297 research outputs found
Effect of Chromatographic Conditions on Supercoiled Plasmid DNA Stability and Bioactivity
Funding: This work was supported by FEDER funds through the POCI—COMPETE 2020 Operational Programme Competitiveness and Internationalization in Axis I—Strengthening research, technological development, and innovation (Project POCI-01-0145-FEDER-007491), and National Funds by FCT Foundation for Science and Technology (Project UID/Multi /00709/2013). This work was also developed within the scope of the project CICECO-Aveiro Institute of Materials, FCT Ref. UID/CTM/50011/2019, financed by national funds through the FCT/MCTES. G.M. Azevedo acknowledges the support and fellowship of Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq/203482/2014-0). J.F.A. Valente acknowledges the PhD fellowship (Ref SFRH/BD/96809/2013) from FCT.Acknowledgments: The authors would like to thank Thomas Roberts for providing the pcDNA3–FLAG–p53
construct through Addgene, ref: 10838.The dysfunction of the tumor suppressor gene TP53 has been associated with the pathogenesis of the majority of the cases of cancer reported to date, leading the cell to acquire different features known as the cancer hallmarks. In normal situations, the protein p53 protects the cells against tumorigenesis. By detecting metabolic stress or DNA damage in response to stress, p53 can lead the cell to senescence, autophagy, cell cycle arrest, DNA repair, and apoptosis. Thus, in the case of p53 mutations, it is reasonable to assume that the reestablishment of its function, may restrain the proliferation of cancer cells. The concept of cancer gene therapy can be based on this assumption, and suitable biotechnological approaches must be explored to assure the preparation of gene-based biopharmaceuticals. Although numerous procedures have already been established to purify supercoiled plasmid DNA (sc pDNA), the therapeutic application is highly dependent on the biopharmaceutical’s activity, which can be affected by the chromatographic conditions used. Thus, the present work aims at comparing quality and in vitro activity of the supercoiled (sc) isoform of the p53 encoding plasmid purified by three different amino acids-based chromatographic strategies, involving histidine–agarose, arginine–macroporous, and histidine–monolith supports. The B-DNA topology was maintained in all purified pDNA samples, but their bioactivity, related to the induction of protein p53 expression and apoptosis in cancer cells, was higher with arginine–macroporous support, followed by histidine–monolith and histidine–agarose. Despite the purity degree of 92% and recovery yield of 43% obtained with arginine–macroporous, the sc pDNA sample led to a higher expression level of the therapeutic p53 protein (58%) and, consequently, induced a slightly higher apoptotic effect (27%) compared with sc pDNA samples obtained with histidine–monolithic support (26%) and histidine–agarose support (24%). This behavior can be related to the mild chromatographic conditions used with arginine–macroporous support, which includes the use of low salt concentrations, at neutral pH and lower temperatures, when compared to the high ionic strength of ammonium sulfate and acidic pH used with histidine-based supports. These results can contribute to field of biopharmaceutical preparation, emphasizing the need to control several experimental conditions while adapting and selecting the methodologies that enable the use of milder conditions as this can have a significant impact on pDNA stability and biological activity.info:eu-repo/semantics/publishedVersio
Solubility of H2S in ammonium-based ionic liquids
This work is inserted in a research program that consists mainly in the experimental and theoretical study of gas/liquid solubility in ionic liquids (ILs). In this study the experimental data of hydrogen sulfide solubility in ammonium-based ionic liquids, namely 2-hydroxyethylammonium acetate [2-HEA][Ace], bis(2-hydroxyethyl)ammonium acetate [B-2-HEA][Ace] and 2-hydroxyethyldiethylammonium hydrogen diacetate [2-HEDEA][H(Ace)2], were determined using a volumetric method in the 298 K to 318 K temperature range and at atmospheric pressure. The ionic liquids are functionalized with the OH group in the ammonium-based cations, in order to study the influence of hydroxyl group in the formation of hydrogen bonds between the IL-IL and IL-gas. The data gathered is modelled with the Cubic Plus Association Equation of State (CPA EoS), considering the association schemes four-sites (4C) for hydrogen sulfide and two-sites (2B) for the ILs ([2-HEA][Ace], [B-2-HEA][Ace] and [2-HEDEA][H(Ace)2]).publishe
Electron spin as a spectrometer of nuclear spin noise and other fluctuations
This chapter describes the relationship between low frequency noise and
coherence decay of localized spins in semiconductors. Section 2 establishes a
direct relationship between an arbitrary noise spectral function and spin
coherence as measured by a number of pulse spin resonance sequences. Section 3
describes the electron-nuclear spin Hamiltonian, including isotropic and
anisotropic hyperfine interactions, inter-nuclear dipolar interactions, and the
effective Hamiltonian for nuclear-nuclear coupling mediated by the electron
spin hyperfine interaction. Section 4 describes a microscopic calculation of
the nuclear spin noise spectrum arising due to nuclear spin dipolar flip-flops
with quasiparticle broadening included. Section 5 compares our explicit
numerical results to electron spin echo decay experiments for phosphorus doped
silicon in natural and nuclear spin enriched samples.Comment: Book chapter in "Electron spin resonance and related phenomena in low
dimensional structures", edited by Marco Fanciulli. To be published by
Springer-Verlag in the TAP series. 35 pages, 9 figure
Theory of nuclear induced spectral diffusion: Spin decoherence of phosphorus donors in Si and GaAs quantum dots
We propose a model for spectral diffusion of localized spins in
semiconductors due to the dipolar fluctuations of lattice nuclear spins. Each
nuclear spin flip-flop is assumed to be independent, the rate for this process
being calculated by a method of moments. Our calculated spin decoherence time
ms for donor electron spins in Si:P is a factor of two longer than
spin echo decay measurements. For P nuclear spins we show that spectral
diffusion is well into the motional narrowing regime. The calculation for GaAs
quantum dots gives s depending on the quantum dot size. Our
theory indicates that nuclear induced spectral diffusion should not be a
serious problem in developing spin-based semiconductor quantum computer
architectures.Comment: 15 pages, 9 figures. Accepted for publication in Phys. Rev.
