characteristic response time and the overall spreading time are predicted to determine different evolution profiles of cell spreading dynamics. These include, a (regular) monotonic increase of cell projected area with time, a non-monotonic (overshooting) profile with a maximum, and damped oscillatory modes. In addition, two populations of myosin II motors are distinguished based on their location in the lamella; those located above the major adhesion zone at the cell periphery are shown to facilitate spreading whereas those in deeper regions of the lamella are shown to oppose spreading. We demonstrate that the attenuation of myosin activity in the two regions may result in reciprocal effects on spreading. These findings provide important new insight into the function of myosin II motors in the course of spreading. PMID:26481613

  20. Synthesis and cancer cell growth inhibitory activity of icaritin derivatives.

    PubMed

    Wang, Chen; Wu, Ping; Shi, Jing-Fang; Jiang, Zi-Hua; Wei, Xiao-Yi

    2015-07-15

    A series of icaritin derivatives bearing carboxylic acid or carboxylic ester groups are synthesized, and their in vitro cytotoxic activity against three cancer cell lines, MCF-7, MDA-MB-435s, and A549, are evaluated by MTT assay. Several derivatives including 2h, 2j, 5b and 5d show higher cytotoxic activity than the parent compound icaritin against these cancer cell lines. Compounds 5b and 5d are even more cytotoxic to MCF-7 cells than the clinic drug tamoxifen. Moreover, compound 5b is found to be non-toxic to normal cells (Vero) and both 5b and 5d exhibit good selectivity towards estrogen receptor positive MCF-7 breast cancer cells over estrogen receptor negative MDA-MB-435s breast cancer cells. The structure activity relationship analysis has revealed that mono-substitution at either C-3 or C-7 hydroxyl group of icaritin could improve the cytotoxicity of icaritin, and the C-3 hydroxyl group may be a preferable site for chemical modification. In addition, the length, the flexibility and the additional branching substituent group of the substitution chain(s) at both C-3 and C-7 hydroxyl groups can all affect the anti-cancer activity of these derivatives. PMID:26079090

  1. Mucosal Regulatory T Cells and T Helper 17 Cells in HIV-Associated Immune Activation

    PubMed Central

    Pandiyan, Pushpa; Younes, Souheil-Antoine; Ribeiro, Susan Pereira; Talla, Aarthi; McDonald, David; Bhaskaran, Natarajan; Levine, Alan D.; Weinberg, Aaron; Sekaly, Rafick P.

    2016-01-01

    Residual mucosal inflammation along with chronic systemic immune activation is an important feature in individuals infected with human immunodeficiency virus (HIV), and has been linked to a wide range of co-morbidities, including malignancy, opportunistic infections, immunopathology, and cardiovascular complications. Although combined antiretroviral therapy (cART) can reduce plasma viral loads to undetectable levels, reservoirs of virus persist, and increased mortality is associated with immune dysbiosis in mucosal lymphoid tissues. Immune-based therapies are pursued with the goal of improving CD4+ T-cell restoration, as well as reducing chronic immune activation in cART-treated patients. However, the majority of research on immune activation has been derived from analysis of circulating T cells. How immune cell alterations in mucosal tissues contribute to HIV immune dysregulation and the associated risk of non-infectious chronic complications is less studied. Given the significant differences between mucosal T cells and circulating T cells, and the immediate interactions of mucosal T cells with the microbiome, more attention should be devoted to mucosal immune cells and their contribution to systemic immune activation in HIV-infected individuals. Here, we will focus on mucosal immune cells with a specific emphasis on CD4+ T lymphocytes, such as T helper 17 cells and CD4+Foxp3+ regulatory T cells (Tregs), which play crucial roles in maintaining mucosal barrier integrity and preventing inflammation, respectively. We hypothesize that pro-inflammatory milieu in cART-treated patients with immune activation significantly contributes to enhanced loss of Th17 cells and increased frequency of dysregulated Tregs in the mucosa, which in turn may exacerbate immune dysfunction in HIV-infected patients. We also present initial evidence to support this hypothesis. A better comprehension of how pro-inflammatory milieu impacts these two types of cells in the mucosa will shed light

  2. Modified agar dilution susceptibility testing method for determining in vitro activities of antifungal agents, including azole compounds.

    PubMed Central

    Yoshida, T; Jono, K; Okonogi, K

    1997-01-01

    In vitro activities of antifungal agents, including azole compounds, against yeasts were easily determined by using RPMI-1640 agar medium and by incubating the plates in the presence of 20% CO2. The end point of inhibition was clear by this method, even in the case of azole compounds, because of the almost complete inhibition of yeast growth at high concentrations which permitted weak growth of some Candida strains by traditional methods. MICs obtained by the agar dilution method were similar to those obtained by the broth dilution method proposed by the National Committee for Clinical Laboratory Standards. PMID:9174197

  3. Endothelial juxtaposition of distinct adult stem cells activates angiogenesis signaling molecules in endothelial cells.

    PubMed

    Mohammadi, Elham; Nassiri, Seyed Mahdi; Rahbarghazi, Reza; Siavashi, Vahid; Araghi, Atefeh

    2015-12-01

    Efficacy of therapeutic angiogenesis needs a comprehensive understanding of endothelial cell (EC) function and biological factors and cells that interplay with ECs. Stem cells are considered the key components of pro- and anti-angiogenic milieu in a wide variety of physiopathological states, and interactions of EC-stem cells have been the subject of controversy in recent years. In this study, the potential effects of three tissue-specific adult stem cells, namely rat marrow-derived mesenchymal stem cells (rBMSCs), rat adipose-derived stem cells (rADSCs) and rat muscle-derived satellite cells (rSCs), on the endothelial activation of key angiogenic signaling molecules, including VEGF, Ang-2, VEGFR-2, Tie-2, and Tie2-pho, were investigated. Human umbilical vein endothelial cells (HUVECs) and rat lung microvascular endothelial cells (RLMECs) were cocultured with the stem cells or incubated with the stem cell-derived conditioned media on Matrigel. Following HUVEC-stem cell coculture, CD31-positive ECs were flow sorted and subjected to western blotting to analyze potential changes in the expression of the pro-angiogenic signaling molecules. Elongation and co-alignment of the stem cells were seen along the EC tubes in the EC-stem cell cocultures on Matrigel, with cell-to-cell dye communication in the EC-rBMSC cocultures. Moreover, rBMSCs and rADSCs significantly improved endothelial tubulogenesis in both juxtacrine and paracrine manners. These two latter stem cells dynamically up-regulated VEGF, Ang-2, VREGR-2, and Tie-2 but down-regulated Tie2-pho and the Tie2-pho/Tie-2 ratio in HUVECs. Induction of pro-angiogenic signaling in ECs by marrow- and adipose-derived MSCs further indicates the significance of stem cell milieu in angiogenesis dynamics. PMID:26068799

  4. Characterization of a serine protease-mediated cell death program activated in human leukemia cells

    SciTech Connect

    O'Connell, A.R.; Holohan, C.; Torriglia, A.; Lee, B.F.; Stenson-Cox, C. . E-mail: catherine.stenson@nuigalway.ie

    2006-01-01

    Tightly controlled proteolysis is a defining feature of apoptosis and caspases are critical in this regard. Significant roles for non-caspase proteases in cell death have been highlighted. Staurosporine causes a rapid induction of apoptosis in virtually all mammalian cell types. Numerous studies demonstrate that staurosporine can activate cell death under caspase-inhibiting circumstances. The aim of this study was to investigate the proteolytic mechanisms responsible for cell death under these conditions. To that end, we show that inhibitors of serine proteases can delay cell death in one such system. Furthermore, through profiling of proteolytic activation, we demonstrate, for the first time, that staurosporine activates a chymotrypsin-like serine protease-dependent cell death in HL-60 cells independently, but in parallel with the caspase controlled systems. Features of the serine protease-mediated system include cell shrinkage and apoptotic morphology, regulation of caspase-3, altered nuclear morphology, generation of an endonuclease and DNA degradation. We also demonstrate a staurosporine-induced activation of a putative 16 kDa chymotrypsin-like protein during apoptosis.

  5. DOCK2 regulates cell proliferation through Rac and ERK activation in B cell lymphoma

    SciTech Connect

    Wang, Lei; Nishihara, Hiroshi; Kimura, Taichi; Kato, Yasutaka; Tanino, Mishie; Nishio, Mitsufumi; Obara, Masato; Endo, Tomoyuki; Koike, Takao; Tanaka, Shinya

    2010-04-23

    DOCK2; a member of the CDM protein family, regulates cell motility and cytokine production through the activation of Rac in mammalian hematopoietic cells and plays a pivotal role in the modulation of the immune system. Here we demonstrated the alternative function of DOCK2 in hematopoietic tumor cells, especially in terms of its association with the tumor progression. Immunostaining for DOCK2 in 20 cases of human B cell lymphoma tissue specimens including diffuse large B cell lymphoma and follicular lymphoma revealed the prominent expression of DOCK2 in all of the lymphoma cells. DOCK2-knockdown (KD) of the B cell lymphoma cell lines, Ramos and Raji, using the lentiviral shRNA system presented decreased cell proliferation compared to the control cells. Furthermore, the tumor formation of DOCK2-KD Ramos cell in nude mice was significantly abrogated. Western blotting analysis and pull-down assay using GST-PAK-RBD kimeric protein suggested the presence of DOCK2-Rac-ERK pathway regulating the cell proliferation of these lymphoma cells. This is the first report to clarify the prominent role of DOCK2 in hematopoietic malignancy.

  6. Strategies to reduce dendritic cell activation through functional biomaterial design

    PubMed Central

    Hume, Patrick S.; He, Jing; Haskins, Kathryn; Anseth, Kristi S.

    2012-01-01

    Dendritic cells play a key role in determining adaptive immunity, and there is growing interest in characterizing and manipulating the interactions between dendritic cells and biomaterial surfaces. Contact with several common biomaterials can induce the maturation of immature dendritic cells, but substrates that reduce dendritic cell maturation are of particular interest within the field of cell-based therapeutics where the goal is to reduce the immune response to cell-laden material carriers. In this study, we use a materials-based strategy to functionalize poly(ethylene glycol) hydrogels with immobilized immunosuppressive factors (TGF-β1 and IL-10) to reduce the maturation of immature dendritic cells. TGF-β1 and IL-10 are commonly employed as soluble factors to program dendritic cells in vitro, and we demonstrate that these proteins retain bioactivity towards dendritic cells when immobilized on hydrogel surfaces. Following stimulation with lipopolysaccharide (LPS) and/or cytokines, a dendritic cell line interacting with the surfaces of immunosuppressive hydrogels expressed reduced markers of maturation, including IL-12 and MHCII. The bioactivity of these immunomodulatory hydrogels was further confirmed with primary bone marrow dendritic cells (BMDCs) isolated from non-obese diabetic (NOD) mice, as quantified by a decrease in activation markers and a significantly reduced capacity to activate T cells. Furthermore, by introducing a second signal to promote BMDC-material interactions combined with the presentation of tolerizing signals, the mulitfunctional PEG hydrogels were found to further increase signaling towards BMDCs, as evidenced by greater reductions in maturation markers. PMID:22361099

  7. Increasing resource allocation and research into tobacco control activities: a comprehensive approach including primary prevention, treatment and brief intervention.

    PubMed

    Richmond, R

    1993-01-01

    The range of tobacco control activities should be viewed as essential parts of a complex multi-component puzzle. Intervention strategies designed to address tobacco control should be comprehensive and include both primary and secondary prevention activities and be multi-faceted and capable of bringing about change at both the individual and broader social and cultural levels. In this paper I argue for a mutually inclusive framework in which the various components contribute in important and different ways. I examine the prevalence of smoking and identify the high risk groups, then I examine the range of available strategies and present the evidence for their success. I discuss the primary prevention approaches such as warning labels, taxes, price increases, workplace bans, education in schools, mass media and self-help materials, as well as brief interventions and treatment strategies which are conducted at the worksite, general practice and specialized cessation clinics. The areas for future research are delineated for increased resource allocation and include: the best ways to disseminate brief interventions to smokers, methods to motivate smokers; training of health professionals to deliver brief interventions; enhancing quitting and access to existing treatment resources among specific disadvantaged minority groups, e.g. migrants, unemployed youth, the effect on smoking prevalence of warning labels on cigarette packets and price rises on cigarettes. PMID:16818330

  8. Numerical power balance and free energy loss analysis for solar cells including optical, thermodynamic, and electrical aspects

    SciTech Connect

    Greulich, Johannes Höffler, Hannes; Würfel, Uli; Rein, Stefan

    2013-11-28

    A method for analyzing the power losses of solar cells is presented, supplying a complete balance of the incident power, the optical, thermodynamic, and electrical power losses and the electrical output power. The involved quantities have the dimension of a power density (units: W/m{sup 2}), which permits their direct comparison. In order to avoid the over-representation of losses arising from the ultraviolet part of the solar spectrum, a method for the analysis of the electrical free energy losses is extended to include optical losses. This extended analysis does not focus on the incident solar power of, e.g., 1000 W/m{sup 2} and does not explicitly include the thermalization losses and losses due to the generation of entropy. Instead, the usable power, i.e., the free energy or electro-chemical potential of the electron-hole pairs is set as reference value, thereby, overcoming the ambiguities of the power balance. Both methods, the power balance and the free energy loss analysis, are carried out exemplarily for a monocrystalline p-type silicon metal wrap through solar cell with passivated emitter and rear (MWT-PERC) based on optical and electrical measurements and numerical modeling. The methods give interesting insights in photovoltaic (PV) energy conversion, provide quantitative analyses of all loss mechanisms, and supply the basis for the systematic technological improvement of the device.

  9. Effects of physical activity on endothelial progenitor cells (EPCs)

    PubMed Central

    De Biase, Chiara; De Rosa, Roberta; Luciano, Rossella; De Luca, Stefania; Capuano, Ernesto; Trimarco, Bruno; Galasso, Gennaro

    2014-01-01

    Physical activity has a therapeutic role in cardiovascular disease (CVD), through its beneficial effects on endothelial function and cardiovascular system. Circulating endothelial progenitor cells (EPCs) are bone marrow (BM) derived cells that represent a novel therapeutic target in CVD patients, because of their ability to home to sites of ischemic injury and repair the damaged vessels. Several studies show that physical activity results in a significant increase in circulating EPCs, and, in particular, there are some evidence of the beneficial exercise-induced effects on EPCs activity in CVD settings, including coronary artery disease (CAD), heart failure (HF), and peripheral artery disease (PAD). The aim of this paper is to review the current evidence about the beneficial effects of physical exercise on endothelial function and EPCs levels and activity in both healthy subjects and patients with CVD. PMID:24550833

  10. Fluorescence activated cell sorting of plant protoplasts.

    PubMed

    Bargmann, Bastiaan O R; Birnbaum, Kenneth D

    2010-01-01

    High-resolution, cell type-specific analysis of gene expression greatly enhances understanding of developmental regulation and responses to environmental stimuli in any multicellular organism. In situ hybridization and reporter gene visualization can to a limited extent be used to this end but for high resolution quantitative RT-PCR or high-throughput transcriptome-wide analysis the isolation of RNA from particular cell types is requisite. Cellular dissociation of tissue expressing a fluorescent protein marker in a specific cell type and subsequent Fluorescence Activated Cell Sorting (FACS) makes it possible to collect sufficient amounts of material for RNA extraction, cDNA synthesis/amplification and microarray analysis. An extensive set of cell type-specific fluorescent reporter lines is available to the plant research community. In this case, two marker lines of the Arabidopsis thaliana root are used: P(SCR;)::GFP (endodermis and quiescent center) and P(WOX5;)::GFP (quiescent center). Large numbers (thousands) of seedlings are grown hydroponically or on agar plates and harvested to obtain enough root material for further analysis. Cellular dissociation of plant material is achieved by enzymatic digestion of the cell wall. This procedure makes use of high osmolarity-induced plasmolysis and commercially available cellulases, pectinases and hemicellulases to release protoplasts into solution. FACS of GFP-positive cells makes use of the visualization of the green versus the red emission spectra of protoplasts excited by a 488 nm laser. GFP-positive protoplasts can be distinguished by their increased ratio of green to red emission. Protoplasts are typically sorted directly into RNA extraction buffer and stored for further processing at a later time. This technique is revealed to be straightforward and practicable. Furthermore, it is shown that it can be used without difficulty to isolate sufficient numbers of cells for transcriptome analysis, even for very scarce

  11. Dopamine Modulates the Activity of Sensory Hair Cells

    PubMed Central

    Toro, Cecilia; Trapani, Josef G.; Pacentine, Itallia; Maeda, Reo; Sheets, Lavinia; Mo, Weike

    2015-01-01

    The senses of hearing and balance are subject to modulation by efferent signaling, including the release of dopamine (DA). How DA influences the activity of the auditory and vestibular systems and its site of action are not well understood. Here we show that dopaminergic efferent fibers innervate the acousticolateralis epithelium of the zebrafish during development but do not directly form synapses with hair cells. However, a member of the D1-like receptor family, D1b, tightly localizes to ribbon synapses in inner ear and lateral-line hair cells. To assess modulation of hair-cell activity, we reversibly activated or inhibited D1-like receptors (D1Rs) in lateral-line hair cells. In extracellular recordings from hair cells, we observed that D1R agonist SKF-38393 increased microphonic potentials, whereas D1R antagonist SCH-23390 decreased microphonic potentials. Using ratiometric calcium imaging, we found that increased D1R activity resulted in larger calcium transients in hair cells. The increase of intracellular calcium requires Cav1.3a channels, as a Cav1 calcium channel antagonist, isradipine, blocked the increase in calcium transients elicited by the agonist SKF-38393. Collectively, our results suggest that DA is released in a paracrine fashion and acts at ribbon synapses, likely enhancing the activity of presynaptic Cav1.3a channels and thereby increasing neurotransmission. SIGNIFICANCE STATEMENT The neurotransmitter dopamine acts in a paracrine fashion (diffusion over a short distance) in several tissues and bodily organs, influencing and regulating their activity. The cellular target and mechanism of the action of dopamine in mechanosensory organs, such as the inner ear and lateral-line organ, is not clearly understood. Here we demonstrate that dopamine receptors are present in sensory hair cells at synaptic sites that are required for signaling to the brain. When nearby neurons release dopamine, activation of the dopamine receptors increases the activity of

  12. Sunitinib activates Axl signaling in renal cell cancer.

    PubMed

    van der Mijn, Johannes C; Broxterman, Henk J; Knol, Jaco C; Piersma, Sander R; De Haas, Richard R; Dekker, Henk; Pham, Thang V; Van Beusechem, Victor W; Halmos, Balazs; Mier, James W; Jiménez, Connie R; Verheul, Henk M W

    2016-06-15

    Mass spectrometry-based phosphoproteomics provides a unique unbiased approach to evaluate signaling network in cancer cells. The tyrosine kinase inhibitor sunitinib is registered as treatment for patients with renal cell cancer (RCC). We investigated the effect of sunitinib on tyrosine phosphorylation in RCC tumor cells to get more insight in its mechanism of action and thereby to find potential leads for combination treatment strategies. Sunitinib inhibitory concentrations of proliferation (IC50) of 786-O, 769-p and A498 RCC cells were determined by MTT-assays. Global tyrosine phosphorylation was measured by LC-MS/MS after immunoprecipitation with the antiphosphotyrosine antibody p-TYR-100. Phosphoproteomic profiling of 786-O cells yielded 1519 phosphopeptides, corresponding to 675 unique proteins including 57 different phosphorylated protein kinases. Compared to control, incubation with sunitinib at its IC50 of 2 µM resulted in downregulation of 86 phosphopeptides including CDK5, DYRK3, DYRK4, G6PD, PKM and LDH-A, while 94 phosphopeptides including Axl, FAK, EPHA2 and p38α were upregulated. Axl- (y702), FAK- (y576) and p38α (y182) upregulation was confirmed by Western Blot in 786-O and A498 cells. Subsequent proliferation assays revealed that inhibition of Axl with a small molecule inhibitor (R428) sensitized 786-O RCC cells and immortalized endothelial cells to sunitinib up to 3 fold. In conclusion, incubation with sunitinib of RCC cells causes significant upregulation of multiple phosphopeptides including Axl. Simultaneous inhibition of Axl improves the antitumor activity of sunitinib. We envision that evaluation of phosphoproteomic changes by TKI treatment enables identification of new targets for combination treatment strategies. PMID:26815723

  13. T Cell-Extrinsic CD18 Attenuates Antigen-Dependent CD4+ T cell Activation In Vivo1

    PubMed Central

    Wu, Xingxin; Lahiri, Amit; Sarin, Ritu; Abraham, Clara

    2015-01-01

    The β2 integrins (CD11/CD18) are heterodimeric leukocyte adhesion molecules expressed on hematopoietic cells. The role of T cell-intrinsic CD18 in trafficking of naïve T cells to secondary lymphoid organs, and in antigen-dependent T cell activation in vitro and in vivo has been well-defined. However, the T cell-extrinsic role for CD18, including on antigen presenting cells (APC), in contributing to T cell activation in vivo is less well understood. We examined the role for T cell-extrinsic CD18 in the activation of WT CD4+ T cells in vivo through the adoptive transfer of DO11.10 Ag-specific CD4+ T cells into CD18−/− mice. We found that T cell-extrinsic CD18 was required for attenuating OVA-induced T cell proliferation in peripheral lymph nodes (PLN). The increased proliferation of WT DO11.10 CD4+ T cells in CD18−/− PLN was associated with a higher percentage of APC, and these APC demonstrated an increased activation profile and increased Ag-uptake, in particular in F4/80+ APC. Depletion of F4/80+ cells both reduced and equalized antigen-dependent T cell proliferation in CD18−/− relative to littermate control PLN, demonstrating that these cells play a critical role in the enhanced T cell proliferation in CD18−/− mice. Consistently, CD11b blockade, which is expressed on F4/80+ macrophages, enhanced the proliferation of DO11.10+ T cells in CD18+/− PLN. Thus, in contrast to the T cell-intrinsic essential role for CD18 in T cell activation, T cell-extrinsic expression of CD18 attenuates antigen-dependent CD4+ T cell activation in PLN in vivo. PMID:25801431

  14. PITBUL: a physics-based modeling package for imaging and tracking of airborne targets for HEL applications including active illumination

    NASA Astrophysics Data System (ADS)

    Van Zandt, Noah R.; McCrae, Jack E.; Fiorino, Steven T.

    2013-05-01

    Aimpoint acquisition and maintenance is critical to high energy laser (HEL) system performance. This study demonstrates the development by the AFIT/CDE of a physics-based modeling package, PITBUL, for tracking airborne targets for HEL applications, including atmospheric and sensor effects and active illumination, which is a focus of this work. High-resolution simulated imagery of the 3D airborne target in-flight as seen from the laser position is generated using the HELSEEM model, and includes solar illumination, laser illumination, and thermal emission. Both CW and pulsed laser illumination are modeled, including the effects of illuminator scintillation, atmospheric backscatter, and speckle, which are treated at a first-principles level. Realistic vertical profiles of molecular and aerosol absorption and scattering, as well as optical turbulence, are generated using AFIT/CDE's Laser Environmental Effects Definition and Reference (LEEDR) model. The spatially and temporally varying effects of turbulence are calculated and applied via a fast-running wave optical method known as light tunneling. Sensor effects, for example blur, sampling, read-out noise, and random photon arrival, are applied to the imagery. Track algorithms, including centroid and Fitts correlation, as a part of a closed loop tracker are applied to the degraded imagery and scored, to provide an estimate of overall system performance. To gauge performance of a laser system against a UAV target, tracking results are presented as a function of signal to noise ratio. Additionally, validation efforts to date involving comparisons between simulated and experimental tracking of UAVs are presented.

  15. Activation of human T-helper/inducer cell, T-cytotoxic/suppressor cell, B-cell, and natural killer (NK)-cells and induction of NK cell activity against K562 chronic myeloid leukemia cells with modified citrus pectin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background Modified citrus pectin (MCP) is known for its anti-cancer effects and its ability to be absorbed and circulated in the human body. In this report we tested the ability of MCP to induce the activation of human blood lymphocyte subsets including T-helper/inducer cell, Tcytotoxic/suppres...

  16. Antimicrobial characterisation of CEM-101 activity against respiratory tract pathogens, including multidrug-resistant pneumococcal serogroup 19A isolates.

    PubMed

    Farrell, David J; Sader, Helio S; Castanheira, Mariana; Biedenbach, Douglas J; Rhomberg, Paul R; Jones, Ronald N

    2010-06-01

    CEM-101 is a novel fluorinated macrolide-ketolide with potent activity against bacterial pathogens that are susceptible or resistant to other macrolide-lincosamide-streptogramin B (MLS(B))-ketolide agents. CEM-101 is being developed for oral and parenteral use in moderate to moderately severe community-acquired bacterial pneumonia. The objective of this study was to assess the activity of CEM-101 and comparators against contemporary respiratory tract infection (RTI) isolates. A worldwide sample of organisms was used, including Streptococcus pneumoniae [n=168; 59.3% erythromycin-resistant and 18 multidrug-resistant (MDR) serogroup 19A strains], Moraxella catarrhalis (n=21; 11 beta-lactamase positive), Haemophilus influenzae (n=100; 48 beta-lactamase positive), Haemophilus parainfluenzae and Haemophilus haemolyticus (n=12), and Legionella pneumophila (n=30). Testing and interpretation were performed using reference Clinical and Laboratory Standards Institute methods. CEM-101 was very potent against S. pneumoniae [minimum inhibitory concentration for 90% of the organisms (MIC90)=0.25 mg/L; highest MIC at 0.5 mg/L] and was 2- and > or =32-fold more active than telithromycin and clindamycin, respectively. CEM-101 also demonstrated potent activity against S. pneumoniae MDR-19A strains (MIC90=0.5 mg/L). CEM-101 was the most potent antimicrobial agent tested against L. pneumophila, with all MIC values at < or = 0.015 mg/L (telithromycin MIC90=0.03 mg/L). CEM-101 was as potent as azithromycin against Haemophilus spp. RTI pathogens (MIC90=2 mg/L), with no variations for beta-lactamase production. CEM-101 MIC values against M. catarrhalis were all at < or =0.5mg/L. Interestingly, CEM-101 potency was ca. 6 log(2) dilutions greater than telithromycin MIC results among 44 beta-haemolytic streptococci having telithromycin MICs > or = 2 mg/L. CEM-101 exhibited the greatest potency and widest spectrum of activity against RTI pathogens among the tested MLS(B)-ketolide agents

  17. The adhesion receptor CD44 promotes atherosclerosis by mediating inflammatory cell recruitment and vascular cell activation

    PubMed Central

    Cuff, Carolyn A.; Kothapalli, Devashish; Azonobi, Ijeoma; Chun, Sam; Zhang, Yuanming; Belkin, Richard; Yeh, Christine; Secreto, Anthony; Assoian, Richard K.; Rader, Daniel J.; Puré, Ellen

    2001-01-01

    Atherosclerosis causes most acute coronary syndromes and strokes. The pathogenesis of atherosclerosis includes recruitment of inflammatory cells to the vessel wall and activation of vascular cells. CD44 is an adhesion protein expressed on inflammatory and vascular cells. CD44 supports the adhesion of activated lymphocytes to endothelium and smooth muscle cells. Furthermore, ligation of CD44 induces activation of both inflammatory and vascular cells. To assess the potential contribution of CD44 to atherosclerosis, we bred CD44-null mice to atherosclerosis-prone apoE-deficient mice. We found a 50–70% reduction in aortic lesions in CD44-null mice compared with CD44 heterozygote and wild-type littermates. We demonstrate that CD44 promotes the recruitment of macrophages to atherosclerotic lesions. Furthermore, we show that CD44 is required for phenotypic dedifferentiation of medial smooth muscle cells to the “synthetic” state as measured by expression of VCAM-1. Finally, we demonstrate that hyaluronan, the principal ligand for CD44, is upregulated in atherosclerotic lesions of apoE-deficient mice and that the low-molecular-weight proinflammatory forms of hyaluronan stimulate VCAM-1 expression and proliferation of cultured primary aortic smooth muscle cells, whereas high-molecular-weight forms of hyaluronan inhibit smooth muscle cell proliferation. We conclude that CD44 plays a critical role in the progression of atherosclerosis through multiple mechanisms. PMID:11581304

  18. Transgelin-2 in B-Cells Controls T-Cell Activation by Stabilizing T Cell - B Cell Conjugates

    PubMed Central

    Chae, Myoung-Won; Kim, Hye-Ran; Kim, Chang-Hyun; Jun, Chang-Duk; Park, Zee-Yong

    2016-01-01

    The immunological synapse (IS), a dynamic and organized junction between T-cells and antigen presenting cells (APCs), is critical for initiating adaptive immunity. The actin cytoskeleton plays a major role in T-cell reorganization during IS formation, and we previously reported that transgelin-2, an actin-binding protein expressed in T-cells, stabilizes cortical F-actin, promoting T-cell activation in response to antigen stimulation. Transgelin-2 is also highly expressed in B-cells, although no specific function has been reported. In this study, we found that deficiency in transgelin-2 (TAGLN2-/-) in B-cells had little effect on B-cell development and activation, as measured by the expression of CD69, MHC class II molecules, and CD80/86. Nevertheless, in B-cells, transgelin-2 accumulated in the IS during the interaction with T-cells. These results led us to hypothesize that transgelin-2 may also be involved in IS stability in B-cells, thereby influencing T-cell function. Notably, we found that transgelin-2 deficiency in B-cells reduced T-cell activation, as determined by the release of IL-2 and interferon-γ and the expression of CD69. Furthermore, the reduced T-cell activation was correlated with reduced B-cell–T-cell conjugate formation. Collectively, these results suggest that actin stability in B-cells during IS formation is critical for the initiation of adaptive T-cell immunity. PMID:27232882

  19. FGF2 activates TRPC and Ca2+ signaling leading to satellite cell activation

    PubMed Central

    Liu, Yewei; Schneider, Martin F.

    2013-01-01

    Satellite cells, as stem cells of adult skeletal muscle, are tightly associated with the differentiated muscle fibers and remain quiescent in the absence of muscle damage. In response to an injury, the quiescent satellite cell is activated by soluble factors, including FGFs released from injured myofibers. Using immunostaining, we here first show that TRPC1 channels are highly expressed in satellite cells attached to muscle fibers. Since CD34, a traditional stem cell marker, was recently found to be expressed in skeletal muscle satellite cells we labeled living satellite cells in their physiological niche associated with host FDB fibers using anti-CD34-FITC antibody. We then monitored intra-cellular calcium in anti-CD34-FITC labeled satellite cells attached to muscle fibers using the calcium sensitive dye X rhod-1 which has little fluorescence cross talk with FITC. FGF2 increased intracellular calcium in satellite cells, which was antagonized by the TRPC channel blocker SKF 96365. Immunostaining showed that NFATc3 is highly expressed in satellite cells, but not in host FDB fibers. Elevation of intracellular calcium by FGF2 is accompanied by nuclear translocation of NFATc3 and NFATc2 and by an increase in the number of MyoD positive cells per muscle fiber, both of which were attenuated by TRPC blocker SKF 96365. Our results suggest a novel pathway of satellite cell activation where FGF2 enhances calcium influx through a TRPC channel, and the increased cytosolic calcium leads to both NFATc3 and NFATc2 nuclear translocation and enhanced number of MyoD positive satellite cells per muscle fiber. PMID:24575047

  20. Cell Micromanipulation with an Active Handheld Micromanipulator

    PubMed Central

    Tabarés, Jaime Cuevas; MacLachlan, Robert A.; Ettensohn, Charles A.

    2012-01-01

    The paper describes the use of an active handheld micromanipulator, known as Micron, for micromanipulation of cells. The device enables users to manipulate objects on the order of tens of microns in size, with the natural ease of use of a fully handheld tool. Micron senses its own position using a purpose-built microscale optical tracker, estimates the erroneous or undesired component of hand motion, and actively corrects it by deflecting its own tool tip using piezoelectric actuators. Benchtop experiments in tip positioning show that active compensation can reduce positioning error by up to 51% compared to unaided performance. Preliminary experiments in bisection of sea urchin embryos exhibit an increased success rate when performed with the help of Micron. PMID:21096452

  1. Master switches of T-cell activation and differentiation.

    PubMed

    Beier, K C; Kallinich, T; Hamelmann, E

    2007-04-01

    T-cells play a central role in allergic airway diseases such as bronchial asthma. The imbalance between allergen-specific pro-inflammatory and pro-allergic T-cell responses on one hand and regulatory or suppressive T-cell responses on the other may best explain the development of unwanted immune responses against environmental allergens, which lead to immunoglobulin E production and airway inflammation. A key role in the fine tuning of any T-cell response is provided by the engagement of so-called co-stimulatory molecules that are required for the full activation of T-cells and the recognition of antigens via the antigen-specific T-cell receptor. Many of these co-stimulatory molecules have been identified only recently, leading to a fundamental change in the overall understanding of T-cell regulation. Due to their pivotal impact on T-cell differentiation and control, co-stimulatory molecules are promising targets for therapeutic intervention in T-cell-regulated or -mediated immune disorders, including allergic diseases and asthma. In the present article, an attempt is made to summarise the current knowledge on the basic concept of co-stimulation, the presently known co-stimulatory molecules and their various functions on T-cell activation or suppression. The mini-series will be completed by two more articles describing the recent experimental studies and preliminary clinical findings regarding the role of co-stimulatory molecules in allergic disorders and bronchial asthma, and a discussion regarding the feasibility of co-stimulatory molecules as potential targets for the treatment of allergic airway disease. Although it is too early for any clinical implication or utilisation at this moment, the authors are convinced that a better understanding of co-stimulation in the context of allergic asthma will finally provide novel and promising approaches for treatment and prevention. PMID:17400879

  2. MST1 activation by curcumin mediates JNK activation, Foxo3a nuclear translocation and apoptosis in melanoma cells

    SciTech Connect

    Yu, Teng; Ji, Jiang; Guo, Yong-li

    2013-11-08

    Highlights: •Curcumin activates MST1 in melanoma cells. •MST1 mediates curcumin-induced apoptosis of melanoma cells. •ROS production is involved in curcumin-induced MST1 activation. •MST1 mediates curcumin-induced JNK activation in melanoma cells. •MST1 mediates curcumin-induced Foxo3a nuclear translocation and Bim expression. -- Abstract: Different groups including ours have shown that curcumin induces melanoma cell apoptosis, here we focused the role of mammalian Sterile 20-like kinase 1 (MST1) in it. We observed that curcumin activated MST1-dependent apoptosis in cultured melanoma cells. MST1 silencing by RNA interference (RNAi) suppressed curcumin-induced cell apoptosis, while MST1 over-expressing increased curcumin sensitivity. Meanwhile, curcumin induced reactive oxygen species (ROS) production in melanoma cells, and the ROS scavenger, N-acetyl-cysteine (NAC), almost blocked MST1 activation to suggest that ROS might be required for MST1 activation by curcumin. c-Jun N-terminal protein kinase (JNK) activation by curcumin was dependent on MST1, since MST1 inhibition by RNAi or NAC largely inhibited curcumin-induced JNK activation. Further, curcumin induced Foxo3 nuclear translocation and Bim-1 (Foxo3 target gene) expression in melanoma cells, such an effect by curcumin was inhibited by MST1 RNAi. In conclusion, we suggested that MST1 activation by curcumin mediates JNK activation, Foxo3a nuclear translocation and apoptosis in melanoma cells.

  3. Immunomodulation of phloretin by impairing dendritic cell activation and function.

    PubMed

    Lin, Chi-Chen; Chu, Ching-Liang; Ng, Chin-Sheng; Lin, Ching-Yen; Chen, Der-Yuan; Pan, I-Hong; Huang, Kao-Jean

    2014-05-01

    Dietary compounds in fruits and vegetables have been shown to exert many biological activities. In addition to antioxidant effects, a number of flavonoids are able to modulate inflammatory responses. Here, we demonstrated that phloretin (PT), a natural dihydrochalcone found in many fruits, suppressed the activation and function of mouse dendritic cells (DCs). Phloretin disturbed the multiple intracellular signaling pathways in DCs induced by the Toll-like receptor 4 (TLR4) agonist lipopolysaccharide (LPS), including ROS, MAPKs (ERK, JNK, p38 MAPK), and NF-κB, and thereby reducing the production of inflammatory cytokines and chemokines. Phloretin also effectively suppressed the activation of DCs treated with different dosages of LPS or various TLR agonists. The LPS-induced DC maturation was attenuated by phloretin because the expression levels of the MHC class II and the co-stimulatory molecules were down-regulated, which then inhibited the LPS-stimulating DCs and the subsequent naïve T cell activation in a mixed lymphocyte reaction. Moreover, in vivo administration of phloretin suppressed the phenotypic maturation of the LPS-challenged splenic DCs and decreased the IFN-γ production from the activated CD4 T cells. Thus, we suggest that phloretin may potentially be an immunomodulator by impairing the activation and function of DCs and phloretin-contained fruits may be helpful in the improvement of inflammation and autoimmune diseases. PMID:24651121

  4. Active stochastic stress fluctuations in the cell cytoskeleton stir the cell and activate primary cilia

    NASA Astrophysics Data System (ADS)

    Schmidt, Christoph F.; Fakhri, Nikta; Battle, Christopher; Ott, Carolyn M.; Wessel, Alok D.; Lippincott-Schwartz, Jennifer; Mackintosh, Frederick C.

    2015-03-01

    Cells are active systems with molecular force generation that drives complex dynamics at the supramolecular scale. Much of cellular dynamics is driven by myosin motors interacting with the actin cytoskeleton. We discovered active random ``stirring'' driven by cytoplasmic myosin as an intermediate mode of transport, different from both thermal diffusion and directed motor activity. We found a further manifestation of cytoskeletal dynamics in the active motion patterns of primary cilia generated by epithelial cells. These cilia were thought to be immotile due to the absence of dynein motors, but it turns out that their anchoring deeper inside the cell in combination with the strongly fluctuating cortex results in clearly measurable non-equilibrium fluctuations.

  5. Comparative analysis of signature genes in PRRSV-infected porcine monocyte-derived dendritic cells at differential activation statuses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Activation statuses of monocytic cells including monocytes, macrophages and dendritic cells (DCs) are critically important for antiviral immunity. In particular, some devastating viruses, including porcine reproductive and respiratory syndrome virus (PRRSV), are capable of directly infecting these c...

