Accurately tracking single-cell movement trajectories in microfluidic cell sorting devices.

Science.gov (United States)

Jeong, Jenny; Frohberg, Nicholas J; Zhou, Enlu; Sulchek, Todd; Qiu, Peng

2018-01-01

Microfluidics are routinely used to study cellular properties, including the efficient quantification of single-cell biomechanics and label-free cell sorting based on the biomechanical properties, such as elasticity, viscosity, stiffness, and adhesion. Both quantification and sorting applications require optimal design of the microfluidic devices and mathematical modeling of the interactions between cells, fluid, and the channel of the device. As a first step toward building such a mathematical model, we collected video recordings of cells moving through a ridged microfluidic channel designed to compress and redirect cells according to cell biomechanics. We developed an efficient algorithm that automatically and accurately tracked the cell trajectories in the recordings. We tested the algorithm on recordings of cells with different stiffness, and showed the correlation between cell stiffness and the tracked trajectories. Moreover, the tracking algorithm successfully picked up subtle differences of cell motion when passing through consecutive ridges. The algorithm for accurately tracking cell trajectories paves the way for future efforts of modeling the flow, forces, and dynamics of cell properties in microfluidics applications.

Accurately tracking single-cell movement trajectories in microfluidic cell sorting devices.

Directory of Open Access Journals (Sweden)

Jenny Jeong

Full Text Available Microfluidics are routinely used to study cellular properties, including the efficient quantification of single-cell biomechanics and label-free cell sorting based on the biomechanical properties, such as elasticity, viscosity, stiffness, and adhesion. Both quantification and sorting applications require optimal design of the microfluidic devices and mathematical modeling of the interactions between cells, fluid, and the channel of the device. As a first step toward building such a mathematical model, we collected video recordings of cells moving through a ridged microfluidic channel designed to compress and redirect cells according to cell biomechanics. We developed an efficient algorithm that automatically and accurately tracked the cell trajectories in the recordings. We tested the algorithm on recordings of cells with different stiffness, and showed the correlation between cell stiffness and the tracked trajectories. Moreover, the tracking algorithm successfully picked up subtle differences of cell motion when passing through consecutive ridges. The algorithm for accurately tracking cell trajectories paves the way for future efforts of modeling the flow, forces, and dynamics of cell properties in microfluidics applications.

β-Arrestin-2 knockout prevents development of cellular μ-opioid receptor tolerance but does not affect opioid-withdrawal-related adaptations in single PAG neurons.

Science.gov (United States)

Connor, M; Bagley, E E; Chieng, B C; Christie, M J

2015-01-01

Tolerance to the behavioural effects of morphine is blunted in β-arrestin-2 knockout mice, but opioid withdrawal is largely unaffected. The cellular mechanisms of tolerance have been studied in some neurons from β-arrestin-2 knockouts, but tolerance and withdrawal mechanisms have not been examined at the cellular level in periaqueductal grey (PAG) neurons, which are crucial for central tolerance and withdrawal phenomena. μ-Opioid receptor (MOPr) inhibition of voltage-gated calcium channel currents (ICa ) was examined by patch-clamp recordings from acutely dissociated PAG neurons from wild-type and β-arrestin-2 knockout mice treated chronically with morphine (CMT) or vehicle. Opioid withdrawal-induced activation of GABA transporter type 1 (GAT-1) currents was determined using perforated patch recordings from PAG neurons in brain slices. MOPr inhibition of ICa in PAG neurons was unaffected by β-arrestin-2 deletion. CMT impaired coupling of MOPrs to ICa in PAG neurons from wild-type mice, but this cellular tolerance was not observed in neurons from CMT β-arrestin-2 knockouts. However, β-arrestin-2 knockouts displayed similar opioid-withdrawal-induced activation of GAT-1 currents as wild-type PAG neurons. In β-arrestin-2 knockout mice, the central neurons involved in the anti-nociceptive actions of opioids also fail to develop cellular tolerance to opioids following chronic morphine. The results also provide the first cellular physiological evidence that opioid withdrawal is not disrupted by β-arrestin-2 deletion. However, the unaffected basal sensitivity to opioids in PAG neurons provides further evidence that changes in basal MOPr sensitivity cannot account for the enhanced acute nociceptive response to morphine reported in β-arrestin-2 knockouts. This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2. © 2014 The British

KnockoutJS blueprints

CERN Document Server

Russo, Carlo

2015-01-01

If you are a JavaScript developer and already know the basics of KnockoutJS and you want to get the most out of it, then this book is for you. This book will help in your transition from a small site to a large web application that is easily maintainable.

Characterization of the first knock-out aldh7a1 zebrafish model for pyridoxine-dependent epilepsy using CRISPR-Cas9 technology.

Science.gov (United States)

Zabinyakov, Nikita; Bullivant, Garrett; Cao, Feng; Fernandez Ojeda, Matilde; Jia, Zheng Ping; Wen, Xiao-Yan; Dowling, James J; Salomons, Gajja S; Mercimek-Andrews, Saadet

2017-01-01

Pyridoxine dependent epilepsy (PDE) is caused by likely pathogenic variants in ALDH7A1 (PDE-ALDH7A1) and inherited autosomal recessively. Neurotoxic alpha-amino adipic semialdehyde (alpha-AASA), piperideine 6-carboxylate and pipecolic acid accumulate in body fluids. Neonatal or infantile onset seizures refractory to anti-epileptic medications are clinical features. Treatment with pyridoxine, arginine and lysine-restricted diet does not normalize neurodevelopmental outcome or accumulation of neurotoxic metabolites. There is no animal model for high throughput drug screening. For this reason, we developed and characterized the first knock-out aldh7a1 zebrafish model using CRISPR-Cas9 technology. Zebrafish aldh7a1 mutants were generated by using a vector free method of CRISPR-Cas9 mutagenesis. Genotype analysis of aldh7a1 knock-out zebrafish was performed by high resolution melt analysis, direct sequencing and QIAxcel system. Electroencephalogram was performed. Alpha-AASA, piperideine 6-carboxylate and pipecolic acid, were measured by liquid chromatography-tandem mass spectrometry. Our knock-out aldh7a1 zebrafish has homozygous 5 base pair (bp) mutation in ALDH7A1. Knock-out aldh7a1 embryos have spontaneous rapid increase in locomotion and a rapid circling swim behavior earliest 8-day post fertilization (dpf). Electroencephalogram revealed large amplitude spike discharges compared to wild type. Knock-out aldh7a1 embryos have elevated alpha-AASA, piperideine 6-carboxylate and pipecolic acid compared to wild type embryos at 3 dpf. Knock-out aldh7a1 embryos showed no aldh7a1 protein by western blot compared to wild type. Our knock-out aldh7a1 zebrafish is a well characterized model for large-scale drug screening using behavioral and biochemical features and accurately recapitulates the human PDE-ALDH7A1 disease.

KnockoutJS web development

CERN Document Server

Farrar, John

2015-01-01

This book is for web developers and designers who work with HTML and JavaScript to help them manage data and interactivity with data using KnockoutJS. Knowledge about jQuery will be useful but is not necessary.

Theoretical analysis of knock-out release of fission products from nuclear fuels

International Nuclear Information System (INIS)

Yamagishi, S.

1975-01-01

The knock-out release of fission products is studied theoretically. The general equations of knock-out release are derived, assuming that a fission fragment passing through the surface of nuclear fuels knocks out a local region of the surface with an effective thickness and an effective cross-sectional area. Using these equations, the knock-out release of fission gases is calculated for various cases. The conditions under which the knock-out coefficients (the average number of uranium atoms knocked out by one fission fragment) is obtainable are clarified by experiments on the knock-out release of fission gases. A method of determining the effective thickness and the effective cross-sectional area of a knock-out region is proposed. (Auth.)

One-neutron knockout from Ne24-28 isotopes

CERN Document Server

Rodriguez-Tajes, C; Caamano, M; Faestermann, T; Cortina-Gil, D; Zhukov, M; Simon, H; Nilsson, T; Borge, M J G; Alvarez-Pol, H; Winkler, M; Prochazka, A; Nociforo, C; Weick, H; Kanungo, R; Perez-Loureiro, D; Kurtukian, T; Suemmerer, K; Eppinger, K; Perea, A; Chatillon, A; Maierbeck, P; Benlliure, J; Pascual-Izarra, C; Gernhaeuser, R; Geissel, H; Aumann, T; Kruecken, R; Larsson, K; Tengblad, O; Benjamim, E; Jonson, B; Casarejos, E

2010-01-01

One-neutron knockout reactions of Ne24-28 in a beryllium target have been studied in the Fragment Separator (FRS), at GSI. The results include inclusive one-neutron knockout cross-sections as well as longitudinal-momentum distributions of the knockout fragments. The ground-state structure of the neutron-rich neon isotopes was obtained from an analysis of the measured momentum distributions. The results indicate that the two heaviest isotopes, Ne-27 and Ne-28, are dominated by a configuration in which a s(1/2) neutron is coupled to an excited state of the Ne-26 and Ne-27 core, respectively. (C) 2010 Elsevier B.V. All rights reserved.

Generation of a heterozygous knockout human embryonic stem cell line for the OCIAD1 locus using CRISPR/CAS9 mediated targeting: BJNhem20-OCIAD1-CRISPR-20

Directory of Open Access Journals (Sweden)

Deeti K. Shetty

2016-03-01

Full Text Available Ovarian carcinoma immuno-reactive antigen domain containing 1(OCIAD1 single copy was knocked out generating an OCIAD1 heterozygous knockout human embryonic stem line named BJNhem20-OCIAD1-CRISPR-20. The line was generated using CRISPR-Cas9D10A double nickase knockout strategy (Mali et al., 2013.

Overlap knock-out effects in the CERN intersecting storage rings (ISR)

CERN Document Server

Gourber, J P; Myers, S

1977-01-01

Overlap knock-out arises from an overlap between frequencies present in a bunched beam and the betatron frequencies in a stack. The 'single ring' effect in the interaction of a bunched beam with a stack in the same ring. Here the coupling forces are fairly linear and are transmitted by machine elements. The 'two-ring' effect is the interaction of a bunched beam with a stack in the other ring. Here the coupling forces are nonlinear since they are produced by the beam-beam interaction. A brief outline of the general theory of these effects is given. The single ring and two-ring dipole effects have been observed and shown to cause a large increase in the transverse size of the stacked beam. (4 refs).

KnockoutJS essentials

CERN Document Server

Ferrando, Jorge

2015-01-01

If you are a JavaScript developer who has been using DOM manipulation libraries such as Mootools or Scriptaculous, and you want go further in modern JavaScript development with a simple and well-documented library, then this book is for you. Learning how to use Knockout will be perfect as your next step towards building JavaScript applications that respond to user interaction.

One-neutron knockout from {sup 24-28}Ne isotopes

Energy Technology Data Exchange (ETDEWEB)

Rodriguez-Tajes, C., E-mail: carme.rodriguez@usc.e [Departamento de Fisica de Particulas, Universidade de Santiago de Compostela, 15782 Santiago de Compostela (Spain); Cortina-Gil, D.; Alvarez-Pol, H. [Departamento de Fisica de Particulas, Universidade de Santiago de Compostela, 15782 Santiago de Compostela (Spain); Aumann, T. [GSI Helmholtzzentrum fuer Schwerionenforschung, 64291 Darmstadt (Germany); Benjamim, E.; Benlliure, J. [Departamento de Fisica de Particulas, Universidade de Santiago de Compostela, 15782 Santiago de Compostela (Spain); Borge, M.J.G. [Instituto de Estructura de la Materia, CSIC, 28006 Madrid (Spain); Caamano, M.; Casarejos, E. [Departamento de Fisica de Particulas, Universidade de Santiago de Compostela, 15782 Santiago de Compostela (Spain); Chatillon, A. [GSI Helmholtzzentrum fuer Schwerionenforschung, 64291 Darmstadt (Germany); Eppinger, K.; Faestermann, T. [Physik Department E12, Technische Universitaet Muenchen, 85748 Garching (Germany); Gascon, M. [Departamento de Fisica de Particulas, Universidade de Santiago de Compostela, 15782 Santiago de Compostela (Spain); Geissel, H. [GSI Helmholtzzentrum fuer Schwerionenforschung, 64291 Darmstadt (Germany); Gernhaeuser, R. [Physik Department E12, Technische Universitaet Muenchen, 85748 Garching (Germany); Jonson, B. [Fundamental Fysik, Chalmers Tekniska Hoegskola, 412 96 Goeteborg (Sweden); PH Department, CERN, 1211 Geneve 23 (Switzerland); Kanungo, R. [Astronomy and Physics Department, Saint Mary' s University, Halifax, NS B3H 3C3 (Canada); Kruecken, R. [Physik Department E12, Technische Universitaet Muenchen, 85748 Garching (Germany); Kurtukian, T. [Departamento de Fisica de Particulas, Universidade de Santiago de Compostela, 15782 Santiago de Compostela (Spain); Larsson, K. [Fundamental Fysik, Chalmers Tekniska Hoegskola, 412 96 Goeteborg (Sweden)

2010-04-05

One-neutron knockout reactions of {sup 24-28}Ne in a beryllium target have been studied in the Fragment Separator (FRS), at GSI. The results include inclusive one-neutron knockout cross-sections as well as longitudinal-momentum distributions of the knockout fragments. The ground-state structure of the neutron-rich neon isotopes was obtained from an analysis of the measured momentum distributions. The results indicate that the two heaviest isotopes, {sup 27}Ne and {sup 28}Ne, are dominated by a configuration in which a s{sub 1/2} neutron is coupled to an excited state of the {sup 26}Ne and {sup 27}Ne core, respectively.

Eliminating graphs by means of parallel knock-out schemes

NARCIS (Netherlands)

Broersma, H.J.; Fomin, F.V.; Královic, R.; Woeginger, G.J.

2007-01-01

In 1997 Lampert and Slater introduced parallel knock-out schemes, an iterative process on graphs that goes through several rounds. In each round of this process, every vertex eliminates exactly one of its neighbors. The parallel knock-out number of a graph is the minimum number of rounds after which

Eliminating graphs by means of parallel knock-out schemes

NARCIS (Netherlands)

Broersma, Haitze J.; Fomin, F.V.; Královič, R.; Woeginger, Gerhard

In 1997 Lampert and Slater introduced parallel knock-out schemes, an iterative process on graphs that goes through several rounds. In each round of this process, every vertex eliminates exactly one of its neighbors. The parallel knock-out number of a graph is the minimum number of rounds after which

Selection-independent generation of gene knockout mouse embryonic stem cells using zinc-finger nucleases.

Directory of Open Access Journals (Sweden)

Anna Osiak

Full Text Available Gene knockout in murine embryonic stem cells (ESCs has been an invaluable tool to study gene function in vitro or to generate animal models with altered phenotypes. Gene targeting using standard techniques, however, is rather inefficient and typically does not exceed frequencies of 10(-6. In consequence, the usage of complex positive/negative selection strategies to isolate targeted clones has been necessary. Here, we present a rapid single-step approach to generate a gene knockout in mouse ESCs using engineered zinc-finger nucleases (ZFNs. Upon transient expression of ZFNs, the target gene is cleaved by the designer nucleases and then repaired by non-homologous end-joining, an error-prone DNA repair process that introduces insertions/deletions at the break site and therefore leads to functional null mutations. To explore and quantify the potential of ZFNs to generate a gene knockout in pluripotent stem cells, we generated a mouse ESC line containing an X-chromosomally integrated EGFP marker gene. Applying optimized conditions, the EGFP locus was disrupted in up to 8% of ESCs after transfection of the ZFN expression vectors, thus obviating the need of selection markers to identify targeted cells, which may impede or complicate downstream applications. Both activity and ZFN-associated cytotoxicity was dependent on vector dose and the architecture of the nuclease domain. Importantly, teratoma formation assays of selected ESC clones confirmed that ZFN-treated ESCs maintained pluripotency. In conclusion, the described ZFN-based approach represents a fast strategy for generating gene knockouts in ESCs in a selection-independent fashion that should be easily transferrable to other pluripotent stem cells.

High energy approximations for nuclear knockout form factors at small momentum transfer

International Nuclear Information System (INIS)

Amado, R.D.; Cannata, F.; Dedonder, J.P.

