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Sample records for modified gec cell

  1. A Planar Probe for Ion Flux and Electron Temperature in the Electrode of a GEC Cell

    NASA Astrophysics Data System (ADS)

    Goodyear, A.; Lubeigt, W.; Verdonck, P. B.; Barroy, P. R. J.; Braithwaite, N. St. J.

    2000-10-01

    A radio frequency (rf) self-biased planar probe technique(J P Booth, N St J Braithwaite, A Goodyear, and P Barroy, Rev. Sci. Instrum., in press (July 2000)) for the measurement of positive ion flux and electron energy distribution functions has been implemented into the Gaseous Electronics Conference (GEC) standard reference cell. Results are presented across a wide range of power-pressure parameter space. Insertion of electrostatic probes into plasmas can lead to perturbation of the plasma and this is particularly problematic in the GEC cell where the inter-electrode distance is small. Planar probes are inherently large, requiring a guard ring (at least as large as typical sheath thicknesses) to ensure that the sheath in front of the probe is truly planar. Once this criterion is satisfied, analysis of current-voltage (IV) characteristics obtained in this planar geometry is relatively straight forward. A planar probe has been engineered into the surface of the showerhead electrode of the GEC cell, aimed at minimal perturbation to the GEC standard, in terms of both physical presence and electrical influence. The probe is self-biased by a burst of rf voltage. It is then allowed to return to its original floating condition under the arrival of charged particles from the plasma. An IV characteristic is recorded during this time, giving positive ion flux and the high energy tail (a few eV and above) of the electron energy distribution function.

  2. Reactor simulations of the GEC reference cell reactor with an ICP source for Ar/Cl2 and BCl3/Cl2 gas mixtures.

    NASA Astrophysics Data System (ADS)

    Veerasingam, Ramana; Choi, Seung J.; Riley, Merle; Hoekstra, Robert; Kushner, Mark

    1996-10-01

    The ICP (inductively coupled plasma) device is a widely researched plasma etching technology to meet the stringent requirements of dielectric and metal etch for the next generation semiconductor wafers. At Sandia, the GEC reference cell has been modified to include a planar coil geometry to couple the RF power to the plasma inductively. Measurements of densities of electrons, CL-, Ar+, and recently of ion current flux have been made. In this paper, we will present results of simulations modeling the GEC cell for Ar/CL2 and BCL3/Cl2 gas mixtures using the HPEM and GEMINI code packages. Results will be parametrized with power, pressure, and gas mixture. In addition, simulations of the ion current flux using a sheath model developed at Sandia and the HPEM will be performed and compared to data.

  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. Phase Transition Studies for Conducting Dust in a GEC Reference Cell

    NASA Astrophysics Data System (ADS)

    Carmona Reyes, Jorge; Hyde, Truell; Matthews, Lorin; George, David; Cook, Mike; Schmoke, Jimmy

    2009-11-01

    Dust particles immersed in plasma typically acquire a negative charge. The resulting Yukawa interaction between grains in a two-dimensional horizontal layer leads to the formation of disordered or ordered structures depending on whether short or long range ordering dominates, as determined by the ratio of the particle's interparticle potential energy to its average kinetic energy. Various stable crystalline phases have been observed experimentally for dust particles residing within such two-dimensionally extended lattice planes with system dynamics driven in large part by particle charge. Although the charging process for insulating materials has been examined in some detail, conducting materials have not yet been fully investigated. This work experimentally examines the phases and phase transitions for both conductive (gold coated) and non-conductive (melamine formaldehyde) particles. Phase maps for each type of particle are obtained using data from pair correlation functions and voronoi diagrams for dust structures formed over a range of pressures and powers within a standard GEC reference cell.

  5. Observation of an electron sheath at a large, transiently biassed surface in the GEC cell

    NASA Astrophysics Data System (ADS)

    Barroy, P. R. J.; Goodyear, A.; Braithwaite, N. St. J.

    2001-10-01

    Sheath reversal has been investigated in front of a biassed surface embedded into the ground electrode of a capacitively coupled GEC cell. Radio frequency bursts (several tens of volts amplitude) were applied to the surface (20 mm diameter including guard ring) and fast, two dimensional observations made of the light emission using an intensified CCD camera synchronously gated within the succession of bursts. A guard ring, biassed to the same potential, ensures sheath planarity. The evolution of the optical emission has been followed during the dc biassing period. If the applied RF is large enough and the timescale short enough a perturbation of the plasma sheath is observed. At the onset of the RF burst periodic, sheath reversal is achieved as the potential of the surface exceeds that of the plasma. After several cycles of RF, the surface acquires enough negative charge to bias itself negatively, below plasma potential; sheath reversal then stops. The overall effect is accompanied by light emision from species excited by electrons accelerated towards the surface. Modelling of the phenomenon has been conducted to account for the observations.

  6. Synthesis and Structural Stability of Ti2GeC

    SciTech Connect

    Phatak, N.; Saxena, S; Fei, Y; Hu, J

    2009-01-01

    In this paper we report the synthesis of a ternary-layered ceramic, Ti2GeC, a 211 Mn+1AXn compound by hot-pressing. X-ray analysis confirmed the hexagonal crystal symmetry. Using a synchrotron radiation and a diamond anvil cell, we measured the pressure dependencies of the lattice parameters. The phase is structurally stable up to the 49 GPa-the maximum experimental pressure reached. The bulk modulus of Ti2GeC calculated using the Birch-Murnaghan equation of state is 211 {+-} 4 GPa with K? = 3.9 {+-} 0.2. Surprisingly the bulk modulus was found to be greater than both Ti2AlC and Ti2SC by 13% and 10%. Also, Ti2GeC has the highest bulk modulus among M2GeC phases where M = Ti, V, Cr.

  7. GEC Student Award for Excellence Finalist: Interaction of Non-Thermal Dielectric Barrier Discharge Plasma with DNA inside Cells

    NASA Astrophysics Data System (ADS)

    Kalghatgi, Sameer; Kelly, Crystal; Fridman, Gregory; Clifford-Azizkhan, Jane; Fridman, Alexander; Friedman, Gary

    2008-10-01

    Direct non-thermal plasma is now being widely considered for various medical applications, viz; cancer treatment, coagulation, wound healing. However, the understanding of the interaction between non-thermal plasma and cells is lacking. Here we study the possibility that effects of the plasma treatment can penetrate though cellular membranes without destroying them. One of the most important of such effects to investigate would be DNA double strand breaks (DSB's) since these are some of the important events in a cell's life cycle. We measured DNA DSB's in mammalian cells using immunofluorescence and western blots. Hydrogen peroxide treatment was used as a positive control since it is known to induce massive DNA double strand breaks. The results indicate that short (5 seconds) direct plasma treatment at low power (0.2 W/cm^2) does produce DNA DSB's in mammalian cells. This means that somehow plasma penetrates inside the cells. Several questions arise about what is the mechanism of penetration and do the cells repair the DNA DSB's. We show that the cells do repair the DNA DSB's produced by short exposure of low power plasma. Although the detailed mechanisms are being investigated we confirmed that reactive oxygen species mediate interaction between plasma and DNA.

  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 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

  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. 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.

  12. 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.

  13. 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.

  14. 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.

  15. 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

  16. 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.

  17. Epitaxial growth and electrical transport properties of Cr{sub 2}GeC thin films

    SciTech Connect

    Eklund, Per; Bugnet, Matthieu; Mauchamp, Vincent; Dubois, Sylvain; Tromas, Christophe; Jaouen, Michel; Cabioc'h, Thierry; Jensen, Jens; Piraux, Luc; Gence, Loiek

    2011-08-15

    Cr{sub 2}GeC thin films were grown by magnetron sputtering from elemental targets. Phase-pure Cr{sub 2}GeC was grown directly onto Al{sub 2}O{sub 3}(0001) at temperatures of 700-800 deg. C. These films have an epitaxial component with the well-known epitaxial relationship Cr{sub 2}GeC(0001)//Al{sub 2}O{sub 3}(0001) and Cr{sub 2}GeC(1120)//Al{sub 2}O{sub 3}(1100) or Cr{sub 2}GeC(1120)//Al{sub 2}O{sub 3}(1210). There is also a large secondary grain population with (1013) orientation. Deposition onto Al{sub 2}O{sub 3}(0001) with a TiN(111) seed layer and onto MgO(111) yielded growth of globally epitaxial Cr{sub 2}GeC(0001) with a virtually negligible (1013) contribution. In contrast to the films deposited at 700-800 deg. C, the ones grown at 500-600 deg. C are polycrystalline Cr{sub 2}GeC with (1010)-dominated orientation; they also exhibit surface segregations of Ge as a consequence of fast Ge diffusion rates along the basal planes. The room-temperature resistivity of our samples is 53-66 {mu}{Omega}cm. Temperature-dependent resistivity measurements from 15-295 K show that electron-phonon coupling is important and likely anisotropic, which emphasizes that the electrical transport properties cannot be understood in terms of ground state electronic structure calculations only.

  18. 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

  19. 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

  20. 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.

  1. Gas-Phase Lasers - a Historical Perspective in Relation to the GEC

    NASA Astrophysics Data System (ADS)

    Hays, Gerry

    1997-10-01

    Understanding of gas-phase lasers inevitably involves an expertise in many of the specialties of the GEC community - especially homogenous and heterogeneous kinetics, collision cross-sections, gas breakdown physics and fundamental swarm parameters. The GEC community decided early in the evolution of gas-phase lasers to include papers on this topic and the result was many years of contributions to the evolution of and improvement in our understanding of this important class of lasers. Many of the ground-breaking results in gas laser technology were presented at the GEC over the last 3 decades as the traditional rare-gas atomic physics and low-temperature plasma groups turned their attention to parameters of interest to the laser modelers and experimenters. This paper will trace the development of this field, especially as it pertained to the GEC. Some of the key results will be highlighted, together with some of the unpublished trivia and anecdotal incidents in order to capture the flavor of the rapid developments in the early days. The talk will include speculation as to the direction this field is taking, and some suggestions as to opportunities. This work supported by the United States Department of Energy under Contract DE-AC04-94AL85000. Sandia is a multiprogram laboratory operated by Sandia Corporation, a Lockheed-Martin Company, for the United States Department of Energy.

  2. 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

  3. Implantation of Vascular Grafts Lined with Genetically Modified Endothelial Cells

    NASA Astrophysics Data System (ADS)

    Wilson, James M.; Birinyi, Louis K.; Salomon, Robert N.; Libby, Peter; Callow, Allan D.; Mulligan, Richard C.

    1989-06-01

    The possibility of using the vascular endothelial cell as a target for gene replacement therapy was explored. Recombinant retroviruses were used to transduce the lacZ gene into endothelial cells harvested from mongrel dogs. Prosthetic vascular grafts seeded with the genetically modified cells were implanted as carotid interposition grafts into the dogs from which the original cells were harvested. Analysis of the graft 5 weeks after implantation revealed genetically modified endothelial cells lining the luminal surface of the graft. This technology could be used in the treatment of atherosclerosis disease and the design of new drug delivery systems.

  4. 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.

  5. Rechargeable cells with modified MnO2 cathodes

    NASA Astrophysics Data System (ADS)

    Dzieciuch, M. A.; Gupta, N.; Wroblowa, H. S.

    1988-10-01

    The recent invention of rechargeable 'modified' manganese oxide materials paves the way to the development of secondary batteries suitable for numerous applications. This includes alternatives to primary dry cells, and secondary lead/acid and nickel-cadmium batteries. Present results describe the performance of cells in which the modified materials are coupled with zinc and iron. As opposed to iron which does not affect the longevity and capacity retention of the modified electrodes, zinc has a pejorative effect on modified MnO2 materials, owing to the formation of heterolite at the positive electrode. Methods to alleviate this effect and produce viable modified MnO2/Zn systems are described. At present, these systems retain about 50 percent of their theoretical one-electron capacity even after two hundred fast charge-discharge cycles.

  6. 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.

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

    PubMed Central

    Sharpe, Michaela; Mount, Natalie

    2015-01-01

    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. PMID:26035842

  8. First-principles study of structural, electronic, and mechanical properties of the nanolaminate compound Ti4GeC3 under pressure

    NASA Astrophysics Data System (ADS)

    Li, Chenliang; Wang, Zhenqing

    2010-06-01

    This paper investigates the pressure dependences of the structural, electronic, mechanical, and optical properties of the nanolaminate Ti4GeC3 compound using the first-principles method based on the density functional theory. The lattice parameters and atom positions of Ti4GeC3 in bulk form were predicted and show that Ti4GeC3 is more compressible in the c direction than along the a direction. The elastic constants, shear modulus and Young's moduli of Ti4GeC3 were then calculated at various pressures. The results indicate that Ti4GeC3 is mechanically stable in the pressure range of 0-70 GPa and that its brittle-ductile transition occurs at 60 GPa. By examining the densities of states, we also found that Ti4GeC3 remains structurally stable for pressures up to 70 GPa. Moreover, the Ti-Ge bonding is softer than all the Ti-C bonding and indicates that the Ti-C bond is more resistant to deformation than the Ti-Ge bond. Finally, the reflectivity spectrum of Ti4GeC3 under various pressures was investigated. Based on the results, we found that Ti4GeC3 can be used as a coating on spacecrafts to avoid solar heating, but it is not suitable for use under high pressure conditions.

  9. Genetically modified T-cell therapy for osteosarcoma.

    PubMed

    DeRenzo, Christopher; Gottschalk, Stephen

    2014-01-01

    T-cell immunotherapy may offer an approach to improve outcomes for patients with osteosarcoma, who fail current therapies. In addition, it has the potential to reduce treatment-related complications for all patients. Generating tumor-specific T cells with conventional antigen presenting cells ex vivo is time consuming and often results in T-cell products with a low frequency of tumor-specific T cells. In addition, the generated T cells remain sensitive to the immunosuppressive tumor microenvironment. Genetic modification of T cells is one strategy to overcome these limitations. For example, T cells can be genetically modified to render them antigen specific, resistant to inhibitory factors, or increase their ability to home to tumor sites. Most genetic modification strategies have only been evaluated in preclinical models, however early phase clinical trials are in progress. In this chapter we review the current status of gene-modified T-cell therapy with special focus on osteosarcoma, highlighting potential antigenic targets, preclinical and clinical studies, and strategies to improve current T-cell therapy approaches.

  10. 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

  11. Modified NASA standard nickel-cadmium cell designs

    NASA Technical Reports Server (NTRS)

    Manzo, Michelle A.

    1992-01-01

    The experimental design, parameters, and testing of a modified NASA standard nickel-cadmium cell are discussed. Modifications regarding positive plate loading levels and nickel attack levels, loading levels for the negative plates, interelectrode spacing, and the positive electrode impregnation process are addressed.

  12. 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.

  13. Electrospun fiber membranes enable proliferation of genetically modified cells

    PubMed Central

    Borjigin, Mandula; Eskridge, Chris; Niamat, Rohina; Strouse, Bryan; Bialk, Pawel; Kmiec, Eric B

    2013-01-01

    Polycaprolactone (PCL) and its blended composites (chitosan, gelatin, and lecithin) are well-established biomaterials that can enrich cell growth and enable tissue engineering. However, their application in the recovery and proliferation of genetically modified cells has not been studied. In the study reported here, we fabricated PCL-biomaterial blended fiber membranes, characterized them using physicochemical techniques, and used them as templates for the growth of genetically modified HCT116-19 colon cancer cells. Our data show that the blended polymers are highly miscible and form homogenous electrospun fiber membranes of uniform texture. The aligned PCL nanofibers support robust cell growth, yielding a 2.5-fold higher proliferation rate than cells plated on standard plastic plate surfaces. PCL-lecithin fiber membranes yielded a 2.7-fold higher rate of proliferation, while PCL-chitosan supported a more modest growth rate (1.5-fold higher). Surprisingly, PCL-gelatin did not enhance cell proliferation when compared to the rate of cell growth on plastic surfaces. PMID:23467983

  14. 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.

  15. Characterization of glomerular epithelial cell matrix receptors.

    PubMed Central

    Adler, S.

    1992-01-01

    Integrin matrix receptors on glomerular epithelial cells (GEC) may play an important role in adhesion of GEC to the glomerular basement membrane (GBM) and in the maintenance of normal glomerular permeability. Therefore, the author determined the types of matrix receptors present on cultured rat GEC and examined their interactions with several components of the extracellular matrix. Beta 1 integrin matrix receptors were detected on all three glomerular cell types in rat kidney in vivo and at areas of cell-cell contact on cultured GEC. Glomerular epithelial cell adhesion to types I and IV collagen was slightly greater than to laminin and fibronectin. Adhesion to fibronectin was significantly inhibited by a synthetic peptide containing the RGD adhesion sequence. Immunoprecipitation of lysates of surface-iodinated GEC showed the presence of alpha 3 beta 1 integrin. Chromatography of lysates on immobilized collagen showed alpha 3 beta 1 integrin and a 70- to 75-kd protein band as the collagen receptors on GEC. Chromatography on the 120-kd cell-binding fragment of fibronectin disclosed only alpha 3 beta 1 as a specific fibronectin receptor. Antibody to the beta 1 integrin chain inhibited adhesion to laminin and collagen. These studies demonstrate that in vitro, as in vivo, GEC appear to express only alpha 3 beta 1 integrin. Furthermore, this matrix receptor is capable of mediating GEC adhesion to collagen, fibronectin, and laminin, components of the GBM, and presumably plays a similar role in promoting GEC adhesion to GBM in vivo. Images Figure 1 Figure 4 Figure 5 Figure 6 Figure 7 PMID:1325740

  16. 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.

  17. 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. PMID:27652340

  18. 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.

  19. 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

  20. Manufacturing genetically modified T cells for clinical trials.

    PubMed

    Gee, A P

    2015-03-01

    Compliance with Food and Drug Administration regulations relating to initiating early phase clinical trials of new cellular therapy products often presents a hurdle to new investigators. One of the biggest obstacles is the requirement to manufacture the therapeutic products under current Good Manufacturing Practices-a system that is usually poorly understood by both basic researchers and clinicians. This article reviews the major points that must be addressed when manufacturing genetically modified T cells for therapeutic use. PMID:25633481

  1. 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

    2009-04-01

    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

  2. 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)

  3. 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.

  4. 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.

  5. 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

  6. 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

  7. 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. PMID:26463253

  8. Mps1 is SUMO-modified during the cell cycle.

    PubMed

    Restuccia, Agnese; Yang, Feikun; Chen, Changyan; Lu, Lou; Dai, Wei

    2016-01-19

    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

  9. 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

  10. Cell research with physically modified microfluidic channels: a review.

    PubMed

    Kim, Sun Min; Lee, Sung Hoon; Suh, Kahp Yang

    2008-07-01

    An overview of the use of physically modified microfluidic channels towards cell research is presented. The physical modification can be realized either by combining embedded physical micro/nanostructures or a topographically patterned substrate at the micro- or nanoscale inside a channel. After a brief description of the background and the importance of the physically modified microfluidic system, various fabrication methods are described based on the materials and geometries of physical structures and channels. Of many operational principles for microfluidics (electrical, magnetic, optical, mechanical, and so on), this review primarily focuses on mechanical operation principles aided by structural modification of the channels. The mechanical forces are classified into (i) hydrodynamic, (ii) gravitational, (iii) capillary, (iv) wetting, and (v) adhesion forces. Throughout this review, we will specify examples where necessary and provide trends and future directions in the field.

  11. Surface modified gold nanowires for mammalian cell transfection

    NASA Astrophysics Data System (ADS)

    Kuo, Chiung-Wen; Lai, Jun-Jung; Wei, Kung Hwa; Chen, Peilin

    2008-01-01

    Aminothiol modified gold nanowires have been used as vectors for the delivery of plasmid DNA into two different types of mammalian cells: 3T3 and HeLa. It was measured that positively charged gold nanowires with a diameter of 200 nm and a length around 5 µm were capable of carrying 1 pg of plasmid DNA per nanowire into cells. Compared with other transfection reagents, the gold nanowires exhibited the highest transfection efficiency while almost no cytotoxicity was observed. In addition, it has been shown that individual nanowires can be visualized with sub-micrometer resolution, which may allow the use of functionalized multi-segment nanowires as local probes for the investigation of the microenvironment inside cells.

  12. 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. PMID:26257187

  13. 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.

  14. An efficient algorithm for function optimization: modified stem cells algorithm

    NASA Astrophysics Data System (ADS)

    Taherdangkoo, Mohammad; Paziresh, Mahsa; Yazdi, Mehran; Bagheri, Mohammad

    2013-03-01

    In this paper, we propose an optimization algorithm based on the intelligent behavior of stem cell swarms in reproduction and self-organization. Optimization algorithms, such as the Genetic Algorithm (GA), Particle Swarm Optimization (PSO) algorithm, Ant Colony Optimization (ACO) algorithm and Artificial Bee Colony (ABC) algorithm, can give solutions to linear and non-linear problems near to the optimum for many applications; however, in some case, they can suffer from becoming trapped in local optima. The Stem Cells Algorithm (SCA) is an optimization algorithm inspired by the natural behavior of stem cells in evolving themselves into new and improved cells. The SCA avoids the local optima problem successfully. In this paper, we have made small changes in the implementation of this algorithm to obtain improved performance over previous versions. Using a series of benchmark functions, we assess the performance of the proposed algorithm and compare it with that of the other aforementioned optimization algorithms. The obtained results prove the superiority of the Modified Stem Cells Algorithm (MSCA).

  15. Discovery of the ternary nanolaminated compound Nb2GeC by a systematic theoretical-experimental approach.

    PubMed

    Eklund, Per; Dahlqvist, Martin; Tengstrand, Olof; Hultman, Lars; Lu, Jun; Nedfors, Nils; Jansson, Ulf; Rosén, Johanna

    2012-07-20

    Since the advent of theoretical materials science some 60 years ago, there has been a drive to predict and design new materials in silicio. Mathematical optimization procedures to determine phase stability can be generally applicable to complex ternary or higher-order materials systems where the phase diagrams of the binary constituents are sufficiently known. Here, we employ a simplex-optimization procedure to predict new compounds in the ternary Nb-Ge-C system. Our theoretical results show that the hypothetical Nb2GeC is stable, and excludes all reasonably conceivable competing hypothetical phases. We verify the existence of the Nb2GeC phase by thin film synthesis using magnetron sputtering. This hexagonal nanolaminated phase has a and c lattice parameters of ∼3.24  Å and 12.82 Å.

  16. Discovery of the Ternary Nanolaminated Compound Nb2GeC by a Systematic Theoretical-Experimental Approach

    NASA Astrophysics Data System (ADS)

    Eklund, Per; Dahlqvist, Martin; Tengstrand, Olof; Hultman, Lars; Lu, Jun; Nedfors, Nils; Jansson, Ulf; Rosén, Johanna

    2012-07-01

    Since the advent of theoretical materials science some 60 years ago, there has been a drive to predict and design new materials in silicio. Mathematical optimization procedures to determine phase stability can be generally applicable to complex ternary or higher-order materials systems where the phase diagrams of the binary constituents are sufficiently known. Here, we employ a simplex-optimization procedure to predict new compounds in the ternary Nb-Ge-C system. Our theoretical results show that the hypothetical Nb2GeC is stable, and excludes all reasonably conceivable competing hypothetical phases. We verify the existence of the Nb2GeC phase by thin film synthesis using magnetron sputtering. This hexagonal nanolaminated phase has a and c lattice parameters of ˜3.24Å and 12.82 Å.

  17. 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.

  18. 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.

  19. 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. PMID:25479396

  20. 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.

  1. 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

  2. Zinc electrowinning analysis in a modified Hull cell

    NASA Astrophysics Data System (ADS)

    McColm, Thomas Dean

    The Hull cell is an analytical cell designed with trapezoidal geometry to incorporate a range of current densities into a single experiment. It was conceived to examine electroplating processes rather than mass production processes. A modified analytical cell was designed, developed and applied to the diagnosis of zinc electrowinning. Emphasis was placed on obtaining the quantitative variation of current efficiency with current density and the associated microscopic variation in deposit morphology. Current density distributions came by placing an insulating baffle in between parallel electrodes. The baffle position and length were easily adjusted, allowing the generation of 12 different distributions for a single applied potential. Ten electrically isolated 1 cm2 segments comprised the cathode. Measurement of the potential drop across I ohm resistors in each of the ten isolated parallel branches permitted direct quantitative determination of current densities. The small segments permitted simple SEM and X-ray analysis of deposits. The cell was designed to allow the continual cycling of electrolyte. In conjunction with experimental analysis, a technique for the determination of current efficiency was tested and developed. The technique involved the comparison of charge passed for the electrodeposition and subsequent electrodissolution of a given mass of zinc and removed the necessity to determine the mass directly. In no prior studies on zinc electrowinning had current efficiencies been determined this way. The cell and technique were developed and verified by the correct diagnosis of industrial zinc electrowinning. Successful determination of the effects of key variables including temperature, acid to zinc ratio and impurity effects on current efficiency and deposit morphology was demonstrated. In parallel with experimental work, cell electrochemistry was modeled. Primary and secondary input parameters were those pertinent to zinc electrowinning. The resultant

  3. Novel eukaryotic enzymes modifying cell-surface biopolymers

    PubMed Central

    2010-01-01

    Background Eukaryotic extracellular matrices such as proteoglycans, sclerotinized structures, mucus, external tests, capsules, cell walls and waxes contain highly modified proteins, glycans and other composite biopolymers. Using comparative genomics and sequence profile analysis we identify several novel enzymes that could be potentially involved in the modification of cell-surface glycans or glycoproteins. Results Using sequence analysis and conservation we define the acyltransferase domain prototyped by the fungal Cas1p proteins, identify its active site residues and unify them to the superfamily of classical 10TM acyltransferases (e.g. oatA). We also identify a novel family of esterases (prototyped by the previously uncharacterized N-terminal domain of Cas1p) that have a similar fold as the SGNH/GDSL esterases but differ from them in their conservation pattern. Conclusions We posit that the combined action of the acyltransferase and esterase domain plays an important role in controlling the acylation levels of glycans and thereby regulates their physico-chemical properties such as hygroscopicity, resistance to enzymatic hydrolysis and physical strength. We present evidence that the action of these novel enzymes on glycans might play an important role in host-pathogen interaction of plants, fungi and metazoans. We present evidence that in plants (e.g. PMR5 and ESK1) the regulation of carbohydrate acylation by these acylesterases might also play an important role in regulation of transpiration and stress resistance. We also identify a subfamily of these esterases in metazoans (e.g. C7orf58), which are fused to an ATP-grasp amino acid ligase domain that is predicted to catalyze, in certain animals, modification of cell surface polymers by amino acid or peptides. Reviewers This article was reviewed by Gaspar Jekely and Frank Eisenhaber PMID:20056006

  4. Electronic and magnetic properties of single-wall GeC nanotubes filled with iron nanowires

    NASA Astrophysics Data System (ADS)

    Wang, Su-Fang; Chen, Li-Yong; Zhang, Jian-Min; Xu, Ke-Wei

    2012-06-01

    Under GGA, the structural, electronic and magnetic properties of single-wall (8, 8) GeC nanotubes filled with iron Fen nanowires (n = 5, 9, 13 and 21) have been investigated systematically using the first-principles PAW potential within DFT. We find that the initial shapes of the Fe5@(8, 8), Fe9@(8, 8) and Fe13@(8, 8) systems are preserved without any visible changes after optimization. But for the Fe21@(8, 8) system, the initial shapes are distorted largely for both nanowire and nanotube. The binding processes of Fen@(8, 8) systems are exothermic, and Fe5@(8, 8) system is the most stable structure. The pristine (8, 8) GeCNT is nonmagnetic and direct semiconductor with a wide band gap of about 2.65 eV. Projected densities of states onto different shell Fe atoms show that the separation between the bonding and antibonding d states is reduced as going from the core Fe atom to the outermost shell Fe atom. The spin polarization of the Fen@(8, 8) systems and free-standing nanowires are higher than that in bulk Fe. And the spin polarization generally decreases with the number n of the Fe atoms increasing for both the Fen@(8, 8) systems and free-standing nanowires. Both the largest spin polarization value itself and not more decrease with respect to value of free-standing Fe5 nanowire suggest the Fe5@(8, 8) system could be of interest for the use in electron spin injection. The magnetism is mainly confined within the inner Fe nanowire for these combined systems. More importantly, the Fe5 nanowire encapsulated inside (8, 8) GeCNT is under the protection of the GeCNT to prevent from oxidation, thus may stably exist in atmosphere for long time and can be expected to have potential applications in building nanodevices.

  5. Ultrastructural co-localisation of vimentin and cytokeratin in visceral glomerular epithelial cells of dogs with glomerulonephritis.

    PubMed

    Vilafranca, M; Ferrer, L; Wohlsein, P; Trautwein, G; Sanchez, J; Navarro, J A

    1995-07-01

    The expression of cytokeratin and vimentin was studied in the glomerular epithelial cells of canine kidneys with and without glomerular abnormalities. Using ultrastructural, immunogold single and double labelling techniques, cytokeratin and vimentin were found together in the visceral glomerular epithelial cells (vGECs) of abnormal kidneys. In normal kidneys, the vGECs expressed only vimentin, and cytokeratin was found exclusively in parietal glomerular epithelial cells (pGECs). These results confirm previous findings in the same animals, obtained by immunohistological staining techniques.

  6. A Modified NK Cell Degranulation Assay Applicable for Routine Evaluation of NK Cell Function

    PubMed Central

    Shabrish, Snehal; Gupta, Maya; Madkaikar, Manisha

    2016-01-01

    Natural killer (NK) cells play important role in innate immunity against tumors and viral infections. Studies show that lysosome-associated membrane protein-1 (LAMP-1, CD107a) is a marker for degranulation of NK and cytotoxic T cells and its expression is a sensitive marker for the cytotoxic activity determination. The conventional methods of determination of CD107a on NK cells involve use of peripheral blood mononuclear cells (PBMC) or pure NK cells and K562 cells as stimulants. Thus, it requires large volume of blood sample which is usually difficult to obtain in pediatric patients and patients with cytopenia and also requires specialized laboratory for maintaining cell line. We have designed a flow cytometric assay to determine CD107a on NK cells using whole blood, eliminating the need for isolation of PBMC or isolate NK cells. This assay uses phorbol-12-myristate-13-acetate (PMA) and calcium ionophore (Ca2+-ionophore) instead of K562 cells for stimulation and thus does not require specialized cell culture laboratory. CD107a expression on NK cells using modified NK cell degranulation assay compared to the conventional assay was significantly elevated (p < 0.0001). It was also validated by testing patients diagnosed with familial hemophagocytic lymphohistiocytosis (FHL) with defect in exocytosis. This assay is rapid, cost effective, and reproducible and requires significantly less volume of blood which is important for clinical evaluation of NK cells. PMID:27413758

  7. Antitumor cell-complex vaccines employing genetically modified tumor cells and fibroblasts.

    PubMed

    Miguel, Antonio; Herrero, María José; Sendra, Luis; Botella, Rafael; Diaz, Ana; Algás, Rosa; Aliño, Salvador F

    2014-02-19

    The present study evaluates the immune response mediated by vaccination with cell complexes composed of irradiated B16 tumor cells and mouse fibroblasts genetically modified to produce GM-CSF. The animals were vaccinated with free B16 cells or cell complexes. We employed two gene plasmid constructions: one high producer (pMok) and a low producer (p2F). Tumor transplant was performed by injection of B16 tumor cells. Plasma levels of total IgG and its subtypes were measured by ELISA. Tumor volumes were measured and survival curves were obtained. The study resulted in a cell complex vaccine able to stimulate the immune system to produce specific anti-tumor membrane proteins (TMP) IgG. In the groups vaccinated with cells transfected with the low producer plasmid, IgG production was higher when we used free B16 cell rather than cell complexes. Nonspecific autoimmune response caused by cell complex was not greater than that induced by the tumor cells alone. Groups vaccinated with B16 transfected with low producer plasmid reached a tumor growth delay of 92% (p ≤ 0.01). When vaccinated with cell complex, the best group was that transfected with high producer plasmid, reaching a tumor growth inhibition of 56% (p ≤ 0.05). Significant survival (40%) was only observed in the groups vaccinated with free transfected B16 cells.

  8. Antitumor Cell-Complex Vaccines Employing Genetically Modified Tumor Cells and Fibroblasts

    PubMed Central

    Miguel, Antonio; Herrero, María José; Sendra, Luis; Botella, Rafael; Diaz, Ana; Algás, Rosa; Aliño, Salvador F.

    2014-01-01

    The present study evaluates the immune response mediated by vaccination with cell complexes composed of irradiated B16 tumor cells and mouse fibroblasts genetically modified to produce GM-CSF. The animals were vaccinated with free B16 cells or cell complexes. We employed two gene plasmid constructions: one high producer (pMok) and a low producer (p2F). Tumor transplant was performed by injection of B16 tumor cells. Plasma levels of total IgG and its subtypes were measured by ELISA. Tumor volumes were measured and survival curves were obtained. The study resulted in a cell complex vaccine able to stimulate the immune system to produce specific anti-tumor membrane proteins (TMP) IgG. In the groups vaccinated with cells transfected with the low producer plasmid, IgG production was higher when we used free B16 cell rather than cell complexes. Nonspecific autoimmune response caused by cell complex was not greater than that induced by the tumor cells alone. Groups vaccinated with B16 transfected with low producer plasmid reached a tumor growth delay of 92% (p ≤ 0.01). When vaccinated with cell complex, the best group was that transfected with high producer plasmid, reaching a tumor growth inhibition of 56% (p ≤ 0.05). Significant survival (40%) was only observed in the groups vaccinated with free transfected B16 cells. PMID:24556729

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

    PubMed

    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

  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. 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.

  12. 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...

  13. 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.

  14. Biosorption of water-soluble dyes on magnetically modified Saccharomyces cerevisiae subsp. uvarum cells.

    PubMed

    Safaríková, M; Ptácková, L; Kibriková, I; Safarík, I

    2005-05-01

    Brewer's yeast (bottom yeast, Saccharomyces cerevisiae subsp. uvarum) cells were magnetically modified using water based magnetic fluid stabilized with perchloric acid. Magnetically modified yeast cells efficiently adsorbed various water soluble dyes. The dyes adsorption can be described by the Langmuir adsorption model. The maximum adsorption capacity of the magnetic cells differed substantially for individual dyes; the highest value was found for aniline blue (approx. 220 mg per g of dried magnetic adsorbent). PMID:15811411

  15. Nanofiber-modified surface directed cell migration and orientation in microsystem

    PubMed Central

    Zhang, Xu; Gao, Xinghua; Jiang, Lei; Zhang, Xulang; Qin, Jianhua

    2011-01-01

    Cell-microscale pattern surface interactions are crucial to understand many fundamental biological questions and develop regenerative medicine and tissue engineering approaches. In this work, we demonstrated a simple method to pattern PDMS surface by sacrificing poly vinyl pyrrolidone (PVP) electrospinning nanofibers and investigated the growth profile of cells on the modified patterned surfaces using stroma cells. The stromal cells were observed to exhibit good viability on this modified surface and the patterned surface with alignment nanofibers could promote cell migration. Furthermore, the modified PDMS surface was integrated with microfluidic channels to create the microscale spatial factor and was used to explore the cell migration and orientation under this microsystem. Both spatial factor and patterned surfaces were found to contribute to the complex cell orientation under the combined dual effects. This established method is simple, fast, and easy for use, demonstrating the potential of this microsystem for applications in addressing biological questions in complex environment. PMID:22662030

  16. 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.

  17. 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.

  18. Ptpn22 Modifies Regulatory T Cell Homeostasis via GITR Upregulation.

    PubMed

    Nowakowska, Dominika J; Kissler, Stephan

    2016-03-01

    PTPN22 gene variation associates with multiple autoimmune diseases, including type 1 diabetes and rheumatoid arthritis. Loss of function studies have demonstrated that PTPN22 impinges on the homeostatic behavior of regulatory T (Treg) cells, a lineage critical for immune tolerance. The frequency and absolute number of Treg cells is increased in Ptpn22-deficient mice, but the mechanism driving this increase is unknown. In this study, we show that Ptpn22 knockdown (KD) promoted the expansion of the Treg cell compartment by upregulating the glucocorticoid-induced TNFR family-related protein (GITR) and increasing GITR signaling. Ptpn22 KD did not accelerate cell division but instead prolonged Treg cell survival, as measured by a decrease in the frequency of apoptotic Treg cells. Loss of Ptpn22 caused a concomitant increase in the proportion of CD44(hi)CD62L(lo) effector Treg cells, at the expense of CD44(lo)CD62L(hi) central Treg cells. The increase in Treg cell numbers, but not their differentiation toward an effector phenotype, was dependent on GITR signaling, because blockade of GITR ligand prevented Treg cell expansion caused by Ptpn22 KD. These findings indicate that GITR plays a key role in regulating the overall size of the Treg cell pool. Our results suggest that the size and composition of the Treg cell compartment are independently controlled and have implications for the design of immunotherapies that seek to improve Treg cell function. PMID:26810223

  19. 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

  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. Poly(L-lysine)-modified iron oxide nanoparticles for stem cell labeling.

    PubMed

    Babic, Michal; Horák, Daniel; Trchová, Miroslava; Jendelová, Pavla; Glogarová, Katerina; Lesný, Petr; Herynek, Vít; Hájek, Milan; Syková, Eva

    2008-03-01

    New surface-modified iron oxide nanoparticles were developed by precipitation of Fe(II) and Fe(III) salts with ammonium hydroxide and oxidation of the resulting magnetite with sodium hypochlorite, followed by the addition of poly( L-lysine) (PLL) solution. PLL of several molecular weights ranging from 146 ( L-lysine) to 579 000 was tested as a coating to boost the intracellular uptake of the nanoparticles. The nanoparticles were characterized by TEM, dynamic light scattering, FTIR, and ultrasonic spectrometry. TEM revealed that the particles were ca. 6 nm in diameter, while FTIR showed that their surfaces were well-coated with PLL. The interaction of PLL-modified iron oxide nanoparticles with DMEM culture medium was verified by UV-vis spectroscopy. Rat bone marrow stromal cells (rMSCs) and human mesenchymal stem cells (hMSC) were labeled with PLL-modified iron oxide nanoparticles or with Endorem (control). Optical microscopy and TEM confirmed the presence of PLL-modified iron oxide nanoparticles inside the cells. Cellular uptake was very high (more than 92%) for PLL-modified nanoparticles that were coated with PLL (molecular weight 388 00) at a concentration of 0.02 mg PLL per milliliter of colloid. The cellular uptake of PLL-modified iron oxide was facilitated by its interaction with the negatively charged cell surface and subsequent endosomolytic uptake. The relaxivity of rMSCs labeled with PLL-modified iron oxide and the amount of iron in the cells were determined. PLL-modified iron oxide-labeled rMSCs were imaged in vitro and in vivo after intracerebral grafting into the contralateral hemisphere of the adult rat brain. The implanted cells were visible on magnetic resonance (MR) images as a hypointense area at the injection site and in the lesion. In comparison with Endorem, nanoparticles modified with PLL of an optimum molecular weight demonstrated a higher efficiency of intracellular uptake by MSC cells.

  2. 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

  3. Counting unstained, confluent cells by modified bright-field microscopy

    PubMed Central

    Drey, L. Louis; Graber, Michael C.; Bieschke, Jan

    2013-01-01

    We present a very simple procedure yielding high-contrast images of adherent, confluent cells such as human neuroblastoma (SH-EP) cells by ordinary bright-field microscopy. Cells are illuminated through a color filter and a pinhole aperture placed between the condenser and the cell culture surface. Refraction by each cell body generates a sharp, bright spot when the image is defocused. The technique allows robust, automatic cell counting from a single bright-field image in a wide range of focal positions; it does this via free, readily available image-analysis tools. Contrast may be enhanced by swelling cell bodies by brief incubation in PBS. The procedure was benchmarked against manual counting and automated counting of fluorescently labeled cell nuclei.. Counts from day-old and freshly seeded plates were compared in a range of densities, from sparse to densely overgrown. On average bright-field images produced the same counts as fluorescent images, with less than 5% error. This method will allow routine cell counting using a plain bright-field microscope, absent cell-line modification or cell staining. PMID:23834382

  4. A strong antibody reacting with enzyme modified E positive red blood cells.

    PubMed

    Heistø, H; Fagerhol, M K

    1979-01-01

    A high titer antibody was discovered in a healthy young man of blood group A1 R2r. The antibody strongly agglutinated all E positive red blood cells including his own, which had been modified by papain, ficin and bromelin, but only very weakly when modified by trypsin. The antibody was shown to be an IgM antibody. It did not react with unmodified red blood cells.

  5. A strong antibody reacting with enzyme modified E positive red blood cells.

    PubMed

    Heistø, H; Fagerhol, M K

    1979-01-01

    A high titer antibody was discovered in a healthy young man of blood group A1 R2r. The antibody strongly agglutinated all E positive red blood cells including his own, which had been modified by papain, ficin and bromelin, but only very weakly when modified by trypsin. The antibody was shown to be an IgM antibody. It did not react with unmodified red blood cells. PMID:116398

  6. Experimental evidence of Cr magnetic moments at low temperature in Cr2A(A=Al, Ge)C.

    PubMed

    Jaouen, M; Bugnet, M; Jaouen, N; Ohresser, P; Mauchamp, V; Cabioc'h, T; Rogalev, A

    2014-04-30

    From x-ray magnetic circular dichroism experiments performed at low temperature on Cr2AlC and Cr2GeC thin films, it is evidenced that Cr atoms carry a net magnetic moment in these ternary phases. It is shown that the Cr magnetization of the Al-based compound nearly vanished at 100 K in agreement with what has been recently observed on bulk. X-ray linear dichroism measurements performed at various angles of incidence and temperatures clearly demonstrate the existence of a charge ordering along the c axis of the structure of Cr2AlC. All these experimental observations support, in part, theoretical calculations claiming that Cr dd correlations have to be considered to correctly describe the structure and properties of these Cr-based ternary phases. PMID:24721758

  7. 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.

  8. 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.

  9. Proteins modified by the lipid peroxidation aldehyde DODE in MCF7 breast cancer cells

    PubMed Central

    Slade, Peter G.; Williams, Michelle V.; Brahmbatt, Viral; Dash, Ajit; Wishnok, John S.; Tannenbaum, Steven R.

    2010-01-01

    The hydroperoxide of linoleic acid (13-HPODE) degrades to 9,12-dioxo-10(E)-dodecenoic acid (DODE) which readily modifies proteins. This study identified the major proteins in MCF7 cells modified by DODE. To reduce false positives, three methods were use to identify DODE-modified proteins. First, cells were treated with a synthetically biotinylated 13-HPODE (13-HPODE-biotin). Modified proteins were enriched by neutravidin affinity and identified by 2D-LC-MS/MS. Second, cells were treated with native 13-HPODE. Protein-carbonyls were biotinylated with an aldehyde reactive probe (ARP) and modified proteins enriched by neutravidin affinity and identified by 2D-LC-MS/MS. Third, using a newly developed DODE antibody, DODE modified proteins were located by 2D-SDS-PAGE and Western blot and identified by in-gel digestion and LC-MS/MS. Analysis of the proteins characterized by all three methods revealed a significant overlap and identified 32 primary proteins modified by DODE in MCF7 cells. These results demonstrated the feasibility for the cellular formation of DODE protein-carbonyl adducts that may be future indicators of oxidative stress. PMID:20131800

  10. 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.

  11. 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.

  12. 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.