  6. Cell cooperation in coelomocyte cytotoxic activity of Paracentrotus lividus coelomocytes.

    PubMed

    Arizza, Vincenzo; Giaramita, Francesca Tiziana; Parrinello, Daniela; Cammarata, Matteo; Parrinello, Nicolò

    2007-06-01

    The coelomic fluid from the sea urchin Paracentrotus lividus contains several coelomocyte types including amoebocytes and uncoloured spherulocytes involved in immune defences. In the present paper, we show a Ca(2+)-dependent cytotoxic activity for the unfractionated coelomocytes assayed in vitro, with rabbit erythrocytes and the K562 tumour cell line. In a plaque-forming assay, whole coelomocyte preparations as well as density gradient separated coelomocyte populations revealed that cell populations enriched in uncoloured spherulocytes, exerted high cytotoxic activity by releasing lysins in the presence of amoebocytes. This cooperative effect could be dependent on soluble factors released by amoebocytes. With regard to this, we show that an enhanced cytotoxic activity was found by adding the supernatant from sonicated amoebocytes or hemocyte culture medium into spherulocyte preparations. PMID:17329136

  7. NK Cell-extrinsic IL-18 Signaling Is Required for Efficient NK Cell Activation to Vaccinia Virus

    PubMed Central

    Brandstadter, Joshua D.; Huang, Xiaopei; Yang, Yiping

    2014-01-01

    Summary NK cells are important for the control of vaccinia virus (VV) in vivo. Recent studies have shown that multiple pathways are required for effective activation of NK cells. These include both TLR-dependent and -independent pathways, as well as the NKG2D activating receptor that recognizes host stress-induced NKG2D ligands. However, it remains largely unknown what controls the upregulation of NKG2D ligands in response to VV infection. In this study, we first showed that IL-18 is critical for NK cell activation and viral clearance. We then demonstrated that IL-18 signaling on both NK cells and DCs is required for efficient NK cell activation upon VV infection in vitro. We further showed in vivo that efficient NK cell activation to VV is dependent on DCs and IL-18 signaling in non-NK cells, suggesting an essential role for NK cell-extrinsic IL-18 signaling in NK cell activation. Mechanistically, IL-18 signaling in DCs promotes expression of Rae-1, an NKG2D ligand. Collectively, our data reveal a previously unrecognized role for NK cell-extrinsic IL-18 signaling in NK cell activation through upregulation of NKG2D ligands. These observations may provide insights into the design of effective NK cell-based therapies for viral infections and cancer. PMID:24846540

  8. Optical Control of Living Cells Electrical Activity by Conjugated Polymers.

    PubMed

    Martino, Nicola; Bossio, Caterina; Vaquero Morata, Susana; Lanzani, Guglielmo; Antognazza, Maria Rosa

    2016-01-01

    Hybrid interfaces between organic semiconductors and living tissues represent a new tool for in-vitro and in-vivo applications. In particular, conjugated polymers display several optimal properties as substrates for biological systems, such as good biocompatibility, excellent mechanical properties, cheap and easy processing technology, and possibility of deposition on light, thin and flexible substrates. These materials have been employed for cellular interfaces like neural probes, transistors for excitation and recording of neural activity, biosensors and actuators for drug release. Recent experiments have also demonstrated the possibility to use conjugated polymers for all-optical modulation of the electrical activity of cells. Several in-vitro study cases have been reported, including primary neuronal networks, astrocytes and secondary line cells. Moreover, signal photo-transduction mediated by organic polymers has been shown to restore light sensitivity in degenerated retinas, suggesting that these devices may be used for artificial retinal prosthesis in the future. All in all, light sensitive conjugated polymers represent a new approach for optical modulation of cellular activity. In this work, all the steps required to fabricate a bio-polymer interface for optical excitation of living cells are described. The function of the active interface is to transduce the light stimulus into a modulation of the cell membrane potential. As a study case, useful for in-vitro studies, a polythiophene thin film is used as the functional, light absorbing layer, and Human Embryonic Kidney (HEK-293) cells are employed as the biological component of the interface. Practical examples of successful control of the cell membrane potential upon stimulation with light pulses of different duration are provided. In particular, it is shown that both depolarizing and hyperpolarizing effects on the cell membrane can be achieved depending on the duration of the light stimulus. The reported

  9. Ligand Mobility Modulates Immunological Synapse Formation and T Cell Activation

    PubMed Central

    Hsu, Chih-Jung; Hsieh, Wan-Ting; Waldman, Abraham; Clarke, Fiona; Huseby, Eric S.; Burkhardt, Janis K.; Baumgart, Tobias

    2012-01-01

    T cell receptor (TCR) engagement induces clustering and recruitment to the plasma membrane of many signaling molecules, including the protein tyrosine kinase zeta-chain associated protein of 70 kDa (ZAP70) and the adaptor SH2 domain-containing leukocyte protein of 76 kDa (SLP76). This molecular rearrangement results in formation of the immunological synapse (IS), a dynamic protein array that modulates T cell activation. The current study investigates the effects of apparent long-range ligand mobility on T cell signaling activity and IS formation. We formed stimulatory lipid bilayers on glass surfaces from binary lipid mixtures with varied composition, and characterized these surfaces with respect to diffusion coefficient and fluid connectivity. Stimulatory ligands coupled to these surfaces with similar density and orientation showed differences in their ability to activate T cells. On less mobile membranes, central supramolecular activation cluster (cSMAC) formation was delayed and the overall accumulation of CD3ζ at the IS was reduced. Analysis of signaling microcluster (MC) dynamics showed that ZAP70 MCs exhibited faster track velocity and longer trajectories as a function of increased ligand mobility, whereas movement of SLP76 MCs was relatively insensitive to this parameter. Actin retrograde flow was observed on all surfaces, but cell spreading and subsequent cytoskeletal contraction were more pronounced on mobile membranes. Finally, increased tyrosine phosphorylation and persistent elevation of intracellular Ca2+ were observed in cells stimulated on fluid membranes. These results point to ligand mobility as an important parameter in modulating T cell responses. PMID:22384241

  10. Telomerase activity in non-small cell lung cancer

    PubMed Central

    Dobija-Kubica, Katarzyna; Bruliński, Krzysztof; Rogoziński, Paweł; Wiczkowski, Andrzej; Gawrychowska, Agata; Gawrychowski, Jacek

    2016-01-01

    Introduction High telomerase activity has been detected in the majority of malignant neoplasms including lung cancer. The purpose of the study was to attempt to use telomerase activity as a prognostic factor in patients with non-small cell lung cancer (NSCLC). Material and methods Telomerase activity was analyzed in 47 tissue specimens taken from patients with NSCLC. The control group consisted of 30 specimens of non-cancerous lung parenchyma. Telomerase activity was measured by means of the telomeric repeat amplification protocol (TRAP). Results Telomerase activity in the neoplastic tissue was significantly higher than in the lung parenchyma that was free from neoplastic infiltration. There was no significant association between telomerase activity and age, gender, tobacco smoking, histological type of the tumor, or staging (pTNM). No association was found between the level of telomerase activity in NSCLC specimens and the two-year survival rate of patients (p = 0.326). A higher level of telomerase activity in poorly differentiated tumors (G3) as compared to moderately differentiated tumors (G2) was detected (p = 0.008). A positive association was identified between telomerase activity in pulmonary parenchyma free from tumor infiltration and the presence of leukocyte infiltration (p = 0.0001). Conclusions No association was found between the level of telomerase activity in NSCLC specimens and the two-year survival rate of patients. The study has revealed a positive association between telomerase activity and the grade of differentiation (G) in NSCLC. PMID:27212973

  11. HPV vaccine stimulates cytotoxic activity of killer dendritic cells and natural killer cells against HPV-positive tumour cells

    PubMed Central

    Van den Bergh, Johan M J; Guerti, Khadija; Willemen, Yannick; Lion, Eva; Cools, Nathalie; Goossens, Herman; Vorsters, Alex; Van Tendeloo, Viggo F I; Anguille, Sébastien; Van Damme, Pierre; Smits, Evelien L J M

    2014-01-01

    Cervarix™ is approved as a preventive vaccine against infection with the human papillomavirus (HPV) strains 16 and 18, which are causally related to the development of cervical cancer. We are the first to investigate in vitro the effects of this HPV vaccine on interleukin (IL)-15 dendritic cells (DC) as proxy of a naturally occurring subset of blood DC, and natural killer (NK) cells, two innate immune cell types that play an important role in antitumour immunity. Our results show that exposure of IL-15 DC to the HPV vaccine results in increased expression of phenotypic maturation markers, pro-inflammatory cytokine production and cytotoxic activity against HPV-positive tumour cells. These effects are mediated by the vaccine adjuvant, partly through Toll-like receptor 4 activation. Next, we demonstrate that vaccine-exposed IL-15 DC in turn induce phenotypic activation of NK cells, resulting in a synergistic cytotoxic action against HPV-infected tumour cells. Our study thus identifies a novel mode of action of the HPV vaccine in boosting innate immunity, including killing of HPV-infected cells by DC and NK cells. PMID:24979331

  12. Mesenchymal stem cells derived from low risk acute lymphoblastic leukemia patients promote NK cell antitumor activity.

    PubMed

    Entrena, Ana; Varas, Alberto; Vázquez, Miriam; Melen, Gustavo J; Fernández-Sevilla, Lidia M; García-Castro, Javier; Ramírez, Manuel; Zapata, Agustín G; Vicente, Ángeles

    2015-07-28

    Mesenchymal stem cells (MSCs) are key components of the bone marrow microenvironment which contribute to the maintenance of the hematopoietic stem cell niche and exert immunoregulatory functions in innate and adaptive immunity. We analyze the immunobiology of MSCs derived from acute lymphoblastic leukemia (ALL) patients and their impact on NK cell function. In contrast to the inhibitory effects on the immune response exerted by MSCs from healthy donors (Healthy-MSCs), we demonstrate that MSCs derived from low/intermediate risk ALL patients at diagnosis (ALL-MSCs) promote an efficient NK cell response including cytokine production, phenotypic activation and most importantly, cytotoxicity. Longitudinal studies indicate that these immunostimulatory effects of ALL-MSCs are progressively attenuated. Healthy-MSCs adopt ALL-MSC-like immunomodulatory features when exposed to leukemia cells, acquiring the ability to stimulate NK cell antitumor function. The mechanisms underlying to these functional changes of ALL-MSCs include reduced production of soluble inhibitory factors, differential expression of costimulatory and coinhibitory molecules, increased expression of specific TLRs and Notch pathway activation. Collectively our findings indicate that, in response to leukemia cells, ALL-MSCs could mediate a host beneficial immunomodulatory effect by stimulating the antitumor innate immune response. PMID:25917077

  13. Blockade of Mast Cell Activation Reduces Cutaneous Scar Formation

    PubMed Central

    Ranzer, Matthew J.; Wilgus, Traci A.; DiPietro, Luisa A.

    2014-01-01

    Damage to the skin initiates a cascade of well-orchestrated events that ultimately leads to repair of the wound. The inflammatory response is key to wound healing both through preventing infection and stimulating proliferation and remodeling of the skin. Mast cells within the tissue are one of the first immune cells to respond to trauma, and upon activation they release pro-inflammatory molecules to initiate recruitment of leukocytes and promote a vascular response in the tissue. Additionally, mast cells stimulate collagen synthesis by dermal fibroblasts, suggesting they may also influence scar formation. To examine the contribution of mast cells in tissue repair, we determined the effects the mast cell inhibitor, disodium cromoglycate (DSCG), on several parameters of dermal repair including, inflammation, re-epithelialization, collagen fiber organization, collagen ultrastructure, scar width and wound breaking strength. Mice treated with DSCG had significantly reduced levels of the inflammatory cytokines IL-1α, IL-1β, and CXCL1. Although DSCG treatment reduced the production of inflammatory mediators, the rate of re-epithelialization was not affected. Compared to control, inhibition of mast cell activity caused a significant decrease in scar width along with accelerated collagen re-organization. Despite the reduced scar width, DSCG treatment did not affect the breaking strength of the healed tissue. Tryptase β1 exclusively produced by mast cells was found to increase significantly in the course of wound healing. However, DSCG treatment did not change its level in the wounds. These results indicate that blockade of mast cell activation reduces scar formation and inflammation without further weakening the healed wound. PMID:24465509

  14. Blockade of mast cell activation reduces cutaneous scar formation.

    PubMed

    Chen, Lin; Schrementi, Megan E; Ranzer, Matthew J; Wilgus, Traci A; DiPietro, Luisa A

    2014-01-01

    Damage to the skin initiates a cascade of well-orchestrated events that ultimately leads to repair of the wound. The inflammatory response is key to wound healing both through preventing infection and stimulating proliferation and remodeling of the skin. Mast cells within the tissue are one of the first immune cells to respond to trauma, and upon activation they release pro-inflammatory molecules to initiate recruitment of leukocytes and promote a vascular response in the tissue. Additionally, mast cells stimulate collagen synthesis by dermal fibroblasts, suggesting they may also influence scar formation. To examine the contribution of mast cells in tissue repair, we determined the effects the mast cell inhibitor, disodium cromoglycate (DSCG), on several parameters of dermal repair including, inflammation, re-epithelialization, collagen fiber organization, collagen ultrastructure, scar width and wound breaking strength. Mice treated with DSCG had significantly reduced levels of the inflammatory cytokines IL-1α, IL-1β, and CXCL1. Although DSCG treatment reduced the production of inflammatory mediators, the rate of re-epithelialization was not affected. Compared to control, inhibition of mast cell activity caused a significant decrease in scar width along with accelerated collagen re-organization. Despite the reduced scar width, DSCG treatment did not affect the breaking strength of the healed tissue. Tryptase β1 exclusively produced by mast cells was found to increase significantly in the course of wound healing. However, DSCG treatment did not change its level in the wounds. These results indicate that blockade of mast cell activation reduces scar formation and inflammation without further weakening the healed wound. PMID:24465509

  15. Probing cell activity in random access modality

    NASA Astrophysics Data System (ADS)

    Sacconi, L.; Crocini, C.; Lotti, J.; Coppini, R.; Ferrantini, C.; Tesi, C.; Yan, P.; Loew, L. M.; Cerbai, E.; Poggesi, C.; Pavone, F. S.

    2013-06-01

    We combined the advantage of an ultrafast random access microscope with novel labelling technologies to study the intra- and inter-cellular action potential propagation in neurons and cardiac myocytes with sub-millisecond time resolution. The random accesses microscopy was used in combination with a new fluorinated voltage sensitive dye with improved photostability to record membrane potential from multiple Purkinje cells with near simultaneous sampling. The RAMP system rapidly scanned between lines drawn in the membranes of neurons to perform multiplex measurements of the TPF signal. This recording was achieved by rapidly positioning the laser excitation with the AOD to sample a patch of membrane from each cell in <100 μs for recording from five cells, multiplexing permits a temporal resolution of 400 μs sufficient to capture every spike. The system is capable to record spontaneous activity over 800 ms from five neighbouring cells simultaneously, showing that spiking is not temporally correlated. The system was also used to investigate the electrical properties of tubular system (TATS) in isolated rat ventricular myocytes.

  16. Aurora A kinase activity influences calcium signaling in kidney cells.

    PubMed

    Plotnikova, Olga V; Pugacheva, Elena N; Golemis, Erica A

    2011-06-13

    Most studies of Aurora A (AurA) describe it as a mitotic centrosomal kinase. However, we and others have recently identified AurA functions as diverse as control of ciliary resorption, cell differentiation, and cell polarity control in interphase cells. In these activities, AurA is transiently activated by noncanonical signals, including Ca(2+)-dependent calmodulin binding. These and other observations suggested that AurA might be involved in pathological conditions, such as polycystic kidney disease (PKD). In this paper, we show that AurA is abundant in normal kidney tissue but is also abnormally expressed and activated in cells lining PKD-associated renal cysts. PKD arises from mutations in the PKD1 or PKD2 genes, encoding polycystins 1 and 2 (PC1 and PC2). AurA binds, phosphorylates, and reduces the activity of PC2, a Ca(2+)-permeable nonselective cation channel and, thus, limits the amplitude of Ca(2+) release from the endoplasmic reticulum. These and other findings suggest AurA may be a relevant new biomarker or target in the therapy of PKD. PMID:21670214

  17. Catalytic activity of baker's yeast in a mediatorless microbial fuel cell.

    PubMed

    Sayed, Enas Taha; Tsujiguchi, Takuya; Nakagawa, Nobuyoshi

    2012-08-01

    The catalytic activity of baker's yeast, Saccharomyces cerevisiae, as a biocatalyst was investigated in a mediatorless microbial fuel cell. The yeast cells that adhered on the anode surface were the active biocatalyst for glucose oxidation in a mediatorless biofuel cell, suggesting that the electron transfer took place through the surface confined species. The species in the anolyte solution including the dispersed yeast cells did not take a part in the electron transfer and thus in the power generation. PMID:22357359

  18. Hymenoptera Allergy and Mast Cell Activation Syndromes.

    PubMed

    Bonadonna, Patrizia; Bonifacio, Massimiliano; Lombardo, Carla; Zanotti, Roberta

    2016-01-01

    Mast cell activation syndrome (MCAS) can be diagnosed in patients with recurrent, severe symptoms from mast cell (MC)-derived mediators, which are transiently increased in serum and are attenuated by mediator-targeting drugs. When KIT-mutated, clonal MC are detected in these patients, a diagnosis of primary MCAS can be made. Severe systemic reactions to hymenoptera venom (HV) represent the most common form of anaphylaxis in patients with mastocytosis. Patients with primary MCAS and HV anaphylaxis are predominantly males and do not have skin lesions in the majority of cases, and anaphylaxis is characterized by hypotension and syncope in the absence of urticaria and angioedema. A normal value of tryptase (≤11.4 ng/ml) in these patients does not exclude a diagnosis of mastocytosis. Patients with primary MCAS and HV anaphylaxis have to undergo lifelong venom immunotherapy, in order to prevent further potentially fatal severe reactions. PMID:26714690

  19. Cell context-dependent activities of parthenolide in primary and metastatic melanoma cells

    PubMed Central

    Czyz, M; Lesiak-Mieczkowska, K; Koprowska, K; Szulawska-Mroczek, A; Wozniak, M

    2010-01-01

    Background and purpose: Growing evidence implicates NF-κB as an important contributor to metastasis and increased chemoresistance of melanoma. Here, we report the effects of parthenolide on either untreated, cisplatin- or TNFα-treated melanoma cell lines A375, 1205Lu and WM793, exhibiting different levels of constitutive NF-κB activity. Experimental approach: Electrophoretic mobility shift assay was used to assess changes in NF-κB activity, and real-time PCR to evaluate expression of NF-κB-regulated genes. Cell cycle arrest and apoptosis were assessed by flow cytometry. Cell death was also visualized by fluorescence microscopy. Migration was determined by scratch assay and invasiveness by Matrigel assay. Key results: Parthenolide suppressed both constitutive and induced NF-κB activity in melanoma cells. This was accompanied by down-regulation of cancer-related genes, with NF-κB-binding sites in their promoters, including: Bcl-XL, survivin, cyclin D1, interleukin 8 and matrix metalloproteinase 9. When the various effects of 6 µM parthenolide were compared, apoptosis associated with loss of mitochondrial membrane potential was most efficiently induced in 1205Lu cells, cell cycle arrest in G0/G1 phase was observed in WM793 cells, and high metastatic potential was markedly reduced in A375 cells. These findings not only reflected differences between melanoma cell lines in basal expression of NF-κB-regulated genes, but also suggested other parthenolide targets involved in cell cycle progression, migration, invasiveness and survival. Conclusions: Inhibition of constitutive and therapeutically induced NF-κB pathway by parthenolide might be useful in the treatment of melanoma, although the diversity of changes induced in melanoma cells with different genetic backgrounds indicate context-dependent poly-pharmacological properties of this compound. PMID:20590608

  20. Sertoli Cells Maintain Leydig Cell Number and Peritubular Myoid Cell Activity in the Adult Mouse Testis

    PubMed Central

    Monteiro, Ana; Milne, Laura; Cruickshanks, Lyndsey; Jeffrey, Nathan; Guillou, Florian; Freeman, Tom C.; Mitchell, Rod T.; Smith, Lee B.

    2014-01-01

    The Sertoli cells are critical regulators of testis differentiation and development. In the adult, however, their known function is restricted largely to maintenance of spermatogenesis. To determine whether the Sertoli cells regulate other aspects of adult testis biology we have used a novel transgenic mouse model in which Amh-Cre induces expression of the receptor for Diphtheria toxin (iDTR) specifically within Sertoli cells. This causes controlled, cell-specific and acute ablation of the Sertoli cell population in the adult animal following Diphtheria toxin injection. Results show that Sertoli cell ablation leads to rapid loss of all germ cell populations. In addition, adult Leydig cell numbers decline by 75% with the remaining cells concentrated around the rete and in the sub-capsular region. In the absence of Sertoli cells, peritubular myoid cell activity is reduced but the cells retain an ability to exclude immune cells from the seminiferous tubules. These data demonstrate that, in addition to support of spermatogenesis, Sertoli cells are required in the adult testis both for retention of the normal adult Leydig cell population and for support of normal peritubular myoid cell function. This has implications for our understanding of male reproductive disorders and wider androgen-related conditions affecting male health. PMID:25144714

  1. Activated spinal cord ependymal stem cells rescue neurological function.

    PubMed

    Moreno-Manzano, Victoria; Rodríguez-Jiménez, Francisco Javier; García-Roselló, Mireia; Laínez, Sergio; Erceg, Slaven; Calvo, Maria Teresa; Ronaghi, Mohammad; Lloret, Maria; Planells-Cases, Rosa; Sánchez-Puelles, Jose María; Stojkovic, Miodrag

    2009-03-01

    Spinal cord injury (SCI) is a major cause of paralysis. Currently, there are no effective therapies to reverse this disabling condition. The presence of ependymal stem/progenitor cells (epSPCs) in the adult spinal cord suggests that endogenous stem cell-associated mechanisms might be exploited to repair spinal cord lesions. epSPC cells that proliferate after SCI are recruited by the injured zone, and can be modulated by innate and adaptive immune responses. Here we demonstrate that when epSPCs are cultured from rats with a SCI (ependymal stem/progenitor cells injury [epSPCi]), these cells proliferate 10 times faster in vitro than epSPC derived from control animals and display enhanced self renewal. Genetic profile analysis revealed an important influence of inflammation on signaling pathways in epSPCi after injury, including the upregulation of Jak/Stat and mitogen activated protein kinase pathways. Although neurospheres derived from either epSPCs or epSPCi differentiated efficiently to oligodendrocites and functional spinal motoneurons, a better yield of differentiated cells was consistently obtained from epSPCi cultures. Acute transplantation of undifferentiated epSPCi or the resulting oligodendrocyte precursor cells into a rat model of severe spinal cord contusion produced a significant recovery of motor activity 1 week after injury. These transplanted cells migrated long distances from the rostral and caudal regions of the transplant to the neurofilament-labeled axons in and around the lesion zone. Our findings demonstrate that modulation of endogenous epSPCs represents a viable cell-based strategy for restoring neuronal dysfunction in patients with spinal cord damage. PMID:19259940

  2. Generation of Insulin-Producing Cells from the Mouse Liver Using β Cell-Related Gene Transfer Including Mafa and Mafb

    PubMed Central

    Oishi, Hisashi; Tai, Pei-Han; Sekiguchi, Yukari; Koshida, Ryusuke; Jung, Yunshin; Kudo, Takashi; Takahashi, Satoru

    2014-01-01

    Recent studies on the large Maf transcription factors have shown that Mafb and Mafa have respective and distinctive roles in β-cell development and maturation. However, whether this difference in roles is due to the timing of the gene expression (roughly, expression of Mafb before birth and of Mafa after birth) or to the specific function of each gene is unclear. Our aim was to examine the functional differences between these genes that are closely related to β cells by using an in vivo model of β-like cell generation. We monitored insulin gene transcription by measuring bioluminescence emitted from the liver of insulin promoter-luciferase transgenic (MIP-Luc-VU) mice. Adenoviral gene transfers of Pdx1/Neurod/Mafa (PDA) and Pdx1/Neurod/Mafb (PDB) combinations generated intense luminescence from the liver that lasted for more than 1 week and peaked at 3 days after transduction. The peak signal intensities of PDA and PDB were comparable. However, PDA but not PDB transfer resulted in significant bioluminescence on day 10, suggesting that Mafa has a more sustainable role in insulin gene activation than does Mafb. Both PDA and PDB transfers ameliorated the glucose levels in a streptozotocin (STZ)-induced diabetic model for up to 21 days and 7 days, respectively. Furthermore, PDA transfer induced several gene expressions necessary for glucose sensing and insulin secretion in the liver on day 9. However, a glucose tolerance test and liver perfusion experiment did not show glucose-stimulated insulin secretion from intrahepatic β-like cells. These results demonstrate that bioluminescence imaging in MIP-Luc-VU mice provides a noninvasive means of detecting β-like cells in the liver. They also show that Mafa has a markedly intense and sustained role in β-like cell production in comparison with Mafb. PMID:25397325

  3. Mitochondrial uncouplers inhibit hepatic stellate cell activation

    PubMed Central

    2012-01-01

    Background Mitochondrial dysfunction participates in the progression of several pathologies. Although there is increasing evidence for a mitochondrial role in liver disease, little is known about its contribution to hepatic stellate cell (HSC) activation. In this study we investigated the role of mitochondrial activity through mild uncoupling during in vitro activation of HSCs. Methods Cultured primary human and mouse HSCs were treated with the chemical uncouplers FCCP and Valinomycin. ATP levels were measured by luciferase assay and production of reactive oxygen species was determined using the fluorescent probe DCFH-DA. Possible cytotoxicity by uncoupler treatment was evaluated by caspase 3/7 activity and cytoplasmic protease leakage. Activation of HSCs and their response to the pro-fibrogenic cytokine TGF-β was evaluated by gene expression of activation markers and signal mediators using RT-qPCR. Proliferation was measured by incorporation of EdU and protein expression of α-smooth muscle actin was analyzed by immunocytochemistry and western blot. Results FCCP and Valinomycin treatment mildly decreased ATP and reactive oxygen species levels. Both uncouplers increased the expression of mitochondrial genes such as Tfam and COXIV while inducing morphological features of quiescent mouse HSCs and abrogating TGF-β signal transduction. Mild uncoupling reduced HSC proliferation and expression of pro-fibrogenic markers of mouse and human HSCs. Conclusions Mild mitochondrial uncoupling inhibits culture-induced HSC activation and their response to pro-fibrogenic cytokines like TGF-β. These results therefore suggest mitochondrial uncoupling of HSCs as a strategy to reduce progression of liver fibrosis. PMID:22686625

  4. Detection of silent cells, synchronization and modulatory activity in developing cellular networks.

    PubMed

    Hjorth, Johannes J J; Dawitz, Julia; Kroon, Tim; Pires, Johny; Dassen, Valerie J; Berkhout, Janna A; Emperador Melero, Javier; Nadadhur, Aish G; Alevra, Mihai; Toonen, Ruud F; Heine, Vivi M; Mansvelder, Huibert D; Meredith, Rhiannon M

    2016-04-01

    Developing networks in the immature nervous system and in cellular cultures are characterized by waves of synchronous activity in restricted clusters of cells. Synchronized activity in immature networks is proposed to regulate many different developmental processes, from neuron growth and cell migration, to the refinement of synapses, topographic maps, and the mature composition of ion channels. These emergent activity patterns are not present in all cells simultaneously within the network and more immature "silent" cells, potentially correlated with the presence of silent synapses, are prominent in different networks during early developmental periods. Many current network analyses for detection of synchronous cellular activity utilize activity-based pixel correlations to identify cellular-based regions of interest (ROIs) and coincident cell activity. However, using activity-based correlations, these methods first underestimate or ignore the inactive silent cells within the developing network and second, are difficult to apply within cell-dense regions commonly found in developing brain networks. In addition, previous methods may ignore ROIs within a network that shows transient activity patterns comprising both inactive and active periods. We developed analysis software to semi-automatically detect cells within developing neuronal networks that were imaged using calcium-sensitive reporter dyes. Using an iterative threshold, modulation of activity was tracked within individual cells across the network. The distribution pattern of both inactive and active, including synchronous cells, could be determined based on distance measures to neighboring cells and according to different anatomical layers. PMID:26097169

  5. Activity of nintedanib in germ cell tumors.

    PubMed

    Steinemann, Gustav; Jacobsen, Christine; Gerwing, Mirjam; Hauschild, Jessica; von Amsberg, Gunhild; Höpfner, Michael; Nitzsche, Bianca; Honecker, Friedemann

    2016-02-01

    Germ cell tumors (GCTs) are the most frequent malignancy in male patients between 15 and 45 years of age. Cisplatin-based chemotherapy shows excellent cure rates, but patients with cisplatin-resistant GCTs have a poor prognosis. Nintedanib (BIBF 1120, Vargatef) inhibits the receptor classes vascular endothelial growth factor receptor, platelet derived growth factor receptor, and fibroblast growth factor receptor, and has shown activity against many tumors, as well as in idiopathic lung fibrosis and bleomycin-induced lung injury. Here, we investigated the antineoplastic and antiangiogenic properties of nintedanib in cisplatin-resistant and cisplatin-sensitive GCT cells, both alone and in combination with classical cytotoxic agents such as cisplatin, etoposide, and bleomycin. The half-maximal inhibitory concentration (IC50) of nintedanib was 4.5 ± 0.43 μmol/l, 3.1 ± 0.45 μmol/l, and 3.6 ± 0.33 μmol/l in cisplatin-sensitive NTERA2, 2102Ep, and NCCIT cells, whereas the IC50 doses of the cisplatin-resistant counterparts were 6.6 ± 0.37 μmol/l (NTERA2-R), 4.5 ± 0.83 μmol/l (2102Ep-R), and 6.1 ± 0.41 μmol/l (NCCIT-R), respectively. Single treatment with nintedanib induced apoptosis and resulted in a sustained reduction in the capacity of colony formation in both cisplatin-sensitive and cisplatin-resistant GCT cells. Cell cycle analysis showed that nintedanib induced a strong G0/G1-phase arrest in all investigated cell lines. Combination treatment with cisplatin did not result in additive, synergistic, or antagonistic effects. The in-vivo activity was studied using the chorioallantoic membrane assay and indicated the antiangiogenic potency of nintedanib with markedly reduced microvessel density. Topical treatment of inoculated tumor plaques resulted in a significant reduction of the tumor size. This indicates that nintedanib might be a promising substance in the treatment of GCT. PMID:26479145

  6. Leukemia-associated activating mutation of Flt3 expands dendritic cells and alters T cell responses.

    PubMed

    Lau, Colleen M; Nish, Simone A; Yogev, Nir; Waisman, Ari; Reiner, Steven L; Reizis, Boris

    2016-03-01

    A common genetic alteration in acute myeloid leukemia is the internal tandem duplication (ITD) in FLT3, the receptor for cytokine FLT3 ligand (FLT3L). Constitutively active FLT3-ITD promotes the expansion of transformed progenitors, but also has pleiotropic effects on hematopoiesis. We analyzed the effect of FLT3-ITD on dendritic cells (DCs), which express FLT3 and can be expanded by FLT3L administration. Pre-leukemic mice with the Flt3(ITD) knock-in allele manifested an expansion of classical DCs (cDCs) and plasmacytoid DCs. The expansion originated in DC progenitors, was cell intrinsic, and was further enhanced in Flt3(ITD/ITD) mice. The mutation caused the down-regulation of Flt3 on the surface of DCs and reduced their responsiveness to Flt3L. Both canonical Batf3-dependent CD8(+) cDCs and noncanonical CD8(+) cDCs were expanded and showed specific alterations in their expression profiles. Flt3(ITD) mice showed enhanced capacity to support T cell proliferation, including a cell-extrinsic expansion of regulatory T (T reg) cells. Accordingly, these mice restricted alloreactive T cell responses during graft-versus-host reaction, but failed to control autoimmunity without T reg cells. Thus, the FLT3-ITD mutation directly affects DC development, indirectly modulating T cell homeostasis and supporting T reg cell expansion. We hypothesize that this effect of FLT3-ITD might subvert immunosurveillance and promote leukemogenesis in a cell-extrinsic manner. PMID:26903243

  7. Isolation of Cells Specialized in Anticancer Alkaloid Metabolism by Fluorescence-Activated Cell Sorting.

    PubMed

    Carqueijeiro, Inês; Guimarães, Ana Luísa; Bettencourt, Sara; Martínez-Cortés, Teresa; Guedes, Joana G; Gardner, Rui; Lopes, Telma; Andrade, Cláudia; Bispo, Cláudia; Martins, Nuno Pimpão; Andrade, Paula; Valentão, Patrícia; Valente, Inês M; Rodrigues, José A; Duarte, Patrícia; Sottomayor, Mariana

    2016-08-01

    Plant specialized metabolism often presents a complex cell-specific compartmentation essential to accomplish the biosynthesis of valuable plant natural products. Hence, the disclosure and potential manipulation of such pathways may depend on the capacity to isolate and characterize specific cell types. Catharanthus roseus is the source of several medicinal terpenoid indole alkaloids, including the low-level anticancer vinblastine and vincristine, for which the late biosynthetic steps occur in specialized mesophyll cells called idioblasts. Here, the optical, fluorescence, and alkaloid-accumulating properties of C. roseus leaf idioblasts are characterized, and a methodology for the isolation of idioblast protoplasts by fluorescence-activated cell sorting is established, taking advantage of the distinctive autofluorescence of these cells. This achievement represents a crucial step for the development of differential omic strategies leading to the identification of candidate genes putatively involved in the biosynthesis, pathway regulation, and transmembrane transport leading to the anticancer alkaloids from C. roseus. PMID:27356972

  8. Antiproliferative Effects of New Dimeric Ellagitannin from Cornus alba in Prostate Cancer Cells Including Apoptosis-Related S-Phase Arrest.

    PubMed

    Park, Kwan Hee; Yin, Jun; Yoon, Ki Hoon; Hwang, Yoon Jeong; Lee, Min Won

    2016-01-01

    Activity-guided isolation of 80% acetone extract of Cornus alba, which is traditionally used as an anti-inflammatory, hemostatic and diuretic in Korea, yielded one novel compound, tentatively designated cornusiin H (13), together with 12 known compounds. The known compounds included four flavonoids (catechin (1), quercetin-3-O-β-D-glucuronide (2), quercetin-3-O-β-D-glucopyranoside (3), kaempferol-3-O-β-D-glucopyranoside (4)) and eight hydrolysable tannins (gallic acid (5), 2,6-di-O-galloyl-hamamelofuranoside (6), 2-galloyl-4-caffeoyl-L-threonic acid (7) 2,3-di-O-galloyl-4-caffeoyl-L-threonic acid (8), 1,2,3,4,6-penta-O-galloyl-β-D-glucopyranoside (9), cornusiin B (10), cornusiin A (11) and camptothin B (12)). All compounds exhibited potent 1,1-diphenyl-2-picrylhydrazyl (DPPH)-free radical scavenging activity. Especially, the radical scavenging activities of 6 and 9-13 were higher than that of vitamin C. Compounds 9, 11, 12 and 13 inhibited the production of nitric oxide (NO) in lipopolysaccharide-stimulated RAW264.7 cells to the same degree as N(G)-Monomethyl-L-arginine (L-NMMA). When the antiproliferative effects of the isolated compounds were assessed in prostate cancer cells, the dimeric ellagitannins (11-13) selectively inhibited LNCaP hormone-dependent prostate cancer cells. Flow cytometry analysis indicated that the dimeric ellagitannins induced apoptosis and S-phase arrest. These results suggest that dimeric ellagitannins from Cornus alba can be developed as functional materials or herbal medicines for prostate tumors such as benign prostate hyperplasia and early-stage prostate cancer. PMID:26805810

  9. Optimizing staining protocols for laser microdissection of specific cell types from the testis including carcinoma in situ.

    PubMed

    Sonne, Si Brask; Dalgaard, Marlene D; Nielsen, John Erik; Hoei-Hansen, Christina E; Rajpert-De Meyts, Ewa; Gjerdrum, Lise Mette; Leffers, Henrik

    2009-01-01

    Microarray and RT-PCR based methods are important tools for analysis of gene expression; however, in tissues containing many different cells types, such as the testis, characterization of gene expression in specific cell types can be severely hampered by noise from other cells. The laser microdissection technology allows for enrichment of specific cell types. However, when the cells are not morphologically distinguishable, it is necessary to use a specific staining method for the target cells. In this study we have tested different fixatives, storage conditions for frozen sections and staining protocols, and present two staining protocols for frozen sections, one for fast and specific staining of fetal germ cells, testicular carcinoma in situ cells, and other cells with embryonic stem cell-like properties that express the alkaline phosphatase, and one for specific staining of lipid droplet-containing cells, which is useful for isolation of the androgen-producing Leydig cells. Both protocols retain a morphology that is compatible with laser microdissection and yield RNA of a quality suitable for PCR and microarray analysis. PMID:19436754

  10. Development and flight evaluation of active controls in the L-1011. [including wing load alleviation and stability augmentation

    NASA Technical Reports Server (NTRS)

    Johnston, J. F.; Urie, D. M.

    1978-01-01

    Active controls in the Lockheed L-1011 for increased energy efficiency are discussed. Active wing load alleviation for extended span, increased aspect ratio, and active stability augmentation with a smaller tail for reduced drag and weight are among the topics considered. Flight tests of active wing load alleviation on the baseline aircraft and moving-base piloted simulation developing criteria for stability augmentation are described.

  11. Cell-cell contact between marrow stromal cells and myeloma cells via VCAM-1 and alpha(4)beta(1)-integrin enhances production of osteoclast-stimulating activity.

    PubMed

    Michigami, T; Shimizu, N; Williams, P J; Niewolna, M; Dallas, S L; Mundy, G R; Yoneda, T

    2000-09-01

    Myeloma is a unique hematologic malignancy that exclusively homes in the bone marrow and induces massive osteoclastic bone destruction presumably by producing cytokines that promote the differentiation of the hematopoietic progenitors to osteoclasts (osteoclastogenesis). It is recognized that neighboring bone marrow stromal cells influence the expression of the malignant phenotype in myeloma cells. This study examined the role of the interactions between myeloma cells and neighboring stromal cells in the production of osteoclastogenic factors to elucidate the mechanism underlying extensive osteoclastic bone destruction. A murine myeloma cell line 5TGM1, which causes severe osteolysis, expresses alpha(4)beta(1)-integrin and tightly adheres to the mouse marrow stromal cell line ST2, which expresses the vascular cell adhesion molecule-1 (VCAM-1), a ligand for alpha(4)beta(1)-integrin. Co-cultures of 5TGM1 with primary bone marrow cells generated tartrate-resistant acid phosphatase-positive multinucleated bone-resorbing osteoclasts. Co-cultures of 5TGM1 with ST2 showed increased production of bone-resorbing activity and neutralizing antibodies against VCAM-1 or alpha(4)beta(1)-integrin inhibited this. The 5TGM1 cells contacting recombinant VCAM-1 produced increased osteoclastogenic and bone-resorbing activity. The activity was not blocked by the neutralizing antibody to known osteoclastogenic cytokines including interleukin (IL)-1, IL-6, tumor necrosis factor, or parathyroid hormone-related peptide. These data suggest that myeloma cells are responsible for producing osteoclastogenic activity and that establishment of direct contact with marrow stromal cells via alpha(4)beta(1)-integrin/VCAM-1 increases the production of this activity by myeloma cells. They also suggest that the presence of stromal cells may provide a microenvironment that allows exclusive colonization of myeloma cells in the bone marrow. (Blood. 2000;96:1953-1960) PMID:10961900

  12. Kinase Activity Studied in Living Cells Using an Immunoassay

    ERIC Educational Resources Information Center

    Bavec, Aljos?a

    2014-01-01

    This laboratory exercise demonstrates the use of an immunoassay for studying kinase enzyme activity in living cells. The advantage over the classical method, in which students have to isolate the enzyme from cell material and measure its activity in vitro, is that enzyme activity is modulated and measured in living cells, providing a more…

  13. Random mitotic activities across human embryonic stem cell colonies.

    SciTech Connect

    Jin, Q.; Duggan, R.; Dasa, S.; Li, F.; Chen, L.

    2010-08-01

    A systemic and quantitative study was performed to examine whether different levels of mitotic activities, assessed by the percentage of S-phase cells at any given time point, existed at different physical regions of human embryonic stem (hES) cell colonies at 2, 4, 6 days after cell passaging. Mitotically active cells were identified by the positive incorporation of 5-bromo-2-deoxyuridine (BrdU) within their newly synthesized DNA. Our data indicated that mitotically active cells were often distributed as clusters randomly across the colonies within the examined growth period, presumably resulting from local deposition of newly divided cells. This latter notion was further demonstrated by the confined growth of enhanced green florescence protein (EGFP) expressing cells amongst non-GFP expressing cells. Furthermore, the overall percentage of mitotically active cells remained constantly at about 50% throughout the 6-day culture period, indicating mitotic activities of hES cell cultures were time-independent under current growth conditions.