1985-01-01

We obtain an explicit approximate expression for the nucleon knockout form factor at small momentum transfer induced by a scalar probe in a single particle model in terms of the momentum space bound state wave function. Our form preserves the orthogonality constraint without using explicitly the final state scattering wave function. We examine the leading large momentum behavior of the momentum space wave function and of correction terms to our expression for the form factor in the case where the bound state is an s state

Quasi-free one nucleon knockout reactions on neutron-rich oxygen isotopes

Energy Technology Data Exchange (ETDEWEB)

Atar, Leyla; Aumann, Thomas [TU Darmstadt, Darmstadt (Germany); GSI, Darmstadt (Germany); Bertulani, Carlos [Texas A and M University-Commerce, Commerse (United States); Paschalis, Stefanos [TU Darmstadt, Darmstadt (Germany); Nociforo, Chiara [GSI, Darmstadt (Germany); Collaboration: R3B-Collaboration

2015-07-01

Recent experiments have shown a reduction of spectroscopic strengths to about 60-70% for stable nuclei. When going to drip lines this tendency is changing, loosely bound nucleons have spectroscopic strengths close unity while deeply bound nucleons have a large reduction of the strength. We aim to make a systematic study of spectroscopic factors (SF) of the Oxygen isotopes using quasi-free (p,2p) and (p,pn) knockout reactions in inverse kinematics. Quasi-free knockout reactions are a direct tool to study the occupancy and the location of valance and deeply bound single particle states. The Oxygen isotopes offer a large variation of separation energies which will allow us to obtain a qualitative and quantitative understanding of SF in a large variation of isospin asymmetry. For this we performed an experiment at the R3B-LAND setup at the GSI with secondary beams containing {sup 14-24}O. The {sup 16-18}O and {sup 21-23}O isotopes have been analyzed and the preliminary results will be presented. The results include the partial cross sections, gamma ray spectra of the residual fragments in coincidence, and the SF obtained via comparison with theory.

On the interpretation of (e,e'p) knock-out reaction

International Nuclear Information System (INIS)

Dieperink, A.E.L.

The basic physics in (e,e'p) knock-out reactions is illustrated assuming that the knocked-out proton can be treated as a plane wave (PWIA). Corrections for distortion and absorption of the outgoing proton can, in principle, be calculated to a good approximation with an optical potential. The spectral function is characterized in terms of its energy moments, the lowest of which can be incorporated in an independent particle shell model (IPSM): occupatiomn probability (zeroth moment) and the mean removal energy (centroid energy). Deviations from IPSM are discussed: binding energy sum rule, A=3 nuclei, 6 Li, and fragmentation of single-particle strength

Impaired social behavior in 5-HT3A receptor knockout mice

Directory of Open Access Journals (Sweden)

Laura A Smit-Rigter

2010-11-01

Full Text Available The 5-HT3 receptor is a ligand-gated ion channel expressed on interneurons throughout the brain. So far, analysis of the 5-HT3A knockout mouse revealed changes in nociceptive processing and a reduction in anxiety related behavior. Recently, it was shown that the 5-HT3 receptor is also expressed on Cajal-Retzius cells which play a key role in cortical development and that knockout mice lacking this receptor showed aberrant growth of the dendritic tree of cortical layer II/III pyramidal neurons. Other mouse models in which serotonergic signaling was disrupted during development showed similar morphological changes in the cortex, and in addition, also deficits in social behavior. Here, we subjected male and female 5-HT3A knockout mice and their non-transgenic littermates to several tests of social behavior. We found that 5-HT3A knockout mice display impaired social communication in the social transmission of food preference task. Interestingly, we showed that in the social interaction test only female 5-HT3A knockout mice spent less time in reciprocal social interaction starting after 5 minutes of testing. Moreover, we observed differences in preference for social novelty for male and female 5-HT3A knockout mice during the social approach test. However, no changes in olfaction, exploratory activity and anxiety were detected. These results indicate that the 5-HT3A knockout mouse displays impaired social behavior with specific changes in males and females, reminiscent to other mouse models in which serotonergic signaling is disturbed in the developing brain.

Interrater agreement of an observational tool to code knockouts and technical knockouts in mixed martial arts.

Science.gov (United States)

Lawrence, David W; Hutchison, Michael G; Cusimano, Michael D; Singh, Tanveer; Li, Luke

2014-09-01

Interrater agreement evaluation of a tool to document and code the situational factors and mechanisms of knockouts (KOs) and technical knockouts (TKOs) in mixed martial arts (MMA). Retrospective case series. Professional MMA matches from the Ultimate Fighting Championship-2006-2012. Two nonmedically trained independent raters. The MMA Knockout Tool (MMA-KT) consists of 20 factors and captures and codes information on match characteristics, situational context preceding KOs and TKOs, as well as describing competitor states during these outcomes. The MMA-KT also evaluates the mechanism of action and subsequent events surrounding a KO. The 2 raters coded 125 unique events for a total of 250 events. The 8 factors of Part A had an average κ of 0.87 (SD = 0.10; range = 0.65-0.98); 7 were considered "substantial" agreement and 1 "moderate." Part B consists of 12 factors with an average κ of 0.84 (SD = 0.16; range = 0.59-1.0); 7 classified as "substantial" agreement, 4 "moderate," and 1 "fair." The majority of the factors in the MMA-KT demonstrated substantial interrater agreement, with an average κ of 0.86 (SD = 0.13; range = 0.59-1.0). The MMA-KT is a reliable tool to extract and code relevant information to investigate the situational factors and mechanism of KOs and TKOs in MMA competitions.

RNaseT2 knockout rats exhibit hippocampal neuropathology and deficits in memory.

Science.gov (United States)

Sinkevicius, Kerstin W; Morrison, Thomas R; Kulkarni, Praveen; Cagliostro, Martha K Caffrey; Iriah, Sade; Malmberg, Samantha; Sabrick, Julia; Honeycutt, Jennifer A; Askew, Kim L; Trivedi, Malav; Ferris, Craig F

2018-05-10

RNASET2 deficiency in humans is associated with infant cystic leukoencephalopathy, which causes psychomotor impairment, spasticity, and epilepsy. A zebrafish mutant model suggests that loss of RNASET2 function leads to neurodegeneration due to the accumulation of non-degraded RNA in the lysosomes. The goal of this study was to characterize the first rodent model of RNASET2 deficiency. The brains of 3- and 12-month-old RNaseT2 knockout rats were studied using multiple magnetic resonance imaging modalities and behavioral tests. While T1 and T2 weighted images of RNaseT2 knockout rats exhibited no evidence of cystic lesions, the prefrontal cortex and hippocampal complex were enlarged in knockout animals. Diffusion weighted imaging showed altered anisotropy and putative gray matter changes in the hippocampal complex of the RNaseT2 knockout rats. Immunohistochemistry for glial fibrillary acidic protein (GFAP) showed the presence of hippocampal neuroinflammation. Decreased levels of lysosome-associated membrane protein 2 (LAMP2) and elevated acid phosphatase and β-N-Acetylglucosaminidase (NAG) activities indicated that the RNASET2 knockout rats likely had altered lysosomal function and potential defects in autophagy. Object recognition tests confirmed the RNaseT2 knockout rats exhibited memory deficits. However, the Barnes maze, and balance beam and rotarod tests, indicated there were no differences in spatial memory or motor impairments, respectively. Overall, patients with RNASET2 deficiency exhibited a more severe neurodegeneration phenotype than was observed in the RNaseT2 knockout rats. However, the vulnerability of the knockout rat hippocampus as evidenced by neuroinflammation, altered lysosomal function, and cognitive defects indicates this is still a useful in vivo model to study RNASET2 function. © 2018. Published by The Company of Biologists Ltd.

PF-KO system for single bunch mode operation of a storage ring

International Nuclear Information System (INIS)

Ohgaki, H.; Sugiyama, S.; Mikado, T.; Chiwaki, M.; Yamada, K.; Suzuki, R.; Sei, N.; Noguchi, T.; Yamazaki, T.

1994-01-01

A new RF-KO (RF knockout) system for the single bunch mode operation of a storage ring has been developed. The knockout signal is modulated by the sum signal of the RF acceleration frequency of the storage ring and a bunch selection signal. We do not need any special device or a timing unit with this method. We obtain a high purity of bunch structure in a short knock out time. The single bunch impurity of 0.2% has been achieved. (author)

One-neutron knockout from {sup 51-55}Sc

Energy Technology Data Exchange (ETDEWEB)

Schwertel, S.; Maierbeck, P.; Gernhaeuser, R.; Bildstein, V.; Boehmer, M.; Eppinger, K.; Faestermann, T.; Friese, J.; Fabbietti, L.; Maier, L.; Winkler, S. [Technische Universitaet Muenchen, Physik Department E12, Garching (Germany); Kruecken, R. [Technische Universitaet Muenchen, Physik Department E12, Garching (Germany); TRIUMF, Vancouver (Canada); University of British Columbia, Department of Physics and Astronomy, Vancouver (Canada); Kroell, T. [Technische Universitaet Muenchen, Physik Department E12, Garching (Germany); Technische Universitaet Darmstadt, Institut fuer Kernphysik, Darmstadt (Germany); Alvarez-Pol, H.; Benjamim, E.A.; Benlliure, J.; Caamano, M.; Cortina-Gil, D.; Gascon, M.; Kurtukian, T.; Perez, D.; Rodriguez-Tajes, C. [Universidade de Santiago de Compostela, Departamento de Fisica de Particulas, Santiago de Compostela (Spain); Aksouh, F.; Aumann, T.; Behr, K.; Boretzky, K.; Bruenle, A.; Chatillon, A.; Chulkov, L.V.; Geissel, H.; Gerl, J.; Gorska, M.; Kojouharov, I.; Klimkiewicz, A.; Kurz, N.; Nociforo, C.; Schaffner, H.; Simon, H.; Stanoiu, M.; Suemmerer, K.; Weick, H. [GSI Helmholtzzentrum fuer Schwerionenforschung, Darmstadt (Germany); Borge, M.J.G.; Pascual-Izarra, C.; Perea, A.; Tengblad, O. [CSIC, Instituto de Estructura de la Materia, Madrid (Spain); Buerger, A. [University of Oslo, SAFE/OCL, Oslo (Norway); CEA, Gif-sur-Yvette (France); Casarejos, E.; Brown, B.A. [University of Vigo, Vigo (Spain); Enders, J.; Schrieder, G. [Technische Universitaet Darmstadt, Institut fuer Kernphysik, Darmstadt (Germany); Hansen, P.G. [Michigan State University, NSCL, East Lansing, Michigan (United States); Jonson, B.; Nyman, G. [Chalmers Tekniska Hoegskola och Goeteborgs Universitet, Experimentell Fysik, Goeteborg (Sweden); Kanungo, R. [TRIUMF, Vancouver (Canada); GSI Helmholtzzentrum fuer Schwerionenforschung, Darmstadt (Germany); Saint Mary' s University, Halifax (Canada); Kiselev, O. [GSI Helmholtzzentrum fuer Schwerionenforschung, Darmstadt (Germany); Johannes Gutenberg Universitaet, Mainz (Germany); Paul Scherrer Institut, Villigen (Switzerland); Larsson, K. [GSI Helmholtzzentrum fuer Schwerionenforschung, Darmstadt (Germany); Chalmers Tekniska Hoegskola och Goeteborgs Universitet, Experimentell Fysik, Goeteborg (Sweden); Le Bleis, T. [GSI Helmholtzzentrum fuer Schwerionenforschung, Darmstadt (Germany); IN2P3-CNRS/Universite Louis Pasteur, Institut Pluridisciplinaire Hubert Curien, Strasbourg Cedex 2 (France); Mahata, K. [GSI Helmholtzzentrum fuer Schwerionenforschung, Darmstadt (Germany); Paul Scherrer Institut, Villigen (Switzerland); Nilsson, T. [Technische Universitaet Darmstadt, Institut fuer Kernphysik, Darmstadt (Germany); Chalmers Tekniska Hoegskola och Goeteborgs Universitet, Experimentell Fysik, Goeteborg (Sweden); Prochazka, A. [GSI Helmholtzzentrum fuer Schwerionenforschung, Darmstadt (Germany); Comenius University, Faculty of Mathematics and Physics, Bratislava (Slovakia); Rossi, D. [Johannes Gutenberg Universitaet, Mainz (Germany); Sitar, B. [Comenius University, Faculty of Mathematics and Physics, Bratislava (Slovakia); Otsuka, T. [University of Tokyo, Hongo, Bunkyo-ku, Department of Physics, Tokyo (Japan); Tostevin, J.A. [University of Surrey, Department of Physics, Faculty of Engineering and Physical Sciences, Guildford (United Kingdom); Rae, W.D.M. [Garsington, Oxfordshire (United Kingdom)

2012-12-15

Results are presented from a one-neutron knockout experiment at relativistic energies of {approx} 420 A MeV on {sup 51-55}Sc using the GSI Fragment Separator as a two-stage magnetic spectrometer and the MINIBALL array for gamma-ray detection. Inclusive longitudinal momentum distributions and cross-sections were measured enabling the determination of the contributions corresponding to knockout from the {nu}p{sub 1/2}, {nu}p{sub 3/2}, (L = 1) and {nu}f{sub 7/2}, {nu}f{sub 5/2} (L = 3) neutron orbitals. The observed L = 1 and L = 3 contributions are compared with theoretical cross-sections using eikonal knockout theory and spectroscopic factors from shell model calculations using the GXPF1A interaction. The measured inclusive knockout cross-sections generally follow the trends expected theoretically and given by the spectroscopic strength predicted from the shell model calculations. However, the deduced L = 1 cross-sections are generally 30-40% higher while the L = 3 contributions are about a factor of two smaller than predicted. This points to a promotion of neutrons from the {nu}f{sub 7/2} to the {nu}p{sub 3/2} orbital indicating a weakening of the N = 28 shell gap in these nuclei. While this is not predicted for the phenomenological GXPF1A interaction such a weakening is predicted by recent calculations using realistic low-momentum interactions V{sub low} {sub k} obtained by evolving a chiral N3LO nucleon-nucleon potential. (orig.)

Accurate quantification of microRNA via single strand displacement reaction on DNA origami motif.

Directory of Open Access Journals (Sweden)

Jie Zhu

Full Text Available DNA origami is an emerging technology that assembles hundreds of staple strands and one single-strand DNA into certain nanopattern. It has been widely used in various fields including detection of biological molecules such as DNA, RNA and proteins. MicroRNAs (miRNAs play important roles in post-transcriptional gene repression as well as many other biological processes such as cell growth and differentiation. Alterations of miRNAs' expression contribute to many human diseases. However, it is still a challenge to quantitatively detect miRNAs by origami technology. In this study, we developed a novel approach based on streptavidin and quantum dots binding complex (STV-QDs labeled single strand displacement reaction on DNA origami to quantitatively detect the concentration of miRNAs. We illustrated a linear relationship between the concentration of an exemplary miRNA as miRNA-133 and the STV-QDs hybridization efficiency; the results demonstrated that it is an accurate nano-scale miRNA quantifier motif. In addition, both symmetrical rectangular motif and asymmetrical China-map motif were tested. With significant linearity in both motifs, our experiments suggested that DNA Origami motif with arbitrary shape can be utilized in this method. Since this DNA origami-based method we developed owns the unique advantages of simple, time-and-material-saving, potentially multi-targets testing in one motif and relatively accurate for certain impurity samples as counted directly by atomic force microscopy rather than fluorescence signal detection, it may be widely used in quantification of miRNAs.

Accurate Quantification of microRNA via Single Strand Displacement Reaction on DNA Origami Motif

Science.gov (United States)

Lou, Jingyu; Li, Weidong; Li, Sheng; Zhu, Hongxin; Yang, Lun; Zhang, Aiping; He, Lin; Li, Can

2013-01-01

DNA origami is an emerging technology that assembles hundreds of staple strands and one single-strand DNA into certain nanopattern. It has been widely used in various fields including detection of biological molecules such as DNA, RNA and proteins. MicroRNAs (miRNAs) play important roles in post-transcriptional gene repression as well as many other biological processes such as cell growth and differentiation. Alterations of miRNAs' expression contribute to many human diseases. However, it is still a challenge to quantitatively detect miRNAs by origami technology. In this study, we developed a novel approach based on streptavidin and quantum dots binding complex (STV-QDs) labeled single strand displacement reaction on DNA origami to quantitatively detect the concentration of miRNAs. We illustrated a linear relationship between the concentration of an exemplary miRNA as miRNA-133 and the STV-QDs hybridization efficiency; the results demonstrated that it is an accurate nano-scale miRNA quantifier motif. In addition, both symmetrical rectangular motif and asymmetrical China-map motif were tested. With significant linearity in both motifs, our experiments suggested that DNA Origami motif with arbitrary shape can be utilized in this method. Since this DNA origami-based method we developed owns the unique advantages of simple, time-and-material-saving, potentially multi-targets testing in one motif and relatively accurate for certain impurity samples as counted directly by atomic force microscopy rather than fluorescence signal detection, it may be widely used in quantification of miRNAs. PMID:23990889

Accurate quantification of microRNA via single strand displacement reaction on DNA origami motif.

Science.gov (United States)

Zhu, Jie; Feng, Xiaolu; Lou, Jingyu; Li, Weidong; Li, Sheng; Zhu, Hongxin; Yang, Lun; Zhang, Aiping; He, Lin; Li, Can

2013-01-01

DNA origami is an emerging technology that assembles hundreds of staple strands and one single-strand DNA into certain nanopattern. It has been widely used in various fields including detection of biological molecules such as DNA, RNA and proteins. MicroRNAs (miRNAs) play important roles in post-transcriptional gene repression as well as many other biological processes such as cell growth and differentiation. Alterations of miRNAs' expression contribute to many human diseases. However, it is still a challenge to quantitatively detect miRNAs by origami technology. In this study, we developed a novel approach based on streptavidin and quantum dots binding complex (STV-QDs) labeled single strand displacement reaction on DNA origami to quantitatively detect the concentration of miRNAs. We illustrated a linear relationship between the concentration of an exemplary miRNA as miRNA-133 and the STV-QDs hybridization efficiency; the results demonstrated that it is an accurate nano-scale miRNA quantifier motif. In addition, both symmetrical rectangular motif and asymmetrical China-map motif were tested. With significant linearity in both motifs, our experiments suggested that DNA Origami motif with arbitrary shape can be utilized in this method. Since this DNA origami-based method we developed owns the unique advantages of simple, time-and-material-saving, potentially multi-targets testing in one motif and relatively accurate for certain impurity samples as counted directly by atomic force microscopy rather than fluorescence signal detection, it may be widely used in quantification of miRNAs.