  13. Regulation of oncogene-induced cell cycle exit and senescence by chromatin modifiers

    PubMed Central

    David, Gregory

    2012-01-01

    Oncogene activation leads to dramatic changes in numerous biological pathways controlling cellular division, and results in the initiation of a transcriptional program that promotes transformation. Conversely, it also triggers an irreversible cell cycle exit called cellular senescence, which allows the organism to counteract the potentially detrimental uncontrolled proliferation of damaged cells. Therefore, a tight transcriptional control is required at the onset of oncogenic signal, coordinating both positive and negative regulation of gene expression. Not surprisingly, numerous chromatin modifiers contribute to the cellular response to oncogenic stress. While these chromatin modifiers were initially thought of as mere mediators of the cellular response to oncogenic stress, recent studies have uncovered a direct and specific regulation of chromatin modifiers by oncogenic signals. We review here the diverse functions of chromatin modifiers in the cellular response to oncogenic stress, and discuss the implications of these findings on the regulation of cell cycle progression and proliferation by activated oncogenes. PMID:22825329

  14. Electroporation chip for adherent cells on photochemically modified polymer surfaces

    NASA Astrophysics Data System (ADS)

    Olbrich, Michael; Rebollar, Esther; Heitz, Johannes; Frischauf, Irene; Romanin, Christoph

    2008-01-01

    We present a polytetrafluoroethylene electroporation microchip with integrated electrodes for transfection of adherent biological cells. For fabrication, UV-surface modification was employed in combination with metal deposition. UV irradiation in reactive atmosphere resulted in introduction of polar chemical groups into the polytetrafluoroethylene surface for significant adhesion enhancement of both biological cells as well as metal electrodes thereon. Electroporation was demonstrated by transfection of human embryonic kidney cells with the enhanced green fluorescent protein. Transparent, working at low voltages, and easy to handle, this chip yields the potential to reduce the amount of sequential working steps necessary for transfection.

  15. Gene-modified hematopoietic stem cells for cancer immunotherapy.

    PubMed

    Larson, Sarah; De Oliveira, Satiro N

    2014-01-01

    The rapid expansion of available cancer immunotherapies has resulted in favorable early outcomes. Specifically the use of gene therapy to introduce chimeric antigen receptors (CARs) and T cell receptors (TCRs) in T cells creates new immunotherapy options for patients. While showing early success with these approaches, limitations remain that can be overcome by the use of modification of hematopoietic stem cells (HSCs) to express CARs and TCRs. With modern gene therapy technologies, increased safety and control of the modification of the HSCs can be achieved through the use of a suicide gene.

  16. 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

  17. Modified Bleomycin Disaccharides Exhibiting Improved Tumor Cell Targeting

    PubMed Central

    2015-01-01

    The bleomycins (BLMs) are a family of antitumor antibiotics used clinically for anticancer chemotherapy. Their antitumor selectivity derives at least in part from their ability to target tumor cells, a property that resides in the carbohydrate moiety of the antitumor agent. In earlier studies, we have demonstrated that the tumor cell selectivity resides in the mannose carbamoyl moiety of the BLM saccharide and that both the BLM disaccharide and monosaccharide containing the carbamoyl moiety were capable of the delivery/uptake of a conjugated cyanine dye into cultured cancer cell lines. Presently, the nature of the participation of the carbamoyl moiety has been explored further to provide compounds of utility for defining the nature of the mechanism of tumor cell recognition and uptake by BLM saccharides and in the hope that more efficient compounds could be identified. A library of seven disaccharide–Cy5** dye conjugates was prepared that are structural analogues of the BLM disaccharide. These differed from the natural BLM disaccharide in the position, orientation, and substitution of the carbamoyl group. Studies of these compounds in four matched sets of tumor and normal cell lines revealed a few that were both tumor cell selective and internalized 2–4-fold more efficiently than the natural BLM disaccharide. PMID:25272367

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

    PubMed Central

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

    2015-01-01

    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. PMID:25054807

  19. Peptide-modified PELCL electrospun membranes for regulation of vascular endothelial cells.

    PubMed

    Zhou, Fang; Jia, Xiaoling; Yang, Yang; Yang, Qingmao; Gao, Chao; Zhao, Yunhui; Fan, Yubo; Yuan, Xiaoyan

    2016-11-01

    The efficiency of biomaterials used in small vascular repair depends greatly on their ability to interact with vascular endothelial cells (VECs). Rapid endothelialization of the vascular grafts is a promising way to prevent thrombosis and intimal hyperplasia. In this work, modification of electrospun membranes of poly(ethylene glycol)-b-poly(l-lactide-co-ε-caprolactone) (PELCL) by three different peptides for regulation of VECs were studied in order to obtain ideal bioactive biomaterials as small diameter vascular grafts. QK (a mimetic peptide to vascular endothelial growth factor), Arg-Glu-Asp-Val (REDV, a specific adhesive peptide to VECs) and Val-Ala-Pro-Gly (VAPG, a specific adhesive peptide to vascular smooth muscle cells) were investigated. Surface properties of the modified membranes and the response of VECs were verified. It was found that protein adsorption and platelet adhesion were effectively suppressed with the introduction of QK, REDV or VAPG peptides on the PELCL electrospun membranes. Both QK- and REDV-modified electrospun membranes could accelerate the proliferation of VECs in the first 9days, and the QK-modified electrospun membrane promoted cell proliferation more significantly than the REDV-modified one. The REDV-modified PELCL membrane was the most favorable for VECs adhesion than QK- and VAPG-modified membranes. It was suggested that QK- or REDV-modified PELCL electrospun membranes may have great potential applications in cardiovascular biomaterials for rapid endothelialization in situ. PMID:27524062

  20. Peptide-modified PELCL electrospun membranes for regulation of vascular endothelial cells.

    PubMed

    Zhou, Fang; Jia, Xiaoling; Yang, Yang; Yang, Qingmao; Gao, Chao; Zhao, Yunhui; Fan, Yubo; Yuan, Xiaoyan

    2016-11-01

    The efficiency of biomaterials used in small vascular repair depends greatly on their ability to interact with vascular endothelial cells (VECs). Rapid endothelialization of the vascular grafts is a promising way to prevent thrombosis and intimal hyperplasia. In this work, modification of electrospun membranes of poly(ethylene glycol)-b-poly(l-lactide-co-ε-caprolactone) (PELCL) by three different peptides for regulation of VECs were studied in order to obtain ideal bioactive biomaterials as small diameter vascular grafts. QK (a mimetic peptide to vascular endothelial growth factor), Arg-Glu-Asp-Val (REDV, a specific adhesive peptide to VECs) and Val-Ala-Pro-Gly (VAPG, a specific adhesive peptide to vascular smooth muscle cells) were investigated. Surface properties of the modified membranes and the response of VECs were verified. It was found that protein adsorption and platelet adhesion were effectively suppressed with the introduction of QK, REDV or VAPG peptides on the PELCL electrospun membranes. Both QK- and REDV-modified electrospun membranes could accelerate the proliferation of VECs in the first 9days, and the QK-modified electrospun membrane promoted cell proliferation more significantly than the REDV-modified one. The REDV-modified PELCL membrane was the most favorable for VECs adhesion than QK- and VAPG-modified membranes. It was suggested that QK- or REDV-modified PELCL electrospun membranes may have great potential applications in cardiovascular biomaterials for rapid endothelialization in situ.

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

    PubMed

    Abay, Nergis; Gurel Pekozer, Gorke; Ramazanoglu, Mustafa; Kose, Gamze Torun

    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

  2. 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

  3. Efficient infection of primitive hematopoietic stem cells by modified adenovirus.

    PubMed

    Yotnda, P; Onishi, H; Heslop, H E; Shayakhmetov, D; Lieber, A; Brenner, M; Davis, A

    2001-06-01

    Almost all studies of adenoviral vector-mediated gene transfer have made use of the adenovirus type 5 (Ad5). Unfortunately, Ad5 has been ineffective at infecting hematopoietic progenitor cells (HPC). Chimeric Ad5/F35 vectors that have been engineered to substitute the shorter-shafted fiber protein from Ad35 can efficiently infect committed hematopoietic cells and we now show highly effective gene transfer to primitive progenitor subsets. An Ad5GFP and Ad5/F35GFP vector was added to CD34(+) and CD34(-)lineage(-) (lin(-)) HPC. Only 5-20% of CD34(+) and CD34(-)lin(-) cells expressed GFP after Ad5 exposure. In contrast, with the Ad5/F35 vector, 30-70% of the CD34(+), 50-70% of the CD34(-)lin(-) and up to 60% of the CD38(-) HPC expressed GFP and there was little evident cellular toxicity. Because of these improved results, we also analyzed the ability of Ad5/F35 virus to infect the hoechst negative 'side population' (SP) of marrow cells, which appear to be among the very earliest multipotent HPC. Between 51% and 80% of marrow SP cells expressed GFP. The infected populations retained their ability to form colonies in two short-term culture systems, with no loss of viability. We also studied the transfer and expression of immunomodulatory genes, CD40L (cell surface expression) and interleukin-2 (secreted). Both were expressed at immunomodulatory levels for >5 days. The ability of Ad5/F35 to deliver transgenes to primitive HPC with high efficiency and low toxicity in the absence of growth factors provides an improved means of studying the consequences of transient gene expression in these cells.

  4. SLC gene-modified dendritic cells mediate T cell-dependent anti-gastric cancer immune responses in vitro.

    PubMed

    Xue, Gang; Cheng, Ying; Ran, Feng; Li, Xianhui; Huang, Tao; Yang, Yong; Zhang, Yanbiao

    2013-02-01

    Dendritic cells (DCs) are potent professional antigen-presenting cells (APCs) with the ability to prime naïve T cells, and play an important role in the initiation and regulation of immune responses. In this study, we constructed a recombinant adenovirus carrying the SLC gene (Ad-SLC), and detected the biological effects of Ad-SLC-modified DCs as an adjuvant for the initiation of gastric cancer immune responses. Human DCs were transfected with Ad-SLC and the recombinant adenovirus carrying the β-galactosidase gene, Ad-LacZ, respectively. Modified DCs were pulsed with the cell lysate antigen of SGC-7901 cells (a type of gastric cancer cell line) and co-cultured with autologous T cells. The T cells were harvested and incubated with SGC-7901 cells and the cytotoxic function of the T cells was detected. Based on the data, the expression of mature DC phenotypes CD83 and CCR7 was upregulated after transfection with Ad-SLC and the chemotaxis function of DCs was augmented after transfection with Ad-SLC. Moreover, the expression of RANTES in DCs was upregulated by Ad-SLC transfection, while expression levels of IL-12p70 and IL-10 were not significantly altered. When co-cultured with autologous T cells, DCs modified with the SLC gene and pulsed with SGC-7901 cell lysates significantly promoted the proliferation of autologous T cells and induced Th1 differentiation, and displayed a strong cytotoxicity to SGC-7901 cells. In conclusion, Ad-SLC promoted DC maturation, enhancing the ability of DCs for T-cell chemotaxis and T-cell stimulation, and induced specific anti-gastric cancer cellular immunity. Recombinant Ad-SLC-modified DCs may be used as an adjuvant to induce an effective anti-gastric cancer immune response.

  5. Modified Laser and Thermos cell calculations on microcomputers

    SciTech Connect

    Shapiro, A.; Huria, H.C.

    1987-01-01

    In the course of designing and operating nuclear reactors, many fuel pin cell calculations are required to obtain homogenized cell cross sections as a function of burnup. In the interest of convenience and cost, it would be very desirable to be able to make such calculations on microcomputers. In addition, such a microcomputer code would be very helpful for educational course work in reactor computations. To establish the feasibility of making detailed cell calculations on a microcomputer, a mainframe cell code was compiled and run on a microcomputer. The computer code Laser, originally written in Fortran IV for the IBM-7090 class of mainframe computers, is a cylindrical, one-dimensional, multigroup lattice cell program that includes burnup. It is based on the MUFT code for epithermal and fast group calculations, and Thermos for the thermal calculations. There are 50 fast and epithermal groups and 35 thermal groups. Resonances are calculated assuming a homogeneous system and then corrected for self-shielding, Dancoff, and Doppler by self-shielding factors. The Laser code was converted to run on a microcomputer. In addition, the Thermos portion of Laser was extracted and compiled separately to have available a stand alone thermal code.

  6. Extracellular proteases modify cell wall turnover in Bacillus subtilis.

    PubMed Central

    Jolliffe, L K; Doyle, R J; Streips, U N

    1980-01-01

    The rate of turnover of peptidoglycan in exponentially growing cultures of Bacillus subtilis was observed to be sensitive to extracellular protease. In protease-deficient mutants the rates of cell wall turnover were greater than that of wild-type strain 168, whereas hyperprotease-producing strains exhibited decreased rates of peptidoglycan turnover. The rate of peptidogylcan turnover in a protease-deficient strain was decreased when the mutant was grown in the presence of a hyperprotease-producing strain. The addition of phenylmethylsulfonyl fluoride, a serine protease inhibitor, to cultures of hyperprotease-producing strains increased their rates of cell wall turnover. Isolated cell walls of all protease mutants contained autolysin levels equal to or greater than that of wild-type strain 168. The presence of filaments, or cells with incomplete septa, was observed in hyperprotease-producing strains or when a protease-deficient strain was grown in the presence of subtilisin. The results suggest that the turnover of cell walls in B. subtilis may be regulated by extracellular proteases. Images PMID:6102558

  7. Adoptive Therapy with Chimeric Antigen Receptor Modified T Cells of Defined Subset Composition

    PubMed Central

    Riddell, Stanley R.; Sommermeyer, Daniel; Berger, Carolina; Liu, Lingfeng (Steven); Balakrishnan, Ashwini; Salter, Alex; Hudecek, Michael; Maloney, David G.; Turtle, Cameron J.

    2014-01-01

    The ability to engineer T cells to recognize tumor cells through genetic modification with a synthetic chimeric antigen receptor has ushered in a new era in cancer immunotherapy. The most advanced clinical applications are in in targeting CD19 on B cell malignancies. The clinical trials of CD19 CAR therapy have thus far not attempted to select defined subsets prior to transduction or imposed uniformity of the CD4 and CD8 cell composition of the cell products. This review will discuss the rationale for and challenges to utilizing adoptive therapy with genetically modified T cells of defined subset and phenotypic composition. PMID:24667960

  8. Low dose of amino-modified nanoparticles induces cell cycle arrest.

    PubMed

    Kim, Jong Ah; Åberg, Christoffer; de Cárcer, Guillermo; Malumbres, Marcos; Salvati, Anna; Dawson, Kenneth A

    2013-09-24

    The interaction of nanoscaled materials with biological systems is currently the focus of a fast-growing area of investigation. Though many nanoparticles interact with cells without acute toxic responses, amino-modified polystyrene nanoparticles are known to induce cell death. We have found that by lowering their dose, cell death remains low for several days while, interestingly, cell cycle progression is arrested. In this scenario, nanoparticle uptake, which we have recently shown to be affected by cell cycle progression, develops differently over time due to the absence of cell division. This suggests that the same nanoparticles can trigger different pathways depending on exposure conditions and the dose accumulated.

  9. 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.

  10. Aptamer-conjugated dendrimer-modified quantum dots for glioblastoma cells imaging

    NASA Astrophysics Data System (ADS)

    Li, Zhiming; Huang, Peng; He, Rong; Zhang, Xiaomin; Bao, Chenchen; Ren, Qiushi; Cui, Daxiang

    2009-09-01

    Targeted quantum dots have shown potential as a platform for development of cancer imaging. Aptamers have recently been demonstrated as ideal candidates for molecular targeting applications. In present work, polyamidoamine dendrimers were used to modify surface of quantum dots and improve their solubility in water solution. Then, dendrimer-modified quantum dots were conjugated with DNA aptamer, GBI-10, can recognize the extracellular matrix protein tenascin-C on the surface of human glioblastoma cells. The dendrimer-modified quantum dots exhibit water-soluble, high quantum yield, and good biocompatibility. Aptamer-conjugated quantum dots can specifically target U251 human glioblastoma cells. High-performance aptamer-conjugated dendrimers modified quantum dot-based nanoprobes have great potential in application such as cancer imaging.

  11. Genetic Modification of Human Pancreatic Progenitor Cells Through Modified mRNA.

    PubMed

    Lu, Song; Chow, Christie C; Zhou, Junwei; Leung, Po Sing; Tsui, Stephen K; Lui, Kathy O

    2016-01-01

    In this chapter, we describe a highly efficient genetic modification strategy for human pancreatic progenitor cells using modified mRNA-encoding GFP and Neurogenin-3. The properties of modified mRNA offer an invaluable platform to drive protein expression, which has broad applicability in pathway regulation, directed differentiation, and lineage specification. This approach can also be used to regulate expression of other pivotal transcription factors during pancreas development and might have potential therapeutic values in regenerative medicine. PMID:27236809

  12. Genetic Modification of Human Pancreatic Progenitor Cells Through Modified mRNA.

    PubMed

    Lu, Song; Chow, Christie C; Zhou, Junwei; Leung, Po Sing; Tsui, Stephen K; Lui, Kathy O

    2016-01-01

    In this chapter, we describe a highly efficient genetic modification strategy for human pancreatic progenitor cells using modified mRNA-encoding GFP and Neurogenin-3. The properties of modified mRNA offer an invaluable platform to drive protein expression, which has broad applicability in pathway regulation, directed differentiation, and lineage specification. This approach can also be used to regulate expression of other pivotal transcription factors during pancreas development and might have potential therapeutic values in regenerative medicine.

  13. 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. PMID:26993532

  14. 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. PMID:26723521

  15. Chromatin states modify network motifs contributing to cell-specific functions

    PubMed Central

    Zhao, Hongying; Liu, Tingting; Liu, Ling; Zhang, Guanxiong; Pang, Lin; Yu, Fulong; Fan, Huihui; Ping, Yanyan; Wang, Li; Xu, Chaohan; Xiao, Yun; Li, Xia

    2015-01-01

    Epigenetic modification can affect many important biological processes, such as cell proliferation and apoptosis. It can alter chromatin conformation and contribute to gene regulation. To investigate how chromatin states associated with network motifs, we assembled chromatin state-modified regulatory networks by combining 269 ChIP-seq data and chromatin states in four cell types. We found that many chromatin states were significantly associated with network motifs, especially for feedforward loops (FFLs). These distinct chromatin state compositions contribute to different expression levels and translational control of targets in FFLs. Strikingly, the chromatin state-modified FFLs were highly cell-specific and, to a large extent, determined cell-selective functions, such as the embryonic stem cell-specific bivalent modification-related FFL with an important role in poising developmentally important genes for expression. Besides, comparisons of chromatin state-modified FFLs between cancerous/stem and primary cell lines revealed specific type of chromatin state alterations that may act together with motif structural changes cooperatively contribute to cell-to-cell functional differences. Combination of these alterations could be helpful in prioritizing candidate genes. Together, this work highlights that a dynamic epigenetic dimension can help network motifs to control cell-specific functions. PMID:26169043

  16. 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

  17. 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

  18. 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

  19. Electrical Impedance Analysis of Mammalian Cells Cultured on Polypyrrole-modified Gold Microlectrodes

    NASA Astrophysics Data System (ADS)

    Chen, Guo; Keese, Charles R.; Giaever, Ivar

    2003-03-01

    In the present study we describe an electrical impedance analysis of BSC cells cultured on gold electrodes (250 im in diameter) that were modified with polypyrrole/heparin composites using electrochemical deposition. Atomic force microscope images show that the composite layer has a porous bulk structure and a very rough surface topology. An electrical technique, referred to as ECIS, was used to measure the impedance of both the cell-covered and the cell-free microelectrodes at frequencies from 25 Hz to 60000 Hz. The electrical characteristics of the system can be modeled with 3 parameters, the intercellular resistance (R_b), the cellular membrane (C_m) and the cell-substrate separation (α). When cells are cultured on the polypyrrole-modified microelectrodes, the contribution to the total resistance from α is decreased, which opens a way to eliminate the contribution arising from α so that Rb and Cm can be directly measured.

  20. 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.

  1. 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. PMID:26716261

  2. 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.

  3. Rauwolfia vomitoria inhibits olfaction and modifies olfactory bulb cells.

    PubMed

    Ekong, Moses B; Peter, Aniekan I; Edagha, Innocent A; Ekpene, Ubong U; Friday, Daniel A

    2016-06-01

    The rising cost of orthodox medication has endeared so many to the use of herbs for the management of neurological conditions. Rauwolfia vomitoria (RV) one of such herbs is a rainforest shrub whose parts are used locally in the management of psychiatry and other medical issues. Its usefulness though not in doubt is wrapped with adverse reports as its active constituents depletes brain monoamine and dopamine stores. This motivated this research on the effects of the root bark extract on olfaction and the olfactory bulb of adult Wistar rats. Eighteen adult Wistar rats (220g average) were divided into three groups (n=6); control (placebo), 200mg/kg and 400mg/kg RV root bark extract, respectively. The oral administration lasted for seven days and on day 8, test of olfaction was carried out and the animals immediately anaesthetized with ketamine hydrochloride (i.p.) and perfuse-fixed with 10% neutral buffered formalin. All the brains were processed for histology and immunoreactivity. Results showed loss of body weights and olfaction in the 200mg/kg and 400mg/kg RV groups. There was hypertrophy and atrophy of mitral cells respectively, in the 200mg/kg and 400mg/kg RV groups, while there was hyperplasia of cells in the internal granular and plexiform layers of both groups. There was decreased neuron specific enolase (NSE) and neurofilament (NF) expression in the 200mg/kg RV group, while NF and glial fibrillary acidic protein (GFAP) expression was decreased in the 400mg/kg RV group. However, NSE expression was enhanced in the 400mg/kg group, while GFAP expression was enhanced in the 200mg/kg RV group. These results suggest that these doses of RV affect olfaction and appetite, and stimulate adverse cellular changes in the olfactory bulb.

  4. Rauwolfia vomitoria inhibits olfaction and modifies olfactory bulb cells.

    PubMed

    Ekong, Moses B; Peter, Aniekan I; Edagha, Innocent A; Ekpene, Ubong U; Friday, Daniel A

    2016-06-01

    The rising cost of orthodox medication has endeared so many to the use of herbs for the management of neurological conditions. Rauwolfia vomitoria (RV) one of such herbs is a rainforest shrub whose parts are used locally in the management of psychiatry and other medical issues. Its usefulness though not in doubt is wrapped with adverse reports as its active constituents depletes brain monoamine and dopamine stores. This motivated this research on the effects of the root bark extract on olfaction and the olfactory bulb of adult Wistar rats. Eighteen adult Wistar rats (220g average) were divided into three groups (n=6); control (placebo), 200mg/kg and 400mg/kg RV root bark extract, respectively. The oral administration lasted for seven days and on day 8, test of olfaction was carried out and the animals immediately anaesthetized with ketamine hydrochloride (i.p.) and perfuse-fixed with 10% neutral buffered formalin. All the brains were processed for histology and immunoreactivity. Results showed loss of body weights and olfaction in the 200mg/kg and 400mg/kg RV groups. There was hypertrophy and atrophy of mitral cells respectively, in the 200mg/kg and 400mg/kg RV groups, while there was hyperplasia of cells in the internal granular and plexiform layers of both groups. There was decreased neuron specific enolase (NSE) and neurofilament (NF) expression in the 200mg/kg RV group, while NF and glial fibrillary acidic protein (GFAP) expression was decreased in the 400mg/kg RV group. However, NSE expression was enhanced in the 400mg/kg group, while GFAP expression was enhanced in the 200mg/kg RV group. These results suggest that these doses of RV affect olfaction and appetite, and stimulate adverse cellular changes in the olfactory bulb. PMID:27208729

  5. 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

    2006-04-01

    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.

  6. 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

  7. Modified hydroxyethyl starch protects cells from oxidative damage.

    PubMed

    Filippov, Sergey K; Sergeeva, Olga Yu; Vlasov, Petr S; Zavyalova, Margarita S; Belostotskaya, Galina B; Garamus, Vasil M; Khrustaleva, Raisa S; Stepanek, Petr; Domnina, Nina S

    2015-12-10

    This article describes the synthesis of novel starch-antioxidant conjugates, which show great potential for biomedical applications to protect cells from oxidative damage. These conjugates were synthesized by the modification of a hydroxyethyl starch (molecular weight=200,000g/mol) with various sterically hindered phenols that differ in radical scavenging activity. They possess substantial radical scavenging activity toward a model free radical. It was found that the polymer conjugate conformation depends on the antioxidant structure and degree of substitution. We constructed the complete conformational phase behavior for the polymers with increasing degrees of substitution from small-angle neutron scattering data. It was observed that the conjugate conformation changes are the result of water shifting from a thermodynamically favorable solvent to an unfavorable one, a process that then leads to compaction of the conjugate. We selected the conjugates that possess high substitution degree but still exhibit coil conformation for biological studies. The high efficiency of the conjugates was confirmed by different in vitro (hypotonic hemolysis of erythrocytes/osmotic resistance of erythrocytes and the change of [Ca(2+)]i inside freshly isolated cardiomyocytes) and in vivo (acute hemorrhage/massive blood loss) methods. PMID:26428130

  8. Gene-modified embryonic stem cell test to characterize chemical risks.

    PubMed

    Kitada, Kohei; Kizu, Akane; Teramura, Takeshi; Takehara, Toshiyuki; Hayashi, Masami; Tachibana, Daisuke; Wanibuchi, Hideki; Fukushima, Shoji; Koyama, Masayasu; Yoshida, Kayo; Morita, Takashi

    2015-11-01

    A high-throughput test of cell growth inhibition was performed using mouse embryonic stem (ES) cells to assess chemical toxicities. We herein demonstrated using a 96-well culture plate approach and the MTT assay that this method was suitable for prioritization of chemicals for their cytotoxic properties. In order to categorize chemicals, we used p53 gene-modified mouse ES cells as well as wild-type ES cells. The p53 gene is a well-known tumor suppressor and controls programmed cell death (apoptosis) and cellular senescence that is triggered by DNA-damaging agents such as alkylating agents and radiation. In the present study, p53-deficient ES cells were found to be more resistant to a tumor initiator, diethylnitrosamine (DEN), than wild-type ES cells, suggesting the inhibition of apoptosis or senescence by a dysfunction in p53. Chromosome aberrations were more frequently detected in p53-deficient ES cells than in wild-type cells, indicating genomic instability due to the deletion of p53. Other tumor initiators, methyl methanesulfonate (MMS) and N-methyl-N-nitrosourea (NMU), did not reveal apparent differences in cytotoxicity between wild-type and p53-deficient ES cells. Thus, ES test system using gene-modified ES cells may be used to categorize chemicals by detecting their characteristic effects on apoptosis, genotoxic potentials as well as general cytotoxicity.

  9. 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

  10. 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.

  11. 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

  12. Cell interaction with modified nanotubes formed on titanium alloy Ti-6Al-4V.

    PubMed

    Moravec, Hynek; Vandrovcova, Marta; Chotova, Katerina; Fojt, Jaroslav; Pruchova, Eva; Joska, Ludek; Bacakova, Lucie

    2016-08-01

    Nanotubes with diameters ranging from 40 to 60nm were prepared by electrochemical oxidation of the Ti-6Al-4V alloy in electrolyte containing ammonium sulphate and ammonium fluoride. The nanotubes were further modified with calcium and phosphate ions or were heat treated. Polished Ti-6Al-4V alloy served as a reference sample. The spreading of human osteoblast-like cells was similar on all nanotube samples but lower than on polished samples. The number of initially adhered cells was higher on non-modified nanotubes, but the final cell number was the highest on Ca-enriched nanotubes and the lowest on heat-treated nanotubes. However, these differences were relatively small and less pronounced than the differences in the concentration of specific molecular markers of cell adhesion and differentiation, estimated by their intensity of immunofluorescence staining. The concentration of vinculin, i.e. a protein of focal adhesion plaques, was the lowest on nanotubes modified with calcium. Collagen I, an early marker of osteogenic cell differentiation, was also the lowest on samples modified with calcium and was highest on polished samples. Alkaline phosphatase, a middle marker of osteogenic differentiation, was observed in lowest concentration on nanotubes modified with phosphorus and the highest on heat-treated samples. Osteocalcin concentrations, a late marker of osteogenic cell differentiation, were similar on all tested samples, although they tended to be the highest on heat-treated samples. Thus, osteogenic differentiation can be modulated by various additional treatments of nanotube coatings on Ti-6Al-4V implants. PMID:27157757

  13. 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

  14. Influence of zeta potential on the flocculation of cyanobacteria cells using chitosan modified soil.

    PubMed

    Li, Liang; Zhang, Honggang; Pan, Gang

    2015-02-01

    Using chitosan modified soil to flocculate and sediment algal cells has been considered as a promising strategy to combat cyanobacteria blooms in natural waters. However, the flocculation efficiency often varies with algal cells with different zeta potential (ZP) attributed to different growth phases or water conditions. This article investigated the relationship between ZP of Microcystis aeruginosa and its influence to the flocculation efficiency using chitosan modified soil. Results suggested that the optimal removal efficiency was obtained when the ZP was between -20.7 and -6.7 mV with a removal efficiency of more than 80% in 30 min and large floc size of >350 μm. When the algal cells were more negatively charged than -20.7 mV, the effect of chitosan modified soil was depressed (<60%) due to the insufficient charge density of chitosan to neutralize and destabilize the algal suspension. When the algal cells were less negative than -6.7 mV or even positively charged, a small floc size (<120 μm) was formed, which may be difficult to sink under natural water conditions. Therefore, manipulation of ZP provided a viable tool to improve the flocculation efficiency of chitosan modified soil and an important guidance for practical engineering of cyanobacteria bloom control.

  15. Novel cellular therapies for leukemia: CAR-modified T cells targeted to the CD19 antigen.

    PubMed

    Brentjens, Renier J; Curran, Kevin J

    2012-01-01

    The ability of immune-competent donor T cells to mediate a beneficial graft-versus-leukemia (GVL) effect was first identified in the setting of allogeneic hematopoietic stem cell transplantation (allo-HSCT) for hematologic malignancies. Unfortunately, with the exception of chronic myelogenous leukemia and EBV-induced lymphoproliferative disease, allo-HSCT GVL lacks the potency to significantly affect disease progression or recurrence in most other hematologic malignancies. The inadequacy of a GVL effect using past approaches is particularly evident in patients with lymphoid malignancies. However, with the advent of improved gene transfer technology, genetically modified tumor-specific immune effectors have extended cellular immunotherapy to lymphoid malignancies. One promising strategy entails the introduction of genes encoding artificial receptors called chimeric antigen receptors (CARs), which redirect the specificity and function of immune effectors. CAR-modified T cells targeted to the B cell-specific CD19 antigen have demonstrated promising results in multiple early clinical trials, supporting further investigation in patients with B-cell cancers. However, disparities in clinical trial design and CAR structure have complicated the discovery of the optimal application of this technology. Recent preclinical studies support additional genetic modifications of CAR-modified T cells to achieve optimal clinical efficacy using this novel adoptive cellular therapy.

  16. 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. PMID:26652045

  17. 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.

  18. 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

  19. Surface-modified hyaluronic acid hydrogels to capture endothelial progenitor cells.

    PubMed

    Camci-Unal, Gulden; Aubin, Hug; Ahari, Amirhossein Farajzadeh; Bae, Hojae; Nichol, Jason William; Khademhosseini, Ali

    2010-10-21

    A major challenge to the effective treatment of injured cardiovascular tissues is the promotion of endothelialization of damaged tissues and implanted devices. For this reason, there is a need for new biomaterials that promote endothelialization to enhance vascular repair. The goal of this work was to develop antibody-modified polysaccharide-based hydrogels that could selectively capture endothelial progenitor cells (EPCs). We showed that CD34 antibody immobilization on hyaluronic acid (HA) hydrogels provides a suitable surface to capture EPCs. The effect of CD34 antibody immobilization on EPC adhesion was found to be dependent on antibody concentration. The highest level of EPC attachment was found to be 52.2 cells per mm(2) on 1% HA gels modified with 25 μg mL(-1) antibody concentration. Macrophages did not exhibit significant attachment on these modified hydrogel surfaces compared to the EPCs, demonstrating the selectivity of the system. Hydrogels containing only HA, with or without immobilized CD34, did not allow for spreading of EPCs 48 h after cell seeding, even though the cells were adhered to the hydrogel surface. To promote spreading of EPCs, 2% (w/v) gelatin methacrylate (GelMA) containing HA hydrogels were synthesized and shown to improve cell spreading and elongation. This strategy could potentially be useful to enhance the biocompatibility of implants such as artificial heart valves or in other tissue engineering applications where formation of vascular structures is required.

  20. 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.

  1. The Control of Mesenchymal Stromal Cell Osteogenic Differentiation through Modified Surfaces.

    PubMed

    Logan, Niall; Brett, Peter

    2013-01-01

    Stem cells continue to receive widespread attention due to their potential to revolutionise treatments in the fields of both tissue engineering and regenerative medicine. Adult stem cells, specifically mesenchymal stromal cells (MSCs), play a vital role in the natural events surrounding bone healing and osseointegration through being stimulated to differentiate along their osteogenic lineage and in doing so, they form new cortical and trabecular bone tissue. Understanding how to control, manipulate, and enhance the intrinsic healing events modulated through osteogenic differentiation of MSCs by the use of modified surfaces and biomaterials could potentially advance the fields of both orthopaedics and dentistry. This could be by either using surface modification to generate greater implant stability and more rapid healing following implantation or the stimulation of MSCs ex vivo for reimplantation. This review aims to gather publications targeted at promoting, enhancing, and controlling the osteogenic differentiation of MSCs through biomaterials, nanotopographies, and modified surfaces for use in implant procedures. PMID:23766768

  2. The Control of Mesenchymal Stromal Cell Osteogenic Differentiation through Modified Surfaces

    PubMed Central

    2013-01-01

    Stem cells continue to receive widespread attention due to their potential to revolutionise treatments in the fields of both tissue engineering and regenerative medicine. Adult stem cells, specifically mesenchymal stromal cells (MSCs), play a vital role in the natural events surrounding bone healing and osseointegration through being stimulated to differentiate along their osteogenic lineage and in doing so, they form new cortical and trabecular bone tissue. Understanding how to control, manipulate, and enhance the intrinsic healing events modulated through osteogenic differentiation of MSCs by the use of modified surfaces and biomaterials could potentially advance the fields of both orthopaedics and dentistry. This could be by either using surface modification to generate greater implant stability and more rapid healing following implantation or the stimulation of MSCs ex vivo for reimplantation. This review aims to gather publications targeted at promoting, enhancing, and controlling the osteogenic differentiation of MSCs through biomaterials, nanotopographies, and modified surfaces for use in implant procedures. PMID:23766768

  3. Behavior of encapsulated MG-63 cells in RGD and gelatine-modified alginate hydrogels.

    PubMed

    Grigore, Alexandra; Sarker, Bapi; Fabry, Ben; Boccaccini, Aldo R; Detsch, Rainer

    2014-08-01

    Achieving cell spreading and proliferation inside hydrogels that are compatible with microencapsulation technology represents a major challenge for tissue engineering scaffolding and for the development of three-dimensional cell culture models. In this study, microcapsules of 650-900 μm in diameter were fabricated from oxidized alginate covalently cross-linked with gelatine (AlGel). Schiff's base bond formed in AlGel, detected by Fourier transform infrared spectroscopy, which confirmed the cross-linking of oxidized alginate with gelatine. Biological properties of alginate based hydrogels were studied by comparing the viability and morphology of MG-63 osteosarcoma cells encapsulated in gelatine and RGD-modified alginate. We hypothesized that the presence of gelatine and RGD will support cell adhesion and spreading inside the microcapsules and finally, also vascular endothelial growth factor (VEGF) secretion. After 4 days of incubation, cells formed extensive cortical protrusions and after 2 weeks they proliferated, migrated, and formed cellular networks through the AlGel material. In contrast, cells encapsulated in pure alginate and in RGD-modified alginate formed spherical aggregates with limited cell mobility and VEGF secretion. Metabolic activity was doubled after 5 days of incubation, making AlGel a promising material for cell encapsulation.

  4. A modified method by differential adhesion for enrichment of bladder cancer stem cells

    PubMed Central

    Zhu, Yong-tong; Pang, Shi-yu; Luo, Yang; Chen, Wei; Bao, Ji-ming; Tan, Wan-long

    2016-01-01

    ABSTRACT Purpose: In a previous study the vaccine was effective against bladder cancer in a mouse model. However, a small portion of tumors regrew because the vaccine could not eliminate bladder cancer stem cells (CSCs). In this study, we showed a modified method for the isolation of bladder CSCs using a combination of differential adhesion method and serum-free culture medium (SFM) method. Materials and Methods: Trypsin-resistant cells and trypsin-sensitive cells were isolated from MB49, EJ and 5637 cells by a combination of differential adhesion method and SFM method. The CSCs characterizations of trypsin-resistant cells were verified by the flow cytometry, the western blotting, the quantitative polymerase chain reaction, the resistance to chemotherapy assay, the transwell assay, and the tumor xenograft formation assay. Results: Trypsin-resistant cells were isolated and identified in CSCs characters, with high expression of CSCs markers, higher resistance to chemotherapy, greater migration in vitro, and stronger tumorigenicity in vivo. Conclusion: Trypsin-resistant cells displayed specific CSCs properties. Our study showed trypsin-resistant cells were isolated successfully with a modified method using a combination of differential adhesion method and SFM method. PMID:27564296

  5. Tolerization of a type I allergic immune response through transplantation of genetically modified hematopoietic stem cells.

    PubMed

    Baranyi, Ulrike; Linhart, Birgit; Pilat, Nina; Gattringer, Martina; Bagley, Jessamyn; Muehlbacher, Ferdinand; Iacomini, John; Valenta, Rudolf; Wekerle, Thomas

    2008-06-15

    Allergy represents a hypersensitivity disease that affects >25% of the population in industrialized countries. The underlying type I allergic immune reaction occurs in predisposed atopic individuals in response to otherwise harmless Ags (i.e., allergens) and is characterized by the production of allergen-specific IgE, an allergen-specific T cell response, and the release of biologically active mediators such as histamine from mast cells and basophils. Regimens permanently tolerizing an allergic immune response still need to be developed. We therefore retrovirally transduced murine hematopoietic stem cells to express the major grass pollen allergen Phl p 5 on their cell membrane. Transplantation of these genetically modified hematopoietic stem cells led to durable multilineage molecular chimerism and permanent immunological tolerance toward the introduced allergen at the B cell, T cell, and effector cell levels. Notably, Phl p 5-specific serum IgE and IgG remained undetectable, and T cell nonresponsiveness persisted throughout follow-up (40 wk). Besides, mediator release was specifically absent in in vitro and in vivo assays. B cell, T cell, and effector cell responses to an unrelated control allergen (Bet v 1) were unperturbed, demonstrating specificity of this tolerance protocol. We thus describe a novel cell-based strategy for the prevention of allergy.

  6. 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

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

    PubMed

    Jin, Li-Fang; Li, Jin-Song

    2016-07-18

    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

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

    PubMed Central

    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. PMID:23982586

  9. 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

  10. Heat-Modified Citrus Pectin Induces Apoptosis-Like Cell Death and Autophagy in HepG2 and A549 Cancer Cells

    PubMed Central

    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. PMID:25794149

  11. PB/PANI-modified electrode used as a novel oxygen reduction cathode in microbial fuel cell.

    PubMed

    Fu, Lei; You, Shi-Jie; Zhang, Guo-Quan; Yang, Feng-Lin; Fang, Xiao-Hong; Gong, Zheng

    2011-01-15

    This study focuses on the preparation of a new type of Prussian Blue/polyaniline (PB/PANI)-modified electrode as oxygen reduction cathode, and its availability in microbial fuel cell (MFC) for biological power generation. The PB/PANI-modified electrode was prepared by electrochemical and chemical methods, both of which exhibited good electrocatalytical reactivity for oxygen reduction in acidic electrolyte. The MFC with PB/PANI-modified cathode aerated by either oxygen or air was shown to yield a maximum power density being the same with that of the MFC with liquid-state ferricyanide cathode, and have an excellent duration as indicated by stable cathode potential for more than eight operating circles. This study suggests a promising potential to utilize this novel electrode as an effective alternative to platinum for oxygen reduction in MFC system without losing sustainability.

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

    NASA Astrophysics Data System (ADS)

    Zdetsis, Aristides D.

    2011-04-01

    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.

  13. Safety of targeting ROR1 in primates with chimeric antigen receptor-modified T cells.

    PubMed

    Berger, Carolina; Sommermeyer, Daniel; Hudecek, Michael; Berger, Michael; Balakrishnan, Ashwini; Paszkiewicz, Paulina J; Kosasih, Paula L; Rader, Christoph; Riddell, Stanley R

    2015-02-01

    Genetic engineering of T cells for adoptive transfer by introducing a tumor-targeting chimeric antigen receptor (CAR) is a new approach to cancer immunotherapy. A challenge for the field is to define cell surface molecules that are both preferentially expressed on tumor cells and can be safely targeted with T cells. The orphan tyrosine kinase receptor ROR1 is a candidate target for T-cell therapy with CAR-modified T cells (CAR-T cells) because it is expressed on the surface of many lymphatic and epithelial malignancies and has a putative role in tumor cell survival. The cell surface isoform of ROR1 is expressed in embryogenesis but absent in adult tissues except for B-cell precursors and low levels of transcripts in adipocytes, pancreas, and lung. ROR1 is highly conserved between humans and macaques and has a similar pattern of tissue expression. To determine if low-level ROR1 expression on normal cells would result in toxicity or adversely affect CAR-T cell survival and/or function, we adoptively transferred autologous ROR1 CAR-T cells into nonhuman primates. ROR1 CAR-T cells did not cause overt toxicity to normal organs and accumulated in bone marrow and lymph node sites, where ROR1-positive B cells were present. The findings support the clinical evaluation of ROR1 CAR-T cells for ROR1(+) malignancies and demonstrate the utility of nonhuman primates for evaluating the safety of immunotherapy with engineered T cells specific for tumor-associated molecules that are homologous between humans and nonhuman primates.

  14. [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.

  15. KRIT1 Protein Depletion Modifies Endothelial Cell Behavior via Increased Vascular Endothelial Growth Factor (VEGF) Signaling*

    PubMed Central

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

    2014-01-01

    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. PMID:25320085

  16. Evaluation of cell behavior on modified polypropylene with swift heavy ion irradiation

    NASA Astrophysics Data System (ADS)

    Arbeitman, Claudia R.; Ibañez, Irene L.; García Bermúdez, Gerardo; Durán, Hebe; del Grosso, Mariela F.; Salguero, Noelia; Mazzei, Rubén

    2012-02-01

    Ion beam irradiation is a well known means to change the physico-chemical properties of polymers, and induced bio and citocompatibility in controlled conditions and in selected areas of surface. However, the enhancement of cell adhesion on a modified substrate does not mean that the surface is adequate for functional cells. The purpose of the present work is to study proliferation, changes in cytoskeleton and cell morphology on substrates as a function of irradiation parameters. We irradiated polypropylene with sulfur (S) ion-beam at energies of 110 MeV with fluences between 1 × 10 6 and 2 × 10 10 ions cm -2. NIH 3T3 cells were cultured on each sample. Cell morphology was observed using phase contrast microscopy and cytoskeleton proteins with fluorescence microscopy. The analysis show different cellular responses as a functions of irradiation parameter, strongly suggests that different underlying substratum can result in distinct types of cytoskeleton reorganization.