  14. Gabapentin-induced mitogenic activity in rat pancreatic acinar cells.

    PubMed

    Dethloff, L; Barr, B; Bestervelt, L; Bulera, S; Sigler, R; LaGattuta, M; de La Iglesia, F

    2000-05-01

    Gabapentin induces pancreatic acinar cell tumors in rats through unknown, yet apparently nongenotoxic mechanisms. The primary objective of this study was to determine whether gabapentin acts as a tumor promoter by stimulating acinar cell proliferation in rat pancreas. To this end, indices of pancreatic growth, including increased pancreatic weight, stimulation of acinar cell proliferation, and/or enhanced expression of immediate-early oncogenes were monitored in rats given gabapentin in the diet at 2 g/kg/day for up to 12 months. Rats fed raw soy flour (RSF), a known inducer of pancreatic acinar cell tumors through cholecystokinin-mediated mitogenic stimulation, were used throughout as positive controls. In addition, recent data suggests that gabapentin binds to the alpha(2)delta subunit of a voltage-gated, L-type calcium channel. Because signaling pathways for proliferative processes in pancreatic acinar cells involve intracellular calcium mobilization, the effects of gabapentin on intracellular calcium mobilization ([Ca(2+)](i)) and (3)H-thymidine incorporation were investigated in pancreatic acinar cells isolated from normal rat pancreas and in the AR42J rat pancreatic tumor cell line. As indicated by BrdU labeling indices, acinar cell proliferation increased 3-fold by Day 3 of RSF treatment and remained slightly greater than controls throughout the experiment. Pancreatic weights of RSF-fed rats were 32 to 56% greater than controls throughout the experiment. In contrast, gabapentin had no effect on pancreatic weight or acinar cell labeling index, and therefore had no apparent effect on pancreatic growth. In isolated pancreatic acinar cells, however, gabapentin induced mobilization of intracellular calcium and caused a slight increase in (3)H-thymidine incorporation. The data suggest that gabapentin may possess low level mitogenic activity, which is not easily detectable in in vivo assays. PMID:10788559

  15. Activity of Debio1452, a FabI Inhibitor with Potent Activity against Staphylococcus aureus and Coagulase-Negative Staphylococcus spp., Including Multidrug-Resistant Strains

    PubMed Central

    Rhomberg, Paul R.; Kaplan, Nachum; Jones, Ronald N.; Farrell, David J.

    2015-01-01

    Staphylococcus aureus and coagulase-negative staphylococci (CoNS) are responsible for a wide variety of human infections. The investigational antibacterial Debio1450 (previously AFN-1720), a prodrug of Debio1452 (previously AFN-1252), specifically targets staphylococci without significant activity against other Gram-positive or Gram-negative species. Debio1452 inhibits FabI, an enzyme critical to fatty acid biosynthesis in staphylococci. The activity of Debio1452 against CoNS, methicillin-susceptible S. aureus (MSSA), and methicillin-resistant S. aureus (MRSA), including significant clones, was determined. A globally diverse collection of 574 patient isolates from 35 countries was tested that included CoNS (6 species, 103 strains), MSSA (154 strains), MRSA (163 strains), and molecularly characterized strains (including spa-typed MRSA clones; 154 strains). The isolates were tested for susceptibility by CLSI broth microdilution methods against Debio1452 and 10 comparators. The susceptibility rates for the comparators were determined using CLSI and EUCAST breakpoint criteria. All S. aureus and CoNS strains were inhibited by Debio1452 concentrations of ≤0.12 and ≤0.5 μg/ml, respectively. The MIC50s for MSSA, MRSA, and molecularly characterized MRSA strains were 0.004 μg/ml, and the MIC90s ranged from 0.008 to 0.03 μg/ml. The MICs were higher for the CoNS isolates (MIC50/90, 0.015/0.12 μg/ml). Among S. aureus strains, resistance was common for erythromycin (61.6%), levofloxacin (49.0%), clindamycin (27.6%), tetracycline (15.7%), and trimethoprim-sulfamethoxazole (7.0%). Debio1452 demonstrated potent activity against MSSA, MRSA, and CoNS. Debio1452 showed significantly greater activity overall (MIC50, 0.004 μg/ml) than the other agents tested against these staphylococcal species, which included dominant MRSA clones and strains resistant to currently utilized antimicrobial agents. PMID:25691627

  16. Enzymatic properties of immobilized Alcaligenes faecalis cells with cell-associated beta-glucosidase activity

    SciTech Connect

    Wheatly, M.A.; Phillips, C.R.

    1984-06-01

    Enzymatic properties of Alcaligenes faecalis cells immobilized in polyacrylamide were characterized and compared with those reported for the extracted enzyme, and with those measured for free cells. Many of the properties reflected those of the extracted enzyme rather than those measured in the free whole cells prior to immobilization, suggesting cell disruption during immobilization. These properties included the pH activity profile, a slightly broader pH stability profile, and the activation energy. Electron micrographs showed evidence of cell debris among the polymer matrix. The immobilized cells were not viable, and did not consume glucose. Thermal stability was less after immobilization with a half-line of 16 h at 45 degrees C, and 3.5 h at 50 degrees C. The immobilized preparation was more stable when stored lyophilized rather than in buffer, losing 23 and 52% activity, respectively, after six months. The enzyme was irreversibly inhibited by both acetate and citrate buffers. If the immobilized enzyme is to be used in conjunction with cellulases from Trichoderma reesei for cellulase saccharification, the optimal conditions would be pH 5.5 and 45 degrees C in a buffer containing no carboxylic acid groups.

  17. Isolation of biologically active nanomaterial (inclusion bodies) from bacterial cells

    PubMed Central

    2010-01-01

    Background In recent years bacterial inclusion bodies (IBs) were recognised as highly pure deposits of active proteins inside bacterial cells. Such active nanoparticles are very interesting for further downstream protein isolation, as well as for many other applications in nanomedicine, cosmetic, chemical and pharmaceutical industry. To prepare large quantities of a high quality product, the whole bioprocess has to be optimised. This includes not only the cultivation of the bacterial culture, but also the isolation step itself, which can be of critical importance for the production process. To determine the most appropriate method for the isolation of biologically active nanoparticles, three methods for bacterial cell disruption were analyzed. Results In this study, enzymatic lysis and two mechanical methods, high-pressure homogenization and sonication, were compared. During enzymatic lysis the enzyme lysozyme was found to attach to the surface of IBs, and it could not be removed by simple washing. As this represents an additional impurity in the engineered nanoparticles, we concluded that enzymatic lysis is not the most suitable method for IBs isolation. During sonication proteins are released (lost) from the surface of IBs and thus the surface of IBs appears more porous when compared to the other two methods. We also found that the acoustic output power needed to isolate the IBs from bacterial cells actually damages proteins structures, thereby causing a reduction in biological activity. High-pressure homogenization also caused some damage to IBs, however the protein loss from the IBs was negligible. Furthermore, homogenization had no side-effects on protein biological activity. Conclusions The study shows that among the three methods tested, homogenization is the most appropriate method for the isolation of active nanoparticles from bacterial cells. PMID:20831775

  18. Utility of hydroxyurea in mast cell activation syndrome

    PubMed Central

    2013-01-01

    Mast cell activation syndrome (MCAS) is a relatively recently recognized cause of chronic multisystem polymorbidity of a generally inflammatory theme. Patients with MCAS often report migratory soft tissue and/or bone pain which frequently responds poorly to typical (narcotic and non-narcotic) analgesics as well as atypical analgesics such as antidepressants and anticonvulsants. Hydroxyurea (HU) is an oral ribonucleotide reductase inhibitor commonly used in the treatment of chronic myeloproliferative neoplasms and sickle cell anemia. HU has been used to treat systemic mastocytosis, sometimes effecting improvement in MC activation symptoms but not tumor burden, suggesting potential utility of the drug in MCAS, too. Reported here are five cases of successful use of relatively low-dose HU in MCAS to reduce symptoms including previously refractory soft tissue and/or bone pain. HU may be useful in treating mediator symptoms in MCAS, but further study is needed to define optimal dosing strategies and patient subpopulations most likely to benefit. PMID:24192267

  19. Intracellular sensing of complement C3 activates cell autonomous immunity

    PubMed Central

    Tam, Jerry C.H.; Bidgood, Susanna R.; McEwan, William A.; James, Leo C.

    2014-01-01

    Pathogens traverse multiple barriers during infection including cell membranes. Here we show that during this transition pathogens carry covalently attached complement C3 into the cell, triggering immediate signalling and effector responses. Sensing of C3 in the cytosol activates MAVS-dependent signalling cascades and induces proinflammatory cytokine secretion. C3 also flags viruses for rapid proteasomal degradation, thereby preventing their replication. This system can detect both viral and bacterial pathogens but is antagonized by enteroviruses, such as rhinovirus and poliovirus, which cleave C3 using their 3C protease. The antiviral Rupintrivir inhibits 3C protease and prevents C3 cleavage, rendering enteroviruses susceptible to intracellular complement sensing. Thus, complement C3 allows cells to detect and disable pathogens that have invaded the cytosol. PMID:25190799

  20. Intracellular sensing of complement C3 activates cell autonomous immunity.

    PubMed

    Tam, Jerry C H; Bidgood, Susanna R; McEwan, William A; James, Leo C

    2014-09-01

    Pathogens traverse multiple barriers during infection, including cell membranes. We found that during this transition, pathogens carried covalently attached complement C3 into the cell, triggering immediate signaling and effector responses. Sensing of C3 in the cytosol activated mitochondrial antiviral signaling (MAVS)-dependent signaling cascades and induced proinflammatory cytokine secretion. C3 also flagged viruses for rapid proteasomal degradation, preventing their replication. This system could detect both viral and bacterial pathogens but was antagonized by enteroviruses, such as rhinovirus and poliovirus, which cleave C3 using their 3C protease. The antiviral rupintrivir inhibited 3C protease and prevented C3 cleavage, rendering enteroviruses susceptible to intracellular complement sensing. Thus, complement C3 allows cells to detect and disable pathogens that have invaded the cytosol. PMID:25190799

  1. Lymphokine-activated killer cell function of peripheral blood mononuclear cells, spleen cells and regional lymph node cells in gastric cancer patients.

    PubMed Central

    Karimine, N; Arinaga, S; Inoue, H; Nanbara, S; Ueo, H; Akiyoshi, T

    1994-01-01

    Lymphokine-activated killer (LAK) cells generated by culture of peripheral blood mononuclear cells (PBMC), spleen cells (SPC) and regional lymph node cells (LNC) with IL-2 for 4 days were examined for their functional capabilities in 29 patients with gastric carcinoma. The cytotoxic activity of LAK cells induced from LNC was significantly lower than that from either PBMC or SPC, although there was no difference between PBMC or SPC. The induction of mRNA of interferon-gamma (IFN-gamma) or tumour necrosis factor-alpha (TNF-alpha) and the production of these cytokines in the non-adherent LAK cells from LNC were also significantly reduced compared with those from PBMC or SPC. Further, the LAK cells from LNC secreted significantly lower levels of these cytokines when stimulated with tumour target, Raji cells, although the production of these cytokines was markedly increased by stimulation with the targets in all three cell populations. Phenotypic analysis of each cell population revealed a decreased proportion of the cells mediating natural killer (NK) activity, including CD16+, CD56+, and CD57+ cells in LNC either before or after culture, although OKIa1+ and CD25+ cells were uniformly increased in all cell populations after culture. Changes in subpopulations of CD4+ and CD8+ cells in LNC were not apparently different from PBMC or SPC. These results indicated the differential reactivity of each lymphocyte population to IL-2 and the reduced LAK cell function of LNC compared with PBMC or SPC in patients with gastric carcinoma. Images Fig. 2 PMID:8004819

  2. SB225002 Induces Cell Death and Cell Cycle Arrest in Acute Lymphoblastic Leukemia Cells through the Activation of GLIPR1

    PubMed Central

    Leal, Paulo C.; Bhasin, Manoj K.; Zenatti, Priscila Pini; Nunes, Ricardo J.; Yunes, Rosendo A.; Nowill, Alexandre E.; Libermann, Towia A.; Zerbini, Luiz Fernando; Yunes, José Andrés

    2015-01-01

    Acute Lymphoblastic Leukemia (ALL) is the most frequent childhood malignancy. In the effort to find new anti-leukemic agents, we evaluated the small drug SB225002 (N-(2-hydroxy-4-nitrophenyl)-N’-(2-bromophenyl)urea). Although initially described as a selective antagonist of CXCR2, later studies have identified other cellular targets for SB225002, with potential medicinal use in cancer. We found that SB225002 has a significant pro-apoptotic effect against both B- and T-ALL cell lines. Cell cycle analysis demonstrated that treatment with SB225002 induces G2-M cell cycle arrest. Transcriptional profiling revealed that SB225002-mediated apoptosis triggered a transcriptional program typical of tubulin binding agents. Network analysis revealed the activation of genes linked to the JUN and p53 pathways and inhibition of genes linked to the TNF pathway. Early cellular effects activated by SB225002 included the up-regulation of GLIPR1, a p53-target gene shown to have pro-apoptotic activities in prostate and bladder cancer. Silencing of GLIPR1 in B- and T-ALL cell lines resulted in increased resistance to SB225002. Although SB225002 promoted ROS increase in ALL cells, antioxidant N-Acetyl Cysteine pre-treatment only modestly attenuated cell death, implying that the pro-apoptotic effects of SB225002 are not exclusively mediated by ROS. Moreover, GLIPR1 silencing resulted in increased ROS levels both in untreated and SB225002-treated cells. In conclusion, SB225002 induces cell cycle arrest and apoptosis in different B- and T-ALL cell lines. Inhibition of tubulin function with concurrent activation of the p53 pathway, in particular, its downstream target GLIPR1, seems to underlie the anti-leukemic effect of SB225002. PMID:26302043

  3. SB225002 Induces Cell Death and Cell Cycle Arrest in Acute Lymphoblastic Leukemia Cells through the Activation of GLIPR1.

    PubMed

    de Vasconcellos, Jaíra Ferreira; Laranjeira, Angelo Brunelli Albertoni; Leal, Paulo C; Bhasin, Manoj K; Zenatti, Priscila Pini; Nunes, Ricardo J; Yunes, Rosendo A; Nowill, Alexandre E; Libermann, Towia A; Zerbini, Luiz Fernando; Yunes, José Andrés

    2015-01-01

    Acute Lymphoblastic Leukemia (ALL) is the most frequent childhood malignancy. In the effort to find new anti-leukemic agents, we evaluated the small drug SB225002 (N-(2-hydroxy-4-nitrophenyl)-N'-(2-bromophenyl)urea). Although initially described as a selective antagonist of CXCR2, later studies have identified other cellular targets for SB225002, with potential medicinal use in cancer. We found that SB225002 has a significant pro-apoptotic effect against both B- and T-ALL cell lines. Cell cycle analysis demonstrated that treatment with SB225002 induces G2-M cell cycle arrest. Transcriptional profiling revealed that SB225002-mediated apoptosis triggered a transcriptional program typical of tubulin binding agents. Network analysis revealed the activation of genes linked to the JUN and p53 pathways and inhibition of genes linked to the TNF pathway. Early cellular effects activated by SB225002 included the up-regulation of GLIPR1, a p53-target gene shown to have pro-apoptotic activities in prostate and bladder cancer. Silencing of GLIPR1 in B- and T-ALL cell lines resulted in increased resistance to SB225002. Although SB225002 promoted ROS increase in ALL cells, antioxidant N-Acetyl Cysteine pre-treatment only modestly attenuated cell death, implying that the pro-apoptotic effects of SB225002 are not exclusively mediated by ROS. Moreover, GLIPR1 silencing resulted in increased ROS levels both in untreated and SB225002-treated cells. In conclusion, SB225002 induces cell cycle arrest and apoptosis in different B- and T-ALL cell lines. Inhibition of tubulin function with concurrent activation of the p53 pathway, in particular, its downstream target GLIPR1, seems to underlie the anti-leukemic effect of SB225002. PMID:26302043

  4. Identification of Multiple DNA Copy Number Alterations Including Frequent 8p11.22 Amplification in Conjunctival Squamous Cell Carcinoma

    PubMed Central

    Asnaghi, Laura; Alkatan, Hind; Mahale, Alka; Othman, Maha; Alwadani, Saeed; Al-Hussain, Hailah; Jastaneiah, Sabah; Yu, Wayne; Maktabi, Azza; Edward, Deepak P.; Eberhart, Charles G.

    2014-01-01

    Purpose. Little is known about the molecular alterations that drive formation and growth of conjunctival squamous cell carcinoma (cSCC). We therefore sought to identify genetic changes that could be used as diagnostic markers or therapeutic targets. Methods. The DNA extracted from 10 snap-frozen cSCC tumor specimens and 2 in situ carcinomas was analyzed using array-based comparative genomic hybridization (aCGH), and further examined with NanoString and quantitative PCR. Results. The number of regions of DNA loss ranged from 1 to 23 per tumor, whereas gains and amplifications ranged from 1 to 15 per tumor. Most large regions of chromosomal gain and loss were confirmed by NanoString karyotype analysis. The commonest alteration was amplification of 8p11.22 in 9 tumors (75%), and quantitative PCR analysis revealed 100-fold or greater overexpression of ADAM3A mRNA from 8p11.22 locus. In addition, recurring losses were observed at 14q13.2 and 22q11.23, both lost in 5 (42%) of the 12 tumors, and at 12p13.31, lost in 4 (33%) of the 12 samples. Of the eight loci associated with the DNA damage repair syndrome xeroderma pigmentosum, three showed loss of at least one allele in our aCGH analysis, including XPA (9q22.33, one tumor), XPE/DDB2 (11p11.2, one tumor) and XPG/ERCC5 (13q33.1, three tumors). Conclusions. Conjunctival SCC contains a range of chromosomal alterations potentially important in tumor formation and growth. Amplification of 8p11.22 and overexpression of ADAM3A suggests a potential role for this protease. Our findings also suggest that defects in DNA repair loci are important in sporadic cSCC. PMID:25491297

  5. A Multimodal Approach Including Craniospinal Irradiation Improves the Treatment Outcome of High-risk Intracranial Nongerminomatous Germ Cell Tumors

    SciTech Connect

    Kim, Jun Won; Kim, Woo Chul; Cho, Jae Ho; Kim, Dong-Seok; Shim, Kyu-Won; Lyu, Chul Joo; Won, Sung Chul; Suh, Chang-Ok

    2012-11-01

    Purpose: To evaluate whether a multimodal approach including craniospinal irradiation (CSI) improves treatment outcome in nongerminomatous germ cell tumor (NGGCT) patients. Methods and Materials: We reviewed the records of 32 patients with NGGCTs. Fourteen patients belonged to the intermediate prognosis group (immature teratoma, teratoma with malignant transformation, and mixed tumors mainly composed of germinoma or teratoma), and 18 patients belonged to the poor prognosis group (other highly malignant tumors). Patients with pure germinoma or mature teratoma were excluded from this study. Nineteen patients were treated with a combination of surgery, chemotherapy, and radiotherapy (RT); 9 patients received chemotherapy plus RT; 3 patients received surgery plus RT; and 1 patient received RT alone. Twenty-seven patients received CSI with a median of 36 Gy (range, 20-41 Gy) plus focal boost of 18-30.6 Gy, and 5 patients received whole-brain RT (WBRT) (20-36 Gy) or focal RT (50.4-54 Gy). The rate of total and subtotal resection was 71.9%. The median follow-up for surviving patients was 121 months. Results: Treatment failed in 7 patients. Three of the 5 patients who received focal RT or WBRT had local failure. Four cerebrospinal fluid (CSF) failures occurred after CSI. No failure occurred in the intermediate prognosis group. Ten-year recurrence-free survival (RFS) and overall survival (OS) for all patients were 77.6% and 74.6%, respectively. Ten-year RFS for the intermediate and poor prognosis groups were 100% and 61.1%, respectively (p = 0.012). OS for the two groups were 85.1% and 66.7%, respectively (p = 0.215). Tumor histology and CSI were significant prognostic factors for RFS, and CSI was significantly associated with OS. Conclusions: A multimodal approach was effective for treating NGGCTs. CSI should be considered for patients with poor prognostic histology.

  6. A shed NKG2D ligand that promotes natural killer cell activation and tumor rejection

    PubMed Central

    Deng, Weiwen; Gowen, Benjamin G.; Zhang, Li; Wang, Lin; Lau, Stephanie; Iannello, Alexandre; Xu, Jianfeng; Rovis, Tihana L.; Xiong, Na; Raulet, David H.

    2016-01-01

    Immune cells, including natural killer (NK) cells, recognize transformed cells and eliminate them in a process termed immunosurveillance. It is thought that tumor cells evade immunosurveillance by shedding membrane ligands that bind to the NKG2D activating receptor on NK cells and/or T cells, and desensitize these cells. In contrast, we show that in mice, shedding of MULT1, a high affinity NKG2D ligand, causes NK cell activation and tumor rejection. Recombinant soluble MULT1 stimulated tumor rejection in mice. Soluble MULT1 functions, at least in part, by competitively reversing a global desensitization of NK cells imposed by engagement of membrane NKG2D ligands on tumor-associated cells, such as myeloid cells. The results overturn conventional wisdom that soluble ligands are inhibitory, and suggest a new approach for cancer immunotherapy. PMID:25745066

  7. Superoxide radical and iron modulate aconitase activity in mammalian cells.

    PubMed

    Gardner, P R; Raineri, I; Epstein, L B; White, C W

    1995-06-01

    Aconitase is a member of a family of iron-sulfur-containing (de)hydratases whose activities are modulated in bacteria by superoxide radical (O2-.)-mediated inactivation and iron-dependent reactivation. The inactivation-reactivation of aconitase(s) in cultured mammalian cells was explored since these reactions may impact important and diverse aconitase functions in the cytoplasm and mitochondria. Conditions which increase O2-. production including exposure to the redox-cycling agent phenazine methosulfate (PMS), inhibitors of mitochondrial ubiquinol-cytochrome c oxidoreductase, or hyperoxia inactivated aconitase in mammalian cells. Overproduction of mitochondrial Mn-superoxide dismutase protected aconitase from inactivation by PMS or inhibitors of ubiquinol-cytochrome c oxidoreductase, but not from normobaric hyperoxia. Aconitase activity was reactivated (t1/2 of 12 +/- 3 min) upon removal of PMS. The iron chelator deferoxamine impaired reactivation and increased net inactivation of aconitase by O2-.. The ability of ubiquinol-cytochrome c oxidoreductase-generated O2-. to inactivate aconitase in several cell types correlated with the fraction of the aconitase activity localized in mitochondria. Extracellular O2-. generated with xanthine oxidase did not affect aconitase activity nor did exogenous superoxide dismutase decrease aconitase inactivation by PMS. The results demonstrate a dynamic and cyclical O2-.-mediated inactivation and iron-dependent reactivation of the mammalian [4Fe-4S] aconitases under normal and stress conditions and provide further evidence for the membrane compartmentalization of O2-.. PMID:7768942

  8. JAK1 Activates STAT3 Activity in Non-Small-Cell Lung Cancer cells and IL-6 Neutralizing Antibodies can Suppress JAK1-STAT3 Signaling

    PubMed Central

    Song, Lanxi; Rawal, Bhupendra; Nemeth, Jeffrey A.; Haura, Eric B.

    2014-01-01

    Members of the signal transducer and activator of transcription (STAT) family of transcription factors are potential targets for the treatment and prevention of cancers including non-small-cell lung cancer. STAT proteins can be phosphorylated and activated by diverse upstream kinases including cytokine receptors and tyrosine kinases. We examined STAT protein activation in lung cancer cell lines including those with activating mutations in the EGFR and examined upstream kinases responsible for STAT3 phosphorylation and activation using small molecules, antibodies, and RNA interference. We found more pronounced STAT3 activation in cells with activating EGFR mutations yet inhibition of EGFR activity had no effect on STAT3 activation. Inhibition of JAK1 with small molecules or RNA interference resulted in loss of STAT3 tyrosine phosphorylation and inhibition of cell growth. An interleukin-6 neutralizing antibody, siltuximab (CNTO 328) could inhibit STAT3 tyrosine phosphorylation in a cell-dependent manner. Siltuximab could completely inhibit STAT3 tyrosine phosphorylation in H1650 cells and this resulted in inhibition of lung cancer cell growth in vivo. Combined EGFR inhibition with erlotinib and siltuximab resulted in dual inhibition of both tyrosine and serine STAT3 phosphorylation, more pronounced inhibition of STAT3 transcriptional activity, and translated into combined effects on lung cancer growth in a mouse model. Our results suggest that JAK1 is responsible for STAT3 activation in lung cancer cells, and that indirect attacks on JAK1-STAT3 using an IL-6 neutralizing antibody with or without EGFR inhibition can inhibit lung cancer growth in lung cancer subsets. PMID:21216930

  9. HEPS Inventory Tool: An Inventory Tool Including Quality Assessment of School Interventions on Healthy Eating and Physical Activity

    ERIC Educational Resources Information Center

    Dadaczynski, Kevin; Paulus, Peter; de Vries, Nanne; de Ruiter, Silvia; Buijs, Goof

    2010-01-01

    The HEPS Inventory Tool aims to support stakeholders working in school health promotion to promote high quality interventions on healthy eating and physical activity. As a tool it provides a step-by-step approach on how to develop a national or regional inventory of existing school based interventions on healthy eating and physical activity. It…

  10. Efficiency enhancement in solid state dye sensitized solar cells by including inverse opals with controlled layer thicknesses

    NASA Astrophysics Data System (ADS)

    Zheng, Hanbin; Shah, Said Karim; Abbas, Mamatimin; Ly, Isabelle; Rivera, Thomas; Almeida, Rui M.; Hirsch, Lionel; Toupance, Thierry; Ravaine, Serge

    2016-09-01

    The photoconversion efficiency of dye sensitized solar cells can be enhanced by the incorporation of light management nanostructures such as photonic crystals. Here, we present a facile route to incorporate titania inverse opals into solid state dye sensitized solar cells and report photoconversion efficiency enhancements of up to 56% compared with a model system without the inverse opal. Our approach is based on the precise design of titania inverse opals with a predetermined thickness that can be controlled at the individual layer level. By choosing an inverse opal exhibiting a photonic bandgap which overlaps the absorption bands of the dye, our results show that there is an optimal thickness of the inverse opal structure for maximum efficiency enhancement of the cell. This is the first experimental proof that the thickness of a titania inverse opal plays a pivotal role in cell efficiency enhancement in solid state dye sensitized solar cells.

  11. MAIT cells are activated during human viral infections.

    PubMed

    van Wilgenburg, Bonnie; Scherwitzl, Iris; Hutchinson, Edward C; Leng, Tianqi; Kurioka, Ayako; Kulicke, Corinna; de Lara, Catherine; Cole, Suzanne; Vasanawathana, Sirijitt; Limpitikul, Wannee; Malasit, Prida; Young, Duncan; Denney, Laura; Moore, Michael D; Fabris, Paolo; Giordani, Maria Teresa; Oo, Ye Htun; Laidlaw, Stephen M; Dustin, Lynn B; Ho, Ling-Pei; Thompson, Fiona M; Ramamurthy, Narayan; Mongkolsapaya, Juthathip; Willberg, Christian B; Screaton, Gavin R; Klenerman, Paul

    2016-01-01

    Mucosal-associated invariant T (MAIT) cells are abundant in humans and recognize bacterial ligands. Here, we demonstrate that MAIT cells are also activated during human viral infections in vivo. MAIT cells activation was observed during infection with dengue virus, hepatitis C virus and influenza virus. This activation-driving cytokine release and Granzyme B upregulation-is TCR-independent but dependent on IL-18 in synergy with IL-12, IL-15 and/or interferon-α/β. IL-18 levels and MAIT cell activation correlate with disease severity in acute dengue infection. Furthermore, HCV treatment with interferon-α leads to specific MAIT cell activation in vivo in parallel with an enhanced therapeutic response. Moreover, TCR-independent activation of MAIT cells leads to a reduction of HCV replication in vitro mediated by IFN-γ. Together these data demonstrate MAIT cells are activated following viral infections, and suggest a potential role in both host defence and immunopathology. PMID:27337592

  12. Bisphenol A (BPA) stimulates the interferon signaling and activates the inflammasome activity in myeloid cells.

    PubMed

    Panchanathan, Ravichandran; Liu, Hongzhu; Leung, Yuet-Kin; Ho, Shuk-mei; Choubey, Divaker

    2015-11-01

    Environmental factors contribute to the development of autoimmune diseases, including systemic lupus erythematosus (SLE), which exhibits a strong female bias (female-to-male ratio 9:1). However, the molecular mechanisms remain largely unknown. Because a feedforward loop between the female sex hormone estrogen (E2) and type I interferon (IFN-α/β)-signaling induces the expression of certain p200-family proteins (such as murine p202 and human IFI16) that regulate innate immune responses and modify lupus susceptibility, we investigated whether treatment of myeloid cells with bisphenol A (BPA), an environmental estrogen, could regulate the p200-family proteins and activate innate immune responses. We found that treatment of murine bone marrow-derived cells (BMCs) and human peripheral blood mononuclear cells with BPA induced the expression of ERα and IFN-β, activated the IFN-signaling, and stimulated the expression of the p202 and IFI16 proteins. Further, the treatment increased levels of the NLRP3 inflammasome and stimulated its activity. Accordingly, BPA-treatment of BMCs from non lupus-prone C57BL/6 and the lupus-prone (NZB×NZW)F1 mice activated the type I IFN-signaling, induced the expression of p202, and activated an inflammasome activity. Our study demonstrates that BPA-induced signaling in the murine and human myeloid cells stimulates the type I IFN-signaling that results in an induction of the p202 and IFI16 innate immune sensors for the cytosolic DNA and activates an inflammasome activity. These observations provide novel molecular insights into the role of environmental BPA exposures in potentiating the development of certain autoimmune diseases such as SLE. PMID:26277401

  13. T cell stimulator cells, an efficient and versatile cellular system to assess the role of costimulatory ligands in the activation of human T cells

    PubMed Central

    Leitner, Judith; Kuschei, Werner; Grabmeier-Pfistershammer, Katharina; Woitek, Ramona; Kriehuber, Ernst; Majdic, Otto; Zlabinger, Gerhard; Pickl, Winfried F.; Steinberger, Peter

    2010-01-01

    It is well established that full activation of T cells requires the interaction of the TCR complex with the peptide–MHC complex (Signal 1) and additional signals (Signal 2). These second signals are generated by the interaction of costimulatory ligands expressed on antigen presenting cells with activating receptors on T cells. In addition, T cell responses are negatively regulated by inhibitory costimulatory pathways. Since professional antigen presenting cells (APC) harbour a plethora of stimulating and inhibitory surface molecules, the contribution of individual costimulatory molecules is difficult to assess on these cells. We have developed a system of stimulator cells that can give signal 1 to human T cells via a membrane bound anti-CD3 antibody fragment. By expressing human costimulatory ligands on these cells, their role in T cell activation processes can readily be analyzed. We demonstrate that T cell stimulator cells are excellent tools to study various aspects of human T cell costimulation, including the effects of immunomodulatory drugs or how costimulatory signals contribute to the in vitro expansion of T cells. T cell stimulator cells are especially suited for the functional evaluation of ligands that are implicated in costimulatory processes. In this study we have evaluated the role of the CD2 family member CD150 (SLAM) and the TNF family member TL1A (TNFSF15) in the activation of human T cells. Whereas our results do not point to a significant role of CD150 in T cell activation we found TL1A to potently costimulate human T cells. Taken together our results demonstrate that T cell stimulator cells are excellent tools to study various aspects of costimulatory processes. PMID:20858499

  14. Chronic lymphocytic leukemia: a disease of activated monoclonal B cells

    PubMed Central

    Damle, Rajendra N.; Calissano, Carlo; Chiorazzi, Nicholas

    2010-01-01

    B-cell type chronic lymphocytic leukemia (CLL) has long been considered a disease of resting lymphocytes. However cell surface and intracellular phenotypes suggest that most CLL cells are activated cells, although only a small subset progresses beyond the G1 stage of the cell cycle. In addition, traditional teaching says that CLL cells divide rarely, and therefore the buildup of leukemic cells is due to an inherent defect in cell death. However, in vivo labeling of CLL cells indicates a much more active rate of cell birth than originally estimated, suggesting that CLL is a dynamic disease. Here we review the observations that have led to these altered views of the activation state and proliferative capacities of CLL cells and also provide our interpretation of these observations in light of their potential impact on patients. PMID:20620969

  15. A multi-stage process including transient polyploidization and EMT precedes the emergence of chemoresistent ovarian carcinoma cells with a dedifferentiated and pro-inflammatory secretory phenotype

    PubMed Central

    Rohnalter, Verena; Roth, Katrin; Finkernagel, Florian; Adhikary, Till; Obert, Julia; Dorzweiler, Kristina; Bensberg, Maike; Müller-Brüsselbach, Sabine; Müller, Rolf

    2015-01-01

    DNA-damaging drugs induce a plethora of molecular and cellular alterations in tumor cells, but their interrelationship is largely obscure. Here, we show that carboplatin treatment of human ovarian carcinoma SKOV3 cells triggers an ordered sequence of events, which precedes the emergence of mitotic chemoresistant cells. The initial phase of cell death after initiation of carboplatin treatment is followed around day 14 by the emergence of a mixed cell population consisting of cycling, cell cycle-arrested and senescent cells. At this stage, giant cells make up >80% of the cell population, p21 (CDKN1A) in strongly induced, and cell numbers remain nearly static. Subsequently, cell death decreases, p21 expression drops to a low level and cell divisions increase, including regular mitoses of giant cells and depolyploidization by multi-daughter divisions. These events are accompanied by the upregulation of stemness markers and a pro-inflammatory secretory phenotype, peaking after approximately 14 days of treatment. At the same time the cells initiate epithelial to mesenchymal transition, which over the subsequent weeks continuously increases, concomitantly with the emergence of highly proliferative, migratory, dedifferentiated, pro-inflammatory and chemoresistant cells (SKOV3-R). These cells are anchorage-independent and grow in a 3D collagen matrix, while cells on day 14 do not survive under these conditions, indicating that SKOV3-R cells were generated thereafter by the multi-stage process described above. This process was essentially recapitulated with the ovarian carcinoma cell line IGROV-1. Our observations suggest that transitory cells characterized by polyploidy, features of stemness and a pro-inflammatory secretory phenotype contribute to the acquisition of chemoresistance. PMID:26503466

  16. Cell growth density modulates cancer cell vascular invasion via Hippo pathway activity and CXCR2 signaling.

    PubMed

    Sharif, G M; Schmidt, M O; Yi, C; Hu, Z; Haddad, B R; Glasgow, E; Riegel, A T; Wellstein, A

    2015-11-26

    Metastasis of cancer cells involves multiple steps, including their dissociation from the primary tumor and invasion through the endothelial cell barrier to enter the circulation and finding their way to distant organ sites where they extravasate and establish metastatic lesions. Deficient contact inhibition is a hallmark of invasive cancer cells, yet surprisingly the vascular invasiveness of commonly studied cancer cell lines is regulated by the density at which cells are propagated in culture. Cells grown at high density were less effective at invading an endothelial monolayer than cells grown at low density. This phenotypic difference was also observed in a zebrafish model of vascular invasion of cancer cells after injection into the yolk sac and extravasation of cancer cells into tissues from the vasculature. The vascular invasive phenotypes were reversible. A kinome-wide RNA interference screen was used to identify drivers of vascular invasion by panning small hairpin RNA (shRNA) library-transduced noninvasive cancer cell populations on endothelial monolayers. The selection of invasive subpopulations showed enrichment of shRNAs targeting the large tumor suppressor 1 (LATS1) kinase that inhibits the activity of the transcriptional coactivator yes-associated protein (YAP) in the Hippo pathway. Depletion of LATS1 from noninvasive cancer cells restored the invasive phenotype. Complementary to this, inhibition or depletion of YAP inhibited invasion in vitro and in vivo. The vascular invasive phenotype was associated with a YAP-dependent upregulation of the cytokines IL6, IL8 and C-X-C motif ligand 1, 2 and 3. Antibody blockade of cytokine receptors inhibited invasion and confirmed that they are rate-limiting drivers that promote cancer cell vascular invasiveness and could provide therapeutic targets. PMID:25772246

  17. Hyperoxia Inhibits T Cell Activation in Mice

    NASA Astrophysics Data System (ADS)

    Hughes-Fulford, M.; Meissler, J.; Aguayo, E. T.; Globus, R.; Aguado, J.; Candelario, T.

    2013-02-01

    , spleens were removed and the splenocytes were isolated and kept as individual biological samples. We have also examined transcription factors (JASPAR) and pathways of the immune system to help us understand the mechanism of regulation. Results: Our recent mouse immunology experiment aboard STS-131 suggests that the early T cell immune response was inhibited in animals that have been exposed to spaceflight, even 24 hours after return to earth. Moreover, recent experiments in hyperoxic mice show that many of the same genes involved in early T cell activation were altered. Specifically, expression of IL-2Rα, Cxcl2, TNFα, FGF2, LTA and BCL2 genes are dysregulated in mice exposed to hyperoxia. Conclusions: If these hyperoxia-induced changes of gene expression in early T cell activation are additive to the changes seen in the microgravity of spaceflight, there could be an increased infection risk to EVA astronauts, which should be addressed prior to conducting a Mars or other long-term mission.

  18. Activation of 2',5'-oligoadenylate synthetase activity on induction of HL-60 leukemia cell differentiation.

    PubMed Central

    Schwartz, E L; Nilson, L A

    1989-01-01

    A 27-fold increase in 2',5'-oligoadenylate synthetase activity, an enzyme associated with the antiproliferative actions of interferon (IFN), was observed after treatment of HL-60 human leukemia cells with dimethyl sulfoxide (DMSO), an inducer of granulocytic differentiation of the cells. Enzyme activity was elevated after 24 h of exposure to DMSO, was maximal at 48 hours, and declined thereafter. A comparable increase was observed after treatment with 1 U of alpha interferon (IFN-alpha) per ml or 8 U of beta interferon (IFN-beta) per ml. Elevated levels of expression of other IFN-inducible genes, including type I histocompatibility antigen (HLA-B) mRNA and 2',5'-oligoadenylate phosphodiesterase activity, were also observed with DMSO treatment. DMSO-treated HL-60 cells had an increased amount of a 1.8-kilobase mRNA for oligoadenylate [oligo(A)] synthetase when compared with that of control cells; both DMSO- and IFN-treated HL-60 cells also expressed 1.6-, 3.4-, and 4.3-kilobase mRNA. The increase in both oligo(A) synthetase activity and mRNA levels was inhibited by polyclonal antiserum to human IFN-alpha; however, no IFN-alpha mRNA could be detected in the cells. Antiserum to IFN-beta or gamma interferon (IFN-gamma) had no effect on oligo(A) synthetase expression or activity nor was there any detectable IFN-beta 1 or IFN-beta 2 mRNA in the cells. The anti-IFN-alpha serum did not block the elevation of HLA-B mRNA in DMSO-treated cells. These observations suggest that the increased expression of oligo(A) synthetase in DMSO-treated cells may be mediated by the release of an IFN-alpha-like factor; however, the levels of any IFN-alpha mRNA produced in the cells were extremely low. Images PMID:2476665

  19. Embryonic Stem Cells Cultured in Microfluidic Chambers Take Control of Their Fate by Producing Endogenous Signals Including LIF.