Accurate Detection of Methicillin-Resistant Staphylococcus aureus in Mixtures by Use of Single-Bacterium Duplex Droplet Digital PCR.

Science.gov (United States)

Luo, Jun; Li, Junhua; Yang, Hang; Yu, Junping; Wei, Hongping

2017-10-01

Accurate and rapid identification of methicillin-resistant Staphylococcus aureus (MRSA) is needed to screen MRSA carriers and improve treatment. The current widely used duplex PCR methods are not able to differentiate MRSA from coexisting methicillin-susceptible S. aureus (MSSA) or other methicillin-resistant staphylococci. In this study, we aimed to develop a direct method for accurate and rapid detection of MRSA in clinical samples from open environments, such as nasal swabs. The new molecular assay is based on detecting the cooccurrence of nuc and mecA markers in a single bacterial cell by utilizing droplet digital PCR (ddPCR) with the chimeric lysin ClyH for cell lysis. The method consists of (i) dispersion of an intact single bacterium into nanoliter droplets, (ii) temperature-controlled release of genomic DNA (gDNA) by ClyH at 37°C, and (iii) amplification and detection of the markers ( nuc and mecA ) using standard TaqMan chemistries with ddPCR. Results were analyzed based on MRSA index ratios used for indicating the presence of the duplex-positive markers in droplets. The method was able to achieve an absolute limit of detection (LOD) of 2,900 CFU/ml for MRSA in nasal swabs spiked with excess amounts of Escherichia coli , MSSA, and other mecA -positive bacteria within 4 h. Initial testing of 104 nasal swabs showed that the method had 100% agreement with the standard culture method, while the normal duplex qPCR method had only about 87.5% agreement. The single-bacterium duplex ddPCR assay is rapid and powerful for more accurate detection of MRSA directly from clinical specimens. Copyright © 2017 American Society for Microbiology.

Nucleon knockout: off-shell effects

International Nuclear Information System (INIS)

Stephenson, G.J. Jr.

1977-01-01

The effect of the off-energy-shell extrapolation of the proton-proton scattering amplitude on the analysis of (p,2p) reactions is discussed. In particular, the range of expected variations in this extrapolation is explored and the possibility of using knock-out reactions to limit models of the p-p amplitude is studied

Easi-CRISPR for creating knock-in and conditional knockout mouse models using long ssDNA donors.

Science.gov (United States)

Miura, Hiromi; Quadros, Rolen M; Gurumurthy, Channabasavaiah B; Ohtsuka, Masato

2018-01-01

CRISPR/Cas9-based genome editing can easily generate knockout mouse models by disrupting the gene sequence, but its efficiency for creating models that require either insertion of exogenous DNA (knock-in) or replacement of genomic segments is very poor. The majority of mouse models used in research involve knock-in (reporters or recombinases) or gene replacement (e.g., conditional knockout alleles containing exons flanked by LoxP sites). A few methods for creating such models have been reported that use double-stranded DNA as donors, but their efficiency is typically 1-10% and therefore not suitable for routine use. We recently demonstrated that long single-stranded DNAs (ssDNAs) serve as very efficient donors, both for insertion and for gene replacement. We call this method efficient additions with ssDNA inserts-CRISPR (Easi-CRISPR) because it is a highly efficient technology (efficiency is typically 30-60% and reaches as high as 100% in some cases). The protocol takes ∼2 months to generate the founder mice.

Characterisation of iunH gene knockout strain from Mycobacterium tuberculosis

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Anne Drumond Villela

Full Text Available BACKGROUND Tuberculosis (TB is an infectious disease caused mainly by the bacillus Mycobacterium tuberculosis. The better understanding of important metabolic pathways from M. tuberculosis can contribute to the development of novel therapeutic and prophylactic strategies to combat TB. Nucleoside hydrolase (MtIAGU-NH, encoded by iunH gene (Rv3393, is an enzyme from purine salvage pathway in M. tuberculosis. MtIAGU-NH accepts inosine, adenosine, guanosine, and uridine as substrates, which may point to a pivotal metabolic role. OBJECTIVES Our aim was to construct a M. tuberculosis knockout strain for iunH gene, to evaluate in vitro growth and the effect of iunH deletion in M. tuberculosis in non-activated and activated macrophages models of infection. METHODS A M. tuberculosis knockout strain for iunH gene was obtained by allelic replacement, using pPR27xylE plasmid. The complemented strain was constructed by the transformation of the knockout strain with pNIP40::iunH. MtIAGU-NH expression was analysed by Western blot and LC-MS/MS. In vitro growth was evaluated in Sauton’s medium. Bacterial load of non-activated and interferon-γ activated RAW 264.7 cells infected with knockout strain was compared with wild-type and complemented strains. FINDINGS Western blot and LC-MS/MS validated iunH deletion at protein level. The iunH knockout led to a delay in M. tuberculosis growth kinetics in Sauton’s medium during log phase, but did not affect bases and nucleosides pool in vitro. No significant difference in bacterial load of knockout strain was observed when compared with both wild-type and complemented strains after infection of non-activated and interferon-γ activated RAW 264.7 cells. MAIN CONCLUSION The disruption of iunH gene does not influence M. tuberculosis growth in both non-activated and activated RAW 264.7 cells, which show that iunH gene is not important for macrophage invasion and virulence. Our results indicated that MtIAGU-NH is not a

Shape and Structure of N=Z Ge64: Electromagnetic Transition Rates from the Application of the Recoil Distance Method to a Knockout Reaction

Science.gov (United States)

Starosta, K.; Dewald, A.; Dunomes, A.; Adrich, P.; Amthor, A. M.; Baumann, T.; Bazin, D.; Bowen, M.; Brown, B. A.; Chester, A.; Gade, A.; Galaviz, D.; Glasmacher, T.; Ginter, T.; Hausmann, M.; Horoi, M.; Jolie, J.; Melon, B.; Miller, D.; Moeller, V.; Norris, R. P.; Pissulla, T.; Portillo, M.; Rother, W.; Shimbara, Y.; Stolz, A.; Vaman, C.; Voss, P.; Weisshaar, D.; Zelevinsky, V.

2007-07-01

Transition rate measurements are reported for the 21+ and 22+ states in N=Z Ge64. The experimental results are in excellent agreement with large-scale shell-model calculations applying the recently developed GXPF1A interactions. The measurement was done using the recoil distance method (RDM) and a unique combination of state-of-the-art instruments at the National Superconducting Cyclotron Laboratory (NSCL). States of interest were populated via an intermediate-energy single-neutron knockout reaction. RDM studies of knockout and fragmentation reaction products hold the promise of reaching far from stability and providing lifetime information for excited states in a wide range of nuclei.

Shape and structure of N=Z 64Ge: electromagnetic transition rates from the application of the recoil distance method to a knockout reaction.

Science.gov (United States)

Starosta, K; Dewald, A; Dunomes, A; Adrich, P; Amthor, A M; Baumann, T; Bazin, D; Bowen, M; Brown, B A; Chester, A; Gade, A; Galaviz, D; Glasmacher, T; Ginter, T; Hausmann, M; Horoi, M; Jolie, J; Melon, B; Miller, D; Moeller, V; Norris, R P; Pissulla, T; Portillo, M; Rother, W; Shimbara, Y; Stolz, A; Vaman, C; Voss, P; Weisshaar, D; Zelevinsky, V

2007-07-27

Transition rate measurements are reported for the 2(1)+ and 2(2)+ states in N=Z 64Ge. The experimental results are in excellent agreement with large-scale shell-model calculations applying the recently developed GXPF1A interactions. The measurement was done using the recoil distance method (RDM) and a unique combination of state-of-the-art instruments at the National Superconducting Cyclotron Laboratory (NSCL). States of interest were populated via an intermediate-energy single-neutron knockout reaction. RDM studies of knockout and fragmentation reaction products hold the promise of reaching far from stability and providing lifetime information for excited states in a wide range of nuclei.

Phytosterol Feeding Causes Toxicity in ABCG5/G8 Knockout Mice

Science.gov (United States)

McDaniel, Allison L.; Alger, Heather M.; Sawyer, Janet K.; Kelley, Kathryn L.; Kock, Nancy D.; Brown, J. Mark; Temel, Ryan E.; Rudel, Lawrence L.

2014-01-01

Plant sterols, or phytosterols, are very similar in structure to cholesterol and are abundant in typical diets. The reason for poor absorption of plant sterols by the body is still unknown. Mutations in the ABC transporters G5 and G8 are known to cause an accumulation of plant sterols in blood and tissues (sitosterolemia). To determine the significance of phytosterol exclusion from the body, we fed wild-type and ABCG5/G8 knockout mice a diet enriched with plant sterols. The high-phytosterol diet was extremely toxic to the ABCG5/G8 knockout mice but had no adverse effects on wild-type mice. ABCG5/G8 knockout mice died prematurely and developed a phenotype that included high levels of plant sterols in many tissues, liver abnormalities, and severe cardiac lesions. This study is the first to report such toxic effects of phytosterol accumulation in ABCG5/G8 knockout mice. We believe these new data support the conclusion that plant sterols are excluded from the body because they are toxic when present at high levels. PMID:23380580

Effect of cra gene knockout together with edd and iclR genes knockout on the metabolism in Escherichia coli.

Science.gov (United States)

Sarkar, Dayanidhi; Siddiquee, Khandaker Al Zaid; Araúzo-Bravo, Marcos J; Oba, Takahiro; Shimizu, Kazuyuki

2008-11-01

To elucidate the physiological adaptation of Escherichia coli due to cra gene knockout, a total of 3,911 gene expressions were investigated by DNA microarray for continuous culture. About 50 genes were differentially regulated for the cra mutant. TCA cycle and glyoxylate shunt were down-regulated, while pentose phosphate (PP) pathway and Entner Doudoroff (ED) pathway were up-regulated in the cra mutant. The glucose uptake rate and the acetate production rate were increased with less acetate consumption for the cra mutant. To identify the genes controlled by Cra protein, the Cra recognition weight matrix from foot-printing data was developed and used to scan the whole genome. Several new Cra-binding sites were found, and some of the result was consistent with the DNA microarray data. The ED pathway was active in the cra mutant; we constructed cra.edd double genes knockout mutant to block this pathway, where the acetate overflowed due to the down-regulation of aceA,B and icd gene expressions. Then we further constructed cra.edd.iclR triple genes knockout mutant to direct the carbon flow through the glyoxylate pathway. The cra.edd.iclR mutant showed the least acetate production, resulting in the highest cell yield together with the activation of the glycolysis pathway, but the glucose consumption rate could not be improved.

Accurate single-scattering simulation of ice cloud using the invariant-imbedding T-matrix method and the physical-geometric optics method

Science.gov (United States)

Sun, B.; Yang, P.; Kattawar, G. W.; Zhang, X.

2017-12-01

The ice cloud single-scattering properties can be accurately simulated using the invariant-imbedding T-matrix method (IITM) and the physical-geometric optics method (PGOM). The IITM has been parallelized using the Message Passing Interface (MPI) method to remove the memory limitation so that the IITM can be used to obtain the single-scattering properties of ice clouds for sizes in the geometric optics regime. Furthermore, the results associated with random orientations can be analytically achieved once the T-matrix is given. The PGOM is also parallelized in conjunction with random orientations. The single-scattering properties of a hexagonal prism with height 400 (in units of lambda/2*pi, where lambda is the incident wavelength) and an aspect ratio of 1 (defined as the height over two times of bottom side length) are given by using the parallelized IITM and compared to the counterparts using the parallelized PGOM. The two results are in close agreement. Furthermore, the integrated single-scattering properties, including the asymmetry factor, the extinction cross-section, and the scattering cross-section, are given in a completed size range. The present results show a smooth transition from the exact IITM solution to the approximate PGOM result. Because the calculation of the IITM method has reached the geometric regime, the IITM and the PGOM can be efficiently employed to accurately compute the single-scattering properties of ice cloud in a wide spectral range.

TRIC-B channels display labile gating: evidence from the TRIC-A knockout mouse model.

Science.gov (United States)

Venturi, Elisa; Matyjaszkiewicz, Antoni; Pitt, Samantha J; Tsaneva-Atanasova, Krasimira; Nishi, Miyuki; Yamazaki, Daiju; Takeshima, Hiroshi; Sitsapesan, Rebecca

2013-08-01

Sarcoplasmic/endoplasmic reticulum (SR) and nuclear membranes contain two related cation channels named TRIC-A and TRIC-B. In many tissues, both subtypes are co-expressed, making it impossible to distinguish the distinct single-channel properties of each subtype. We therefore incorporated skeletal muscle SR vesicles derived from Tric-a-knockout mice into bilayers in order to characterise the biophysical properties of native TRIC-B without possible misclassification of the channels as TRIC-A, and without potential distortion of functional properties by detergent purification protocols. The native TRIC-B channels were ideally selective for cations. In symmetrical 210 mM K(+), the maximum (full) open channel level (199 pS) was equivalent to that observed when wild-type SR vesicles were incorporated into bilayers. Analysis of TRIC-B gating revealed complex and variable behaviour. Four main sub-conductance levels were observed at approximately 80 % (161 pS), 60 % (123 pS), 46 % (93 pS), and 30 % (60 pS) of the full open state. Seventy-five percent of the channels were voltage sensitive with Po being markedly reduced at negative holding potentials. The frequent, rapid transitions between TRIC-B sub-conductance states prevented development of reliable gating models using conventional single-channel analysis. Instead, we used mean-variance plots to highlight key features of TRIC-B gating in a more accurate and visually useful manner. Our study provides the first biophysical characterisation of native TRIC-B channels and indicates that this channel would be suited to provide counter current in response to Ca(2+) release from the SR. Further experiments are required to distinguish the distinct functional properties of TRIC-A and TRIC-B and understand their individual but complementary physiological roles.

Gene Knockout Identification Using an Extension of Bees Hill Flux Balance Analysis

Directory of Open Access Journals (Sweden)

Yee Wen Choon

2015-01-01

Full Text Available Microbial strain optimisation for the overproduction of a desired phenotype has been a popular topic in recent years. Gene knockout is a genetic engineering technique that can modify the metabolism of microbial cells to obtain desirable phenotypes. Optimisation algorithms have been developed to identify the effects of gene knockout. However, the complexities of metabolic networks have made the process of identifying the effects of genetic modification on desirable phenotypes challenging. Furthermore, a vast number of reactions in cellular metabolism often lead to a combinatorial problem in obtaining optimal gene knockout. The computational time increases exponentially as the size of the problem increases. This work reports an extension of Bees Hill Flux Balance Analysis (BHFBA to identify optimal gene knockouts to maximise the production yield of desired phenotypes while sustaining the growth rate. This proposed method functions by integrating OptKnock into BHFBA for validating the results automatically. The results show that the extension of BHFBA is suitable, reliable, and applicable in predicting gene knockout. Through several experiments conducted on Escherichia coli, Bacillus subtilis, and Clostridium thermocellum as model organisms, extension of BHFBA has shown better performance in terms of computational time, stability, growth rate, and production yield of desired phenotypes.

Knockouts of high-ranking males have limited impact on baboon social networks.

Science.gov (United States)

Franz, Mathias; Altmann, Jeanne; Alberts, Susan C

Social network structures can crucially impact complex social processes such as collective behaviour or the transmission of information and diseases. However, currently it is poorly understood how social networks change over time. Previous studies on primates suggest that `knockouts' (due to death or dispersal) of high-ranking individuals might be important drivers for structural changes in animal social networks. Here we test this hypothesis using long-term data on a natural population of baboons, examining the effects of 29 natural knockouts of alpha or beta males on adult female social networks. We investigated whether and how knockouts affected (1) changes in grooming and association rates among adult females, and (2) changes in mean degree and global clustering coefficient in these networks. The only significant effect that we found was a decrease in mean degree in grooming networks in the first month after knockouts, but this decrease was rather small, and grooming networks rebounded to baseline levels by the second month after knockouts. Taken together our results indicate that the removal of high-ranking males has only limited or no lasting effects on social networks of adult female baboons. This finding calls into question the hypothesis that the removal of high-ranking individuals has a destabilizing effect on social network structures in social animals.

Parallel knock-out schemes in networks

NARCIS (Netherlands)

Broersma, H.J.; Fomin, F.V.; Woeginger, G.J.