  17. Screening for Glycosylphosphatidylinositol-Modified Cell Wall Proteins in Pichia pastoris and Their Recombinant Expression on the Cell Surface

    PubMed Central

    Zhang, Li; Liang, Shuli; Zhou, Xinying; Jin, Zi; Jiang, Fengchun; Han, Shuangyan; Zheng, Suiping

    2013-01-01

    Glycosylphosphatidylinositol (GPI)-anchored glycoproteins have various intrinsic functions in yeasts and different uses in vitro. In the present study, the genome of Pichia pastoris GS115 was screened for potential GPI-modified cell wall proteins. Fifty putative GPI-anchored proteins were selected on the basis of (i) the presence of a C-terminal GPI attachment signal sequence, (ii) the presence of an N-terminal signal sequence for secretion, and (iii) the absence of transmembrane domains in mature protein. The predicted GPI-anchored proteins were fused to an alpha-factor secretion signal as a substitute for their own N-terminal signal peptides and tagged with the chimeric reporters FLAG tag and mature Candida antarctica lipase B (CALB). The expression of fusion proteins on the cell surface of P. pastoris GS115 was determined by whole-cell flow cytometry and immunoblotting analysis of the cell wall extracts obtained by β-1,3-glucanase digestion. CALB displayed on the cell surface of P. pastoris GS115 with the predicted GPI-anchored proteins was examined on the basis of potential hydrolysis of p-nitrophenyl butyrate. Finally, 13 proteins were confirmed to be GPI-modified cell wall proteins in P. pastoris GS115, which can be used to display heterologous proteins on the yeast cell surface. PMID:23835174

  18. Chitosan-modified porous silicon microparticles for enhanced permeability of insulin across intestinal cell monolayers.

    PubMed

    Shrestha, Neha; Shahbazi, Mohammad-Ali; Araújo, Francisca; Zhang, Hongbo; Mäkilä, Ermei M; Kauppila, Jussi; Sarmento, Bruno; Salonen, Jarno J; Hirvonen, Jouni T; Santos, Hélder A

    2014-08-01

    Porous silicon (PSi) based particulate systems are emerging as an important drug delivery system due to its advantageous properties such as biocompatibility, biodegradability and ability to tailor the particles' physicochemical properties. Here, annealed thermally hydrocarbonized PSi (AnnTHCPSi) and undecylenic acid modified AnnTHCPSi (AnnUnTHCPSi) microparticles were developed as a PSi-based platform for oral delivery of insulin. Chitosan (CS) was used to modify the AnnUnTHCPSi microparticles to enhance the intestinal permeation of insulin. Surface modification with CS led to significant increase in the interaction of PSi microparticles with Caco-2/HT-29 cell co-culture monolayers. Compared to pure insulin, the CS-conjugated microparticles significantly improved the permeation of insulin across the Caco-2/HT-29 cell monolayers, with ca. 20-fold increase in the amount of insulin permeated and ca. 7-fold increase in the apparent permeability (P(app)) value. Moreover, among all the investigated particles, the CS-conjugated microparticles also showed the highest amount of insulin associated with the mucus layer and the intestinal Caco-2 cells and mucus secreting HT-29 cells. Our results demonstrate that CS-conjugated AnnUnTHCPSi microparticles can efficiently enhance the insulin absorption across intestinal cells, and thus, they are promising microsystems for the oral delivery of proteins and peptides across the intestinal cell membrane.

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

    PubMed Central

    Wu, Min-fei; Zhang, Shu-quan; Gu, Rui; Liu, Jia-bei; Li, Ye; Zhu, Qing-san

    2015-01-01

    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

  20. 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

  1. Ultrastructure of modified root-tip cells in Ficus carica, induced by the ectoparasitic nematode Xiphinema index.

    PubMed

    Wyss, U; Lehmann, H; Jank-Ladwig, R

    1980-02-01

    The migratory ectoparasitic root nematode Xiphinema index, added to Ficus carica seedlings in sterile agar culture, fed exclusively on the tips of the roots. As a response the tips started to swell and became transformed into terminal galls as long as feeding was continued. When the cytology of swollen root-tips was examined 24 h after the first nematode attack, necrotic cells, scattered singly or in small groups within the root apex, were found in ultrathin sections. These cells, whose protoplasts showed features of a hypersensitive reaction, were most probably those fed upon by nematodes. Each necrotic cell was surrounded by several enlarged, mostly binucleate cells with dense cytoplasm. One day later the binucleate cells were multinucleate, containing 4 or even 8 nuclei. The clear-cut demarcation between necrotic and modified cells indicated that only the stimulus for the induction of modified cells but not the stimulus for cell necrosis passed into neighbouring cells. Root-tip galls that provided the appropriate food for egg production in nematodes contained greatly enlarged multinucleate cells between necrotic cells. The modified cells showed features of high metabolic activities, expressed in nuclear and nucleolar hypertrophy, invagination of the nuclear envelope, increased cytoplasmic density, abundance of mitochondria, plastids and rough endoplasmic reticulum. Wall ingrowths, typical of transfer cells, were rare and if present occurred only adjacent to necrotic cells. In older modified cells new cell plates, surrounded by phragmoplasts, were formed.

  2. 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...

  3. Chemically crosslinked alginate porous microcarriers modified with bioactive molecule for expansion of human hepatocellular carcinoma cells.

    PubMed

    Li, Chunge; Zhao, Shuang; Zhao, Yanyan; Qian, Yufeng; Li, Junjie; Yin, Yuji

    2014-11-01

    Microcarrier is an essential matrix for the large-scale culture of anchorage-dependent cells. In this study, chemical cross-linked alginate porous microcarriers (AMC) were prepared using microemulsion and freeze-drying technology. Moreover, chitosan was coated on the surface of microcarriers (AMC-CS) via electrostatic interactions to improve the mechanical strength. The size of AMC can be modulated through adjusting the concentration of alginate, amount of dispersant and stirring rate. The surface chemical characteristics and morphology of AMC-CS were evaluated by Fourier transformed infrared, X-ray photoelectron spectroscopy, and scanning electron microscope. Fibronectin (Fn) or heparin/basic fibroblast growth factor (bFGF) was then immobilized on the surface of microcarriers via layer-by-layer technology to improve the cytocompatibility. Our data suggested that the size of AMC can be accurately modulated from 90 μm to 900 μm with a narrow size distribution. Micropore structures of AMC-CS were relatively disordered and the pore size ranged between 20 μm and 100 μm. Using AMC after modified with Fn or bFGF as the cell expansion microcarriers, we showed that the proliferation rates of HepG2 cells increased significantly, reaching to more than 30-fold of cell expansion after 10 days of culture, with minor cellular damage caused by the microcarriers. Moreover, the AMC microcarriers modified with Fn or bFGF can increase albumin secretion of HepG2. We suggest that our new modified AMC-based microcarriers will be an attractive candidate for the large-scale cell culture of therapeutic cells.

  4. Enhanced performance of dye-sensitized solar cells using gold nanoparticles modified fluorine tin oxide electrodes

    NASA Astrophysics Data System (ADS)

    Zhang, Dingwen; Wang, Milton; Brolo, Alexandre G.; Shen, Jie; Li, Xiaodong; Huang, Sumei

    2013-01-01

    We have investigated plasmon-assisted energy conversion in dye-sensitized solar cells (DSCs) applying gold nanoparticles (NPs) modified fluorine tin oxide (FTO) electrodes. A series of Au NPs with different sizes (15-80 nm) were synthesized and immobilized onto FTO glass slides. Photoanodes were prepared on these Au modified FTO substrates using P25 TiO2 powders and by the screen-printing method. The size effects of Au NPs on the photovoltaic performance of the formed DSCs were investigated systematically. Structural and photoelectrochemical properties of the formed photoanodes were examined by field emission scanning electron microscopy and electrochemical impedance spectroscopy. It was found that the energy conversion efficiency of the DSC was highly dependent on the Au particle size. When the particle size was not greater than 60 nm, the DSC based on the Au NP-FTO composite electrode showed a higher short-circuit current density and better photovoltaic (PV) performance than the cell based on the bare FTO. The best cell was achieved using 25 nm sized Au NPs modified FTO. It exhibited a conversion efficiency of 6.69%, which was 15% higher than that of DSCs without Au NPs. The related PV performance enhancement mechanisms, photoelectrochemical processes and surface-plasmon resonances in DSCs with Au nanostructures are analysed and discussed.

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

    PubMed

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

    2016-03-01

    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.

  6. 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

  7. 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. PMID:27338712

  8. 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.

  9. 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.

  10. 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.

  11. 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.

  12. Loss of the HVEM Tumor Suppressor in Lymphoma and Restoration by Modified CAR-T Cells.

    PubMed

    Boice, Michael; Salloum, Darin; Mourcin, Frederic; Sanghvi, Viraj; Amin, Rada; Oricchio, Elisa; Jiang, Man; Mottok, Anja; Denis-Lagache, Nicolas; Ciriello, Giovanni; Tam, Wayne; Teruya-Feldstein, Julie; de Stanchina, Elisa; Chan, Wing C; Malek, Sami N; Ennishi, Daisuke; Brentjens, Renier J; Gascoyne, Randy D; Cogné, Michel; Tarte, Karin; Wendel, Hans-Guido

    2016-10-01

    The HVEM (TNFRSF14) receptor gene is among the most frequently mutated genes in germinal center lymphomas. We report that loss of HVEM leads to cell-autonomous activation of B cell proliferation and drives the development of GC lymphomas in vivo. HVEM-deficient lymphoma B cells also induce a tumor-supportive microenvironment marked by exacerbated lymphoid stroma activation and increased recruitment of T follicular helper (TFH) cells. These changes result from the disruption of inhibitory cell-cell interactions between the HVEM and BTLA (B and T lymphocyte attenuator) receptors. Accordingly, administration of the HVEM ectodomain protein (solHVEM((P37-V202))) binds BTLA and restores tumor suppression. To deliver solHVEM to lymphomas in vivo, we engineered CD19-targeted chimeric antigen receptor (CAR) T cells that produce solHVEM locally and continuously. These modified CAR-T cells show enhanced therapeutic activity against xenografted lymphomas. Hence, the HVEM-BTLA axis opposes lymphoma development, and our study illustrates the use of CAR-T cells as "micro-pharmacies" able to deliver an anti-cancer protein.

  13. Derivation of LIF-independent mouse iPS cells with modified Oct4

    PubMed Central

    Hirai, Hiroyuki; Firpo, Meri; Kikyo, Nobuaki

    2015-01-01

    It has been very difficult, if not impossible, to establish mouse induced pluripotent stem cells (iPSCs) from differentiated cells, such as fibroblasts, without leukemia inhibitory factor (LIF). We have established and maintained LIF-independent iPSCs for longer than 120 days with modified Oct4 along with Sox2, Klf4, and c-Myc. The iPSCs will provide a novel tool to investigate the roles of the LIF-Stat3 signaling pathway in mouse pluripotent stem cells. Resource TableBiological reagent: induced pluripotent stem (iPS) cellName of Stem Cell constructM3O-lenti-iPSC-LIF(−)InstitutionUniversity of MinnesotaPerson who created resourceHiroyuki HiraiContact person and emailNobuaki Kikyo, kikyo001@umn.eduDate archived/stock date2014–2015OriginMouse embryonic fibroblastsType of resourceBiological reagent: induced pluripotent stem (iPS) cellSub-typeCell lineKey transcription factorsM3O, Sox2, Klf4, c-MycAuthenticationIdentity and purity of cell line confirmed (Figure 1)Link to related literature (direct URL links and full references)Hirai H., et al. In press, Stem Cell ResearchInformation in public databases PMID:26318720

  14. Impact of Pdx1-associated chromatin modifiers on islet β-cells.

    PubMed

    Spaeth, J M; Walker, E M; Stein, R

    2016-09-01

    Diabetes mellitus arises from insufficient insulin secretion from pancreatic islet β-cells. In type 2 diabetes (T2D), β-cell dysfunction is associated with inactivation and/or loss of transcription factor (TF) activity, including Pdx1. Notably, this particular TF is viewed as a master regulator of pancreas development and islet β-cell formation, identity and function. TFs, like Pdx1, recruit coregulators to transduce activating and/or repressing signals to the general transcriptional machinery for controlling gene expression, including modifiers of DNA, histones and nucleosome architecture. These coregulators impart a secondary layer of control that can be exploited to modulate TF activity. In this review, we describe Pdx1-recruited coregulators that impact chromatin structure, consequently influencing normal β-cell function and likely Pdx1 activity in pathophysiological settings. PMID:27615141

  15. 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

  16. Ovalbumin Modified with Pyrraline, a Maillard Reaction Product, shows Enhanced T-cell Immunogenicity*

    PubMed Central

    Heilmann, Monika; Wellner, Anne; Gadermaier, Gabriele; Ilchmann, Anne; Briza, Peter; Krause, Maren; Nagai, Ryoji; Burgdorf, Sven; Scheurer, Stephan; Vieths, Stefan; Henle, Thomas; Toda, Masako

    2014-01-01

    The Maillard reaction (also referred to as “glycation”) takes place between reducing sugars and compounds with free amino groups during thermal processing of foods. In the final stage of the complex reaction cascade, the so-called advanced glycation end products (AGEs) are formed, including proteins with various glycation structures. It has been suggested that some AGEs could have immunostimulatory effects. Here, we aimed to identify specific glycation structure(s) that could influence the T-cell immunogenicity and potential allergenicity of food allergens, using ovalbumin (OVA, an egg white allergen) as a model allergen. OVA was specifically modified with representative glycation structures: Nϵ-carboxymethyl lysine (CM-OVA), Nϵ-carboxyethyl lysine (CE-OVA), pyrraline (Pyr-OVA), or methylglyoxal-derived arginine derivatives (MGO-OVA). As well as AGE-OVA, a crude glycation product in thermal incubation of OVA with glucose, only Pyr-OVA, and not other modified OVAs, was efficiently taken up by bone marrow-derived murine dendritic cells (BMDCs). The uptake of Pyr-OVA was reduced in scavenger receptor class A (SR-A)-deficient BMDCs, but not in cells treated with inhibitors of scavenger receptor class B, galectin-3, or blocking antibodies against CD36, suggesting that pyrraline binds to SR-A. Compared with other modified OVAs, Pyr-OVA induced higher activation of OVA-specific CD4+ T-cells in co-culture with BMDCs. Furthermore, compared with native OVA, AGE-OVA and Pyr-OVA induced higher IgE production in mice. Pyrraline could induce better allergen uptake by DCs via association with SR-A and subsequently enhance CD4+ T-cell activation and IgE production. Our findings help us to understand how Maillard reaction enhances the potential allergenicity of food allergens. PMID:24505139

  17. 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

  18. 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

  19. 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.

  20. Improved Photovoltaic Properties of Dye-Sensitized Solar Cells with KNO3-Modified Photoelectrodes.

    PubMed

    Oh, Ju Hee; Lee, Sang-Ju; Kim, Dae-Hwan; Sung, Shi-Joon; Lee, Min Ho; Han, Yoon Soo

    2015-11-01

    The surface of a TiO2 photoelectrode was modified through a dip-coating process using an aqueous potassium nitrate (KNO3) solution to increase the power-conversion efficiency of dye-sensitized solar cells (DSSCs). The KNO3-modified TiO2 electrode was applied to the photoanode of the DSSCs. The DSSC with the KNO3-modified TiO2 electrode exhibited a short-circuit current (J(sc)) of 15.26 mA/cm2 and an open-circuit voltage (V(oc)) of 671 mV, compared with a J(sc) of 13.74 mA/cm2 and V(oc) of 654 mV for a reference device with a pristine TiO2 electrode. The results in combination with relevant data from electrochemical impedance spectroscopy, open-circuit voltage decay, and dark current measurements revealed that the modification of the TiO2 surface using the surface modifier (KNO3) led to a longer electron lifetime by the suppression of the charge recombination between injected electrons and I3- ions, resulting in an increase in both J(sc) and V(oc), compared with those of the reference device without surface modification. PMID:26726607

  1. 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. PMID:27287173

  2. The small ubiquitin-like modifier (SUMO) is essential in cell cycle regulation in Trypanosoma brucei.

    PubMed

    Liao, Shanhui; Wang, Tao; Fan, Kai; Tu, Xiaoming

    2010-03-10

    SUMO, a reversible post-translational protein modifier, plays important roles in many processes of higher eukaryotic cell life. Although SUMO has been identified in many eukaryotes, SUMO and SUMO system are still unknown in some eukaryotic unicellular organisms, such as Trypanosoma brucei (T. brucei). In this study, only one SUMO homologue (TbSUMO) was identified in T. brucei. Expression of TbSUMO was knocked down by using RNA interference technique in procyclic-form T. brucei. The growth of TbSUMO-deficient cells was significantly inhibited. TbSUMO-deficient cells were arrested in G2/M phase accompanied with an obvious increase of 0N1K cells (zoids), and failed in chromosome segregation. These results indicate that TbSUMO is essential in cell cycle regulation, with one important role in mitosis. Meanwhile, the enrichment of zoids suggests the inhibition of mitosis does not prevent the cell division in procyclic-form T. brucei. HA-tagged TbSUMO was overexpressed in T. brucei and was shown to be localized to the nucleus through the whole cell cycle, further revealing its distinguished functions in nucleus. All these accumulated data imply that a SUMO system essential for regulating cell cycle progression might exist in the procyclic-form T. brucei.

  3. A genetically modified protein-based hydrogel for 3D culture of AD293 cells.

    PubMed

    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

  4. A Single-Cell-Type Real-Time PCR Method Based on a Modified Patch-Pipette Cell Harvesting System.

    PubMed

    Song, Yuanlong; Zhang, Miaomiao; Tao, Xiaoqing; Xu, Zifen; Zhang, Liangpin; Zheng, Yunjie; Zhu, Minjie; Gao, Linlin

    2016-09-01

    Real-time PCR is a powerful tool for quantifying nucleic acid expression. Real-time PCR is conventionally performed at the tissue level to guarantee an abundance of nucleic acid for detection. The precision and reliability of this method, however, is limited by usually being composed of a mixture of different cell types. Single-cell PCR, in contrast, eliminates the purity problem of the cell source. However, use of this method is usually impeded by difficulties in cell harvesting and stringent requirements for processing of very small quantities of nucleic acids. In this study, we combined the advantages of the high purity of selected cells in single-cell PCR with the greater nucleic acid quantities and thus greater ease of tissue-level PCR. The key aspect of our method is to use a modified patch-clamp pipette to harvest several selected cells of the same type. This method is therefore especially useful for cells that can be morphologically or histologically identified such as primary sensory neurons, striated muscle fibers and cells labeled with fluorescent makers. PMID:27271017

  5. 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). PMID:21104198

  6. 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).

  7. Use of Genetically Modified Mesenchymal Stem Cells to Treat Neurodegenerative Diseases

    PubMed Central

    Wyse, Robert D.; Dunbar, Gary L.; Rossignol, Julien

    2014-01-01

    The transplantation of mesenchymal stem cells (MSCs) for treating neurodegenerative disorders has received growing attention recently because these cells are readily available, easily expanded in culture, and when transplanted, survive for relatively long periods of time. Given that such transplants have been shown to be safe in a variety of applications, in addition to recent findings that MSCs have useful immunomodulatory and chemotactic properties, the use of these cells as vehicles for delivering or producing beneficial proteins for therapeutic purposes has been the focus of several labs. In our lab, the use of genetic modified MSCs to release neurotrophic factors for the treatment of neurodegenerative diseases is of particular interest. Specifically, glial cell-derived neurotrophic factor (GDNF), nerve growth factor (NGF), and brain derived neurotrophic factor (BDNF) have been recognized as therapeutic trophic factors for Parkinson’s, Alzheimer’s and Huntington’s diseases, respectively. The aim of this literature review is to provide insights into: (1) the inherent properties of MSCs as a platform for neurotrophic factor delivery; (2) the molecular tools available for genetic manipulation of MSCs; (3) the rationale for utilizing various neurotrophic factors for particular neurodegenerative diseases; and (4) the clinical challenges of utilizing genetically modified MSCs. PMID:24463293

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

    PubMed

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

    2015-01-01

    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. PMID:25569516

  9. Tumor immunotherapy using gene-modified human mesenchymal stem cells loaded into synthetic extracellular matrix scaffolds.

    PubMed

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

    2009-03-01

    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 (MSC(luc)), 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 alpha-carcinoembryonic antigen (alphaCEA)/alphaCD3 diabody (MSC(dAb)) 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 alphaCEA/alphaCD3 diabody activated T cells and promoted tumor cell lysis. Reduction of tumor growth in MSC(dAb)-treated mice was statistically significant compared with animals that only received MSC(luc). 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

  10. 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.

  11. Use of genetically modified mesenchymal stem cells to treat neurodegenerative diseases.

    PubMed

    Wyse, Robert D; Dunbar, Gary L; Rossignol, Julien

    2014-01-23

    The transplantation of mesenchymal stem cells (MSCs) for treating neurodegenerative disorders has received growing attention recently because these cells are readily available, easily expanded in culture, and when transplanted, survive for relatively long periods of time. Given that such transplants have been shown to be safe in a variety of applications, in addition to recent findings that MSCs have useful immunomodulatory and chemotactic properties, the use of these cells as vehicles for delivering or producing beneficial proteins for therapeutic purposes has been the focus of several labs. In our lab, the use of genetic modified MSCs to release neurotrophic factors for the treatment of neurodegenerative diseases is of particular interest. Specifically, glial cell-derived neurotrophic factor (GDNF), nerve growth factor (NGF), and brain derived neurotrophic factor (BDNF) have been recognized as therapeutic trophic factors for Parkinson's, Alzheimer's and Huntington's diseases, respectively. The aim of this literature review is to provide insights into: (1) the inherent properties of MSCs as a platform for neurotrophic factor delivery; (2) the molecular tools available for genetic manipulation of MSCs; (3) the rationale for utilizing various neurotrophic factors for particular neurodegenerative diseases; and (4) the clinical challenges of utilizing genetically modified MSCs.

  12. 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.

  13. 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

  14. 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.

  15. A Modified Method of Insulin Producing Cells' Generation from Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Czubak, Paweł; Putowski, Lechosław

    2014-01-01

    Type 1 diabetes mellitus is a result of autoimmune destruction of pancreatic insulin producing β-cells and so far it can be cured only by insulin injection, by pancreas transplantation, or by pancreatic islet cells' transplantation. The methods are, however, imperfect and have a lot of disadvantages. Therefore new solutions are needed. The best one would be the use of differentiated mesenchymal stem cells (MSCs). In the present study, we investigated the potential of the bone marrow-derived MSCs line for in vitro differentiation into insulin producing cells (IPSs). We applied an 18-day protocol to differentiate MSCs. Differentiating cells formed cell clusters some of which resembled pancreatic islet-like cells. Using dithizone we confirmed the presence of insulin in the cells. What is more, the expression of proinsulin C-peptide in differentiated IPCs was analyzed by flow cytometry. For the first time, we investigated the influence of growth factors' concentration on IPCs differentiation efficiency. We have found that an increase in the concentration of growth factors up to 60 ng/mL of β-FGF/EGF and 30 ng/mL of activin A/β-cellulin increases the percentage of IPCs. Further increase of growth factors does not show any increase of the percentage of differentiated cells. Our findings suggest that the presented protocol can be adapted for differentiation of insulin producing cells from stem cells. PMID:25405207

  16. A modified method of insulin producing cells' generation from bone marrow-derived mesenchymal stem cells.

    PubMed

    Czubak, Paweł; Bojarska-Junak, Agnieszka; Tabarkiewicz, Jacek; Putowski, Lechosław

    2014-01-01

    Type 1 diabetes mellitus is a result of autoimmune destruction of pancreatic insulin producing β-cells and so far it can be cured only by insulin injection, by pancreas transplantation, or by pancreatic islet cells' transplantation. The methods are, however, imperfect and have a lot of disadvantages. Therefore new solutions are needed. The best one would be the use of differentiated mesenchymal stem cells (MSCs). In the present study, we investigated the potential of the bone marrow-derived MSCs line for in vitro differentiation into insulin producing cells (IPSs). We applied an 18-day protocol to differentiate MSCs. Differentiating cells formed cell clusters some of which resembled pancreatic islet-like cells. Using dithizone we confirmed the presence of insulin in the cells. What is more, the expression of proinsulin C-peptide in differentiated IPCs was analyzed by flow cytometry. For the first time, we investigated the influence of growth factors' concentration on IPCs differentiation efficiency. We have found that an increase in the concentration of growth factors up to 60 ng/mL of β-FGF/EGF and 30 ng/mL of activin A/β-cellulin increases the percentage of IPCs. Further increase of growth factors does not show any increase of the percentage of differentiated cells. Our findings suggest that the presented protocol can be adapted for differentiation of insulin producing cells from stem cells. PMID:25405207

  17. A modified method of insulin producing cells' generation from bone marrow-derived mesenchymal stem cells.

    PubMed

    Czubak, Paweł; Bojarska-Junak, Agnieszka; Tabarkiewicz, Jacek; Putowski, Lechosław

    2014-01-01

    Type 1 diabetes mellitus is a result of autoimmune destruction of pancreatic insulin producing β-cells and so far it can be cured only by insulin injection, by pancreas transplantation, or by pancreatic islet cells' transplantation. The methods are, however, imperfect and have a lot of disadvantages. Therefore new solutions are needed. The best one would be the use of differentiated mesenchymal stem cells (MSCs). In the present study, we investigated the potential of the bone marrow-derived MSCs line for in vitro differentiation into insulin producing cells (IPSs). We applied an 18-day protocol to differentiate MSCs. Differentiating cells formed cell clusters some of which resembled pancreatic islet-like cells. Using dithizone we confirmed the presence of insulin in the cells. What is more, the expression of proinsulin C-peptide in differentiated IPCs was analyzed by flow cytometry. For the first time, we investigated the influence of growth factors' concentration on IPCs differentiation efficiency. We have found that an increase in the concentration of growth factors up to 60 ng/mL of β-FGF/EGF and 30 ng/mL of activin A/β-cellulin increases the percentage of IPCs. Further increase of growth factors does not show any increase of the percentage of differentiated cells. Our findings suggest that the presented protocol can be adapted for differentiation of insulin producing cells from stem cells.

  18. Engineering antigen-specific T cells from genetically modified human hematopoietic stem cells in immunodeficient mice.

    PubMed

    Kitchen, Scott G; Bennett, Michael; Galić, Zoran; Kim, Joanne; Xu, Qing; Young, Alan; Lieberman, Alexis; Joseph, Aviva; Goldstein, Harris; Ng, Hwee; Yang, Otto; Zack, Jerome A

    2009-01-01

    There is a desperate need for effective therapies to fight chronic viral infections. The immune response is normally fastidious at controlling the majority of viral infections and a therapeutic strategy aimed at reestablishing immune control represents a potentially powerful approach towards treating persistent viral infections. We examined the potential of genetically programming human hematopoietic stem cells to generate mature CD8+ cytotoxic T lymphocytes that express a molecularly cloned, "transgenic" human anti-HIV T cell receptor (TCR). Anti-HIV TCR transduction of human hematopoietic stem cells directed the maturation of a large population of polyfunctional, HIV-specific CD8+ cells capable of recognizing and killing viral antigen-presenting cells. Thus, through this proof-of-concept we propose that genetic engineering of human hematopoietic stem cells will allow the tailoring of effector T cell responses to fight HIV infection or other diseases that are characterized by the loss of immune control.

  19. 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...

  20. Plasmons in metallic nanospheres: Towards efficiency enhancement of metallic nano-modified solar cells

    NASA Astrophysics Data System (ADS)

    Jacak, W.; Krasnyj, J.; Jacak, J.; Jacak, L.

    2011-07-01

    An explanation of a large plasmon-induced PV efficiency enhancement of metallically surface-modified photo-cell is given by inclusion of all indirect inter-band electron transitions in semiconductor due to near-field coupling with plasmon radiation of a nano-scale metallic components. The model of nanosphere plasmon is formulated (of RPA-type, adjusted to large clusters) for both surface and volume modes. Damping of plasmons is analyzed including irradiation losses due to the Lorentz friction. Probability of the interband transition in substrate semiconductor caused by the coupling with plasmons in near-field regime turns out to be larger by 4-orders (in an idealised atomic regime) than for coupling of electrons with planar-wave photons. Inclusion of proximity and interference effects allows for explanation of photo-current growth measured in experimental metallically modified photo-diode systems.

  1. Corrosive characteristics of surface-modified stainless steel bipolar plate in solid polymer fuel cell

    NASA Astrophysics Data System (ADS)

    Zhang, Xiaowen; Wang, Lixia; Sun, Juncai

    2015-03-01

    In this paper, corrosion behavior of an AISI 304 stainless steel modified by niobium or niobium nitride (denoted as niobized 304 SS and Nb-N 304 SS, respectively) is investigated in simulated solid polymer fuel cell (SPFC) operating conditions. Potentiodynamic polarizations show that the corrosion potentials of surface modified 304 SS shift to positive direction while the corrosion current densities decrease greatly comparing with the bare 304 SS in simulated anodic SPFC environments. The order of corrosive resistance in corrosive potential, corrosive current density and pitting potential is: Nb-N 304 SS > niobized 304 SS > bare 304 SS. In the methanol-fueled SPFC operating conditions, the results show that the corrosion resistance of bare and niobized 304 SS increases with the methanol concentration increasing in the test solutions.

  2. Interleukin-2 gene-modified allogeneic tumor cells for treatment of relapsed neuroblastoma.

    PubMed

    Bowman, L C; Grossmann, M; Rill, D; Brown, M; Zhong, W Y; Alexander, B; Leimig, T; Coustan-Smith, E; Campana, D; Jenkins, J; Woods, D; Brenner, M

    1998-06-10

    Tumor cells that have been genetically modified to express immunostimulatory genes will induce effective antitumor responses in a range of syngeneic animal models. For human applications, transduced autologous tumor cell lines are often difficult or impossible to prepare, so that there are strong incentives for substituting a standardized allogeneic tumor cell line. However, such lines may be inferior immunogens if they differ from host tumors in the antigens they express. We have evaluated the safety, immunostimulatory, and antitumor activity of an interleukin-2-secreting allogeneic neuroblastoma cell line in 12 children with relapsed stage IV neuroblastoma. They received two to four subcutaneous injections of cells in a dose-escalating schedule, up to a maximum of 10(8) cells per injection. There was induration and pruritus at the injection site, and skin biopsies revealed mild panniculitis with CD3+ cells surrounding scanty residual tumor cells. There was a limited but significant peripheral monocytosis. No patient showed any increase in direct cytotoxic effector function against the immunizing cell line, but 3 patients had a rise in the frequency of neuroblastoma-reactive cytotoxic T lymphocyte precursor cells. One child had > 90% tumor response (PR), 7 had stable disease, and 4 had progressive disease in response to vaccine alone. Although these results offer some encouragement for the continued pursuit of allogeneic vaccine strategies in human cancer, the antitumor immune responses we observed are inferior to those obtained in an earlier immunization study using autologous neuroblastoma cells. Hence, we suggest that this earlier approach remains preferable, its difficulties notwithstanding.

  3. Modifying akt signaling in B-cell chronic lymphocytic leukemia cells.

    PubMed

    Hofbauer, Sebastian W; Piñón, Josefina D; Brachtl, Gabriele; Haginger, Lucia; Wang, Wei; Jöhrer, Karin; Tinhofer, Ingeborg; Hartmann, Tanja Nicole; Greil, Richard

    2010-09-15

    Emerging evidence suggests that the survival of B-cell chronic lymphocytic leukemia (CLL) cells is dependent on microenvironmental influences such as antigenic stimulation and support by stromal cells. Akt, also known as protein kinase B, is a central component in prosurvival signaling downstream of these events. We investigated the role of Akt and its modulation by the protooncogene T-cell leukemia 1a (Tcl1a) in the survival pathways of primary CLL samples and CLL-derived prolymphocytic cell lines MEC-1 and MEC-2. Akt activation was increased by the protective presence of human bone marrow stromal cells and B-cell receptor mimicking signals but antagonized by direct Akt blockade with the novel specific inhibitor AiX, with preferential apoptosis induction in CLL cells with an unmutated immunoglobulin status, which predicts poor clinical outcome. In addition, we found a direct interaction of Akt with Tcl1a in an endogenous coimmunoprecipitation assay. Confirming the critical role of Tcl1a in modulating Akt signaling, Akt activation was enhanced by overexpressing Tcl1a in CLL. In contrast, decreasing Tcl1a levels by small interfering RNA reduced Akt activation in the fludarabine-insensitive CLL cell line MEC-2 and sensitized the malignant cells to fludarabine treatment. In summary, our data reveal a significant role for the Akt-Tcl1a axis in CLL survival and propose a further evaluation of this interplay for targeting chemoresistance phenomena.

  4. Oncolytic virus expressing RANTES and IL-15 enhances function of CAR-modified T cells in solid tumors

    PubMed Central

    Nishio, Nobuhiro; Dotti, Gianpietro

    2015-01-01

    We improved the migration and survival of chimeric antigen receptor (CAR)-modified T cells in solid tumors by combining CAR-T cells with an armed oncolytic virus. Local delivery of the chemokine RANTES and the cytokine IL-15 by the oncolytic virus enhanced the trafficking and persistence of the CAR-T cells, resulting in improved antitumor effects. PMID:25949885

  5. Mechanism of poly-l-lysine-modified iron oxide nanoparticles uptake into cells.

    PubMed

    Li, Zheng; Shuai, Cijun; Li, Xiayu; Li, Xiaoling; Xiang, Juanjuan; Li, Guiyuan

    2013-10-01

    Poly-l-lysine-modified iron oxide nanoparticle (IONP-PLL), which is formed by modifying poly- l-lysine to the surface of iron oxide nanoparticles, can deliver exogenous genes to cells in vitro and in vivo. However, there is relatively little information available about how is IONP-PLL uptaken by cells. In this study, we are focusing on the transferrin receptor (TFR) mediated and TFR-independent cellular internalization of IONP-PLL. The cells were incubated with 1 µM of IONP-PLL with or without transferrin bound. Transferrin-TFR pathway blockers, such as NH4 Cl, CH3 NH2 , or trypsin, were added to the media and their effects were observed. Atomic absorption spectrophotometer was used to quantify the cellular concentration of iron. The cellular concentrations of iron were evaluated at 37°C or 4°C. (1) Transferrin-IONP-PLL uptake into cells was reliant on time and temperature. (2) The addition of blockers, either NH4 CL, CH3 NH2 , or trypsin, decreased the cellular transferrin-dependent IONP-PLL uptake, but not completely blocked the entry of IONP-PLL. (3) When the cells were culture at pH 6.5, under conditions which the binding of iron and transferrin were inhibited, IONP-PLL still had the capacity to enter into cells with time and temperature-dependent manner. These results suggest that the cellular internalization of IONP-PLL, much like iron ion, were mediated by TFR-dependent endocytosis and TFR-free uptake.

  6. T cell epitopes and post-translationally modified epitopes in type 1 diabetes.

    PubMed

    McGinty, John W; Marré, Meghan L; Bajzik, Veronique; Piganelli, Jon D; James, Eddie A

    2015-11-01

    Type 1 diabetes (T1D) is an autoimmune disease in which progressive loss of self-tolerance, evidenced by accumulation of auto-antibodies and auto-reactive T cells that recognize diverse self-proteins, leads to immune-mediated destruction of pancreatic beta cells and loss of insulin secretion. In this review, we discuss antigens and epitopes in T1D and the role that post-translational modifications play in circumventing tolerance mechanisms and increasing antigenic diversity. Emerging data suggest that, analogous to other autoimmune diseases such as rheumatoid arthritis and celiac disease, enzymatically modified epitopes are preferentially recognized in T1D. Modifying enzymes such as peptidyl deiminases and tissue transglutaminase are activated in response to beta cell stress, providing a mechanistic link between post-translational modification and interactions with the environment. Although studies of such responses in the at-risk population have been limited, current data suggests that breakdown in tolerance through post-translational modification represents an important checkpoint in the development of T1D. PMID:26370701

  7. Somatosensory inputs modify auditory spike timing in dorsal cochlear nucleus principal cells

    PubMed Central

    Koehler, Seth D; Pradhan, Shashwati; Manis, Paul B; Shore, Susan E

    2010-01-01

    In addition to auditory inputs, dorsal cochlear nucleus (DCN) pyramidal cells in the guinea pig receive and respond to somatosensory inputs and perform multisensory integration. DCN pyramidal cells respond to sounds with characteristic spike-timing patterns that are partially controlled by rapidly inactivating potassium conductances. Deactivating these conductances can modify both spike rate and spike timing of responses to sound. Somatosensory pathways are known to modify response rates to subsequent acoustic stimuli, but their effect on spike timing is unknown. Here, we demonstrate that preceding tonal stimulation with spinal trigeminal nucleus (Sp5) stimulation significantly alters the first spike latency, the first interspike interval, and the average discharge regularity of firing evoked by the tone. These effects occur whether the neuron is excited or inhibited by Sp5 stimulation alone. Our results demonstrate that multisensory integration in DCN alters spike-timing representations of acoustic stimuli in pyramidal cells. These changes likely occur through synaptic modulation of intrinsic excitability or synaptic inhibition. PMID:21198989

  8. 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.

  9. Differentiation of PDX1 gene-modified human umbilical cord mesenchymal stem cells into insulin-producing cells in vitro.

    PubMed

    He, Dongmei; Wang, Juan; Gao, Yangjun; Zhang, Yuan

    2011-12-01

    Mesenchymal stem cells (MSCs) have significant advantages over other stem cell types, and greater potential for immediate clinical application. MSCs would be an interesting cellular source for treatment of type 1 diabetes. In this study, MSCs from human umbilical cord were differentiated into functional insulin-producing cells in vitro by introduction of the pancreatic and duodenal homeobox factor 1 (PDX1) and in the presence of induction factors. The expressions of cell surface antigens were detected by flow cytometry. After induction in an adipogenic medium or an osteogenic medium, the cells were observed by Oil Red O staining and alkaline phosphatase staining. Recombinant adenovirus carrying the PDX1 gene was constructed and MSCs were infected by the recombinant adenovirus, then treated with several inducing factors for differentiation into islet β-like cells. The expression of the genes and protein related to islet β-cells was detected by immunocytochemistry, RT-PCR and Western blot analysis. Insulin and C-peptide secretion were assayed. Our results show that the morphology and immunophenotype of MSCs from human umbilical cord were similar to those present in human bone marrow. The MSCs could be induced to differentiate into osteocytes and adipocytes. After induction by recombined adenovirus vector with induction factors, MSCs were aggregated and presented islet-like bodies. Dithizone staining of these cells was positive. The genes' expression related to islet β-cells was found. After induction, insulin and C-peptide secretion in the supernatant were significantly increased. In conclusion, our results demonstrated that PDX1 gene-modified human umbilical cord mesenchymal stem cells could be differentiated into insulin-producing cells in vitro. PMID:21837359

  10. RA induces the neural-like cells generated from epigenetic modified NIH/3T3 cells.

    PubMed

    Zhang, Xi-Mei; Li, Qiu-Ming; Su, Dong-Ju; Wang, Ning; Shan, Zhi-Yan; Jin, Lian-Hong; Lei, Lei

    2010-03-01

    Recently, differentiated somatic cells had been reprogrammed to pluripotential state in vitro, and various tissue cells had been elicited from those cells. Epigenetic modifications allow differentiated cells to perpetuate the molecular memory needed for the cells to retain their identity. DNA methylation and histone deacetylation are important patterns involved in epigenetic modification, which take critical roles in regulating DNA expression. In this study, we dedifferentiated NIH/3T3 fibroblasts by 5-aza-2-deoxycytidine (5-aza-dC) and Trichstatin A (TSA) combination, and detected gene expression pattern, DNA methylation level, and differentiation potential of reprogrammed cells. As the results, embryonic marker Sox2, klf4, c-Myc and Oct4 were expressed in reprogrammed NIH/3T3 fibroblasts. Total DNA methylation level was significant decreased after the treatment. Moreover, exposure of the reprogrammed cells to all trans-retinoic acid (RA) medium elicited the generation of neuronal class IIIbeta-tubulin-positive, neuron-specific enolase (NSE)-positive, nestin-positive, and neurofilament light chain (NF-L)-positive neural-like cells. PMID:19263240

  11. Modifying an Insect Cell N-Glycan Processing Pathway Using CRISPR-Cas Technology.

    PubMed

    Mabashi-Asazuma, Hideaki; Kuo, Chu-Wei; Khoo, Kay-Hooi; Jarvis, Donald L

    2015-10-16

    Fused lobes (FDL) is an enzyme that simultaneously catalyzes a key trimming reaction and antagonizes elongation reactions in the insect N-glycan processing pathway. Accordingly, FDL function accounts, at least in part, for major differences in the N-glycosylation patterns of glycoproteins produced by insect and mammalian cells. In this study, we used the CRISPR-Cas9 system to edit the fdl gene in Drosophila melanogaster S2 cells. CRISPR-Cas9 editing produced a high frequency of site-specific nucleotide insertions and deletions, reduced the production of insect-type, paucimannosidic products (Man3GlcNAc2), and led to the production of partially elongated, mammalian-type complex N-glycans (GlcNAc2Man3GlcNAc2) in S2 cells. As CRISPR-Cas9 has not been widely used to analyze or modify protein glycosylation pathways or edit insect cell genes, these results underscore its broad utility as a tool for these purposes. Our results also confirm the key role of FDL at the major branch point distinguishing insect and mammalian N-glycan processing pathways. Finally, the new FDL-deficient S2 cell derivative produced in this study will enable future bottom-up glycoengineering efforts designed to isolate insect cell lines that can efficiently produce recombinant glycoproteins with chemically predefined oligosaccharide side-chain structures. PMID:26241388

  12. 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. PMID:25953566

  13. Heck products of parthenolide and melampomagnolide-B as anticancer modulators that modify cell cycle progression

    PubMed Central

    Penthala, Narsimha R.; Bommagani, Shobanbabu; Janganati, Venumadhav; MacNicol, Kenzie B.; Cragle, Chad E.; Madadi, Nikhil R.; Hardy, Linda L.; MacNicol, Angus M.; Crooks, Peter A.

    2014-01-01

    (E)-13-(Aryl/heteroaryl)parthenolides (5a–i and 6a–i) were synthesized and evaluated for their ability to modify cell cycle progression during progesterone-stimulated Xenopus oocyte maturation and screened for their anticancer activity against a panel of 60 human cancer cell lines. (E)-13-(4-aminophenyl) parthenolide (5b) caused a significant inhibition of progesterone-stimulated oocyte maturation, and was determined to function downstream of MAP kinase signaling, but upstream of the activation of the universal G2/M regulator, M-phase promoting factor (MPF, cyclin B/Cyclin-dependent kinase (CDK). The compound (E)-13-(2-bromo-phenyl)parthenolide (5c) activates oocyte maturation independently of progesterone stimulation. Compounds 5b and 5c displayed modest growth inhibition on select cancer cell lines at 10 micromolar dose when tested on the panel of 60 cancer cell lines. By contrast, compounds (5f and 7) did not modulate oocyte maturation but did exhibit micromolar level growth inhibition against most of the human cancer cell lines over a range of doses. Together, our findings indicate that screening of compounds in the oocyte maturation assay may identify additional effective cell cycle regulatory compounds that do not necessarily exert overt cytotoxicity as assessed in traditional drug screening assays. PMID:25117652

  14. Modifying an Insect Cell N-Glycan Processing Pathway Using CRISPR-Cas Technology.

    PubMed

    Mabashi-Asazuma, Hideaki; Kuo, Chu-Wei; Khoo, Kay-Hooi; Jarvis, Donald L

    2015-10-16

    Fused lobes (FDL) is an enzyme that simultaneously catalyzes a key trimming reaction and antagonizes elongation reactions in the insect N-glycan processing pathway. Accordingly, FDL function accounts, at least in part, for major differences in the N-glycosylation patterns of glycoproteins produced by insect and mammalian cells. In this study, we used the CRISPR-Cas9 system to edit the fdl gene in Drosophila melanogaster S2 cells. CRISPR-Cas9 editing produced a high frequency of site-specific nucleotide insertions and deletions, reduced the production of insect-type, paucimannosidic products (Man3GlcNAc2), and led to the production of partially elongated, mammalian-type complex N-glycans (GlcNAc2Man3GlcNAc2) in S2 cells. As CRISPR-Cas9 has not been widely used to analyze or modify protein glycosylation pathways or edit insect cell genes, these results underscore its broad utility as a tool for these purposes. Our results also confirm the key role of FDL at the major branch point distinguishing insect and mammalian N-glycan processing pathways. Finally, the new FDL-deficient S2 cell derivative produced in this study will enable future bottom-up glycoengineering efforts designed to isolate insect cell lines that can efficiently produce recombinant glycoproteins with chemically predefined oligosaccharide side-chain structures.