    PubMed

    Guild, Joshua; Haque, Amranul; Gheibi, Pantea; Gao, Yandong; Son, Kyung Jin; Foster, Elena; Dumont, Sophie; Revzin, Alexander

    2016-06-01

    It is important to understand the role played by endogenous signals in shaping stem cell fate decisions to develop better culture systems and to improve understanding of development processes. In this study, we describe the behavior of mouse embryonic stem cells (mESCs) inside microfluidic chambers (microchambers) operated under conditions of minimal perfusion. mESCs inside microchambers formed colonies and expressed markers of pluripotency in the absence of feeders or pluripotency-inducing signals such as leukemia inhibitory factor (LIF), while mESCs in standard cultureware differentiated rapidly. In a series of experiments, we demonstrate that remarkable differences in stem cell phenotype are due to endogenous production of LIF and other growth factors brought upon by cultivation in confines of a microchamber in the absence of perfusion (dilution). At the protein level, mESCs produced ∼140 times more LIF inside microchambers than under standard culture conditions. In addition, we demonstrate that pluripotent phenotype of stem cells could be degraded by increasing the height (volume) of the microchamber. Furthermore, we show that inhibition of LIF in microchambers, via the JAK/STAT3 pathway, leads to preferential differentiation into mesoderm that is driven by bone morphogenetic protein (BMP)-4. Collectively, we demonstrate for the first time that it is possible to design a cell culture system where stem cell fate is controlled solely by the endogenous signals. Our study may help shift the paradigm of stem cell cultivation away from relying on expensive exogenous molecules such as growth factors and toward designing culture chambers for harnessing endogenous signals. Stem Cells 2016;34:1501-1512. PMID:26865369

  20. Elasticity of adherent active cells on a compliant substrate

    NASA Astrophysics Data System (ADS)

    Banerjee, Shiladitya; Mertz, Aaron F.; Dufresne, Eric R.; Marchetti, M. Cristina

    2012-02-01

    We present a continuum mechanical model of rigidity sensing by livings cells adhering to a compliant substrate. The cell or cell colony is modeled as an elastic active gel, adapting recently developed continuum theories of active viscoelastic fluids. The coupling to the substrate enters as a boundary condition that relates the cell's deformation field to local stress gradients. In the presence of activity, the substrate induces spatially inhomogeneous contractile stresses and deformations, with a power law dependence of the total traction forces on cell or colony size. This is in agreement with recent experiments on keratinocyte colonies adhered to fibronectin coated surfaces. In the presence of acto-myosin activity, the substrate also enhances the cell polarization, breaking the cell's front-rear symmetry. Maximal polarization is observed when the substrate stiffness matches that of the cell, in agreement with experiments on stem cells.

  1. Effects of Lactobacillus salivarius, Lactobacillus reuteri, and Pediococcus acidilactici on the nematode Caenorhabditis elegans include possible antitumor activity.

    PubMed

    Fasseas, Michael K; Fasseas, Costas; Mountzouris, Konstantinos C; Syntichaki, Popi

    2013-03-01

    This study examined the effects of three lactic acid bacteria (LAB) strains on the nematode Caenorhabditis elegans. Lactobacillus salivarius, Lactobacillus reuteri, and Pediococcus acidilactici were found to inhibit the development and growth of the worm. Compared to Escherichia coli used as the control, L. reuteri and P. acidilactici reduced the lifespan of wild-type and short-lived daf-16 worms. On the contrary, L. salivarius extended the lifespan of daf-16 worms when used live, but reduced it as UV-killed bacteria. The three LAB induced the expression of genes involved in pathogen response and inhibited the growth of tumor-like germ cells, without affecting DAF16 localization or increasing corpse cells. Our results suggest the possible use of C. elegans as a model for studying the antitumor attributes of LAB. The negative effects of these LAB strains on the nematode also indicate their potential use against parasitic nematodes. PMID:22923095

  2. MAIT cells are activated during human viral infections

    PubMed Central

    van Wilgenburg, Bonnie; Scherwitzl, Iris; Hutchinson, Edward C.; Leng, Tianqi; Kurioka, Ayako; Kulicke, Corinna; de Lara, Catherine; Cole, Suzanne; Vasanawathana, Sirijitt; Limpitikul, Wannee; Malasit, Prida; Young, Duncan; Denney, Laura; Barnes, Eleanor; Ball, Jonathan; Burgess, Gary; Cooke, Graham; Dillon, John; Gore, Charles; Foster, Graham; Guha, Neil; Halford, Rachel; Herath, Cham; Holmes, Chris; Howe, Anita; Hudson, Emma; Irving, William; Khakoo, Salim; Koletzki, Diana; Martin, Natasha; Mbisa, Tamyo; McKeating, Jane; McLauchlan, John; Miners, Alec; Murray, Andrea; Shaw, Peter; Simmonds, Peter; Spencer, Chris; Targett-Adams, Paul; Thomson, Emma; Vickerman, Peter; Zitzmann, Nicole; Moore, Michael D.; Fabris, Paolo; Giordani, Maria Teresa; Oo, Ye Htun; Laidlaw, Stephen M.; Dustin, Lynn B.; Ho, Ling-Pei; Thompson, Fiona M.; Ramamurthy, Narayan; Mongkolsapaya, Juthathip; Willberg, Christian B.; Screaton, Gavin R.; Klenerman, Paul

    2016-01-01

    Mucosal-associated invariant T (MAIT) cells are abundant in humans and recognize bacterial ligands. Here, we demonstrate that MAIT cells are also activated during human viral infections in vivo. MAIT cells activation was observed during infection with dengue virus, hepatitis C virus and influenza virus. This activation—driving cytokine release and Granzyme B upregulation—is TCR-independent but dependent on IL-18 in synergy with IL-12, IL-15 and/or interferon-α/β. IL-18 levels and MAIT cell activation correlate with disease severity in acute dengue infection. Furthermore, HCV treatment with interferon-α leads to specific MAIT cell activation in vivo in parallel with an enhanced therapeutic response. Moreover, TCR-independent activation of MAIT cells leads to a reduction of HCV replication in vitro mediated by IFN-γ. Together these data demonstrate MAIT cells are activated following viral infections, and suggest a potential role in both host defence and immunopathology. PMID:27337592

  3. Functionally Active Gap Junctions between Connexin 43-Positive Mesenchymal Stem Cells and Glioma Cells.

    PubMed

    Gabashvili, A N; Baklaushev, V P; Grinenko, N F; Levinskii, A B; Mel'nikov, P A; Cherepanov, S A; Chekhonin, V P

    2015-05-01

    The formation of functional gap junctions between mesenchymal stem cells and cells of low-grade rat glioma C6 cells was studied in in vitro experiments. Immunocytochemical analysis with antibodies to connexin 43 extracellular loop 2 showed that mesenchymal stem cells as well as C6 glioma cells express the main astroglial gap junction protein connexin 43. Analysis of migration activity showed that mesenchymal stem cells actively migrate towards C6 glioma cells. During co-culturing, mesenchymal stem cells and glioma C6 form functionally active gap junctions mediating the transport of cytoplasmic dye from glioma cells to mesenchymal stem cells in the opposite direction. Fluorometry showed that the intensity of transport of low-molecular substances through heterologous gap junctions between mesenchymal stem cells and glioma cells is similar to that through homologous gap junctions between glioma cells. This phenomenon can be used for the development of new methods of cell therapy of high-grade gliomas. PMID:26033611

  4. Activation-induced and damage-induced cell death in aging human T cells.

    PubMed

    Sikora, Ewa

    2015-11-01

    In multicellular organisms the proper system functionality is ensured by the balance between cell division, differentiation, senescence and death. This balance is changed during aging. Immunosenescence plays a crucial role in aging and leads to the shrinkage of T cell repertoire and the propensity to apoptosis. The elimination of expanded T cells at the end of immune response is crucial to maintain homeostasis and avoid any uncontrolled inflammation. Resting mature T lymphocytes, when activated via their antigen-specific receptor (TCR) and CD28 co-receptor, start to proliferate and then undergo the so called activation induced cell death (AICD), which mechanistically is triggered by the death receptor and leads to apoptosis. T lymphocytes, like other cells, are also exposed to damage, which can trigger the so called damage-induced cell death (DICD). It was hypothesized that oxidative stress and chronic antigenic load increasing with age reduced lymphocyte susceptibility to DICD and enhanced a proinflamatory status leading to increased AICD. However, data collected so far are inconsistent and does not support this assumption. Systematic and comprehensive studies are still needed for conclusive elucidation of the role of AICD and DICD in human immunosenescence, including the role of autophagy and necroptosis in the processes. PMID:25843236

  5. Evaluation of Cell Binding Activities of Leptospira ECM Adhesins

    PubMed Central

    Robbins, Gregory T.; Hahn, Beth L.; Evangelista, Karen V.; Padmore, Lavinia; Aranda, Patrick S.; Coburn, Jenifer

    2015-01-01

    Pathogenic spirochetes of the genus Leptospira are the causative agents of leptospirosis, a zoonotic infection that occurs globally. The bacteria colonize the renal proximal tubules of many animals and are shed in the urine. Contact with the urine, or with water contaminated with the urine of infected animals can cause infection of new host animals, including humans. Mechanisms of colonization of the proximal tubule and other tissues are not known, but specific interactions between bacterial adhesins and host substrates are likely to be critical in this process. Several extracellular matrix (ECM) adhesins have been previously identified, but more recently, it has been shown that Leptospira bind more efficiently to cells than ECM. In this work, recombinant forms of five putative Leptospira ECM adhesins, namely LipL32, Loa22, OmpL1, p31/LipL45, and LenA were evaluated for binding to cells as well as an expanded variety of ECM components. Reproducible and significant adhesin activity was demonstrated only for OmpL1, which bound to both mammalian cell lines tested and to glycosaminoglycans (GAGs). While determination of biologically significant bacterial adhesion activity will require generation of site-directed mutant strains, our results suggest that OmpL1 is a strong candidate for future evaluation regarding the roles of the adhesin activity of the protein during L. interrogans infection. PMID:25875373

  6. How-to-Do-It: Hands-on Activities that Relate Mendelian Genetics to Cell Division.

    ERIC Educational Resources Information Center

    McKean, Heather R.; Gibson, Linda S.

    1989-01-01

    Presented is an activity designed to connect Mendelian laws with the physical processes of cell division. Included are materials production, procedures and worksheets for the meiosis-mitosis game and a genetics game. (CW)

  7. Langerhans Cells Serve as Immunoregulatory Cells by Activating NKT Cells1

    PubMed Central

    Fukunaga, Atsushi; Khaskhely, Noor M.; Ma, Ying; Sreevidya, Coimbatore S.; Taguchi, Kumiko; Nishigori, Chikako; Ullrich, Stephen E.

    2010-01-01

    UV exposure alters the morphology and function of epidermal Langerhans cells, which plays a role in UV-induced immune suppression. It is generally believed that UV exposure triggers the migration of immature Langerhans cells (LC) from the skin to the draining lymph nodes, where they induce tolerance. However, because most of the previous studies employed in vitro UV-irradiated LC, the data generated may not adequately reflect what is happening in vivo. In this study we isolated migrating Langerhans cells from the lymph nodes of UV-irradiated mice and studied their function. We found prolonged LC survival in the lymph nodes of UV-irradiated mice. LC were necessary for UV-induced immune suppression because no immune suppression was observed in Langerhans cells-deficient mice. Transferring LC from UV-irradiated mice into normal recipient animals transferred immune suppression and induced tolerance. We found that LC co-localized with lymph node Natural Killer T (NKT) cells. No immune suppression was observed when LC were transferred from UV-irradiated mice into NKT cell-deficient mice. NKT cells isolated from the lymph nodes of UV-irradiated mice secreted significantly more IL-4 than NKT cells isolated from non-irradiated controls. Injecting the wild type mice with anti-IL-4 blocked the induction of immune suppression. Our findings indicate that UV exposure activates the migration of mature LC to the skin draining lymph nodes where they induce immune regulation in vivo by activating NKT cells. PMID:20844203

  8. Fuel cell integral bundle assembly including ceramic open end seal and vertical and horizontal thermal expansion control

    SciTech Connect

    Zafred, Paolo R.; Gillett, James E.

    2012-04-24

    A plurality of integral bundle assemblies contain a top portion with an inlet fuel plenum and a bottom portion containing a base support, the base supports a dense, ceramic air exhaust manifold having four supporting legs, the manifold is below and connects to air feed tubes located in a recuperator zone, the air feed tubes passing into the center of inverted, tubular, elongated, hollow electrically connected solid oxide fuel cells having an open end above a combustion zone into which the air feed tubes pass and a closed end near the inlet fuel plenum, where the open end of the fuel cells rest upon and within a separate combination ceramic seal and bundle support contained in a ceramic support casting, where at least one flexible cushion ceramic band seal located between the recuperator and fuel cells protects and controls horizontal thermal expansion, and where the fuel cells operate in the fuel cell mode and where the base support and bottom ceramic air exhaust manifolds carry from 85% to all of the weight of the generator.

  9. Chitosan-Based Film of Tyrothricin for Enhanced Antimicrobial Activity against Common Skin Pathogens Including Staphylococcus aureus.

    PubMed

    Han, Sang Duk; Sung, Hyun Jung; Lee, Ga Hyeon; Jun, Joon-Ho; Son, Miwon; Kang, Myung Joo

    2016-05-28

    Chitosan-based film-forming gel is regarded as a promising vehicle for topical delivery of antimicrobial agents to skin wounds, since it protects from microbial infection and the cationic polymer itself possesses antibacterial activity. In this study, possible synergistic interaction against common skin pathogens between the cationic polymer and tyrothricin (TRC), a cyclic polypeptide antibiotic, was investigated, by determining the concentration to inhibit 90% of bacterial isolates (MIC). The addition of the polysaccharide to TRC dramatically reduced the MIC values of TRC by 1/33 and 1/4 against both methicillin-resistant and methicillinsusceptible Staphylococcus aureus, respectively. The synergism of TRC and chitosan combination against both strains was demonstrated by the checkerboard method, with a fractional inhibitory concentration index below 0.5. Moreover, co-treatment of TRC and chitosan exhibited antibacterial activity against Pseudomonas aeruginosa, due to the antibacterial activity of chitosan, whereas TRC itself did not inhibit the gram-negative bacterial growth. These findings suggested that the use of chitosan-based film for topical delivery of TRC could be an alternative to improve TRC antimicrobial activity against strains that are abundant in skin wounds. PMID:26907760

  10. The Azorhizobium caulinodans nifA gene: identification of upstream-activating sequences including a new element, the 'anaerobox'.

    PubMed Central

    Nees, D W; Stein, P A; Ludwig, R A

    1988-01-01

    From nucleotide sequencing analyses, the A. caulinodans nifA gene seems to be under dual control by the Ntr (in response to available N) and Fnr (in response to available O2) transcriptional activation/repression systems. Because it fixes N2 in two contexts, the Ntr system might regulate A. caulinodans nif gene expression ex planta, while the Fnr system might similarly regulate in planta. As nifA upstream-activating elements, we have identified: (i) a gpNifA binding site allowing autogenous nifA regulation, (ii) an Ntr-dependent transcription start, presumably the target of gpNifA activation, and (iii) an "anaerobox" tetradecameric nucleotide sequence that is precisely conserved among O2 regulated enteric bacterial genes controlled by the gpFnr transcriptional activator. Because it is precisely positioned upstream of enteric bacterial transcriptional starts, the "anaerobox" sequence may constitute the gpFnr DNA binding site. If so, then a second, Ntr-independent nifA transcription start may exist. We have also deduced the A. caulinodans nifA open reading frame and have compared the gene product (gpNifA) with those of other N2-fixing organisms. These proteins exhibit strongly conserved motifs: (i) sites conserved among ATP-binding proteins, (ii) an interdomain linker region, and (iii) a C-terminal alpha-helix-turn-alpha-helix DNA binding site. PMID:3186446

  11. T cells conditioned with MDSC show an increased anti-tumor activity after adoptive T cell based immunotherapy

    PubMed Central

    Raber, Patrick L.; Sierra, Rosa A.; Thevenot, Paul T.; Shuzhong, Zhang; Wyczechowska, Dorota D.; Kumai, Takumi; Celis, Esteban; Rodriguez, Paulo C.

    2016-01-01

    The success of adoptive T cell-based immunotherapy (ACT) in cancer is limited in part by the accumulation of myeloid-derived suppressor cells (MDSC), which block several T cell functions, including T cell proliferation and the expression of various cytotoxic mediators. Paradoxically, the inhibition of CD8+ T cell differentiation into cytotoxic populations increased their efficacy after ACT into tumor-bearing hosts. Therefore, we aimed to test the impact of conditioning CD8+ T cells with MDSC on their differentiation potential and ACT efficacy. Our results indicate that MDSC impaired the progression of CD8+ T cells into effector populations, without altering their activation status, production of IL-2, or signaling through the T cell receptor. In addition, culture of CD8+ T cells with MDSC resulted in an increased ACT anti-tumor efficacy, which correlated with a higher frequency of the transferred T cells and elevated IFNγ production. Interestingly, activated CD62L+ CD8+ Tcells were responsible for the enhanced anti-tumor activity showed by MDSC-exposed T cells. Additional results showed a decreased protein synthesis rate and lower activity of the mammalian/mechanistic target of rapamycin (mTOR) in T cells conditioned with MDSC. Silencing of the negative mTOR regulator tuberous sclerosis complex-2 in T cells co-cultured with MDSC restored mTOR activity, but resulted in T cell apoptosis. These results indicate that conditioning of T cells with MDSC induces stress survival pathways mediated by a blunted mTOR signaling, which regulated T cell differentiation and ACT efficacy. Continuation of this research will enable the development of better strategies to increase ACT responses in cancer. PMID:27007050

  12. Differential activation of mitogen-activated protein kinases following high and low LET radiation in murine macrophage cell line.

    PubMed

    Narang, Himanshi; Bhat, Nagesh; Gupta, S K; Santra, S; Choudhary, R K; Kailash, S; Krishna, Malini

    2009-04-01

    Mitogen-activated protein kinases have been shown to respond to various stimuli including cytokines, mitogens and gamma irradiation, leading to cell proliferation, differentiation, or death. The duration of their activation determines the specificity of response to each stimulus in various cells. In this study, the crucial intracellular kinases, ERK, JNK, and p38 kinase involved in cell survival, death, or damage and repair were examined for their activity in RAW 264.7 cells at various time points after irradiation with 2 Gy doses of proton ions or X-rays. This is the first report that shows that the MAPK signaling induced after heavy ion or X-ray exposure is not the same. Unlike gamma irradiation, there was prolonged but marginal activation of prosurvival ERK pathway and significant activation of proapoptotic p38 pathway in response to high LET radiation. PMID:19112558

  13. Quality of life, reproduction and sexuality after stem cell transplantation with partially T-cell-depleted grafts and after conditioning with a regimen including total body irradiation.

    PubMed

    Claessens, J J M; Beerendonk, C C M; Schattenberg, A V M B

    2006-05-01

    Thirty-four men and 36 women (median age 43 and 45 years, respectively) underwent stem cell transplantation (SCT) for acute leukaemia in first complete remission or chronic myelogenous leukaemia in first chronic phase between 1981 and 2001 from HLA-identical siblings. The conditioning regimen included TBI and all grafts were partially depleted of T cells. Changes in quality of life (QOL), reproduction and sexuality were studied using a questionnaire, and the previously given information related to these problems was assessed. In addition, endocrine status was assessed and semen analysis was performed. After SCT, patients reported less energy (n=50) and a deterioration in the job situation (n=31). Patients experienced a negative change in sexual relations (n=41). Important problems of sexual dysfunction were vaginal dryness in women (n=19) and erectile dysfunction in men (n=16). None of the patients was fertile based on their gonadotrophin levels, sperm concentrations and reproductive outcomes. Women experienced climacteric symptoms (n=24). Quality of life was negatively influenced by these changes. One-fifth of the patients were not satisfied with the information given with regard to reproduction, premature menopause and sexual problems. PMID:16547485

  14. Activated Muscle Satellite Cells Chase Ghosts.

    PubMed

    Mourikis, Philippos; Relaix, Frédéric

    2016-02-01

    The in vivo behaviors of skeletal muscle stem cells, i.e., satellite cells, during homeostasis and after injury are poorly understood. In this issue of Cell Stem Cell, Webster et al. (2016) now perform a tour de force intravital microscopic analysis of this population, showing that "ghost fiber" remnants act as scaffolds to guide satellite cell divisions after injury. PMID:26849298

  15. A Simple Laboratory Exercise Illustrating Active Transport in Yeast Cells.

    ERIC Educational Resources Information Center

    Stambuk, Boris U.

    2000-01-01

    Describes a simple laboratory activity illustrating the chemiosmotic principles of active transport in yeast cells. Demonstrates the energy coupling mechanism of active a-glucoside uptake by Saccaromyces cerevisiae cells with a colorimetric transport assay using very simple equipment. (Contains 22 references.) (Author/YDS)

  16. Small Molecule Antagonizes Autoinhibition and Activates AMP-activated Protein Kinase in Cells*

    PubMed Central

    Pang, Tao; Zhang, Zhen-Shan; Gu, Min; Qiu, Bei-Ying; Yu, Li-Fang; Cao, Peng-Rong; Shao, Wei; Su, Ming-Bo; Li, Jing-Ya; Nan, Fa-Jun; Li, Jia

    2008-01-01

    AMP-activated protein kinase (AMPK) serves as an energy sensor and is considered a promising drug target for treatment of type II diabetes and obesity. A previous report has shown that mammalian AMPK α1 catalytic subunit including autoinhibitory domain was inactive. To test the hypothesis that small molecules can activate AMPK through antagonizing the autoinhibition in α subunits, we screened a chemical library with inactive human α1394 (α1, residues 1-394) and found a novel small-molecule activator, PT1, which dose-dependently activated AMPK α1394, α1335, α2398, and even heterotrimer α1β1γ1. Based on PT1-docked AMPK α1 subunit structure model and different mutations, we found PT1 might interact with Glu-96 and Lys-156 residues near the autoinhibitory domain and directly relieve autoinhibition. Further studies using L6 myotubes showed that the phosphorylation of AMPK and its downstream substrate, acetyl-CoA carboxylase, were dose-dependently and time-dependently increased by PT1 with-out an increase in cellular AMP:ATP ratio. Moreover, in HeLa cells deficient in LKB1, PT1 enhanced AMPK phosphorylation, which can be inhibited by the calcium/calmodulin-dependent protein kinase kinases inhibitor STO-609 and AMPK inhibitor compound C. PT1 also lowered hepatic lipid content in a dose-dependent manner through AMPK activation in HepG2 cells, and this effect was diminished by compound C. Taken together, these data indicate that this small-molecule activator may directly activate AMPK via antagonizing the autoinhibition in vitro and in cells. This compound highlights the effort to discover novel AMPK activators and can be a useful tool for elucidating the mechanism responsible for conformational change and autoinhibitory regulation of AMPK. PMID:18321858

  17. 2', 3'-cyclic nucleotide 3'-phosphodiesterase is expressed in dissociated rat cerebellar cells and included in the postsynaptic density fraction.

    PubMed

    Cho, Sun-Jung; Jung, Jae Seob; Jin, IngNyol; Moon, Il Soo

    2003-08-31

    We have shown by protein sequencing that the phosphotyrosine-containing 48 kDa protein band of the rat cerebellar postsynaptic density fraction (CBL-PSD) is 2', 3'-cyclic nucleotide 3'-phosphodiesterase 2 (CNP2). Immunoblot analysis indicated that both CNP1 and CNP2 isoforms are present in the CBL-PSD fraction, whereas there is little CNP2 in the forebrain (FB)-PSD fraction. Both isoforms in the CBL-PSD fraction were tyrosine-phosphorylated to a basal extent. They were efficiently dissociated from the complexes in the PSD fraction by salt, but not by non-ionic detergents such as n-octyl glucoside (OG) and Triton X-100. Immunocytochemistry of dissociated cerebellar cultures revealed patchy CNP staining in oligodendrocytes (OLs), Purkinje cells (PCs), and unidentified PSD95-positive cells, but no staining in granule cells (GCs). Our results indicate that both CNP1 and CNP2 are expressed in cerian populations of cerebellar cells in addition to OL, and that they are associated with complexes that are co-isolated with the PSD. PMID:14503857

  18. Fast serial analysis of active cholesterol at the plasma membrane in single cells.

    PubMed

    Tian, Chunxiu; Zhou, Junyu; Wu, Zeng-Qiang; Fang, Danjun; Jiang, Dechen

    2014-01-01

    Previously, our group has utilized the luminol electrochemiluminescence to analyze the active cholesterol at the plasma membrane in single cells by the exposure of one cell to a photomultiplier tube (PMT) through a pinhole. In this paper, fast analysis of active cholesterol at the plasma membrane in single cells was achieved by a multimicroelectrode array without the pinhole. Single cells were directly located on the microelectrodes using cell-sized microwell traps. A cycle of voltage was applied on the microelectrodes sequentially to induce a peak of luminescence from each microelectrode for the serial measurement of active membrane cholesterol. A minimal time of 1.60 s was determined for the analysis of one cell. The simulation and the experimental data exhibited a semisteady-state distribution of hydrogen peroxide on the microelectrode after the reaction of cholesterol oxidase with the membrane cholesterol, which supported the relative accuracy of the serial analysis. An eight-microelectrode array was demonstrated to analyze eight single cells in 22 s serially, including the channel switching time. The results from 64 single cells either activated by low ion strength buffer or the inhibition of intracellular acyl-coA/cholesterol acyltransferase (ACAT) revealed that most of the cells analyzed had the similar active membrane cholesterol, while few cells had more active cholesterol resulting in the cellular heterogeneity. The fast single-cell analysis platform developed will be potentially useful for the analysis of more molecules in single cells using proper oxidases. PMID:24328095

  19. The MRX Complex Ensures NHEJ Fidelity through Multiple Pathways Including Xrs2-FHA-Dependent Tel1 Activation.

    PubMed

    Iwasaki, Daichi; Hayashihara, Kayoko; Shima, Hiroki; Higashide, Mika; Terasawa, Masahiro; Gasser, Susan M; Shinohara, Miki

    2016-03-01

    Because DNA double-strand breaks (DSBs) are one of the most cytotoxic DNA lesions and often cause genomic instability, precise repair of DSBs is vital for the maintenance of genomic stability. Xrs2/Nbs1 is a multi-functional regulatory subunit of the Mre11-Rad50-Xrs2/Nbs1 (MRX/N) complex, and its function is critical for the primary step of DSB repair, whether by homologous recombination (HR) or non-homologous end joining. In human NBS1, mutations result truncation of the N-terminus region, which contains a forkhead-associated (FHA) domain, cause Nijmegen breakage syndrome. Here we show that the Xrs2 FHA domain of budding yeast is required both to suppress the imprecise repair of DSBs and to promote the robust activation of Tel1 in the DNA damage response pathway. The role of the Xrs2 FHA domain in Tel1 activation was independent of the Tel1-binding activity of the Xrs2 C terminus, which mediates Tel1 recruitment to DSB ends. Both the Xrs2 FHA domain and Tel1 were required for the timely removal of the Ku complex from DSB ends, which correlates with a reduced frequency of imprecise end-joining. Thus, the Xrs2 FHA domain and Tel1 kinase work in a coordinated manner to maintain DSB repair fidelity. PMID:26990569

  20. Age-related changes in trunk neuromuscular activation patterns during a controlled functional transfer task include amplitude and temporal synergies.

    PubMed

    Quirk, D Adam; Hubley-Kozey, Cheryl L

    2014-12-01

    While healthy aging is associated with physiological changes that can impair control of trunk motion, few studies examine how spinal muscle responses change with increasing age. This study examined whether older (over 65 years) compared to younger (20-45 years) adults had higher overall amplitude and altered temporal recruitment patterns of trunk musculature when performing a functional transfer task. Surface electromyograms from twelve bilateral trunk muscle (24) sites were analyzed using principal component analysis, extracting amplitude and temporal features (PCs) from electromyographic waveforms. Two PCs explained 96% of the waveform variance. Three factor ANOVA models tested main effects (group, muscle and reach) and interactions for PC scores. Significant (p<.0125) group interactions were found for all PC scores. Post hoc analysis revealed that relative to younger adults, older adults recruited higher agonist and antagonistic activity, demonstrated continuous activation levels in specific muscle sites despite changing external moments, and had altered temporal synergies within abdominal and back musculature. In summary both older and younger adults recruit highly organized activation patterns in response to changing external moments. Differences in temporal trunk musculature recruitment patterns suggest that older adults experience different dynamic spinal stiffness and loading compared to younger adults during a functional lifting task. PMID:25457424

  1. Mycoplasma pneumoniae induces cytotoxic activity in guinea pig bronchoalveolar cells

    SciTech Connect

    Kist, M.; Koester, H.; Bredt, W.

    1985-06-01

    Precultured guinea pig alveolar macrophages (AM) and freshly harvested alveolar cells (FHAC) activated by interaction with Mycoplasma pneumoniae were cytotoxic for xenogeneic /sup 75/selenomethionine-labeled tumor target cells. Phagocytosis of whole opsonized or nonopsonized M. pneumoniae cells was more effective in eliciting cytotoxicity than uptake of sonicated microorganisms. The addition of living mycoplasma cells to the assay system enhanced the cytotoxic effect considerably. Target cells were significantly more susceptible to the cytotoxic action of phagocytes if they were coated with mycoplasma antigen or cocultured together with M. pneumoniae. The activation of the phagocytes could be inhibited by 2-deoxy-D-glucose but not by antimicrobial substances suppressing mycoplasma protein synthesis. It was accompanied by /sup 51/Cr release without detectable signs of cell damage. The supernatants of activated cells were cytotoxic for approximately 24 h. Inhibition, release, and cytotoxic activity indicate the necessity of an intact metabolism of the effector cells and suggest a secretion of cytotoxic substances.

  2. Human Liver Stem Cells Suppress T-Cell Proliferation, NK Activity, and Dendritic Cell Differentiation

    PubMed Central

    Bruno, Stefania; Grange, Cristina; Tapparo, Marta; Pasquino, Chiara; Romagnoli, Renato; Dametto, Ennia; Amoroso, Antonio; Tetta, Ciro; Camussi, Giovanni

    2016-01-01

    Human liver stem cells (HLSCs) are a mesenchymal stromal cell-like population resident in the adult liver. Preclinical studies indicate that HLSCs could be a good candidate for cell therapy. The aim of the present study was to evaluate the immunogenicity and the immunomodulatory properties of HLSCs on T-lymphocytes, natural killer cells (NKs), and dendritic cells (DCs) in allogeneic experimental settings. We found that HLSCs inhibited T-cell proliferation by a mechanism independent of cell contact and dependent on the release of prostaglandin E2 (PGE2) and on indoleamine 2,3-dioxygenase activity. When compared with mesenchymal stromal cells (MSCs), HLSCs were more efficient in inhibiting T-cell proliferation. At variance with MSCs, HLSCs did not elicit NK degranulation. Moreover, HLSCs inhibited NK degranulation against K562, a NK-sensitive target, by a mechanism dependent on HLA-G release. When tested on DC generation from monocytes, HLSCs were found to impair DC differentiation and DCs ability to induce T-cell proliferation through PGE2. This study shows that HLSCs have immunomodulatory properties similar to MSCs, but, at variance with MSCs, they do not elicit a NK response. PMID:27127520

  3. Simultaneous BVI noise and vibration reduction in rotorcraft using actively-controlled flaps and including performance considerations

    NASA Astrophysics Data System (ADS)

    Patt, Daniel A.

    This work presents the development and application of an active control approach for reduction of both vibration and noise induced by helicopter rotor blade vortex interaction (BVI). Control is implemented through single or dual actively controlled flaps (ACFs) on each blade. Low-speed helicopter flight is prone to severe BVI, resulting in elevated vibration and noise levels. Existing research has suggested that when some form of active control is used to reduce vibration, noise will increase and vice versa. The present research achieves simultaneous reduction of noise and vibration, and also investigates the physical sources of the observed reduction. The initial portion of this work focused on developing a tool for simulating helicopter noise and vibrations in the BVI flight regime. A method for predicting compressible unsteady blade surface pressure distribution on rotor blades was developed and combined with an enhanced free-wake model and an acoustic prediction tool with provisions for blade flexibility. These elements were incorporated within an aeroelastic analysis featuring fully coupled flap-lag-torsional blade dynamics. Subsequently, control algorithms were developed that were effective for reducing noise and vibration even in the nonlinear BVI flight regime; saturation limits were incorporated constraining flap deflections to specified limits. The resulting simulation was also validated with a wide range of experimental data, achieving excellent correlation. Finally, a number of active control studies were performed. Multi-component vibration reductions of 40--80% could be achieved, while incurring a small noise penalty. Noise was reduced using an onboard feedback microphone; reductions of 4--10 dB on the advancing side were observed on a plane beneath the rotor when using dual flaps. Finally, simultaneous noise and vibration reduction was studied. A reduction of about 5 dB in noise on the advancing side combined with a 60% reduction in vibration was

  4. PTP1B triggers integrin-mediated repression of myosin activity and modulates cell contractility

    PubMed Central

    González Wusener, Ana E.; González, Ángela; Nakamura, Fumihiko; Arregui, Carlos O.

    2016-01-01

    ABSTRACT Cell contractility and migration by integrins depends on precise regulation of protein tyrosine kinase and Rho-family GTPase activities in specific spatiotemporal patterns. Here we show that protein tyrosine phosphatase PTP1B cooperates with β3 integrin to activate the Src/FAK signalling pathway which represses RhoA-myosin-dependent contractility. Using PTP1B null (KO) cells and PTP1B reconstituted (WT) cells, we determined that some early steps following cell adhesion to fibronectin and vitronectin occurred robustly in WT cells, including aggregation of β3 integrins and adaptor proteins, and activation of Src/FAK-dependent signalling at small puncta in a lamellipodium. However, these events were significantly impaired in KO cells. We established that cytoskeletal strain and cell contractility was highly enhanced at the periphery of KO cells compared to WT cells. Inhibition of the Src/FAK signalling pathway or expression of constitutive active RhoA in WT cells induced a KO cell phenotype. Conversely, expression of constitutive active Src or myosin inhibition in KO cells restored the WT phenotype. We propose that this novel function of PTP1B stimulates permissive conditions for adhesion and lamellipodium assembly at the protruding edge during cell spreading and migration. PMID:26700725

  5. Denbinobin induces apoptosis in human lung adenocarcinoma cells via Akt inactivation, Bad activation, and mitochondrial dysfunction.

    PubMed

    Kuo, Chen-Tzu; Hsu, Ming-Jen; Chen, Bing-Chang; Chen, Chien-Chih; Teng, Che-Ming; Pan, Shiow-Lin; Lin, Chien-Huang

    2008-02-28

    Increasing evidence demonstrated that denbinobin, isolated from Ephemerantha lonchophylla, exert cytotoxic effects in cancer cells. The purpose of this study was to investigate whether denbinobin induces apoptosis and the apoptotic mechanism of denbinobin in human lung adenocarcinoma cells (A549). Denbinobin (1-20microM) caused cell death in a concentration-dependent manner. Flow cytometric analysis and annexin V labeling demonstrated that denbinobin increased the percentage of apoptotic cells. A549 cells treated with denbinobin showed typical characteristics of apoptosis including morphological changes and DNA fragmentation. Denbinobin induced caspase 3 activation, and N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (zVAD-fmk), a broad-spectrum caspase inhibitor, prevented denbinobin-induced cell death. Denbinobin induced the loss of the mitochondrial membrane potential and the release of mitochondrial apoptotic proteins including cytochrome c, second mitochondria derived activator of caspase (Smac), and apoptosis-inducing factor (AIF). In addition, denbinobin-induced Bad activation was accompanied by the dissociation of Bad with 14-3-3 and the association of Bad with Bcl-xL. Furthermore, denbinobin induced Akt inactivation in a time-dependent manner. Transfection of A549 cells with both wild-type and constitutively active Akt significantly suppressed denbinobin-induced Bad activation and cell apoptosis. These results suggest that Akt inactivation, followed by Bad activation, mitochondrial dysfunction, caspase 3 activation, and AIF release, contributes to denbinobin-induced cell apoptosis. PMID:18262737

  6. TAE226, a Bis-Anilino Pyrimidine Compound, Inhibits the EGFR-Mutant Kinase Including T790M Mutant to Show Anti-Tumor Effect on EGFR-Mutant Non-Small Cell Lung Cancer Cells.

    PubMed

    Otani, Hiroki; Yamamoto, Hiromasa; Takaoka, Munenori; Sakaguchi, Masakiyo; Soh, Junichi; Jida, Masaru; Ueno, Tsuyoshi; Kubo, Takafumi; Asano, Hiroaki; Tsukuda, Kazunori; Kiura, Katsuyuki; Hatakeyama, Shinji; Kawahara, Eiji; Naomoto, Yoshio; Miyoshi, Shinichiro; Toyooka, Shinichi

    2015-01-01

    TAE226, a bis-anilino pyrimidine compound, has been developed as an inhibitor of focal adhesion kinase (FAK) and insulin-like growth factor-I receptor (IGF-IR). In this study, we investigated the effect of TAE226 on non-small-cell lung cancer (NSCLC), especially focusing on the EGFR mutational status. TAE226 was more effective against cells with mutant EGFR, including the T790M mutant, than against cells with wild-type one. TAE226 preferentially inhibited phospho-EGFR and its downstream signaling mediators in the cells with mutant EGFR than in those with wild-type one. Phosphorylation of FAK and IGF-IR was not inhibited at the concentration at which the proliferation of EGFR-mutant cells was inhibited. Results of the in vitro binding assay indicated significant differences in the affinity for TAE226 between the wild-type and L858R (or delE746_A750) mutant, and the reduced affinity of ATP to the L858R (or delE746_A750) mutant resulted in good responsiveness of the L858R (or delE746_A750) mutant cells to TAE226. Of interest, the L858R/T790M or delE746_A750/T790M mutant enhanced the binding affinity for TAE226 compared with the L858R or delE746_A750 mutant, resulting in the effectiveness of TAE226 against T790M mutant cells despite the T790M mutation restoring the ATP affinity for the mutant EGFR close to that for the wild-type. TAE226 also showed higher affinity of about 15-fold for the L858R/T790M mutant than for the wild-type one by kinetic interaction analysis. The anti-tumor effect against EGFR-mutant tumors including T790M mutation was confirmed in mouse models without any significant toxicity. In summary, we showed that TAE226 inhibited the activation of mutant EGFR and exhibited anti-proliferative activity against NSCLCs carrying EGFR mutations, including T790M mutation. PMID:26090892

  7. TAE226, a Bis-Anilino Pyrimidine Compound, Inhibits the EGFR-Mutant Kinase Including T790M Mutant to Show Anti-Tumor Effect on EGFR-Mutant Non-Small Cell Lung Cancer Cells

    PubMed Central

    Otani, Hiroki; Yamamoto, Hiromasa; Takaoka, Munenori; Sakaguchi, Masakiyo; Soh, Junichi; Jida, Masaru; Ueno, Tsuyoshi; Kubo, Takafumi; Asano, Hiroaki; Tsukuda, Kazunori; Kiura, Katsuyuki; Hatakeyama, Shinji; Kawahara, Eiji; Naomoto, Yoshio; Miyoshi, Shinichiro; Toyooka, Shinichi

    2015-01-01

    TAE226, a bis-anilino pyrimidine compound, has been developed as an inhibitor of focal adhesion kinase (FAK) and insulin-like growth factor-I receptor (IGF-IR). In this study, we investigated the effect of TAE226 on non-small-cell lung cancer (NSCLC), especially focusing on the EGFR mutational status. TAE226 was more effective against cells with mutant EGFR, including the T790M mutant, than against cells with wild-type one. TAE226 preferentially inhibited phospho-EGFR and its downstream signaling mediators in the cells with mutant EGFR than in those with wild-type one. Phosphorylation of FAK and IGF-IR was not inhibited at the concentration at which the proliferation of EGFR-mutant cells was inhibited. Results of the in vitro binding assay indicated significant differences in the affinity for TAE226 between the wild-type and L858R (or delE746_A750) mutant, and the reduced affinity of ATP to the L858R (or delE746_A750) mutant resulted in good responsiveness of the L858R (or delE746_A750) mutant cells to TAE226. Of interest, the L858R/T790M or delE746_A750/T790M mutant enhanced the binding affinity for TAE226 compared with the L858R or delE746_A750 mutant, resulting in the effectiveness of TAE226 against T790M mutant cells despite the T790M mutation restoring the ATP affinity for the mutant EGFR close to that for the wild-type. TAE226 also showed higher affinity of about 15-fold for the L858R/T790M mutant than for the wild-type one by kinetic interaction analysis. The anti-tumor effect against EGFR-mutant tumors including T790M mutation was confirmed in mouse models without any significant toxicity. In summary, we showed that TAE226 inhibited the activation of mutant EGFR and exhibited anti-proliferative activity against NSCLCs carrying EGFR mutations, including T790M mutation. PMID:26090892

  8. Radiation exposure induces inflammasome pathway activation in immune cells.

    PubMed

    Stoecklein, Veit M; Osuka, Akinori; Ishikawa, Shizu; Lederer, Madeline R; Wanke-Jellinek, Lorenz; Lederer, James A

    2015-02-01

    Radiation exposure induces cell and tissue damage, causing local and systemic inflammatory responses. Because the inflammasome pathway is triggered by cell death and danger-associated molecular patterns, we hypothesized that the inflammasome may signal acute and chronic immune responses to radiation. Using a mouse radiation model, we show that radiation induces a dose-dependent increase in inflammasome activation in macrophages, dendritic cells, NK cells, T cells, and B cells as judged by cleaved caspase-1 detection in cells. Time course analysis showed the appearance of cleaved caspase-1 in cells by day 1 and sustained expression until day 7 after radiation. Also, cells showing inflammasome activation coexpressed the cell surface apoptosis marker annexin V. The role of caspase-1 as a trigger for hematopoietic cell losses after radiation was studied in caspase-1(-/-) mice. We found less radiation-induced cell apoptosis and immune cell loss in caspase-1(-/-) mice than in control mice. Next, we tested whether uric acid might mediate inflammasome activation in cells by treating mice with allopurinol and discovered that allopurinol treatment completely blocked caspase-1 activation in cells. Finally, we demonstrate that radiation-induced caspase-1 activation occurs by a Nod-like receptor family protein 3-independent mechanism because radiation-exposed Nlrp3(-/-) mice showed caspase-1 activation profiles that were indistinguishable from those of wild-type mice. In summary, our data demonstrate that inflammasome activation occurs in many immune cell types following radiation exposure and that allopurinol prevented radiation-induced inflammasome activation. These results suggest that targeting the inflammasome may help control radiation-induced inflammation. PMID:25539818

  9. Ex Vivo Activity of Endoperoxide Antimalarials, Including Artemisone and Arterolane, against Multidrug-Resistant Plasmodium falciparum Isolates from Cambodia

    PubMed Central

    Chaorattanakawee, Suwanna; Lon, Chanthap; Saunders, David L.; Rutvisuttinunt, Wiriya; Yingyuen, Kritsanai; Bathurst, Ian; Ding, Xavier C.; Tyner, Stuart D.