2004-01-01

We consider parallel knock-out schemes, a procedure on graphs introduced by Lampert and Slater in 1997 in which each vertex eliminates exactly one of its neighbors in each round. We are considering cases in which after a finite number of rounds, where the minimimum number is called the parallel

Formation of a single-bunch beam in the booster synchrotron at SPring-8

CERN Document Server

Suzuki, H; Ego, H; Hara, M; Hosoda, N; Kawashima, Y; Ohashi, Y; Ohshima, T; Tani, N; Yabashi, M; Yonehara, H

2000-01-01

In order to fill a radio frequency (rf) bucket with an electron beam in the storage ring at SPring-8, an rf knockout system was installed in the booster synchrotron. With this system, the energy of the electron beam injected from the linac was increased from 1 to 8 GeV. The time width of multi-bunch beams from the linac operated at 2856 MHz rf can be selected as 1 or 40 ns. The beam injected from the linac is distributed in rf buckets of the booster synchrotron operated at 508.58 MHz rf. To fill a single rf bucket with a beam, the rf knockout system is operated at a minimum beam energy of 1 GeV. By using the rf knockout system, the electron beam is effectively kept in a single rf bucket. Then the beam is injected into a targeted rf bucket in the storage ring with a precise timing system. The beam intensity of satellite rf buckets in the storage ring was measured with a photon counting method and determined to be 10 sup - sup 6 less than that of the main rf bucket. In this paper, we describe the rf knockout sy...

Bile Salt Homeostasis in Normal and Bsep Gene Knockout Rats with Single and Repeated Doses of Troglitazone.

Science.gov (United States)

Cheng, Yaofeng; Chen, Shenjue; Freeden, Chris; Chen, Weiqi; Zhang, Yueping; Abraham, Pamela; Nelson, David M; Humphreys, W Griffith; Gan, Jinping; Lai, Yurong

2017-09-01

The interference of bile acid secretion through bile salt export pump (BSEP) inhibition is one of the mechanisms for troglitazone (TGZ)-induced hepatotoxicity. Here, we investigated the impact of single or repeated oral doses of TGZ (200 mg/kg/day, 7 days) on bile acid homoeostasis in wild-type (WT) and Bsep knockout (KO) rats. Following oral doses, plasma exposures of TGZ were not different between WT and KO rats, and were similar on day 1 and day 7. However, plasma exposures of the major metabolite, troglitazone sulfate (TS), in KO rats were 7.6- and 9.3-fold lower than in WT on day 1 and day 7, respectively, due to increased TS biliary excretion. With Bsep KO, the mRNA levels of multidrug resistance-associated protein 2 (Mrp2), Mrp3, Mrp4, Mdr1, breast cancer resistance protein (Bcrp), sodium taurocholate cotransporting polypeptide, small heterodimer partner, and Sult2A1 were significantly altered in KO rats. Following seven daily TGZ treatments, Cyp7A1 was significantly increased in both WT and KO rats. In the vehicle groups, plasma exposures of individual bile acids demonstrated variable changes in KO rats as compared with WT. WT rats dosed with TGZ showed an increase of many bile acid species in plasma on day 1, suggesting the inhibition of Bsep. Conversely, these changes returned to base levels on day 7. In KO rats, alterations of most bile acids were observed after seven doses of TGZ. Collectively, bile acid homeostasis in rats was regulated through bile acid synthesis and transport in response to Bsep deficiency and TGZ inhibition. Additionally, our study is the first to demonstrate that repeated TGZ doses can upregulate Cyp7A1 in rats. Copyright © 2017 by The American Society for Pharmacology and Experimental Therapeutics.

Effects of HAb18G/CD147 knockout on hepatocellular carcinoma cells in vitro using a novel zinc-finger nuclease-targeted gene knockout approach.

Science.gov (United States)

Li, Hong-Wei; Yang, Xiang-Min; Tang, Juan; Wang, Shi-Jie; Chen, Zhi-Nan; Jiang, Jian-Li

2015-03-01

HAb18G/CD147 belongs to the immunoglobulin superfamily and predominantly functions as an inducer of matrix metalloproteinase secretion for tumor invasion and metastasis. This study was designed to investigate the effects of HAb18G/CD147 knockout on hepatocellular carcinoma cells using zinc-finger nuclease (ZFNs)-targeted gene knockout approach. The HCC cell line SMMC-7721 was used for ZFNs-targeted cleavage of the HAb18G/CD147 gene. RT-PCR and Western blot assays were used to detect HAb18G/CD147 expression. HAb18G phenotypic changes following HAb18G/CD147 knockout in SMMC-K7721 cells were assessed using tumor cell adhesion, invasion, migration and colony formation and flow cytometric assays. These data demonstrated that tumor cell adhesion, invasion, migration, and colony formation capabilities of SMMC-K7721 were significantly reduced compared to parental cells or SMMC-7721 with re-expression of HAb18G/CD147 protein transfected with HAb18G/CD147 cDNA. Moreover, knockout of HAb18G/CD147 expression also induced SMMC-K7721 cells to undergo apoptosis compared to SMMC-7721 and SMMC-R7721 (P CD147 reduced p53 levels in SMMC-R7721 cells, possibly through inhibition of the PI3K-Akt-MDM2 signaling pathway. The findings provide a novel insight into the mechanisms underlying HAb18G/CD147-induced progression of HCC cells.

Site-Directed Genome Knockout in Chicken Cell Line and Embryos Can Use CRISPR/Cas Gene Editing Technology

Directory of Open Access Journals (Sweden)

Qisheng Zuo

2016-06-01

Full Text Available The present study established an efficient genome editing approach for the construction of stable transgenic cell lines of the domestic chicken (Gallus gallus domesticus. Our objectives were to facilitate the breeding of high-yield, high-quality chicken strains, and to investigate gene function in chicken stem cells. Three guide RNA (gRNAs were designed to knockout the C2EIP gene, and knockout efficiency was evaluated in DF-1 chicken fibroblasts and chicken ESCs using the luciferase single-strand annealing (SSA recombination assay, T7 endonuclease I (T7EI assay, and TA clone sequencing. In addition, the polyethylenimine-encapsulated Cas9/gRNA plasmid was injected into fresh fertilized eggs. At 4.5 d later, frozen sections of the embryos were prepared, and knockout efficiency was evaluated by the T7EI assay. SSA assay results showed that luciferase activity of the vector expressing gRNA-3 was double that of the control. Results of the T7EI assay and TA clone sequencing indicated that Cas9/gRNA vector-mediated gene knockdown efficiency was approximately 27% in both DF-1 cells and ESCs. The CRISPR/Cas9 vector was also expressed in chicken embryos, resulting in gene knockdown in three of the 20 embryos (gene knockdown efficiency 15%. Taken together, our results indicate that the CRISPR/Cas9 system can mediate stable gene knockdown at the cell and embryo levels in domestic chickens.

Empirical Accurate Masses and Radii of Single Stars with TESS and Gaia

Science.gov (United States)

Stassun, Keivan G.; Corsaro, Enrico; Pepper, Joshua A.; Gaudi, B. Scott

2018-01-01

We present a methodology for the determination of empirical masses of single stars through the combination of three direct observables with Gaia and Transiting Exoplanet Survey Satellite (TESS): (i) the surface gravity via granulation-driven variations in the TESS light curve, (ii) the bolometric flux at Earth via the broadband spectral energy distribution, and (iii) the distance via the Gaia parallax. We demonstrate the method using 525 Kepler stars for which these measures are available in the literature, and show that the stellar masses can be measured with this method to a precision of ∼25%, limited by the surface-gravity precision of the granulation “flicker” method (∼0.1 dex) and by the parallax uncertainties (∼10% for the Kepler sample). We explore the impact of expected improvements in the surface gravity determinations—through the application of granulation background fitting and the use of recently published granulation-metallicity relations—and improvements in the parallaxes with the arrival of the Gaia second data release. We show that the application of this methodology to stars that will be observed by TESS should yield radii good to a few percent and masses good to ≈10%. Importantly, the method does not require the presence of an orbiting, eclipsing, or transiting body, nor does it require spatial resolution of the stellar surface. Thus, we can anticipate the determination of fundamental, accurate stellar radii and masses for hundreds of thousands of bright single stars—across the entire sky and spanning the Hertzsprung–Russell diagram—including those that will ultimately be found to host planets.

Fully integrated free-running InGaAs/InP single-photon detector for accurate lidar applications.

Science.gov (United States)

Yu, Chao; Shangguan, Mingjia; Xia, Haiyun; Zhang, Jun; Dou, Xiankang; Pan, Jian-Wei

2017-06-26

We present a fully integrated InGaAs/InP negative feedback avalanche diode (NFAD) based free-running single-photon detector (SPD) designed for accurate lidar applications. A free-piston Stirling cooler is used to cool down the NFAD with a large temperature range, and an active hold-off circuit implemented in a field programmable gate array is applied to further suppress the afterpulsing contribution. The key parameters of the free-running SPD including photon detection efficiency (PDE), dark count rate (DCR), afterpulse probability, and maximum count rate (MCR) are dedicatedly optimized for lidar application in practice. We then perform a field experiment using a Mie lidar system with 20 kHz pulse repetition frequency to compare the performance between the free-running InGaAs/InP SPD and a commercial superconducting nanowire single-photon detector (SNSPD). Our detector exhibits good performance with 1.6 Mcps MCR (0.6 μs hold-off time), 10% PDE, 950 cps DCR, and 18% afterpulse probability over 50 μs period. Such performance is worse than the SNSPD with 60% PDE and 300 cps DCR. However, after performing a specific algorithm that we have developed for afterpulse and count rate corrections, the lidar system performance in terms of range-corrected signal (Pr 2 ) distribution using our SPD agrees very well with the result using the SNSPD, with only a relative error of ∼2%. Due to the advantages of low-cost and small size of InGaAs/InP NFADs, such detector provides a practical solution for accurate lidar applications.

An accurate behavioral model for single-photon avalanche diode statistical performance simulation

Science.gov (United States)

Xu, Yue; Zhao, Tingchen; Li, Ding

2018-01-01

An accurate behavioral model is presented to simulate important statistical performance of single-photon avalanche diodes (SPADs), such as dark count and after-pulsing noise. The derived simulation model takes into account all important generation mechanisms of the two kinds of noise. For the first time, thermal agitation, trap-assisted tunneling and band-to-band tunneling mechanisms are simultaneously incorporated in the simulation model to evaluate dark count behavior of SPADs fabricated in deep sub-micron CMOS technology. Meanwhile, a complete carrier trapping and de-trapping process is considered in afterpulsing model and a simple analytical expression is derived to estimate after-pulsing probability. In particular, the key model parameters of avalanche triggering probability and electric field dependence of excess bias voltage are extracted from Geiger-mode TCAD simulation and this behavioral simulation model doesn't include any empirical parameters. The developed SPAD model is implemented in Verilog-A behavioral hardware description language and successfully operated on commercial Cadence Spectre simulator, showing good universality and compatibility. The model simulation results are in a good accordance with the test data, validating high simulation accuracy.

ANTXR2 Knock-Out Does Not Result in the Development of Hypertension in Rats.

Science.gov (United States)

Liu, Xiaoyan; Yuan, Wen; Li, Jing; Yang, Lei; Cai, Jun

2017-02-01

Our recent genetic study as well as robust evidences reported by previous genome-wide association studies (GWASs) have indicated that the single nucleotide polymorphism rs16998073, located near gene anthrax toxin receptor 2 (ANTXR2), was significantly associated with hypertension in Asians and Europeans. The aim of the present study was to determine whether ANTXR2 is the causal gene of hypertension at the 4q21 locus using an ANTXR2 knock-out model. Relative expression of ANTXR2 in Wistar-Kyoto rats (WKYs) and spontaneously hypertensive rats (SHRs) were determined by real-time quantitative polymerase chain reaction and western blot analysis. ANTXR2 knock-out rats were created using CRISPR/Cas9-mediated genome editing and blood pressure values were measured in ANTXR2 -/- and wild type (WT) rats by tail-cuff method and carotid arterial catheterization method. Neither the mRNA nor protein levels of ANTXR2 were significantly different between tissues from SHRs and WKYs. To create ANTXR2 -/- rats, 67 base pairs were deleted in exon 1 of ANTXR2 using CRISPR/Cas9-mediated genome editing. ANTXR2 protein decreased significantly in aortas of ANTXR2 -/- rats, suggesting sufficient efficiency of ANTXR2 knock-out in this model. However, ANTXR2 -/- rats exhibited nearly the same blood pressure as WT rats at baseline conditions as well as during Angiotensin II (400ng/kg/min) infusion or high-salt diet treatment. These findings suggest that ANTXR2 might not be associated with hypertension and thus further functional analysis is warranted to identify the causal gene at this locus. © American Journal of Hypertension, Ltd 2016. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Hyperactivity of newborn Pten knock-out neurons results from increased excitatory synaptic drive.

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Williams, Michael R; DeSpenza, Tyrone; Li, Meijie; Gulledge, Allan T; Luikart, Bryan W

2015-01-21

Developing neurons must regulate morphology, intrinsic excitability, and synaptogenesis to form neural circuits. When these processes go awry, disorders, including autism spectrum disorder (ASD) or epilepsy, may result. The phosphatase Pten is mutated in some patients having ASD and seizures, suggesting that its mutation disrupts neurological function in part through increasing neuronal activity. Supporting this idea, neuronal knock-out of Pten in mice can cause macrocephaly, behavioral changes similar to ASD, and seizures. However, the mechanisms through which excitability is enhanced following Pten depletion are unclear. Previous studies have separately shown that Pten-depleted neurons can drive seizures, receive elevated excitatory synaptic input, and have abnormal dendrites. We therefore tested the hypothesis that developing Pten-depleted neurons are hyperactive due to increased excitatory synaptogenesis using electrophysiology, calcium imaging, morphological analyses, and modeling. This was accomplished by coinjecting retroviruses to either "birthdate" or birthdate and knock-out Pten in granule neurons of the murine neonatal dentate gyrus. We found that Pten knock-out neurons, despite a rapid onset of hypertrophy, were more active in vivo. Pten knock-out neurons fired at more hyperpolarized membrane potentials, displayed greater peak spike rates, and were more sensitive to depolarizing synaptic input. The increased sensitivity of Pten knock-out neurons was due, in part, to a higher density of synapses located more proximal to the soma. We determined that increased synaptic drive was sufficient to drive hypertrophic Pten knock-out neurons beyond their altered action potential threshold. Thus, our work contributes a developmental mechanism for the increased activity of Pten-depleted neurons. Copyright © 2015 the authors 0270-6474/15/350943-17$15.00/0.

Fast beam cut-off method in RF-knockout extraction for spot-scanning

CERN Document Server

Furukawa, T

2002-01-01

An irradiation method with magnetic scanning has been developed in order to provide accurate irradiation even for an irregular target shape. The scanning method has strongly required a lower ripple of the beam spill and a faster response to beam-on/off in slow extraction from a synchrotron ring. At HIMAC, RF-knockout extraction has utilized a bunched beam to reduce the beam-spill ripple. Therefore, particles near the resonance can be spilled out from the separatrices by synchrotron oscillation as well as by a transverse RF field. From this point of view, a fast beam cut-off method has been proposed and verified by both simulations and experiments. The maximum delay from the beam cut-off signal to beam-off has been improved to around 60 mu s from 700 mu s by a usual method. Unwanted dose has been considerably reduced by around a factor of 10 compared with that by the usual method.

Highly segmented CVD diamond detectors and high-resolution momentum measurements in knockout reactions; Hochsegmentierte CVD Diamant Detektoren und hochaufloesende Impulsmessungen in Knockout Reaktionen

Energy Technology Data Exchange (ETDEWEB)

Schwertel, Sabine

2009-11-26

In recent years knockout reactions have proven to be important tools for investigations of the structure of light exotic nuclei. In spring 2006 an experiment was performed with the fragment separator at GSI in Darmstadt to extend this method to medium-mass nuclei with energies of about 400 AMeV. An experiment with a stable and well-known {sup 48}Ca primary beam was performed as a reference. The FRS was set for the reaction {sup 56}Ti{yields}{sup 55}Ti. Because of the high acceptance of the FRS, mother and daughter nuclei of one-neutron knockout reactions in the Sc isotopes {sup 51,52,53,54,55}Sc were also transported with high efficiency. These are investigated in the first part of this thesis. Inclusive cross sections of 77(10) mbarn for one-neutron knockout from {sup 48}Ca and 78(12) mbarn, 99(15) mbarn, 101(15) mbarn, 113(17) mbarn and 72(14) mbarn for knockout from {sup 51,52,53,54,55}Sc, respectively, were measured for the first time. For the Sc isotopes the reduction factors are close to 1. For the one-neutron knockout reactions in {sup 48}Ca and the Sc isotopes, respectively, the momentum distributions could be measured with a relative resolution of 0.17-0.27 %. From the momentum distributions spectroscopic factors of the involved orbitals could be extracted. In the future, further knockout experiments should be performed using the R{sup 3}B setup at FAIR. The available beam intensity will be up to four orders of magnitude higher. As the beam has to be tracked from the dispersive plane of the Super-FRS up to the R{sup 3}B target, radiation hard detectors are needed. In the framework of this thesis extensive measurements were performed at the tandem accelerator in Munich with numerous small (10 x 10 mm{sup 2}) test detectors. Samples using new manufacturing methods were characterized. A dose of some 10{sup 11} ions/mm{sup 2} was determined as a limit for the exposure of the material with heavy ions of high ionisation density. It could be shown that even

Test of distorted wave kinematic coupling approximation calculations for knockout reactions

International Nuclear Information System (INIS)

Jain, A.K.