  15. 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.

  16. Heck products of parthenolide and melampomagnolide-B as anticancer modulators that modify cell cycle progression.

    PubMed

    Penthala, Narsimha R; Bommagani, Shobanbabu; Janganati, Venumadhav; MacNicol, Kenzie B; Cragle, Chad E; Madadi, Nikhil R; Hardy, Linda L; MacNicol, Angus M; Crooks, Peter A

    2014-10-01

    (E)-13-(Aryl/heteroaryl)parthenolides (5a-i and 6a-i) were synthesized and evaluated for their ability to modify cell cycle progression during progesterone-stimulated Xenopus oocyte maturation and screened for their anticancer activity against a panel of 60 human cancer cell lines. (E)-13-(4-aminophenyl) parthenolide (5b) caused a significant inhibition of progesterone-stimulated oocyte maturation, and was determined to function downstream of MAP kinase signaling, but upstream of the activation of the universal G2/M regulator, M-phase promoting factor (MPF), cyclin B/Cyclin-dependent kinase (CDK). The compound (E)-13-(2-bromo-phenyl)parthenolide (5c) activates oocyte maturation independently of progesterone stimulation. Compounds 5b and 5c displayed modest growth inhibition on select cancer cell lines at 10 μM dose when tested on the panel of 60 cancer cell lines. By contrast, compounds (5f and 7) did not modulate oocyte maturation but did exhibit micromolar level growth inhibition against most of the human cancer cell lines over a range of doses. Together, our findings indicate that screening of compounds in the oocyte maturation assay may identify additional effective cell cycle regulatory compounds that do not necessarily exert overt cytotoxicity as assessed in traditional drug screening assays.

  17. Accelerated cell-surface interlocking on plasma polymer-modified porous ceramics.

    PubMed

    Rebl, Henrike; Finke, Birgit; Schmidt, Jürgen; Mohamad, Heba S; Ihrke, Roland; Helm, Christiane A; Nebe, J Barbara

    2016-12-01

    Excellent osseointegration of permanent implants is crucial for the long lasting success of the implantation. To improve the osseointegrative potential, bio-inert titanium alloy surfaces (Ti6Al4V) are modified by plasma chemical oxidation (PCO®) of the titanium-oxide layer to a non-stoichiometric, amorphous calcium phosphate layer. The native titanium-oxide film measuring only a few nanometers is converted by PCO® to a thick porous calcium phosphate layer of about 10μm. In a second step the PCO surface is combined with a cell adhesive plasma-polymerized allylamine (PPAAm) nano film (5 and 50nm). Independent of the PPAAm coating homogeneity, the human osteoblast-like MG-63 cells show a remarkable increase in cell size and well-developed filopodia. Analyses of the actin cytoskeleton reveal that the cells mold to the pore shape of the PPAAm-covered PCO, thereby establishing a strong attachment to the surface. Interestingly, we could demonstrate that even though our untreated PCO shows excellent hydrophilicity, this alone is not sufficient to facilitate fast cell spreading, but the positive surface charges mediated by PPAAm. This multilayer composite material guarantees enhanced interlocking of the cells with the porous surface.

  18. Accelerated cell-surface interlocking on plasma polymer-modified porous ceramics.

    PubMed

    Rebl, Henrike; Finke, Birgit; Schmidt, Jürgen; Mohamad, Heba S; Ihrke, Roland; Helm, Christiane A; Nebe, J Barbara

    2016-12-01

    Excellent osseointegration of permanent implants is crucial for the long lasting success of the implantation. To improve the osseointegrative potential, bio-inert titanium alloy surfaces (Ti6Al4V) are modified by plasma chemical oxidation (PCO®) of the titanium-oxide layer to a non-stoichiometric, amorphous calcium phosphate layer. The native titanium-oxide film measuring only a few nanometers is converted by PCO® to a thick porous calcium phosphate layer of about 10μm. In a second step the PCO surface is combined with a cell adhesive plasma-polymerized allylamine (PPAAm) nano film (5 and 50nm). Independent of the PPAAm coating homogeneity, the human osteoblast-like MG-63 cells show a remarkable increase in cell size and well-developed filopodia. Analyses of the actin cytoskeleton reveal that the cells mold to the pore shape of the PPAAm-covered PCO, thereby establishing a strong attachment to the surface. Interestingly, we could demonstrate that even though our untreated PCO shows excellent hydrophilicity, this alone is not sufficient to facilitate fast cell spreading, but the positive surface charges mediated by PPAAm. This multilayer composite material guarantees enhanced interlocking of the cells with the porous surface. PMID:27612809

  19. Osteoblast cell response to a CO 2 laser modified polymeric material

    NASA Astrophysics Data System (ADS)

    Waugh, D. G.; Lawrence, J.; Brown, E. M.

    2012-02-01

    Lasers are an efficient technology, which can be applied for the surface treatment of polymeric biomaterials to enhance insufficient surface properties. That is, the surface chemistry and topography of biomaterials can be modulated to increase the biofunctionality of that material. By employing CO 2 laser patterning and whole area processing of nylon 6,6 this paper details how the surface properties were significantly modified. Samples, which had undergone whole area processing, followed the current theory in which the advancing contact angle, θ, with water decreased and the polar component, γp, increased upon an increase in surface roughness. For the patterned samples it was observed that θ increased and γP decreased. This did not follow the current theory and can be explained by a mixed-state wetting regime. By seeding osteoblast cells onto the samples for 24 h and 4 days the laser surface treatment gave rise to modulated cell response. For the laser whole area processing, θ and γP correlated with the observed cell count and cover density. Owed to the wetting regime, the patterned samples did not give rise to any correlative trend. As a result, CO 2 laser whole area processing is more likely to allow one to predict biofunctionality prior to cell seeding. Moreover, for all samples, cell differentiation was evidenced. On account of this and the modulation in cell response, it has been shown that laser surface treatment lends itself to changing the biofunctional properties of nylon 6,6.

  20. 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.

  1. Modulation of chromatin modifying factors' gene expression in embryonic and induced pluripotent stem cells.

    PubMed

    Luzzani, Carlos; Solari, Claudia; Losino, Noelia; Ariel, Waisman; Romorini, Leonardo; Bluguermann, Carolina; Sevlever, Gustavo; Barañao, Lino; Miriuka, Santiago; Guberman, Alejandra

    2011-07-15

    Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) are a promising source of cells for regenerative medicine because of their potential of self renew and differentiation. Multiple evidences highlight the relationship of chromatin remodeling with stem cell properties, differentiation programs and reprogramming for iPSC obtention. With the purpose of finding chromatin modifying factors relevant to these processes, and based on ChIP on chip studies, we selected several genes that could be modulated by Oct4, Sox2 and Nanog, critical transcription factors in stem cells, and studied their expression profile along the differentiation in mouse and human ESCs, and in mouse iPSCs. In this work, we analyzed the expression of Gcn5l2, GTF3C3, TAF15, ATF7IP, Myst2, HDAC2, HDAC3, HDAC5, HDAC10, SUV39H2, Jarid2, and Bmi-1. We found some genes from different functional groups that were highly modulated, suggesting that they could be relevant both in the undifferentiated state and during differentiation. These findings could contribute to the comprehension of molecular mechanisms involved in pluripotency, early differentiation and reprogramming. We believe that a deeper knowledge of the epigenetic regulation of ESC will allow improving somatic cell reprogramming for iPSC obtention and differentiation protocols optimization.

  2. Highly efficient inverted organic solar cells using amino acid modified indium tin oxide as cathode

    SciTech Connect

    Li, Aiyuan; Nie, Riming; Deng, Xianyu; Wei, Huaixin; Li, Yanqing; Tang, Jianxin; Zheng, Shizhao; Wong, King-Young

    2014-03-24

    In this paper, we report that highly efficient inverted organic solar cells were achieved by modifying the surface of indium tin oxide (ITO) using an amino acid, Serine (Ser). With the modification of the ITO surface, device efficiency was significantly enhanced from 0.63% to 4.17%, accompanied with an open circuit voltage (Voc) that was enhanced from 0.30 V to 0.55 V. Ultraviolet and X-ray photoelectron spectroscopy studies indicate that the work function reduction induced by the amino acid modification resulting in the decreased barrier height at the ITO/organic interface played a crucial role in the enhanced performances.

  3. Genetically modified murine adipose-derived mesenchymal stem cells producing interleukin-2 favor B16F10 melanoma cell proliferation.

    PubMed

    Bahrambeigi, Vahid; Ahmadi, Nafiseh; Salehi, Rasoul; Javanmard, Shaghayegh Haghjooy

    2015-01-01

    Adipose-derived mesenchymal stem cells (ADSCs) are attractive tools for cancer gene therapy due to their intrinsic tropism to the tumor environment. Interleukin-2 (IL2) is recognized as a key regulatory molecule, which enhances the activity and growth of the immune effector cell function. High-Dose IL2 Therapy is an option for treatment of malignant melanoma but has frequent, often serious and sometimes life-threatening side effects. Here we investigated the effect of genetically modified ADSCs (GM-ADSCs) expressing IL2 in immunocompetent mouse models of subcutaneous and lung metastatic melanoma. Prior to in vivo studies, we demonstrated that IL2 produced by GM-ADSCs may act as a growth factor for melanoma cells due to the increased viability and reduced apoptosis of melanoma cells after in vitro treatment. Subcutaneous co-injection of IL2-expressing ADSCs with melanoma cells significantly enhanced the melanoma tumor growth. Furthermore, histological analysis of subcutaneous tumors for IL2 and Melan-A (a melanocytic differentiation marker) confirmed that most of cells in melanoma/IL2-ADSC co-injected tumors are melanoma cells, not IL2-ADSCs. In pulmonary metastases model, melanoma cells were injected intravenously and 10 days later mice were treated by systematical injection of GM-ADSCs. Intravenously injected IL2-ADSCs engrafted into melanoma lung tumors but were unable to reduce melanoma lung metastases. Besides, administered IL2-ADSCs significantly reduced systemic CD4+ cells and did not impact the total survival of lung metastases melanoma bearing mice. In conclusion, this study showed that IL2-producing ADSCs can favor B16F10 melanoma cell proliferation. Therefore, therapies utilizing IL2 have to be taken into careful consideration.

  4. Ion Energy and Ion Flux Distributions of CF4/Ar/O2 Inductively Coupled Plasmas in a GEC Cell

    NASA Technical Reports Server (NTRS)

    Rao, M. V. V. S.; Cruden, Brett; Sharma, Surendra; Meyyappan, Meyya

    2001-01-01

    Knowledge of ion kinetics in plasma processing gas mixtures, such as CF4:Ar:O2, is important for understanding plasma assisted etching and deposition of materials. Ion energies and ion fluxes were measured in this mixture for 80:10:10, 60:20:20, and 40:30:30 mixture ratios in the pressure range of 10-50 mTorr, and at 200 and 300 W of RF power. Ions from plasma, sampled through a 10 micron orifice in the center of the lower plane electrode, were energy and mass analyzed by a combination of electrostatic energy and quadrupole mass filters. CFx(+) (x = 1 - 3), F2(+), F(+), C(+) from CF4, Ar(+) from Ar, and O2(+) and O(+) from O2, and by-product ions SiFx(+)(x = 1 - 3) from etching of quartz coupling window, COFx(+)(x = 1 - 3), CO(+), CO2(+), and OF(+) were detected. In all conditions ion flux decreases with increase of pressure but increase with increase of RF power. Ar(+) signal decreases with increase of pressure while CF3(+), which is the dominant ion at all conditions, increases with increase in pressure. The loss mechanism for Ar(+) and increase of CF3(+) is due to large cross section for Ar(+) + CF4 yields Ar + CF3(+) + F. Ion energies, which range from 15-25 eV depending on plasma operating conditions, are nearly Gaussian. By-product ion signals are higher at lower pressures indicating stronger plasma interaction with quartz window.

  5. 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

  6. 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...

  7. Complement modulates the function of the ubiquitin-proteasome system and endoplasmic reticulum-associated degradation in glomerular epithelial cells.

    PubMed

    Kitzler, Thomas M; Papillon, Joan; Guillemette, Julie; Wing, Simon S; Cybulsky, Andrey V

    2012-05-01

    In experimental membranous nephropathy, complement C5b-9 induces sublethal glomerular epithelial cell (GEC) injury and proteinuria. C5b-9 also activates mechanisms that restrict injury or facilitate recovery. The ubiquitin-proteasome system (UPS) selectively degrades damaged or abnormal proteins, while misfolded proteins in the endoplasmic reticulum (ER) undergo ER-associated degradation (ERAD). In this study, we investigated the effect of complement on the UPS and ERAD. We monitored UPS function by transfection of rat GECs with a UPS reporter, GFP(u) (CL1 degron fused with green fluorescent protein). By analogy, CD3δ-yellow fluorescent protein (YFP) was employed as a reporter of ERAD. We demonstrated decreased GFP(u) levels in GECs after incubation with antibody and complement, compared with control. Using C8-deficient serum with or without purified C8, cycloheximide (an inhibitor of protein synthesis), and the proteasome inhibitor, MG132, we confirmed that the decrease of GFP(u) was mediated by C5b-9, and subsequent proteasomal degradation of the reporter. Inhibition of the c-Jun N-terminal kinase attenuated the effect of complement on GFP(u) degradation. Complement, however, increased the level of CD3δ-YFP in GECs, implying an impairment of ERAD, likely due to an overabundance of misfolded proteins in the ER. The overall ubiquitination of proteins was enhanced in complement-treated GECs and in glomeruli of rats with experimental membranous nephropathy, although ubiquitin mRNA was unchanged in GECs. Proteasome inhibition with MG132 increased the cytotoxic effect of complement in GECs. Complement-stimulated UPS function, by accelerating removal of damaged proteins, may be a novel mechanism to limit complement-induced injury.

  8. Magnetically modified polymer electrolyte fuel cells and low temperature effects on polymer electrolyte Nafion

    NASA Astrophysics Data System (ADS)

    Dunwoody, Drew Christian

    Polymer electrolyte fuel cell power systems are a promising technology which may provide clean and efficient electrical power in the future. Though the technology is promising, challenging technical issues must be overcome to make the technology a practical alternative to existing power sources. Fuel cell power systems are often touted as possible replacements for combustion engines. Materials used in these power systems must be able to withstand a wide range of environmental conditions including both extremes of heat and cold. An evaluation of fuel cell materials under these extreme conditions is warranted. Also, the expense of a fuel cell power system with comparable power output to more traditional power sources such as batteries and combustion engines is on the order of seven fold. Reductions in the costs of materials and methods to increase the overall efficiency of the power systems need to be identified. Here, the electrochemical behavior of the polymer electrolyte Nafion is investigated as a function of temperature from room temperature to significantly below the freezing point of water. The type of intercalant and electrolyte solution dictates the temperature at which electrochemical properties change and also impacts polymer integrity. Magnetic modification of electrode surfaces has been shown to enhance electrochemical flux up to 3600% over that of nonmagnetic electrodes. Here, polymer electrolyte fuel cells are tested for enhanced performance over nonmagnetic cells. Under certain conditions, an increase in performance is observed for magnetic modification. Descriptions of the methods used to construct magnetically modified fuel cells and special considerations to heed when operating these cells are included.

  9. 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

  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. Discrete element simulation of dense granular flow in a modified Couette cell.

    SciTech Connect

    Lechman, Jeremy B.; Grest, Gary Stephen

    2005-02-01

    Large-scale three dimensional Discrete Element simulations of granular flow in a modified split-bottom Couette cell for packs of up to 180,000 mono-disperse spheres are presented and compared with experiments. We find that the velocity profiles collapse onto a universal curve not only at the surface but also in the bulk of the pack until slip between layers becomes significant. In agreement with experiment, we find similar relations between the cell geometry and parameters involved in rescaling the velocities at the surface and in the bulk. Likewise, a change in the shape of the shear zone is observed as predicted for tall packs once the center of the shear zone is correctly defined; although the transition does not appear to be first order. Finally, the effect of cohesion is considered as a means to test the theoretical predictions.

  12. [Development and challenge of modified hemoglobins as red blood cell substitutes].

    PubMed

    Lu, Xiu-Ling

    2006-01-01

    The problems of blood shortage and the virus infection risk of blood transfusion have promoted the study of blood substitutes. Modified hemoglobin has become the focus of the challenges research because of its excellent oxygen carrying ability. To overcome the toxicity effect on direct use of purified native hemoglobin, various modification technologies have been developed, including diaspirin cross-linking, glutaraldehyde polymerization, O-raffinose polymerization, polyethylene glycol conjugation, liposome encapsulation and biodegradable polymer encapsulation. Some of the products have been in clinical trials, and one of the products has been approved in a country for clinical use. Research on red blood cell substitutes in China has also developed fast. This paper provides an overview of the history and current status in development of different hemoglobin-based red blood cell substitutes, especially the problems encountered, the challenges faced, and the prospects in future.

  13. Human cDNA clones that modify radiomimetic sensitivity of Ataxia-telangiectasia (Group A) cells

    SciTech Connect

    Ziv, Y.; Bar-Shira, A.; Sartiel, A.

    1995-03-01

    Genes responsible for genetic diseases with increased sensitivity to DNA-damaging agents can be identified using complementation cloning. This strategy is based on in vitro complementation of the cellular sensitivity by gene transfer. Ataxia-telangiectasia (A-T) is a multisystem autosomal recessive disorder involving cellular sensitivity to ionizing radiation and radiomimetic drugs. A-T is genetically heterogeneous, with four complementation groups. We attempted to identify cDNA clones that modify the radiomimetic sensitivity of A-T cells assigned to complementation group [A-T(A)]. The cells were transfected with human cDNA libraries clones in episomal vectors, and various protocols by radiomimetic selection were applied. Thirteen cDNAs rescued from survivor cells were found to confer various degrees of radiomimetic resistance to A-T(A) cells upon repeated introduction, and one of them also partially influenced another feature of the A-T phenotype, radioresistant DNA synthesis. None of the clones mapped to the A-T locus on chromosome 11q22-23. Nine of the clones were derived from known genes, some of which are involved in cellular stress responses. We concluded that a number of different genes, not necessarily associated with A-T, can influence the response of A-T cells to radiomimetic drugs, and hence the complementation cloning approach may be less applicable to A-T than to other diseases involving abnormal processing of DNA damage. 57 refs., 5 figs., 2 tabs.

  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. Surface properties and early murine pre-osteoblastic cell responses of phosphoric acid modified titanium surface

    PubMed Central

    Osathanon, Thanaphum; Sawangmake, Chenphop; Ruangchainicom, Nanticha; Wutikornwipak, Pavitra; Kantukiti, Panisa; Nowwarote, Nunthawan; Pavasant, Prasit

    2015-01-01

    Aims The present study investigated the surface properties and murine pre-osteoblast cell (MC3T3-E1) responses of phosphoric acid (H3PO4) treated commercially pure titanium. Methods Titanium discs were treated with various concentration of H3PO4 (5%, 10%, and 20%; v/v) at 90 °C for 30 min. Surface properties were evaluated by profilometer, contact angle meter, and scanning electron microscopy (SEM) with energy dispersive X-rays. MC3T3-E1 attachment and spreading were evaluated by SEM and phalloidin immunohistochemistry staining. Results Surface roughness and wettability were not statistically difference among all experimental and control groups. Phosphate and oxygen were detected on H3PO4 treated surfaces. At 20 min, cell attachment was significantly higher in 10% and 20% H3PO4 treated groups compared to the control. Cells exhibited orientated-cytoskeleton fibers on 20% H3PO4 modified titanium surface. Though, there was no difference in cell spreading stage among all treatment groups. Conclusion H3PO4 treatment on titanium may influence early cell response, particularly on attachment and spreading. PMID:26937362

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

    SciTech Connect

    Marina, Olga A; Stevenson, Jeffry W

    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.

  17. 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.

  18. 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.

  19. CD4(+) T cells epigenetically modified by oxidative stress cause lupus-like autoimmunity in mice.

    PubMed

    Strickland, Faith M; Li, YePeng; Johnson, Kent; Sun, Zhichao; Richardson, Bruce C

    2015-08-01

    Lupus develops when genetically predisposed people encounter environmental agents such as UV light, silica, infections and cigarette smoke that cause oxidative stress, but how oxidative damage modifies the immune system to cause lupus flares is unknown. We previously showed that oxidizing agents decreased ERK pathway signaling in human T cells, decreased DNA methyltransferase 1 and caused demethylation and overexpression of genes similar to those from patients with active lupus. The current study tested whether oxidant-treated T cells can induce lupus in mice. We adoptively transferred CD4(+) T cells treated in vitro with oxidants hydrogen peroxide or nitric oxide or the demethylating agent 5-azacytidine into syngeneic mice and studied the development and severity of lupus in the recipients. Disease severity was assessed by measuring anti-dsDNA antibodies, proteinuria, hematuria and by histopathology of kidney tissues. The effect of the oxidants on expression of CD40L, CD70, KirL1 and DNMT1 genes and CD40L protein in the treated CD4(+) T cells was assessed by Q-RT-PCR and flow cytometry. H2O2 and ONOO(-) decreased Dnmt1 expression in CD4(+) T cells and caused the upregulation of genes known to be suppressed by DNA methylation in patients with lupus and animal models of SLE. Adoptive transfer of oxidant-treated CD4(+) T cells into syngeneic recipients resulted in the induction of anti-dsDNA antibody and glomerulonephritis. The results show that oxidative stress may contribute to lupus disease by inhibiting ERK pathway signaling in T cells leading to DNA demethylation, upregulation of immune genes and autoreactivity.

  20. 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.

  1. 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...

  2. Glucocorticosteroids modify Langerhans cells to produce TGF-β and expand regulatory T cells.

    PubMed

    Stary, Georg; Klein, Irene; Bauer, Wolfgang; Koszik, Frieder; Reininger, Bärbel; Kohlhofer, Sabine; Gruber, Kristina; Skvara, Hans; Jung, Thomas; Stingl, Georg

    2011-01-01

    Although glucocorticosteroids (GCSs) have been used for many decades in transplantation and (auto)inflammatory diseases, the exact mechanisms responsible for their immunosuppressive properties are not fully understood. The purpose of this study was to characterize the effects of oral GCSs on the cutaneous immune response. We analyzed, by immunofluorescence staining and quantitative RT-PCR, residual skin biopsy material from a clinical study in which we had used oral GCS as positive control for determining the effects of candidate anti-inflammatory compounds on epicutaneous patch tests of Ni-allergic patients. Expectedly, oral GCS treatment led to a reduction of clinical symptoms and infiltrating leukocytes. Notably, we observed increased numbers of dermal FOXP3(+)CD25(+) T cells and epidermal Langerhans cells (LCs) that were associated with upregulated mRNA expression of TGF-β in lesions of GCS-treated Ni-allergic patients. To investigate this phenomenon further, we exposed purified LCs to GCS. They exhibited, in contrast to GCS-nonexposed LCs, 1) a more immature phenotype, 2) higher intracellular amounts of TGF-β, and 3) increased receptor activator for NF-κB expression, conditions that reportedly favor the expansion of regulatory T cells (Tregs). Indeed, we observed an enhancement of functionally suppressive FOXP3(+) T cells when CD3(+) cells were incubated with GCS-pretreated LCs. The expansion of Tregs was inhibited by TGF-β blockage alone, and their suppressive activity was neutralized by a combination of anti-TGF-β and anti-IL-10 Abs. Our data show that systemically applied GCSs endow LCs with Treg-promoting properties and thus shed new light on the mechanisms of GCS-mediated immunosuppression.

  3. Immune-modifying properties of topical vitamin D: Focus on dendritic cells and T cells.

    PubMed

    Gorman, Shelley; Judge, Melinda A; Hart, Prue H

    2010-07-01

    Topical creams containing the active form of vitamin D (1,25-dihydroxyvitamin D3; 1,25(OH)2D3) or analogues of this compound are currently used with some success to treat skin conditions including psoriasis and vitiligo. As well as targeting inflammatory processes in the skin, topical application of 1,25(OH)2D3 also affects the function of immune cells in the skin and draining lymph nodes. Topically applied 1,25(OH)2D3 reduces the number of dendritic cells in the skin, resulting in suppressed immunity and in particular reduced contact hypersensitivity (CHS) responses. Topical 1,25(OH)2D3 may also promote the migration of dendritic cells from the skin to the draining lymph nodes. Skin application of 1,25(OH)2D3 prevented the inflammatory effects of UVB irradiation on lymph node hypertrophy, when cell numbers were examined 4 days after skin treatment. In contrast, when 1,25(OH)2D3 was applied to UVB irradiated skin, there was no reversal in the suppression of CHS responses caused by UVB irradiation. Instead, 1,25(OH)2D3 had an additive effect with UVB to suppress CHS responses to a greater degree than UVB alone. In these studies, 1,25(OH)2D3 was applied to the treated skin of BALB/c mice immediately following UVB irradiation. Finally, topical 1,25(OH)2D3 also enhanced the number and suppressive activity of CD4+CD25+ regulatory T cells in the lymphatic tissue draining skin.

  4. The effect of modified polysialic acid based hydrogels on the adhesion and viability of primary neurons and glial cells.

    PubMed

    Haile, Yohannes; Berski, Silke; Dräger, Gerald; Nobre, Andrè; Stummeyer, Katharina; Gerardy-Schahn, Rita; Grothe, Claudia

    2008-04-01

    In this study we present the enzymatic and biological analysis of polysialic acid (polySia) based hydrogel in terms of its degradation and cytocompatibility. PolySia based hydrogel is completely degradable by endosialidase enzyme which may avoid second surgery after tissue recovery. Viability assay showed that soluble components of polySia hydrogel did not cause any toxic effect on cultured Schwann cells. Moreover, green fluorescence protein transfected neonatal and adult Schwann cells, neural stem cells and dorsal root ganglionic cells (unlabelled) were seeded on polySia hydrogel modified with poly-L-lysine (Pll), poly-L-ornithine-laminin (porn-laminin) or collagen. Water soluble tetrazolium salt assay revealed that modification of the hydrogel significantly improved cell adhesion and viability. These results infer that polySia based scaffolds in combination with cell adhesion molecules and cells genetically modified to express growth factors would potentially be promising alternative in reconstructive therapeutic strategies. PMID:18255143

  5. The effect of modified polysialic acid based hydrogels on the adhesion and viability of primary neurons and glial cells.

    PubMed

    Haile, Yohannes; Berski, Silke; Dräger, Gerald; Nobre, Andrè; Stummeyer, Katharina; Gerardy-Schahn, Rita; Grothe, Claudia

    2008-04-01

    In this study we present the enzymatic and biological analysis of polysialic acid (polySia) based hydrogel in terms of its degradation and cytocompatibility. PolySia based hydrogel is completely degradable by endosialidase enzyme which may avoid second surgery after tissue recovery. Viability assay showed that soluble components of polySia hydrogel did not cause any toxic effect on cultured Schwann cells. Moreover, green fluorescence protein transfected neonatal and adult Schwann cells, neural stem cells and dorsal root ganglionic cells (unlabelled) were seeded on polySia hydrogel modified with poly-L-lysine (Pll), poly-L-ornithine-laminin (porn-laminin) or collagen. Water soluble tetrazolium salt assay revealed that modification of the hydrogel significantly improved cell adhesion and viability. These results infer that polySia based scaffolds in combination with cell adhesion molecules and cells genetically modified to express growth factors would potentially be promising alternative in reconstructive therapeutic strategies.

  6. 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

  7. High level protein expression in mammalian cells using a safe viral vector: modified vaccinia virus Ankara.

    PubMed

    Hebben, Matthias; Brants, Jan; Birck, Catherine; Samama, Jean-Pierre; Wasylyk, Bohdan; Spehner, Danièle; Pradeau, Karine; Domi, Arban; Moss, Bernard; Schultz, Patrick; Drillien, Robert

    2007-12-01

    Vaccinia virus vectors are attractive tools to direct high level protein synthesis in mammalian cells. In one of the most efficient strategies developed so far, the gene to be expressed is positioned downstream of a bacteriophage T7 promoter within the vaccinia genome and transcribed by the T7 RNA polymerase, also encoded by the vaccinia virus genome. Tight regulation of transcription and efficient translation are ensured by control elements of the Escherichia coli lactose operon and the encephalomyocarditis virus leader sequence, respectively. We have integrated such a stringently controlled expression system, previously used successfully in a standard vaccinia virus backbone, into the modified vaccinia virus Ankara strain (MVA). In this manner, proteins of interest can be produced in mammalian cells under standard laboratory conditions because of the inherent safety of the MVA strain. Using this system for expression of beta-galactosidase, about 15 mg protein could be produced from 10(8) BHK21 cells over a 24-h period, a value 4-fold higher than the amount produced from an identical expression system based on a standard vaccinia virus strain. In another application, we employed the MVA vector to produce human tubulin tyrosine ligase and demonstrate that this protein becomes a major cellular protein upon induction conditions and displays its characteristic enzymatic activity. The MVA vector should prove useful for many other applications in which mammalian cells are required for protein production. PMID:17892951

  8. Crosslinking Liposomes/Cells Using Cholesteryl Group-Modified Tilapia Gelatin

    PubMed Central

    Taguchi, Tetsushi; Endo, Yoshiaki

    2014-01-01

    Cholesteryl group-modified tilapia gelatins (Chol-T-Gltns) with various Chol contents from 3 to 69 mol % per amino group of Gltn were prepared for the assembly of liposomes and cells. Liposomes were physically crosslinked by anchoring Chol groups of Chol-T-Gltns into lipid membranes. The resulting liposome gels were enzymatically degraded by addition of collagenase. Liposome gels prepared using Chol-T-Gltn with high Chol content (69Chol-T-Gltn) showed slower enzymatic degradation when compared with gels prepared using Chol-T-Gltn with low Chol content (3Chol-T-Gltn). The hepatocyte cell line HepG2 showed good assembly properties and no cytotoxic effects after addition of 69Chol-T-Gltns. In addition, the number of HepG2 cells increased with concentration of 69Chol-T-Gltns. Therefore, Chol-T-Gltn, particularly, 69Chol-T-Gltn, can be used as an assembling material for liposomes and various cell types. The resulting organization can be applied to various biomedical fields, such as drug delivery systems, tissue engineering and regenerative medicine. PMID:25056548

  9. 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.

  10. Interview: bioreactors and surfaced-modified 3D-scaffolds for stem cell research.

    PubMed

    Weibezahn, Karl-Friedrich

    2008-01-01

    A Nature Editorial in 2003 asked the question "Good-bye, flat biology?" What does this question imply? In the past, many in vitro culture systems, mainly monolayer cultures, often suffered from the disadvantage that differentiated primary cells had a relatively short life-span and de-differentiated during culture. As a consequence, most of their organ-specific functions were lost rapidly. Thus, in order to reproduce better conditions for these cells in vitro, modifications and adaptations have been made to conventional monolayer cultures. The last generation of CellChips--micro-thermoformed containers--a specific technology was developed, which offers the additional possibility to modify the whole surface of the 3D formed containers. This allows a surface-patterning on a submicron scale with distinct signalling molecules. Sensors and signal electrodes may be incorporated. Applications range from basic research in cell biology to toxicology and pharmacology. Using biodegradable polymers, clinical applications become a possibility. Furthermore, the last generation of micro-thermoformed chips has been optimized to allow for cheap mass production.

  11. Interview: Bioreactors and Surfaced-Modified 3D-Scaffolds for Stem Cell Research

    PubMed Central

    Weibezahn, Karl-Friedrich

    2008-01-01

    A Nature Editorial in 2003 asked the question "Good-bye, flat biology?" What does this question imply? In the past, many in vitro culture systems, mainly monolayer cultures, often suffered from the disadvantage that differentiated primary cells had a relatively short life-span and de-differentiated during culture. As a consequence, most of their organ-specific functions were lost rapidly. Thus, in order to reproduce better conditions for these cells in vitro, modifications and adaptations have been made to conventional monolayer cultures. The last generation of CellChips -- micro-thermoformed containers -- a specific technology was developed, which offers the additional possibility to modify the whole surface of the 3D formed containers. This allows a surface-patterning on a submicron scale with distinct signalling molecules. Sensors and signal electrodes may be incorporated. Applications range from basic research in cell biology to toxicology and pharmacology. Using biodegradable polymers, clinical applications become a possibility. Furthermore, the last generation of micro-thermoformed chips has been optimized to allow for cheap mass production. PMID:19066581

  12. Angiopep-2 and activatable cell penetrating peptide dual modified nanoparticles for enhanced tumor targeting and penetrating.

    PubMed

    Mei, Ling; Zhang, Qianyu; Yang, Yuting; He, Qin; Gao, Huile

    2014-10-20

    Delivering chemotherapeutics by nanoparticles into tumor was influenced by at least two factors: specific targeting and highly efficient penetrating of the nanoparticles. In this study, two targeting ligands, angiopep-2 and activatable cell penetrating peptide (ACP), were functionalized onto nanoparticles for tumor targeting delivery. In this system, angiopep-2 is a ligand of low-density lipoprotein receptor-related protein-1 (LRP1) which was highly expressed on tumor cells, and the ACP was constructed by the conjugation of RRRRRRRR (R8) with EEEEEEEE through a matrix metalloproteinase-2 (MMP-2) sensitive linker, enabling the ACP with tumor microenvironment-responsive cell penetrating property. 4h incubation of ACP with MMP-2 leads to over 80% cleavage of ACP, demonstrating ACP indeed possessed MMP-2 responsive property. The constructed dual targeting nanoparticles (AnACNPs) were approximately 110 nm with a polydispersity index of 0.231. In vitro, ACP modification and angiopep-2 modification could both enhance the U-87 MG cell uptake because of the high expression of MMP-2 and LRP-1 on C6 cells. AnACNPs showed higher uptake level than the single ligand modified nanoparticles. The uptake of all particles was time- and concentration-dependent and endosomes were involved. In vivo, AnACNPs showed best tumor targeting efficiency. The distribution of AnACNPs in tumor was higher than all the other particles. After microvessel staining with anti-CD31 antibody, the fluorescent distribution demonstrated AnACNPs could distribute in the whole tumor with the highest intensity. In conclusion, a novel drug delivery system was developed for enhanced tumor dual targeting and elevated cell internalization.

  13. 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

  14. 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.

  15. 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

  16. Combinatorial Treatment of DNA and Chromatin-Modifying Drugs Cause Cell Death in Human and Canine Osteosarcoma Cell Lines

    PubMed Central

    Thayanithy, Venugopal; Park, ChangWon; Sarver, Aaron L.; Kartha, Reena V.; Korpela, Derek M.; Graef, Ashley J.; Steer, Clifford J.; Modiano, Jaime F.; Subramanian, Subbaya

    2012-01-01

    Downregulation of microRNAs (miRNAs) at the 14q32 locus stabilizes the expression of cMYC, thus significantly contributing to osteosarcoma (OS) pathobiology. Here, we show that downregulation of 14q32 miRNAs is epigenetically regulated. The predicted promoter regions of miRNA clusters at 14q32 locus showed no recurrent patterns of differential methylation, but Saos2 cells showed elevated histone deacetylase (HDAC) activity. Treatment with 4-phenylbutyrate increased acetylation of histones associated with 14q32 miRNAs, but interestingly, robust restoration of 14q32 miRNA expression, attenuation of cMYC expression, and induction of apoptosis required concomitant treatment with 5-Azacytidine, an inhibitor of DNA methylation. These events were associated with genome-wide gene expression changes including induction of pro-apoptotic genes and downregulation of cell cycle genes. Comparable effects were achieved in human and canine OS cells using the HDAC inhibitor suberoylanilide hydroxamic acid (SAHA/Vorinostat) and the DNA methylation inhibitor Zebularine (Zeb), with significantly more pronounced cytotoxicity in cells whose molecular phenotypes were indicative of aggressive biological behavior. These results suggested that the combination of these chromatin-modifying drugs may be a useful adjuvant in the treatment of rapidly progressive OS. PMID:22957032

  17. Adoptive Immunotherapy for Hematological Malignancies Using T Cells Gene-Modified to Express Tumor Antigen-Specific Receptors

    PubMed Central

    Fujiwara, Hiroshi

    2014-01-01

    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. PMID:25517545

  18. CD133 is a modifier of hematopoietic progenitor frequencies but is dispensable for the maintenance of mouse hematopoietic stem cells

    PubMed Central

    Arndt, Kathrin; Grinenko, Tatyana; Mende, Nicole; Reichert, Doreen; Portz, Melanie; Ripich, Tatsiana; Carmeliet, Peter; Corbeil, Denis; Waskow, Claudia

    2013-01-01

    Pentatransmembrane glycoprotein prominin-1 (CD133) is expressed at the cell surface of multiple somatic stem cells, and it is widely used as a cell surface marker for the isolation and characterization of human hematopoietic stem cells (HSCs) and cancer stem cells. CD133 has been linked on a cell biological basis to stem cell-fate decisions in human HSCs and emerges as an important physiological regulator of stem cell maintenance and expansion. Its expression and physiological relevance in the murine hematopoietic system is nevertheless elusive. We show here that CD133 is expressed by bone marrow-resident murine HSCs and myeloid precursor cells with the developmental propensity to give rise to granulocytes and monocytes. However, CD133 is dispensable for the pool size and function of HSCs during steady-state hematopoiesis and after transplantation, demonstrating a substantial species difference between mouse and man. Blood cell numbers in the periphery are normal; however, CD133 appears to be a modifier for the development of growth-factor responsive myeloerythroid precursor cells in the bone marrow under steady state and mature red blood cells after hematopoietic stress. Taken together, these studies show that CD133 is not a critical regulator of hematopoietic stem cell function in mouse but that it modifies frequencies of growth-factor responsive hematopoietic progenitor cells during steady state and after myelotoxic stress in vivo. PMID:23509298

  19. An intravenous implantable glucose/dioxygen biofuel cell with modified flexible carbon fiber electrodes.

    PubMed

    Sales, Fernanda C P F; Iost, Rodrigo M; Martins, Marccus V A; Almeida, Maria C; Crespilho, Frank N

    2013-02-01

    An intravenous implantable glucose/dioxygen hybrid enzyme-Pt micro-biofuel cell (BFC) was investigated. In this miniaturized BFC, a flexible carbon fiber (FCF) microelectrode modified with neutral red redox mediator and glucose oxidase was used as the bioanode, and an FCF modified with platinum nanoparticles stabilized on PAMAM-G4 dendrimer was used as the cathode. In vitro experiments conducted using the BFC in a phosphate buffer solution (50 mmol L(-1), pH = 7.2) and glucose (47 mmol L(-1)) showed high electrocatalytic performance with an open circuit voltage (OCV) of 400 mV, a maximum current density of 2700 μA cm(-2) at 0.0 V and a maximum output power of 200 μW cm(-2) at 250 mV. Under physiological conditions, glucose from rat blood is used as a fuel in anodic reactions and dissolved molecular oxygen is used as the oxidizing agent on the cathode. For in vivo experiments, the BFC was inserted into the jugular vein of a living rat (Rattus novergicus) using a catheter (internal diameter 0.5 mm). The power density of the implantable BFC was evaluated over a period of 24 h, and an OCV of 125 mV with a maximum power density of 95 μW cm(-2) was obtained at 80 mV.

  20. 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.

  1. Subcellular compartmentalization in protoplasts from Artemisia annua cell cultures: engineering attempts using a modified SNARE protein.

    PubMed

    Di Sansebastiano, Gian Pietro; Rizzello, Francesca; Durante, Miriana; Caretto, Sofia; Nisi, Rossella; De Paolis, Angelo; Faraco, Marianna; Montefusco, Anna; Piro, Gabriella; Mita, Giovanni

    2015-05-20

    Plants are ideal bioreactors for the production of macromolecules but transport mechanisms are not fully understood and cannot be easily manipulated. Several attempts to overproduce recombinant proteins or secondary metabolites failed. Because of an independent regulation of the storage compartment, the product may be rapidly degraded or cause self-intoxication. The case of the anti-malarial compound artemisinin produced by Artemisia annua plants is emblematic. The accumulation of artemisinin naturally occurs in the apoplast of glandular trichomes probably involving autophagy and unconventional secretion thus its production by undifferentiated tissues such as cell suspension cultures can be challenging. Here we characterize the subcellular compartmentalization of several known fluorescent markers in protoplasts derived from Artemisia suspension cultures and explore the possibility to modify compartmentalization using a modified SNARE protein as molecular tool to be used in future biotechnological applications. We focused on the observation of the vacuolar organization in vivo and the truncated form of AtSYP51, 51H3, was used to induce a compartment generated by the contribution of membrane from endocytosis and from endoplasmic reticulum to vacuole trafficking. The artificial compartment crossing exocytosis and endocytosis may trap artemisinin stabilizing it until extraction; indeed, it is able to increase total enzymatic activity of a vacuolar marker (RGUSChi), probably increasing its stability. Exploring the 51H3-induced compartment we gained new insights on the function of the SNARE SYP51, recently shown to be an interfering-SNARE, and new hints to engineer eukaryote endomembranes for future biotechnological applications.

  2. Cell Death Pathways in Astrocytes with a Modified Model of Oxygen-Glucose Deprivation

    PubMed Central

    Zou, Liang yu; Cao, Xu; Chu, Xiaofan

    2013-01-01

    Traditional oxygen-glucose deprivation (OGD) models do not produce sufficiently stable and continuous deprivation to induce cell death in the ischemic core. Therefore, we modified the OGD model to mimic the observed damage in the ischemic core following stroke and utilized this new model to study cell death pathways in astrocytes. The PO2 and pH levels in the astrocyte culture medium were compared between a physical OGD group, a chemical OGD group and a mixed OGD group. The mixed OGD group was able to maintain anaerobic conditions in astrocyte culture medium for 6 h, while the physical and the chemical groups failed to maintain such conditions. Astrocyte viability decreased and LDH release into in the medium increased as a function of exposure to OGD. Compared to the control group, the expression of active caspase-3 in the mixed OGD group increased within 2 h after OGD, but decreased after 2 h of OGD. Additionally, porimin mRNA levels did not significantly increase during the first 2 h of OGD, while bcl-2 mRNA levels decreased at 1 h. However, both porimin and bcl-2 mRNA levels increased after 2 h of OGD; interestingly, they both suddenly decreased at 4 h of OGD. Taken together, these results indicate that apoptosis and oncosis are the two cell death pathways responsible for astrocyte death in the ischemic core. However, the main death pathway varies depending on the OGD period. PMID:23637816

  3. Extracellular matrix proteins interact with cell-signaling pathways in modifying risk of achilles tendinopathy.

    PubMed

    Saunders, Colleen J; van der Merwe, Lize; Cook, Jill; Handley, Christopher J; Collins, Malcolm; September, Alison V

    2015-06-01

    The aim of this study was to investigate interactions between variants within genes encoding components of the collagen fibril and components of cell-signaling pathways within the extracellular matrix, and determine the relative contribution of these variants to Achilles tendinopathy risk in a polygenic model. A total of 339 asymptomatic control participants and 179 participants clinically diagnosed with Achilles tendinopathy were genotyped for variants within six genes encoding components of the collagen fibril and three genes encoding components of cell-signaling pathways. Logistic regression, stepwise selection, and receiver operating characteristic curve (ROC) analysis was used to select and evaluate genetic interactions and determine the relative contribution of these variants to overall genetic risk. The strongest, best fit polygenic risk model included the variables sex, three COL27A1 variants (rs4143245; rs1249744; rs946053), COL5A1 rs12722, CASP8 rs1045485, and CASP8 rs2824129 with an area under the ROC curve of 0.737 and the maximum sum of sensitivity and specificity indicators equal to 134%. Significant interactions between genes encoding components of the collagen fibril and genes encoding components of the cell-signaling pathways modify risk of Achilles tendinopathy.