    2014-01-01

    Novel synthetic endoperoxides are being evaluated as new components of artemisinin combination therapies (ACTs) to treat artemisinin-resistant Plasmodium falciparum malaria. We conducted blinded ex vivo activity testing of fully synthetic (OZ78 and OZ277) and semisynthetic (artemisone, artemiside, artesunate, and dihydroartemisinin) endoperoxides in the histidine-rich protein 2 enzyme-linked immunosorbent assay against 200 P. falciparum isolates from areas of artemisinin-resistant malaria in western and northern Cambodia in 2009 and 2010. The order of potency and geometric mean (GM) 50% inhibitory concentrations (IC50s) were as follows: artemisone (2.40 nM) > artesunate (8.49 nM) > dihydroartemisinin (11.26 nM) > artemiside (15.28 nM) > OZ277 (31.25 nM) > OZ78 (755.27 nM). Ex vivo activities of test endoperoxides positively correlated with dihydroartemisinin and artesunate. The isolates were over 2-fold less susceptible to dihydroartemisinin than the artemisinin-sensitive P. falciparum W2 clone and showed sensitivity comparable to those with test endoperoxides and artesunate, with isolate/W2 IC50 susceptibility ratios of <2.0. All isolates had P. falciparum chloroquine resistance transporter mutations, with negative correlations in sensitivity to endoperoxides and chloroquine. The activities of endoperoxides (artesunate, dihydroartemisinin, OZ277, and artemisone) significantly correlated with that of the ACT partner drug, mefloquine. Isolates had mutations associated with clinical resistance to mefloquine, with 35% prevalence of P. falciparum multidrug resistance gene 1 (pfmdr1) amplification and 84.5% occurrence of the pfmdr1 Y184F mutation. GM IC50s for mefloquine, lumefantrine, and endoperoxides (artesunate, dihydroartemisinin, OZ277, OZ78, and artemisone) correlated with pfmdr1 copy number. Given that current ACTs are failing potentially from reduced sensitivity to artemisinins and partner drugs, newly identified mutations associated with artemisinin resistance

  10. Bisphosphonamidate Clodronate Prodrug Exhibits Selective Cytotoxic Activity Against Melanoma Cell Lines

    PubMed Central

    Webster, Marie R.; Kamat, Chandrashekhar; Connis, Nick; Zhao, Ming; Weeraratna, Ashani T.; Rudek, Michelle A.; Hann, Christine L.; Freel Meyers, Caren L.

    2014-01-01

    Bisphosphonates are used clinically to treat disorders of calcium metabolism and malignant bone disease and are known to inhibit cancer cell growth, adhesion, and invasion. However, clinical use of these agents for the treatment of extraskeletal disease is limited due to low cell permeability. We recently described a bisphosphonamidate prodrug strategy for efficient intracellular release of bisphosphonates, including clodronate (CLO), in NSCLC cells. To evaluate anticancer activity of this prodrug class across many cancer cell types, the bisphosphonamidate clodronate prodrug (CLO prodrug) was screened against the NCI-60 cell line panel, and was found to exhibit selectivity toward melanoma cell lines. Here, we confirm efficient cellular uptake and intracellular activation of this prodrug class in melanoma cells. We further demonstrate inhibition of melanoma cell proliferation, induction of apoptosis, and an anti-tumor effect of CLO prodrug in a xenograft model. These data suggest a novel therapeutic application for the CLO prodrug and potential to selectively target melanoma cells. PMID:24310621

  11. AMP-Activated Protein Kinase Regulates the Cell Surface Proteome and Integrin Membrane Traffic

    PubMed Central

    Thavarajah, Thanusi; Medvedev, Sergei; Bowden, Peter; Marshall, John G.; Antonescu, Costin N.

    2015-01-01

    The cell surface proteome controls numerous cellular functions including cell migration and adhesion, intercellular communication and nutrient uptake. Cell surface proteins are controlled by acute changes in protein abundance at the plasma membrane through regulation of endocytosis and recycling (endomembrane traffic). Many cellular signals regulate endomembrane traffic, including metabolic signaling; however, the extent to which the cell surface proteome is controlled by acute regulation of endomembrane traffic under various conditions remains incompletely understood. AMP-activated protein kinase (AMPK) is a key metabolic sensor that is activated upon reduced cellular energy availability. AMPK activation alters the endomembrane traffic of a few specific proteins, as part of an adaptive response to increase energy intake and reduce energy expenditure. How increased AMPK activity during energy stress may globally regulate the cell surface proteome is not well understood. To study how AMPK may regulate the cell surface proteome, we used cell-impermeable biotinylation to selectively purify cell surface proteins under various conditions. Using ESI-MS/MS, we found that acute (90 min) treatment with the AMPK activator A-769662 elicits broad control of the cell surface abundance of diverse proteins. In particular, A-769662 treatment depleted from the cell surface proteins with functions in cell migration and adhesion. To complement our mass spectrometry results, we used other methods to show that A-769662 treatment results in impaired cell migration. Further, A-769662 treatment reduced the cell surface abundance of β1-integrin, a key cell migration protein, and AMPK gene silencing prevented this effect. While the control of the cell surface abundance of various proteins by A-769662 treatment was broad, it was also selective, as this treatment did not change the cell surface abundance of the transferrin receptor. Hence, the cell surface proteome is subject to acute

  12. Dengue Virus Directly Stimulates Polyclonal B Cell Activation

    PubMed Central

    Papa, Michelle Premazzi; de Morais, Ana Theresa Silveira; Peçanha, Ligia Maria Torres; de Arruda, Luciana Barros

    2015-01-01

    Dengue infection is associated to vigorous inflammatory response, to a high frequency of activated B cells, and to increased levels of circulating cross-reactive antibodies. We investigated whether direct infection of B cells would promote activation by culturing primary human B lymphocytes from healthy donors with DENV in vitro. B cells were susceptible, but poorly permissive to infection. Even though, primary B cells cultured with DENV induced substantial IgM secretion, which is a hallmark of polyclonal B cell activation. Notably, DENV induced the activation of B cells obtained from either DENV immune or DENV naïve donors, suggesting that it was not dependent on DENV-specific secondary/memory response. B cell stimulation was dependent on activation of MAPK and CD81. B cells cultured with DENV also secreted IL-6 and presented increased expression of CD86 and HLA-DR, which might contribute to B lymphocyte co-stimulatory function. Indeed, PBMCs, but not isolated B cells, secreted high amounts of IgG upon DENV culture, suggesting that interaction with other cell types in vivo might promote Ig isotype switching and IgG secretion from different B cell clones. These findings suggest that activation signaling pathways triggered by DENV interaction with non-specific receptors on B cells might contribute to the exacerbated response observed in dengue patients. PMID:26656738

  13. The Influence of Organized Physical Activity (including Gymnastics) on Young Adult Skeletal Traits: Is Maturity Phase Important?

    PubMed Central

    Bernardoni, Brittney; Scerpella, Tamara A.; Rosenbaum, Paula F.; Kanaley, Jill A.; Raab, Lindsay N.; Li, Quefeng; Wang, Sijian; Dowthwaite, Jodi N.

    2015-01-01

    We prospectively evaluated adolescent organized physical activity (PA) as a factor in adult female bone traits. Annual DXA scans accompanied semi-annual records of anthropometry, maturity and PA for 42 participants in this preliminary analysis (criteria: appropriately timed DXA scans at ~1 year pre-menarche [predictor] and ~5 years post-menarche [dependent variable]). Regression analysis evaluated total adolescent inter-scan PA and PA over 3 maturity sub-phases as predictors of young adult bone outcomes: 1) bone mineral content (BMC), geometry and strength indices at non-dominant distal radius and femoral neck; 2) sub-head BMC; 3) lumbar spine BMC. Analyses accounted for baseline gynecological age (years pre- or post-menarche), baseline bone status, adult body size and inter-scan body size change. Gymnastics training was evaluated as a potentially independent predictor, but did not improve models for any outcomes (p<0.07). Pre-menarcheal bone traits were strong predictors of most adult outcomes (semi-partial r2 = 0.21-0.59, p≤0.001). Adult 1/3 radius and sub-head BMC were predicted by both total PA and PA 1-3 years post-menarche (p<0.03). PA 3-5 years post-menarche predicted femoral narrow neck width, endosteal diameter and buckling ratio (p<0.05). Thus, participation in organized physical activity programs throughout middle and high school may reduce lifetime fracture risk in females. PMID:25386845

  14. The Influence of Organized Physical Activity (Including Gymnastics) on Young Adult Skeletal Traits: Is Maturity Phase Important?

    PubMed

    Bernardoni, Brittney; Scerpella, Tamara A; Rosenbaum, Paula F; Kanaley, Jill A; Raab, Lindsay N; Li, Quefeng; Wang, Sijian; Dowthwaite, Jodi N

    2015-05-01

    We prospectively evaluated adolescent organized physical activity (PA) as a factor in adult female bone traits. Annual DXA scans accompanied semiannual records of anthropometry, maturity, and PA for 42 participants in this preliminary analysis (criteria: appropriately timed DXA scans at ~1 year premenarche [predictor] and ~5 years postmenarche [dependent variable]). Regression analysis evaluated total adolescent interscan PA and PA over 3 maturity subphases as predictors of young adult bone outcomes: 1) bone mineral content (BMC), geometry, and strength indices at nondominant distal radius and femoral neck; 2) subhead BMC; 3) lumbar spine BMC. Analyses accounted for baseline gynecological age (years pre- or postmenarche), baseline bone status, adult body size and interscan body size change. Gymnastics training was evaluated as a potentially independent predictor, but did not improve models for any outcomes (p > .07). Premenarcheal bone traits were strong predictors of most adult outcomes (semipartial r2 = .21-0.59, p ≤ .001). Adult 1/3 radius and subhead BMC were predicted by both total PA and PA 1-3 years postmenarche (p < .03). PA 3-5 years postmenarche predicted femoral narrow neck width, endosteal diameter, and buckling ratio (p < .05). Thus, participation in organized physical activity programs throughout middle and high school may reduce lifetime fracture risk in females. PMID:25386845

  15. Shape control and compartmentalization in active colloidal cells.

    PubMed

    Spellings, Matthew; Engel, Michael; Klotsa, Daphne; Sabrina, Syeda; Drews, Aaron M; Nguyen, Nguyen H P; Bishop, Kyle J M; Glotzer, Sharon C

    2015-08-25

    Small autonomous machines like biological cells or soft robots can convert energy input into control of function and form. It is desired that this behavior emerges spontaneously and can be easily switched over time. For this purpose we introduce an active matter system that is loosely inspired by biology and which we term an active colloidal cell. The active colloidal cell consists of a boundary and a fluid interior, both of which are built from identical rotating spinners whose activity creates convective flows. Similarly to biological cell motility, which is driven by cytoskeletal components spread throughout the entire volume of the cell, active colloidal cells are characterized by highly distributed energy conversion. We demonstrate that we can control the shape of the active colloidal cell and drive compartmentalization by varying the details of the boundary (hard vs. flexible) and the character of the spinners (passive vs. active). We report buckling of the boundary controlled by the pattern of boundary activity, as well as formation of core-shell and inverted Janus phase-separated configurations within the active cell interior. As the cell size is increased, the inverted Janus configuration spontaneously breaks its mirror symmetry. The result is a bubble-crescent configuration, which alternates between two degenerate states over time and exhibits collective migration of the fluid along the boundary. Our results are obtained using microscopic, non-momentum-conserving Langevin dynamics simulations and verified via a phase-field continuum model coupled to a Navier-Stokes equation. PMID:26253763

  16. Shape control and compartmentalization in active colloidal cells

    PubMed Central

    Spellings, Matthew; Engel, Michael; Klotsa, Daphne; Sabrina, Syeda; Drews, Aaron M.; Nguyen, Nguyen H. P.; Bishop, Kyle J. M.; Glotzer, Sharon C.

    2015-01-01

    Small autonomous machines like biological cells or soft robots can convert energy input into control of function and form. It is desired that this behavior emerges spontaneously and can be easily switched over time. For this purpose we introduce an active matter system that is loosely inspired by biology and which we term an active colloidal cell. The active colloidal cell consists of a boundary and a fluid interior, both of which are built from identical rotating spinners whose activity creates convective flows. Similarly to biological cell motility, which is driven by cytoskeletal components spread throughout the entire volume of the cell, active colloidal cells are characterized by highly distributed energy conversion. We demonstrate that we can control the shape of the active colloidal cell and drive compartmentalization by varying the details of the boundary (hard vs. flexible) and the character of the spinners (passive vs. active). We report buckling of the boundary controlled by the pattern of boundary activity, as well as formation of core–shell and inverted Janus phase-separated configurations within the active cell interior. As the cell size is increased, the inverted Janus configuration spontaneously breaks its mirror symmetry. The result is a bubble–crescent configuration, which alternates between two degenerate states over time and exhibits collective migration of the fluid along the boundary. Our results are obtained using microscopic, non–momentum-conserving Langevin dynamics simulations and verified via a phase-field continuum model coupled to a Navier–Stokes equation. PMID:26253763

  17. Remote Control of T Cell Activation Using Magnetic Janus Particles.

    PubMed

    Lee, Kwahun; Yi, Yi; Yu, Yan

    2016-06-20

    We report a strategy for using magnetic Janus microparticles to control the stimulation of T cell signaling with single-cell precision. To achieve this, we designed Janus particles that are magnetically responsive on one hemisphere and stimulatory to T cells on the other side. By manipulating the rotation and locomotion of Janus particles under an external magnetic field, we could control the orientation of the particle-cell recognition and thereby the initiation of T cell activation. This study demonstrates a step towards employing anisotropic material properties of Janus particles to control single-cell activities without the need of complex magnetic manipulation devices. PMID:27144475

  18. Remote Activation of Host Cell DNA Synthesis in Uninfected Cells Signaled by Infected Cells in Advance of Virus Transmission

    PubMed Central

    Schmidt, Nora; Hennig, Thomas; Serwa, Remigiusz A.; Marchetti, Magda

    2015-01-01

    ABSTRACT Viruses modulate cellular processes and metabolism in diverse ways, but these are almost universally studied in the infected cell itself. Here, we study spatial organization of DNA synthesis during multiround transmission of herpes simplex virus (HSV) using pulse-labeling with ethynyl nucleotides and cycloaddition of azide fluorophores. We report a hitherto unknown and unexpected outcome of virus-host interaction. Consistent with the current understanding of the single-step growth cycle, HSV suppresses host DNA synthesis and promotes viral DNA synthesis in spatially segregated compartments within the cell. In striking contrast, during progressive rounds of infection initiated at a single cell, we observe that infection induces a clear and pronounced stimulation of cellular DNA replication in remote uninfected cells. This induced DNA synthesis was observed in hundreds of uninfected cells at the extended border, outside the perimeter of the progressing infection. Moreover, using pulse-chase analysis, we show that this activation is maintained, resulting in a propagating wave of host DNA synthesis continually in advance of infection. As the virus reaches and infects these activated cells, host DNA synthesis is then shut off and replaced with virus DNA synthesis. Using nonpropagating viruses or conditioned medium, we demonstrate a paracrine effector of uninfected cell DNA synthesis in remote cells continually in advance of infection. These findings have significant implications, likely with broad applicability, for our understanding of the ways in which virus infection manipulates cell processes not only in the infected cell itself but also now in remote uninfected cells, as well as of mechanisms governing host DNA synthesis. IMPORTANCE We show that during infection initiated by a single particle with progressive cell-cell virus transmission (i.e., the normal situation), HSV induces host DNA synthesis in uninfected cells, mediated by a virus-induced paracrine

  19. Rapid Feedforward Inhibition and Asynchronous Excitation Regulate Granule Cell Activity in the Mammalian Main Olfactory Bulb

    PubMed Central

    Burton, Shawn D.

    2015-01-01

    Granule cell-mediated inhibition is critical to patterning principal neuron activity in the olfactory bulb, and perturbation of synaptic input to granule cells significantly alters olfactory-guided behavior. Despite the critical role of granule cells in olfaction, little is known about how sensory input recruits granule cells. Here, we combined whole-cell patch-clamp electrophysiology in acute mouse olfactory bulb slices with biophysical multicompartmental modeling to investigate the synaptic basis of granule cell recruitment. Physiological activation of sensory afferents within single glomeruli evoked diverse modes of granule cell activity, including subthreshold depolarization, spikelets, and suprathreshold responses with widely distributed spike latencies. The generation of these diverse activity modes depended, in part, on the asynchronous time course of synaptic excitation onto granule cells, which lasted several hundred milliseconds. In addition to asynchronous excitation, each granule cell also received synchronous feedforward inhibition. This inhibition targeted both proximal somatodendritic and distal apical dendritic domains of granule cells, was reliably recruited across sniff rhythms, and scaled in strength with excitation as more glomeruli were activated. Feedforward inhibition onto granule cells originated from deep short-axon cells, which responded to glomerular activation with highly reliable, short-latency firing consistent with tufted cell-mediated excitation. Simulations showed that feedforward inhibition interacts with asynchronous excitation to broaden granule cell spike latency distributions and significantly attenuates granule cell depolarization within local subcellular compartments. Collectively, our results thus identify feedforward inhibition onto granule cells as a core feature of olfactory bulb circuitry and establish asynchronous excitation and feedforward inhibition as critical regulators of granule cell activity. SIGNIFICANCE

  20. The role of versican G3 domain in regulating breast cancer cell motility including effects on osteoblast cell growth and differentiation in vitro – evaluation towards understanding breast cancer cell bone metastasis

    PubMed Central

    2012-01-01

    Background Versican is detected in the interstitial tissues at the invasive margins of breast carcinoma, is predictive of relapse, and negatively impacts overall survival rates. The versican G3 domain is important in breast cancer cell growth, migration and bone metastasis. However, mechanistic studies evaluating versican G3 enhanced breast cancer bone metastasis are limited. Methods A versican G3 construct was exogenously expressed in the 66c14 and the MC3T3-E1 cell line. Cells were observed through light microscopy and viability analyzed by Coulter Counter or determined with colorimetric proliferation assays. The Annexin V-FITC apoptosis detection kit was used to detect apoptotic activity. Modified Chemotactic Boyden chamber migration invasion assays were applied to observe tumor migration and invasion to bone stromal cells and MC3T3-E1 cells. Alkaline phosphatase (ALP) staining and ALP ELISA assays were performed to observe ALP activity in MC3T3-E1 cells. Results In the four mouse breast cancer cell lines 67NR, 66c14, 4T07, and 4T1, 4T1 cells expressed higher levels of versican, and showed higher migration and invasion ability to MC3T3-E1 cells and primary bone stromal cells. 4T1 conditioned medium (CM) inhibited MC3T3-E1 cell growth, and even lead to apoptosis. Only 4T1 CM prevented MC3T3-E1 cell differentiation, noted by inhibition of alkaline phosphatase (ALP) activity. We exogenously expressed a versican G3 construct in a cell line that expresses low versican levels (66c14), and observed that the G3-expressing 66c14 cells showed enhanced cell migration and invasion to bone stromal and MC3T3-E1 cells. This observation was prevented by selective EGFR inhibitor AG1478, selective MEK inhibitor PD 98059, and selective AKT inhibitor Triciribine, but not by selective JNK inhibitor SP 600125. Versican G3 enhanced breast cancer cell invasion to bone stromal cells or osteoblast cells appears to occur through enhancing EGFR/ERK or AKT signaling. G3 expressing MC3T3-E1

  1. Rhomboid domain-containing protein 3 is a negative regulator of TLR3-triggered natural killer cell activation.

    PubMed

    Liu, Juan; Liu, Shuxun; Xia, Meng; Xu, Sheng; Wang, Chunmei; Bao, Yan; Jiang, Minghong; Wu, Yue; Xu, Tian; Cao, Xuetao

    2013-05-01

    Rhomboid domain-containing protein 3 (Rhbdd3), which belongs to a family of proteins with rhomboid domain, is widely expressed in immune cells; however, the roles of the Rhbdd members, including Rhbdd3, in immunity remain unknown. Natural killer (NK) cells are critical for host immune defense and also can mediate inflammatory diseases such as hepatitis. Although much is known about how NK cells are activated, the detailed mechanisms for negative regulation of NK cell activation remain to be fully understood. Using Rhbdd3-deficient mice, we reveal that Rhbdd3, selectively up-regulated in NK cells upon Toll-like receptor 3 (TLR3) stimulation, negatively regulates TLR3-mediated NK cell activation in a feedback manner. Rhbdd3 inhibits TLR3-triggered IFN-γ and granzyme B expression of NK cells in cell-cell contact dependence of accessory cells such as dendritic cells and Kupffer cells. Rhbdd3 interacts with DNAX activation protein of 12 kDa and promotes its degradation, inhibiting MAPK activation in TLR3-triggered NK cells. Furthermore, Rhbdd3 plays a critical role in attenuating TLR3-triggered acute inflammation by controlling NK cell activation and accumulation in liver and disrupting NK cell-Kupffer cell interaction. Therefore, Rhbdd3 is a feedback inhibitor of TLR3-triggered NK cell activation. Our study outlines a mechanism for the negative regulation of NK cell activation and also provides clues for the function of the rhomboid proteins in immunity. PMID:23610400

  2. Activation of B cells by antigens on follicular dendritic cells

    PubMed Central

    El Shikh, Mohey Eldin M.; El Sayed, Rania M.; Sukumar, Selvakumar; Szakal, Andras K.; Tew, John G.

    2010-01-01

    A need for antigen-processing and presentation to B cells is not widely appreciated. However, cross-linking of multiple B cell receptors (BCRs) by T-independent antigens delivers a potent signal that induces antibody responses. Such BCR cross-linking also occurs in germinal centers where follicular dendritic cells (FDCs) present multimerized antigens as periodically arranged antigen-antibody complexes (ICs). Unlike T cells that recognize antigens as peptide-MHC complexes, optimal B cell-responses are induced by multimerized FDC-ICs that simultaneously engage multiple BCRs. FDC-FcγRIIB mediates IC-periodicity and FDC-BAFF, -IL-6 and -C4bBP are co-stimulators. Remarkably, specific antibody responses can be induced by FDC-ICs in the absence of T cells, opening up the exciting possibility that people with T cell insufficiencies may be immunized with T-dependent vaccines via FDC-ICs. PMID:20418164

  3. Cell trapping in activated micropores for functional analysis.

    PubMed

    Talasaz, AmirAli H; Powell, Ashley A; Stahl, Patrik; Ronaghi, Mostafa; Jeffrey, Stefanie S; Mindrinos, Michael; Davis, Ronald W

    2006-01-01

    This paper presents a novel device which provides the opportunity to perform high-throughput biochemical assays on different individual cells. In particular, the proposed device is suited to screen the rare cells in biological samples for early stage cancer diagnosis and explore their biochemical functionality. In the process, single cells are precisely positioned and captured in activated micropores. To show the performance of the proposed device, cultured yeast cells and human epithelial circulating tumor cells are successfully captured. PMID:17945673

  4. IL-2 induces STAT4 activation in primary NK cells and NK cell lines, but not in T cells.

    PubMed

    Wang, K S; Ritz, J; Frank, D A

    1999-01-01

    IL-2 exerts potent but distinct functional effects on two critical cell populations of the immune system, T cells and NK cells. Whereas IL-2 leads to proliferation in both cell types, it enhances cytotoxicity primarily in NK cells. In both T cells and NK cells, IL-2 induces the activation of STAT1, STAT3, and STAT5. Given this similarity in intracellular signaling, the mechanism underlying the distinct response to IL-2 in T cells and NK cells is not clear. In this study, we show that in primary NK cells and NK cell lines, in addition to the activation of STAT1 and STAT5, IL-2 induces tyrosine phosphorylation of STAT4, a STAT previously reported to be activated only in response to IL-12 and IFN-alpha. This activation of STAT4 in response to IL-2 is not due to the autocrine production of IL-12 or IFN-alpha. STAT4 activated in response to IL-2 is able to bind to a STAT-binding DNA sequence, suggesting that in NK cells IL-2 is capable of activating target genes through phosphorylation of STAT4. IL-2 induces the activation of Jak2 uniquely in NK cells, which may underlie the ability of IL-2 to activate STAT4 only in these cells. Although the activation of STAT4 in response to IL-2 occurs in primary resting and activated NK cells, it does not occur in primary resting T cells or mitogen-activated T cells. The unique activation of the STAT4-signaling pathway in NK cells may underlie the distinct functional effect of IL-2 on this cell population. PMID:9886399

  5. Thermodynamic Activity Measurements with Knudsen Cell Mass Spectrometry

    NASA Technical Reports Server (NTRS)

    Copland, Evan H.; Jacobson, Nathan S.

    2001-01-01

    Coupling the Knudsen effusion method with mass spectrometry has proven to be one of the most useful experimental techniques for studying the equilibrium between condensed phases and complex vapors. The Knudsen effusion method involves placing a condensed sample in a Knudsen cell, a small "enclosure", that is uniformly heated and held until equilibrium is attained between the condensed and vapor phases. The vapor is continuously sampled by effusion through a small orifice in the cell. A molecular beam is formed from the effusing vapor and directed into a mass spectrometer for identification and pressure measurement of the species in the vapor phase. Knudsen cell mass spectrometry (KCMS) has been used for nearly fifty years now and continues to be a leading technique for obtaining thermodynamic data. Indeed, much of the well-established vapor specie data in the JANAF tables has been obtained from this technique. This is due to the extreme versatility of the technique. All classes of materials can be studied and all constituents of the vapor phase can be measured over a wide range of pressures (approximately 10(exp -4) to 10(exp -11) bar) and temperatures (500-2800 K). The ability to selectively measure different vapor species makes KCMS a very powerful tool for the measurement of component activities in metallic and ceramic solutions. Today several groups are applying KCMS to measure thermodynamic functions in multicomponent metallic and ceramic systems. Thermodynamic functions, especially component activities, are extremely important in the development of CALPHAD (Calculation of Phase Diagrams) type thermodynamic descriptions. These descriptions, in turn, are useful for modeling materials processing and predicting reactions such as oxide formation and fiber/matrix interactions. The leading experimental methods for measuring activities are the Galvanic cell or electro-motive force (EMF) technique and the KCMS technique. Each has specific advantages, depending on

  6. Optimization of protocols for derivation of mouse embryonic stem cell lines from refractory strains, including the non obese diabetic mouse.

    PubMed

    Davies, Timothy J; Fairchild, Paul J

    2012-07-01

    The derivation of pluripotent embryonic stem cells (ESCs) from a variety of genetic backgrounds remains a desirable objective in the generation of mice functionally deficient in genes of interest and the modeling of human disease. Nevertheless, disparity in the ease with which different strains of mice yield ESC lines has long been acknowledged. Indeed, the generation of bona fide ESCs from the non obese diabetic (NOD) mouse, a well-characterized model of human type I diabetes, has historically proved especially difficult to achieve. Here, we report the development of protocols for the derivation of novel ESC lines from C57Bl/6 mice based on the combined use of high concentrations of leukemia inhibitory factor and serum-replacement, which is equally applicable to fresh and cryo-preserved embryos. Further, we demonstrate the success of this approach using Balb/K and CBA/Ca mice, widely considered to be refractory strains. CBA/Ca ESCs contributed to the somatic germ layers of chimeras and displayed a very high competence at germline transmission. Importantly, we were able to use the same protocol for the derivation of ESC lines from nonpermissive NOD mice. These ESCs displayed a normal karyotype that was robustly stable during long-term culture, were capable of forming teratomas in vivo and germline competent chimeras after injection into recipient blastocysts. Further, these novel ESC lines efficiently formed embryoid bodies in vitro and could be directed in their differentiation along the dendritic cell lineage, thus illustrating their potential application to the generation of cell types of relevance to the pathogenesis of type I diabetes. PMID:21933027

  7. Lithium Iron Phosphate Cell Performance Evaluations for Lunar Extravehicular Activities

    NASA Technical Reports Server (NTRS)

    Reid, Concha

    2007-01-01

    Lithium-ion battery cells are being evaluated for their ability to provide primary power and energy storage for NASA s future Exploration missions. These missions include the Orion Crew Exploration Vehicle, the Ares Crew Launch Vehicle Upper Stage, Extravehicular Activities (EVA, the advanced space suit), the Lunar Surface Ascent Module (LSAM), and the Lunar Precursor and Robotic Program (LPRP), among others. Each of these missions will have different battery requirements. Some missions may require high specific energy and high energy density, while others may require high specific power, wide operating temperature ranges, or a combination of several of these attributes. EVA is one type of mission that presents particular challenges for today s existing power sources. The Portable Life Support System (PLSS) for the advanced Lunar surface suit will be carried on an astronaut s back during eight hour long sorties, requiring a lightweight power source. Lunar sorties are also expected to occur during varying environmental conditions, requiring a power source that can operate over a wide range of temperatures. Concepts for Lunar EVAs include a primary power source for the PLSS that can recharge rapidly. A power source that can charge quickly could enable a lighter weight system that can be recharged while an astronaut is taking a short break. Preliminary results of Al23 Ml 26650 lithium iron phosphate cell performance evaluations for an advanced Lunar surface space suit application are discussed in this paper. These cells exhibit excellent recharge rate capability, however, their specific energy and energy density is lower than typical lithium-ion cell chemistries. The cells were evaluated for their ability to provide primary power in a lightweight battery system while operating at multiple temperatures.

  8. Senescence of activated stellate cells limits liver fibrosis

    PubMed Central

    Krizhanovsky, Valery; Yon, Monica; Dickins, Ross A.; Hearn, Stephen; Simon, Janelle; Miething, Cornelius; Yee, Herman; Zender, Lars; Lowe, Scott W.

    2011-01-01

    Summary Cellular senescence acts as a potent mechanism of tumor suppression; however, its functional contribution to non-cancer pathologies has not been examined. Here we show that senescent cells accumulate in murine livers treated to produce fibrosis, a precursor pathology to cirrhosis. The senescent cells are derived primarily from activated hepatic stellate cells, which initially proliferate in response to liver damage and produce the extracellular matrix deposited in the fibrotic scar. In mice lacking key senescence regulators, stellate cells continue to proliferate, leading to excessive liver fibrosis. Furthermore, senescent activated stellate cells exhibit gene expression profile consistent with cell cycle exit, reduced secretion of extracellular matrix components, enhanced secretion of extracellular matrix degrading enzymes, and enhanced immune surveillance. Accordingly natural killer cells preferentially kill senescent activated stellate cells in vitro and in vivo, thereby facilitating the resolution of fibrosis. Therefore, the senescence program limits the fibrogenic response to acute tissue damage. PMID:18724938

  9. Mast cells and their activation in lung disease.

    PubMed

    Virk, Harvinder; Arthur, Greer; Bradding, Peter

    2016-08-01

    Mast cells and their activation contribute to lung health via innate and adaptive immune responses to respiratory pathogens. They are also involved in the normal response to tissue injury. However, mast cells are involved in disease processes characterized by inflammation and remodeling of tissue structure. In these diseases mast cells are often inappropriately and chronically activated. There is evidence for activation of mast cells contributing to the pathophysiology of asthma, pulmonary fibrosis, and pulmonary hypertension. They may also play a role in chronic obstructive pulmonary disease, acute respiratory distress syndrome, and lung cancer. The diverse mechanisms through which mast cells sense and interact with the external and internal microenvironment account for their role in these diseases. Newly discovered mechanisms of redistribution and interaction between mast cells, airway structural cells, and other inflammatory cells may offer novel therapeutic targets in these disease processes. PMID:26845625

  10. Enhancement of anti-leukemia activity of NK cells in vitro and in vivo by inhibition of leukemia cell-induced NK cell damage

    PubMed Central

    Arriga, Roberto; Caratelli, Sara; Coppola, Andrea; Spagnoli, Giulio Cesare; Venditti, Adriano; Amadori, Sergio; Lanzilli, Giulia; Lauro, Davide; Palomba, Patrizia; Sconocchia, Tommaso; Del Principe, Maria Ilaria; Maurillo, Luca; Buccisano, Francesco; Capuani, Barbara; Ferrone, Soldano; Sconocchia, Giuseppe

    2016-01-01

    Acute myeloid leukemia (AML) cells induce, in vitro, NK cell abnormalities (NKCAs) including apoptosis and activating receptor down-regulation. The potential negative impact of AML cells on the therapeutic efficacy of NK cell-based strategies prompted us to analyze the mechanisms underlying NKCAs and to develop approaches to protect NK cells from NKCAs. NKCA induction by the AML leukemia cells target a subpopulation of peripheral blood NK cells and is interleukin-2 independent but is abrogated by a long-term culture of NK (LTNK) cells at 37°C. LTNK cells displayed a significantly enhanced ability to damage AML cells in vitro and inhibited the subcutaneous growth of ML-2 cells grafted into CB17 SCID mice. Actinomycin D restored the susceptibility of LTNK cells to NKCAs while TAPI-0, a functional analog of the tissue inhibitor of metalloproteinase (TIMP) 3, inhibits ML-2 cell-induced NKCAs suggesting that the generation of NK cell resistance to NKCAs involves RNA transcription and metalloproteinase (MPP) inactivation. This conclusion is supported by the reduced susceptibility to AML cell-induced NKCAs of LTNK cells in which TIMP3 gene and protein are over-expressed. This information may contribute to the rational design of targeted strategies to enhance the efficacy of NK cell-based-immunotherapy of AML with haploidentical NK cells. PMID:26655503

  11. Cisplatin-induced Casepase-3 activation in different tumor cells

    NASA Astrophysics Data System (ADS)

    Shi, Hua; Li, Xiao; Su, Ting; Zhang, Yu-Hai

    2008-12-01

    Apoptosis plays an essential role in normal organism development which is one of the main types of programmed cell death to help tissues maintain homeostasis. Defective apoptosis can result in cell accumulation and therefore effects on tumor pathogenesis, progression and therapy resistance. A family of proteins, known as caspases, is typically activated in the early stages of apoptosis. Therefore, studying the kinetics of activation of caspases induced by antitumor drugs can contribute to antitumor drug discovery and explanation of the molecular mechanisms. This paper detected the Caspase-3 activity induced by cisplatin in human adenoid cystic carcinoma cell line (ACC-M), human hepatocellular liver carcinoma cell line (HepG2) and human epithelial carcinoma cell line (Hela) with stably expressing ECFP-DEVDDsRed (CD3) probe, a fluorescent probe consisting of Enhanced Cyan Fluorescent Protein (ECFP), red fluorescent protein (DsRed) and a linker with a recognition site of Caspase-3, by using the capillary electrophoresis (CE) and fluorescence resonance energy transfer (FRET) imaging system. Under the same concentration of cisplatin, ACC-M cells responded the most rapidly, and then HepG2 cells and Hela cells, respectively, in the early 30 hours. Later, HepG2 cells represented acceleration in the Caspase-3 activation speed and reached full activation the earliest comparing to other two cell types. The results demonstrated that ACC-M cell is more sensitive than the other two cell types under the treatment of cisplatin.

  12. Macrophages contribute to the cyclic activation of adult hair follicle stem cells.

    PubMed

    Castellana, Donatello; Paus, Ralf; Perez-Moreno, Mirna

    2014-12-01

    Skin epithelial stem cells operate within a complex signaling milieu that orchestrates their lifetime regenerative properties. The question of whether and how immune cells impact on these stem cells within their niche is not well understood. Here we show that skin-resident macrophages decrease in number because of apoptosis before the onset of epithelial hair follicle stem cell activation during the murine hair cycle. This process is linked to distinct gene expression, including Wnt transcription. Interestingly, by mimicking this event through the selective induction of macrophage apoptosis in early telogen, we identify a novel involvement of macrophages in stem cell activation in vivo. Importantly, the macrophage-specific pharmacological inhibition of Wnt production delays hair follicle growth. Thus, perifollicular macrophages contribute to the activation of skin epithelial stem cells as a novel, additional cue that regulates their regenerative activity. This finding may have translational implications for skin repair, inflammatory skin diseases and cancer. PMID:25536657

  13. Macrophages Contribute to the Cyclic Activation of Adult Hair Follicle Stem Cells

    PubMed Central

    Castellana, Donatello; Paus, Ralf; Perez-Moreno, Mirna

    2014-01-01

    Skin epithelial stem cells operate within a complex signaling milieu that orchestrates their lifetime regenerative properties. The question of whether and how immune cells impact on these stem cells within their niche is not well understood. Here we show that skin-resident macrophages decrease in number because of apoptosis before the onset of epithelial hair follicle stem cell activation during the murine hair cycle. This process is linked to distinct gene expression, including Wnt transcription. Interestingly, by mimicking this event through the selective induction of macrophage apoptosis in early telogen, we identify a novel involvement of macrophages in stem cell activation in vivo. Importantly, the macrophage-specific pharmacological inhibition of Wnt production delays hair follicle growth. Thus, perifollicular macrophages contribute to the activation of skin epithelial stem cells as a novel, additional cue that regulates their regenerative activity. This finding may have translational implications for skin repair, inflammatory skin diseases and cancer. PMID:25536657

  14. Stathmin Activity Influences Sarcoma Cell Shape, Motility, and Metastatic Potential

    PubMed Central

    Belletti, Barbara; Nicoloso, Milena S.; Schiappacassi, Monica; Berton, Stefania; Lovat, Francesca; Wolf, Katarina; Canzonieri, Vincenzo; D'Andrea, Sara; Zucchetto, Antonella; Friedl, Peter; Colombatti, Alfonso

    2008-01-01

    The balanced activity of microtubule-stabilizing and -destabilizing proteins determines the extent of microtubule dynamics, which is implicated in many cellular processes, including adhesion, migration, and morphology. Among the destabilizing proteins, stathmin is overexpressed in different human malignancies and has been recently linked to the regulation of cell motility. The observation that stathmin was overexpressed in human recurrent and metastatic sarcomas prompted us to investigate stathmin contribution to tumor local invasiveness and distant dissemination. We found that stathmin stimulated cell motility in and through the extracellular matrix (ECM) in vitro and increased the metastatic potential of sarcoma cells in vivo. On contact with the ECM, stathmin was negatively regulated by phosphorylation. Accordingly, a less phosphorylable stathmin point mutant impaired ECM-induced microtubule stabilization and conferred a higher invasive potential, inducing a rounded cell shape coupled with amoeboid-like motility in three-dimensional matrices. Our results indicate that stathmin plays a significant role in tumor metastasis formation, a finding that could lead to exploitation of stathmin as a target of new antimetastatic drugs. PMID:18305103

  15. Product and rate determinations with chemically activated nucleotides in the presence of various prebiotic materials, including other mono- and polynucleotides

    NASA Technical Reports Server (NTRS)

    Kanavarioti, A.; Alberas, D. J.; Rosenbach, M. T.; Bernasconi, C. F.; Chang, S.

    1991-01-01

    We are investigating the reactions of ImpN's in the presence of a number of prebiotically plausible materials, such as metal ions, phosphate, amines and other nucleotides and hope to learn more about the stability/reactivity of ImpN's in a prebiotic aqueous environment. We find that, in the presence of phosphate, ImpN's form substantial amounts of diphosphate nucleotides. These diphosphate nucleotides are not very good substrates for template directed reactions, but are chemically activated and are known to revert to the phosphoimidazolides in the presence of imidazole under solid state conditions. With respect to our studies of the oligomerization reaction, the determination of the dimerization rate constant of a specific ImpN (guanosine 5'-phospho 2 methylimidazolide) both in the absence and the presence of the template leads to the conclusion that at 37 C the dimerization is not template directed, although the subsequent polymerization steps are. In other words, this specific polynucleotide synthesizing system favors the elongation of oligonucleotides as compared with the formation of dimers and trimers. This favoring of the synthesis of long as opposed to short oligonucleotides may be regarded as a rudimentary example of natural selection at the molecular level.

  16. Immobilization of Pichia pastoris cells containing alcohol oxidase activity

    PubMed Central

    Maleknia, S; Ahmadi, H; Norouzian, D

    2011-01-01

    Background and Objectives The attempts were made to describe the development of a whole cell immobilization of P. pastoris by entrapping the cells in polyacrylamide gel beads. The alcohol oxidase activity of the whole cell Pichia pastoris was evaluated in comparison with yeast biomass production. Materials and Methods Methylotrophic yeast P. pastoris was obtained from Collection of Standard Microorganisms, Department of Bacterial Vaccines, Pasteur Institute of Iran (CSMPI). Stock culture was maintained on YPD agar plates. Alcohol oxidase was strongly induced by addition of 0.5% methanol as the carbon source. The cells were harvested by centrifugation then permeabilized. Finally the cells were immobilized in polyacrylamide gel beads. The activity of alcohol oxidase was determined by method of Tane et al. Results At the end of the logarithmic phase of cell culture, the alcohol oxidase activity of the whole cell P. Pastoris reached the highest level. In comparison, the alcohol oxidase activity was measured in an immobilized P. pastoris when entrapped in polyacrylamide gel beads. The alcohol oxidase activity of cells was induced by addition of 0.5% methanol as the carbon source. The cells were permeabilized by cetyltrimethylammonium bromide (CTAB) and immobilized. CTAB was also found to increase the gel permeability. Alcohol oxidase activity of immobilized cells was then quantitated by ABTS/POD spectrophotometric method at OD 420. There was a 14% increase in alcohol oxidase activity in immobilized cells as compared with free cells. By addition of 2-butanol as a substrate, the relative activity of alcohol oxidase was significantly higher as compared with other substrates added to the reaction media. Conclusion Immobilization of cells could eliminate lengthy and expensive procedures of enzyme separation and purification, protect and stabilize enzyme activity, and perform easy separation of the enzyme from the reaction media. PMID:22530090

  17. Gli1 Mediates Lung Cancer Cell Proliferation and Sonic Hedgehog-Dependent Mesenchymal Cell Activation

    PubMed Central

    Bermudez, Olga; Hennen, Elisabeth; Koch, Ina; Lindner, Michael; Eickelberg, Oliver

    2013-01-01

    Non-Small-Cell-Lung-Cancer (NSCLC) represents approximately 85% of all lung cancers and remains poorly understood. While signaling pathways operative during organ development, including Sonic Hedgehog (Shh) and associated Gli transcription factors (Gli1-3), have recently been found to be reactivated in NSCLC, their functional role remains unclear. Here, we hypothesized that Shh/Gli1-3 could mediate NSCLC autonomous proliferation and epithelial/stromal signaling in the tumoral tissue. In this context, we have investigated the activity of Shh/Gli1-3 signaling in NSCLC in both, cancer and stromal cells. We report here that inhibition of Shh signaling induces a significant decrease in the proliferation of NSCLC cells. This effect is mediated by Gli1 and Gli2, but not Gli3, through regulation of cyclin D1 and cyclin D2 expression. While exogenous Shh was unable to induce signaling in either A549 lung adenocarcinoma or H520 lung squamous carcinoma cells, both cells were found to secrete Shh ligand, which induced fibroblast proliferation, survival, migration, invasion, and collagen synthesis. Furthermore, Shh secreted by NSCLC mediates the production of proangiogenic and metastatic factors in lung fibroblasts. Our results thus provide evidence that Shh plays an important role in mediating epithelial/mesenchymal crosstalk in NSCLC. While autonomous Gli activity controls NSCLC proliferation, increased Shh expression by NSCLC is associated with fibroblast activation in tumor-associated stroma. Our study highlights the relevance of studying stromal-associated cells in the context of NSCLC regarding new prognosis and therapeutic options. PMID:23667589

  18. Gli1 mediates lung cancer cell proliferation and Sonic Hedgehog-dependent mesenchymal cell activation.

    PubMed

    Bermudez, Olga; Hennen, Elisabeth; Koch, Ina; Lindner, Michael; Eickelberg, Oliver

    2013-01-01

    Non-Small-Cell-Lung-Cancer (NSCLC) represents approximately 85% of all lung cancers and remains poorly understood. While signaling pathways operative during organ development, including Sonic Hedgehog (Shh) and associated Gli transcription factors (Gli1-3), have recently been found to be reactivated in NSCLC, their functional role remains unclear. Here, we hypothesized that Shh/Gli1-3 could mediate NSCLC autonomous proliferation and epithelial/stromal signaling in the tumoral tissue. In this context, we have investigated the activity of Shh/Gli1-3 signaling in NSCLC in both, cancer and stromal cells. We report here that inhibition of Shh signaling induces a significant decrease in the proliferation of NSCLC cells. This effect is mediated by Gli1 and Gli2, but not Gli3, through regulation of cyclin D1 and cyclin D2 expression. While exogenous Shh was unable to induce signaling in either A549 lung adenocarcinoma or H520 lung squamous carcinoma cells, both cells were found to secrete Shh ligand, which induced fibroblast proliferation, survival, migration, invasion, and collagen synthesis. Furthermore, Shh secreted by NSCLC mediates the production of proangiogenic and metastatic factors in lung fibroblasts. Our results thus provide evidence that Shh plays an important role in mediating epithelial/mesenchymal crosstalk in NSCLC. While autonomous Gli activity controls NSCLC proliferation, increased Shh expression by NSCLC is associated with fibroblast activation in tumor-associated stroma. Our study highlights the relevance of studying stromal-associated cells in the context of NSCLC regarding new prognosis and therapeutic options. PMID:23667589

  19. An adenoviral vector for probing promoter activity in primary immune cells

    PubMed Central

    Tripathi, Pulak; Madan, Rajat; Chougnet, Claire; Divanovic, Senad; Ma, Xiaojing; Wahl, Larry M.; Gajewski, Thomas; Karp, Christopher L.; Hildeman, David A.

    2010-01-01

    Functional analysis of the DNA regulatory regions that control gene expression has largely been performed through transient transfection of promoter–reporter constructs into transformed cells. However, transformed cells are often poor models of primary cells. To directly analyze DNA regulatory regions in primary cells, we generated a novel adenoviral luciferase reporter vector, pShuttle-luciferase-GFP (pSLUG) that contains a promoterless luciferase cassette (with an upstream cloning site) for probing promoter activity, and a GFP expression cassette that allows for the identification of transduced cells. Recombinant adenoviruses generated from this vector can transduce a wide range of primary immune cells with high efficiency, including human macrophages, dendritic cells and T cells; and mouse T cells transgenic for the coxsackie and adenoviral receptor (CAR). In primary T cells, we show inducible nuclear factor of activated T cells (NF-AT) activity using a recombinant pSLUG adenovirus containing a consensus NF-AT promoter. We further show inducible IL-12/23 p40 promoter activity in primary macrophages and dendritic cells using a recombinant pSLUG adenovirus containing the proximal human IL-12/23 p40 promoter. The pSLUG system promises to be a powerful tool for the analysis of DNA regulatory regions in diverse types of primary immune cells. PMID:16563424

  20. Two Unexpected Tumors in a Laparoscopic Nephrectomy Specimen, Including a Rare Tubulocystic Renal-Cell Carcinoma: A Case Report

    PubMed Central

    Jipp, Jacob; Defrain, Chad; Schwartz, Bradley

    2015-01-01

    Abstract We present a case of a 52-year-old Caucasian male who underwent a laparoscopic nephrectomy for an atrophic kidney and was found to have two unexpected, synchronous kidney cancers. He had a remote history of testicular cancer complicated by lymphadenopathy and external ureteral compression. Over time, he developed an atrophic left kidney from obstructive uropathy. Years later, due to flank pain and renal scintigraphy showing minimal function, a laparoscopic nephrectomy was performed. Final pathology demonstrated papillary renal-cell carcinoma (RCC) and tubulocystic RCC. Tubulocystic RCC is a rare neoplasm thought to be an indolent subset of collecting duct carcinoma, but was identified as a unique entity in 2004. Currently, there are ∼100 cases of this neoplasm in the literature.

  1. A constitutive model for the compressive response of metallic closed-cell foams including micro-inertia effects

    NASA Astrophysics Data System (ADS)

    Barthélémy, Romain; Jacques, Nicolas; Vermeersch, François; Kerampran, Steven

    2015-09-01

    Metallic foams have known a keen interest in the last decades. Their ability to undergo very large deformations while transmitting low stress levels make them capable of performing functions of protective layers against intense loadings and of energy absorbers, for instance. The behaviour of metal foams varies considerably between quasi-static and dynamic regimes. Those differences can be linked to the strain-rate sensitivity of the skeleton material and to micro-inertial effects (induced by the crushing of the foam cells). In the present work, a micromechanical model has been developed to take into account micro-inertia effects on the macroscopic behaviour of closed-cell foams under dynamic loading conditions. The proposed modelling is based on the dynamic homogenisation procedure introduced by Molinari and Mercier (J. Mech. Phys. Solids 49 (2001) 1497-1516). Within this framework, the macrostress is the sum of two terms. The first one is a static stress, that can be described with any existing model of metal foam. The second contribution is a dynamic stress related to micro-inertia effects. Considering an initially spherical shell as a Representative Volume Element (RVE) of the foam material, a closed-form expression of the dynamic stress was obtained. The proposed modelling was applied to shock propagation in aluminium foams (it should however be noted that the present theory is not restricted to uniaxial deformation but can be applied to arbitrary loadings). From experimental data of the literature, it is observed that incorporating micro-inertia effects allows one to achieve a better description of the foam shock response. This indicates that micro-inertia may have a significant influence on the dynamic behaviour of metallic foams.

  2. Managing misaligned paternity findings in research including sickle cell disease screening in Kenya: 'consulting communities' to inform policy.

    PubMed

    Marsh, Vicki; Kombe, Francis; Fitzpatrick, Ray; Molyneux, Sassy; Parker, Michael

    2013-11-01

    The management of misaligned paternity findings raises important controversy worldwide. It has mainly, however, been discussed in the context of high-income countries. Genetic and genomics research, with the potential to show misaligned paternity, are becoming increasingly common in Africa. During a genomics study in Kenya, a dilemma arose over testing and sharing information on paternal sickle cell disease status. This dilemma may be paradigmatic of challenges in sharing misaligned paternity findings in many research and health care settings. Using a deliberative approach to community consultation to inform research practice, we explored residents' views on paternal testing and sharing misaligned paternity information. Between December 2009 and November 2010, 63 residents in Kilifi County were engaged in informed deliberative small group discussions, structured to support normative reflection within the groups, with purposive selection to explore diversity. Analysis was based on a modified framework analysis approach, drawing on relevant social science and bioethics literature. The methods generated in-depth individual and group reflection on morally important issues and uncovered wide diversity in views and values. Fundamental and conflicting values emerged around the importance of family interests and openness, underpinned by disagreement on the moral implications of marital infidelity and withholding truth. Wider consideration of ethical issues emerging in these debates supports locally-held reasoning that paternal sickle cell testing should not be undertaken in this context, in contrast to views that testing should be done with or without the disclosure of misaligned paternity information. The findings highlight the importance of facilitating wider testing of family members of affected children, contingent on the development and implementation of national policies for the management of this inherited disorder. Their richness also illustrates the potential for

  3. Interacting active elastic dimers: Two cells moving on a rigid track

    NASA Astrophysics Data System (ADS)

    Das, Moumita; Mayett, David; Schwarz, J. M.

    2015-03-01

    Cell migration in morphogenesis and cancer metastasis typically involves an interplay between different cell types. The rules governing such interplay remain largely unknown, however, a recent experiment studying the interaction between neural crest (NC) cells and placodal cells reveals an example of such rules. The study found that NC cells chase the placodal cells by chemotaxis, while placodal cells run away from NC cells when contacted by them. Motivated by this observation, we construct and study a minimal one-dimensional cell-cell model comprised of two cells with each cell represented by two-beads-connected-by-an-active spring. The active spring for each moving cell models the stress fibers with their myosin-driven contractility (and alpha-actinin extendability), while the friction coefficients of the beads describe the catch/slip bond behavior of the integrins in focal adhesions. We also include a dynamic contact interaction between the two cells, as well as a chemotactic potential, to decipher the chase-and-run dynamics observed in the experiment. We then use our modeling to further generalize the rules governing the interplay between different cell types during collective cell migration.

  4. Variable susceptibility of ovarian cancer cells to non-thermal plasma-activated medium.

    PubMed

    Utsumi, Fumi; Kajiyama, Hiroaki; Nakamura, Kae; Tanaka, Hiromasa; Mizuno, Masaaki; Toyokuni, Shinnya; Hori, Masaru; Kikkawa, Fumitaka

    2016-06-01

    Non-thermal atmospheric pressure plasma has been widely studied in recent years in many fields, including cancer treatment. However, its efficiency for inducing apoptosis sometimes varies depending on the cell species and experimental conditions. The aim of this study was to elucidate what causes these differences in responses to plasma treatment. Using four ovarian cancer cell lines, the cell density had a markedly negative impact on the proliferation inhibition rate (PIR) and it was more obvious in OVCAR-3 and NOS2 cells. Furthermore, TOV21G and ES-2 cells were drastically sensitive to plasma‑activated medium (PAM) compared with the other two cell lines. We demonstrated that the proportion of reactive oxygen species and cell number had a marked impact on the effect of PAM against ovarian cancer cells. Additionally it was suggested that the morphological features of cells were also closely related to the sensitivity of cancer cells to the plasma treatment. PMID:27035127

  5. The role of Rac1 in the regulation of NF-kB activity, cell proliferation, and cell migration in non-small cell lung carcinoma

    PubMed Central

    Gastonguay, Adam; Berg, Tracy; Hauser, Andrew D.; Schuld, Nathan; Lorimer, Ellen; Williams, Carol L.

    2012-01-01

    The small GTPase Rac1 regulates many cellular processes, including cytoskeletal reorganization, cell migration, proliferation, and survival. Additionally, Rac1 plays a major role in activating NF-κB-mediated transcription. Both Rac1 and NF-κB regulate many properties of the malignant phenotype, including anchorage-independent proliferation and survival, metastasis, and angiogenesis. Despite these findings, the roles of Rac1and NF-κB in non-small cell lung carcinoma, a leading cause of cancer deaths, have not been thoroughly investigated. Here, we compared the effects of Rac1 siRNA to that of the Rac1 inhibitor NSC23766 on multiple features of the NSCLC malignant phenotype, including NF-κB activity. We show that the siRNA-mediated silencing of Rac1 in lung cancer cells results in decreased cell proliferation and migration. The decrease in proliferation was observed in both anchorage-dependent and anchorage-independent assays. Furthermore, cells with decreased Rac1 expression have a slowed progression through the G1 phase of the cell cycle. These effects induced by Rac1 siRNA correlated with a decrease in NF-κB transcriptional activity. Additionally, inhibition of NF-κB signaling with BAY 11–7082 inhibited proliferation; indicating that the loss of cell proliferation and migration induced by the silencing of Rac1 expression may be attributed in part to loss of NF-κB activity. Interestingly, treatment with the Rac1 inhibitor NSC23766 strongly inhibits cell proliferation, cell cycle progression, and NF-κB activity in lung cancer cells, to an even greater extent than the inhibition induced by Rac1 siRNA. These findings indicate that Rac1 plays an important role in lung cancer cell proliferation and migration, most likely through its ability to promote NF-κB activity, and highlight Rac1 pathways as therapeutic targets for the treatment of lung cancer. PMID:22549160

  6. Oligomeric Procyanidins Interfere with Glycolysis of Activated T Cells. A Novel Mechanism for Inhibition of T Cell Function.

    PubMed

    Goto, Masao; Wakagi, Manabu; Shoji, Toshihiko; Takano-Ishikawa, Yuko

    2015-01-01

    Procyanidins, which are flavonoids that are found in a variety of plant species, reduce or prevent immune disorders, such as allergy and autoimmune diseases, through an unknown mechanism. In the present study, we investigated the effects of procyanidins on the T cell receptor (TCR)-mediated responses of CD4⁺ T cells in vitro. Apple procyanidins strongly suppressed the proliferation of splenic CD4⁺ T cells that were stimulated by an anti-CD3ε antibody, as well as splenocytes stimulated by antigen, but did not alter interleukin (IL)-2 secretion from these cells. Furthermore, we found that oligomeric procyanidins strongly suppressed, in a degree of polymerization dependent manner, the proliferation of activated CD4⁺ T cells, as well as their production of effector cytokines, including glycolysis associated-cytokines, without affecting IL-2 secretion. Additionally, we investigated the inhibitory effects of oligomeric procyanidins on the glycolytic activity of activated CD4⁺ T cells. We show that pentameric procyanidin suppressed L-lactate production and glucose uptake in activated CD4⁺ T cells. These results suggest that oligomeric procyanidins suppress the functions of activated CD4⁺ T cells by interfering with glycolysis. PMID:26492229

  7. AUGMENTATION OF MURINE NATURAL KILLER CELL ACTIVITY BY MANGANESE CHLORIDE

    EPA Science Inventory

    Natural Killer (NK) cell activity of spleen cells from male CBA/J mice was augmented by a single parenteral injection of MnCl2 administered 1 day prior to testing by in vitro and in vivo isotope release assays. Increased cytotoxic activity was observed in vitro against both NK-se...

  8. Evaluation of the damage of cell wall and cell membrane for various extracellular polymeric substance extractions of activated sludge.

    PubMed

    Guo, Xuesong; Liu, Junxin; Xiao, Benyi

    2014-10-20

    Extracellular polymeric substances (EPS) are susceptible to contamination by intracellular substances released during the extraction of EPS owing to the damage caused to microbial cell structures. The damage to cell walls and cell membranes in nine EPS extraction processes of activated sludge was evaluated in this study. The extraction of EPS (including proteins, carbohydrates and DNA) was the highest using the NaOH extraction method and the lowest using formaldehyde extraction. All nine EPS extraction methods in this study resulted in cell wall and membrane damage. The damage to cell walls, evaluated by 2-keto-3-deoxyoctonate (KDO) and N-acetylglucosamine content changes in extracted EPS, was the most significant in the NaOH extraction process. Formaldehyde extraction showed a similar extent of damage to cell walls to those detected in the control method (centrifugation), while those in the formaldehyde-NaOH and cation exchange resin extractions were slightly higher than those detected in the control. N-acetylglucosamine was more suitable than KDO for the evaluation of cell wall damage in the EPS extraction of activated sludge. The damage to cell membranes was characterized by two fluorochromes (propidium iodide and FITC Annexin V) with flow cytometry (FCM) measurement. The highest proportion of membrane-damaged cells was detected in NaOH extraction (26.54% of total cells) while membrane-damaged cells comprised 8.19% of total cells in the control. PMID:25173614

  9. Transcriptional activity around bacterial cell death reveals molecular biomarkers for cell viability

    PubMed Central

    Kort, Remco; Keijser, Bart J; Caspers, Martien PM; Schuren, Frank H; Montijn, Roy

    2008-01-01

    Background In bacteriology, the ability to grow in selective media and to form colonies on nutrient agar plates is routinely used as a retrospective criterion for the detection of living bacteria. However, the utilization of indicators for bacterial viability-such as the presence of specific transcripts or membrane integrity-would overcome bias introduced by cultivation and reduces the time span of analysis from initiation to read out. Therefore, we investigated the correlation between transcriptional activity, membrane integrity and cultivation-based viability in the Gram-positive model bacterium Bacillus subtilis. Results We present microbiological, cytological and molecular analyses of the physiological response to lethal heat stress under accurately defined conditions through systematic sampling of bacteria from a single culture exposed to gradually increasing temperatures. We identified a coherent transcriptional program including known heat shock responses as well as the rapid expression of a small number of sporulation and competence genes, the latter only known to be active in the stationary growth phase. Conclusion The observed coordinated gene expression continued even after cell death, in other words after all bacteria permanently lost their ability to reproduce. Transcription of a very limited number of genes correlated with cell viability under the applied killing regime. The transcripts of the expressed genes in living bacteria – but silent in dead bacteria-include those of essential genes encoding chaperones of the protein folding machinery and can serve as molecular biomarkers for bacterial cell viability. PMID:19061518

  10. mTOR and metabolic pathways in T cell quiescence and functional activation

    PubMed Central

    Yang, Kai; Chi, Hongbo

    2013-01-01

    The mechanistic target of rapamycin (mTOR), an evolutionally conserved serine and threonine kinase, plays a critical role in the promotion of cell growth and proliferation via integration of cellular and environmental cues. In adaptive immunity, the mTOR pathway orchestrates multiple physiological processes including the development and homeostasis of T cells under steady state, and their subsequent activation and differentiation upon antigen recognition. Associated with such fate decisions is the dynamic reprogramming of T cell metabolic pathways, as naïve, activated and memory cells are defined by distinct bioenergetic and biosynthetic activities. Emerging evidence indicates that mTOR signaling intersects with T cell metabolism at two major levels to constitute a critical control mechanism of T cell fate decisions. First, as a central environmental sensor, mTOR links immune signaling and the availability of nutrients, especially amino acids. Second, mTOR activates specific metabolic pathways in T cells such as aerobic glycolysis (also known as the “Warburg effect”) in a process dependent upon the induction of transcription factors MYC and HIF1α. Understanding how mTOR interplays with T cell metabolism to dictate T cell fates and functions will provide fundamental insights into the mechanism of immune responses and the development of novel therapeutics against immune-mediated diseases. In this review, we summarize the current advances on mTOR signaling and T cell metabolism in the control of development, homeostasis, activation and differentiation of T cells. PMID:23375549

  11. CD8+CD122+CD49dlow regulatory T cells maintain T-cell homeostasis by killing activated T cells via Fas/FasL-mediated cytotoxicity.

    PubMed

    Akane, Kazuyuki; Kojima, Seiji; Mak, Tak W; Shiku, Hiroshi; Suzuki, Haruhiko

    2016-03-01

    The Fas/FasL (CD95/CD178) system is required for immune regulation; however, it is unclear in which cells, when, and where Fas/FasL molecules act in the immune system. We found that CD8(+)CD122(+) cells, which are mostly composed of memory T cells in comparison with naïve cells in the CD8(+)CD122(-) population, were previously shown to include cells with regulatory activity and could be separated into CD49d(low) cells and CD49d(high) cells. We established in vitro and in vivo experimental systems to evaluate the regulatory activity of CD122(+) cells. Regulatory activity was observed in CD8(+)CD122(+)CD49d(low) but not in CD8(+)CD122(+)CD49d(high) cells, indicating that the regulatory cells in the CD8(+)CD122(+) population could be narrowed down to CD49d(low) cells. CD8(+)CD122(-) cells taken from lymphoproliferation (lpr) mice were resistant to regulation by normal CD122(+) Tregs. CD122(+) Tregs taken from generalized lymphoproliferative disease (gld) mice did not regulate wild-type CD8(+)CD122(-) cells, indicating that the regulation by CD122(+) Tregs is Fas/FasL-dependent. CD122(+) Tregs taken from IL-10-deficient mice could regulate CD8(+)CD122(-) cells as equally as wild-type CD122(+) Tregs both in vitro and in vivo. MHC class I-missing T cells were not regulated by CD122(+) Tregs in vitro. CD122(+) Tregs also regulated CD4(+) cells in a Fas/FasL-dependent manner in vitro. These results suggest an essential role of Fas/FasL as a terminal effector of the CD122(+) Tregs that kill activated T cells to maintain immune homeostasis. PMID:26869716

  12. Pacemaker activity resulting from the coupling with nonexcitable cells

    NASA Astrophysics Data System (ADS)

    Jacquemet, Vincent

    2006-07-01

    Fibroblasts are nonexcitable cells that are sometimes coupled with excitable cells (cardiomyocytes). Due to a higher resting potential, these cells may act as a current source or sink and therefore disturb the electrical activity of the surrounding excitable cells. The possible occurrence of spontaneous pacemaker activity resulting from these electrotonic interactions was investigated in a theoretical model of two coupled cells as well as in a multicellular fiber model based on the Courtemanche kinetics. The results indicate that repeated spontaneous activations can be observed after an alteration in the activation and recovery properties of the sodium current (changes in excitability properties), provided that the difference in the resting potential as well as the coupling between the excitable and nonexcitable cells is sufficiently high. This may constitute a mechanism of focal sources triggering arrhythmias such as atrial fibrillation.

  13. Oncolytic Activity of a Recombinant Measles Virus, Blind to Signaling Lymphocyte Activation Molecule, Against Colorectal Cancer Cells

    PubMed Central

    Amagai, Yosuke; Fujiyuki, Tomoko; Yoneda, Misako; Shoji, Koichiro; Furukawa, Yoichi; Sato, Hiroki; Kai, Chieko

    2016-01-01

    Oncolytic virotherapy is a distinctive antitumor therapy based on the cancer-cell-specific infectivity and killing activity of viruses, which exert a considerable antitumor effect with only a few treatments. Because colorectal cancer cells often acquire resistance to the molecular-targeted therapies and alternative treatments are called for, in this study, we evaluated the oncolytic activity against colorectal cancer cells of a recombinant measles virus (rMV-SLAMblind), which is blind to signaling lymphocytic activation molecule (SLAM) and infects target cells via nectin-4/poliovirus receptor-related 4 protein. We examined 10 cell lines including 8 cell lines that were resistant to epidermal-growth-factor-receptor (EGFR) targeted therapy. rMV-SLAMblind infected and lysed the nectin-4-positive cell lines dependently on nectin-4 expression, in spite of mutation in EGFR cascade. Tumour progression in xenograft models was also abrogated by the virus, and the infection of cancer cells in vivo by the virus was demonstrated with both flow cytometry and a histological analysis. Therefore, rMV-SLAMblind is considered a novel therapeutic agent for colorectal cancers, including those resistant to molecular-targeted therapies. PMID:27090874

  14. Oncolytic Activity of a Recombinant Measles Virus, Blind to Signaling Lymphocyte Activation Molecule, Against Colorectal Cancer Cells.

    PubMed

    Amagai, Yosuke; Fujiyuki, Tomoko; Yoneda, Misako; Shoji, Koichiro; Furukawa, Yoichi; Sato, Hiroki; Kai, Chieko

    2016-01-01

    Oncolytic virotherapy is a distinctive antitumor therapy based on the cancer-cell-specific infectivity and killing activity of viruses, which exert a considerable antitumor effect with only a few treatments. Because colorectal cancer cells often acquire resistance to the molecular-targeted therapies and alternative treatments are called for, in this study, we evaluated the oncolytic activity against colorectal cancer cells of a recombinant measles virus (rMV-SLAMblind), which is blind to signaling lymphocytic activation molecule (SLAM) and infects target cells via nectin-4/poliovirus receptor-related 4 protein. We examined 10 cell lines including 8 cell lines that were resistant to epidermal-growth-factor-receptor (EGFR) targeted therapy. rMV-SLAMblind infected and lysed the nectin-4-positive cell lines dependently on nectin-4 expression, in spite of mutation in EGFR cascade. Tumour progression in xenograft models was also abrogated by the virus, and the infection of cancer cells in vivo by the virus was demonstrated with both flow cytometry and a histological analysis. Therefore, rMV-SLAMblind is considered a novel therapeutic agent for colorectal cancers, including those resistant to molecular-targeted therapies. PMID:27090874

  15. Activated mouse CD4(+)Foxp3(-) T cells facilitate melanoma metastasis via Qa-1-dependent suppression of NK-cell cytotoxicity.

    PubMed

    Wang, Xiaojuan; Cui, Yanyan; Luo, Gaoxing; Wang, Qinghong; Hu, Jie; He, Weifeng; Yuan, Jun; Zhou, Junyi; Wu, Yan; Sun, Xiaofeng; Robson, Simon C; Li, Xianchang; Tan, Jiangling; Peng, Yanmeng; Xue, Gang; Lu, Linrong; Gao, Wenda; Wu, Jun

    2012-12-01

    The regulatory activities of mouse CD4(+)Foxp3(+) T cells on various immune cells, including NK cells, have been well documented. Under some conditions, conventional CD4(+)Foxp3(-) T cells in the periphery are able to acquire inhibitory function on other T cells, but their roles in controlling innate immune cells are poorly defined. As a potential cellular therapy for cancer, ex vivo activated CD4(+)Foxp3(-) effector T cells are often infused back in vivo to suppress tumor growth and metastasis. Whether such activated T cells could affect NK-cell control of tumorigenesis is unclear. In the present study, we found that mitogen-activated CD4(+)Foxp3(-) T cells exhibited potent suppressor function on NK-cell proliferation and cytotoxicity in vitro, and notably facilitated B16 melanoma metastasis in vivo. Suppression of NK cells by activated CD4(+)Foxp3(-) T cells is cell-cell contact dependent and is mediated by Qa-1:NKG2A interaction, as administration of antibodies blocking either Qa-1 or NKG2A could completely reverse this suppression, and significantly inhibited otherwise facilitated melanoma metastasis. Moreover, activated CD4(+)Foxp3(-) cells from Qa-1 knockout mice completely lost the suppressor activity on NK cells, and failed to facilitate melanoma metastasis when transferred in vivo. Taken together, our findings indicate that innate anti-tumor response is counter regulated by the activation of adaptive immunity, a phenomenon we term as "activation-induced inhibition". Thus, the regulatory role of activated CD4(+)Foxp3(-) T cells in NK-cell activity must be taken into consideration in the future design of cancer therapies. PMID:22945357

  16. High water-stressed population estimated by world water resources assessment including human activities under SRES scenarios

    NASA Astrophysics Data System (ADS)

    Kiguchi, M.; Shen, Y.; Kanae, S.; Oki, T.

    2009-04-01

    In an argument of the reduction and the adaptation for the climate change, the evaluation of the influence by the climate change is important. When we argue in adaptation plan from a damage scale and balance with the cost, it is particularly important. Parry et al (2001) evaluated the risks in shortage of water, malaria, food, the risk of the coast flood by temperature function and clarified the level of critical climate change. According to their evaluation, the population to be affected by the shortage of water suddenly increases in the range where temperature increases from 1.5 to 2.0 degree in 2080s. They showed how much we need to reduce emissions in order to draw-down significantly the number at risk. This evaluation of critical climate change threats and targets of water shortage did not include the water withdrawal divided by water availability. Shen et al (2008a) estimated the water withdrawal of projection of future world water resources according to socio-economic driving factors predicted for scenarios A1b, A2, B1, and B2 of the Special Report on Emission Scenarios (SRES). However, these results were in function of not temperature but time. The assessment of the highly water-stressed population considered the socioeconomic development is necessary for a function of the temperature. Because of it is easy to understand to need to reduce emission. We present a multi-GCM analysis of the global and regional populations lived in highly water-stressed basin for a function of the temperature using the socioeconomic data and the outputs of GCMs. In scenario A2, the population increases gradually with warming. On the other hand, the future projection population in scenario A1b and B1 increase gradually until the temperature anomaly exceeds around from +1 to +1.5 degree. After that the population is almost constant. From Shen et al (2008b), we evaluated the HWSP and its ratio in the world with temperature function for scenarios A1B, A2, and B1 by the index of W

  17. Cancer cell-associated fatty acid synthase activates endothelial cells and promotes angiogenesis in colorectal cancer.

    PubMed

    Zaytseva, Yekaterina Y; Elliott, Victoria A; Rychahou, Piotr; Mustain, W Conan; Kim, Ji Tae; Valentino, Joseph; Gao, Tianyan; O'Connor, Kathleen L; Neltner, Janna M; Lee, Eun Y; Weiss, Heidi L; Evers, B Mark

    2014-06-01

    Upregulation of fatty acid synthase (FASN), a key enzyme of de novo lipogenesis, is associated with metastasis in colorectal cancer (CRC). However, the mechanisms of regulation are unknown. Since angiogenesis is crucial for metastasis, we investigated the role of FASN in the neovascularization of CRC. The effect of FASN on tumor vasculature was studied in orthotopic CRCs, the chick embryo chorioallantoic membrane (CAM) and Matrigel plug models using immunohistochemistry, immunofluorescent staining and confocal microscopy. Cell secretion was evaluated by ELISA and antibody arrays. Proliferation, migration and tubulogenesis of endothelial cells (ECs) were assessed in CRC-EC coculture models. In this study, we found that stable knockdown of FASN decreased microvessel density in HT29 and HCT116 orthotopic CRCs and resulted in 'normalization' of tumor vasculature in both orthotopic and CAM models. Furthermore, FASN regulated secretion of pro- and antiangiogenic factors, including vascular endothelial growth factor-A (VEGF-A). Mechanisms associated with the antiangiogenic activity noted with knockdown of FASN included: downregulation of VEGF(189), upregulation of antiangiogenic isoform VEGF(165b) and a decrease in expression and activity of matrix metalloproteinase-9. Furthermore, conditioned medium from FASN knockdown CRC cells inhibited activation of vascular endothelial growth factor receptor-2 and its downstream signaling and decreased proliferation, migration and tubulogenesis of ECs as compared with control medium. Together, these results suggest that cancer cell-associated FASN regulates tumor vasculature through alteration of the profile of secreted angiogenic factors and regulation of their bioavailability. Inhibition of FASN upstream of VEGF-A and other angiogenic pathways can be a novel therapeutic strategy to prevent or inhibit metastasis in CRC. PMID:24510238

  18. Cancer cell-associated fatty acid synthase activates endothelial cells and promotes angiogenesis in colorectal cancer

    PubMed Central

    Evers, B.Mark

    2014-01-01

    Upregulation of fatty acid synthase (FASN), a key enzyme of de novo lipogenesis, is associated with metastasis in colorectal cancer (CRC). However, the mechanisms of regulation are unknown. Since angiogenesis is crucial for metastasis, we investigated the role of FASN in the neovascularization of CRC. The effect of FASN on tumor vasculature was studied in orthotopic CRCs, the chick embryo chorioallantoic membrane (CAM) and Matrigel plug models using immunohistochemistry, immunofluorescent staining and confocal microscopy. Cell secretion was evaluated by ELISA and antibody arrays. Proliferation, migration and tubulogenesis of endothelial cells (ECs) were assessed in CRC–EC coculture models. In this study, we found that stable knockdown of FASN decreased microvessel density in HT29 and HCT116 orthotopic CRCs and resulted in ‘normalization’ of tumor vasculature in both orthotopic and CAM models. Furthermore, FASN regulated secretion of pro- and antiangiogenic factors, including vascular endothelial growth factor-A (VEGF-A). Mechanisms associated with the antiangiogenic activity noted with knockdown of FASN included: downregulation of VEGF189, upregulation of antiangiogenic isoform VEGF165b and a decrease in expression and activity of matrix metalloproteinase-9. Furthermore, conditioned medium from FASN knockdown CRC cells inhibited activation of vascular endothelial growth factor receptor-2 and its downstream signaling and decreased proliferation, migration and tubulogenesis of ECs as compared with control medium. Together, these results suggest that cancer cell-associated FASN regulates tumor vasculature through alteration of the profile of secreted angiogenic factors and regulation of their bioavailability. Inhibition of FASN upstream of VEGF-A and other angiogenic pathways can be a novel therapeutic strategy to prevent or inhibit metastasis in CRC. PMID:24510238

  19. GATA3 inhibits GCM1 activity and trophoblast cell invasion.

    PubMed

    Chiu, Yueh Ho; Chen, Hungwen

    2016-01-01

    Development of human placenta involves the invasion of trophoblast cells from anchoring villi into the maternal decidua. Placental transcription factor GCM1 regulates trophoblast cell invasion via transcriptional activation of HtrA4 gene, which encodes a serine protease enzyme. The GATA3 transcription factor regulates trophoblast cell differentiation and is highly expressed in invasive murine trophoblast giant cells. The regulation of trophoblastic invasion by GCM1 may involve novel cellular factors. Here we show that GATA3 interacts with GCM1 and inhibits its activity to suppress trophoblastic invasion. Immunohistochemistry demonstrates that GATA3 and GCM1 are coexpressed in villous cytotrophoblast cells, syncytiotrophoblast layer, and extravillous trophoblast cells of human placenta. Interestingly, GATA3 interacts with GCM1, but not the GCM2 homologue, through the DNA-binding domain and first transcriptional activation domain in GCM1 and the transcriptional activation domains and zinc finger 1 domain in GATA3. While GATA3 did not affect DNA-binding activity of GCM1, it suppressed transcriptional activity of GCM1 and therefore HtrA4 promoter activity. Correspondingly, GATA3 knockdown elevated HtrA4 expression in BeWo and JEG-3 trophoblast cell lines and enhanced the invasion activities of both lines. This study uncovered a new GATA3 function in placenta as a negative regulator of GCM1 activity and trophoblastic invasion. PMID:26899996

  20. GATA3 inhibits GCM1 activity and trophoblast cell invasion

    PubMed Central

    Chiu, Yueh Ho; Chen, Hungwen

    2016-01-01

    Development of human placenta involves the invasion of trophoblast cells from anchoring villi into the maternal decidua. Placental transcription factor GCM1 regulates trophoblast cell invasion via transcriptional activation of HtrA4 gene, which encodes a serine protease enzyme. The GATA3 transcription factor regulates trophoblast cell differentiation and is highly expressed in invasive murine trophoblast giant cells. The regulation of trophoblastic invasion by GCM1 may involve novel cellular factors. Here we show that GATA3 interacts with GCM1 and inhibits its activity to suppress trophoblastic invasion. Immunohistochemistry demonstrates that GATA3 and GCM1 are coexpressed in villous cytotrophoblast cells, syncytiotrophoblast layer, and extravillous trophoblast cells of human placenta. Interestingly, GATA3 interacts with GCM1, but not the GCM2 homologue, through the DNA-binding domain and first transcriptional activation domain in GCM1 and the transcriptional activation domains and zinc finger 1 domain in GATA3. While GATA3 did not affect DNA-binding activity of GCM1, it suppressed transcriptional activity of GCM1 and therefore HtrA4 promoter activity. Correspondingly, GATA3 knockdown elevated HtrA4 expression in BeWo and JEG-3 trophoblast cell lines and enhanced the invasion activities of both lines. This study uncovered a new GATA3 function in placenta as a negative regulator of GCM1 activity and trophoblastic invasion. PMID:26899996

  1. Fuel cell system including a unit for electrical isolation of a fuel cell stack from a manifold assembly and method therefor

    DOEpatents

    Kelley; Dana A. , Farooque; Mohammad , Davis; Keith

    2007-10-02

    A fuel cell system with improved electrical isolation having a fuel cell stack with a positive potential end and a negative potential, a manifold for use in coupling gases to and from a face of the fuel cell stack, an electrical isolating assembly for electrically isolating the manifold from the stack, and a unit for adjusting an electrical potential of the manifold such as to impede the flow of electrolyte from the stack across the isolating assembly.

  2. Anticancer activity of cryptotanshinone on acute lymphoblastic leukemia cells.

    PubMed

    Wu, Ching-Fen; Klauck, Sabine M; Efferth, Thomas

    2016-09-01

    Cryptotanshinone, a well-known diterpene quinone from a widely used traditional Chinese herb named Salvia miltiorrhiza, has been reported for its therapeutical potentials on diverse activities. In this study, pharmacological effects of cryptotanshinone on acute lymphoblastic leukemia cells were investigated. IC50 values of 5.0 and 4.8 were obtained in CEM/ADR5000 and CCRF-CEM. Microarray-based mRNA expression revealed that cryptotanshinone regulated genes associated with cell cycle, DNA damage, reactive oxygen species (ROS), NFκB signaling and cellular movement. The involvement of these pathways in the mode of action of cryptotanshinone was subsequently validated by additional independent in vitro studies. Cryptotanshinone stimulated ROS generation and induced DNA damage. It arrested cells in G2/M phase of the cell cycle and induced apoptosis as measured by annexin V-FITC-conjugating fluorescence. The induction of the intrinsic apoptotic pathway by cryptotanshinone was proved by loss of mitochondrial membrane potential and increased cleavage of caspase 3/7, caspase 9 and poly ADP ribose polymerase (PARP). DNA-binding motif analysis of the microarray-retrieved deregulated genes in the promoter region revealed NFκB as potential transcription factor involved in cryptotanshinone's mode of action. Molecular docking and Western blotting provided supportive evidence, suggesting that cryptotanshinone binds to IKK-β and inhibits the translocation of p65 from the cytosol to the nucleus. In addition, cryptotanshinone inhibited cellular movement as shown by a fibronectin-based cellular adhesion assay, indicating that this compound exerts anti-invasive features. In conclusion, cryptotanshinone exerts profound cytotoxicity, which is caused by multispecific modes of actions, including G2/M arrest, apoptosis and inhibition of cellular movement. The inhibitory activities of this compound may be explained by inhibition of NFκB, which orchestrates all these mechanisms. PMID

  3. Tenascin‐C Aggravates Autoimmune Myocarditis via Dendritic Cell Activation and Th17 Cell Differentiation

    PubMed Central

    Machino‐Ohtsuka, Tomoko; Tajiri, Kazuko; Kimura, Taizo; Sakai, Satoshi; Sato, Akira; Yoshida, Toshimichi; Hiroe, Michiaki; Yasutomi, Yasuhiro; Aonuma, Kazutaka; Imanaka‐Yoshida, Kyoko

    2014-01-01

    Background Tenascin‐C (TN‐C), an extracellular matrix glycoprotein, appears at several important steps of cardiac development in the embryo, but is sparse in the normal adult heart. TN‐C re‐expresses under pathological conditions including myocarditis, and is closely associated with tissue injury and inflammation in both experimental and clinical settings. However, the pathophysiological role of TN‐C in the development of myocarditis is not clear. We examined how TN‐C affects the initiation of experimental autoimmune myocarditis, immunologically. Methods and Results A model of experimental autoimmune myocarditis was established in BALB/c mice by immunization with murine α‐myosin heavy chains. We found that TN‐C knockout mice were protected from severe myocarditis compared to wild‐type mice. TN‐C induced synthesis of proinflammatory cytokines, including interleukin (IL)‐6, in dendritic cells via activation of a Toll‐like receptor 4, which led to T‐helper (Th)17 cell differentiation and exacerbated the myocardial inflammation. In the transfer experiment, dendritic cells loaded with cardiac myosin peptide acquired the functional capacity to induce myocarditis when stimulated with TN‐C; however, TN‐C‐stimulated dendritic cells generated from Toll‐like receptor 4 knockout mice did not induce myocarditis in recipients. Conclusions Our results demonstrated that TN‐C aggravates autoimmune myocarditis by driving the dendritic cell activation and Th17 differentiation via Toll‐like receptor 4. The blockade of Toll‐like receptor 4‐mediated signaling to inhibit the proinflammatory effects of TN‐C could be a promising therapeutic strategy against autoimmune myocarditis. PMID:25376187

  4. Signal transduction pathways in mast cell granule-mediated endothelial cell activation.

    PubMed Central

    Chi, Luqi; Stehno-Bittel, Lisa; Smirnova, Irina; Stechschulte, Daniel J; Dileepan, Kottarappat N

    2003-01-01

    BACKGROUND: We have previously shown that incubation of human endothelial cells with mast cell granules results in potentiation of lipopolysaccharide-induced production of interleukin-6 and interleukin-8. AIMS: The objective of the present study was to identify candidate molecules and signal transduction pathways involved in the synergy between mast cell granules and lipopolysaccharide on endothelial cell activation. METHODS: Human umbilical vein endothelial cells were incubated with rat mast cell granules in the presence and absence of lipopolysaccharide, and IL-6 production was quantified. The status of c-Jun amino-terminal kinase and extracellular signal-regulated kinase 1/2 activation, nuclear factor-kappaB translocation and intracellular calcium levels were determined to identify the mechanism of synergy between mast cell granules and lipopolysaccaride. RESULTS: Mast cell granules induced low levels of interleukin-6 production by endothelial cells, and this effect was markedly enhanced by lipopolysaccharide. The results revealed that both serine proteases and histamine present in mast cell granules were involved in this activation process. Mast cell granules increased intracellular calcium, and activated c-Jun amino-terminal kinase and extracellular signal-regulated kinase 1/2. The combination of lipopolysaccharide and mast cell granules prolonged c-Jun amino-terminal kinase activity beyond the duration of induction by either stimulant alone and was entirely due to active proteases. However, both proteases and histamine contributed to calcium mobilization and extracellular signal-regulated kinase 1/2 activation. The nuclear translocation of nuclear factor-kappaB proteins was of greater magnitude in endothelial cells treated with the combination of mast cell granules and lipopolysaccharide. CONCLUSIONS:Mast cell granule serine proteases and histamine can amplify lipopolysaccharide-induced endothelial cell activation, which involves calcium mobilization, mitogen-activated

  5. Imaging the coordination of multiple signaling activities in living cells

    PubMed Central

    Welch, Christopher M.; Elliott, Hunter; Danuser, Gaudenz; Hahn, Klaus M.

    2013-01-01

    Preface Cellular signal transduction occurs in complex and redundant interaction networks that are best examined at the level of single cells by simultaneously monitoring the activation dynamics of multiple components. Recent advances in biosensor technology have made it possible to visualize and quantify the activation of multiple network nodes in the same living cell. The precision and scope of this approach has been greatly extended by novel computational approaches to determine the relationships between different networks, studied in separate cells. PMID:22016058

  6. Effects of Neuroendocrine CB1 Activity on Adult Leydig Cells

    PubMed Central

    Cobellis, Gilda; Meccariello, Rosaria; Chianese, Rosanna; Chioccarelli, Teresa; Fasano, Silvia; Pierantoni, Riccardo

    2016-01-01

    Endocannabinoids control male reproduction acting at central and local level via cannabinoid receptors. The cannabinoid receptor CB1 has been characterized in the testis, in somatic and germ cells of mammalian and non-mammalian animal models, and its activity related to Leydig cell differentiation, steroidogenesis, spermiogenesis, sperm quality, and maturation. In this short review, we provide a summary of the insights concerning neuroendocrine CB1 activity in male reproduction focusing on adult Leydig cell ontogenesis and steroid biosynthesis. PMID:27375550

  7. Effects of Neuroendocrine CB1 Activity on Adult Leydig Cells.

    PubMed

    Cobellis, Gilda; Meccariello, Rosaria; Chianese, Rosanna; Chioccarelli, Teresa; Fasano, Silvia; Pierantoni, Riccardo

    2016-01-01

    Endocannabinoids control male reproduction acting at central and local level via cannabinoid receptors. The cannabinoid receptor CB1 has been characterized in the testis, in somatic and germ cells of mammalian and non-mammalian animal models, and its activity related to Leydig cell differentiation, steroidogenesis, spermiogenesis, sperm quality, and maturation. In this short review, we provide a summary of the insights concerning neuroendocrine CB1 activity in male reproduction focusing on adult Leydig cell ontogenesis and steroid biosynthesis. PMID:27375550

  8. Microwave-induced thermogenetic activation of single cells

    SciTech Connect

    Safronov, N. A.; Fedotov, I. V.; Ermakova, Yu. G.; Matlashov, M. E.; Belousov, V. V.; Sidorov-Biryukov, D. A.; Fedotov, A. B.; Zheltikov, A. M.

    2015-04-20

    Exposure to a microwave field is shown to enable thermogenetic activation of individual cells in a culture of cell expressing thermosensitive ion channels. Integration of a microwave transmission line with an optical fiber and a diamond quantum thermometer has been shown to allow thermogenetic single-cell activation to be combined with accurate local online temperature measurements based on an optical detection of electron spin resonance in nitrogen–vacancy centers in diamond.

  9. Tumor cell-specific bioluminescence platform to identify stroma-induced changes to anticancer drug activity.

    PubMed

    McMillin, Douglas W; Delmore, Jake; Weisberg, Ellen; Negri, Joseph M; Geer, D Corey; Klippel, Steffen; Mitsiades, Nicholas; Schlossman, Robert L; Munshi, Nikhil C; Kung, Andrew L; Griffin, James D; Richardson, Paul G; Anderson, Kenneth C; Mitsiades, Constantine S

    2010-04-01

    Conventional anticancer drug screening is typically performed in the absence of accessory cells of the tumor microenvironment, which can profoundly alter antitumor drug activity. To address this limitation, we developed the tumor cell-specific in vitro bioluminescence imaging (CS-BLI) assay. Tumor cells (for example, myeloma, leukemia and solid tumors) stably expressing luciferase are cultured with nonmalignant accessory cells (for example, stromal cells) for selective quantification of tumor cell viability, in presence versus absence of stromal cells or drug treatment. CS-BLI is high-throughput scalable and identifies stroma-induced chemoresistance in diverse malignancies, including imatinib resistance in leukemic cells. A stroma-induced signature in tumor cells correlates with adverse clinical prognosis and includes signatures for activated Akt, Ras, NF-kappaB, HIF-1alpha, myc, hTERT and IRF4; for biological aggressiveness; and for self-renewal. Unlike conventional screening, CS-BLI can also identify agents with increased activity against tumor cells interacting with stroma. One such compound, reversine, shows more potent activity in an orthotopic model of diffuse myeloma bone lesions than in conventional subcutaneous xenografts. Use of CS-BLI, therefore, enables refined screening of candidate anticancer agents to enrich preclinical pipelines with potential therapeutics that overcome stroma-mediated drug resistance and can act in a synthetic lethal manner in the context of tumor-stroma interactions. PMID:20228816

  10. Differential thioredoxin reductase activity from human normal hepatic and hepatoma cell lines.

    PubMed

    Jung, Haeng-Im; Lim, Hye-Won; Kim, Byung-Chul; Park, Eun-Hee; Lim, Chang-Jin

    2004-04-30

    Thioredoxin reductase (TrxR), a component of the thioredoxin system, including thioredoxin (Trx) and NADPH, catalyzes the transfer of electrons from NADPH to Trx, acts as a reductant of disulfide-containing proteins and participates in the defense system against oxidative stresses. In this study, the regulation pattern of TrxR in the presence of various stressful reagents was compared between Chang (human normal hepatic cell) and HepG2 (human hepatoma cell) cell lines. Aluminum chloride (0.5 mM) and zinc chloride (0.5 mM) enhanced the TrxR activity in the Chang cell line to a higher degree than in the HepG2 cell line, but cupric chloride (0.2 mM) and cadmium chloride (0.1 mM) enhanced the TrxR activity in the HepG2 cell line to a greater degree. The TrxR activities in both Chang and HepG2 cell lines were similarly induced by treatment with sodium selenite (0.02 mM) and menadione (0.5 and 1.0 mM). Lipopolysaccharide (2 micro g/m1) increased the TrxR activity upto 4.02- and 2.2-fold in the Chang and HepG2 cell lines, respectively, in time-dependent manners. Hydrogen peroxide (5 mM) markedly enhanced the TrxR activity in the HepG2 cell line, but not in the Chang cell line. NO-generating sodium nitroprusside (3.0 and 6.0 mM) induced TrxR activities in both human liver cell lines. The TrxR activity was also induced in human liver cells under limited growth conditions by serum deprivation. These results imply that the TrxR activities in normal hepatic and hepatoma cell lines are subject to different regulatory responses to various stresses. PMID:15118998

  11. Effects of dexamethasone on palate mesenchymal cell phospholipase activity

    SciTech Connect

    Bulleit, R.F.; Zimmerman, E.F.

    1984-09-15

    Corticosteroids will induce cleft palate in mice. One suggested mechanism for this effect is through inhibition of phospholipase activity. This hypothesis was tested by measuring the effects of dexamethasone, a synthetic corticosteroid, on phospholipase activity in cultures of palate mesenchymal cells. Palate mesenchymal cells were prelabeled with (3H)arachidonic acid. The cells were subsequently treated with various concentrations of dexamethasone. Concurrently, cultures of M-MSV-transformed 3T3 cells were prepared identically. After treatment, phospholipase activity was stimulated by the addition of serum or epidermal growth factor (EGF), and radioactivity released into the medium was taken as a measure of phospholipase activity. Dexamethasone (1 X 10(-5) or 1 X 10(-4) M) could inhibit serum-stimulated phospholipase activity in transformed 3T3 cells after 1 to 24 hr of treatment. However, no inhibition of activity was measured in palate mesenchymal cells following this period of treatment. Not until 120 hr of treatment with dexamethasone (1 X 10(-4) M) was any significant inhibition of serum-stimulated phospholipase activity observed in palate mesenchymal cells. When EGF was used to stimulate phospholipase activity, dexamethasone (1 X 10(-5) M) caused an increase in phospholipase activity in palate mesenchymal cells. These observations suggested that phospholipase in transformed 3T3 cells was sensitive to inhibition by dexamethasone. However, palate mesenchymal cell phospholipase is only minimally sensitive to dexamethasone, and in certain instances can be enhanced. These results cannot support the hypothesis that corticosteroids mediate their teratogenic effect via inhibition of phospholipase activity.

  12. Mitogen-activated Tasmanian devil blood mononuclear cells kill devil facial tumour disease cells.

    PubMed

    Brown, Gabriella K; Tovar, Cesar; Cooray, Anne A; Kreiss, Alexandre; Darby, Jocelyn; Murphy, James M; Corcoran, Lynn M; Bettiol, Silvana S; Lyons, A Bruce; Woods, Gregory M

    2016-08-01

    Devil facial tumour disease (DFTD) is a transmissible cancer that has brought the host species, the Tasmanian devil, to the brink of extinction. The cancer cells avoid allogeneic immune recognition by downregulating cell surface major histocompatibility complex (MHC) I expression. This should prevent CD8(+) T cell, but not natural killer (NK) cell, cytotoxicity. The reason why NK cells, normally reactive to MHC-negative cells, are not activated to kill DFTD cells has not been determined. The immune response of wild devils to DFTD, if it occurs, is uncharacterised. To investigate this, we tested 12 wild devils with DFTD, and found suggestive evidence of low levels of antibodies against DFTD cells in one devil. Eight of these devils were also analysed for cytotoxicity, however, none showed evidence for cytotoxicity against cultured DFTD cells. To establish whether mimicking activation of antitumour responses could induce cytotoxic activity against DFTD, Tasmanian devil peripheral blood mononuclear cells (PBMCs) were treated with either the mitogen Concanavalin A, the Toll-like receptor agonist polyinosinic:polycytidylic acid or recombinant Tasmanian devil IL-2. All induced the PBMC cells to kill cultured DFTD cells, suggesting that activation does not occur after encounter with DFTD cells in vivo, but can be induced. The identification of agents that activate cytotoxicity against DFTD target cells is critical for developing strategies to protect against DFTD. Such agents could function as adjuvants to induce functional immune responses capable of targeting DFTD cells and tumours in vivo. PMID:27089941

  13. A Rapid and Sensitive Method for Measuring N-Acetylglucosaminidase Activity in Cultured Cells

    PubMed Central

    Mauri, Victor; Lotfi, Parisa; Segatori, Laura; Sardiello, Marco

    2013-01-01

    A rapid and sensitive method to quantitatively assess N-acetylglucosaminidase (NAG) activity in cultured cells is highly desirable for both basic research and clinical studies. NAG activity is deficient in cells from patients with Mucopolysaccharidosis type IIIB (MPS IIIB) due to mutations in NAGLU, the gene that encodes NAG. Currently available techniques for measuring NAG activity in patient-derived cell lines include chromogenic and fluorogenic assays and provide a biochemical method for the diagnosis of MPS IIIB. However, standard protocols require large amounts of cells, cell disruption by sonication or freeze-thawing, and normalization to the cellular protein content, resulting in an error-prone procedure that is material- and time-consuming and that produces highly variable results. Here we report a new procedure for measuring NAG activity in cultured cells. This procedure is based on the use of the fluorogenic NAG substrate, 4-Methylumbelliferyl-2-acetamido-2-deoxy-alpha-D-glucopyranoside (MUG), in a one-step cell assay that does not require cell disruption or post-assay normalization and that employs a low number of cells in 96-well plate format. We show that the NAG one-step cell assay greatly discriminates between wild-type and MPS IIIB patient-derived fibroblasts, thus providing a rapid method for the detection of deficiencies in NAG activity. We also show that the assay is sensitive to changes in NAG activity due to increases in NAGLU expression achieved by either overexpressing the transcription factor EB (TFEB), a master regulator of lysosomal function, or by inducing TFEB activation chemically. Because of its small format, rapidity, sensitivity and reproducibility, the NAG one-step cell assay is suitable for multiple procedures, including the high-throughput screening of chemical libraries to identify modulators of NAG expression, folding and activity, and the investigation of candidate molecules and constructs for applications in enzyme replacement

  14. B Cell Infection and Activation by Rabies Virus-Based Vaccines

    PubMed Central

    Lytle, Andrew G.; Norton, James E.; Dorfmeier, Corin L.; Shen, Shixue

    2013-01-01

    Replication-deficient rabies viruses (RABV) are promising rabies postexposure vaccines due to their prompt and potent stimulation of protective virus neutralizing antibody titers, which are produced in mice by both T-dependent and T-independent mechanisms. To promote such early and robust B cell stimulation, we hypothesized that live RABV-based vaccines directly infect B cells, thereby activating a large pool of antigen-presenting cells (APCs) capable of providing early priming and costimulation to CD4+ T cells. In this report, we show that live RABV-based vaccine vectors efficiently infect naive primary murine and human B cells ex vivo. Infection of B cells resulted in the significant upregulation of early markers of B cell activation and antigen presentation, including CD69, major histocompatibility complex class II (MHC-II), and CD40 in murine B cells or HLA-DR and CD40 in human B cells compared to mock-infected cells or cells treated with an inactivated RABV-based vaccine. Furthermore, primary B cells infected with a live RABV expressing ovalbumin were able to prime and stimulate naive CD4+ OT-II T cells to proliferate and to secrete interleukin-2 (IL-2), demonstrating a functional consequence of B cell infection and activation by live RABV-based vaccine vectors. We propose that this direct B cell stimulation by live RABV-based vaccines is a potential mechanism underlying their induction of early protective T cell-dependent B cell responses, and that designing live RABV-based vaccines to infect and activate B cells represents a promising strategy to develop a single-dose postexposure rabies vaccine where the generation of early protective antibody titers is critical. PMID:23760241

  15. B cell infection and activation by rabies virus-based vaccines.

    PubMed

    Lytle, Andrew G; Norton, James E; Dorfmeier, Corin L; Shen, Shixue; McGettigan, James P

    2013-08-01

    Replication-deficient rabies viruses (RABV) are promising rabies postexposure vaccines due to their prompt and potent stimulation of protective virus neutralizing antibody titers, which are produced in mice by both T-dependent and T-independent mechanisms. To promote such early and robust B cell stimulation, we hypothesized that live RABV-based vaccines directly infect B cells, thereby activating a large pool of antigen-presenting cells (APCs) capable of providing early priming and costimulation to CD4(+) T cells. In this report, we show that live RABV-based vaccine vectors efficiently infect naive primary murine and human B cells ex vivo. Infection of B cells resulted in the significant upregulation of early markers of B cell activation and antigen presentation, including CD69, major histocompatibility complex class II (MHC-II), and CD40 in murine B cells or HLA-DR and CD40 in human B cells compared to mock-infected cells or cells treated with an inactivated RABV-based vaccine. Furthermore, primary B cells infected with a live RABV expressing ovalbumin were able to prime and stimulate naive CD4(+) OT-II T cells to proliferate and to secrete interleukin-2 (IL-2), demonstrating a functional consequence of B cell infection and activation by live RABV-based vaccine vectors. We propose that this direct B cell stimulation by live RABV-based vaccines is a potential mechanism underlying their induction of early protective T cell-dependent B cell responses, and that designing live RABV-based vaccines to infect and activate B cells represents a promising strategy to develop a single-dose postexposure rabies vaccine where the generation of early protective antibody titers is critical. PMID:23760241

  16. Selective activation of functional suppressor cells by human seminal fluid.

    PubMed Central

    Witkin, S S

    1986-01-01

    The ability of seminal fluid (SF) to induce suppressor cell activity from peripheral blood mononuclear cells (PBMN) was examined. PBMN were incubated with SF for 48 h, washed to remove SF components, treated with mitomycin C (mit C) and co-cultured with Raji cells, a lymphoblastoid cell line. Raji cell proliferation was inhibited by SF-treated PBMN proportionally to SF concentration. SF (50-200 micrograms), mit C-treated Raji cells or mit C-treated PBMN pre-incubated with phytohaemagglutinin were without effect on Raji cell growth. Suppressor T lymphocytes generated by incubation of PBMN with concanavalin A inhibited Raji cells to the same extent as did SF-treated PBMN. All activity was lost following heating at 56 degrees C for 30 min; freezing and thawing reduced the ability of SF to induce suppression by 50%. Dialysis of SF or treatment with antibody to prostaglandin E2 led to a 50% reduction in suppression. PMID:2943541

  17. Platelet activating factor: regulation by mast cells and aspirin.

    PubMed

    Denburg, J A; Williams, D B; Kinlough-Rathbone, R L; Cazenave, J P; Bienenstock, J

    1984-02-01

    We have investigated some aspects of the regulation of production of rat platelet activating factor (PAF)2 in vitro. Suspensions of unseparated (PLC1), mast cell-depleted (PLC2), or mast cell (MC)-enriched rat peritoneal lavage cells (PLC) were analyzed for PAF content by extraction at alkaline pH. PAF activity extracted from PLC1 varied inversely with viable cell concentration: at 1 X 10(6) cells/ml, 32 +/- 9.3 PAF units, decreasing to 11.2 +/- 9.5 units at 10 X 10(6) cells/ml, and no activity at higher concentrations. Incubation of PLC1 in Tyrode's buffer or acetylsalicylic acid (ASA), but not salicylate, resulted in a time-dependent loss of PAF activity. Mean PAF activity of PLC2 was similar to that in PLC1, while no PAF activity was extractable from MC. Co-incubation with MC extracts inhibited PAF activity of PLC1 extracts in a dose-dependent fashion. Ultracentrifugation of PAF-containing samples led to a loss of all PAF activity in PLC1 extracts, suggesting the association of PAF activity with subcellular components. PAF appears to be derived from a non-MC population of rat PLC, is not extractable from rat PLC in the presence of ASA and is inhibited by MC extracts. These studies suggest that ASA regulates PAF availability unrelated to its effect on cyclooxygenase and that MC membrane products directly inhibit PAF activity from rat PLC. PMID:6711391

  18. Effect of substrate mechanical properties on T cell activation

    NASA Astrophysics Data System (ADS)

    Hui, King; Upadhyaya, Arpita

    2013-03-01

    T cell activation is a key process in cell-mediated immunity, and engagement of T cell receptors by peptides on antigen presenting cells leads to activation of signaling cascades as well as cytoskeletal reorganization and large scale membrane deformations. While significant advances have been made in understanding the biochemical signaling pathways, the effects imposed by the physical environment and the role of mechanical forces on cell activation are not well understood. In this study, we have used anti-CD3 coated elastic polyacrylamide gels as stimulatory substrates to enable the spreading of Jurkat T cells and the measurement of cellular traction forces. We have investigated the effect of substrate stiffness on the dynamics of T cell spreading and cellular force generation. We found that T cells display more active and sustained edge dynamics on softer gels and that they exert increased traction stresses with increasing gel stiffness. A dynamic actin cytoskeleton was required to maintain the forces generated during activation, as inferred from small molecule inhibition experiments. Our results indicate an important role for physical properties of the antigen presenting cell as well as cytoskeleton-driven forces in signaling activation.

  19. BMP2 Transfer to Neighboring Cells and Activation of Signaling.

    PubMed

    Alborzinia, Hamed; Shaikhkarami, Marjan; Hortschansky, Peter; Wölfl, Stefan

    2016-09-01

    Morphogen gradients and concentration are critical features during early embryonic development and cellular differentiation. Previously we reported the preparation of biologically active, fluorescently labeled BMP2 and quantitatively analyzed their binding to the cell surface and followed BMP2 endocytosis over time on the level of single endosomes. Here we show that this internalized BMP2 can be transferred to neighboring cells and, moreover, also activates downstream BMP signaling in adjacent cells, indicated by Smad1/5/8 phosphorylation and activation of the downstream target gene id1. Using a 3D matrix to modulate cell-cell contacts in culture we could show that direct cell-cell contact significantly increased BMP2 transfer. Using inhibitors of vesicular transport, transfer was strongly inhibited. Interestingly, cotreatment with the physiological BMP inhibitor Noggin increased BMP2 uptake and transfer, albeit activation of Smad signaling in neighboring cells was completely suppressed. Our findings present a novel and interesting mechanism by which morphogens such as BMP2 can be transferred between cells and how this is modulated by BMP antagonists such as Noggin, and how this influences activation of Smad signaling by BMP2 in neighboring cells. PMID:27306974

  20. Functional Implications of Plasma Membrane Condensation for T Cell Activation

    PubMed Central

    Quinn, Carmel M.; Engelhardt, Karin; Williamson, David; Grewal, Thomas; Jessup, Wendy; Harder, Thomas; Gaus, Katharina

    2008-01-01

    The T lymphocyte plasma membrane condenses at the site of activation but the functional significance of this receptor-mediated membrane reorganization is not yet known. Here we demonstrate that membrane condensation at the T cell activation sites can be inhibited by incorporation of the oxysterol 7-ketocholesterol (7KC), which is known to prevent the formation of raft-like liquid-ordered domains in model membranes. We enriched T cells with 7KC, or cholesterol as control, to assess the importance of membrane condensation for T cell activation. Upon 7KC treatment, T cell antigen receptor (TCR) triggered calcium fluxes and early tyrosine phosphorylation events appear unaltered. However, signaling complexes form less efficiently on the cell surface, fewer phosphorylated signaling proteins are retained in the plasma membrane and actin restructuring at activation sites is impaired in 7KC-enriched cells resulting in compromised downstream activation responses. Our data emphasizes lipids as an important medium for the organization at T cell activation sites and strongly indicates that membrane condensation is an important element of the T cell activation process. PMID:18509459

  1. Mast cells and dendritic cells form synapses that facilitate antigen transfer for T cell activation

    PubMed Central

    Carroll-Portillo, Amanda; Cannon, Judy L.; te Riet, Joost; Holmes, Anna; Kawakami, Yuko; Kawakami, Toshiaki; Cambi, Alessandra

    2015-01-01

    Mast cells (MCs) produce soluble mediators such as histamine and prostaglandins that are known to influence dendritic cell (DC) function by stimulating maturation and antigen processing. Whether direct cell–cell interactions are important in modulating MC/DC function is unclear. In this paper, we show that direct contact between MCs and DCs occurs and plays an important role in modulating the immune response. Activation of MCs through FcεRI cross-linking triggers the formation of stable cell–cell interactions with immature DCs that are reminiscent of the immunological synapse. Direct cellular contact differentially regulates the secreted cytokine profile, indicating that MC modulation of DC populations is influenced by the nature of their interaction. Synapse formation requires integrin engagement and facilitates the transfer of internalized MC-specific antigen from MCs to DCs. The transferred material is ultimately processed and presented by DCs and can activate T cells. The physiological outcomes of the MC–DC synapse suggest a new role for intercellular crosstalk in defining the immune response. PMID:26304724

  2. Activating Receptor Signals Drive Receptor Diversity in Developing Natural Killer Cells.

    PubMed

    Freund, Jacquelyn; May, Rebecca M; Yang, Enjun; Li, Hongchuan; McCullen, Matthew; Zhang, Bin; Lenvik, Todd; Cichocki, Frank; Anderson, Stephen K; Kambayashi, Taku

    2016-08-01

    It has recently been appreciated that NK cells exhibit many features reminiscent of adaptive immune cells. Considerable heterogeneity exists with respect to the ligand specificity of individual NK cells and as such, a subset of NK cells can respond, expand, and differentiate into memory-like cells in a ligand-specific manner. MHC I-binding inhibitory receptors, including those belonging to the Ly49 and KIR families, are expressed in a variegated manner, which creates ligand-specific diversity within the NK cell pool. However, how NK cells determine which inhibitory receptors to express on their cell surface during a narrow window of development is largely unknown. In this manuscript, we demonstrate that signals from activating receptors are critical for induction of Ly49 and KIR receptors during NK cell development; activating receptor-derived signals increased the probability of the Ly49 bidirectional Pro1 promoter to transcribe in the forward versus the reverse direction, leading to stable expression of Ly49 receptors in mature NK cells. Our data support a model where the balance of activating and inhibitory receptor signaling in NK cells selects for the induction of appropriate inhibitory receptors during development, which NK cells use to create a diverse pool of ligand-specific NK cells. PMID:27500644

  3. Activating Receptor Signals Drive Receptor Diversity in Developing Natural Killer Cells

    PubMed Central

    Freund, Jacquelyn; May, Rebecca M.; Li, Hongchuan; McCullen, Matthew; Zhang, Bin; Lenvik, Todd; Cichocki, Frank; Anderson, Stephen K.; Kambayashi, Taku

    2016-01-01

    It has recently been appreciated that NK cells exhibit many features reminiscent of adaptive immune cells. Considerable heterogeneity exists with respect to the ligand specificity of individual NK cells and as such, a subset of NK cells can respond, expand, and differentiate into memory-like cells in a ligand-specific manner. MHC I-binding inhibitory receptors, including those belonging to the Ly49 and KIR families, are expressed in a variegated manner, which creates ligand-specific diversity within the NK cell pool. However, how NK cells determine which inhibitory receptors to express on their cell surface during a narrow window of development is largely unknown. In this manuscript, we demonstrate that signals from activating receptors are critical for induction of Ly49 and KIR receptors during NK cell development; activating receptor-derived signals increased the probability of the Ly49 bidirectional Pro1 promoter to transcribe in the forward versus the reverse direction, leading to stable expression of Ly49 receptors in mature NK cells. Our data support a model where the balance of activating and inhibitory receptor signaling in NK cells selects for the induction of appropriate inhibitory receptors during development, which NK cells use to create a diverse pool of ligand-specific NK cells. PMID:27500644

  4. Tubular solid oxide fuel cell demonstration activities

    SciTech Connect

    Veyo, S.E.

    1995-08-01

    The development of a viable fuel cell driven electrical power generation system involves not only the development of cell and stack technology, but also the development of the overall system concept, the strategy for control, and the ancillary subsystems. The design requirements used to guide system development must reflect a customer focus in order to evolve a commercial product. In order to obtain useful customer feedback, Westinghouse has practiced the deployment with customers of fully integrated, automatically controlled, packaged solid oxide fuel cell power generation systems. These field units have served to demonstrate to customers first hand the beneficial attributes of the SOFC, to expose deficiencies through experience in order to guide continued development, and to garner real world feedback and data concerning not only cell and stack parameters, but also transportation, installation, permitting and licensing, start-up and shutdown, system alarming, fault detection, fault response, and operator interaction.

  5. Interleukin-7 and Toll-Like Receptor 7 Induce Synergistic B Cell and T Cell Activation

    PubMed Central

    Bikker, Angela; Kruize, Aike A.; van der Wurff-Jacobs, Kim M. G.; Peters, Rogier P.; Kleinjan, Marije; Redegeld, Frank; de Jager, Wilco; Lafeber, Floris P. J. G.; van Roon, Joël A. G.

    2014-01-01

    Objectives To investigate the potential synergy of IL-7-driven T cell-dependent and TLR7-mediated B cell activation and to assess the additive effects of monocyte/macrophages in this respect. Methods Isolated CD19 B cells and CD4 T cells from healthy donors were co-cultured with TLR7 agonist (TLR7A, Gardiquimod), IL-7, or their combination with or without CD14 monocytes/macrophages (T/B/mono; 1 : 1 : 0,1). Proliferation was measured using 3H-thymidine incorporation and Ki67 expression. Activation marker (CD19, HLA-DR, CD25) expression was measured by FACS analysis. Immunoglobulins were measured by ELISA and release of cytokines was measured by Luminex assay. Results TLR7-induced B cell activation was not associated with T cell activation. IL-7-induced T cell activation alone and together with TLR7A synergistically increased numbers of both proliferating (Ki67+) B cells and T cells, which was further increased in the presence of monocytes/macrophages. This was associated by up regulation of activation markers on B cells and T cells. Additive or synergistic induction of production of immunoglobulins by TLR7 and IL-7 was associated by synergistic induction of T cell cytokines (IFNγ, IL-17A, IL-22), which was only evident in the presence of monocytes/macrophages. Conclusions IL-7-induced CD4 T cell activation and TLR7-induced B cell activation synergistically induce T helper cell cytokine and B cell immunoglobulin production, which is critically dependent on monocytes/macrophages. Our results indicate that previously described increased expression of IL-7 and TLR7 together with increased numbers of macrophages at sites of inflammation in autoimmune diseases like RA and pSS significantly contributes to enhanced lymphocyte activation. PMID:24740301

  6. Expression of executioner procaspases and their activation by a procaspase-activating compound in chronic lymphocytic leukemia cells

    PubMed Central

    Patel, Viralkumar; Balakrishnan, Kumudha; Keating, Michael J.; Wierda, William G.

    2015-01-01

    Intrinsic and extrinsic apoptotic pathways converge to activate common downstream executioner caspases (caspase-3, -6, and -7), resulting in cell death. In chronic lymphocytic leukemia (CLL), neoplastic B cells evade apoptosis owing to the overexpression of survival proteins. We hypothesized that direct activation of procaspases could bypass the apoptosis resistance induced by the upstream prosurvival proteins. The procaspase-activating compounds (PAC-1), including B-PAC-1 (L14R8), convert inactive executioner procaspases to their active cleaved forms by chelation of labile zinc ions. Both at transcript and protein levels, primary CLL cells express high levels of latent procaspases (3, -7, and -9). B-PAC-1 treatment induced CLL lymphocyte death which was higher than that in normal peripheral blood mononuclear cells or B cells, and was independent of prognostic markers and microenvironmental factors. Mechanistically, B-PAC-1 treatment activated executioner procaspases and not other Zn-dependent enzymes. Exogenous zinc completely, and pancaspase inhibitors partially, reversed B-PAC-1–induced apoptosis, elucidating the zinc-mediated mechanism of action. The cell demise relied on the presence of caspase-3/7 but not caspase-8 or Bax/Bak proteins. B-PAC-1 in combination with an inhibitor of apoptosis protein antagonist (Smac066) synergistically induced apoptosis in CLL samples. Our investigations demonstrated that direct activation of executioner procaspases via B-PAC-1 treatment bypasses apoptosis resistance and is a novel approach for CLL therapeutics. PMID:25538042

  7. Sox2 Activates Cell Proliferation and Differentiation in the Respiratory Epithelium

    PubMed Central

    Tompkins, David H.; Besnard, Valérie; Lange, Alexander W.; Keiser, Angela R.; Wert, Susan E.; Bruno, Michael D.; Whitsett, Jeffrey A.

    2011-01-01

    Sox2, a transcription factor critical for the maintenance of embryonic stem cells and induction of pluripotent stem cells, is expressed exclusively in the conducting airway epithelium of the lung, where it is required for differentiation of nonciliated, goblet, and ciliated cells. To determine the role of Sox2 in respiratory epithelial cells, Sox2 was selectively and conditionally expressed in nonciliated airway epithelial cells and in alveolar type II cells in the adult mouse. Sox2 induced epithelial cell proliferation within 3 days of expression. Epithelial cell proliferation was associated with increased Ki-67 and cyclin D1 staining. Expression of cell cycle genes, including FoxM1, Ccna2 (Cyclin A2), Ccnb2 (Cyclin B2), and Ccnd1 (Cyclin D1), was increased. Consistent with a role in cell proliferation, Sox2 activated the transcription of FoxM1 in vitro. In alveoli, Sox2 caused hyperplasia and ectopic differentiation of epithelial cells to those with morphologic and molecular characteristics of conducting airway epithelium. Sox2 induced the expression of conducting airway epithelial specific genes, including Scgb1a1, Foxj1, Tubb3, and Cyp2f2. Although prolonged expression of Sox2 caused cell proliferation and epithelial hyperplasia, Sox2 did not induce pulmonary tumors. Sox2 induces proliferation of respiratory epithelial cells and, subsequently, partially reprograms alveolar epithelial cells into cells with characteristics of the conducting airways. PMID:20855650

  8. Cutting edge: cell surface linker for activation of T cells is recruited to microclusters and is active in signaling.

    PubMed

    Balagopalan, Lakshmi; Barr, Valarie A; Kortum, Robert L; Park, Anna K; Samelson, Lawrence E

    2013-04-15

    A controversy has recently emerged regarding the location of the cellular pool of the adapter linker for activation of T cells (LAT) that participates in propagation of signals downstream of the TCR. In one model phosphorylation and direct recruitment of cell surface LAT to activation-induced microclusters is critical for T cell activation, whereas in the other model vesicular, but not surface, LAT participates in these processes. By using a chimeric version of LAT that can be tracked via an extracellular domain, we provide evidence that LAT located at the cell surface can be recruited efficiently to activation-induced microclusters within seconds of TCR engagement. Importantly, we also demonstrate that this pool of LAT at the plasma membrane is rapidly phosphorylated. Our results provide support for the model in which the cell utilizes LAT from the cell surface for rapid responses to TCR stimulation. PMID:23487428

  9. Calpain secreted by activated human lymphoid cells degrades myelin.

    PubMed

    Deshpande, R V; Goust, J M; Hogan, E L; Banik, N L

    1995-10-01

    Calpain secreted by lymphoid (MOLT-3, M.R.) or monocytic (U-937, THP-1) cell lines activated with PMA and A23187 degraded myelin antigens. The degradative effect of enzymes released in the extracellular medium was tested on purified myelin basic protein and rat central nervous system myelin in vitro. The extent of protein degradation was determined by SDS-PAGE and densitometric analysis. Various proteinase inhibitors were used to determine to what extent protein degradation was mediated by calpain and/or other enzymes. Lysosomal and serine proteinase inhibitors inhibited 20-40% of the myelin-degradative activity found in the incubation media of cell lines, whereas the calcium chelator (EGTA), the calpain-specific inhibitor (calpastatin), and a monoclonal antibody to m calpain blocked myelin degradation by 60-80%. Since breakdown products of MBP generated by calpain may include fragments with antigenic epitopes, this enzyme may play an important role in the initiation of immune-mediated demyelination. PMID:8568927

  10. Activation of natural killer cells and dendritic cells upon recognition of a novel CD99-like ligand by paired immunoglobulin-like type 2 receptor.

    PubMed

    Shiratori, Ikuo; Ogasawara, Kouetsu; Saito, Takashi; Lanier, Lewis L; Arase, Hisashi

    2004-02-16

    Paired receptors that consist of highly related activating and inhibitory receptors are widely involved in the regulation of the immune system. Here, we report a mouse orthologue of the human activating paired immunoglobulin-like type 2 receptor (PILR) beta, which was cloned from a cDNA library of natural killer (NK) cells based on its ability to associate with the DAP12 signaling adaptor protein. The activating PILRbeta was expressed not only on NK cells but also on dendritic cells and macrophages. Furthermore, we have identified a novel CD99-like molecule as a ligand for the activating PILRbeta and inhibitory PILRalpha receptors. Transcripts of PILR ligand are present in many tissues, including some T cell lines. Cells expressing the PILR ligand specifically activated NK cells and dendritic cells that express the activating PILRbeta. Our findings reveal a new regulatory mechanism of innate immunity by PILR and its CD99-like ligand. PMID:14970179

  11. Comparative analysis of signature genes in porcine reproductive and respiratory syndrome virus (PRRSV)-infected porcine monocyte-derived dendritic cells at differential activation statuses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Activation statuses of monocytic cells, e.g. monocytes, macrophages and dendritic cells (DCs), are critically important for antiviral immunity. In particular, some devastating viruses, including porcine reproductive and respiratory syndrome virus (PRRSV), are capable of directly infecting these cell...

  12. T Cell Activation Thresholds are Affected by Gravitational

    NASA Technical Reports Server (NTRS)

    Adams, Charley; Gonzalez, M.; Nelman-Gonzalez, M.

    1999-01-01

    T cells stimulated in space flight by various mitogenic signals show a dramatic reduction in proliferation and expression of early activation markers. Similar results are also obtained in a ground based model of microgravity, clinorotation, which provides a vector-averaged reduction of the apparent gravity on cells without significant shear force. Here we demonstrate that T cell inhibition is due to an increase in the required threshold for activation. Dose response curves indicate that cells activated during clinorotation require higher stimulation to achieve the same level of activation, as measured by CD69 expression. Interleukin 2 receptor expression, and DNA synthesis. The amount of stimulation necessary for 50% activation is 5 fold in the clinostat relative to static. Correlation of TCR internalization with activation also exhibit a dramatic right shift in clinorotation, demonstrating unequivocally that signal transduction mechanism independent of TCR triggering account for the increased activation threshold. Previous results from space flight experiments are consistent with the dose response curves obtained for clinorotation. Activation thresholds are important aspects of T cell memory, autoimmunity and tolerance Clinorotation is a useful, noninvasive tool for the study of cellular and biochemical event regulating T cell activation threshold and the effects of gravitation forces on these systems.

  13. Retinal Pigment Epithelial Cell Line Suppression of Phagolysosome Activation

    PubMed Central

    Taylor, AW; Dixit, S; Yu, J

    2015-01-01

    The eye is an immune privileged tissue with multiple mechanisms of immunosuppression to protect the light gathering tissues from the damage of inflammation. One of theses mechanisms involves retinal pigment epithelial cell suppression of phagosome activation in macrophages. The objective of this work is to determine if the human RPE cell line ARPE-19 is capable of suppressing the activation of the phagolysosome in macrophages in a manner similar to primary RPE. The conditioned media of RPE eyecups, sub-confluent, just confluent cultures, or established confluent cultures of human ARPE-19 cells were generated. These condition media were used to treat macrophages phagocytizing pHrodo bioparticles. After 24 hours incubation the macrophages were imaged by fluorescent microscopy, and fluorescence was measured. The fluorescent intensity is proportional to the amount of bioparticles phagocytized and are in an activated phagolysosome. The conditioned media of in situ mouse RPE eyecups significantly suppressed the activation of phagolysosome. The conditioned media from cultures of human ARPE-19 cells, grown to sub-confluence (50%) or grown to confluence had no effect on phagolysosome activation. In contrast, the conditioned media from established confluent cultures significantly suppressed phagolysosome activation. The neuropeptides alpha-MSH and NPY were depleted from the conditioned media of established confluent ARPE-19 cell cultures. This depleted conditioned media had diminished suppression of phagolysosome activation while promoting macrophage cell death. In addition, the condition media from cultures of ARPE-19 monolayers wounded with a bisecting scrape was diminished in suppressing phagolysosome activation. This technical report suggests that like primary RPE monolayers, established confluent cultures of ARPE-19 cells produce soluble factors that suppress the activation of macrophages, and can be used to study the molecular mechanisms of retinal immunobiology. In

  14. Variable susceptibility of ovarian cancer cells to non-thermal plasma-activated medium

    PubMed Central

    UTSUMI, FUMI; KAJIYAMA, HIROAKI; NAKAMURA, KAE; TANAKA, HIROMASA; MIZUNO, MASAAKI; TOYOKUNI, SHINNYA; HORI, MASARU; KIKKAWA, FUMITAKA

    2016-01-01

    Non-thermal atmospheric pressure plasma has been widely studied in recent years in many fields, including cancer treatment. However, its efficiency for inducing apoptosis sometimes varies depending on the cell species and experimental conditions. The aim of this study was to elucidate what causes these differences in responses to plasma treatment. Using four ovarian cancer cell lines, the cell density had a markedly negative impact on the proliferation inhibition rate (PIR) and it was more obvious in OVCAR-3 and NOS2 cells. Furthermore, TOV21G and ES-2 cells were drastically sensitive to plasma-activated medium (PAM) compared with the other two cell lines. We demonstrated that the proportion of reactive oxygen species and cell number had a marked impact on the effect of PAM against ovarian cancer cells. Additionally it was suggested that the morphological features of cells were also closely related PMID:27035127

  15. Identification of aromatase activity in rodent pituitary cell strains.

    PubMed

    Callard, G V; Petro, Z; Tashjian, A H

    1983-07-01

    To date, biochemical evidence has been presented for hypophysial aromatization in only one species, a teleost fish, although the pituitary glands of several mammals have been reported to be aromatase negative. To reinvestigate this problem, established clonal strains of rodent pituitary cells (GH3, GH4C1, and AtT20/D16) were incubated at 37 C for 6-48 h in serum-less medium containing [7-3H]androstenedione. Radiolabeled metabolites were isolated by solvent extraction, thin layer chromatography, and phenolic partition. The authenticity of the estrogenic products in both cells and incubation medium was verified by methylation and recrystallization to constant specific activity. Measurement of androgen metabolites was also validated by recrystallization of selected samples. Authentic estrone and 17 beta-estradiol were identified in cultures of the two PRL- and GH-secreting clones, and there were strain differences in the quantity of estrogen produced (GH3 greater than GH4C1). Under the same conditions, aromatization was not detectable in the ACTH-secreting line (AtT20/D16). A time-yield analysis of androgen metabolism in GH4C1 cells showed that aromatization was linear for 12 h after labeling, but that substrate was diverted mainly to 5 alpha-reducing pathways. Large amounts of highly polar metabolites accumulated 24 and 48 h after the addition of [3H]androgen, and subsequent hydrolysis revealed that these were sulfo- and glucuronoconjugates. The metabolic fate of estrogen in GH4C1 cultures was investigated indirectly by adding a radioinert estrone trap together with the radiolabeled androgen substrate and was also tested in separate cultures by adding [3H]estrone and [3H]estradiol directly. Although the two estrogens were interconverted, there was no evidence that formed or added estrogen was extensively metabolized or conjugated. We conclude that the expression of aromatase activity in hypophysial cells is not a property of all transformed lines but may be dictated

  16. ACTIVATION OF THE EGF RECEPTOR SIGNALING PATHWAY IN HUMAN AIRWAY EPITHELIAL CELLS EXPOSED TO METALS

    EPA Science Inventory

    We have previously shown that exposure to combustion-derived metals rapidly (within 20 min) activated mitogen-activated protein kinases (MAPK), including extracellular signal-regulated kinase (ERK), in the human bronchial epithelial cell line BEAS. To study the mechanisms respons...

  17. Activation of human inflammatory cells by secreted phospholipases A2.

    PubMed

    Triggiani, Massimo; Granata, Francescopaolo; Frattini, Annunziata; Marone, Gianni

    2006-11-01

    Secreted phospholipases A(2) (sPLA(2)s) are enzymes detected in serum and biological fluids of patients with various inflammatory, autoimmune and allergic disorders. Different isoforms of sPLA(2)s are expressed and released by human inflammatory cells, such as neutrophils, eosinophils, T cells, monocytes, macrophages and mast cells. sPLA(2)s generate arachidonic acid and lysophospholipids thus contributing to the production of bioactive lipid mediators in inflammatory cells. However, sPLA(2)s also activate human inflammatory cells by mechanisms unrelated to their enzymatic activity. Several human and non-human sPLA(2)s induce degranulation of mast cells, neutrophils and eosinophils and activate exocytosis in macrophages. In addition some, but not all, sPLA(2) isoforms promote cytokine and chemokine production from macrophages, neutrophils, eosinophils, monocytes and endothelial cells. These effects are primarily mediated by binding of sPLA(2)s to specific membrane targets (heparan sulfate proteoglycans, M-type, N-type or mannose receptors) expressed on effector cells. Thus, sPLA(2)s may play an important role in the initiation and amplification of inflammatory reactions by at least two mechanisms: production of lipid mediators and direct activation of inflammatory cells. Selective inhibitors of sPLA(2)-enzymatic activity and specific antagonists of sPLA(2) receptors are current being tested for pharmacological treatment of inflammatory and autoimmune diseases. PMID:16952481

  18. Resveratrol activates autophagic cell death in prostate cancer cells via downregulation of STIM1 and the mTOR pathway.

    PubMed

    Selvaraj, Senthil; Sun, Yuyang; Sukumaran, Pramod; Singh, Brij B

    2016-05-01

    Resveratrol (RSV), a natural polyphenol, has been suggested to induce cell cycle arrest and activate apoptosis-mediated cell death in several cancer cells, including prostate cancer. However, several molecular mechanisms have been proposed on its chemopreventive action, the precise mechanisms by which RSV exerts its anti-proliferative effect in androgen-independent prostate cancer cells remain questionable. In the present study, we show that RSV activates autophagic cell death in PC3 and DU145 cells, which was dependent on stromal interaction molecule 1 (STIM1) expression. RSV treatment decreases STIM1 expression in a time-dependent manner and attenuates STIM1 association with TRPC1 and Orai1. Furthermore, RSV treatment also decreases ER calcium storage and store operated calcium entry (SOCE), which induces endoplasmic reticulum (ER) stress, thereby, activating AMPK and inhibiting the AKT/mTOR pathway. Similarly, inhibition of SOCE by SKF-96365 decreases the survival and proliferation of PC3 and DU145 cells and inhibits AKT/mTOR pathway and induces autophagic cell death. Importantly, SOCE inhibition and subsequent autophagic cell death caused by RSV was reversed by STIM1 overexpression. STIM1 overexpression restored SOCE, prevents the loss of mTOR phosphorylation and decreased the expression of CHOP and LC3A in PC3 cells. Taken together, for the first time, our results revealed that RSV induces autophagy-mediated cell death in PC3 and DU145 cells through regulation of SOCE mechanisms, including downregulating STIM1 expression and trigger ER stress by depleting ER calcium pool. © 2015 The Authors. Molecular Carcinogenesis, published by Wiley Periodicals, Inc. PMID:25917875

  19. The individual-cell-based cryo-chip for the cryopreservation, manipulation and observation of spatially identifiable cells. II: Functional activity of cryopreserved cells

    PubMed Central

    2010-01-01

    Background The cryopreservation and thawing processes are known to induce many deleterious effects in cells and might be detrimental to several cell types. There is an inherent variability in cellular responses among cell types and within individual cells of a given population with regard to their ability to endure the freezing and thawing process. The aim of this study was to evaluate the fate of cryopreserved cells within an optical cryo apparatus, the individual-cell-based cryo-chip (i3C), by monitoring several basic cellular functional activities at the resolution of individual cells. Results In the present study, U937 cells underwent the freezing and thawing cycle in the i3C device. Then a panel of vital tests was performed, including the number of dead cells (PI staining), apoptotic rate (Annexin V staining), mitochondrial membrane potential (TMRM staining), cytoplasm membrane integrity and intracellular metabolism (FDA staining), as well as post-thawing cell proliferation assays. Cells that underwent the freezing - thawing cycle in i3C devices exhibited the same functional activity as control cells. Moreover, the combination of the multi-parametric analysis at a single cell resolution and the optical and biological features of the device enable an accurate determination of the functional status of individual cells and subsequent retrieval and utilization of the most valuable cells. Conclusions The means and methodologies described here enable the freezing and thawing of spatially identifiable cells, as well as the efficient detection of viable, specific, highly biologically active cells for future applications. PMID:20973993

  20. Active elastic dimers: cells moving on rigid tracks.

    PubMed

    Lopez, J H; Das, Moumita; Schwarz, J M

    2014-09-01

    Experiments suggest that the migration of some cells in the three-dimensional extracellular matrix bears strong resemblance to one-dimensional cell migration. Motivated by this observation, we construct and study a minimal one-dimensional model cell made of two beads and an active spring moving along a rigid track. The active spring models the stress fibers with their myosin-driven contractility and α-actinin-driven extendability, while the friction coefficients of the two beads describe the catch and slip-bond behaviors of the integrins in focal adhesions. In the absence of active noise, net motion arises from an interplay between active contractility (and passive extendability) of the stress fibers and an asymmetry between the front and back of the cell due to catch-bond behavior of integrins at the front of the cell and slip-bond behavior of integrins at the back. We obtain reasonable cell speeds with independently estimated parameters. We also study the effects of hysteresis in the active spring, due to catch-bond behavior and the dynamics of cross linking, and the addition of active noise on the motion of the cell. Our model highlights the role of α-actinin in three-dimensional cell motility and does not require Arp2/3 actin filament nucleation for net motion. PMID:25314473

  1. Active elastic dimers: Cells moving on rigid tracks

    NASA Astrophysics Data System (ADS)

    Lopez, J. H.; Das, Moumita; Schwarz, J. M.

    2014-09-01

    Experiments suggest that the migration of some cells in the three-dimensional extracellular matrix bears strong resemblance to one-dimensional cell migration. Motivated by this observation, we construct and study a minimal one-dimensional model cell made of two beads and an active spring moving along a rigid track. The active spring models the stress fibers with their myosin-driven contractility and α-actinin-driven extendability, while the friction coefficients of the two beads describe the catch and slip-bond behaviors of the integrins in focal adhesions. In the absence of active noise, net motion arises from an interplay between active contractility (and passive extendability) of the stress fibers and an asymmetry between the front and back of the cell due to catch-bond behavior of integrins at the front of the cell and slip-bond behavior of integrins at the back. We obtain reasonable cell speeds with independently estimated parameters. We also study the effects of hysteresis in the active spring, due to catch-bond behavior and the dynamics of cross linking, and the addition of active noise on the motion of the cell. Our model highlights the role of α-actinin in three-dimensional cell motility and does not require Arp2/3 actin filament nucleation for net motion.

  2. STAT3 Impairs STAT5 Activation in the Development of IL-9-Secreting T Cells.

    PubMed

    Olson, Matthew R; Verdan, Felipe Fortino; Hufford, Matthew M; Dent, Alexander L; Kaplan, Mark H

    2016-04-15

    Th cell subsets develop in response to multiple activating signals, including the cytokine environment. IL-9-secreting T cells develop in response to the combination of IL-4 and TGF-β, although they clearly require other cytokine signals, leading to the activation of transcription factors including STAT5. In Th17 cells, there is a molecular antagonism of STAT5 with STAT3 signaling, although whether this paradigm exists in other Th subsets is not clear. In this paper, we demonstrate that STAT3 attenuates the ability of STAT5 to promote the development of IL-9-secreting T cells. We demonstrate that production of IL-9 is increased in the absence of STAT3 and cytokines that result in a sustained activation of STAT3, including IL-6, have the greatest potency in repressing IL-9 production in a STAT3-dependent manner. Increased IL-9 production in the absence of STAT3 correlates with increased endogenous IL-2 production and STAT5 activation, and blocking IL-2 responses eliminates the difference in IL-9 production between wild-type and STAT3-deficient T cells. Moreover, transduction of developing Th9 cells with a constitutively active STAT5 eliminates the ability of IL-6 to reduce IL-9 production. Thus, STAT3 functions as a negative regulator of IL-9 production through attenuation of STAT5 activation and function. PMID:26976954

  3. A fluorescence-based demonstration of intestinal villi and epithelial cell in chickens fed dietary silicic acid powder including bamboo vinegar compound liquid.

    PubMed

    Ruttanavut, J; Matsumoto, Y; Yamauchi, K

    2012-10-01

    This study investigates the combined effect of silicic acid and bamboo vinegar compound liquid (SPV) on the growth and intestinal histological alterations in poultry. Forty-eight 7-day-old male Sanuki Cochin chickens were fed a commercial mash diet supplemented with SPV at 0, 0.1, 0.2, and 0.3% level ad libitum for 112 days. Body weight gain tended to improve with increased concentrations of dietary SPV, although these results were not statistically significant (P<0.1). Tissue observation by light microscopy revealed that the jejunal villus height (P<0.01) and duodenal and jejunal villus area (P<0.05) increased in the 0.2 and 0.3% SPV groups, respectively, compared with the control. Cell mitosis within the duodenum and jejunum also increased in the 0.2 and 0.3% SPV groups. Scanning electron microscopy revealed a prominent increase in the number of protuberant cells on the villus apical surface of the duodenum and jejunum for the 0.2 and 0.3% SPV groups compared with the control. Poultry in the 0.3% SPV group had the highest body weight gain and hypertrophied histological alterations of intestinal villi. Fluorescent microscopic images of cell mitosis and protuberant cells in the duodenal crypt clearly confirmed positive reactions for the activator protein 2α (AP-2α) and proliferating cell nuclear antigen (PCNA), compared with the control. The present results indicate that dietary SPV stimulates adsorption by the epithelial cells, which activate cell proliferation and self-renewal and regulate the expression of cell cycle regulators AP-2α and PCNA, resulting in higher body weight gain. Thus, we can conclude that a concentration of 0.3% dietary SPV is ideal for promoting growth in poultry. PMID:22936452

  4. Emergent cell and tissue dynamics from subcellular modeling of active biomechanical processes

    NASA Astrophysics Data System (ADS)

    Sandersius, S. A.; Weijer, C. J.; Newman, T. J.

    2011-08-01

    Cells and the tissues they form are not passive material bodies. Cells change their behavior in response to external biochemical and biomechanical cues. Behavioral changes, such as morphological deformation, proliferation and migration, are striking in many multicellular processes such as morphogenesis, wound healing and cancer progression. Cell-based modeling of these phenomena requires algorithms that can capture active cell behavior and their emergent tissue-level phenotypes. In this paper, we report on extensions of the subcellular element model to model active biomechanical subcellular processes. These processes lead to emergent cell and tissue level phenotypes at larger scales, including (i) adaptive shape deformations in cells responding to slow stretching, (ii) viscous flow of embryonic tissues, and (iii) streaming patterns of chemotactic cells in epithelial-like sheets. In each case, we connect our simulation results to recent experiments.

  5. Nuclear Envelope Lamin-A Couples Actin Dynamics with Immunological Synapse Architecture and T Cell Activation

    PubMed Central

    González-Granado, José María; Trigueros-Motos, Laia; Cibrián, Danay; Morlino, Giulia; Blanco-Berrocal, Marta; Osorio, Fernando Garcia; Freije, José María Pérez; López-Otín, Carlos; Sánchez-Madrid, Francisco; Andrés, Vicente

    2014-01-01

    In many cell types, nuclear A-type lamins have been implicated in structural and functional activities, including higher-order genome organization, DNA replication and repair, gene transcription, and signal transduction. However, their role in specialized immune cells remains largely unexplored. Here, we showed that the abundance of A-type lamins is almost negligible in resting naïve T lymphocytes, but that it is substantially increased upon activation of the T cell receptor (TCR), and is an early event that accelerates formation of the immunological synapse between T cells and antigen-presenting cells. We found that lamin-A enhanced the polymerization of F-actin in T cells, a critical step for immunological synapse formation, by physically connecting the nucleus to the plasma membrane through the linker of nucleoskeleton and cytoskeleton (LINC) complex. We also showed that lamin-A played a key role in other membrane, cytoplasmic, and nuclear events related to TCR activation, including receptor-clustering, downstream signaling, and target gene expression. Notably, the presence of lamin-A was associated with enhanced extracellular signal–regulated kinase 1/2 signaling, and pharmacological inhibition of this pathway reduced the extent of lamin-A–dependent T cell activation. Moreover, mice deficient in lamin-A exhibited impaired T cell responses in vivo. These findings underscore the importance of A-type lamins for TCR activation, and identify lamin-A as a previously unappreciated regulator of the immune response. PMID:24757177

  6. Activating β-catenin signaling in CD133-positive dermal papilla cells increases hair inductivity.

    PubMed

    Zhou, Linli; Yang, Kun; Xu, Mingang; Andl, Thomas; Millar, Sarah E; Boyce, Steven; Zhang, Yuhang

    2016-08-01

    Bioengineering hair follicles using cells isolated from human tissue remains a difficult task. Dermal papilla (DP) cells are known to guide the growth and cycling activities of hair follicles by interacting with keratinocytes. However, DP cells quickly lose their inductivity during in vitro passaging. Rodent DP cell cultures need external addition of growth factors, including WNT and BMP molecules, to maintain the hair inductive property. CD133 is expressed by a subpopulation of DP cells that are capable of inducing hair follicle formation in vivo. We report here that expression of a stabilized form of β-catenin promoted clonal growth of CD133-positive (CD133+) DP cells in in vitro three-dimensional hydrogel culture while maintaining expression of DP markers, including alkaline phosphatase (AP), CD133, and integrin α8. After a 2-week in vitro culture, cultured CD133+ DP cells with up-regulated β-catenin activity led to an accelerated in vivo hair growth in reconstituted skin compared to control cells. Further analysis showed that matrix cell proliferation and differentiation were significantly promoted in hair follicles when β-catenin signaling was up-regulated in CD133+ DP cells. Our data highlight an important role for β-catenin signaling in promoting the inductive capability of CD133+ DP cells for in vitro expansion and in vivo hair follicle regeneration, which could potentially be applied to cultured human DP cells. PMID:27312243

  7. Human Immunodeficiency Syndromes Affecting Human Natural Killer Cell Cytolytic Activity

    PubMed Central

    Ham, Hyoungjun; Billadeau, Daniel D.

    2013-01-01

    Natural killer (NK) cells are lymphocytes of the innate immune system that secrete cytokines upon activation and mediate the killing of tumor cells and virus-infected cells, especially those that escape the adaptive T cell response caused by the down regulation of MHC-I. The induction of cytotoxicity requires that NK cells contact target cells through adhesion receptors, and initiate activation signaling leading to increased adhesion and accumulation of F-actin at the NK cell cytotoxic synapse. Concurrently, lytic granules undergo minus-end directed movement and accumulate at the microtubule-organizing center through the interaction with microtubule motor proteins, followed by polarization of the lethal cargo toward the target cell. Ultimately, myosin-dependent movement of the lytic granules toward the NK cell plasma membrane through F-actin channels, along with soluble N-ethylmaleimide-sensitive factor attachment protein receptor-dependent fusion, promotes the release of the lytic granule contents into the cleft between the NK cell and target cell resulting in target cell killing. Herein, we will discuss several disease-causing mutations in primary immunodeficiency syndromes and how they impact NK cell-mediated killing by disrupting distinct steps of this tightly regulated process. PMID:24478771

  8. EFFECT OF NICKEL AND MANGANESE ON NATURAL KILLER CELL ACTIVITY

    EPA Science Inventory

    A single intramuscular injection of NiCl2 causes a suppression of natural killer (NK) cell activity, while a single injection of MnCl2 enhances NK activity. When injected together Mn preempts the suppressive effect of Ni on NK activity.

  9. Evidence of a retinoid signaling alteration involving the activator protein 1 complex in tumorigenic human bronchial epithelial cells and non-small cell lung cancer cells.

    PubMed

    Lee, H Y; Dawson, M I; Claret, F X; Chen, J D; Walsh, G L; Hong, W K; Kurie, J M

    1997-03-01

    Retinoids, including retinol and retinoic acid derivatives, inhibit the growth of normal human bronchial epithelial (HBE) cells. Using a lung carcinogenesis model consisting of normal, immortalized, and tumorigenic HBE cells, we showed previously that, compared to normal HBE cells, the tumorigenic HBE cell line 11701 is resistant to the growth-inhibitory effects of all-trans-retinoic acid (t-RA). Retinoid receptor function is preserved in tumorigenic 11701 cells, suggesting that other retinoid signaling components are altered. The activator protein 1 (AP-1) complex is a component of the retinoid signaling pathway and has demonstrated importance in cellular growth and differentiation. Therefore, we investigated whether AP-1 is involved in a retinoid signaling defect in tumorigenic 11701 cells and in retinoid-resistant non-small cell lung cancer (NSCLC) cell lines. We found that t-RA treatment inhibited AP-1 transcriptional activity in normal HBE cells but not in tumorigenic 11701 cells nor in the NSCLC cell lines Calu-1, Calu-6, SKMES-1, and ChaGo K1. We sought mechanisms for this retinoid signaling alteration involving AP-1 in tumorigenic 11701 cells. Basal AP-1 transcriptional activity; AP-1 DNA-binding activity; and the mRNA levels of c-fos, the AP-1 coactivator Jun activation domain-binding protein 1, and the retinoid receptor corepressor, the silencing mediator for retinoid and thyroid hormone receptors (SMRT), were lower in tumorigenic 11701 cells than in normal HBE cells. Transient transfection of tumorigenic 11701 cells with c-fos or CREB binding protein, which is a coactivator of AP-1 and retinoid receptors, enhanced basal AP-1 transcriptional activity but did not alter the effects of t-RA on AP-1 transcriptional activity. These findings provide evidence of a retinoid signaling alteration involving AP-1 in tumorigenic 11701 and NSCLC cells. Furthermore, the inhibitory effect of t-RA on AP-1 transcriptional activity was not restored in tumorigenic 11701

  10. Microchamber Device for Detection of Transporter Activity of Adherent Cells

    PubMed Central

    Tsugane, Mamiko; Uejima, Etsuko; Suzuki, Hiroaki

    2015-01-01

    We present a method to detect the transporter activity of intact adherent cells using a microchamber device. When adherent cells are seeded onto the poly-di-methyl siloxane substrate having microchambers with openings smaller than the size of a cell, the cells form a confluent layer that covers the microchambers, creating minute, confined spaces. As substances exported across the cell membrane accumulate, transporter activity can be detected by observing the fluorescence intensity increase in the microchamber. We tested the microchamber device with HeLa cells over-expressing MDR1, an ATP-binding cassette transporter, and succeeded in detecting the transport of fluorescence-conjugated paclitaxel, the anti-cancer drug, at the single-cell level. PMID:25853126

  11. ACTIVATION ASSAY FOR PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR- ALPHA (PPARÁ) BY PERFLUOROALKYL ACIDS (PFAAS) IN COS-1 CELLS

    EPA Science Inventory

    PFAAs have been found to elicit various physiological effects including peroxisome proliferation, indicating the mechanism of action for these chemicals could involve PPAR. This study investigates the ability of PFAAs to bind and activate mouse and human PPARα in COS-1 cell...

  12. Peptide mini-scaffold facilitates JNK3 activation in cells

    PubMed Central

    Zhan, Xuanzhi; Stoy, Henriette; Kaoud, Tamer S.; Perry, Nicole A.; Chen, Qiuyan; Perez, Alejandro; Els-Heindl, Sylvia; Slagis, Jack V.; Iverson, Tina M.; Beck-Sickinger, Annette G.; Gurevich, Eugenia V.; Dalby, Kevin N.; Gurevich, Vsevolod V.

    2016-01-01

    Three-kinase mitogen-activated protein kinase (MAPK) signaling cascades are present in virtually all eukaryotic cells. MAPK cascades are organized by scaffold proteins, which assemble cognate kinases into productive signaling complexes. Arrestin-3 facilitates JNK activation in cells, and a short 25-residue arrestin-3 peptide was identified as the critical JNK3-binding element. Here we demonstrate that this peptide also binds MKK4, MKK7, and ASK1, which are upstream JNK3-activating kinases. This peptide is sufficient to enhance JNK3 activity in cells. A homologous arrestin-2 peptide, which differs only in four positions, binds MKK4, but not MKK7 or JNK3, and is ineffective in cells at enhancing activation of JNK3. The arrestin-3 peptide is the smallest MAPK scaffold known. This peptide or its mimics can regulate MAPKs, affecting cellular decisions to live or die. PMID:26868142

  13. Twin Knudsen Cell Configuration for Activity Measurements by Mass Spectrometry

    NASA Technical Reports Server (NTRS)

    Jacobson, N. S.

    1996-01-01

    A twin Knudsen cell apparatus for alloy activity measurements by mass spectrometry is described. Two Knudsen cells - one containing an alloy and one containing a pure component - are mounted on a single flange and translated into the sampling region via a motorized x-y table. Mixing of the molecular beams from the cells is minimized by a novel system of shutters. Activity measurements were taken on two well-characterized alloys to verify the operation of the system. Silver activity measurements are reported for Ag-Cu alloys and aluminum activity measurements are reported for Fe-Al alloys. The temperature dependence of activity for a 0.474 mol fraction Al-Fe alloy gives a partial molar heat of aluminum. Measurements taken with the twin cell show good agreement with literature values for these alloys.

  14. EspC, an Autotransporter Protein Secreted by Enteropathogenic Escherichia coli, Causes Apoptosis and Necrosis through Caspase and Calpain Activation, Including Direct Procaspase-3 Cleavage

    PubMed Central

    Serapio-Palacios, Antonio

    2016-01-01

    ABSTRACT Enteropathogenic Escherichia coli (EPEC) has the ability to antagonize host apoptosis during infection through promotion and inhibition of effectors injected by the type III secretion system (T3SS), but the total number of these effectors and the overall functional relationships between these effectors during infection are poorly understood. EspC produced by EPEC cleaves fodrin, paxillin, and focal adhesion kinase (FAK), which are also cleaved by caspases and calpains during apoptosis. Here we show the role of EspC in cell death induced by EPEC. EspC is involved in EPEC-mediated cell death and induces both apoptosis and necrosis in epithelial cells. EspC induces apoptosis through the mitochondrial apoptotic pathway by provoking (i) a decrease in the expression levels of antiapoptotic protein Bcl-2, (ii) translocation of the proapoptotic protein Bax from cytosol to mitochondria, (iii) cytochrome c release from mitochondria to the cytoplasm, (iv) loss of mitochondrial membrane potential, (v) caspase-9 activation, (vi) cleavage of procaspase-3 and (vii) an increase in caspase-3 activity, (viii) PARP proteolysis, and (ix) nuclear fragmentation and an increase in the sub-G1 population. Interestingly, EspC-induced apoptosis was triggered through a dual mechanism involving both independent and dependent functions of its EspC serine protease motif, the direct cleavage of procaspase-3 being dependent on this motif. This is the first report showing a shortcut for induction of apoptosis by the catalytic activity of an EPEC protein. Furthermore, this atypical intrinsic apoptosis appeared to induce necrosis through the activation of calpain and through the increase of intracellular calcium induced by EspC. Our data indicate that EspC plays a relevant role in cell death induced by EPEC. PMID:27329750

  15. Cytotoxic activity of allogeneic natural killer cells on U251 glioma cells in vitro.

    PubMed

    Guo, Meng; Wu, Tingting; Wan, Lixin

    2016-07-01

    The present study aimed to observe the cytotoxic activity of allogeneic natural killer (NK) cells on U251 glioma cells and to investigate their mechanism of action to establish an effective treatment strategy for neuroglioma. Cell survival curves, colony formation assays and karyotype analysis were performed to investigate the characteristics of U251 glioma cells. The present study demonstrated that natural killer group 2, member D (NKG2D)‑major histocompatibility complex class I‑related chain A/B (MICA/B) interactions contributed to the cytotoxic effect of NK cells on K562 and U251 cells. In antibody‑blocking assays to inhibit NKG2D ligands, the cytotoxic activity was not completely attenuated, which suggested that other signaling pathways contribute to the cytotoxic activity of NK cells on tumor cells in addition to the NKG2D‑mediated activity. The present study identified that the expression levels of NKG2D ligands on the surface of target cells influenced the strength of the NK cell immune response. Furthermore, allogeneic NK cells were observed to kill glioma cells in vitro, and this anticancer activity is associated with the rate of NKG2D expression on the surface of glioma cells. PMID:27175912

  16. Active Cellular Mechanics and Information Processing in the Living Cell

    NASA Astrophysics Data System (ADS)

    Rao, M.

    2014-07-01

    I will present our recent work on the organization of signaling molecules on the surface of living cells. Using novel experimental and theoretical approaches we have found that many cell surface receptors are organized as dynamic clusters driven by active currents and stresses generated by the cortical cytoskeleton adjoining the cell surface. We have shown that this organization is optimal for both information processing and computation. In connecting active mechanics in the cell with information processing and computation, we bring together two of the seminal works of Alan Turing.

  17. The effects of selenium on glutathione peroxidase activity and radioprotection in mammalian cells

    SciTech Connect

    Diamond, A.M.; Murray, J.L.; Dale, P.; Tritz, R.; Grdina, D.J.

    1995-09-05

    The media of representative mammalian cell lines were supplemented with low levels of selenium in the form of sodium selenite in order to investigate the effects of selenium on mammalian cells. Following incubation in 30 nM sodium selenite, these cells were assayed for changes in glutathione peroxidase (GPx) activity. The cells examined included NIH 3T3 mouse fibroblasts, PC12 rat sympathetic precursor cells, SupT-1 human lymphocytes, MCF-7{sup adr} human breast carcinoma cells and AA8 Chinese hamster ovary cells. Selenium supplementation resulted in a marginal increase in GPx activity for the NIH 3T3, MCF-7{sup adr} and Supt-1 cells but stimulated GPx activity approximately 5-fold in PC12 and AA8 cells. AA8 cells were selected to evaluate whether selenium supplementation was radioprotective against {sup 60}cobalt gamma irradiation. Protection against radiation-induced mutation was measured by evaluating mutation frequency at the hprt locus. In this assay, preincubation of AA8 CHO cells significantly protected these cells from exposure to 8 Gy.

  18. Mapping the polarity and stimulus density requirements for T-cell activation

    NASA Astrophysics Data System (ADS)

    Wei, Xunbin; Krasieva, Tatiana B.; Zhang, Zhanxiang; Negulescu, Paul A.; Sun, Chung-Ho; Berns, Michael W.; Cahalan, Michael D.; Tromberg, Bruce J.

    1998-08-01

    T-cell contact with antigen-presenting cells (APC) initiates an activation cascade which includes an increase in T-cell intracellular calcium [(Ca2+)i] and leads to T-cell proliferation and differentiation. Although T-cell/APC physical contact is required for an immune response, little is known about the patterns of cellular interaction and their relation to activation. We have combined fluorescence spectroscopy and imaging with optical manipulation to investigate the contact requirements for T-cell activation, using optical tweezers to control the orientation of T- cell/APC pairs and fluorescence microscopy to measure the subsequent (Ca2+)i response, detected as an emission shift from the combination of fura-red and oregon- green, two cytoplasmic (Ca2+) indicators. APCs or beads coated with antibodies to the T-cell receptor (TCR) are trapped with a near-infrared titanium-sapphire laser and placed at different locations along the T-cell, which has a polarized appearance defined by the shape and direction of crawling (2-5 micrometers /min). T cells contacted with antigen- presenting cells or antibody-coated beads entered a dynamic and reproducible program in the first 10 - 20 mins, including (Ca2+)i increase, changes in shape and motility, engulfment, and stable contact. T cells presented with antigen at the leading edge had a higher probability of responding (85%) and a shorter latency of response (50 secs) than those contacting APCs or beads with their trailing end (APCs: 30%, 150 secs; beads: 6%, 300 secs). Alterations in antibody density, quantified by FACS analysis, and bead size were used to determine the spatial requirements for T cell activation and the minimum number of receptors which must be engaged in order to transmit a positive signal. Preliminary data show that T cell responses [response percentage, latency and (Ca2+)i pattern] depend on both antibody density and bead size.

  19. Aldehyde dehydrogenase activity promotes survival of human muscle precursor cells

    PubMed Central

    Jean, Elise; Laoudj-Chenivesse, Dalila; Notarnicola, Cécile; Rouger, Karl; Serratrice, Nicolas; Bonnieu, Anne; Gay, Stéphanie; Bacou, Francis; Duret, Cédric; Carnac, Gilles

    2011-01-01

    Abstract Aldehyde dehydrogenases (ALDH) are a family of enzymes that efficiently detoxify aldehydic products generated by reactive oxygen species and might therefore participate in cell survival. Because ALDH activity has been used to identify normal and malignant cells with stem cell properties, we asked whether human myogenic precursor cells (myoblasts) could be identified and isolated based on their levels of ALDH activity. Human muscle explant-derived cells were incubated with ALDEFLUOR, a fluorescent substrate for ALDH, and we determined by flow cytometry the level of enzyme activity. We found that ALDH activity positively correlated with the myoblast-CD56+ fraction in those cells, but, we also observed heterogeneity of ALDH activity levels within CD56-purified myoblasts. Using lentiviral mediated expression of shRNA we demonstrated that ALDH activity was associated with expression of Aldh1a1 protein. Surprisingly, ALDH activity and Aldh1a1 expression levels were very low in mouse, rat, rabbit and non-human primate myoblasts. Using different approaches, from pharmacological inhibition of ALDH activity by diethylaminobenzaldehyde, an inhibitor of class I ALDH, to cell fractionation by flow cytometry using the ALDEFLUOR assay, we characterized human myoblasts expressing low or high levels of ALDH. We correlated high ALDH activity ex vivo to resistance to hydrogen peroxide (H2O2)-induced cytotoxic effect and in vivo to improved cell viability when human myoblasts were transplanted into host muscle of immune deficient scid mice. Therefore detection of ALDH activity, as a purification strategy, could allow non-toxic and efficient isolation of a fraction of human myoblasts resistant to cytotoxic damage. PMID:19840193

  20. Enzyme Activities in Polarized Cell Membranes

    PubMed Central

    Bass, L.; McIlroy, D. K.

    1968-01-01

    The theoretical pH dependence of enzyme activities in membranes of low dielectric constant is estimated. It is shown that in biological membranes some types of enzymes may attain a limiting pH sensitivity such that an increment of only 0.2 pH unit (sufficient to induce action potentials in squid axons) causes a relative activity change of over 25%. The transients of enzyme activity generated by membrane depolarization and by pH increments in the bathing solution are discussed in relation to the transients of nervous excitation. PMID:5641405

  1. Activated microglia cause reversible apoptosis of pheochromocytoma cells, inducing their cell death by phagocytosis

    PubMed Central

    Hornik, Tamara C.; Vilalta, Anna; Brown, Guy C.

    2016-01-01

    ABSTRACT Some apoptotic processes, such as phosphatidylserine exposure, are potentially reversible and do not necessarily lead to cell death. However, phosphatidylserine exposure can induce phagocytosis of a cell, resulting in cell death by phagocytosis: phagoptosis. Phagoptosis of neurons by microglia might contribute to neuropathology, whereas phagoptosis of tumour cells by macrophages might limit cancer. Here, we examined the mechanisms by which BV-2 microglia killed co-cultured pheochromocytoma (PC12) cells that were either undifferentiated or differentiated into neuronal cells. We found that microglia activated by lipopolysaccharide rapidly phagocytosed PC12 cells. Activated microglia caused reversible phosphatidylserine exposure on and reversible caspase activation in PC12 cells, and caspase inhibition prevented phosphatidylserine exposur and decreased subsequent phagocytosis. Nitric oxide was necessary and sufficient to induce the reversible phosphatidylserine exposure and phagocy