1990-01-01

A test has been devised to check the validity of conventional distorted-wave impulse approximation (DWIA) treatment of knockout reactions. The conventional DWIA formalism separates the three-body final state Schroedinger equation for a knockout reaction into two two-body Schroedinger equations by assuming an asymptotic constant value for the three-body coupling term commonly known as the kinematic coupling approximation (KCA). In the test case, which consists of an extreme asymmetric situation where one of the distorting optical potentials is assumed to vanish, the three-body final state Schroedinger equation can be solved exactly as a product of two two-body solutions using one particular set of relative coordinates. Large influence of the three-body coupling term is seen in the comparison of the exact and KCA results for (α,2α) and (p,pα) knockout reactions when the distorting optical potentials are weakly absorbing

Hepatic changes in metabolic gene expression in old ghrelin and ghrelin receptor knockout mice

Science.gov (United States)

Ghrelin knockout (GKO) and ghrelin receptor (growth hormone secretagogue receptor) knockout (GHSRKO) mice exhibit enhanced insulin sensitivity, but the mechanism is unclear. Insulin sensitivity declines with age and is inversely associated with accumulation of lipid in liver, a key glucoregulatory ...

Deconstructing mammalian reproduction: using knockouts to define fertility pathways.

Science.gov (United States)

Roy, Angshumoy; Matzuk, Martin M

2006-02-01

Reproduction is the sine qua non for the propagation of species and continuation of life. It is a complex biological process that is regulated by multiple factors during the reproductive life of an organism. Over the past decade, the molecular mechanisms regulating reproduction in mammals have been rapidly unraveled by the study of a vast number of mouse gene knockouts with impaired fertility. The use of reverse genetics to generate null mutants in mice through targeted disruption of specific genes has enabled researchers to identify essential regulators of spermatogenesis and oogenesis in vivo and model human disorders affecting reproduction. This review focuses on the merits, utility, and the variations of the knockout technology in studies of reproduction in mammals.

Efficient generation of P53 biallelic knockout Diannan miniature pigs via TALENs and somatic cell nuclear transfer

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Youfeng Shen

2017-11-01

Full Text Available Abstract Background Pigs have many features that make them attractive as biomedical models for various diseases, including cancer. P53 is an important tumor suppressor gene that exerts a central role in protecting cells from oncogenic transformation and is mutated in a large number of human cancers. P53 mutations occur in almost every type of tumor and in over 50% of all tumors. In a recent publication, pigs with a mutated P53 gene were generated that resulted in lymphoma and renal and osteogenic tumors. However, approximately 80% of human tumors have dysfunctional P53. A P53-deficient pig model is still required to elucidate. Methods Transcription activator-like effector nucleases (TALENs were designed to target porcine P53 exon 4. The targeting activity was evaluated using a luciferase SSA recombination assay. P53 biallelic knockout (KO cell lines were established from single-cell colonies of fetal fibroblasts derived from Diannan miniature pigs followed by electroporation with TALENs plasmids. One cell line was selected as the donor cell line for somatic cell nuclear transfer (SCNT for the generation of P53 KO pigs. P53 KO stillborn fetuses and living piglets were obtained. Gene typing of the collected cloned individuals was performed by T7EI assay and sequencing. Fibroblast cells from Diannan miniature piglets with a P53 biallelic knockout or wild type were analyzed for the P53 response to doxorubicin treatment by confocal microscopy and western blotting. Results The luciferase SSA recombination assay revealed that the targeting activities of the designed TALENs were 55.35-fold higher than those of the control. Eight cell lines (8/19 were mutated for P53, and five of them were biallelic knockouts. One of the biallelic knockout cell lines was selected as nuclear donor cells for SCNT. The cloned embryos were transferred into five recipient gilts, three of them becoming pregnant. Five live fetuses were obtained from one surrogate by caesarean

Knockout of endothelial cell-derived endothelin-1 attenuates skin fibrosis but accelerates cutaneous wound healing.

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Katsunari Makino

Full Text Available Endothelin (ET-1 is known for the most potent vasoconstrictive peptide that is released mainly from endothelial cells. Several studies have reported ET-1 signaling is involved in the process of wound healing or fibrosis as well as vasodilation. However, little is known about the role of ET-1 in these processes. To clarify its mechanism, we compared skin fibrogenesis and wound repair between vascular endothelial cell-specific ET-1 knockout mice and their wild-type littermates. Bleomycin-injected fibrotic skin of the knockout mice showed significantly decreased skin thickness and collagen content compared to that of wild-type mice, indicating that bleomycin-induced skin fibrosis is attenuated in the knockout mice. The mRNA levels of transforming growth factor (TGF-β were decreased in the bleomycin-treated skin of ET-1 knockout mice. On the other hand, skin wound healing was accelerated in ET-1 knockout mice, which was indicated by earlier granulation tissue reduction and re-epithelialization in these mice. The mRNA levels of TGF-β, tumor necrosis factor (TNF-α and connective tissue growth factor (CTGF were reduced in the wound of ET-1 knockout mice. In endothelial ET-1 knockout mouse, the expression of TNF-α, CTGF and TGF-β was down-regulated. Bosentan, an antagonist of dual ET receptors, is known to attenuate skin fibrosis and accelerate wound healing in systemic sclerosis, and such contradictory effect may be mediated by above molecules. The endothelial cell-derived ET-1 is the potent therapeutic target in fibrosis or wound healing, and investigations of the overall regulatory mechanisms of these pathological conditions by ET-1 may lead to a new therapeutic approach.

Akt2/LDLr double knockout mice display impaired glucose tolerance and develop more complex atherosclerotic plaques than LDLr knockout mice

NARCIS (Netherlands)

Rensing, Katrijn L.; de Jager, Saskia C. A.; Stroes, Erik S.; Vos, Mariska; Twickler, Marcel Th B.; Dallinga-Thie, Geesje M.; de Vries, Carlie J. M.; Kuiper, Johan; Bot, Ilze; von der Thüsen, Jan H.

2014-01-01

To characterize the phenotype of Akt2/low-density-lipoprotein receptor double knockout (dKO) (Akt2/LDLr dKO) mice with respect to insulin resistance and features of atherosclerotic plaque progression. Metabolic profile and atherosclerotic plaque progression were compared between LDLr KO mice and

FMR1 Knockout mice: A model to study fragile X mental retardation

Energy Technology Data Exchange (ETDEWEB)

Oostra, B.A.; Bakker, C.E.; Reyniers, E. [Erasmus Univ., Rotterdam (Netherlands)] [and others

1994-09-01

The fragile X syndrome is the most frequent form of inherited mental retardation in humans with an incidence of 1 in 1250 males and 1 in 2500 females. The clinical syndrome includes moderate to severe mental retardation, autistic behavior, macroorchidism, and facial features, such as long face with mandibular prognathism and large, everted ears. The molecular basis for this disease is a large expansion of a triplet repeat (CGG){sub n} in the 5{prime} untranslated region of the FMR1 gene. Due to this large expansion of the CGG repeat, the promoter region becomes methylated and the FMR1 gene is subsequently silenced. Hardly anything is known about the physiologic function of FMR1 and the pathologic mechanisms leading to these symptoms. Since the FMR1 gene is highly conserved in the mouse, we used the mouse to design a knockout model for the fragile X syndrome. These knockout mice lacking Fmrp have normal litter size suggesting that FMR1 is not essential in human gametogenesis and embryonic development. The knockout mice show the abnormalities also seen in the affected organs of human patients. Mutant mice show a gradual development through time of macroorchidism. In the knockout mice we observed cognitive defects in the form of deficits in learning (as shown by the hidden platform Morris water maze task) and behavioral abnormalities such as increased exploratory behavior and hyperactivity. Therefore this knockout mouse may serve as a valuable tool in studying the role of FMR1 in the fragile X syndrome and may serve as a model to elucidate the mechanisms involved in macroorchidism, abnormal behavior, and mental retardation.

Shape and structure of N=Z ^64Ge; Electromagnetic transition rates from the application of the Recoil Distance Method to knock-out reactions.

Science.gov (United States)

Starosta, K.; Dewald, A.

2007-04-01

Transition rate measurements are reported for the 2^+1 and 2^+2 states in the N=Z nucleus ^64Ge. The measurement was done utilizing the Recoil Distance Method (RDM) and a unique combination of state of the art instruments at the National Superconducting Cyclotron Laboratory (NSCL). States of interest were populated via an intermediate energy single neutron knock-out reaction. RDM studies of knock-out and fragmentation reaction products hold the promise of reaching far from stability and providing lifetime information for intermediate-spin excited states in a wide range of exotic nuclei. The large-scale Shell Model calculations applying the recently developed GXPF1A interaction are in excellent agreement with the above results. Theoretical analysis suggests that ^64Ge is a collective γ-soft anharmonic vibrator.

Pauli blocking and medium effects in nucleon knockout reactions

International Nuclear Information System (INIS)

Bertulani, C. A.; De Conti, C.

2010-01-01

We study medium modifications of the nucleon-nucleon (NN) cross sections and their influence on the nucleon knockout reactions. Using the eikonal approximation, we compare the results obtained with free NN cross sections with those obtained with a purely geometrical treatment of Pauli blocking and with NN obtained with more elaborated Dirac-Bruecker methods. The medium effects are parametrized in terms of the baryon density. We focus on symmetric nuclear matter, although the geometrical Pauli blocking also allows for the treatment of asymmetric nuclear matter. It is shown that medium effects can change the nucleon knockout cross sections and momentum distributions up to 10% in the energy range E lab =50-300 MeV/nucleon. The effect is more evident in reactions involving halo nuclei.

Post-irradiation studies on knock-out and pseudo-recoil releases of fission products from fissioning UO2

International Nuclear Information System (INIS)

Yamagishi, S.; Tanifuji, T.

1976-01-01

By using post-irradiation techniques, in-pile releases of 133 Xe, sup(85m)Kr, 88 Kr, 87 Kr and 138 Xe from UO 2 fissioning at low temperatures below about 200 0 C are studied: these are analyzed into a time-dependent knock-out and time-independent pseudo-recoil releases. For the latter, a 'self knock-out' mechanism is proposed: when a fission fragment loses thoroughly its energy near the UO 2 surface and stops there, it will knock out the surface substances and accordingly the fragment (i.e. the fission product) will be released. The effective thickness of the layer where the self knock-out occurs is found to be approximately 7A. As for the knock-out release, the following is estimated from its dependence on various factors: the knock-out release of fission products occurs from the surface layer with the effective thickness of approximately 20A: the shape of UO 2 matrix knocked out by one fission fragment passing through the surface is equivalent to a cylinder approximately 32A diameter by approximately 27A thick, (i.e. the knock-out coefficient for UO 2 is approximately 660 uranium atoms per knock-out event). On the basis of the above estimations, the conclusions derived from the past in-pile studies of fission gas releases are evaluated. (Auth.)

Study of 19C by One-Neutron Knockout

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Hwang Jongwon

2016-01-01

Full Text Available The spectroscopic structure of 19C, a prominent one-neutron halo nucleus, has been studied with a 20C secondary beam at 290 MeV/nucleon and a carbon target. Neutron-unbound states populated by the one-neutron knockout reaction were investigated by means of the invariant mass method. The preliminary relative energy spectrum and parallel momentum distribution of the knockout residue, 19C*, were reconstructed from the measured four momenta of the 18C fragment, neutron, and beam. Three resonances were observed in the spectrum, which correspond to the states at Ex = 0.62(9, 1.42(10, and 2.89(10 MeV. The parallel momentum distributions for the 0.62-MeV and 2.89-MeV states suggest spin-parity assignments of 5/2+ and 1/2−, respectively. The 1.42-MeV state is in line with the reported 5/22+ state.

High-temperature expansion and knock-out properties of moulding sands with water glass

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Major-Gabryś K.

2007-01-01

Full Text Available The article focuses on the topic of improving the knock-out properties of moulding sand with water glass and ester hardener. It is settled that the cause of worse knock-out properties of moulding sand can be brought by their thermal expansion in increased temperatures. There is a presentation of the influence of different additives, containing Al2O3, on moulding sands’ expansion in increased temperatures. Within the frames of research, there was an elaboration of the influence of authors own additive- Glassex, on the expansion phenomenon of moulding sands with water glass and ester hardener. It is concluded, that the new additive stops the expansion of moulding sands and as well it improves their knock-out properties.

Describing the role of Drosophila melanogaster ABC transporters in insecticide biology using CRISPR-Cas9 knockouts.

Science.gov (United States)

Denecke, Shane; Fusetto, Roberto; Batterham, Philip

2017-12-01

ABC transporters have a well-established role in drug resistance, effluxing xenobiotics from cells and tissues within the organism. More recently, research has been dedicated to understanding the role insect ABC transporters play in insecticide toxicity, but progress in understanding the contribution of specific transporters has been hampered by the lack of functional genetic tools. Here, we report knockouts of three Drosophila melanogaster ABC transporter genes, Mdr49, Mdr50, and Mdr65, that are homologous to the well-studied mammalian ABCB1 (P-glycoprotein). Each knockout mutant was created in the same wild type background and tested against a panel of insecticides representing different chemical classes. Mdr65 knockouts were more susceptible to all neuroactive insecticides tested, but Mdr49 and Mdr50 knockouts showed increased susceptibility or resistance depending on the insecticide used. Mdr65 was chosen for further analysis. Calculation of LC 50 values for the Mdr65 knockout allowed the substrate specificity of this transporter to be examined. No obvious distinguishing structural features were shared among MDR65 substrates. A role for Mdr65 in insecticide transport was confirmed by testing the capacity of the knockout to synergize with the ABC inhibitor verapamil and by measuring the levels of insecticide retained in the body of knockout flies. These data unambiguously establish the influence of ABC transporters on the capacity of wild type D. melanogaster to tolerate insecticide exposure and suggest that both tissue and substrate specificity underpin this capacity. Copyright © 2017 Elsevier Ltd. All rights reserved.

Proton-induced knockout reactions with netron-rich oxygen isotopes at R{sup 3}B

Energy Technology Data Exchange (ETDEWEB)

Atar, Leyla [IKP, TU Darmstadt (Germany); GSI (Germany); Collaboration: R3B-Collaboration

2014-07-01

Proton-induced knockout reactions are one of the main goal of the experimental program at the future R{sup 3}B (Reactions with Relativistic Radioactive Beams) Experiment at FAIR. It allows us to obtain spectroscopic information about valence and deeply bound single-nucleon states and to study their evolution over a large variation in isospin. Recent studies have shown that the occupancies of loosely bound valence nucleons in neutron- or proton-rich nuclei have a spectroscopic factor close to unity, whereas single-particle strength for deeply bound nucleons is suppressed in isospin asymmetric systems compared to the predictions of the many-body shell model. Further experimental and theoretical studies are needed for a qualitative and quantitative understanding. For this aim a series of measurements have been performed on the complete oxygen isotopic chain using the existing experimental setup LAND/R{sup 3}B at GSI. We present the main scientific goals, the concepts of the experiment and the preliminary results.

Sdhd and SDHD/H19 knockout mice do not develop paraganglioma or pheochromocytoma.

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Jean-Pierre Bayley

Full Text Available BACKGROUND: Mitochondrial succinate dehydrogenase (SDH is a component of both the tricarboxylic acid cycle and the electron transport chain. Mutations of SDHD, the first protein of intermediary metabolism shown to be involved in tumorigenesis, lead to the human tumors paraganglioma (PGL and pheochromocytoma (PC. SDHD is remarkable in showing an 'imprinted' tumor suppressor phenotype. Mutations of SDHD show a very high penetrance in man and we postulated that knockout of Sdhd would lead to the development of PGL/PC, probably in aged mice. METHODOLOGY/PRINCIPAL FINDINGS: We generated a conventional knockout of Sdhd in the mouse, removing the entire third exon. We also crossed this mouse with a knockout of H19, a postulated imprinted modifier gene of Sdhd tumorigenesis, to evaluate if loss of these genes together would lead to the initiation or enhancement of tumor development. Homozygous knockout of Sdhd results in embryonic lethality. No paraganglioma or other tumor development was seen in Sdhd KO mice followed for their entire lifespan, in sharp contrast to the highly penetrant phenotype in humans. Heterozygous Sdhd KO mice did not show hyperplasia of paraganglioma-related tissues such as the carotid body or of the adrenal medulla, or any genotype-related pathology, with similar body and organ weights to wildtype mice. A cohort of Sdhd/H19 KO mice developed several cases of profound cardiac hypertrophy, but showed no evidence of PGL/PC. CONCLUSIONS: Knockout of Sdhd in the mouse does not result in a disease phenotype. H19 may not be an initiator of PGL/PC tumorigenesis.