  4. 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.

  5. Investigation of modified p-n junctions in crystalline silicon on glass solar cells

    NASA Astrophysics Data System (ADS)

    Lausch, D.; Werner, M.; Naumann, V.; Schneider, J.; Hagendorf, C.

    2011-04-01

    In this paper various methods for studying p-n junctions in thin film solar cells are applied with the aim to localize and investigate defects on a microscopic scale. Different electron and ion beam characterization methods are introduced to determine the p-n junction position using two different examples from crystalline silicon on glass thin film technology. In a first example, planview and cross section electron beam induced current measurements revealed that oxygen rich columnar growth at textured substrates strongly disturbs the p-n junction. In a second example, diffusion from glass substrate is identified by ToF-SIMS to influence the electrical and structural characteristics of the thin Si layer resulting in a modified p-n junction. A model describing the formation of both defect structures is introduced.

  6. Carbon Nanofibers Modified Graphite Felt for High Performance Anode in High Substrate Concentration Microbial Fuel Cells

    PubMed Central

    Shen, Youliang; Zhou, Yan; Chen, Shuiliang; Yang, Fangfang; Zheng, Suqi; Hou, Haoqing

    2014-01-01

    Carbon nanofibers modified graphite fibers (CNFs/GF) composite electrode was prepared for anode in high substrate concentration microbial fuel cells. Electrochemical tests showed that the CNFs/GF anode generated a peak current density of 2.42 mA cm−2 at a low acetate concentration of 20 mM, which was 54% higher than that from bare GF. Increase of the acetate concentration to 80 mM, in which the peak current density of the CNFs/GF anode greatly increased and was up to 3.57 mA cm−2, was seven times as that of GF anode. Morphology characterization revealed that the biofilms in the CNFs/GF anode were much denser than those in the bare GF. This result revealed that the nanostructure in the anode not only enhanced current generation but also could tolerate high substrate concentration. PMID:24883348

  7. Carbon nanofibers modified graphite felt for high performance anode in high substrate concentration microbial fuel cells.

    PubMed

    Shen, Youliang; Zhou, Yan; Chen, Shuiliang; Yang, Fangfang; Zheng, Suqi; Hou, Haoqing

    2014-01-01

    Carbon nanofibers modified graphite fibers (CNFs/GF) composite electrode was prepared for anode in high substrate concentration microbial fuel cells. Electrochemical tests showed that the CNFs/GF anode generated a peak current density of 2.42 mA cm(-2) at a low acetate concentration of 20 mM, which was 54% higher than that from bare GF. Increase of the acetate concentration to 80 mM, in which the peak current density of the CNFs/GF anode greatly increased and was up to 3.57 mA cm(-2), was seven times as that of GF anode. Morphology characterization revealed that the biofilms in the CNFs/GF anode were much denser than those in the bare GF. This result revealed that the nanostructure in the anode not only enhanced current generation but also could tolerate high substrate concentration.

  8. Methanol electro-oxidation on platinum modified tungsten carbides in direct methanol fuel cells: a DFT study.

    PubMed

    Sheng, Tian; Lin, Xiao; Chen, Zhao-Yang; Hu, P; Sun, Shi-Gang; Chu, You-Qun; Ma, Chun-An; Lin, Wen-Feng

    2015-10-14

    In exploration of low-cost electrocatalysts for direct methanol fuel cells (DMFCs), Pt modified tungsten carbide (WC) materials are found to be great potential candidates for decreasing Pt usage whilst exhibiting satisfactory reactivity. In this work, the mechanisms, onset potentials and activity for electrooxidation of methanol were studied on a series of Pt-modified WC catalysts where the bare W-terminated WC(0001) substrate was employed. In the surface energy calculations of a series of Pt-modified WC models, we found that the feasible structures are mono- and bi-layer Pt-modified WCs. The tri-layer Pt-modified WC model is not thermodynamically stable where the top layer Pt atoms tend to accumulate and form particles or clusters rather than being dispersed as a layer. We further calculated the mechanisms of methanol oxidation on the feasible models via methanol dehydrogenation to CO involving C-H and O-H bonds dissociating subsequently, and further CO oxidation with the C-O bond association. The onset potentials for the oxidation reactions over the Pt-modified WC catalysts were determined thermodynamically by water dissociation to surface OH* species. The activities of these Pt-modified WC catalysts were estimated from the calculated kinetic data. It has been found that the bi-layer Pt-modified WC catalysts may provide a good reactivity and an onset oxidation potential comparable to pure Pt and serve as promising electrocatalysts for DMFCs with a significant decrease in Pt usage. PMID:26351805

  9. A modified Nafion membrane with in situ polymerized polypyrrole for the direct methanol fuel cell

    NASA Astrophysics Data System (ADS)

    Smit, M. A.; Ocampo, A. L.; Espinosa-Medina, M. A.; Sebastián, P. J.

    Nafion membranes were modified by the in situ electrodeposition of polypyrrole inside the membrane pores and on the anode side only, in order to prevent the cross-over of methanol in the direct methanol fuel cell (DMFC). Pretreated Nafion membranes were first immersed in 0.1 M sulphuric acid containing the pyrrole monomer and subsequently removed from this solution and placed in a two-electrode solid-state electrochemical cell, where the polypyrrole was formed galvanostatically. The modified membranes were studied in terms of morphology, electrochemical characteristics and methanol permeability. FTIR and SEM confirmed the presence of the polypyrrole on the anode side of the Nafion membrane. SEM shows the polymer to be present both on the membrane surface and inside the membrane pores. It was found to be deposited as small grains, with two distinct sizes, the smallest particles have a diameter of around 100 nm, while the larger particles have diameters of around 700 nm. Methanol permeability was determined electrochemically and was shown to be effectively reduced. Cyclic voltammetry was performed in sulphuric acid, in pure methanol and in 50 vol.% methanol. The untreated Nafion membrane showed CV curves which were similar in all electrolytes with electroactivity only at the extreme ends of the curve. The Nafion/Ppy membrane showed typical polypyrrole curves, with current densities lowest in sulphuric acid, and highest in the 50 vol.% methanol, respectively. For the methanol containing electrolytes, an additional oxidative peak appears in the CV, which may be related to electrocatalytic activity of the polypyrrole for methanol oxidation.

  10. A modified host-cell reactivation assay to quantify DNA repair capacity in cryopreserved peripheral lymphocytes.

    PubMed

    Mendez, Pedro; Taron, Miquel; Moran, Teresa; Fernandez, Marco A; Requena, Gerard; Rosell, Rafael

    2011-06-10

    The host-cell reactivation assay (HCRA) is a functional assay that allows the identification of the genes responsible for DNA repair-deficient syndromes, such as Xeroderma pigmentosum, by cross-complementation experiments. It has also been used in molecular epidemiology studies to correlate the low nucleotide excision repair pathway function in peripheral blood lymphocytes with an increased risk of bladder, head and neck, skin and lung cancers. Herein, we present the technical validation of a newly modified HCRA, where nucleofection is used for the transfection of the pmaxGFP plasmid into cryopreserved peripheral blood lymphocytes (PBLs) or lymphoblastoid cell lines. In each sample, 20-24h after transfection, the relative DNA repair capacity (DRC) was quantified by flow cytometry, comparing the transfection efficiency of nucleoporated cells with undamaged plasmid to those transfected with UV-light damaged plasmid in the seven cell lines that were characterized by different DNA repair phenotypes. Dead cells were excluded from the analysis. We observed a high reproducibility of the relative DRC, transfection efficiency and cell viability. The inter-experimental normalization of the flow cytometry resulted in an increased data accuracy and reproducibility. The amount of cells required for each transfection reaction was reduced fourfold, without affecting the final relative DRC. Furthermore, our HCRA demonstrated strong discrimination power in the UV-light dose-response, both in lymphoblastoid cell lines and cryopreserved PBLs. We also observed a strong correlation of the relative DRC data, when samples were measured against two independent batches of both damaged and undamaged plasmid DNA. The relative DRC variable shows a normal distribution when analyzed in the cryopreserved PBLs from a cohort of 35 lung cancer patients and a 5.59-fold variation in the relative DRC is identified among our patients. The mitotic dynamic was discarded as a confounding factor for the

  11. Uses of thermoresponsive and RGD/insulin-modified poly(vinyl ether)-based hydrogels in cell cultures.

    PubMed

    Gümüşderelioğlu, Menemşe; Karakeçili, Ayşe Gönen

    2003-01-01

    Thermoresponsive hydrogels were synthesized by radiation copolymerization of ethylene glycol vinyl ether (1) and butyl vinyl ether (2) in the presence of cross-linking agent diethylene glycol divinyl ether. The comonomer ratio (monomer 1/monomer 2) and the cross-linker concentration were kept constant at 60:40 (mole percentage in the monomeric mixture) and 4% (mole basis), respectively. The hydrogels showed a volume-phase transition in the temperature range 10-25 degrees C and their swelling behaviour was reversible. The gels were modified by a cell adhesion factor, the RGD sequence of fibronectin, and a cell growth factor, insulin. However, they lost their thermoresponsive character after modification. The use of the gels in cell culture was investigated without using a proteolytic enzyme or serum. Cell culture studies realized by human skin fibroblasts (HS An1) showed that the cells can attach and proliferate on the surface of a thermoresponsive polymer. 80% of the cultured cells were readily detached from the polymer surface by lowering the incubation temperature from 37 degrees C to 10 degrees C for 30 min. In the studies carried out with RGD or insulin-modified hydrogels in serum-free cultures, higher values of cell proliferation (9 x 10(5) cells/ml) were obtained on the insulin-modified hydrogels, whereas higher values of cell attachment were obtained on the RGD-immobilized surfaces.

  12. High frequency detection of different T-cell subsets in mice by a modified virus plaque assay.

    PubMed Central

    Fujisawa, H; Kumazawa, Y; Ohtani, A; Nishimura, C

    1983-01-01

    Different T-cell subsets participating in immune responses were detected at a high frequency by a modified virus plaque assay (VPA). By using the modified VPA, different activated T-cell subsets generated in primary immune responses, helper and suppressor T cells participating in antibody formation, and effector T cells involved in the delayed-type hypersensitivity (DTH) reaction were enumerated directly without in vitro antigen stimulation. The frequency of detection in immune systems used was 7.5-17.7 V-PFC/10(3) spleen cells. Although neither helper T cells for antibody formation nor effector T cells for DTH reaction were detected as V-PFC at a high frequency by the original VPA, it was also found in secondary immune response that Lyt 1 positive, antigen-specific helper and effector T-cell subsets, and cyclophosphamide (CY)-resistant precursors were enumerated at a high frequency by the modified VPA when the received in vitro antigen stimulation, and that the proliferative stage of these cells was critical for the development of V-PFC. PMID:6601613

  13. 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.

  14. 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

  15. Organically Modified Silica Nanoparticles Interaction with Macrophage Cells: Assessment of Cell Viability on the Basis of Physicochemical Properties.

    PubMed

    Kumar, Dhiraj; Mutreja, Isha; Keshvan, Prashant C; Bhat, Madhusudan; Dinda, Amit K; Mitra, Susmita

    2015-11-01

    Silica nanoparticles have drawn a lot of attention for nanomedicine application, and this is attributed to their biocompatibility and ease of surface functionalization. However, successful utilization of these inorganic systems for biomedical application depends on their physicochemical properties. This study, therefore, discusses in vitro toxicity of organically modified silica nanoparticles on the basis of size, shape, and surface properties of silica nanoparticles. Spherical- and oval-shaped nanoparticles having hydroxyl and amine groups were synthesized in Tween 80 micelles using different organosilanes. Nanoparticles of similar size and morphology were considered for comparative assessment. "As-prepared" nanoparticles were characterized in terms of size, shape, and surface properties using ZetaSizer, transmission electron microscopy, and Fourier transform infrared to establish the above parameters. In vitro analysis in terms of nanoparticle-based toxicity was performed on J-774 (macrophage) cell line using propidium iodide-4',6-diamidino-2-phenylindol and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays. Fluorescent dye-entrapped nanoparticles were used to visualize the uptake of the nanoparticles by macrophage cells. Results from cell studies suggested low levels of toxicity for different nanoparticle formulations studied, therefore are suitable for nanocarrier application for poorly soluble molecules. On the contrary, the nanoparticles of similar size and shape, having amine groups and low net negative charge, do not exhibit any in vitro cytotoxicity.

  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. 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.

  18. 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. PMID:27433617

  19. 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

  20. 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.

  1. 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.

  2. 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. PMID:25046528

  3. Synthetically Modified Viral Capsids as Versatile Carriers for Use in Antibody-Based Cell Targeting.

    PubMed

    ElSohly, Adel M; Netirojjanakul, Chawita; Aanei, Ioana L; Jager, Astraea; Bendall, Sean C; Farkas, Michelle E; Nolan, Garry P; Francis, Matthew B

    2015-08-19

    The present study describes an efficient and reliable method for the preparation of MS2 viral capsids that are synthetically modified with antibodies using a rapid oxidative coupling strategy. The overall protocol delivers conjugates in high yields and recoveries, requires a minimal excess of antibody to achieve modification of more than 95% of capsids, and can be completed in a short period of time. Antibody-capsid conjugates targeting extracellular receptors on human breast cancer cell lines were prepared and characterized. Notably, conjugation to the capsid did not significantly perturb the binding of the antibodies, as indicated by binding affinities similar to those obtained for the parent antibodies. An array of conjugates was synthesized with various reporters on the interior surface of the capsids to be used in cell studies, including fluorescence-based flow cytometry, confocal microscopy, and mass cytometry. The results of these studies lay the foundation for further exploration of these constructs in the context of clinically relevant applications, including drug delivery and in vivo diagnostics. PMID:26076186

  4. 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.

  5. 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. PMID:19712996

  6. 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. PMID:26784658

  7. Paracrine action of HO-1-modified mesenchymal stem cells mediates cardiac protection and functional improvement.

    PubMed

    Zeng, Bin; Ren, Xiaofeng; Lin, Guosheng; Zhu, Chengang; Chen, Honglei; Yin, Jiechao; Jiang, Hong; Yang, Bo; Ding, Danhua

    2008-10-01

    The aim has been to determine whether the supernatants of mesenchymal stem cells (MSCs) transfected with adenovirus carrying human heme oxygenase-1 (hHO-1) gene protect cardiomyocytes from ischemic injury. We have found that hHO-1 infected MSCs (hHO-1-MSCs) increased expression of hHO-1 protein. Apoptosis of cultured hHO-1-MSCs exposed to hypoxia was suppressed. Several cytokines, including HGF, bFGF, TGF-beta, VEGF and IL-1beta, were produced by hHO-1-MSCs, some being significantly enhanced under hypoxia stimulation. Meanwhile, those cytokines reduced caspase-3 level and activity in cultured adult rat ventricular cardiomyocytes (ARVCs) exposed to hypoxia. Supernatants obtained from hHO-1-MSCs improved left ventricular function, limited myocardial infarct size, increased microvessel density, and inhibited apoptosis of cardiomyocytes in rat myocardial infarction. It can be concluded hHO-1-modified MSCs prevent myocardial cell injury via secretion of paracrine-acting mediators.

  8. 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

  9. 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

  10. 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.

  11. In Vivo Proof of Concept of Adoptive Immunotherapy for Hepatocellular Carcinoma Using Allogeneic Suicide Gene-modified Killer Cells

    PubMed Central

    Leboeuf, Céline; Mailly, Laurent; Wu, Tao; Bour, Gaetan; Durand, Sarah; Brignon, Nicolas; Ferrand, Christophe; Borg, Christophe; Tiberghien, Pierre; Thimme, Robert; Pessaux, Patrick; Marescaux, Jacques; Baumert, Thomas F.; Robinet, Eric

    2014-01-01

    Cell therapy based on alloreactivity has completed clinical proof of concept against hematological malignancies. However, the efficacy of alloreactivity as a therapeutic approach to treat solid tumors is unknown. Using cell culture and animal models, we aimed to investigate the efficacy and safety of allogeneic suicide gene-modified killer cells as a cell-based therapy for hepatocellular carcinoma (HCC), for which treatment options are limited. Allogeneic killer cells from healthy donors were isolated, expanded, and phenotypically characterized. Antitumor cytotoxic activity and safety were studied using a panel of human or murine HCC cell lines engrafted in immunodeficient or immunocompetent mouse models. Human allogeneic suicide gene-modified killer cells (aSGMKCs) exhibit a high, rapid, interleukin-2–dependent, and non–major histocompatibility complex class I-restricted in vitro cytotoxicity toward human hepatoma cells, mainly mediated by natural killer (NK) and NK-like T cells. In vivo evaluation of this cell therapy product demonstrates a marked, rapid, and sustained regression of HCC. Preferential liver homing of effector cells contributed to its marked efficacy. Calcineurin inhibitors allowed preventing rejection of allogeneic lymphocytes by the host immune system without impairing their antitumor activity. Our results demonstrate proof of concept for aSGMKCs as immunotherapy for HCC and open perspectives for the clinical development of this approach. PMID:24445938

  12. Regulation of Epithelial Cell Morphology and Functions Approaching To More In Vivo-Like by Modifying Polyethylene Glycol on Polysulfone Membranes

    PubMed Central

    Shen, Chong; Zhang, Guoliang; Meng, Qin

    2012-01-01

    Cytocompatibility is critically important in design of biomaterials for application in tissue engineering. However, the currently well-accepted “cytocompatible" biomaterials are those which promote cells to sustain good attachment/spreading. The cells on such materials usually lack the self-assembled cell morphology and high cell functions as in vivo. In our view, biomaterials that can promote the ability of cells to self-assemble and demonstrate cell-specific functions would be cytocompatible. This paper examined the interaction of polyethylene glycol (PEG) modified polysulfone (PSf) membranes with four epithelial cell types (primary liver cells, a liver tumor cell line, and two renal tubular cell lines). Our results show that PSf membranes modified with proper PEG promoted the aggregation of both liver and renal cells, but the liver cells more easily formed aggregates than the renal tubular cells. The culture on PEG-modified PSf membranes also enhanced cell-specific functions. In particular, the cells cultured on F127 membranes with the proper PEG content mimicked the in vivo ultrastructure of liver cells or renal tubules cells and displayed the highest cell functions. Gene expression data for adhesion proteins suggest that the PEG modification impaired cell-membrane interactions and increased cell-cell interactions, thus facilitating cell self-assembly. In conclusion, PEG-modified membrane could be a cytocompatible material which regulates the morphology and functions of epithelial cells in mimicking cell performance in vivo. PMID:22558349

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

    PubMed

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

    2016-07-01

    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

  14. 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

  15. Maternal nutrition modifies trophoblast giant cell phenotype and fetal growth in mice

    PubMed Central

    Watkins, Adam J; Lucas, Emma S; Marfy-Smith, Stephanie; Bates, Nicola; Kimber, Susan J; Fleming, Tom P

    2015-01-01

    Mammalian placentation is dependent upon the action of trophoblast cells at the time of implantation. Appropriate fetal growth, regulated by maternal nutrition and nutrient transport across the placenta, is a critical factor for adult offspring long-term health. We have demonstrated that a mouse maternal low-protein diet (LPD) fed exclusively during preimplantation development (Emb-LPD) increases offspring growth but programmes adult cardiovascular and metabolic disease. In this study, we investigate the impact of maternal nutrition on post-implantation trophoblast phenotype and fetal growth. Ectoplacental cone explants were isolated at day 8 of gestation from female mice fed either normal protein diet (NPD: 18% casein), LPD (9% casein) or Emb-LPD and cultured in vitro. We observed enhanced spreading and cell division within proliferative and secondary trophoblast giant cells (TGCs) emerging from explants isolated from LPD-fed females when compared with NPD and Emb-LPD explants after 24 and 48 h. Moreover, both LPD and Emb-LPD explants showed substantial expansion of TGC area during 24–48 h, not observed in NPD. No difference in invasive capacity was observed between treatments using Matrigel transwell migration assays. At day 17 of gestation, LPD- and Emb-LPD-fed conceptuses displayed smaller placentas and larger fetuses respectively, resulting in increased fetal:placental ratios in both groups compared with NPD conceptuses. Analysis of placental and yolk sac nutrient signalling within the mammalian target of rapamycin complex 1 pathway revealed similar levels of total and phosphorylated downstream targets across groups. These data demonstrate that early post-implantation embryos modify trophoblast phenotype to regulate fetal growth under conditions of poor maternal nutrition. PMID:25755287

  16. 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.

  17. Nuclear transfer of adult and genetically modified fetal cells of the rat.

    PubMed

    Hayes, E; Galea, S; Verkuylen, A; Pera, M; Morrison, J; Lacham-Kaplan, O; Trounson, A

    2001-04-27

    The present study examines the handling, activation, and micromanipulation of rat eggs in an attempt to produce live young using nuclear transfer (NT) of adult and genetically modified rat fetal cells. Mature rat eggs cultured in calcium-free medium showed reduced rates (24%) of chromosomal dispersion ("spontaneous activation" characteristic of this species) compared with eggs cultured in calcium-containing medium (47%), but failed to survive micromanipulation procedures. High rates of parthenogenetic cleavage were obtained with chemical activation using ethanol/cycloheximide (65%) compared with other standard chemical activation methods (4-28%). This type of activation was also effective in reestablishing cleavage capability (19-71%), in a time-dependent manner, of spontaneously activated eggs arrested at a second prophase-like state. At most, two of four tested micromanipulation procedures were effective in producing NT embryos capable of morula or blastocyst development (14-16%) in vivo following transfer to mouse oviducts. NT blastocysts produced from cumulus cells and transfected rat fetal fibroblasts appeared morphologically and karyotypically normal (2n = 42). Nocodazole-assisted metaphase enucleation and piezoelectric-assisted donor cell injection produced significant and equivocal effects on survival and cleavage rates of reconstructed embryos but failed to significantly improve in vivo morula/blastocyst development rates (16-28%) compared with unassisted micromanipulation (16%). Live births have not yet been obtained from early cleavage stage embryos (n = 269) transferred to pseudopregnant recipient rat oviducts. Improvements in reconstituted NT embryo culture and transfer are required for these methods to be an effective means of transgenic rat production.

  18. Cross-resistance to UV radiation of a cisplatin-resistant human cell line: Overexpression of cellular factors that recognize UV-modified DNA

    SciTech Connect

    Chao, C.C.; Huang, S.L.; Huang, H.M.; Lin-Chao, S. )

    1991-04-01

    A human cell line selected for cisplatin resistance (CPR) was irradiated with UV light and showed cross-resistance to UV light. Applying a modified chloramphenicol acetyltransferase assay, we observed that CPR cells acquired enhanced host cell reactivation of a transfected plasmid carrying UV damage. Gel mobility shift analysis indicated that two nuclear factors that recognize UV-modified DNA were overexpressed in CPR cells. In addition, factors that bind UV-modified DNA were independent from the factors that bind cisplatin-modified DNA. The significance of the identified binding factors, possibly DNA repair enzymes, is discussed.

  19. Modifying the 5'-Cap for Click Reactions of Eukaryotic mRNA and To Tune Translation Efficiency in Living Cells.

    PubMed

    Holstein, Josephin M; Anhäuser, Lea; Rentmeister, Andrea

    2016-08-26

    The 5'-cap is a hallmark of eukaryotic mRNAs and plays fundamental roles in RNA metabolism, ranging from quality control to export and translation. Modifying the 5'-cap may thus enable modulation of the underlying processes and investigation or tuning of several biological functions. A straightforward approach is presented for the efficient production of a range of N7-modified caps based on the highly promiscuous methyltransferase Ecm1. We show that these, as well as N(2) -modified 5'-caps, can be used to tune translation of the respective mRNAs both in vitro and in cells. Appropriate modifications allow subsequent bioorthogonal chemistry, as demonstrated by intracellular live-cell labeling of a target mRNA. The efficient and versatile N7 manipulation of the mRNA cap makes mRNAs amenable to both modulation of their biological function and intracellular labeling, and represents a valuable addition to the chemical biology toolbox. PMID:27511141

  20. Generation of human lactoferrin transgenic cloned goats using donor cells with dual markers and a modified selection procedure.

    PubMed

    An, Li-You; Yuan, Yu-Guo; Yu, Bao-Li; Yang, Ting-Jia; Cheng, Yong

    2012-10-01

    The objective was to use dual markers to accurately select genetically modified donor cells and ensure that the resulting somatic cell nuclear transfer kids born were transgenic. Fetal fibroblast cells were transfected with dual marking gene vector (pCNLF-ng) that contained the red-shifted variant of the jellyfish green fluorescent protein (LGFP) and neomycin resistance (Neo) markers. Cell clones that were G418-resistant and polymerase chain reaction-positive were subcultured for several passages; individual cells of the clones were examined with fluorescence microscopy to confirm transgenic integration. Clones in which every cell had bright green fluorescence were used as donor cells for nuclear transfer. In total, 86.7% (26/30) cell clones were confirmed to have transgenic integration of the markers by polymerase chain reaction, 76.7% (23/30) exhibited fluorescence, but only 40% (12/30) of these fluorescent cell clones had fluorescence in all cell populations. Moreover, through several cell passages, only 20% (6/30) of the cell clones exhibited stable LGFP expression. Seven transgenic cloned offspring were produced from these cells by nuclear transfer. Overall, the reconstructed embryo fusion rate was 76.6%, pregnancy rates at 35 and 60 days were 39.1% and 21.7%, respectively, and the offspring birth rate was 1.4%. There were no significant differences between nuclear transfer with dual versus a single (Neo) marker (overall, 73.8% embryo fusion rate, 53.8% and 26.9% pregnancy rates, and 1.9% birth rate with five offspring). In conclusion, the use of LGFP/Neo dual markers and an optimized selection procedure reliably screened genetically modified donor cells, excluded pseudotransgenic cells, and led to production of human lactoferrin transgenic goats. Furthermore, the LGFP/Neo markers had no adverse effects on the efficiency of somatic cell nuclear transfer.

  1. Biological characteristics of adipose tissue-derived stem cells labeled with amine-surface-modified superparamagnetic iron oxide nanoparticles.

    PubMed

    Wang, Nan; Zhao, Jing-Yuan; Guan, Xin; Dong, Yue; Liu, Yang; Zhou, Xiang; Wu, Ren'an; Du, Yue; Zhao, Liang; Zou, Wei; Han, Chao; Song, Lin; Sun, Bo; Liu, Yan; Liu, Jing

    2015-08-01

    Cell labeling and tracking are becoming increasingly important areas within the field of stem cell transplantation. The ability to track the migration and distribution of implanted cells is critical to understanding the beneficial effects and mechanisms of stem cell therapy. The present study investigated the effects of amine-surface-modified superparamagnetic iron oxide (SPIO) nanoparticles on the biological properties of human adipose tissue-derived stem cells (hADSCs). Monodisperse hydrophobic magnetite (Fe3 O4 ) nanoparticles were prepared using silicon and surface-modified with amine coating. Cell viability, proliferation, differentiation potential, and surface marker expression were evaluated. The magnetic particles (10-18 nm) displayed high labeling efficiency and stability in hADSCs. SPIO-labeled cells produced a hypointense signal and were effectively visualized by MRI for up to 21 days. The results of MTT proliferation assays and flow cytometry analysis demonstrated that SPIOs were biocompatible, viz. the labeling process did not cause cell death or apoptosis and had no side effects on cell proliferation. In vivo experiments showed that the magnetic particles did not affect liver and kidney function. The successful and stable labeling of hADSCs combined with efficient magnetic tropism demonstrates that SPIOs are promising candidates for hADSC tracking in hADSC-based cell therapy applications.

  2. Reduced in vitro T-cell responses induced by glutaraldehyde-modified allergen extracts are caused mainly by retarded internalization of dendritic cells.

    PubMed

    Heydenreich, Bärbel; Bellinghausen, Iris; Lorenz, Steffen; Henmar, Helene; Strand, Dennis; Würtzen, Peter A; Saloga, Joachim

    2012-06-01

    Although allergen-specific immunotherapy is a clinically effective therapy for IgE-mediated allergic diseases, the risk of IgE-mediated adverse effects still exists. For this reason, chemically modified allergoids have been introduced, which may destroy IgE-binding sites while T-cell activation should be retained. The aim of the study was to analyse the differences between intact allergens and differently modified/aggregated allergoids concerning their internalization as well as T-cell and basophil activation. For this purpose human monocyte-derived immature dendritic cells (DC) were incubated with Phleum pratense or Betula verrucosa pollen extract or with the corresponding allergoids, modified with formaldehyde or glutaraldehyde. After an additional maturation process, the antigen-loaded mature DC were co-cultured with autologous CD4(+) T cells. Allergenicity was tested by leukotriene release from basophils. In addition, the uptake of intact allergens and allergoids by immature DC was analysed. The proliferation of, as well as the interleukin-4 (IL-4), IL-10, IL-13 and interferon-γ production by, CD4(+) T cells which had been stimulated with glutaraldehyde allergoid-treated DC was reduced compared with CD4(+) T cells stimulated with intact allergen-treated or formaldehyde allergoid-treated DC. In line with this, glutaraldehyde-modified allergoids were more aggregated and were internalized more slowly. Furthermore, only the allergoids modified with glutaraldehyde induced a decreased leukotriene release by activated basophils. These findings suggest that IgE-reactive epitopes were destroyed more efficiently by modification with glutaraldehyde than with formaldehyde under the conditions chosen for these investigations. Glutaraldehyde-modified allergoids also displayed lower T-cell stimulatory capacity, which is mainly the result of greater modification/aggregation and diminished uptake by DC.

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

    PubMed Central

    2011-01-01

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

  4. Extensive cell migration, axon regeneration and improved function with polysialic acid-modified Schwann cells after spinal cord injury

    PubMed Central

    Ghosh, Mousumi; Tuesta, Luis M.; Puentes, Rocio; Patel, Samik; Melendez, Kiara; Maarouf, Abderrahman El; Rutishauser, Urs; Pearse, Damien Daniel

    2015-01-01

    Schwann cells (SC) implantation after spinal cord injury (SCI) promotes axonal regeneration, remyelination repair and functional recovery. Reparative efficacy, however, may be limited due to the inability of SCs to migrate outward from the lesion-implant site. Altering SC cell surface properties by over-expressing polysialic acid (PSA) has been shown to promote SC migration. In the current study, a SCI contusion was used to evaluate the migration, supraspinal axon growth support and functional recovery associated with polysialyltransferase (PST)-over-expressing SCs (PST-GFP SCs) or controls (GFP SCs). Compared to GFP SCs, which remained confined to the injection site at the injury center, PST-GFP SCs migrated across the lesion:host cord interface for distances of up to 4.4 mm within adjacent host tissue. In addition, with PST-GFP SCs, there was extensive serotonergic and corticospinal axon in-growth within the implants that was limited in the GFP SC controls. The enhanced migration of PST-GFP SCs was accompanied by significant growth of these axons caudal to lesion. Animals receiving PST-GFP SCs exhibited improved functional outcome, both in the open-field and on the gridwalk test, over modest improvements provided by GFP SC controls. The current study for the first time demonstrates that a lack of migration by SC may hinder their reparative benefits and that cell surface overexpression of PSA enhances the ability of implanted SCs to associate with and support the growth of corticospinal axons. These results provide further promise that PSA modified SCs will be a potent reparative approach for SCI. PMID:22460918

  5. Extensive cell migration, axon regeneration, and improved function with polysialic acid-modified Schwann cells after spinal cord injury.

    PubMed

    Ghosh, Mousumi; Tuesta, Luis M; Puentes, Rocio; Patel, Samik; Melendez, Kiara; El Maarouf, Abderrahman; Rutishauser, Urs; Pearse, Damien Daniel

    2012-05-01

    Schwann cell (SC) implantation after spinal cord injury (SCI) promotes axonal regeneration, remyelination repair, and functional recovery. Reparative efficacy, however, may be limited because of the inability of SCs to migrate outward from the lesion-implant site. Altering SC cell surface properties by overexpressing polysialic acid (PSA) has been shown to promote SC migration. In this study, a SCI contusion model was used to evaluate the migration, supraspinal axon growth support, and functional recovery associated with polysialyltransferase (PST)-overexpressing SCs [PST-green fluorescent protein (GFP) SCs] or controls (GFP SCs). Compared with GFP SCs, which remained confined to the injection site at the injury center, PST-GFP SCs migrated across the lesion:host cord interface for distances of up to 4.4 mm within adjacent host tissue. In addition, with PST-GFP SCs, there was extensive serotonergic and corticospinal axon in-growth within the implants that was limited in the GFP SC controls. The enhanced migration of PST-GFP SCs was accompanied by significant growth of these axons caudal to lesion. Animals receiving PST-GFP SCs exhibited improved functional outcome, both in the open-field and on the gridwalk test, beyond the modest improvements provided by GFP SC controls. This study for the first time demonstrates that a lack of migration by SCs may hinder their reparative benefits and that cell surface overexpression of PSA enhances the ability of implanted SCs to associate with and support the growth of corticospinal axons. These results provide further promise that PSA-modified SCs will be a potent reparative approach for SCI. © 2012 Wiley Periodicals, Inc.

  6. Extensive cell migration, axon regeneration, and improved function with polysialic acid-modified Schwann cells after spinal cord injury.

    PubMed

    Ghosh, Mousumi; Tuesta, Luis M; Puentes, Rocio; Patel, Samik; Melendez, Kiara; El Maarouf, Abderrahman; Rutishauser, Urs; Pearse, Damien Daniel

    2012-05-01

    Schwann cell (SC) implantation after spinal cord injury (SCI) promotes axonal regeneration, remyelination repair, and functional recovery. Reparative efficacy, however, may be limited because of the inability of SCs to migrate outward from the lesion-implant site. Altering SC cell surface properties by overexpressing polysialic acid (PSA) has been shown to promote SC migration. In this study, a SCI contusion model was used to evaluate the migration, supraspinal axon growth support, and functional recovery associated with polysialyltransferase (PST)-overexpressing SCs [PST-green fluorescent protein (GFP) SCs] or controls (GFP SCs). Compared with GFP SCs, which remained confined to the injection site at the injury center, PST-GFP SCs migrated across the lesion:host cord interface for distances of up to 4.4 mm within adjacent host tissue. In addition, with PST-GFP SCs, there was extensive serotonergic and corticospinal axon in-growth within the implants that was limited in the GFP SC controls. The enhanced migration of PST-GFP SCs was accompanied by significant growth of these axons caudal to lesion. Animals receiving PST-GFP SCs exhibited improved functional outcome, both in the open-field and on the gridwalk test, beyond the modest improvements provided by GFP SC controls. This study for the first time demonstrates that a lack of migration by SCs may hinder their reparative benefits and that cell surface overexpression of PSA enhances the ability of implanted SCs to associate with and support the growth of corticospinal axons. These results provide further promise that PSA-modified SCs will be a potent reparative approach for SCI. © 2012 Wiley Periodicals, Inc. PMID:22460918

  7. Genetically modified T cells targeting interleukin-11 receptor α-chain kill human osteosarcoma cells and induce the regression of established osteosarcoma lung metastases.

    PubMed

    Huang, Gangxiong; Yu, Ling; Cooper, Laurence J N; Hollomon, Mario; Huls, Helen; Kleinerman, Eugenie S

    2012-01-01

    The treatment of osteosarcoma pulmonary metastases remains a challenge. T cells genetically modified to express a chimeric antigen receptor (CAR), which recognizes a tumor-associated antigen, have shown activity against hematopoietic malignancies in clinical trials, but this requires the identification of a specific receptor on the tumor cell. In the current study, we found that interleukin (IL)-11Rα was selectively expressed on 14 of 16 osteosarcoma patients' lung metastases and four different human osteosarcoma cell lines, indicating that IL-11Rα may be a novel target for CAR-specific T-cell therapy. IL-11Rα expression was absent or low in normal organ tissues, with the exception of the gastrointestinal tract. IL-11Rα-CAR-specific T cells were obtained by non-viral gene transfer of Sleeping Beauty DNA plasmids and selectively expanded ex vivo using artificial antigen-presenting cells derived from IL-11Rα + K562 cells genetically modified to coexpress T-cell costimulatory molecules. IL-11Rα-CAR(+) T cells killed all four osteosarcoma cell lines in vitro; cytotoxicity correlated with the level of IL-11Rα expression on the tumor cells. Intravenous injection of IL-11Rα-CAR(+) T cells into mice resulted in the regression of osteosarcoma pulmonary metastases with no organ toxicity. Together, the data suggest that IL-11Rα-CAR T cells may represent a new therapy for patients with osteosarcoma pulmonary metastases. PMID:22075555

  8. [A Modified Procedure to Isolate Synchronous Cells from Yeasts with Continuous Percoll Density Gradient and Their Raman Discrimination].

    PubMed

    Huang, Shu-shi; Lai, Jun-zhuo; Lu, Ming-qian; Cheng, Qin; Liao, Wei; Chen, Li-mei

    2015-08-01

    A modified procedure of Percoll density gradient centrifugation was developed to isolate and fractionate synchronous cells from stationary phase (sp) cultures of different yeast strains, as well as Raman spectra discrimination of single yeast cells was reported. About 1.75 mL Percoll solution in 2 mL polypropylene centrifugal tube was centrifuged at 19,320 g, 20 °C with an angle rotor for 15 min to form continuous densities gradient (1.00~1.31 g · mL(-1)), approximately 100 μL sample was overlaid onto the preformed continuous density gradient carefully, subsequently, centrifuged at 400 g for 60 min in a tabletop centrifuge equipped with a angle rotor at 25 °C. Yeast samples could be observed that the suspensions were separated into two cell fractions obviously. Both fractions of different yeast strains were respectively determined by differential interference contrast (DIC), phase contrast microscope and synchronous culture to distinguish their morphological and growth trait. The results showed that the lower fraction cells were unbudded, mostly unicellular, highly refractive, homogeneous and uniform in size, and represented growth characteristic synchronously; Their protoplasm had relatively high density, and contained significant concentrations of glycogen; all of which were accordant with description of quiescent yeast cells and G0 cells in previously published paper. It was shown that lower fraction was quiescent cells, synchronous G0 cells as well. A Raman tweezers setup was used to investigate the differences between two fractions, G0 cells and non G0 cells, at a single cell level. The result showed that both G0 cells and the non G0 cells had the same characteristic peaks corresponding biological macromolecules including proteins, carbohydrates and nucleic acids, but all characteristic peak intensities of G0 cells were higher than that of non G0 cells, implied that the macromolecular substance content of G0 cells was more higher. Principal component

  9. 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

  10. 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

  11. 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.

  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. [EVALUATION OF THE CYTOGENETIC AND MUTAGEN-MODIFYING ACTIVITY OF CAFFEINE IN MOUSE BONE MARROW CELLS].

    PubMed

    Durnev, A D; Kulakova, A V; Zhanataev, A K; Oganesiants, L A

    2015-01-01

    The cytogenetic and mutagen-modifying activity of caffeine was studied with the method of chromosomal aberrations in bone marrow cells of mice hybrids F1 CBAxC57BL/6. Caffeine per se was administered intragastrically or intraperitoneally, and in combination with mutagens--intragastrically. Mutagens injected intraperitoneally. Caffeine at doses of 10 and 100 mg/kg (single dose) and 10 mg/kg (five days) in parenteral administration and oral introduction failed to possess cytogenetic activity. In combination with mutagens caffeine (1, 10 and 100 mg/kg) had no effect on the cytogenetic activity of dioxydine (200 mg/kg/intraperitoneally) for a single coadministration, five-day pre or five-day coadministration. In combination with other mutagens under the same processing conditions caffeine at doses of 10 and 100 mg/kg significantly increased cytogenetic effects of cyclophosphamide (20 mg/kg) in the pretreatment of the animals and at the dose of 100 mg/kg significantly attenuated the cytogenetic effect of cisplatin (5 mg/kg) in single and repeated co-administration. Thus we have shown the absence of caffeine cytogenetic activity in vivo and showed the multidirectional effect of caffeine in doses far exceeding its daily consumption, to the manifestation ofcytogenetic effects of certain chemical mutagens in some modes of processing animals.

  14. 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

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

    PubMed

    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

  16. Enhanced Salt Removal in an Inverted Capacitive Deionization Cell Using Amine Modified Microporous Carbon Cathodes.

    PubMed

    Gao, Xin; Omosebi, Ayokunle; Landon, James; Liu, Kunlei

    2015-09-15

    Microporous SpectraCarb carbon cloth was treated using nitric acid to enhance negative surface charges of COO(-) in a neutral solution. This acid-treated carbon was further modified by ethylenediamine to attach -NH2 surface functional groups, resulting in positive surface charges of -NH3(+) via pronation in a neutral solution. Through multiple characterizations, in comparison to pristine SpectraCarb carbon, amine-treated SpectraCarb carbon displays a decreased potential of zero charge but an increased point of zero charge, which is opposed to the effect obtained for acid-treated SpectraCarb carbon. An inverted capacitive deionization cell was constructed using amine-treated cathodes and acid-treated anodes, where the cathode is the negatively polarized electrode and the anode is the positively polarized electrode. Constant-voltage switching operation using NaCl solution showed that the salt removal capacity was approximately 5.3 mg g(-1) at a maximum working voltage of 1.1/0 V, which is an expansion in both the salt capacity and potential window from previous i-CDI results demonstrated for carbon xerogel materials. This improved performance is accounted for by the enlarged cathodic working voltage window through ethylenediamine-derived functional groups, and the enhanced microporosity of the SpectraCarb electrodes for salt adsorption. These results expand the use of i-CDI for efficient desalination applications.

  17. Loss of Modifier of Cell Adhesion Reveals a Pathway Leading to Axonal Degeneration

    PubMed Central

    Chen, Qi; Peto, Charles A.; Shelton, G. Diane; Mizisin, Andrew; Sawchenko, Paul E.; Schubert, David

    2009-01-01

    Axonal dysfunction is the major phenotypic change in many neurodegenerative diseases, but the processes underlying this impairment are not clear. Modifier of cell adhesion (MOCA) is a presenilin binding protein that functions as a guanine nucleotide exchange factor for Rac1. The loss of MOCA in mice leads to axonal degeneration and causes sensorimotor impairments by decreasing cofilin phosphorylation and altering its upstream signaling partners LIM kinase and p21-activated kinase, an enzyme directly downstream of Rac1. The dystrophic axons found in MOCA-deficient mice are associated with abnormal aggregates of neurofilament protein, the disorganization of the axonal cytoskeleton, and the accumulation of autophagic vacuoles and polyubiquitinated proteins. Furthermore, MOCA deficiency causes an alteration in the actin cytoskeleton and the formation of cofilin-containing rod-like structures. The dystrophic axons show functional abnormalities, including impaired axonal transport. These findings demonstrate that MOCA is required for maintaining the functional integrity of axons and define a model for the steps leading to axonal degeneration. PMID:19129390

  18. 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. PMID:25982723

  19. Toxic effects of a modified montmorillonite clay on the human intestinal cell line Caco-2.

    PubMed

    Maisanaba, Sara; Gutiérrez-Praena, Daniel; Pichardo, Silvia; Moreno, F Javier; Jordá, María; Cameán, Ana M; Aucejo, Susana; Jos, Angeles

    2014-06-01

    The incorporation of the natural mineral clay montmorillonite into polymeric systems enhances their barrier properties as well as their thermal and mechanical resistance, making them suitable for a wide range of industrial applications, e.g., in the food industry. Considering humans could easily be exposed to these clays due to migration into food, toxicological and health effects of clay exposure should be studied. In the present work, the cytotoxic effects induced by two different clays (the unmodified clay Cloisite(®) Na(+) , and the organically modified Cloisite(®) 30B) on Caco-2 cells were studied after 24 and 48 h of exposure. The basal cytotoxicity endpoints assessed were total protein content, neutral red uptake and a tetrazolium salt reduction. Our results showed that only Cloisite(®) 30B induced toxic effects. Therefore, the effects of subcytotoxic concentrations of this clay on the generation of intracellular reactive oxygen species, glutathione content and DNA damage (comet assay) were investigated. Results indicate that oxidative stress may be implicated in the toxicity induced by Closite(®) 30B, in regards of the increases in intracellular reactive oxygen species production and glutathione content at the highest concentration assayed, while no damage was observed in DNA. The most remarkable morphological alterations observed were dilated cisternae edge in the Golgi apparatus and nucleolar segregation, suggesting impairment in the secretory functions, which could be related to inhibition in the synthesis of proteins.

  20. 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

  1. Intracoronary transplantation of genetically modified mesenchymal stem cells, a novel method to close muscular ventricular septal defects.

    PubMed

    Yang, Qing; Zhang, Juqian; Jiang, Jian

    2011-10-01

    Muscular ventricular septal defects remain a challenge despite the progress in surgical and interventional closure of ventricular septal defects. Our hypothesis was inspired by the fact that more than two thirds of children with muscular ventricular septal defects experienced spontaneous closure. Therefore, we intend to induce the spontaneous closure of muscular ventricular septal defects by means of targeted intracoronary injection of mesenchymal stem cells which are genetically modified to enhance myocardial hypertrophy. The transplantation of bone marrow derived cells has been observed to be effective in improving tissue recovery and ameliorating cardiac function in patients and animal models with ischemic heart disease, acute myocarditis and dilated cardiomyopathy. We expect that the targeted intracoronary transplantation of genetically modified mesenchymal stem cells could enhance the tissue generation and myocardial hypertrophy simultaneously, which may lead to the closure of muscular ventricular septal defects in a way that imitate the spontaneous closure of ventricular septal defects.

  2. 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.

  3. 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. PMID:26572430

  4. The B-cell tumor–associated antigen ROR1 can be targeted with T cells modified to express a ROR1-specific chimeric antigen receptor

    PubMed Central

    Schmitt, Thomas M.; Baskar, Sivasubramanian; Lupo-Stanghellini, Maria Teresa; Nishida, Tetsuya; Yamamoto, Tori N.; Bleakley, Marie; Turtle, Cameron J.; Chang, Wen-Chung; Greisman, Harvey A.; Wood, Brent; Maloney, David G.; Jensen, Michael C.; Rader, Christoph; Riddell, Stanley R.

    2010-01-01

    Monoclonal antibodies and T cells modified to express chimeric antigen receptors specific for B-cell lineage surface molecules such as CD20 exert antitumor activity in B-cell malignancies, but deplete normal B cells. The receptor tyrosine kinase-like orphan receptor 1 (ROR1) was identified as a highly expressed gene in B-cell chronic lymphocytic leukemia (B-CLL), but not normal B cells, suggesting it may serve as a tumor-specific target for therapy. We analyzed ROR1-expression in normal nonhematopoietic and hematopoietic cells including B-cell precursors, and in hematopoietic malignancies. ROR1 has characteristics of an oncofetal gene and is expressed in undifferentiated embryonic stem cells, B-CLL and mantle cell lymphoma, but not in major adult tissues apart from low levels in adipose tissue and at an early stage of B-cell development. We constructed a ROR1-specific chimeric antigen receptor that when expressed in T cells from healthy donors or CLL patients conferred specific recognition of primary B-CLL and mantle cell lymphoma, including rare drug effluxing chemotherapy resistant tumor cells that have been implicated in maintaining the malignancy, but not mature normal B cells. T-cell therapies targeting ROR1 may be effective in B-CLL and other ROR1-positive tumors. However, the expression of ROR1 on some normal tissues suggests the potential for toxi-city to subsets of normal cells. PMID:20702778

  5. Regulation of the complement-mediated elimination of red blood cells modified with biotin and streptavidin.

    PubMed

    Muzykantov, V R; Murciano, J C; Taylor, R P; Atochina, E N; Herraez, A

    1996-10-01

    Red blood cells (RBC) modified with biotin and streptavidin (SA) present an interesting potential drug delivery system. Biotinylation and SA attachment, however, alter the biocompatibility of RBC. We have reported that polyvalent SA attachment induces lysis of biotinylated RBC (b-RBC) by homologous complement via the alternative pathway. Lysis occurs due to inactivation of the membrane regulators of complement, DAF and CD59, cross-linked by SA. However, monovalent SA attachment does not induce lysis. On the basis of these findings we hypothesized that reduction of the biotin surface density on b-RBC would allow for monovalent SA attachment to b-RBC and that such SA/b-RBC should then be stable in the circulation. In the present work we injected into rats several different radiolabeled RBC probes: rat RBC biotinylated to varying degrees (bn-RBC, where bn represents the input micromolar concentration of biotinylating agent), as well as SA/bn-RBC. Extensively biotinylated rat RBC (b700-RBC, stable in serum in vitro) were rapidly cleared from the bloodstream. We further found that extensively biotinylated human b1000-RBC bound C3b from serum in vitro without detectable lysis, and that rat b700-RBC bound to isolated macrophages in a complement-dependent fashion. Therefore, nonlytic C3b flxation and uptake of C3b-carrying b700-RBC by macrophages appears to be the mechanism leading to clearance of b700-RBC in vivo. Moderately biotinylated RBC (b70-RBC and b240-RBC) were stable in serum in vitro. SA attachment to b240-RBC led to their rapid lysis in serum in vitro, lysis in the bloodstream, and clearance by the liver and spleen. SA attachment to b70-RBC led to fast elimination of SA/b70-RBC from the bloodstream, while in vitro SA/ b70-RBC were stable in serum. Modestly biotinylated RBC (b22-RBC) demonstrated only marginally decreased 60-min survival in the bloodstream regardless of SA attachment. Our in vitro studies indicate that b23-RBC bound approximately 10(5) SA

  6. Library synthesis, screening, and discovery of modified Zinc(II)-Bis(dipicolylamine) probe for enhanced molecular imaging of cell death.

    PubMed

    Plaunt, Adam J; Harmatys, Kara M; Wolter, William R; Suckow, Mark A; Smith, Bradley D

    2014-04-16

    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.

  7. Armed oncolytic virus enhances immune functions of chimeric antigen receptor-modified T cells in solid tumors.

    PubMed

    Nishio, Nobuhiro; Diaconu, Iulia; Liu, Hao; Cerullo, Vincenzo; Caruana, Ignazio; Hoyos, Valentina; Bouchier-Hayes, Lisa; Savoldo, Barbara; Dotti, Gianpietro

    2014-09-15

    The clinical efficacy of chimeric antigen receptor (CAR)-redirected T cells remains marginal in solid tumors compared with leukemias. Failures have been attributed to insufficient T-cell migration and to the highly immunosuppressive milieu of solid tumors. To overcome these obstacles, we have combined CAR-T cells with an oncolytic virus armed with the chemokine RANTES and the cytokine IL15, reasoning that the modified oncolytic virus will both have a direct lytic effect on infected malignant cells and facilitate migration and survival of CAR-T cells. Using neuroblastoma as a tumor model, we found that the adenovirus Ad5Δ24 exerted a potent, dose-dependent, cytotoxic effect on tumor cells, whereas CAR-T cells specific for the tumor antigen GD2 (GD2.CAR-T cells) were not damaged. When used in combination, Ad5Δ24 directly accelerated the caspase pathways in tumor cells exposed to CAR-T cells, whereas the intratumoral release of both RANTES and IL15 attracted CAR-T cells and promoted their local survival, respectively, increasing the overall survival of tumor-bearing mice. These preclinical data support the use of this innovative biologic platform of immunotherapy for solid tumors. Cancer Res; 74(18); 5195-205. ©2014 AACR.

  8. Development of Genetically Modified Chinese Hamster Ovary Host Cells for the Enhancement of Recombinant Tissue Plasminogen Activator Expression

    PubMed Central

    Rahimpour, Azam; Ahani, Roshanak; Najaei, Azita; Adeli, Ahmad; Barkhordari, Farzaneh; Mahboudi, Fereidoun

    2016-01-01

    Background Chinese hamster ovary (CHO) cells are the most commonly used host system for the expression of high quality recombinant proteins. However, the development of stable, high-yielding CHO cell lines is a major bottleneck in the industrial manufacturing of therapeutic proteins. Therefore, different strategies such as the generation of more efficient expression vectors and establishment of genetically engineered host cells have been employed to increase the efficiency of cell line development. In order to examine the possibility of generating improved CHO host cells, cell line engineering approaches were developed based on ceramide transfer protein (CERT), and X-box binding protein 1s (XBP1s). Methods CHO cells were transfected with CERT S132A, a mutant variant of CERT which is resistant to phosphorylation, or XBP1s expression plasmids, and then stable cell pools were generated. Transient expression of t-PA was examined in engineered cell pools in comparison to un-modified CHO host cells. Results Overexpression of CERT S132A led to the enhancement of recombinant tissue plasminogen activator (t-PA) expression in transient expression by 50%. On the other hand, it was observed that the ectopic expression of the XBP1s, did not improve the t-PA expression level. Conclusion The results obtained in this study indicate successful development of the improved CHO host cells through CERT S132A overexpression. PMID:27547109

  9. 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.

  10. 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.

  11. 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. PMID:27226436

  12. 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.

  13. Modifying the NH2 and COOH termini of aquaporin-5: effects on localization in polarized epithelial cells.

    PubMed

    Wellner, Robert B; Hong, Sohee; Cotrim, Ana P; Swaim, William D; Baum, Bruce J

    2005-01-01

    To reengineer polarized epithelial cell functions directly in situ, or ex vivo in the fabrication of an artificial organ, it is necessary to understand mechanisms that account for polarized membrane sorting. We have used the aquaporins (AQPs), a family of homotetrameric water channel proteins, as model membrane proteins for this purpose. AQP monomers contain six transmembrane-spanning domains linked by five interconnecting loops, with the NH2 and COOH termini residing in the cytosol. AQP5 is localized in the apical membranes of several different epithelia in vivo, and in stably transfected MDCK-II cells grown as a polarized monolayer. We wished to identify a structural region(s) within rat AQP5 (rAQP5) important for apical localization, and to study the MDCK-II cell localization of rAQP5s modified in either their NH2 or COOH terminus. We show that the NH2- terminal region does not play a major role in apical localization as deletion of the NH2 terminus produced a modified rAQP5 construct (AQP5-NT(del)) that was stably expressed and localized primarily to the apical membranes of MDCK-II cells. Attachment of a FLAG epitope to the NH2 terminus of AQP5 (AQP5(flag) construct) also did not perturb apical localization. In addition, we found that the exchange of NH2-terminal regions between rAQP5 and human AQP1 (hAQP1; a nonpolarized AQP isoform) produced a modified rAQP5 construct (AQP5-1NT) and a modified hAQP1 construct (AQP1-5NT), each of which localized as the parental AQP (apically, and to both apical and basolateral membranes, respectively). In contrast, we found that deletion of the COOH terminus resulted in a modified rAQP5 construct (AQP5-CT(del)) that was unstably expressed and localized to intracellular site(s) in MDCK-II cells. Substitution of the COOH terminus of AQP1 with the COOH terminus of AQP5 also produced a construct (AQP1-5CT) transiently expressed in intracellular compartment(s). However, substitution of the COOH terminus of rAQP5 with the COOH

  14. Genomic Modifiers of Natural Killer Cells, Immune Responsiveness and Lymphoid Tissue Remodeling Together Increase Host Resistance to Viral Infection.

    PubMed

    Gillespie, Alyssa Lundgren; Teoh, Jeffrey; 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

    2016-02-01

    The MHC class I D(k) molecule supplies vital host resistance during murine cytomegalovirus (MCMV) infection. Natural killer (NK) cells expressing the Ly49G2 inhibitory receptor, which specifically binds D(k), are required to control viral spread. The extent of D(k)-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 D(k)-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

  15. Genomic Modifiers of Natural Killer Cells, Immune Responsiveness and Lymphoid Tissue Remodeling Together Increase Host Resistance to Viral Infection.

    PubMed

    Gillespie, Alyssa Lundgren; Teoh, Jeffrey; 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

    2016-02-01

    The MHC class I D(k) molecule supplies vital host resistance during murine cytomegalovirus (MCMV) infection. Natural killer (NK) cells expressing the Ly49G2 inhibitory receptor, which specifically binds D(k), are required to control viral spread. The extent of D(k)-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 D(k)-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.

  16. 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. PMID:26952437

  17. A modified approach to identification of the sickle cell anemia mutation by means of allele-specific polymerase chain reaction.

    PubMed

    Birikh, K R; Plutalov, O V; Schwartz, E I; Devi, P S; Berlin, Y A

    1992-01-01

    The allele-specific PCR approach has been modified by introducing a second mismatch at the 3'-penultimate link of the primer and used to identify the sickle cell anemia mutation (A-->T transversion in the sixth codon of the human beta-globin gene causing Glu-->Val substitution in the protein), thus obviating the problem of an interpretationally ambiguous 3'-terminal mismatch including T residue. PMID:1301951

  18. A modified approach to identification of the sickle cell anemia mutation by means of allele-specific polymerase chain reaction.

    PubMed

    Birikh, K R; Plutalov, O V; Schwartz, E I; Devi, P S; Berlin, Y A

    1992-01-01

    The allele-specific PCR approach has been modified by introducing a second mismatch at the 3'-penultimate link of the primer and used to identify the sickle cell anemia mutation (A-->T transversion in the sixth codon of the human beta-globin gene causing Glu-->Val substitution in the protein), thus obviating the problem of an interpretationally ambiguous 3'-terminal mismatch including T residue.

  19. Regulation of Noxa-mediated apoptosis in Helicobacter pylori–infected gastric epithelial cells

    PubMed Central

    Rath, Suvasmita; Das, Lopamudra; Kokate, Shrikant Babanrao; Pratheek, B. M.; Chattopadhyay, Subhasis; Goswami, Chandan; Chattopadhyay, Ranajoy; Crowe, Sheila Eileen; Bhattacharyya, Asima

    2015-01-01

    Helicobacter pylori induces the antiapoptotic protein myeloid cell leukemia 1 (Mcl1) in human gastric epithelial cells (GECs). Apoptosis of oncogenic protein Mcl1-expressing cells is mainly regulated by Noxa-mediated degradation of Mcl1. We wanted to elucidate the status of Noxa in H. pylori–infected GECs. For this, various GECs such as AGS, MKN45, and KATO III were either infected with H. pylori or left uninfected. The effect of infection was examined by immunoblotting, immunoprecipitation, chromatin immunoprecipitation assay, in vitro binding assay, flow cytometry, and confocal microscopy. Infected GECs, surgical samples collected from patients with gastric adenocarcinoma as well as biopsy samples from patients infected with H. pylori showed significant up-regulation of both Mcl1 and Noxa compared with noninfected samples. Coexistence of Mcl1 and Noxa was indicative of an impaired Mcl-Noxa interaction. We proved that Noxa was phosphorylated at Ser13 residue by JNK in infected GECs, which caused cytoplasmic retention of Noxa. JNK inhibition enhanced Mcl1-Noxa interaction in the mitochondrial fraction of infected cells, whereas overexpression of nonphosphorylatable Noxa resulted in enhanced mitochondria-mediated apoptosis in the infected epithelium. Because phosphorylation-dephosphorylation can regulate the apoptotic function of Noxa, this could be a potential target molecule for future treatment approaches for H. pylori–induced gastric cancer.—Rath, S., Das, L., Kokate, S. B., Pratheek, B. M., Chattopadhyay, S., Goswami, C., Chattopadhyay, R., Crowe, S. E., Bhattacharyya, A. Regulation of Noxa-mediated apoptosis in Helicobacter pylori–infected gastric epithelial cells. PMID:25404713

  20. 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

    2015-03-01

    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. PMID:25658033

  1. 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.

  2. Overexpression of the carbohydrate binding module of strawberry expansin2 in Arabidopsis thaliana modifies plant growth and cell wall metabolism.

    PubMed

    Nardi, Cristina F; Villarreal, Natalia M; Rossi, Franco R; Martínez, Santiago; Martínez, Gustavo A; Civello, Pedro M

    2015-05-01

    Several cell wall enzymes are carbohydrate active enzymes that contain a putative Carbohydrate Binding Module (CBM) in their structures. The main function of these non-catalitic modules is to facilitate the interaction between the enzyme and its substrate. Expansins are non-hydrolytic proteins present in the cell wall, and their structure includes a CBM in the C-terminal that bind to cell wall polymers such as cellulose, hemicelluloses and pectins. We studied the ability of the Expansin2 CBM (CBMFaEXP2) from strawberry (Fragaria x ananassa, Duch) to modify the cell wall of Arabidopsis thaliana. Plants overexpressing CBMFaEXP2 were characterized phenotypically and biochemically. Transgenic plants were taller than wild type, possibly owing to a faster growth of the main stem. Cell walls of CBMFaEXP2-expressing plants were thicker and contained higher amount of pectins. Lower activity of a set of enzymes involved in cell wall degradation (PG, β-Gal, β-Xyl) was found, and the expression of the corresponding genes (AtPG, Atβ-Gal, Atβ-Xyl5) was reduced also. In addition, a decrease in the expression of two A. thaliana Expansin genes (AtEXP5 and AtEXP8) was observed. Transgenic plants were more resistant to Botrytis cinerea infection than wild type, possibly as a consequence of higher cell wall integrity. Our results support the hypothesis that the overexpression of a putative CBM is able to modify plant cell wall structure leading to modulation of wall loosening and plant growth. These findings might offer a tool to controlling physiological processes where cell wall disassembly is relevant, such as fruit softening. PMID:25837738

  3. Overexpression of the carbohydrate binding module of strawberry expansin2 in Arabidopsis thaliana modifies plant growth and cell wall metabolism.

    PubMed

    Nardi, Cristina F; Villarreal, Natalia M; Rossi, Franco R; Martínez, Santiago; Martínez, Gustavo A; Civello, Pedro M

    2015-05-01

    Several cell wall enzymes are carbohydrate active enzymes that contain a putative Carbohydrate Binding Module (CBM) in their structures. The main function of these non-catalitic modules is to facilitate the interaction between the enzyme and its substrate. Expansins are non-hydrolytic proteins present in the cell wall, and their structure includes a CBM in the C-terminal that bind to cell wall polymers such as cellulose, hemicelluloses and pectins. We studied the ability of the Expansin2 CBM (CBMFaEXP2) from strawberry (Fragaria x ananassa, Duch) to modify the cell wall of Arabidopsis thaliana. Plants overexpressing CBMFaEXP2 were characterized phenotypically and biochemically. Transgenic plants were taller than wild type, possibly owing to a faster growth of the main stem. Cell walls of CBMFaEXP2-expressing plants were thicker and contained higher amount of pectins. Lower activity of a set of enzymes involved in cell wall degradation (PG, β-Gal, β-Xyl) was found, and the expression of the corresponding genes (AtPG, Atβ-Gal, Atβ-Xyl5) was reduced also. In addition, a decrease in the expression of two A. thaliana Expansin genes (AtEXP5 and AtEXP8) was observed. Transgenic plants were more resistant to Botrytis cinerea infection than wild type, possibly as a consequence of higher cell wall integrity. Our results support the hypothesis that the overexpression of a putative CBM is able to modify plant cell wall structure leading to modulation of wall loosening and plant growth. These findings might offer a tool to controlling physiological processes where cell wall disassembly is relevant, such as fruit softening.

  4. The use of substrate materials and topography to modify growth patterns and rates of differentiation of muscle cells.

    PubMed

    Murray, L M; Nock, V; Evans, J J; Alkaisi, M M

    2016-07-01

    Cells are cultured on platforms made of a variety of materials with selected topographies during studies of cell response and behavior. Understanding the effects of substrates is essential for such applications as developing effective interfaces between body cells and implanted materials and devices. In this study, the effects of substrate surface properties on cell differentiation and alignment on C2C12 myoblasts cultured on conventional or fabricated polymeric cell culture substrates were investigated. Comparisons were made between cells cultured on tissue culture grade polystyrene (TCPS), glass, Permanox, and cured polydimethylsiloxane (PDMS) substrates. Fluorescent immunohistochemistry of cell markers was used to analyse the extent of differentiation. Alignment and guidance of cell growth and spread were studied using patterned platforms. Gratings were made on polystyrene (PS) and PDMS and differentiation was facilitated after 5 days by media exchange. Differences in cell morphology were observed between cells cultured on TCPS and PDMS substrates. Fully differentiated myotubes were observed in highest numbers on TCPS substrates and were non-detectable on PDMS substrates in the time frame of 144 h. Muscle cell alignment and their differentiation followed along the grating patterns on PS and elongated along the pattern length. On the other hand, on PDMS cells formed sheets of tissue and peeled from the substrate. We have revealed the potential for the combinations of surface materials and topography on cell behavior to induce accelerated differentiation and coordinated alignment. The results demonstrate that culture environment can be designed or engineered to modify or regulate muscle cell functions. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1638-1645, 2016.

  5. 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. PMID:27106502

  6. 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

  7. 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

    2015-05-01

    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.

  8. Designed Stem Cell Aggregates: Enhanced Biological Functions of Human Mesenchymal Stem-Cell Aggregates Incorporating E-Cadherin-Modified PLGA Microparticles (Adv. Healthcare Mater. 15/2016).

    PubMed

    Zhang, Yan; Mao, Hongli; Gao, Chao; Li, Suhua; Shuai, Qizhi; Xu, Jianbin; Xu, Ke; Cao, Lei; Lang, Ren; Gu, Zhongwei; Akaike, Toshihiro; Yang, Jun

    2016-08-01

    E-cadherin-modified poly(lactic-co-glycolic acid) (hE-cad-PLGA) microparticles were fabricated and then mediated the 3D cell aggregates of human mesenchymal stem cells (MSCs) on page 1949 by Jun Yang and co-workers. The hE-cad-Fc matrix and the PLGA microparticles synergistically regulate the proliferation and bioactive factors secretions of MSCs by activating EGFR, AKT and ERK1/2 signaling pathways. The hE-cad-PLGA microparticles offer a novel route to expand multipotent stem cell-based clinical applications. PMID:27511954

  9. Automatic recognition of myeloma cells in microscopic images using bottleneck algorithm, modified watershed and SVM classifier.

    PubMed

    Saeedizadeh, Z; Mehri Dehnavi, A; Talebi, A; Rabbani, H; Sarrafzadeh, O; Vard, A

    2015-01-01

    Plasma cells are developed from B lymphocytes, a type of white blood cells that is generated in the bone marrow. The plasma cells produce antibodies to fight with bacteria and viruses and stop infection and disease. Multiple myeloma is a cancer of plasma cells that collections of abnormal plasma cells (myeloma cells) accumulate in the bone marrow. The definitive diagnosis of multiple myeloma is done by searching for myeloma cells in the bone marrow slides through a microscope. Diagnosis of myeloma cells from bone marrow smears is a subjective and time-consuming task for pathologists. Also, because of depending on final decision on human eye and opinion, error risk in decision may occur. Sometimes, existence of infection in body causes plasma cell's increment which could be diagnosed wrongly as multiple myeloma. The computer diagnostic process will reduce the diagnostic time and also can be worked as a second opinion for pathologists. This study presents a computer-aided diagnostic method for myeloma cells diagnosis from bone marrow smears. At first, white blood cells consist of plasma cells and other marrow cells are separated from the red blood cells and background. Then, plasma cells are detected from other marrow cells by feature extraction and series of decision rules. Finally, normal plasma cells and myeloma cells could be classified easily by a classifier. This algorithm is applied on 50 digital images that are provided from bone marrow aspiration smears. These images contain 678 cells: 132 normal plasma cells, 256 myeloma cells and 290 other types of marrow cells. Applying the computer-aided diagnostic method for identifying myeloma cells on provided database showed a sensitivity of 96.52%; specificity of 93.04% and precision of 95.28%. PMID:26457371

  10. 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). PMID:25863381

  11. [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

  12. Carbon foam anode modified by urea and its higher electrochemical performance in marine benthic microbial fuel cell

    NASA Astrophysics Data System (ADS)

    Fu, Yubin; Lu, Zhikai; Zai, Xuerong; Wang, Jian

    2015-08-01

    Electrode materials have an important effect on the property of microbial fuel cell (MFC). Carbon foam is utilized as an anode and further modified by urea to improve its performance in marine benthic microbial fuel cell (BMFC) with higher voltage and output power. The electrochemical properties of plain carbon foam (PC) and urea-modified carbon foam (UC) are measured respectively. Results show that the UC obtains better wettability after its modification and higher anti-polarization ability than the PC. A novel phenomenon has been found that the electrical potential of the modified UC anode is nearly 100 mV lower than that of the PC, reaching -570 ±10 mV ( vs. SCE), and that it also has a much higher electron transfer kinetic activity, reaching 9399.4 mW m-2, which is 566.2-fold higher than that from plain graphite anode (PG). The fuel cell containing the UC anode has the maximum power density (256.0 mW m-2) among the three different BMFCs. Urea would enhance the bacteria biofilm formation with a more diverse microbial community and maintain more electrons, leading to a lower anodic redox potential and higher power output. The paper primarily analyzes why the electrical potential of the modified anode becomes much lower than that of others after urea modification. These results can be utilized to construct a novel BMFC with higher output power and to design the conditioner of voltage booster with a higher conversion ratio. Finally, the carbon foam with a bigger pore size would be a potential anodic material in conventional MFC.

  13. The role of the cell cycle in the cellular uptake of folate-modified poly(l-amino acid) micelles in a cell population

    NASA Astrophysics Data System (ADS)

    Tang, Jihui; Liu, Ziwei; Ji, Fenqi; Li, Yao; Liu, Junjie; Song, Jian; Li, Jun; Zhou, Jianping

    2015-12-01

    Nanoparticles are widely recognized as a vehicle for tumor-targeted therapies. There are many factors that can influence the uptake of nanoparticles, such as the size of the nanoparticles, and/or their shape, elasticity, surface charge and even the cell cycle phase. However, the influence of the cell cycle on the active targeting of a drug delivery system has been unknown until now. In this study, we initially investigated the folate receptor α (FR-α) expression in different phases of HeLa cells by flow cytometric and immunocytochemical methods. The results obtained showed that FR-α expression was cell cycle-dependent, i.e. the S cells' folate receptor expression was the highest as the cell progressed through its cycle. Then, we used folate modified poly(l-amino acid) micelles (FA-PM) as an example to investigate the influence of the cell cycle on the active targeting drug delivery system. The results obtained indicated that the uptake of FA-PM by cells was influenced by the cell cycle phase, and the S cells took up the greatest number of folate conjugated nanoparticles. Our findings suggest that future studies on ligand-mediated active targeting preparations should consider the cell cycle, especially when this system is used for a cell cycle-specific drug.

  14. Genetically Modified T Cells in Treating Patients With Stage III-IV Non-small Cell Lung Cancer or Mesothelioma

    ClinicalTrials.gov

    2016-11-01

    Advanced Pleural Malignant Mesothelioma; HLA-A*0201 Positive Cells Present; Recurrent Non-Small Cell Lung Carcinoma; Recurrent Pleural Malignant Mesothelioma; 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

  15. 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.

  16. 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

  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.

    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

  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. Differences in Virulence of Marine and Freshwater Isolates of Viral Hemorrhagic Septicemia Virus In Vivo Correlate with In Vitro Ability To Infect Gill Epithelial Cells and Macrophages of Rainbow Trout (Oncorhynchus mykiss)▿

    PubMed Central

    Brudeseth, Bjørn E.; Skall, Helle F.; Evensen, Øystein

    2008-01-01

    Two strains of viral hemorrhagic septicemia virus (VHSV) with known different virulence characteristics in vivo were studied (by a time course approach) for their abilities to infect and translocate across a primary culture of gill epithelial cells (GEC) of rainbow trout (RBT; Oncorhynchus mykiss). The strains included one low-virulence marine VHSV (ma-VHSV) strain, ma-1p8, and a highly pathogenic freshwater VHSV (fw-VHSV) strain, fw-DK-3592B. Infectivities toward trout head kidney macrophages were also studied (by a time course method), and differences in in vivo virulence were reconfirmed, the aim being to determine any correlation between in vivo virulence and in vitro infectivity. The in vitro studies showed that the fw-VHSV isolate infected and caused a cytotoxic effect in monolayers of GEC (demonstrating virulence) at an early time point (2 h postinoculation) and that the same virus strain had translocated over a confluent, polarized GEC layer by 2 h postinoculation. The marine isolate did not infect monolayers of GEC, and delayed translocation across polarized GEC was seen by 48 h postinoculation. Primary cultures of head kidney macrophages were also infected with fw-VHSV, with a maximum of 9.5% virus-positive cells by 3 days postinfection, while for the ma-VHSV strain, only 0.5% of the macrophages were positive after 3 days of culture. In vivo studies showed that the fw-VHSV strain was highly virulent for RBT fry and caused high mortality, with classical features of viral hemorrhagic septicemia. The ma-VHSV showed a very low level of virulence (only one pool of samples from the dead fish was VHSV positive). This study has shown that the differences in virulence between marine and freshwater strains of VHSV following the in vivo infection of RBT correlate with in vitro abilities to infect primary cultures of GEC and head kidney macrophages of the same species. PMID:18753199

  1. Antigen presentation by chemically modified splenocytes induces antigen- specific T cell unresponsiveness in vitro and in vivo

    PubMed Central

    1987-01-01

    We investigated the antigen specificity and presentation requirements for inactivation of T lymphocytes in vitro and in vivo. In vitro studies revealed that splenocytes treated with the crosslinker 1-ethyl- 3-(3-dimethylaminopropyl)-carbodiimide (ECDI) and soluble antigen fragments failed to stimulate significant proliferation by normal pigeon cytochrome c-specific T cell clones, suggesting that the chemical treatment inactivated full antigen presentation function. However, T cell clones exposed to ECDI-treated splenocytes and antigen in vitro were rendered unresponsive for at least 8 d to subsequent antigen stimulation with normal presenting cells. As predicted by the in vitro results, specific T cell unresponsiveness was also induced in vivo in B10.A mice injected intravenously with B10.A, but not B10.A(4R), splenocytes coupled with pigeon cytochrome c via ECDI. The antigen and MHC specificity of the induction of this T cell unresponsiveness in vitro and in vivo was identical to that required for T cell activation. These results suggest that nonmitogenic T cell recognition of antigen/MHC on ECDI-modified APCs results in the functional inactivation of T cell clones. PMID:3029267

  2. Efficient delivery and functional expression of transfected modified mRNA in human embryonic stem cell-derived retinal pigmented epithelial cells.

    PubMed

    Hansson, Magnus L; Albert, Silvia; González Somermeyer, Louisa; Peco, Rubén; Mejía-Ramírez, Eva; Montserrat, Núria; Izpisua Belmonte, Juan Carlos

    2015-02-27

    Gene- and cell-based therapies are promising strategies for the treatment of degenerative retinal diseases such as age-related macular degeneration, Stargardt disease, and retinitis pigmentosa. Cellular engineering before transplantation may allow the delivery of cellular factors that can promote functional improvements, such as increased engraftment or survival of transplanted cells. A current challenge in traditional DNA-based vector transfection is to find a delivery system that is both safe and efficient, but using mRNA as an alternative to DNA can circumvent these major roadblocks. In this study, we show that both unmodified and modified mRNA can be delivered to retinal pigmented epithelial (RPE) cells with a high efficiency compared with conventional plasmid delivery systems. On the other hand, administration of unmodified mRNA induced a strong innate immune response that was almost absent when using modified mRNA. Importantly, transfection of mRNA encoding a key regulator of RPE gene expression, microphthalmia-associated transcription factor (MITF), confirmed the functionality of the delivered mRNA. Immunostaining showed that transfection with either type of mRNA led to the expression of roughly equal levels of MITF, primarily localized in the nucleus. Despite these findings, quantitative RT-PCR analyses showed that the activation of the expression of MITF target genes was higher following transfection with modified mRNA compared with unmodified mRNA. Our findings, therefore, show that modified mRNA transfection can be applied to human embryonic stem cell-derived RPE cells and that the method is safe, efficient, and functional.

  3. Efficient delivery and functional expression of transfected modified mRNA in human embryonic stem cell-derived retinal pigmented epithelial cells.

    PubMed

    Hansson, Magnus L; Albert, Silvia; González Somermeyer, Louisa; Peco, Rubén; Mejía-Ramírez, Eva; Montserrat, Núria; Izpisua Belmonte, Juan Carlos

    2015-02-27

    Gene- and cell-based therapies are promising strategies for the treatment of degenerative retinal diseases such as age-related macular degeneration, Stargardt disease, and retinitis pigmentosa. Cellular engineering before transplantation may allow the delivery of cellular factors that can promote functional improvements, such as increased engraftment or survival of transplanted cells. A current challenge in traditional DNA-based vector transfection is to find a delivery system that is both safe and efficient, but using mRNA as an alternative to DNA can circumvent these major roadblocks. In this study, we show that both unmodified and modified mRNA can be delivered to retinal pigmented epithelial (RPE) cells with a high efficiency compared with conventional plasmid delivery systems. On the other hand, administration of unmodified mRNA induced a strong innate immune response that was almost absent when using modified mRNA. Importantly, transfection of mRNA encoding a key regulator of RPE gene expression, microphthalmia-associated transcription factor (MITF), confirmed the functionality of the delivered mRNA. Immunostaining showed that transfection with either type of mRNA led to the expression of roughly equal levels of MITF, primarily localized in the nucleus. Despite these findings, quantitative RT-PCR analyses showed that the activation of the expression of MITF target genes was higher following transfection with modified mRNA compared with unmodified mRNA. Our findings, therefore, show that modified mRNA transfection can be applied to human embryonic stem cell-derived RPE cells and that the method is safe, efficient, and functional. PMID:25555917

  4. Guanine-modified inhibitory oligonucleotides efficiently impair TLR7- and TLR9-mediated immune responses of human immune cells.

    PubMed

    Römmler, Franziska; Hammel, Monika; Waldhuber, Anna; Müller, Tina; Jurk, Marion; Uhlmann, Eugen; Wagner, Hermann; Vollmer, Jörg; Miethke, Thomas

    2015-01-01

    Activation of TLR7 and TLR9 by endogenous RNA- or DNA-containing ligands, respectively, is thought to contribute to the complicated pathophysiology of systemic lupus erythematosus (SLE). These ligands induce the release of type-I interferons by plasmacytoid dendritic cells and autoreactive antibodies by B-cells, both responses being key events in perpetuating SLE. We recently described the development of inhibitory oligonucleotides (INH-ODN), which are characterized by a phosphorothioate backbone, a CC(T)XXX3-5GGG motif and a chemical modification of the G-quartet to avoid the formation of higher order structures via intermolecular G-tetrads. These INH-ODNs were equally or significantly more efficient to impair TLR7- and TLR9-stimulated murine B-cells, macrophages, conventional and plasmacytoid dendritic cells than the parent INH-ODN 2088, which lacks G-modification. Here, we evaluate the inhibitory/therapeutic potential of our set of G-modified INH-ODN on human immune cells. We report the novel finding that G-modified INH-ODNs efficiently inhibited the release of IFN-α by PBMC stimulated either with the TLR7-ligand oligoribonucleotide (ORN) 22075 or the TLR9-ligand CpG-ODN 2216. G-modification of INH-ODNs significantly improved inhibition of IL-6 release by PBMCs and purified human B-cells stimulated with the TLR7-ligand imiquimod or the TLR9-ligand CpG-ODN 2006. Furthermore, inhibition of B-cell activation analyzed by expression of activation markers and intracellular ATP content was significantly improved by G-modification. As observed with murine B-cells, high concentrations of INH-ODN 2088 but not of G-modified INH-ODNs stimulated IL-6 secretion by PBMCs in the absence of TLR-ligands thus limiting its blocking efficacy. In summary, G-modification of INH-ODNs improved their ability to impair TLR7- and TLR9-mediated signaling in those human immune cells which are considered as crucial in the pathophysiology of SLE.

  5. 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.

  6. tRNA Modifying Enzymes, NSUN2 and METTL1, Determine Sensitivity to 5-Fluorouracil in HeLa Cells

    PubMed Central

    Hori, Masato; Okada, Kaoru; Yazama, Futoshi; Konishi, Hiroaki; Xiao, Yegui; Qi, Guangying; Shimamoto, Fumio; Ota, Takahide; Temme, Achim; Tatsuka, Masaaki

    2014-01-01

    Nonessential tRNA modifications by methyltransferases are evolutionarily conserved and have been reported to stabilize mature tRNA molecules and prevent rapid tRNA decay (RTD). The tRNA modifying enzymes, NSUN2 and METTL1, are mammalian orthologs of yeast Trm4 and Trm8, which are required for protecting tRNA against RTD. A simultaneous overexpression of NSUN2 and METTL1 is widely observed among human cancers suggesting that targeting of both proteins provides a novel powerful strategy for cancer chemotherapy. Here, we show that combined knockdown of NSUN2 and METTL1 in HeLa cells drastically potentiate sensitivity of cells to 5-fluorouracil (5-FU) whereas heat stress of cells revealed no effects. Since NSUN2 and METTL1 are phosphorylated by Aurora-B and Akt, respectively, and their tRNA modifying activities are suppressed by phosphorylation, overexpression of constitutively dephosphorylated forms of both methyltransferases is able to suppress 5-FU sensitivity. Thus, NSUN2 and METTL1 are implicated in 5-FU sensitivity in HeLa cells. Interfering with methylation of tRNAs might provide a promising rationale to improve 5-FU chemotherapy of cancer. PMID:25233213

  7. 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

  8. Neurotrophin-3 gene modified mesenchymal stem cells promote remyelination and functional recovery in the demyelinated spinal cord of rats.

    PubMed

    Zhang, Yu-Jiao; Zhang, Wei; Lin, Cheng-Guang; Ding, Ying; Huang, Si-Fan; Wu, Jin-Lang; Li, Yan; Dong, Hongxin; Zeng, Yuan-Shan

    2012-02-15

    Multiple sclerosis (MS) is a debilitating neurodegenerative disease characterized by axonal/neuronal damage that may be caused by defective remyelination. Current therapies aim to slow the rate of degeneration, however there are no treatment options that can stop or reverse the myelin sheath damage. Bone marrow mesenchymal stem cells (MSCs) are a potential candidate for the cell implantation-targeted therapeutic strategies, but the pro-remyelination effects of MSCs when directly injected into a demyelinated cord lesion have been questioned. Neurotrophin-3 (NT-3) has been shown to serve a crucial role in the proliferation, differentiation and maturation of oligodendrocyte lineages. Here, we showed that implantation of NT-3 gene-modified MSCs via a recombinant adenoviral vector (Adv) into a region of ethidium bromide (EB)-induced demyelination in the spinal cord resulted in significant improvement of locomotor function and restoration of electrophysiological properties in rats. The morphological basis of this recovery was evidenced by robust myelin basic protein (MBP) expression and the extensive remyelination. AdvNT-3-MSC implants promote the endogenous remyelinating cells to participate directly in myelination, which was confirmed under light and electron microscopy. Our study suggested that genetically modified MSCs could be a potential therapeutic avenue for improving the efficacy of stem cell treatment for neurodegenerative diseases such as MS.

  9. 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

    2012-01-01

    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

  10. Antibody-free magnetic cell sorting of genetically modified primary human CD4+ T cells by one-step streptavidin affinity purification.

    PubMed

    Matheson, Nicholas J; Peden, Andrew A; Lehner, Paul J

    2014-01-01

    Existing methods for phenotypic selection of genetically modified mammalian cells suffer disadvantages of time, cost and scalability and, where antibodies are used to bind exogenous cell surface markers for magnetic selection, typically yield cells coated with antibody-antigen complexes and beads. To overcome these limitations we have developed a method termed Antibody-Free Magnetic Cell Sorting in which the 38 amino acid Streptavidin Binding Peptide (SBP) is displayed at the cell surface by the truncated Low Affinity Nerve Growth Receptor (LNGFRF) and used as an affinity tag for one-step selection with streptavidin-conjugated magnetic beads. Cells are released through competition with the naturally occurring vitamin biotin, free of either beads or antibody-antigen complexes and ready for culture or use in downstream applications. Antibody-Free Magnetic Cell Sorting is a rapid, cost-effective, scalable method of magnetic selection applicable to either viral transduction or transient transfection of cell lines or primary cells. We have optimised the system for enrichment of primary human CD4+ T cells expressing shRNAs and exogenous genes of interest to purities of >99%, and used it to isolate cells following Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 genome editing.

  11. 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.

  12. Modified low density lipoproteins suppress production of a platelet-derived growth factor-like protein by cultured endothelial cells.

    PubMed Central

    Fox, P L; DiCorleto, P E

    1986-01-01

    Cultured endothelial cells (EC) produce a platelet-derived growth factor-like protein (PDGF-c) that stimulates the growth of cultured cells of mesenchymal origin. We have examined the effect of native plasma low density lipoprotein (LDL) and chemically modified LDL on production of PDGF-c by EC. Acetyl-LDL, but not native LDL, suppressed the production of PDGF-c by bovine aortic EC. Half-maximal inhibition was observed at a concentration of 25-75 micrograms of cholesterol per ml, and maximal inhibition (0-25% of control) at 150 micrograms of cholesterol per ml. EC treated with acetyl-LDL showed no morphological damage, there was no change in cell number, and the effect on production of PDGF-c was substantially reversed upon removal of the acetyl-LDL. The observed inhibition of PDGF-c production was specific, since total cellular and secreted protein synthesis were unaffected by acetyl-LDL. Acetyl-LDL suppressed PDGF-c production in both bovine aortic and human umbilical vein EC, but not in rat heart EC. This cell specificity correlated with the presence of scavenger receptors as measured by degradation of 125I-labeled acetyl-LDL and uptake of fluorescently labeled acetyl-LDL. Dimethylpropanediamine-LDL, a cationic modified lipoprotein, also inhibited PDGF-c production. The inhibition by both types of modified LDL was accompanied by significant intracellular cholesterol accumulation, suggesting a role for EC lipid composition in the regulation of production of PDGF-c. PMID:3460071

  13. GE11-modified liposomes for non-small cell lung cancer targeting: preparation, ex vitro and in vivo evaluation.

    PubMed

    Cheng, Liang; Huang, Fa-Zhen; Cheng, Li-Fang; Zhu, Ya-Qin; Hu, Qing; Li, Ling; Wei, Lin; Chen, Da-Wei

    2014-01-01

    Non-small cell lung cancer (NSCLC) is a serious threat to human health, and 40%-80% of NSCLCs express high levels of epidermal growth factor receptor (EGFR). GE11 is a novel peptide and exhibits high affinity for EGFR binding. The aim of this study was to construct and evaluate GE11-modified liposomes for targeted drug delivery to EGFR-positive NSCLC. Doxorubicin, a broad-spectrum antitumor agent, was chosen as the payload. GE11 was conjugated to the distal end of DSPE-PEG2000-Mal by an addition reaction with a conjugation efficiency above 90%. Doxorubicin-loaded liposomes containing GE11 (GE11-LP/DOX) at densities ranging from 0% to 15% were prepared by combination of a thin film hydration method and a post insertion method. Irrespective of GE11 density, the physicochemical properties of these targeted liposomes, including particle size, zeta potential, and drug entrapment efficiency, were nearly identical. Interestingly, the cytotoxic effect of the liposomes on A549 tumor cells was closely related to GE11 density, and liposomes with 10% GE11 had the highest tumor cell killing activity and a 2.6-fold lower half maximal inhibitory concentration than that of the nontargeted counterpart (PEG-LP/DOX). Fluorescence microscopy and flow cytometry analysis revealed that GE11 significantly increased cellular uptake of the liposomes, which could be ascribed to specific EGFR-mediated endocytosis. It was found that multiple endocytic pathways were involved in entry of GE11-LP/DOX into cells, but GE11 assisted in cellular internalization mainly via the clathrin-mediated endocytosis pathway. Importantly, the GE11-modified liposomes showed enhanced accumulation and prolonged retention in tumor tissue, as evidenced by a 2.2-fold stronger mean fluorescence intensity in tumor tissue than the unmodified liposomes at 24 hours. In summary, GE11-modified liposomes may be a promising platform for targeted delivery of chemotherapeutic drugs in NSCLC. PMID:24611009

  14. GE11-modified liposomes for non-small cell lung cancer targeting: preparation, ex vitro and in vivo evaluation

    PubMed Central

    Cheng, Liang; Huang, Fa-Zhen; Cheng, Li-Fang; Zhu, Ya-Qin; Hu, Qing; Li, Ling; Wei, Lin; Chen, Da-Wei

    2014-01-01

    Non-small cell lung cancer (NSCLC) is a serious threat to human health, and 40%–80% of NSCLCs express high levels of epidermal growth factor receptor (EGFR). GE11 is a novel peptide and exhibits high affinity for EGFR binding. The aim of this study was to construct and evaluate GE11-modified liposomes for targeted drug delivery to EGFR-positive NSCLC. Doxorubicin, a broad-spectrum antitumor agent, was chosen as the payload. GE11 was conjugated to the distal end of DSPE-PEG2000-Mal by an addition reaction with a conjugation efficiency above 90%. Doxorubicin-loaded liposomes containing GE11 (GE11-LP/DOX) at densities ranging from 0% to 15% were prepared by combination of a thin film hydration method and a post insertion method. Irrespective of GE11 density, the physicochemical properties of these targeted liposomes, including particle size, zeta potential, and drug entrapment efficiency, were nearly identical. Interestingly, the cytotoxic effect of the liposomes on A549 tumor cells was closely related to GE11 density, and liposomes with 10% GE11 had the highest tumor cell killing activity and a 2.6-fold lower half maximal inhibitory concentration than that of the nontargeted counterpart (PEG-LP/DOX). Fluorescence microscopy and flow cytometry analysis revealed that GE11 significantly increased cellular uptake of the liposomes, which could be ascribed to specific EGFR-mediated endocytosis. It was found that multiple endocytic pathways were involved in entry of GE11-LP/DOX into cells, but GE11 assisted in cellular internalization mainly via the clathrin-mediated endocytosis pathway. Importantly, the GE11-modified liposomes showed enhanced accumulation and prolonged retention in tumor tissue, as evidenced by a 2.2-fold stronger mean fluorescence intensity in tumor tissue than the unmodified liposomes at 24 hours. In summary, GE11-modified liposomes may be a promising platform for targeted delivery of chemotherapeutic drugs in NSCLC. PMID:24611009

  15. 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.

  16. Specific combinations of the chromatin-modifying enzyme modulators significantly attenuate glioblastoma cell proliferation and viability while exerting minimal effect on normal adult stem cells growth.

    PubMed

    Alexanian, Arshak R; Huang, Yi-Wen

    2015-11-01

    The discoveries of recent decade showed that all critical changes in cancer cells, such as silencing of tumor-suppressor genes and activation of oncogenes, are caused not only by genetic but also by epigenetic mechanisms. Although epigenetic changes are somatically heritable, in contrast to genetic changes, they are potentially reversible, making them good targets for therapeutic intervention. Covalent modifications of chromatin such as methylation and acetylation of histones and methylation of DNA are the important components of epigenetic machinery. In this study, we investigated the effect of different modulators of DNA and histone covalent-modifying enzymes on the proliferation and viability of normal adult stem cells, such as human bone marrow mesenchymal stem cells (hMSCs), and on malignant tumor cells, such as glioblastoma (GB) D54 cells. Results demonstrated that specific combinations of histone methyltransferases and deacetylases inhibitors significantly attenuated D54 cells viability but having only a small effect on hMSCs growth. Taken together, these studies suggest that specific combinations of histone covalent modifiers could be an effective treatment option for the most aggressive type of primary brain tumors such as glioblastoma multiforme.

  17. Cell Competition Modifies Adult Stem Cell and Tissue Population Dynamics in a JAK-STAT-Dependent Manner

    PubMed Central

    Kolahgar, Golnar; Suijkerbuijk, Saskia J.E.; Kucinski, Iwo; Poirier, Enzo Z.; Mansour, Sarah; Simons, Benjamin D.; Piddini, Eugenia

    2015-01-01

    Summary Throughout their lifetime, cells may suffer insults that reduce their fitness and disrupt their function, and it is unclear how these potentially harmful cells are managed in adult tissues. We address this question using the adult Drosophila posterior midgut as a model of homeostatic tissue and ribosomal Minute mutations to reduce fitness in groups of cells. We take a quantitative approach combining lineage tracing and biophysical modeling and address how cell competition affects stem cell and tissue population dynamics. We show that healthy cells induce clonal extinction in weak tissues, targeting both stem and differentiated cells for elimination. We also find that competition induces stem cell proliferation and self-renewal in healthy tissue, promoting selective advantage and tissue colonization. Finally, we show that winner cell proliferation is fueled by the JAK-STAT ligand Unpaired-3, produced by Minute−/+ cells in response to chronic JNK stress signaling. PMID:26212135

  18. Enhanced Biological Functions of Human Mesenchymal Stem-Cell Aggregates Incorporating E-Cadherin-Modified PLGA Microparticles.

    PubMed

    Zhang, Yan; Mao, Hongli; Gao, Chao; Li, Suhua; Shuai, Qizhi; Xu, Jianbin; Xu, Ke; Cao, Lei; Lang, Ren; Gu, Zhongwei; Akaike, Toshihiro; Yang, Jun

    2016-08-01

    Mesenchymal stem cells (MSCs) have emerged as a promising source of multipotent cells for various cell-based therapies due to their unique properties, and formation of 3D MSC aggregates has been explored as a potential strategy to enhance therapeutic efficacy. In this study, poly(lactic-co-glycolic acid) (PLGA) microparticles modified with human E-cadherin fusion protein (hE-cad-PLGA microparticles) have been fabricated and integrated with human MSCs to form 3D cell aggregates. The results show that, compared with the plain PLGA, the hE-cad-PLGA microparticles distribute within the aggregates more evenly and further result in a more significant improvement of cellular proliferation and secretion of a series of bioactive factors due to the synergistic effects from the bioactive E-cadherin fragments and the PLGA microparticles. Meanwhile, the hE-cad-PLGA microparticles incorporated in the aggregates upregulate the phosphorylation of epidermal growth factor receptors and activate the AKT and ERK1/2 signaling pathways in the MSCs. Additionally, the E-cadherin/β-catenin cellular membrane complex in the MSCs is markedly stimulated by the hE-cad-PLGA microparticles. Therefore, engineering 3D cell aggregates with hE-cad-PLGA microparticles can be a promising method for ex vivo multipotent stem-cell expansion with enhanced biological functions and may offer a novel route to expand multipotent stem-cell-based clinical applications. PMID:27245478

  19. 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.

  20. 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. PMID:26494511

  1. 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.

  2. Genetically modified anthrax lethal toxin safely delivers whole HIV protein antigens into the cytosol to induce T cell immunity

    PubMed Central

    Lu, Yichen; Friedman, Rachel; Kushner, Nicholas; Doling, Amy; Thomas, Lawrence; Touzjian, Neal; Starnbach, Michael; Lieberman, Judy

    2000-01-01

    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. PMID:10884430

  3. Nature of tumor control by permanently and transiently modified GD2 chimeric antigen receptor T cells in xenograft models of neuroblastoma.

    PubMed

    Singh, Nathan; Liu, Xiaojun; Hulitt, Jessica; Jiang, Shuguang; June, Carl H; Grupp, Stephan A; Barrett, David M; Zhao, Yangbing

    2014-11-01

    Chimeric antigen receptor (CAR) therapy has begun to demonstrate success as a novel treatment modality for hematologic malignancies. The success observed thus far has been with T cells permanently engineered to express chimeric receptors. T cells engineered using RNA electroporation represent an alternative with the potential for similar efficacy and greater safety when initially targeting novel antigens. Neuroblastoma is a common pediatric solid tumor with the potential to be targeted using immunotherapy. We performed xenograft studies in NSG mice in which we assessed the efficacy of both permanently modified and transiently modified CAR T cells directed against the neuroblastoma antigen GD2 in both local and disseminated disease models. Disease response was monitored by tumor volume measurement and histologic examination, as well as in vivo bioluminescence. RNA-modified GD2 CAR T cells mediated rapid tumor destruction when delivered locally. A single infusion of lentivirally modified GD2 CAR T cells resulted in long-term control of disseminated disease. Multiple infusions of RNA GD2 CAR T cells slowed the progression of disseminated disease and improved survival, but did not result in long-term disease control. Histologic examination revealed that the transiently modified cells were unable to significantly penetrate the tumor environment when delivered systemically, despite multiple infusions of CAR T cells. Thus, we demonstrate that RNA-modified GD2 CAR T cells can mediate effective antitumor responses in vivo, and permanently modified cells are able to control disseminated neuroblastoma in xenograft mice. Lack of long-term disease control by RNA-engineered cells resulted from an inability to penetrate the tumor microenvironment.

  4. Genetically Modified T-Cell Therapy in Treating Patients With Advanced ROR1+ Malignancies

    ClinicalTrials.gov

    2016-08-29

    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. Chitosan-modified cobalt oxide nanoparticles stimulate TNF-α-mediated apoptosis in human leukemic cells.

    PubMed

    Chattopadhyay, Sourav; Dash, Sandeep Kumar; Kar Mahapatra, Santanu; Tripathy, Satyajit; Ghosh, Totan; Das, Balaram; Das, Debasis; Pramanik, Panchanan; Roy, Somenath

    2014-03-01

    The objective of this study was to develop chitosan-based delivery of cobalt oxide nanoparticles to human leukemic cells and investigate their specific induction of apoptosis. The physicochemical properties of the chitosan-coated cobalt oxide nanoparticles were characterized using transmission electron microscopy, dynamic light scattering, X-ray diffraction, and Fourier transform infrared spectroscopy. The solubility of chitosan-coated cobalt oxide nanoparticles was higher at acidic pH, which helps to release more cobalt ions into the medium. Chitosan-coated cobalt oxide nanoparticles showed good compatibility with normal cells. However, our results showed that exposure of leukemic cells (Jurkat cells) to chitosan-coated cobalt oxide nanoparticles caused an increase in reactive oxygen species generation that was abolished by pretreatment of cells with the reactive oxygen species scavenger N-acetyl-L-cysteine. The apoptosis of Jurkat cells was confirmed by flow-cytometric analysis. Induction of TNF-α secretion was observed from stimulation of Jurkat cells with chitosan-coated cobalt oxide nanoparticles. We also tested the role of TNF-α in the induction of Jurkat cell death in the presence of TNF-α and caspase inhibitors. Treatment of leukemic cells with a blocker had a greater effect on cancer cell viability. From our findings, oxidative stress and caspase activation are involved in cancer cell death induced by chitosan-coated cobalt oxide nanoparticles.

  6. 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

    2012-04-01

    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.

  7. Detection of circulating tumor cells in prostate cancer based on carboxylated graphene oxide modified light addressable potentiometric sensor.

    PubMed

    Gu, Yajun; Ju, Cheng; Li, Yanjun; Shang, Zhiqun; Wu, Yudong; Jia, Yunfang; Niu, Yuanjie

    2015-04-15

    Circulating tumor cells (CTCs) are a group of rare cancer cells that have detached from a primary tumor and circulate in the bloodstream. Herein, light addressable potentiometric sensor (LAPS) was exploited in the label-free detection of CTCs in the prostate cancer. To this end, the mouse anti-human epithelial cell adhesion molecule (anti-EpCAM) monoclonal antibody was selected as the probe to capture CTCs according to our western blot experiments, and therefore the anti-EpCAM was immobilized on the surface of carboxylated graphene oxide (GO-COOH) modified LAPS. Spiking experiments confirmed that LAPS' voltage decreased with the increasing of CTCs' concentration both in phosphate buffer (PBS) and blood, and as few as 10 CTCs in 1ml of blood could be detected, illustrating the high sensitivity of the proposed strategy. The analysis of healthy blood samples revealed no change in electrical signal, confirming the specificity of the system. Ultraviolet-visible (UV-vis) spectroscopy, scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS) and immunofluorescent assay (IFA) were conducted to characterize GO-COOH, testify its existence on LAPS and validate CTCs' capturing by anti-EpCAM grafted on GO-COOH modified substrates. It is indicated that LAPS could be a potential platform for CTCs detection and may provide a powerful tool for downstream analysis.

  8. Electron spin resonance studies on intact cells and isolated lipid droplets from fatty acid-modified L1210 murine leukemia.

    PubMed

    Simon, I; Burns, C P; Spector, A A

    1982-07-01

    It has been suggested that the formation of cytoplasmic lipid droplets may produce an artifact and be responsible for the differences in membrane physical properties detected in lipid-modified cells using fluorescence polarization or spin label probes. To investigate this, the electron spin resonance spectra of lipid droplets isolated from the cytoplasm of L1210 leukemia cells were compared with spectra obtained from the intact cell. Mice bearing the L1210 leukemia were fed diets containing either 16% sunflower oil or 16% coconut oil in order to modify the fatty acid composition of the tumor. A microsome-rich fraction prepared from L1210 cells grown in animals fed the sunflower oil-rich diet contained more polyenoic fatty acids (52 versus 29%), while microsomes from L1210 cells grown in animals fed the coconut oil-rich diets contained more monoenoic fatty acids (37 versus 12%). The order parameter calculated for lipid droplets labeled with the 5-nitroxystearic acid spin probe was only about one-half that of intact cells, whereas it was similar to that obtained for pure triolein droplets suspended in buffer. Order parameters of the inner hyperfine splittings calculated from the spectra of cells grown in the sunflower oil-fed animals [0.543 +/- 0.001 (S.E.)] were lower than those from the cells grown in animals fed the coconut oil diets (0.555 +/- 0.002) (p less than 0.005). In contrast, the order parameters of the lipid droplets isolated from the cells grown in animals fed sunflower oil (0.303 +/- 0.029) or coconut oil (0.295 +/- 0.021) were not significantly different, indicating that motion of a spin label probe in the highly fluid cytoplasmic lipid droplets is not affected by these types of modifications in cellular fatty acid composition. Therefore, the electron spin resonance changes that are observed in the intact cells cannot be due to localization of the probe in cytoplasmic lipid droplets. These results support the conclusion that the electron spin

  9. Chimeric antigen receptor (CAR) and T cell receptor (TCR) Modified T cells Enter Main Street and Wall Street

    PubMed Central

    Barrett, David M; Grupp, Stephan A; June, Carl H

    2015-01-01

    The field of adoptive cell transfer (ACT) is currently comprised of 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 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. Here, we discuss some of the challenges and opportunities that face the field of ACT. PMID:26188068

  10. Foxj1 regulates floor plate cilia architecture and modifies the response of cells to sonic hedgehog signalling

    PubMed Central

    Cruz, Catarina; Ribes, Vanessa; Kutejova, Eva; Cayuso, Jordi; Lawson, Victoria; Norris, Dominic; Stevens, Jonathan; Davey, Megan; Blight, Ken; Bangs, Fiona; Mynett, Anita; Hirst, Elizabeth; Chung, Rachel; Balaskas, Nikolaos; Brody, Steven L.; Marti, Elisa; Briscoe, James

    2010-01-01

    Sonic hedgehog signalling is essential for the embryonic development of many tissues including the central nervous system, where it controls the pattern of cellular differentiation. A genome-wide screen of neural progenitor cells to evaluate the Shh signalling-regulated transcriptome identified the forkhead transcription factor Foxj1. In both chick and mouse Foxj1 is expressed in the ventral midline of the neural tube in cells that make up the floor plate. Consistent with the role of Foxj1 in the formation of long motile cilia, floor plate cells produce cilia that are longer than the primary cilia found elsewhere in the neural tube, and forced expression of Foxj1 in neuroepithelial cells is sufficient to increase cilia length. In addition, the expression of Foxj1 in the neural tube and in an Shh-responsive cell line attenuates intracellular signalling by decreasing the activity of Gli proteins, the transcriptional mediators of Shh signalling. We show that this function of Foxj1 depends on cilia. Nevertheless, floor plate identity and ciliogenesis are unaffected in mouse embryos lacking Foxj1 and we provide evidence that additional transcription factors expressed in the floor plate share overlapping functions with Foxj1. Together, these findings identify a novel mechanism that modifies the cellular response to Shh signalling and reveal morphological and functional features of the amniote floor plate that distinguish these cells from the rest of the neuroepithelium. PMID:21098568

  11. New modified polyetheretherketone membrane for liver cell culture in biohybrid systems: adhesion and specific functions of isolated hepatocytes.

    PubMed

    De Bartolo, L; Morelli, S; Rende, M; Gordano, A; Drioli, E

    2004-08-01

    There has been growing interest in innovative materials with physico-chemical properties that provide improved blood/cell compatibility. We propose new polymeric membranes made of modified polyetheretherketone (PEEK-WC) as materials with potential for use in biohybrid devices. PEEK-WC exhibits high chemical, thermal stability and mechanical resistance. Owing to its lack of crystallinity this polymer can be used for preparing membranes with cheap and flexible methods. We compared the properties of PEEK-WC membranes to polyurethane membranes prepared using the same phase inverse technique and commercial membranes. The physico-chemical properties of the membranes were characterised by contact angle measurements. The different parameters acid (gamma+), base (gamma-) and Lifshitz-van der Waals (gammaLW) of the surface free energy were calculated according to Good-van Oss's model. We evaluated the cytocompatibility of PEEK-WC membranes by culturing hepatocytes isolated from rat liver. Cell adhesion and metabolic behaviour in terms of ammonia elimination, urea synthesis and protein synthesis were evaluated during the first days of culture. Liver cells adhered and formed three-dimensional aggregates on the most tested membranes. PEEK-WC membranes promoted hepatocyte adhesion most effectively. Urea synthesis, ammonia elimination and protein synthesis improved significantly when cells adhered to PEEK-WC membrane. The considerable metabolic activities of cells cultured on this membrane confirmed the good structural and physico-chemical properties of the PEEK-WC membrane that could be a promising biomaterial for cell culture in biohybrid devices. PMID:15020136

  12. 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. PMID:26864442

  13. 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)

  14. CD4(+) T Cells Modified by the Endoribonuclease MazF Are Safe and Can Persist in SHIV-infected Rhesus Macaques.

    PubMed

    Saito, Naoki; Chono, Hideto; Shibata, Hiroaki; Ageyama, Naohide; Yasutomi, Yasuhiro; Mineno, Junichi

    2014-06-10

    MazF, an endoribonuclease encoded by Escherichia coli, specifically cleaves the ACA (adenine-cytosine-adenine) sequence of single-stranded RNAs. Conditional expression of MazF under the control of the HIV-1 LTR promoter rendered CD4(+) T cells resistant to HIV-1 replication without affecting cell growth. To investigate the safety, persistence and efficacy of MazF-modified CD4(+) T cells in a nonhuman primate model in vivo, rhesus macaques were infected with a pathogenic simian/human immunodeficiency virus (SHIV) and transplanted with autologous MazF-modified CD4(+) T cells. MazF-modified CD4(+) T cells were clearly detected throughout the experimental period of more than 6 months. The CD4(+) T cell count values increased in all four rhesus macaques. Moreover, the transplantation of the MazF-modified CD4(+) T cells was not immunogenic, and did not elicit cellular or humoral immune responses. These data suggest that the autologous transplantation of MazF-modified CD4(+) T cells in the presence of SHIV is effective, safe and not immunogenic, indicating that this is an attractive strategy for HIV-1 gene therapy.

  15. CD4+ T Cells Modified by the Endoribonuclease MazF Are Safe and Can Persist in SHIV-infected Rhesus Macaques

    PubMed Central

    Saito, Naoki; Chono, Hideto; Shibata, Hiroaki; Ageyama, Naohide; Yasutomi, Yasuhiro; Mineno, Junichi

    2014-01-01

    MazF, an endoribonuclease encoded by Escherichia coli, specifically cleaves the ACA (adenine–cytosine–adenine) sequence of single-stranded RNAs. Conditional expression of MazF under the control of the HIV-1 LTR promoter rendered CD4+ T cells resistant to HIV-1 replication without affecting cell growth. To investigate the safety, persistence and efficacy of MazF-modified CD4+ T cells in a nonhuman primate model in vivo, rhesus macaques were infected with a pathogenic simian/human immunodeficiency virus (SHIV) and transplanted with autologous MazF-modified CD4+ T cells. MazF-modified CD4+ T cells were clearly detected throughout the experimental period of more than 6 months. The CD4+ T cell count values increased in all four rhesus macaques. Moreover, the transplantation of the MazF-modified CD4+ T cells was not immunogenic, and did not elicit cellular or humoral immune responses. These data suggest that the autologous transplantation of MazF-modified CD4+ T cells in the presence of SHIV is effective, safe and not immunogenic, indicating that this is an attractive strategy for HIV-1 gene therapy. PMID:24914931

  16. The perfect host: a mouse host embryo facilitating more efficient germ line transmission of genetically modified embryonic stem cells.

    PubMed

    Taft, Robert A; Low, Benjamin E; Byers, Shannon L; Murray, Stephen A; Kutny, Peter; Wiles, Michael V

    2013-01-01

    There is a continual need to improve efficiency in creating precise genetic modifications in mice using embryonic stem cells (ESCs). We describe a novel approach resulting in 100% germline transmission from competent injected ESCs. We developed an F1 mouse host embryo (Perfect Host, PH) that selectively ablates its own germ cells via tissue-specific induction of diphtheria toxin. This approach allows competent microinjected ESCs to fully dominate the germline, eliminating competition for this critical niche in the developing and adult animal. This is in contrast to conventional methods, where competition from host germ cells results in offspring derived from host cells and ESCs, necessitating extensive breeding of chimeras and genotyping to identify germline. The germline transmission process is also complicated by variability in the actual number of ESCs that colonize the germline niche and the proportion that are germline competent. To validate the PH approach we used ESC lines derived from 129 F1, BALB/cByJ, and BTBR backgrounds as well as an iPS line. Resulting chimeric males produced 194 offspring, all paternally derived from the introduced stem cells, with no offspring being derived from the host genome. We further tested this approach using eleven genetically modified C57BL/6N ESC lines (International Knockout Mouse Consortium). ESC germline transmission was observed in 9/11 (82%) lines using PH blastocysts, compared to 6/11 (55%) when conventional host blastocysts were used. Furthermore, less than 35% (83/240) of mice born in the first litters from conventional chimeras were confirmed to be of ESC-origin. By comparison, 100% (137/137) of the first litter offspring of PH chimeras were confirmed as ESC-derived. Together, these data demonstrate that the PH approach increases the probability of germline transmission and speeds the generation of ESC derived animals from chimeras. Collectively, this approach reduces the time and costs inherent in the production

  17. Issues in the manufacture and transplantation of genetically modified hematopoietic stem cells.

    PubMed

    Sadelain, M; Frassoni, F; Rivière, I

    2000-11-01

    The advent of safe and practical means to correct, enhance or protect blood cells at the genetic level offers tantalizing therapeutic perspectives. At present, gene delivery using a replication-defective retrovirus is the most efficient method to stably transduce hematopoietic cells. The successful adaptation of retroviral infection to hematopoietic stem cells requires optimized transduction conditions that maximize gene transfer while preserving the cells' potential for engraftment and longterm hematopoiesis. The successful establishment of effective transduction protocols hinges on retrovirus biology as well as stem cell and transplantation biology. Interestingly, the genetic approach could permit novel strategies to promote host repopulation by transplanted stem cells. However, regulated and predictable expression of any transgene integrated at random chromosomal locations cannot be taken for granted. Investigation of the control of transgene expression and prevention of vector silencing will become increasingly important.

  18. Vascularization of hollow channel-modified porous silk scaffolds with endothelial cells for tissue regeneration.

    PubMed

    Zhang, Wenjie; Wray, Lindsay S; Rnjak-Kovacina, Jelena; Xu, Ling; Zou, Duohong; Wang, Shaoyi; Zhang, Maolin; Dong, Jiachen; Li, Guanglong; Kaplan, David L; Jiang, Xinquan

    2015-07-01

    Despite the promise for stem cell-based tissue engineering for regenerative therapy, slow and insufficient vascularization of large tissue constructs negatively impacts the survival and function of these transplanted cells. A combination of channeled porous silk scaffolds and prevascularization with endothelial cells was investigated to test the ability of this tissue engineering strategy to support rapid and extensive vascularization process. We report that hollow channels promote in vitro prevascularization by facilitating endothelial cell growth, VEGF secretion, and capillary-like tube formation. When implanted in vivo, the pre-established vascular networks in the hollow channel scaffolds anastomose with host vessels and exhibit accelerated vascular infiltration throughout the whole tissue construct, which provides timely and sufficient nutrients to ensure the survival of the transplanted stem cells. This tissue engineering strategy can promote the effective application of stem cell-based regeneration to improve future clinical applications.

  19. Brood cell size of Apis mellifera modifies the reproductive behavior of Varroa destructor.

    PubMed

    Maggi, Matías; Damiani, Natalia; Ruffinengo, Sergio; De Jong, David; Principal, Judith; Eguaras, Martín

    2010-03-01

    We undertook a field study to determine whether comb cell size affects the reproductive behavior of Varroa destructor under natural conditions. We examined the effect of brood cell width on the reproductive behavior of V. destructor in honey bee colonies, under natural conditions. Drone and worker brood combs were sampled from 11 colonies of Apis mellifera. A Pearson correlation test and a Tukey test were used to determine whether mite reproduction rate varied with brood cell width. Generalized additive model analysis showed that infestation rate increased positively and linearly with the width of worker and drone cells. The reproduction rate for viable mother mites was 0.96 viable female descendants per original invading female. No significant correlation was observed between brood cell width and number of offspring of V. destructor. Infertile mother mites were more frequent in narrower brood cells.

  20. 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.

  1. 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. PMID:27010252

  2. Gene-modified T-cell therapy using chimeric antigen receptors for pediatric hematologic malignancies.

    PubMed

    Nakazawa, Yozo

    2016-06-01

    Chimeric antigen receptor (CAR) is the generic name for synthetic T cell receptors redirected to tumor-associated antigens. Most CARs consist of an ectodomain (scFv or ligand), a hinge region, a transmembrane domain, and signaling endodomains derived from one or two co-stimulatory molecules (CD28, 4-1BB, etc) and from a CD3-ζ chain. CD19-targeted CAR T cell therapy has achieved major success in the treatment of B cell malignancies. CD19 CAR-T cells elicited complete remission in 70-90% of adult and pediatric patients with relapsed/refractory acute lymphoblastic leukemia (ALL). CD19 CAR T cell therapy from allogeneic donors including third party donors is a potential option for B-cell malignancies. CAR T cell therapies for myeloma, acute myeloid leukemia, and T-cell leukemia are still under development. Our group is currently preparing a phase I study of CD19 CAR T cell therapy in pediatric and young adult patients with ALL using a non-viral gene transfer method, the piggyBac-transposon system. PMID:27384848

  3. Modified Li/SO/sub 2/ cells for long-life applications

    SciTech Connect

    Levy, S C

    1980-01-01

    A 6-year test program to evaluate an ambient-temperature Li primary battery was initiated on cells from two manufacturers. Three modes of premature failure were observed in Li/SO/sub 2/ cells: corrosion of the lithium-to-can contact, corrosion of the glass in the glass-to-metal seal in the header, and corrosion at the Al-Ta weld in the positive lead. Potential fixes for each of these failure modes were incorporated into cells from two suppliers, and accelerated tests of these cells have begun. 2 figures, 1 table. (RWR)

  4. 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

  5. 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

  6. Scavenger receptors on sinusoidal liver endothelial cells are involved in the uptake of aldehyde-modified proteins.

    PubMed

    Duryee, Michael J; Freeman, Thomas L; Willis, Monte S; Hunter, Carlos D; Hamilton, Bartlett C; Suzuki, Hiroshi; Tuma, Dean J; Klassen, Lynell W; Thiele, Geoffrey M

    2005-11-01

    Scavenger receptors on sinusoidal liver endothelial cells (SECs) eliminate potentially harmful modified proteins circulating through the liver. It was shown recently that aldehyde-modified proteins bind to scavenger receptors and are associated with the development/progression of alcoholic liver diseases. For these studies, rat livers were perfused in situ with 125I-formaldehyde-bovine serum albumin (f-Alb) or 125I-malondialdehyde-acetaldehyde-bovine serum albumin (MAA-Alb) in the presence of known scavenger receptor ligands as inhibitors. Reverse transcription-polymerase chain reaction (RT-PCR) analysis and scavenger receptor Type A (SRA) knock-out mice were used to assess the role of these receptors in mediating immune responses. The degradation of 125I-f-Alb or 125I-MAA-Alb in whole livers and isolated SECs can be inhibited by known scavenger receptor ligands, including f-Alb, maleylated bovine albumin, and fucoidan. 125I-f-Alb could not be completely inhibited by MAA-Alb. In contrast, 125I-MAA-Alb was only partially inhibited with advanced glycosylated endproduct albumin. RT-PCR data show the presence of a number of scavenger receptors on SECs that may be responsible for the binding of MAA-modified proteins. SRA seems to be one of these receptors involved in the effects mediated by MAA-modified proteins. In a study using SRA knockout mice, it was shown that a decreased antibody response to MAA-Alb resulted. By RT-PCR, CD36, LOX-1, and SR-AI are the scavenger receptors most likely involved in the degradation of MAA-Alb.

  7. Scavenger receptors on sinusoidal liver endothelial cells are involved in the uptake of aldehyde-modified proteins.

    PubMed

    Duryee, Michael J; Freeman, Thomas L; Willis, Monte S; Hunter, Carlos D; Hamilton, Bartlett C; Suzuki, Hiroshi; Tuma, Dean J; Klassen, Lynell W; Thiele, Geoffrey M

    2005-11-01

    Scavenger receptors on sinusoidal liver endothelial cells (SECs) eliminate potentially harmful modified proteins circulating through the liver. It was shown recently that aldehyde-modified proteins bind to scavenger receptors and are associated with the development/progression of alcoholic liver diseases. For these studies, rat livers were perfused in situ with 125I-formaldehyde-bovine serum albumin (f-Alb) or 125I-malondialdehyde-acetaldehyde-bovine serum albumin (MAA-Alb) in the presence of known scavenger receptor ligands as inhibitors. Reverse transcription-polymerase chain reaction (RT-PCR) analysis and scavenger receptor Type A (SRA) knock-out mice were used to assess the role of these receptors in mediating immune responses. The degradation of 125I-f-Alb or 125I-MAA-Alb in whole livers and isolated SECs can be inhibited by known scavenger receptor ligands, including f-Alb, maleylated bovine albumin, and fucoidan. 125I-f-Alb could not be completely inhibited by MAA-Alb. In contrast, 125I-MAA-Alb was only partially inhibited with advanced glycosylated endproduct albumin. RT-PCR data show the presence of a number of scavenger receptors on SECs that may be responsible for the binding of MAA-modified proteins. SRA seems to be one of these receptors involved in the effects mediated by MAA-modified proteins. In a study using SRA knockout mice, it was shown that a decreased antibody response to MAA-Alb resulted. By RT-PCR, CD36, LOX-1, and SR-AI are the scavenger receptors most likely involved in the degradation of MAA-Alb. PMID:16105988

  8. 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.

  9. 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.

  10. 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 ...

  11. IGF-1 C Domain-Modified Hydrogel Enhances Cell Therapy for AKI.

    PubMed

    Feng, Guowei; Zhang, Jimin; Li, Yang; Nie, Yan; Zhu, Dashuai; Wang, Ran; Liu, Jianfeng; Gao, Jie; Liu, Na; He, Ningning; Du, Wei; Tao, Hongyan; Che, Yongzhe; Xu, Yong; Kong, Deling; Zhao, Qiang; Li, Zongjin

    2016-08-01

    Low cell retention and engraftment after transplantation limit the successful application of stem cell therapy for AKI. Engineered microenvironments consisting of a hydrogel matrix and growth factors have been increasingly successful in controlling stem cell fate by mimicking native stem cell niche components. Here, we synthesized a bioactive hydrogel by immobilizing the C domain peptide of IGF-1 (IGF-1C) on chitosan, and we hypothesized that this hydrogel could provide a favorable niche for adipose-derived mesenchymal stem cells (ADSCs) and thereby enhance cell survival in an AKI model. In vitro studies demonstrated that compared with no hydrogel or chitosan hydrogel only, the chitosan-IGF-1C hydrogel increased cell viability through paracrine effects. In vivo, cotransplantation of the chitosan-IGF-1C hydrogel and ADSCs in ischemic kidneys ameliorated renal function, likely by the observed promotion of stem cell survival and angiogenesis, as visualized by bioluminescence imaging and attenuation of fibrosis. In conclusion, IGF-1C immobilized on a chitosan hydrogel provides an artificial microenvironment for ADSCs and may be a promising therapeutic approach for AKI. PMID:26869006

  12. Positive selection of gene-modified cells increases the efficacy of pancreatic cancer suicide gene therapy.

    PubMed

    Martinez-Quintanilla, Jordi; Cascallo, Manel; Gros, Alena; Fillat, Cristina; Alemany, Ramon

    2009-11-01

    Thymidine kinase (TK)-mediated suicide gene therapy has been considered for the treatment of pancreatic cancer. However, despite a bystander effect, the proportion of transduced tumor cells has proven too low to result in efficacy. We propose the use of a drug-selectable marker (MDR1) to enrich TK-expressing cells using chemotherapy. This enrichment or positive selection phase may increase the efficacy of suicide gene therapy. To test this strategy, we generated stable NP18MDR/TK-GFP transfectants and showed docetaxel resistance in vivo. Mixed tumors of MDR/TK-expressing cells and parental NP18 cells were established and docetaxel was used to increase the proportion of TK-expressing cells. After this positive selection phase, suicide gene therapy with ganciclovir was applied. Upon positive selection, the proportion of TK-expressing cells increased from 4% to 22%. Subsequent suicide gene therapy was more effective compared with a control group without positive selection. Starting with 10% of TK-expressing cells the positive-negative selection strategy completely inhibited tumor growth. Taken together, these results suggest that a positive-negative selection strategy based on MDR and TK genes represents an efficient way to increase the proportion of TK-expressing cells in the tumor and the efficacy of TK-mediated suicide gene therapy.

  13. Diesel exhaust particles modify natural killer cell function and cytokine release

    PubMed Central

    2013-01-01

    Background Natural killer (NK) cells are an important lymphocyte population in the nasal mucosa and play important roles in linking the innate and the adaptive immune response. Their two main functions are direct cell-mediated cytotoxicity and the release of cytokines. They are important during viral infections and cancer. Due to their location in the nasal mucosa, NK cells are likely exposed to inhaled pollutants, such as diesel exhaust. Whether and how exposure to diesel exhaust particles (DEP) affects NK cell function in the context of viral infections has not been investigated. Methods NK cells were isolated from peripheral blood obtained from normal healthy volunteers and subsequently stimulated with the viral mimetic polyinosinic:polycytidylic acid (pI:C), DEP, or pI:C+DEP for 18 hours. NK cells were subsequently analyzed for changes in surface marker expression, cytokine production, gene expression changes, and cytotoxic function using flow cytometry, ELISA, qRT-PCR, and cell-mediated cytotoxicity assay, respectively. Results Stimulation of NK cells with pI:C and pI:C+DEP, but not DEP alone, increased the release of IL-1β, IL-2, IL-4, IL-8, IL-10, IL-12p70, IFN-γ and TNF-α. As compared to pI:C alone or pI:C+DEP, the release of IL-1β, IL-8 and TNF-α was significantly lower after DEP stimulation alone. Stimulation with pI:C alone increased the gene and protein expression of granzyme B and perforin, which was completely blunted by adding DEP. Addition of DEP further reduced CD16 expression in pI:C stimulated cells. Similarly, cell-mediated cytotoxicity was significantly reduced by the addition of DEP. Conclusions In the context of viral infection, DEP potentially reduces NK cells' ability to kill virus-infected host cells, in spite of normal cytokine levels, and this may increase susceptibility to viral infections . This reduction in the potential ability of NK cells to kill virus-infected host cells may increase the susceptibility to viral infections

  14. Graphene Oxide Nanosheets Modified with Single-Domain Antibodies for Rapid and Efficient Capture of Cells.

    PubMed

    Chen, Guan-Yu; Li, Zeyang; Theile, Christopher S; Bardhan, Neelkanth M; Kumar, Priyank V; Duarte, Joao N; Maruyama, Takeshi; Rashidfarrokh, Ali; Belcher, Angela M; Ploegh, Hidde L

    2015-11-23

    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

  15. Induction of primary mixed leukocyte reactions with ultraviolet B or chemically modified stimulator cells

    SciTech Connect

    Mincheff, M.S.; Meryman, H.T. )

    1989-12-01

    Treatment of stimulator cells with paraformaldehyde for 60 sec or ultraviolet-B (UV-B) irradiation eliminates their ability to elicit T cell proliferation in a primary mixed leukocyte reaction. However, a T cell response equal to 20-40% of control value could be elicited by paraformaldehyde fixed or UV-B irradiated cells providing the latter are incubated at 37 degrees C for 18 hr prior to treatment. The incubation also induces a one-log increase in the density of fluorescence when the cells are stained with monoclonal antibodies against class II molecules DR and DP as well as the intercellular adhesion molecule -1. We interpret this as an increase in the membrane expression of these structures following incubation. Chloroquine and cerulenin, known to inhibit protein degradation and antigen processing and presentation do not influence the upregulation in membrane expression of these class II and adhesion molecules, but do prevent incubation from overriding the effect of paraformaldehyde treatment. Colchicine, which reduces the traffic through tubular lysosomes, also has no effect on the upregulation but enhances allopresentation. We propose that incubation of stimulator cells in the presence of chloroquine and cerulenin results in the membrane expression of class II molecules without associated peptides. The inability of stimulator cells expressing such nude MHC molecules to elicit T cell proliferation after chemical modification could be due to easier crosslinking of the allodeterminants by paraformaldehyde when the binding site is empty but could also mean that nude MHC molecules are not per se immunogenic and become so only after acquisition of a peptide. It is also possible that chloroquine, NH4Cl, and cerulenin block the expression of signals other than the class II and cell adhesion molecules that are essential for induction of T cell proliferation.

  16. Microtubule nucleating and severing enzymes for modifying microtubule array organization and cell morphogenesis in response to environmental cues.

    PubMed

    Nakamura, Masayoshi

    2015-02-01

    In higher plants, reorientation of cortical microtubule arrays has been postulated to be of importance for modifying cell growth to adapt to environmental conditions. However, the process of microtubule reorientation is largely unknown. Recent genetic and live cell imaging studies of microtubule dynamics shed light on the regulatory mechanisms of microtubule molecular nucleation and severing apparatuses, which are required for array reorientation in response to blue light signaling. Branching nucleation from γ-tubulin complexes creates a small population of discordant microtubules that are acted on by KATANIN-mediated severing in two ways. KATANIN releases microtubules from nucleation sites and rapidly amplifies discordant microtubules by severing at microtubule crossovers. In this review, I focus on the molecular details of these two enzymes, which enable microtubule array transition.

  17. High-bendability flexible dye-sensitized solar cell with a nanoparticle-modified ZnO-nanowire electrode

    NASA Astrophysics Data System (ADS)

    Jiang, C. Y.; Sun, X. W.; Tan, K. W.; Lo, G. Q.; Kyaw, A. K. K.; Kwong, D. L.

    2008-04-01

    We report a high-bendability flexible dye-sensitized solar cell (DSSC) based on a ZnO-nanowire photoelectrode, which was fabricated on polyethylene terephtalate/indium tin oxide substrate by low-temperature hydrothermal growth. Nanowire morphology shows preferable in crack resistance due to its efficient release of bending stress. The ZnO-nanowire film can be bended to an extreme radius of 2mm with no crack observed. Flexible DSSCs based on this kind of ZnO-nanowire photoelectrodes showed good bending stability. With a ZnO-nanoparticle modification on the nanowires, the flexible DSSC fabricated showed a much improved power conversion efficiency. Meanwhile, the good bendablility of this nanoparticle-modified nanowire electrode is maintained. The results demonstrate that high quality ZnO nanowires fabricated by the low-temperature method is promising for efficient and flexible plastic solar cells.

  18. Osteogenic Potential of Dental Mesenchymal Stem Cells in Preclinical Studies: A Systematic Review Using Modified ARRIVE and CONSORT Guidelines

    PubMed Central

    Ramamoorthi, Murali; Bakkar, Mohammed; Jordan, Jack; Tran, Simon D.

    2015-01-01

    Background and Objective. Dental stem cell-based tissue engineered constructs are emerging as a promising alternative to autologous bone transfer for treating bone defects. The purpose of this review is to systematically assess the preclinical in vivo and in vitro studies which have evaluated the efficacy of dental stem cells on bone regeneration. Methods. A literature search was conducted in Ovid Medline, Embase, PubMed, and Web of Science up to October 2014. Implantation of dental stem cells in animal models for evaluating bone regeneration and/or in vitro studies demonstrating osteogenic potential of dental stem cells were included. The preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines were used to ensure the quality of the search. Modified ARRIVE (Animal research: reporting in invivo experiments) and CONSORT (Consolidated reporting of trials) were used to critically analyze the selected studies. Results. From 1914 citations, 207 full-text articles were screened and 137 studies were included in this review. Because of the heterogeneity observed in the studies selected, meta-analysis was not possible. Conclusion. Both in vivo and in vitro studies indicate the potential use of dental stem cells in bone regeneration. However well-designed randomized animal trials are needed before moving into clinical trials. PMID:26106427

  19. 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.

  20. 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

  1. The epigenetic modifiers 5-aza-2'-deoxycytidine and trichostatin A influence adipocyte differentiation in human mesenchymal stem cells.

    PubMed

    Zych, J; Stimamiglio, M A; Senegaglia, A C; Brofman, P R S; Dallagiovanna, B; Goldenberg, S; Correa, A

    2013-05-01

    Epigenetic mechanisms such as DNA methylation and histone modification are important in stem cell differentiation. Methylation is principally associated with transcriptional repression, and histone acetylation is correlated with an active chromatin state. We determined the effects of these epigenetic mechanisms on adipocyte differentiation in mesenchymal stem cells (MSCs) derived from bone marrow (BM-MSCs) and adipose tissue (ADSCs) using the chromatin-modifying agents trichostatin A (TSA), a histone deacetylase inhibitor, and 5-aza-2'-deoxycytidine (5azadC), a demethylating agent. Subconfluent MSC cultures were treated with 5, 50, or 500 nM TSA or with 1, 10, or 100 µM 5azadC for 2 days before the initiation of adipogenesis. The differentiation was quantified and expression of the adipocyte genes PPARG and FABP4 and of the anti-adipocyte gene GATA2 was evaluated. TSA decreased adipogenesis, except in BM-MSCs treated with 5 nM TSA. Only treatment with 500 nM TSA decreased cell proliferation. 5azadC treatment decreased proliferation and adipocyte differentiation in all conditions evaluated, resulting in the downregulation of PPARG and FABP4 and the upregulation of GATA2. The response to treatment was stronger in ADSCs than in BM-MSCs, suggesting that epigenetic memories may differ between cells of different origins. As epigenetic signatures affect differentiation, it should be possible to direct the use of MSCs in cell therapies to improve process efficiency by considering the various sources available.

  2. [Transplantation of genetically modified cells in the treatment of children with SCID: great hopes and recent disappointments].

    PubMed

    Smogorzewska, Elzbieta Monika; Weinberg, Kenneth I; Kohn, Donald B

    2003-01-01

    Children with severe combined immunodeficiency (SCID) die within 2 years of age if untreated. The only effective treatment for SCID since 1968 is a hematopoetic stem cells (HSC) transplantation. Only 25% of patients have an HLA matched related donor, while the rest have to be transplanted with T cells depleted haploidentical parental bone marrow, unrelated bone marrow or unrelated umbilical cord blood. In many cases, however, despite a positive outcome, children are not achieving B cell reconstitution and require regular IV Ig infusion. Gene therapy with genetically modified autologous cells offers a cure with no immunological complications such as graft rejection, graft versus host disease (GVHD) or post-transplantation immunosuppressive therapy. The first gene therapy trials were introduced in 1990 for adenosine deaminase (ADA) deficient patients who had failed to respond to PEG-ADA. Since then, three clinical trials have evaluated the transplantation of ex-vivo transduced autologous haematopoietic stem cells (HSC) to treat ADA deficiency. One trial used only bone marrow HSC, a second used bone marrow plus peripheral blood T lymphocytes, and a third used umbilical cord blood HSC. These trials give promise but also define the present limitations of gene therapy. Future protocols might be adjusted according to the new observations that ADA-expressing T cells have a strong selective advantage over ADA-deficient T cells. PEG-ADA enzyme therapy might be therefore contraindicated. Another new strategy might involve moderate conditioning prior to the reinfusion of genetically modified CD34+ cells, "making space" for transplanted HSC. The first successful gene therapy was reported for treatment of X-linked severe combined immunodeficiency (SCID-X1) in Science 2000. Since then, the group at the Hopital Necker in Paris has treated 11 patients with ex-vivo gene therapy for the deficiency of the common g chain. All eleven boys are alive, however, one of them recently

  3. Selective biological response of human pulmonary microvascular endothelial cells and human pulmonary artery smooth muscle cells on cold-plasma-modified polyester vascular prostheses.

    PubMed

    Blanchemain, N; Aguilar, M R; Chai, F; Jimenez, M; Jean-Baptiste, E; El-Achari, A; Martel, B; Hildebrand, H F; Roman, J San

    2011-12-01

    The aim of this work was to improve the hemocompatibility and the selectivity according to cells of non-woven poly(ethylene terephthalate) (PET) membranes. Non-woven PET membranes were modified by a combined plasma-chemical process. The surface of these materials was pre-activated by cold-plasma treatment and poly(acrylic acid) (PAA) was grafted by the in situ free radical polymerization of acrylic acid (AA). The extent of this reaction and the number of carboxylic groups incorporated were evaluated by colorimetric titration using toluidine blue O. All samples were characterized by SEM, AFM and thermogravimetric analysis, and the mechanical properties of the PAA grafted sample were determined. A selective cell response was observed when human pulmonary artery smooth muscle cells (HPASMC) or human pulmonary micro vascular endothelial cells (HPMEC) were seeded on the modified surfaces. HPASMC proliferation decreased about 60%, while HPMEC proliferation was just reduced about 10%. PAA grafted samples did not present hemolytic activity and the platelet adhesion decreased about 28% on PAA grafted surfaces. PMID:22002636

  4. Increased apoptosis in cancer cells in vitro and in vivo by ceramides in transferrin-modified liposomes

    PubMed Central

    Koshkaryev, Alexander; Piroyan, Aleksandr

    2012-01-01

    Lysosomes are a promising therapeutic target for induction apoptosis in cancer cells due to lysosomal membrane permeabilization (LMP) leading to leakage of hydrolytic enzymes, especially the cathepsins, into the cytoplasm. We hypothesized that with the modification of the ceramide-loaded liposomes with transferrin (Tf), we would achieve both tumor targeting and increased delivery of lysosome-destabilizing agents, such as ceramides to lysosomes, to initiate LMP-induced apoptosis. We prepared Tf-modified (TL) and plain (PL) liposomes loaded with short (C6) or long (C16) N-acyl chain ceramides. Uptake, intracellular localization of liposomes, stability of the lysosomal membrane and release of cathepsin D were investigated on Hela cells by fluorescence microscopy and flow cytometry. Apoptosis was evaluated by binding of fluorescently-labeled Annexin V. Antitumor and pro-apoptotic effects of C6Cer-loaded Tf-liposomes were demonstrated in vivo in an A2780-ovarian carcinoma xenograft mouse model. TL were internalized specifically via the TfR-dependent endocytic pathway and localized within the endosomelysosomal compartment. Ceramide-loaded Tf-liposomes significantly increased apoptosis compared with ceramide-free and ceramide-loaded non-modified liposomes. The treatment of cancer cells with TL led to increased LMP and cytoplasmic relocation of the intralysosomal cathepsin D. A strong antitumor and pro-apoptotic effect of C6Cer-loaded TL was also demonstrated in vivo in an A2780-ovarian carcinoma xenograft mouse model. The lysosomal accumulation of ceramides delivered by Tf-liposomes initiates the permeabilization of the lysosomal membranes required for the release of lysosomal cathepsins into the cytoplasm and initiation of the cancer cell apoptosis both in vitro and in vivo. PMID:22336588

  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. Angiopoietin-1 gene-modified human mesenchymal stem cells promote angiogenesis and reduce acute pancreatitis in rats.

    PubMed

    Hua, Jie; He, Zhi-Gang; Qian, Dao-Hai; Lin, Sheng-Ping; Gong, Jian; Meng, Hong-Bo; Yang, Ting-Song; Sun, Wei; Xu, Bin; Zhou, Bo; Song, Zhen-Shun

    2014-01-01

    Mesenchymal stem cells (MSCs) can serve as a vehicle for gene therapy. Angiopoietin-1 (ANGPT1) plays an important role in the regulation of endothelial cell survival, vascular stabilization, and angiogenesis. We hypothesized that ANGPT1 gene-modified MSCs might be a potential therapeutic approach for severe acute pancreatitis (SAP) in rats. Human umbilical cord-derived MSCs with or without transfection with lentiviral vectors containing the ANGPT1 gene were delivered through the tail vein of rats 12 h after induction of SAP. Administration of MSCs alone significantly reduced pancreatic injury and inflammation, as reflected by reductions in pancreatitis severity scores and serum amylase and lipase levels as well as reducing the serum levels of proinflammatory cytokines (TNF-α, IFN-γ, IL-1β, and IL-6). Furthermore, administration of ANGPT1-transfected MSCs resulted in not only further reductions in pancreatic injury and serum levels of proinflammatory cytokines, but also promotion of pancreatic angiogenesis. These results suggest that MSCs and ANGPT1 have a synergistic role in the treatment of SAP. ANGPT1 gene-modified MSCs may be developed as a potential novel therapy strategy for the treatment of SAP. PMID:25120736

  7. 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. PMID:27462342

  8. 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

  9. Biodistribution and in vivo efficacy of genetically modified human mesenchymal stem cells systemically transplanted into a mouse bone fracture model.

    PubMed

    Kang, Jin Wook; Park, Ki Dae; Choi, Youngju; Baek, Dae Hyun; Cho, Wan-Seob; Choi, Mina; Park, Jae Hyun; Choi, Kyoung Suk; Kim, Hyung Soo; Yoo, Tae Moo

    2013-08-01

    Human mesenchymal stem cells (hMSCs) have generated a great deal of interest in clinical application due to their ability to undergo multi-lineage differentiation. Recently, ex vivo genetic modification of hMSCs was attempted to increase their differentiation potential. The present study was conducted to evaluate the biodistribution and in vivo efficacy of genetically modified hMSCs. To accomplish this, Runx2, which is a key transcription factor associated with osteoblast differentiation, was transduced into hMSCs using lentiviral vectors expressing green fluorescent protein (GFP) or luciferase. Here, we developed an experimental fracture in mice femur to investigate the effects of Runx2-transduced hMSCs on bone healing and migration into injury site. We conducted bio-luminescence imaging (BLI) using luciferase-tagged vector and quantitative real-time PCR using GFP probe to investigate the biodistribution of Runx2-transduced hMSCs in the fracture model. The biodistribution of hMSC cells in the fractured femur was observed at 14 days post-transplantation upon both BLI imaging and real-time PCR. Moreover, the fractured mice transplanted with Runx2-transduced hMSCs showed superior bone healing when compared to mock-transduced hMSC and MRC5 fibroblasts which were used as control. These data suggested that transplanted genetically modified hMSCs systemically migrate to the fractured femur, where they contribute to bone formation in vivo.

  10. 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

  11. Proteomes of host cell membranes modified by intracellular activities of Salmonella enterica.

    PubMed

    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.

  12. 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

  13. Improved lysis of single bacterial cells by a modified alkaline-thermal shock procedure.

    PubMed

    He, Jian; Du, Shiyu; Tan, Xiaohua; Arefin, Ayesha; Han, Cliff S

    2016-01-01

    Single-cell genomics (SCG) is a recently developed tool to study the genomes of unculturable bacterial species. SCG relies on multiple-strand displacement amplification (MDA), PCR, and next-generation sequencing (NGS); however, obtaining sufficient amounts of high-quality DNA from samples is a major challenge when performing this technique. Here we present an improved bacterial cell lysing procedure that combines incubation in an alkaline buffer with a thermal shock (freezing/heating) treatment to yield highly intact genomic DNA with high efficiency. This procedure is more efficient in lysing Bacillus subtilis and Synechocystis cells compared with two other frequently used lysis methods. Furthermore, 16S ribosomal RNA gene and overall genome recovery were found to be improved by this method using single cells from a Utah desert soil community or Escherichia coli single cells, respectively. The efficiency of genome recovery for E. coli single cells using our procedure is comparable with that of the REPLI-g Single Cell (sc) Kit, but our method is much more economical. By providing high-quality genome templates suitable for downstream applications, our procedure will be a promising improvement for SCG research. PMID:26956090

  14. Modifying Risk of Aneuploidy with a Positive Cell-Free Fetal DNA Result.

    PubMed

    Long, A Ashleigh; Abuhamad, Alfred Z; Warsof, Steven L

    2016-06-01

    Noninvasive genomic assessments of the fetus while in utero have been made possible by the analysis of cell-free fetal DNA fragments from the serum of pregnant women, as part of a noninvasive prenatal testing screening strategy. Between 7% and 10% of total cell-free DNA in the maternal blood comes from placental trophoblasts, allowing for identification of the DNA associated with the fetal component of the placenta. Using simple venipuncture in the outpatient setting, this cell-free, extracellular fetal DNA can be isolated in the maternal serum from a single blood draw as early as the seventh week of gestation. PMID:27235910

  15. Specific deletion of AMP-activated protein kinase (α1AMPK) in mouse Sertoli cells modifies germ cell quality.

    PubMed

    Bertoldo, Michael J; Guibert, Edith; Faure, Melanie; Guillou, Florian; Ramé, Christelle; Nadal-Desbarats, Lydie; Foretz, Marc; Viollet, Benoit; Dupont, Joëlle; Froment, Pascal

    2016-03-01

    The AMP-activated protein kinase (AMPK) is an important regulator of cellular energy homeostasis which plays a role in fertility. Complete disruption of the AMPK catalytic subunit α1 gene (α1AMPK KO) in male mice results in a decrease in litter size which is associated with the production of altered sperm morphology and motility. Because of the importance of Sertoli cells in the formation of germ cells, we have chosen to selectively disrupt α1AMPK only in the Sertoli cells in mice (Sc-α1AMPK-KO mice). Specific deletion of the α1AMPK gene in Sertoli cells resulted in a 25% reduction in male fertility associated with abnormal spermatozoa with a thin head. No clear alterations in testis morphology or modification in the number of Sertoli cells in vivo were observed, but a dysregulation in energy metabolism in Sertoli cells occurred. We have reported an increase in lactate production, in lipid droplets, and a reduction in ATP production in Sc-α1AMPK-KO Sertoli cells. These perturbations were associated with lower expression of mitochondrial markers (cytochrome c and PGC1-α). In addition another metabolic sensor, the deacetylase SIRT1, had a reduction in expression which is correlated with a decline in deacetylase activity. Finally, expression and localization of junctions forming the blood-testis barrier between Sertoli cells themselves and with germ cells were deregulated in Sc-α1AMPK-KO. In conclusion, these results suggest that dysregulation of the energy sensing machinery exclusively through disruption of α1AMPK in Sertoli cells translates to a reduction in the quality of germ cells and fertility. PMID:26772142

  16. Sialic acid-modified antigens impose tolerance via inhibition of T-cell proliferation and de novo induction of regulatory T cells

    PubMed Central

    Perdicchio, Maurizio; Ilarregui, Juan M.; Verstege, Marleen I.; Cornelissen, Lenneke A. M.; Schetters, Sjoerd T. T.; Engels, Steef; Ambrosini, Martino; Kalay, Hakan; Veninga, Henrike; den Haan, Joke M. M.; van Berkel, Lisette A.; Samsom, Janneke N.; Crocker, Paul R.; Sparwasser, Tim; Berod, Luciana; Garcia-Vallejo, Juan J.; van Kooyk, Yvette; Unger, Wendy W. J.

    2016-01-01

    Sialic acids are negatively charged nine-carbon carboxylated monosaccharides that often cap glycans on glycosylated proteins and lipids. Because of their strategic location at the cell surface, sialic acids contribute to interactions that are critical for immune homeostasis via interactions with sialic acid-binding Ig-type lectins (siglecs). In particular, these interactions may be of importance in cases where sialic acids may be overexpressed, such as on certain pathogens and tumors. We now demonstrate that modification of antigens with sialic acids (Sia-antigens) regulates the generation of antigen-specific regulatory T (Treg) cells via dendritic cells (DCs). Additionally, DCs that take up Sia-antigen prevent formation of effector CD4+ and CD8+ T cells. Importantly, the regulatory properties endowed on DCs upon Sia-antigen uptake are antigen-specific: only T cells responsive to the sialylated antigen become tolerized. In vivo, injection of Sia-antigen–loaded DCs increased de novo Treg-cell numbers and dampened effector T-cell expansion and IFN-γ production. The dual tolerogenic features that Sia-antigen imposed on DCs are Siglec-E–mediated and maintained under inflammatory conditions. Moreover, loading DCs with Sia-antigens not only inhibited the function of in vitro–established Th1 and Th17 effector T cells but also significantly dampened ex vivo myelin-reactive T cells, present in the circulation of mice with experimental autoimmune encephalomyelitis. These data indicate that sialic acid-modified antigens instruct DCs in an antigen-specific tolerogenic programming, enhancing Treg cells and reducing the generation and propagation of inflammatory T cells. Our data suggest that sialylation of antigens provides an attractive way to induce antigen-specific immune tolerance. PMID:26941238

  17. Efficiency of CD19 chimeric antigen receptor-modified T cells for treatment of B cell malignancies in phase I clinical trials: a meta-analysis

    PubMed Central

    Shi, Ni; Zhang, Zhen; Luo, Zhenzhen; Yue, Dongli; Zhang, Zimeng; Wang, Liping; Han, Weidong; Xu, Zhongwei; Chen, Hu; Zhang, Yi

    2015-01-01

    Chimeric antigen receptor (CAR) modified T cells targeted CD19 showed promising clinical outcomes in treatment of B cell malignances such as chronic lymphocytic leukemia (CLL), acute lymphoblastic leukemia (ALL) and other indolent lymphomas. However, the clinical benefit varies tremendously among different trials. This meta-analysis investigated the efficacy (response rates and survival time) of CD19-CAR T cells in refractory B cell malignances in Phase I clinical trials. We searched publications between 1991 and 2014 from PubMed and Web of Science. Pooled response rates were calculated using random-effects models. Heterogeneity was investigated by subgroup analysis and meta-regression. Fourteen clinical trials including 119 patients were eligible for response rate evaluation, 62 patients in 12 clinical trials were eligible for progression-free survival analysis. The overall pooled response rate of CD19-CAR T cells was 73% (95% confidence interval [CI]: 46-94%). Significant heterogeneity across estimates of response rates was observed (p < 0.001, I2=88.3%). ALL patients have higher response rate (93%, 95% CI: 65-100%) than CLL (62%, 95% CI: 27-93%) and lymphoma patients (36%, 95% CI: 1-83%). Meta-regression analysis identified lymphodepletion and no IL-2 administrated T cells as two key factors associated with better clinical response. Lymphodepletion and higher infused CAR T cell number were associated with better prognosis. In conclusion, this meta-analysis showed a high clinical response rate of CD19-CAR T cell-based immunotherapy in treatment of refractory B cell malignancies. Lymphodepletion and increasing number of infused CD19-CAR T cells have positive correlations with the clinical efficiency, on the contrary, IL-2 administration to T cells is not recommended. PMID:26376680

  18. Do trichothecenes reduce viability of circulating blood cells and modify haemostasis parameters?

    PubMed

    Froquet, R; Arnold, F; Batina, P; Parent-Massin, D

    2003-01-01

    This manuscript describes the results of experiments conducted using human blood cells to determine the ability of T-2 toxin and DON to cause changes in clotting time, platelet aggregation, red blood cell haemolysis, RBC glucose content, lactate release, glutathione depletion, as well as white blood cell viability. In vitro results showed that haemostasis parameters and erythrocytes were not affected at concentrations able to induce inhibition of haematopoietic progenitor proliferation. In the presence of 10(-8) M and 10(-6) M T-2, the leucocyte number decreased at 24 h by 30% and 50% respectively. A 50% decrease in leucocyte number was observed for 10(-5) M DON. Results were compared with haematopoietic progenitor sensitivities. Due to the differences in sensitivities between mature blood cells and haematopoietic progenitors, haematological problems associated with trichothecene intoxication could be attributed to haematopoiesis inhibition.

  19. Modulation of the heterogeneous senescence of human mesenchymal stem cells on chemically-modified surfaces.

    PubMed

    Kim, Sung Hoon; Lee, Byung Man; Min, Seul Ki; Song, Sun U; Cho, Jeong Ho; Cho, Kilwon; Shin, Hwa Sung

    2012-02-01

    Human mesenchymal stem cells (hMSCs) are multipotent and have been recognized as a source for tissue engineering or cell therapy. It is, therefore, imperative to develop methods to acquire enough hMSCs that maintain self-renewal and differentiation potential. However, aged hMSCs are prone to have a gradual decline in differentiation and proliferation potential with continual cell cycle divisions during in vitro culture. The physiochemical properties of hMSCs are highly dependent on their micro-environment, i.e. the 'stem cell niche'. In this study, the heterogeneous aging of hMSC was examined on chemically defined self-assembly monolayer surfaces. Surface energy was shown to regulate aged hMSC morphology, survival, and proteoglycan expression. High surface energy supplied a preferable environment for hMSC survival and expression of proteoglycans. These results will prove valuable to the design of scaffolds for tissue engineering or for the modulation of implantation environments.

  20. Surface-modified complex SU-8 microstructures for indirect optical manipulation of single cells.

    PubMed

    Aekbote, Badri L; Fekete, Tamás; Jacak, Jaroslaw; Vizsnyiczai, Gaszton; Ormos, Pál; Kelemen, Lóránd

    2016-01-01

    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

  1. 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

  2. Distribution of oxidized and HNE-modified proteins in U87 cells.

    PubMed

    Jung, Tobias; Engels, Martina; Kaiser, Barbara; Grune, Tilman

    2005-01-01

    Protein modification is one of the important processes during oxidative stress. This modification of proteins is either due to direct oxidation of proteins by various oxidants or due to secondary modification by lipid peroxidation products, e.g. 4-hydroxynonenal. In the here presented work we compare the intracellular distribution of protein modification products after treatment of human U87 astrocytoma cells with hydrogen peroxide or HNE. The treatment with hydrogen peroxide leads mainly to a cytosolic formation of oxidized proteins whereas HNE treatment is forming HNE-adducts throughout the cell. Therefore, we concluded that HNE diffusion distance in cells enables this lipid peroxidation product to act as a second messenger within the cell and on the other hand is the reason for the genotoxic properties of this compound.

  3. 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.

  4. A modified neural network model of tumor cell interactions and subpopulation dynamics.

    PubMed

    Prideaux, J A; Mikulecky, D C; Clarke, A M; Ware, J L

    1993-01-01

    Tumors consist of phenotypically heterogeneous subpopulations of cells which are frequently affected by both autocrine and paracrine factors. Applying concepts from neural network theory, we have developed a computer model of chemical communication among hypothetical tumor cells, which simulates some of the complex epigenetic behavior of real tumors. Deletion of subpopulations often destabilized the whole population. The impact of deletion of specific subpopulations was affected by (a) which subpopulation was deleted, and (b) the timing of the deletion during tumor progression.

  5. 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.

  6. Tumor necrosis factor-alpha modifies adhesion properties of rat islet B cells.

    PubMed Central

    Cirulli, V; Halban, P A; Rouiller, D G

    1993-01-01

    The characteristic three-dimensional cell type organization of islets of Langerhans is perturbed in animal models of diabetes, suggesting that it may be important for islet function. Rat islet cells in culture are able to form aggregates with an architecture similar to native islets (pseudoislets), thus providing a good model to study the molecular basis of islet architecture and its role in islet function. Sorted islet B cells and non-B cells were permanently labeled with two different fluorescent dyes (DiO and DiI), mixed, and allowed to form aggregates during a 5-d culture in the presence or absence of TNF-alpha (100 U/ml), a cytokine suggested to be implicated in the early physiological events leading to insulin-dependent diabetes mellitus. Confocal microscopy of aggregates revealed that TNF-alpha reversibly perturbs the typical segregation between B and non-B cells. Insulin secretion, was altered in the disorganized aggregates, and returned towards normal when pseudoislets had regained their typical architecture. The homotypic adhesion properties of sorted B and non-B cells cultured for 20 h in the presence or absence of TNF-alpha were studied in a short term aggregation assay. TNF-alpha induced a significant rise in Ca(2+)-independent adhesion of B cells (from 24 +/- 1.1% to 44.3 +/- 1.2%; n = 4, P < 0.001). These findings raise the possibility that the increased expression of Ca(2+)-independent adhesion molecules on B cells leads to altered islet architecture, which might be a factor in the perturbation of islet function induced by TNF-alpha. Images PMID:8098044

  7. 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. PMID:26700650

  8. Endostatin and irradiation modifies the activity of ADAM10 and neprilysin in breast cancer cells.

    PubMed

    Aydemir, Esra Arslan; Şimşek, Ece; Korcum, Aylin Fidan; Fişkin, Kayahan

    2016-09-01

    Angiogenesis, the formation of new blood vessels, is regarded as a key cancer cell property. Endostatin (ES) is a potential antiangiogenic agent and it may be useful when implemented in combination with other cancer therapeutic strategies. The present study investigated the in vitro effects of ES, radiotherapy (RT) or combination therapy (ES + RT) on two important proteases, a disintegrin and metalloproteinase domain‑containing protein 10 (ADAM10) and neprilysin (NEP) in 4T1 mouse breast cancer cells and the more metastatic phenotype of 4THMpc breast cancer cells. 4T1 and 4THMpc cells were treated with recombinant murine ES (4 µg/ml) alone, RT (45 Gy) alone or with ES + RT. ADAM10 enzyme activity was determined using a tumor necrosis factor‑α converting enzyme (α‑secretase) activity assay kit, and NEP enzyme activity was measured with a fluorometric assay based on the generation of free dansyl‑D‑Ala‑Gly from N-dansyl-Ala-Gly-D-nitro-Phe-Gly, the substrate of NEP. Western blotting analysis was performed to determine whether the altered enzyme activity levels of the two cell lines occurred due to changes in expression level. These data indicate that ES independently potentiates the activity of ADAM10 and NEP enzymes in 4T1 and 4THMpc breast cancer cells. PMID:27430992

  9. Impact of differently modified nanocrystalline diamond on the growth of neuroblastoma cells.

    PubMed

    Vaitkuviene, Aida; McDonald, Matthew; Vahidpour, Farnoosh; Noben, Jean-Paul; Sanen, Kathleen; Ameloot, Marcel; Ratautaite, Vilma; Kaseta, Vytautas; Biziuleviciene, Gene; Ramanaviciene, Almira; Nesladek, Milos; Ramanavicius, Arunas

    2015-01-25

    The aim of this study was to assess the impact of nanocrystalline diamond (NCD) thin coatings on neural cell adhesion and proliferation. NCD was fabricated on fused silica substrates by microwave plasma chemical vapor deposition (MPCVD) method. Different surface terminations were performed through exposure to reactive hydrogen and by UV induced oxidation during ozone treatment. Boron doped NCD coatings were also prepared and investigated. NCD surface wettability was determined by contact angle measurement. To assess biocompatibility of the NCD coatings, the neuroblastoma SH-SY5Y cell line was used. Cells were plated directly onto diamond surfaces and cultured in medium with or without fetal bovine serum (FBS), in order to evaluate the ability of cells to adhere and to proliferate. The obtained results showed that these cells adhered and proliferated better on NCD surfaces than on the bare fused silica. The cell proliferation on NCD in medium with and without FBS after 48h from plating was on average, respectively, 20 and 58% higher than that on fused silica, irrespective of NCD surface modification. Our results showed that the hydrogenated, oxygenated and boron-doped NCD coatings can be used for biomedical purposes, especially where good optical transparency is required.

  10. 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

  11. Adhesion, Proliferation and Migration of NIH/3T3 Cells on Modified Polyaniline Surfaces.

    PubMed

    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

  12. 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.

  13. Vitamin B6 Modifies the Immune Cross-Talk between Mononuclear and Colon Carcinoma Cells.

    PubMed

    Bessler, H; Djaldetti, M

    2016-01-01

    The role of vitamin B6 as a key component in a number of biological events has been well established. Based on the relationship between chronic inflammation and carcinogenesis on the one hand, and the interaction between immune and cancer cells expressed by modulated cytokine production on the other hand, the aim of the present work was to examine the possibility that vitamin B6 affects cancer development by an interference in the cross-talk between human peripheral blood mononuclear cells (PBMC) and those from two colon carcinoma cell lines. Both non-stimulated PBMC and mononuclear cells induced for cytokine production by HT-29 and RKO cells from human colon carcinoma lines were incubated without and with 4, 20 and 100 μg/ml of pyridoxal hydrochloride (vitamin B6) and secretion of TNF-α, IL-1β, IL-6, IFN-γ, IL-10, and IL-1ra was examined. Vit B6 caused a dose-dependent decrease in production of all cytokines examined, except for that of IL-1ra. The results indicate that vitamin B6 exerts an immunomodulatory effect on human PBMC. The finding that production of inflammatory cytokines is more pronounced when PBMC are in contact with malignant cells and markedly inhibited by the vitamin suggests an additional way by which vitamin B6 may exert its carcinopreventive effect. PMID:27085010

  14. Adhesion, Proliferation and Migration of NIH/3T3 Cells on Modified Polyaniline Surfaces.

    PubMed

    Rejmontová, Petra; Capáková, Zdenka; Mikušová, Nikola; Maráková, Nela; Kašpárková, Věra; Lehocký, Marián; Humpolíček, Petr

    2016-09-15

    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.

  15. Comparative antitumor effect among GM-CSF, IL-12 and GM-CSF+IL-12 genetically modified tumor cell vaccines.

    PubMed

    Miguel, A; Herrero, M J; Sendra, L; Botella, R; Algás, R; Sánchez, M; Aliño, S F

    2013-10-01

    Genetically modified cells have been shown to be one of the most effective cancer vaccine strategies. An evaluation is made of the efficacy of both preventive and therapeutic antitumor vaccines against murine melanoma, using C57BL/6 mice and irradiated B16 tumor cells expressing granulocyte and macrophage colony-stimulating factor (GM-CSF), interleukin-12 (IL-12) or both. Tumor was transplanted by the injection of wild-type B16 cells. Tumor growth and survival were measured to evaluate the efficacy of vaccination. Specific humoral response and immunoglobulin G (IgG) switch were evaluated measuring total IgG and IgG1 and IgG2a subtypes against tumor membrane proteins of B16 cells. In preventive vaccination, all treated groups showed delayed tumor growth. In addition, the group vaccinated to express only GM-CSF achieved 100% animal survival (P<0.005). Vaccination with GM-CSF+IL-12-producing B16 cells yielded lesser results (60% survival, P<0.005). Furthermore, all surviving animals remained disease-free after second tumor implantation 1 year later. The therapeutic vaccination strategies resulted in significantly delayed tumor growth, mainly using B16 cells producing GM-CSF+IL-12 cytokines, with 70% tumor growth inhibition (P<0.001)-although none of the animals reached overall survival. The results obtained suggest that the GM-CSF+IL-12 combination only increases the efficacy of therapeutic vaccines. No differences in classical regulatory T cells were found among the different groups.

  16. Utilization of modified surfactant-associated protein B for delivery of DNA to airway cells in culture.

    PubMed Central

    Baatz, J E; Bruno, M D; Ciraolo, P J; Glasser, S W; Stripp, B R; Smyth, K L; Korfhagen, T R

    1994-01-01

    Pulmonary surfactant lines the airway epithelium and creates a potential barrier to successful transfection of the epithelium in vivo. Based on the functional properties of pulmonary surfactant protein B (SP-B) and the fact that this protein is neither toxic nor immunogenic in the airway, we hypothesized that SP-B could be modified to deliver DNA to airway cells. We have modified native bovine SP-B by the covalent linkage of poly(lysine) (average molecular mass of 3.3 or 10 kDa) to the N terminus of SP-B and formed complexes between a test plasmid and the modified SP-B. Transfection efficiency was determined by transfection of pulmonary adenocarcinoma cells (H441) in culture with the test plasmid pCPA-RSV followed by measurement of activity of the reporter gene encoding chloramphenicol acetyltransferase (CAT). Transfections were performed with DNA.protein complexes using poly(lysine)10kDa-SP-B ([Lys]10kDa-SP-B) or poly(lysine)3.3kDa-SP-B ([Lys]3.3kDa-SP-B), and results were compared with transfections using unmodified poly(lysine).DNA, unmodified SP-B.DNA, or DNA only. For [Lys]10kDa-SP-B.pCPA-RSV preparations, CAT activity was readily detectable above the background of [Lys]3.3kDa-SP-B or unmodified SP-B. The SP-B-poly(lysine) conjugates were effective over a broad range of protein-to-DNA molar ratios, although they were optimal at approximately 500:1-1000:1. Transfection efficiency varied with the tested cell line but was not specific to airway cells. Addition of replication-defective adenovirus to the [Lys]10kDa-SP-B.pCPA-RSV complex enhanced CAT activity about 30-fold with respect to that produced by the [Lys]10kDa-SP-B.pCPA-RSV complex alone. This increase suggests routing of the adenoviral.[Lys]10kDa-SP-B.pCPA-RSV complex through an endosomal pathway. Effects of covalent modification on the secondary structure of SP-B were examined by Fourier transform infrared spectrometry (FTIR). Results of FTIR indicated that the conformation of [Lys]10kDa-SP-B was

  17. Mesenchymal stem cell response to topographically modified CoCrMo.

    PubMed

    Logan, Niall; Bozec, Laurent; Traynor, Alison; Brett, Peter

    2015-12-01

    Surface roughness on implant materials has been shown to be highly influential on the behavior of osteogenic cells. Four surface topographies were engineered on cobalt chromium molybdenum (CoCrMo) in order to examine this influence on human mesenchymal stem cells (MSC). These treatments were smooth polished (SMO), acid etched (AE) using HCl 7.4% and H2SO4 76% followed by HNO3 30%, sand blasted, and acid etched using either 50 μm Al2O3 (SLA50) or 250 μm Al2 O3 grit (SLA250). Characterization of the surfaces included energy dispersive X-ray analysis (EDX), contact angle, and surface roughness analysis. Human MSCs were cultured onto the four CoCrMo substrates and markers of cell attachment, retention, proliferation, cytotoxicity, and osteogenic differentiation were studied. Residual aluminum was observed on both SLA surfaces although this appeared to be more widely spread on SLA50, whilst SLA250 was shown to have the roughest topography with an Ra value greater than 1 μm. All substrates were shown to be largely non-cytotoxic although both SLA surfaces were shown to reduce cell attachment, whilst SLA50 also delayed cell proliferation. In contrast, SLA250 stimulated a good rate of proliferation resulting in the largest cell population by day 21. In addition, SLA250 stimulated enhanced cell retention, calcium deposition, and hydroxyapatite formation compared to SMO (p < 0.05). The enhanced response stimulated by SLA250 surface modification may prove advantageous for increasing the bioactivity of implants formed of CoCrMo. PMID:26015290

  18. [In vitro targeting effect of lactoferrin modified PEGylated liposomes for hepatoma cells].

    PubMed

    Wei, Min-yan; Zou, Qi; Wu, Chuan-bin; Xu, Yue-hong

    2015-10-01

    A lactoferrin-containing PEGylated liposome system (Lf-PLS) was developed and tested in vitro as a hepatoma-targeting drug delivery system. PEGylated liposomes (PLS) were successfully prepared using the thin film hydration method with peglipid post insertion. Lf was covalently conjugated onto the carboxyl terminal of DSPE-PEG2000-COOH on liposomes. Coumarin-6 was used to trace Lf-PLS with fluorescence. The cellular uptake of this system was carried out in asialoglycoprotein receptor (ASGPR) positive HepG2 cells via confocal microscopy and flow cytometry. The Lf-PLS liposome was observed as spherical or oval vesicles with the particle size around 130 nm, zeta potential about -30 mV and encapsulation efficiency more than 80%. The confocal microscopy images and flow cytometry data demonstrated that Lf-PLS resulted in significantly higher cell association by ASGPR positive HepG2 cells compared to PLS. The association between Lf-PLS and cells were dependent on the concentration, time and temperature, which was inhibited by pre-incubation with excessive free Lf. The results suggest that Lf-PLS has a good targeting effect on HepG2 cells in vitro. The targeting mechanism may be related to the specific binding of Lf and ASGPR on HepG2 cells, which guides Lf-PLS to the cell surface to induce an active endocytosis process. All these results demonstrated that Lf-PLS might be a potential drug delivery system in targeting hepatocellular carcinoma, which deserves more research on its targeting ability, antitumor efficiency, and metabolism in vivo for treatment of hepatomacellular carcinoma.

  19. Mesenchymal stem cell response to topographically modified CoCrMo

    PubMed Central

    Logan, Niall; Bozec, Laurent; Traynor, Alison

    2015-01-01

    Abstract Surface roughness on implant materials has been shown to be highly influential on the behavior of osteogenic cells. Four surface topographies were engineered on cobalt chromium molybdenum (CoCrMo) in order to examine this influence on human mesenchymal stem cells (MSC). These treatments were smooth polished (SMO), acid etched (AE) using HCl 7.4% and H2SO4 76% followed by HNO3 30%, sand blasted, and acid etched using either 50 μm Al2O3 (SLA50) or 250 μm Al2O3 grit (SLA250). Characterization of the surfaces included energy dispersive X‐ray analysis (EDX), contact angle, and surface roughness analysis. Human MSCs were cultured onto the four CoCrMo substrates and markers of cell attachment, retention, proliferation, cytotoxicity, and osteogenic differentiation were studied. Residual aluminum was observed on both SLA surfaces although this appeared to be more widely spread on SLA50, whilst SLA250 was shown to have the roughest topography with an R a value greater than 1 μm. All substrates were shown to be largely non‐cytotoxic although both SLA surfaces were shown to reduce cell attachment, whilst SLA50 also delayed cell proliferation. In contrast, SLA250 stimulated a good rate of proliferation resulting in the largest cell population by day 21. In addition, SLA250 stimulated enhanced cell retention, calcium deposition, and hydroxyapatite formation compared to SMO (p < 0.05). The enhanced response stimulated by SLA250 surface modification may prove advantageous for increasing the bioactivity of implants formed of CoCrMo. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 103A: 3747–3756, 2015. PMID:26015290

  20. Cell-penetrating peptide-doxorubicin conjugate loaded NGR-modified nanobubbles for ultrasound triggered drug delivery.

    PubMed

    Lin, Wen; Xie, Xiangyang; Deng, Jianping; Liu, Hui; Chen, Ying; Fu, Xudong; Liu, Hong; Yang, Yang

    2016-01-01

    A new drug-targeting system for CD13(+) tumors has been developed, based on ultrasound-sensitive nanobubbles (NBs) and cell-permeable peptides (CPPs). Here, the CPP-doxorubicin conjugate (CPP-DOX) was entrapped in the asparagine-glycine-arginine (NGR) peptide modified NB (CPP-DOX/NGR-NB) and the penetration of CPP-DOX was temporally masked; local ultrasound stimulation could trigger the CPP-DOX release from NB and activate its penetration. The CPP-DOX/NGR-NBs had particle sizes of about 200 nm and drug entrapment efficiency larger than 90%. In vitro release results showed that over 85% of the encapsulated DOX or CPP-DOX would release from NBs in the presence of ultrasound, while less than 1.5% of that (30 min) without ultrasound. Cell experiments showed the higher cellular CPP-DOX uptake of CPP-DOX/NGR-NB among the various NB formulations in Human fibrosarcoma cells (HT-1080, CD13(+)). The CPP-DOX/NGR-NB with ultrasound treatment exhibited an increased cytotoxic activity than the one without ultrasound. In nude mice xenograft of HT-1080 cells, CPP-DOX/NGR-NB with ultrasound showed a higher tumor inhibition effect (3.1% of T/C%, day 24), longer median survival time (50 days) and excellent body safety compared with the normal DOX injection group. These results indicate that the constructed vesicle would be a promising drug delivery system for specific cancer treatment.

  1. Cyclodextrin-Modified Porous Silicon Nanoparticles for Efficient Sustained Drug Delivery and Proliferation Inhibition of Breast Cancer Cells.

    PubMed

    Correia, Alexandra; Shahbazi, Mohammad-Ali; Mäkilä, Ermei; Almeida, Sérgio; Salonen, Jarno; Hirvonen, Jouni; Santos, Hélder A

    2015-10-21

    Over the past decade, the potential of polymeric structures has been investigated to overcome many limitations related to nanosized drug carriers by modulating their toxicity, cellular interactions, stability, and drug-release kinetics. In this study, we have developed a successful nanocomposite consisting of undecylenic acid modified thermally hydrocarbonized porous silicon nanoparticles (UnTHCPSi NPs) loaded with an anticancer drug, sorafenib, and surface-conjugated with heptakis(6-amino-6-deoxy)-β-cyclodextrin (HABCD) to show the impact of the surface polymeric functionalization on the physical and biological properties of the drug-loaded nanoparticles. Cytocompatibility studies showed that the UnTHCPSi-HABCD NPs were not toxic to breast cancer cells. HABCD also enhanced the suspensibility and both the colloidal and plasma stabilities of the UnTHCPSi NPs. UnTHCPSi-HABCD NPs showed a significantly increased interaction with breast cancer cells compared to bare NPs and also sustained the drug release. Furthermore, the sorafenib-loaded UnTHCPSi-HABCD NPs efficiently inhibited cell proliferation of the breast cancer cells.

  2. Cyclodextrin-Modified Porous Silicon Nanoparticles for Efficient Sustained Drug Delivery and Proliferation Inhibition of Breast Cancer Cells.

    PubMed

    Correia, Alexandra; Shahbazi, Mohammad-Ali; Mäkilä, Ermei; Almeida, Sérgio; Salonen, Jarno; Hirvonen, Jouni; Santos, Hélder A

    2015-10-21

    Over the past decade, the potential of polymeric structures has been investigated to overcome many limitations related to nanosized drug carriers by modulating their toxicity, cellular interactions, stability, and drug-release kinetics. In this study, we have developed a successful nanocomposite consisting of undecylenic acid modified thermally hydrocarbonized porous silicon nanoparticles (UnTHCPSi NPs) loaded with an anticancer drug, sorafenib, and surface-conjugated with heptakis(6-amino-6-deoxy)-β-cyclodextrin (HABCD) to show the impact of the surface polymeric functionalization on the physical and biological properties of the drug-loaded nanoparticles. Cytocompatibility studies showed that the UnTHCPSi-HABCD NPs were not toxic to breast cancer cells. HABCD also enhanced the suspensibility and both the colloidal and plasma stabilities of the UnTHCPSi NPs. UnTHCPSi-HABCD NPs showed a significantly increased interaction with breast cancer cells compared to bare NPs and also sustained the drug release. Furthermore, the sorafenib-loaded UnTHCPSi-HABCD NPs efficiently inhibited cell proliferation of the breast cancer cells. PMID:26440739

  3. Sustained Delivery Growth Factors with Polyethyleneimine‐Modified Nanoparticles Promote Embryonic Stem Cells Differentiation and Liver Regeneration

    PubMed Central

    Wang, Meiyan; Yang, Xiaomei; Zhang, Peng; Cai, Lei; Yang, Xibin; Chen, Youwei; Jing, Yuanya; Kong, Jilie

    2016-01-01

    Stem‐cell‐derived hepatocyte transplantation is considered as a potential method for the therapy of acute and chronic liver failure. However, the low efficiency of differentiation into mature and functional hepatocytes remains a major challenge for clinical applications. By using polyethyleneimine‐modified silica nanoparticles, this study develops a system for sustained delivery of growth factors, leading to induce hepatocyte‐like cells (iHeps) from mouse embryonic stem cells (mESCs) and improve the expression of endoderm and hepatocyte‐specific genes and proteins significantly, thus producing a higher population of functional hepatocytes in vitro. When transplanted into liver‐injured mice after four weeks, mESC‐derived definitive endoderm cells treated with this delivery system show higher integration efficiency into the host liver, differentiated into iHeps in vivo and significantly restored the injured liver. Therefore, these findings reveal the multiple advantages of functionalized nanoparticles to serve as efficient delivery platforms to promote stem cell differentiation in the regenerative medicine.

  4. Gene expression profiles of human adipose tissue-derived mesenchymal stem cells are modified by cell culture density.

    PubMed

    Kim, Dae Seong; Lee, Myoung Woo; 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/cm(2). 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.