Targeting Tyrosine Phosphatases by 3-Bromopyruvate Overcomes Hyperactivation of Platelets from Gastrointestinal Cancer Patients
Venous thromboembolism (VTE) is one of the most common causes of cancer related
mortality. It has been speculated that hypercoagulation in cancer patients is triggered by direct or
indirect contact of platelets with tumor cells, however the underlying molecular mechanisms involved
are currently unknown. Unraveling these mechanisms may provide potential avenues for preventing
platelet-tumor cell aggregation. Here, we investigated the role of protein tyrosine phosphatases
in the functionality of platelets in both healthy individuals and patients with gastrointestinal
cancer, and determined their use as a target to inhibit platelet hyperactivity. This is the first study
to demonstrate that platelet agonists selectively activate low molecular weight protein tyrosine
phosphatase (LMWPTP) and PTP1B, resulting in activation of Src, a tyrosine kinase known to
contribute to several platelet functions. Furthermore, we demonstrate that these phosphatases are
a target for 3-bromopyruvate (3-BP), a lactic acid analog currently investigated for its use in the
treatment of various metabolic tumors. Our data indicate that 3-BP reduces Src activity, platelet
aggregation, expression of platelet activation makers and platelet-tumor cell interaction. Thus,
Search for direct production of charginos and neutralinos in events with three leptons and missing transverse momentum in √s = 7 TeV pp collisions with the ATLAS detector
A search for the direct production of charginos and neutralinos in final states with three electrons or muons and missing transverse momentum is presented. The analysis is based on 4.7 fb−1 of proton–proton collision data delivered by the Large Hadron Collider and recorded with the ATLAS detector. Observations are consistent with Standard Model expectations in three signal regions that are either depleted or enriched in Z-boson decays. Upper limits at 95% confidence level are set in R-parity conserving phenomenological minimal supersymmetric models and in simplified models, significantly extending previous results
Jet size dependence of single jet suppression in lead-lead collisions at sqrt(s(NN)) = 2.76 TeV with the ATLAS detector at the LHC
Measurements of inclusive jet suppression in heavy ion collisions at the LHC
provide direct sensitivity to the physics of jet quenching. In a sample of
lead-lead collisions at sqrt(s) = 2.76 TeV corresponding to an integrated
luminosity of approximately 7 inverse microbarns, ATLAS has measured jets with
a calorimeter over the pseudorapidity interval |eta| < 2.1 and over the
transverse momentum range 38 < pT < 210 GeV. Jets were reconstructed using the
anti-kt algorithm with values for the distance parameter that determines the
nominal jet radius of R = 0.2, 0.3, 0.4 and 0.5. The centrality dependence of
the jet yield is characterized by the jet "central-to-peripheral ratio," Rcp.
Jet production is found to be suppressed by approximately a factor of two in
the 10% most central collisions relative to peripheral collisions. Rcp varies
smoothly with centrality as characterized by the number of participating
nucleons. The observed suppression is only weakly dependent on jet radius and
transverse momentum. These results provide the first direct measurement of
inclusive jet suppression in heavy ion collisions and complement previous
measurements of dijet transverse energy imbalance at the LHC.Comment: 15 pages plus author list (30 pages total), 8 figures, 2 tables,
submitted to Physics Letters B. All figures including auxiliary figures are
available at
http://atlas.web.cern.ch/Atlas/GROUPS/PHYSICS/PAPERS/HION-2011-02
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