Development of the Multiple Gene Knockout System with One-Step PCR in Thermoacidophilic Crenarchaeon Sulfolobus acidocaldarius

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Shoji Suzuki

2017-01-01

Full Text Available Multiple gene knockout systems developed in the thermoacidophilic crenarchaeon Sulfolobus acidocaldarius are powerful genetic tools. However, plasmid construction typically requires several steps. Alternatively, PCR tailing for high-throughput gene disruption was also developed in S. acidocaldarius, but repeated gene knockout based on PCR tailing has been limited due to lack of a genetic marker system. In this study, we demonstrated efficient homologous recombination frequency (2.8 × 104 ± 6.9 × 103 colonies/μg DNA by optimizing the transformation conditions. This optimized protocol allowed to develop reliable gene knockout via double crossover using short homologous arms and to establish the multiple gene knockout system with one-step PCR (MONSTER. In the MONSTER, a multiple gene knockout cassette was simply and rapidly constructed by one-step PCR without plasmid construction, and the PCR product can be immediately used for target gene deletion. As an example of the applications of this strategy, we successfully made a DNA photolyase- (phr- and arginine decarboxylase- (argD- deficient strain of S. acidocaldarius. In addition, an agmatine selection system consisting of an agmatine-auxotrophic strain and argD marker was also established. The MONSTER provides an alternative strategy that enables the very simple construction of multiple gene knockout cassettes for genetic studies in S. acidocaldarius.

Fluctuation localization imaging-based fluorescence in situ hybridization (fliFISH) for accurate detection and counting of RNA copies in single cells

Energy Technology Data Exchange (ETDEWEB)

Cui, Yi; Hu, Dehong; Markillie, Lye Meng; Chrisler, William B.; Gaffrey, Matthew J.; Ansong, Charles; Sussel, Lori; Orr, Galya

2017-10-04

Quantitative gene expression analysis in intact single cells can be achieved using single molecule- based fluorescence in situ hybridization (smFISH). This approach relies on fluorescence intensity to distinguish between true signals, emitted from an RNA copy hybridized with multiple FISH sub-probes, and background noise. Thus, the precision in smFISH is often compromised by partial or nonspecific binding of sub-probes and tissue autofluorescence, limiting its accuracy. Here we provide an accurate approach for setting quantitative thresholds between true and false signals, which relies on blinking frequencies of photoswitchable dyes. This fluctuation localization imaging-based FISH (fliFISH) uses blinking frequency patterns, emitted from a transcript bound to multiple sub-probes, which are distinct from blinking patterns emitted from partial or nonspecifically bound sub-probes and autofluorescence. Using multicolor fliFISH, we identified radial gene expression patterns in mouse pancreatic islets for insulin, the transcription factor, NKX2-2, and their ratio (Nkx2-2/Ins2). These radial patterns, showing higher values in β cells at the islet core and lower values in peripheral cells, were lost in diabetic mouse islets. In summary, fliFISH provides an accurate, quantitative approach for detecting and counting true RNA copies and rejecting false signals by their distinct blinking frequency patterns, laying the foundation for reliable single-cell transcriptomics.

Optimal knockout strategies in genome-scale metabolic networks using particle swarm optimization.

Science.gov (United States)

Nair, Govind; Jungreuthmayer, Christian; Zanghellini, Jürgen

2017-02-01

Knockout strategies, particularly the concept of constrained minimal cut sets (cMCSs), are an important part of the arsenal of tools used in manipulating metabolic networks. Given a specific design, cMCSs can be calculated even in genome-scale networks. We would however like to find not only the optimal intervention strategy for a given design but the best possible design too. Our solution (PSOMCS) is to use particle swarm optimization (PSO) along with the direct calculation of cMCSs from the stoichiometric matrix to obtain optimal designs satisfying multiple objectives. To illustrate the working of PSOMCS, we apply it to a toy network. Next we show its superiority by comparing its performance against other comparable methods on a medium sized E. coli core metabolic network. PSOMCS not only finds solutions comparable to previously published results but also it is orders of magnitude faster. Finally, we use PSOMCS to predict knockouts satisfying multiple objectives in a genome-scale metabolic model of E. coli and compare it with OptKnock and RobustKnock. PSOMCS finds competitive knockout strategies and designs compared to other current methods and is in some cases significantly faster. It can be used in identifying knockouts which will force optimal desired behaviors in large and genome scale metabolic networks. It will be even more useful as larger metabolic models of industrially relevant organisms become available.

IKKε knockout prevents high fat diet induced arterial atherosclerosis and NF-κB signaling in mice.

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Changchun Cao

Full Text Available AIMS: Atherosclerosis is a public health concern affecting many worldwide, but its pathogenesis remains unclear. In this study we investigated the role of IKKε during the formation of atherosclerosis and its molecular mechanism in the mouse aortic vessel wall. METHODS AND RESULTS: C57BL/6 wild-type or IKKε knockout mice bred into the ApoE knockout genetic background were divided into 4 groups: (1 wild-type (WT, (2 ApoE knockout (AK, (3 IKKε knockout (IK, (4 or both ApoE and IKKε knockout (DK. Each group of mice were fed with a high fat diet (HFD for 12 weeks from 8 weeks of age. Immunohistochemistry and Western blotting analysis demonstrated obvious increases in the expression of IKKε in the AK group compared with the WT group, especially in the intima. Serum lipid levels were significantly higher in the AK and DK groups than in the other two groups. Staining with hematoxylin-eosin and Oil Red, as well as scanning electron microscopy revealed less severe atherosclerotic lesions in the DK group than in the AK group. Immunofluorescence and Western blot analysis demonstrated obvious increases in the expression of NF-κB pathway components and downstream factors in the AK group, especially in the intima, while these increases were blocked in the DK group. CONCLUSION: The knockout of IKKε prevented significant atherosclerosis lesions in the mouse aorta from in both wild-type and ApoE knockout mice fed a HFD, suggesting that IKKε may play a vital role in HFD-induced atherosclerosis and would be an important target for the treatment of atherosclerosis.

Fetal growth retardation and lack of hypotaurine in ezrin knockout mice.

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Tomohiro Nishimura

Full Text Available Ezrin is a membrane-associated cytoplasmic protein that serves to link cell-membrane proteins with the actin-based cytoskeleton, and also plays a role in regulation of the functional activities of some transmembrane proteins. It is expressed in placental trophoblasts. We hypothesized that placental ezrin is involved in the supply of nutrients from mother to fetus, thereby influencing fetal growth. The aim of this study was firstly to clarify the effect of ezrin on fetal growth and secondly to determine whether knockout of ezrin is associated with decreased concentrations of serum and placental nutrients. Ezrin knockout mice (Ez(-/- were confirmed to exhibit fetal growth retardation. Metabolome analysis of fetal serum and placental extract of ezrin knockout mice by means of capillary electrophoresis-time-of-flight mass spectrometry revealed a markedly decreased concentration of hypotaurine, a precursor of taurine. However, placental levels of cysteine and cysteine sulfinic acid (precursors of hypotaurine and taurine were not affected. Lack of hypotaurine in Ez(-/- mice was confirmed by liquid chromatography with tandem mass spectrometry. Administration of hypotaurine to heterogenous dams significantly decreased the placenta-to-maternal plasma ratio of hypotaurine in wild-type fetuses but only slightly decreased it in ezrin knockout fetuses, indicating that the uptake of hypotaurine from mother to placenta is saturable and that disruption of ezrin impairs the uptake of hypotaurine by placental trophoblasts. These results indicate that ezrin is required for uptake of hypotaurine from maternal serum by placental trophoblasts, and plays an important role in fetal growth.

Unimpaired dendritic cell functions in MVP/LRP knockout mice.

Science.gov (United States)

Mossink, Marieke H; de Groot, Jan; van Zon, Arend; Fränzel-Luiten, Erna; Schoester, Martijn; Scheffer, George L; Sonneveld, Pieter; Scheper, Rik J; Wiemer, Erik A C

2003-09-01

Dendritic cells (DCs) act as mobile sentinels of the immune system. By stimulating T lymphocytes, DCs are pivotal for the initiation of both T- and B-cell-mediated immune responses. Recently, ribonucleoprotein particles (vaults) were found to be involved in the development and/or function of human DCs. To further investigate the role of vaults in DCs, we examined the effects of disruption of the major vault protein (MVP/LRP) on the development and antigen-presenting capacity of DCs, using our MVP/LRP knockout mouse model. Mononuclear bone marrow cells were isolated from wild-type and knockout mice and stimulated to differentiate to DCs. Like human DCs, the wild-type murine DC cultures strongly expressed MVP/LRP. Nevertheless, the MVP/LRP-deficient DCs developed normally and showed similar expression levels of several DC surface markers. No differences were observed in in vitro studies on the antigen uptake and presenting capacities of the wild-type and MVP/LRP knockout DCs. Moreover, immunization of the MVP/LRP-deficient mice with several T-cell antigens led to responses similar to those observed in the wild-type mice, indicating that the in vivo DC migration and antigen-presentation capacities are intact. Moreover, no differences were observed in the induction of the T cell-dependent humoral responses and orally induced peripheral T-cell tolerance. In conclusion, vaults are not required for primary DC functions. Their abundance in DCs may, however, still reflect basic roles in myeloid cell proliferation and DC development.

Human Genetic Disorders and Knockout Mice Deficient in Glycosaminoglycan

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Shuji Mizumoto

2014-01-01

Full Text Available Glycosaminoglycans (GAGs are constructed through the stepwise addition of respective monosaccharides by various glycosyltransferases and maturated by epimerases and sulfotransferases. The structural diversity of GAG polysaccharides, including their sulfation patterns and sequential arrangements, is essential for a wide range of biological activities such as cell signaling, cell proliferation, tissue morphogenesis, and interactions with various growth factors. Studies using knockout mice of enzymes responsible for the biosynthesis of the GAG side chains of proteoglycans have revealed their physiological functions. Furthermore, mutations in the human genes encoding glycosyltransferases, sulfotransferases, and related enzymes responsible for the biosynthesis of GAGs cause a number of genetic disorders including chondrodysplasia, spondyloepiphyseal dysplasia, and Ehlers-Danlos syndromes. This review focused on the increasing number of glycobiological studies on knockout mice and genetic diseases caused by disturbances in the biosynthetic enzymes for GAGs.

Phenotypic assessment of THC discriminative stimulus properties in fatty acid amide hydrolase knockout and wildtype mice.

Science.gov (United States)

Walentiny, D Matthew; Vann, Robert E; Wiley, Jenny L

2015-06-01

A number of studies have examined the ability of the endogenous cannabinoid anandamide to elicit Δ(9)-tetrahydrocannabinol (THC)-like subjective effects, as modeled through the THC discrimination paradigm. In the present study, we compared transgenic mice lacking fatty acid amide hydrolase (FAAH), the enzyme primarily responsible for anandamide catabolism, to wildtype counterparts in a THC discrimination procedure. THC (5.6 mg/kg) served as a discriminative stimulus in both genotypes, with similar THC dose-response curves between groups. Anandamide fully substituted for THC in FAAH knockout, but not wildtype, mice. Conversely, the metabolically stable anandamide analog O-1812 fully substituted in both groups, but was more potent in knockouts. The CB1 receptor antagonist rimonabant dose-dependently attenuated THC generalization in both groups and anandamide substitution in FAAH knockouts. Pharmacological inhibition of monoacylglycerol lipase (MAGL), the primary catabolic enzyme for the endocannabinoid 2-arachidonoylglycerol (2-AG), with JZL184 resulted in full substitution for THC in FAAH knockout mice and nearly full substitution in wildtypes. Quantification of brain endocannabinoid levels revealed expected elevations in anandamide in FAAH knockout mice compared to wildtypes and equipotent dose-dependent elevations in 2-AG following JZL184 administration. Dual inhibition of FAAH and MAGL with JZL195 resulted in roughly equipotent increases in THC-appropriate responding in both groups. While the notable similarity in THC's discriminative stimulus effects across genotype suggests that the increased baseline brain anandamide levels (as seen in FAAH knockout mice) do not alter THC's subjective effects, FAAH knockout mice are more sensitive to the THC-like effects of pharmacologically induced increases in anandamide and MAGL inhibition (e.g., JZL184). Copyright © 2015 Elsevier Ltd. All rights reserved.

Quasi-free knockout reactions with the proton-dripline nucleus {sup 17}Ne

Energy Technology Data Exchange (ETDEWEB)

Wamers, Felix; Aumann, Thomas [Institut fuer Kernphysik, TU, Darmstadt (Germany); Heil, Michael [Kernreaktionen und Nukleare Astrophysik, GSI, Darmstadt (Germany); Marganiec, Justyna [ExtreMe Matter Institute EMMI, GSI, Darmstadt (Germany); Plag, Ralf [Kernreaktionen und Nukleare Astrophysik, GSI, Darmstadt (Germany); Goethe Universitaet, Frankfurt a.M. (Germany); Collaboration: R3B-Collaboration

2011-07-01

{sup 17}Ne is a proton-dripline nucleus that has raised special interest in nuclear-structure physics in recent years. As a ({sup 15}O+2p) Borromean 3-body system, it is often considered to be a 2-proton-halo nucleus, yet lacking concluding experimental evidence about its structure. We have studied breakup reactions of 500 AMeV {sup 17}Ne secondary beams using the R{sup 3}B-LAND setup at GSI. One focus was on the quasi-free one-proton knockout in a proton-rich paraffin (CH{sub 2}) target in inverse kinematics, i.e., {sup 17}Ne(p,2p){sup 16}F{yields}{sup 15}O+p, in comparison to the one-proton knockout with a carbon target. Recoil protons have been detected with Si-Strip detectors and the surrounding 4{pi} NaI spectrometer ''Crystal Ball'', thus providing a clean signature for quasi-free knockout. First results on two-proton removal cross sections with CH{sub 2} and C targets will be presented, as well as transverse momentum distributions of the {sup 15}O core in {sup 17}Ne. Projectile-like forward protons after one-proton knockout from {sup 17}Ne have been measured in coincidence with the {sup 15}O residual core, leading to the relative-energy spectrum of the unbound {sup 16}F. Possible interpretations and implications regarding the structure of {sup 17}Ne are discussed.

Generation of knockout rabbits using transcription activator-like effector nucleases.

Science.gov (United States)

Wang, Yu; Fan, Nana; Song, Jun; Zhong, Juan; Guo, Xiaogang; Tian, Weihua; Zhang, Quanjun; Cui, Fenggong; Li, Li; Newsome, Philip N; Frampton, Jon; Esteban, Miguel A; Lai, Liangxue

2014-01-01

Zinc-finger nucleases and transcription activator-like effector nucleases are novel gene-editing platforms contributing to redefine the boundaries of modern biological research. They are composed of a non-specific cleavage domain and a tailor made DNA-binding module, which enables a broad range of genetic modifications by inducing efficient DNA double-strand breaks at desired loci. Among other remarkable uses, these nucleases have been employed to produce gene knockouts in mid-size and large animals, such as rabbits and pigs, respectively. This approach is cost effective, relatively quick, and can produce invaluable models for human disease studies, biotechnology or agricultural purposes. Here we describe a protocol for the efficient generation of knockout rabbits using transcription activator-like effector nucleases, and a perspective of the field.

Global gene expression profiling in PAI-1 knockout murine heart and kidney: molecular basis of cardiac-selective fibrosis.

Directory of Open Access Journals (Sweden)

Asish K Ghosh

Full Text Available Fibrosis is defined as an abnormal matrix remodeling due to excessive synthesis and accumulation of extracellular matrix proteins in tissues during wound healing or in response to chemical, mechanical and immunological stresses. At present, there is no effective therapy for organ fibrosis. Previous studies demonstrated that aged plasminogen activator inhibitor-1 (PAI-1 knockout mice develop spontaneously cardiac-selective fibrosis without affecting any other organs. We hypothesized that differential expressions of profibrotic and antifibrotic genes in PAI-1 knockout hearts and unaffected organs lead to cardiac selective fibrosis. In order to address this prediction, we have used a genome-wide gene expression profiling of transcripts derived from aged PAI-1 knockout hearts and kidneys. The variations of global gene expression profiling were compared within four groups: wildtype heart vs. knockout heart; wildtype kidney vs. knockout kidney; knockout heart vs. knockout kidney and wildtype heart vs. wildtype kidney. Analysis of illumina-based microarray data revealed that several genes involved in different biological processes such as immune system processing, response to stress, cytokine signaling, cell proliferation, adhesion, migration, matrix organization and transcriptional regulation were affected in hearts and kidneys by the absence of PAI-1, a potent inhibitor of urokinase and tissue-type plasminogen activator. Importantly, the expressions of a number of genes, involved in profibrotic pathways including Ankrd1, Pi16, Egr1, Scx, Timp1, Timp2, Klf6, Loxl1 and Klotho, were deregulated in PAI-1 knockout hearts compared to wildtype hearts and PAI-1 knockout kidneys. While the levels of Ankrd1, Pi16 and Timp1 proteins were elevated during EndMT, the level of Timp4 protein was decreased. To our knowledge, this is the first comprehensive report on the influence of PAI-1 on global gene expression profiling in the heart and kidney and its implication

Knockout mutations of insulin-like peptide genes enhance sexual receptivity in Drosophila virgin females.

Science.gov (United States)

Watanabe, Kazuki; Sakai, Takaomi

2016-01-01

In the fruitfly Drosophila melanogaster, females take the initiative to mate successfully because they decide whether to mate or not. However, little is known about the molecular and neuronal mechanisms regulating sexual receptivity in virgin females. Genetic tools available in Drosophila are useful for identifying molecules and neural circuits involved in the regulation of sexual receptivity. We previously demonstrated that insulin-producing cells (IPCs) in the female brain are critical to the regulation of female sexual receptivity. Ablation and inactivation of IPCs enhance female sexual receptivity, suggesting that neurosecretion from IPCs inhibits female sexual receptivity. IPCs produce and release insulin-like peptides (Ilps) that modulate various biological processes such as metabolism, growth, lifespan and behaviors. Here, we report a novel role of the Ilps in sexual behavior in Drosophila virgin females. Compared with wild-type females, females with knockout mutations of Ilps showed a high mating success rate toward wild-type males, whereas wild-type males courted wild-type and Ilp-knockout females to the same extent. Wild-type receptive females retard their movement during male courtship and this reduced female mobility allows males to copulate. Thus, it was anticipated that knockout mutations of Ilps would reduce general locomotion. However, the locomotor activity in Ilp-knockout females was significantly higher than that in wild-type females. Thus, our findings indicate that the high mating success rate in Ilp-knockout females is caused by their enhanced sexual receptivity, but not by improvement of their sex appeal or by general sluggishness.

Quasi-free one nucleon knockout reactions on neutron-rich oxygen isotopes at the R3B-LAND setup

Energy Technology Data Exchange (ETDEWEB)

Atar, Leyla; Aumann, Thomas [TU Darmstadt, Darmstadt (Germany); GSI, Darmstadt (Germany); Bertulani, Carlos [Texas A and M University-Commerce, Commerce (United States); Paschalis, Stefanos [TU Darmstadt, Darmstadt (Germany); Nociforo, Chiara [GSI, Darmstadt (Germany); Collaboration: R3B-Collaboration

2016-07-01

Recent experiments have showed a reduction of spectroscopic strengths of about 60-70% for stable nuclei. When going to driplines this tendency is changing, loosely bound nucleons have spectroscopic strengths close unity while deeply bound nucleons have a large reduction of the strength. We aim to make a systematic study of spectroscopic factors (SF) of the Oxygen isotopes using quasi-free (p,2p) and (p,pn) knockout reactions in inverse kinematics. Quasi-free knockout reactions are a direct tool to study the occupancy and the location of valance and deeply bound single particle states. The Oxygen isotopes offer a large variation of separation energies which will allow us to obtain a qualitative and quantitative understanding of SF in a large variation of isospin asymmetry. For this we performed an experiment at the R3B-LAND setup at the GSI with a secondary beam {sup 14-24}O. The {sup 16-18}O and {sup 21-23}O isotopes have been analyzed and the preliminary results will be presented. The results include the partial cross sections, gamma ray spectra of the residual fragments in coincidence, and the SF obtained via comparison with theory.

Robust and sensitive analysis of mouse knockout phenotypes.

Directory of Open Access Journals (Sweden)

Natasha A Karp

Full Text Available A significant challenge of in-vivo studies is the identification of phenotypes with a method that is robust and reliable. The challenge arises from practical issues that lead to experimental designs which are not ideal. Breeding issues, particularly in the presence of fertility or fecundity problems, frequently lead to data being collected in multiple batches. This problem is acute in high throughput phenotyping programs. In addition, in a high throughput environment operational issues lead to controls not being measured on the same day as knockouts. We highlight how application of traditional methods, such as a Student's t-Test or a 2-way ANOVA, in these situations give flawed results and should not be used. We explore the use of mixed models using worked examples from Sanger Mouse Genome Project focusing on Dual-Energy X-Ray Absorptiometry data for the analysis of mouse knockout data and compare to a reference range approach. We show that mixed model analysis is more sensitive and less prone to artefacts allowing the discovery of subtle quantitative phenotypes essential for correlating a gene's function to human disease. We demonstrate how a mixed model approach has the additional advantage of being able to include covariates, such as body weight, to separate effect of genotype from these covariates. This is a particular issue in knockout studies, where body weight is a common phenotype and will enhance the precision of assigning phenotypes and the subsequent selection of lines for secondary phenotyping. The use of mixed models with in-vivo studies has value not only in improving the quality and sensitivity of the data analysis but also ethically as a method suitable for small batches which reduces the breeding burden of a colony. This will reduce the use of animals, increase throughput, and decrease cost whilst improving the quality and depth of knowledge gained.

Accurate determination of genetic identity for a single cacao bean, using molecular markers with a nanofluidic system, ensures cocoa authentication.

Science.gov (United States)

Fang, Wanping; Meinhardt, Lyndel W; Mischke, Sue; Bellato, Cláudia M; Motilal, Lambert; Zhang, Dapeng

2014-01-15

Cacao (Theobroma cacao L.), the source of cocoa, is an economically important tropical crop. One problem with the premium cacao market is contamination with off-types adulterating raw premium material. Accurate determination of the genetic identity of single cacao beans is essential for ensuring cocoa authentication. Using nanofluidic single nucleotide polymorphism (SNP) genotyping with 48 SNP markers, we generated SNP fingerprints for small quantities of DNA extracted from the seed coat of single cacao beans. On the basis of the SNP profiles, we identified an assumed adulterant variety, which was unambiguously distinguished from the authentic beans by multilocus matching. Assignment tests based on both Bayesian clustering analysis and allele frequency clearly separated all 30 authentic samples from the non-authentic samples. Distance-based principle coordinate analysis further supported these results. The nanofluidic SNP protocol, together with forensic statistical tools, is sufficiently robust to establish authentication and to verify gourmet cacao varieties. This method shows significant potential for practical application.

In Vivo Knockout of the Vegfa Gene by Lentiviral Delivery of CRISPR/Cas9 in Mouse Retinal Pigment Epithelium Cells

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Andreas Holmgaard

2017-12-01

Full Text Available Virus-based gene therapy by CRISPR/Cas9-mediated genome editing and knockout may provide a new option for treatment of inherited and acquired ocular diseases of the retina. In support of this notion, we show that Streptococcus pyogenes (Sp Cas9, delivered by lentiviral vectors (LVs, can be used in vivo to selectively ablate the vascular endothelial growth factor A (Vegfa gene in mice. By generating LVs encoding SpCas9 targeted to Vegfa, and in parallel the fluorescent eGFP marker protein, we demonstrate robust knockout of Vegfa that leads to a significant reduction of VEGFA protein in transduced cells. Three of the designed single-guide RNAs (sgRNAs induce in vitro indel formation at high frequencies (44%–93%. A single unilateral subretinal injection facilitates RPE-specific localization of the vector and disruption of Vegfa in isolated eGFP+ RPE cells obtained from mice five weeks after LV administration. Notably, sgRNA delivery results in the disruption of Vegfa with an in vivo indel formation efficacy of up to 84%. Sequencing of Vegfa-specific amplicons reveals formation of indels, including 4-bp deletions and 2-bp insertions. Taken together, our data demonstrate the capacity of lentivirus-delivered SpCas9 and sgRNAs as a developing therapeutic path in the treatment of ocular diseases, including age-related macular degeneration.

Quasifree (p , 2 p ) Reactions on Oxygen Isotopes: Observation of Isospin Independence of the Reduced Single-Particle Strength

Science.gov (United States)

Atar, L.; Paschalis, S.; Barbieri, C.; Bertulani, C. A.; Díaz Fernández, P.; Holl, M.; Najafi, M. A.; Panin, V.; Alvarez-Pol, H.; Aumann, T.; Avdeichikov, V.; Beceiro-Novo, S.; Bemmerer, D.; Benlliure, J.; Boillos, J. M.; Boretzky, K.; Borge, M. J. G.; Caamaño, M.; Caesar, C.; Casarejos, E.; Catford, W.; Cederkall, J.; Chartier, M.; Chulkov, L.; Cortina-Gil, D.; Cravo, E.; Crespo, R.; Dillmann, I.; Elekes, Z.; Enders, J.; Ershova, O.; Estrade, A.; Farinon, F.; Fraile, L. M.; Freer, M.; Galaviz Redondo, D.; Geissel, H.; Gernhäuser, R.; Golubev, P.; Göbel, K.; Hagdahl, J.; Heftrich, T.; Heil, M.; Heine, M.; Heinz, A.; Henriques, A.; Hufnagel, A.; Ignatov, A.; Johansson, H. T.; Jonson, B.; Kahlbow, J.; Kalantar-Nayestanaki, N.; Kanungo, R.; Kelic-Heil, A.; Knyazev, A.; Kröll, T.; Kurz, N.; Labiche, M.; Langer, C.; Le Bleis, T.; Lemmon, R.; Lindberg, S.; Machado, J.; Marganiec-Gałązka, J.; Movsesyan, A.; Nacher, E.; Nikolskii, E. Y.; Nilsson, T.; Nociforo, C.; Perea, A.; Petri, M.; Pietri, S.; Plag, R.; Reifarth, R.; Ribeiro, G.; Rigollet, C.; Rossi, D. M.; Röder, M.; Savran, D.; Scheit, H.; Simon, H.; Sorlin, O.; Syndikus, I.; Taylor, J. T.; Tengblad, O.; Thies, R.; Togano, Y.; Vandebrouck, M.; Velho, P.; Volkov, V.; Wagner, A.; Wamers, F.; Weick, H.; Wheldon, C.; Wilson, G. L.; Winfield, J. S.; Woods, P.; Yakorev, D.; Zhukov, M.; Zilges, A.; Zuber, K.; R3B Collaboration

2018-01-01

Quasifree one-proton knockout reactions have been employed in inverse kinematics for a systematic study of the structure of stable and exotic oxygen isotopes at the R3B /LAND setup with incident beam energies in the range of 300 - 450 MeV /u . The oxygen isotopic chain offers a large variation of separation energies that allows for a quantitative understanding of single-particle strength with changing isospin asymmetry. Quasifree knockout reactions provide a complementary approach to intermediate-energy one-nucleon removal reactions. Inclusive cross sections for quasifree knockout reactions of the type O A (p ,2 p )N-1A have been determined and compared to calculations based on the eikonal reaction theory. The reduction factors for the single-particle strength with respect to the independent-particle model were obtained and compared to state-of-the-art ab initio predictions. The results do not show any significant dependence on proton-neutron asymmetry.

Quasifree (p, 2p) Reactions on Oxygen Isotopes: Observation of Isospin Independence of the Reduced Single-Particle Strength.

Science.gov (United States)

Atar, L; Paschalis, S; Barbieri, C; Bertulani, C A; Díaz Fernández, P; Holl, M; Najafi, M A; Panin, V; Alvarez-Pol, H; Aumann, T; Avdeichikov, V; Beceiro-Novo, S; Bemmerer, D; Benlliure, J; Boillos, J M; Boretzky, K; Borge, M J G; Caamaño, M; Caesar, C; Casarejos, E; Catford, W; Cederkall, J; Chartier, M; Chulkov, L; Cortina-Gil, D; Cravo, E; Crespo, R; Dillmann, I; Elekes, Z; Enders, J; Ershova, O; Estrade, A; Farinon, F; Fraile, L M; Freer, M; Galaviz Redondo, D; Geissel, H; Gernhäuser, R; Golubev, P; Göbel, K; Hagdahl, J; Heftrich, T; Heil, M; Heine, M; Heinz, A; Henriques, A; Hufnagel, A; Ignatov, A; Johansson, H T; Jonson, B; Kahlbow, J; Kalantar-Nayestanaki, N; Kanungo, R; Kelic-Heil, A; Knyazev, A; Kröll, T; Kurz, N; Labiche, M; Langer, C; Le Bleis, T; Lemmon, R; Lindberg, S; Machado, J; Marganiec-Gałązka, J; Movsesyan, A; Nacher, E; Nikolskii, E Y; Nilsson, T; Nociforo, C; Perea, A; Petri, M; Pietri, S; Plag, R; Reifarth, R; Ribeiro, G; Rigollet, C; Rossi, D M; Röder, M; Savran, D; Scheit, H; Simon, H; Sorlin, O; Syndikus, I; Taylor, J T; Tengblad, O; Thies, R; Togano, Y; Vandebrouck, M; Velho, P; Volkov, V; Wagner, A; Wamers, F; Weick, H; Wheldon, C; Wilson, G L; Winfield, J S; Woods, P; Yakorev, D; Zhukov, M; Zilges, A; Zuber, K

2018-02-02

Quasifree one-proton knockout reactions have been employed in inverse kinematics for a systematic study of the structure of stable and exotic oxygen isotopes at the R^{3}B/LAND setup with incident beam energies in the range of 300-450  MeV/u. The oxygen isotopic chain offers a large variation of separation energies that allows for a quantitative understanding of single-particle strength with changing isospin asymmetry. Quasifree knockout reactions provide a complementary approach to intermediate-energy one-nucleon removal reactions. Inclusive cross sections for quasifree knockout reactions of the type ^{A}O(p,2p)^{A-1}N have been determined and compared to calculations based on the eikonal reaction theory. The reduction factors for the single-particle strength with respect to the independent-particle model were obtained and compared to state-of-the-art ab initio predictions. The results do not show any significant dependence on proton-neutron asymmetry.

Generation of knockout rabbits using transcription activator-like effector nucleases

Directory of Open Access Journals (Sweden)

Yu Wang

2014-01-01

Full Text Available Zinc-finger nucleases and transcription activator-like effector nucleases are novel gene-editing platforms contributing to redefine the boundaries of modern biological research. They are composed of a non-specific cleavage domain and a tailor made DNA-binding module, which enables a broad range of genetic modifications by inducing efficient DNA double-strand breaks at desired loci. Among other remarkable uses, these nucleases have been employed to produce gene knockouts in mid-size and large animals, such as rabbits and pigs, respectively. This approach is cost effective, relatively quick, and can produce invaluable models for human disease studies, biotechnology or agricultural purposes. Here we describe a protocol for the efficient generation of knockout rabbits using transcription activator-like effector nucleases, and a perspective of the field.

The Knockout of Secretin in Cerebellar Purkinje Cells Impairs Mouse Motor Coordination and Motor Learning

Science.gov (United States)

Zhang, Li; Chung, Sookja Kim; Chow, Billy Kwok Chong

2014-01-01

Secretin (SCT) was first considered to be a gut hormone regulating gastrointestinal functions when discovered. Recently, however, central actions of SCT have drawn intense research interest and are supported by the broad distribution of SCT in specific neuronal populations and by in vivo physiological studies regarding its role in water homeostasis and food intake. The direct action of SCT on a central neuron was first discovered in cerebellar Purkinje cells in which SCT from cerebellar Purkinje cells was found to potentiate GABAergic inhibitory transmission from presynaptic basket cells. Because Purkinje neurons have a major role in motor coordination and learning functions, we hypothesize a behavioral modulatory function for SCT. In this study, we successfully generated a mouse model in which the SCT gene was deleted specifically in Purkinje cells. This mouse line was tested together with SCT knockout and SCT receptor knockout mice in a full battery of behavioral tasks. We found that the knockout of SCT in Purkinje neurons did not affect general motor ability or the anxiety level in open field tests. However, knockout mice did exhibit impairments in neuromuscular strength, motor coordination, and motor learning abilities, as shown by wire hanging, vertical climbing, and rotarod tests. In addition, SCT knockout in Purkinje cells possibly led to the delayed development of motor neurons, as supported by the later occurrence of key neural reflexes. In summary, our data suggest a role in motor coordination and motor learning for SCT expressed in cerebellar Purkinje cells. PMID:24356714

Highly accurate surface maps from profilometer measurements

Science.gov (United States)

Medicus, Kate M.; Nelson, Jessica D.; Mandina, Mike P.

2013-04-01

Many aspheres and free-form optical surfaces are measured using a single line trace profilometer which is limiting because accurate 3D corrections are not possible with the single trace. We show a method to produce an accurate fully 2.5D surface height map when measuring a surface with a profilometer using only 6 traces and without expensive hardware. The 6 traces are taken at varying angular positions of the lens, rotating the part between each trace. The output height map contains low form error only, the first 36 Zernikes. The accuracy of the height map is ±10% of the actual Zernike values and within ±3% of the actual peak to valley number. The calculated Zernike values are affected by errors in the angular positioning, by the centering of the lens, and to a small effect, choices made in the processing algorithm. We have found that the angular positioning of the part should be better than 1?, which is achievable with typical hardware. The centering of the lens is essential to achieving accurate measurements. The part must be centered to within 0.5% of the diameter to achieve accurate results. This value is achievable with care, with an indicator, but the part must be edged to a clean diameter.

Knockout and fragmentation reactions using a broad range of tin isotopes

Science.gov (United States)

Rodríguez-Sánchez, J. L.; Benlliure, J.; Bertulani, C. A.; Vargas, J.; Ayyad, Y.; Alvarez-Pol, H.; Atkinson, J.; Aumann, T.; Beceiro-Novo, S.; Boretzky, K.; Caamaño, M.; Casarejos, E.; Cortina-Gil, D.; Díaz-Cortes, J.; Fernández, P. Díaz; Estrade, A.; Geissel, H.; Kelić-Heil, A.; Litvinov, Yu. A.; Mostazo, M.; Paradela, C.; Pérez-Loureiro, D.; Pietri, S.; Prochazka, A.; Takechi, M.; Weick, H.; Winfield, J. S.

2017-09-01

Production cross sections of residual nuclei obtained by knockout and fragmentation reactions of different tin isotopes accelerated at 1 A GeV have been measured with the fragment separator (FRS) at GSI, Darmstadt. The new measurements are used to investigate the neutron-excess dependence of the neutron- and proton-knockout cross sections. These cross sections are compared to Glauber model calculations coupled to a nuclear de-excitation code in order to investigate the role of the remnant excitations. This bench marking shows an overestimation of the cross sections for the removal of deeply bound nucleons. A phenomenological increase in the excitation energy induced in the remnants produced in these cases allows us to reproduce the measured cross sections.

Neutron knockout from 68,70Ni ground and isomeric states.

Science.gov (United States)

Recchia, F.; Weisshaar, D.; Gade, A.; Tostevin, J. A.; Janssens, R. V. F.; Albers, M.; Bader, V. M.; Baugher, T.; Bazin, D.; Berryman, J. S.; Brown, B. A.; Campbell, C. M.; Carpenter, M. P.; Chen, J.; Chiara, C. J.; Crawford, H. L.; Hoffman, C. R.; Kondev, F. G.; Korichi, A.; Langer, C.; Lauritsen, T.; Liddick, S. N.; Lunderberg, E.; Noji, S.; Prokop, C.; Stroberg, S. R.; Suchyta, S.; Wimmer, K.; Zhu, S.

2018-02-01

Neutron-rich isotopes are an important source of new information on nuclear physics. Specifically, the spin-isospin components in the nucleon-nucleon (NN) interaction, e.g., the proton-neutron tensor force, are expected to modify shell structure in exotic nuclei. These potential changes in the intrinsic shell structure are of fundamental interest. The study of the excitation energy of states corresponding to specific configurations in even-even isotopes, together with the single-particle character of the first excited states of odd-A, neutron-rich Ni isotopes, probes the evolution of the neutron orbitals around the Fermi surface as a function of the neutron number a step forward in the understanding of the region and the nature of the NN interaction at large N/Z ratios. In an experiment carried out at the National Superconducting Cyclotron Laboratory [1], new spectroscopic information was obtained for 68Ni and the distribution of single-particle strengths in 67,69Ni was characterized by means of single-neutron knockout from 68,70Ni secondary beams. The spectroscopic strengths, deduced from the measured partial cross sections to the individual states tagged by their de-exciting gamma rays, is used to identify and quantify configurations that involve neutron excitations across the N = 40 harmonic oscillator shell closure. The de-excitation γ rays were measured with the GRETINA tracking array [2]. The results challenge the validity of the most current shell-model Hamiltonians and effective interactions, highlighting shortcomings that cannot yet be explained. These results suggest that our understanding of the low-energy states in such nuclei is not complete and requires further investigation.

The importance of immunohistochemical analyses in evaluating the phenotype of Kv channel knockout mice.

Science.gov (United States)

Menegola, Milena; Clark, Eliana; Trimmer, James S

2012-06-01

To gain insights into the phenotype of voltage-gated potassium (Kv)1.1 and Kv4.2 knockout mice, we used immunohistochemistry to analyze the expression of component principal or α subunits and auxiliary subunits of neuronal Kv channels in knockout mouse brains. Genetic ablation of the Kv1.1 α subunit did not result in compensatory changes in the expression levels or subcellular distribution of related ion channel subunits in hippocampal medial perforant path and mossy fiber nerve terminals, where high levels of Kv1.1 are normally expressed. Genetic ablation of the Kv4.2 α subunit did not result in altered neuronal cytoarchitecture of the hippocampus. Although Kv4.2 knockout mice did not exhibit compensatory changes in the expression levels or subcellular distribution of the related Kv4.3 α subunit, we found dramatic decreases in the cellular and subcellular expression of specific Kv channel interacting proteins (KChIPs) that reflected their degree of association and colocalization with Kv4.2 in wild-type mouse and rat brains. These studies highlight the insights that can be gained by performing detailed immunohistochemical analyses of Kv channel knockout mouse brains. Wiley Periodicals, Inc. © 2012 International League Against Epilepsy.

Accurate optical vector network analyzer based on optical single-sideband modulation and balanced photodetection.

Science.gov (United States)

Xue, Min; Pan, Shilong; Zhao, Yongjiu

2015-02-15

A novel optical vector network analyzer (OVNA) based on optical single-sideband (OSSB) modulation and balanced photodetection is proposed and experimentally demonstrated, which can eliminate the measurement error induced by the high-order sidebands in the OSSB signal. According to the analytical model of the conventional OSSB-based OVNA, if the optical carrier in the OSSB signal is fully suppressed, the measurement result is exactly the high-order-sideband-induced measurement error. By splitting the OSSB signal after the optical device-under-test (ODUT) into two paths, removing the optical carrier in one path, and then detecting the two signals in the two paths using a balanced photodetector (BPD), high-order-sideband-induced measurement error can be ideally eliminated. As a result, accurate responses of the ODUT can be achieved without complex post-signal processing. A proof-of-concept experiment is carried out. The magnitude and phase responses of a fiber Bragg grating (FBG) measured by the proposed OVNA with different modulation indices are superimposed, showing that the high-order-sideband-induced measurement error is effectively removed.

Knock-out of a mitochondrial sirtuin protects neurons from degeneration in Caenorhabditis elegans.

Science.gov (United States)

Sangaletti, Rachele; D'Amico, Massimo; Grant, Jeff; Della-Morte, David; Bianchi, Laura

2017-08-01

Sirtuins are NAD⁺-dependent deacetylases, lipoamidases, and ADP-ribosyltransferases that link cellular metabolism to multiple intracellular pathways that influence processes as diverse as cell survival, longevity, and cancer growth. Sirtuins influence the extent of neuronal death in stroke. However, different sirtuins appear to have opposite roles in neuronal protection. In Caenorhabditis elegans, we found that knock-out of mitochondrial sirtuin sir-2.3, homologous to mammalian SIRT4, is protective in both chemical ischemia and hyperactive channel induced necrosis. Furthermore, the protective effect of sir-2.3 knock-out is enhanced by block of glycolysis and eliminated by a null mutation in daf-16/FOXO transcription factor, supporting the involvement of the insulin/IGF pathway. However, data in Caenorhabditis elegans cell culture suggest that the effects of sir-2.3 knock-out act downstream of the DAF-2/IGF-1 receptor. Analysis of ROS in sir-2.3 knock-out reveals that ROS become elevated in this mutant under ischemic conditions in dietary deprivation (DD), but to a lesser extent than in wild type, suggesting more robust activation of a ROS scavenging system in this mutant in the absence of food. This work suggests a deleterious role of SIRT4 during ischemic processes in mammals that must be further investigated and reveals a novel pathway that can be targeted for the design of therapies aimed at protecting neurons from death in ischemic conditions.

Voluntary exercise decreases atherosclerosis in nephrectomised ApoE knockout mice.

Directory of Open Access Journals (Sweden)

Cecilia M Shing

Full Text Available Cardiovascular disease is the main cause of morbidity and mortality in patients with kidney disease. The effectiveness of exercise for cardiovascular disease that is accelerated by the presence of chronic kidney disease remains unknown. The present study utilized apolipoprotein E knockout mice with 5/6 nephrectomy as a model of combined kidney disease and cardiovascular disease to investigate the effect of exercise on aortic plaque formation, vascular function and systemic inflammation. Animals were randomly assigned to nephrectomy or control and then to either voluntary wheel running exercise or sedentary. Following 12-weeks, aortic plaque area was significantly (p0.05. Nephrectomy increased IL-6 and TNF-α concentrations compared with control mice (p0.05. Exercise was an effective non-pharmacologic approach to slow cardiovascular disease in the presence of kidney disease in the apolipoprotein E knockout mouse.

Accurate X-ray diffraction studies of KTiOPO{sub 4} single crystals doped with niobium

Energy Technology Data Exchange (ETDEWEB)

Novikova, N. E., E-mail: natnov@ns.crys.ras.ru; Sorokina, N. I.; Alekseeva, O. A.; Verin, I. A. [Russian Academy of Sciences, Shubnikov Institute of Crystallography, Crystallography and Photonics Federal Scientific Research Center (Russian Federation); Kharitonova, E. P.; Orlova, E. I.; Voronkova, V. I. [Moscow State University, Faculty of Physics (Russian Federation)

2017-01-15

Single crystals of potassium titanyl phosphate doped with 4% of niobium (КТР:4%Nb) and 6% of niobium (KTP:6%Nb) are studied by accurate X-ray diffraction at room temperature. The niobium atoms are localized near the Ti1 and Ti2 atomic positions, and their positions are for the first time refined independent of the titanium atomic positions. Maps of difference electron density in the vicinity of K1 and K2 atomic positions are analyzed. It is found that in the structure of crystal КТР:4%Nb, additional positions of K atoms are located farther from the main positions and from each other than in КТР and KTP:6%Nb crystals. The nonuniform distribution of electron density found in the channels of the КТР:4%Nb structure is responsible for ~20% increase in the signal of second harmonic generation.

The role of nuclear factor E2-Related factor 2 and uncoupling protein 2 in glutathione metabolism: Evidence from an in vivo gene knockout study

International Nuclear Information System (INIS)

Chen, Yanyan; Xu, Yuanyuan; Zheng, Hongzhi; Fu, Jingqi; Hou, Yongyong; Wang, Huihui; Zhang, Qiang; Yamamoto, Masayuki; Pi, Jingbo

2016-01-01

Nuclear factor E2-related factor 2 (NRF2) and uncoupling protein 2 (UCP2) are indicated to protect from oxidative stress. They also play roles in the homeostasis of glutathione. However, the detailed mechanisms are not well understood. In the present study, we found Nrf2-knockout (Nrf2-KO) mice exhibited altered glutathione homeostasis and reduced expression of various genes involved in GSH biosynthesis, regeneration, utilization and transport in the liver. Ucp2-knockout (Ucp2-KO) mice exhibited altered glutathione homeostasis in the liver, spleen and blood, as well as increased transcript of cystic fibrosis transmembrane conductance regulator in the liver, a protein capable of mediating glutathione efflux. Nrf2-Ucp2-double knockout (DKO) mice showed characteristics of both Nrf2-KO and Ucp2-KO mice. But no significant difference was observed in DKO mice when compared with Nrf2-KO or Ucp2-KO mice, except in blood glutathione levels. These data suggest that ablation of Nrf2 and Ucp2 leads to disrupted GSH balance, which could result from altered expression of genes involved in GSH metabolism. DKO may not evoke more severe oxidative stress than the single gene knockout. - Highlights: • Nrf2/Ucp2 deficiency leads to alteration of glutathione homeostasis. • Nrf2 regulates expression of genes in glutathione generation and utilization. • Ucp2 affects glutathione metabolism by regulating hepatic efflux of glutathione. • Nrf2 deficiency may not aggravate oxidative stress in Ucp2-deficient mice.

The role of nuclear factor E2-Related factor 2 and uncoupling protein 2 in glutathione metabolism: Evidence from an in vivo gene knockout study

Energy Technology Data Exchange (ETDEWEB)

Chen, Yanyan [The First Affiliated Hospital, China Medical University, Shenyang, Liaoning (China); The Hamner Institutes for Health Sciences, Research Triangle Park, NC (United States); Xu, Yuanyuan, E-mail: yyxu@cmu.edu.cn [School of Public Health, China Medical University, Shenyang, Liaoning (China); Zheng, Hongzhi [The First Affiliated Hospital, China Medical University, Shenyang, Liaoning (China); The Hamner Institutes for Health Sciences, Research Triangle Park, NC (United States); Fu, Jingqi; Hou, Yongyong; Wang, Huihui [School of Public Health, China Medical University, Shenyang, Liaoning (China); Zhang, Qiang [Rollins School of Public Health, Emory University, Atlanta, GA (United States); Yamamoto, Masayuki [Graduate School of Medicine, Tohoku University, Sendai (Japan); Pi, Jingbo, E-mail: jbpi@cmu.edu.cn [School of Public Health, China Medical University, Shenyang, Liaoning (China); The Hamner Institutes for Health Sciences, Research Triangle Park, NC (United States)

2016-09-09

Nuclear factor E2-related factor 2 (NRF2) and uncoupling protein 2 (UCP2) are indicated to protect from oxidative stress. They also play roles in the homeostasis of glutathione. However, the detailed mechanisms are not well understood. In the present study, we found Nrf2-knockout (Nrf2-KO) mice exhibited altered glutathione homeostasis and reduced expression of various genes involved in GSH biosynthesis, regeneration, utilization and transport in the liver. Ucp2-knockout (Ucp2-KO) mice exhibited altered glutathione homeostasis in the liver, spleen and blood, as well as increased transcript of cystic fibrosis transmembrane conductance regulator in the liver, a protein capable of mediating glutathione efflux. Nrf2-Ucp2-double knockout (DKO) mice showed characteristics of both Nrf2-KO and Ucp2-KO mice. But no significant difference was observed in DKO mice when compared with Nrf2-KO or Ucp2-KO mice, except in blood glutathione levels. These data suggest that ablation of Nrf2 and Ucp2 leads to disrupted GSH balance, which could result from altered expression of genes involved in GSH metabolism. DKO may not evoke more severe oxidative stress than the single gene knockout. - Highlights: • Nrf2/Ucp2 deficiency leads to alteration of glutathione homeostasis. • Nrf2 regulates expression of genes in glutathione generation and utilization. • Ucp2 affects glutathione metabolism by regulating hepatic efflux of glutathione. • Nrf2 deficiency may not aggravate oxidative stress in Ucp2-deficient mice.

A norm knockout method on indirect reciprocity to reveal indispensable norms

Science.gov (United States)

Yamamoto, Hitoshi; Okada, Isamu; Uchida, Satoshi; Sasaki, Tatsuya

2017-03-01

Although various norms for reciprocity-based cooperation have been suggested that are evolutionarily stable against invasion from free riders, the process of alternation of norms and the role of diversified norms remain unclear in the evolution of cooperation. We clarify the co-evolutionary dynamics of norms and cooperation in indirect reciprocity and also identify the indispensable norms for the evolution of cooperation. Inspired by the gene knockout method, a genetic engineering technique, we developed the norm knockout method and clarified the norms necessary for the establishment of cooperation. The results of numerical investigations revealed that the majority of norms gradually transitioned to tolerant norms after defectors are eliminated by strict norms. Furthermore, no cooperation emerges when specific norms that are intolerant to defectors are knocked out.

Probing short-range correlations in asymmetric nuclei with quasi-free pair knockout reactions

Science.gov (United States)

Stevens, Sam; Ryckebusch, Jan; Cosyn, Wim; Waets, Andreas

2018-02-01

Short-range correlations (SRC) in asymmetric nuclei with an unusual neutron-to-proton ratio can be studied with quasi-free two-nucleon knockout processes following the collision between accelerated ions and a proton target. We derive an approximate factorized cross section for those SRC-driven p (A ,p‧N1N2) reactions. Our reaction model hinges on the factorization properties of SRC-driven A (e ,e‧N1N2) reactions for which strong indications are found in theory-experiment comparisons. In order to put our model to the test we compare its predictions with results of 12C (p ,p‧ pn) measurements conducted at Brookhaven National Laboratory (BNL) and find a fair agreement. The model can also reproduce characteristic features of SRC-driven two-nucleon knockout reactions, like back-to-back emission of the correlated nucleons. We study the asymmetry dependence of nuclear SRC by providing predictions for the ratio of proton-proton to proton-neutron knockout cross sections for the carbon isotopes 9-15C thereby covering neutron excess values (N - Z) / Z between -0.5 and +0.5.

Trinucleon cluster knockout from 6Li

International Nuclear Information System (INIS)

Connelly, J.P.; Berman, B.L.; Briscoe, W.J.; Dhuga, K.S.; Mokhtari, A.; Zubanov, D.; Blok, H.P.; Ent, R.; Mitchell, J.H.; Lapikas, L.

1998-01-01

The momentum-transfer dependence of the 3 H and 3 He knockout reactions from 6 Li via exclusive electron scattering has been measured, and the two reactions are compared. In the absence of two-step processes, the ratio of the fivefold cross sections for these mirror reactions should simply scale by the ratio of the 3 H and 3 He electron-scattering cross sections. A significant deviation from this simple expectation is seen at low momentum transfer. Possible explanations for this dramatic difference in cross sections for these mirror reactions are discussed. copyright 1998 The American Physical Society

Transthyretin knockout mice display decreased susceptibility to AMPA-induced neurodegeneration

DEFF Research Database (Denmark)

Nunes, Ana Filipa; Montero, Maria; Franquinho, Filipa

2009-01-01

Transthyretin (TTR) has been regarded as a neuroprotective protein given that TTR knockout (KO) mice display increased susceptibility for amyloid beta deposition and memory deficits during aging. In parallel, TTR KO mice have increased levels of neuropeptide Y (NPY), which promotes neuroprotectio...

Glutamate transporter type 3 knockout leads to decreased heart rate possibly via parasympathetic mechanism

OpenAIRE

Deng, Jiao; