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Sample records for modified gec cell

  1. Temperature Measurement for Dust Particles in a GEC Reference Cell

    NASA Astrophysics Data System (ADS)

    Kong, Jie; Qiao, Ke; Matthews, Lorin; Hyde, Truell

    2016-10-01

    The thermal motion of a dust particle levitated in a plasma chamber is similar to that described by Brownian motion in many ways. The primary differences between a dust particle in a plasma system and a free Brownian particle is that in addition to the random collisions between the dust particle and the neutral gas atoms, there are electric field fluctuations, dust charge fluctuations, and correlated motions from unwanted continuous signals originating within the plasma system itself. Correlated motion cannot be qualified as random motion, and therefore should not be included in a measurement of the dust temperature. In this presentation, we discuss how to separate random and coherent motion of a dust particle confined in a glass box within a GEC radio frequency reference cell. Dust particle fluctuation data are obtained experimentally and analyzed using the mean square displacement and other techniques, and temperatures obtained by various methods are compared. NSF / DOE funding is gratefully acknowledged - PHY1414523 & PHY1262031.

  2. Diode Laser Measurements in an Inductively Coupled GEC Cell for Oxide Etching

    NASA Astrophysics Data System (ADS)

    Perry, Lee; Deering, Glen; Koltunski, Laure; Anderson, Harold

    1998-10-01

    Diode laser absorption measurements have been made on CF, CF2 and CO radicals in an inductively coupled GEC reference cell. The GEC reference cell was modified with a quartz confinement ring around the source region to stabilize the plasma. Optical emission and Langmuir probe studies indicated this modification resulted in fluorocarbon discharges with a plasma chemistry similar to that found in commercial etch tools. The experiments in this study focused on radical concentrations found in the reactor under typical high-density plasma etching conditions. In a 10 mTorr C_2F6 discharge at 300 W source power and 100 W bias power, etching proceeded at about 5000 Åmin-1. A range of source power and bias power conditions, from 100 to 400 W and from 0 to 130 W, respectively, was employed. The time evolution of CF, CF2 and CO in a C_2F6 plasma was monitored during an approximate 2 minute etch cycle. Chamber cleanliness and bias was found to exert a strong influence on radical densities. The data is expected to provide an important database for models of oxide etching in inductively coupled plasma tools. (This work has been supported by SEMATECH)

  3. Langmuir Probe Measurements in an Inductively Coupled GEC Reference Cell Plasma

    NASA Technical Reports Server (NTRS)

    Ji, J. S.; Kim, J. S.; Cappelli, M. A.; Sharma, S. P.; Arnold, J. O. (Technical Monitor)

    1998-01-01

    Measurements of electron number density, electron temperature, and electron energy distribution function (EEDF) using a compensated Langmuir probe have been performed on an inductively (transformer ) coupled Gaseous Electronics Conference (GEC) reference cell plasma. The plasma source is operated with CH4, CF4, or their mixtures with argon. The effect of independently driving the electrode supporting the wafer on the probe data is studied. In particular, we find that the plasma structure depends on the phase in addition to the magnitude of the power coupled to the electrode relative to that of the transformer coil. The Langmuir probe is translated in a plane parallel to the electrode to investigate the spatial structure of the plasma. The probe data is also compared with fluid model predictions.

  4. Temperature measurement of a dust particle in a RF plasma GEC reference cell

    NASA Astrophysics Data System (ADS)

    Kong, Jie; Qiao, Ke; Matthews, Lorin S.; Hyde, Truell W.

    2016-10-01

    The thermal motion of a dust particle levitated in a plasma chamber is similar to that described by Brownian motion in many ways. The primary difference between a dust particle in a plasma system and a free Brownian particle is that in addition to the random collisions between the dust particle and the neutral gas atoms, there are electric field fluctuations, dust charge fluctuations, and correlated motions from the unwanted continuous signals originating within the plasma system itself. This last contribution does not include random motion and is therefore separable from the random motion in a `normal' temperature measurement. In this paper, we discuss how to separate random and coherent motions of a dust particle confined in a glass box in a Gaseous Electronic Conference (GEC) radio-frequency (RF) reference cell employing experimentally determined dust particle fluctuation data analysed using the mean square displacement technique.

  5. Fourier Transform Infrared Spectroscopy of CF4 on the GEC Reference Cell

    NASA Technical Reports Server (NTRS)

    Rao, M. V. V. S.; Sharma, S. P.; Meyyappan, M.; Cruden, Brett A.; Arnold, Jim (Technical Monitor)

    2001-01-01

    Fourier Transform Infrared Spectroscopy (FTIR) has been used to characterize inductively coupled CF4 plasmas in a GEC Reference Cell in-situ In examining these FTIR spectra, several assumptions and approximations of FTIR analysis are addressed. This includes the density dependence of cross-sections, non-linear effects in the addition of overlapping bands and the effect of spatial variations in density and temperature, This analysis demonstrates that temperatures extracted from MR spectra may provide a poor estimate of the true neutral plasma temperature. The FTIR spectra are dominated by unreacted CF, accounting for 40-60% of the gas products. The amount of CF4 consumption is found to have a marked dependence on power, and is nearly independent of pressure in the range of 10-50 mtorr. Small amounts of C2F6 are observed at low power. Also observed are etching products from the quartz window SiF4 COF2 and CO which occur in approximately equal ratios and together account for less than 10% of the gas. The concentrations of these species are nearly independent of pressure. CFx radicals are below the detection limit of this apparatus (approx. 1012/cc).

  6. GEC-ESTRO recommendations for brachytherapy for head and neck squamous cell carcinomas.

    PubMed

    Mazeron, Jean-Jacques; Ardiet, Jean-Michel; Haie-Méder, Christine; Kovács, György; Levendag, Peter; Peiffert, Didier; Polo, Alfredo; Rovirosa, Angels; Strnad, Vratislav

    2009-05-01

    Both primary and recurrent squamous cell carcinoma of the head and neck are classic indications for brachytherapy. A high rate of local tumor control at the cost of limited morbidity can be achieved with brachytherapy through good patient selection, meticulous source implantation and careful treatment planning. However, no randomized trials have been performed, and there is scant evidence in the literature especially regarding practical clinical recommendations for brachytherapy for head and neck subsites. The Head and Neck Working Group of the European Brachytherapy Group (Groupe Européen de Curiethérapie-European Society for Therapeutic Radiology and Oncology (GEC-ESTRO) therefore decided to formulate the present consensus recommendations for low-dose rate, pulsed-dose rate and high-dose rate brachytherapy. The use of brachytherapy in combination with external beam radiotherapy and/or surgery is also covered as well as the use of brachytherapy in previously irradiated patients. Given the paucity of evidence in the literature, these recommendations are mainly based on clinical experience accumulated by the members of the working group over several decades and the respective publications. The recommendations cover in a general part (1) patient selection, the pre-treatment work up and patient care, (2) treatment strategy, (3) target definition, (4) implant techniques, (5) dose and dose rate prescription, (6) treatment planning and reporting, (7) treatment monitoring (8) catheter removal, and (9) post-treatment patient care and follow-up. The recommendations are then specified for the classical brachytherapy tumor sites following an analogue more focussed structure (patient selection, implant technique, target definition, dose and dose rate prescription, results): lip, oral mucosa, mobile tongue, floor of mouth, oropharynx, nasopharynx, paranasal sinuses.

  7. Measurement of the Interaction Force between Dust Particles within a Glass Box in a GEC RF Reference Cell

    NASA Astrophysics Data System (ADS)

    Chen, Mudi; Kong, Jie; Qiao, Ke; Carmona-Reyes, Jorge; Harris, Brandon; Matthews, Lorin; Hyde, Truell

    2013-10-01

    A wide variety of structural states (for example, Coulomb balls, one-dimensional vertically aligned dust particle chains, helical dust structures, etc.) have been observed for dust particles confined within a glass box placed on the lower electrode of a GEC rf reference cell. Both the interparticle interaction force and the confinement force play important roles in the formation of these structures. Unfortunately, since the exact nature of the confinement force produced by the walls of the glass box is yet unclear, it is difficult to differentiate between the effects produced by the interparticle interaction force and the effects created by the confinement force. In this experiment, a free-falling dust particle in the box acts as an in-situ probe, providing information on the structure of the confinement force. It will be shown that the data provided by this mapping procedure allows the interaction force between particles within various dust particle structures to be measured through perturbation of individual particles employing a diode pumped solid state laser (Coherent VERDI).

  8. GEC Plasma Data Exchange Project

    NASA Astrophysics Data System (ADS)

    Pitchford, L. C.

    2013-08-01

    In 2010 the Gaseous Electronics Conference (GEC), a major international conference for the low temperature plasma science (LTPS) community, initiated the Plasma Data Exchange Project (PDEP). The PDEP is an informal, community-based project that aims to address, at least in part, the well-recognized needs for the community to organize the means of collecting, evaluating and sharing data both for modelling and for interpretation of experiments. The emphasis to date in the PDEP has been on data related to the electron and ion components of these plasmas rather than on the plasma chemistry. At the heart of the PDEP is the open-access website, LXCat [1], developed by researchers at LAPLACE (Laboratoire Plasma et Conversion d'Energie, Toulouse, France). LXCat is a platform for archiving and manipulating collections of data related to electron scattering and transport in cold, neutral gases, organized in databases set-up by individual members or institutions of the LTPS community. At present, 15 databases of electron scattering data, contributed by groups around the world, can be accessed on LXCat. These databases include complete sets of electron cross sections, over an energy range from thermal to nominally 1 keV, for almost 40 ground-state neutral species and partial sets of data for about 30 other neutral, excited and ionized species. 'Complete' implies that all the major electron momentum and energy loss processes are well described in the dataset. Such 'complete' datasets can be used as input to a Boltzmann calculation of the electron energy distribution function (generally non-Maxwellian), and electron transport and rate coefficients can be obtained in pure gases or mixtures by averaging over the distribution function. Online tools enable importing and exporting data, plotting and comparing different sets of data. An online version of the Boltzmann equation solver BOLSIG+ [2] is also available on the LXCat site. Other members of the community have contributed their

  9. GEC-ESTRO ACROP recommendations for head & neck brachytherapy in squamous cell carcinomas: 1st update - Improvement by cross sectional imaging based treatment planning and stepping source technology.

    PubMed

    Kovács, György; Martinez-Monge, Rafael; Budrukkar, Ashwini; Guinot, Jose Luis; Johansson, Bengt; Strnad, Vratislav; Skowronek, Janusz; Rovirosa, Angeles; Siebert, Frank-André

    2017-02-01

    The Head and Neck Working Group of the GEC-ESTRO (Groupe Européen de Curiethérapie - European Society for Therapeutic Radiology and Oncology) published in 2009 the consensus recommendations for low-dose rate, pulsed-dose rate and high-dose rate brachytherapy in head & neck cancers. The use of brachytherapy in combination with external beam radiotherapy and/or surgery was also covered as well as the use of brachytherapy in previously irradiated patients. Given the developments in the field, these recommendations needed to be updated to reflect up-to-date knowledge. The present update does not repeat basic knowledge which was published in the first recommendation but covers in a general part developments in (1) dose and fractionation, (2) aspects of treatment selection for brachytherapy alone versus combined BT+EBRT and (3) quality assurance issues. Detailed expert committee opinion intends to help the clinical practice in lip-, oral cavity-, oropharynx-, nasopharynx-, and superficial cancers. Different aspects of adjuvant treatment techniques and their results are discussed, as well the possibilities of salvage brachytherapy applications.

  10. GEC Alsthom diesel applications in Far East

    SciTech Connect

    Mullins, P.

    1996-07-01

    Recent achievements in the Far East for GEC Alsthom Diesels follow a drive to extend its market presence in these fast-growing markets. Ruston has supplied Samsung Engineering Co. with three medium-speed 16RK270 diesel engines for base-load generating sets. Paxman has won a contract to re-engine four locomotives for Sri Lankan Government Railways, as well as supplying six of its latest VPI85 high-speed diesels for new Taiwanese fast petrol vessels. This paper describes briefly the specifications of these diesels.

  11. Orbit Optimization for the Geospace Electrodynamics Connections (GEC) Mission

    NASA Technical Reports Server (NTRS)

    Mesarch, Michael A.

    2003-01-01

    The Geospace Electrodynamics Connections (GEC) mission plan is to launch multiple spacecraft to perform in-situ atmospheric science in the lower ionosphere. There is limited experience in this low altitude region with the Atmospheric Explorer-C (AE-C) being the last spacecraft to explore this region in 1973. AE-C flew an eccentric orbit using maneuvers to lower its perigee to near 130 km at various times during its mission. GEC will advance the science performed by AE-C by performing multiple low-perigee, atmospheric dipping campaigns for extended durations. AE-C kept its perigee near 130 km for only a total of roughly 1 day. Furthermore, GEC plans to carry a more diverse suite of instruments and will be able to capture different temporal and spatial phenomena through the use of multiple spacecraft flying in a string of pearls formation. The mission analysis for GEC has been broken into two parts: the analysis of the parking orbit with the dipping campaigns and the examination of the multi-satellite dynamics of the GEC constellation. The analysis described in this paper examines the capability to meet the requirements necessary to support the 10 dipping campaigns using a single spacecraft as a representative of all three in the constellation. Further analysis is being performed to analyze the multi-satellite nature of the GEC mission.

  12. GEC-derived SFRP5 inhibits Wnt5a-induced macrophage chemotaxis and activation.

    PubMed

    Zhao, Chenghai; Bu, Xianmin; Wang, Wei; Ma, Tingxian; Ma, Haiying

    2014-01-01

    Aberrant macrophage infiltration and activation has been implicated in gastric inflammation and carcinogenesis. Overexpression of Wnt5a and downregulation of SFRP5, a Wnt5a antagonist, were both observed in gastric cancers recently. This study attempted to explore whether Wnt5a/SFRP5 axis was involved in macrophage chemotaxis and activation. It was found that both Wnt5a transfection and recombinant Wnt5a (rWnt5a) treatment upregulated CCL2 expression in macrophages, involving JNK and NFκB signals. Conditioned medium from Wnt5a-treated macrophages promoted macrophage chemotaxis mainly dependent on CCL2. SFRP5 from gastric epithelial cells (GECs) inhibited Wnt5a-induced CCL2 expression and macrophage chemotaxis. In addition, Wnt5a treatment stimulated macrophages to produce inflammatory cytokines and COX-2/PGE2, which was also suppressed by SFRP5 from GECs. These results demonstrate that Wnt5a induces macrophage chemotaxis and activation, which can be blocked by GEC-derived SFRP5, suggesting that Wnt5a overproduction and SFRP5 deficiency in gastric mucosa may together play an important role in gastric inflammation and carcinogenesis.

  13. [Therapeutic approaches using genetically modified cells].

    PubMed

    Anliker, Brigitte; Renner, Matthias; Schweizer, Matthias

    2015-11-01

    Medicinal products containing genetically modified cells are, in most cases, classified as gene therapy and cell therapy medicinal products. Although no medicinal product containing genetically modified cells has been licensed in Europe yet, a variety of therapeutic strategies using genetically modified cells are in different stages of clinical development for the treatment of acquired and inherited diseases. In this chapter, several examples of promising approaches are presented, with an emphasis on gene therapy for inherited immunodeficiencies and on tumour immunotherapy with genetically modified T-cells expressing a chimeric antigen receptor or a recombinant T-cell receptor.

  14. Blade System Design Study. Part II, final project report (GEC).

    SciTech Connect

    Griffin, Dayton A.

    2009-05-01

    As part of the U.S. Department of Energy's Low Wind Speed Turbine program, Global Energy Concepts LLC (GEC)1 has studied alternative composite materials for wind turbine blades in the multi-megawatt size range. This work in one of the Blade System Design Studies (BSDS) funded through Sandia National Laboratories. The BSDS program was conducted in two phases. In the Part I BSDS, GEC assessed candidate innovations in composite materials, manufacturing processes, and structural configurations. GEC also made recommendations for testing composite coupons, details, assemblies, and blade substructures to be carried out in the Part II study (BSDS-II). The BSDS-II contract period began in May 2003, and testing was initiated in June 2004. The current report summarizes the results from the BSDS-II test program. Composite materials evaluated include carbon fiber in both pre-impregnated and vacuum-assisted resin transfer molding (VARTM) forms. Initial thin-coupon static testing included a wide range of parameters, including variation in manufacturer, fiber tow size, fabric architecture, and resin type. A smaller set of these materials and process types was also evaluated in thin-coupon fatigue testing, and in ply-drop and ply-transition panels. The majority of materials used epoxy resin, with vinyl ester (VE) resin also used for selected cases. Late in the project, testing of unidirectional fiberglass was added to provide an updated baseline against which to evaluate the carbon material performance. Numerous unidirectional carbon fabrics were considered for evaluation with VARTM infusion. All but one fabric style considered suffered either from poor infusibility or waviness of fibers combined with poor compaction. The exception was a triaxial carbon-fiberglass fabric produced by SAERTEX. This fabric became the primary choice for infused articles throughout the test program. The generally positive results obtained in this program for the SAERTEX material have led to its being

  15. Chemically modified graphite for electrochemical cells

    DOEpatents

    Greinke, Ronald Alfred; Lewis, Irwin Charles

    1998-01-01

    This invention relates to chemically modified graphite particles: (a) that are useful in alkali metal-containing electrode of a electrochemical cell comprising: (i) the electrode, (ii) a non-aqueous electrolytic solution comprising an organic aprotic solvent which solvent tends to decompose when the electrochemical cell is in use, and an electrically conductive salt of an alkali metal, and (iii) a counterelectrode; and (b) that are chemically modified with fluorine, chlorine, iodine or phosphorus to reduce such decomposition. This invention also relates to electrodes comprising such chemically modified graphite and a binder and to electrochemical cells containing such electrodes.

  16. Chemically modified graphite for electrochemical cells

    DOEpatents

    Greinke, R.A.; Lewis, I.C.

    1998-05-26

    This invention relates to chemically modified graphite particles: (a) that are useful in alkali metal-containing electrode of a electrochemical cell comprising: (1) the electrode, (2) a non-aqueous electrolytic solution comprising an organic aprotic solvent which solvent tends to decompose when the electrochemical cell is in use, and an electrically conductive salt of an alkali metal, and (3) a counter electrode; and (b) that are chemically modified with fluorine, chlorine, iodine or phosphorus to reduce such decomposition. This invention also relates to electrodes comprising such chemically modified graphite and a binder and to electrochemical cells containing such electrodes. 3 figs.

  17. Understanding Plasma Interactions with the Atmosphere: The Geospace Electrodynamic Connections (GEC) Mission

    NASA Technical Reports Server (NTRS)

    2001-01-01

    The Geospace Electrodynamic Connections (GEC) mission is a multispacecraft Solar-Terrestrial Probe that has been specifically designed to advance the level of physical insight of our understanding of the coupling among the ionosphere, thermosphere, and magnetosphere. GEC is NASA's fifth Solar-Terrestrial Probe. Through multipoint measurements in the Earth's ionosphere-thermosphere (I-T) system, GEC will (i) discover the spatial and temporal scales on which magnetospheric energy input into the I-T region occurs, (ii) determine the spatial and temporal scales for the response of the I-T system to this input of energy, and (iii) quantify the altitude dependence of the response.

  18. Modified host cells with efflux pumps

    DOEpatents

    Dunlop, Mary J.; Keasling, Jay D.; Mukhopadhyay, Aindrila

    2016-08-30

    The present invention provides for a modified host cell comprising a heterologous expression of an efflux pump capable of transporting an organic molecule out of the host cell wherein the organic molecule at a sufficiently high concentration reduces the growth rate of or is lethal to the host cell.

  19. Cell microarrays on photochemically modified polytetrafluoroethylene.

    PubMed

    Mikulikova, Regina; Moritz, Sieglinde; Gumpenberger, Thomas; Olbrich, Michael; Romanin, Christoph; Bacakova, Lucie; Svorcik, Vaclav; Heitz, Johannes

    2005-09-01

    We studied the adhesion, proliferation, and viability of human umbilical vein endothelial cells (HUVEC) and human embryonic kidney cells (HEK) on modified spots at polytetrafluoroethylene (PTFE) surfaces. The viability of the cells was assessed using an aqueous non-radioactive cell proliferation assay. Round spots with a diameter of 100 microm were modified by exposure to the ultraviolet (UV) light of a Xe(2)(*)-excimer lamp at a wavelength of 172 nm in an ammonia atmosphere employing a contact mask. The spots were arranged in a quadratic pattern with 300 microm center-to-center spot distances. With optimized degree of modification, the cells adhered to the modified spots with a high degree of selectivity (70-90%). The adhered cells on the spots proliferated. This resulted in a significant increase in the number of adhering HUVECS or HEK cells after seeding and in the formation of confluent cell clusters after 3-4 days. With higher start seeding density, these clusters were not only confined to the modified spots but extended several micrometer to the neighborhood. The high potential of the cell microarrays for gene analysis in living cells was demonstrated with HEK cells transfected by yellow fluorescent protein (YFP).

  20. Orbit Optimization For The Geospace Electrodynamics Connections (GEC) Mission

    NASA Technical Reports Server (NTRS)

    Mesarch, Michael A.

    2004-01-01

    Part of NASA's Solar Terrestrial Probe line of missions, the Geospace Electrodynamics Connections (GEC) mission will deploy a formation of three spacecraft to perform in-situ atmospheric research in the low Ionosphere-Thermosphere region. These spacecraft will fly together in a %tring-of-pearls formation with variable spacings ranging from 10 seconds to one-quarter of an orbit at perigee. Over the course of its two-year mission, the three spacecraft will perform ten, 1-week dipping campaigns whereby they maneuver to lower their perigee to near 134 km. Using available launch vehicle performance data, an optimal parking orbit of 222 x 1525 km was found to maximize the dry mass available while providing enough propellant to perform the ten deep-dipping campaigns over its two-year mission. The results were used to create multi-variable contour plots containing the orbit perigee, the orbit apogee, spacecraft dry mass, propellant mass, and T500 (a science data collection figure of merit that tabulates the cumulative time spent below 500 km). These plots illustrate how the mission can trade off science return relative to the cost in dry mass and propellant. Other optimal solutions such as minimum propellant or maximum T500 were found to either limit the science data collection or to be dry mass limiting, respectively. Sensitivity analyses were performed to find new optimal (maximum dry mass) solutions if the number of campaigns changed, if the coefficient of drag (CD) were different, and if the propellant specific impulse were increased. A surprising result showed that the dry mass and T500 were both increased if the number of campaigns decreased. Changes in CD provided the expected results - raising CD lowered both the dry mass and T500 while lowering CD raised both the dry mass and T500. Increases in the propellant specific impulse had the expected outcome of raising the dry mass and lowering the propellant load but there was no change in the T500 figure of merit. The

  1. Visualization of Space-Time Ambiguities to be Explored by NASA GEC Mission with a Critique of Synthesized Measurements for Different GEC Mission Scenarios

    NASA Technical Reports Server (NTRS)

    Sojka, Jan J.

    2003-01-01

    The Grant supported research addressing the question of how the NASA Solar Terrestrial Probes (STP) Mission called Geospace electrodynamics Connections (GEC) will resolve space-time structures as well as collect sufficient information to solve the coupled thermosphere-ionosphere- magnetosphere dynamics and electrodynamics. The approach adopted was to develop a high resolution in both space and time model of the ionosphere-thermosphere (I-T) over altitudes relevant to GEC, especially the deep-dipping phase. This I-T model was driven by a high- resolution model of magnetospheric-ionospheric (M-I) coupling electrodynamics. Such a model contains all the key parameters to be measured by GEC instrumentation, which in turn are the required parameters to resolve present-day problems in describing the energy and momentum coupling between the ionosphere-magnetosphere and ionosphere-thermosphere. This model database has been successfully created for one geophysical condition; winter, solar maximum with disturbed geophysical conditions, specifically a substorm. Using this data set, visualizations (movies) were created to contrast dynamics of the different measurable parameters. Specifically, the rapidly varying magnetospheric E and auroral electron precipitation versus the slower varying ionospheric F-region electron density, but rapidly responding E-region density.

  2. Ozone exposed epithelial cells modify cocultured natural killer cells

    PubMed Central

    Müller, Loretta; Brighton, Luisa E.

    2013-01-01

    Ozone (O3) causes significant adverse health effects worldwide. Nasal epithelial cells (NECs) are among the first sites within the respiratory system to be exposed to inhaled air pollutants. They recruit, activate, and interact with immune cells via soluble mediators and direct cell-cell contacts. Based on our recent observation demonstrating the presence of natural killer (NK) cells in nasal lavages, the goal of this study was to establish a coculture model of NECs and NK cells and examine how exposure to O3 modifies this interaction. Flow cytometry analysis was used to assess immunophenotypes of NK cells cocultured with either air- or O3-exposed NECs. Our data show that coculturing NK cells with O3-exposed NECs decreased intracellular interferon-γ (IFN-γ), enhanced, albeit not statistically significant, IL-4, and increased CD16 expression on NK cells compared with air controls. Additionally, the cytotoxicity potential of NK cells was reduced after coculturing with O3-exposed NECs. To determine whether soluble mediators released by O3-exposed NECs caused this shift, apical and basolateral supernatants of air- and O3-exposed NECs were used to stimulate NK cells. While the conditioned media of O3-exposed NECs alone did not reduce intracellular IFN-γ, O3 enhanced the expression of NK cell ligands ULBP3 and MICA/B on NECs. Blocking ULBP3 and MICA/B reversed the effects of O3-exposed NECs on IFN-γ production in NK cells. Taken together, these data showed that interactions between NECs and NK cells in the context of O3 exposure changes NK cell activity via direct cell-cell interactions and is dependent on ULBP3/MICA/B expressed on NECs. PMID:23241529

  3. Visualization of Space-Time Ambiguities to be Explored by the NASA GEC Mission with a Critique of Synthesized Measurements for Different GEC Mission Scenarios

    NASA Technical Reports Server (NTRS)

    Sojka, Jan J.; Zhu, Lie; Fuller-Rowell, Timothy J.

    2005-01-01

    The objective of this grant was to study how a multi-satellite mission configuration can be optimized for maximum exploratory scientific return. NASA's Solar Terrestrial Probe (STP) concept mission Geospace Electrodynamic Connections (GEC) was the target mission for this pilot study. GEC prime mission characteristics were two fold: (i) a series of three satellites in the same orbit plane with differential spacing, and (ii) a deep-dipping phase in which these satellites could dip to altitudes as low as 130 km to explore the lower ionosphere and thermosphere. Each satellite would carry a full suite of plasma and neutral in-situ sensors and have the same dipping capability. This latter aspect would be envisaged as a series, up to 10, of deep-dipping campaigns, each lasting 10 days during which the perigee would be lowered to the desired probing depth. The challenge in optimization is to establish the scientific problems that can best be addressed by varying or selecting satellite spacing during a two-year mission while also interspersing, in this two year time frame, the deep-dipping campaigns. Although this sounds like a straightforward trade-off situation, it is complicated by the orbit precession in local time, the location of perigee, and that even the dipping campaigns will have preferred satellite spacing requirements.

  4. Heterochrony as Diachronically Modified Cell-Cell Interactions

    PubMed Central

    Torday, John S.

    2016-01-01

    Heterochrony is an enabling concept in evolution theory that metaphorically captures the mechanism of biologic change due to mechanisms of growth and development. The spatio-temporal patterns of morphogenesis are determined by cell-to-cell signaling mediated by specific soluble growth factors and their cognate receptors on nearby cells of different germline origins. Subsequently, down-stream production of second messengers generates patterns of form and function. Environmental upheavals such as Romer’s hypothesized drying up of bodies of water globally caused the vertebrate water-land transition. That transition caused physiologic stress, modifying cell-cell signaling to generate terrestrial adaptations of the skeleton, lung, skin, kidney and brain. These tissue-specific remodeling events occurred as a result of the duplication of the Parathyroid Hormone-related Protein Receptor (PTHrPR) gene, expressed in mesodermal fibroblasts in close proximity to ubiquitously expressed endodermal PTHrP, amplifying this signaling pathway. Examples of how and why PTHrPR amplification affected the ontogeny, phylogeny, physiology and pathophysiology of the lung are used to substantiate and further our understanding through insights to the heterochronic mechanisms of evolution, such as the fish swim bladder evolving into the vertebrate lung, interrelated by such functional homologies as surfactant and mechanotransduction. Instead of the conventional description of this phenomenon, lung evolution can now be understood as adaptive changes in the cellular-molecular signaling mechanisms underlying its ontogeny and phylogeny. PMID:26784244

  5. Genetically Modified T Cells to Target Glioblastoma

    PubMed Central

    Krebs, Simone; Rodríguez-Cruz, Tania G.; DeRenzo, Christopher; Gottschalk, Stephen

    2013-01-01

    Despite advances in surgical procedures, radiation, and chemotherapy the outcome for patients with glioblastoma (GBM) remains poor. While GBM cells express antigens that are potentially recognized by T cells, GBMs prevent the induction of GBM-specific immune responses by creating an immunosuppressive microenvironment. The advent of gene transfer has allowed the rapid generation of antigen-specific T cells as well as T cells with enhanced effector function. Here we review recent advances in the field of cell therapy with genetically modified T cells and how these advances might improve outcomes for patients with GBM in the future. PMID:24427741

  6. The renal microenvironment modifies dendritic cell phenotype.

    PubMed

    Chessa, Federica; Mathow, Daniel; Wang, Shijun; Hielscher, Thomas; Atzberger, Ann; Porubsky, Stefan; Gretz, Norbert; Burgdorf, Sven; Gröne, Hermann-Josef; Popovic, Zoran V

    2016-01-01

    Renal dendritic cells are a major component of the renal mononuclear phagocytic system. In the renal interstitium, these cells are exposed to an osmotic gradient, mainly sodium, whose concentration progressively increases towards inner medulla. Renal allograft rejection affects predominantly the cortex, suggesting a protective role of the renal medullary micromilieu. Whether osmolar variations can modulate the function of renal dendritic cells is currently undefined. Considering the central role of dendritic cells in promoting allorejection, we tested whether the biophysical micromilieu, particularly the interstitial osmotic gradient, influences their alloreactivity. There was a progressive depletion of leukocytes towards the medulla of homeostatic kidney. Only macrophages opposed this tendency. Flow cytometry of homeostatic and post-transplant medullary dendritic cells revealed a switch towards a macrophage-like phenotype. Similarly, bone marrow-derived dendritic cells developed ex vivo in sodium chloride-enriched medium acquired a M2-like signature. Microarray analysis of allotransplant dendritic cells posed a medullary downregulation of genes mainly involved in alloantigen recognition. Gene expression profiles of both medullary dendritic cells and bone marrow-derived dendritic cells matured in hyperosmolar medium had an overlap with the macrophage M2 signature. Thus, the medullary environment inhibits an alloimmune response by modulating the phenotype and function of dendritic cells.

  7. Cell behavior on surface modified polydimethylsiloxane (PDMS).

    PubMed

    Stanton, Morgan M; Rankenberg, Johanna M; Park, Byung-Wook; McGimpsey, W Grant; Malcuit, Christopher; Lambert, Christopher R

    2014-07-01

    Designing complex tissue culture systems requires cell alignment and directed extracellular matrix (ECM) and gene expression. Here, a micro-rough, polydimethylsiloxane (PDMS) surface, that also integrates a micro-pattern of 50 µm wide lines of fibronectin (FN) separated by 60 µm wide lines of bovine serum albumin (BSA), is developed. Human fibroblasts cultured on the rough, patterned substrate have aligned growth and a significant change in morphology when compared to cells on a flat, patterned surface. The rough PDMS topography significantly decreases cell area and induces the upregulation of several ECM related genes by two-fold when compared to cells cultured on flat PDMS. This study describes a simple surface engineering procedure for creating surface architecture for scaffolds to design and control the cell-surface interface.

  8. Glomerular endothelial cell injury and cross talk in diabetic kidney disease.

    PubMed

    Fu, Jia; Lee, Kyung; Chuang, Peter Y; Liu, Zhihong; He, John Cijiang

    2015-02-15

    Diabetic kidney disease (DKD) remains a leading cause of new-onset end-stage renal disease (ESRD), and yet, at present, the treatment is still very limited. A better understanding of the pathogenesis of DKD is therefore necessary to develop more effective therapies. Increasing evidence suggests that glomerular endothelial cell (GEC) injury plays a major role in the development and progression of DKD. Alteration of the glomerular endothelial cell surface layer, including its major component, glycocalyx, is a leading cause of microalbuminuria observed in early DKD. Many studies suggest a presence of cross talk between glomerular cells, such as between GEC and mesangial cells or GEC and podocytes. PDGFB/PDGFRβ is a major mediator for GEC and mesangial cell cross talk, while vascular endothelial growth factor (VEGF), angiopoietins, and endothelin-1 are the major mediators for GEC and podocyte communication. In DKD, GEC injury may lead to podocyte damage, while podocyte loss further exacerbates GEC injury, forming a vicious cycle. Therefore, GEC injury may predispose to albuminuria in diabetes either directly or indirectly by communication with neighboring podocytes and mesangial cells via secreted mediators. Identification of novel mediators of glomerular cell cross talk, such as microRNAs, will lead to a better understanding of the pathogenesis of DKD. Targeting these mediators may be a novel approach to develop more effective therapy for DKD.

  9. Glomerular endothelial cell injury and cross talk in diabetic kidney disease

    PubMed Central

    Fu, Jia; Lee, Kyung; Chuang, Peter Y.; Liu, Zhihong

    2014-01-01

    Diabetic kidney disease (DKD) remains a leading cause of new-onset end-stage renal disease (ESRD), and yet, at present, the treatment is still very limited. A better understanding of the pathogenesis of DKD is therefore necessary to develop more effective therapies. Increasing evidence suggests that glomerular endothelial cell (GEC) injury plays a major role in the development and progression of DKD. Alteration of the glomerular endothelial cell surface layer, including its major component, glycocalyx, is a leading cause of microalbuminuria observed in early DKD. Many studies suggest a presence of cross talk between glomerular cells, such as between GEC and mesangial cells or GEC and podocytes. PDGFB/PDGFRβ is a major mediator for GEC and mesangial cell cross talk, while vascular endothelial growth factor (VEGF), angiopoietins, and endothelin-1 are the major mediators for GEC and podocyte communication. In DKD, GEC injury may lead to podocyte damage, while podocyte loss further exacerbates GEC injury, forming a vicious cycle. Therefore, GEC injury may predispose to albuminuria in diabetes either directly or indirectly by communication with neighboring podocytes and mesangial cells via secreted mediators. Identification of novel mediators of glomerular cell cross talk, such as microRNAs, will lead to a better understanding of the pathogenesis of DKD. Targeting these mediators may be a novel approach to develop more effective therapy for DKD. PMID:25411387

  10. Accelerated Tumor Cell Death by Angiogenic Modifiers

    DTIC Science & Technology

    2004-08-01

    neuroendocrine factors. They can guide cancer cell perineural invasion and dissemination through the release of soluble and solid matrix factors (see review (32...Ooshima, A. Targeted disruption of TGF-betal/Smad3 signaling protects against renal tubulointerstitial fibrosis induced by unilateral ureteral

  11. Compound 13, an α1-selective small molecule activator of AMPK, inhibits Helicobacter pylori-induced oxidative stresses and gastric epithelial cell apoptosis

    SciTech Connect

    Zhao, Hangyong; Zhu, Huanghuang; Lin, Zhou; Lin, Gang; Lv, Guoqiang

    2015-08-07

    Half of the world's population experiences Helicobacter pylori (H. pylori) infection, which is a main cause of gastritis, duodenal and gastric ulcer, and gastric cancers. In the current study, we investigated the potential role of compound 13 (C13), a novel α1-selective small molecule activator of AMP-activated protein kinase (AMPK), against H. pylori-induced cytotoxicity in cultured gastric epithelial cells (GECs). We found that C13 induced significant AMPK activation, evidenced by phosphorylation of AMPKα1 and ACC (acetyl-CoA carboxylase), in both primary and transformed GECs. Treatment of C13 inhibited H. pylori-induced GEC apoptosis. AMPK activation was required for C13-mediated GEC protection. Inhibition of AMPK kinase activity by the AMPK inhibitor Compound C, or silencing AMPKα1 expression by targeted-shRNAs, alleviated C13-induced GEC protective activities against H. pylori. Significantly, C13 inhibited H. pylori-induced reactive oxygen species (ROS) production in GECs. C13 induced AMPK-dependent expression of anti-oxidant gene heme oxygenase (HO-1) in GECs. Zinc protoporphyrin (ZnPP) and tin protoporphyrin (SnPP), two HO-1 inhibitors, not only suppressed C13-mediated ROS scavenging activity, but also alleviated its activity in GECs against H. pylori. Together, these results indicate that C13 inhibits H. pylori-induced ROS production and GEC apoptosis through activating AMPK–HO–1 signaling. - Highlights: • We synthesized compound 13 (C13), a α1-selective small molecule AMPK activator. • C13-induced AMPK activation requires α1 subunit in gastric epithelial cells (GECs). • C13 enhances Helicobacter pylori-induced pro-survival AMPK activation to inhibit GEC apoptosis. • C13 inhibits H. pylori-induced reactive oxygen species (ROS) production in GECs. • AMPK-heme oxygenase (HO-1) activation is required for C13-mediated anti-oxidant activity.

  12. Accelerated Tumor Cell Death by Angiogenic Modifiers

    DTIC Science & Technology

    2003-08-01

    form an active autocrine loop. fibrosis . Luminal epithelial cells of PIA lesions have elev- A recent study indicated the increase of both IL-6 and...its receptor dothelin-3 (ET-3), and endothelin-4 (ET-4) (Cun- ( CXCR4 ), may play a role as prostate cancer bone meta- ningham et al., 1997). All...members of the endothelin stasis homing signals. The level of CXCR4 increased family contain two essential disulfide bridges and six with the malignancy of

  13. Magnetic studies of ferrofluid-modified microbial cells.

    PubMed

    Mosiniewicz-Szablewska, Ewa; Safarikova, Mirka; Safarik, Ivo

    2010-04-01

    Microbial cells (Kluyveromyces fragilis and Chlorella vulgaris) efficiently interacted with maghemite nanoparticles stabilized as low-pH ionic magnetic fluid, leading to the formation of magnetically labeled cells. This simple procedure allows to use the prepared materials as new cheap and easy to get magnetic affinity adsorbents to the removal of water-soluble dyes from polluted water sources using magnetic separation techniques. Magnetically modified cells were investigated by means of electron spin resonance spectroscopy and conventional magnetic methods over the temperature range 4-300 K. The magnetic behavior of these materials was dominated by the superparamagnetic relaxation of isolated single domain maghemite particles although a little amount of agglomerates was also present on the cell surface. However, these agglomerates were sufficiently small to show at static conditions the superparamagnetic behavior at room temperature. Therefore, the ferrofluid-modified microbial cells represent new interesting magnetic affinity adsorbents which could be applied for large-scale magnetic separation processes.

  14. Surface-modified gold nanorods for specific cell targeting

    NASA Astrophysics Data System (ADS)

    Wang, Chan-Ung; Arai, Yoshie; Kim, Insun; Jang, Wonhee; Lee, Seonghyun; Hafner, Jason H.; Jeoung, Eunhee; Jung, Deokho; Kwon, Youngeun

    2012-05-01

    Gold nanoparticles (GNPs) have unique properties that make them highly attractive materials for developing functional reagents for various biomedical applications including photothermal therapy, targeted drug delivery, and molecular imaging. For in vivo applications, GNPs need to be prepared with very little or negligible cytotoxicitiy. Most GNPs are, however, prepared using growth-directing surfactants such as cetyl trimethylammonium bromide (CTAB), which are known to have considerable cytotoxicity. In this paper, we describe an approach to remove CTAB to a non-toxic concentration. We optimized the conditions for surface modification with methoxypolyethylene glycol thiol (mPEG), which replaced CTAB and formed a protective layer on the surface of gold nanorods (GNRs). The cytotoxicities of pristine and surface-modified GNRs were measured in primary human umbilical vein endothelial cells and human cell lines derived from hepatic carcinoma cells, embryonic kidney cells, and thyroid papillary carcinoma cells. Cytotoxicity assays revealed that treating cells with GNRs did not significantly affect cell viability except for thyroid papillary carcinoma cells. Thyroid cancer cells were more susceptible to residual CTAB, so CTAB had to be further removed by dialysis in order to use GNRs for thyroid cell targeting. PEGylated GNRs are further modified to present monoclonal antibodies that recognize a specific surface marker, Na-I symporter, for thyroid cells. Antibody-conjugated GNRs specifically targeted human thyroid cells in vitro.

  15. Genetically modified T cells in cancer therapy: opportunities and challenges.

    PubMed

    Sharpe, Michaela; Mount, Natalie

    2015-04-01

    Tumours use many strategies to evade the host immune response, including downregulation or weak immunogenicity of target antigens and creation of an immune-suppressive tumour environment. T cells play a key role in cell-mediated immunity and, recently, strategies to genetically modify T cells either through altering the specificity of the T cell receptor (TCR) or through introducing antibody-like recognition in chimeric antigen receptors (CARs) have made substantial advances. The potential of these approaches has been demonstrated in particular by the successful use of genetically modified T cells to treat B cell haematological malignancies in clinical trials. This clinical success is reflected in the growing number of strategic partnerships in this area that have attracted a high level of investment and involve large pharmaceutical organisations. Although our understanding of the factors that influence the safety and efficacy of these therapies has increased, challenges for bringing genetically modified T-cell immunotherapy to many patients with different tumour types remain. These challenges range from the selection of antigen targets and dealing with regulatory and safety issues to successfully navigating the routes to commercial development. However, the encouraging clinical data, the progress in the scientific understanding of tumour immunology and the improvements in the manufacture of cell products are all advancing the clinical translation of these important cellular immunotherapies.

  16. Total Ozone Mapping Spectrometer (TOMS) Derived Data, Global Earth Coverage (GEC) from NASA's Earth Probe Satellite

    DOE Data Explorer

    This is data from an external datastream processed through the ARM External Data Center (XDC) at Brookhaven National Laboratory. The XDC identifies sources and acquires data, called "external data", to augment the data being generated within the ARM program. The external data acquired are usually converted from native format to either netCDF or HDF formats. The GEC collection contains global data derived from the Total Ozone Mapping Spectrometer (TOMS) instrument on the Earth Probe satellite, consisting of daily values of aerosol index, ozone and reflectivity remapped into a regular 1x1.25 deg grid. Data are available from July 25, 1996 - December 31, 2005, but have been updated or replaced as of September 2007. See the explanation on the ARM web site at http://www.arm.gov/xds/static/toms.stm and the information at the NASA/TOMS web site: http://toms.gsfc.nasa.gov/ (Registration required)

  17. Properties of reactively deposited SiC and GeC alloys

    NASA Astrophysics Data System (ADS)

    Martin, Peter M.; Johnston, John W.; Bennett, Wendy D.

    1990-12-01

    Thin-film silicon carbide (SiCi) and germanium carbon (Ge,Ci) alloy coatings with low ›ifrared optical absorption have been fabricated by DC- and RF-reactive magnetron sputtering. The optical and mechanical properties of the coatings depend on composition determined by deposition conditions. The refractive index and optical absorption coefficient of SiCi. alloys were varied from those of amorphous Si to those near diamond-like carbon (DLC) by increasing C content. The band edge shifted below 1.2 eV with C content as high as 0.8. The useful range of the SiCi coatings was extended to wavelengths as low as 1 jim. The useful transparency range of GeCi coatings is from 3 to 12 jim. The refractive index of GeCi coatings was varied from 4.2 of amorphous Ge to near 3.4 by increasing x from 0 to 0.5. The optical absorption coefficient was a complex function of composition and C-H, Ge-H, and Ge-C bonding. Mechanical stress in both materials was generally moderate, and increased with increasing C content for the GeC alloys and decreased with increasing C for the SiC alloys. The wide range of optical properties obtainable for both coating types makes them useful in many types of multilayer designs. Abrasion-resistant infrared (IR) multispectral antireflection coatings on zinc sulfide (ZnS) were demonstrated using Geij"9C and DLC layers.

  18. Engineering chemically modified viruses for prostate cancer cell recognition.

    PubMed

    Mohan, K; Weiss, G A

    2015-12-01

    Specific detection of circulating tumor cells and characterization of their aggressiveness could improve cancer diagnostics and treatment. Metastasis results from such tumor cells, and causes the majority of cancer deaths. Chemically modified viruses could provide an inexpensive and efficient approach to detect tumor cells and quantitate their cell surface biomarkers. However, non-specific adhesion between the cell surface receptors and the virus surface presents a challenge. This report describes wrapping the virus surface with different PEG architectures, including as fusions to oligolysine, linkers, spacers and scaffolded ligands. The reported PEG wrappers can reduce by >75% the non-specific adhesion of phage to cell surfaces. Dynamic light scattering verified the non-covalent attachment by the reported wrappers as increased sizes of the virus particles. Further modifications resulted in specific detection of prostate cancer cells expressing PSMA, a key prostate cancer biomarker. The approach allowed quantification of PSMA levels on the cell surface, and could distinguish more aggressive forms of the disease.

  19. RGD modified polymers: biomaterials for stimulated cell adhesion and beyond.

    PubMed

    Hersel, Ulrich; Dahmen, Claudia; Kessler, Horst

    2003-11-01

    Since RGD peptides (R: arginine; G: glycine; D: aspartic acid) have been found to promote cell adhesion in 1984 (Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule, Nature 309 (1984) 30), numerous materials have been RGD functionalized for academic studies or medical applications. This review gives an overview of RGD modified polymers, that have been used for cell adhesion, and provides information about technical aspects of RGD immobilization on polymers. The impacts of RGD peptide surface density, spatial arrangement as well as integrin affinity and selectivity on cell responses like adhesion and migration are discussed.

  20. Visceral glomerular epithelial cells can proliferate in vivo and synthesize platelet-derived growth factor B-chain.

    PubMed Central

    Floege, J.; Johnson, R. J.; Alpers, C. E.; Fatemi-Nainie, S.; Richardson, C. A.; Gordon, K.; Couser, W. G.

    1993-01-01

    In glomerular diseases associated with antibody- and complement-mediated injury to endothelial and mesangial cells, cell proliferation is an important early response that precedes matrix accumulation and glomerulosclerosis. In contrast, in diseases in which the visceral glomerular epithelial cell (vGEC) is the principal target of injury, cell proliferation is not a recognized consequence, although vGECs respond in vitro to a variety of growth factors and inflammatory mediators. To explore the possibility that low levels of vGEC proliferation may occur and participate in such diseases, serial studies were done in the passive Heymann nephritis model of membranous nephropathy, in which the vGEC is the primary target of antibody- and C5b-9-mediated injury. The results showed mitotic figures and positive staining for the proliferating cell nuclear antigen in cells whose location defined them as vGECs. The proliferating cell nuclear antigen-positive cells at the edge of the capillary wall were confirmed to be vGECs by double-immunostaining with antibodies to SPARC/osteonectin, which preferentially label vGECs in passive Heymann nephritis. Proliferation of vGECs in vivo was preceded by increased glomerular expression of platelet-derived growth factor (PDGF) B-chain protein and messenger RNA, both of which localized to vGECs. PDGF B-chain protein and messenger RNA were also detected in cultured vGECs. PDGF receptor beta-subunit protein or messenger RNA could not be demonstrated in vGECs in vivo or in vitro, and no growth response of cultured vGECs to PDGF was noted. These results suggest that proliferation of vGECs does occur in a model of glomerular injury induced by antibody and C5b-9 on vGECs. VGEC proliferation and production of PDGF may be involved in the restoration of the capillary wall but could also contribute to local capillary wall injury and proliferation of other cells in Bowman's capsule, interstitium, and tubules. Images Figure 1 Figure 2 Figure 4 Figure 5

  1. Endothelial cell migration on surfaces modified with immobilized adhesive peptides.

    PubMed

    Kouvroukoglou, S; Dee, K C; Bizios, R; McIntire, L V; Zygourakis, K

    2000-09-01

    Endothelial cell (EC) migration has been studied on aminophase surfaces with covalently bound RGDS and YIGSRG cell adhesion peptides. The fluorescent marker dansyl chloride was used to quantify the spatial distribution of the peptides on the modified surfaces. Peptides appeared to be distributed in uniformly dispersed large clusters separated by areas of lower peptide concentrations. We employed digital time-lapse video microscopy and image analysis to monitor EC migration on the modified surfaces and to reconstruct the cell trajectories. The persistent random walk model was then applied to analyze the cell displacement data and compute the mean root square speed, the persistence time, and the random motility coefficient of EC. We also calculated the time-averaged speed of cell locomotion. No differences in the speed of cell locomotion on the various substrates were noted. Immobilization of the cell adhesion peptides (RGDS and YIGSRG), however, significantly increased the persistence of cell movement and, thus, the random motility coefficient. These results suggest that immobilization of cell adhesion peptides on the surface of implantable biomaterials may lead to enhanced endothelization rates.

  2. Carboxybetaine methacrylate oligomer modified nylon for circulating tumor cells capture.

    PubMed

    Dong, Chaoqun; Wang, Huiyu; Zhang, Zhuo; Zhang, Tao; Liu, Baorui

    2014-10-15

    Circulating tumor cells (CTC) capture is one of the most effective approaches in diagnosis and treatment of cancers in the field of personalized cancer medicine. In our study, zwitterionic carboxybetaine methacrylate (CBMA) oligomers were grafted onto nylon via atomic transfer random polymerization (ATRP) which would serve as a novel material for the development of convenient CTC capture interventional medical devices. The chemical, physical and biological properties of pristine and modified nylon surfaces were assessed by Fourier transform infrared spectra, atomic force microscope, water contact angle measurements, X-ray photoelectron spectroscopy, protein adsorption, platelet adhesion, and plasma recalcification time (PRT) determinations, etc. The results, including the significant decrease of proteins adsorption and platelets adhesion, as well as prolonged PRTs demonstrated the extraordinary biocompatibility and blood compatibility of the modified surface. Furthermore, we showed that upon immobilization of anti-epithelial cell adhesion molecular (anti-EpCAM) antibody onto the CBMA moiety, the modified nylon surface can selectively capture EpCAM positive tumor cells from blood with high efficiency, indicating the potential of the modified nylon in the manufacture of convenient interventional CTC capture medical devices.

  3. First-principles calculations of the structural, elastic, electronic, chemical bonding and optical properties of zinc-blende and rocksalt GeC

    NASA Astrophysics Data System (ADS)

    Liu, Qi-Jun; Liu, Zheng-Tang; Che, Xing-Sen; Feng, Li-Ping; Tian, Hao

    2011-12-01

    Structural parameters, elastic, electronic, bonding and optical properties of zinc-blende and rocksalt GeC have been investigated using the plane-wave ultrasoft pseudopotential technique based on the first-principles density-functional theory (DFT). The ground-state properties obtained by minimizing the total energy are in favorable agreement with the previous work. Two phases of GeC are found to be elastically stable and we have obtained the bulk, shear and Young's modulus, Poisson's coefficient and Lamé's constants for zinc-blende and rocksalt GeC. We estimated the Debye temperature of zinc-blende and rocksalt GeC from the acoustic velocity. Electronic and chemical bonding properties have been studied. Moreover, the complex dielectric function, refractive index, extinction coefficient, optical reflectivity, absorption coefficient, energy-loss spectrum and the complex conductivity function are calculated.

  4. Genetically modified mesenchymal stem cells for improved islet transplantation.

    PubMed

    Wu, Hao; Ye, Zhaoyang; Mahato, Ram I

    2011-10-03

    The use of adult stem cells for therapeutic purposes has met with great success in recent years. Among several types of adult stem cells, mesenchymal stem cells (MSCs) derived from bone marrow (BM) and other sources have gained popularity for basic research and clinical applications because of their therapeutic potential in treating a variety of diseases. Because of their tissue regeneration potential and immune modulation effect, MSCs were recently used as cell-based therapy to promote revascularization, increase pancreatic β-cell proliferation, and avoid allograft rejection in islet transplantation. Taking advantage of the recent progress in gene therapy, genetically modified MSCs can further enhance and expand the therapeutic benefit of primary MSCs while retaining their stem-cell-like properties. This review aims to gain a thorough understanding of the current obstacles to successful islet transplantation and discusses the potential role of primary MSCs before or after genetic modification in islet transplantation.

  5. Interaction of carbohydrate modified boron nitride nanotubes with living cells.

    PubMed

    Emanet, Melis; Şen, Özlem; Çobandede, Zehra; Çulha, Mustafa

    2015-10-01

    Boron nitride nanotubes (BNNTs) are composed of boron and nitrogen atoms and they show significantly different properties from their carbon analogues (carbon nanotubes, CNTs). Due to their unique properties including low electrical conductivity, and imaging contrast and neutron capture properties; they can be used in biomedical applications. When their use in biological fields is considered, the route of their toxic effect should be clarified. Therefore, the study of interactions between BNNTs and living systems is important in envisaging biological applications at both cellular and sub-cellular levels to fully gain insights of their potential adverse effects. In this study, BNNTs were modified with lactose, glucose and starch and tested for their cytotoxicity. First, the interactions and the behavior of BNNTs with bovine serum albumin (BSA), Dulbecco's Modified Eagle's Medium (DMEM) and DMEM/Nutrient Mixture F-12Ham were investigated. Thereafter, their cellular uptake and the cyto- and genotoxicity on human dermal fibroblasts (HDFs) and adenocarcinoma human alveolar basal epithelial cells (A549) were evaluated. HDFs and A549 cells internalized the modified and unmodified BNNTs, and BNNTs were found to not cause significant viability change and DNA damage. A higher uptake rate of BNNTs by A549 cells compared to HDFs was observed. Moreover, a concentration-dependent cytotoxicity was observed on A549 cells while they were safer for HDFs in the same concentration range. Based on these findings, it can be concluded that BNNTs and their derivatives made with biomacromolecules might be good candidates for several applications in medicine and biomedical applications.

  6. Somatic cell reprogramming-free generation of genetically modified pigs.

    PubMed

    Tanihara, Fuminori; Takemoto, Tatsuya; Kitagawa, Eri; Rao, Shengbin; Do, Lanh Thi Kim; Onishi, Akira; Yamashita, Yukiko; Kosugi, Chisato; Suzuki, Hitomi; Sembon, Shoichiro; Suzuki, Shunichi; Nakai, Michiko; Hashimoto, Masakazu; Yasue, Akihiro; Matsuhisa, Munehide; Noji, Sumihare; Fujimura, Tatsuya; Fuchimoto, Dai-Ichiro; Otoi, Takeshige

    2016-09-01

    Genetically modified pigs for biomedical applications have been mainly generated using the somatic cell nuclear transfer technique; however, this approach requires complex micromanipulation techniques and sometimes increases the risks of both prenatal and postnatal death by faulty epigenetic reprogramming of a donor somatic cell nucleus. As a result, the production of genetically modified pigs has not been widely applied. We provide a simple method for CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 gene editing in pigs that involves the introduction of Cas9 protein and single-guide RNA into in vitro fertilized zygotes by electroporation. The use of gene editing by electroporation of Cas9 protein (GEEP) resulted in highly efficient targeted gene disruption and was validated by the efficient production of Myostatin mutant pigs. Because GEEP does not require the complex methods associated with micromanipulation for somatic reprogramming, it has the potential for facilitating the genetic modification of pigs.

  7. Somatic cell reprogramming-free generation of genetically modified pigs

    PubMed Central

    Tanihara, Fuminori; Takemoto, Tatsuya; Kitagawa, Eri; Rao, Shengbin; Do, Lanh Thi Kim; Onishi, Akira; Yamashita, Yukiko; Kosugi, Chisato; Suzuki, Hitomi; Sembon, Shoichiro; Suzuki, Shunichi; Nakai, Michiko; Hashimoto, Masakazu; Yasue, Akihiro; Matsuhisa, Munehide; Noji, Sumihare; Fujimura, Tatsuya; Fuchimoto, Dai-ichiro; Otoi, Takeshige

    2016-01-01

    Genetically modified pigs for biomedical applications have been mainly generated using the somatic cell nuclear transfer technique; however, this approach requires complex micromanipulation techniques and sometimes increases the risks of both prenatal and postnatal death by faulty epigenetic reprogramming of a donor somatic cell nucleus. As a result, the production of genetically modified pigs has not been widely applied. We provide a simple method for CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 gene editing in pigs that involves the introduction of Cas9 protein and single-guide RNA into in vitro fertilized zygotes by electroporation. The use of gene editing by electroporation of Cas9 protein (GEEP) resulted in highly efficient targeted gene disruption and was validated by the efficient production of Myostatin mutant pigs. Because GEEP does not require the complex methods associated with micromanipulation for somatic reprogramming, it has the potential for facilitating the genetic modification of pigs. PMID:27652340

  8. Attachment of human primary osteoblast cells to modified polyethylene surfaces.

    PubMed

    Poulsson, Alexandra H C; Mitchell, Stephen A; Davidson, Marcus R; Johnstone, Alan J; Emmison, Neil; Bradley, Robert H

    2009-04-09

    Ultra-high-molecular-weight polyethylene (UHMWPE) has a long history of use in medical devices, primarily for articulating surfaces due to its inherent low surface energy which limits tissue integration. To widen the applications of UHMWPE, the surface energy can be increased. The increase in surface energy would improve the adsorption of proteins and attachment of cells to allow tissue integration, thereby allowing UHMWPE to potentially be used for a wider range of implants. The attachment and function of human primary osteoblast-like (HOB) cells to surfaces of UHMWPE with various levels of incorporated surface oxygen have been investigated. The surface modification of the UHMWPE was produced by exposure to a UV/ozone treatment. The resulting surface chemistry was studied using X-ray photoelectron spectroscopy (XPS), and the topography and surface structure were probed by atomic force microscopy (AFM) and scanning electron microscopy (SEM), which showed an increase in surface oxygen from 11 to 26 atom % with no significant change to the surface topography. The absolute root mean square roughness of both untreated and UV/ozone-treated surfaces was within 350-450 nm, and the water contact angles decreased with increasing oxygen incorporation, i.e., showing an increase in surface hydrophilicity. Cell attachment and functionality were assessed over a 21 day period for each cell-surface combination studied; these were performed using SEM and the alamarBlue assay to study cell attachment and proliferation and energy-dispersive X-ray (EDX) analysis to confirm extracellular mineral deposits, and total protein assay to examine the intra- and extracellular protein expressed by the cells. HOB cells cultured for 21 days on the modified UHMWPE surfaces with 19 and 26 atom % oxygen incorporated showed significantly higher cell densities compared to cells cultured on tissue culture polystyrene (TCPS) from day 3 onward. This indicated that the cells attached and proliferated more

  9. Immobilization of chlorine dioxide modified cells for uranium absorption.

    PubMed

    He, Shengbin; Ruan, Binbiao; Zheng, Yueping; Zhou, Xiaobin; Xu, Xiaoping

    2014-11-01

    There has been a trend towards the use of microorganisms to recover metals from industrial wastewater, for which various methods have been reported to be used to improve microorganism adsorption characteristics such as absorption capacity, tolerance and reusability. In present study, chlorine dioxide(ClO2), a high-efficiency, low toxicity and environment-benign disinfectant, was first reported to be used for microorganism surface modification. The chlorine dioxide modified cells demonstrated a 10.1% higher uranium adsorption capacity than control ones. FTIR analysis indicated that several cell surface groups are involved in the uranium adsorption and cell surface modification. The modified cells were further immobilized on a carboxymethylcellulose(CMC) matrix to improve their reusability. The cell-immobilized adsorbent could be employed either in a high concentration system to move vast UO2(2+) ions or in a low concentration system to purify UO2(2+) contaminated water thoroughly, and could be repeatedly used in multiple adsorption-desorption cycles with about 90% adsorption capacity maintained after seven cycles.

  10. Antitumor efficacy of vaccinia virus-modified tumor cell vaccine

    SciTech Connect

    Ito, T.; Wang, D.Q.; Maru, M.; Nakajima, K.; Kato, S.; Kurimura, T.; Wakamiya, N. )

    1990-11-01

    The antitumor efficacies of vaccinia virus-modified tumor cell vaccines were examined in murine syngeneic MH134 and X5563 tumor cells. UV-inactivated vaccinia virus was inoculated i.p. into C3H/HeN mice that had received whole body X-irradiation at 150 rads. After 3 weeks, the vaccines were administered i.p. 3 times at weekly intervals. One week after the last injection, mice were challenged i.p. with various doses of syngeneic MH134 or X5563 viable tumor cells. Four methods were used for preparing tumor cell vaccines: X-ray irradiation; fixation with paraformaldehyde for 1 h or 3 months; and purification of the membrane fraction. All four vaccines were effective, but the former two vaccines were the most effective. A mixture of the membrane fraction of untreated tumor cells and UV-inactivated vaccinia virus also had an antitumor effect. These results indicate that vaccine with the complete cell structure is the most effective. The membrane fraction of UV-inactivated vaccinia virus-absorbed tumor cells was also effective. UV-inactivated vaccinia virus can react with not only intact tumor cells but also the purified membrane fraction of tumor cells and augment antitumor activity.

  11. Geothermal Economics Calculator (GEC) - additional modifications to final report as per GTP's request.

    SciTech Connect

    Gowda, Varun; Hogue, Michael

    2015-07-17

    This report will discuss the methods and the results from economic impact analysis applied to the development of Enhanced Geothermal Systems (EGS), conventional hydrothermal, low temperature geothermal and coproduced fluid technologies resulting in electric power production. As part of this work, the Energy & Geoscience Institute (EGI) has developed a web-based Geothermal Economics Calculator (Geothermal Economics Calculator (GEC)) tool that is aimed at helping the industry perform geothermal systems analysis and study the associated impacts of specific geothermal investments or technological improvements on employment, energy and environment. It is well-known in the industry that geothermal power projects will generate positive economic impacts for their host regions. Our aim in the assessment of these impacts includes quantification of the increase in overall economic output due to geothermal projects and of the job creation associated with this increase. Such an estimate of economic impacts of geothermal investments on employment, energy and the environment will also help us understand the contributions that the geothermal industry will have in achieving a sustainable path towards energy production.

  12. An efficient magnetically modified microbial cell biocomposite for carbazole biodegradation

    NASA Astrophysics Data System (ADS)

    Li, Yufei; Du, Xiaoyu; Wu, Chao; Liu, Xueying; Wang, Xia; Xu, Ping

    2013-12-01

    Magnetic modification of microbial cells enables to prepare smart biocomposites in bioremediation. In this study, we constructed an efficient biocomposite by assembling Fe3O4 nanoparticles onto the surface of Sphingomonas sp. XLDN2-5 cells. The average particle size of Fe3O4 nanoparticles was about 20 nm with 45.5 emu g-1 saturation magnetization. The morphology of Sphingomonas sp. XLDN2-5 cells before and after Fe3O4 nanoparticle loading was verified by scanning electron microscopy and transmission electronic microscopy. Compared with free cells, the microbial cell/Fe3O4 biocomposite had the same biodegradation activity but exhibited remarkable reusability. The degradation activity of the microbial cell/Fe3O4 biocomposite increased gradually during recycling processes. Additionally, the microbial cell/Fe3O4 biocomposite could be easily separated and recycled by an external magnetic field due to the super-paramagnetic properties of Fe3O4 nanoparticle coating. These results indicated that magnetically modified microbial cells provide a promising technique for improving biocatalysts used in the biodegradation of hazardous compounds.

  13. An efficient magnetically modified microbial cell biocomposite for carbazole biodegradation

    PubMed Central

    2013-01-01

    Magnetic modification of microbial cells enables to prepare smart biocomposites in bioremediation. In this study, we constructed an efficient biocomposite by assembling Fe3O4 nanoparticles onto the surface of Sphingomonas sp. XLDN2-5 cells. The average particle size of Fe3O4 nanoparticles was about 20 nm with 45.5 emu g-1 saturation magnetization. The morphology of Sphingomonas sp. XLDN2-5 cells before and after Fe3O4 nanoparticle loading was verified by scanning electron microscopy and transmission electronic microscopy. Compared with free cells, the microbial cell/Fe3O4 biocomposite had the same biodegradation activity but exhibited remarkable reusability. The degradation activity of the microbial cell/Fe3O4 biocomposite increased gradually during recycling processes. Additionally, the microbial cell/Fe3O4 biocomposite could be easily separated and recycled by an external magnetic field due to the super-paramagnetic properties of Fe3O4 nanoparticle coating. These results indicated that magnetically modified microbial cells provide a promising technique for improving biocatalysts used in the biodegradation of hazardous compounds. PMID:24330511

  14. Mps1 is SUMO-modified during the cell cycle

    PubMed Central

    Chen, Changyan; Lu, Lou; Dai, Wei

    2016-01-01

    Mps1 is a dual specificity protein kinase that regulates the spindle assembly checkpoint and mediates proper microtubule attachment to chromosomes during mitosis. However, the molecular mechanism that controls Mps1 protein level and its activity during the cell cycle remains unclear. Given that sumoylation plays an important role in mitotic progression, we investigated whether Mps1 was SUMO-modified and whether sumoylation affects its activity in mitosis. Our results showed that Mps1 was sumoylated in both asynchronized and mitotic cell populations. Mps1 was modified by both SUMO-1 and SUMO-2. Our further studies revealed that lysine residues including K71, K287, K367 and K471 were essential for Mps1 sumoylation. Sumoylation appeared to play a role in mediating kinetochore localization of Mps1, thus affecting normal mitotic progression. Furthermore, SUMO-resistant mutants of Mps1 interacted with BubR1 more efficiently than it did with the wild-type control. Combined, our results indicate that Mps1 is SUMO-modified that plays an essential role in regulating Mps1 functions during mitosis. PMID:26675261

  15. Hydrodynamic modeling of granular flows in a modified Couette cell.

    PubMed

    Jop, Pierre

    2008-03-01

    We present simulations of granular flows in a modified Couette cell, using a continuum model recently proposed for dense granular flows. Based on a friction coefficient, which depends on an inertial number, the model captures the positions of the wide shear bands. We show that a smooth transition in velocity-profile shape occurs when the height of the granular material is increased, leading to a differential rotation of the central part close to the surface. The numerical predictions are in qualitative agreement with previous experimental results. The model provides predictions for the increase of the shear band width when the rotation rate is increased.

  16. Release testing of retroviral vectors and gene-modified cells.

    PubMed

    Nordling, Diana; Kaiser, Anne; Reeves, Lilith

    2009-01-01

    This chapter will review the design and execution of release testing requirements for retroviral vectors and gene-modified cells consistent with ensuring the success of the clinical trial on the basis of current US regulatory requirements. It is the ethical and legal responsibility of the clinical trial sponsor(s) to ensure safety of the patients through proper evaluation of the drug products prior to use. Any clinical trial drug product used in human subjects must be produced and evaluated for safety, quality, purity, and effectiveness according to Current Good Manufacturing Practices appropriate for the stage of clinical development.

  17. Stretchable biofuel cell with enzyme-modified conductive textiles.

    PubMed

    Ogawa, Yudai; Takai, Yuki; Kato, Yuto; Kai, Hiroyuki; Miyake, Takeo; Nishizawa, Matsuhiko

    2015-12-15

    A sheet-type, stretchable biofuel cell was developed by laminating three components: a bioanode textile for fructose oxidation, a hydrogel sheet containing fructose as fuel, and a gas-diffusion biocathode textile for oxygen reduction. The anode and cathode textiles were prepared by modifying carbon nanotube (CNT)-decorated stretchable textiles with fructose dehydrogenase (FDH) and bilirubin oxidase (BOD), respectively. Enzymatic reaction currents of anode and cathode textiles were stable for 30 cycles of 50% stretching, with initial loss of 20-30% in the first few cycles due to the partial breaking of the CNT network at the junction of textile fibers. The assembled laminate biofuel cell showed power of ~0.2 mW/cm(2) with 1.2 kΩ load, which was stable even at stretched, twisted, and wrapped forms.

  18. Aldosterone does not modify gene expression in human endothelial cells.

    PubMed

    Verhovez, A; Williams, T A; Morello, F; Monticone, S; Brizzi, M F; Dentelli, P; Fallo, F; Fabris, B; Amenta, F; Gomez-Sanchez, C; Veglio, F; Mulatero, P

    2012-03-01

    The toxic effects of aldosterone on the vasculature, and in particular on the endothelial layer, have been proposed as having an important role in the cardiovascular pathology observed in mineralocorticoid-excess states. In order to characterize the genomic molecular mechanisms driving the aldosterone-induced endothelial dysfunction, we performed an expression microarray on transcripts obtained from both human umbilical vein endothelial cells and human coronary artery endothelial cells stimulated with 10 - 7 M aldosterone for 18 h. The results were then subjected to qRT-PCR confirmation, also including a group of genes known to be involved in the control of the endothelial function or previously described as regulated by aldosterone. The state of activation of the mineralocorticoid receptor was investigated by means of a luciferase-reporter assay using a plasmid encoding a mineralocorticoid and glucocorticoid-sensitive promoter. Aldosterone did not determine any significant change in gene expression in either cell type both in the microarray and in the qRT-PCR analysis. The luciferase-reporter assay showed no activation of the mineralocorticoid receptor following aldosterone stimulation. The status of nonfunctionality of the mineralocorticoid receptor expressed in cultured human umbilical and coronary artery endothelial cells does not allow aldosterone to modify gene expression and provides evidence against either a beneficial or harmful genomic effect of aldosterone on healthy endothelial cells.

  19. Ge/C nanowires as high-capacity and long-life anode materials for Li-ion batteries.

    PubMed

    Liu, Jun; Song, Kepeng; Zhu, Changbao; Chen, Chia-Chin; van Aken, Peter A; Maier, Joachim; Yu, Yan

    2014-07-22

    Germanium-based materials (Ge and GeOx) have recently demonstrated excellent lithium-ion storage ability and are being considered as the most promising candidates to substitute commercial carbon-based anodes of lithium-ion batteries. Nevertheless, practical implementation of Ge-based materials to lithium-ion batteries is greatly hampered by the poor cyclability that resulted from the huge volume variation during lithiation/delithiation processes. Herein, uniform carbon-encapsulated Ge and GeOx nanowires were synthesized by a one-step controlled pyrolysis of organic-inorganic hybrid GeOx/ethylenediamine (GeOx/EDA) nanowires in H2/Ar and Ar atmospheres, respectively. The as-obtained Ge/C and GeOx/C nanowires possess well-defined 0D-in-1D morphology and homogeneous carbon encapsulation, which exhibit excellent Li storage properties including high specific capacities (approximate 1200 and 1000 mA h g(-1) at 0.2C for Ge/C and GeOx/C, respectively). The Ge/C nanowires, in particular, demonstrate superior rate capability with excellent capacity retention and stability (producing high stable discharge capacities of about 770 mA h g(-1) after 500 cycles at 10C), making them promising candidates for future electrodes for high-power Li-ion batteries. The improved electrochemical performance arises from synergistic effects of 0D-in-1D morphology and uniform carbon coating, which could effectively accommodate the huge volume change of Ge/GeOx during cycling and maintain perfect electrical conductivity throughout the electrode.

  20. Cysteine modified polyaniline films improve biocompatibility for two cell lines.

    PubMed

    Yslas, Edith I; Cavallo, Pablo; Acevedo, Diego F; Barbero, César A; Rivarola, Viviana A

    2015-06-01

    This work focuses on one of the most exciting application areas of conjugated conducting polymers, which is cell culture and tissue engineering. To improve the biocompatibility of conducting polymers we present an easy method that involves the modification of the polymer backbone using l-cysteine. In this publication, we show the synthesis of polyaniline (PANI) films supported onto Polyethylene terephthalate (PET) films, and modified using cysteine (PANI-Cys) in order to generate a biocompatible substrate for cell culture. The PANI-Cys films are characterized by Fourier Transform infrared and UV-visible spectroscopy. The changes in the hydrophilicity of the polymer films after and before the modification were tested using contact angle measurements. After modification the contact angle changes from 86°±1 to 90°±1, suggesting a more hydrophylic surface. The adhesion properties of LM2 and HaCaT cell lines on the surface of PANI-Cys films in comparison with tissue culture plastic (TCP) are studied. The PANI-Cys film shows better biocompatibility than PANI film for both cell lines. The cell morphologies on the TCP and PANI-Cys film were examined by florescence and Atomic Force Microscopy (AFM). Microscopic observations show normal cellular behavior when PANI-Cys is used as a substrate of both cell lines (HaCaT and LM2) as when they are cultured on TCP. The ability of these PANI-Cys films to support cell attachment and growth indicates their potential use as biocompatible surfaces and in tissue engineering.

  1. Comparative analysis of the biological and physical properties of Enterococcus faecalis bacteriophage vB_EfaS_GEC-EfS_3 and Streptococcus mitis bacteriophage vB_SmM_GEC-SmitisM_2.

    PubMed

    Rigvava, Sophio; Tchgkonia, Irina; Jgenti, Darejan; Dvalidze, Teona; Carpino, James; Goderdzishvili, Marina

    2013-01-01

    Enterococcus faecalis and Streptococcus mitis are common commensal inhabitants of the human gastrointestinal and genitourinary tracts. However, both species can be opportunistic pathogens and cause disease in nosocomial settings. These infections can be difficult to treat because of the frequency of antibiotic resistance among these strains. Bacteriophages are often suggested as an alternative therapeutic agent against these infections. In this study, E. faecalis and S. mitis strains were isolated from female patients with urinary tract infections. Bacteriophages active against these strains were isolated from sewage water from the Mtkvari River. Two phages, designated vB_EfaS_GEC-EfS_3 (Syphoviridae) and vB_SmM_GEC-SmitisM_2 (Myoviridae), were specific for E. faecalis and S. mitis, respectively. Each phage's growth patterns and adsorption rates were quantified. Sensitivity to ultraviolet light and temperature was determined, as was host range and serology. The S. mitis bacteriophage was found to be more resistant to ultraviolet light and exposure to high temperatures than the E. faecalis bacteriophage, despite having a much greater rate of replication. While each phage was able to infect a broad range of strains of the same species as the host species from which they were isolated, they were unable to infect other host species tested.

  2. Surface modified stainless steels for PEM fuel cell bipolar plates

    DOEpatents

    Brady, Michael P [Oak Ridge, TN; Wang, Heli [Littleton, CO; Turner, John A [Littleton, CO

    2007-07-24

    A nitridation treated stainless steel article (such as a bipolar plate for a proton exchange membrane fuel cell) having lower interfacial contact electrical resistance and better corrosion resistance than an untreated stainless steel article is disclosed. The treated stainless steel article has a surface layer including nitrogen-modified chromium-base oxide and precipitates of chromium nitride formed during nitridation wherein oxygen is present in the surface layer at a greater concentration than nitrogen. The surface layer may further include precipitates of titanium nitride and/or aluminum oxide. The surface layer in the treated article is chemically heterogeneous surface rather than a uniform or semi-uniform surface layer exclusively rich in chromium, titanium or aluminum. The precipitates of titanium nitride and/or aluminum oxide are formed by the nitriding treatment wherein titanium and/or aluminum in the stainless steel are segregated to the surface layer in forms that exhibit a low contact resistance and good corrosion resistance.

  3. Perchlorate reduction in microbial electrolysis cell with polyaniline modified cathode.

    PubMed

    Li, Jia-Jia; Gao, Ming-Ming; Zhang, Gang; Wang, Xin-Hua; Wang, Shu-Guang; Song, Chao; Xu, Yan-Yan

    2015-02-01

    Excellent perchlorate reduction was obtained under various initial concentrations in a non-membrane microbial electrolysis cell with polyaniline (PANI) modified graphite cathode as sole electron donor. PANI modification is conducive to the formation of biofilm due to its porous structure and good electrocatalytic performance. Compared with cathode without biofilm, over 12% higher reduction rates were acquired in the presence of biocathode. The study demonstrates that, instead of perchlorate reduction, the main contribution of biofilm is involved in facilitate electron transfer from cathode to electrolyte. Interestingly, hairlike structure, referred as to pili-like, was observed in the biofilm as well as in the electrolyte. Additionally, the results show that pili were prone to formation under the condition of external electron field as sole electron donor. Analysis of microbial community suggests that perchlorate reduction bacteria community was most consistent with Azospiraoryzae strain DSM 13638 in the subdivision of the class Proteobacteria.

  4. Automatic Biological Cell Counting Using a Modified Gradient Hough Transform.

    PubMed

    Denimal, Emmanuel; Marin, Ambroise; Guyot, Stéphane; Journaux, Ludovic; Molin, Paul

    2017-02-01

    We present a computational method for pseudo-circular object detection and quantitative characterization in digital images, using the gradient accumulation matrix as a basic tool. This Gradient Accumulation Transform (GAT) was first introduced in 1992 by Kierkegaard and recently used by Kaytanli & Valentine. In the present article, we modify the approach by using the phase coding studied by Cicconet, and by adding a "local contributor list" (LCL) as well as a "used contributor matrix" (UCM), which allow for accurate peak detection and exploitation. These changes help make the GAT algorithm a robust and precise method to automatically detect pseudo-circular objects in a microscopic image. We then present an application of the method to cell counting in microbiological images.

  5. Review of clinical brachytherapy uncertainties: Analysis guidelines of GEC-ESTRO and the AAPM☆

    PubMed Central

    Kirisits, Christian; Rivard, Mark J.; Baltas, Dimos; Ballester, Facundo; De Brabandere, Marisol; van der Laarse, Rob; Niatsetski, Yury; Papagiannis, Panagiotis; Hellebust, Taran Paulsen; Perez-Calatayud, Jose; Tanderup, Kari; Venselaar, Jack L.M.; Siebert, Frank-André

    2014-01-01

    treatment course, taking into account the fractionation schedule and level of image guidance for adaptation. Conclusions This report on brachytherapy clinical uncertainties represents a working project developed by the Brachytherapy Physics Quality Assurances System (BRAPHYQS) subcommittee to the Physics Committee within GEC-ESTRO. Further, this report has been reviewed and approved by the American Association of Physicists in Medicine. PMID:24299968

  6. Antigenically Modified Human Pluripotent Stem Cells Generate Antigen-Presenting Dendritic Cells

    PubMed Central

    Zeng, Jieming; Wu, Chunxiao; Wang, Shu

    2015-01-01

    Human pluripotent stem cells (hPSCs) provide a promising platform to produce dendritic cell (DC) vaccine. To streamline the production process, we investigated a unique antigen-loading strategy that suits this novel platform. Specifically, we stably modified hPSCs using tumour antigen genes in the form of a full-length tumour antigen gene or an artificial tumour antigen epitope-coding minigene. Such antigenically modified hPSCs were able to differentiate into tumour antigen-presenting DCs. Without conventional antigen-loading, DCs derived from the minigene-modified hPSCs were ready to prime a tumour antigen-specific T cell response and further expand these specific T cells in restimulation processes. These expanded tumour antigen-specific T cells were potent effectors with central memory or effector memory phenotype. Thus, we demonstrated that immunocompetent tumour antigen-loaded DCs can be directly generated from antigenically modified hPSCs. Using such strategy, we can completely eliminate the conventional antigen-loading step and significantly simplify the production of DC vaccine from hPSCs. PMID:26471005

  7. Fluorapatite-modified scaffold on dental pulp stem cell mineralization.

    PubMed

    Guo, T; Li, Y; Cao, G; Zhang, Z; Chang, S; Czajka-Jakubowska, A; Nör, J E; Clarkson, B H; Liu, J

    2014-12-01

    In previous studies, fluorapatite (FA) crystal-coated surfaces have been shown to stimulate the differentiation and mineralization of human dental pulp stem cells (DPSCs) in two-dimensional cell culture. However, whether the FA surface can recapitulate these properties in three-dimensional culture is still unknown. This study examined the differences in behavior of human DPSCs cultured on electrospun polycaprolactone (PCL) NanoECM nanofibers with or without the FA crystals. Under near-physiologic conditions, the FA crystals were synthesized on the PCL nanofiber scaffolds. The FA crystals were evenly distributed on the scaffolds. DPSCs were cultured on the PCL+FA or the PCL scaffolds for up to 28 days. Scanning electron microscope images showed that DPSCs attached well to both scaffolds after the initial seeding. However, it appeared that more multicellular aggregates formed on the PCL+FA scaffolds. After 14 days, the cell proliferation on the PCL+FA was slower than that on the PCL-only scaffolds. Interestingly, even without any induction of mineralization, from day 7, the upregulation of several pro-osteogenic molecules (dmp1, dspp, runx2, ocn, spp1, col1a1) was detected in cells seeded on the PCL+FA scaffolds. A significant increase in alkaline phosphatase activity was also seen on FA-coated scaffolds compared with the PCL-only scaffolds at days 14 and 21. At the protein level, osteocalcin expression was induced only in the DPSCs on the PCL+FA surfaces at day 21 and then significantly enhanced at day 28. A similar pattern was observed in those specimens stained with Alizarin red and Von Kossa after 21 and 28 days. These data suggest that the incorporation of FA crystals within the three-dimensional PCL nanofiber scaffolds provided a favorable extracellular matrix microenvironment for the growth, differentiation, and mineralization of human DPSCs. This FA-modified PCL nanofiber scaffold shows promising potential for future bone, dental, and orthopedic regenerative

  8. Detection of Chamber Conditioning by CF4 in the GEC Cell

    NASA Technical Reports Server (NTRS)

    Cruden, Brett A.; Rao, M. V. V. S.; Sharma, S. P.; Meyyappan, M.; Arnold, James (Technical Monitor)

    2001-01-01

    During oxide etch processes, buildup of fluorocarbon residues on reactor sidewalks can cause to drift and will necessitate time for conditioning and cleaning of the reactor. Various measurements in CF4 and Ar plasmas are made in an attempt to identify a metric able to indicate the chamber condition. Mass spectrometry and a Langmuir probe shows that the buildup of fluorocarbon films on the reactor surface causes a decrease in plasma floating potential, plasma potential, and ion energy in argon plasmas. This change in floating potential is also observed in CF4 plasma operation, and occurs primarily during the first hour and a half of plasma operation. A slight rise in electron density is also observed in the argon plasmas. Because the change is seen in an argon plasma, it is indicative of altered physical, not chemical, plasma-surface interactions. Specifically, the insulating films deposited on metal surfaces alter the electromagnetic fields seen by the plasma, affecting various parameters including the floating potential and electron density. An impedance probe placed on the inductive coil shows a slight reduction in plasma impedance due to this rising electron density. The optical emission of several species, including CF, C2, atomic Si and atomic C, is also monitored for changes in density resulting from the buildup of film on the chamber wall. Changes in the optical emission spectrum are comparable to the noise levels in their measurement.

  9. Antibody-Modified Reduced Graphene Oxide Films with Extreme Sensitivity to Circulating Tumor Cells.

    PubMed

    Li, Yingying; Lu, Qihang; Liu, Hongliang; Wang, Jianfeng; Zhang, Pengchao; Liang, Huageng; Jiang, Lei; Wang, Shutao

    2015-11-18

    An antibody-modified reduced graphene oxide (rGO) film with unexpected -extreme sensitivity to circulating tumor cells (CTCs) is reported. The antibody--modified rGO films efficiently capture CTCs from billions of blood cells and minimize the background of white blood cells, without complex microfluidic operations.

  10. Epigenetic Heterogeneity of B-Cell Lymphoma: Chromatin Modifiers

    PubMed Central

    Hopp, Lydia; Nersisyan, Lilit; Löffler-Wirth, Henry; Arakelyan, Arsen; Binder, Hans

    2015-01-01

    We systematically studied the expression of more than fifty histone and DNA (de)methylating enzymes in lymphoma and healthy controls. As a main result, we found that the expression levels of nearly all enzymes become markedly disturbed in lymphoma, suggesting deregulation of large parts of the epigenetic machinery. We discuss the effect of DNA promoter methylation and of transcriptional activity in the context of mutated epigenetic modifiers such as EZH2 and MLL2. As another mechanism, we studied the coupling between the energy metabolism and epigenetics via metabolites that act as cofactors of JmjC-type demethylases. Our study results suggest that Burkitt’s lymphoma and diffuse large B-cell Lymphoma differ by an imbalance of repressive and poised promoters, which is governed predominantly by the activity of methyltransferases and the underrepresentation of demethylases in this regulation. The data further suggest that coupling of epigenetics with the energy metabolism can also be an important factor in lymphomagenesis in the absence of direct mutations of genes in metabolic pathways. Understanding of epigenetic deregulation in lymphoma and possibly in cancers in general must go beyond simple schemes using only a few modes of regulation. PMID:26506391

  11. A Modified Approach to Inducing Bone Marrow Stromal Cells to Differentiate into Cells with Mature Schwann Cell Phenotypes.

    PubMed

    Liu, Yutian; Chen, Jianghai; Liu, Wei; Lu, Xiaocheng; Liu, Zhenyu; Zhao, Xiaobo; Li, Gongchi; Chen, Zhenbing

    2016-02-15

    Marrow stromal cells (MSCs) can be induced to differentiate into Schwann-like cells under classical induction conditions. However, cells derived from this method are unstable, exhibiting a low neurotrophin expression level after the induction conditions are removed. In Schwann cell (SC) culture, progesterone (PROG) enhances neurotrophic synthesis and myelination, specifically regulating the expression of the myelin protein zero (P0)- and peripheral myelin protein 22 (PMP22)-encoding genes by acting in concert or in synergy with insulin and glucocorticoids (GLUCs). In the present study, we investigated whether combined PROG, GLUC, and insulin therapy induced MSCs to differentiate into modified SC-like cells with phenotypes similar to those of mature SCs. After being cultured for 2 weeks in modified differentiation medium, the modified SC-like cells showed increased expression of P0 and PMP22. In addition, morphological and phenotypic characterizations were conducted over a period of over 2 weeks, and functional characteristics persisted for more than 3 weeks after the induction reagents were withdrawn. The transplantation of green fluorescent protein-labeled, modified SC-like cells into transected sciatic nerves with a 10-mm gap significantly increased the proliferation of the original SCs and improved axon regeneration and myelination compared with original BM-SCs. Immunostaining for P0 revealed that more of the transplanted modified SC-like cells retained the phenotypic characteristics of SCs. Taken together, these results reveal that the combined application of PROG, GLUC, and insulin induces MSCs to differentiate into cells with phenotypic, molecular, and functional properties of mature SCs.

  12. Cell Penetrating Peptide-Modified Poly(Lactic-co-Glycolic Acid) Nanoparticles with Enhanced Cell Internalization

    PubMed Central

    Steinbach, Jill M.; Seo, Young-Eun; Saltzman, W. Mark

    2015-01-01

    The surface modification of nanoparticles (NPs) can enhance the intracellular delivery of drugs, proteins, and genetic agents. Here we studied the effect of different surface ligands, including cell penetrating peptides (CPPs), on the cell binding and internalization of poly(lactic-co-glycolic) (PLGA) NPs. Relative to unmodified NPs, we observed that surface-modified NPs greatly enhanced cell internalization. Using one CPP, MPG (unabbreviated notation), that achieved the highest degree of internalization at both low and high surface modification densities, we evaluated the effect of two different NP surface chemistries on cell internalization. After 2 hr, avidin-MPG NPs enhanced cellular internalization by 5 to 26-fold relative to DSPE-MPG NP formulations. Yet, despite a 5-fold increase in MPG density on DSPE- relative to Avidin-NPs, after 24 hr., both formulations resulted in similar internalization levels (48 and 64-fold, respectively). Regardless of surface modification, all NPs were internalized through an energy-dependent, clathrin-mediated process, and became dispersed throughout the cell. Overall both Avidin- and DSPE-CPP modified NPs significantly increased internalization and offer promising delivery options for applications in which internalization presents challenges to efficacious delivery. PMID:26602822

  13. Cell penetrating peptide-modified poly(lactic-co-glycolic acid) nanoparticles with enhanced cell internalization.

    PubMed

    Steinbach, Jill M; Seo, Young-Eun; Saltzman, W Mark

    2016-01-01

    The surface modification of nanoparticles (NPs) can enhance the intracellular delivery of drugs, proteins, and genetic agents. Here we studied the effect of different surface ligands, including cell penetrating peptides (CPPs), on the cell binding and internalization of poly(lactic-co-glycolic) (PLGA) NPs. Relative to unmodified NPs, we observed that surface-modified NPs greatly enhanced cell internalization. Using one CPP, MPG (unabbreviated notation), that achieved the highest degree of internalization at both low and high surface modification densities, we evaluated the effect of two different NP surface chemistries on cell internalization. After 2h, avidin-MPG NPs enhanced cellular internalization by 5 to 26-fold relative to DSPE-MPG NP formulations. Yet, despite a 5-fold increase in MPG density on DSPE compared to Avidin NPs, both formulations resulted in similar internalization levels (48 and 64-fold, respectively) after 24h. Regardless of surface modification, all NPs were internalized through an energy-dependent, clathrin-mediated process, and became dispersed throughout the cell. Overall both Avidin- and DSPE-CPP modified NPs significantly increased internalization and offer promising delivery options for applications in which internalization presents challenges to efficacious delivery.

  14. Elimination of progressive mammary cancer by repeated administrations of chimeric antigen receptor-modified T cells.

    PubMed

    Globerson-Levin, Anat; Waks, Tova; Eshhar, Zelig

    2014-05-01

    Continuous oncogenic processes that generate cancer require an on-going treatment approach to eliminate the transformed cells, and prevent their further development. Here, we studied the ability of T cells expressing a chimeric antibody-based receptor (CAR) to offer a therapeutic benefit for breast cancer induced by erbB-2. We tested CAR-modified T cells (T-bodies) specific to erbB-2 for their antitumor potential in a mouse model overexpressing a human erbB-2 transgene that develops mammary tumors. Comparing the antitumor reactivity of CAR-modified T cells under various therapeutic settings, either prophylactic, prior to tumor development, or therapeutically. We found that repeated administration of CAR-modified T cells is required to eliminate spontaneously developing mammary cancer. Systemic, as well as intratumoral administered CAR-modified T cells accumulated at tumor sites and eventually eliminated the malignant cells. Interestingly, within a few weeks after a single CAR T cells' administration, and rejection of primary lesion, tumors usually relapsed both in treated mammary gland and at remote sites; however, repeated injections of CAR-modified T cells were able to control the secondary tumors. Since spontaneous tumors can arise repeatedly, especially in the case of syndromes characterized by specific susceptibility to cancer, multiple administrations of CAR-modified T cells can serve to control relapsing disease.

  15. Interaction with Epithelial Cells Modifies Airway Macrophage Response to Ozone

    EPA Science Inventory

    The initial innate immune response to ozone (03) in the lung is orchestrated by structural cells, such as epithelial cells, and resident immune cells, such as airway macrophages (Macs). We developed an epithelial cell-Mac coculture model to investigate how epithelial cell-derived...

  16. Interaction of human endothelial cells and nickel-titanium materials modified with silicon ions

    SciTech Connect

    Lotkov, Aleksandr I. Kashin, Oleg A.; Kudryavtseva, Yuliya A. Antonova, Larisa V. Matveeva, Vera G. Sergeeva, Evgeniya A.; Kudryashov, Andrey N.

    2015-10-27

    The paper studies the influence of chemical and phase compositions of NiTi surface layers modified with Si ions by plasma immersion implantation on their interaction with endothelial cells. It is shown that certain technological modes of Si ion implantation enhance the adhesion, proliferation, and viability of endothelial cells. It is found that the Si-modified NiTi surface is capable of stimulating the formation of capillary-like structures in the cell culture.

  17. Interleukin-27 expression modifies prostate cancer cell crosstalk with bone and immune cells in vitro.

    PubMed

    Zolochevska, Olga; Diaz-Quiñones, Adriana O; Ellis, Jayne; Figueiredo, Marxa L

    2013-05-01

    Prostate cancer is frequently associated with bone metastases, where the crosstalk between tumor cells and key cells of the bone microenvironment (osteoblasts, osteoclasts, immune cells) amplifies tumor growth. We have explored the potential of a novel cytokine, interleukin-27 (IL-27), for inhibiting this malignant crosstalk, and have examined the effect of autocrine IL-27 on prostate cancer cell gene expression, as well as the effect of paracrine IL-27 on gene expression in bone and T cells. In prostate tumor cells, IL-27 upregulated genes related to its signaling pathway while downregulating malignancy-related receptors and cytokine genes involved in gp130 signaling, as well as several protease genes. In both undifferentiated and differentiated osteoblasts, IL-27 modulated upregulation of genes related to its own signaling pathway as well as pro-osteogenic genes. In osteoclasts, IL-27 downregulated several genes typically involved in malignancy and also downregulated osteoclastogenesis-related genes. Furthermore, an osteogenesis-focused real-time PCR array revealed a more extensive profile of pro-osteogenic gene changes in both osteoblasts and osteoclasts. In T-lymphocyte cells, IL-27 upregulated several activation-related genes and also genes related to the IL-27 signaling pathway and downregulated several genes that could modulate osteoclastogenesis. Overall, our results suggest that IL-27 may be able to modify interactions between prostate tumor and bone microenvironment cells and thus could be used as a multifunctional therapeutic for restoring bone homeostasis while treating metastatic prostate tumors.

  18. Small cell foams containing a modified dense star polymer or dendrimer as a nucleating agent

    DOEpatents

    Hedstrand, D.M.; Tomalia, D.A.

    1995-02-28

    A small cell foam having a modified dense star polymer or dendrimer is described. This modified dense star polymer or dendrimer has a highly branched interior of one monomeric composition and an exterior structure of a different monomeric composition capable of providing a hydrophobic outer shell and a particle diameter of from about 5 to about 1,000 nm with a matrix polymer.

  19. Small cell foams containing a modified dense star polymer or dendrimer as a nucleating agent

    DOEpatents

    Hedstrand, David M.; Tomalia, Donald A.

    1995-01-01

    A small cell foam having a modified dense star polymer or dendrimer is described. This modified dense star polymer or dendrimer has a highly branched interior of one monomeric composition and an exterior structure of a different monomeric composition capable of providing a hydrophobic outer shell and a particle diameter of from about 5 to about 1,000 nm with a matrix polymer.

  20. Chimeric Antigen Receptors Modified T-Cells for Cancer Therapy.

    PubMed

    Dai, Hanren; Wang, Yao; Lu, Xuechun; Han, Weidong

    2016-07-01

    The genetic modification and characterization of T-cells with chimeric antigen receptors (CARs) allow functionally distinct T-cell subsets to recognize specific tumor cells. The incorporation of costimulatory molecules or cytokines can enable engineered T-cells to eliminate tumor cells. CARs are generated by fusing the antigen-binding region of a monoclonal antibody (mAb) or other ligand to membrane-spanning and intracellular-signaling domains. They have recently shown clinical benefit in patients treated with CD19-directed autologous T-cells. Recent successes suggest that the modification of T-cells with CARs could be a powerful approach for developing safe and effective cancer therapeutics. Here, we briefly review early studies, consider strategies to improve the therapeutic potential and safety, and discuss the challenges and future prospects for CAR T-cells in cancer therapy.

  1. Chimeric Antigen Receptors Modified T-Cells for Cancer Therapy

    PubMed Central

    Dai, Hanren; Wang, Yao; Lu, Xuechun

    2016-01-01

    The genetic modification and characterization of T-cells with chimeric antigen receptors (CARs) allow functionally distinct T-cell subsets to recognize specific tumor cells. The incorporation of costimulatory molecules or cytokines can enable engineered T-cells to eliminate tumor cells. CARs are generated by fusing the antigen-binding region of a monoclonal antibody (mAb) or other ligand to membrane-spanning and intracellular-signaling domains. They have recently shown clinical benefit in patients treated with CD19-directed autologous T-cells. Recent successes suggest that the modification of T-cells with CARs could be a powerful approach for developing safe and effective cancer therapeutics. Here, we briefly review early studies, consider strategies to improve the therapeutic potential and safety, and discuss the challenges and future prospects for CAR T-cells in cancer therapy. PMID:26819347

  2. Gene Transfection toward Spheroid Cells on Micropatterned Culture Plates for Genetically-modified Cell Transplantation.

    PubMed

    Itaka, Keiji; Uchida, Satoshi; Matsui, Akitsugu; Yanagihara, Kayoko; Ikegami, Masaru; Endo, Taisuke; Ishii, Takehiko; Kataoka, Kazunori

    2015-07-31

    To improve the therapeutic effectiveness of cell transplantation, a transplantation system of genetically modified, injectable spheroids was developed. The cell spheroids are prepared in a culture system on micropatterned plates coated with a thermosensitive polymer. A number of spheroids are formed on the plates, corresponding to the cell adhesion areas of 100 µm diameter that are regularly arrayed in a two-dimensional manner, surrounded by non-adhesive areas that are coated by a polyethylene glycol (PEG) matrix. The spheroids can be easily recovered as a liquid suspension by lowering the temperature of the plates, and their structure is well maintained by passing them through injection needles with a sufficiently large caliber (over 27 G). Genetic modification is achieved by gene transfection using the original non-viral gene carrier, polyplex nanomicelle, which is capable of introducing genes into cells without disrupting the spheroid structure. For primary hepatocyte spheroids transfected with a luciferase-expressing gene, the luciferase is sustainably obtained in transplanted animals, along with preserved hepatocyte function, as indicated by albumin expression. This system can be applied to a variety of cell types including mesenchymal stem cells.

  3. A fluorene-modified porphyrin for efficient dye-sensitized solar cells.

    PubMed

    Wu, Cheng-Hua; Pan, Tsung-Yu; Hong, Shang-Hao; Wang, Chin-Li; Kuo, Hshin-Hui; Chu, Yang-Yun; Diau, Eric Wei-Guang; Lin, Ching-Yao

    2012-05-07

    Porphyrins bearing a polyaromatic or a heterocyclic group are prepared to study their fundamental and photovoltaic properties. Solar cells sensitized with a fluorene-modified porphyrin outperform other dyes in the series, reaching ~90% efficiency of N719 dye.

  4. Studies demonstrate modified T cells effective in treating blood-borne cancers

    Cancer.gov

    At the 2013 American Society of Hematology meeting in Dec. 2013, James Kochenderfer, M.D., NCI, presented findings from two clinical trials evaluating the use of genetically modified immune system T cells as cancer therapy. These reports represent import

  5. AAPM and GEC-ESTRO guidelines for image-guided robotic brachytherapy: Report of Task Group 192

    SciTech Connect

    Podder, Tarun K.; Beaulieu, Luc; Caldwell, Barrett; Cormack, Robert A.; Crass, Jostin B.; Dicker, Adam P.; Yu, Yan; Fenster, Aaron; Fichtinger, Gabor; Meltsner, Michael A.; Moerland, Marinus A.; Nath, Ravinder; Rivard, Mark J.; Salcudean, Tim; Song, Danny Y.; Thomadsen, Bruce R.

    2014-10-15

    In the last decade, there have been significant developments into integration of robots and automation tools with brachytherapy delivery systems. These systems aim to improve the current paradigm by executing higher precision and accuracy in seed placement, improving calculation of optimal seed locations, minimizing surgical trauma, and reducing radiation exposure to medical staff. Most of the applications of this technology have been in the implantation of seeds in patients with early-stage prostate cancer. Nevertheless, the techniques apply to any clinical site where interstitial brachytherapy is appropriate. In consideration of the rapid developments in this area, the American Association of Physicists in Medicine (AAPM) commissioned Task Group 192 to review the state-of-the-art in the field of robotic interstitial brachytherapy. This is a joint Task Group with the Groupe Européen de Curiethérapie-European Society for Radiotherapy and Oncology (GEC-ESTRO). All developed and reported robotic brachytherapy systems were reviewed. Commissioning and quality assurance procedures for the safe and consistent use of these systems are also provided. Manual seed placement techniques with a rigid template have an estimated in vivo accuracy of 3–6 mm. In addition to the placement accuracy, factors such as tissue deformation, needle deviation, and edema may result in a delivered dose distribution that differs from the preimplant or intraoperative plan. However, real-time needle tracking and seed identification for dynamic updating of dosimetry may improve the quality of seed implantation. The AAPM and GEC-ESTRO recommend that robotic systems should demonstrate a spatial accuracy of seed placement ≤1.0 mm in a phantom. This recommendation is based on the current performance of existing robotic brachytherapy systems and propagation of uncertainties. During clinical commissioning, tests should be conducted to ensure that this level of accuracy is achieved. These tests

  6. AAPM and GEC-ESTRO guidelines for image-guided robotic brachytherapy: report of Task Group 192.

    PubMed

    Podder, Tarun K; Beaulieu, Luc; Caldwell, Barrett; Cormack, Robert A; Crass, Jostin B; Dicker, Adam P; Fenster, Aaron; Fichtinger, Gabor; Meltsner, Michael A; Moerland, Marinus A; Nath, Ravinder; Rivard, Mark J; Salcudean, Tim; Song, Danny Y; Thomadsen, Bruce R; Yu, Yan

    2014-10-01

    In the last decade, there have been significant developments into integration of robots and automation tools with brachytherapy delivery systems. These systems aim to improve the current paradigm by executing higher precision and accuracy in seed placement, improving calculation of optimal seed locations, minimizing surgical trauma, and reducing radiation exposure to medical staff. Most of the applications of this technology have been in the implantation of seeds in patients with early-stage prostate cancer. Nevertheless, the techniques apply to any clinical site where interstitial brachytherapy is appropriate. In consideration of the rapid developments in this area, the American Association of Physicists in Medicine (AAPM) commissioned Task Group 192 to review the state-of-the-art in the field of robotic interstitial brachytherapy. This is a joint Task Group with the Groupe Européen de Curiethérapie-European Society for Radiotherapy & Oncology (GEC-ESTRO). All developed and reported robotic brachytherapy systems were reviewed. Commissioning and quality assurance procedures for the safe and consistent use of these systems are also provided. Manual seed placement techniques with a rigid template have an estimated in vivo accuracy of 3-6 mm. In addition to the placement accuracy, factors such as tissue deformation, needle deviation, and edema may result in a delivered dose distribution that differs from the preimplant or intraoperative plan. However, real-time needle tracking and seed identification for dynamic updating of dosimetry may improve the quality of seed implantation. The AAPM and GEC-ESTRO recommend that robotic systems should demonstrate a spatial accuracy of seed placement ≤1.0 mm in a phantom. This recommendation is based on the current performance of existing robotic brachytherapy systems and propagation of uncertainties. During clinical commissioning, tests should be conducted to ensure that this level of accuracy is achieved. These tests should

  7. In Vitro Generation of Human NK cells Expressing Chimeric Antigen Receptor through Differentiation of Gene-Modified Hematopoietic Stem Cells

    PubMed Central

    Lowe, Emily; Truscott, Laurel C.; De Oliveira, Satiro N.

    2016-01-01

    Summary NK cells represent a very promising source for adoptive cellular approaches for cancer immunotherapy, and extensive research has been conducted, including clinical trials. Gene modification of NK cells can direct their specificity and enhance their function, but the efficiency of gene transfer techniques is very limited. Here we describe two protocols designed to generate mature human NK cells from gene-modified hematopoietic stem cells. These protocols use chimeric antigen receptor as the transgene, but could potentially be modified for the expression any particular transgene in human NK cells. PMID:27177671

  8. Enhanced Fuel Cell Catalyst Durability with Nitrogen Modified Carbon Supports

    DTIC Science & Technology

    2013-02-12

    materials. enrichment in ruthenium with the N-modified samples as compared to the non-implanted commercial and in-house sputtered samples. Over- all we...found a major difference between commercial and sputtered samples with respect to their ruthenium compositions with the results summarized in Table I. In...commercial catalysts, surface ruthenium is distributed between metallic ruthenium (Ru(0), Ru(II), Ru(IV), ruthe- nium oxide RuO2 and hydrous ruthenium

  9. Adhesion and proliferation of human endothelial cells on photochemically modified polytetrafluoroethylene.

    PubMed

    Gumpenberger, T; Heitz, J; Bäuerle, D; Kahr, H; Graz, I; Romanin, C; Svorcik, V; Leisch, F

    2003-12-01

    We studied the adhesion and proliferation of human endothelial cells on photochemically modified polytetrafluoroethylene samples. The polymer surfaces were modified by exposure to the ultraviolet light of a Xe(2)(*)-excimer lamp at a wavelength of 172 nm in an ammonia atmosphere. Treatment times were between 10 and 20 min. The endothelial cell density was determined 1, 3 and 8 days after seeding by image analysis. Surface modification of the samples resulted in a significant increase in the number of adhering cells and in the formation of a confluent cell layer after 3-8 days. The results were comparable than those obtained on polystyrene Petri dishes, which are used as standard substrates in cell cultivation. Thus modified PTFE appears to be a promising material for the fabrication of artificial vascular prostheses coated with endothelial cells.

  10. A novel modified graphene oxide/chitosan composite used as an adsorbent for Cr(VI) in aqueous solutions.

    PubMed

    Zhang, Li; Luo, Hanjin; Liu, Peipei; Fang, Wei; Geng, Junjie

    2016-06-01

    A novel adsorbent for removal of hexavalent chromium (Cr(VI)) from aqueous solutions has been successfully prepared by modifying graphene oxide/chitosan composite with disodium ethylenediaminetetraacetate (EDTA-2Na) (GEC). This modified composite was characterized by various technologies; including scanning electron microscopy (SEM), Transmission Electron Microscopy (TEM), X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR). Batch adsorption experiments were carried out to evaluate the adsorption of Cr(VI) by GEC under different conditions. The results indicate that the adsorption of Cr(VI) on GEC was highly pH-dependent, with the highest adsorption capacity (86.17mg/g) occurring at pH 2. The kinetics of adsorption exhibited pseudo-second-order behavior. The adsorption data were well described by the Freundlich isotherm model. The adsorption capacity increased with increasing temperature. The calculated thermodynamic parameters indicate that the adsorption is a spontaneous, endothermic and feasible process. The further regeneration experiments showed the adsorption capacity of GEC for Cr(VI) decreased 5% after 7 times reuse, indicating the potential of the as-prepared material for practical application.

  11. Interaction with epithelial cells modifies airway macrophage response to ozone.

    PubMed

    Bauer, Rebecca N; Müller, Loretta; Brighton, Luisa E; Duncan, Kelly E; Jaspers, Ilona

    2015-03-01

    The initial innate immune response to ozone (O3) in the lung is orchestrated by structural cells, such as epithelial cells, and resident immune cells, such as airway macrophages (Macs). We developed an epithelial cell-Mac coculture model to investigate how epithelial cell-derived signals affect Mac response to O3. Macs from the bronchoalveolar lavage (BAL) of healthy volunteers were cocultured with the human bronchial epithelial (16HBE) or alveolar (A549) epithelial cell lines. Cocultures, Mac monocultures, and epithelial cell monocultures were exposed to O3 or air, and Mac immunophenotype, phagocytosis, and cytotoxicity were assessed. Quantities of hyaluronic acid (HA) and IL-8 were compared across cultures and in BAL fluid from healthy volunteers exposed to O3 or air for in vivo confirmation. We show that Macs in coculture had increased markers of alternative activation, enhanced cytotoxicity, and reduced phagocytosis compared with Macs in monoculture that differed based on coculture with A549 or 16HBE. Production of HA by epithelial cell monocultures was not affected by O3, but quantities of HA in the in vitro coculture and BAL fluid from volunteers exposed in vivo were increased with O3 exposure, indicating that O3 exposure impairs Mac regulation of HA. Together, we show epithelial cell-Mac coculture models that have many similarities to the in vivo responses to O3, and demonstrate that epithelial cell-derived signals are important determinants of Mac immunophenotype and response to O3.

  12. [Flocculation and removal of water bloom cells Microcystis aeruginosa by chitosan-modified clays].

    PubMed

    Zou, Hua; Pan, Gang; Chen, Hao

    2004-11-01

    The kinetics of flocculation and removal of Microcystis aeruginosa by chitosan-modified clays was studied. The efficiency of flocculating and removing of algal cells was greatly improved after the modification of the clays. About 80% of algae cell was removed in 0.5 hour, and 90% in 2 hours, when 11 mg/L modified sepiolite was added. Algae-removal capacities of different clays were all improved to a similar level of >90% at a total loading of 11 mg/L after being modified with chitosan. The efficiency of algae-removing was reduced when the clay loading was larger or smaller than the optimum loading.

  13. Micro-organism and cell viability on antimicrobially modified titanium.

    PubMed

    Omori, S; Shibata, Y; Arimoto, T; Igarashi, T; Baba, K; Miyazaki, T

    2009-10-01

    When titanium is anodized by discharge in NaCl solution, both antimicrobial activity and osteoconductivity are conferred. The viability of adherent micro-organisms and cells on antimicrobial titanium remains uncertain. We hypothesized that a thin peroxidation barrier would efficiently destroy adherent bacteria, whereas adherent osteoblastic cells would be viable, since these cells adhere to the surface indirectly though serum proteins. The efficacy of antimicrobial titanium appears to be based on peroxidation, since peroxidation products were detected in parallel with the destruction of bacterial cell-surface structures. The peroxidation effect of antimicrobial titanium was confined to the surface within narrow limits. The viability of osteoblastic cells on the surface was strongly dependent on the presence of serum protein, whereas that of adherent Streptococcus mutans was not affected by the presence of serum proteins. Therefore, differences in the adherent systems used by bacteria and osteoblastic cells are important determinants of their viability on antimicrobial titanium.

  14. Equivalence of Gyn GEC-ESTRO guidelines for image guided cervical brachytherapy with EUD-based dose prescription

    PubMed Central

    2013-01-01

    Background To establish a generalized equivalent uniform dose (gEUD) -based prescription method for Image Guided Brachytherapy (IGBT) that reproduces the Gyn GEC-ESTRO WG (GGE) prescription for cervix carcinoma patients on CT images with limited soft tissue resolution. Methods The equivalence of two IGBT planning approaches was investigated in 20 patients who received external beam radiotherapy (EBT) and 5 concomitant high dose rate IGBT treatments. The GGE planning strategy based on dose to the most exposed 2 cm3 (D2cc) was used to derive criteria for the gEUD-based planning of the bladder and rectum. The safety of gEUD constraints in terms of GGE criteria was tested by maximizing dose to the gEUD constraints for individual fractions. Results The gEUD constraints of 3.55 Gy for the rectum and 5.19 Gy for the bladder were derived. Rectum and bladder gEUD-maximized plans resulted in D2cc averages very similar to the initial GGE criteria. Average D2ccs and EUDs from the full treatment course were comparable for the two techniques within both sets of normal tissue constraints. The same was found for the tumor doses. Conclusions The derived gEUD criteria for normal organs result in GGE-equivalent IGBT treatment plans. The gEUD-based planning considers the entire dose distribution of organs in contrast to a single dose-volume-histogram point. PMID:24225184

  15. GEC-ESTRO/ACROP recommendations for performing bladder-sparing treatment with brachytherapy for muscle-invasive bladder carcinoma.

    PubMed

    Pieters, Bradley R; van der Steen-Banasik, Elzbieta; Smits, Geert A; De Brabandere, Marisol; Bossi, Alberto; Van Limbergen, Erik

    2016-12-31

    The standard treatment for muscle-invasive bladder cancer (MIBC) is a radical cystectomy with pelvic lymph node dissection with or without neoadjuvant chemotherapy. In selected cases a bladder sparing approach is possible, for example a limited surgical excision combined with external beam radiotherapy and brachytherapy. To perform brachytherapy flexible catheters have to be implanted in the bladder wall. The implantation is done either by the open retropubic approach or the endoscopic surgical approach. The largest experience for brachytherapy is with low-dose rate and pulsed-dose rate, although some short-term experience with high-dose rate is also reported. The main advantage for this technique is the conservation of bladder function, with comparable local control rates as for cystectomy series in selected cases. The GEC-ESTRO/ACROP (Groupe Européen de Curiethérapie-European Society for Radiotherapy and Oncology / Advisory Committee on Radiation Oncology Practice) recommendations to perform bladder implantations and brachytherapy as a treatment option for MIBC are described.

  16. Surface modified alginate microcapsules for 3D cell culture

    NASA Astrophysics Data System (ADS)

    Chen, Yi-Wen; Kuo, Chiung Wen; Chueh, Di-Yen; Chen, Peilin

    2016-06-01

    Culture as three dimensional cell aggregates or spheroids can offer an ideal platform for tissue engineering applications and for pharmaceutical screening. Such 3D culture models, however, may suffer from the problems such as immune response and ineffective and cumbersome culture. This paper describes a simple method for producing microcapsules with alginate cores and a thin shell of poly(L-lysine)-graft-poly(ethylene glycol) (PLL-g-PEG) to encapsulate mouse induced pluripotent stem (miPS) cells, generating a non-fouling surface as an effective immunoisolation barrier. We demonstrated the trapping of the alginate microcapsules in a microwell array for the continuous observation and culture of a large number of encapsulated miPS cells in parallel. miPS cells cultured in the microcapsules survived well and proliferated to form a single cell aggregate. Droplet formation of monodisperse microcapsules with controlled size combined with flow cytometry provided an efficient way to quantitatively analyze the growth of encapsulated cells in a high-throughput manner. The simple and cost-effective coating technique employed to produce the core-shell microcapsules could be used in the emerging field of cell therapy. The microwell array would provide a convenient, user friendly and high-throughput platform for long-term cell culture and monitoring.

  17. Improving T cell responses to modified peptides in tumor vaccines.

    PubMed

    Buhrman, Jonathan D; Slansky, Jill E

    2013-03-01

    Immune recognition and elimination of cancerous cells is the primary goal of cancer immunotherapy. However, obstacles including immune tolerance and tumor-induced immunosuppression often limit beneficial immune responses. Vaccination is one proposed intervention that may help to overcome these issues and is an active area of study in cancer immunotherapy. Immunizing with tumor antigenic peptides is a promising, straight-forward vaccine strategy hypothesized to boost preexisting antitumor immunity. However, tumor antigens are often weak T cell agonists, attributable to several mechanisms, including immune self-tolerance and poor immunogenicity of self-derived tumor peptides. One strategy for overcoming these mechanisms is vaccination with mimotopes, or peptide mimics of tumor antigens, which alter the antigen presentation and/or T cell activation to increase the expansion of tumor-specific T cells. Evaluation of mimotope vaccine strategies has revealed that even subtle alterations in peptide sequence can dramatically alter antigen presentation and T cell receptor recognition. Most of this research has been performed using T cell clones, which may not be accurate representations of the naturally occurring antitumor response. The relationship between clones generated after mimotope vaccination and the polyclonal T cell repertoire is unclear. Our work with mimotopes in a mouse model of colon carcinoma has revealed important insights into these issues. We propose that the identification of mimotopes based on stimulation of the naturally responding T cell repertoire will dramatically improve the efficacy of mimotope vaccination.

  18. Efficient inverted polymer solar cells based on conjugated polyelectrolyte and zinc oxide modified ITO electrode

    SciTech Connect

    Yuan, Tao; Zhu, Xiaoguang; Tu, Guoli; Zhou, Lingyu; Zhang, Jian

    2015-02-23

    Efficient inverted polymer solar cells (PSCs) were constructed by utilizing a conjugated polyelectrolyte PF{sub EO}SO{sub 3}Na and zinc oxide to modify the indium tin oxide (ITO) electrode. The ITO electrode modified by PF{sub EO}SO{sub 3}Na and zinc oxide possesses high transparency, increased electron mobility, smoothened surface, and lower work function. PTB7:PC{sub 71}BM inverted PSCs containing the modified ITO electrode achieved a high power conversion efficiency (PCE) of 8.49%, exceeding that of the control device containing a ZnO modified ITO electrode (7.48%). Especially, PCE-10:PC{sub 71}BM inverted polymer solar cells achieved a high PCE up to 9.4%. These results demonstrate a useful approach to improve the performance of inverted polymer solar cells.

  19. Genetically modified cells in regenerative medicine and tissue engineering.

    PubMed

    Sheyn, Dima; Mizrahi, Olga; Benjamin, Shimon; Gazit, Zulma; Pelled, Gadi; Gazit, Dan

    2010-06-15

    Regenerative medicine appears to take as its patron, the Titan Prometheus, whose liver was able to regenerate daily, as the field attempts to restore lost, damaged, or aging cells and tissues. The tremendous technological progress achieved during the last decade in gene transfer methods and imaging techniques, as well as recent increases in our knowledge of cell biology, have opened new horizons in the field of regenerative medicine. Genetically engineered cells are a tool for tissue engineering and regenerative medicine, albeit a tool whose development is fraught with difficulties. Gene-and-cell therapy offers solutions to severe problems faced by modern medicine, but several impediments obstruct the path of such treatments as they move from the laboratory toward the clinical setting. In this review we provide an overview of recent advances in the gene-and-cell therapy approach and discuss the main hurdles and bottlenecks of this approach on its path to clinical trials and prospective clinical practice.

  20. FTIR characterization of animal lung cells: normal and precancerous modified e10 cell line

    NASA Astrophysics Data System (ADS)

    Zezell, D. M.; Pereira, T. M.; Mennecier, G.; Bachmann, L.; Govone, A. B.; Dagli, M. L. Z.

    2012-06-01

    The chemical carcinogens from tobacco are related to over 90% of lung cancers around the world. The risk of death of this kind of cancer is high because the diagnosis usually is made only in advanced stages. Therefore, it is necessary to develop new diagnostic methods for detecting the lung cancer in earlier stages. The Fourier Transform Infrared Spectroscopy (FTIR) can offer high sensibility and accuracy to detect the minimal chemical changes into the biological sample. The aim of this study is to evaluate the differences on infrared spectra between normal lung cells and precancerous lung cells transformed by NNK. Non-cancerous lung cell line e10 (ATCC) and NNK-transformed e10 cell lines were maintained in complete culture medium (1:1 mixture of Dulbecco's modified Eagle's medium and Ham's F12 [DMEM/Ham's F12], supplemented with 100 ng/ml cholera enterotoxin, 10 lg/ml insulin, 0.5 lg/ml. hydrocortisol, 20 ng/ml epidermal growth factor, and 5% horse serum. The cultures were maintained in alcohol 70%. The infrared spectra were acquired on ATR-FTIR Nicolet 6700 spectrophotometer at 4 cm-1 resolution, 30 scans, in the 1800-900 cm-1 spectral range. Each sample had 3 spectra recorded, 30 infrared spectra were obtained from each cell line. The second derivate of spectra indicates that there are displacement in 1646 cm-1 (amine I) and 1255 cm-1(DNA), allowing the possibility to differentiate the two king of cells, with accuracy of 89,9%. These preliminary results indicate that ATR-FTIR is useful to differentiate normal e10 lung cells from precancerous e10 transformed by NNK.

  1. Primordial germ cell differentiation of nuclear transfer embryonic stem cells using surface modified electroconductive scaffolds.

    PubMed

    Eslami-Arshaghi, Tarlan; Vakilian, Saeid; Seyedjafari, Ehsan; Ardeshirylajimi, Abdolreza; Soleimani, Masoud; Salehi, Mohammad

    2016-12-30

    A combination of nanotopographical cues and surface modification of collagen and fibronectin is a potential platform in primordial germ cells (PGCs) differentiation. In the present study, the synergistic effect of nanotopography and surface modification on differentiation of nuclear transfer embryonic stem cells (nt-ESCs) toward PGC lineage was investigated. In order to achieve this goal, poly-anyline (PANi) was mix within poly-L-lactic acid (PLLA). Afterward, the random composite mats were fabricated using PLLA and PANi mix solution. The nanofiber topography notably upregulated the expressions of prdm14, mvh and c-kit compared with tissue culture polystyrene (TCP). Moreover, the combination of nanofiber topography and surface modification resulted in more enhancement of PGCs differentiation compared with non-modified nanofibrous scaffold. Additionally, gene expression results showed that mvh and c-kit were expressed at higher intensity in cells exposed to collagen and fibronectin rather than collagen or fibronectin solitary. These results demonstrated the importance of combined effect of collagen and fibronectin in order to develop a functional extracellular matrix (ECM) mimic in directing stem cell fate and the potential of such biofunctional scaffolds for treatment of infertility.

  2. Drug Delivery via Cell Membrane Fusion Using Lipopeptide Modified Liposomes

    PubMed Central

    2016-01-01

    Efficient delivery of drugs to living cells is still a major challenge. Currently, most methods rely on the endocytotic pathway resulting in low delivery efficiency due to limited endosomal escape and/or degradation in lysosomes. Here, we report a new method for direct drug delivery into the cytosol of live cells in vitro and invivo utilizing targeted membrane fusion between liposomes and live cells. A pair of complementary coiled-coil lipopeptides was embedded in the lipid bilayer of liposomes and cell membranes respectively, resulting in targeted membrane fusion with concomitant release of liposome encapsulated cargo including fluorescent dyes and the cytotoxic drug doxorubicin. Using a wide spectrum of endocytosis inhibitors and endosome trackers, we demonstrate that the major site of cargo release is at the plasma membrane. This method thus allows for the quick and efficient delivery of drugs and is expected to have many invitro, ex vivo, and invivo applications. PMID:27725960

  3. Bone Formation from Porcine Dental Germ Stem Cells on Surface Modified Polybutylene Succinate Scaffolds

    PubMed Central

    2016-01-01

    Designing and providing a scaffold are very important for the cells in tissue engineering. Polybutylene succinate (PBS) has high potential as a scaffold for bone regeneration due to its capacity in cell proliferation and differentiation. Also, stem cells from 3rd molar tooth germs were favoured in this study due to their developmentally and replicatively immature nature. In this study, porcine dental germ stem cells (pDGSCs) seeded PBS scaffolds were used to investigate the effects of surface modification with fibronectin or laminin on these scaffolds to improve cell attachment, proliferation, and osteogenic differentiation for tissue engineering applications. The osteogenic potentials of pDGSCs on these modified and unmodified foams were examined to heal bone defects and the effects of fibronectin or laminin modified PBS scaffolds on pDGSC differentiation into bone were compared for the first time. For this study, MTS assay was used to assess the cytotoxic effects of modified and unmodified surfaces. For the characterization of pDGSCs, flow cytometry analysis was carried out. Besides, alkaline phosphatase (ALP) assay, von Kossa staining, real-time PCR, CM-Dil, and immunostaining were applied to analyze osteogenic potentials of pDGSCs. The results of these studies demonstrated that pDGSCs were differentiated into osteogenic cells on fibronectin modified PBS foams better than those on unmodified and laminin modified PBS foams. PMID:27413380

  4. Nerve cells culture from lumbar spinal cord on surfaces modified by plasma pyrrole polymerization.

    PubMed

    Zuñiga-Aguilar, E; Olayo, R; Ramírez-Fernández, O; Morales, J; Godínez, R

    2014-01-01

    Currently, there are several techniques for modified cell culture surfaces under research to improve cell growth and adhesion. Recently, different methods have been used for surface coating, using biomolecules that enhance cell attachment and growth of nerve cells from spinal cord, such as the use of Poly-DL-Ornithine/Laminin. Plasma-polymerized pyrrole (PPy)-treated surfaces have showed improvement on surfaces biocompatibility with the cells in culture since they do not interfere with any of the biological cell functions. In the present work, we present a novel mouse nerve cell culture technique, using PPy-treated cell culture surfaces. A comparative study of cell survival using Poly-DL-Ornithine/Laminin-treated surfaces was performed. Our results of cell survival when compared with data already reported by other investigators, show that cells cultured on the PPy-modified surface increased survival up to 21 days when compared with Poly-DL-Ornithine/Laminin-coated culture, where 8 days cell survival was obtained. There were electrical and morphological differences in the nerve cells grown in the different surfaces. By comparing the peak ion currents of Poly-DL-Ornithine/Laminin-seeded cells for 8 days with cells grown for 21 days on PPy, an increase of 516% in the Na(+) current and 127% in K(+) currents in cells seeded on PPy were observed. Immunofluorescence techniques showed the presence of cell synapses and culture viability after 21 days. Our results then showed that PPy-modified surfaces are an alternative culture method that increases nerve cells survival from lumbar spinal cord cell culture by preserving its electrical and morphological features.

  5. Conformal nano-thin modified polyelectrolyte coatings for encapsulation of cells.

    PubMed

    Granicka, L H; Antosiak-Iwańska, M; Godlewska, E; Strawski, M; Szklarczyk, M; Maranowski, B; Kowalewski, C; Wiśniewsk, J

    2011-10-01

    Encapsulation of cells in polymeric shells allows for separation of biological material from produced factors, which may find biotechnological and biomedical applications. Human T-lymphocyte cell line Jurkat as well as rat pancreatic islets were encapsulated using LbL technique within shells of polyelectrolyte modified by incorporation of biotin complexed with avidin to improve cell coating and to create the potential ability to elicit specific biochemical responses. The coating with nano-thin modified shells allowed for maintenance of the evaluated cells' integrity and viability during the 8-day culture. The different PE impact may be observed on different biological materials. The islets exhibited lower mitochondrial activity than the Jurkat cells. Nevertheless, coating of cells with polyelectrolyte modified membrane allowed for functioning of both model cell types: 10 μm leukemia cells or 150 μm islets during the culture. Applied membranes maintained the molecular structure during the culture period. The conclusion is that applied modified membrane conformation may be recommended for coating shells for biomedical purposes.

  6. Oxide modified air electrode surface for high temperature electrochemical cells

    DOEpatents

    Singh, Prabhakar; Ruka, Roswell J.

    1992-01-01

    An electrochemical cell is made having a porous cermet electrode (16) and a porous lanthanum manganite electrode (14), with solid oxide electrolyte (15) between them, where the lanthanum manganite surface next to the electrolyte contains a thin discontinuous layer of high surface area cerium oxide and/or praseodymium oxide, preferably as discrete particles (30) in contact with the air electrode and electrolyte.

  7. Cancer Cells Hijack PRC2 to Modify Multiple Cytokine Pathways

    PubMed Central

    Zhao, Michael; Song, Lan; Yu, Tao; Liu, Yu; Liu, Jeffrey C.; McCurdy, Sean; Ma, Anqi; Wither, Joan; Jin, Jian; Zacksenhaus, Eldad; Wrana, Jeffrey L.; Bremner, Rod

    2015-01-01

    Polycomb Repressive Complex 2 (PRC2) is an epigenetic regulator induced in many cancers. It is thought to drive tumorigenesis by repressing division, stemness, and/or developmental regulators. Cancers evade immune detection, and diverse immune regulators are perturbed in different tumors. It is unclear how such cell-specific effects are coordinated. Here, we show a profound and cancer-selective role for PRC2 in repressing multiple cytokine pathways. We find that PRC2 represses hundreds of IFNγ stimulated genes (ISGs), cytokines and cytokine receptors. This target repertoire is significantly broadened in cancer vs non-cancer cells, and is distinct in different cancer types. PRC2 is therefore a higher order regulator of the immune program in cancer cells. Inhibiting PRC2 with either RNAi or EZH2 inhibitors activates cytokine/cytokine receptor promoters marked with bivalent H3K27me3/H3K4me3 chromatin, and augments responsiveness to diverse immune signals. PRC2 inhibition rescues immune gene induction even in the absence of SWI/SNF, a tumor suppressor defective in ~20% of human cancers. This novel PRC2 function in tumor cells could profoundly impact the mechanism of action and efficacy of EZH2 inhibitors in cancer treatment. PMID:26030458

  8. Laser-modified nanostructures of PET films and cell behavior.

    PubMed

    Mirzadeh, Hamid; Moghadam, Ehsan Vahedi; Mivehchi, Houri

    2011-07-01

    The surface of polyethylene terephthalate (PET) films was irradiated using KrF excimer laser (λ = 248 nm) with different number of pulses at constant repetition rate. The adhesion behavior of L-929 fibroblast cells on the irradiated surface was investigated. The changes in films' morphology were characterized by atomic force microscopy (AFM) and scanning electron microscopy (SEM). The hydrophilicity and both polar and dispersion components of the surface tension of the treated films were evaluated by contact angle and surface tension measurement techniques. The films roughness was evaluated by atomic force microscopy. AFM and SEM observations showed that a specific nanostructure was created on the laser-treated polyethylene terephthalate surface. Contact angle and surface energy measurements have indicated an increase in wettability of the laser treated samples up to 5 pulses as optimum result; while, by increasing the laser pulses beyond 5 pulses the hydrophilicity of laser treated samples dropped and the surface energy of the treated films was leveled off. Data from in vitro assays showed significant cell attachment and cell growth onto laser treated samples in comparison with the untreated films. Moreover, a number of fibroblast cells attached and proliferated onto treated PET films were achieved under optimum condition of 5 pulses which was significantly higher than the other treated samples.

  9. A methyl-deficient diet modifies early B cell development.

    PubMed

    Kurogi, Toshiaki; Inoue, Hiroko; Guo, Yun; Nobukiyo, Asako; Nohara, Keiko; Kanno, Masamoto

    2012-01-01

    A functional methyl group donor is essential for the epigenetic regulation of all biological events due to the importance of DNA methylation and histone methylation as an epigenetic marker. However, the epigenetic alterations in the immune system due to methyl donor deficiency are not well known. In this study, we tried to address this question by studying the lymphocyte development and DNA methylation changes caused by a methyl-deficient diet (MDD). We fed one group of C57BL/6J mice with a methyl-sufficient diet (MSD) and the other group with an MDD for 5 months. Flow cytometry analyses of their immune systems showed a decrease in B220+ IgM+ (immature B) cells and an increase in B220+ IgM- (pro/pre-B) cells in the bone marrow of mice fed an MDD. By means of an in vitro OP9 coculture system, we recognized that this B220+ IgM- cell fraction from the MDD has an intrinsic developmental defect. When we quantitatively measured the mRNA expression levels of transcription factors and recombination machinery related to B cell development in the B220+ IgM- cell fraction of their bone marrow, we found that ADA, EBF1, DNTT and Pax5 mRNA expression levels were significantly downregulated in mice fed with an MDD. In addition, there was a drastic decrease in histone methylation profile H3K4me3 in the Pax5 and EBF1 promoters in these B220+ IgM- B cells. However, CpG-DNA methylation profiles had not changed and this revealed that these two promoters are demethylated even under an MSD condition. We also found changed expression levels of the Polycomb group genes (mel18, bmi1, Pc1, Pc2, Ring1A, Ring1B, Ph1) on semi-quantitative RT-PCR. These results indicate that under an MDD condition, early B cell development in bone marrow is easily affected by epigenetic alterations.

  10. Suitability of polyelectrolyte shells modified with fullerene derivate for immunoisolation of cells. Experimental study.

    PubMed

    Borkowska, M; Godlewska, E; Antosiak-Iwańska, M; Kinasiewicz, J; Strawski, M; Szklarczyk, M; Granicka, L H

    2012-12-01

    The polymeric permiselective membranes application for immunoisolation of cells separating the transplanted cells from the host immunological system may eliminate immunosuppressive therapy during transplantation. The suitability of polyelectrolyte modified nanocoatings for immunoisolation of cells was assessed. The polymeric shells modified with incorporated fullerene derivate were applied for encapsulation of human T-lymphocyte cell line Jurkat or rat pancreatic islets of Langerhans using layer-by-layer technique. Hydroxylated fullerene was incorporated to the polyelectrolyte shell for hydrophility increase as well as for layer stability improvement. Evaluation with AFM, FTIR, fluorescence microscopy confirmed the nanocoating presence on the encapsulated cells. It was observed that polylysine-polyethyleneimine membrane with incorporated fullerenol allowed for encapsulated cells functioning in vitro. Membrane conformation applied for encapsulation of pancreatic rat islets allowed for glucose level decline during xenotransplantation into mice. The elaborated nanocoating may be recommended as the possible alternative to the space consuming microencapsulation for biomedical purposes.

  11. Flocculation of cyanobacterial cells using coal fly ash modified chitosan.

    PubMed

    Yuan, Yuting; Zhang, Honggang; Pan, Gang

    2016-06-15

    Harmful algal blooms (HABs) have increasingly occurred worldwide and pose serious threats to water environment safety. In this study, a compound flocculant (CFAL-Chitosan) was developed for HABs mitigation where chitosan was modified by coal fly ash leachate (CFAL). When using optimized dosage of CFAL-Chitosan flocculant, the zeta potential of Microcystis aeruginosa (M.A.) flocs stayed close to zero and algal removal efficiency plateaued over 90% in a wide dosage range from 3 to 6 mg L(-1). For chitosan without CFAL, removal efficiency peaked at 3 mg L(-1) with a maximum removal efficiency of 81%, which quickly decreased as the dosage increased (>3 mg L(-1)) due to the fast reversal of zeta potential. This indicated that CFAL-Chitosan could maintain a better removal efficiency over a wide dosage range as a result of improved charge neutralization compared with the chitosan only treatment. The flocs of CFAL-Chitosan were larger and denser than produced in the presence of chitosan without CFAL. However, excessive CFAL beyond the optimized dose inhibited M.A. removal due to hydrolysis and declining molecular weight of chitosan that weakened the bridging-netting properties, where surface charge reversal happened within a narrow dosage range and the removal-dosage curve became parabolic. The pH and metal residuals that were assumed to pose a threat to the aquatic environment were not significantly affected by adding optimized dosage of CFAL-Chitosan. The study provides a HABs control method using a cheap material of CFA. Further studies are needed to check the potential influence of leachable metals and persistent organic pollutants in CFA under a wide range of environmental condition.

  12. Designing novel Sn-Bi, Si-C and Ge-C nanostructures, using simple theoretical chemical similarities

    PubMed Central

    2011-01-01

    A framework of simple, transparent and powerful concepts is presented which is based on isoelectronic (or isovalent) principles, analogies, regularities and similarities. These analogies could be considered as conceptual extensions of the periodical table of the elements, assuming that two atoms or molecules having the same number of valence electrons would be expected to have similar or homologous properties. In addition, such similar moieties should be able, in principle, to replace each other in more complex structures and nanocomposites. This is only partly true and only occurs under certain conditions which are investigated and reviewed here. When successful, these concepts are very powerful and transparent, leading to a large variety of nanomaterials based on Si and other group 14 elements, similar to well known and well studied analogous materials based on boron and carbon. Such nanomaterias designed in silico include, among many others, Si-C, Sn-Bi, Si-C and Ge-C clusters, rings, nanowheels, nanorodes, nanocages and multidecker sandwiches, as well as silicon planar rings and fullerenes similar to the analogous sp2 bonding carbon structures. It is shown that this pedagogically simple and transparent framework can lead to an endless variety of novel and functional nanomaterials with important potential applications in nanotechnology, nanomedicine and nanobiology. Some of the so called predicted structures have been already synthesized, not necessarily with the same rational and motivation. Finally, it is anticipated that such powerful and transparent rules and analogies, in addition to their predictive power, could also lead to far-reaching interpretations and a deeper understanding of already known results and information. PMID:21711875

  13. Controlling cell-material interactions with polymer nanocomposites by use of surface modifying additives

    NASA Astrophysics Data System (ADS)

    Poole-Warren, L. A.; Farrugia, B.; Fong, N.; Hume, E.; Simmons, A.

    2008-11-01

    Polymer nanocomposites (NC) are fabricated by incorporating well dispersed nanoscale particles within a polymer matrix. This study focuses on elastomeric polyurethane (PU) based nanocomposites, containing organically modified silicates (OMS), as bioactive materials. Nanocomposites incorporating chlorhexidine diacetate as an organic modifier (OM) were demonstrated to be antibacterial with a dose dependence related to both the silicate loading and the loading of OM. When the non-antibacterial OM dodecylamine was used, both cell and platelet adhesion were decreased on the nanocomposite surface. These results suggest that OM is released from the polymer and can impact on cell behaviour at the interface. Nanocomposites have potential use as bioactive materials in a range of biomedical applications.

  14. Recent Progress of Nanostructure Modified Anodes in Microbial Fuel Cells.

    PubMed

    Kim, Marie; Kim, Hyeon Woo; Nam, Joo-Youn; In, Su-Il

    2015-09-01

    Microbial fuel cell (MFC) is a bio-electrochemical system which converts chemical energy into electrical energy by catalytic activity of microorganisms. Electrons produced by microbial oxidation from substrates such as organic matter, complex or renewable biomass are transferred to the anode. Protons produced at the anode migrate to the cathode via the wire and combine with oxygen to form water. Therefore MFC technologies are promising approach for generating electricity or hydrogen gas and wastewater treatment. Electrode materials are one of the keys to increase the power output of MFCs. To improve the cost effective performance of MFCs, various electrodes materials, modifications and configurations have been developed. In this paper, among other recent advances of nanostructured electrodes, especially carbon based anodes, are highlighted. The properties of these electrodes, in terms of surface characteristics, conductivity, modifications, and options were reviewed. The applications, challenges and perspectives of the current MFCs electrode for future development in bio or medical field are briefly discussed.

  15. Technical aspects of the integration of three-dimensional treatment planning dose parameters (GEC-ESTRO Working Group) into pre-implant planning for LDR gynecological interstitial brachytherapy.

    PubMed

    Chi, A; Gao, M; Nguyen, N P; Albuquerque, K

    2009-06-01

    This study investigates the technical feasibility of pre-implant image-based treatment planning for LDR GYN interstitial brachytherapy(IB) based on the GEC-ESTRO guidelines. Initially, a virtual plan is generated based on the prescription dose and GEC-ESTRO defined OAR dose constraints with a pre-implant CT. After the actual implant, a regular diagnostic CT was obtained and fused with our pre-implant scan/initial treatment plan in our planning software. The Flexi-needle position changes, and treatment plan modifications were made if needed. Dose values were normalized to equivalent doses in 2 Gy fractions (LQED 2 Gy) derived from the linear-quadratic model with alpha/beta of 3 for late responding tissues and alpha/beta of 10 for early responding tissues. D(90) to the CTV, which was gross tumor (GTV) at the time of brachytherapy with a margin to count for microscopic disease, was 84.7 +/- 4.9% of the prescribed dose. The OAR doses were evaluated by D(2cc) (EBRT+IB). Mean D(2cc) values (LQED(2Gy)) for the rectum, bladder, sigmoid, and small bowel were the following: 63.7 +/- 8.4 Gy, 61.2 +/- 6.9 Gy, 48.0 +/- 3.5 Gy, and 49.9 +/- 4.2 Gy. This study confirms the feasibility of applying the GEC-ESTRO recommended dose parameters in pre-implant CT-based treatment planning in GYN IB. In the process, this pre-implant technique also demonstrates a good approximation of the target volume dose coverage, and doses to the OARs.

  16. Genetically modified whole-cell bioreporters for environmental assessment

    PubMed Central

    Xu, Tingting; Close, Dan M.; Sayler, Gary S.; Ripp, Steven

    2015-01-01

    Living whole-cell bioreporters serve as environmental biosentinels that survey their ecosystems for harmful pollutants and chemical toxicants, and in the process act as human and other higher animal proxies to pre-alert for unfavorable, damaging, or toxic conditions. Endowed with bioluminescent, fluorescent, or colorimetric signaling elements, bioreporters can provide a fast, easily measured link to chemical contaminant presence, bioavailability, and toxicity relative to a living system. Though well tested in the confines of the laboratory, real-world applications of bioreporters are limited. In this review, we will consider bioreporter technologies that have evolved from the laboratory towards true environmental applications, and discuss their merits as well as crucial advancements that still require adoption for more widespread utilization. Although the vast majority of environmental monitoring strategies rely upon bioreporters constructed from bacteria, we will also examine environmental biosensing through the use of less conventional eukaryotic-based bioreporters, whose chemical signaling capacity facilitates a more human-relevant link to toxicity and health-related consequences. PMID:26594130

  17. Modified SPEEK membranes for direct ethanol fuel cell

    NASA Astrophysics Data System (ADS)

    Maab, Husnul; Nunes, Suzana Pereira

    Membranes with low ethanol crossover were prepared aiming their application for direct ethanol fuel cell (DEFC). They were based on (1) sulfonated poly(ether ether ketone) (SPEEK) coated with carbon molecular sieves (CMS) and (2) on SPEEK/PI homogeneous blends. The membranes were characterized concerning their water and ethanol solution uptake, water and ethanol permeability in pervaporation experiments and their performance in DEFC tests. The ethanol permeabilities for the CMS-coated (180 nm and 400 nm thick layers) SPEEK were 8.5 and 3.1 × 10 -10 kg m s -1 m -2 and for the homogeneous SPEEK/PI blends membranes with 10, 20 and 30 wt.% of PI were 4.4, 1.0 and 0.4 × 10 -10 kg m s -1 m -2 respectively, which is 2- to 50-fold lower than that for plain SPEEK (19 × 10 -10 kg m s -1 m -2). Particularly the SPEEK/PI membranes had substantially better performance than Nafion 117 ® membranes in DEFC tests at 60 °C and 90 °C.

  18. Optimization of the cell seeding density and modeling of cell growth and metabolism using the modified Gompertz model for microencapsulated animal cell culture.

    PubMed

    Wen-tao, Qi; Ying, Zhang; Juan, Ma; Xin, Guo; Yu-bing, Xie; Wei, Wang; Xiaojun, Ma

    2006-04-05

    Cell microencapsulation is one of the promising strategies for the in vitro production of proteins or in vivo delivery of therapeutic products. In order to design and fabricate the optimized microencapsulated cell system, the Gompertz model was applied and modified to describe the growth and metabolism of microencapsulated cell, including substrate consumption and product formation. The Gompertz model successfully described the cell growth kinetics and the modified Gompertz models fitted the substrate consumption and product formation well. It was demonstrated that the optimal initial cell seeding density was about 4-5 x 10(6) cells/mL of microcapsule, in terms of the maximum specific growth rate, the glucose consumption potential and the product formation potential calculated by the Gompertz and modified Gompertz models. Modeling of cell growth and metabolism in microcapsules provides a guideline for optimizing the culture of microencapsulated cells.

  19. HTCC-Modified Nanoclay for Tissue Engineering Applications: A Synergistic Cell Growth and Antibacterial Efficiency

    PubMed Central

    Aliabadi, Majid; Dastjerdi, Roya; Kabiri, Kourosh

    2013-01-01

    This paper deals with the synthesis of a biocompatible chitosan ammonium salt N-(2-hydroxy) propyl-3-trimethylammonium chitosan chloride (HTCC) and using it in montmorillonite ion-exchange process. HTCC-modified montmorillonite (Mt) with different chemical ratios was successfully synthesized, and their characteristics have been verified by XRD and FTIR analyses. Produced samples have been evaluated in terms of antibacterial efficiency and biocompatibility (cell culture test). Antibacterial efficiency of synthesized HTCC/Mt samples has been confirmed against both gram negative bacteria (Escherichia coli) and gram positive bacteria (Staphylococcus aureus). The results disclosed that the antibacterial efficiency of HTCC-modified montmorillonite was unexpectedly even more than HTCC. This excellent synergistic effect has been referred to entrapping bacteria between the intercalated structures of HTCC-modified montmorillonite. Then HTCC on clay layers can seriously attack and damage the entrapped bacteria. An extraordinary biocompatibility, cell attachment, and cell growth even more than tissue culture polystyrene (TCPS) have been recorded in the case of this novel kind of modified clay. Due to existing concerns about serious and chronic infections after implant placement, this natural-based bioactive and antibacterial modified clay can be used in electrospun nanofibers and other polymeric implants with promising mechanical properties for tissue engineering applications. PMID:23998128

  20. New Strategies in Engineering T-Cell Receptor Gene-Modified T Cells to More Effectively Target Malignancies

    PubMed Central

    Schmitt, Thomas M.; Stromnes, Ingunn M.; Chapuis, Aude G.; Greenberg, Philip D.

    2016-01-01

    The immune system, and T cells in particular, have the ability to target and destroy malignant cells. However, anti-tumor immune responses induced from the endogenous T cell repertoire are often insufficient for the eradication of established tumors, as illustrated by the failure of cancer vaccination strategies or checkpoint blockade for most tumors. Genetic modification of T cells to express a defined T cell receptor (TCR) can provide the means to rapidly generate large numbers of tumor-reactive T cells capable of targeting tumor cells in vivo. However, cell-intrinsic factors as well as immunosuppressive factors in the tumor microenvironment can limit the function of such gene-modified T cells. New strategies currently being developed are refining and enhancing this approach, resulting in cellular therapies that more effectively target tumors and that are less susceptible to tumor immune-evasion. PMID:26463711

  1. New Strategies in Engineering T-cell Receptor Gene-Modified T cells to More Effectively Target Malignancies.

    PubMed

    Schmitt, Thomas M; Stromnes, Ingunn M; Chapuis, Aude G; Greenberg, Philip D

    2015-12-01

    The immune system, T cells in particular, have the ability to target and destroy malignant cells. However, antitumor immune responses induced from the endogenous T-cell repertoire are often insufficient for the eradication of established tumors, as illustrated by the failure of cancer vaccination strategies or checkpoint blockade for most tumors. Genetic modification of T cells to express a defined T-cell receptor (TCR) can provide the means to rapidly generate large numbers of tumor-reactive T cells capable of targeting tumor cells in vivo. However, cell-intrinsic factors as well as immunosuppressive factors in the tumor microenvironment can limit the function of such gene-modified T cells. New strategies currently being developed are refining and enhancing this approach, resulting in cellular therapies that more effectively target tumors and that are less susceptible to tumor immune evasion.

  2. Efficient Gene Transduction of Dispersed Islet Cells in Culture Using Fiber-Modified Adenoviral Vectors

    PubMed Central

    Hanayama, Hiroyuki; Ohashi, Kazuo; Utoh, Rie; Shimizu, Hirofumi; Ise, Kazuya; Sakurai, Fuminori; Mizuguchi, Hiroyuki; Tsuchiya, Hiroyuki; Okano, Teruo; Gotoh, Mitsukazu

    2015-01-01

    To establish novel islet-based therapies, our group has recently developed technologies for creating functional neo-islet tissues in the subcutaneous space by transplanting monolithic sheets of dispersed islet cells (islet cell sheets). Improving cellular function and viability are the next important challenges for enhancing the therapeutic effects. This article describes the adenoviral vector-mediated gene transduction of dispersed islet cells under culture conditions. Purified pancreatic islets were obtained from Lewis rats and dissociated into single islet cells. Cells were plated onto laminin-5-coated temperature-responsive polymer poly(N-isopropylacrylamide)-immobilized plastic dishes. At 0 h, islet cells were infected for 1 h with either conventional type 5 adenoviral vector (Ad-CA-GFP) or fiber-modified adenoviral vector (AdK7-CA-GFP) harboring a polylysine (K7) peptide in the C terminus of the fiber knob. We investigated gene transduction efficiency at 48 h after infection and found that AdK7-CA-GFP yielded higher transduction efficiencies than Ad-CA-GFP at a multiplicity of infection (MOI) of 5 and 10. For AdK7-CA-GFP at MOI = 10, 84.4 ± 1.5% of islet cells were found to be genetically transduced without marked vector infection-related cellular damage as determined by viable cell number and lactate dehydrogenase (LDH) release assay. After AdK7-CA-GFP infection at MOI = 10, cells remained attached and expanded to nearly full confluency, showing that this adenoviral infection protocol is a feasible approach for creating islet cell sheets. We have shown that dispersed and cultured islet cells can be genetically modified efficiently using fiber-modified adenoviral vectors. Therefore, this gene therapy technique could be used for cellular modification or biological assessment of dispersed islet cells. PMID:26858906

  3. Magnet-Bead Based MicroRNA Delivery System to Modify CD133+ Stem Cells

    PubMed Central

    Wiekhorst, Frank; Steinhoff, Gustav

    2016-01-01

    Aim. CD133+ stem cells bear huge potential for regenerative medicine. However, low retention in the injured tissue and massive cell death reduce beneficial effects. In order to address these issues, we intended to develop a nonviral system for appropriate cell engineering. Materials and Methods. Modification of human CD133+ stem cells with magnetic polyplexes carrying microRNA was studied in terms of efficiency, safety, and targeting potential. Results. High microRNA uptake rates (~80–90%) were achieved without affecting CD133+ stem cell properties. Modified cells can be magnetically guided. Conclusion. We developed a safe and efficient protocol for CD133+ stem cell modification. Our work may become a basis to improve stem cell therapeutical effects as well as their monitoring with magnetic resonance imaging. PMID:27795713

  4. Gene Expression in Mammalian Cells Using BacMam, a Modified Baculovirus System.

    PubMed

    Fornwald, James A; Lu, Quinn; Boyce, Frederick M; Ames, Robert S

    2016-01-01

    BacMams are modified baculoviruses that contain mammalian expression cassettes for gene delivery and expression in mammalian cells. BacMams have become an integral part of the recombinant mammalian gene expression toolbox in research labs worldwide. Construction of transfer vectors is straightforward using basic molecular biology protocols. Virus generation is based on common methods used with the baculovirus insect cell expression system. BacMam transduction of mammalian cells requires minimal modifications to familiar cell culture methods. This chapter highlights the BacMam transfer vector pHTBV.

  5. A Phase I Study on Adoptive Immunotherapy Using Gene-Modified T Cells for Ovarian Cancer

    PubMed Central

    Kershaw, Michael H.; Westwood, Jennifer A.; Parker, Linda L.; Wang, Gang; Eshhar, Zelig; Mavroukakis, Sharon A.; White, Donald E.; Wunderlich, John R.; Canevari, Silvana; Rogers-Freezer, Linda; Chen, Clara C.; Yang, James C.; Rosenberg, Steven A.; Hwu, Patrick

    2007-01-01

    Purpose A phase I study was conducted to assess the safety of adoptive immunotherapy using gene-modified autologous T cells for the treatment of metastatic ovarian cancer. Experimental Design T cells with reactivity against the ovarian cancer – associated antigen α-folate receptor (FR) were generated by genetic modification of autologous T cells with a chimeric gene incorporating an anti-FR single-chain antibody linked to the signaling domain of the Fc receptor γ chain. Patients were assigned to one of two cohorts in the study. Eight patients in cohort 1received a dose escalation of T cells in combination with high-dose interleukin-2, and six patients in cohort 2 received dual-specific T cells (reactive with both FR and allogeneic cells) followed by immunization with allogeneic peripheral blood mononuclear cells. Results Five patients in cohort 1 experienced some grade 3 to 4 treatment-related toxicity that was probably due to interleukin-2 administration, which could be managed using standard measures. Patients in cohort 2 experienced relatively mild side effects with grade 1to 2 symptoms. No reduction in tumor burden was seen in any patient. Tracking 111In-labeled adoptively transferred T cells in cohort 1revealed a lack of specific localization of T cells to tumor except in one patient where some signal was detected in a peritoneal deposit. PCR analysis showed that gene-modified T cells were present in the circulation in large numbers for the first 2 days after transfer, but these quickly declined to be barely detectable 1month later in most patients. An inhibitory factor developed in the serum of three of six patients tested over the period of treatment, which significantly reduced the ability of gene-modified T cells to respond against FR+ tumor cells. Conclusions Large numbers of gene-modified tumor-reactive T cells can be safely given to patients, but these cells do not persist in large numbers long term. Future studies need to employ strategies to

  6. c-Jun Gene-Modified Schwann Cells: Upregulating Multiple Neurotrophic Factors and Promoting Neurite Outgrowth

    PubMed Central

    Huang, Liangliang; Quan, Xin; Liu, Zhongyang; Ma, Teng; Wu, Yazhen; Ge, Jun; Zhu, Shu; Yang, Yafeng; Liu, Liang; Sun, Zhen

    2015-01-01

    Genetically modified Schwann cells (SCs) that overexpress neurotrophic factors (NFs), especially those that overexpress multiple NFs, hold great potential for promoting nerve regeneration. Currently, only one NF can be upregulated in most genetically modified SCs, and simultaneously upregulating multiple NFs in SCs remains challenging. In this study, we found that the overexpression of c-Jun, a component of the AP-1 transcription factor, effectively upregulated the expression and secretion of multiple NFs, including glial cell line-derived neurotrophic factor, brain-derived neurotrophic factor, artemin, leukemia inhibitory factor, and nerve growth factor. The c-Jun gene-modified SCs showed a normal morphology in scanning electron microscopy and fluorescent staining analysis. In addition, the c-Jun-modified SCs showed enhanced proliferation and migration abilities compared with vector control cells. We used transwell chambers to establish coculture systems imitating the in vivo conditions in which transplanted SCs might influence native SCs and neurons. We found that the c-Jun-modified SCs enhanced native SC migration and promoted the proliferation of native SCs in the presence of axons. Further analysis revealed that in the c-Jun group, the average length and the total area of neurites divided by the total area of the explant body were μm 1180±25 and 6.4±0.4, respectively, which were significantly greater compared with the other groups. These findings raise the possibility of constructing an optimal therapeutic alternative for nerve repair using c-Jun-modified SCs, which have the potential to promote axonal regeneration and functional recovery by upregulating multiple NFs. In addition, these cells exhibit enhanced migration and proliferation abilities, enhance the biological functions of native SCs, and promote neurite outgrowth. PMID:25588149

  7. Enhanced ambient stability of efficient perovskite solar cells by employing a modified fullerene cathode interlayer

    DOE PAGES

    Zhu, Zonglong; Chueh, Chu -Chen; Lin, Francis; ...

    2016-03-22

    A novel fullerene cathode interlayer is employed to facilitate the fabrication of stable and efficient perovskite solar cells. Here, this modified fullerene surfactant significantly increases air stability of the derived devices due to its hydrophobic characteristics to enable 80% of the initial PCE to be retained after being exposed in ambient condition with 20% relative humidity for 14 days.

  8. Cell adhesion on polytetrafluoroethylene modified by UV-irradiation in an ammonia atmosphere.

    PubMed

    Heitz, J; Svorcík, V; Bacáková, L; Rocková, K; Ratajová, E; Gumpenberger, T; Bäuerle, D; Dvoránková, B; Kahr, H; Graz, I; Romanin, C

    2003-10-01

    We report on the modification of polytetrafluoroethylene (PTFE) by exposure to the ultraviolet (UV) light of a Xe(2)*-excimer lamp at a wavelength of 172 nm in an ammonia atmosphere. Typical treatment times were up to 30 min. Subsequently, the samples were grafted with the amino acid alanine from an aqueous solution. The samples were characterized by means of optical transmission spectroscopy, laser-induced fluorescence and contact-angle measurements. We studied the adhesion of rat aortic smooth muscle cells (SMC) and mouse fibroblasts (3T3 cells) to the modified polymer samples using an in vitro technique, where the population density and spread of adhering cells is determined 24 h after seeding by image analysis. For both cell types the exposure of PTFE to UV-light in an ammonia atmosphere resulted in a significant increase in the number of adhering cells and in the size of their spreading area. The grafting with alanine enhanced this effect. Additional experiments with human endothelial cells (HEC) also demonstrated improved adhesion to modified PTFE. Thus, PTFE modified by our method appears to be a promising material for fabrication of artificial vascular prostheses and implants or for cultivation of skin substitutes.

  9. CAR-modified T-cell therapy for cancer: an updated review.

    PubMed

    Haji-Fatahaliha, Mostafa; Hosseini, Maryam; Akbarian, Asiye; Sadreddini, Sanam; Jadidi-Niaragh, Farhad; Yousefi, Mehdi

    2016-09-01

    The use of chimeric antigen receptor (CAR)-modified T cells is a promising approach for cancer immunotherapy. These genetically modified receptors contain an antigen-binding moiety, a hinge region, a transmembrane domain, and an intracellular costimulatory domain resulting in T-cell activation subsequent to antigen binding. Optimal tumor removal through CAR-modified T cells requires suitable target antigen selection, co-stimulatory signaling domain, and the ability of CAR T cells to traffic, persist, and retain antitumor function after adoptive transfer. There are several elements which can improve antitumor function of CAR T cells, including signaling, conditioning chemotherapy and irradiation, tumor burden of the disease, T-cell phenotype, and supplementary cytokine usage. This review outlines four generations of CAR. The pre-clinical and clinical studies showed that this technique has a great potential for treatment of solid and hematological malignancies. The main purpose of the current review is to focus on the pre-clinical and clinical developments of CAR-based immunotherapy.

  10. IL-10/TGF-beta-modified macrophages induce regulatory T cells and protect against adriamycin nephrosis.

    PubMed

    Cao, Qi; Wang, Yiping; Zheng, Dong; Sun, Yan; Wang, Ya; Lee, Vincent W S; Zheng, Guoping; Tan, Thian Kui; Ince, Jon; Alexander, Stephen I; Harris, David C H

    2010-06-01

    IL-10/TGF-beta-modified macrophages, a subset of activated macrophages, produce anti-inflammatory cytokines, suggesting that they may protect against inflammation-mediated injury. Here, macrophages modified ex vivo by IL-10/TGF-beta (IL-10/TGF-beta Mu2) significantly attenuated renal inflammation, structural injury, and functional decline in murine adriamycin nephrosis (AN). These cells deactivated effector macrophages and inhibited CD4+ T cell proliferation. IL-10/TGF-beta Mu2 expressed high levels of the regulatory co-stimulatory molecule B7-H4, induced regulatory T cells from CD4+CD25- T cells in vitro, and increased the number of regulatory T cells in lymph nodes draining the kidneys in AN. The phenotype of IL-10/TGF-beta Mu2 did not switch to that of effector macrophages in the inflamed kidney, and these cells did not promote fibrosis. Taken together, these data demonstrate that IL-10/TGF-beta-modified macrophages effectively protect against renal injury in AN and may become part of a therapeutic strategy for chronic inflammatory disease.

  11. Cell Adhesion and Proliferation on Sulfonated and Non-Modified Chitosan Films.

    PubMed

    Martínez-Campos, Enrique; Civantos, Ana; Redondo, Juan Alfonso; Guzmán, Rodrigo; Pérez-Perrino, Mónica; Gallardo, Alberto; Ramos, Viviana; Aranaz, Inmaculada

    2016-09-15

    Three types of chitosan-based films have been prepared and evaluated: a non-modified chitosan film bearing cationizable aliphatic amines and two films made of N-sulfopropyl chitosan derivatives bearing both aliphatic amines and negative sulfonate groups at different ratios. Cell adhesion and proliferation on chitosan films of C2C12 pre-myoblastic cells and B16 cells as tumoral model have been tested. A differential cell behavior has been observed on chitosan films due to their different surface modification. B16 cells have shown lower vinculin expression when cultured on sulfonated chitosan films. This study shows how the interaction among cells and material surface can be modulated by physicochemical characteristics of the biomaterial surface, altering tumoral cell adhesion and proliferation processes.

  12. [Automatic segmentation of clustered breast cancer cells based on modified watershed algorithm and concavity points searching].

    PubMed

    Tong, Zhen; Pu, Lixin; Dong, Fangjie

    2013-08-01

    As a common malignant tumor, breast cancer has seriously affected women's physical and psychological health even threatened their lives. Breast cancer has even begun to show a gradual trend of high incidence in some places in the world. As a kind of common pathological assist diagnosis technique, immunohistochemical technique plays an important role in the diagnosis of breast cancer. Usually, Pathologists isolate positive cells from the stained specimen which were processed by immunohistochemical technique and calculate the ratio of positive cells which is a core indicator of breast cancer in diagnosis. In this paper, we present a new algorithm which was based on modified watershed algorithm and concavity points searching to identify the positive cells and segment the clustered cells automatically, and then realize automatic counting. By comparison of the results of our experiments with those of other methods, our method can exactly segment the clustered cells without losing any geometrical cell features and give the exact number of separating cells.

  13. Platelet Lysate-Modified Porous Silicon Microparticles for Enhanced Cell Proliferation in Wound Healing Applications.

    PubMed

    Fontana, Flavia; Mori, Michela; Riva, Federica; Mäkilä, Ermei; Liu, Dongfei; Salonen, Jarno; Nicoletti, Giovanni; Hirvonen, Jouni; Caramella, Carla; Santos, Hélder A

    2016-01-13

    The new frontier in the treatment of chronic nonhealing wounds is the use of micro- and nanoparticles to deliver drugs or growth factors into the wound. Here, we used platelet lysate (PL), a hemoderivative of platelets, consisting of a multifactorial cocktail of growth factors, to modify porous silicon (PSi) microparticles and assessed both in vitro and ex vivo the properties of the developed microsystem. PL-modified PSi was assessed for its potential to induce proliferation of fibroblasts. The wound closure-promoting properties of the microsystem were then assessed in an in vitro wound healing assay. Finally, the PL-modified PSi microparticles were evaluated in an ex vivo experiment over human skin. It was shown that PL-modified PSi microparticles were cytocompatible and enhanced the cell proliferation in different experimental settings. In addition, this microsystem promoted the closure of the gap between the fibroblast cells in the wound healing assay, in periods of time comparable with the positive control, and induced a proliferation and regeneration process onto the human skin in an ex vivo experiment. Overall, our results show that PL-modified PSi microparticles are suitable microsystems for further development toward applications in the treatment of chronic nonhealing wounds.

  14. Characterization and Testing the Efficiency of Acinetobacter baumannii Phage vB-GEC_Ab-M-G7 as an Antibacterial Agent

    PubMed Central

    Kusradze, Ia; Karumidze, Natia; Rigvava, Sophio; Dvalidze, Teona; Katsitadze, Malkhaz; Amiranashvili, Irakli; Goderdzishvili, Marina

    2016-01-01

    Acinetobacter baumannii is a gram-negative, non-motile bacterium that, due to its multidrug resistance, has become a major nosocomial pathogen. The increasing number of multidrug resistant (MDR) strains has renewed interest in phage therapy. The aim of our study was to assess the effectiveness of phage administration in Acinetobacter baumannii wound infections in an animal model to demonstrate phage therapy as non-toxic, safe and alternative antibacterial remedy. Using classical methods for the study of bacteriophage properties, we characterized phage vB-GEC_Ab-M-G7 as a dsDNA myovirus with a 90 kb genome size. Important characteristics of vB-GEC_Ab-M-G7include a short latent period and large burst size, wide host range, resistance to chloroform and thermal and pH stability. In a rat wound model, phage application effectively decreased the number of bacteria isolated from the wounds of successfully treated animals. This study highlights the effectiveness of the phage therapy and provides further insight into treating infections caused by MDR strains using phage administration. PMID:27757110

  15. Overview on the dosimetric uncertainty analysis for photon-emitting brachytherapy sources, in the light of the AAPM Task Group No 138 and GEC-ESTRO report

    NASA Astrophysics Data System (ADS)

    DeWerd, Larry A.; Venselaar, Jack L. M.; Ibbott, Geoffrey S.; Meigooni, Ali S.; Stump, Kurt E.; Thomadsen, Bruce R.; Rivard, Mark J.

    2012-10-01

    In 2011, the American Association of Physicists in Medicine (AAPM) and the Groupe Européen de Curiethérapie-European Society for Radiotherapy and Oncology (GEC-ESTRO) published a report pertaining to uncertainties in brachytherapy single-source dosimetry preceding clinical use. The International Organization for Standardization's Guide to the Expression of Uncertainty in Measurement and Technical Note 1297 by the National Institute of Standards and Technology are taken as reference standards for uncertainty formalism. Uncertainties involved in measurements or Monte Carlo methods to estimate brachytherapy dose distributions are provided with discussion of the components intrinsic to the overall dosimetric assessment. The uncertainty propagation from the primary calibration standard through transfer to the clinic for air-kerma strength is given with uncertainties in each of the brachytherapy dosimetry parameters of the AAPM TG-43 dose-calculation formalism. For low-energy and high-energy brachytherapy sources of low dose-rate and high dose-rate, a combined dosimetric uncertainty <5% (k = 1) is estimated, which is consistent with prior literature estimates. Recommendations are provided for clinical medical physicists, dosimetry investigators, and manufacturers of brachytherapy sources and treatment planning systems. These recommendations reflect the guidance of the AAPM and GEC-ESTRO for their members, and may also be used as guidance to manufacturers and regulatory agencies in developing good manufacturing practices for conventional brachytherapy sources used in routine clinical treatments.

  16. Overexpression of AQP3 Modifies the Cell Cycle and the Proliferation Rate of Mammalian Cells in Culture.

    PubMed

    Galán-Cobo, Ana; Ramírez-Lorca, Reposo; Serna, Ana; Echevarría, Miriam

    2015-01-01

    Abnormal AQP3 overexpression in tumor cells of different origins has been reported and a role for this enhanced AQP3 expression in cell proliferation and tumor processess has been indicated. To further understand the role AQP3 plays in cell proliferation we explore the effect that stable over expression of AQP3 produces over the proliferation rate and cell cycle of mammalian cells. The cell cycle was analyzed by flow cytometry with propidium iodide (PI) and the cell proliferation rate measured through cell counting and BrdU staining. Cells with overexpression of AQP3 (AQP3-o) showed higher proliferation rate and larger percentage of cells in phases S and G2/M, than wild type cells (wt). Evaluation of the cell response against arresting the cell cycle with Nocodazole showed that AQP3-o exhibited a less modified cell cycle pattern and lower Annexin V specific staining than wt, consistently with a higher resistance to apoptosis of AQP3-overexpressing cells. The cell volume and complexity were also larger in AQP3-o compared to wt cells. After transcriptomic analysis, RT-qPCR was performed to highlight key molecules implicated in cell proliferation which expression may be altered by overexpression of AQP3 and the comparative analysis between both type of cells showed significant changes in the expression of Zeb2, Jun, JunB, NF-kβ, Cxcl9, Cxcl10, TNF, and TNF receptors. We conclude that the role of AQP3 in cell proliferation seems to be connected to increments in the cell cycle turnover and changes in the expression levels of relevant genes for this process. Larger expression of AQP3 may confer to the cell a more tumor like phenotype and contributes to explain the presence of this protein in many different tumors.

  17. Overexpression of AQP3 Modifies the Cell Cycle and the Proliferation Rate of Mammalian Cells in Culture

    PubMed Central

    Galán-Cobo, Ana; Ramírez-Lorca, Reposo; Serna, Ana; Echevarría, Miriam

    2015-01-01

    Abnormal AQP3 overexpression in tumor cells of different origins has been reported and a role for this enhanced AQP3 expression in cell proliferation and tumor processess has been indicated. To further understand the role AQP3 plays in cell proliferation we explore the effect that stable over expression of AQP3 produces over the proliferation rate and cell cycle of mammalian cells. The cell cycle was analyzed by flow cytometry with propidium iodide (PI) and the cell proliferation rate measured through cell counting and BrdU staining. Cells with overexpression of AQP3 (AQP3-o) showed higher proliferation rate and larger percentage of cells in phases S and G2/M, than wild type cells (wt). Evaluation of the cell response against arresting the cell cycle with Nocodazole showed that AQP3-o exhibited a less modified cell cycle pattern and lower Annexin V specific staining than wt, consistently with a higher resistance to apoptosis of AQP3-overexpressing cells. The cell volume and complexity were also larger in AQP3-o compared to wt cells. After transcriptomic analysis, RT-qPCR was performed to highlight key molecules implicated in cell proliferation which expression may be altered by overexpression of AQP3 and the comparative analysis between both type of cells showed significant changes in the expression of Zeb2, Jun, JunB, NF-kβ, Cxcl9, Cxcl10, TNF, and TNF receptors. We conclude that the role of AQP3 in cell proliferation seems to be connected to increments in the cell cycle turnover and changes in the expression levels of relevant genes for this process. Larger expression of AQP3 may confer to the cell a more tumor like phenotype and contributes to explain the presence of this protein in many different tumors. PMID:26367709

  18. Biocompatibility of pure titanium modified by human endothelial cell-derived extracellular matrix

    NASA Astrophysics Data System (ADS)

    Xue, Xiaoqing; Wang, Jin; Zhu, Ying; Tu, Qiufen; Huang, Nan

    2010-04-01

    Extracellular matrix (ECM) used to modify biomaterial surface is a promising method for improving cardiovascular material hemocompatibility. In the present work, human umbilical vein endothelial cells (HUVECs) are cultured and native ECM is obtained on pure titanium surface. Fourier infrared spectrum (FTIR) test proves the existence of amide I and amide II band on the modified titanium surface. X-ray photoelectron spectroscopy (XPS) further confirms the chemical composition and binding types of the ECM proteins on the titanium substrate. The results of light microscopy and atomic force microscopy (AFM) exhibit the morphology of HUVEC derived ECM. There are higher water contact angles on the ECM modified samples. Furthermore, some ECM components, including fibronectin (FN), laminin (LN) and type IV collagen (IV-COL) are presented on ECM-covered titanium surface by immunofluorescence staining. The biological behavior of cultured HUVECs and adherent platelets on different samples are investigated by in vitro HUVECs culture and platelet adhesion. Cells exhibit better morphology and their proliferation ability greatly improve on the ECM-covered titanium. At the same time, the platelet adhesion and spreading are inhibited on ECM-covered titanium surface. These investigations demonstrate that ECM produced by HUVECs cannot only improve adhesion and proliferation ability of endothelial cell but also inhibit adhesion and activation of platelets. Thus, the approach described here may provide a basis for preparation of modified surface in cardiovascular implants application.

  19. Generation of genetically modified mice using CRISPR/Cas9 and haploid embryonic stem cell systems

    PubMed Central

    JIN, Li-Fang; LI, Jin-Song

    2016-01-01

    With the development of high-throughput sequencing technology in the post-genomic era, researchers have concentrated their efforts on elucidating the relationships between genes and their corresponding functions. Recently, important progress has been achieved in the generation of genetically modified mice based on CRISPR/Cas9 and haploid embryonic stem cell (haESC) approaches, which provide new platforms for gene function analysis, human disease modeling, and gene therapy. Here, we review the CRISPR/Cas9 and haESC technology for the generation of genetically modified mice and discuss the key challenges in the application of these approaches. PMID:27469251

  20. Increased endothelial cell adhesion on plasma modified nanostructured polymeric and metallic surfaces for vascular stent applications.

    PubMed

    Pareta, Rajesh A; Reising, Alexander B; Miller, Tiffany; Storey, Dan; Webster, Thomas J

    2009-06-15

    Techniques to regenerate the vasculature have risen considerably over the last few decades due to the increased clinical diagnosis of artery narrowing and blood vessel blockage. Although initially re-establishing blood flow, current small diameter vascular regenerative materials often eventually cause thrombosis and restenosis due to a lack of initial endothelial cell coverage on such materials. The objective of this in vitro study was to evaluate commonly used vascular materials (specifically, polyethylene terephthalate, polytetrafluoroethylene, polyvinyl chloride, polyurethane, nylon, commercially pure titanium, and a titanium alloy (Ti6Al4V)) modified using an ionic plasma deposition (IPD) process and a nitrogen ion implantation plasma deposition (NIIPD) process. Such surface modifications have been previously shown to create nanostructured surface features which mimic the natural nanostructured surface features of blood vessels. The modified and unmodified surfaces were characterized by scanning electron microscopy, atomic force microscopy and surface energy measurements. Furthermore, in vitro endothelial cell adhesion tests (a key first step for vascular material endothelialization) demonstrated increased endothelial cell adhesion on many modified (with IPD and NIIPD + IPD) compared to unmodified samples. In general, endothelial cell adhesion increased with nanoroughness and surface energy but demonstrated a decreased endothelial cell adhesion trend after an optimal coating surface energy value was reached. Thus, results from this study provided materials and a versatile surface modification process that can potentially increase endothelialization faster than current unmodified (conventional) polymer and metallic vascular materials.

  1. Monitoring of dopamine release in single cell using ultrasensitive ITO microsensors modified with carbon nanotubes.

    PubMed

    Shi, Bao-Xian; Wang, Yu; Zhang, Kai; Lam, Tin-Lun; Chan, Helen Lai-Wa

    2011-02-15

    The study of single cell dynamics has been greatly adapted in biological and medical research and applications. In this work a novel microfluidic electrochemical sensor with carbon nanotubes (CNTs) modified indium tin oxide (ITO) microelectrode was developed for single cells release monitoring. The sensitivity of the electrochemical sensor after CNTs surface modification was improved by 2.5-3 orders of magnitude. The developed CNTs modified ITO sensor was successfully employed to monitor the dopamine release from single living rat pheochromocytoma (PC 12) cells. Its ultrahigh sensitivity, transparency and need for fewer agents enable this smart electrochemical sensor to become a powerful tool in recording dynamic release from various living tissues and organs optically and electrically.

  2. Genetically Modified T-Cell-Based Adoptive Immunotherapy in Hematological Malignancies

    PubMed Central

    Ye, Baixin; Gao, Qingping; Wang, Qiongyu; Zeng, Zhi

    2017-01-01

    A significant proportion of hematological malignancies remain limited in treatment options. Immune system modulation serves as a promising therapeutic approach to eliminate malignant cells. Cytotoxic T lymphocytes (CTLs) play a central role in antitumor immunity; unfortunately, nonspecific approaches for targeted recognition of tumor cells by CTLs to mediate tumor immune evasion in hematological malignancies imply multiple mechanisms, which may or may not be clinically relevant. Recently, genetically modified T-cell-based adoptive immunotherapy approaches, including chimeric antigen receptor (CAR) T-cell therapy and engineered T-cell receptor (TCR) T-cell therapy, promise to overcome immune evasion by redirecting the specificity of CTLs to tumor cells. In clinic trials, CAR-T-cell- and TCR-T-cell-based adoptive immunotherapy have produced encouraging clinical outcomes, thereby demonstrating their therapeutic potential in mitigating tumor development. The purpose of the present review is to (1) provide a detailed overview of the multiple mechanisms for immune evasion related with T-cell-based therapies; (2) provide a current summary of the applications of CAR-T-cell- as well as neoantigen-specific TCR-T-cell-based adoptive immunotherapy and routes taken to overcome immune evasion; and (3) evaluate alternative approaches targeting immune evasion via optimization of CAR-T and TCR-T-cell immunotherapies. PMID:28116322

  3. Effect of deoxynivalenol (DON) on growing pigs and its modification by modified yeast cell wall or modified yeast cell wall and bentonite.

    PubMed

    Shehata, S; Richter, W; Schuster, M; Lindermayer, H

    2004-03-01

    The study examined effect of two adsorbents on the toxicity of Deoxynivalenol (DON) in growing pigs in a feeding trial. 24 male growing pigs (average initial body weight 11.5 kg) were assigned to one of six dietary treatments: control (uncontaminated diet); control + 0.5% adsorbent I; DON contaminated diet (1.73 mg/kg); DON contaminated diet + 0.5% adsorbent I; control + 0.5% adsorbent II and DON contaminated diet + 0.5% adsorbent II. Two digestibility trials were conducted on the second and fourth week of the feeding period with a sampling period of 7 days to determine the digestibility of the nutrients and the amounts of DON in faeces and urine. At the end of the experiments, the pigs were slaughtered, followed by blood haematology and biochemi analys. These data suggest that the addition of 0.5% modified yeast cell wall or a combination of modified yeast cell wall and bentonite to the naturally DON - contaminated diets reduce the effect of DON on the immune system of pigs but do not play an significant role in detoxification of DON in growing pigs.

  4. Genetically Modified Caco-2 Cells With Improved Cytochrome P450 Metabolic Capacity.

    PubMed

    Küblbeck, Jenni; Hakkarainen, Jenni J; Petsalo, Aleksanteri; Vellonen, Kati-Sisko; Tolonen, Ari; Reponen, Petri; Forsberg, Markus M; Honkakoski, Paavo

    2016-02-01

    The human intestinal Caco-2 cell line has been extensively used as a model of small intestinal absorption but it lacks expression and function of cytochrome P450 enzymes, particularly CYP3A4 and CYP2C9, which are normally expressed in the intestinal epithelium. In order to increase the expression and activity of CYP isozymes in these cells, we created 2 novel Caco-2 sublines expressing chimeric constitutive androstane or pregnane X receptors and characterized these cells for their metabolic and absorption properties. In spite of elevated mRNA expression of transporters and differentiation markers, the permeation properties of the modified cell lines did not significantly differ from those of the wild-type cells. In contrast, the metabolic activity was increased beyond the currently used models. Specifically, CYP3A4 activity was increased up to 20-fold as compared to vitamin D treated wild-type Caco-2 cells.

  5. Ligand modified nanoparticles increases cell uptake, alters endocytosis and elevates glioma distribution and internalization.

    PubMed

    Gao, Huile; Yang, Zhi; Zhang, Shuang; Cao, Shijie; Shen, Shun; Pang, Zhiqing; Jiang, Xinguo

    2013-01-01

    Nanoparticles (NPs) were widely used in drugs/probes delivery for improved disease diagnosis and/or treatment. Targeted delivery to cancer cells is a highly attractive application of NPs. However, few studies have been performed on the targeting mechanisms of these ligand-modified delivery systems. Additional studies are needed to understand the transport of nanoparticles in the cancer site, the interactions between nanoparticles and cancer cells, the intracellular trafficking of nanoparticles within the cancer cells and the subcellular destiny and potential toxicity. Interleukin 13 (IL-13) peptide can specifically bind IL-13Rα2, a receptor that is highly expressed on glioma cells but is expressed at low levels on other normal cells. It was shown that the nanoparticels modification with the IL-13 peptide could improve glioma treatment by selectively increasing cellular uptake, facilitating cell internalization, altering the uptake pathway and increasing glioma localization.

  6. Brain-derived neurotrophic factor gene-modified bone marrow mesenchymal stem cells

    PubMed Central

    HAN, ZHONG-MIN; HUANG, HE-MEI; WANG, FEI-FEI

    2015-01-01

    The present study aimed to investigate the effects of human brain-derived neurotrophic factor (hBDNF) on the differentiation of bone marrow mesenchymal stem cells (MSCs) into neuron-like cells. Lentiviral vectors carrying the hBDNF gene were used to modify the bone marrow stromal cells (BMSCs) of Sprague-Dawley (SD) rats. The rat BMSCs were isolated, cultured and identified. A lentivirus bearing hBDNF and enhanced green fluorescent protein (eGFP) genes was subcultured and used to infect the SD rat BMSCs. The expression of eGFP was observed under a fluorescence microscope to determine the infection rate and growth of the transfected cells. Methylthiazolyldiphenyl-tetrazolium bromide (MTT) was used to detect the proliferation rate of cells following transfection. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blot analysis were used to detect the expression levels of hBDNF. Differentiation of neuron-like cells was induced in vitro and the differentiation rate of the induced neural-like cells was compared with that in control groups and analyzed statistically. In the cultured cells, flow cytometry demonstrated positive expression of cluster of differentiation (CD)90 and CD44, and negative expression of CD34 and CD45. The proliferation rate of the rat BMSCs increased following gene transfection. The expression of hBDNF-eGFP was detected in the BMSCs of the experimental group. The differentiation rate of hBDNF-modified cells into neuron-like cells in the experimental group was higher compared with that in empty plasmid and untransfected negative control groups. The difference was statistically significant (P<0.05). Thus, BDNF gene transfection is able to promote the differentiation of BMSCs into neuron-like cells. BDNF may play an important role in the differentiation of MSCs into neuron-like cells. PMID:25574226

  7. Heat-modified citrus pectin induces apoptosis-like cell death and autophagy in HepG2 and A549 cancer cells.

    PubMed

    Leclere, Lionel; Fransolet, Maude; Cote, Francois; Cambier, Pierre; Arnould, Thierry; Van Cutsem, Pierre; Michiels, Carine

    2015-01-01

    Cancer is still one of the leading causes of death worldwide, and finding new treatments remains a major challenge. Previous studies showed that modified forms of pectin, a complex polysaccharide present in the primary plant cell wall, possess anticancer properties. Nevertheless, the mechanism of action of modified pectin and the pathways involved are unclear. Here, we show that citrus pectin modified by heat treatment induced cell death in HepG2 and A549 cells. The induced cell death differs from classical apoptosis because no DNA cleavage was observed. In addition, Z-VAD-fmk, a pan-caspase inhibitor, did not influence the observed cell death in HepG2 cells but appeared to be partly protective in A549 cells, indicating that heat-modified citrus pectin might induce caspase-independent cell death. An increase in the abundance of the phosphatidylethanolamine-conjugated Light Chain 3 (LC3) protein and a decrease in p62 protein abundance were observed in both cell types when incubated in the presence of heat-modified citrus pectin. These results indicate the activation of autophagy. To our knowledge, this is the first time that autophagy has been revealed in cells incubated in the presence of a modified form of pectin. This autophagy activation appears to be protective, at least for A549 cells, because its inhibition with 3-methyladenine increased the observed modified pectin-induced cytotoxicity. This study confirms the potential of modified pectin to improve chemotherapeutic cancer treatments.

  8. Genetically modified Schwann cells producing glial cell line-derived neurotrophic factor inhibit neuronal apoptosis in rat spinal cord injury.

    PubMed

    Liu, Guomin; Wang, Xukai; Shao, Guoxi; Liu, Qinyi

    2014-04-01

    Schwann cells (SCs) are the major cells constituting the peripheral nerve structure and function, and also secret a variety of neurotrophic factors. Schwann cell (SC) transplantation has recently emerged as a promising therapeutic strategy for spinal cord injury (SCI). In the present study, the ability of genetically modified SCs producing high levels of glial cell line‑derived neurotrophic factor (GDNF) to promote spinal cord repair was assessed. The GDNF gene was transduced into SCs. The engineered SCs were characterized by their ability to express and secrete biologically active GDNF, which was shown to inhibit apoptosis of primary rat neurons induced by radiation, and upregulate the expression of B‑cell lymphoma 2 (Bcl‑2) and downregulate the expression of Bcl‑2 associated X protein (Bax) in vitro. Following SC implantation into the spinal cord of adult rats with SCI induced by weight‑drop impact, the survival of rats with transplanted SCs, histology of the spinal cord and expression levels of Bcl‑2 and Bax were examined. Transplantation of unmodified and genetically modified SCs producing GDNF attenuated SCI by inhibiting apoptosis via the Bcl‑2/Bax pathways. The genetically modified SCs demonstrated markedly improved recovery of SCI as compared with unmodified SCs. The present study combined the outgrowth‑promoting property of SCs with the neuroprotective effects of overexpressed GDNF and identified this as a potential novel therapeutic strategy for SCI.

  9. [Cloning goat producing human lactoferrin with genetically modified donor cells selected by single or dual markers].

    PubMed

    An, Liyou; Yuan, Yuguo; Yu, Baoli; Yang, Tingjia; Cheng, Yong

    2012-12-01

    We compared the efficiency of cloning goat using human lactoferrin (hLF) with genetically modified donor cells marked by single (Neo(r)) or double (Neo(r)/GFP) markers. Single marker expression vector (pBLC14) or dual markers expression vector (pAPLM) was delivered to goat fetal fibroblasts (GFF), and then the transgenic GFF was used as donor cells to produce transgenic goats. Respectively, 58.8% (20/34) and 86.7% (26/30) resistant cell lines confirmed the transgenic integration by PCR. Moreover, pAPLM cells lines were subcultured with several passages, only 20% (6/30) cell lines was observed fluorescence from each cell during the cell passage. Somatic cell nuclear transfer using the donor cells harbouring pBLC14 or pAPLM construct, resulting in a total of 806 reconstructed embryos, a pregnancy rate at 35 d (53.8%, 39.1%) and 60 d (26.9%, 21.7%), and an offspring birth rate (1.9%, 1.4%) with 5 and 7 newborn cloned goats, respectively. Transgene was confirmed by PCR and southern-blot in all cloned offspring. There were no significant differences at the reconstructed embryo fusion rates, pregnancy rates and the birth rate (P > 0.05) between single and double markers groups. The Neo(r)/GFP double markers could improve the reliability for accurately and efficiently selecting the genetically modified donor cells. No adverse effect was observed on the efficiency of transgenic goat production by SCNT using somatic cells transfected with double (Neo(r)/GFP) markers vector.

  10. Neurotransplantation of stem cells genetically modified to express human dopamine transporter reduces alcohol consumption

    PubMed Central

    2010-01-01

    Introduction Regulated neurotransmitter actions in the mammalian central nervous system determine brain function and control peripheral organs and behavior. Although drug-seeking behaviors, including alcohol consumption, depend on central neurotransmission, modification of neurotransmitter actions in specific brain nuclei remains challenging. Herein, we report a novel approach for neurotransmission modification in vivo by transplantation of stem cells engineered to take up the neurotransmitter dopamine (DA) efficiently through the action of the human dopamine transporter (hDAT). As a functional test in mice, we used voluntary alcohol consumption, which is known to release DA in nucleus accumbens (NAC), an event hypothesized to help maintain drug-seeking behavior. We reasoned that reducing extracellular DA levels, by engrafting into NAC DA-sequestering stem cells expressing hDAT, would alter alcohol intake. Methods We have generated a neural stem cell line stably expressing the hDAT. Uptake kinetics of DA were determined to select a clone for transplantation. These genetically modified stem cells (or cells transfected with a construct lacking the hDAT sequence) were transplanted bilaterally into the NAC of wild-type mice trained to consume 10% alcohol in a two-bottle free-choice test for alcohol consumption. Alcohol intake was then ascertained for 1 week after transplantation, and brain sections through the NAC were examined for surviving grafted cells. Results Modified stem cells expressed hDAT and uptaken DA selectively via hDAT. Mice accustomed to drinking 10% ethanol by free choice reduced their alcohol consumption after being transplanted with hDAT-expressing stem cells. By contrast, control stem cells lacked that effect. Histologic examination revealed surviving stem cells in the NAC of all engrafted brains. Conclusions Our findings represent proof of principle suggesting that genetically engineered stem cells can be useful for exploring the role of

  11. KRIT1 protein depletion modifies endothelial cell behavior via increased vascular endothelial growth factor (VEGF) signaling.

    PubMed

    DiStefano, Peter V; Kuebel, Julia M; Sarelius, Ingrid H; Glading, Angela J

    2014-11-21

    Disruption of endothelial cell-cell contact is a key event in many cardiovascular diseases and a characteristic of pathologically activated vascular endothelium. The CCM (cerebral cavernous malformation) family of proteins (KRIT1 (Krev-interaction trapped 1), PDCD10, and CCM2) are critical regulators of endothelial cell-cell contact and vascular homeostasis. Here we show novel regulation of vascular endothelial growth factor (VEGF) signaling in KRIT1-depleted endothelial cells. Loss of KRIT1 and PDCD10, but not CCM2, increases nuclear β-catenin signaling and up-regulates VEGF-A protein expression. In KRIT1-depleted cells, increased VEGF-A levels led to increased VEGF receptor 2 (VEGFR2) activation and subsequent alteration of cytoskeletal organization, migration, and barrier function and to in vivo endothelial permeability in KRIT1-deficient animals. VEGFR2 activation also increases β-catenin phosphorylation but is only partially responsible for KRIT1 depletion-dependent disruption of cell-cell contacts. Thus, VEGF signaling contributes to modifying endothelial function in KRIT1-deficient cells and microvessel permeability in Krit1(+/-) mice; however, VEGF signaling is likely not the only contributor to disrupted endothelial cell-cell contacts in the absence of KRIT1.

  12. Chemically Modified Plastic Tube for High Volume Removal and Collection of Circulating Tumor Cells

    PubMed Central

    Gaitas, Angelo; Kim, Gwangseong

    2015-01-01

    In this preliminary effort, we use a commercially available and chemically modified tube to selectively capture circulating tumor cells (CTCs) from the blood stream by immobilizing human anti-EpCAM antibodies on the tube's interior surface. We describe the requisite and critical steps required to modify a tube into a cancer cell-capturing device. Using these simple modifications, we were able to capture or entrap about 85% of cancer cells from suspension and 44% of cancer cells from spiked whole blood. We also found that the percentage of cells captured was dependent on the tube's length and also the number of cancer cells present. It is our strong belief that with the utilization of appropriate tube lengths and procedures, we can ensure capture and removal of nearly the entire CTC population in whole blood. Importantly after a patient’s entire blood volume has circulated through the tube, the tube can then be trypsinized to release the captured live CTCs for further analysis and testing. PMID:26176235

  13. Monitoring of microbial cell viability using nanostructured electrodes modified with Graphene/Alumina nanocomposite.

    PubMed

    Hassan, Rabeay Y A; Mekawy, Moataz M; Ramnani, Pankaj; Mulchandani, Ashok

    2017-05-15

    Microbial infections are rapidly increasing; however most of the existing microbiological and molecular detection methods are time consuming and/or cannot differentiate between the viable and dead cells which may overestimate the risk of infections. Therefore, a bioelectrochemical sensing platform with a high potential to the microbial-electrode interactions was designed based on decorated graphene oxide (GO) sheet with alumina (Al2O3) nanocrystals. GO-Al2O3 nanocomposite was synthesized using self-assembly of GO and Al2O3 and characterized using the scanning electron microscopy (SEM), transmission electron microscopy (TEM), x-ray diffraction (XRD), Raman-spectroscopy, electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). Enhancement of electrocatalytic activity of the composite-modified electrode was demonstrated. Thus, using the GO-Al2O3 nanocomposite modified electrode, the cell viability was determined by monitoring the bioelectrochemical response of the living microbial cells (bacteria and yeast) upon stimulation with carbon source. The bioelectrochemical assay was optimized to obtain high sensitivity and the method was applied to monitor cell viability and screen susceptibility of metabolically active cells (E. coli, B. subtilis, Enterococcus, P. aeruginosa and Salmonella typhi) to antibiotics such as ampicillin and kanamycin. Therefore, the developed assay is suitable for cell proliferation and cytotoxicity testing.

  14. Mitochondria modify exercise-induced development of stem cell-derived neurons in the adult brain.

    PubMed

    Steib, Kathrin; Schäffner, Iris; Jagasia, Ravi; Ebert, Birgit; Lie, D Chichung

    2014-05-07

    Neural stem cells in the adult mammalian hippocampus continuously generate new functional neurons, which modify the hippocampal network and significantly contribute to cognitive processes and mood regulation. Here, we show that the development of new neurons from stem cells in adult mice is paralleled by extensive changes to mitochondrial mass, distribution, and shape. Moreover, exercise-a strong modifier of adult hippocampal neurogenesis-accelerates neuronal maturation and induces a profound increase in mitochondrial content and the presence of mitochondria in dendritic segments. Genetic inhibition of the activity of the mitochondrial fission factor dynamin-related protein 1 (Drp1) inhibits neurogenesis under basal and exercise conditions. Conversely, enhanced Drp1 activity furthers exercise-induced acceleration of neuronal maturation. Collectively, these results indicate that adult hippocampal neurogenesis requires adaptation of the mitochondrial compartment and suggest that mitochondria are targets for enhancing neurogenesis-dependent hippocampal plasticity.

  15. Transplantation of erythropoietin gene-modified neural stem cells improves the repair of injured spinal cord.

    PubMed

    Wu, Min-Fei; Zhang, Shu-Quan; Gu, Rui; Liu, Jia-Bei; Li, Ye; Zhu, Qing-San

    2015-09-01

    The protective effects of erythropoietin on spinal cord injury have not been well described. Here, the eukaryotic expression plasmid pcDNA3.1 human erythropoietin was transfected into rat neural stem cells cultured in vitro. A rat model of spinal cord injury was established using a free falling object. In the human erythropoietin-neural stem cells group, transfected neural stem cells were injected into the rat subarachnoid cavity, while the neural stem cells group was injected with non-transfected neural stem cells. Dulbecco's modified Eagle's medium/F12 medium was injected into the rats in the spinal cord injury group as a control. At 1-4 weeks post injury, the motor function in the rat lower limbs was best in the human erythropoietin-neural stem cells group, followed by the neural stem cells group, and lastly the spinal cord injury group. At 72 hours, compared with the spinal cord injury group, the apoptotic index and Caspase-3 gene and protein expressions were apparently decreased, and the bcl-2 gene and protein expressions were noticeably increased, in the tissues surrounding the injured region in the human erythropoietin-neural stem cells group. At 4 weeks, the cavities were clearly smaller and the motor and somatosensory evoked potential latencies were remarkably shorter in the human erythropoietin-neural stem cells group and neural stem cells group than those in the spinal cord injury group. These differences were particularly obvious in the human erythropoietin-neural stem cells group. More CM-Dil-positive cells and horseradish peroxidase-positive nerve fibers and larger amplitude motor and somatosensory evoked potentials were found in the human erythropoietin-neural stem cells group and neural stem cells group than in the spinal cord injury group. Again, these differences were particularly obvious in the human erythropoietin-neural stem cells group. These data indicate that transplantation of erythropoietin gene-modified neural stem cells into the

  16. 78 FR 69429 - Prospective Grant of Exclusive License: The Development of Modified T-cells for the Treatment of...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-19

    ... Modified T-cells for the Treatment of Multiple Myeloma AGENCY: National Institutes of Health, HHS. ACTION.../ 622,6008 entitled, ``Chimeric Antigen Receptors Targeting B-cell Maturation Antigen'' . The patent... human T-cells directed against B-cell Maturation Antigen (BCMA) for the treatment of multiple...

  17. Edge-Modified Phosphorene Nanoflake Heterojunctions as Highly Efficient Solar Cells.

    PubMed

    Hu, Wei; Lin, Lin; Yang, Chao; Dai, Jun; Yang, Jinlong

    2016-03-09

    We propose to use edge-modified phosphorene nanoflakes (PNFs) as donor and acceptor materials for heterojunction solar cells. By using density functional theory based calculations, we show that heterojunctions consisting of hydrogen- and fluorine-passivated PNFs have a number of desired optoelectronic properties that are suitable for use in a solar cell. We explain why these properties hold for these types of heterojunctions. Our calculations also predict that the maximum energy conversion efficiency of these type of heterojunctions, which can be easily fabricated, can be as high as 20%, making them extremely competitive with other types of two-dimensional heterojunctions.

  18. Poly(imide)/Organically-Modified Montmorillonite Nanocomposite as a Potential Membrane for Alkaline Fuel Cells

    PubMed Central

    Battirola, Liliane C.; Gasparotto, Luiz H. S.; Rodrigues-Filho, Ubirajara P.; Tremiliosi-Filho, Germano

    2012-01-01

    In this work we evaluated the potentiality of a poly(imide) (PI)/organically-modified montmorillonite (O-MMT) nanocomposite membrane for the use in alkaline fuel cells. Both X-ray diffraction and scanning electron microscopy revealed a good dispersion of O-MMT into the PI matrix and preservation of the O-MMT layered structure. When compared to the pure PI, the addition of O-MMT improved thermal stability and markedly increased the capability of absorbing electrolyte and ionic conductivity of the composite. The results show that the PI/O-MMT nanocomposite is a promising candidate for alkaline fuel cell applications. PMID:24958290

  19. Comparative in vitro cytotoxicity of modified deoxynivalenol on porcine intestinal epithelial cells.

    PubMed

    Broekaert, Nathan; Devreese, Mathias; Demeyere, Kristel; Berthiller, Franz; Michlmayr, Herbert; Varga, Elisabeth; Adam, Gerhard; Meyer, Evelyne; Croubels, Siska

    2016-09-01

    The gastrointestinal tract is the first target after ingestion of the mycotoxin deoxynivalenol (DON) via feed and food. Deoxynivalenol is known to affect the proliferation and viability of animal and human intestinal epithelial cells. In addition to DON, feed and food is often co-contaminated with modified forms of DON, such as 3-acetyldeoxynivalenol (3ADON), 15-acetyl-deoxynivalenol (15ADON) and deoxynivalenol-3-β-D-glucoside (DON3G). The goal of this study was to determine the in vitro intrinsic cytotoxicity of these modified forms towards differentiated and proliferative porcine intestinal epithelial cells by means of flow cytometry. Cell death was assessed by dual staining with Annexin-V-fluorescein isothiocyanate (FITC) and propidium iodide (PI), which allows the discrimination of viable (FITC-/PI-), apoptotic (FITC+/PI-) and necrotic cells (FITC+/PI+). Based on the data from the presented pilot in vitro study, it is concluded that cytotoxicity for proliferative cells can be ranked as follows: DON3G ≪ 3ADON < DON≈15ADON.

  20. Fabrication of endothelial progenitor cell capture surface via DNA aptamer modifying dopamine/polyethyleneimine copolymer film

    NASA Astrophysics Data System (ADS)

    Li, Xin; Deng, Jinchuan; Yuan, Shuheng; Wang, Juan; Luo, Rifang; Chen, Si; Wang, Jin; Huang, Nan

    2016-11-01

    Endothelial progenitor cells (EPCs) are mainly located in bone marrow and circulate, and play a crucial role in repairmen of injury endothelium. One of the most promising strategies of stents designs were considered to make in-situ endothelialization in vivo via EPC-capture biomolecules on a vascular graft to capture EPCs directly from circulatory blood. In this work, an EPC specific aptamer with a 34 bases single strand DNA sequence was conjugated onto the stent surface via dopamine/polyethyleneimine copolymer film as a platform and linker. The assembled density of DNA aptamer could be regulated by controlling dopamine percentage in this copolymer film. X-ray photoelectron spectroscopy (XPS), water contact angle (WCA) and fluorescence test confirmed the successful immobilization of DNA aptamer. To confirm its biofunctionality and cytocompatibility, the capturing cells ability of the aptamer modified surface and the effects on the growth behavior of human umbilical vein endothelial cells (HUVECs), smooth muscle cells (SMCs) were investigated. The aptamer functionalized sample revealed a good EPC-capture ability, and had a cellular friendly feature for both EPC and EC growth, while not stimulated the hyperplasia of SMCs. And, the co-culture experiment of three types of cells confirmed the specificity capturing of EPCs to aptamer modified surface, rather than ECs and SMCs. These data suggested that this aptamer functionalized surface may have a large potentiality for the application of vascular grafts with targeted endothelialization.

  1. Growth control of genetically modified cells using an antibody/c-Kit chimera.

    PubMed

    Kaneko, Etsuji; Kawahara, Masahiro; Ueda, Hiroshi; Nagamune, Teruyuki

    2012-05-01

    Gene therapy has been regarded as an innovative potential treatment against serious congenital diseases. However, applications of gene therapy remain limited, partly because its clinical success depends on therapeutic gene-transduced cells acquiring a proliferative advantage. To address this problem, we have developed the antigen-mediated genetically modified cell amplification (AMEGA) system, which uses chimeric receptors to enable the selective proliferation of gene-transduced cells. In this report, we describe mimicry of c-Kit signaling and its application to the AMEGA system. We created an antibody/c-Kit chimera in which the extracellular domain of c-Kit is replaced with an anti-fluorescein single-chain Fv antibody fragment and the extracellular D2 domain of the erythropoietin receptor. A genetically modified mouse pro-B cell line carrying this chimera showed selective expansion in the presence of fluorescein-conjugated BSA (BSA-FL) as a growth inducer. By further engineering the transmembrane domain of the chimera to reduce interchain interaction we attained stricter ligand-dependency. Since c-Kit is an important molecule in the expansion of hematopoietic stem cells (HSCs), this antibody/c-Kit chimera could be a promising tool for gene therapy targeting HSCs.

  2. Hyaluronic acid-modified multiwalled carbon nanotubes for targeted delivery of doxorubicin into cancer cells.

    PubMed

    Cao, Xueyan; Tao, Lei; Wen, Shihui; Hou, Wenxiu; Shi, Xiangyang

    2015-03-20

    Development of novel drug carriers for targeted cancer therapy with high efficiency and specificity is of paramount importance and has been one of the major topics in current nanomedicine. Here we report a general approach to using multifunctional multiwalled carbon nanotubes (MWCNTs) as a platform to encapsulate an anticancer drug doxorubicin (DOX) for targeted cancer therapy. In this approach, polyethyleneimine (PEI)-modified MWCNTs were covalently conjugated with fluorescein isothiocyanate (FI) and hyaluronic acid (HA). The formed MWCNT/PEI-FI-HA conjugates were characterized via different techniques and were used as a new carrier system to encapsulate the anticancer drug doxorubicin for targeted delivery to cancer cells overexpressing CD44 receptors. We show that the formed MWCNT/PEI-FI-HA/DOX complexes with a drug loading percentage of 72% are water soluble and stable. In vitro release studies show that the drug release rate under an acidic condition (pH 5.8, tumor cell microenvironment) is higher than that under physiological condition (pH 7.4). Cell viability assay demonstrates that the carrier material has good biocompatibility in the tested concentration range, and the MWCNT/PEI-FI-HA/DOX complexes can specifically target cancer cells overexpressing CD44 receptors and exert growth inhibition effect to the cancer cells. The developed HA-modified MWCNTs hold a great promise to be used as an efficient anticancer drug carrier for tumor-targeted chemotherapy.

  3. Review: Current clinical applications of chimeric antigen receptor (CAR) modified T cells.

    PubMed

    Geyer, Mark B; Brentjens, Renier J

    2016-11-01

    The past several years have been marked by extraordinary advances in clinical applications of immunotherapy. In particular, adoptive cellular therapy utilizing chimeric antigen receptor (CAR)-modified T cells targeted to CD19 has demonstrated substantial clinical efficacy in children and adults with relapsed or refractory B-cell acute lymphoblastic leukemia (B-ALL) and durable clinical benefit in a smaller subset of patients with relapsed or refractory chronic lymphocytic leukemia (CLL) or B-cell non-Hodgkin lymphoma (B-NHL). Early-phase clinical trials are currently assessing CAR T-cell safety and efficacy in additional malignancies. Here, we discuss clinical results from the largest series to date investigating CD19-targeted CAR T cells in B-ALL, CLL, and B-NHL, including discussion of differences in CAR T-cell design and production and treatment approach, as well as clinical efficacy, nature of severe cytokine release syndrome and neurologic toxicities, and CAR T-cell expansion and persistence. We additionally review the current and forthcoming use of CAR T cells in multiple myeloma and several solid tumors and highlight challenges and opportunities afforded by the current state of CAR T-cell therapies, including strategies to overcome inhibitory aspects of the tumor microenvironment and enhance antitumor efficacy.

  4. [Cancer vaccine therapy using genetically modified induced pluripotent stem cell-derived dendritic cells expressing the TAA gene].

    PubMed

    Iwamoto, Hiromitsu; Ojima, Toshiyasu; Nakamori, Mikihito; Nakamura, Masaki; Hayata, Keiji; Katsuda, Masahiro; Iida, Takeshi; Miyazawa, Motoki; Iwahashi, Makoto; Yamaue, Hiroki

    2013-11-01

    It is generally accepted that the difficulty in obtaining a sufficient number of functional dendritic cells (DCs) poses a serious problem in DC-based immunotherapy. Therefore, we used induced pluripotent stem (iPS) cell-derived DCs (iPSDCs) instead. If the therapeutic efficacy of iPSDCs was equivalent to that of bone marrow-derived DCs( BMDCs), then the above-mentioned problems may be solved. In this study, we generated iPSDCs from iPS cells and compared their capacity to mature and migrate to the regional lymph nodes with that of BMDCs. We adenovirally transduced the hgp100 gene, which codes for a natural tumor antigen, into the DCs and immunized the mice with these genetically modified DCs. The cytotoxic activity of CD8( +) cytotoxic T lymphocytes( CTLs) was assayed using a 51Cr-release assay. The therapeutic efficacy of the vaccination was examined in a subcutaneous tumor model. Our results demonstrated that iPSDCs equaled BMDCs in terms of their maturation and migration capacity. Furthermore, hgp100-specific CTLs were generated in mice that were immunized with the genetically modified iPSDCs. These CTLs exhibited a high level of cytotoxicity against B16 cells, which is similar to that exhibited by CTLs generated in BMDCs immunized mice. Moreover, vaccination with genetically modified iPSDCs elicited a high level of therapeutic efficacy equaling that of vaccination with BMDCs. This study clarified experimentally that genetically modified iPSDCs are equivalent to BMDCs in terms of tumor-associated antigen-specific therapeutic antitumor immunity. This vaccination strategy may therefore be useful for future clinical application as a cancer vaccine.

  5. Polysaccharide-degrading Enzymes are Unable to Attack Plant Cell Walls without Prior Action by a "Wall-modifying Enzyme".

    PubMed

    Karr, A L; Albersheim, P

    1970-07-01

    A study of the degradation of plant cell walls by the mixture of enzymes present in Pectinol R-10 is described. A "wall-modifying enzyme" has been purified from this mixture by a combination of diethylaminoethyl cellulose, Bio Gel P-100, and carboxymethyl cellulose chromatography. Treatment of cell walls with the "wall-modifying enzyme" is shown to be a necessary prerequisite to wall degradation catalyzed by a mixture of polysaccharide-degrading enzymes prepared from Pectinol R-10 or by an alpha-galactosidase secreted by the pathogenic fungus Colletotrichum lindemuthianum. The action of the "wall-modifying enzyme" on cell walls is shown to result in both a release of water-soluble, 70% ethanol-insoluble polymers and an alteration of the residual cell wall. A purified preparation of the "wall-modifying enzyme" is unable to degrade a wide variety of polysaccharide, glycoside, and peptide substrates. However, the purified preparation of wall-modifying enzyme has a limited ability to degrade polygalacturonic acid. The fact that polygalacturonic acid inhibits the ability of the "wall-modifying enzyme" to affect cell walls suggests that the "wall-modifying enzyme" may be responsible for the limited polygalacturonic acid-degrading activity present in the purified preparation. The importance of a wall-modifying enzyme in developmental processes and in pathogenesis is discussed.

  6. Lenalidomide enhances antitumor functions of chimeric antigen receptor modified T cells.

    PubMed

    Otáhal, Pavel; Průková, Dana; Král, Vlastimil; Fabry, Milan; Vočková, Petra; Latečková, Lucie; Trněný, Marek; Klener, Pavel

    2016-04-01

    Tumor immunotherapy based on the use of chimeric antigen receptor modified T cells (CAR T cells) is a promising approach for the treatment of refractory hematological malignancies. However, a robust response mediated by CAR T cells is observed only in a minority of patients and the expansion and persistence of CAR T cells in vivo is mostly unpredictable.Lenalidomide (LEN) is an immunomodulatory drug currently approved for the treatment of multiple myeloma (MM) and mantle cell lymphoma, while it is clinically tested in the therapy of diffuse large B-cell lymphoma of activated B cell immunophenotype. LEN was shown to increase antitumor immune responses at least partially by modulating the activity of E3 ubiquitin ligase Cereblon, which leads to increased ubiquitinylation of Ikaros and Aiolos transcription factors, which in turn results in changed expression of various receptors on the surface of tumor cells. In order to enhance the effectiveness of CAR-based immunotherapy, we assessed the anti-lymphoma efficacy of LEN in combination with CAR19 T cells or CAR20 T cells in vitro and in vivo using various murine models of aggressive B-cell non-Hodgkin lymphomas (B-NHL).Immunodeficient NSG mice were transplanted with various human B-NHL cells followed by treatment with CAR19 or CAR20 T cells with or without LEN. Next, CAR19 T cells were subjected to series of tests in vitro to evaluate their response and signaling capacity following recognition of B cell in the presence or absence of LEN.Our data shows that LEN significantly enhances antitumor functions of CAR19 and CAR20 T cells in vivo. Additionally, it enhances production of interferon gamma by CAR19 T cells and augments cell signaling via CAR19 protein in T cells in vitro. Our data further suggests that LEN works through direct effects on T cells but not on B-NHL cells. The biochemical events underlying this costimulatory effect of LEN are currently being investigated. In summary, our data supports the use of LEN for

  7. Lenalidomide enhances antitumor functions of chimeric antigen receptor modified T cells

    PubMed Central

    Otáhal, Pavel; Průková, Dana; Král, Vlastimil; Fabry, Milan; Vočková, Petra; Latečková, Lucie; Trněný, Marek; Klener, Pavel

    2016-01-01

    ABSTRACT Tumor immunotherapy based on the use of chimeric antigen receptor modified T cells (CAR T cells) is a promising approach for the treatment of refractory hematological malignancies. However, a robust response mediated by CAR T cells is observed only in a minority of patients and the expansion and persistence of CAR T cells in vivo is mostly unpredictable.Lenalidomide (LEN) is an immunomodulatory drug currently approved for the treatment of multiple myeloma (MM) and mantle cell lymphoma, while it is clinically tested in the therapy of diffuse large B-cell lymphoma of activated B cell immunophenotype. LEN was shown to increase antitumor immune responses at least partially by modulating the activity of E3 ubiquitin ligase Cereblon, which leads to increased ubiquitinylation of Ikaros and Aiolos transcription factors, which in turn results in changed expression of various receptors on the surface of tumor cells. In order to enhance the effectiveness of CAR-based immunotherapy, we assessed the anti-lymphoma efficacy of LEN in combination with CAR19 T cells or CAR20 T cells in vitro and in vivo using various murine models of aggressive B-cell non-Hodgkin lymphomas (B-NHL).Immunodeficient NSG mice were transplanted with various human B-NHL cells followed by treatment with CAR19 or CAR20 T cells with or without LEN. Next, CAR19 T cells were subjected to series of tests in vitro to evaluate their response and signaling capacity following recognition of B cell in the presence or absence of LEN.Our data shows that LEN significantly enhances antitumor functions of CAR19 and CAR20 T cells in vivo. Additionally, it enhances production of interferon gamma by CAR19 T cells and augments cell signaling via CAR19 protein in T cells in vitro. Our data further suggests that LEN works through direct effects on T cells but not on B-NHL cells. The biochemical events underlying this costimulatory effect of LEN are currently being investigated. In summary, our data supports the use

  8. Time resolved study of cell death mechanisms induced by amine-modified polystyrene nanoparticles

    NASA Astrophysics Data System (ADS)

    Wang, Fengjuan; Bexiga, Mariana G.; Anguissola, Sergio; Boya, Patricia; Simpson, Jeremy C.; Salvati, Anna; Dawson, Kenneth A.

    2013-10-01

    Positively charged polymers and nanoparticles (NPs) can be toxic to cells in various systems. Using human astrocytoma cells, we have previously shown that 50 nm amine-modified polystyrene NPs damage mitochondria and induce cell death by apoptosis. Here we provide comprehensive details of the cellular events occurring after exposure to the NPs in a time-resolved manner. We demonstrate that the accumulation of NPs in lysosomes plays a central role in the observed cell death, leading to swelling of the lysosomes and release of cathepsins into the cytosol, which ultimately propagates the damage to the mitochondria with subsequent activation of apoptosis. This is accompanied and sustained by other events, such as increasing ROS levels and autophagy. Using various inhibitors, we also show the interplay between apoptosis and autophagy as a response to NP accumulation in lysosomes.Positively charged polymers and nanoparticles (NPs) can be toxic to cells in various systems. Using human astrocytoma cells, we have previously shown that 50 nm amine-modified polystyrene NPs damage mitochondria and induce cell death by apoptosis. Here we provide comprehensive details of the cellular events occurring after exposure to the NPs in a time-resolved manner. We demonstrate that the accumulation of NPs in lysosomes plays a central role in the observed cell death, leading to swelling of the lysosomes and release of cathepsins into the cytosol, which ultimately propagates the damage to the mitochondria with subsequent activation of apoptosis. This is accompanied and sustained by other events, such as increasing ROS levels and autophagy. Using various inhibitors, we also show the interplay between apoptosis and autophagy as a response to NP accumulation in lysosomes. Electronic supplementary information (ESI) available: additional analysis of flow cytometry results, western blots and experiments with cathepsin inhibitors. See DOI: 10.1039/c3nr03249c

  9. The blood and vascular cell compatibility of heparin-modified ePTFE vascular grafts

    PubMed Central

    Hoshi, Ryan A.; Van Lith, Robert; Jen, Michele C.; Allen, Josephine B.; Lapidos, Karen A.; Ameer, Guillermo

    2014-01-01

    Prosthetic vascular grafts do not mimic the antithrombogenic properties of native blood vessels and therefore have higher rates of complications that involve thrombosis and restenosis. We developed an approach for grafting bioactive heparin, a potent anticoagulant glycosaminoglycan, to the lumen of ePTFE vascular grafts to improve their interactions with blood and vascular cells. Heparin was bound to aminated poly(1,8-octanediol-co-citrate) (POC) via its carboxyl functional groups onto POC-modified ePTFE grafts. The bioactivity and stability of the POC-immobilized heparin (POC–Heparin) were characterized via platelet adhesion and clotting assays. The effects of POC–Heparin on the adhesion, viability and phenotype of primary endothelial cells (EC), blood outgrowth endothelial cells (BOECs) obtained from endothelial progenitor cells (EPCs) isolated from human peripheral blood, and smooth muscle cells were also investigated. POC–Heparin grafts maintained bioactivity under physiologically relevant conditions in vitro for at least one month. Specifically, POC–Heparin-coated ePTFE grafts significantly reduced platelet adhesion and inhibited whole blood clotting kinetics. POC–Heparin supported EC and BOEC adhesion, viability, proliferation, NO production, and expression of endothelial cell-specific markers von Willebrand factor (vWF) and vascular endothelial-cadherin (VE-cadherin). Smooth muscle cells cultured on POC–Heparin showed increased expression of α-actin and decreased cell proliferation. This approach can be easily adapted to modify other blood contacting devices such as stents where antithrombogenicity and improved endothelialization are desirable properties. PMID:23069711

  10. Transient delivery of modified mRNA encoding TERT rapidly extends telomeres in human cells

    PubMed Central

    Ramunas, John; Yakubov, Eduard; Brady, Jennifer J.; Corbel, Stéphane Y.; Holbrook, Colin; Brandt, Moritz; Stein, Jonathan; Santiago, Juan G.; Cooke, John P.; Blau, Helen M.

    2015-01-01

    Telomere extension has been proposed as a means to improve cell culture and tissue engineering and to treat disease. However, telomere extension by nonviral, nonintegrating methods remains inefficient. Here we report that delivery of modified mRNA encoding TERT to human fibroblasts and myoblasts increases telomerase activity transiently (24–48 h) and rapidly extends telomeres, after which telomeres resume shortening. Three successive transfections over a 4 d period extended telomeres up to 0.9 kb in a cell type-specific manner in fibroblasts and myoblasts and conferred an additional 28 ± 1.5 and 3.4 ± 0.4 population doublings (PDs), respectively. Proliferative capacity increased in a dose-dependent manner. The second and third transfections had less effect on proliferative capacity than the first, revealing a refractory period. However, the refractory period was transient as a later fourth transfection increased fibroblast proliferative capacity by an additional 15.2 ± 1.1 PDs, similar to the first transfection. Overall, these treatments led to an increase in absolute cell number of more than 1012-fold. Notably, unlike immortalized cells, all treated cell populations eventually stopped increasing in number and expressed senescence markers to the same extent as untreated cells. This rapid method of extending telomeres and increasing cell proliferative capacity without risk of insertional mutagenesis should have broad utility in disease modeling, drug screening, and regenerative medicine.—Ramunas, J., Yakubov, E., Brady, J. J., Corbel, S. Y., Holbrook, C., Brandt, M., Stein, J., Santiago, J. G., Cooke, J. P., Blau, H. M. Transient delivery of modified mRNA encoding TERT rapidly extends telomeres in human cells. PMID:25614443

  11. Human Dendritic Cells Derived From Embryonic Stem Cells Stably Modified With CD1d Efficiently Stimulate Antitumor Invariant Natural Killer T Cell Response

    PubMed Central

    2014-01-01

    Invariant natural killer T (iNKT) cells are a unique lymphocyte subpopulation that mediates antitumor activities upon activation. A current strategy to harness iNKT cells for cancer treatment is endogenous iNKT cell activation using patient-derived dendritic cells (DCs). However, the limited number and functional defects of patient DCs are still the major challenges for this therapeutic approach. In this study, we investigated whether human embryonic stem cells (hESCs) with an ectopically expressed CD1d gene could be exploited to address this issue. Using a lentivector carrying an optimized expression cassette, we generated stably modified hESC lines that consistently overexpressed CD1d. These modified hESC lines were able to differentiate into DCs as efficiently as the parental line. Most importantly, more than 50% of such derived DCs were CD1d+. These CD1d-overexpressing DCs were more efficient in inducing iNKT cell response than those without modification, and their ability was comparable to that of DCs generated from monocytes of healthy donors. The iNKT cells expanded by the CD1d-overexpressing DCs were functional, as demonstrated by their ability to lyse iNKT cell-sensitive glioma cells. Therefore, hESCs stably modified with the CD1d gene may serve as a convenient, unlimited, and competent DC source for iNKT cell-based cancer immunotherapy. PMID:24292792

  12. Localized electroporation effect on adherent cells in modified electric cell-substrate impedance sensing circuits

    NASA Astrophysics Data System (ADS)

    Kim, Yu Jin; Ram Song, Ka; Kim, Hee-Dae; Park, Bum Chul; Kim, Young Keun; Kang, Chi Jung

    2016-10-01

    Electroporation is a physical transfection method for introducing foreign genes or drugs into cells. It does not require toxic reagents or transfection vectors. However, its applications have been limited because of cell damage and nonspecific transport. Here, we present an effective method for selective and localized electroporation using atomic force microscopy. This electroporation method is applied to adherent cells on substrates, instead of conventionally used suspended cells, and offers relatively effective cell transfection. Moreover, this method enables localized transfection into targeted areas at the single-cell level.

  13. In vitro mesenchymal stem cell response to a CO2 laser modified polymeric material.

    PubMed

    Waugh, D G; Hussain, I; Lawrence, J; Smith, G C; Cosgrove, D; Toccaceli, C

    2016-10-01

    With an ageing world population it is becoming significantly apparent that there is a need to produce implants and platforms to manipulate stem cell growth on a pharmaceutical scale. This is needed to meet the socio-economic demands of many countries worldwide. This paper details one of the first ever studies in to the manipulation of stem cell growth on CO2 laser surface treated nylon 6,6 highlighting its potential as an inexpensive platform to manipulate stem cell growth on a pharmaceutical scale. Through CO2 laser surface treatment discrete changes to the surfaces were made. That is, the surface roughness of the nylon 6,6 was increased by up to 4.3μm, the contact angle was modulated by up to 5° and the surface oxygen content increased by up to 1atom %. Following mesenchymal stem cell growth on the laser treated samples, it was identified that CO2 laser surface treatment gave rise to an enhanced response with an increase in viable cell count of up to 60,000cells/ml when compared to the as-received sample. The effect of surface parameters modified by the CO2 laser surface treatment on the mesenchymal stem cell response is also discussed along with potential trends that could be identified to govern the mesenchymal stem cell response.

  14. A Genetically Modified Protein-Based Hydrogel for 3D Culture of AD293 Cells

    PubMed Central

    Du, Xiao; Wang, Jingyu; Diao, Wentao; Wang, Ling; Long, Jiafu; Zhou, Hao

    2014-01-01

    Hydrogels have strong application prospects for drug delivery, tissue engineering and cell therapy because of their excellent biocompatibility and abundant availability as scaffolds for drugs and cells. In this study, we created hybrid hydrogels based on a genetically modified tax interactive protein-1 (TIP1) by introducing two or four cysteine residues in the primary structure of TIP1. The introduced cysteine residues were crosslinked with a four-armed poly (ethylene glycol) having their arm ends capped with maleimide residues (4-armed-PEG-Mal) to form hydrogels. In one form of the genetically modification, we incorporated a peptide sequence ‘GRGDSP’ to introduce bioactivity to the protein, and the resultant hydrogel could provide an excellent environment for a three dimensional cell culture of AD293 cells. The AD293 cells continued to divide and displayed a polyhedron or spindle-shape during the 3-day culture period. Besides, AD293 cells could be easily separated from the cell-gel constructs for future large-scale culture after being cultured for 3 days and treating hydrogel with trypsinase. This work significantly expands the toolbox of recombinant proteins for hydrogel formation, and we believe that our hydrogel will be of considerable interest to those working in cell therapy and controlled drug delivery. PMID:25233088

  15. Allogeneic mesenchymal stem cells, but not culture modified monocytes, improve burn wound healing.

    PubMed

    Clover, Anthony J P; Kumar, Arun H S; Isakson, Matthew; Whelan, Derek; Stocca, Alecia; Gleeson, Birgitta M; Caplice, Noel M

    2015-05-01

    The use of cell therapy to improve burn wound healing is limited as a validated cell source is not rapidly available after injury. Progenitor cells have shown potential to drive the intrinsic wound regeneration. Two sources of cells, allogeneic mesenchymal stem cells (MSC) and autologous culture modified monocytes (CMM), were assessed for their ability to influence burn wound healing. Both could be widely available shortly after injury. Cells were delivered in a fibrin matrix following contact burns in a porcine burns model. Application of MSC significantly decreased the area of unhealed burn compared to CMM or delivery matrix alone (6% MSC, 27% CMM, 24% Matrix, p<0.001). MSC treated wounds showed histological evidence of improved wound healing with increased collagen content (MSC 49%, CMM 42%, p<0.01), increased epidermal area (MSC 8.8%, CMM 6.1%, p<0.01) and dermal thickness (MSC 1108 μm, CMM 1007 μm, p<0.01) compared to CMM treated wounds. Labelled MSC and CMM were identified in the wounds after 2 weeks by immunohistochemistry and FACS. A single application of allogeneic MSC improves the rate of burn wound healing and improves the histological appearance of the burn wound. These cells show potential as a cell therapy that is rapidly available following burn.

  16. Assessment of tolerance to multistresses and in vitro cell adhesion in genetically modified Lactobacillus plantarum 590.

    PubMed

    Liu, Haiyan; Xu, Wentao; Luo, Yunbo; Tian, Hongtao; Wang, Hongxin; Guo, Xing; Yuan, Yanfang; Huang, Kunlun

    2011-03-01

    Lactobacillus plantarum (Lp) is a lactic acid bacterium that has many excellent traits that meet the needs of industrial production. Genetically modified (GM) Lp590 was obtained from Lp that was modified by the insertion of the gene nisI, which can confer resistance to nisin and play a role as a bio-preservative. Here, explorations were made to assess the safety of GM Lp590 and establish an in vitro evaluation model. The ability of Lp590 to tolerate both environmental stresses (such as temperatures ranging from 52 to 4 °C, or exposure to ethanol, oxygen, and osmotic stresses) and gastrointestinal transit was assessed. Lp590 showed a tolerance to 4 °C and ethanol (20%) within a period of 240 min that was similar to Lp. Notably, Lp590 can tolerate higher temperature (52 °C) and higher levels of H(2)O(2) (2%) and NaCl (4.0 M) than Lp. In contrast, Lp590 has the same gastrointestinal transit tolerance as Lp. In addition, Lp590 can adhere to Caco-2 cells, and it has no adverse effect on the cell membrane in vitro. These results indicate that GM Lp590 has many desirable biological characteristics and has good prospects for industrial applications. A useful and comprehensive exploration has been undertaken to establish a new in vitro evaluation model for genetically modified microorganisms (GMMs).

  17. A novel mechanism of action for salidroside to alleviate diabetic albuminuria: effects on albumin transcytosis across glomerular endothelial cells.

    PubMed

    Wu, Dan; Yang, Xiaoyan; Zheng, Tao; Xing, Shasha; Wang, Jianghong; Chi, Jiangyang; Bian, Fang; Li, Wenjing; Xu, Gao; Bai, Xiangli; Wu, Guangjie; Jin, Si

    2016-02-01

    Salidroside (SAL) is a phenylethanoid glycoside isolated from the medicinal plant Rhodiola rosea. R. rosea has been reported to have beneficial effects on diabetic nephropathy (DN) and high-glucose (HG)-induced mesangial cell proliferation. Given the importance of caveolin-1 (Cav-1) in transcytosis of albumin across the endothelial barrier, the present study was designed to elucidate whether SAL could inhibit Cav-1 phosphorylation and reduce the albumin transcytosis across glomerular endothelial cells (GECs) to alleviate diabetic albuminuria as well as to explore its upstream signaling pathway. To assess the therapeutic potential of SAL and the mechanisms involved in DN albuminuria, we orally administered SAL to db/db mice, and the effect of SAL on the albuminuria was measured. The albumin transcytosis across GECs was explored in a newly established in vitro cellular model. The ratio of albumin to creatinine was significantly reduced upon SAL treatment in db/db mice. SAL decreased the albumin transcytosis across GECs in both normoglycemic and hyperglycemic conditions. SAL reversed the HG-induced downregulation of AMP-activated protein kinase and upregulation of Src kinase and blocked the upregulation Cav-1 phosphorylation. Meanwhile, SAL decreased mitochondrial superoxide anion production and moderately depolarized mitochondrial membrane potential. We conclude that SAL exerts its proteinuria-alleviating effects by downregulation of Cav-1 phosphorylation and inhibition of albumin transcytosis across GECs. These studies provide the first evidence of interference with albumin transcytosis across GECs as a novel approach to the treatment of diabetic albuminuria.

  18. Ex Vivo Expansion of Human Mobilized Peripheral Blood Stem Cells Using Epigenetic Modifiers

    PubMed Central

    Saraf, Santosh; Araki, Hiroto; Petro, Benjamin; Park, Youngmin; Taioli, Simona; Yoshinaga, Kazumi G; Koca, Emre; Rondelli, Damiano; Mahmud, Nadim

    2014-01-01

    Background Epigenetic modifications likely control fate of hematopoietic stem cells (HSC). The chromatin modifying agents (CMA), 5-aza-2’-deoxyctidine (5azaD) and trichostatin A (TSA) have previously been shown to expand HSC from cord blood and bone marrow. Here we assessed whether CMA can also expand HSCs present in growth factor mobilized human peripheral blood (MPB). Study Design & Methods 5azaD and TSA were sequentially added to CD34+ MPB cells in the presence of cytokines and the cells were cultured for nine days. Results Following culture, a 3.6 ± 0.5 fold expansion of CD34+CD90+ cells, a 10.1 ± 0.5 fold expansion of primitive colony forming unit (CFU)-mix, and a 2.2 ± 0.5 fold expansion of long-term cobble stone-area forming cells (CAFC) was observed in 5azaD/TSA expanded cells. By contrast, cells cultured in cytokines without 5azaD/TSA displayed no expansion; rather a reduction in CD34+CD90+ cells (0.7 ± 0.1 fold) and CAFCs (0.3 ± 0.1) from their initial numbers was observed. Global hypomethylation corresponding with increased transcript levels of several genes implicated in HSC self-renewal, including HOXB4, GATA2, and EZH2, was observed in 5azaD/TSA expanded MPB cells in contrast to controls. 5azaD/TSA expanded MPB cells retained in vivo hematopoietic engraftment capacity. Conclusion MPB CD34+ cells from donors can be expanded using 5azaD/TSA and these expanded cells retain in vivo hematopoietic reconstitution capacity. This strategy may prove to be potentially useful to augment HSCs numbers for patients who fail to mobilize. PMID:25363624

  19. Tumor Immunotherapy Using Gene-Modified Human Mesenchymal Stem Cells Loaded into Synthetic Extracellular Matrix Scaffolds

    PubMed Central

    Compte, Marta; Cuesta, Ángel M; Sánchez-Martín, David; Alonso-Camino, Vanesa; Vicario, José Luís; Sanz, Laura; Álvarez-Vallina, Luís

    2009-01-01

    Mesenchymal stem cells (MSCs) are appealing as gene therapy cell vehicles given their ease of expansion and transduction. However, MSCs exhibit immunomodulatory and proangiogenic properties that may pose a risk in their use in anticancer therapy. For this reason, we looked for a strategy to confine MSCs to a determined location, compatible with a clinical application. Human MSCs genetically modified to express luciferase (MSCluc), seeded in a synthetic extracellular matrix (sECM) scaffold (sentinel scaffold) and injected subcutaneously in immunodeficient mice, persisted for more than 40 days, as assessed by bioluminescence imaging in vivo. MSCs modified to express a bispecific α-carcinoembryonic antigen (αCEA)/αCD3 diabody (MSCdAb) and seeded in an sECM scaffold (therapeutic scaffolds) supported the release of functional diabody into the bloodstream at detectable levels for at least 6 weeks after implantation. Furthermore, when therapeutic scaffolds were implanted into CEA-positive human colon cancer xenograft-bearing mice and human T lymphocytes were subsequently transferred, circulating αCEA/αCD3 diabody activated T cells and promoted tumor cell lysis. Reduction of tumor growth in MSCdAb-treated mice was statistically significant compared with animals that only received MSCluc. In summary, we report here for the first time that human MSCs genetically engineered to secrete a bispecific diabody, seeded in an sECM scaffold and implanted in a location distant from the primary tumor, induce an effective antitumor response and tumor regression. PMID:19096041

  20. Preparation, cell compatibility and degradability of collagen-modified poly(lactic acid).

    PubMed

    Cui, Miaomiao; Liu, Leili; Guo, Ning; Su, Ruixia; Ma, Feng

    2015-01-05

    Poly(lactic acid) (PLA) was modified using collagen through a grafting method to improve its biocompatibility and degradability. The carboxylic group at the open end of PLA was transferred into the reactive acylchlorided group by a reaction with phosphorus pentachloride. Then, collagen-modified PLA (collagen-PLA) was prepared by the reaction between the reactive acylchlorided group and amino/hydroxyl groups on collagen. Subsequently, the structure of collagen-PLA was confirmed by Fourier transform infrared spectroscopy, fluorescein isothiocyanate-labeled fluorescence spectroscopy, X-ray photoelectron spectroscopy, and DSC analyses. Finally, some properties of collagen-PLA, such as hydrophilicity, cell compatibility and degradability were characterized. Results showed that collagen had been grafted onto the PLA with 5% graft ratio. Water contact angle and water absorption behavior tests indicated that the hydrophilicity of collagen-PLA was significantly higher than that of PLA. The cell compatibility of collagen-PLA with mouse embryonic fibroblasts (3T3) was also significantly better than PLA in terms of cell morphology and cell proliferation, and the degradability of PLA was also improved after introducing collagen. Results suggested that collagen-PLA was a promising candidate for biomedical applications.

  1. Nur77 is differentially modified in PC12 cells upon membrane depolarization and growth factor treatment.

    PubMed Central

    Hazel, T G; Misra, R; Davis, I J; Greenberg, M E; Lau, L F

    1991-01-01

    The rat pheochromocytoma cell line PC12 can be induced by growth factors to undergo proliferation and neuronal differentiation. These cells also have excitable membranes that can be depolarized by neurotransmitters or elevated levels of extracellular KCl. Treatment of PC12 cells with growth factors or membrane-depolarizing agents rapidly activates the expression of specific genes whose products are thought to mediate the subsequent biological responses. One such gene, nur77, is a member of the steroid and thyroid hormone receptor gene superfamily. We have identified the Nur77 protein and shown that it is synthesized rapidly and transiently in PC12 cells following stimulation, has a short half-life of 30 to 40 min, and is located in both the nucleus and the cytoplasm. Nur77 is posttranslationally modified, primarily by phosphorylation on serine residues. Phosphopeptide analysis reveals that Nur77 is modified differently upon membrane depolarization than after treatment with growth factors. We hypothesize that the activity of Nur77 is regulated by both differential gene expression and posttranslational modification and that these modes of regulation contribute to distinct downstream responses specific to membrane depolarization and growth factor treatment. Images PMID:1645447

  2. Carboxybetaine methacrylate-modified nylon surface for circulating tumor cell capture.

    PubMed

    Wang, Huiyu; Yue, Guofeng; Dong, Chaoqun; Wu, Fenglei; Wei, Jia; Yang, Yang; Zou, Zhengyun; Wang, Lifeng; Qian, Xiaoping; Zhang, Tao; Liu, Baorui

    2014-03-26

    Conventional in vitro circulating tumor cell (CTC) detection methods are always limited by blood sample volume because of the requirement of a large amount of blood. The aim of this study was to overcome the limitation by designing and making an in vivo CTC capture device. In this study, we designed and prepared a kind of proper material to serve the purpose of intervention. A method employing 3-aminopropyltriethoxysilane (γ-APS) as the coupling reagent to graft carboxybetaine methacrylate (CBMA) and to immobilize an anti-epithelial cell adhesion molecular (EpCAM) antibody on Nylon was developed. The results of X-ray photoelectron spectroscopy and Fourier transform infrared spectroscopy proved the successful graft of γ-APS and CBMA to Nylon. Furthermore, the predicted improvement in the biocompatibilities of our modified Nylon was confirmed by water contact angle measurement, bovine serum albumin adhesion, platelet adhesion, plasma recalcification time determination, and cytotoxicity tests. The tumor cells adhesion experiment revealed that Nylon with the antibody immobilized on it had an affinity for EpCAM positive tumor cells higher than that of pristine Nylon. Additionally, the capture ability of the CTCs was demonstrated in a nude mouse tumor model using the interventional device made of the modified Nylon wire. The positive results suggest that CBMA-grafted and anti-EpCAM antibody-immobilized Nylon is a promising new material for in vivo CTC capture devices.

  3. Synthesization of SnO2-modified carbon nanotubes and their application in microbial fuel cell

    NASA Astrophysics Data System (ADS)

    Wang, Zi-Bo; Xiong, Shi-Chang; Guan, Yu-Jiang; Zhu, Xue-Qiang

    2016-03-01

    The aim of this work was to study the synthesization of SnO2-modified carbon nanotubes and their application in microbial fuel cell. With the chemical vapor deposition technique, carbon nanotubes growing in situ on a carbon felt are obtained. A SnO2 sol was applied to the carbon felt to prepare a SnO2-modified carbon nanotubes. X-ray diffraction and energy-dispersive X-ray analysis confirmed that SnO2 existed in the prepared samples. Using the prepared samples as anode electrodes, flexible graphite as cathode, and glucose solution as substrate in microbial fuel cell, the effects of the temperature, substrate concentration, and electrodes on removal rates for chemical oxygen demand and the performance of microbial fuel cell have been analyzed. With substrate concentration of 1500 mg L-1, the microbial fuel cell had an optimal output voltage of 563 mV and a removal rate of 78 % for chemical oxygen demand at 311 K. The composite electrodes are stable and reusable.

  4. An evaluation of the robustness of organ-at-risk recommendations made by GEC/ESTRO according to interobserver variability: a single-center experience

    PubMed Central

    Celada-Alvarez, Francisco; Roldán, Susana; Torregrosa, Asunción; Betancourt, Jesus; Bautista-Ballesteros, Juan; Farga, Dolores; Ibañez, Blanca; Tormo, Alejandro; Perez-Calatayud, Jose

    2016-01-01

    Purpose Groupe Européen de Curiethérapie (GEC) and European Society for Radiotherapy & Oncology (ESTRO) has proposed a rectal dose constraint of the most exposed 2-cc volume (D2cc of ≤ 75 Gy EQD2α/β = 3) during external-beam plus high-dose-rate brachytherapy (HDR-BT) in localized prostate cancer patients. This study aimed to evaluate D2cc for rectal contouring via interobserver variability. Material and methods Four blinded observers contoured rectums of 5 patients. Rectal contouring anatomical limits were determined through previous consensus. Dose-volume histogram (DVH) dosimetric parameters (D0.1cc, D1cc, and D2cc) were analyzed according to GEC/ESTRO recommendations and subjected to intra- and interobserver comparisons. Latter comparisons involved coefficients of variation. For each parameter, the mean, standard deviation (SD), and range were evaluated. The effect of interobserver variation on total dose was analyzed by estimating the biologically equivalent rectal dose (EQD2α/β = 3). Results Interobserver coefficients of variation for D0.1cc, D1cc, and D2cc were 5.7%, 4.5%, and 4%, respectively. The highest interobserver rectal delineation variation yielded a rectal dose difference up to 5.8 Gy EQD2. Estimated intraobserver variation for the reported D2cc was 5.5% in the worst-case scenario (non-significant). Conclusions We observed acceptable interobserver variability in EQD2 for D2cc, with strong impacts on clinical threshold levels (D2cc ≤ 75 Gy EQD2) in some cases. This small, single-center analysis will be extended in a multicenter study. PMID:27648090

  5. Bio-active molecules modified surfaces enhanced mesenchymal stem cell adhesion and proliferation.

    PubMed

    Mobasseri, Rezvan; Tian, Lingling; Soleimani, Masoud; Ramakrishna, Seeram; Naderi-Manesh, Hossein

    2017-01-29

    Surface modification of the substrate as a component of in vitro cell culture and tissue engineering, using bio-active molecules including extracellular matrix (ECM) proteins or peptides derived ECM proteins can modulate the surface properties and thereby induce the desired signaling pathways in cells. The aim of this study was to evaluate the behavior of human bone marrow mesenchymal stem cells (hBM-MSCs) on glass substrates modified with fibronectin (Fn), collagen (Coll), RGD peptides (RGD) and designed peptide (R-pept) as bio-active molecules. The glass coverslips were coated with fibronectin, collagen, RGD peptide and R-peptide. Bone marrow mesenchymal stem cells were cultured on different substrates and the adhesion behavior in early incubation times was investigated using scanning electron microscopy (SEM) and confocal microscopy. The MTT assay was performed to evaluate the effect of different bio-active molecules on MSCs proliferation rate during 24 and 72 h. Formation of filopodia and focal adhesion (FA) complexes, two steps of cell adhesion process, were observed in MSCs cultured on bio-active molecules modified coverslips, specifically in Fn coated and R-pept coated groups. SEM image showed well adhesion pattern for MSCs cultured on Fn and R-pept after 2 h incubation, while the shape of cells cultured on Coll and RGD substrates indicated that they might experience stress condition in early hours of culture. Investigation of adhesion behavior, as well as proliferation pattern, suggests R-peptide as a promising bio-active molecule to be used for surface modification of substrate in supporting and inducing cell adhesion and proliferation.

  6. Modified sugar beet pectin induces apoptosis of colon cancer cells via interaction with the neutral sugar side-chains

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pectins extracted from a variety of sources and modified with heat and/or pH have previously been shown to exhibit activity towards several cancer cell lines. However, the structural basis for the anti-cancer activity of modified pectin requires clarification. Sugar beet and citrus pectin extracts h...

  7. Surface Modified Biodegradable Electrospun Membranes as a Carrier for Human Embryonic Stem Cell-Derived Retinal Pigment Epithelial Cells.

    PubMed

    Sorkio, Anni; Porter, Patrick J; Juuti-Uusitalo, Kati; Meenan, Brian J; Skottman, Heli; Burke, George A

    2015-09-01

    Human embryonic stem cell-derived retinal pigment epithelial (hESC-RPE) cells are currently undergoing clinical trials to treat retinal degenerative diseases. Transplantation of hESC-RPE cells in conjuction with a supportive biomaterial carrier holds great potential as a future treatment for retinal degeneration. However, there has been no such biodegradable material that could support the growth and maturation of hESC-RPE cells so far. The primary aim of this work was to create a thin porous poly (L-lactide-co-caprolactone) (PLCL) membrane that could promote attachment, proliferation, and maturation of the hESC-RPE cells in serum-free culture conditions. The PLCL membranes were modified by atmospheric pressure plasma processing and coated with collagen IV to enhance cell growth and maturation. Permeability of the membranes was analyzed with an Ussing chamber system. Analysis with scanning electron microscopy, contact angle measurement, atomic force microscopy, and X-ray photoelectron spectroscopy demonstrated that plasma surface treatment augments the surface properties of the membrane, which enhances the binding and conformation of the protein. Cell proliferation assays, reverse transcription-polymerase chain reaction, indirect immunofluoresence staining, trans-epithelial electrical resistance measurements, and in vitro phagocytosis assay clearly demonstrated that the plasma treated PLCL membranes supported the adherence, proliferation, maturation and functionality of hESC-RPE cells in serum-free culture conditions. Here, we report for the first time, how PLCL membranes can be modified with atmospheric pressure plasma processing to enable the formation of a functional hESC-RPE monolayer on a porous biodegradable substrate, which have a potential as a tissue-engineered construct for regenerative retinal repair applications.

  8. A graphene modified anode to improve the performance of microbial fuel cells

    NASA Astrophysics Data System (ADS)

    Zhang, Yezhen; Mo, Guangquan; Li, Xuwen; Zhang, Weide; Zhang, Jiaqi; Ye, Jianshan; Huang, Xiaodan; Yu, Chengzhong

    Graphene with a Brunauer-Emmett-Teller (BET) specific surface area of 264 m 2 g -1 has been used as anodic catalyst of microbial fuel cells (MFCs) based on Escherichia coli (ATCC 25922). The electrochemical activities of plain stainless steel mesh (SSM), polytetrafluoroethylene (PTFE) modified SSM (PMS) and graphene modified SSM (GMS) have been investigated by cyclic voltammetry (CV), discharge experiment and polarization curve measurement. The GMS shows better electrochemical performance than those of SSM and PMS. The MFC equipped with GMS anode delivers a maximum power density of 2668 mW m -2, which is 18 times larger than that obtained from the MFC with the SSM anode and is 17 times larger than that obtained from the MFC with the PMS anode. Scanning electron microscopy (SEM) results indicate that the increase in power generation could be attributed to the high surface area of anode and an increase in the number of bacteria attached to anode.

  9. [Modified method for whole bone marrow adherent culture of human bone marrow mesenchymal stem cells].

    PubMed

    Wang, Xiao-Qing; Zhong, Zhao-Dong; Chen, Zhi-Chao; Zou, Ping

    2014-04-01

    This study was aimed to investigate a more convenient and efficient method to cultivate the human bone marrow mesenchymal stem cells by means of natural erythrocyte sedimentation principle, based on the whole bone marrow adherent method. The bone marrow was cultured with a six-well plate instead of the flasks.Firstly, the bone marrow specimen was cultivated with the MSC complete medium for 48 h, then the upper RBC-free supernatant layer was drawn and placed into the new wells to isolate MSC. Inverted microscope was used to observe the cell morphology and to record the adherent time of first cell passage, first passaging time. The traditional whole bone marrow adherent method was used as the control. The cell cycle and cell surface markers were detected by flow cytometry,and the differentiative capacity of MSC into osteocyte and adipocyte was identified by alkaline phosphatase kit and oil red O, respectively. Besides, the proliferative curve of P1,P3,P5 of BMSC was depicted by counting method. The results showed that MSC cultured by the modified method highly expressed CD90, CD105, CD13, CD44 and lowly expressed CD14, CD45, CD34. Concerning the cell cycle feature, it was found that most of the cells were in G0/G1 phase (88.76%) , followed by G2/M phase (3.04%) and S phase (8.2%), which was in accordance with stem cell cycle characteristics. The proliferative curve showed a typical "S" type, and both the oil red O and alkaline phosphatase staining of MSC were positive. Compared with the traditional method, the modified method had the advantage of high adherence rate (P = 0.0001) and shorter passaging time for the first passage (P = 0.001), with the statistically significant difference. It is concluded that there is a large number of adherent, active and suspended MSC in the RBC-free supernatant layer after the culture of bone marrow for 48 h. Isolating MSC by the modified method is more convenient and efficient than the traditional whole bone marrow adherent method.

  10. In vitro studies of interaction of modified silica nanoparticles with different types of immunocompetent cells.

    PubMed

    Kulikova, Galina A; Parfenyuk, Elena V; Ryabinina, Irina V; Antsiferova, Yuliya S; Sotnikova, Nataliya Yu; Posiseeva, Lubov V; Eliseeva, Mariya A

    2010-11-01

    Interactions between different types of immune cells and organically-modified silica nanoparticles were studied. The silica particles functionalized with amine groups were prepared by sol-gel technique. Sheep immunoglobulin labeled with fluoresceine isothiocyanate was immobilized by adsorption onto the nanoparticles. The presence of the functional groups was confirmed by infrared absorption measurements. The level of immunocompetent cells interacting with the silica nanoparticles was estimated as the amount of fluorescence-bright cells by flow cytometry method. A low level of interaction of the peripheral blood lymphocytes with the silica nanoparticles was found. On the contrary, the macrophages are actively involved in interaction with the silica nanoparticles. The influence of different size of the silica nanoparticles and incubation time on viability and functional activity of peripheral blood lymphocytes and peritoneal macrophages were investigated.

  11. FACTORS WHICH MODIFY THE EFFECT OF SODIUM AND POTASSIUM ON BACTERIAL CELL MEMBRANES.

    PubMed

    HENNEMAN, D H; UMBREIT, W W

    1964-06-01

    Henneman, Dorothy H. (Rutgers, The State University, New Brunswick, N.J.), and W. W. Umbreit. Factors which modify the effect of sodium and potassium on bacterial cell membranes. J. Bacteriol. 87:1266-1273. 1964.-Suspensions of Escherichia coli B, when placed in 0.2 to 0.5 m solutions of NaCl, KCl, or LiCl, show an increased turbidity. With NaCl, this increased turbidity is stable with time; with KCl and LiCl, it is gradually lost. The stability to NaCl with time is due to substances removable from the cell by incubation in phosphate buffer; these materials exist in water washings from such phosphate-incubated cells.

  12. Improved morphology control using a modified two-step method for efficient perovskite solar cells.

    PubMed

    Bi, Dongqin; El-Zohry, Ahmed M; Hagfeldt, Anders; Boschloo, Gerrit

    2014-11-12

    A two-step wet chemical synthesis method for methylammonium lead(II) triiodide (CH3NH3PbI3) perovskite is further developed for the preparation of highly reproducible solar cells, with the following structure: fluorine-doped tin oxide (FTO)/TiO2 (compact)/TiO2 (mesoporous)/CH3NH3PbI3/spiro-OMeTAD/Ag. The morphology of the perovskite layer could be controlled by careful variation of the processing conditions. Specifically, by modifying the drying process and inclusion of a dichloromethane treatment, more uniform films could be prepared, with longer emission lifetime in the perovskite material and longer electron lifetime in solar cell devices, as well as faster electron transport and enhanced charge collection at the selective contacts. Solar cell efficiencies up to 13.5% were obtained.

  13. Enhancing the Detection of Dysmorphic Red Blood Cells and Renal Tubular Epithelial Cells with a Modified Urinalysis Protocol

    PubMed Central

    Chu-Su, Yu; Shukuya, Kenichi; Yokoyama, Takashi; Lin, Wei-Chou; Chiang, Chih-Kang; Lin, Chii-Wann

    2017-01-01

    Urinary sediment is used to evaluate patients with possible urinary tract diseases. Currently, numerous protocols are applied to detect dysmorphic red blood cells (RBCs) and renal tubular epithelial cells (RTECs) in urinary sediment. However, distinct protocols are used by nephrologists and medical technologists for specimen concentration and observation, which leads to major discrepancies in the differential counts of formed elements such as dysmorphic RBCs and RTECs and might interfere with an accurate clinical diagnosis. To resolve these problems, we first tested a modified urinalysis protocol with an increased relative centrifuge force and concentration factor in 20 biopsy-confirmed glomerulonephritis patients with haematuria. We successfully improved the recovery ratio of dysmorphic RBCs in clinical specimens from 34.7% to 42.0% (P < 0.001). Furthermore, we confirmed the correlation between counts by the modified urinary protocol and Sysmex UF-1000i urinary flow cytometer (r ≥ 0.898, P < 0.001). A total of 28 types of isomorphic and dysmorphic RBCs were detected using a bright field microscope, with results comparable to those using a standard phase contrast microscope. Finally, we applied Sternheimer stain to enhance the contrast of RTECs in the urinary sediments. We concluded that this modified urinalysis protocol significantly enhanced the quality of urinalysis. PMID:28074941

  14. Fucoidan reduced the invasion of oral squamous cell carcinoma cells and modified their effects to macrophages.

    PubMed

    Lin, Junda; Wang, Ketao; Wang, Huayang; Shao, Qianqian; Luan, Yijun; Xu, Yan; Song, Xiaobin; Tan, Wanye; Liu, Shaohua; Wei, Fengcai; Qu, Xun

    2017-01-01

    Fucoidan is a complex of polysaccharides showing antitumor and immunomodulation properties. Our previous studies found its regulation to myeloid immune cells, including macrophages. Aberrant infiltration and functions of macrophages are commonly found in oral squamous cell carcinoma (OSCC). In this study, we analyzed the effects of fucoidan on invasion of OSCC cells, and their regulation to macrophages, trying to evaluate its role as a potential therapy for OSCC. CAL27 and THP-1-derived macrophages were used as models for OSCC cells and tumor-infiltrated macrophages in the in vitro study, respectively. The effects of fucoidan on invasion of OSCC cells and their recruitment to macrophages were analyzed by transwell assay. KIF4A siRNA transfection was performed to investigate its role in fucoidan-modulated OSCC cells invasion. CCL3-neutralizing antibody was added into the conditioned medium of OSCC cells to evaluate its role in fucoidan-mediated macrophages recruitment and re-education. Fucoidan reduced the invasive potential of CAL27 cells with a decrease of MMP-2 and KIF4A transcription. KIF4A knockdown in CAL27 cells led to decreased invasion and MMP-2 expression. The conditioned medium of fucoidan-treated CAL27 cells promoted recruitment and inflammatory cytokines secretion on THP-1-derived macrophages. Further analysis found that fucoidan increased CCL3 production in CAL27 cells. Blocking CCL3 expression reversed the effects of fucoidan on macrophage recruitment and re-education. Our study found that fucoidan regulated the invasion of OSCC cells and also their recruiting and re-educating effects on macrophages, suggesting it could be a complementary approach in the treatment of OSCC.

  15. Time course of bronchial cell inflammation following exposure to diesel particulate matter using a modified EAVES.

    PubMed

    Hawley, Brie; McKenna, Dave; Marchese, Anthony; Volckens, John

    2014-08-01

    Electrostatic deposition of particles onto the surface of well-differentiated airway cells is a rapid and efficient means to screen for toxicity associated with exposure to fine and ultrafine particulate air pollution. This work describes the development and application of an electrostatic aerosol in vitro exposure system (EAVES) with increased throughput and ease-of-use. The modified EAVES accommodates standard tissue culture plates and uses an alternating electric field to deposit a net neutral charge of aerosol onto air-interface cell cultures. Using this higher-throughput design, we were able to examine the time-course (1, 3, 6, 9, and 24 h post-exposure) of transcript production and cytotoxicity in well-differentiated human bronchial cells exposed to diesel particulate matter at levels of 'real-world' significance. Statistically significant responses were observed at exposure levels (∼0.4 μg/cm(2)) much lower than typically reported in vitro using traditional submerged/resuspended techniques. Levels of HO-1, IL-8, CYP1A1, COX-2, and HSP-70 transcripts increased immediately following diesel particulate exposure and persisted for several hours; cytotoxicity was increased at 24h. The modified EAVES provides a platform for higher throughput, more efficient and representative testing of aerosol toxicity in vitro.

  16. Genetically modified mesenchymal stem cells (MSCs) promote axonal regeneration and prevent hypersensitivity after spinal cord injury.

    PubMed

    Kumagai, Gentaro; Tsoulfas, Pantelis; Toh, Satoshi; McNiece, Ian; Bramlett, Helen M; Dietrich, W Dalton

    2013-10-01

    Neurotrophins and the transplantation of bone marrow-derived stromal cells (MSCs) are both candidate therapies targeting spinal cord injury (SCI). While some studies have suggested the ability of MSCs to transdifferentiate into neural cells, other SCI studies have proposed anti-inflammatory and other mechanisms underlying established beneficial effects. We grafted rat MSCs genetically modified to express MNTS1, a multineurotrophin that binds TrkA, TrkB and TrkC, and p75(NTR) receptors or MSC-MNTS1/p75(-) that binds mainly to the Trk receptors. Seven days after contusive SCI, PBS-only, GFP-MSC, MSC-MNTS1/GFP or MSC-MNTS1/p75(-)/GFP were delivered into the injury epicenter. All transplanted groups showed reduced inflammation and cystic cavity size compared to control SCI rats. Interestingly, transplantation of the MSC-MNTS1 and MSC-MNTS1/p75(-), but not the naïve MSCs, enhanced axonal growth and significantly prevented cutaneous hypersensitivity after SCI. Moreover, transplantation of MSC-MNTS1/p75(-) promoted angiogenesis and modified glial scar formation. These findings suggest that MSCs transduced with a multineurotrophin are effective in promoting cell growth and improving sensory function after SCI. These novel data also provide insight into the neurotrophin-receptor dependent mechanisms through which cellular transplantation leads to functional improvement after experimental SCI.

  17. Thiols as interfacial modifiers to enhance the performance and stability of perovskite solar cells.

    PubMed

    Cao, Jing; Yin, Jun; Yuan, Shangfu; Zhao, Yun; Li, Jing; Zheng, Nanfeng

    2015-06-07

    Modifying the interfaces of CH3NH3PbI3 with TiO2 and hole transport layers using two different types of thiols leads to enhanced performance and stability of perovskite solar cells. The incorporation of HOOC-Ph-SH at the TiO2/perovskite interface facilitates electron transfer from perovskite to TiO2 and also alters the morphology of perovskite crystal growth to increase the power conversion efficiency. The modification of pentafluorobenzenethiol at the perovskite/hole transport layer interface improves the stability.

  18. [Application of Chimeric Antigen Receptor-Modified CAR-T/NK Cells to Treatment of Multiple Myeloma].

    PubMed

    Wang, Lei; Ou, jian-Feng; Bai, Hai

    2015-04-01

    Chimeric antigen receptor(CAR) is a synthesized transmembrane protein, which redirects the modified cells through specific or associated antigen on tumor cells. CAR-modified T/NK cells, especially CAR-T cells, are a new tool of rapidly developing of adoptive immunotherapy of tumor in recent years, they give T/NK cells the targeting cytotoxic activity and can overcome the tumor immunosuppressive microenvironment and break the state of the host immune tolerance. CAR combines the single-chain antibody to tumor-associated antigen with T/NK cells' activated motifs, giving T/NK cells' tumor targeting activity, so enhancing their cytotoxic activity and lasting the vitality by gene transduction. In this article the CAR development, comparison of CAR-T and CAR-NK cells, surface markers on MM cells and use of CAR in MM, and CAR perspectives are summarized.

  19. Pathway of cytotoxicity induced by folic acid modified selenium nanoparticles in MCF-7 cells.

    PubMed

    Pi, Jiang; Jin, Hua; Liu, Ruiying; Song, Bing; Wu, Qing; Liu, Li; Jiang, Jinhuan; Yang, Fen; Cai, Huaihong; Cai, Jiye

    2013-02-01

    Selenium nanoparticles (Se NPs) have been recognized as promising materials for biomedical applications. To prepare Se NPs which contained cancer targeting methods and to clarify the cellular localization and cytotoxicity mechanisms of these Se NPs against cancer cells, folic acid protected/modified selenium nanoparticles (FA-Se NPs) were first prepared by a one-step method. Some morphologic and spectroscopic methods were obtained to prove the successfully formation of FA-Se NPs while free folate competitive inhibition assay, microscope, and several biological methods were used to determine the in vitro uptake, subcellular localization, and cytotoxicity mechanism of FA-Se NPs in MCF-7 cells. The results indicated that the 70-nm FA-Se NPs were internalized by MCF-7 cells through folate receptor-mediated endocytosis and targeted to mitochondria located regions through endocytic vesicles transporting. Then, the FA-Se NPs entered into mitochondria; triggered the mitochondria-dependent apoptosis of MCF-7 cells which involved oxidative stress, Ca(2)+ stress changes, and mitochondrial dysfunction; and finally caused the damage of mitochondria. FA-Se NPs released from broken mitochondria were transported into nucleus and further into nucleolus which then induced MCF-7 cell cycle arrest. In addition, FA-Se NPs could induce cytoskeleton disorganization and induce MCF-7 cell membrane morphology alterations. These results collectively suggested that FA-Se NPs could be served as potential therapeutic agents and organelle-targeted drug carriers in cancer therapy.

  20. Enhanced adherence of mouse fibroblast and vascular cells to plasma modified polyethylene.

    PubMed

    Reznickova, Alena; Novotna, Zdenka; Kolska, Zdenka; Kasalkova, Nikola Slepickova; Rimpelova, Silvie; Svorcik, Vaclav

    2015-01-01

    Since the last decade, tissue engineering has shown a sensational promise in providing more viable alternatives to surgical procedures for harvested tissues, implants and prostheses. Biomedical polymers, such as low-density polyethylene (LDPE), high-density polyethylene (HDPE) and ultra-high molecular weight polyethylene (UHMWPE), were activated by Ar plasma discharge. Degradation of polymer chains was examined by determination of the thickness of ablated layer. The amount of an ablated polymer layer was measured by gravimetry. Contact angle, measured by goniometry, was studied as a function of plasma exposure and post-exposure aging times. Chemical structure of modified polymers was characterized by angle resolved X-ray photoelectron spectroscopy. Surface chemistry and polarity of the samples were investigated by electrokinetic analysis. Changes in surface morphology were followed using atomic force microscopy. Cytocompatibility of plasma activated polyethylene foils was studied using two distinct model cell lines; VSMCs (vascular smooth muscle cells) as a model for vascular graft testing and connective tissue cells L929 (mouse fibroblasts) approved for standardized material cytotoxicity testing. Specifically, the cell number, morphology, and metabolic activity of the adhered and proliferated cells on the polyethylene matrices were studied in vitro. It was found that the plasma treatment caused ablation of the polymers, resulting in dramatic changes in their surface morphology and roughness. ARXPS and electrokinetic measurements revealed oxidation of the polymer surface. It was found that plasma activation has a positive effect on the adhesion and proliferation of VSMCs and L929 cells.

  1. The Chromatin Modifier MSK1/2 Suppresses Endocrine Cell Fates during Mouse Pancreatic Development

    PubMed Central

    Bhat, Neha; Park, Jeehye; Zoghbi, Huda Y.; Arthur, J. Simon C.; Zaret, Kenneth S.

    2016-01-01

    Type I diabetes is caused by loss of insulin-secreting beta cells. To identify novel, pharmacologically-targetable histone-modifying proteins that enhance beta cell production from pancreatic progenitors, we performed a screen for histone modifications induced by signal transduction pathways at key pancreatic genes. The screen led us to investigate the temporal dynamics of ser-28 phosphorylated histone H3 (H3S28ph) and its upstream kinases, MSK1 and MSK2 (MSK1/2). H3S28ph and MSK1/2 were enriched at the key endocrine and acinar promoters in E12.5 multipotent pancreatic progenitors. Pharmacological inhibition of MSK1/2 in embryonic pancreatic explants promoted the specification of endocrine fates, including the beta-cell lineage, while depleting acinar fates. Germline knockout of both Msk isoforms caused enhancement of alpha cells and a reduction in acinar differentiation, while monoallelic loss of Msk1 promoted beta cell mass. Our screen of chromatin state dynamics can be applied to other developmental contexts to reveal new pathways and approaches to modulate cell fates. PMID:27973548

  2. Early postweaning social isolation but not environmental enrichment modifies vermal Purkinje cell dendritic outgrowth in rats.

    PubMed

    Pascual, Rodrigo; Bustamante, Carlos

    2013-01-01

    In the present study, we analyzed the effects of enriched, social and isolated experiences on vermal Purkinje cell of the rat, together with anxiety-like behavior in the elevated-plus maze. Sprague-Dawley male rats were randomly submitted to either enriched, social, or isolated environments during the early postweaning period (postnatal days 22-32) and were then behaviorally evaluated in the elevated-plus maze and euthanized for histological analysis. Vermal Purkinje cells (sub-lobules VIa and VIb) were sampled, drawn under camera lucida and morphometrically assessed using the Sholl's concentric ring method. Data obtained indicate that environmental enrichment did not significantly modify the Purkinje cell dendritic branching. On the contrary, Purkinje cell of animals reared in social isolation exhibited a significant reduction in dendritic arborization, which was closely associated with anxiety-like behaviors. The data obtained indicate that, although environmental stimulation in normal animals does not produce significant changes in vermal Purkinje cell dendritic arborization, these cells are vulnerable to early stressful experiences, which is in close association with anxiety-like behaviors.

  3. Minimally modified low density lipoprotein induces monocyte chemotactic protein 1 in human endothelial cells and smooth muscle cells

    SciTech Connect

    Cushing, S.D.; Berliner, J.A.; Valente, A.J.; Territo, M.C.; Navab, M.; Parhami, F.; Gerrity, R.; Schwartz, C.J.; Fogelman, A.M.

    1990-07-01

    After exposure to low density lipoprotein (LDL) that had been minimally modified by oxidation (MM-LDL), human endothelial cells (EC) and smooth muscle cells (SMC) cultured separately or together produced 2- to 3-fold more monocyte chemotactic activity than did control cells or cells exposed to freshly isolated LDL. This increase in monocyte chemotactic activity was paralleled by increases in mRNA levels for a monocyte chemotactic protein 1 (MCP-1) that is constitutively produced by the human glioma U-105MG cell line. Antibody that had been prepared against cultured baboon smooth muscle cell chemotactic factor (anti-SMCF) did not inhibit monocyte migration induced by the potent bacterial chemotactic factor f-Met-Leu-Phe. However, anti-SMCF completely inhibited the monocyte chemotactic activity found in the media of U-105MG cells, EC, and SMC before and after exposure to MM-LDL. Moreover, monocyte migration into the subendothelial space of a coculture of EC and SMC that had been exposed to MM-LDL was completely inhibited by anti-SMCF. Anti-SMCF specifically immunoprecipitated 10-kDa and 12.5-kDa proteins from EC. Incorporation of (35S)methionine into the immunoprecipitated proteins paralleled the monocyte chemotactic activity found in the medium of MM-LDL stimulated EC and the levels of MCP-1 mRNA found in the EC. We conclude that SMCF is in fact MCP-1 and MCP-1 is induced by MM-LDL.

  4. Multi-scale cell/surface interaction on modified titanium aluminum vanadium surfaces

    NASA Astrophysics Data System (ADS)

    Chen, Jianbo

    This dissertation presents a series of experimental studies of the effects of multi-scale cell/surface interactions on modified Ti-6Al-4V surfaces. These include laser-grooved surfaces; porous structures and RGD-coated laser-grooved surfaces. A nano-second DPSS UV lasers with a Gaussian pulse energy profile was used to introduce the desired micro-groove geometries onto Ti-6Al-4V surfaces. This was done without inducing micro-cracks or significant changes in surface chemistry within the heat affected zones. The desired 8-12 mum groove depths and widths were achieved by the control of pulse frequency, scan speed, and the lens focal length that controls spot size. The interactions between human osteosarcoma (HOS) cells and laser-grooved Ti-6Al-4V surfaces were investigated after 48 hours of cell culture. The cell behavior, including cell spreading, alignment and adhesion, was elucidated using scanning electronic microscopy (SEM), immuno-fluorescence staining and enzymatic detachment. Contact guidance was shown to increase as grooved spacing decreased. For the range of micro-groove geometries studied, micro-grooves with groove spacings of 20 mum provided the best combination of cell orientation and adhesion. Short-term adhesion experiments (15 mins to 1 day) also revealed that there is a positive correlation between cell orientation and cell adhesion. Contact guidance on the micro-grooved surfaces is shown to be enhanced by nano- and micro-scale asperities that provide sites for the attachment of lamellopodia during cell locomotion and spreading. Contact guidance is also promoted by the geometrical confinement provided by laser grooves. An experimental study of initial cell spreading and ingrowth into Ti-6Al-4V porous structures was also carried out on porous structures with different pore sizes and geometries. A combination of SEM, the tetrazolium salt (MTT) colorimetric assay and enzymatic detachment were used to study cell spreading and adhesion. The extent of cell

  5. Epithelial cells from smokers modify dendritic cell responses in the context of influenza infection

    EPA Science Inventory

    Epidemiologic evidence suggests that cigarette smoking is a risk factor for infection with influenza, but the mechanisms underlying this susceptibility remain unknown. To ascertain if airway epithelial cells from smokers demonstrate a decreased ability to orchestrate an influenza...

  6. Mediatorless sugar/oxygen enzymatic fuel cells based on gold nanoparticle-modified electrodes.

    PubMed

    Wang, Xiaoju; Falk, Magnus; Ortiz, Roberto; Matsumura, Hirotoshi; Bobacka, Johan; Ludwig, Roland; Bergelin, Mikael; Gorton, Lo; Shleev, Sergey

    2012-01-15

    We report on the fabrication and characterisation of a gold-nanoparticle (AuNP)-based mediatorless sugar/oxygen biofuel cell (BFC) operating in neutral sugar-containing buffers and human physiological fluids, such as blood and plasma. First, Corynascus thermophilus cellobiose dehydrogenase (CtCDH) and Myrothecium verrucaria bilirubin oxidase (MvBOx), used as anodic and cathodic bioelements, respectively, were immobilised on gold electrodes modified with 20 nm AuNPs. Detailed characterisation and optimisation of a new CDH/AuNP-based bioanode were performed and the following fundamental parameters were obtained: (i) the redox potential of the haem-containing centre of the enzyme was measured to be 75 mV vs. NHE, (ii) the surface coverage of CtCDH was found to be 0.65 pmol cm(-2) corresponding to a sub-monolayer coverage of the thiol-modified AuNPs by the enzyme, (iii) a turnover number for CtCDH immobilised on thiol-modified AuNPs was calculated to be ca. 0.5 s(-1), and (iv) the maximal current densities as high as 40 μA cm(-2) were registered in sugar-containing neutral buffers. Second, both biomodified electrodes, namely the CtCDH/AuNP-based bioanode and the MvBOx/AuNP-based biocathode, were combined into a functional BFC and the designed biodevices were carefully investigated. The following characteristics of the mediator-, separator- and membrane-less, miniature BFC were obtained: in phosphate buffer; an open-circuit voltage of 0.68 V, a maximum power density of 15 μW cm(-2) at a cell voltage of 0.52 V and in human blood; an open-circuit voltage of 0.65 V, a maximum power density of 3 μW cm(-2) at a cell voltage of 0.45 V, respectively. The estimated half-lives of the biodevices were found to be >12, <8, and <2 h in a sugar-containing buffer, human plasma, and blood, respectively. The basic characteristics of mediatorless sugar/oxygen BFCs were significantly improved compared with previously designed biodevices, because of the usage of three-dimensional AuNP-modified

  7. Epithelial cell adhesion molecule independent capture of non-small lung carcinoma cells with peptide modified microfluidic chip.

    PubMed

    Pu, Kefeng; Li, Chunlin; Zhang, Nengpan; Wang, Hui; Shen, Wenjiang; Zhu, Yimin

    2017-03-15

    Circulating tumor cells (CTCs) present in the blood of patients with non-hematological cancers are accessible sources for diagnosis and monitoring of cancers. By the aid of the ability of the anti-EpCAM antibody to recognize the epithelial cells, microsystem-based technologies provide robust means for effectively detecting CTCs in vitro. Considering the EpCAM expression is down-regulated during epithelial-mesenchymal transition (EMT) process, the amount of CTCs detected based on anti-EpCAM antibody is underestimated. In our study, the A549 cells targeting peptide (A-1 peptide), as the substitute of anti-EpCAM antibody, was introduced to microfluidic chip to capture A549 cells. Our results showed that both epithelial-like and mesenchymal-like A549 cells could efficiently be captured by the A-1 peptide modified microfluidic chip, and the capture efficiency for epithelial-like cells is comparable to that captured by the EpCAM antibody. Thus, we concluded that the peptide could be a better supplement to the EpCAM antibody for capturing CTCs in microfluidic system with broader spectrum.

  8. Chemically modified guide RNAs enhance CRISPR-Cas genome editing in human primary cells

    PubMed Central

    Clark, Joseph T; Kennedy, Andrew B; Ryan, Daniel E; Roy, Subhadeep; Steinfeld, Israel; Lunstad, Benjamin D; Kaiser, Robert J; Wilkens, Alec B; Bacchetta, Rosa; Tsalenko, Anya; Dellinger, Douglas; Bruhn, Laurakay; Porteus, Matthew H

    2016-01-01

    CRISPR-Cas-mediated genome editing relies on guide RNAs that direct site-specific DNA cleavage facilitated by the Cas endonuclease. Here we report that chemical alterations to synthesized single guide RNAs (sgRNAs) enhance genome editing efficiency in human primary T cells and CD34+ hematopoietic stem and progenitor cells. Co-delivering chemically modified sgRNAs with Cas9 mRNA or protein is an efficient RNA- or ribonucleoprotein (RNP)-based delivery method for the CRISPR-Cas system, without the toxicity associated with DNA delivery. This approach is a simple and effective way to streamline the development of genome editing with the potential to accelerate a wide array of biotechnological and therapeutic applications of the CRISPR-Cas technology. PMID:26121415

  9. miRNAs modified by dietary lipids in Caco-2 cells. A microarray screening

    PubMed Central

    Daimiel, Lidia; Ordovás, Jose Mª.; Dávalos, Alberto

    2015-01-01

    We performed a screening of miRNAs regulated by dietary lipids in a cellular model of enterocytes, Caco-2 cells. Our aim was to describe new lipid-modified miRNAs with an implication in lipid homeostasis and cardiovascular disease [1], [2]. For that purpose, we treated differentiated Caco-2 cells with micelles containing the assayed lipids (cholesterol, conjugated linoleic acid and docosahexaenoic acid) and the screening of miRNAs was carried out by microarray using the μParaflo®Microfluidic Biochip Technology of LC Sciences (Huston, TX, USA). Experimental design, microarray description and raw data have been made available in the GEO database with the reference number of GSE59153. Here we described in detail the experimental design and methods used to obtain the relative expression data. PMID:26484250

  10. Labeling of human hepatocellular carcinoma cells by hexamethylene diamine modified fluorescent carbon dots

    NASA Astrophysics Data System (ADS)

    Dong, Wei; Dong, Yan; Wang, Ying; Zhou, Shiqi; Ge, Xin; Sui, Lili; Wang, Jingwen

    2013-12-01

    Fluorescent carbon dots (CDs) were synthesized by a solvothermal method with glucose as carbon source and surface-modified with 1,6-hexamethylene diamine. In this hybrid CDs, the modification played important role for improving the fluorescent performance by introducing nitrogenous compound to passivate CD's surface, making the CDs emit strong fluorescence. The as-prepared CDs were linked with mouse anti-human Alpha fetoprotein (AFP) antibody and goat anti-mouse immunoglobulin (IgG) to directly and indirectly label fixed human hepatocellular carcinoma cells, respectively. The cytotoxicity of these CDs were also tested using the human hepatocellular carcinoma cells. No apparent cytotoxicity was observed, which suggested the potential application of the as-prepared CDs in bioimaging.

  11. Antigen-receptor gene-modified T cells for treatment of glioma.

    PubMed

    Ikeda, Hiroaki; Shiku, Hiroshi

    2012-01-01

    Immunological effector cells and molecules have been shown to access intracranial tumor sites despite the existence of blood brain barrier (BBB) or immunosuppressive mechanisms associated with brain tumors. Recent progress in T-cell biology and tumor immunology made possible to develop strategies of tumor-associated antigen-specific immunotherapeutic approaches such as vaccination with defined antigens and adoptive T-cell therapy with antigen-specific T cells including gene-modified T cells for the treatment of patients with brain tumors. An array of recent reports on the trials of active and passive immunotherapy for patients with brain tumors have documented safety and some preliminary clinical efficacy, although the ultimate judgment for clinical benefits awaits rigorous evaluation in trials of later phases. Nevertheless, treatment with lymphocytes that are engineered to express tumor-specific receptor genes is a promising immunotherapy against glioma, based on the significant efficacy reported in the trials for patients with other types of malignancy. Overcoming the relative difficulty to apply immunotherapeutic approach to intracranial region, current advances in the understanding of human tumor immunology and the gene-therapy methodology will address the development of effective immunotherapy of brain tumors.

  12. Genetically modified endothelial progenitor cells in the therapy of cardiovascular disease and pulmonary hypertension.

    PubMed

    Lavoie, Jessie R; Stewart, Duncan J

    2012-05-01

    Since their initial discovery, endothelial progenitor cells (EPCs) have held tremendous promise for cell therapy for a variety of cardiovascular diseases including pulmonary hypertension. The clinical experience to date suggests that circulating or bone marrow mononuclear cells and EPCs can induce neovascularization, and enhance cardiac repair after myocardial function, as well as improvements in the hemodynamic and functional status of patients with idiopathic pulmonary arterial hypertension. Although these results are promising, the overall magnitude of the clinical benefits seen in these trials appear to be rather modest. Indeed, strong experimental evidence points towards a reduction in mobilization and impairment in function of EPCs in preclinical models and patients with cardiac disease or with cardiovascular risk factors such as advanced age, type I and II diabetes, hypercholesterolemia, coronary artery disease, as well as other conditions such as pulmonary hypertension. Genetic engineering of EPCs ex vivo, prior to transplantation, is a promising cell-enhancement strategy for restoring the angiogenic potential of autologous, patient-derived cells. This review provides an update of the experimental studies that have used gene-modified EPC therapy to treat ischemic cardiovascular disease and pulmonary hypertension.

  13. The effect of CD47 modified polymer surfaces on inflammatory cell attachment and activation

    PubMed Central

    Stachelek, Stanley J.; Finley, Matthew J.; Alferiev, Ivan S.; Wang, Fengxiang; Tsai, Richard; Eckells, Edward C.; Tomczyk, Nancy; Connolly, Jeanne M.; Discher, Dennis E.; Eckmann, David M.; Levy, Robert J.

    2011-01-01

    CD47 is a transmembrane protein that is a marker of “self”. CD47 binding to its cognate receptor in leukocytes and macrophages, signal regulatory protein alpha (SIRPα), causes inhibition of inflammatory cell attachment. We hypothesized that immobilization of recombinant CD47 on polymeric surfaces would reduce inflammation. Recombinant CD47 was appended to polyvinyl chloride (PVC) or polyurethane (PU) surfaces via photoactivation chemistry. Cell culture studies showed that CD47 immobilization significantly reduced human neutrophil (HL-60) and human monocyte derived macrophage (MDM) (THP-1) attachment to PVC and PU respectively. A neutralizing antibody, directed against SIRPα, inhibited THP-1 and HL-60 binding to PU and PVC surfaces respectively. This antibody also increased the level of SIRPα tyrosine phosphorylation, thereby indicating a direct role for SIRPα mediated signaling in preventing inflammatory cell attachment. Studies using human blood in an ex vivo flow-loop showed that CD47 modified PVC tubing significantly reduced cell binding and neutrophil activation compared to unmodified tubing or poly-2-methoxy-ethylacrylate (PMEA) coated tubing. In ten-week rat subdermal implants, CD47 functionalized PU films showed a significant reduction in markers of MDM mediated oxidative degradation compared to unmodified PU. In conclusion, CD47 functionalized surfaces can resist inflammatory cell interactions both in vitro and in vivo. PMID:21429575

  14. Hyaluronic acid modified mesoporous silica nanoparticles for targeted drug delivery to CD44-overexpressing cancer cells

    NASA Astrophysics Data System (ADS)

    Yu, Meihua; Jambhrunkar, Siddharth; Thorn, Peter; Chen, Jiezhong; Gu, Wenyi; Yu, Chengzhong

    2012-12-01

    In this paper, a targeted drug delivery system has been developed based on hyaluronic acid (HA) modified mesoporous silica nanoparticles (MSNs). HA-MSNs possess a specific affinity to CD44 over-expressed on the surface of a specific cancer cell line, HCT-116 (human colon cancer cells). The cellular uptake performance of fluorescently labelled MSNs with and without HA modification has been evaluated by confocal microscopy and fluorescence-activated cell sorter (FACS) analysis. Compared to bare MSNs, HA-MSNs exhibit a higher cellular uptake via HA receptor mediated endocytosis. An anticancer drug, doxorubicin hydrochloride (Dox), has been loaded into MSNs and HA-MSNs as drug delivery vehicles. Dox loaded HA-MSNs show greater cytotoxicity to HCT-116 cells than free Dox and Dox-MSNs due to the enhanced cell internalization behavior of HA-MSNs. It is expected that HA-MSNs have a great potential in targeted delivery of anticancer drugs to CD44 over-expressing tumors.

  15. Inhibitory impacts of chemically modified tetracycline-3 and underlying mechanism in human cervical cancer cells.

    PubMed

    Zhao, Lin; Xu, Jiaying; Yang, Yang; Chong, Yu; Liu, Chang; Jiao, Yang; Fan, Saijun

    2013-09-01

    Chemically modified tetracyclines (CMTs) have been rationally designed from tetracyclines. The CMTs that show the antimicrobial properties are eliminated, whereas matrix metalloproteinase inhibitory properties are retained. Interestingly, CMT-3 (COL-3, by eliminating the dimethylamino, methyl, and hydroxyl functionalities on the basic tetracycline structure), one of the CMTs, has shown strong anticancer activity. In this study, we found that CMT-3 showed dose-dependent and time-dependent cytotoxicity in HeLa and Siha cells, two human cervical cancer cell lines. HeLa cells were more sensitive to CMT-3 compared with Siha cells. The antiproliferation potential of CMT-3 was associated with the mitochondrial apoptosis pathway, increasing reactive oxygen species level, and proapoptosis protein (e.g. caspase-3) expression, but decreasing antiapoptosis protein expression (e.g. Bcl-2). N-acetylcysteine (a reactive oxygen species inhibitor) and Z-LEHD-FMK significantly reduced or blocked the apoptosis event resulting from cytotoxic effect of CMT-3. CMT-3 also induced G0/G1 phase arrest with the reduction of cell cycle regulatory protein cyclin E and the translocation of NF-κB from the cytoplasm to the nucleus. Our findings provide the important foundation for further investigation of the underlying mechanism for the anticancer activity of CMT-3 and the potential application of CMT-3 as a new therapeutic candidate for clinical cervical cancer therapy.

  16. Expression of a fungal ferulic acid esterase in alfalfa modifies cell wall digestibility

    PubMed Central

    2014-01-01

    Background Alfalfa (Medicago sativa) is an important forage crop in North America owing to its high biomass production, perennial nature and ability to fix nitrogen. Feruloyl esterase (EC 3.1.1.73) hydrolyzes ester linkages in plant cell walls and has the potential to further improve alfalfa as biomass for biofuel production. Results In this study, faeB [GenBank:AJ309807] was synthesized at GenScript and sub-cloned into a novel pEACH vector containing different signaling peptides to target type B ferulic acid esterase (FAEB) proteins to the apoplast, chloroplast, endoplasmic reticulum and vacuole. Four constructs harboring faeB were transiently expressed in Nicotiana leaves, with FAEB accumulating at high levels in all target sites, except chloroplast. Stable transformed lines of alfalfa were subsequently obtained using Agrobacterium tumefaciens (LBA4404). Out of 136 transgenic plants regenerated, 18 independent lines exhibited FAEB activity. Subsequent in vitro digestibility and Fourier transformed infrared spectroscopy (FTIR) analysis of FAEB-expressing lines showed that they possessed modified cell wall morphology and composition with a reduction in ester linkages and elevated lignin content. Consequently, they were more recalcitrant to digestion by mixed ruminal microorganisms. Interestingly, delignification by alkaline peroxide treatment followed by exposure to a commercial cellulase mixture resulted in higher glucose release from transgenic lines as compared to the control line. Conclusion Modifying cell wall crosslinking has the potential to lower recalcitrance of holocellulose, but also exhibited unintended consequences on alfalfa cell wall digestibility due to elevated lignin content. The combination of efficient delignification treatment (alkaline peroxide) and transgenic esterase activity complement each other towards efficient and effective digestion of transgenic lines. PMID:24650274

  17. Analysis of transgene-specific immune responses that limit the in vivo persistence of adoptively transferred HSV-TK–modified donor T cells after allogeneic hematopoietic cell transplantation

    PubMed Central

    Berger, Carolina; Flowers, Mary E.; Warren, Edus H.; Riddell, Stanley R.

    2006-01-01

    The introduction of an inducible suicide gene such as the herpes simplex virus thymidine kinase (HSV-TK) might allow exploitation of the antitumor activity of donor T cells after allogeneic hematopoietic cell transplantation (HCT) without graft versus host disease. However, HSV-TK is foreign, and immune responses to gene-modified T cells could lead to their premature elimination. We show that after the infusion of HSV-TK–modified donor T cells to HCT recipients, CD8+ and CD4+ T-cell responses to HSV-TK are rapidly induced and coincide with the disappearance of transferred cells. Cytokine flow cytometry using an overlapping panel of HSV-TK peptides allowed rapid detection and quantitation of HSV-TK–specific T cells in the blood and identified multiple immunogenic epitopes. Repeated infusion of modified T cells boosted the induced HSV-TK–specific T cells, which persisted as memory cells. These studies demonstrate the need for nonimmunogenic suicide genes and identify a strategy for detection of CD4+ and CD8+ T-cell responses to transgene products that should be generally applicable to monitoring patients on gene therapy trials. The potency of gene-modified T cells to elicit robust and durable immune responses imply this approach might be used for vaccination to elicit T-cell responses to viral or tumor antigens. PMID:16282341

  18. Cerium-modified doped strontium titanate compositions for solid oxide fuel cell anodes and electrodes for other electrochemical devices

    DOEpatents

    Marina, Olga A [Richland, WA; Stevenson, Jeffry W [Richland, WA

    2010-11-23

    The present invention provides novel compositions that find advantageous use in making electrodes for electrochemical cells and electrochemical devices such as solid oxide fuel cells, electrolyzers, sensors, pumps and the like, the compositions comprising cerium-modified doped strontium titanate. The invention also provides novel methods for making and using anode material compositions and solid oxide fuel cells and solid oxide fuel cell assemblies having anodes comprising the compositions.

  19. Cerium-modified doped strontium titanate compositions for solid oxide fuel cell anodes and electrodes for other electrochemical devices

    DOEpatents

    Marina, Olga A [Richland, WA; Stevenson, Jeffry W [Richland, WA

    2010-03-02

    The present invention provides novel compositions that find advantageous use in making electrodes for electrochemical cells and electrochemical devices such as solid oxide fuel cells, electrolyzers, sensors, pumps and the like, the compositions comprising cerium-modified doped strontium titanate. The invention also provides novel methods for making and using anode material compositions and solid oxide fuel cells and solid oxide fuel cell assemblies having anodes comprising the compositions.

  20. In vitro response of hFOB cells to pamidronate modified sodium silicate coated cellulose scaffolds.

    PubMed

    Ponader, Sabine; Brandt, Heike; Vairaktaris, Eleftherios; von Wilmowsky, Cornelius; Nkenke, Emeka; Schlegel, Karl A; Neukam, Friedrich W; Holst, Stefan; Müller, Frank A; Greil, Peter

    2008-07-15

    The aim of the present study was to evaluate the suitability of cellulose-based scaffolds coated with pure sodium silicate gel and sodium silicate gels accumulated with different concentrations of the bisphosphonate pamidronate as scaffolds for attachment, proliferation and differentiation of human fetal osteoblasts (hFOB 1.19). Human osteoblasts were cultured in vitro for a period up to 14 days on different cellulose scaffolds. Unmodified and sodium silicate coated cellulose scaffolds were used as control. Two surface-coated modifications of cellulose were applied. The scaffolds were coated in a modified two-step dip coating process with pure sodium silicate gel and pamidronate enriched sodium silicate gel, respectively. In order to investigate the influence of the pamidronate, concentrations of 0.667 mg Na-pamidronate/ml sodium silicate solution, 0.333 mg Na-pamidronate/ml sodium silicate solution and 3.33 x 10(-3) mg Na-pamidronate/ml sodium silicate solution were used for the coating process. Cell proliferation, vitality and attachment were examined by means of cell counting, WST-1 test, fluorescence and scanning electron microscopy. The relative grade of differentiation of hFOB cells was examined by using quantitative real-time polymerase chain reaction (qRT-PCR) analysis for the gene expression of alkaline phosphatase and osteocalcin. Proliferation and differentiation of human osteoblasts was enhanced by the sodium silicate coatings accumulated with pamidronate compared to pure sodium silicate coatings. There was a reciprocal correlation of vitality with the concentration of pamidronate. The highest vitality was found on surfaces with the lowest pamidronate accumulation. Alkaline phosphatase, an early differentiation marker, was overexpressed after 7 days in cells on all pamidronate-containing surfaces (up to 350% compared to untreated cellulose). Osteocalcin, a late differentiation marker, was overexpressed after 14 days in cells on all coated surfaces (up

  1. Altered folate metabolism modifies cell proliferation and progesterone secretion in human placental choriocarcinoma JEG-3 cells.

    PubMed

    Moussa, Carolyne; Ross, Nikia; Jolette, Philippe; MacFarlane, Amanda J

    2015-09-28

    Folate is an essential B vitamin required for de novo purine and thymidylate synthesis, and for the remethylation of homocysteine to form methionine. Folate deficiency has been associated with placenta-related pregnancy complications, as have SNP in genes of the folate-dependent enzymes, methionine synthase (MTR) and methylenetetrahydrofolate dehydrogenase 1 (MTHFD1). We aimed to determine the effect of altered folate metabolism on placental cell proliferation, viability and invasive capacity and on progesterone and human chorionic gonadotropin (hCG) secretion. Human placental choriocarcinoma (JEG-3) cells cultured in low folic acid (FA) (2 nM) demonstrated 13% (P<0.001) and 26% (P<0.001) lower proliferation, 5.5% (P=0.025) and 7.5% (P=0.004) lower invasion capacity, and 5 to 7.5% (P=0.004-0.025) lower viability compared with control (20 nM) or supplemented (100 nM) cells, respectively. FA concentration had no effect on progesterone or hCG secretion. Small interfering RNA (siRNA) knockdown of MTR gene and protein expression resulted in 17.7% (P<0.0001) lower proliferation and 61% (P=0.014) higher progesterone secretion, but had no effect on cell invasion and hCG secretion. siRNA knockdown of MTHFD1 gene expression in the absence of detectable changes in protein expression resulted in 10.3% (P=0.001) lower cell proliferation, but had no effect on cell invasion and progesterone or hCG secretion. Our data indicate that impaired folate metabolism can result in lower trophoblast proliferation, and could alter viability, invasion capacity and progesterone secretion, which may explain in part the observed associations between folate and placenta-related complications.

  2. Modified arabinoxylan rice bran (MGN-3/Biobran) sensitizes human T cell leukemia cells to death receptor (CD95)-induced apoptosis.

    PubMed

    Ghoneum, Mamdooh; Gollapudi, Sastry

    2003-11-10

    MGN-3, an arabinoxylan extracted from rice bran that is treated enzymatically with an extract from Shiitaki mushrooms, is an effective biological response modifier that increases NK cell activity, and potentiates the activity of conventional chemotherapeutic agents. In this study, we investigated the effect of MGN-3 on death receptor-induced apoptosis in the human leukemic HUT 78 cell line. HUT 78 cells were pre-treated with MGN-3, and then were incubated with the agonistic antibody against death receptor (Fas, CD95). Apoptosis was determined by the propidium iodide technique using FACScan. Activation of caspase 3, caspase 8, and caspase 9 was determined by flow cytometry. Mitochondrial membrane potential was measured with DIOC(6) dye using FACScan. Expression of CD95 and Bcl-2 were measured by flow cytometry. In a dose-dependent manner, MGN-3 enhanced anti-CD95 antibody-induced apoptosis. Increased cell death was correlated with increased depolarization of mitochondrial membrane potential and increased activation of caspase 3, caspase 8, and caspase 9. MGN-3 treatment had no effect on the level of expression of CD95, but it caused down regulation of Bcl-2 expression. These results suggest that MGN-3 increases the susceptibility of cancer cells to undergo apoptosis mediated by death ligands, which may be relevant for anti-cancer activities.

  3. 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...

  4. Multidentate zwitterionic chitosan oligosaccharide modified gold nanoparticles: stability, biocompatibility and cell interactions

    NASA Astrophysics Data System (ADS)

    Liu, Xiangsheng; Huang, Haoyuan; Liu, Gongyan; Zhou, Wenbo; Chen, Yangjun; Jin, Qiao; Ji, Jian

    2013-04-01

    Surface engineering of nanoparticles plays an essential role in their colloidal stability, biocompatibility and interaction with biosystems. In this study, a novel multidentate zwitterionic biopolymer derivative is obtained from conjugating dithiolane lipoic acid and zwitterionic acryloyloxyethyl phosphorylcholine to the chitosan oligosaccharide backbone. Gold nanoparticles (AuNPs) modified by this polymer exhibit remarkable colloidal stabilities under extreme conditions including high salt conditions, wide pH range and serum or plasma containing media. The AuNPs also show strong resistance to competition from dithiothreitol (as high as 1.5 M). Moreover, the modified AuNPs demonstrate low cytotoxicity investigated by both MTT and LDH assays, and good hemocompatibility evaluated by hemolysis of human red blood cells. In addition, the intracellular fate of AuNPs was investigated by ICP-MS and TEM. It showed that the AuNPs are uptaken by cells in a concentration dependent manner, and they can escape from endosomes/lysosomes to cytosol and tend to accumulate around the nucleus after 24 h incubation but few of them are excreted out of the cells. Gold nanorods are also stabilized by this ligand, which demonstrates robust dispersion stability and excellent hemocompatibility. This kind of multidentate zwitterionic chitosan derivative could be widely used for stabilizing other inorganic nanoparticles, which will greatly improve their performance in a variety of bio-related applications.Surface engineering of nanoparticles plays an essential role in their colloidal stability, biocompatibility and interaction with biosystems. In this study, a novel multidentate zwitterionic biopolymer derivative is obtained from conjugating dithiolane lipoic acid and zwitterionic acryloyloxyethyl phosphorylcholine to the chitosan oligosaccharide backbone. Gold nanoparticles (AuNPs) modified by this polymer exhibit remarkable colloidal stabilities under extreme conditions including high salt

  5. Sex Hormone Binding Globulin Modifies Testosterone Action and Metabolism in Prostate Cancer Cells.

    PubMed

    Li, Huika; Pham, Thy; McWhinney, Brett C; Ungerer, Jacobus P; Pretorius, Carel J; Richard, Derek J; Mortimer, Robin H; d'Emden, Michael C; Richard, Kerry

    2016-01-01

    Sex Hormone Binding Globulin (SHBG) is the major serum carrier of sex hormones. However, growing evidence suggests that SHBG is internalised and plays a role in regulating intracellular hormone action. This study was to determine whether SHBG plays a role in testosterone uptake, metabolism, and action in the androgen sensitive LNCaP prostate cancer cell line. Internalisation of SHBG and testosterone, the effects of SHBG on testosterone uptake, metabolism, regulation of androgen responsive genes, and cell growth were assessed. LNCaP cells internalised SHBG by a testosterone independent process. Testosterone was rapidly taken up and effluxed as testosterone-glucuronide; however this effect was reduced by the presence of SHBG. Addition of SHBG, rather than reducing testosterone bioavailability, further increased testosterone-induced expression of prostate specific antigen and enhanced testosterone-induced reduction of androgen receptor mRNA expression. Following 38 hours of testosterone treatment cell morphology changed and growth declined; however, cotreatment with SHBG abrogated these inhibitory effects. These findings clearly demonstrate that internalised SHBG plays an important regulatory and intracellular role in modifying testosterone action and this has important implications for the role of SHBG in health and disease.

  6. Modified titanium surface with gelatin nano gold composite increases osteoblast cell biocompatibility

    NASA Astrophysics Data System (ADS)

    Lee, Young-Hee; Bhattarai, Govinda; Aryal, Santosh; Lee, Nan-Hee; Lee, Min-Ho; Kim, Tae-Gun; Jhee, Eun-Chung; Kim, Hak-Yong; Yi, Ho-Keun

    2010-08-01

    This study examined the gelatin nano gold (GnG) composite for surface modification of titanium in addition to insure biocompatibility on dental implants or biomaterials. The GnG composite was constructed by gelatin and hydrogen tetrachloroaurate in presence of reducing agent, sodium borohydrate (NabH 4). The GnG composite was confirmed by UV-VIS spectroscopy and transmission electron microscopy (TEM). A dipping method was used to modify the titanium surface by GnG composite. Surface was characterized by scanning electron microscopy (SEM) and energy dispersive X-ray (EDX). The MC-3T3 E1 cell viability was assessed by trypan blue and the expression of proteins to biocompatibility were analyzed by Western blotting. The GnG composite showed well dispersed character, the strong absorption at 530 nm, roughness, regular crystal and clear C, Na, Cl, P, and Au signals onto titanium. Further, this composite allowed MC-3T3 E1 growth and viability compared to gelatin and pure titanium. It induced ERK activation and the expression of cell adherent molecules, FAK and SPARC, and growth factor, VEGF. However, GnG decreased the level of SAPK/JNK. This shows that GnG composite coated titanium surfaces have a good biocompatibility for osteoblast growth and attachment than in intact by simple and versatile dipping method. Furthermore, it offers good communication between cell and implant surfaces by regulating cell signaling and adherent molecules, which are useful to enhance the biocompatibility of titanium surfaces.

  7. Sex Hormone Binding Globulin Modifies Testosterone Action and Metabolism in Prostate Cancer Cells

    PubMed Central

    Li, Huika; Ungerer, Jacobus P.; Pretorius, Carel J.; Mortimer, Robin H.; d'Emden, Michael C.

    2016-01-01

    Sex Hormone Binding Globulin (SHBG) is the major serum carrier of sex hormones. However, growing evidence suggests that SHBG is internalised and plays a role in regulating intracellular hormone action. This study was to determine whether SHBG plays a role in testosterone uptake, metabolism, and action in the androgen sensitive LNCaP prostate cancer cell line. Internalisation of SHBG and testosterone, the effects of SHBG on testosterone uptake, metabolism, regulation of androgen responsive genes, and cell growth were assessed. LNCaP cells internalised SHBG by a testosterone independent process. Testosterone was rapidly taken up and effluxed as testosterone-glucuronide; however this effect was reduced by the presence of SHBG. Addition of SHBG, rather than reducing testosterone bioavailability, further increased testosterone-induced expression of prostate specific antigen and enhanced testosterone-induced reduction of androgen receptor mRNA expression. Following 38 hours of testosterone treatment cell morphology changed and growth declined; however, cotreatment with SHBG abrogated these inhibitory effects. These findings clearly demonstrate that internalised SHBG plays an important regulatory and intracellular role in modifying testosterone action and this has important implications for the role of SHBG in health and disease. PMID:27990161

  8. Lipid-modified G4-decoy oligonucleotide anchored to nanoparticles: delivery and bioactivity in pancreatic cancer cells

    PubMed Central

    Cogoi, S.; Jakobsen, U.; Pedersen, E. B.; Vogel, S.; Xodo, L. E.

    2016-01-01

    KRAS is mutated in >90% of pancreatic ductal adenocarcinomas. As its inactivation leads to tumour regression, mutant KRAS is considered an attractive target for anticancer drugs. In this study we report a new delivery strategy for a G4-decoy oligonucleotide that sequesters MAZ, a transcription factor essential for KRAS transcription. It is based on the use of palmitoyl-oleyl-phosphatidylcholine (POPC) liposomes functionalized with lipid-modified G4-decoy oligonucleotides and a lipid-modified cell penetrating TAT peptide. The potency of the strategy in pancreatic cancer cells is demonstrated by cell cytometry, confocal microscopy, clonogenic and qRT-PCR assays. PMID:27929127

  9. Phospholipase signaling is modified differentially by phytoregulators in Capsicum chinense J. cells.

    PubMed

    Muñoz-Sánchez, J Armando; Altúzar-Molina, Alma; Hérnandez-Sotomayor, S M Teresa

    2012-09-01

    Plant defense mechanisms respond to diverse environmental factors and play key roles in signaling pathways. The phospholipidic signaling pathway forms part of the plant response to several phytoregulators, such as salicylic acid (SA) and methyl jasmonate (MJ), which have been widely used to stimulate secondary metabolite production in cell cultures. ( 1) Furthermore, it has been reported that the levels of such phytoregulators as SA and MJ can increase in response to stressful conditions. ( 2) (,) ( 3) The phospholipidic signal transduction system involves the generation of second messengers by the hydrolysis of phospholipids. In this study, we examined how phospholipidic signaling can be modulated depending on the growth stage of the culture, and we focused on two key lipases having relevant roles in the signaling cascades in plants. An evaluation was made of the effects of SA and MJ on the phospholipase activities in Capsicum chinense Jacq. suspension cells at different phases of the culture cycle. The treatment with SA differentially modified the phospholipase C (PLC) (EC: 3.1.4.3) and phospholipase D (PLD) (EC: 3.1.4.4) activities in a dose-dependent manner that also depended on the day of the culture cycle. In contrast, the treatment with MJ resulted in a biphasic behavior of the PLC and PLD activities. We conclude that the enzymatic activities in the phospholipidic signaling pathways are modified differentially depending on the day of the culture's growth cycle; accordingly, the response capacity to such environmental factors as phytoregulators is variable at different stages of growth and the physiology of the cells.

  10. Phospholipase signaling is modified differentially by phytoregulators in Capsicum chinense J. cells

    PubMed Central

    Muñoz-Sánchez, J. Armando; Altúzar-Molina, Alma; Hérnandez-Sotomayor, S. M. Teresa

    2012-01-01

    Plant defense mechanisms respond to diverse environmental factors and play key roles in signaling pathways. The phospholipidic signaling pathway forms part of the plant response to several phytoregulators, such as salicylic acid (SA) and methyl jasmonate (MJ), which have been widely used to stimulate secondary metabolite production in cell cultures.1 Furthermore, it has been reported that the levels of such phytoregulators as SA and MJ can increase in response to stressful conditions.2,3 The phospholipidic signal transduction system involves the generation of second messengers by the hydrolysis of phospholipids. In this study, we examined how phospholipidic signaling can be modulated depending on the growth stage of the culture, and we focused on two key lipases having relevant roles in the signaling cascades in plants. An evaluation was made of the effects of SA and MJ on the phospholipase activities in Capsicum chinense Jacq. suspension cells at different phases of the culture cycle. The treatment with SA differentially modified the phospholipase C (PLC) (EC: 3.1.4.3) and phospholipase D (PLD) (EC: 3.1.4.4) activities in a dose-dependent manner that also depended on the day of the culture cycle. In contrast, the treatment with MJ resulted in a biphasic behavior of the PLC and PLD activities. We conclude that the enzymatic activities in the phospholipidic signaling pathways are modified differentially depending on the day of the culture’s growth cycle; accordingly, the response capacity to such environmental factors as phytoregulators is variable at different stages of growth and the physiology of the cells. PMID:22899070

  11. Simulation of fluid-solid coexistence via thermodynamic integration using a modified cell model.

    PubMed

    Nayhouse, Michael; Amlani, Ankur M; Heng, Vincent R; Orkoulas, G

    2012-04-18

    Despite recent advances, precise simulation of fluid-solid transitions still remains a challenging task. Thermodynamic integration techniques are the simplest methods to study fluid-solid coexistence. These methods are based on the calculation of the free energies of the fluid and the solid phases, starting from a state of known free energy which is usually an ideal-gas state. Despite their simplicity, the main drawback of thermodynamic integration techniques is the large number of states that must be simulated. In the present work, a thermodynamic integration technique, which reduces the number of simulated states, is proposed and tested on a system of particles interacting via an inverse twelfth-power potential energy function. The simulations are implemented at constant pressure and the solid phase is modeled according to the constrained cell model of Hoover and Ree. The fluid and the solid phases are linked together by performing constant-pressure simulations of a modified cell model. The modified cell model, which was originally proposed by Hoover and Ree, facilitates transitions between the fluid and the solid phase by tuning a homogeneous external field. This model is simulated on a constant-pressure path for a series of progressively increasing values of the field, thus allowing for direct determination of the free energy difference between the fluid and the solid phase via histogram reweighting. The size-dependent results are analyzed using histogram reweighting and finite-size scaling techniques. The scaling analysis is based on studying the size-dependent behavior of the second- and higher-order derivatives of the free energy as well as the dimensionless moment ratios of the order parameter. The results clearly demonstrate the importance of accounting for size effects in simulation studies of fluid-solid transitions.

  12. Quantum dots modified with quaternized poly(dimethylaminoethyl methacrylate) for selective recognition and killing of bacteria over mammalian cells.

    PubMed

    Tu, Qin; Ma, Chao; Tian, Chang; Yuan, Maosen; Han, Xiang; Wang, Dong-En; Cao, Chenyu; Wang, Jinyi

    2016-05-23

    Copper-free click chemistry has been used to graft quaternized poly(dimethylaminoethyl methacrylate) (QPA) modified with azide to the quantum dots (QDs) derived with dibenzocyclooctynes (DBCO). The success of the quaternary ammonium polymer-modified QDs was confirmed by ultraviolet-visible spectrophotometry (UV-Vis), fluorescence spectroscopy, zeta (ζ) potential, size distribution, and transmission electron microscopy (TEM). The QPA-modified QDs exhibited properties of selective recognition and killing of bacteria. The novelty of this study lies in fact that the synthesis method of the antimicrobial QPA-modified QDs is simple. Moreover, from another standpoint, QPA-modified QDs simultaneously possess abilities of selective recognition and killing of bacteria over mammalian cells, which is very different from the currently designed multifunctional antimicrobial systems composed of complicated systematic compositions.

  13. Recommendations from Gynaecological (GYN) GEC-ESTRO Working Group (IV): Basic principles and parameters for MR imaging within the frame of image based adaptive cervix cancer brachytherapy

    PubMed Central

    Dimopoulos, Johannes C.A.; Petrow, Peter; Tanderup, Kari; Petric, Primoz; Berger, Daniel; Kirisits, Christian; Pedersen, Erik M.; van Limbergen, Erik; Haie-Meder, Christine; Pötter, Richard

    2012-01-01

    The GYN GEC-ESTRO working group issued three parts of recommendations and highlighted the pivotal role of MRI for the successful implementation of 3D image-based cervical cancer brachytherapy (BT). The main advantage of MRI as an imaging modality is its superior soft tissue depiction quality. To exploit the full potential of MRI for the better ability of the radiation oncologist to make the appropriate choice for the BT application technique and to accurately define the target volumes and the organs at risk, certain MR imaging criteria have to be fulfilled. Technical requirements, patient preparation, as well as image acquisition protocols have to be tailored to the needs of 3D image-based BT. The present recommendation is focused on the general principles of MR imaging for 3D image-based BT. Methods and parameters have been developed and progressively validated from clinical experience from different institutions (IGR, Universities of Vienna, Leuven, Aarhus and Ljubljana) and successfully applied during expert meetings, contouring workshops, as well as within clinical and interobserver studies. It is useful to perform pelvic MRI scanning prior to radiotherapy (“Pre-RT-MRI examination”) and at the time of BT (“BT MRI examination”) with one MR imager. Both low and high-field imagers, as well as both open and close magnet configurations conform to the requirements of 3D image-based cervical cancer BT. Multiplanar (transversal, sagittal, coronal and oblique image orientation) T2-weighted images obtained with pelvic surface coils are considered as the golden standard for visualisation of the tumour and the critical organs. The use of complementary MRI sequences (e.g. contrast-enhanced T1-weighted or 3D isotropic MRI sequences) is optional. Patient preparation has to be adapted to the needs of BT intervention and MR imaging. It is recommended to visualise and interpret the MR images on dedicated DICOM-viewer workstations, which should also assist the contouring

  14. Recommendations from Gynaecological (GYN) GEC-ESTRO Working Group (IV): Basic principles and parameters for MR imaging within the frame of image based adaptive cervix cancer brachytherapy.

    PubMed

    Dimopoulos, Johannes C A; Petrow, Peter; Tanderup, Kari; Petric, Primoz; Berger, Daniel; Kirisits, Christian; Pedersen, Erik M; van Limbergen, Erik; Haie-Meder, Christine; Pötter, Richard

    2012-04-01

    The GYN GEC-ESTRO working group issued three parts of recommendations and highlighted the pivotal role of MRI for the successful implementation of 3D image-based cervical cancer brachytherapy (BT). The main advantage of MRI as an imaging modality is its superior soft tissue depiction quality. To exploit the full potential of MRI for the better ability of the radiation oncologist to make the appropriate choice for the BT application technique and to accurately define the target volumes and the organs at risk, certain MR imaging criteria have to be fulfilled. Technical requirements, patient preparation, as well as image acquisition protocols have to be tailored to the needs of 3D image-based BT. The present recommendation is focused on the general principles of MR imaging for 3D image-based BT. Methods and parameters have been developed and progressively validated from clinical experience from different institutions (IGR, Universities of Vienna, Leuven, Aarhus and Ljubljana) and successfully applied during expert meetings, contouring workshops, as well as within clinical and interobserver studies. It is useful to perform pelvic MRI scanning prior to radiotherapy ("Pre-RT-MRI examination") and at the time of BT ("BT MRI examination") with one MR imager. Both low and high-field imagers, as well as both open and close magnet configurations conform to the requirements of 3D image-based cervical cancer BT. Multiplanar (transversal, sagittal, coronal and oblique image orientation) T2-weighted images obtained with pelvic surface coils are considered as the golden standard for visualisation of the tumour and the critical organs. The use of complementary MRI sequences (e.g. contrast-enhanced T1-weighted or 3D isotropic MRI sequences) is optional. Patient preparation has to be adapted to the needs of BT intervention and MR imaging. It is recommended to visualise and interpret the MR images on dedicated DICOM-viewer workstations, which should also assist the contouring

  15. Transferrin-modified PLGA nanoparticles significantly increase the cytotoxicity of paclitaxel in bladder cancer cells by increasing intracellular retention

    NASA Astrophysics Data System (ADS)

    Jin, Shihua; Zhang, Yi; Yu, Chengfan; Wang, Gang; Zhang, Zhihong; Li, Ningchen; Na, Yanqun

    2014-10-01

    To improve the anticancer effects of paclitaxel (Tax) on bladder cancer, transferrin-modified and unmodified poly( d,l lactide- co-glycolide) nanoparticles (NPs) were generated to deliver Tax. The characteristics of the NPs and the drug-release profiles were evaluated. The cytotoxicity levels of blank NPs and Tax-loaded NPs in the bladder cancer cell lines MBT-2, J-82, and TCC Sup were determined. The uptakes and retentions of the NPs by the cell lines and the intracellular distribution of the NPs were also studied. The results showed similar NPs characteristics and drug-release profiles for NPs with and without transferrin modification. The sizes of NPs with and without transferrin modification were 206 and 278 nm, respectively; the Z-potentials were -23.5 and -24.3 mV, respectively; the drug loadings were 6.5 and 6.7 % w/w, respectively. No cytotoxicity was observed in the bladder cancer cells exposed to blank NPs. Both types of Tax-loaded NPs, however, had significantly higher cytotoxicity levels compared with the Tax solution in the bladder cancer cells. The transferrin-modified, Tax-loaded NPs were significantly more cytotoxic than the Tax-loaded NPs without modification in the MBT-2 and TCC Sup cells. There were no significant differences in NP uptakes between transferrin-modified and unmodified NPs in any of the three studied bladder cancer cell lines; however, the retentions of the modified NPs were significantly higher in the MBT-2 and TCC Sup cells. These findings suggest that NPs can significantly improve the anticancer effect of Tax in bladder cells. Furthermore, transferrin-modified NPs can improve the anticancer effect by increasing intracellular retention and not by increasing uptake. The transferrin-modified NPs are promising drug delivery vehicle for bladder cancer treatment.

  16. The potential role of genetically-modified pig mesenchymal stromal cells in xenotransplantation.

    PubMed

    Li, Jiang; Ezzelarab, Mohamed B; Ayares, David; Cooper, David K C

    2014-02-01

    Mesenchymal stromal cells (MSCs) are known to have regenerative, anti-inflammatory, and immunodulatory effects. There are extensive indications that pig MSCs function satisfactorily across species barriers. Pig MSCs might have considerable therapeutic potential, particularly in xenotransplantation, where they have several potential advantages. (i) pMSCs can be obtained from the specific organ- or cell-source donor pig or from an identical (cloned) pig. (ii) They are easy to obtain in large numbers, negating the need for prolonged ex vivo expansion. (iii) They can be obtained from genetically-engineered pigs, and the genetic modification can be related to the therapeutic goal of the MSCs. We have reviewed our own studies on MSCs from genetically-engineered pigs, and summarize them here. We have successfully harvested and cultured MSCs from wild-type and genetically-engineered pig bone marrow and adipose tissue. We have identified several pig (p)MSC surface markers (positive for CD29, CD44, CD73, CD105, CD166, and negative for CD31, CD45), have demonstrated their proliferation and differentiation (into adipocytes, osteoblasts, and chondroblasts), and evaluated their antigenicity and immune suppressive effects on human peripheral blood mononuclear cells and CD4(+)T cells. They have identical or very similar characteristics to MSCs from other mammals. Genetically-modified pMSCs are significantly less immunogenic than wild-type pMSCs, and downregulate the human T cell response to pig antigens as efficiently as do human MSCs. We hypothesized that pMSCs can immunomodulate human T cells through induction of apoptosis or anergy, or cause T cell phenotype switching with induction of regulatory T cells, but we could find no evidence for these mechanisms. However, pMSCs upregulated the expression of CD69 on human CD4(+) and CD8(+) T cells, the relevance of which is currently under investigation. We conclude that MSCs from genetically-engineered pigs should continue to be

  17. Analysis of trunk neural crest cell migration using a modified Zigmond chamber assay.

    PubMed

    Walheim, Christopher C; Zanin, Juan Pablo; de Bellard, Maria Elena

    2012-01-19

    Neural crest cells (NCCs) are a transient population of cells present in vertebrate development that emigrate from the dorsal neural tube (NT) after undergoing an epithelial-mesenchymal transition. Following EMT, NCCs migrate large distances along stereotypic pathways until they reach their targets. NCCs differentiate into a vast array of cell types including neurons, glia, melanocytes, and chromaffin cells. The ability of NCCs to reach and recognize their proper target locations is foundational for the appropriate formation of all structures containing trunk NCC-derived components. Elucidating the mechanisms of guidance for trunk NCC migration has therefore been a matter of great significance. Numerous molecules have been demonstrated to guide NCC migration. For instance, trunk NCCs are known to be repelled by negative guidance cues such as Semaphorin, Ephrin, and Slit ligands. However, not until recently have any chemoattractants of trunk NCCs been identified. Conventional in vitro approaches to studying the chemotactic behavior of adherent cells work best with immortalized, homogenously distributed cells, but are more challenging to apply to certain primary stem cell cultures that initially lack a homogenous distribution and rapidly differentiate (such as NCCs). One approach to homogenize the distribution of trunk NCCs for chemotaxis studies is to isolate trunk NCCs from primary NT explant cultures, then lift and replate them to be almost 100% confluent. However, this plating approach requires substantial amounts of time and effort to explant enough cells, is harsh, and distributes trunk NCCs in a dissimilar manner to that found in in vivo conditions. Here, we report an in vitro approach that is able to evaluate chemotaxis and other migratory responses of trunk NCCs without requiring a homogenous cell distribution. This technique utilizes time-lapse imaging of primary, unperturbed trunk NCCs inside a modified Zigmond chamber (a standard Zigmond chamber is

  18. Manufacture of gene-modified human T-cells with a memory stem/central memory phenotype.

    PubMed

    Gomez-Eerland, Raquel; Nuijen, Bastiaan; Heemskerk, Bianca; van Rooij, Nienke; van den Berg, Joost H; Beijnen, Jos H; Uckert, Wolfgang; Kvistborg, Pia; Schumacher, Ton N; Haanen, John B A G; Jorritsma, Annelies

    2014-10-01

    Advances in genetic engineering have made it possible to generate human T-cell products that carry desired functionalities, such as the ability to recognize cancer cells. The currently used strategies for the generation of gene-modified T-cell products lead to highly differentiated cells within the infusion product, and on the basis of data obtained in preclinical models, this is likely to impact the efficacy of these products. We set out to develop a good manufacturing practice (GMP) protocol that yields T-cell receptor (TCR) gene-modified T-cells with more favorable properties for clinical application. Here, we show the robust clinical-scale production of human peripheral blood T-cells with an early memory phenotype that express a MART-1-specific TCR. By combining selection and stimulation using anti-CD3/CD28 beads for retroviral transduction, followed by expansion in the presence of IL-7 and IL-15, production of a well-defined clinical-scale TCR gene-modified T-cell product could be achieved. A major fraction of the T-cells generated in this fashion were shown to coexpress CD62L and CD45RA, and express CD27 and CD28, indicating a central memory or memory stemlike phenotype. Furthermore, these cells produced IFNγ, TNFα, and IL-2 and displayed cytolytic activity against target cells expressing the relevant antigen. The T-cell products manufactured by this robust and validated GMP production process are now undergoing testing in a phase I/IIa clinical trial in HLA-A*02:01 MART-1-positive advanced stage melanoma patients. To our knowledge, this is the first clinical trial protocol in which the combination of IL-7 and IL-15 has been applied for the generation of gene-modified T-cell products.

  19. Manufacture of Gene-Modified Human T-Cells with a Memory Stem/Central Memory Phenotype

    PubMed Central

    Gomez-Eerland, Raquel; Nuijen, Bastiaan; Heemskerk, Bianca; van Rooij, Nienke; van den Berg, Joost H.; Beijnen, Jos H.; Uckert, Wolfgang; Kvistborg, Pia; Schumacher, Ton N.; Jorritsma, Annelies

    2014-01-01

    Abstract Advances in genetic engineering have made it possible to generate human T-cell products that carry desired functionalities, such as the ability to recognize cancer cells. The currently used strategies for the generation of gene-modified T-cell products lead to highly differentiated cells within the infusion product, and on the basis of data obtained in preclinical models, this is likely to impact the efficacy of these products. We set out to develop a good manufacturing practice (GMP) protocol that yields T-cell receptor (TCR) gene-modified T-cells with more favorable properties for clinical application. Here, we show the robust clinical-scale production of human peripheral blood T-cells with an early memory phenotype that express a MART-1-specific TCR. By combining selection and stimulation using anti-CD3/CD28 beads for retroviral transduction, followed by expansion in the presence of IL-7 and IL-15, production of a well-defined clinical-scale TCR gene-modified T-cell product could be achieved. A major fraction of the T-cells generated in this fashion were shown to coexpress CD62L and CD45RA, and express CD27 and CD28, indicating a central memory or memory stemlike phenotype. Furthermore, these cells produced IFNγ, TNFα, and IL-2 and displayed cytolytic activity against target cells expressing the relevant antigen. The T-cell products manufactured by this robust and validated GMP production process are now undergoing testing in a phase I/IIa clinical trial in HLA-A*02:01 MART-1-positive advanced stage melanoma patients. To our knowledge, this is the first clinical trial protocol in which the combination of IL-7 and IL-15 has been applied for the generation of gene-modified T-cell products. PMID:25143008

  20. Genetically modified neural stem cells for a local and sustained delivery of neuroprotective factors to the dystrophic mouse retina.

    PubMed

    Jung, Gila; Sun, Jing; Petrowitz, Bettina; Riecken, Kristoffer; Kruszewski, Katharina; Jankowiak, Wanda; Kunst, Frank; Skevas, Christos; Richard, Gisbert; Fehse, Boris; Bartsch, Udo

    2013-12-01

    A continuous intraocular delivery of neurotrophic factors (NFs) is being explored as a strategy to rescue photoreceptor cells and visual functions in degenerative retinal disorders that are currently untreatable. To establish a cell-based intraocular delivery system for a sustained administration of NFs to the dystrophic mouse retina, we used a polycistronic lentiviral vector to genetically modify adherently cultivated murine neural stem (NS) cells. The vector concurrently encoded a gene of interest, a reporter gene, and a resistance gene and thus facilitated the selection, cloning, and in vivo tracking of the modified cells. To evaluate whether modified NS cells permit delivery of functionally relevant quantities of NFs to the dystrophic mouse retina, we expressed a secretable variant of ciliary neurotrophic factor (CNTF) in NS cells and grafted the cells into the vitreous space of Pde6b(rd1) and Pde6b(rd10) mice, two animal models of retinitis pigmentosa. In both mouse lines, grafted cells attached to the retina and lens, where they differentiated into astrocytes and some neurons. Adverse effects of the transplanted cells on the morphology of host retinas were not observed. Importantly, the CNTF-secreting NS cells significantly attenuated photoreceptor degeneration in both mutant mouse lines. The neuroprotective effect was significantly more pronounced when clonally derived NS cell lines selected for high expression levels of CNTF were grafted into Pde6b(rd1) mice. Intravitreal transplantations of modified NS cells may thus represent a useful method for preclinical studies aimed at evaluating the therapeutic potential of a cell-based intraocular delivery of NFs in mouse models of photoreceptor degeneration.

  1. Adoptive immunotherapy for hematological malignancies using T cells gene-modified to express tumor antigen-specific receptors.

    PubMed

    Fujiwara, Hiroshi

    2014-12-15

    Accumulating clinical evidence suggests that adoptive T-cell immunotherapy could be a promising option for control of cancer; evident examples include the graft-vs-leukemia effect mediated by donor lymphocyte infusion (DLI) and therapeutic infusion of ex vivo-expanded tumor-infiltrating lymphocytes (TIL) for melanoma. Currently, along with advances in synthetic immunology, gene-modified T cells retargeted to defined tumor antigens have been introduced as "cellular drugs". As the functional properties of the adoptive immune response mediated by T lymphocytes are decisively regulated by their T-cell receptors (TCRs), transfer of genes encoding target antigen-specific receptors should enable polyclonal T cells to be uniformly redirected toward cancer cells. Clinically, anticancer adoptive immunotherapy using genetically engineered T cells has an impressive track record. Notable examples include the dramatic benefit of chimeric antigen receptor (CAR) gene-modified T cells redirected towards CD19 in patients with B-cell malignancy, and the encouraging results obtained with TCR gene-modified T cells redirected towards NY-ESO-1, a cancer-testis antigen, in patients with advanced melanoma and synovial cell sarcoma. This article overviews the current status of this treatment option, and discusses challenging issues that still restrain the full effectiveness of this strategy, especially in the context of hematological malignancy.

  2. Tuning the work functions of graphene quantum dot-modified electrodes for polymer solar cell applications.

    PubMed

    Zhang, L; Ding, Z C; Tong, T; Liu, J

    2017-03-09

    The graphene quantum dot (GQD) is a new kind of anode/cathode interlayer material for polymer solar cells (PSCs). The key requirement for a cathode interlayer (CIL) is a low work function. In this article, aiming at application as a CIL for PSCs, we report a general approach to tune the work function of GQD-modified electrodes using alkali metal cations, e.g. Li(+), Na(+), K(+), Rb(+) and Cs(+). For ITO electrodes modified with these GQDs containing alkali metal cations, the work function can be finely tuned within the range of 4.0-4.5 eV. Owing to their low work function, GQDs containing K(+), Rb(+) and Cs(+) can be used as CILs for PSCs. Their device performance is fairly comparable to that of the state-of-the-art CIL material ZnO. This work provides a rational approach to tune the properties of GQD and to design solution-processable electrode interlayer materials for organic electronic devices.

  3. The O2 reduction at the IFC modified O2 fuel cell electrode

    NASA Technical Reports Server (NTRS)

    Fielder, William L.; Singer, Joseph; Baldwin, Richard S.; Johnson, Richard E.

    1992-01-01

    The International Fuel Corporation (IFC) state of the art (SOA) O2 electrode (Au-10 percent Pt electrocatalyst by weight) is currently being used in the alkaline H2-O2 fuel cell in the NASA Space Shuttle. Recently, IFC modified O2 electrode, as a possible replacement for the SOA electrode. In the present study, O2 reduction data were obtained for the modified electrode at temperatures between 23.3 and 91.7 C. BET measurements gave an electrode BET surface area of about 2070 sq. cm/sq. cm of geometric surface area. The Tafel data could be fitted to two straight line regions. The slope for the lower region, designated as the 0.04 V/decade region, was temperature dependent, and the transfer coefficient was about 1.5. The 'apparent' energy of activation for this region was about 19 kcal/mol. An O2 reduction mechanism for this 0.04 region is presented. In the upper region, designated as the 0.08 V/decade region, diffusion may be the controlling process. Tafel data are presented to illustrate the increase in performance with increasing temperature.

  4. Endothelial cell functions in vitro cultured on poly(L-lactic acid) membranes modified with different methods.

    PubMed

    Zhu, Yabin; Gao, Changyou; Liu, Yunxiao; Shen, Jiacong

    2004-06-01

    We recently developed several methods to enhance the cell-polymer interactions. Optimal conditions for each method have been revealed separately by in vitro cell culture. As a practical consideration for construction of tissue-engineered organs, it is necessary to consider which is the most suitable and convenient in clinical applications. To compare the efficiency of these methods with respect to cell functions, poly-L-lactic acid (PLLA) was selected as matrix being modified by 1) aminolysis (PLLA-NH(2)), 2) collagen immobilization with GA (PLLA-GA-Col), 3) chondroitin sulfate (CS)/collagen layer-by-layer (LBL) assembly (PLLA-CS/Col), 4) photo-induced grafting copolymerization of hydrophilic methacrylic acid (MAA) (PLLA-g-PMAA), and 5) further immobilization of collagen with 1-ethyl-3-(3-dimethylamino propyl) carbodiimide hydrochloride (EDAC) (PLLA-g-PMAA-Col). The surface wettability of the modified PLLA was determined by water contact angle measurements. The cell response to the modified PLLA was quantitatively assessed and compared by using human umbilical endothelial cells (HUVECs) culture. Our results indicate that all the modifications can improve the cytocompatibility of PLLA (e.g., cells can attach with spreading morphology, proliferate and secret vWF and 6-keto-PGF(1 alpha)). All the collagen-modified PLLA showed more positive cell response than those purely aminolyzed or PMAA grafted. Among all the methods, collagen immobilization by LBL assembly or GA bridging after aminolysis is more acceptable for the convenience and applicability to scaffolds.

  5. Effects of Foxp3 gene modified dendritic cells on mouse corneal allograft rejection

    PubMed Central

    Gong, Yu-Bo; Hu, Lian-Na; Liu, Yong; Han, Gen-Cheng; Guo, Hui-Ling; Luo, Ling; Wang, Li-Qiang; Li, Yan; Huang, Yi-Fei

    2015-01-01

    Objective: To investigate the effect of Foxp3 gene modified dendritic cells (Foxp3 + DC) on allogeneic T cells proliferation and to study the effect of Foxp3 + DC on corneal allograft rejection. Methods: Lentivirus-Foxp3 was transfected into DC2.4 cells, as Foxp3 + DC cells. 42 BALB/c mice were randomly divided into: Group A (n = 6), normal group; Group B (n = 12), Group C (n = 12) and Group D (n = 12), allograft groups, were treated with normal saline, DC2.4, Foxp3 + DC by intraperitoneal injection, respectively. Results: Compared with the control group, Foxp3 protein in the Foxp3 + DC cells increased significantly (P < 0.05); the expressions of CD80 and CD86 immunophenotypes of Foxp3 + DC cells decreased significantly (P < 0.05); IL-12 secretion reduced (P < 0.05), but IL-10 secretion was promoted (P < 0.05). The average transplant survival time in Group B was (14.833 ± 1.472) d, and Group C and Group D led to a statistically significant prolongation of transplant survival to (17.667 ± 1.366, 23.000 ± 2.000) d (P < 0.05) respectively. 14 d after transplantation, as compared with Group C and D, the expressions of IFN-γ in grafts markedly increased in Group B. 14 d after transplantation, as compared with Group B, the expressions of Foxp3 mRNA, IDO mRNA in grafts decreased remarkably in Group C and D (P < 0.05); as compared with Group C, the expressions of Foxp3 mRNA, IDO mRNA in grafts decreased remarkably in Group D (P < 0.05). Conclusion: Foxp3 + DC cells reduce the expression of costimulatory factors, reduce the secretion of IL-12, promote IL-10 production and inhibit the stimulation of alloreactive T cell proliferation response capacity. Foxp3 + DC cells play important roles in inhibiting corneal allograft immune response and prolonging graft survival time. PMID:26064298

  6. Glucose deprivation induces G2/M transition-arrest and cell death in N-GlcNAc2-modified protein-producing renal carcinoma cells.

    PubMed

    Isono, Takahiro; Chano, Tokuhiro; Kitamura, Asuka; Yuasa, Takeshi

    2014-01-01

    Some cancer cells can survive under glucose deprivation within the microenvironment of a tumor. Recently, we reported that N-linked (β-N-acetylglucosamine)2 [N-GlcNAc2]-modified proteins induce G2/M arrest and cell death under glucose deprivation. Here, we investigated whether such a response to glucose deprivation contributes to the survival of renal cell carcinomas, which are sensitive to nutritional stress. Specifically, we analyzed seven renal carcinoma cell lines. Four of these cell lines produced N-GlcNAc2-modified proteins and led G2/M-phase arrest under glucose deprivation, leading to cell death. The remaining three cell lines did not produce N-GlcNAc2-modified proteins and undergo G1/S-phase arrest under glucose deprivation, leading to survival. The four dead cell lines displayed significant up-regulation in the UDP-GlcNAc biosynthesis pathway as well as increased phosphorylation of p53, which was not observed in the surviving three cell lines. In addition, the four dead cell lines showed prolonged up-regulated expression of ATF3, which is related to unfolded protein response (UPR), while the surviving three cell lines showed only transient up-regulation of ATF3. In this study, we demonstrated that the renal carcinoma cells which accumulate N-GlcNAc2-modified proteins under glucose deprivation do not survive with abnormaly prolonged UPR pathway. By contrast, renal carcinoma cells that do not accumulate N-GlcNAc2-modified proteins under these conditions survive. Morover, we demonstrated that buformin, a UPR inhibitor, efficiently reduced cell survival under conditions of glucose deprivation for both sensitive and resistant phenotypes. Further studies to clarify these findings will lead to the development of novel chemotherapeutic treatments for renal cancer.

  7. Glucose Deprivation Induces G2/M Transition-Arrest and Cell Death in N-GlcNAc2-Modified Protein-Producing Renal Carcinoma Cells

    PubMed Central

    Isono, Takahiro; Chano, Tokuhiro; Kitamura, Asuka; Yuasa, Takeshi

    2014-01-01

    Some cancer cells can survive under glucose deprivation within the microenvironment of a tumor. Recently, we reported that N-linked (β-N-acetylglucosamine)2 [N-GlcNAc2]-modified proteins induce G2/M arrest and cell death under glucose deprivation. Here, we investigated whether such a response to glucose deprivation contributes to the survival of renal cell carcinomas, which are sensitive to nutritional stress. Specifically, we analyzed seven renal carcinoma cell lines. Four of these cell lines produced N-GlcNAc2-modified proteins and led G2/M-phase arrest under glucose deprivation, leading to cell death. The remaining three cell lines did not produce N-GlcNAc2-modified proteins and undergo G1/S-phase arrest under glucose deprivation, leading to survival. The four dead cell lines displayed significant up-regulation in the UDP-GlcNAc biosynthesis pathway as well as increased phosphorylation of p53, which was not observed in the surviving three cell lines. In addition, the four dead cell lines showed prolonged up-regulated expression of ATF3, which is related to unfolded protein response (UPR), while the surviving three cell lines showed only transient up-regulation of ATF3. In this study, we demonstrated that the renal carcinoma cells which accumulate N-GlcNAc2-modified proteins under glucose deprivation do not survive with abnormaly prolonged UPR pathway. By contrast, renal carcinoma cells that do not accumulate N-GlcNAc2-modified proteins under these conditions survive. Morover, we demonstrated that buformin, a UPR inhibitor, efficiently reduced cell survival under conditions of glucose deprivation for both sensitive and resistant phenotypes. Further studies to clarify these findings will lead to the development of novel chemotherapeutic treatments for renal cancer. PMID:24796485

  8. Development of biologically modified anodes for energy harvesting using microbial fuel cells

    NASA Astrophysics Data System (ADS)

    Sumner, James J.; Ganguli, Rahul; Chmelka, Brad

    2012-06-01

    Biological fuel cells hold promise as an alternative energy source to batteries for unattended ground sensor applications due to the fact that they can be extremely long lived. This lifetime can be extended over batteries by scavenging fuel from the deployed environment. Microbial fuel cells (MFC) are one class of such sources that produce usable energy from small organic compounds (i.e. sugars, alcohols, organic acids, and biopolymers) which can be easily containerized or scavenged from the environment. The use of microorganisms as the anodic catalysts is what makes these systems unique from other biofuel cell designs. One of the main drawbacks of engineering a sensor system powered by an MFC is that power densities and current flux are extremely low in currently reported systems. The power density is limited by the mass transfer of the fuel source to the catalyst, the metabolism of the microbial catalysts and the electron transfer from the organism to the anode. This presentation will focus on the development of a new style of microbially-modified anodes which will increase power density to a level where a practical power source can be engineered. This is being achieved by developing a three dimensional matrix as an artificial, conductive biofilm. These artificial biofilms will allow the capture of a consortium of microbes designed for efficient metabolism of the available fuel source. Also it will keep the microbes close to the electrode allowing ready access by fuel and providing a low resistance passage of the liberated electrons from fuel oxidation.

  9. Quercetin-Based Modified Porous Silicon Nanoparticles for Enhanced Inhibition of Doxorubicin-Resistant Cancer Cells.

    PubMed

    Liu, Zehua; Balasubramanian, Vimalkumar; Bhat, Chinmay; Vahermo, Mikko; Mäkilä, Ermei; Kemell, Marianna; Fontana, Flavia; Janoniene, Agne; Petrikaite, Vilma; Salonen, Jarno; Yli-Kauhaluoma, Jari; Hirvonen, Jouni; Zhang, Hongbo; Santos, Hélder A

    2017-02-01

    One of the most challenging obstacles in nanoparticle's surface modification is to achieve the concept that one ligand can accomplish multiple purposes. Upon such consideration, 3-aminopropoxy-linked quercetin (AmQu), a derivative of a natural flavonoid inspired by the structure of dopamine, is designed and subsequently used to modify the surface of thermally hydrocarbonized porous silicon (PSi) nanoparticles. This nanosystem inherits several advanced properties in a single carrier, including promoted anticancer efficiency, multiple drug resistance (MDR) reversing, stimuli-responsive drug release, drug release monitoring, and enhanced particle-cell interactions. The anticancer drug doxorubicin (DOX) is efficiently loaded into this nanosystem and released in a pH-dependent manner. AmQu also effectively quenches the fluorescence of the loaded DOX, thereby allowing the use of the nanosystem for monitoring the intracellular drug release. Furthermore, a synergistic effect with the presence of AmQu is observed in both normal MCF-7 and DOX-resistant MCF-7 breast cancer cells. Due to the similar structure as dopamine, AmQu may facilitate both the interaction and internalization of PSi into the cells. Overall, this PSi-based platform exhibits remarkable superiority in both multifunctionality and anticancer efficiency, making this nanovector a promising system for anti-MDR cancer treatment.

  10. Sulfatase modifying factor 1 trafficking through the cells: from endoplasmic reticulum to the endoplasmic reticulum.

    PubMed

    Zito, Ester; Buono, Mario; Pepe, Stefano; Settembre, Carmine; Annunziata, Ida; Surace, Enrico Maria; Dierks, Thomas; Monti, Maria; Cozzolino, Marianna; Pucci, Piero; Ballabio, Andrea; Cosma, Maria Pia

    2007-05-16

    Sulfatase modifying factor 1 (SUMF1) is the gene mutated in multiple sulfatase deficiency (MSD) that encodes the formylglycine-generating enzyme, an essential activator of all the sulfatases. SUMF1 is a glycosylated enzyme that is resident in the endoplasmic reticulum (ER), although it is also secreted. Here, we demonstrate that upon secretion, SUMF1 can be taken up from the medium by several cell lines. Furthermore, the in vivo engineering of mice liver to produce SUMF1 shows its secretion into the blood serum and its uptake into different tissues. Additionally, we show that non-glycosylated forms of SUMF1 can still be secreted, while only the glycosylated SUMF1 enters cells, via a receptor-mediated mechanism. Surprisingly, following its uptake, SUMF1 shuttles from the plasma membrane to the ER, a route that has to date only been well characterized for some of the toxins. Remarkably, once taken up and relocalized into the ER, SUMF1 is still active, enhancing the sulfatase activities in both cultured cells and mice tissues.

  11. Effect of microinjected N-ethylmaleimide-modified heavy meromyosin on cell division in amphibian eggs

    PubMed Central

    1980-01-01

    N-Ethylmaleimide-modified heavy meromyosin (NEM-HMM) microinjected into amphibian eggs inhibits cytokinesis and the cortical contractions associated with wound closure. Injection of NEM-HMM into two-cell Rana pipiens embryos produces a zone of cleavage inhibition around the point of injection. Early furrows followed by time-lapse microcinematography are seen to slow and stop as they enter the NEM-HMM-injected zone. Arrested furrows slowly regress, leaving a large region of cytoplasm uncleaved. Few nuclei are found in these regions of cleavage inhibition. Wound closure is often inhibited by NEM-HMM, especially when this inhibitor is injected just beneath the egg cortex. We observe that the surface of an unfertilized Rana egg is covered with microvilli that disappear during the course of development. The surfaces of NEM- HMM-inhibited zones remain covered with microvilli and resemble the unfertilized egg surface. PMID:6997323

  12. Cell proliferation on UV-excimer lamp modified and grafted polytetrafluoroethylene

    NASA Astrophysics Data System (ADS)

    Švorčík, V.; Ročková, K.; Ratajová, E.; Heitz, J.; Huber, N.; Bäuerle, D.; Bačáková, L.; Dvořánková, B.; Hnatowicz, V.

    2004-04-01

    Polytetrafluoroethylene (PTFE) was modified by irradiation with 172 nm light from an UV-excimer lamp in reactive ammonia atmosphere. Then, the samples were exposed to aqueous solutions of amino acids (glycine, alanine and leucine). The samples were characterized by water contact angle measurement, UV-VIS spectroscopy and laser-induced fluorescence (LIF). The results indicate, that the amino acids are bound onto the polymer chain. The amount of incorporated amino acid is a decreasing function of its molecular size. By in vitro experiments it was shown that the incorporated amino acids support adhesion and proliferation of rat aortic smooth muscle cells and mouse embryonic 3T3 fibroblasts on the polymer surface. The maximum effect was observed for smallest molecule of glycine.

  13. Dry eye modifies the thermal and menthol responses in rat corneal primary afferent cool cells.

    PubMed

    Kurose, Masayuki; Meng, Ian D

    2013-07-01

    Dry eye syndrome is a painful condition caused by inadequate or altered tear film on the ocular surface. Primary afferent cool cells innervating the cornea regulate the ocular fluid status by increasing reflex tearing in response to evaporative cooling and hyperosmicity. It has been proposed that activation of corneal cool cells via a transient receptor potential melastatin 8 (TRPM8) channel agonist may represent a potential therapeutic intervention to treat dry eye. This study examined the effect of dry eye on the response properties of corneal cool cells and the ability of the TRPM8 agonist menthol to modify these properties. A unilateral dry eye condition was created in rats by removing the left lacrimal gland. Lacrimal gland removal reduced tears in the dry eye to 35% compared with the contralateral eye and increased the number of spontaneous blinks in the dry eye by over 300%. Extracellular single-unit recordings were performed 8-10 wk following surgery in the trigeminal ganglion of dry eye animals and age-matched controls. Responses of corneal cool cells to cooling were examined after the application of menthol (10 μM-1.0 mM) to the ocular surface. The peak frequency of discharge to cooling was higher and the cooling threshold was warmer in dry eye animals compared with controls. The dry condition also altered the neuronal sensitivity to menthol, causing desensitization to cold-evoked responses at concentrations that produced facilitation in control animals. The menthol-induced desensitization of corneal cool cells would likely result in reduced tearing, a deleterious effect in individuals with dry eye.

  14. Glucose and sucrose differentially modify cell proliferation in maize during germination.

    PubMed

    Lara-Núñez, Aurora; García-Ayala, Brendy B; Garza-Aguilar, Sara M; Flores-Sánchez, Jesús; Sánchez-Camargo, Victor A; Bravo-Alberto, Carlos E; Vázquez-Santana, Sonia; Vázquez-Ramos, Jorge M

    2017-04-01

    Glucose and sucrose play a dual role: as carbon and energy sources and as signaling molecules. In order to address the impact that sugars may have on maize seeds during germination, embryo axes were incubated with or without either of the two sugars. Expression of key cell cycle markers and protein abundance, cell patterning and de novo DNA synthesis in root meristem zones were analyzed. Embryo axes without added sugars in imbibition medium were unable to grow after 7 days; in sucrose, embryo axes developed seminal and primary roots with numerous root hairs, whereas in glucose axes showed a twisted morphology, no root hair formation but callus-like structures on adventitious and primary seminal roots. More and smaller cells were observed with glucose treatment in root apical meristems. de novo DNA synthesis was stimulated more by glucose than by sucrose. At 24 h of imbibition, expression of ZmCycD2;2a and ZmCycD4;2 was increased by sucrose and reduced by glucose. CDKA1;1 and CDKA2;1 expression was stimulated equally by both sugars. Protein abundance patterns were modified by sugars: ZmCycD2 showed peaks on glucose at 12 and 36 h of imbibition whereas sucrose promoted ZmCycD3 protein accumulation. In presence of glucose ZmCycD3, ZmCycD4 and ZmCycD6 protein abundance was reduced after 24 h. Finally, both sugars stimulated ZmCDKA protein accumulation but at different times. Overall, even though glucose appears to act as a stronger mitogen stimulator, sucrose stimulated the expression of more cell cycle markers during germination. This work provides evidence of a differential response of cell cycle markers to sucrose and glucose during maize germination that may affect the developmental program during plantlet establishment.

  15. RGD peptide-modified multifunctional dendrimer platform for drug encapsulation and targeted inhibition of cancer cells.

    PubMed

    He, Xuedan; Alves, Carla S; Oliveira, Nilsa; Rodrigues, João; Zhu, Jingyi; Bányai, István; Tomás, Helena; Shi, Xiangyang

    2015-01-01

    Development of multifunctional nanoscale drug-delivery systems for targeted cancer therapy still remains a great challenge. Here, we report the synthesis of cyclic arginine-glycine-aspartic acid (RGD) peptide-conjugated generation 5 (G5) poly(amidoamine) dendrimers for anticancer drug encapsulation and targeted therapy of cancer cells overexpressing αvβ3 integrins. In this study, amine-terminated G5 dendrimers were used as a platform to be sequentially modified with fluorescein isothiocyanate (FI) via a thiourea linkage and RGD peptide via a polyethylene glycol (PEG) spacer, followed by acetylation of the remaining dendrimer terminal amines. The developed multifunctional dendrimer platform (G5.NHAc-FI-PEG-RGD) was then used to encapsulate an anticancer drug doxorubicin (DOX). We show that approximately six DOX molecules are able to be encapsulated within each dendrimer platform. The formed complexes are water-soluble, stable, and able to release DOX in a sustained manner. One- and two-dimensional NMR techniques were applied to investigate the interaction between dendrimers and DOX, and the impact of the environmental pH on the release rate of DOX from the dendrimer/DOX complexes was also explored. Furthermore, cell biological studies demonstrate that the encapsulation of DOX within the G5.NHAc-FI-PEG-RGD dendrimers does not compromise the anticancer activity of DOX and that the therapeutic efficacy of the dendrimer/DOX complexes is solely related to the encapsulated DOX drug. Importantly, thanks to the role played by RGD-mediated targeting, the developed dendrimer/drug complexes are able to specifically target αvβ3 integrin-overexpressing cancer cells and display specific therapeutic efficacy to the target cells. The developed RGD peptide-targeted multifunctional dendrimers may thus be used as a versatile platform for targeted therapy of different types of αvβ3 integrin-overexpressing cancer cells.

  16. Specific growth inhibition of ErbB2‑expressing human breast cancer cells by genetically modified NK‑92 cells.

    PubMed

    Liu, Hui; Yang, Bo; Sun, Tingting; Lin, Lin; Hu, Yi; Deng, Muhong; Yang, Junlan; Liu, Tianyi; Li, Jinyu; Sun, Shengjie; Jiao, Shunchang

    2015-01-01

    The natural killer cell line NK‑92 shows great cytotoxicity against various types of cancer. Several types of solid tumor cells, however, can effectively resist NK-mediated lysis by interaction of major histocompatibility complex (MHC) molecules with NK cell inhibitory receptors. To generate a eukaryotic expression vector encoding chimeric antigen receptor scFv anti-erbB2-CD28-ζ and to investigate the expression and action of this chimeric antigen receptor in cancer cells both in vitro and in vivo, NK‑92 cells were genetically modified with an scFv anti-erbB2-CD28-ζ chimeric recep-tor by optimized electro-poration using the Amaxa Nucleofector system. The expression of the chimeric receptor was evaluated by RT-PCR and immunofluorescence. The ability of the genetically modified NK‑92 cells to induce cell death in tumor targets was assessed in vitro and in vivo. The transduced NK‑92-anti-erbB2 scFv-CD28-ζ cells expressing high levels of the fusion protein on the cell surface were analyzed by fluorescence-activated cell-sorting (FACS) analysis. These cells specifically enhanced the cell death of the erbB2‑expressing human breast cancer cell lines MDA-MB-453 and SKBr3. Furthermore, adoptive transfer of genetically modified NK‑92 cells specifically reduced tumor size and lung metastasis of nude mice bearing established MDA-MB-453 cells, and significantly enhanced the survival period of these mice. The genetically modified NK‑92 cells significantly enhanced the killing of erbB2‑expressing cancer and may be a novel therapeutic strategy for erbB2‑expressing cancer cells.

  17. Immunohistochemical analysis of IA-2 family of protein tyrosine phosphatases in rat gastrointestinal endocrine cells.

    PubMed

    Gomi, Hiroshi; Kubota-Murata, Chisato; Yasui, Tadashi; Tsukise, Azuma; Torii, Seiji

    2013-02-01

    Islet-associated protein-2 (IA-2) and IA-2β (also known as phogrin) are unique neuroendocrine-specific protein tyrosine phosphatases (PTPs). The IA-2 family of PTPs was originally identified from insulinoma cells and discovered to be major autoantigens in type 1 diabetes. Despite its expression in the neural and canonical endocrine tissues, data on expression of the IA-2 family of PTPs in gastrointestinal endocrine cells (GECs) are limited. Therefore, we immunohistochemically investigated the expression of the IA-2 family of PTPs in the rat gastrointestinal tract. In the stomach, IA-2 and IA-2β were expressed in GECs that secrete serotonin, somatostatin, and cholecystokinin/gastrin-1. In addition to these hormones, secretin, gastric inhibitory polypeptide (also known as the glucose-dependent insulinotropic peptide), glucagon-like peptide-1, and glucagon, but not ghrelin were coexpressed with IA-2 or IA-2β in duodenal GECs. Pancreatic islet cells that secrete gut hormones expressed the IA-2 family of PTPs. The expression patterns of IA-2 and IA-2β were comparable. These results reveal that the IA-2 family of PTPs is expressed in a cell type-specific manner in rat GECs. The extensive expression of the IA-2 family of PTPs in pancreo-gastrointestinal endocrine cells and in the enteric plexus suggests their systemic contribution to nutritional control through a neuroendocrine signaling network.

  18. Fibrin patch-based insulin-like growth factor-1 gene-modified stem cell transplantation repairs ischemic myocardium

    PubMed Central

    Li, Jun; Zhu, Kai; Yang, Shan; Wang, Yulin; Guo, Changfa; Yin, Kanhua; Wang, Chunsheng

    2015-01-01

    Bone marrow mesenchymal stem cells (BMSCs), tissue-engineered cardiac patch, and therapeutic gene have all been proposed as promising therapy strategies for cardiac repair after myocardial infarction. In our study, BMSCs were modified with insulin-like growth factor-1 (IGF-1) gene, loaded into a fibrin patch, and then transplanted into a porcine model of ischemia/reperfusion (I/R) myocardium injury. The results demonstrated that IGF-1 gene overexpression could promote proliferation of endothelial cells and cardiomyocyte-like differentiation of BMSCs in vitro. Four weeks after transplantation of fibrin patch loaded with gene-modified BMSCs, IGF-1 overexpression could successfully promote angiogenesis, inhibit remodeling, increase grafted cell survival and reduce apoptosis. In conclusion, the integrated strategy, which combined fibrin patch with IGF-1 gene modified BMSCs, could promote the histological cardiac repair for a clinically relevant porcine model of I/R myocardium injury. PMID:25767192

  19. Up-up-down-down magnetic chain structure of the spin-1/2 tetragonally distorted spinel GeC u2O4

    NASA Astrophysics Data System (ADS)

    Zou, T.; Cai, Y.-Q.; dela Cruz, C. R.; Garlea, V. O.; Mahanti, S. D.; Cheng, J.-G.; Ke, X.

    2016-12-01

    GeC u2O4 spinel exhibits a tetragonal structure due to the strong Jahn-Teller distortion associated with C u2 + ions. We show that its magnetic structure can be described as slabs composed of a pair of layers with orthogonally oriented spin-1/2 Cu chains in the basal a b plane. The spins between the two layers within a slab are collinearly aligned while the spin directions of neighboring slabs are perpendicular to each other. Interestingly, we find that spins along each chain form an unusual up-up-down-down (UUDD) pattern, suggesting a non-negligible nearest-neighbor biquadratic exchange interaction in the effective classical spin Hamiltonian. We hypothesize that spin-orbit coupling and orbital mixing of C u2 + ions in this system are non-negligible, which calls for future calculations using perturbation theory with extended Hilbert (spin and orbital) space and calculations based on density functional theory including spin-orbit coupling and looking at the global stability of the UUDD state.

  20. Bone marrow mesenchymal stem cells are an attractive donor cell type for production of cloned pigs as well as genetically modified cloned pigs by somatic cell nuclear transfer.

    PubMed

    Li, Zicong; He, Xiaoyan; Chen, Liwen; Shi, Junsong; Zhou, Rong; Xu, Weihua; Liu, Dewu; Wu, Zhenfang

    2013-10-01

    The somatic cell nuclear transfer (SCNT) technique has been widely applied to clone pigs or to produce genetically modified pigs. Currently, this technique relies mainly on using terminally differentiated fibroblasts as donor cells. To improve cloning efficiency, only partially differentiated multipotent mesenchymal stem cells (MSCs), thought to be more easily reprogrammed to a pluripotent state, have been used as nuclear donors in pig SCNT. Although in vitro-cultured embryos cloned from porcine MSCs (MSCs-embryos) were shown to have higher preimplantation developmental ability than cloned embryos reconstructed from fibroblasts (Fs-embryos), the difference in in vivo full-term developmental rate between porcine MSCs-embryos and Fs-embryos has not been investigated so far. In this study, we demonstrated that blastocyst total cell number and full-term survival abilities of MSCs-embryos were significantly higher than those of Fs-embryos cloned from the same donor pig. The enhanced developmental potential of MSCs-embryos may be associated with their nuclear donors' DNA methylation profile, because we found that the methylation level of imprinting genes and repeat sequences differed between MSCs and fibroblasts. In addition, we showed that use of transgenic porcine MSCs generated from transgene plasmid transfection as donor cells for SCNT can produce live transgenic cloned pigs. These results strongly suggest that porcine bone marrow MSCs are a desirable donor cell type for production of cloned pigs and genetically modified cloned pigs via SCNT.

  1. Inhibition of NF-kappaB with Dehydroxymethylepoxyquinomicin modifies the function of human peritoneal mesothelial cells

    PubMed Central

    Sosińska, Patrycja; Baum, Ewa; Maćkowiak, Beata; Staniszewski, Ryszard; Jasinski, Tomasz; Umezawa, Kazuo; Bręborowicz, Andrzej

    2016-01-01

    Peritoneal mesothelial cells exposed to bioincompatible dialysis fluids contribute to damage of the peritoneum during chronic dialysis. Inflammatory response triggered in the mesothelium leading to neovascularization and fibrosis plays an important role in that process. We studied the effects of Dehydroxymethyepoxyquinmicin (DHMEQ)-an NF-κB inhibitor on function of human peritoneal mesothelial cells (HPMC) in in vitro culture. DHMEQ studied in concentrations of 1-10 µg/ml was not toxic to HPMC. Synthesis of IL-6, MCP-1 and hyaluronan in unstimulated and stimulated with interleukin-1 (100 pg/ml) HPMC was inhibited in the presence of DHMEQ and the effect was proportional to the dose of the drug. DHMEQ (10 µg/ml) reduced in unstimulated HPMC synthesis of IL-6 (-55%), MCP-1 (-58%) and hyaluronan (-41%). Respective values for stimulated HMPC were: -63% for IL-6, -57% for MCP-1 and -67% for hyaluronan. The observed effects were due to the suppression of the expression of genes responsible for the synthesis of these molecules. DHMEQ modified the effects of the effluent dialysates from CAPD patients on the function of HMPC. Dialysate induced accelerated growth of these cells, and synthesis of collagen was inhibited in the presence of DHMEQ 10 µg/ml, by 69% and 40%, respectively. The results of our study show that DHMEQ effectively reduces inflammatory response in HMPC and prevents excessive dialysate induced proliferation and collagen synthesis in these cells. All of these effects may be beneficial during chronic peritoneal dialysis and prevents progressive dialysis-induced damage to the peritoneum. PMID:28078047

  2. Retention Curve Measurement for Sands Using a TDR-based Long Column and Modified Tempe Cell

    NASA Astrophysics Data System (ADS)

    Sakaki, T.; Illangasekare, T. H.

    2006-12-01

    Long column and Tempe cell are typical methods for measuring the water retention curves for soils. In the conventional long column method utilizing a stack of rings, water saturation profile is determined gravimetrically. X-ray or gamma ray attenuation are non-destructive methods but require complex and expensive devices and involve the use of photon sources. Time domain reflectometry (TDR) is an alternative to these radioactive methods for measuring water content profile along the column. Typical Tempe cells have a sample height of 3 to 6 cm. Suction is applied to the sample to induce drainage and monitored outflow is used to calculate the average water saturation of the sample, which potentially leads to obscuring the distinct displacement pressure value and results in a smoothed retention curve. In this study, we assumed that direct point-wise measurements provide retention curves that represent the physical behavior of the porous medium. We first determined retention curves for a number of well-sorted industrial silica sands using a long column that allows such point-wise measurements by TDR probes horizontally installed at eleven different elevations. Then, we modified a commercially available Tempe cell so that water saturation and capillary pressure head at a physical point in the cell, as well as the conventional height-averaged water saturation, can be measured simultaneously. Comparison of the conventional and point- measured retention curves that were obtained simultaneously for the identical sand samples revealed that 1) point-measured retention curves were identical to the ones measured in the long column, 2) the artifact of using height-averaged saturation values as pointed out by Dane et al. [1992] was experimentally confirmed. We further show that the displacement pressure head can possibly be underestimated especially for coarse soils when height-averaged water saturation is used. This is more significant for oil-water and DNAPL-water systems where

  3. Biological Behavior of Osteoblast Cell and Apatite Forming Ability of the Surface Modified Ti Alloys.

    PubMed

    Zhao, Jingming; Hwang, K H; Choi, W S; Shin, S J; Lee, J K

    2016-02-01

    Titanium as one kind of biomaterials comes in direct contact with the body, making evaluation of biocompatibility an important aspect to biomaterials development. Surface chemistry of titanium plays an important role in osseointegration. Different surface modification alters the surface chemistry and result in different biological response. In this study, three kinds of mixed acid solutions were used to treat Ti specimens to induce Ca-P formation. Following a strong mixed acid activation process, Ca-P coating successfully formed on the Ti surfaces in simulated body fluid. Strong mixed acid increased the roughness of the metal surface, because the porous and rough surface allows better adhesion between Ca-P coatings and substrates. After modification of titanium surface by mixed acidic solution and subsequently H2O2/HCL treatment evaluation of biocompatibility was conducted from hydroxyapatite formation by biomimetic process and cell viability on modified titanium surface. Nano-scale modification of titanium surfaces can alter cellular and tissue responses, which may benefit osseointegration and dental implant therapy. Results from this study indicated that surface treatment methods affect the surface morphology, type of TiO2 layer formed and subsequent apatite deposition and biological responses. The thermo scientific alamarblue cell viability assay reagent is used to quantitatively measure the viability of mammalian cell lines, bacteria and fungi by incorporating a rapid, sensitive and reliable fluorometric/colorimetric growth indicator, without any toxic and side effect to cell line. In addition, mixed acid treatment uses a lower temperature and shorter time period than widely used alkali treatment.

  4. The Role of Genetically Modified Mesenchymal Stem Cells in Urinary Bladder Regeneration

    PubMed Central

    Fuller, Natalie J.; Hannick, Jessica H.; Ahmad, Nida; Sharma, Arun K.

    2015-01-01

    Recent studies have demonstrated that mesenchymal stem cells (MSCs) combined with CD34+ hematopoietic/stem progenitor cells (HSPCs) can function as surrogate urinary bladder cells to synergistically promote multi-faceted bladder tissue regeneration. However, the molecular pathways governing these events are unknown. The pleiotropic effects of Wnt5a and Cyr61 are known to affect aspects of hematopoiesis, angiogenesis, and muscle and nerve regeneration. Within this study, the effects of Cyr61 and Wnt5a on bladder tissue regeneration were evaluated by grafting scaffolds containing modified human bone marrow derived MSCs. These cell lines were engineered to independently over-express Wnt5a or Cyr61, or to exhibit reduced expression of Cyr61 within the context of a nude rat bladder augmentation model. At 4 weeks post-surgery, data demonstrated increased vessel number (~250 vs ~109 vessels/mm2) and bladder smooth muscle content (~42% vs ~36%) in Cyr61OX (over-expressing) vs Cyr61KD (knock-down) groups. Muscle content decreased to ~25% at 10 weeks in Cyr61KD groups. Wnt5aOX resulted in high numbers of vessels and muscle content (~206 vessels/mm2 and ~51%, respectively) at 4 weeks. Over-expressing cell constructs resulted in peripheral nerve regeneration while Cyr61KD animals were devoid of peripheral nerve regeneration at 4 weeks. At 10 weeks post-grafting, peripheral nerve regeneration was at a minimal level for both Cyr61OX and Wnt5aOX cell lines. Blood vessel and bladder functionality were evident at both time-points in all animals. Results from this study indicate that MSC-based Cyr61OX and Wnt5aOX cell lines play pivotal roles with regards to increasing the levels of functional vasculature, influencing muscle regeneration, and the regeneration of peripheral nerves in a model of bladder augmentation. Wnt5aOX constructs closely approximated the outcomes previously observed with the co-transplantation of MSCs with CD34+ HSPCs and may be specifically targeted as an

  5. Chemically and compositionally modified solid solution disordered multiphase nickel hydroxide positive electrode for alkaline rechargeable electrochemical cells

    DOEpatents

    Ovshinsky, Stanford R.; Corrigan, Dennis; Venkatesan, Srini; Young, Rosa; Fierro, Christian; Fetcenko, Michael A.

    1994-01-01

    A high capacity, long cycle life positive electrode for use in an alkaline rechargeable electrochemical cell comprising: a solid solution nickel hydroxide material having a multiphase structure that comprises at least one polycrystalline .gamma.-phase including a polycrystalline .gamma.-phase unit cell comprising spacedly disposed plates with at least one chemical modifier incorporated around the plates, the plates having a range of stable intersheet distances corresponding to a 2.sup.+ oxidation state and a 3.5.sup.+, or greater, oxidation state; and at least one compositional modifier incorporated into the solid solution nickel hydroxide material to promote the multiphase structure.

  6. Cell separation by immunoaffinity partitioning with polyethylene glycol-modified Protein A in aqueous polymer two-phase systems

    NASA Technical Reports Server (NTRS)

    Karr, Laurel J.; Van Alstine, James M.; Snyder, Robert S.; Shafer, Steven G.; Harris, J. Milton

    1988-01-01

    Previous work has shown that polyethylene glycol (PEG)-bound antibodies can be used as affinity ligands in PEG-dextran two-phase systems to provide selective partitioning of cells to the PEG-rich phase. In the present work it is shown that immunoaffinity partitioning can be simplified by use of PEG-modified Protein A which complexes with unmodified antibody and cells and shifts their partitioning into the PEG-rich phase, thus eliminating the need to prepare a PEG-modified antibody for each cell type. In addition, the paper provides a more rigorous test of the original technique with PEG-bound antibodies by showing that it is effective at shifting the partitioning of either cell type of a mixture of two cell populations.

  7. Transferrin surface-modified PLGA nanoparticles-mediated delivery of a proteasome inhibitor to human pancreatic cancer cells.

    PubMed

    Frasco, Manuela F; Almeida, Gabriela M; Santos-Silva, Filipe; Pereira, Maria do Carmo; Coelho, Manuel A N

    2015-04-01

    The aim of this study was to develop a drug delivery system based on poly(lactic-co-glycolic acid) (PLGA) nanoparticles for an efficient and targeted action of the proteasome inhibitor bortezomib against pancreatic cancer cells. The PLGA nanoparticles were formulated with a poloxamer, and further surface-modified with transferrin for tumor targeting. The nanoparticles were characterized as polymer carriers of bortezomib, and the cellular uptake and growth inhibitory effects were evaluated in pancreatic cells. Cellular internalization of nanoparticles was observed in normal and cancer cells, but with higher uptake by cancer cells. The sustained release of the loaded bortezomib from PLGA nanoparticles showed cytotoxic effects against pancreatic normal and cancer cells. Noteworthy differential cytotoxicity was attained by transferrin surface-modified PLGA nanoparticles since significant cell growth inhibition by delivered bortezomib was only observed in cancer cells. These findings demonstrate that the ligand transferrin enhanced the targeted delivery of bortezomib-loaded PLGA nanoparticles to pancreatic cancer cells. These in vitro results highlight the transferrin surface-modified PLGA nanoparticles as a promising system for targeted delivery of anticancer drugs.

  8. Modified bacterial cellulose scaffolds for localized doxorubicin release in human colorectal HT-29 cells.

    PubMed

    Cacicedo, Maximiliano L; León, Ignacio E; Gonzalez, Jimena S; Porto, Luismar M; Alvarez, Vera A; Castro, Guillermo R

    2016-04-01

    Bacterial cellulose (BC) films modified by the in situ method with the addition of alginate (Alg) during the microbial cultivation of Gluconacetobacter hansenii under static conditions increased the loading of doxorubicin by at least three times. Biophysical analysis of BC-Alg films by scanning electron microscopy, thermogravimetry, X-ray diffraction and FTIR showed a highly homogeneous interpenetrated network scaffold without changes in the BC crystalline structure but with an increased amorphous phase. The main molecular interactions determined by FTIR between both biopolymers clearly suggest high compatibility. These results indicate that alginate plays a key role in the biophysical properties of the hybrid BC matrix. BC-Alg scaffold analysis by nitrogen adsorption isotherms revealed by the Brunauer-Emmett-Teller (BET) method an increase in surface area of about 84% and in pore volume of more than 200%. The Barrett-Joyner-Halenda (BJH) model also showed an increase of about 25% in the pore size compared to the BC film. Loading BC-Alg scaffolds with different amounts of doxorubicin decreased the cell viability of HT-29 human colorectal adenocarcinoma cell line compared to the free Dox from around 95-53% after 24h and from 63% to 37% after 48 h. Dox kinetic release from the BC-Alg nanocomposite displayed hyperbolic curves related to the different amounts of drug payload and was stable for at least 14 days. The results of the BC-Alg nanocomposites show a promissory potential for anticancer therapies of solid tumors.

  9. Zinc and zinc chelators modify taurine transport in rat retinal cells.

    PubMed

    Márquez, Asarí; Urbina, Mary; Lima, Lucimey

    2014-11-01

    Zinc regulates Na(+)/Cl(-)-dependent transporters, similar to taurine one, such as those for dopamine, serotonin and norepinephrine. This study examined the ex vivo effect of zinc (ZnSO4), N,N,N,N-tetraquis-(2-piridilmetil)etilendiamino (TPEN) and diethylenetriaminepenta-acetic acid (DTPA), intracellular and extracellular zinc chelators, respectively, on rat retina [(3)H]taurine transport. Isolated cells were incubated in Locke solution with 100 nM of [(3)H]taurine for 25 s. Different concentrations of ZnSO4 (0.5-200 μM) were used. Low concentrations of ZnSO4 (30 and 40 μM) increased the transport, while higher concentrations (100, 150 and 200 μM) decreased it. Various concentrations of TPEN (1-200 μM) were added. Intermediate concentrations of TPEN (10-60 μM) significantly decreased [(3)H]taurine transport. The presence of TPEN, 20 μM, plus ZnSO4 reversed the effect of TPEN alone. Several concentrations of DTPA (1-500 μM) were also investigated. Reduction of transport took place at high concentrations of the chelator (100, 250 and 500 μM). DTPA, 500 μM, plus ZnSO4, did not modify the effect of it. These results indicate that zinc modulates taurine transport in a concentration-dependent manner, directly acting on the transporter or by forming taurine-zinc complexes in cell membranes.

  10. Miraculin, a taste-modifying protein is secreted into intercellular spaces in plant cells.

    PubMed

    Hirai, Tadayoshi; Sato, Mayuko; Toyooka, Kiminari; Sun, Hyeon-Jin; Yano, Megumu; Ezura, Hiroshi

    2010-02-15

    A taste-modifying protein, miraculin, is highly accumulated in ripe fruit of miracle fruit (Richadella dulcifica) and the content can reach up to 10% of the total soluble protein in these fruits. Although speculated for decades that miraculin is secreted into intercellular spaces in miracle fruit, no evidence exists of its cellular localization. To study the cellular localization of miraculin in plant cells, using miracle fruit and transgenic tomato that constitutively express miraculin, immunoelectron microscopy, imaging GFP fusion proteins, and immunological detection of secreted proteins in culture medium of transgenic tomato were carried out. Immunoelectron microscopy showed the specific accumulation of miraculin in the intercellular layers of both miracle fruit and transgenic tomato. Imaging GFP fusion protein demonstrated that the miraculin-GFP fusion protein was accumulated in the intercellular spaces of tomato epidermal cells. Immunological detection of secreted proteins in culture medium of transgenic tomato indicated that miraculin was secreted from the roots of transgenic tomato expressing miraculin. This study firstly showed the evidences of the intercellular localization of miraculin, and provided a new insight of biological roles of miraculin in plants.

  11. Drug diffusion from disperse systems with a hydrophobically modified polysaccharide: Enhancer vs Franz cells.

    PubMed

    Lucero, María Jesús; Claro, Carmen; Casas, Marta; Jiménez-Castellanos, María Rosa

    2013-01-30

    This study assesses the capacity of a new hydrophobically modified polysaccharide -hydroxypropyl cellulose-methyl methacrylate - to control drug release in semisolid formulations. The dispersed systems contain the new polymer, Igepal CO520 as surfactant and theophylline as model drug at three concentrations (0.5, 1 and 1.5%, w/w). Drug release study shows that the systems containing 0.5% (w/w) of drug have faster release and higher diffusion coefficient than the other two concentrations. These results can be explained by two different structures ("relaxed" and "structured") found from a rheological point of view. Also, this paper compares two different devices for testing drug release and diffusion. It has been obtained more reliable and reproducible results with Enhancer Cell respect to Franz diffusion cell. In both cases, Fickian diffusion was the mechanism predominant for all systems. Finally, the utility of this polymer has been demonstrated to make three-dimensional gel structure and control theophylline release from systems in topical application.

  12. A new synthesis route for Os-complex modified redox polymers for potential biofuel cell applications.

    PubMed

    Pöller, Sascha; Beyl, Yvonne; Vivekananthan, Jeevanthi; Guschin, Dmitrii A; Schuhmann, Wolfgang

    2012-10-01

    A new synthesis route for Os-complex modified redox polymers was developed. Instead of ligand exchange reactions for coordinative binding of suitable precursor Os-complexes at the polymer, Os-complexes already exhibiting the final ligand shell containing a suitable functional group were bound to the polymer via an epoxide opening reaction. By separation of the polymer synthesis from the ligand exchange reaction at the Os-complex, the modification of the same polymer backbone with different Os-complexes or the binding of the same Os-complex to a number of different polymer backbones becomes feasible. In addition, the Os-complex can be purified and characterized prior to its binding to the polymer. In order to further understand and optimize suitable enzyme/redox polymer systems concerning their potential application in biosensors or biofuel cells, a series of redox polymers was synthesized and used as immobilization matrix for Trametes hirsuta laccase. The properties of the obtained biofuel cell cathodes were compared with similar biocatalytic interfaces derived from redox polymers obtained via ligand exchange reaction of the parent Os-complex with a ligand integrated into the polymer backbone during the polymer synthesis.

  13. Predicting Cell Association of Surface-Modified Nanoparticles Using Protein Corona Structure - Activity Relationships (PCSAR).

    PubMed

    Kamath, Padmaja; Fernandez, Alberto; Giralt, Francesc; Rallo, Robert

    2015-01-01

    Nanoparticles are likely to interact in real-case application scenarios with mixtures of proteins and biomolecules that will absorb onto their surface forming the so-called protein corona. Information related to the composition of the protein corona and net cell association was collected from literature for a library of surface-modified gold and silver nanoparticles. For each protein in the corona, sequence information was extracted and used to calculate physicochemical properties and statistical descriptors. Data cleaning and preprocessing techniques including statistical analysis and feature selection methods were applied to remove highly correlated, redundant and non-significant features. A weighting technique was applied to construct specific signatures that represent the corona composition for each nanoparticle. Using this basic set of protein descriptors, a new Protein Corona Structure-Activity Relationship (PCSAR) that relates net cell association with the physicochemical descriptors of the proteins that form the corona was developed and validated. The features that resulted from the feature selection were in line with already published literature, and the computational model constructed on these features had a good accuracy (R(2)LOO=0.76 and R(2)LMO(25%)=0.72) and stability, with the advantage that the fingerprints based on physicochemical descriptors were independent of the specific proteins that form the corona.

  14. Development of a Biomimetic Chondroitin Sulfate-modified Hydrogel to Enhance the Metastasis of Tumor Cells

    PubMed Central

    Liu, Yang; Wang, Shujun; Sun, Dongsheng; Liu, Yongdong; Liu, Yang; Wang, Yang; Liu, Chang; Wu, Hao; Lv, Yan; Ren, Ying; Guo, Xin; Sun, Guangwei; Ma, Xiaojun

    2016-01-01

    Tumor metastasis with resistance to anticancer therapies is the main cause of death in cancer patients. It is necessary to develop reliable tumor metastasis models that can closely recapitulate the pathophysiological features of the native tumor tissue. In this study, chondroitin sulfate (CS)-modified alginate hydrogel beads (ALG-CS) are developed to mimic the in vivo tumor microenvironment with an abnormally increased expression of CS for the promotion of tumor cell metastasis. The modification mechanism of CS on alginate hydrogel is due to the cross-linking between CS and alginate molecules via coordination of calcium ions, which enables ALG-CS to possess significantly different physical characteristics than the traditional alginate beads (ALG). And quantum chemistry calculations show that in addition to the traditional egg-box structure, novel asymmetric egg-box-like structures based on the interaction between these two kinds of polymers are also formed within ALG-CS. Moreover, tumor cell metastasis is significantly enhanced in ALG-CS compared with that in ALG, as confirmed by the increased expression of MMP genes and proteins and greater in vitro invasion ability. Therefore, ALG-CS could be a convenient and effective 3D biomimetic scaffold that would be used to construct standardized tumor metastasis models for tumor research and anticancer drug screening. PMID:27432752

  15. 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

  16. A highly efficient modified human serum albumin signal peptide to secrete proteins in cells derived from different mammalian species.

    PubMed

    Attallah, Carolina; Etcheverrigaray, Marina; Kratje, Ricardo; Oggero, Marcos

    2017-01-10

    Signal peptides (SPs) are key elements in the production of recombinant proteins; however, little information is available concerning different SP in mammalian cells other than CHO. In order to study the efficiency of different SPs to direct the traffic along the secretory pathway of the green fluorescence protein (GFP) and a scFv-Fc fusion protein; CHO-K1, HEK293 and NS0 cell lines were transfected in a transient and stable way. SP of human azurocidin (AZ), modified human albumin (mSA), modified Cricetulus griseus Ig kappa chain V III region MOPC 63 like (mIgκ C) and modified human Ig kappa chain V III region VG (mIgκ H) were evaluated. The efficiency of SPs to translocate a propeptide across the ER membrane was evaluated by fluorescence microscopy and flow cytometry for the GFP inside the secretory pathway, and by antigen-specific indirect ELISA for the scFv-Fc outside the cell. The mSA SP was successful in directing the secretion of the active proteins in these different types of mammalian cells, regardless of the transgene copy number. The goal of this work was to demonstrate that a modified version of SA SP might be used in different mammalian cells employing the same expression vector.

  17. Migration and proliferation of intact and genetically modified primordial germ cells and the generation of a transgenic chicken.

    PubMed

    Kim, Jin Nam; Park, Tae Sub; Park, Sang Hyun; Park, Kyung Je; Kim, Tae Min; Lee, Seul Ki; Lim, Jeong Mook; Han, Jae Yong

    2010-02-01

    This study evaluated gonadal migration and postmigratory proliferation of intact and genetically modified chicken primordial germ cells (PGCs). A randomized, controlled trial was conducted with the gonadal population of PGCs and transgenic chicken production as major parameters. PGCs (0, 90, 900, 1800, or 3000 cells) were transferred into 53-h-old embryos. The percentage of PGCs migrating on Day 6 of development was highest (35.8%) following the transfer of 900 PGCs and did not change with increases in transferred PGCs. The number of migrating PGCs gradually increased (P = 0.0001) as the number of transferred PGCs was increased. Gonadal migration was detected after the transfer of intact and genetically modified PGCs, but prominent decreases in PGC migration (from 21.9% to 0.38%) and chimera ratio (from 0.4 to 0.007) occurred with genetically modified PGCs. However, subsequent vigorous proliferation of the modified PGCs (3.67-fold increase from transferred number) led to the derivation of a germline chimera and produced a transgenic hatchling. In conclusion, the number of migrating PGCs increased as the number of transferred cells increased. Vigorous proliferation after transfer compensated for the decreased migration capacity of genetically modified PGCs and resulted in the production of a transgenic chicken.

  18. Comb-type grafted poly(N-isopropylacrylamide) gel modified surfaces for rapid detachment of cell sheet.

    PubMed

    Tang, Zhonglan; Akiyama, Yoshikatsu; Yamato, Masayuki; Okano, Teruo

    2010-10-01

    A comb-type grafted poly(N-isopropylacrylamide) (PIPAAm) gel modified surface was newly developed for providing a rapid cell sheet recovery for tissue engineering. PIPAAm macromonomer was prepared by the etherification reaction of the hydroxyl terminal moieties of PIPAAm with acryloyl chloride, followed by the radical telomerization reaction of N-isopropylacrylamide (IPAAm) monomer using 2-mercaptoethanol as a chain transfer agent. Solution containing IPAAm monomer and PIPAAm macromonomer was spread on the surface of tissue culture polystyrene (TCPS), and then the surface was subjected to electron beam irradiation for grafting the monomer and macromonomer on the surfaces, resulting in comb-type grafted PIPAAm gel modified TCPS (GG-TCPS). Besides the difference of the amount of the modified PIPAAm, no distinct difference was found between the properties of GG-TCPSs and normal-type PIPAAm gel modified TCPS (NG-TCPS) through XPS, AFM and a contact angle measurement. At 37 degrees C, bovine aortic endothelial cells (BAECs) were well adhered and spread on GG-TCPS as well as NG-TCPS regardless of the macromonomer concentration. By lowering temperature to 20 degrees C, BAECs detached themselves more rapidly from GG-TCPS compared with NG-TCPS. Upon lowering temperature, the grafted polymer was speculated to accelerate the hydration of modified PIPAAm gel, resulting in a rapid cell sheet detachment.

  19. Identification of Genes Mediating Drosophila Follicle Cell Progenitor Differentiation by Screening for Modifiers of GAL4::UAS Variegation

    PubMed Central

    Lee, Ming-Chia; Skora, Andrew D.; Spradling, Allan C.

    2016-01-01

    The Drosophila melanogaster ovarian follicle cell lineage provides a powerful system for investigating how epigenetic changes contribute to differentiation. Downstream from an epithelial stem cell, follicle progenitors undergo nine mitotic cell cycles before transitioning to the endocycle and initiating differentiation. During their proliferative phase, follicle progenitors experience Lsd1-dependent changes in epigenetic stability that can be monitored using GAL4::UAS variegation. Eventually, follicle progenitors acquire competence to respond to Delta, a Notch ligand present in the environment, which signals them to cease division and initiate differentiation. The time required to acquire competence determines the duration of mitotic cycling and hence the final number of follicle cells. We carried out a screen for dominant modifiers of variegation spanning nearly 70% of Drosophila euchromatin to identify new genes influencing follicle progenitor epigenetic maturation. The eight genes found include chromatin modifiers, but also cell cycle regulators and transcription factors. Five of the modifier genes accelerate the acquisition of progenitor competence and reduce follicle cell number, however, the other three genes affect follicle cell number in an unexpected manner. PMID:27866148

  20. Identification of Genes Mediating Drosophila Follicle Cell Progenitor Differentiation by Screening for Modifiers of GAL4::UAS Variegation.

    PubMed

    Lee, Ming-Chia; Skora, Andrew D; Spradling, Allan C

    2017-01-05

    The Drosophila melanogaster ovarian follicle cell lineage provides a powerful system for investigating how epigenetic changes contribute to differentiation. Downstream from an epithelial stem cell, follicle progenitors undergo nine mitotic cell cycles before transitioning to the endocycle and initiating differentiation. During their proliferative phase, follicle progenitors experience Lsd1-dependent changes in epigenetic stability that can be monitored using GAL4::UAS variegation. Eventually, follicle progenitors acquire competence to respond to Delta, a Notch ligand present in the environment, which signals them to cease division and initiate differentiation. The time required to acquire competence determines the duration of mitotic cycling and hence the final number of follicle cells. We carried out a screen for dominant modifiers of variegation spanning nearly 70% of Drosophila euchromatin to identify new genes influencing follicle progenitor epigenetic maturation. The eight genes found include chromatin modifiers, but also cell cycle regulators and transcription factors. Five of the modifier genes accelerate the acquisition of progenitor competence and reduce follicle cell number, however, the other three genes affect follicle cell number in an unexpected manner.

  1. Ranitidine modifies myeloid cell populations and inhibits breast tumor development and spread in mice

    PubMed Central

    Vila-Leahey, Ava; Oldford, Sharon A.; Marignani, Paola A.; Wang, Jun; Haidl, Ian D.; Marshall, Jean S.

    2016-01-01

    ABSTRACT Histamine receptor 2 (H2) antagonists are widely used clinically for the control of gastrointestinal symptoms, but also impact immune function. They have been reported to reduce tumor growth in established colon and lung cancer models. Histamine has also been reported to modify populations of myeloid-derived suppressor cells (MDSCs). We have examined the impact of the widely used H2 antagonist ranitidine, on both myeloid cell populations and tumor development and spread, in three distinct models of breast cancer that highlight different stages of cancer progression. Oral ranitidine treatment significantly decreased the monocytic MDSC population in the spleen and bone marrow both alone and in the context of an orthotopic breast tumor model. H2 antagonists ranitidine and famotidine, but not H1 or H4 antagonists, significantly inhibited lung metastasis in the 4T1 model. In the E0771 model, ranitidine decreased primary tumor growth while omeprazole treatment had no impact on tumor development. Gemcitabine treatment prevented the tumor growth inhibition associated with ranitidine treatment. In keeping with ranitidine-induced changes in myeloid cell populations in non-tumor-bearing mice, ranitidine also delayed the onset of spontaneous tumor development, and decreased the number of tumors that developed in LKB1−/−/NIC mice. These results indicate that ranitidine alters monocyte populations associated with MDSC activity, and subsequently impacts breast tumor development and outcome. Ranitidine has potential as an adjuvant therapy or preventative agent in breast cancer and provides a novel and safe approach to the long-term reduction of tumor-associated immune suppression. PMID:27622015

  2. microRNA-142 is upregulated by tumor necrosis factor-alpha and triggers apoptosis in human gingival epithelial cells by repressing BACH2 expression

    PubMed Central

    Li, Song; Song, Zhongchen; Dong, Jiachen; Shu, Rong

    2017-01-01

    Tumor necrosis factor-alpha (TNF-α) has been shown to cause apoptosis of gingival epithelial cells (GECs) in periodontitis. However, the underlying molecular mechanism is still unclear. In this study, we showed that miR-142 expression was significantly elevated in human GECs after exposure to TNF-α. Such induction was in a time- and concentration-dependent manner. Serum miR-142 levels were positively correlated with serum TNF-α levels in patients with chronic periodontitis (r = 0.314, P = 0.0152). Depletion of miR-142 was found to attenuate TNF-α-induced apoptosis, as determined by TUNEL staining and caspase-3 activity assays. In contrast, overexpression of miR-142 significantly reduced viability and induced apoptosis in GECs. Basic leucine zipper transcription factor 2 (BACH2) was identified to be a functional target of miR-142. Overexpression of miR-142 caused a 3-fold reduction of BACH2 protein in primary GECs. Overexpression of BACH2 significantly reversed miR-142- or TNF-α-induced apoptosis of GECs. Similar to the findings with miR-142 mimic, depletion of BACH2 significantly promoted apoptosis in GECs, which was accompanied by decreased expression of Bcl-2 and Bcl-xL and increased expression of Bax and Bim. Overall, miR-142 mediates TNF-α-induced apoptosis in gingival epithelial cells by targeting BACH2 and may represent a potential therapeutic target for periodontitis. PMID:28123644

  3. Patterned cell arrays and patterned co-cultures on polydopamine-modified poly(vinyl alcohol) hydrogels.

    PubMed

    Beckwith, Kai M; Sikorski, Pawel

    2013-12-01

    Live cell arrays are an emerging tool that expand traditional 2D in vitro cell culture, increasing experimental precision and throughput. A patterned cell system was developed by combining the cell-repellent properties of polyvinyl alcohol hydrogels with the cell adhesive properties of self-assembled films of dopamine (polydopamine). It was shown that polydopamine could be patterned onto spin-cast polyvinyl alcohol hydrogels by microcontact printing, which in turn effectively patterned the growth of several cell types (HeLa, human embryonic kidney, human umbilical vein endothelial cells (HUVEC) and prostate cancer). The cells could be patterned in geometries down to single-cell confinement, and it was demonstrated that cell patterns could be maintained for at least 3 weeks. Furthermore, polydopamine could be used to modify poly(vinyl alcohol) in situ using a cell-compatible deposition buffer (1 mg mL(-1) dopamine in 25 mM tris with a physiological salt balance). The treatment switched the PVA hydrogel from cell repellent to cell adhesive. Finally, by combining microcontact printing and in situ deposition of polydopamine, patterned co-cultures of the same cell type (HeLa/HeLa) and dissimilar cell types (HeLa/HUVEC) were realized through simple chemistry and could be studied over time. The combination of polyvinyl alcohol and polydopamine was shown to be an attractive route to versatile, patterned cell culture experiments with minimal infrastructure requirements and low complexity.

  4. The Role of Histone Protein Modifications and Mutations in Histone Modifiers in Pediatric B-Cell Progenitor Acute Lymphoblastic Leukemia

    PubMed Central

    Janczar, Szymon; Janczar, Karolina; Pastorczak, Agata; Harb, Hani; Paige, Adam J. W.; Zalewska-Szewczyk, Beata; Danilewicz, Marian; Mlynarski, Wojciech

    2017-01-01

    While cancer has been long recognized as a disease of the genome, the importance of epigenetic mechanisms in neoplasia was acknowledged more recently. The most active epigenetic marks are DNA methylation and histone protein modifications and they are involved in basic biological phenomena in every cell. Their role in tumorigenesis is stressed by recent unbiased large-scale studies providing evidence that several epigenetic modifiers are recurrently mutated or frequently dysregulated in multiple cancers. The interest in epigenetic marks is especially due to the fact that they are potentially reversible and thus druggable. In B-cell progenitor acute lymphoblastic leukemia (BCP-ALL) there is a relative paucity of reports on the role of histone protein modifications (acetylation, methylation, phosphorylation) as compared to acute myeloid leukemia, T-cell ALL, or other hematologic cancers, and in this setting chromatin modifications are relatively less well studied and reviewed than DNA methylation. In this paper, we discuss the biomarker associations and evidence for a driver role of dysregulated global and loci-specific histone marks, as well as mutations in epigenetic modifiers in BCP-ALL. Examples of chromatin modifiers recurrently mutated/disrupted in BCP-ALL and associated with disease outcomes include MLL1, CREBBP, NSD2, and SETD2. Altered histone marks and histone modifiers and readers may play a particular role in disease chemoresistance and relapse. We also suggest that epigenetic regulation of B-cell differentiation may have parallel roles in leukemogenesis. PMID:28054944

  5. Bioinspired phosphorylcholine-modified polyplexes as an effective strategy for selective uptake and transfection of cancer cells.

    PubMed

    Chen, Lina; Wang, Haibo; Zhang, Yuanfeng; Wang, Youxiang; Hu, Qiaoling; Ji, Jian

    2013-11-01

    We demonstrated here that the phosphorylcholine-modified polyplexes can be explored as effective gene vector for selective uptake and high transfection of cancer cells. 12-acryloyloxy dodecyl phosphorylcholine modified polyethyleneimine (PEI-ADPC) with grafting level about 13%, 8.3% and 4.5% was successfully synthesized. Gel retardation assay indicated that ADPC modification did not affect the DNA condensation ability. The PEI-ADPC13%/DNA and PEI-ADPC8.3%/DNA polyplexes were under 100nm with a beneficial neutral surface at N/P ratio of 30. Sufficient ADPC shell endowed the polyplexes with high colloidal stability and low cytotoxicity. Compared to PEGylated polyplexes, it was interesting to find out that the PEI-ADPC/DNA polyplexes were selectively uptaked by liver cancer HepG2 cells. At the presence of chloroquine to exclude the limitation of lysosome escape, the ADPC-modified polyplexes showed more effective gene transfection in cancer cells than in normal cells because of the selective cell uptake. In conclusion, the convenient PC-modification modality was found to have both the function of biostability in the physiological environment and targetability toward cancer cells uniquely, which might have great potential use in cancer gene therapy.

  6. Superparamagnetic poly(methyl methacrylate) nanoparticles surface modified with folic acid presenting cell uptake mediated by endocytosis

    NASA Astrophysics Data System (ADS)

    Feuser, Paulo Emilio; Jacques, Amanda Virtuoso; Arévalo, Juan Marcelo Carpio; Rocha, Maria Eliane Merlin; dos Santos-Silva, Maria Claudia; Sayer, Claudia; de Araújo, Pedro H. Hermes

    2016-04-01

    The encapsulation of superparamagnetic nanoparticles (MNPs) in polymeric nanoparticles (NPs) with modified surfaces can improve targeted delivery and induce cell death by hyperthermia. The goals of this study were to synthesize and characterize surface modified superparamagnetic poly(methyl methacrylate) with folic acid (FA) prepared by miniemulsion polymerization (MNPsPMMA-FA) and to evaluate their in vitro cytotoxicity and cellular uptake in non-tumor cells, murine fibroblast (L929) cells and tumor cells that overexpressed folate receptor (FR) β, and chronic myeloid leukemia cells in blast crisis (K562). Lastly, hemolysis assays were performed on human red blood cells. MNPsPMMA-FA presented an average mean diameter of 135 nm and a saturation magnetization (Ms) value of 37 emu/g of iron oxide, as well as superparamagnetic behavior. The MNPsPMMA-FA did not present cytotoxicity in L929 and K562 cells. Cellular uptake assays showed a higher uptake of MNPsPMMA-FA than MNPsPMMA in K562 cells when incubated at 37 °C. On the other hand, MNPsPMMA-FA showed a low uptake when endocytosis mechanisms were blocked at low temperature (4 °C), suggesting that the MNPsPMMA-FA uptake was mediated by endocytosis. High concentrations of MNPsPMMA-FA showed hemocompatibility when incubated for 24 h in human red blood cells. Therefore, our results suggest that these carrier systems can be an excellent alternative in targeted drug delivery via FR.

  7. Development of nanostructured and surface modified semiconductors for hybrid organic-inorganic solar cells.

    SciTech Connect

    Hsu, Julia, W. P.

    2008-09-01

    Solar energy conversion is increasingly being recognized as one of the principal ways to meet future energy needs without causing detrimental environmental impact. Hybrid organic-inorganic solar cells (SCs) are attracting particular interest due to the potential for low cost manufacturing and for use in new applications, such as consumer electronics, architectural integration and light-weight sensors. Key materials advantages of these next generation SCs over conventional semiconductor SCs are in design opportunities--since the different functions of the SCs are carried out by different materials, there are greater materials choices for producing optimized structures. In this project, we explore the hybrid organic-inorganic solar cell system that consists of oxide, primarily ZnO, nanostructures as the electron transporter and poly-(3-hexylthiophene) (P3HT) as the light-absorber and hole transporter. It builds on our capabilities in the solution synthesis of nanostructured semiconducting oxide arrays to this photovoltaic (PV) technology. The three challenges in this hybrid material system for solar applications are (1) achieving inorganic nanostructures with critical spacing that matches the exciton diffusion in the polymer, {approx} 10 nm, (2) infiltrating the polymer completely into the dense nanostructure arrays, and (3) optimizing the interfacial properties to facilitate efficient charge transfer. We have gained an understanding and control over growing oriented ZnO nanorods with sub-50 nm diameters and the required rod-to-rod spacing on various substrates. We have developed novel approaches to infiltrate commercially available P3HT in the narrow spacing between ZnO nanorods. Also, we have begun to explore ways to modify the interfacial properties. In addition, we have established device fabrication and testing capabilities at Sandia for prototype devices. Moreover, the control synthesis of ZnO nanorod arrays lead to the development of an efficient anti

  8. Zinc-modified titanium surface enhances osteoblast differentiation of dental pulp stem cells in vitro

    PubMed Central

    Yusa, Kazuyuki; Yamamoto, Osamu; Takano, Hiroshi; Fukuda, Masayuki; Iino, Mitsuyoshi

    2016-01-01

    Zinc is an essential trace element that plays an important role in differentiation of osteoblasts and bone modeling. This in vitro study aimed to evaluate the osteoblast differentiation of human dental pulp stem cells (DPSCs) on zinc-modified titanium (Zn-Ti) that releases zinc ions from its surface. Based on real-time PCR, alkaline phosphatase (ALP) activity and Western blot analysis data, we investigated osteoblast differentiation of DPSCs cultured on Zn-Ti and controls. DPSCs cultured on Zn-Ti exhibited significantly up-regulated gene expression levels of osteoblast-related genes of type I collagen (Col I), bone morphogenetic protein 2 (BMP2), ALP, runt-related transcription factor 2 (Runx2), osteopontin (OPN), and vascular endothelial growth factor A (VEGF A), as compared with controls. We also investigated extracellular matrix (ECM) mineralization by Alizarin Red S (ARS) staining and found that Zn-Ti significantly promoted ECM mineralization when compared with controls. These findings suggest that the combination of Zn-Ti and DPSCs provides a novel approach for bone regeneration therapy. PMID:27387130

  9. Cellular uptake pathways of lipid-modified cationic polymers in gene delivery to primary cells.

    PubMed

    Hsu, Charlie Y M; Uludağ, Hasan

    2012-11-01

    Hydrophobic modifications have emerged as a promising approach to improve the efficiency of non-viral gene delivery vectors (GDV). Functional GDVs from non-toxic polymers have been created with this approach but the mechanism(s) behind lipid-mediated enhancement in transfection remains to be clarified. Using a linoleic acid-substituted 2 kDa polyethylenimine (PEI2LA), we aimed to define the cellular uptake pathways and intracellular trafficking of plasmid DNA in normal human foreskin fibroblast cells. Several pharmacological compounds were applied to selectively inhibit uptake by clathrin-mediated endocytosis (CME), caveolin-mediated endocytosis (CvME) and macropinocytosis. We found that PEI2LA complexes were taken up predominantly through CME, and to a lesser extent by CvME. In contrast, its precursor molecule, PEI2 complexes was internalized primarily by CvME and macropinocytosis. The commonly used 25 kDa PEI 25 complexes utilized all endocytic pathways, suggesting its efficiency is derived from a different set of transfection pathways than PEI2LA. We further applied several endosome disruptive agents and found that hypertonic media enhanced the transfection of PEI2LA by 6.5-fold. We infer that lipid substitution changes the normal uptake pathways significantly and transfection with hydrophobically modified GDVs may be further enhanced by incorporating endosome disruptive elements into vector design.

  10. Effect of Graphene-Graphene Oxide Modified Anode on the Performance of Microbial Fuel Cell

    PubMed Central

    Yang, Na; Ren, Yueping; Li, Xiufen; Wang, Xinhua

    2016-01-01

    The inferior hydrophilicity of graphene is an adverse factor to the performance of the graphene modified anodes (G anodes) in microbial fuel cells (MFCs). In this paper, different amounts of hydrophilic graphene oxide (GO) were doped into the modification layers to elevate the hydrophilicity of the G anodes so as to further improve their performance. Increasing the GO doped ratio from 0.15 mg·mg−1 to 0.2 mg·mg−1 and 0.25 mg·mg−1, the static water contact angle (θc) of the G-GO anodes decreased from 74.2 ± 0.52° to 64.6 ± 2.75° and 41.7 ± 3.69°, respectively. The G-GO0.2 anode with GO doped ratio of 0.2 mg·mg−1 exhibited the optimal performance and the maximum power density (Pmax) of the corresponding MFC was 1100.18 mW·m−2, 1.51 times higher than that of the MFC with the G anode. PMID:28335302

  11. Minimally Modified LDL Upregulates Endothelin Type A Receptors in Rat Coronary Arterial Smooth Muscle Cells

    PubMed Central

    Li, Jie; Cao, Lei; Xu, Cang-Bao; Wang, Jun-Jie

    2013-01-01

    Minimally modified low-density lipoprotein (mmLDL) is a risk factor for cardiovascular disease. The present study investigated the effects of mmLDL on the expression of endothelin type A (ETA) receptors in coronary arteries. Rat coronary arteries were organ-cultured for 24 h. The contractile responses were recorded using a myographic system. ETA receptor mRNA and protein expressions were determined using real-time PCR and western blotting, respectively. The results showed that organ-culturing in the presence of mmLDL enhanced the arterial contractility mediated by the ETA receptor in a concentration-dependent and time-dependent manner. Culturing with mmLDL (10 μg/mL) for 24 h shifted the concentration-contractile curves toward the left significantly with increased Emax of 228% ± 20% from control of 100% ± 10% and significantly increased ETA receptor mRNA and protein levels. Inhibition of the protein kinase C, extracellular signal-related kinases 1 and 2 (ERK1/2), or NF-κB activities significantly attenuated the effects of mmLDL. The c-Jun N-terminal kinase inhibitor or the p38 pathway inhibitor, however, had no such effects. The results indicate that mmLDL upregulates the ETA receptors in rat coronary arterial smooth muscle cells mainly via activating protein kinase C, ERK1/2, and the downstream transcriptional factor, NF-κB. PMID:23861561

  12. New biosensor for detection of copper ions in water based on immobilized genetically modified yeast cells.

    PubMed

    Vopálenská, Irena; Váchová, Libuše; Palková, Zdena

    2015-10-15

    Contamination of water by heavy metals represents a potential risk for both aquatic and terrestrial organisms, including humans. Heavy metals in water resources can come from various industrial activities, and drinking water can be ex-post contaminated by heavy metals such as Cu(2+) from house fittings (e.g., water reservoirs) and pipes. Here, we present a new copper biosensor capable of detecting copper ions at concentrations of 1-100 μM. This biosensor is based on cells of a specifically modified Saccharomyces cerevisiae strain immobilized in alginate beads. Depending on the concentration of copper, the biosensor beads change color from white, when copper is present in concentrations below the detection limit, to pink or red based on the increase in copper concentration. The biosensor was successfully tested in the determination of copper concentrations in real samples of water contaminated with copper ions. In contrast to analytical methods or other biosensors based on fluorescent proteins, the newly designed biosensor does not require specific equipment and allows the quick detection of copper in many parallel samples.

  13. Colloidal PbSe Solar Cells with Molybdenum Oxide Modified Graphene Anodes.

    PubMed

    Wu, Hua; Zhang, Xiaoyu; Zhang, Yu; Yan, Long; Gao, Wenzhu; Zhang, Tieqiang; Wang, Yiding; Zhao, Jun; Yu, William W

    2015-09-30

    With good electrical conductivity, optical transparency, and mechanical compliance, graphene films have shown great potential in application for photovoltaic devices as electrodes. However, photovoltaic devices employing graphene anodes usually suffer from poor hole collection efficiency because of the mismatch of energy levels between the anode and light-harvesting layers. Here, a simple solution treatment and a low-cost solution-processed molybdenum oxide (MoOx) film were used to modify the work function of graphene and the interfacial morphology, respectively, yielding highly efficient hole transfer. As a result, the graphene/MoOx anodes demonstrated low surface roughness and high electrical conductivity. Using the graphene/MoOx anodes in PbSe nanocrystal solar cells, we achieved 1 sun power conversion efficiency of 3.56%. Compared to the control devices with indium tin oxide anodes, the graphene/MoOx-based devices show excellent performance, demonstrating the great potential of the graphene/MoOx anodes for use in optoelectronics.

  14. Characterization of direct methanol fuel cell (DMFC) applications with H 2SO 4 modified chitosan membrane

    NASA Astrophysics Data System (ADS)

    Osifo, Peter O.; Masala, Aluwani

    Chitosan (Chs) flakes were prepared from chitin materials that were extracted from the exoskeleton of Cape rock lobsters in South Africa. The Chs flakes were prepared into membranes and the Chs membranes were modified by cross-linking with H 2SO 4. The cross-linked Chs membranes were characterized for the application in direct methanol fuel cells. The Chs membrane characteristics such as water uptake, thermal stability, proton resistance and methanol permeability were compared to that of high performance conventional Nafion 117 membranes. Under the temperature range studied 20-60 °C, the membrane water uptake for Chs was found to be higher than that of Nafion. Thermal analysis revealed that Chs membranes could withstand temperature as high as 230 °C whereas Nafion 117 membranes were stable to 320 °C under nitrogen. Nafion 117 membranes were found to exhibit high proton resistance of 284 s cm -1 than Chs membranes of 204 s cm -1. The proton fluxes across the membranes were 2.73 mol cm -2 s -1 for Chs- and 1.12 mol cm -2 s -1 Nafion membranes. Methanol (MeOH) permeability through Chs membrane was less, 1.4 × 10 -6 cm 2 s -1 for Chs membranes and 3.9 × 10 -6 cm 2 s -1 for Nafion 117 membranes at 20 °C. Chs and Nafion membranes were fabricated into membrane electrode assemblies (MAE) and their performances measure in a free-breathing commercial single cell DMFC. The Nafion membranes showed a better performance as the power density determined for Nafion membranes of 0.0075 W cm -2 was 2.7 times higher than in the case of Chs MEA.

  15. Cell surface adrenergic receptor stimulation modifies the endothelial response to SIRS. Systemic Inflammatory Response Syndrome.

    PubMed

    Tighe, D; Moss, R; Bennett, D

    1996-11-01

    The complex pathway seen in patients with the systemic inflammatory response syndrome (SIRS) does not readily respond to mediator blockade. All such trials conducted in SIRS patients have shown no benefit in reducing mortality. We have shown experimentally that in sepsis, the administration of beta 2-adrenoceptor agonists reduces hepatic cellular injury, whereas administration of an alpha 1-adrenoceptor agonist increases hepatic cellular injury. Inflammatory mediators can cause a dose-related reversible change in target endothelial cells (ECs). There is a substantial body of literature describing the anti-inflammatory effects of beta 2-adrenoceptor agonists. They reduce both the increased permeability and the production of inflammatory mediators from ECs. Cellular transduction processes are involved when adrenergic receptor agonists modify either the anti-inflammatory or proinflammatory response to sepsis in ECs. Inflammatory mediators and alpha 1-adrenoceptor agonists stimulate their trimeric G protein-linked receptors to produce diacylglycerol (DAG) and increase the intracellular concentration of calcium. DAG is involved in the production of both inflammatory proteins and lipids. In addition, mitogen-activated protein kinase (MAPK) is activated which is also involved in the production of inflammatory proteins and lipids. beta 2-adrenoceptor agonists activate their trimeric G protein-linked receptors to produce the stimulatory G protein (Gs). Gs stimulates adenyl cyclase to form cyclic adenosine monophosphate (cAMP) and activate protein kinase A (PKA). PKA is involved in activating gene transcription agents to produce anti-inflammatory proteins such as interleukin-10. PKA also inhibits phospholipase C and MAPK. Although promising, the use of beta-adrenoceptor agonists or agonists that increase cellular cAMP to activate the cells' endogenous anti-inflammatory pathway requires further study.

  16. Salivary gland acinar cells regenerate functional glandular structures in modified hydrogels

    NASA Astrophysics Data System (ADS)

    Pradhan, Swati

    Xerostomia, a condition resulting from irradiation of the head and neck, affects over 40,000 cancer patients each year in the United States. Direct radiation damage of the acinar cells that secrete fluid and protein results in salivary gland hypofunction. Present medical management for xerostomia for patients treated for upper respiratory cancer is largely ineffective. Patients who have survived their terminal diagnosis are often left with a diminished quality of life and are unable to enjoy the simple pleasures of eating and drinking. This project aims to ultimately reduce human suffering by developing a functional implantable artificial salivary gland. The goal was to create an extracellular matrix (ECM) modified hyaluronic acid (HA) based hydrogel culture system that allows for the growth and differentiation of salivary acinar cells into functional acini-like structures capable of secreting large amounts of protein and fluid unidirectionally and to ultimately engineer a functional artificial salivary gland that can be implanted into an animal model. A tissue collection protocol was established and salivary gland tissue was obtained from patients undergoing head and neck surgery. The tissue specimen was assessed by histology and immunohistochemistry to establish the phenotype of normal salivary gland cells including the native basement membranes. Hematoxylin and eosin staining confirmed normal glandular tissue structures including intercalated ducts, striated ducts and acini. alpha-Amylase and periodic acid schiff stain, used for structures with a high proportion of carbohydrate macromolecules, preferentially stained acinar cells in the tissue. Intercalated and striated duct structures were identified using cytokeratins 19 and 7 staining. Myoepithelial cells positive for cytokeratin 14 were found wrapped around the serous and mucous acini. Tight junction components including ZO-1 and E-cadherin were present between both ductal and acinar cells. Ductal and acinar

  17. A dosimetric uncertainty analysis for photon-emitting brachytherapy sources: report of AAPM Task Group No. 138 and GEC-ESTRO.

    PubMed

    DeWerd, Larry A; Ibbott, Geoffrey S; Meigooni, Ali S; Mitch, Michael G; Rivard, Mark J; Stump, Kurt E; Thomadsen, Bruce R; Venselaar, Jack L M

    2011-02-01

    This report addresses uncertainties pertaining to brachytherapy single-source dosimetry preceding clinical use. The International Organization for Standardization (ISO) Guide to the Expression of Uncertainty in Measurement (GUM) and the National Institute of Standards and Technology (NIST) Technical Note 1297 are taken as reference standards for uncertainty formalism. Uncertainties in using detectors to measure or utilizing Monte Carlo methods to estimate brachytherapy dose distributions are provided with discussion of the components intrinsic to the overall dosimetric assessment. Uncertainties provided are based on published observations and cited when available. The uncertainty propagation from the primary calibration standard through transfer to the clinic for air-kerma strength is covered first. Uncertainties in each of the brachytherapy dosimetry parameters of the TG-43 formalism are then explored, ending with transfer to the clinic and recommended approaches. Dosimetric uncertainties during treatment delivery are considered briefly but are not included in the detailed analysis. For low- and high-energy brachytherapy sources of low dose rate and high dose rate, a combined dosimetric uncertainty <5% (k=1) is estimated, which is consistent with prior literature estimates. Recommendations are provided for clinical medical physicists, dosimetry investigators, and source and treatment planning system manufacturers. These recommendations include the use of the GUM and NIST reports, a requirement of constancy of manufacturer source design, dosimetry investigator guidelines, provision of the lowest uncertainty for patient treatment dosimetry, and the establishment of an action level based on dosimetric uncertainty. These recommendations reflect the guidance of the American Association of Physicists in Medicine (AAPM) and the Groupe Européen de Curiethérapie-European Society for Therapeutic Radiology and Oncology (GEC-ESTRO) for their members and may also be used as

  18. A dosimetric uncertainty analysis for photon-emitting brachytherapy sources: Report of AAPM Task Group No. 138 and GEC-ESTRO

    SciTech Connect

    DeWerd, Larry A.; Ibbott, Geoffrey S.; Meigooni, Ali S.; Mitch, Michael G.; Rivard, Mark J.; Stump, Kurt E.; Thomadsen, Bruce R.; Venselaar, Jack L. M.

    2011-02-15

    This report addresses uncertainties pertaining to brachytherapy single-source dosimetry preceding clinical use. The International Organization for Standardization (ISO) Guide to the Expression of Uncertainty in Measurement (GUM) and the National Institute of Standards and Technology (NIST) Technical Note 1297 are taken as reference standards for uncertainty formalism. Uncertainties in using detectors to measure or utilizing Monte Carlo methods to estimate brachytherapy dose distributions are provided with discussion of the components intrinsic to the overall dosimetric assessment. Uncertainties provided are based on published observations and cited when available. The uncertainty propagation from the primary calibration standard through transfer to the clinic for air-kerma strength is covered first. Uncertainties in each of the brachytherapy dosimetry parameters of the TG-43 formalism are then explored, ending with transfer to the clinic and recommended approaches. Dosimetric uncertainties during treatment delivery are considered briefly but are not included in the detailed analysis. For low- and high-energy brachytherapy sources of low dose rate and high dose rate, a combined dosimetric uncertainty <5% (k=1) is estimated, which is consistent with prior literature estimates. Recommendations are provided for clinical medical physicists, dosimetry investigators, and source and treatment planning system manufacturers. These recommendations include the use of the GUM and NIST reports, a requirement of constancy of manufacturer source design, dosimetry investigator guidelines, provision of the lowest uncertainty for patient treatment dosimetry, and the establishment of an action level based on dosimetric uncertainty. These recommendations reflect the guidance of the American Association of Physicists in Medicine (AAPM) and the Groupe Europeen de Curietherapie-European Society for Therapeutic Radiology and Oncology (GEC-ESTRO) for their members and may also be used as

  19. A dosimetric uncertainty analysis for photon-emitting brachytherapy sources: Report of AAPM Task Group No. 138 and GEC-ESTRO

    PubMed Central

    DeWerd, Larry A.; Ibbott, Geoffrey S.; Meigooni, Ali S.; Mitch, Michael G.; Rivard, Mark J.; Stump, Kurt E.; Thomadsen, Bruce R.; Venselaar, Jack L. M.

    2011-01-01

    This report addresses uncertainties pertaining to brachytherapy single-source dosimetry preceding clinical use. The International Organization for Standardization (ISO) Guide to the Expression of Uncertainty in Measurement (GUM) and the National Institute of Standards and Technology (NIST) Technical Note 1297 are taken as reference standards for uncertainty formalism. Uncertainties in using detectors to measure or utilizing Monte Carlo methods to estimate brachytherapy dose distributions are provided with discussion of the components intrinsic to the overall dosimetric assessment. Uncertainties provided are based on published observations and cited when available. The uncertainty propagation from the primary calibration standard through transfer to the clinic for air-kerma strength is covered first. Uncertainties in each of the brachytherapy dosimetry parameters of the TG-43 formalism are then explored, ending with transfer to the clinic and recommended approaches. Dosimetric uncertainties during treatment delivery are considered briefly but are not included in the detailed analysis. For low- and high-energy brachytherapy sources of low dose rate and high dose rate, a combined dosimetric uncertainty <5% (k=1) is estimated, which is consistent with prior literature estimates. Recommendations are provided for clinical medical physicists, dosimetry investigators, and source and treatment planning system manufacturers. These recommendations include the use of the GUM and NIST reports, a requirement of constancy of manufacturer source design, dosimetry investigator guidelines, provision of the lowest uncertainty for patient treatment dosimetry, and the establishment of an action level based on dosimetric uncertainty. These recommendations reflect the guidance of the American Association of Physicists in Medicine (AAPM) and the Groupe Européen de Curiethérapie–European Society for Therapeutic Radiology and Oncology (GEC-ESTRO) for their members and may also be used

  20. Modified PHBV scaffolds by in situ UV polymerization: structural characteristic, mechanical properties and bone mesenchymal stem cell compatibility.

    PubMed

    Ke, Y; Wang, Y J; Ren, L; Zhao, Q C; Huang, W

    2010-04-01

    An ideal scaffold provides an interface for cell adhesion and maintains enough biomechanical support during tissue regeneration. Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) scaffolds with pore sizes ranging from 100 to 500 microm and porosity approximately 90% were prepared by the particulate-leaching method, and then modified by the introduction of polyacrylamide (PAM) on the inner surface of scaffolds using in situ UV polymerization, with the aim of enhancing the biological and mechanical properties of the PHBV scaffolds. The modified PHBV scaffolds had interconnected pores with porosity of 75.4-78.6% and pore sizes at peak volume from 20 to 50 microm. The compressive load and modulus were up to 62.45 N and 1.06 MPa, respectively. The water swelling percentage (WSP) of the modified PHBV scaffolds increased notably compared with that of the PHBV scaffolds, with the maximum WSP at 537%. Sheep bone mesenchymal stem cells (BMSC) were cultured on the PHBV and modified PHBV. The hydrophilic PAM chains did not influence BMSC viability or proliferation index, but the initial cell adhesion at 1h of culture was enhanced significantly. Framing PHBV scaffold along with gel-like PAM chains inside is a novel model of inner surface modification for PHBV scaffolds, which shows potential in tissue engineering applications.

  1. Modified sugar beet pectin induces apoptosis of colon cancer cells via an interaction with the neutral sugar side-chains.

    PubMed

    Maxwell, Ellen G; Colquhoun, Ian J; Chau, Hoa K; Hotchkiss, Arland T; Waldron, Keith W; Morris, Victor J; Belshaw, Nigel J

    2016-01-20

    Pectins extracted from a variety of sources and modified with heat and/or pH have previously been shown to exhibit activity towards several cancer cell lines. However, the structural basis for the anti-cancer activity of modified pectin requires clarification. Sugar beet and citrus pectin extracts have been compared. Pectin extracted from sugar beet pulp only weakly affected the viability of colon cancer cells. Alkali treatment increased the anti-cancer effect of sugar beet pectin via an induction of apoptosis. Alkali treatment decreased the degree of esterification (DE) and increased the ratio of rhamnogalacturonan I (RGI) to homogalacturonan. Low DE per se did not play a significant role in the anti-cancer activity. However, the enzymatic removal of galactose and, to a lesser extent, arabinose from the pectin decreased the effect on cancer cells indicating that the neutral sugar-containing RGI regions are important for pectin bioactivity.

  2. A modified chronic ocular hypertension rat model for retinal ganglion cell neuroprotection.

    PubMed

    Zhong, Lichun

    2013-09-01

    This study aimed to modify a chronic ocular hypertension (OHT) rat model to screen for potential compounds to protect retinal ganglion cells (RGCs) from responding to increased intraocular pressure (IOP). A total of 266 rats were prepared and randomly grouped according to different time-points, namely, weeks 3, 8, 16, and 24. Rats were sedated and eye examination was performed to score as the corneal damage on a scale of 1 to 4. The OHT rat model was created via the injection of a hypertonic saline solution into the episcleral veins once weekly for two weeks. OHT was identified when the IOP at week 0 was [Symbol: see text] 6 mmHg than that at week -2 for the same eye. Viable RGCs were labeled by injecting 4% FluoroGold. Rats were sacrificed, and the eyes were enucleated and fixed. The fixed retinas were dissected to prepare flat whole-mounts. The viable RGCs were visualized and imaged. The IOP (mean ± SD) was calculated, and data were analyzed by the paired t-test and one-way ANOVA. The OHT model was created in 234 of 266 rats (87.97%), whereas 32 rats (12.03%) were removed from the study because of the absence of IOP elevation (11.28%) and/or corneal damage scores over 4 (0.75%). IOP was elevated by as much as 81.35% for 24 weeks. The average IOP was (16.68 ± 0.98) mmHg in non-OHT eyes (n = 234), but was (27.95 ± 0.97) mmHg in OHTeyes (n = 234). Viable RGCs in the OHT eyes were significantly decreased in a time-dependent manner by 29.41%, 38.24%, 55.32%, and 59.30% at weeks 3, 8, 16, and 24, respectively, as compared to viable RGCs in the non-OHT eyes (P < 0.05). The OHT model was successfully created in 88% of the rats. The IOP in the OHT eyes was elevated by approximately 81% for 24 weeks. The number of viable RGCs was decreased by 59% of the rats in a time-dependent manner. The modified OHT model may provide an effective and reliable method for screening drugs to protect RGCs from glaucoma.

  3. Increased phosphorylation of eIF2α in chronic myeloid leukemia cells stimulates secretion of matrix modifying enzymes

    PubMed Central

    Podszywalow-Bartnicka, Paulina; Cmoch, Anna; Wolczyk, Magdalena; Bugajski, Lukasz; Tkaczyk, Marta; Dadlez, Michal; Nieborowska-Skorska, Margaret; Koromilas, Antonis E.; Skorski, Tomasz; Piwocka, Katarzyna

    2016-01-01

    Recent studies underscore the role of the microenvironment in therapy resistance of chronic myeloid leukemia (CML) cells and leukemia progression. We previously showed that sustained mild activation of endoplasmic reticulum (ER) stress in CML cells supports their survival and resistance to chemotherapy. We now demonstrate, using dominant negative non-phosphorylable mutant of eukaryotic initiation factor 2 α subunit (eIF2α), that phosphorylation of eIF2α (eIF2α-P), which is a hallmark of ER stress in CML cells, substantially enhances their invasive potential and modifies their ability to secrete extracellular components, including the matrix-modifying enzymes cathepsins and matrix metalloproteinases. These changes are dependent on the induction of activating transcription factor-4 (ATF4) and facilitate extracellular matrix degradation by CML cells. Conditioned media from CML cells with constitutive activation of the eIF2α-P/ATF4 pathway induces invasiveness of bone marrow stromal fibroblasts, suggesting that eIF2α-P may be important for extracellular matrix remodeling and thus leukemia cells-stroma interactions. Our data show that activation of stress response in CML cells may contribute to the disruption of bone marrow niche components by cancer cells and in this way support CML progression. PMID:27802179

  4. Selective biophysical interactions of surface modified nanoparticles with cancer cell lipids improve tumor targeting and gene therapy.

    PubMed

    Sharma, Blanka; Peetla, Chiranjeevi; Adjei, Isaac M; Labhasetwar, Vinod

    2013-07-01

    Targeting gene- or drug-loaded nanoparticles (NPs) to tumors and ensuring their intratumoral retention after systemic administration remain key challenges to improving the efficacy of NP-based therapeutics. Here, we investigate a novel targeting approach that exploits changes in lipid metabolism and cell membrane biophysics that occur during malignancy. We hypothesized that modifications to the surface of NPs that preferentially increase their biophysical interaction with the membrane lipids of cancer cells will improve intratumoral retention and in vivo efficacy upon delivery of NPs loaded with a therapeutic gene. We have demonstrated that different surfactants, incorporated onto the NPs' surface, affect the biophysical interactions of NPs with the lipids of cancer cells and normal endothelial cells. NPs surface modified with didodecyldimethylammoniumbromide (DMAB) demonstrated greater interaction with cancer cell lipids, which was 6.7-fold greater than with unmodified NPs and 5.5-fold greater than with endothelial cell lipids. This correlated with increased uptake of DMAB-modified NPs with incubation time by cancer cells compared to other formulations of NPs and to uptake by endothelial cells. Upon systemic injection, DMAB-NPs demonstrated a 4.6-fold increase in tumor accumulation compared to unmodified NPs which also correlated to improved efficacy of p53 gene therapy. Characterization of the biophysical interactions between NPs and lipid membranes of tumors or other diseased tissues/organs may hold promise for engineering targeted delivery of therapeutics.

  5. Modified arabinoxylan rice bran (MGN-3/Biobran) enhances yeast-induced apoptosis in human breast cancer cells in vitro.

    PubMed

    Ghoneum, Mamdooh; Gollapudi, Sastry

    2005-01-01

    We have recently reported that phagocytosis of killed Saccharomyces cerevisiae, baker's yeast, induced apoptosis in human breast cancer cell lines MCF-7, ZR-75-1 and HCC70. In this study we have evaluated the effect of treatment with MGN-3, a modified arabinoxylan from rice bran, on phagocytosis and yeast-induced apoptosis in breast cancer cells. Cancer cells were cultured with yeast at a ratio of 1:10 in the absence or presence of MGN-3, and the percentages of phagocytic and apoptotic cancer cells were examined by flow cytometry and by cytospin preparations. Cancer cells treated with MGN-3 exhibited increased percentages of attachment (200%) and uptake of yeast (313%) by MCF-7 cells at 0.5 hr, as compared with cells without MGN-3. In addition, treatment with MGN-3 resulted in a 2 fold increase in the percentage of apoptotic MCF-7 cells, 2.5 fold for ZR-75 cells and 1.8 fold for HCC70 cells. MGN-3 effect was dose-dependent and associated with increased activation of caspases 8 and 9 in MCF-7 cells, and caspases 8, 9 and 3 in HCC70 cells. This data demonstrates that MGN-3 accelerates phagocytosis-induced apoptosis of cancer cells, which may represent a novel therapeutic strategy for the treatment of breast cancer.

  6. Crown ether-modified electrodes for the simultaneous stripping voltammetric determination of Cd(II), Pb(II) and Cu(II).

    PubMed

    Serrano, Núria; González-Calabuig, Andreu; del Valle, Manel

    2015-06-01

    This work describes the immobilization of 4-carboxybenzo-18-crown-6 (CB-18-crown-6) and 4-carboxybenzo-15-crown-5 (CB-15-crown-5) assisted by lysine on aryl diazonium salt monolayers anchored to the surface of graphite-epoxy composite electrodes (GEC), and their use for the simultaneous determination of Cd(II), Pb(II) and Cu(II) by differential pulse anodic stripping voltammetry (DPASV). These modified electrodes display a good repeatability and reproducibility with detection and quantification limits at levels of µg L(-1) (ppb), confirming their suitability for the determination of Cd(II), Pb(II) and Cu(II) ions in environmental samples. The overlapped nature of the multimetal stripping measurements was resolved by employing the two-sensor array CB-15-crown-5-GEC and CB-18-crown-6-GEC, since the metal complex selectivity exhibited by the considered ligands could add some discrimination power. For the processing of the voltammograms, Discrete Wavelet Transform and Causal Index were selected as preprocessing tools for data compression coupled with an artificial neural network for the modeling of the obtained responses, allowing the resolution of mixtures of these metals with good prediction of their concentrations (correlation with expected values for an external test subset better than 0.942).

  7. The target gene carrying validity to HePG2 cells with the brush-like glutathione modified chitosan compound.

    PubMed

    Li, Congxin; Zhou, Dezhong; Hu, Yuling; Zhou, Hao; Chen, Jiatong; Zhang, Zhengpu; Guo, Tianying

    2012-06-05

    The grafting modified chitosan with L-glutathione (GSH) end capped PEG brush-like poly [poly(ethylene glycol) methacrylate] (PMPEG), CS-PMPEG-GSH, as the pDNA condensed vector material could result in a much higher transfection efficiency and lower cytotoxity for NIH3T3 cells. In this work, we have further examined the morphology stabilities of CS-PMPEG-GSH/pDNA vectors at different medium pH values and in the presence of serum protein in detail. And then the targeted characters for HepG2 cells have been probed by tracing the cell uptake behavior and transfection efficiency.

  8. Myristoylation increases the CD8+T-cell response to a GFP prototype antigen delivered by modified vaccinia virus Ankara.

    PubMed

    Marr, Lisa; Lülf, Anna-Theresa; Freudenstein, Astrid; Sutter, Gerd; Volz, Asisa

    2016-04-01

    Activation of CD8(+)T-cells is an essential part of immune responses elicited by recombinant modified vaccinia virus Ankara (MVA). Strategies to enhance T-cell responses to antigens may be particularly necessary for broadly protective immunization against influenza A virus infections or for candidate vaccines targeting chronic infections and cancer. Here, we tested recombinant MVAs that targeted a model antigen, GFP, to different localizations in infected cells. In vitro characterization demonstrated that GFP accumulated in the nucleus (MVA-nls-GFP), associated with cellular membranes (MVA-myr-GFP) or was equally distributed throughout the cell (MVA-GFP). On vaccination, we found significantly higher levels of GFP-specific CD8(+)T-cells in MVA-myr-GFP-vaccinated BALB/c mice than in those immunized with MVA-GFP or MVA-nls-GFP. Thus, myristoyl modification may be a useful strategy to enhance CD8(+)T-cell responses to MVA-delivered target antigens.

  9. The protective effect of modified intravenous immunoglobulin in LPS sepsis model is associated with an increased IRA B cells response.

    PubMed

    Djoumerska-Alexieva, Iglika; Pashova, Shina; Vassilev, Tchavdar; Pashov, Anastas

    2013-04-01

    Intravenous immunoglobulin preparations (IVIg) that have undergone a mild oxidizing treatment with ferrous ions have an increased polyspecificity, which is not associated with a higher propensity to form aggregates. Among other biological properties of the modified IVIg, a protective effect in LPS sepsis model stands out as the native preparation is totally devoid of it or even exacerbates sepsis. A recent finding identified an LPS induced subset of B1 lymphocytes that migrate from the peritoneal cavity to the spleen acquiring the expression of CD93, GM-CSF as well as the capacity to control sepsis. This report demonstrates that modified IVIg, but not the native preparation, causes a further increase in this population during LPS sepsis. Partial targeted suppression of the peritoneal B cell proliferation by an intracellular dye abrogates this effect and the clinical benefit of modified IVIg.

  10. Library Synthesis, Screening, and Discovery of Modified Zinc(II)-Bis(dipicolylamine) Probe for Enhanced Molecular Imaging of Cell Death

    PubMed Central

    2015-01-01

    Zinc(II)-bis(dipicolylamine) (Zn-BDPA) coordination complexes selectively target the surfaces of dead and dying mammalian cells, and they have promise as molecular probes for imaging cell death. A necessary step toward eventual clinical imaging applications is the development of next-generation Zn-BDPA complexes with enhanced affinity for the cell death membrane biomarker, phosphatidylserine (PS). This study employed an iterative cycle of library synthesis and screening, using a novel rapid equilibrium dialysis assay, to discover a modified Zn-BDPA structure with high and selective affinity for vesicles containing PS. The lead structure was converted into a deep-red fluorescent probe and its targeting and imaging performance was compared with an unmodified control Zn-BDPA probe. The evaluation process included a series of FRET-based vesicle titration studies, cell microscopy experiments, and rat tumor biodistribution measurements. In all cases, the modified probe exhibited comparatively higher affinity and selectivity for the target membranes of dead and dying cells. The results show that this next-generation deep-red fluorescent Zn-BDPA probe is well suited for preclinical molecular imaging of cell death in cell cultures and animal models. Furthermore, it should be possible to substitute the deep-red fluorophore with alternative reporter groups that enable clinically useful, deep-tissue imaging modalities, such as MRI and nuclear imaging. PMID:24575875

  11. Cadm1 is a metastasis susceptibility gene that suppresses metastasis by modifying tumor interaction with the cell-mediated immunity.

    PubMed

    Faraji, Farhoud; Pang, Yanli; Walker, Renard C; Nieves Borges, Rosan; Yang, Li; Hunter, Kent W

    2012-09-01

    Metastasis is a complex process utilizing both tumor-cell-autonomous properties and host-derived factors, including cellular immunity. We have previously shown that germline polymorphisms can modify tumor cell metastatic capabilities through cell-autonomous mechanisms. However, how metastasis susceptibility genes interact with the tumor stroma is incompletely understood. Here, we employ a complex genetic screen to identify Cadm1 as a novel modifier of metastasis. We demonstrate that Cadm1 can specifically suppress metastasis without affecting primary tumor growth. Unexpectedly, Cadm1 did not alter tumor-cell-autonomous properties such as proliferation or invasion, but required the host's adaptive immune system to affect metastasis. The metastasis-suppressing effect of Cadm1 was lost in mice lacking T cell-mediated immunity, which was partially phenocopied by depleting CD8(+) T cells in immune-competent mice. Our data show a novel function for Cadm1 in suppressing metastasis by sensitizing tumor cells to immune surveillance mechanisms, and this is the first report of a heritable metastasis susceptibility gene engaging tumor non-autonomous factors.

  12. Determination of glutathione in single HepG2 cells by capillary electrophoresis with reduced graphene oxide modified microelectrode.

    PubMed

    Wang, Xiaolei; Wang, Jun; Fu, Hongyan; Liu, Dongju; Chen, Zhenzhen

    2014-12-01

    Determination of intracellular bioactive species will afford beneficial information related to cell metabolism, signal transduction, cell function, and disease treatment. In this study, the electrochemically reduced graphene oxide modified carbon fiber microdisk electrode (ER-GOME) was used as a detector of CZE-electrochemical detection and developed to detect glutathione (GSH). The electrocatalytic activity of the modified microelectrode was characterized by cyclic voltammetry. Under optimized experimental conditions, the concentration linear range of GSH was from 1 to 60 μM. When the S/N ratio was 3, the concentration detection limit was 1 μM. Compared with the unmodified carbon fiber microdisk electrode, the sensitivity was enhanced more than five times. With the use of this method, the average contents of GSH in single HepG2 cells were found to be 7.13 ± 1.11 fmol (n = 10). Compared with gold/mercury amalgam microelectrode, which was usually used in determining GSH, the electrochemically reduced graphene oxide modified carbon fiber microdisk electrode was friendly to environment for free mercury. Furthermore, there were several merits of the novel electrochemical detector coupled with CE, such as comparative repeatability, easy fabrication, and high sensitivity, hold great potential for the single-cell assay.

  13. Treating congenital megacolon by transplanting GDNF and GFRα-1 double genetically modified rat bone marrow mesenchymal stem cells.

    PubMed

    Zhou, C B; Peng, C H; Pang, W B; Zhang, D; Chen, Y J

    2015-08-14

    We studied the survival and gene expression of glial cell line-derived neurotrophic factor (GDNF) and GDNF receptor α-1 (GFRα-1) double-genetically modified rat bone marrow mesenchymal stem cells (BMSCs) transplanted into the intestinal walls of the rat models with congenital megacolon and determine the feasibility of treatment by transplantation of double-genetically modified rat BMSCs. The rat colorectal intestinal wall nerve plexus was treated with the cationic surface active agent benzalkonium chloride to establish an experimental megacolon model. The rat target genes GDNF and GFRα-1 were extracted and ligated into pEGFP-N1. Eukaryotic fluorescent expression vectors carrying the GDNF and GFRα-1 genes were transfected into BMSCs by in vitro culture. We treated congenital megacolon by transplanting double-genetically modified rat bone marrow mesenchymal stem cells. The pEGFP-EGFP-GDNF-GFRα-1 double-gene co-expressing the eukaryotic expression plasmid vector was successfully established. Protein gene protein 9.5 and vasoactive intestinal peptide-positive ganglion cells showed no positive expression in the phosphate-buffered saline transplantation group based on an immunofluorescence test at 1, 2, and 4 weeks after transplantation of BMSCs. Additionally, compared with the phosphate-buffered saline transplantation group, the expression of rearranged during transfection, GDNF, and GFRα-1 mRNA in the stem cell transplantation group increased gradually. The double-genetically modified BMSCs colonized and survived in the intestinal wall of the experimental megacolon rat model and expressed related genes, partially recovering the colonic neuromuscular regulatory functions and thus providing an experimental basis for treating congenital megacolon by cellular transplantation.

  14. Adoptive T-cell therapy for hematological malignancies using T cells gene-modified to express tumor antigen-specific receptors.

    PubMed

    Fujiwara, Hiroshi

    2014-02-01

    The functional properties of the adoptive immune response mediated by effector T lymphocytes are decisively regulated by their T-cell receptors (TCRs). Transfer of genes encoding target antigen-specific receptors enables polyclonal T cells to redirect toward cancer cells and virally infected cells expressing those defined antigens. Using this technology, a large population of redirected T cells displaying uniform therapeutic properties has been produced, powerfully advancing their clinical application as "cellular drugs" for adoptive immunotherapy against cancer. Clinically, anticancer adoptive immunotherapy using these genetically engineered T cells has an impressive and proven track record. Notable examples include the dramatic benefit of chimeric antigen receptor gene-modified T cells redirected towards B-cell lineage antigen CD19 in patients with chronic lymphocytic leukemia, and the impressive outcomes in the use of TCR gene-modified T cells redirected towards NY-ESO-1, a representative cancer-testis antigen, in patients with advanced melanoma and synovial cell sarcoma. In this review, we briefly overview the current status of this treatment option in the context of hematological malignancy, and discuss a number of challenges that still pose an obstacle to the full effectiveness of this strategy.

  15. An efficient genotyping method for genome-modified animals and human cells generated with CRISPR/Cas9 system.

    PubMed

    Zhu, Xiaoxiao; Xu, Yajie; Yu, Shanshan; Lu, Lu; Ding, Mingqin; Cheng, Jing; Song, Guoxu; Gao, Xing; Yao, Liangming; Fan, Dongdong; Meng, Shu; Zhang, Xuewen; Hu, Shengdi; Tian, Yong

    2014-09-19

    The rapid generation of various species and strains of laboratory animals using CRISPR/Cas9 technology has dramatically accelerated the interrogation of gene function in vivo. So far, the dominant approach for genotyping of genome-modified animals has been the T7E1 endonuclease cleavage assay. Here, we present a polyacrylamide gel electrophoresis-based (PAGE) method to genotype mice harboring different types of indel mutations. We developed 6 strains of genome-modified mice using CRISPR/Cas9 system, and utilized this approach to genotype mice from F0 to F2 generation, which included single and multiplexed genome-modified mice. We also determined the maximal detection sensitivity for detecting mosaic DNA using PAGE-based assay as 0.5%. We further applied PAGE-based genotyping approach to detect CRISPR/Cas9-mediated on- and off-target effect in human 293T and induced pluripotent stem cells (iPSCs). Thus, PAGE-based genotyping approach meets the rapidly increasing demand for genotyping of the fast-growing number of genome-modified animals and human cell lines created using CRISPR/Cas9 system or other nuclease systems such as TALEN or ZFN.

  16. Transplantation of NGF-gene-modified bone marrow stromal cells into a rat model of Alzheimer' disease.

    PubMed

    Li, Li-Yan; Li, Jin-Tao; Wu, Qing-Ying; Li, Jin; Feng, Zhong-Tang; Liu, Su; Wang, Ting-Hua

    2008-02-01

    It is well known that bone marrow stromal cells (BMSC) grafted into the hippocampus of the rat model of Alzheimer's disease (AD) could survive and differentiate into cholinergic neurons as well as contribute towards functional restoration. The present study evaluated the effects of BMSC as a seed cell modified by nerve growth factor (NGF) gene into the hippocampus of AD rats. The beta-amyloid protein was injected bilaterally into the rat hippocampus to reproduce the AD model. After the human total RNA was extracted, the NGF gene was amplified by reverse transcription-polymerase chain reaction, then cloned into the pcDNA3. BMSC derived from a green fluorescence protein transgenic mouse were isolated, cultured, identified, and transfected by the NGF recombinant. The NGF-gene-modified BMSC were then transplanted into the hippocampus of AD rats. The results showed that implanted BMSC survived, migrated and expressed NGF as well as differentiated into ChAT-positive neurons. A significant improvement in learning and memory in AD rats was also seen in NGF-gene-modified BMSC group, when compared with the BMSC group. The present findings suggested that BMSC provided an effective carrier for delivery of NGF into AD rats, and the administration of NGF-gene-modified BMSC may be considered as a potential strategy for the development of effective therapies for the treatment of AD.

  17. An autoantibody is modified for use as a delivery system to target the cell nucleus: therapeutic implications.

    PubMed

    Weisbart, R H; Stempniak, M; Harris, S; Zack, D J; Ferreri, K

    1998-10-01

    A murine monoclonal anti-dsDNA antibody was found to penetrate living cells and localize in the nucleus without pathologic effects. A single mutation in VH markedly enhanced cellular penetration. The mutant antibody was produced as recombinant Fab and single chain antibody fragments to investigate its use as a delivery system to target the cell nucleus. Complexes were made containing Fab fragments and alkaline phosphatase conjugated goat antibodies to mouse |gk chains. Fab fragments transported 305 kDa goat antibody-enzyme complexes into the nucleus in COS-7 and CHO cells. A single chain antibody cDNA was constructed by splice overlap extension PCR and expressed in COS-7 cells. Binding of the single chain antibody to dsDNA was shown by ELISA, and cellular penetration and nuclear localization were demonstrated in COS-7 and CHO cells. The single chain antibody cDNA was ligated into the expression vector, pEGFP, to produce a fusion protein with green fluorescent protein. The fusion protein penetrated COS-7 cells and localized in the cell nucleus. The single chain antibody produced during sustained expression in CHO cells re-entered antibody-producing cells and localized in the nucleus without affecting cell viability. Our results demonstrate the potential use of a modified autoantibody as a delivery system to target the cell nucleus.

  18. Generation of clinical-grade CD19-specific CAR-modified CD8+ memory stem cells for the treatment of human B-cell malignancies.

    PubMed

    Sabatino, Marianna; Hu, Jinhui; Sommariva, Michele; Gautam, Sanjivan; Fellowes, Vicki; Hocker, James D; Dougherty, Sean; Qin, Haiying; Klebanoff, Christopher A; Fry, Terry J; Gress, Ronald E; Kochenderfer, James N; Stroncek, David F; Ji, Yun; Gattinoni, Luca

    2016-07-28

    Long-lived, self-renewing, multipotent T memory stem cells (TSCM) can trigger profound and sustained tumor regression but their rareness poses a major hurdle to their clinical application. Presently, clinically compliant procedures to generate relevant numbers of this T-cell population are undefined. Here, we provide a strategy for deriving large numbers of clinical-grade tumor-redirected TSCM starting from naive precursors. CD8(+)CD62L(+)CD45RA(+) naive T cells enriched by streptamer-based serial-positive selection were activated by CD3/CD28 engagement in the presence of interleukin-7 (IL-7), IL-21, and the glycogen synthase-3β inhibitor TWS119, and genetically engineered to express a CD19-specific chimeric antigen receptor (CD19-CAR). These conditions enabled the generation of CD19-CAR-modified CD8(+) TSCM that were phenotypically, functionally, and transcriptomically equivalent to their naturally occurring counterpart. Compared with CD8(+) T cells generated with clinical protocols currently under investigation, CD19-CAR-modified CD8(+) TSCM exhibited enhanced metabolic fitness and mediated robust, long-lasting antitumor responses against systemic acute lymphoblastic leukemia xenografts. This clinical-grade platform provides the basis for a phase 1 trial evaluating the activity of CD19-CAR-modified CD8(+) TSCM in patients with B-cell malignancies refractory to prior allogeneic hematopoietic stem cell transplantation.

  19. Quasi-solid electrolyte with polyamidoamine dendron modified-talc applied to dye-sensitized solar cells

    NASA Astrophysics Data System (ADS)

    Andrade, Marcos A. S.; Nogueira, Ana F.; Miettunen, Kati; Tiihonen, Armi; Lund, Peter D.; Pastore, Heloise O.

    2016-09-01

    A sequence of generations of polyamidoamine dendron modified-talc, PAMAM-talc-Gn (n = 1, 3, 5 and 7), is proposed as additive in a composite gel electrolyte for dye-sensitized solar cells. Polyiodides are intercalated into the organotalc interlamellar space by adsorption of iodine vapor, producing triiodide and polyiodides. We investigate the effect of organotalc content on the charge transport in the electrolyte and solar cell performance and optimize the organotalc content. Without the previous adsorption of iodine molecules, the organotalcs appear to remove iodine from the electrolyte solution decreasing device's performance significantly. Instead, the samples with additional iodide had higher Jsc and efficiency approaching the values of the reference cells containing liquid, which suggests that this kind of gelling method would be suitable for dye solar cells. Charge transport in the gel electrolyte is investigated with electrochemical impedance spectroscopy and cyclic voltammetry analyses using symmetrical CE-CE electrochemical cells.

  20. Genomic Modifiers of Natural Killer Cells, Immune Responsiveness and Lymphoid Tissue Remodeling Together Increase Host Resistance to Viral Infection

    PubMed Central

    Lee, Heather; Prince, Jessica; Stadnisky, Michael D.; Anderson, Monique; Nash, William; Rival, Claudia; Wei, Hairong; Gamache, Awndre; Farber, Charles R.; Tung, Kenneth; Brown, Michael G.

    2016-01-01

    The MHC class I Dk molecule supplies vital host resistance during murine cytomegalovirus (MCMV) infection. Natural killer (NK) cells expressing the Ly49G2 inhibitory receptor, which specifically binds Dk, are required to control viral spread. The extent of Dk-dependent host resistance, however, differs significantly amongst related strains of mice, C57L and MA/My. As a result, we predicted that relatively small-effect modifier genetic loci might together shape immune cell features, NK cell reactivity, and the host immune response to MCMV. A robust Dk-dependent genetic effect, however, has so far hindered attempts to identify additional host resistance factors. Thus, we applied genomic mapping strategies and multicolor flow cytometric analysis of immune cells in naive and virus-infected hosts to identify genetic modifiers of the host immune response to MCMV. We discovered and validated many quantitative trait loci (QTL); these were mapped to at least 19 positions on 16 chromosomes. Intriguingly, one newly discovered non-MHC locus (Cmv5) controlled splenic NK cell accrual, secondary lymphoid organ structure, and lymphoid follicle development during MCMV infection. We infer that Cmv5 aids host resistance to MCMV infection by expanding NK cells needed to preserve and protect essential tissue structural elements, to enhance lymphoid remodeling and to increase viral clearance in spleen. PMID:26845690

  1. The labeling of stem cells by superparamagnetic iron oxide nanoparticles modified with PEG/PVP or PEG/PEI.

    PubMed

    Yang, Gao; Ma, Weiqiong; Zhang, Baolin; Xie, Qi

    2016-05-01

    Poly(ethylene glycol) (PEG) and poly(vinyl pyrrolidone) (PVP) co-modified superparamagnetic iron oxide nanoparticles (SPIONs) (PEG/PVP-SPIONs), and PEG and poly(ethylene imine) (PEI) co-modified SPIONs (PEG/PEI-SPIONs) synthesized by thermal decomposition have been used as magnetic resonance imaging (MRI) contrast agents to label adipose-derived stem cells (ADSCs). Efficient cell labeling was achieved after incubation with PEG/PVP-SPIONs and PEG/PEI-SPIONs for 12h, and the MRI of labeled cells was evaluated. The cell viability tests showed the low cytotoxicity of PEG/PVP-SPIONs and PEG/PEI-SPIONs. The cellular iron content incubated with PEG/PVP-SPIONs at a concentration of 25 μg/ml was 6.96 pg/cell, the cellular iron contents incubated with PEG/PEI-SPIONs at concentrations of 12 and 25 μg/ml were 20.16, 35.4 pg/cell, respectively. The SPIONs were located predominantly in the intracellular vesicles. The cellular iron oxide uptake was significantly high after incubation with PEG/PEI-SPIONs as compared with the commercial iron oxide agents (Feridex, Feridex@PLL, Resovist and Resovist@PLL) reported. This work demonstrates that PEG/PEI-SPIONs are the competent agents for the labeling of ADSCs.

  2. On-line observation of cell growth in a three-dimensional matrix on surface-modified microelectrode arrays.

    PubMed

    Lin, Shu-Ping; Kyriakides, Themis R; Chen, Jia-Jin J

    2009-06-01

    Despite many successful applications of microelectrode arrays (MEAs), typical two-dimensional in-vitro cultures do not project the full scale of the cell growth environment in the three-dimensional (3D) in-vivo setting. This study aims to on-line monitor in-vitro cell growth in a 3D matrix on the surface-modified MEAs with a dynamic perfusion culture system. A 3D matrix consisting of poly(ethylene glycol) hydrogel supplemented with poly-D-lysine was subsequently synthesized in situ on the self-assembled monolayer modified MEAs. FTIR spectrum analysis revealed a peak at 2100 cm(-1) due to the degradation of the structure of the 3D matrix. After 2 wks, microscopic examination revealed that the non-degraded area was around 1500 microm(2) and provided enough space for cell growth. Fluorescence microscopy revealed that the degraded 3D matrix was non-cytotoxic allowing the growth of NIH3T3 fibroblasts and cortical neurons in vitro. Time-course changes of total impedance including resistance and reactance were recorded for 8 days to evaluate the cell growth in the 3D matrix on the MEA. A consistent trend reflecting changes of reactance and total impedance was observed. These in-vitro assays demonstrate that our 3D matrix can construct a biomimetic system for cell growth and analysis of cell surface interactions.

  3. A modified multiparametric assay using HepaRG cells for predicting the degree of drug-induced liver injury risk.

    PubMed

    Tomida, Takafumi; Okamura, Hayao; Yokoi, Tsuyoshi; Konno, Yoshihiro

    2017-03-01

    The approach for predicting the degree of drug-induced liver injury (DILI) risk was investigated quantitatively in a modified multiparametric assay using HepaRG cells. Thirty-eight drugs were classified by DILI risk into five categories based on drug labels approved by the Food and Drug Administration (FDA) as follows: withdrawn (WDN), boxed warning (BW), warnings and precautions (WP), adverse reactions (AR), and no match (NM). Also, WP was classified into two categories: high and low concern. Differentiated HepaRG cells were treated with drugs for 24 h. The maximum concentration was set at 100-fold the therapeutic maximum plasma concentration (Cmax ). After treatment with drugs, the cell viability, glutathione content, caspase 3/7 activity, lactate dehydrogenase leakage and albumin secretion were measured. As modified cut-off values of each parameter, the TC50 (toxic concentration that decreased the response by 50%) and EC200 (effective concentration giving a response equal to 200% of controls) were calculated. In addition, the toxicity score (total sum score of the cytotoxic level of each parameter) was calculated. This modified multiparametric assay showed an 87% sensitivity and 87% specificity for predicting the DILI risk. The toxicity score showed a good predictive performance for WDN, BW and WP (high concern) categories [cut-off: score ≥ 1; area under a receiver operating characteristic curve (ROC-AUC): 0.88], and for WDN and BW categories (cut-off: score ≥ 3; ROC-AUC: 0.88). This study newly indicated that the degree of DILI risk might be predictable quantitatively by assessing the toxicity score in the modified multiparametric assay using HepaRG cells. Copyright © 2016 John Wiley & Sons, Ltd.

  4. Demineralized Bone Matrix Scaffolds Modified by CBD-SDF-1α Promote Bone Regeneration via Recruiting Endogenous Stem Cells.

    PubMed

    Shi, Jiajia; Sun, Jie; Zhang, Wen; Liang, Hui; Shi, Qin; Li, Xiaoran; Chen, Yanyan; Zhuang, Yan; Dai, Jianwu

    2016-10-07

    The reconstruction of bone usually depends on substitute transplantation, which has drawbacks including the limited bone substitutes available, comorbidity, immune rejection, and limited endogenous bone regeneration. Here, we constructed a functionalized bone substitute by combining application of the demineralized bone matrix (DBM) and collagen-binding stromal-cell-derived factor-1α (CBD-SDF-1α). DBM was a poriferous and biodegradable bone substitute, derived from bovine bone and consisting mainly of collagen. CBD-SDF-1α could bind to collagen and be controllably released from the DBM to mobilize stem cells. In a rat femur defect model, CBD-SDF-1α-modified DBM scaffolds could efficiently mobilize CD34(+) and c-kit(+) endogenous stem cells homing to the injured site at 3 days after implantation. According to the data from micro-CT, CBD-SDF-1α-modified DBM scaffolds could help the bone defects rejoin with mineralization accumulated and bone volume expanded. Interestingly, osteoprotegerin (OPG) and osteopontin (OPN) were highly expressed in CBD-SDF-1α group at an early time after implantation, while osteocalcin (OCN) was more expanded. H&E and Masson's trichrome staining showed that the CBD-SDF-1α-modified DBM scaffold group had more osteoblasts and that the bone defect rejoined earlier. The ultimate strength of the regenerated bone was investigated by three-point bending, showing that the CBD-SDF-1α group had superior strength. In conclusion, CBD-SDF-1α-modified DBM scaffolds could promote bone regeneration by recruiting endogenous stem cells.

  5. Activated Ion ETD Performed in a Modified Collision Cell on a Hybrid QLT-Oribtrap Mass Spectrometer

    NASA Astrophysics Data System (ADS)

    Ledvina, Aaron R.; Rose, Christopher M.; McAlister, Graeme C.; Syka, John E. P.; Westphall, Michael S.; Griep-Raming, Jens; Schwartz, Jae C.; Coon, Joshua J.

    2013-11-01

    We describe the implementation and characterization of activated ion electron transfer dissociation (AI-ETD) on a hybrid QLT-Orbitrap mass spectrometer. AI-ETD was performed using a collision cell that was modified to enable ETD reactions, in addition to normal collisional activation. The instrument manifold was modified to enable irradiation of ions along the axis of this modified cell with IR photons from a CO2 laser. Laser power settings were optimized for both charge (z) and mass to charge ( m/z) and the instrument control firmware was updated to allow for automated adjustments to the level of irradiation. This implementation of AI-ETD yielded 1.6-fold more unique identifications than ETD in an nLC-MS/MS analysis of tryptic yeast peptides. Furthermore, we investigated the application of AI-ETD on large scale analysis of phosphopeptides, where laser power aids ETD, but can produce b- and y-type ions because of the phosphoryl moiety's high IR adsorption. nLC-MS/MS analysis of phosphopeptides derived from human embryonic stem cells using AI-ETD yielded 2.4-fold more unique identifications than ETD alone, demonstrating a promising advance in ETD sequencing of PTM containing peptides.

  6. Activated ion ETD performed in a modified collision cell on a hybrid QLT-Oribtrap mass spectrometer.

    PubMed

    Ledvina, Aaron R; Rose, Christopher M; McAlister, Graeme C; Syka, John E P; Westphall, Michael S; Griep-Raming, Jens; Schwartz, Jae C; Coon, Joshua J

    2013-11-01

    We describe the implementation and characterization of activated ion electron transfer dissociation (AI-ETD) on a hybrid QLT-Orbitrap mass spectrometer. AI-ETD was performed using a collision cell that was modified to enable ETD reactions, in addition to normal collisional activation. The instrument manifold was modified to enable irradiation of ions along the axis of this modified cell with IR photons from a CO2 laser. Laser power settings were optimized for both charge (z) and mass to charge (m/z) and the instrument control firmware was updated to allow for automated adjustments to the level of irradiation. This implementation of AI-ETD yielded 1.6-fold more unique identifications than ETD in an nLC-MS/MS analysis of tryptic yeast peptides. Furthermore, we investigated the application of AI-ETD on large scale analysis of phosphopeptides, where laser power aids ETD, but can produce b- and y-type ions because of the phosphoryl moiety's high IR adsorption. nLC-MS/MS analysis of phosphopeptides derived from human embryonic stem cells using AI-ETD yielded 2.4-fold more unique identifications than ETD alone, demonstrating a promising advance in ETD sequencing of PTM containing peptides.

  7. Activated Ion ETD Performed in a Modified Collision Cell on a Hybrid QLT-Oribtrap Mass Spectrometer

    PubMed Central

    Ledvina, Aaron R.; Rose, Christopher M.; McAlister, Graeme C.; Syka, John E.P.; Westphall, Michael S.; Griep-Raming, Jens; Schwartz, Jae C.; Coon, Joshua J.

    2013-01-01

    We describe the implementation and characterization of activated ion electron transfer dissociation (AI-ETD) on a hybrid QLT-Orbitrap mass spectrometer. AI-ETD was performed using a collision cell that was modified to enable ETD reactions, in addition to normal collisional activation. The instrument manifold was modified to enable irradiation of ions along the axis of this modified cell with IR photons from a CO2 laser. Laser power settings were optimized for both charge (z) and mass to charge (m/z) and the instrument control firmware was updated to allow for automated adjustments to the level of irradiation. This implementation of AI-ETD yielded 1.6 fold more unique identifications than ETD in an nLC-MS/MS analysis of tryptic yeast peptides. Furthermore, we investigated the application of AI-ETD on large scale analysis of phosphopeptides, where laser power aids ETD, but can produce b- and y-type ions due to the phosphoryl moiety's high IR adsorption. nLC-MS/MS analysis of phosphopeptides derived from human embryonic stem cells using AI-ETD yielded 2.4 fold more unique identifications than ETD alone, demonstrating a promising advance in ETD sequencing of PTM containing peptides. PMID:23677544

  8. RGD peptide-modified dendrimer-entrapped gold nanoparticles enable highly efficient and specific gene delivery to stem cells.

    PubMed

    Kong, Lingdan; Alves, Carla S; Hou, Wenxiu; Qiu, Jieru; Möhwald, Helmuth; Tomás, Helena; Shi, Xiangyang

    2015-03-04

    We report the use of arginine-glycine-aspartic (Arg-Gly-Asp, RGD) peptide-modified dendrimer-entrapped gold nanoparticles (Au DENPs) for highly efficient and specific gene delivery to stem cells. In this study, generation 5 poly(amidoamine) dendrimers modified with RGD via a poly(ethylene glycol) (PEG) spacer and with PEG monomethyl ether were used as templates to entrap gold nanoparticles (AuNPs). The native and the RGD-modified PEGylated dendrimers and the respective well characterized Au DENPs were used as vectors to transfect human mesenchymal stem cells (hMSCs) with plasmid DNA (pDNA) carrying both the enhanced green fluorescent protein and the luciferase (pEGFPLuc) reporter genes, as well as pDNA encoding the human bone morphogenetic protein-2 (hBMP-2) gene. We show that all vectors are capable of transfecting the hMSCs with both pDNAs. Gene transfection using pEGFPLuc was demonstrated by quantitative Luc activity assay and qualitative evaluation by fluorescence microscopy. For the transfection with hBMP-2, the gene delivery efficiency was evaluated by monitoring the hBMP-2 concentration and the level of osteogenic differentiation of the hMSCs via alkaline phosphatase activity, osteocalcin secretion, calcium deposition, and von Kossa staining assays. Our results reveal that the stem cell gene delivery efficiency is largely dependent on the composition and the surface functionality of the dendrimer-based vectors. The coexistence of RGD and AuNPs rendered the designed dendrimeric vector with specific stem cell binding ability likely via binding of integrin receptor on the cell surface and improved three-dimensional conformation of dendrimers, which is beneficial for highly efficient and specific stem cell gene delivery applications.

  9. Guidelines by the AAPM and GEC-ESTRO on the use of innovative brachytherapy devices and applications: Report of Task Group 167.

    PubMed

    Nath, Ravinder; Rivard, Mark J; DeWerd, Larry A; Dezarn, William A; Thompson Heaton, H; Ibbott, Geoffrey S; Meigooni, Ali S; Ouhib, Zoubir; Rusch, Thomas W; Siebert, Frank-André; Venselaar, Jack L M

    2016-06-01

    Although a multicenter, Phase III, prospective, randomized trial is the gold standard for evidence-based medicine, it is rarely used in the evaluation of innovative devices because of many practical and ethical reasons. It is usually sufficient to compare the dose distributions and dose rates for determining the equivalence of the innovative treatment modality to an existing one. Thus, quantitative evaluation of the dosimetric characteristics of innovative radiotherapy devices or applications is a critical part in which physicists should be actively involved. The physicist's role, along with physician colleagues, in this process is highlighted for innovative brachytherapy devices and applications and includes evaluation of (1) dosimetric considerations for clinical implementation (including calibrations, dose calculations, and radiobiological aspects) to comply with existing societal dosimetric prerequisites for sources in routine clinical use, (2) risks and benefits from a regulatory and safety perspective, and (3) resource assessment and preparedness. Further, it is suggested that any developed calibration methods be traceable to a primary standards dosimetry laboratory (PSDL) such as the National Institute of Standards and Technology in the U.S. or to other PSDLs located elsewhere such as in Europe. Clinical users should follow standards as approved by their country's regulatory agencies that approved such a brachytherapy device. Integration of this system into the medical source calibration infrastructure of secondary standard dosimetry laboratories such as the Accredited Dosimetry Calibration Laboratories in the U.S. is encouraged before a source is introduced into widespread routine clinical use. The American Association of Physicists in Medicine and the Groupe Européen de Curiethérapie-European Society for Radiotherapy and Oncology (GEC-ESTRO) have developed guidelines for the safe and consistent application of brachytherapy using innovative devices and

  10. Genetically Modified T Cells in Treating Patients With Stage III-IV Non-small Cell Lung Cancer or Mesothelioma

    ClinicalTrials.gov

    2017-01-04

    Advanced Pleural Malignant Mesothelioma; HLA-A*0201 Positive Cells Present; Recurrent Non-Small Cell Lung Carcinoma; Recurrent Pleural Malignant Mesothelioma; Stage III Non-Small Cell Lung Cancer; Stage III Pleural Mesothelioma; Stage IIIA Non-Small Cell Lung Cancer; Stage IIIB Non-Small Cell Lung Cancer; Stage IV Non-Small Cell Lung Cancer; Stage IV Pleural Mesothelioma

  11. Improved mucoadhesion and cell uptake of chitosan and chitosan oligosaccharide surface-modified polymer nanoparticles for mucosal delivery of proteins.

    PubMed

    Dyawanapelly, Sathish; Koli, Uday; Dharamdasani, Vimisha; Jain, Ratnesh; Dandekar, Prajakta

    2016-08-01

    The main aim of the present study was to compare mucoadhesion and cellular uptake efficiency of chitosan (CS) and chitosan oligosaccharide (COS) surface-modified polymer nanoparticles (NPs) for mucosal delivery of proteins. We have developed poly (D, L-lactide-co-glycolide) (PLGA) NPs, surface-modified COS-PLGA NPs and CS-PLGA NPs, by using double emulsion solvent evaporation method, for encapsulating bovine serum albumin (BSA) as a model protein. Surface modification of NPs was confirmed using physicochemical characterization methods such as particle size and zeta potential, SEM, TEM and FTIR analysis. Both surface-modified PLGA NPs displayed a slow release of protein compared to PLGA NPs. Furthermore, we have explored the mucoadhesive property of COS as a material for modifying the surface of polymeric NPs. During in vitro mucoadhesion test, positively charged COS-PLGA NPs and CS-PLGA NPs exhibited enhanced mucoadhesion, compared to negatively charged PLGA NPs. This interaction was anticipated to improve the cell interaction and uptake of NPs, which is an important requirement for mucosal delivery of proteins. All nanoformulations were found to be safe for cellular delivery when evaluated in A549 cells. Moreover, intracellular uptake behaviour of FITC-BSA loaded NPs was extensively investigated by confocal laser scanning microscopy and flow cytometry. As we hypothesized, positively charged COS-PLGA NPs and CS-PLGA NPs displayed enhanced intracellular uptake compared to negatively charged PLGA NPs. Our results demonstrated that CS- and COS-modified polymer NPs could be promising carriers for proteins, drugs and nucleic acids via nasal, oral, buccal, ocular and vaginal mucosal routes.

  12. Semi-automated closed system manufacturing of lentivirus gene-modified haematopoietic stem cells for gene therapy

    PubMed Central

    Adair, Jennifer E.; Waters, Timothy; Haworth, Kevin G.; Kubek, Sara P.; Trobridge, Grant D.; Hocum, Jonah D.; Heimfeld, Shelly; Kiem, Hans-Peter

    2016-01-01

    Haematopoietic stem cell (HSC) gene therapy has demonstrated potential to treat many diseases. However, current state of the art requires sophisticated ex vivo gene transfer in a dedicated Good Manufacturing Practices facility, limiting availability. An automated process would improve the availability and standardized manufacture of HSC gene therapy. Here, we develop a novel program for semi-automated cell isolation and culture equipment to permit complete benchtop generation of gene-modified CD34+ blood cell products for transplantation. These cell products meet current manufacturing quality standards for both mobilized leukapheresis and bone marrow, and reconstitute human haematopoiesis in immunocompromised mice. Importantly, nonhuman primate autologous gene-modified CD34+ cell products are capable of stable, polyclonal multilineage reconstitution with follow-up of more than 1 year. These data demonstrate proof of concept for point-of-care delivery of HSC gene therapy. Given the many target diseases for gene therapy, there is enormous potential for this approach to treat patients on a global scale. PMID:27762266

  13. Hysteretic Behavior upon Light Soaking in Perovskite Solar Cells Prepared via Modified Vapor-Assisted Solution Process.

    PubMed

    Liu, Chong; Fan, Jiandong; Zhang, Xing; Shen, Yanjiao; Yang, Lin; Mai, Yaohua

    2015-05-06

    Recently, the organic-inorganic hybrid perovskite solar cells exhibit rapidly rising efficiencies, while anomalous hysteresis in perovskite solar cells remains unsolvable. Herein, a high-quality perovskite thin film is prepared by a modified vapor-assisted solution process, which is a simple but well-controllable method proven to be capable of producing a thin film with full surface coverage and grain size up to micrometers. The as-fabricated perovskite solar cell has efficiency as high as 10.2%. The hysteresis effects of both planar and mesoscopic TiO2-based perovskite solar cells have been comprehensively studied upon illumination. The results demonstrate that mesoporous-based perovskite cells combined with remarkable grain size are subject to alleviating the hysteresis effects in comparison to the planar cells. Likewise, mesoscopic TiO2-based perovskite cells perform independently of illumination and bias conditions prior to the measurements, whereas the planar cells display a reversible behavior of illumination and applied bias-dependent I-V curves. The present study would refer strip road for the stability study of the perovskite solar cells.

  14. Control of growth and squamous differentiation in normal human bronchial epithelial cells by chemical and biological modifiers and transferred genes.

    PubMed Central

    Pfeifer, A M; Lechner, J F; Masui, T; Reddel, R R; Mark, G E; Harris, C C

    1989-01-01

    The majority of human lung cancers arise from bronchial epithelial cells. The normal pseudostratified bronchial epithelium is composed of basal, mucous, and ciliated cells. This multi-differentiated epithelium usually responds to xenobiotics and physical injury by undergoing basal cell hyperplasia, mucous cell hyperplasia, and squamous metaplasia. One step of the multistage process of carcinogenesis is thought to involve aberrations in control of the squamous metaplastic processes. Decreased responsiveness to regulators of terminal squamous differentiation may confer a selective clonal expansion advantage to an initiated cell. We studied the effects of endogenous [e.g., transforming growth factor beta 1 (TGF-beta 1) and serum] and exogenous [e.g., 12-O-tetradecanoyl-13-phorbol-acetate (TPA), tobacco smoke condensate, and aldehydes] modifiers of normal human bronchial epithelial (NHBE) cell in a serum-free culture system. NHBE cells are growth inhibited by all of these compounds and induced to undergo squamous differentiation by TGF-beta 1 or TPA. In contrast, lung carcinoma cell lines are relatively resistant to inducers of terminal squamous differentiation which may provide them with a selective growth advantage. Chemical agents and activated protooncogenes (ras,raf,myc) altered the response to endogenous and exogenous inducers of squamous differentiation and caused extended cellular lifespan, aneuploidy, and/or tumorigenicity. The data suggest a close relationship between dysregulation of terminal differentiation pathways and neoplastic transformation of human bronchial epithelial cells. PMID:2538323

  15. Polysaccharide-degrading Enzymes are Unable to Attack Plant Cell Walls without Prior Action by a “Wall-modifying Enzyme” 1

    PubMed Central

    Karr, Arthur L.; Albersheim, Peter

    1970-01-01

    A study of the degradation of plant cell walls by the mixture of enzymes present in Pectinol R-10 is described. A “wall-modifying enzyme” has been purified from this mixture by a combination of diethylaminoethyl cellulose, Bio Gel P-100, and carboxymethyl cellulose chromatography. Treatment of cell walls with the “wall-modifying enzyme” is shown to be a necessary prerequisite to wall degradation catalyzed by a mixture of polysaccharide-degrading enzymes prepared from Pectinol R-10 or by an α-galactosidase secreted by the pathogenic fungus Colletotrichum lindemuthianum. The action of the “wall-modifying enzyme” on cell walls is shown to result in both a release of water-soluble, 70% ethanol-insoluble polymers and an alteration of the residual cell wall. A purified preparation of the “wall-modifying enzyme” is unable to degrade a wide variety of polysaccharide, glycoside, and peptide substrates. However, the purified preparation of wall-modifying enzyme has a limited ability to degrade polygalacturonic acid. The fact that polygalacturonic acid inhibits the ability of the “wall-modifying enzyme” to affect cell walls suggests that the “wall-modifying enzyme” may be responsible for the limited polygalacturonic acid-degrading activity present in the purified preparation. The importance of a wall-modifying enzyme in developmental processes and in pathogenesis is discussed. PMID:16657425

  16. T Lymphocyte Antigen 4-Modified Dendritic Cell Therapy for Asthmatic Mice Guided by the CCR7 Chemokine Receptor

    PubMed Central

    Chen, Yan; Wang, Yongming; Fu, Zhou

    2014-01-01

    The CD80/CD86-CD28 axis is a critical pathway for immuno-corrective therapy, and the cytotoxic T lymphocyte antigen 4 (CTLA4) is a promising immunosuppressor targeting the CD80/CD86-CD28 axis; however, its use for asthma therapy needs further optimization. A human CTLA4 fused with the IgCγ Fc (CTLA4Ig) and mouse CC chemokine receptor type7 (CCR7) coding sequences were inserted into a recombinant adenovirus (rAdV) vector to generate rAdV-CTLA4Ig and rAdV-CCR7. The naive dendritic cells (DCs) were infected with these rAdVs to ensure CCR7 and CTLA4Ig expression. The therapeutic effects of modified DCs were evaluated. rAdV-CTLA4Ig and rAdV-CCR7 infected DCs improved all asthma symptoms. Inflammatory cell infiltration and cytokine analysis showed that rAdV-CTLA4Ig and rAdV-CCR7-modified DC therapy reduced the number of eosinophils and lymphocyte and neutrophil infiltration in the lung. Interestingly, assessment of the humoral immunity showed that the IL-4 and IFNγ levels of the rAdV-CTLA4Ig and rAdV-CCR7-modified DC-treated mice decreased significantly and did not reverse the Th1/Th2 balance. DCs expressing CCR7 displayed guidance ability for DC migration, primarily for DCs in the inflammatory lung. Additionally, the rAdVs caused an inflammatory response by inducing DC differentiation, inflammatory cell infiltration and changes in cytokines; however, mice transplanted with rAdV-green fluorescent protein (GFP)-infected DCs displayed no asthma manifestations. In conclusion, CTLA4Ig-modified DCs exhibited a therapeutic effect on asthma, and CCR7 may guide DC homing. The combination of these two molecules may be a model for precision-guided immunotherapy. PMID:25177863

  17. T lymphocyte antigen 4-modified dendritic cell therapy for asthmatic mice guided by the CCR7 chemokine receptor.

    PubMed

    Chen, Yan; Wang, Yongming; Fu, Zhou

    2014-08-29

    The CD80/CD86-CD28 axis is a critical pathway for immuno-corrective therapy, and the cytotoxic T lymphocyte antigen 4 (CTLA4) is a promising immunosuppressor targeting the CD80/CD86-CD28 axis; however, its use for asthma therapy needs further optimization. A human CTLA4 fused with the IgCγ Fc (CTLA4Ig) and mouse CC chemokine receptor type7 (CCR7) coding sequences were inserted into a recombinant adenovirus (rAdV) vector to generate rAdV-CTLA4Ig and rAdV-CCR7. The naive dendritic cells (DCs) were infected with these rAdVs to ensure CCR7 and CTLA4Ig expression. The therapeutic effects of modified DCs were evaluated. rAdV-CTLA4Ig and rAdV-CCR7 infected DCs improved all asthma symptoms. Inflammatory cell infiltration and cytokine analysis showed that rAdV-CTLA4Ig and rAdV-CCR7-modified DC therapy reduced the number of eosinophils and lymphocyte and neutrophil infiltration in the lung. Interestingly, assessment of the humoral immunity showed that the IL-4 and IFNγ levels of the rAdV-CTLA4Ig and rAdV-CCR7-modified DC-treated mice decreased significantly and did not reverse the Th1/Th2 balance. DCs expressing CCR7 displayed guidance ability for DC migration, primarily for DCs in the inflammatory lung. Additionally, the rAdVs caused an inflammatory response by inducing DC differentiation, inflammatory cell infiltration and changes in cytokines; however, mice transplanted with rAdV-green fluorescent protein (GFP)-infected DCs displayed no asthma manifestations. In conclusion, CTLA4Ig-modified DCs exhibited a therapeutic effect on asthma, and CCR7 may guide DC homing. The combination of these two molecules may be a model for precision-guided immunotherapy.

  18. [Modified Mechanism of Cell Walls from Chinese Fir Treated with Low-Molecular-Weight Phenol Formaldehyde Resin].

    PubMed

    Huang, Yan-hui; Fei, Ben-hua; Zhao, Rong-jun

    2015-12-01

    Study on the modified mechanism of wood cell walls, it is very important for improving treatment reagents, optimizing treatment technology, and enhancing wood density, mechanical properties, dimensional stability, and so on. Samples of plantation Chinese fir were treated gradually with synthesized water-soluble low-molecular-weight phenol formaldehyde (PF) resins under vacuum and pressure. The correlated physical and chemical properties of the treated and untreated reference samples were determined by X-ray diffractometer (XRD), Fourier transform infrared spectrometer (FTIR), and nuclear magnetic resonance spectrometer(NMR) (Using method of Cross Polarization/Magic Angle Spinning for continuous testing) with high precision and resolution. The results showed that, after treated with water-soluble low-molecular-weight PF resin, the average values of crystallinity from the treated samples were decreased obviously, and the average reduction rate was 12.67%, 11.91% and 6.26%, respectively. Comparing water-soluble, low-molecular-weight PF resin modified Chinese fir with untreated reference samples, no new chemical shifts and characteristic peaks of functional groups from esters, ethers, etc. were present by using FTIR and ¹³C NMR spectrum. It was considered that there was no distinct chemical reaction between the water-soluble low-molecular-weight PF resin and Chinese Fir cell walls. But water-soluble low-molecular-weight PF resin could enter into the structure relatively loose, large size spaces, relatively area large amorphous regions in cell walls of Chinese fir tracheids, and form physical filling, which resulting in the decreasing of relative crystallinity. This study has important reference value for the development of new wood modification reagents and the optimization of wood modification process. The findings also provide important theoretical foundation for further proving the modification mechanisms of wood cell walls and enriching the modified theories of

  19. Regulated apoptosis of genetically modified hematopoietic stem and progenitor cells via an inducible caspase-9 suicide gene in rhesus macaques.

    PubMed

    Barese, Cecilia N; Felizardo, Tania C; Sellers, Stephanie E; Keyvanfar, Keyvan; Di Stasi, Antonio; Metzger, Mark E; Krouse, Allen E; Donahue, Robert E; Spencer, David M; Dunbar, Cynthia E

    2015-01-01

    The high risk of insertional oncogenesis reported in clinical trials using integrating retroviral vectors to genetically modify hematopoietic stem and progenitor cells (HSPCs) requires the development of safety strategies to minimize risks associated with novel cell and gene therapies. The ability to ablate genetically modified cells in vivo is desirable, should an abnormal clone emerge. Inclusion of "suicide genes" in vectors to facilitate targeted ablation of vector-containing abnormal clones in vivo is one potential safety approach. We tested whether the inclusion of the "inducible Caspase-9" (iCasp9) suicide gene in a gamma-retroviral vector facilitated efficient elimination of vector-containing HSPCs and their hematopoietic progeny in vivo long-term, in an autologous non-human primate transplantation model. Following stable engraftment of iCasp9 expressing hematopoietic cells in rhesus macaques, administration of AP1903, a chemical inducer of dimerization able to activate iCasp9, specifically eliminated vector-containing cells in all hematopoietic lineages long-term, suggesting activity at the HSPC level. Between 75% and 94% of vector-containing cells were eliminated by well-tolerated AP1903 dosing, but lack of complete ablation was linked to lower iCasp9 expression in residual cells. Further investigation of resistance mechanisms demonstrated upregulation of Bcl-2 in hematopoietic cell lines transduced with the vector and resistant to AP1903 ablation. These results demonstrate both the potential and the limitations of safety approaches using iCasp9 to HSPC-targeted gene therapy settings, in a model with great relevance to clinical development.

  20. Effect of novel chitosan-fluoroaluminosilicate resin modified glass ionomer cement supplemented with translationally controlled tumor protein on pulp cells.

    PubMed

    Wanachottrakul, Nattaporn; Chotigeat, Wilaiwan; Kedjarune-Leggat, Ureporn

    2014-04-01

    Dental materials that can promote cell proliferation and function is required for regenerative pulp therapy. Resin modified glass ionomer cement (RMGIC), a broadly used liner or restorative material, can cause apoptosis to pulp cells mainly due to HEMA (2-hydroxyethyl methacrylate), the released residual monomer. Recent studies found that chitosan and albumin could promote release of protein in GIC while translationally controlled tumor protein (TCTP) has an anti-apoptotic activity against HEMA. The aim of this study was to examine the effect of chitosan and albumin modified RMGIC (Exp-RMGIC) supplemented with TCTP on pulp cell viability and mineralization. Exp-RMGIC+TCTP was composed of RMGIC powder incorporated with 15 % of chitosan, 5 % albumin and supplemented with TCTP mixed with the same liquid components of RMGIC. The effect of each specimen on pulp cells was examined using the Transwell plate. From the MTT assay, Exp-RMGIC+TCTP had the highest percentages of viable cells (P < 0.05) at both 24 and 74 h. Flow cytometry revealed that, after 24 h, Exp-RMGIC+TCTP gave the lowest percentages of apoptotic cells compared to other groups. There was no difference in alkaline phosphatase (ALP) activity among different formula of the specimens, while cells cultured in media with TCTP had higher ALP activity. Von Kossa staining revealed that RMGIC+TCTP, and Exp-RMGIC+TCTP had higher percentages of calcium deposit area compared to those without TCTP. It was concluded that Exp-RMGIC supplemented with TCTP had less cytotoxicity than RMGIC and can protect cells from apoptosis better than RMGIC supplemented with TCTP.

  1. Gentamicin modified chitosan film with improved antibacterial property and cell biocompatibility.

    PubMed

    Liu, Yang; Ji, Peihong; Lv, Huilin; Qin, Yong; Deng, Linhong

    2017-05-01

    Gentamicin modified chitosan film (CS-GT) was produced using a three-step procedure comprising: (i) the chitosan solution was air-dried to form a chitosan (CS) film, (ii) using citric acid to generate the amide and carboxyl groups on the surface of CS, (iii) the CS with surface carboxyl groups was modified by grafting of gentamicin. After modification, this CS-GT film has excellent hydrophilicity and biocompatibility. It is very evident that the gentamicin grafting treatment significantly improves the antibacterial properties of the CS film. Our preliminary results suggest that this novel gentamicin modified chitosan film, which can be prepared in large quantities and at low cost, should have potential application in biomedical applications.

  2. Surface science and electrochemical studies of metal-modified carbides for fuel cells and hydrogen production

    NASA Astrophysics Data System (ADS)

    Kelly, Thomas Glenn

    Carbides of the early transition metals have emerged as low-cost catalysts that are active for a wide range of reactions. The surface chemistry of carbides can be altered by modifying the surface with small amounts of admetals. These metal-modified carbides can be effective replacements for Pt-based bimetallic systems, which suffer from the drawbacks of high cost and low thermal stability. In this dissertation, metal-modified carbides were studied for reactions with applications to renewable energy technologies. It is demonstrated that metal-modified carbides possess high activity for alcohol reforming and electrochemical hydrogen production. First, the surface chemistry of carbides towards alcohol decomposition is studied using density functional theory (DFT) and surface science experiments. The Vienna Ab initio Simulation Package (VASP) was used to calculate the binding energies of alcohols and decomposition intermediates on metal-modified carbides. The calculated binding energies were then correlated to reforming activity determined experimentally using temperature programmed desorption (TPD). In the case of methanol decomposition, it was found that tungsten monocarbide (WC) selectively cleaved the C-O bond to produce methane. Upon modifying the surface with a single layer of metal such as Ni, Pt, or Rh, the selectivity shifted towards scission of the C-H bonds while leaving the C-O bond intact, producing carbon monoxide (CO) and H2. High resolution energy loss spectroscopy (HREELS) was used to examine the bond breaking sequence as a function of temperature. From HREELS, it was shown that the surfaces followed an activity trend of Rh > Ni > Pt. The Au-modified WC surface possessed too low of a methanol binding energy, and molecular desorption of methanol was the most favorable pathway on this surface. Next, the ability of Rh-modified WC to break the C-C bond of C2 and C3 alcohols was demonstrated. HREELS showed that ethanol decomposed through an acetaldehyde

  3. Genetic modifiers of sickle cell anemia in the BABY HUG cohort: influence on laboratory and clinical phenotypes.

    PubMed

    Sheehan, Vivien A; Luo, Zhaoyu; Flanagan, Jonathan M; Howard, Thad A; Thompson, Bruce W; Wang, Winfred C; Kutlar, Abdullah; Ware, Russell E

    2013-07-01

    The recently completed BABY HUG trial investigated the safety and efficacy of hydroxyurea in infants with sickle cell anemia (SCA). To investigate the effects of known genetic modifiers, genomic DNA on 190 randomized subjects were analyzed for alpha thalassemia, beta-globin haplotype, polymorphisms affecting endogenous fetal hemoglobin (HbF) levels (XmnI, BCL11A, and HBS1L-MYB), UGT1A1 promoter polymorphisms, and the common G6PD A(-) mutation. At study entry, infants with alpha thalassemia trait had significantly lower mean corpuscular volume, total bilirubin, and absolute reticulocyte count. Beta-globin haplotypes associated with milder disease had significantly higher hemoglobin and %HbF. BCL11A and XmnI polymorphisms had significant effects on baseline HbF, while UGT1A1 promoter polymorphisms significantly influenced baseline serum bilirubin. At study exit, subjects randomized to placebo still exhibited laboratory effects of alpha thalassemia and other modifiers, while those assigned hydroxyurea had treatment effects that exceeded most genetic influences. The pain phenotype was influenced by HbF modifiers in both treatment groups. These data document that genetic polymorphisms do modify laboratory and clinical phenotypes even in very young patients with SCA. The hydroxyurea effects are more potent, however, indicating that treatment criteria should not be limited to certain genetic subsets, and supporting the use of hydroxyurea for all young patients with SCA.

  4. Transcellular transport and membrane insertion of the C5b-9 membrane attack complex of complement by glomerular epithelial cells in experimental membranous nephropathy.

    PubMed

    Kerjaschki, D; Schulze, M; Binder, S; Kain, R; Ojha, P P; Susani, M; Horvat, R; Baker, P J; Couser, W G

    1989-07-15

    Deposition of the C5b-9 complex of C in glomeruli of rats with experimental membranous nephropathy (MN) is essential for the development of proteinuria. In this investigation C5b-9 was localized in the passive Heymann nephritis (PHN) by immunoelectron microscopy with a mAb specific for C5b-9(m) neoantigen. Its distribution was compared with that in another model of MN induced by successive injections of cationic human IgG and rabbit anti-human IgG into rats. In PHN C5b-9 was found: 1) in the immune deposits (ID), and on the cell membranes of foot processes close to the ID; 2) in clathrin-coated pits of the glomerular epithelial cells (GEC) close to the ID and in membrane vesicles in the cytoplasm, separated from sheep IgG and the gp330 Ag; 3) in high concentration in multivesicular bodies of GEC; and 4) in association with membrane vesicles in the urinary space which presumably are the exocytosed content of membrane vesicular bodies. By contrast, in the cationic IgG-MN model C5b-9 was found mostly in ID, but rarely within the GEC. By freeze-fracture electron microscopy we have further identified 200- to 250-A intramembrane particles in PHN in the cell membranes of the "soles" of the foot processes which resemble membrane inserted human C5b-9(m). Degradation products of C5b-9 were further detected by immunoblotting of a 100,000 x g pellet of PHN rat urine. These results indicate that, in PHN, C5b-9 is inserted into the cell membranes of GEC, and that it is selectively endocytosed and transported across GEC by a cellular mechanism which apparently protects the cell from accumulation of membrane-inserted C5b-9.

  5. Synthesis of E7 peptide-modified biodegradable polyester with the improving affinity to mesenchymal stem cells.

    PubMed

    Li, Qian; Xing, Dongming; Ma, Lie; Gao, Changyou

    2017-04-01

    As the most promising stem cell, bone marrow-derived mesenchymal stem cells (BMSCs) has attracted many attentions and applied widely in regenerative medicine. A biodegradable polyester with tunable affinity to BMSCs plays critical role in determining the properties of the BMSCs-based constructs. In this study, maleimide functionalized biodegradable polyester (P(MTMC-LA)) was synthesized through ring-opening copolymerization between l-lactide (LA) and furan-maleimide functionalized trimethylene carbonate (FMTMC) and a subsequent retro Diels-Alder reaction. P(MTMC-LA) was modified by different amounts of BMSCs specific affinity peptide (EPLQLKM, E7) through click-chemistry to investigate the effect on BMSCs. The E7 peptide modified P(MTMC-LA) was casted into films on glass slides and BMSCs were seeded onto the films. In vitro study showed that E7 peptide modified P(MTMC-LA) films supported BMSCs adhesion and proliferation compared to unmodified P(MTMC-LA) film. Besides, the adhesion and proliferation were enhanced by the increasing peptide grafting ratio. These results indicated that the novel biodegradable polyester can serve as a biomaterial with great potential application in tissue engineering and regenerative medicine.

  6. Control-oriented thermal management of solid oxide fuel cells based on a modified Takagi-Sugeno fuzzy model

    NASA Astrophysics Data System (ADS)

    Yang, Jie; Li, Xi; Mou, Hong-Gang; Jian, Li

    Thermal management for a solid oxide fuel cell (SOFC) is actually temperature control, due to the importance of cell temperature for the performance of an SOFC. An SOFC stack is a nonlinear and multi-variable system which is difficult to model by traditional methods. A modified Takagi-Sugeno (T-S) fuzzy model that is suitable for nonlinear systems is built to model the SOFC stack. The model parameters are initialized by the fuzzy c-means clustering method, and learned using an off-line back-propagation algorithm. In order to obtain the training data to identify the modified T-S model, a SOFC physical model via MATLAB is established. The temperature model is the center of the physical model and is developed by enthalpy-balance equations. It is shown that the modified T-S fuzzy model is sufficiently accurate to follow the temperature response of the stack, and can be conveniently utilized to design temperature control strategies.

  7. Plasma membranes modified by plasma treatment or deposition as solid electrolytes for potential application in solid alkaline fuel cells.

    PubMed

    Reinholdt, Marc; Ilie, Alina; Roualdès, Stéphanie; Frugier, Jérémy; Schieda, Mauricio; Coutanceau, Christophe; Martemianov, Serguei; Flaud, Valérie; Beche, Eric; Durand, Jean

    2012-07-30

    In the highly competitive market of fuel cells, solid alkaline fuel cells using liquid fuel (such as cheap, non-toxic and non-valorized glycerol) and not requiring noble metal as catalyst seem quite promising. One of the main hurdles for emergence of such a technology is the development of a hydroxide-conducting membrane characterized by both high conductivity and low fuel permeability. Plasma treatments can enable to positively tune the main fuel cell membrane requirements. In this work, commercial ADP-Morgane® fluorinated polymer membranes and a new brand of cross-linked poly(aryl-ether) polymer membranes, named AMELI-32®, both containing quaternary ammonium functionalities, have been modified by argon plasma treatment or triallylamine-based plasma deposit. Under the concomitant etching/cross-linking/oxidation effects inherent to the plasma modification, transport properties (ionic exchange capacity, water uptake, ionic conductivity and fuel retention) of membranes have been improved. Consequently, using plasma modified ADP-Morgane® membrane as electrolyte in a solid alkaline fuel cell operating with glycerol as fuel has allowed increasing the maximum power density by a factor 3 when compared to the untreated membrane.

  8. Plasma Membranes Modified by Plasma Treatment or Deposition as Solid Electrolytes for Potential Application in Solid Alkaline Fuel Cells

    PubMed Central

    Reinholdt, Marc; Ilie, Alina; Roualdès, Stéphanie; Frugier, Jérémy; Schieda, Mauricio; Coutanceau, Christophe; Martemianov, Serguei; Flaud, Valérie; Beche, Eric; Durand, Jean

    2012-01-01

    In the highly competitive market of fuel cells, solid alkaline fuel cells using liquid fuel (such as cheap, non-toxic and non-valorized glycerol) and not requiring noble metal as catalyst seem quite promising. One of the main hurdles for emergence of such a technology is the development of a hydroxide-conducting membrane characterized by both high conductivity and low fuel permeability. Plasma treatments can enable to positively tune the main fuel cell membrane requirements. In this work, commercial ADP-Morgane® fluorinated polymer membranes and a new brand of cross-linked poly(aryl-ether) polymer membranes, named AMELI-32®, both containing quaternary ammonium functionalities, have been modified by argon plasma treatment or triallylamine-based plasma deposit. Under the concomitant etching/cross-linking/oxidation effects inherent to the plasma modification, transport properties (ionic exchange capacity, water uptake, ionic conductivity and fuel retention) of membranes have been improved. Consequently, using plasma modified ADP-Morgane® membrane as electrolyte in a solid alkaline fuel cell operating with glycerol as fuel has allowed increasing the maximum power density by a factor 3 when compared to the untreated membrane. PMID:24958295

  9. Synthetically modified mRNA for efficient and fast human iPS cell generation and direct transdifferentiation to myoblasts.

    PubMed

    Preskey, David; Allison, Thomas F; Jones, Mark; Mamchaoui, Kamel; Unger, Christian

    2016-05-06

    Synthetic mRNA transfection enables efficient and controlled gene expression in human cells, without genome integrations. Further, modifications to the mRNA and transfection protocol now allow for repeated transfection and long-term gene expression of an otherwise short-lived mRNA expression. This is mainly achieved through introducing modified nucleosides and interferon suppression. In this study we provide an overview and details of the advanced synthesis and modifications of mRNA originally developed for reprogramming. This mRNA allows for very efficient transfection of fibroblasts enabling the generation of high quality human iPS cells with a six-factor mRNA cocktail in 9 days. Furthermore, we synthesised and transfected modified MYOD1 mRNA to transdifferentiate human fibroblasts into myoblast-like cells without a transgene footprint. This efficient and integration-free mRNA technology opens the door for safe and controlled gene expression to reverse or redirect cell fate.

  10. Induction of B cell immune tolerance by antigen-modified CTL

    PubMed Central

    Nguyen, Phuong; Geiger, Terrence L.

    2010-01-01

    Background Third party-specific CTL, or veto CTL, are being assessed as a cellular therapeutic for the induction of T cell tolerance during transplantation. Conceptually, veto cell expressed antigens may induce B cell immune responses, and this may have deleterious consequences. Whether veto cells induce immunity, tolerance, or are ignored by B lymphocytes has, however, not been addressed. Methods CTL were retrovirally transduced with a model cell surface antigen to generate veto CTL. The impact of CTL-specific antigen expression on the activation and tolerization of antigen specific B cells was assessed in vitro and, using adoptive transfer models, in vivo. Results In vitro, CTL-expressed antigen induced an abortive proliferative response in specific B lymphocytes, whereby an initial proliferative burst was followed by cell death. In vivo, the administration of veto CTL also induced B cell tolerance. Specific immunoglobulin was not detected after subsequent immunization with a veto cell-expressed antigen. Modeling of this effect with antigen-specific B cell receptor (BCR) transgenic B lymphocytes demonstrated that antigen-specific B cells were eliminated by the veto CTL; cell division was accompanied by the exhaustion and depletion of responding cells. Veto-induced B cell tolerance could be wholly abrogated by treatment with the toll-like receptor ligand LPS, implying that this tolerance resulted from the absence of adequate supplemental signals during antigenic stimulation. Conclusions Veto CTL are effective promoters of B cell tolerance. Further assessment of their therapeutic potential in this regard is warranted. PMID:20065917

  11. Quick chip assay using locked nucleic acid modified epithelial cell adhesion molecule and nucleolin aptamers for the capture of circulating tumor cells

    PubMed Central

    Maremanda, Nihal G.; Roy, Kislay; Kanwar, Rupinder K.; Shyamsundar, Vidyarani; Ramshankar, Vijayalakshmi; Krishnamurthy, Arvind; Krishnakumar, Subramanian; Kanwar, Jagat R.

    2015-01-01

    The role of circulating tumor cells (CTCs) in disease diagnosis, prognosis, monitoring of the therapeutic efficacy, and clinical decision making is immense and has attracted tremendous focus in the last decade. We designed and fabricated simple, flat channel microfluidic devices polydimethylsiloxane (PDMS based) functionalized with locked nucleic acid (LNA) modified aptamers (targeting epithelial cell adhesion molecule (EpCAM) and nucleolin expression) for quick and efficient capture of CTCs and cancer cells. With optimized flow rates (10 μl/min), it was revealed that the aptamer modified devices offered reusability for up to six times while retaining optimal capture efficiency (>90%) and specificity. High capture sensitivity (92%) and specificity (100%) was observed in whole blood samples spiked with Caco-2 cells (10–100 cells/ml). Analysis of blood samples obtained from 25 head and neck cancer patients on the EpCAM LNA aptamer functionalized chip revealed that an average count of 5 ± 3 CTCs/ml of blood were captured from 22/25 samples (88%). EpCAM intracellular domain (EpICD) immunohistochemistry on 9 oral squamous cell carcinomas showed the EpICD positivity in the tumor cells, confirming the EpCAM expression in CTCs from head and neck cancers. These microfluidic devices also maintained viability for in vitro culture and characterization. Use of LNA modified aptamers provided added benefits in terms of cost effectiveness due to increased reusability and sustainability of the devices. Our results present a robust, quick, and efficient CTC capture platform with the use of simple PDMS based devices that are easy to fabricate at low cost and have an immense potential in cancer diagnosis, prognosis, and therapeutic planning. PMID:26487896

  12. Fetal cell carcinogenesis of the thyroid: a modified theory based on recent evidence.

    PubMed

    Takano, Toru

    2014-01-01

    Thyroid cancer cells were believed to be generated by multi-step carcinogenesis, in which cancer cells are derived from thyrocytes, via multiple incidences of damage to their genome, especially in oncogenes or anti-oncogenes that accelerate proliferation or foster malignant phenotypes, such as the ability to invade the surrounding tissue or metastasize to distant organs, until a new hypothesis, fetal cell carcinogenesis, was presented. In fetal cell carcinogenesis, thyroid tumor cells are assumed to be derived from three types of fetal thyroid cell which only exist in fetuses or young children, namely, thyroid stem cells (TSCs), thyroblasts and prothyrocytes, by proliferation without differentiation. Genomic alternations, such as RET/PTC and PAX8-PPARγ1 rearrangements and a mutation in the BRAF gene, play an oncogenic role by preventing thyroid fetal cells from differentiating. Fetal cell carcinogenesis effectively explains recent molecular and clinical evidence regarding thyroid cancer, including thyroid cancer initiating cells (TCICs), and it underscores the importance of identifying a stem cells and clarifying the molecular mechanism of organ development in cancer research. It introduces three important concepts, the reverse approach, stem cell crisis and mature and immature cancers. Further, it implies that analysis of a small population of cells in a cancer tissue will be a key technique in establishing future laboratory tests. In the contrary, mass analysis such as gene expression profiling, whole genomic scan, and proteomics analysis may have definite limitations since they can only provide information based on many cells.

  13. Zinc oxide modified with benzylphosphonic acids as transparent electrodes in regular and inverted organic solar cell structures

    SciTech Connect

    Lange, Ilja; Reiter, Sina; Kniepert, Juliane; Piersimoni, Fortunato; Brenner, Thomas; Neher, Dieter; Pätzel, Michael; Hildebrandt, Jana; Hecht, Stefan

    2015-03-16

    An approach is presented to modify the work function of solution-processed sol-gel derived zinc oxide (ZnO) over an exceptionally wide range of more than 2.3 eV. This approach relies on the formation of dense and homogeneous self-assembled monolayers based on phosphonic acids with different dipole moments. This allows us to apply ZnO as charge selective bottom electrodes in either regular or inverted solar cell structures, using poly(3-hexylthiophene):phenyl-C71-butyric acid methyl ester as the active layer. These devices compete with or even surpass the performance of the reference on indium tin oxide/poly(3,4-ethylenedioxythiophene) polystyrene sulfonate. Our findings highlight the potential of properly modified ZnO as electron or hole extracting electrodes in hybrid optoelectronic devices.

  14. Photoabsorption-enhanced dye-sensitized solar cell by using localized surface plasmon of silver nanoparticles modified with polymer

    NASA Astrophysics Data System (ADS)

    Ihara, Manabu; Kanno, Masami; Inoue, Shiho

    2010-09-01

    Photoelectric conversion efficiency ( Eff) of a dye-sensitized solar cell (DSSC) was improved by localized surface plasmon on silver (Ag) nanoparticles modified with polyacrylate-based comb-shaped block copolymer. The polymer-modified Ag nanoparticles were inserted into pores of a TiO 2 film in a DSSC. Then, the characteristics of this photoabsorption enhanced DSSC (PE-DSSC) with the Ag nanoparticles were evaluated. To clarify the effect of Ag nanoparticles on the characteristics, the thickness of the TiO 2 film in the DSSC was 2 μm, thinner than that for a typical DSSC (10 μm). A PE-DSSC showed an improved Eff of 2.5%, compared with about 1.5% for a DSSC with no Ag nanoparticles.

  15. DBD atmospheric plasma-modified, electrospun, layer-by-layer polymeric scaffolds for L929 fibroblast cell cultivation.

    PubMed

    Surucu, Seda; Turkoglu Sasmazel, Hilal

    2016-01-01

    This paper reported a study related to atmospheric pressure dielectric barrier discharge (DBD) Ar + O2 and Ar + N2 plasma modifications to alter surface properties of 3D PCL/Chitosan/PCL layer-by-layer hybrid scaffolds and to improve mouse fibroblast (L929 ATCC CCL-1) cell attachment, proliferation, and growth. The scaffolds were fabricated using electrospinning technique and each layer was electrospun sequentially on top of the other. The surface modifications were performed with an atmospheric pressure DBD plasma under different gas flow rates (50, 60, 70, 80, 90, and 100 sccm) and for different modification times (0.5-7 min), and then the chemical and topographical characterizations of the modified samples were done by contact angle (CA) measurements, scanning electron microscopy (SEM), atomic force microscopy, and X-ray photoelectron spectroscopy. The samples modified with Ar + O2 plasma for 1 min under 70 cm(3)/min O2 flow rate (71.077° ± 3.578) showed a 18.83% decrease compare to unmodified samples' CA value (84.463° ± 3.864). Comparing with unmodified samples, the average fiber diameter values for plasma-modified samples by Ar + O2 (1 min 70 sccm) and Ar + N2 (40 s 70 sccm) increased 40.756 and 54.295%, respectively. Additionally, the average inter-fiber pore size values exhibited decrease of 37.699 and 48.463% for the same Ar + O2 and Ar + N2 plasma-modified samples, respectively, compare to unmodified samples. Biocompatibility performance was determined with MTT assay, fluorescence, Giemsa, and confocal imaging as well as SEM. The results showed that Ar + O2-based plasma modification increased the hydrophilicity and oxygen functionality of the surface, thus affecting the cell viability and proliferation on/within scaffolds.

  16. Displacement damage analysis and modified electrical equivalent circuit for electron and photon-irradiated silicon solar cells

    NASA Astrophysics Data System (ADS)

    Arjhangmehr, Afshin; Feghhi, Seyed Amir Hossein

    2014-10-01

    Solar modules and arrays are the conventional energy resources of space satellites. Outside the earth's atmosphere, solar panels experience abnormal radiation environments and because of incident particles, photovoltaic (PV) parameters degrade. This article tries to analyze the electrical performance of electron and photon-irradiated mono-crystalline silicon (mono-Si) solar cells. PV cells are irradiated by mono-energetic electrons and poly-energetic photons and immediately characterized after the irradiation. The mean degradation of the maximum power (Pmax) of silicon solar cells is presented and correlated using the displacement damage dose (Dd) methodology. This method simplifies evaluation of cell performance in space radiation environments and produces a single characteristic curve for Pmax degradation. Furthermore, complete analysis of the results revealed that the open-circuit voltage (Voc) and the filling factor of mono-Si cells did not significantly change during the irradiation and were independent of the radiation type and fluence. Moreover, a new technique is developed that adapts the irradiation-induced effects in a single-cell equivalent electrical circuit and adjusts its elements. The "modified circuit" is capable of modeling the "radiation damage" in the electrical behavior of mono-Si solar cells and simplifies the designing of the compensation circuits.

  17. Genetically modified anthrax lethal toxin safely delivers whole HIV protein antigens into the cytosol to induce T cell immunity

    NASA Astrophysics Data System (ADS)

    Lu, Yichen; Friedman, Rachel; Kushner, Nicholas; Doling, Amy; Thomas, Lawrence; Touzjian, Neal; Starnbach, Michael; Lieberman, Judy

    2000-07-01

    Bacillus anthrax lethal toxin can be engineered to deliver foreign proteins to the cytosol for antigen presentation to CD8 T cells. Vaccination with modified toxins carrying 8-9 amino acid peptide epitopes induces protective immunity in mice. To evaluate whether large protein antigens can be used with this system, recombinant constructs encoding several HIV antigens up to 500 amino acids were produced. These candidate HIV vaccines are safe in animals and induce CD8 T cells in mice. Constructs encoding gag p24 and nef stimulate gag-specific CD4 proliferation and a secondary cytotoxic T lymphocyte response in HIV-infected donor peripheral blood mononuclear cells in vitro. These results lay the foundation for future clinical vaccine studies.

  18. Dye-sensitized solar cells with modified TiO2 surface chemical states: The role of Ti3+

    NASA Astrophysics Data System (ADS)

    Yu, Yang; Wu, Kunjie; Wang, Deliang

    2011-11-01

    Dye-sensitized solar cells (DSSCs) with TiO2 electrodes, which were modified to have different surface chemical states, were fabricated. The DSSCs had an ultra-flat TiO2 electrode, on which only a mono-layer dye was attached. The cell I-V measurement showed that Ti3+ ion had a dramatic effect on cell performance. The efficiency of a DSSC was shown to have a strong correlation with the concentration of the Ti3+ surface state, it decreased almost linearly with increased Ti3+ concentration. The oxygen vacancy-Ti3+ defect served as electron recombination center and decreased both the open-circuit voltage and the fill factor.

  19. Serotonergic chemosensory neurons modify the C. elegans immune response by regulating G-protein signaling in epithelial cells.

    PubMed

    Anderson, Alexandra; Laurenson-Schafer, Henry; Partridge, Frederick A; Hodgkin, Jonathan; McMullan, Rachel

    2013-01-01

    The nervous and immune systems influence each other, allowing animals to rapidly protect themselves from changes in their internal and external environment. However, the complex nature of these systems in mammals makes it difficult to determine how neuronal signaling influences the immune response. Here we show that serotonin, synthesized in Caenorhabditis elegans chemosensory neurons, modulates the immune response. Serotonin released from these cells acts, directly or indirectly, to regulate G-protein signaling in epithelial cells. Signaling in these cells is required for the immune response to infection by the natural pathogen Microbacterium nematophilum. Here we show that serotonin signaling suppresses the innate immune response and limits the rate of pathogen clearance. We show that C. elegans uses classical neurotransmitters to alter the immune response. Serotonin released from sensory neurons may function to modify the immune system in response to changes in the animal's external environment such as the availability, or quality, of food.

  20. Enhancement of natural killer cell activity of aged mice by modified arabinoxylan rice bran (MGN-3/Biobran).

    PubMed

    Ghoneum, Mamdooh; Abedi, Sarah

    2004-12-01

    The present study is aimed to examine the possibility of enhancement of natural killer (NK) cell activity in aged C57BL/6 and C3H mice using MGN-3, a modified arabinoxylan from rice bran. Intraperitoneal injection of MGN-3 (10 mg kg(-1) per day) caused a remarkable increase in the peritoneal NK activity as early as 2 days (35.2 lytic units), and the level remained elevated through day 14. The control aged mice had a level of 5.8 lytic units. Enhancement in NK activity was associated with an increase in both the binding capacity of NK cells to tumour targets and in the granular content as measured by BLT-esterase activity. Treatment did not alter the percentage of peritoneal NK cells. Data showed that peritoneal macrophages inhibit NK activity. In conclusion, MGN-3 enhances murine NK activity of aged mice and may be useful for enhancing NK function in aged humans.

  1. Chimeric Antigen Receptor- and TCR-Modified T Cells Enter Main Street and Wall Street.

    PubMed

    Barrett, David M; Grupp, Stephan A; June, Carl H

    2015-08-01

    The field of adoptive cell transfer (ACT) is currently comprised of chimeric Ag receptor (CAR)- and TCR-engineered T cells and has emerged from principles of basic immunology to paradigm-shifting clinical immunotherapy. ACT of T cells engineered to express artificial receptors that target cells of choice is an exciting new approach for cancer, and it holds equal promise for chronic infection and autoimmunity. Using principles of synthetic biology, advances in immunology, and genetic engineering have made it possible to generate human T cells that display desired specificities and enhanced functionalities. Clinical trials in patients with advanced B cell leukemias and lymphomas treated with CD19-specific CAR T cells have induced durable remissions in adults and children. The prospects for the widespread availability of engineered T cells have changed dramatically given the recent entry of the pharmaceutical industry to this arena. In this overview, we discuss some of the challenges and opportunities that face the field of ACT.

  2. Angiopoietin-1-expressing adipose stem cells genetically modified with baculovirus nanocomplex: investigation in rat heart with acute infarction.

    PubMed

    Paul, Arghya; Nayan, Madhur; Khan, Afshan Afsar; Shum-Tim, Dominique; Prakash, Satya

    2012-01-01

    The objective of this study was to develop angiopoietin-1 (Ang1)-expressing genetically modified human adipose tissue derived stem cells (hASCs) for myocardial therapy. For this, an efficient gene delivery system using recombinant baculovirus complexed with cell penetrating transactivating transcriptional activator TAT peptide/deoxyribonucleic acid nanoparticles (Bac-NP), through ionic interactions, was used. It was hypothesized that the hybrid Bac- NP(Ang1) system can efficiently transduce hASCs and induces favorable therapeutic effects when transplanted in vivo. To evaluate this hypothesis, a rat model with acute myocardial infarction and intramyocardially transplanted Ang1-expressing hASCs (hASC-Ang1), genetically modified by Bac-NP(Ang1), was used. Ang1 is a crucial pro-angiogenic factor for vascular maturation and neovasculogenesis. The released hAng1 from hASC-Ang1 demonstrated profound mitotic and anti-apoptotic activities on endothelial cells and cardiomyocytes. The transplanted hASC-Ang1 group showed higher cell retention compared to hASC and control groups. A significant increase in capillary density and reduction in infarct sizes were noted in the infarcted hearts with hASC-Ang1 treatment compared to infarcted hearts treated with hASC or the untreated group. Furthermore, the hASC-Ang1 group showed significantly higher cardiac performance in echocardiography (ejection fraction 46.28% ± 6.3%, P < 0.001 versus control, n = 8) than the hASC group (36.35% ± 5.7%, P < 0.01, n = 8), 28 days post-infarction. The study identified Bac-NP complex as an advanced gene delivery vehicle for stem cells and demonstrated its potential to treat ischemic heart disease with high therapeutic index for combined stem cell-gene therapy strategy.

  3. Modifying dielectrophoretic response of nonviable yeast cells by ionic surfactant treatment.

    PubMed

    Tang, Shi-Yang; Zhang, Wei; Baratchi, Sara; Nasabi, Mahyar; Kalantar-Zadeh, Kourosh; Khoshmanesh, Khashayar

    2013-07-02

    Nonviable cells are essential biosystems, due to the functionalities they offer and their effects on viable cells. Therefore, the separation and immobilization of nonviable cells separately or in the vicinity of viable cells is of great importance for many fundamentals investigations in cell biology. However, most nonviable cells become less polarizable than the surrounding medium at conductivities above 0.01 S/m. This means that in such a medium, dielectrophoresis, despite its great versatilities for manipulation of cells, cannot be employed for immobilizing nonviable cells. Here, we present a novel approach to change the dielectrophoretic (DEP) response of nonviable yeast cells by treating them with low concentrations of ionic surfactants such as sodium dodecyl sulfate. After this treatment, they exhibit a strong positive DEP response, even at high medium conductivities. The capability of this treatment is demonstrated in two proof-of-concept experiments. First, we show the sorting and immobilization of viable and nonviable yeast cells, along consecutive microelectrode arrays. Second, we demonstrate the immobilization of viable and nonviable cells in the vicinity of each other along the same microelectrode array. The proposed technique allows DEP platforms to be utilized for the immobilization and subsequent postanalysis of both viable and nonviable cells with and without the presence of each other.

  4. Genetically Modified T-Cell Therapy in Treating Patients With Advanced ROR1+ Malignancies

    ClinicalTrials.gov

    2017-03-15

    Estrogen Receptor Negative; HER2/Neu Negative; Progesterone Receptor Negative; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Breast Carcinoma; Recurrent Chronic Lymphocytic Leukemia; Recurrent Mantle Cell Lymphoma; Recurrent Non-Small Cell Lung Carcinoma; Refractory Chronic Lymphocytic Leukemia; Stage IV Breast Cancer; Stage IV Non-Small Cell Lung Cancer; Triple-Negative Breast Carcinoma

  5. The preoperative sensitive-modified Glasgow prognostic score is superior to the modified Glasgow prognostic score in predicting long-term survival for esophageal squamous cell carcinoma

    PubMed Central

    Tian, Rui; Zhang, Fei; Sun, Peng; Wu, Jing; Yan, Hong; Wu, Ai-Ran; Zhang, Min; Jiang, Yu-Lu; Lu, Yan-Hong; Xu, Qiu-Yan; Zhan, Xiao-Hong; Zhang, Rong-Xin; Qian, Li-Ting; He, Jie

    2016-01-01

    The present study was designed to investigate the prognostic significance of the preoperative sensitive-modified Glasgow prognostic score (S-mGPS) and its superiority in esophageal squamous cell carcinoma (ESCC). Clinicopathologic characteristics, preoperative albumin and C-reactive protein (CRP) levels were retrospectively collected in 442 patients who underwent transthoracic esophagectomy. The S-mGPS was calculated before surgery based on optimal cutoff values of 45.6 g/L for albumin and 10.0 mg/L for CRP. 360, 74 and 8 cases were assigned an mGPS of 0, 1 and 2, respectively. In contrast, the S-mGPS was 0 in 114, 1 in 258 and 2 in 70 patients. Of the 360 patients with an mGPS of 0, 246 migrated to the S-mGPS-1 group. Both mGPS and S-mGPS were significantly correlated with tumor length, depth of invasion, pathological tumor-node-metastasis (pTNM) stage and adjuvant treatment. In addition, they were significantly associated with disease free survival (DFS) and overall survival (OS) in univariate analysis. Furthermore, multivariate Cox regression analysis identified S-mGPS as an independent prognostic indicator for both DFS [hazard ratio (HR), 1.577; 95% confidence interval (CI), 1.149-2.163; P = 0.005] and OS (HR, 1.762; 95% CI, 1.250-2.484; P = 0.001), but not mGPS (HR, 0.957; 95% CI, 0.692-1.323; P = 0.790 for DFS and HR, 1.089; 95% CI, 0.781-1.517; P = 0.615 for OS, respectively). Moreover, subgroup analysis revealed that the prognostic impact of the S-mGPS was especially striking in pTNM stage II patients. The preoperative S-mGPS is superior to the mGPS as a prognostic predictor in patients with resectable ESCC. PMID:27528228

  6. Protein, cell and bacterial fouling resistance of polypeptoid-modified surfaces: effect of side-chain chemistry†

    PubMed Central

    Statz, Andrea R.; Barron, Annelise E.; Messersmith, Phillip B.

    2011-01-01

    Peptidomimetic polymers consisting of poly-N-substituted glycine oligomers (polypeptoids) conjugated to biomimetic adhesive polypeptides were investigated as antifouling surface coatings. The polymers were immobilized onto TiO2 surfaces via an anchoring peptide consisting of alternating residues of 3,4-dihydroxyphenylalanine (DOPA) and lysine. Three polypeptoid side-chain compositions were investigated for antifouling performance and stability toward enzymatic degradation. Ellipsometry and XPS analysis confirmed that purified polymers adsorbed strongly to TiO2 surfaces, and the immobilized polymers were resistant to enzymatic degradation as demonstrated by mass spectrometry. All polypeptoid-modified surfaces exhibited significant reductions in adsorption of lysozyme, fibrinogen and serum proteins, and were resistant to 3T3 fibroblast cell attachment for up to seven days. Long-term in vitro cell attachment studies conducted for six weeks revealed the importance of polypeptoid side-chain composition, with a methoxyethyl side chain providing superior long-term fouling resistance compared to hydroxyethyl and hydroxypropyl side chains. Finally, attachment of both gram-positive and gram-negative bacteria for up to four days under continuous-flow conditions was significantly reduced on the polypeptoid-modified surfaces compared to unmodified TiO2 surfaces. The results reveal the influence of polypeptoid side-chain chemistry on short-term and long-term protein, cell and bacterial fouling resistance. PMID:21472038

  7. Polymer films with surfaces unmodified and modified by non-thermal plasma as new substrates for cell adhesion.

    PubMed

    Borges, A M G; Benetoli, L O; Licínio, M A; Zoldan, V C; Santos-Silva, M C; Assreuy, J; Pasa, A A; Debacher, N A; Soldi, V

    2013-04-01

    The surface properties of biomaterials, such as wettability, polar group distribution, and topography, play important roles in the behavior of cell adhesion and proliferation. Gaseous plasma discharges are among the most common means to modify the surface of a polymer without affecting its properties. Herein, we describe the surface modification of poly(styrene) (PS) and poly(methyl methacrylate) (PMMA) films using atmospheric pressure plasma processing through exposure to a dielectric barrier discharge (DBD). After treatment the film surface showed significant changes from hydrophobic to hydrophilic as the water contact angle decreasing from 95° to 37°. All plasma-treated films developed more hydrophilic surfaces compared to untreated films, although the reasons for the change in the surface properties of PS and PMMA differed, that is, the PS showed chemical changes and in the case of PMMA they were topographical. Excellent adhesion and cell proliferation were observed in all films. In vitro studies employing flow cytometry showed that the proliferation of L929 cells was higher in the film formed by a 1:1 mixture of PS/PMMA, which is consistent with the results of a previous study. These findings suggest better adhesion of L929 onto the 1:1 PS/PMMA modified film, indicating that this system is a new candidate biomaterial for tissue engineering.

  8. Aptamer-modified gold nanoparticles for targeting breast cancer cells through light scattering

    NASA Astrophysics Data System (ADS)

    Huang, Yu-Fen; Lin, Yang-Wei; Lin, Zong-Hong; Chang, Huan-Tsung

    2009-05-01

    In this study, we demonstrated the potential use of nucleic acid ligand (aptamers) conjugated gold nanoparticles (AuNPs) for cancer cell detection. Through specific binding of the aptamers toward platelet-derived growth factor (PDGF), MDA-MB-231 and Hs578T cells (cancer cells) that over-express PDGF, interact with Apt-AuNPs to a greater extent than do H184B5F5/M10 cells (normal cells). These results were confirmed through inductively coupled plasma mass spectrometry measurements of the gold ion concentrations within these cells. Aggregation of the Apt-AuNPs in the cytoplasm of the cancer cells led to the generation of an intense scattered light upon photo-illumination; this phenomenon allows the differentiation of cancer cells from normal cells using a dark field optical microscope. The presence of Apt-AuNPs suppressed the proliferation of MDA-MB-231 cancer cells, but not H184B5F5/M10 cells.

  9. X-box-binding protein 1-modified neural stem cells for treatment of Parkinson's disease.

    PubMed

    Si, Lihui; Xu, Tianmin; Wang, Fengzhang; Liu, Qun; Cui, Manhua

    2012-04-05

    X-box-binding protein 1-transfected neural stem cells were transplanted into the right lateral ventricles of rats with rotenone-induced Parkinson's disease. The survival capacities and differentiation rates of cells expressing the dopaminergic marker tyrosine hydroxylase were higher in X-box-binding protein 1-transfected neural stem cells compared to non-transfected cells. Moreover, dopamine and 3,4-dihydroxyphenylacetic acid levels in the substantia nigra were significantly increased, α-synuclein expression was decreased, and neurological behaviors were significantly ameliorated in rats following transplantation of X-box-binding protein 1-transfected neural stem cells. These results indicate that transplantation of X-box-binding protein 1-transfected neural stem cells can promote stem cell survival and differentiation into dopaminergic neurons, increase dopamine and 3,4-dihydroxyphenylacetic acid levels, reduce α-synuclein aggregation in the substantia nigra, and improve the symptoms of Parkinson's disease in rats.

  10. Tissue kallikrein-modified human endothelial progenitor cell implantation improves cardiac function via enhanced activation of akt and increased angiogenesis.

    PubMed

    Yao, Yuyu; Sheng, Zulong; Li, YeFei; Fu, Cong; Ma, Genshan; Liu, Naifeng; Chao, Julie; Chao, Lee

    2013-05-01

    Endothelial progenitor cells (EPCs) have been shown to enhance angiogenesis not only by incorporating into the vasculature but also by secreting cytokines, thereby serving as an ideal vehicle for gene transfer. As tissue kallikrein (TK) has pleiotropic effects in inhibiting apoptosis and oxidative stress, and promoting angiogenesis, we evaluated the salutary potential of kallikrein-modified human EPCs (hEPCs; Ad.hTK-hEPCs) after acute myocardial infarction (MI). We genetically modified hEPCs with a TK gene and evaluated cell survival, engraftment, revascularization, and functional improvement in a nude mouse left anterior descending ligation model. hEPCs were manipulated to overexpress the TK gene. In vitro, the antiapoptotic and paracrine effects were assessed under oxidative stress. TK protects hEPCs from oxidative stress-induced apoptosis via inhibition of activation of caspase-3 and -9, induction of Akt phosphorylation, and secretion of vascular endothelial growth factor. In vivo, the Ad.hTK-hEPCs were transplanted after MI via intracardiac injection. The surviving cells were tracked after transplantation using near-infrared optical imaging. Left ventricular (LV) function was evaluated by transthoracic echocardiography. Capillary density was quantified using immunohistochemical staining. Engrafted Ad.hTK-hEPCs exhibited advanced protection against ischemia by increasing LV ejection fraction. Compared with Ad.Null-hEPCs, transplantation with Ad.hTK-hEPCs significantly decreased cardiomyocyte apoptosis in association with increased retention of transplanted EPCs in the myocardium. Capillary density and arteriolar density in the infarct border zone was significantly higher in Ad.hTK-hEPC-transplanted mice than in Ad.Null-hEPC-treated mice. Transplanted hEPCs were clearly incorporated into CD31(+) capillaries. These results indicate that implantation of kallikrein-modified EPCs in the heart provides advanced benefits in protection against ischemia-induced MI by

  11. [Vitro study on gene transfection efficiency of hyaluronic acid modified core-shell liponanoparticles in human retinal pigment epithelium cells].

    PubMed

    Zhao, Ya-Nan; Gan, Li; Wang, Jing; Chen, Xi; Jia, Zheng; Gan, Yong; Liu, Jian-Ping

    2014-05-01

    The aim of this study is to prepare hyaluronic acid (HA) modified core-shell liponanoparticles (pHA-LCS-NPs) as gene delivery system and investigate its gene transfection efficiency in human retinal pigment epithelium (ARPE-19) cells in vitro. The pHA-LCS-NPs was prepared by firstly hydrating dry lipid film with CS-NPs suspension to get LCS-NPs, then modifying the lipid bilayer with HA by amidation reaction between HA and dioleoyl phosphatidylethanolamine (DOPE). Its morphology, particle size and zeta potential were investigated. XTT assay was used to evaluate the cell safety of different vectors in vitro. The gene transfection efficiency of pHA-LCS-NPs modified with different contents of HA was investigated in ARPE-19 cells with green fluorescent protein (pEGFP) as the reporter gene. The results showed that the obtained pHA-LCS-NPs exhibited a clear core-shell structure with the average particles size of (214.9 +/- 7.2) nm and zeta potential of (-35 +/- 3.7) mV. The 24 h cumulative release of gene from pHA-LCS-NPs was less than 30%. After 48 h incubation, gene transfection efficiency of pHA-LCS-NPs/pEGFP was 1.81 times and 3.75 times higher than that of CS-NPs/pEGFP and naked pEGFP, respectively. Also no obvious cytotoxicity was observed on pHA-LCS-NPs. It suggested that the pHA-LCS-NPs might be promising non-viral gene delivery systems with high efficiency and low cytotoxicity.

  12. Optical data recording by laser pulses in liquid-crystal cells with an azo-modified surface

    SciTech Connect

    Serak, S V; Agashkov, A V; Reshetnyak, V Yu

    2001-03-31

    The effect of trans-cis photoisomerisation of azofragments of a polymer film on the molecular reorientation of a liquid crystal is studied. It is shown that, using nanosecond laser pulses, one can perform both the reversible and static data recording in liquid-crystal cells with an azo-modified surface. The rise time of the reorientation is measured by the methods of dynamic holography to be about {approx} 30 {mu}s, and the grating efficiency achieves 15 %. (laser applications and other topics in quantum electronics)

  13. C3N4 Nanosheet Modified Microwell Array with Enhanced Electrochemiluminescence for Total Analysis of Cholesterol at Single Cells.

    PubMed

    Xu, Jingjing; Jiang, Depeng; Qin, Yanling; Xia, Juan; Jiang, Dechen; Chen, Hong-Yuan

    2017-02-21

    Here, a g-C3N4 nanosheet modified microwell array providing enhanced electrochemiluminescence (ECL) and better visible sensitivity was prepared to simultaneously analyze total (membrane and intracellular) cholesterol at single cells. The detection limit for ECL visualization of hydrogen peroxide at microwell array was improved to be 500 nM that guaranteed the detection of low concentration cholesterol at single cells in parallel. To achieve single cell cholesterol analysis, the individual cells cultured at the microwell array were exposed to cholesterol oxidase generating hydrogen peroxide for luminescence analysis of membrane cholesterol, and then treated with triton X-100, cholesterol esterase, and cholesterol oxidase to produce hydrogen peroxide from intracellular cholesterol for luminescence determination. The observation of the luminescence spots at microwells in these two steps confirmed the codetection of membrane and intracellular cholesterol at single cells. The inhibition of intracellular acyl-coA/cholesterol acyltransferase (ACAT) resulted in less intracellular cholesterol storage (less luminescence) and more membrane cholesterol (more luminescence). The correlation of the luminescence intensity with the amount of cholesterol confirmed that our assay could simultaneously monitor membrane and intracellular cholesterol pools at different cellular states, which should offer more information for the study of cholesterol-related pathways at single cells.

  14. Shifting the balance: cell-based therapeutics as modifiers of the amyotrophic lateral sclerosis-specific neuronal microenvironment.

    PubMed

    Riley, Jonathan; Sweeney, Walter; Boulis, Nicholas

    2008-01-01

    Recent advances in the laboratory have improved the current understanding of neurobiological mechanisms underlying the initiating events and pathological progression observed in amyotrophic lateral sclerosis (ALS). Whereas initial studies have revealed the late-stage intracellular cascades contributing to neuronal dysfunction and cell death, more recently collected data have begun to elucidate the presence and importance of a "non-cell autonomous" component indicating that affected glial cell subtypes may serve distinct and required roles. Pharmacological interventions for ALS have largely been disappointing likely in part because they have failed to address either the proximate events contributing to neuronal dysfunction and death or the deleterious contributions of non-neuronal cells within the local microenvironment. Alternatively, cell-based therapeutics offer the potential of a multifaceted approach oriented toward the dual ends of protecting remaining viable neurons and attempting to restore neuronal function lost as a manifestation of disease progression. The authors review the evolving knowledge of disease initiation and progression, with specific emphasis on the role of affected glia as crucial contributors to the observed ALS phenotype. This basis is used to underscore the potential roles of cell-based therapeutics as modifiers of the ALS-specific microenvironment.

  15. Nucleoside-Diphosphate-Kinase of P. gingivalis is Secreted from Epithelial Cells In the Absence of a Leader Sequence Through a Pannexin-1 Interactome

    PubMed Central

    Atanasova, Kalina; Lee, Jungnam; Roberts, JoAnn; Lee, Kyulim; Ojcius, David M; Yilmaz, Özlem

    2016-01-01

    Nucleoside-diphosphate-kinases (NDKs) are leaderless, multifunctional enzymes. The mode(s) of NDK secretion is currently undefined, while extracellular translocation of bacterial NDKs is critical for avoidance of host pathogen clearance by opportunistic pathogens such as Porphyromonas gingivalis. P. gingivalis-NDK during infection inhibits extracellular-ATP (eATP)/P2X7-receptor mediated cell death in gingival epithelial cells (GECs) via eATP hydrolysis. Furthermore, depletion of pannexin-1-hemichannel (PNX1) coupled with P2X7-receptor blocks the infection-induced eATP release in GECs, and P. gingivalis-NDK impacts this pathway. Ultrastructural and confocal microscopy of P. gingivalis-co-cultured GECs or green-fluorescent-protein (GFP)-P. gingivalis-NDK transfected GECs revealed a perinuclear/cytoplasmic localization of NDK. eATP stimulation induced NDK recruitment to the cell periphery. Depletion of PNX1 by siRNA or inhibition by probenecid resulted in significant blocking of extracellular NDK activity and secretion using ATPase and ELISA assays. Co-immunoprecipitation-coupled Mass-spectrometry method revealed association of P. gingivalis-NDK to the myosin-9 motor molecule. Interestingly, inhibition of myosin-9, actin, and lipid-rafts, shown to be involved in PNX1-hemichannel function, resulted in marked intracellular accumulation of NDK and decreased NDK secretion from infected GECs. These results elucidate for the first time PNX1-hemichannels as potentially main extracellular translocation pathway for NDKs from an intracellular pathogen, suggesting that PNX1-hemichannels may represent a therapeutic target for chronic opportunistic infections. PMID:27883084

  16. Nuclear localisation of endogenous SUMO-1-modified PDGF-C in human thyroid tissue and cell lines

    SciTech Connect

    Reigstad, Laila J.; Martinez, Aurora; Varhaug, Jan Erik; Lillehaug, Johan R. . E-mail: johan.lillehaug@mbi.uib.no

    2006-04-01

    We investigated post-translational modification and subcellular localisation of endogenous platelet-derived growth factor-C (PDGF-C) in human thyroid papillary carcinomas (PTC), non-neoplastic thyroid tissues, and a selection of cultured cell lines. PDGF-C expressed nuclear localisation in 95% of all tested cell types in culture and in 10% of the thyrocytes from both PTC and non-neoplastic tissue. The cell lines expressed two forms of full-length PDGF-C, {approx}39 and {approx}55 kDa, in cell membrane and cytosol, while the {approx}55 kDa form dominated in the nucleus where it was partly chromatin-associated. The {approx}55 kDa form was post-translationally modified by SUMO-1. The putative PDGF-C SUMOylation site is the surface exposed {sup 314}lysine part of a positively charged loop ({sup 312}RPKTGVRGLHK{sup 322}) with characteristics of a nuclear localisation signal. The tissue thyrocytes expressed a non-SUMOylated {approx}43 kDa and the 55 kDa PDGF-C. The SUMO-1 modified {approx}55 kDa PDGF-C expression was low in PTC where the {approx}43 kDa PDGF-C dominated. This is in contrast to non-neoplastic tissue and cultured cells where the SUMOylated {approx}55 kDa PDGF-C was strongly expressed. Our data provide novel evidence for nuclear localisation of PDGF-C, post-translational modification by SUMOylation and the expression of a novel form of PDGF-C in human papillary thyroid carcinomas.

  17. Modified citrus pectin stops progression of liver fibrosis by inhibiting galectin-3 and inducing apoptosis of stellate cells.

    PubMed

    Abu-Elsaad, Nashwa M; Elkashef, Wagdi Fawzi

    2016-05-01

    Modified citrus pectin (MCP) is a pH modified form of the dietary soluble citrus peel fiber known as pectin. The current study aims at testing its effect on liver fibrosis progression. Rats were injected with CCl4 (1 mL/kg, 40% v/v, i.p., twice a week for 8 weeks). Concurrently, MCP (400 or 1200 mg/kg) was administered daily in drinking water from the first week in groups I and II (prophylactic model) and in the beginning of week 5 in groups III and IV (therapeutic model). Liver function biomarkers (ATL, AST, and ALP), fibrosis markers (laminin and hyaluronic acid), and antioxidant biomarkers (reduced glutathione (GSH) and superoxide dismutase (SOD)) were measured. Stained liver sections were scored for fibrosis and necroinflammation. Additionally, expression of galectin-3 (Gal-3), α-smooth muscle actin (SMA), tissue inhibitor metalloproteinase (TIMP)-1, collagen (Col)1A1, caspase (Cas)-3, and apoptosis related factor (FAS) were assigned. Modified pectin late administration significantly (p < 0.05) decreased malondialdehyde (MDA), TIMP-1, Col1A1, α-SMA, and Gal-3 levels and increased levels of FAS, Cas-3, GSH, and SOD. It also decreased percentage of fibrosis and necroinflammation significantly (p < 0.05). It can be concluded that MCP can attenuate liver fibrosis through an antioxidant effect, inhibition of Gal-3 mediated hepatic stellate cells activation, and induction of apoptosis.

  18. Modifiers of muscle and heart cell fate specification identified by gain-of-function screen in Drosophila.

    PubMed

    Bidet, Yannick; Jagla, Teresa; Da Ponte, Jean-Philippe; Dastugue, Bernard; Jagla, Krzysztof

    2003-09-01

    The homeobox genes ladybird in Drosophila and their vertebrate counterparts Lbx1 genes display restricted expression patterns in a subset of muscle precursors and are both implicated in diversification of muscle cell fates. In order to gain new insights into mechanisms controlling conserved aspects of cell fate specification, we have performed a gain-of-function (GOF) screen for modifiers of the mesodermal expression of ladybird genes using a collection of EP element carrying Drosophila lines. Amongst the identified genes, several have been previously implicated in cell fate specification processes, thus validating the strategy of our screen. Observed GOF phenotypes have led us to identification of an important number of candidate genes, whose myogenic and/or cardiogenic functions remain to be investigated. Amongst them, the EP insertions close to rhomboid, yan and rac2 suggest new roles for these genes in diversification of muscle and/or heart cell lineages. The analysis of loss and GOF of rhomboid and yan reveals their new roles in specification of ladybird-expressing precursors of adult muscles (LaPs) and ladybird/tinman-positive pericardial cells. Observed phenotypes strongly suggest that rhomboid and yan act at the level of progenitor and founder cells and contribute to the diversification of mesodermal fates. Our analysis of rac2 phenotypes clearly demonstrates that the altered mesodermal level of Rho-GTPase Rac2 can influence specification of a number of cardiac and muscular cell types including those expressing ladybird. Finding that in rac2 mutants ladybird and even skipped-positive muscle founders are overproduced, indicate a new early function for this gene during segregation of muscle progenitors and/or specification of founder cells. Intriguingly, rhomboid, yan and rac2 act as conserved components of Receptor Tyrosine Kinases (RTKs) signalling pathways, suggesting that RTK signalling constitutes a part of a conserved regulatory network governing

  19. Cells responsible for tumor surveillance in man: effects of radiotherapy, chemotherapy, and biologic response modifiers

    SciTech Connect

    Reizenstein, P.; Ogier, C.; Blomgren, H.; Petrini, B.; Wasserman, J.

    1985-01-01

    Currently, the most probable theory of tumor surveillance is neither the existence of any tumor-specific, antigen-dependent, T-cell-mediated cytotoxic effect that could eliminate spontaneous tumors in man and that could be used for some kind of vaccination against tumors, nor the complete absence of any surveillance or defense systems against tumors. What is probable is the cooperation of a number of antigen-independent, relatively weakly cytotoxic or possibly only cytostatic humoral and cellular effects, including nutritional immunity, tumor necrosis factor, certain cytokines, and the cytotoxic effects mediated by macrophages, NK cells, NK-like cells, and certain stimulated T-cells. One question remaining to be solved is why these antigen-independent effects do not attack normal cells. A number of plausible hypotheses are discussed. The hypothetical surveillance system is modulated both by traditional cancer treatment and by attempts at immunomodulation. Radiotherapy reduced the T-helper cell function for almost a decade, but not those of macrophages or NK cells. T-cell changes have no prognostic implication, supporting, perhaps, the suggestion of a major role for macrophages and NK cells. Cyclic adjuvant chemotherapy reduces the peripheral lymphocyte population and several lymphocyte functions but not NK activity. Most of the parameters were normalized some years following treatment, but NK activity remained elevated and Th/Ts cell ratio was still decreased. This might possibly be taken to support the surveillance role of NK cells. Bestatin increases the frequency of lymphocytes forming rosettes with sheep red blood cells (but not their mitogenic responses), enhances NK activity, and augments the phagocytic capacity of granulocytes and monocytes (but not their cytotoxic activity). 154 references.

  20. Separation of hematopoietic stem cells from human peripheral blood through modified polyurethane foaming membranes.

    PubMed

    Higuchi, Akon; Sekiya, Mayu; Gomei, Yumiko; Sakurai, Masaru; Chen, Wen-Yih; Egashira, Satsuki; Matsuoka, Yuki

    2008-06-15

    Cell separation from peripheral blood was investigated using polyurethane (PU) foam membranes having 5.2 mum pore size and coated with Pluronic F127 or hyaluronic acid. The permeation ratio of hematopoietic stem cells (CD34(+) cells) and lymphocytes through the membranes was lower than for red blood cells and platelets. Adhered cells were detached from membrane surfaces using human serum albumin (HSA) solution after permeation of blood through the membranes, allowing isolation of CD34(+) cells in the permeate (recovery) solution. High-yield isolation of CD34(+) cells was achieved using Pluronic-coated membranes. This was because the Pluronic coating dissolved into the recovery solution at 4 degrees C, releasing adhered cells from the surfaces of the membranes during permeation of HSA solution through these membranes. Dextran and/or bovine serum albumin solutions were also evaluated for use as recovery solutions after blood permeation. A high recovery ratio of CD34(+) cells was achieved at 4 degrees C in a process using 20% dextran solution through PU membranes having carboxylic acid groups.

  1. The Submerged Printing of Cells onto a Modified Surface Using a Continuous Flow Microspotter

    PubMed Central

    Davidoff, Sherry N.; Miles, Adam R.; Romanov, Valentin; Gale, Bruce K.; Eckman, Josh W.; Brooks, Benjamin D.

    2014-01-01

    The printing of cells for microarray applications possesses significant challenges including the problem of maintaining physiologically relevant cell phenotype after printing, poor organization and distribution of desired cells, and the inability to deliver drugs and/or nutrients to targeted areas in the array. Our 3D microfluidic printing technology is uniquely capable of sealing and printing arrays of cells onto submerged surfaces in an automated and multiplexed manner. The design of the microfluidic cell array (MFCA) 3D fluidics enables the printhead tip to be lowered into a liquid-filled well or dish and compressed against a surface to form a seal. The soft silicone tip of the printhead behaves like a gasket and is able to form a reversible seal by applying pressure or backing away. Other cells printing technologies such as pin or ink-jet printers are unable to print in submerged applications. Submerged surface printing is essential to maintain phenotypes of cells and to monitor these cells on a surface without disturbing the material surface characteristics. By printing onto submerged surfaces, cell microarrays are produced that allow for drug screening and cytotoxicity assessment in a multitude of areas including cancer, diabetes, inflammation, infections, and cardiovascular disease. PMID:24796939

  2. The submerged printing of cells onto a modified surface using a continuous flow microspotter.

    PubMed

    Davidoff, Sherry N; Miles, Adam R; Romanov, Valentin; Gale, Bruce K; Eckman, Josh W; Brooks, Benjamin D

    2014-04-22

    The printing of cells for microarray applications possesses significant challenges including the problem of maintaining physiologically relevant cell phenotype after printing, poor organization and distribution of desired cells, and the inability to deliver drugs and/or nutrients to targeted areas in the array. Our 3D microfluidic printing technology is uniquely capable of sealing and printing arrays of cells onto submerged surfaces in an automated and multiplexed manner. The design of the microfluidic cell array (MFCA) 3D fluidics enables the printhead tip to be lowered into a liquid-filled well or dish and compressed against a surface to form a seal. The soft silicone tip of the printhead behaves like a gasket and is able to form a reversible seal by applying pressure or backing away. Other cells printing technologies such as pin or ink-jet printers are unable to print in submerged applications. Submerged surface printing is essential to maintain phenotypes of cells and to monitor these cells on a surface without disturbing the material surface characteristics. By printing onto submerged surfaces, cell microarrays are produced that allow for drug screening and cytotoxicity assessment in a multitude of areas including cancer, diabetes, inflammation, infections, and cardiovascular disease.

  3. Microtopography of metal surfaces influence fibroblast growth by modifying cell shape, cytoskeleton, and adhesion.

    PubMed

    Meredith, David O; Eschbach, Lukas; Riehle, Mathis O; Curtis, Adam S G; Richards, Robert G

    2007-11-01

    Stainless Steel (SS), titanium (cpTi), and Ti-6Al-7Nb (TAN) are frequently used metals in fracture fixation, which contact not only bone, but also soft tissue. In previous soft tissue cytocompatibility studies, TAN was demonstrated to inhibit cell growth in its "standard" micro-roughened state. To elucidate a possible mechanism for this inhibition, cell area, shape, adhesion, and cytoskeletal integrity was studied. Only minor changes in spreading were observed for cells on electropolished SS, cpTi, and TAN. Cells on "standard" cpTi were similarly spread in comparison with electropolished cpTi and TAN, although the topography influenced the cell periphery and also resulted in lower numbers and shorter length of focal adhesions. On "standard" microrough TAN, cell spreading was significantly lower than all other surfaces, and cell morphology differed by being more elongated. In addition, focal adhesion numbers and mean length were significantly lower on standard TAN than on all other surfaces, with 80% of the measured adhesions below a 2-microm threshold. Focal adhesion site location and maturation and microtubule integrity were compromised by the presence of protruding beta-phase microspikes found solely on the surface of standard TAN. This led us to propose that the impairment of focal adhesion numbers, maturation (length), and cell spreading to a possibly sufficient threshold observed on standard TAN blocks cell cycle progress and eventually cell growth on the surface. We believe, as demonstrated with standard cpTi and TAN, that a difference in surface morphology is influential for controlling cell behavior on implant surfaces.

  4. Achieving high efficiency laminated polymer solar cell with interfacial modified metallic electrode and pressure induced crystallization

    NASA Astrophysics Data System (ADS)

    Yuan, Yongbo; Bi, Yu; Huang, Jinsong

    2011-02-01

    We report efficient laminated organic photovoltaic device with efficiency approach the optimized device by regular method based on Poly(3-hexylthiophene-2,5-diyl) and [6,6]-phenyl-C61-butyric acid methyl ester (PCBM). The high efficiency is mainly attributed to the formation of a concrete polymer/metal interface mechanically and electrically by the use of electronic-glue, and using the highly conductive and flexible silver film as anode to reduce photovoltage loss and modifying its work function for efficiency hole extraction by ultraviolet/ozone treatment, and the pressure induced crystallization of PCBM.

  5. Ghrelin promotes oral tumor cell proliferation by modifying GLUT1 expression.

    PubMed

    Kraus, Dominik; Reckenbeil, Jan; Wenghoefer, Matthias; Stark, Helmut; Frentzen, Matthias; Allam, Jean-Pierre; Novak, Natalija; Frede, Stilla; Götz, Werner; Probstmeier, Rainer; Meyer, Rainer; Winter, Jochen

    2016-03-01

    In our study, ghrelin was investigated with respect to its capacity on proliferative effects and molecular correlations on oral tumor cells. The presence of all molecular components of the ghrelin system, i.e., ghrelin and its receptors, was analyzed and could be detected using real-time PCR and immunohistochemistry. To examine cellular effects caused by ghrelin and to clarify downstream-regulatory mechanisms, two different oral tumor cell lines (BHY and HN) were used in cell culture experiments. Stimulation of either cell line with ghrelin led to a significantly increased proliferation. Signal transduction occurred through phosphorylation of GSK-3β and nuclear translocation of β-catenin. This effect could be inhibited by blocking protein kinase A. Glucose transporter1 (GLUT1), as an important factor for delivering sufficient amounts of glucose to tumor cells having high requirements for this carbohydrate (Warburg effect) was up-regulated by exogenous and endogenous ghrelin. Silencing intracellular ghrelin concentrations using siRNA led to a significant decreased expression of GLUT1 and proliferation. In conclusion, our study describes the role for the appetite-stimulating peptide hormone ghrelin in oral cancer proliferation under the particular aspect of glucose uptake: (1) tumor cells are a source of ghrelin. (2) Ghrelin affects tumor cell proliferation through autocrine and/or paracrine activity. (3) Ghrelin modulates GLUT1 expression and thus indirectly enhances tumor cell proliferation. These findings are of major relevance, because glucose uptake is assumed to be a promising target for cancer treatment.

  6. Prostaglandins modify phosphorylation of specific proteins in the insect cell line BCIRL-HzAM1

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Prostaglandins (PGs) play crucial roles in vertebrate biology, particularly in immune functions. Because PGs also mediate specific cell functions in insect immunity, we are investigating how these signaling molecules affect insect cells. We reported that PGs, notably PGA1, PGA2, and PGE1, up and/or ...

  7. Rechargeable silver-modified mercuric oxide-zinc cell for cardiac pacemakers.

    PubMed

    Tyers, G F; Hughes, H C; Brownlee, R R; Manley, N J; Gorman, I N

    1976-11-04

    Tests were conducted on rechargeable mercury-zinc pacemaker batteries under simulated and actual biologic conditions, using a variety of discharge rates and charging schedules. In tests on 96 cells at a 6.4 milliampere (ma) discharge, recharging once every 15 months of simulated pacing at a 25 microampere (mua) drain, the earliest cell failure occurred after an equivalent of 50 years of pacing. The mean pacing equivalent for all 96 cells was more than 140 years. In 6.4 ma discharge tests on 24 cells, recharging once every 8 days of simulated pacing, only 1 cell in 24 failed after an equivalent of more than 500 years of pacing (actual time 2 years). In tests on 13 cells pacing at a 200 mua drain without recharging, the simulated mean duration of pacing before total discharge was 4.8 years. Seven other cells at a 200 mua drain with periodic recharging continue to function normally after more than 7 years of actual time, simulating 56 years of pacing at a 25 mua drain. Cardiac pacemakers using the rechargeable mercury-zinc cell have been implanted in animals for more than 2 1/2 years and in patients for more than 1 year with all units continuing to function satisfactorily. It has been demonstrated unequivocally that a rechargeable mercury-zinc pacemaker will function continuously for more than 4 years without recharging and that periodic recharging will extend pacing life far beyond that predicted for lithium and nuclear primary power sources.

  8. Epithelial cell morphology and adhesion on diamond films deposited and chemically modified by plasma processes.

    PubMed

    Rezek, Bohuslav; Ukraintsev, Egor; Krátká, Marie; Taylor, Andrew; Fendrych, Frantisek; Mandys, Vaclav

    2014-09-01

    The authors show that nanocrystalline diamond (NCD) thin films prepared by microwave plasma enhanced chemical vapor deposition apparatus with a linear antenna delivery system are well compatible with epithelial cells (5637 human bladder carcinoma) and significantly improve the cell adhesion compared to reference glass substrates. This is attributed to better adhesion of adsorbed layers to diamond as observed by atomic force microscopy (AFM) beneath the cells. Moreover, the cell morphology can be adjusted by appropriate surface treatment of diamond by using hydrogen and oxygen plasma. Cell bodies, cytoplasmic rims, and filopodia were characterized by Peakforce AFM. Oxidized NCD films perform better than other substrates under all conditions (96% of cells adhered well). A thin adsorbed layer formed from culture medium and supplemented with fetal bovine serum (FBS) covered the diamond surface and played an important role in the cell adhesion. Nevertheless, 50-100 nm large aggregates formed from the RPMI medium without FBS facilitated cell adhesion also on hydrophobic hydrogenated NCD (increase from 23% to 61%). The authors discuss applicability for biomedical uses.

  9. Cell adhesion and growth on surfaces modified by plasma and ion implantation

    NASA Astrophysics Data System (ADS)

    Araujo, W. W. R.; Teixeira, F. S.; da Silva, G. N.; Salvadori, D. M. F.; Salvadori, M. C.

    2014-04-01

    In this study, we show and discuss the results of the interaction of living CHO (Chinese Hamster Ovary) cells, in terms of adhesion and growth on glass, SU-8 (epoxi photoresist), PDMS (polydimethylsiloxane), and DLC (hydrogen free diamond-like carbon) surfaces. Glass, SU-8, and DLC but not PDMS showed to be good surfaces for cell growth. DLC surfaces were treated by oxygen plasma (DLC-O) and sulfur hexafluoride plasma (DLC-F). After 24 h of cell culture, the number of cells on DLC-O was higher than on DLC-F surface. SU-8 with silver implanted, creating nanoparticles 12 nm below the surface, increased significantly the number of cells per unit area.

  10. Graphene Oxide Nanosheets Modified with Single Domain Antibodies for Rapid and Efficient Capture of Cells**

    PubMed Central

    Chen, Guan-Yu; Li, Zeyang; Theile, Christopher S.; Bardhan, Neelkanth M.; Kumar, Priyank V.; Duarte, Joao N.; Maruyama, Takeshi; Rashidfarrokh, Ali; Belcher, Angela M.

    2015-01-01

    Peripheral blood can provide valuable information on an individual’s immune status. Cell-based assays typically target leukocytes and their products. Characterization of leukocytes from whole blood requires their separation from the far more numerous red blood cells.[1] Current methods to classify leukocytes, such as recovery on antibody-coated beads or fluorescence-activated cell sorting require long sample preparation times and relatively large sample volumes.[2] A simple method that enables the characterization of cells from a small peripheral whole blood sample could overcome limitations of current analytical techniques. We describe the development of a simple graphene oxide surface coated with single domain antibody fragments. This format allows quick and efficient capture of distinct WBC subpopulations from small samples (~30 μL) of whole blood in a geometry that does not require any specialized equipment such as cell sorters or microfluidic devices. PMID:26472062

  11. Effect of 2′-O-methyl/thiophosphonoacetate-modified antisense oligonucleotides on huntingtin expression in patient-derived cells

    PubMed Central

    Matsui, Masayuki; Threlfall, Richard N; Caruthers, Marvin H; Corey, David R

    2014-01-01

    ABSTRACT Optimizing oligonucleotides as therapeutics will require exploring how chemistry can be used to enhance their effects inside cells. To achieve this goal it will be necessary to fully explore chemical space around the native DNA/RNA framework to define the potential of diverse chemical modifications. In this report we examine the potential of thiophosphonoacetate (thioPACE)-modified 2′-O-methyl oligoribonucleotides as inhibitors of human huntingtin (HTT) expression. Inhibition occurred, but was less than with analogous locked nucleic acid (LNA)-substituted oligomers lacking the thioPACE modification. These data suggest that thioPACE oligonucleotides have the potential to control gene expression inside cells. However, advantages relative to other modifications were not demonstrated. Additional modifications are likely to be necessary to fully explore any potential advantages of thioPACE substitutions. PMID:26865404

  12. Induction of apoptosis in cancer cells through N-acetyl-l-leucine-modified polyethylenimine-mediated p53 gene delivery.

    PubMed

    Li, Zhiyuan; Zhang, Liu; Li, Quanshun

    2015-11-01

    Herein, N-acetyl-L-leucine-modified polyethylenimine was successfully constructed through the EDC/NHS-mediated coupling reaction and employed as vectors to accomplish p53 gene delivery using HeLa (p53wt) and PC-3 cells (p53null) as models. Compared with PEI25K, the derivatives exhibited lower cytotoxicity, protein adsorption and hemolytic activity, together with satisfactory pDNA condensation capability and gene transfection efficiency. After p53 transfection, MTT analysis confirmed that the cell proliferation was inhibited. Flow cytometric analysis showed that the derivative-mediated p53 delivery could induce stronger early apoptosis than PEI25K and Lipofectamine(2000). Further, PC-3 cells showed higher sensitivity to the exogenous p53 transfection than HeLa cells. The mechanism for inducing apoptosis was determined to be up-regulation of p53 expression at both mRNA and protein levels using RT-PCR and western blotting analysis. Expression level and activity analysis of caspase-3, -8 and -9, and mitochondrial membrane potential measurement revealed that p53 transfection mediated by these derivatives facilitated early apoptosis of tumor cells via a mitochondria-dependent apoptosis pathway. Thus, the derivatives showed potential as biocompatible carriers for realizing effective tumor gene therapy.

  13. Data set for comparison of cellular dynamics between human AAVS1 locus-modified and wild-type cells

    PubMed Central

    Mizutani, Takeomi; Haga, Hisashi; Kawabata, Kazushige

    2016-01-01

    This data article describes cellular dynamics, such as migration speed and mobility of the cytoskeletal protein, of wild-type human fibroblast cells and cells with a modified adeno-associated virus integration site 1 (AAVS1) locus on human chromosome 19. Insertion of exogenous gene into the AAVS1 locus has been conducted in recent biological researches. Previously, our data showed that the AAVS1-modification changes cellular contractile force (Mizutani et al., 2015 [1]). To assess if this AAVS1-modification affects cell migration, we compared cellular migration speed and turnover of cytoskeletal protein in human fibroblasts and fibroblasts with a green fluorescent protein gene knocked-in at the AAVS1 locus in this data article. Cell nuclei were stained and changes in their position attributable to cell migration were analyzed. Fluorescence recovery was observed after photobleaching for the fluorescent protein-tagged myosin regulatory light chain. Data here are related to the research article “Transgene Integration into the Human AAVS1 Locus Enhances Myosin II-Dependent Contractile Force by Reducing Expression of Myosin Binding Subunit 85” [1]. PMID:26937449

  14. Effect of native and modified bull seminal ribonuclease on tumour and testicular cells and phytohaemagglutinin-stimulated pig lymphocytes.

    PubMed

    Matousek, J; Stanĕk, R; Dostál, J; Mácha, J

    1979-01-01

    The carboxymethylated, oxidized and reduced forms of AS RNase inhibited transplantability and DNA synthesis of tumour cells BP-8 and EL-4 incubated in vitro. With tumour cells EL-4 the results under in vitro conditions did not not correspond to those obtained under the conditions in vivo. The survival of mice given injections of EL-4 cells and of the native and carboxymethylated AS RNase was only slightly prolonged. Mice that received intra-abdominally BP-8 cells and both carboxymethylated and oxidized and reduced forms of AS RNase survived two or three times longer than the controls. Succinylation and maleylation of AS RNase eliminated any antitumoral effect. Aspermatogenic activity of AS RNase was abolished by any modification of the molecule which had substantially reduced, or removed, the RNase activity. Neither native nor modified forms of AS RNase had an inhibitory effect on unstimulated pig lymphocytes. The DNA synthesis of PHA-stimulated lymphocytes was inhibited by the native and carboxymethylated AS RNase only. Bovine pancreatic A RNase had any inhibitory effect on neither tumour nor testicular cells.

  15. C(5) modified uracil derivatives showing antiproliferative and erythroid differentiation inducing activities on human chronic myelogenous leukemia K562 cells

    PubMed Central

    Brognara, Eleonora; Lampronti, Ilaria; Breveglieri, Giulia; Accetta, Alessandro; Corradini, Roberto; Manicardi, Alex; Borgatti, Monica; Canella, Alessandro; Multineddu, Chiara; Marchelli, Rosangela; Gambari, Roberto

    2011-01-01

    The K562 cell line has been proposed as a useful experimental system to identify anti-tumor compounds acting by inducing terminal erythroid differentiation. K562 cells exhibit a low proportion of hemoglobin-synthesizing cells under standard cell growth conditions, but are able to undergo terminal erythroid differentiation when treated with a variety of anti-tumor compounds. In this paper we report a screening study on a set of different modified C(5) uracil derivatives for the evaluation of their antiproliferative effect in connection with erythroid differentiation pathways, and for defining a new class of drug candidates for the treatment of chronic myelogenous leukemia. Activity of the derivatives tested can be classified in two effect: an antiproliferative effect linked to a high level of erythroid differentiation activity and an antiproliferative effect without activation of gamma globin genes The highest antiproliferative effect and erythroid induction was shown by compound 9, a thymine derivative bearing a n-octyl chain on nitrogen N(1), whereas thymine did not show any effect, suggesting the importance of the linear alkyl chain in position N(1). To our knowledge this compound should be considered among the most efficient inducers of erythroid differentiation of K562 cells. This work is the starting point for the quest of more effective and specific drugs for the induction of terminal erythroid differentiation, for leading new insights in the treatment of neoplastic diseases with molecules acting by inducing differentiation rather than by simply exerting cytotoxic effects. PMID:21958870

  16. HUMAN T CELLS UPREGULATE CD69 AFTER COCULTURE WITH XENOGENEIC GENETICALLY-MODIFIED PIG MESENCHYMAL STROMAL CELLS

    PubMed Central

    Li, Jiang; Andreyev, Oleg; Chen, Man; Marco, Michael; Iwase, Hayato; Long, Cassandra; Ayares, David; Shen, Zhongyang; Cooper, David K.C.; Ezzelarab, Mohamed B.

    2013-01-01

    Mesenchymal stromal cells (MSC) obtained from α1,3-galactosyltransferase gene knock-out pigs transgenic for the human complement-regulatory protein CD46 (GTKO/CD46 pMSC) suppress in vitro human anti-pig cellular responses as efficiently as allogeneic human MSC. We investigated the immunoregulatory effects of GTKO/CD46 pMSC on human CD4+ and CD8+ T cell proliferation in response to pig aortic endothelial cells (pAEC). pMSC efficiently suppressed T cell proliferation, which was associated with downregulation of granzyme B expression. No induction of CD4+CD25+Foxp3hi regulatory T cells or T cell apoptosis was documented. In correlation with T cell proliferation, CD25 expression was upregulated on T cells in response to pAEC but not to pMSC. In contrast, CD69 expression was upregulated on T cells in response to both pMSC and pAEC, which was associated with a significant increase in the phosphorylation of STAT5. GTKO/CD46 pMSC possibly regulate human T cell responses through modulation of CD69 expression and STAT5 signaling. PMID:24044963

  17. A Novel Approach to Decrease Sialic Acid Expression in Cells by a C-3-modified N-Acetylmannosamine*

    PubMed Central

    Wratil, Paul R.; Rigol, Stephan; Solecka, Barbara; Kohla, Guido; Kannicht, Christoph; Reutter, Werner; Giannis, Athanassios; Nguyen, Long D.

    2014-01-01

    Due to its position at the outermost of glycans, sialic acid is involved in a myriad of physiological and pathophysiological cell functions such as host-pathogen interactions, immune regulation, and tumor evasion. Inhibitors of cell surface sialylation could be a useful tool in cancer, immune, antibiotic, or antiviral therapy. In this work, four different C-3 modified N-acetylmannosamine analogs were tested as potential inhibitors of cell surface sialylation. Peracetylated 2-acetylamino-2-deoxy-3-O-methyl-d-mannose decreases cell surface sialylation in Jurkat cells in a dose-dependent manner up to 80%, quantified by flow cytometry and enzyme-linked lectin assays. High-performance liquid chromatography experiments revealed that not only the concentration of membrane bound but also of cytosolic sialic acid is reduced in treated cells. We have strong evidence that the observed reduction of sialic acid expression in cells is caused by the inhibition of the bifunctional enzyme UDP-GlcNAc-2-epimerase/ManNAc kinase. 2-Acetylamino-2-deoxy-3-O-methyl-d-mannose inhibits the human ManNAc kinase domain of the UDP-GlcNAc-2-epimerase/ManNAc kinase. Binding kinetics of the inhibitor and human N-acetylmannosamine kinase were evaluated using surface plasmon resonance. Specificity studies with human N-acetylglucosamine kinase and hexokinase IV indicated a high specificity of 2-acetylamino-2-deoxy-3-O-methyl-d-mannose for MNK. This substance represents a novel class of inhibitors of sialic acid expression in cells, targeting the key enzyme of sialic acid de novo biosynthesis. PMID:25278018

  18. Lipopolysaccharide (LPS) processing by Kupffer cells releases a modified LPS with increased hepatocyte binding and decreased tumor necrosis factor-alpha stimulatory capacity.

    PubMed

    Treon, S P; Thomas, P; Broitman, S A

    1993-02-01

    Normal physiological clearance of gut-derived endotoxin lipopolysaccharide [LPS] has been described previously; initially, there is uptake by Kupffer cells (KC), then release of modified LPS, followed by hepatocyte uptake. Previous work in our laboratories indicated that LPS is structurally modified with loss of carbohydrate prior to its release by KC. In this study, we functionally characterize KC modified LPS. KC-modified 125I-LPS was prepared from primary rat KC. Escherichia coli 0127:B8 native 125I-LPS or KC-modified 125I-LPS (40 ng) was incubated for 1 hr with 1 x 10E6 primary hepatocytes. The binding of KC-modified LPS was 4.33-fold higher than native LPS (P = 0.0024). Binding analysis studies were conducted to determine the region of KC-modified LPS responsible for enhanced hepatocyte binding. KC-modified Salmonella minnesota LPS was competed with 100-fold excess native or mutant (Ra, Rc, Rd, or Re) strains of LPS or Lipid A with no decrease to hepatocyte binding. S. minnesota-native 125I-LPS was compared with KC-modified 125I-LPS in a study to assess induction of tumor necrosis factor (TNF)-gamma by rat peritoneal macrophages. Native or KC-modified 125I-LPS (100 ng) was presented to 1 x 10E7 peritoneal macrophages for 6 hr. TNF-alpha was measured in supernatants using the WEHI-164 cytotoxicity assay. Native LPS induced 5.7-fold higher TNF-alpha levels than KC-modified LPS (P < 0.0001). The above data suggest that structural alterations in KC-modified LPS are accompanied by functional alterations resulting in enhanced hepatocyte binding and decreased TNF-alpha release. The latter result implies that an early step in LPS detoxification occurs in the KC in which LPS is modified to prevent elicitation of biologically active cytokines.

  19. Platelet-derived growth factor is a potent biologic response modifier of T cells

    PubMed Central

    1991-01-01

    Freshly isolated lymph node (LN) cells cultured in serum-containing medium were restricted to produce primarily interleukin 2 (IL-2) subsequent to T cell activation. Only minimal amounts of IL-4, IL-5, or interferon gamma (IFN-gamma) were produced under these conditions. Similar populations of LN cells cultured in serum-free medium were able to produce a variety of lymphokines after T cell activation, with the relative quantities of each species being dependent upon the lymphoid organ source of the lymphocytes. A similar relationship in the patterns of lymphokines produced by activated T cell hybridomas maintained under serum-free conditions was also observed, whereas activation in serum- supplemented media resulted in a predominant restriction to the secretion of IL-2. Additional studies determined that the entity in serum responsible for restricting T cell function in vitro was platelet- derived growth factor (PDGF). The PDGF-BB isoform was established to be the most active in the regulation of T cell function, enhancing IL-2 while depressing the production of IL-4, IL-5, and IFN-gamma at concentrations below 1 ng/ml. PDGF-AB was also found to be quite active, however, this isoform of PDGF was incapable of influencing IFN- gamma production at the concentrations tested. PDGF-AA was very weakly active. It therefore appears that PDGF, acting primarily through a beta receptor subunit (either alpha/beta- or beta/beta-type receptors) is able to influence profoundly the behavior of T cells, with some of its modulatory effects exhibiting isoform specificity. This is reflected by an enhancement in the production of IL-2, while simultaneously depressing the secretion of IL-4, IL-5, and IFN-gamma (PDGF-BB only) after T cell activation. Kinetic studies, where cell supernatants were analyzed both 24 and 48 h after T cell activation, suggested that "desensitization" to PDGF influences can occur naturally in vitro. Those species of lymphokines that were inhibited by PDGF over

  20. Proteomes of Host Cell Membranes Modified by Intracellular Activities of Salmonella enterica*

    PubMed Central

    Vorwerk, Stephanie; Krieger, Viktoria; Deiwick, Jörg; Hensel, Michael; Hansmeier, Nicole

    2015-01-01

    Intracellular pathogens need to establish a growth-stimulating host niche for survival and replication. A unique feature of the gastrointestinal pathogen Salmonella enterica serovar Typhimurium is the creation of extensive membrane networks within its host. An understanding of the origin and function of these membranes is crucial for the development of new treatment strategies. However, the characterization of this compartment is very challenging, and only fragmentary knowledge of its composition and biogenesis exists. Here, we describe a new proteome-based approach to enrich and characterize Salmonella-modified membranes. Using a Salmonella mutant strain that does not form this unique membrane network as a reference, we identified a high-confidence set of host proteins associated with Salmonella-modified membranes. This comprehensive analysis allowed us to reconstruct the interactions of Salmonella with host membranes. For example, we noted that Salmonella redirects endoplasmic reticulum (ER) membrane trafficking to its intracellular niche, a finding that has not been described for Salmonella previously. Our system-wide approach therefore has the potential to rapidly close gaps in our knowledge of the infection process of intracellular pathogens and demonstrates a hitherto unrecognized complexity in the formation of Salmonella host niches. PMID:25348832

  1. The Banana Transcriptional Repressor MaDEAR1 Negatively Regulates Cell Wall-Modifying Genes Involved in Fruit Ripening

    PubMed Central

    Fan, Zhong-qi; Kuang, Jian-fei; Fu, Chang-chun; Shan, Wei; Han, Yan-chao; Xiao, Yun-yi; Ye, Yu-jie; Lu, Wang-jin; Lakshmanan, Prakash; Duan, Xue-wu; Chen, Jian-ye

    2016-01-01

    Ethylene plays an essential role in many biological processes including fruit ripening via modulation of ethylene signaling pathway. Ethylene Response Factors (ERFs) are key transcription factors (TFs) involved in ethylene perception and are divided into AP2, RAV, ERF, and DREB sub-families. Although a number of studies have implicated the involvement of DREB sub-family genes in stress responses, little is known about their roles in fruit ripening. In this study, we identified a DREB TF with a EAR motif, designated as MaDEAR1, which is a nucleus-localized transcriptional repressor. Expression analysis indicated that MaDEAR1 expression was repressed by ethylene, with reduced levels of histone H3 and H4 acetylation at its regulatory regions during fruit ripening. In addition, MaDEAR1 promoter activity was also suppressed in response to ethylene treatment. More importantly, MaDEAR1 directly binds to the DRE/CRT motifs in promoters of several cell wall-modifying genes including MaEXP1/3, MaPG1, MaXTH10, MaPL3, and MaPME3 associated with fruit softening during ripening and represses their activities. These data suggest that MaDEAR1 acts as a transcriptional repressor of cell wall-modifying genes, and may be negatively involved in ethylene-mediated ripening of banana fruit. Our findings provide new insights into the involvement of DREB TFs in the regulation of fruit ripening. PMID:27462342

  2. Effects of surface-modified photoelectrode on the power conversion efficiency of dye-sensitized solar cells

    NASA Astrophysics Data System (ADS)

    Kim, Jong Tae; Han, Yoon Soo

    2014-05-01

    The effects of Na2SO4 as a surface modification material on the performance of dye-sensitized solar cells (DSSCs) were studied. The surfaces of TiO2 films were firstly modified with aqueous Na2SO4 solution by a dip coating process, and then the resulting electrode was applied to the photoelectrode of a DSSC. The DSSC with the Na2SO4-modified photoelectrode had a power conversion efficiency of 9.01% compared with that (7.97%) of the reference cell, which corresponds to an increase of about 13.0% in the efficiency due to an enhancement in short-circuit current ( J sc ) and open-circuit voltage ( V oc ). A series of measurements such as UV-visible absorption, electrochemical impedance, incident photon to current conversion (IPCE) efficiency and dark current revealed that incorporation of Na2SO4 onto the TiO2 film led to an increase of dye adsorption and a longer lifetime of electrons injected from dyes to the TiO2 electrodes, resulting in the improvement in both J sc and V oc , compared to those of a reference device without surface modification.

  3. ATP activates P2x receptors and requires extracellular Ca(++) participation to modify outer hair cell nonlinear capacitance.

    PubMed

    Yu, Ning; Zhao, Hong-Bo

    2008-11-01

    Intracochlear ATP is an important mediator in regulating hearing function. ATP can activate ionotropic purinergic (P2x) and metabotropic purinergic (P2y) receptors to influence cell functions. In this paper, we report that ATP can activate P2x receptors directly to modify outer hair cell (OHC) electromotility, which is an active cochlear amplifier determining hearing sensitivity and frequency selectivity in mammals. We found that ATP, but not UTP, a P2y receptor agonist, reduced the OHC electromotility-associated nonlinear capacitance (NLC) and shifted its voltage dependence to the right (depolarizing) direction. Blockage of the activation of P2x receptors by pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS), suramin, and 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid (DIDS) could block the ATP effect. This modification also required extracellular Ca(++) participation. Removal of extracellular Ca(++) abolished the ATP effect. However, chelation of intracellular Ca(++) concentration by a fast calcium-chelating reagent 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA, 10 mM) did not affect the effect of ATP on NLC. The effect is also independent of K(+) ions. Substitution of Cs(+) for intracellular or extracellular K(+) did not affect the ATP effect. Our findings indicate that ATP activates P2x receptors instead of P2y receptors to modify OHC electromotility. Extracellular Ca(++) is required for this modification.

  4. Genetically modified canine Schwann cells--In vitro and in vivo evaluation of their suitability for peripheral nerve tissue engineering.

    PubMed

    Schmitte, Ruth; Tipold, Andrea; Stein, Veronika M; Schenk, Henning; Flieshardt, Cornelia; Grothe, Claudia; Haastert, Kirsten

    2010-02-15

    After peripheral nerve injury, Schwann cells (SC) guarantee for a regeneration-promoting milieu and are crucially involved in axonal regeneration. For extended nerve defects, bridging with an autologous nerve transplant is the gold standard therapy. Artificial biohybrid nerve transplants which combine a synthetic conduit with autologous SC genetically modified to express regeneration-promoting proteins may provide an alternative therapy to autotransplantation. The dog seems to be an ideal translational animal model for new treatment strategies. In the present study, utilizing a new transfection protocol, we transplanted enhanced green fluorescent protein (EGFP)-expressing adult canine SC (cSC) into a 5mm epineural pouch in the sciatic nerve of adult rats (n=9). The epineurial pouch technique serves as proof of principle to follow the fate of the transplanted cSC for up to 14 days after surgery. Fluorescence microscopy and immunohistochemistry revealed survival and integration of EGFP-expressing cSC into the regenerating host nerve tissue. We demonstrate that transplanted cSC contribute to the formation of bands of Büngner and are located in close vicinity to growth-associated protein-43 (GAP-43) expressing regenerating nerve fibers. This provides first evidence that transplanted genetically modified Schwann cells do successfully integrate into the host tissue where they could actively contribute to the regeneration process.

  5. The Banana Transcriptional Repressor MaDEAR1 Negatively Regulates Cell Wall-Modifying Genes Involved in Fruit Ripening.

    PubMed

    Fan, Zhong-Qi; Kuang, Jian-Fei; Fu, Chang-Chun; Shan, Wei; Han, Yan-Chao; Xiao, Yun-Yi; Ye, Yu-Jie; Lu, Wang-Jin; Lakshmanan, Prakash; Duan, Xue-Wu; Chen, Jian-Ye

    2016-01-01

    Ethylene plays an essential role in many biological processes including fruit ripening via modulation of ethylene signaling pathway. Ethylene Response Factors (ERFs) are key transcription factors (TFs) involved in ethylene perception and are divided into AP2, RAV, ERF, and DREB sub-families. Although a number of studies have implicated the involvement of DREB sub-family genes in stress responses, little is known about their roles in fruit ripening. In this study, we identified a DREB TF with a EAR motif, designated as MaDEAR1, which is a nucleus-localized transcriptional repressor. Expression analysis indicated that MaDEAR1 expression was repressed by ethylene, with reduced levels of histone H3 and H4 acetylation at its regulatory regions during fruit ripening. In addition, MaDEAR1 promoter activity was also suppressed in response to ethylene treatment. More importantly, MaDEAR1 directly binds to the DRE/CRT motifs in promoters of several cell wall-modifying genes including MaEXP1/3, MaPG1, MaXTH10, MaPL3, and MaPME3 associated with fruit softening during ripening and represses their activities. These data suggest that MaDEAR1 acts as a transcriptional repressor of cell wall-modifying genes, and may be negatively involved in ethylene-mediated ripening of banana fruit. Our findings provide new insights into the involvement of DREB TFs in the regulation of fruit ripening.

  6. Enhancement of radiotherapy by ceria nanoparticles modified with neogambogic acid in breast cancer cells

    PubMed Central

    Chen, Feng; Zhang, Xiao Hong; Hu, Xiao Dan; Zhang, Wei; Lou, Zhi Chao; Xie, Li Hua; Liu, Pei Dang; Zhang, Hai Qian

    2015-01-01

    Radiotherapy is one of the main strategies for cancer treatment but has significant challenges, such as cancer cell resistance and radiation damage to normal tissue. Radiosensitizers that selectively increase the susceptibility of cancer cells to radiation can enhance the effectiveness of radiotherapy. We report here the development of a novel radiosensitizer consisting of monodispersed ceria nanoparticles (CNPs) covered with the anticancer drug neogambogic acid (NGA-CNPs). These were used in conjunction with radiation in MCF-7 breast cancer cells, and the efficacy and mechanisms of action of this combined treatment approach were evaluated. NGA-CNPs potentiated the toxic effects of radiation, leading to a higher rate of cell death than either treatment used alone and inducing the activation of autophagy and cell cycle arrest at the G2/M phase, while pretreatment with NGA or CNPs did not improve the rate of radiation-induced cancer cells death. However, NGA-CNPs decreased both endogenous and radiation-induced reactive oxygen species formation, unlike other nanomaterials. These results suggest that the adjunctive use of NGA-CNPs can increase the effectiveness of radiotherapy in breast cancer treatment by lowering the radiation doses required to kill cancer cells and thereby minimizing collateral damage to healthy adjacent tissue. PMID:26316742

  7. RAD001 (everolimus) induces dose-dependent changes to cell cycle regulation and modifies the cell cycle response to vincristine.

    PubMed

    Saunders, P O; Weiss, J; Welschinger, R; Baraz, R; Bradstock, K F; Bendall, L J

    2013-10-01

    More than 50% of adults and ~20% of children with pre-B acute lymphoblastic leukemia (ALL) relapse following treatment. Dismal outcomes for patients with relapsed or refractory disease mandate novel approaches to therapy. We have previously shown that the combination of the mTOR inhibitor RAD001 (everolimus) and the chemotherapeutic agent vincristine increases the survival of non-obese diabetic/severe combined immuno-deficient (NOD/SCID) mice bearing human ALL xenografts. We have also shown that 16 μM RAD001 synergized with agents that cause DNA damage or microtubule disruption in pre-B ALL cells in vitro. Here, we demonstrate that RAD001 has dose-dependent effects on the cell cycle in ALL cells, with 1.5 μM RAD001 inhibiting pRb, Ki67 and PCNA expression and increasing G0/1 cell cycle arrest, whereas 16 μM RAD001 increases pRb, cyclin D1, Ki67 and PCNA, with no evidence of an accumulation of cells in G0/1. Transition from G2 into mitosis was promoted by 16 μM RAD001 with reduced phosphorylation of cdc2 in cells with 4 N DNA content. However, 16 μM RAD001 preferentially induced cell death in cells undergoing mitosis. When combined with vincristine, 16 μM RAD001 reduced the vincristine-induced accumulation of cells in mitosis, probably as a result of increased death in this population. Although 16 μM RAD001 weakly activated Chk1 and Chk2, it suppressed strong vincristine-induced activation of these cell cycle checkpoint regulators. We conclude that RAD001 enhances chemosensitivity at least in part through suppression of cell cycle checkpoint regulation in response to vincristine and increased progression from G2 into mitosis.

  8. Inkjet Gene Printing: A Novel Approach to Achieve Gene Modified Cells for Tissue Engineering

    DTIC Science & Technology

    2008-12-01

    regeneration. ACKNOWLEDGMENTS The authors wish to thank Dr. Jennifer Olson for editorial assistance with this manuscript. The authors thank Mary ...and D. Gospodarowicz , 1987: Capillary endothelial cells express basic fibroblast growth factor, a mitogen that 8 promotes their own growth

  9. Antibody-modified T cells: CARs take the front seat for hematologic malignancies

    PubMed Central

    Maus, Marcela V.; Grupp, Stephan A.; Porter, David L.

    2014-01-01

    T cells redirected to specific antigen targets with engineered chimeric antigen receptors (CARs) are emerging as powerful therapies in hematologic malignancies. Various CAR designs, manufacturing processes, and study populations, among other variables, have been tested and reported in over 10 clinical trials. Here, we review and compare the results of the reported clinical trials and discuss the progress and key emerging factors that may play a role in effecting tumor responses. We also discuss the outlook for CAR T-cell therapies, including managing toxicities and expanding the availability of personalized cell therapy as a promising approach to all hematologic malignancies. Many questions remain in the field of CAR T cells directed to hematologic malignancies, but the encouraging response rates pave a wide road for future investigation. PMID:24578504

  10. Antibody-modified T cells: CARs take the front seat for hematologic malignancies.

    PubMed

    Maus, Marcela V; Grupp, Stephan A; Porter, David L; June, Carl H

    2014-04-24

    T cells redirected to specific antigen targets with engineered chimeric antigen receptors (CARs) are emerging as powerful therapies in hematologic malignancies. Various CAR designs, manufacturing processes, and study populations, among other variables, have been tested and reported in over 10 clinical trials. Here, we review and compare the results of the reported clinical trials and discuss the progress and key emerging factors that may play a role in effecting tumor responses. We also discuss the outlook for CAR T-cell therapies, including managing toxicities and expanding the availability of personalized cell therapy as a promising approach to all hematologic malignancies. Many questions remain in the field of CAR T cells directed to hematologic malignancies, but the encouraging response rates pave a wide road for future investigation.

  11. Surface-modified complex SU-8 microstructures for indirect optical manipulation of single cells

    PubMed Central

    Aekbote, Badri L.; Fekete, Tamás; Jacak, Jaroslaw; Vizsnyiczai, Gaszton; Ormos, Pál; Kelemen, Lóránd

    2015-01-01

    We introduce a method that combines two-photon polymerization (TPP) and surface functionalization to enable the indirect optical manipulation of live cells. TPP-made 3D microstructures were coated specifically with a multilayer of the protein streptavidin and non-specifically with IgG antibody using polyethylene glycol diamine as a linker molecule. Protein density on their surfaces was quantified for various coating methods. The streptavidin-coated structures were shown to attach to biotinated cells reproducibly. We performed basic indirect optical micromanipulation tasks with attached structure-cell couples using complex structures and a multi-focus optical trap. The use of such extended manipulators for indirect optical trapping ensures to keep a safe distance between the trapping beams and the sensitive cell and enables their 6 degrees of freedom actuation. PMID:26819816

  12. Chimeric antigen receptor-modified T cells for the immunotherapy of patients with EGFR-expressing advanced relapsed/refractory non-small cell lung cancer.

    PubMed

    Feng, Kaichao; Guo, Yelei; Dai, Hanren; Wang, Yao; Li, Xiang; Jia, Hejin; Han, Weidong

    2016-05-01

    The successes achieved by chimeric antigen receptor-modified T (CAR-T) cells in hematological malignancies raised the possibility of their use in non-small lung cancer (NSCLC). In this phase I clinical study (NCT01869166), patients with epidermal growth factor receptor (EGFR)-positive (>50% expression), relapsed/refractory NSCLC received escalating doses of EGFR-targeted CAR-T cell infusions. The EGFR-targeted CAR-T cells were generated from peripheral blood after a 10 to 13-day in vitro expansion. Serum cytokines in peripheral blood and copy numbers of CAR-EGFR transgene in peripheral blood and in tissue biopsy were monitored periodically. Clinical responses were evaluated with RECIST1.1 and immune- related response criteria, and adverse events were graded with CTCAE 4.0. The EGFR-targeted CAR-T cell infusions were well-tolerated without severe toxicity. Of 11 evaluable patients, two patients obtained partial response and five had stable disease for two to eight months. The median dose of transfused CAR(+) T cells was 0.97×10(7) cells kg(-1) (interquartile range (IQR), 0.45 to 1.09×10(7) cells kg(-1)). Pathological eradication of EGFR positive tumor cells after EGFR-targeted CAR-T cell treatment can be observed in tumor biopsies, along with the CAR-EGFR gene detected in tumor-infiltrating T cells in all four biopsied patients. The EGFR-targeted CAR-T cell therapy is safe and feasible for EGFR-positive advanced relapsed/refractory NSCLC.

  13. Enhanced compatibility of chemically modified titanium surface with periodontal ligament cells

    NASA Astrophysics Data System (ADS)

    Kado, T.; Hidaka, T.; Aita, H.; Endo, K.; Furuichi, Y.

    2012-12-01

    A simple chemical modification method was developed to immobilize cell-adhesive molecules on a titanium surface to improve its compatibility with human periodontal ligament cells (HPDLCs).The polished titanium disk was immersed in 1% (v/v) p-vinylbenzoic acid solution for 2 h to introduce carboxyl groups onto the surface. After rinsing with distilled deionized water, the titanium disk was dipped into 1.47% 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide solution containing 0.1 mg/ml Gly-Arg-Gly-Asp-Ser (GRGDS), human plasma fibronectin (pFN), or type I collagen from calf skin (Col) to covalently immobilize the cell-adhesive molecules on the titanium surface via formation of peptide bonds. X-ray photoelectron spectroscopy analyses revealed that cell-adhesive molecules were successfully immobilized on the titanium surfaces. The Col-immobilized titanium surface revealed higher values regarding nano rough characteristics than the as-polished titanium surface under scanning probe microscopy. The number of HPDLCs attached to both the pFN- and Col-immobilized titanium surfaces was twice that attached to the as-polished titanium surfaces. The cells were larger with the cellular processes that stretched to a greater extent on the pFN- and Col-immobilized titanium surfaces than on the as-polished titanium surface (p < 0.05). HPDLCs on the Col-immobilized titanium surfaces showed more extensive expression of vinculin at the tips of cell projections and more contiguously along the cell outline than on the as-polished, GRGDS-immobilized and pFN-immobilized titanium surfaces. It was concluded that cell-adhesive molecules successfully immobilized on the titanium surface and improved the compatibility of the surface with HPDLCs. The Col-immobilized titanium surface could be used for forming ligament-like tissues around titanium dental implants.

  14. Transvers Impedance Measurements of the Modified DARHT-2Accelerator Cell Design

    SciTech Connect

    Briggs, Dick; Waldron, Will

    2005-11-30

    The DARHT-2 accelerator cells have been redesigned to make their high voltage performance more robust. At the outset of the DARHT-2 development program about 8 years ago, an extensive campaign was mounted to minimize the transverse impedance of the original cell design. Since the initial spec on the machine was a beam current of 4 kA, the control of beam-breakup (BBU) amplification with a 2 microsecond pulse length was considered to be one the most critical issues in the design. Even after advances in detector technology allowed the beam current requirement to be lowered to 2 kA, the goal for the standard cell impedance was kept at {approx}300 ohms/meter to allow for the possibility of future beam current upgrades to 4 kA without any modifications in the cells. The results of this campaign to minimize the transverse impedance are described in detail in Reference 1. After several iterations in the design of ferrite dampers and the anode finger stock shape, the measured (peak) impedance of the original standard cell was determined to be about 280 ohms/meter. (As a reference point, the measured impedance of the DARHT-1 cell is about 880 ohms/meter). This impedance provided such a wide safety margin against BBU amplification at 2 kA that it was felt that the cell redesign could focus on voltage holding without any detailed considerations of impacts on the transverse impedance. Now that a baseline design for the DARHT-2 cell has been established and tested, however, it was felt that a measurement of its impedance would be prudent. The results of these impedance measurements are presented in this note. The objective was mainly to do a ''quick check'' to ensure that there were no surprises, and to provide an estimate of the BBU frequencies and growth rates to the experimental test program.

  15. Deregulation of polycomb repressor complex 1 modifier AUTS2 in T-cell leukemia

    PubMed Central

    Nagel, Stefan; Pommerenke, Claudia; Meyer, Corinna; Kaufmann, Maren; Drexler, Hans G.; MacLeod, Roderick A.F.

    2016-01-01

    Recently, we identified deregulated expression of the B-cell specific transcription factor MEF2C in T-cell acute lymphoid leukemia (T-ALL). Here, we performed sequence analysis of a regulatory upstream section of MEF2C in T-ALL cell lines which, however, proved devoid of mutations. Unexpectedly, we found strong conservation between the regulatory upstream region of MEF2C (located at chromosomal band 5q14) and an intergenic stretch at 7q11 located between STAG3L4 and AUTS2, covering nearly 20 kb. While the non-coding gene STAG3L4 was inconspicuously expressed, AUTS2 was aberrantly upregulated in 6% of T-ALL patients (public dataset GSE42038) and in 3/24 T-ALL cell lines, two of which represented very immature differentiation stages. AUTS2 expression was higher in normal B-cells than in T-cells, indicating lineage-specific activity in lymphopoiesis. While excluding chromosomal aberrations, examinations of AUTS2 transcriptional regulation in T-ALL cells revealed activation by IL7-IL7R-STAT5-signalling and MEF2C. AUTS2 protein has been shown to interact with polycomb repressor complex 1 subtype 5 (PRC1.5), transforming this particular complex into an activator. Accordingly, expression profiling and functional analyses demonstrated that AUTS2 activated while PCGF5 repressed transcription of NKL homeobox gene MSX1 in T-ALL cells. Forced expression and pharmacological inhibition of EZH2 in addition to H3K27me3 analysis indicated that PRC2 repressed MSX1 as well. Taken together, we found that AUTS2 and MEF2C, despite lying on different chromosomes, share strikingly similar regulatory upstream regions and aberrant expression in T-ALL subsets. Our data implicate chromatin complexes PRC1/AUTS2 and PRC2 in a gene network in T-ALL regulating early lymphoid differentiation. PMID:27322685

  16. The Influence of Modified Silica Nanomaterials on Adult Stem Cell Culture

    PubMed Central

    Tarpani, Luigi; Morena, Francesco; Gambucci, Marta; Zampini, Giulia; Massaro, Giuseppina; Argentati, Chiara; Emiliani, Carla; Martino, Sabata; Latterini, Loredana

    2016-01-01

    The preparation of tailored nanomaterials able to support cell growth and viability is mandatory for tissue engineering applications. In the present work, silica nanoparticles were prepared by a sol-gel procedure and were then functionalized by condensation of amino groups and by adsorption of silver nanoparticles. Transmission electron microscopy (TEM) imaging was used to establish the morphology and the average dimensions of about 130 nm, which were not affected by the functionalization. The three silica samples were deposited (1 mg/mL) on cover glasses, which were used as a substrate to culture adult human bone marrow-mesenchymal stem cells (hBM-MSCs) and human adipose-derived stem cells (hASCs). The good cell viability over the different silica surfaces was evaluated by monitoring the mitochondrial dehydrogenase activity. The analysis of the morphological parameters (aspect ratio, cell length, and nuclear shape Index) yielded information about the interactions of stem cells with the surface of three different nanoparticles. The data are discussed in terms of chemical properties of the surface of silica nanoparticles.

  17. Surface-modified particles loaded with CaMKII inhibitor protect cardiac cells against mitochondrial injury.

    PubMed

    Wongrakpanich, Amaraporn; Morris, Angie S; Geary, Sean M; Joiner, Mei-Ling A; Salem, Aliasger K

    2017-03-30

    An excess of calcium (Ca(2+)) influx into mitochondria during mitochondrial re-energization is one of the causes of myocardial cell death during ischemic/reperfusion injury. This overload of Ca(2+) triggers the mitochondrial permeability transition pore (mPTP) opening which leads to programmed cell death. During the ischemic/reperfusion stage, the activated Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) enzyme is responsible for Ca(2+) influx. To reduce CaMKII-related cell death, sub-micron particles composed of poly(lactic-co-glycolic acid) (PLGA), loaded with a CaMKII inhibitor peptide were fabricated. The CaMKII inhibitor peptide-loaded (CIP) particles were coated with a mitochondria targeting moiety, triphenylphosphonium cation (TPP), which allowed the particles to accumulate and release the peptide inside mitochondria to inhibit CaMKII activity. The fluorescently labeled TPP-CIP was taken up by mitochondria and successfully reduced reactive oxygen species (ROS) caused by Isoprenaline (ISO) in a differentiated rat cardiomyocyte-like cell line. When cells were treated with TPP-CIP prior to ISO exposure, they maintained mitochondrial membrane potential. The TPP-CIP protected cells from ISO-induced ROS production and decreased mitochondrial membrane potential. Thus, TPP-CIP has the potential to be used in protection against ischemia/reperfusion injury.

  18. Cathepsin E Deficiency Ameliorates Graft-versus-Host Disease and Modifies Dendritic Cell Motility

    PubMed Central

    Mengwasser, Jörg; Babes, Liane; Cordes, Steffen; Mertlitz, Sarah; Riesner, Katarina; Shi, Yu; McGearey, Aleixandria; Kalupa, Martina; Reinheckel, Thomas; Penack, Olaf

    2017-01-01

    Microbial products influence immunity after allogeneic hematopoietic stem cell transplantation (allo-SCT). In this context, the role of cathepsin E (Ctse), an aspartate protease known to cleave bacterial peptides for antigen presentation in dendritic cells (DCs), has not been studied. During experimental acute graft-versus-host disease (GVHD), we found infiltration by Ctse-positive immune cells leading to higher Ctse RNA- and protein levels in target organs. In Ctse-deficient allo-SCT recipients, we found ameliorated GVHD, improved survival, and lower numbers of tissue-infiltrating DCs. Donor T cell proliferation was not different in Ctse-deficient vs. wild-type allo-SCT recipients in MHC-matched and MHC-mismatched models. Furthermore, Ctse-deficient DCs had an intact ability to induce allogeneic T cell proliferation, suggesting that its role in antigen presentation may not be the main mechanism how Ctse impacts GVHD. We found that Ctse deficiency significantly decreases DC motility in vivo, reduces adhesion to extracellular matrix (ECM), and diminishes invasion through ECM. We conclude that Ctse has a previously unrecognized role in regulating DC motility that possibly contributes to reduced DC counts and ameliorated inflammation in GVHD target organs of Ctse-deficient allo-SCT recipients. However, our data do not provide definite proof that the observed effect of Ctse−/− deficiency is exclusively mediated by DCs. A contribution of Ctse−/−-mediated functions in other recipient cell types, e.g., macrophages, cannot be excluded. PMID:28298913

  19. Melanoma Proteoglycan Modifies Gene Expression to Stimulate Tumor Cell Motility, Growth and Epithelial to Mesenchymal Transition

    PubMed Central

    Yang, Jianbo; Price, Matthew A.; Li, GuiYuan; Bar-Eli, Menashe; Salgia, Ravi; Jagedeeswaran, Ramasamy; Carlson, Jennifer H.; Ferrone, Soldano; Turley, Eva A.; McCarthy, James B.

    2009-01-01

    Melanoma chondroitin sulfate proteoglycan (MCSP) is a plasma membrane-associated proteoglycan that facilitates the growth, motility and invasion of tumor cells. MCSP expression in melanoma cells enhances integrin function and constitutive activation of Erk 1,2. The current studies were performed to determine the mechanism by which MCSP expression promotes tumor growth and motility. The results demonstrate that MCSP expression in radial growth phase (RGP), vertical growth phase (VGP) or metastatic cell lines causes sustained activation of Erk 1,2, enhanced growth and motility which all require the cytoplasmic domain of the MCSP core protein. MCSP expression in an RGP cell line also promotes an epithelial to mesenchymal transition (EMT) based on changes in cell morphology and the expression of several EMT markers. Finally MCSP enhances the expression of c-Met and HGF, and inhibiting c-Met expression or activation limits the increased growth and motility of multiple melanoma cell lines. The studies collectively demonstrate an importance for MCSP in promoting progression by an epigenetic mechanism and they indicate that MCSP could be targeted to delay or inhibit tumor progression in patients. PMID:19738072

  20. Adhesion, Proliferation and Migration of NIH/3T3 Cells on Modified Polyaniline Surfaces

    PubMed Central

    Rejmontová, Petra; Capáková, Zdenka; Mikušová, Nikola; Maráková, Nela; Kašpárková, Věra; Lehocký, Marián; Humpolíček, Petr

    2016-01-01

    Polyaniline shows great potential and promises wide application in the biomedical field thanks to its intrinsic conductivity and material properties, which closely resemble natural tissues. Surface properties are crucial, as these predetermine any interaction with biological fluids, proteins and cells. An advantage of polyaniline is the simple modification of its surface, e.g., by using various dopant acids. An investigation was made into the adhesion, proliferation and migration of mouse embryonic fibroblasts on pristine polyaniline films and films doped with sulfamic and phosphotungstic acids. In addition, polyaniline films supplemented with poly (2-acrylamido-2-methyl-1-propanesulfonic) acid at various ratios were tested. Results showed that the NIH/3T3 cell line was able to adhere, proliferate and migrate on the pristine polyaniline films as well as those films doped with sulfamic and phosphotungstic acids; thus, utilization of said forms in biomedicine appears promising. Nevertheless, incorporating poly (2-acrylamido-2-methyl-1-propanesulfonic) acid altered the surface properties of the polyaniline films and significantly affected cell behavior. In order to reveal the crucial factor influencing the surface/cell interaction, cell behavior is discussed in the context of the surface energy of individual samples. It was clearly demonstrated that the lesser the difference between the surface energy of the sample and cell, the more cyto-compatible the surface is. PMID:27649159

  1. Cell Attachment and Viability Study of PCL Nano-fiber Modified by Cold Atmospheric Plasma.

    PubMed

    Atyabi, Seyed Mohammad; Sharifi, Fereshteh; Irani, Shiva; Zandi, Mojgan; Mivehchi, Houri; Nagheh, Zahra

    2016-06-01

    The field of tissue engineering is an emerging discipline which applies the basic principles of life sciences and engineering to repair and restore living tissues and organs. The purpose of this study was to investigate the effect of cold and non-thermal plasma surface modification of poly (ϵ-caprolactone) (PCL) scaffolds on fibroblast cell behavior. Nano-fiber PCL was fabricated through electrospinning technique, and some fibers were then treated by cold and non-thermal plasma. The cell-biomaterial interactions were studied by culturing the fibroblast cells on nano-fiber PCL. Scaffold biocompatibility test was assessed using an inverted microscope. The growth and proliferation of fibroblast cells on nano-fiber PCL were analyzed by MTT viability assay. Cellular attachment on the nano-fiber and their morphology were evaluated using scanning electron microscope. The result of cell culture showed that nano-fiber could support the cellular growth and proliferation by developing three-dimensional topography. The present study demonstrated that the nano-fiber surface modification with cold plasma sharply enhanced the fibroblast cell attachment. Thus, cold plasma surface modification greatly raised the bioactivity of scaffolds.

  2. Localization of collagen modifying enzymes on fibroblastic reticular cells and follicular dendritic cells in non-neoplastic and neoplastic lymphoid tissues.

    PubMed

    Ohe, Rintaro; Aung, Naing Ye; Meng, Hongxue; Kabasawa, Takanobu; Suto, Aya; Tamazawa, Nobuyuki; Yang, Suran; Kato, Tomoya; Yamakawa, Mitsunori

    2016-07-01

    The aim of this study was to evaluate the localization of collagen modifying enzymes (CMEs) on fibroblastic reticular cells (FRCs) and follicular dendritic cells (FDCs) in non-neoplastic lymphoid tissues and various malignant lymphomas. The expression of prolyl 4-hydroxylase 1 (P4H1), lysyl hydroxylase 3 (LH3), and protein disulfide isomerase (PDI) was frequently observed on FRCs and FDCs in the germinal center (GC), except for the mantle zone. The expression of CMEs was lower in most lymphomas than in their respective postulated normal counterparts. The ratio of transglutaminase II(+) FRCs/CD35(+) FDCs was also lower in follicular lymphomas (FL) than in other lymphomas. The mRNAs of some CMEs (P4H1, prolyl 4-hydroxylase 3, LH3, and heat shock protein 47) were confirmed in almost all lymphomas. These results indicate that lymphoma cell proliferation suppresses/decreases the number of CMEs expressing FRCs and FDCs in most lymphomas.

  3. Localization of collagen modifying enzymes on fibroblastic reticular cells and follicular dendritic cells in non-neoplastic and neoplastic lymphoid tissues

    PubMed Central

    Ohe, Rintaro; Aung, Naing Ye; Meng, Hongxue; Kabasawa, Takanobu; Suto, Aya; Tamazawa, Nobuyuki; Yang, Suran; Kato, Tomoya; Yamakawa, Mitsunori

    2016-01-01

    Abstract The aim of this study was to evaluate the localization of collagen modifying enzymes (CMEs) on fibroblastic reticular cells (FRCs) and follicular dendritic cells (FDCs) in non-neoplastic lymphoid tissues and various malignant lymphomas. The expression of prolyl 4-hydroxylase 1 (P4H1), lysyl hydroxylase 3 (LH3), and protein disulfide isomerase (PDI) was frequently observed on FRCs and FDCs in the germinal center (GC), except for the mantle zone. The expression of CMEs was lower in most lymphomas than in their respective postulated normal counterparts. The ratio of transglutaminase II+ FRCs/CD35+ FDCs was also lower in follicular lymphomas (FL) than in other lymphomas. The mRNAs of some CMEs (P4H1, prolyl 4-hydroxylase 3, LH3, and heat shock protein 47) were confirmed in almost all lymphomas. These results indicate that lymphoma cell proliferation suppresses/decreases the number of CMEs expressing FRCs and FDCs in most lymphomas. PMID:26700650

  4. Effect of surface wettability and topography on the adhesion of osteosarcoma cells on plasma-modified polystyrene.

    PubMed

    Dowling, Denis P; Miller, Ian S; Ardhaoui, Malika; Gallagher, William M

    2011-09-01

    Biomaterials interact with the biological environment at their surface, making accurate biophysical characterization of the surface crucially important for understanding subsequent biological effects. In this study, the surface of polystyrene (PS) was systematically altered in order to determine the effect of plasma treatment and surface roughness on cell adhesion and spreading. Surfaces with water contact angle from hydrophilic (12°) to superhydrophobic (155°) were obtained through a combination of modifying surface roughness (R (a)), the deposition of siloxane coatings and the fluorination of the PS surface. R (a) values in the range of 19-2365 nm were obtained by grinding the PS surface. The nanometer-thick siloxane coatings were deposited using an atmospheric pressure plasma system, while the fluorination of the PS was carried out using a low-pressure radio frequency (RF) plasma. The siloxane coatings were obtained using a liquid poly(dimethylsiloxane) precursor that was nebulized into helium or helium/oxygen plasmas. Water contact angles in the range of 12-122° were obtained with these coatings. Cell adhesion studies were carried out using human MG63 osteosarcoma cells. It was observed that higher polymer surface roughness enhanced cell adhesion, but had a negative effect on cell spreading. Optimum cell adhesion was observed at ∼64° for the siloxane coatings, with a decrease in adhesion observed for the more hydrophilic and hydrophobic coatings. This decrease in cell adhesion with an increase in hydrophobicity was also observed for the fluorinated PS surfaces with water contact angles in the range of 110-155°.

  5. Driving vascular endothelial cell fate of human multipotent Isl1+ heart progenitors with VEGF modified mRNA.

    PubMed

    Lui, Kathy O; Zangi, Lior; Silva, Eduardo A; Bu, Lei; Sahara, Makoto; Li, Ronald A; Mooney, David J; Chien, Kenneth R

    2013-10-01

    Distinct families of multipotent heart progenitors play a central role in the generation of diverse cardiac, smooth muscle and endothelial cell lineages during mammalian cardiogenesis. The identification of precise paracrine signals that drive the cell-fate decision of these multipotent progenitors, and the development of novel approaches to deliver these signals in vivo, are critical steps towards unlocking their regenerative therapeutic potential. Herein, we have identified a family of human cardiac endothelial intermediates located in outflow tract of the early human fetal hearts (OFT-ECs), characterized by coexpression of Isl1 and CD144/vWF. By comparing angiocrine factors expressed by the human OFT-ECs and non-cardiac ECs, vascular endothelial growth factor (VEGF)-A was identified as the most abundantly expressed factor, and clonal assays documented its ability to drive endothelial specification of human embryonic stem cell (ESC)-derived Isl1+ progenitors in a VEGF receptor-dependent manner. Human Isl1-ECs (endothelial cells differentiated from hESC-derived ISL1+ progenitors) resemble OFT-ECs in terms of expression of the cardiac endothelial progenitor- and endocardial cell-specific genes, confirming their organ specificity. To determine whether VEGF-A might serve as an in vivo cell-fate switch for human ESC-derived Isl1-ECs, we established a novel approach using chemically modified mRNA as a platform for transient, yet highly efficient expression of paracrine factors in cardiovascular progenitors. Overexpression of VEGF-A promotes not only the endothelial specification but also engraftment, proliferation and survival (reduced apoptosis) of the human Isl1+ progenitors in vivo. The large-scale derivation of cardiac-specific human Isl1-ECs from human pluripotent stem cells, coupled with the ability to drive endothelial specification, engraftment, and survival following transplantation, suggest a novel strategy for vascular regeneration in the heart.

  6. Cerium Oxide Nanoparticle Modified Scaffold Interface Enhances Vascularization of Bone Grafts by Activating Calcium Channel of Mesenchymal Stem Cells.

    PubMed

    Xiang, Junyu; Li, Jianmei; He, Jian; Tang, Xiangyu; Dou, Ce; Cao, Zhen; Yu, Bo; Zhao, Chunrong; Kang, Fei; Yang, Lu; Dong, Shiwu; Yang, Xiaochao

    2016-02-01

    Insufficient blood perfusion is one of the critical problems that hamper the clinical application of tissue engineering bone (TEB). Current methods for improving blood vessel distribution in TEB mainly rely on delivering exogenous angiogenic factors to promote the proliferation, migration, differentiation, and vessel formation of endothelial cells (ECs) and/or endothelial progenitor cells (EPCs). However, obstacles including limited activity preservation, difficulty in controlled release, and high cost obstructed the practical application of this strategy. In this study, TEB scaffold were modified with cerium oxide nanoparticles (CNPs) and the effects of CNPs existed at the scaffold surface on the growth and paracrine behavior of mesenchymal stem cells (MSCs) were investigated. The CNPs could improve the proliferation and inhibit the apoptosis of MSCs. Meanwhile, the interaction between the cell membrane and the nanoparticle surface could activate the calcium channel of MSCs leading to the rise of intracellular free Ca(2+) level, which subsequently augments the stability of HIF-1α. These chain reactions finally resulted in high expression of angiogenic factor VEGF. The improved paracrine of VEGF could thereby promote the proliferation, differentiation, and tube formation ability of EPCs. Most importantly, in vivo ectopic bone formation experiment demonstrated this method could significantly improve the blood vessel distribution inside of TEB.

  7. Human Schwann Cells Seeded on a Novel Collagen-Based Microstructured Nerve Guide Survive, Proliferate, and Modify Neurite Outgrowth

    PubMed Central

    van Neerven, Sabien G. A.; Haastert-Talini, Kirsten; Tolba, René H.; Pallua, Norbert; Bozkurt, Ahmet

    2014-01-01

    A variety of new bioartificial nerve guides have been tested preclinically for their safety and nerve regeneration supporting properties. So far, only a limited number of biomaterials have been tested in humans since the step from preclinical work to a clinical application is challenging. We here present an in vitro model with human Schwann cells (hSCs) as an intermediate step towards clinical application of the nerve guide Perimaix, a collagen-based microstructured 3D scaffold containing numerous longitudinal guidance channels for directed axonal growth. hSCs were seeded onto different prototypes of Perimaix and cultivated for 14 days. hSC adhered to the scaffold, proliferated, and demonstrated healthy Schwann cell morphology (spindle shaped cell bodies, bipolar oriented processes) not only at the surface of the material, but also in the deeper layers of the scaffold. The general well-being of the cells was quantitatively confirmed by low levels of lactate dehydrogenase release into the culture medium. Moreover, conditioned medium of hSCs that were cultivated on Perimaix was able to modify neurite outgrowth from sensory dorsal root ganglion neurons. Overall these data indicate that Perimaix is able to provide a matrix that can promote the attachment and supports process extension, migration, and proliferation of hSC. PMID:24895582

  8. Enhanced cellular uptake and anti-proliferating effect of chitosan hydrochlorides modified genistein loaded NLC on human lens epithelial cells.

    PubMed

    Zhang, Wenji; Liu, Jinlu; Zhang, Qi; Li, Xuedong; Yu, Shihui; Yang, Xinggang; Kong, Jun; Pan, Weisan

    2014-08-25

    This study was attempted to increase the cellular uptake of developed genistein loaded nanostructured lipid carriers (NLC) into human lens epithelial (HLE) cells by chitosan hydrochlorides coatings when applied in post lens capsule (PCO) treatment, and to provide further understanding of the uptake and anti-proliferation mechanisms inside. NLCs were produced using melt-emulsification method and were subsequently coated with chitosan hydrochlorides by adsorption. The uptake of various particle sizes were evaluated and visualized by confocal laser scanning microscopy (CLSM), showing a size-dependent manner. The uptake of NLC was proved to be endocytosed in an energy dependent and clathrin-mediated endocytosis to HLE cells by the decrease in uptake at lower temperature, when pre-saturated by blank NLC and in the presence of NaN3 and sucrose. CH coating improved the uptake percentage of NLC irrespective of the particle size, without influencing the uptake mechanism. Cell apoptosis was tested using PI and Annexin V-FITC/PI staining, followed by flow cytometer analysis. Higher anti-proliferation effect was observed for CH-NLC in inhibiting the growth of HLE cells by causing more apoptosis. Results above indicate that GEN-NLC surface modified by chitosan hydrochlorides could enhance the trans-cellular performance and anti-proliferating effect as PCO therapy.

  9. In vivo invasion of modified chorioallantoic membrane by tumor cells: the role of cell surface-bound urokinase

    PubMed Central

    1988-01-01

    The ability of the chick embryo chorioallantoic membrane (CAM) to withstand invasion by tumor cells can be intentionally compromised by altering its morphological integrity. Using a newly developed quantitative assay of invasion we showed that intact CAMs were completely resistant to invasion by tumor cells, wounded CAMs did not pose a barrier to penetration, and CAMs that were wounded and then allowed to reseal displayed partial susceptibility to invasion. The invasion of resealed CAMs required catalytically active plasminogen activator (PA) of the urokinase type (uPA); the invasive efficiency of tumor cells was reduced by 75% when tumor uPA activity or tumor uPA production was inhibited. The invasive ability of human tumor cells, which have surface uPA receptors but which do not produce the enzyme, could be augmented by saturating their receptors with exogenous uPA. The mere stimulation of either uPA or tissue plasminogen activator production, in absence of binding to cell receptors, did not result in an enhancement of invasiveness. These findings suggest that the increased invasive potential of tumor cells is correlated with cell surface-associated proteolytic activity stemming from the interaction between uPA and its surface receptor. PMID:2848851

  10. Comparison of Behaviour in Different Liquids and in Cells of Gold Nanorods and Spherical Nanoparticles Modified by Linear Polyethyleneimine and Bovine Serum Albumin

    PubMed Central

    Pyshnaya, Inna A.; Razum, Kristina V.; Poletaeva, Julia E.; Pyshnyi, Dmitrii V.; Zenkova, Marina A.; Ryabchikova, Elena I.

    2014-01-01

    Gold nanorods (GNRs) are considered one of the most promising forms of nanoparticles for nanobiotechnology; however, the problem of their toxicity is currently not resolved. We synthesised GNRs, modified with linear polyethyleneimine (PEI-GNRs), and examined their physicochemical and some biological properties in comparison with GNRs modified with BSA and spherical gold nanoparticles (sGNPs) modified with the same agents. The influence of the buffer, cell culture media, and serum on hydrodynamic diameter and zeta potential of all GNPs was studied. Simultaneously, the size, shape, and formation of a corona were examined by transmission electron microscopy (TEM). PEI-GNRs and GNPs were nontoxic for BHK-21 and HeLa cells (MTT test). Penetration of all GNPs into BHK-21, melanoma B16, and HeLa cells was examined after 30 min, 3 h, and 24 h of incubation using TEM ultrathin sections. PEI-GNRs and PEI-sGNPs demonstrated fast and active penetration into cells by caveolin-dependent and lipid raft-mediated endocytosis and accumulated in endosomes and lysosomes. BSA-modified GNPs showed prolonged flotation and a significant delay in cell penetration. The results show that the charge of initial NPs determines penetration into cells. Thus, the designed PEI-GNRs were nontoxic and stable in cell culture media and could efficiently penetrate cells. PMID:25093190

  11. Modified Sagnac interferometer for contact-free length measurement of a direct absorption cell.

    PubMed

    Elandaloussi, Hadj; Rouillé, Christian; Marie-Jeanne, Patrick; Janssen, Christof

    2016-03-10

    Accurate path length measurements in absorption cells are recurrent requirements in quantitative molecular absorption spectroscopy. A new twin path laser interferometer for length measurements in a simple direct path absorption geometry is presented, along with a full uncertainty budget. The path in an absorption cell is determined by measuring the optical path length change due to the diminution of the refractive index when the cell originally filled with nitrogen gas is evacuated. The performance of the instrument based on a stabilized HeNe laser is verified by comparison with the results of direct mechanical length measurements of a roughly 45 mm long, specially designed absorption cell. Due to a resolution of about 1/300 of a HeNe fringe, an expanded (coverage factor k=2) uncertainty of 16 μm in the length measurement is achieved, providing an expanded relative uncertainty of 3.6·10⁻⁴ for the length of our test absorption cell. This value is about 8 times lower than what has been reported previously. The instrument will be useful for precision measurements of absorption cross sections of strong absorbers which require short light paths, such as ozone, halogen oxides, sulfur dioxide, and volatile organic compounds in the UV.

  12. A commercially available cell culture device modified for dentin barrier tests.

    PubMed

    Schmalz, G; Garhammer, P; Schweiki, H

    1996-05-01

    The suitability of a dentin barrier test based on a commercially available cell culture chamber was evaluated by testing the cytotoxicity of dental cements. The two chambers of the culture device as produced are separated by a membrane. This was replaced by a bovine dentin disk (500 micrometers thick). Mouse fibroblasts were grown on the "pulpal" side of the dentin for 24 h; test materials were then placed into the "cavity" side of the upper chamber. The number of viable cells was determined after 24 h. After exposure to zinc phosphate cement at a powder/liquid ratio of 2:1, approximately 100% of cells survived. A ratio of 1:1 yielded 81% survival. Only 24% and 28% of the cells survived after exposure to Ketac Fil and Ketac Silver, respectively. The light-curing glass ionomer cement (vitrebond) and zinc oxide-eugenol killed all cells. These results agree with those obtained from a previous study, wherein the dentin barrier test device was constructed in our laboratory.

  13. Oryzalin-modified disruption of microtubular cytoskeleton in Arabidopsis thaliana root cells under clinorotation

    NASA Astrophysics Data System (ADS)

    Kalinina, Ia.; Shevchenko, G.; Kordyum, E.

    There are data on gravisensitivity of cells not specialized to perceive a gravity vector but the molecular processes by which gravity affects not graviperceptive cells are still unclear Spaceflight experiments show that the microtubule self-organization in vitro is gravity-dependent Confocal microscopic analysis of the microtubule spatial organization under altered gravity with combination of approach drugs that disrupt normal microtubule behavior should give us a better understanding of the possible role of microtubule cytoskeleton in gravisensing on cellular level With this aim we examined influence of horizontal clinorotation 2 rpm on the spatial organization of microtubules in the root cortical and epidermal cells by means of LSM 5 PASCAL Zeiss Germany Microtubules were visualized by using stably transformed line of transgenic Arabidopsis thaliana expressing a green fluorescent protein-MAP4 fusion protein We inhibited microtubule function applying 5 956 M L oryzalin microtubule inhibitor in control and clinorotated seedlings Preliminary investigations show that cortical microtubule arrays were dense and predominantly transverse to the root long axis in the meristem and distal elongation zone in control and they got oblique direction when rapid cell elongation is finishing In the differentiation zone microtubules reorient with respect to the longitudinal growth axis of cell Under clinorotation cortical microtubules have the same configuration in the meristem central elongation zone and differentiation zone but it is observed appearances of several

  14. Fabricating upconversion fluorescent nanoparticles modified substrate for dynamical control of cancer cells and pathogenic bacteria.

    PubMed

    Li, Huanhuan; Chen, Quansheng; Zhao, Jiewen; Urmila, Khulal

    2016-09-07

    Lanthanide-doped upconversion nanoparticles (UCNPs) have attracted widespread interests in the field of biomedicine because of their unique upconverting capability by converting near infrared (NIR) excitation to visible or ultraviolet (UV) emission. Here, we developed a novel UCNP-based substrate for dynamic capture and release of cancer cells and pathogenic bacteria under NIR-control. The UCNPs harvest NIR light and convert it to ultraviolet light, which subsequently result in the cleavage of photoresponsive linker (PR linker) from the substrate, and on demand allows the release of a captured cell. The results show that after seeding cells for 5 h, the cells were efficiently captured on the surface of the substrate and ˜89.4% of the originally captured S. aureus was released from the surface after exposure to 2 W/cm(2) NIR light for 30 min, and ˜92.1% of HepG2 cells. These findings provide a unique platform for exploring an entirely new application field for this promising luminescent nanomaterial.

  15. An OGA-Resistant Probe Allows Specific Visualization and Accurate Identification of O-GlcNAc-Modified Proteins in Cells.

    PubMed

    Li, Jing; Wang, Jiajia; Wen, Liuqing; Zhu, He; Li, Shanshan; Huang, Kenneth; Jiang, Kuan; Li, Xu; Ma, Cheng; Qu, Jingyao; Parameswaran, Aishwarya; Song, Jing; Zhao, Wei; Wang, Peng George

    2016-11-18

    O-linked β-N-acetyl-glucosamine (O-GlcNAc) is an essential and ubiquitous post-translational modification present in nucleic and cytoplasmic proteins of multicellular eukaryotes. The metabolic chemical probes such as GlcNAc or GalNAc analogues bearing ketone or azide handles, in conjunction with bioorthogonal reactions, provide a powerful approach for detecting and identifying this modification. However, these chemical probes either enter multiple glycosylation pathways or have low labeling efficiency. Therefore, selective and potent probes are needed to assess this modification. We report here the development of a novel probe, 1,3,6-tri-O-acetyl-2-azidoacetamido-2,4-dideoxy-d-glucopyranose (Ac34dGlcNAz), that can be processed by the GalNAc salvage pathway and transferred by O-GlcNAc transferase (OGT) to O-GlcNAc proteins. Due to the absence of a hydroxyl group at C4, this probe is less incorporated into α/β 4-GlcNAc or GalNAc containing glycoconjugates. Furthermore, the O-4dGlcNAz modification was resistant to the hydrolysis of O-GlcNAcase (OGA), which greatly enhanced the efficiency of incorporation for O-GlcNAcylation. Combined with a click reaction, Ac34dGlcNAz allowed the selective visualization of O-GlcNAc in cells and accurate identification of O-GlcNAc-modified proteins with LC-MS/MS. This probe represents a more potent and selective tool in tracking, capturing, and identifying O-GlcNAc-modified proteins in cells and cell lysates.

  16. Gene Expression Profiles of Human Adipose Tissue-Derived Mesenchymal Stem Cells Are Modified by Cell Culture Density

    PubMed Central

    Yoo, Keon Hee; Lee, Tae-Hee; Kim, Hye Jin; Jang, In Keun; Chun, Yong Hoon; Kim, Hyung Joon; Park, Seung Jo; Lee, Soo Hyun; Son, Meong Hi; Jung, Hye Lim; Sung, Ki Woong; Koo, Hong Hoe

    2014-01-01

    Previous studies conducted cell expansion ex vivo using low initial plating densities for optimal expansion and subsequent differentiation of mesenchymal stem cells (MSCs). However, MSC populations are heterogeneous and culture conditions can affect the characteristics of MSCs. In this study, differences in gene expression profiles of adipose tissue (AT)-derived MSCs were examined after harvesting cells cultured at different densities. AT-MSCs from three different donors were plated at a density of 200 or 5,000 cells/cm2. After 7 days in culture, detailed gene expression profiles were investigated using a DNA chip microarray, and subsequently validated using a reverse transcription polymerase chain reaction (RT-PCR) analysis. Gene expression profiles were influenced primarily by the level of cell confluence at harvest. In MSCs harvested at ∼90% confluence, 177 genes were up-regulated and 102 genes down-regulated relative to cells harvested at ∼50% confluence (P<0.05, FC>2). Proliferation-related genes were highly expressed in MSCs harvested at low density, while genes that were highly expressed in MSCs harvested at high density (∼90% confluent) were linked to immunity and defense, cell communication, signal transduction and cell motility. Several cytokine, chemokine and growth factor genes involved in immunosuppression, migration, and reconstitution of damaged tissues were up-regulated in MSCs harvested at high density compared with MSCs harvested at low density. These results imply that cell density at harvest is a critical factor for modulating the specific gene-expression patterns of heterogeneous MSCs. PMID:24400072

  17. Effective capture of proteins inside living cells by antibodies indirectly linked to a novel cell-penetrating polymer-modified protein A derivative.

    PubMed

    Itakura, Shoko; Hama, Susumu; Ikeda, Hisafumi; Mitsuhashi, Naoto; Majima, Eiji; Kogure, Kentaro

    2015-01-01

    Antibodies against cytoplasmic proteins are useful tools that can control cellular function and clarify signaling mechanisms. However, it is difficult to capture proteins inside living cells, and thus appropriate methods for antibody delivery to the cytoplasm of living cells are required. Cell-penetrating materials, such as the TAT-peptide, have received attention for their ability to deliver various cargos into living cells. However, the direct modification of cargos with cell-penetrating materials is time-consuming and lacks versatility. Therefore, we conceived that protein A, which can bind to the fragment crystallizable region of an antibody, could indirectly link antibodies with cell-penetrating materials, creating an efficient and simple antibody delivery system. Here, we constructed a novel antibody delivery system using a cell-penetrating polymer-modified protein A derivative (CPP-pAd). Living cells treated with CPP-pAd/antibody complexes showed significantly higher antibody levels than those achieved with the commercially available reagent HVJ-E. Pre-treatment with sucrose prevented cellular uptake of the CPP-pAd/antibody complex, suggesting that the CPP-pAd/antibody internalization mechanism occurs through clathrin-dependent endocytosis. Interestingly, intracellularly delivered antibodies did not colocalize with endosome/lysosome markers, further suggesting that antibodies were delivered to the cytoplasm by escape from endosome/lysosome. Moreover, we observed that anti-nuclear pore complex antibodies, delivered to cells using CPP-pAd, localized to the nuclear membrane and inhibited nuclear factor κB dependent luciferase activity. Together, these results suggest that the antibodies delivered by CPP-pAd captured functional proteins, making CPP-pAd a promising strategy for effective capture of proteins inside living cells.

  18. Suicide gene-modified killer cells as an allogeneic alternative to autologous cytokine-induced killer cell immunotherapy of hepatocellular carcinoma.

    PubMed

    Wu, Tao; Leboeuf, Céline; Durand, Sarah; Su, Bin; Deschamps, Marina; Zhang, Xiaowen; Ferrand, Christophe; Pessaux, Patrick; Robinet, Eric

    2016-03-01

    Adoptive immunotherapy using autologous cytokine-induced killer (CIK) cells reduces the recurrence rate of hepatocellular carcinoma (HCC) in association with transarterial chemoembolization or radiofrequency. However, a large‑scale development of this immunotherapy remains difficult to consider in an autologous setting, considering the logistical hurdles associated with the production of this cell therapy product. A previous study has provided the in vitro and in vivo proof‑of‑concept that allogeneic suicide gene‑modified killer cells (aSGMKCs) from healthy blood donors (a cell therapy product previously demonstrated to provide anti‑leukemic effects to patients receiving allogeneic hematopoietic transplantation) may exert a potent anti‑tumor effect towards HCC. Therefore, the development of a bank of 'ready‑for‑use' aSGMKCs was proposed as an approach allowing for the development of immunotherapies that are more convenient and on a broader scale than that of autologous therapies. In the present study, aSGMKCs were compared with CIK cells generated according to three different protocols. Similar to CIK cells, the cytotoxic activity of aSGMKCs toward the Huh‑7 HCC cell line was mediated by tumor necrosis factor‑related apoptosis‑inducing ligand, tumor necrosis factor‑α and interferon‑γ. Furthermore, the frequency of natural killer (NK), NK‑like T and T cells, and their in vitro and in vivo cytotoxicity activities were similar between aSGMKCs and CIK cells. Thus, the present study demonstrated that aSGMKCs are similar to CIK cells, further suggesting the possibility for future use of aSGMKCs in the treatment of solid tumors, including HCC.

  19. P2X7 receptors trigger ATP exocytosis and modify secretory vesicle dynamics in neuroblastoma cells.

    PubMed

    Gutiérrez-Martín, Yolanda; Bustillo, Diego; Gómez-Villafuertes, Rosa; Sánchez-Nogueiro, Jesús; Torregrosa-Hetland, Cristina; Binz, Thomas; Gutiérrez, Luis Miguel; Miras-Portugal, María Teresa; Artalejo, Antonio R

    2011-04-01

    Previously, we reported that purinergic ionotropic P2X7 receptors negatively regulate neurite formation in Neuro-2a (N2a) mouse neuroblastoma cells through a Ca(2+)/calmodulin-dependent kinase II-related mechanism. In the present study we used this cell line to investigate a parallel though faster P2X7 receptor-mediated signaling pathway, namely Ca(2+)-regulated exocytosis. Selective activation of P2X7 receptors evoked exocytosis as assayed by high resolution membrane capacitance measurements. Using dual-wavelength total internal reflection microscopy, we have observed both the increase in near-membrane Ca(2+) concentration and the exocytosis of fluorescently labeled vesicles in response to P2X7 receptor stimulation. Moreover, activation of P2X7 receptors also affects vesicle motion in the vertical and horizontal directions, thus, involving this receptor type in the control of early steps (docking and priming) of the secretory pathway. Immunocytochemical and RT-PCR experiments evidenced that N2a cells express the three neuronal SNAREs as well as vesicular nucleotide and monoamine (VMAT-1 and VMAT-2) transporters. Biochemical measurements indicated that ionomycin induced a significant release of ATP from N2a cells. Finally, P2X7 receptor stimulation and ionomycin increased the incidence of small transient inward currents, reminiscent of postsynaptic quantal events observed at synapses. Small transient inward currents were dependent on extracellular Ca(2+) and were abolished by Brilliant Blue G, suggesting they were mediated by P2X7 receptors. Altogether, these results suggest the existence of a positive feedback mechanism mediated by P2X7 receptor-stimulated exocytotic release of ATP that would act on P2X7 receptors on the same or neighbor cells to further stimulate its own release and negatively control N2a cell differentiation.

  20. P2X7 Receptors Trigger ATP Exocytosis and Modify Secretory Vesicle Dynamics in Neuroblastoma Cells*

    PubMed Central

    Gutiérrez-Martín, Yolanda; Bustillo, Diego; Gómez-Villafuertes, Rosa; Sánchez-Nogueiro, Jesús; Torregrosa-Hetland, Cristina; Binz, Thomas; Gutiérrez, Luis Miguel; Miras-Portugal, María Teresa; Artalejo, Antonio R.

    2011-01-01

    Previously, we reported that purinergic ionotropic P2X7 receptors negatively regulate neurite formation in Neuro-2a (N2a) mouse neuroblastoma cells through a Ca2+/calmodulin-dependent kinase II-related mechanism. In the present study we used this cell line to investigate a parallel though faster P2X7 receptor-mediated signaling pathway, namely Ca2+-regulated exocytosis. Selective activation of P2X7 receptors evoked exocytosis as assayed by high resolution membrane capacitance measurements. Using dual-wavelength total internal reflection microscopy, we have observed both the increase in near-membrane Ca2+ concentration and the exocytosis of fluorescently labeled vesicles in response to P2X7 receptor stimulation. Moreover, activation of P2X7 receptors also affects vesicle motion in the vertical and horizontal directions, thus, involving this receptor type in the control of early steps (docking and priming) of the secretory pathway. Immunocytochemical and RT-PCR experiments evidenced that N2a cells express the three neuronal SNAREs as well as vesicular nucleotide and monoamine (VMAT-1 and VMAT-2) transporters. Biochemical measurements indicated that ionomycin induced a significant release of ATP from N2a cells. Finally, P2X7 receptor stimulation and ionomycin increased the incidence of small transient inward currents, reminiscent of postsynaptic quantal events observed at synapses. Small transient inward currents were dependent on extracellular Ca2+ and were abolished by Brilliant Blue G, suggesting they were mediated by P2X7 receptors. Altogether, these results suggest the existence of a positive feedback mechanism mediated by P2X7 receptor-stimulated exocytotic release of ATP that would act on P2X7 receptors on the same or neighbor cells to further stimulate its own release and negatively control N2a cell differentiation. PMID:21292765

  1. Genetically modified CHO cells for studying the genotoxicity of heterocyclic amines from cooked foods

    SciTech Connect

    Thompson, L.H.; Wu, R.W.; Felton, J.S.

    1995-07-01

    We have developed metabolically competent CHO cells to evaluate the genotoxicity associated with heterocyclic amines, such as those that are present in cooked foods. Into repair-deficient UV5 cells we introduced cDNAs for expressing cytochrome P450IA2 and acetyltransferases. We then genetically reverted these transformed lines to obtain matched metabolically competent repair-deficient/proficient lines. For a high mutagenic response, we find a requirement for acetyltransferase with IQ but not with PhIP. This system allows for both quantifying mutagenesis and analyzing the mutational spectra produced by heterocyclic amines.

  2. Fluorophore-tagged pharmacophores for antitumor cytotoxicity: Modified chiral lipidic dialkynylcarbinols for cell imaging.

    PubMed

    Listunov, Dymytrii; Mazères, Serge; Volovenko, Yulian; Joly, Etienne; Génisson, Yves; Maraval, Valérie; Chauvin, Remi

    2015-10-15

    Chiral lipidic dialkynylcarbinols (DACs), recently highlighted as antitumoral pharmacophores, have been conjugated to difluoroboron-dipyrromethene (bodipy), 7-hydroxy-coumarine, and 7-nitro-benzoxadiazole (NBD) fluorophore motifs through triazole clips. The labeled lipids preserve cytotoxic activity against HCT116 cells, and fluorescence microscopy of the stained cells showed clear signals in the intra-cellular membrane system. While the bodipy conjugate also labels lipid droplets very brightly, as expected, the coumarine and NBD probes appear as promising specific tools for the identification of the intra-cellular targets of DACs' cytotoxicity.

  3. Control of Oxygen Delamination in Solid Oxide Electrolyzer Cells via Modifying Operational Regime

    SciTech Connect

    Sergey N. Rashkeev; Michael V. Glazoff

    2011-10-01

    Possible modifications of operational regimes for solid oxide fuel cell (SOEC) devices for hydrogen production are discussed. It is shown that applying alternating current (AC) voltage pulses at a certain frequency range to SOECs could reduce oxygen delamination degradation in these devices and significantly increase their lifetime. This operational scheme provides wide possibilities to increase longevity of SOEC devices required for their use in commercial hydrogen production processes, without any significant modification of used materials and/or cell design. Developed simulation method possesses a broad generality and be employed in a number of other industrial processes.

  4. Visual detection of STAT5B gene expression in living cell using the hairpin DNA modified gold nanoparticle beacon.

    PubMed

    Xue, Jianpeng; Shan, Lingling; Chen, Haiyan; Li, Yang; Zhu, Hongyan; Deng, Dawei; Qian, Zhiyu; Achilefu, Samuel; Gu, Yueqing

    2013-03-15

    Signal transducer and activator of transcription 5B (STAT5B) is an important protein in JAK-STAT signaling pathway that is responsible for the metastasis and proliferation of tumor cells. Determination of the STAT5B messenger Ribonucleic Acid (mRNA) relating to the STAT5B expression provides insight into the mechanism of tumor progression. In this study, we designed and used a special hairpin deoxyribonucleic acid (DNA) for human STAT5B mRNA to functionalize gold nanoparticles, which served as a beacon for detecting human STAT5B expression. Up to 90% quenching efficiency was achieved. Upon hybridizing with the target mRNA, the hairpin DNA modified gold nanoparticle beacons (hDAuNP beacons) release the fluorophores attached at 5' end of the oligonucleotide sequence. The fluorescence properties of the beacon before and after the hybridization with the complementary DNA were confirmed in vitro. The stability of hDAuNP beacons against degradation by DNase I and GSH indicated that the prepared beacon is stable inside cells. The detected fluorescence in MCF-7 cancer cells correlates with the specific STAT5B mRNA expression, which is consistent with the result from PCR measurement. Fluorescence microscopy showed that the hDAuNP beacons internalized in cells without using transfection agents, with intracellular distribution in the cytoplasm rather than the nucleus. The results demonstrated that this beacon could directly provide quantitative measurement of the intracellular STAT5B mRNA in living cells. Compared to the previous approaches, this beacon has advantages of higher target to background ratio of detection and an increased resistance to nuclease degradation. The strategy reported in this study is a promising approach for the intracellular measurement of RNA or protein expression in living cells, and has great potential in the study of drug screening and discovery.

  5. Safety and immunogenicity of mammalian cell derived and Modified Vaccinia Ankara vectored African swine fever subunit antigens in swine.

    PubMed

    Lopera-Madrid, Jaime; Osorio, Jorge E; He, Yongqun; Xiang, Zuoshuang; Adams, L Garry; Laughlin, Richard C; Mwangi, Waithaka; Subramanya, Sandesh; Neilan, John; Brake, David; Burrage, Thomas G; Brown, William Clay; Clavijo, Alfonso; Bounpheng, Mangkey A

    2017-03-01

    A reverse vaccinology system, Vaxign, was used to identify and select a subset of five African Swine Fever (ASF) antigens that were successfully purified from human embryonic kidney 293 (HEK) cells and produced in Modified vaccinia virus Ankara (MVA) viral vectors. Three HEK-purified antigens [B646L (p72), E183L (p54), and O61R (p12)], and three MVA-vectored antigens [B646L, EP153R, and EP402R (CD2v)] were evaluated using a prime-boost immunization regimen swine safety and immunogenicity study. Antibody responses were detected in pigs following prime-boost immunization four weeks apart with the HEK-293-purified p72, p54, and p12 antigens. Notably, sera from the vaccinees were positive by immunofluorescence on ASFV (Georgia 2007/1)-infected primary macrophages. Although MVA-vectored p72, CD2v, and EP153R failed to induce antibody responses, interferon-gamma (IFN-γ(+)) spot forming cell responses against all three antigens were detected one week post-boost. The highest IFN-γ(+) spot forming cell responses were detected against p72 in pigs primed with MVA-p72 and boosted with the recombinant p72. Antigen-specific (p12, p72, CD2v, and EP153R) T-cell proliferative responses were also detected post-boost. Collectively, these results are the first demonstration that ASFV subunit antigens purified from mammalian cells or expressed in MVA vectors are safe and can induce ASFV-specific antibody and T-cell responses following a prime-boost immunization regimen in swine.

  6. Antiproliferative Activity of Double Point Modified Analogs of 1,25-Dihydroxyvitamin D2 Against Human Malignant Melanoma Cell Lines

    PubMed Central

    Piotrowska, Anna; Wierzbicka, Justyna; Nadkarni, Sharmin; Brown, Geoffrey; Kutner, Andrzej; Żmijewski, Michał A.

    2016-01-01

    Vitamin D is a lipid soluble steroid hormone with pleiotropic biological properties, including regulation of cell proliferation, differentiation and apoptosis. As to these desirable anticancer actions, 1,25-dihydroxyvitamins D and analogs have been reported to inhibit the proliferation and to induce differentiation of a wide variety of cancer cell types, including human malignant melanoma. However, there is a need for novel and more efficacious vitamin D analogs, and how best to design such is still an open issue. A series of double point modified (DPM) analogs of 1,25-dihydroxyvitamin D2 (1,25(OH)2D2) induced differentiation of the vitamin D receptor (VDR) positive A375 and VDR negative SK-MEL 188b human malignant melanoma cell lines. Surprisingly, the dose of 1,25(OH)2D2 required to inhibit the proliferation of the A375 melanoma cell line by was several fold lower than that required in the case of 1,25(OH)2D3. To evaluate the impact of the modification in the side chain (additional 22-hydroxyl) and in the A-ring (5,6-trans modification), the regular side-chain of vitamin D2 or D3 was retained in the structure of our analogs. As expected, 5,6-trans modification was advantageous to enhancing the anti-proliferative activity of analogs, but not as a single point modification (SPM). Very unexpectedly, the additional 22-hydroxyl in the side-chain reduced significantly the anti-proliferative activity of both the natural and 5,6-trans series analogs. Finally, an induction of pigmentation in melanoma SK-MEL 188b cells was observed to sensitized cells to the effect of vitamin D analogs. PMID:26760999

  7. Antitumor efficacy of DMSA modified Fe3O4 magnetic nanoparticles combined with arsenic trioxide and adriamycin in Raji cells.

    PubMed

    Cai, Xiaohui; Cai, Xiaohui; Wang, Chunling; Chen, Baoan; Hua, Weijun; Shen, Fei; Yu, Liang; He, Zhengmei; Shi, Yuye; Chen, Yue; Xia, Guohua; Bao, Wen; Zhang, Yu; Wang, Xuemei

    2014-02-01

    The objective of the present study was to investigate the anticancer efficacy of dimercaptosuccinic acid modified iron oxide (DMSA-Fe3O4) magnetic nanoparticles (MNPs) combined with arsenic trioxide (As2O3) and doxorubicin (ADM) in non-Hodgkin's lymphoma (NHL) cell line (Raji cells). The growth inhibition rate of Raji cells was determined by MTT assay. Characteristics of DMSA-Fe3O4 MNPs and distribution of nanoparticles taken up by Raji cells were observed under a transmission electron microscopy (TEM). Further, apoptosis of cells and intracellular concentration of ADM were detected by flow cytometry (FCM). DAPI staining was used to view apoptotic cellular morphology. Subsequently, transcription and protein expression levels of bcl-2, NFKB, survivin, bax, p53 and caspase-3 were determined by reverse transciptase polymerase chain reaction (RT-PCR) and Western blotting analysis, respectively. The results of MTT assay indicated that the inhibition of Raji cells by the combined form of ADM and As2O3 was significantly higher than either ADM or As2O3 alone. However, ADM-As2O3 MNPs proved superior over all other groups. TEM observation revealed that the majority of MNPs were quasi-spherical with an average diameter of about 18 nm and the MNPs taken up by cells were located in the endosome vesicles of cytoplasm. The apoptotic rate and accumulation of intracellular ADM in ADM-As2O3 MNPs group were significantly higher than those in control, ADM, As2O3 and ADM+As2O3, groups. In addition, DAPI staining of Raji cells from ADM-As,O3 MNPs group clearly exhibited more morphological changes (severe structural alterations) than other groups. Moreover, transcription and protein expression of bcl-2, NFKB, survivin, bax, p53 and caspase-3 of Raji cells were regulated at the most remarkable extent in ADM-As2O3, MNPs group as compared with other groups. These findings suggest that the antitumor efficacy of the combination of novel ADM-As2O3, MNPs on Raji cells would be a promising

  8. β-Diketone modified trastuzumab: a next-generation of Herceptin for resistant breast cancer cells?

    PubMed

    Lu, Jianguo; Pu, Jun; Lu, Xiaozhao; Fu, Haiyan; Wei, Mengying; Yang, Guodong

    2012-11-01

    Despite the initial efficacy of trastuzumab (commercially named Herceptin), acquired resistance in a majority of patients remains the biggest hurdle in breast cancer therapy. Recently, the Scripps Research Institute developed a method termed "instant immunity", in which antibodies (chemical programmed antibody) are rapidly induced by chemicals like β-diketone. When β-diketone is chemically linked to peptides specifically targeting cancer cells, the instant chemical programmed antibody would clear the cancer cells through antibody-dependent cellular cytotoxicity (ADCC) and complement-directed cytotoxicity (CDC). This novel strategy has a super advantage over passive immunization or immunization with recombinant or vectored vaccines because it induces a universal immune response and memory. Theoretically, combination of the cancer cell specific recognition advantage of trastuzumab and cancerous cell clearance of active immunization would be an option for trastuzumab resistant patients, which harbors both the advantages of cancer specific targeting of trastuzumab and active immunization of the "instant immunity", implicating a better clinical outcome. Further studies are needed to verify our hypothesis, which is worth validating.

  9. Generation of genetically modified mice by oocyte injection of androgenetic haploid embryonic stem cells.

    PubMed

    Yang, Hui; Shi, Linyu; Wang, Bang-An; Liang, Dan; Zhong, Cuiqing; Liu, Wei; Nie, Yongzhan; Liu, Jie; Zhao, Jing; Gao, Xiang; Li, Dangsheng; Xu, Guo-Liang; Li, Jinsong

    2012-04-27

    Haploid cells are amenable for genetic analysis. Recent success in the derivation of mouse haploid embryonic stem cells (haESCs) via parthenogenesis has enabled genetic screening in mammalian cells. However, successful generation of live animals from these haESCs, which is needed to extend the genetic analysis to the organism level, has not been achieved. Here, we report the derivation of haESCs from androgenetic blastocysts. These cells, designated as AG-haESCs, partially maintain paternal imprints, express classical ESC pluripotency markers, and contribute to various tissues, including the germline, upon injection into diploid blastocysts. Strikingly, live mice can be obtained upon injection of AG-haESCs into MII oocytes, and these mice bear haESC-carried genetic traits and develop into fertile adults. Furthermore, gene targeting via homologous recombination is feasible in the AG-haESCs. Our results demonstrate that AG-haESCs can be used as a genetically tractable fertilization agent for the production of live animals via injection into oocytes.

  10. Endothelial cell palmitoylproteomics identifies novel lipid modified targets and potential substrates for protein acyl transferases

    PubMed Central

    Marin, Ethan P.; Derakhshan, Behrad; Lam, TuKiet T.; Davalos, Alberto; Sessa, William C.

    2012-01-01

    Rationale Protein S-palmitoylation is the post-translational attachment of a saturated 16-carbon palmitic acid to a cysteine side chain via a thioester bond. Palmitoylation can affect protein localization, trafficking, stability, and function. The extent and roles of palmitoylation in endothelial cell (EC) biology is not well understood, in part due to technological limits on palmitoylprotein detection. Objective To develop a method using acyl-biotinyl exchange (ABE) technology coupled with mass spectrometry to globally isolate and identify palmitoylproteins in EC. Methods and Results More than 150 putative palmitoyl proteins were identified in EC using ABE and mass spectrometry. Among the novel palmitoylproteins identified is superoxide dismutase 1 (SOD1), an intensively studied enzyme that protects all cells from oxidative damage. Mutation of cysteine 6 prevents palmitoylation, leads to reduction in SOD1 activity in vivo and in vitro, and inhibits nuclear localization, thereby supporting a functional role for SOD1 palmitoylation. Moreover, we used ABE to search for substrates of particular protein acyl transferases in EC. We found that palmitoylation of the cell adhesion protein PECAM1 is dependent on the protein acyl transferase ZDHHC21. We show that knockdown of ZDHHC21 leads to reduced levels of PECAM1 at the cell surface. Conclusions Our data demonstrate the utility of EC palmitoylproteomics to reveal new insights into the role of this important post-translational lipid modification in EC biology. PMID:22496122

  11. Technological Implications of Modifying the Extent of Cell Wall-Proanthocyanidin Interactions Using Enzymes.

    PubMed

    Bautista-Ortín, Ana Belén; Ben Abdallah, Rim; Castro-López, Liliana Del Rocío; Jiménez-Martínez, María Dolores; Gómez-Plaza, Encarna

    2016-01-18

    The transference and reactivity of proanthocyanidins is an important issue that affects the technological processing of some fruits, such as grapes and apples. These processes are affected by proanthocyanidins bound to cell wall polysaccharides, which are present in high concentrations during the processing of the fruits. Therefore, the effective extraction of proanthocyanidins from fruits to their juices or derived products will depend on the ability to manage these associations, and, in this respect, enzymes that degrade these polysaccharides could play an important role. The main objective of this work was to test the role of pure hydrolytic enzymes (polygalacturonase and cellulose) and a commercial enzyme containing these two activities on the extent of proanthocyanidin-cell wall interactions. The results showed that the modification promoted by enzymes reduced the amount of proanthocyanidins adsorbed to cell walls since they contributed to the degradation and release of the cell wall polysaccharides, which diffused into the model solution. Some of these released polysaccharides also presented some reactivity towards the proanthocyanidins present in a model solution.

  12. Technological Implications of Modifying the Extent of Cell Wall-Proanthocyanidin Interactions Using Enzymes

    PubMed Central

    Bautista-Ortín, Ana Belén; Ben Abdallah, Rim; Castro-López, Liliana del Rocío; Jiménez-Martínez, María Dolores; Gómez-Plaza, Encarna

    2016-01-01

    The transference and reactivity of proanthocyanidins is an important issue that affects the technological processing of some fruits, such as grapes and apples. These processes are affected by proanthocyanidins bound to cell wall polysaccharides, which are present in high concentrations during the processing of the fruits. Therefore, the effective extraction of proanthocyanidins from fruits to their juices or derived products will depend on the ability to manage these associations, and, in this respect, enzymes that degrade these polysaccharides could p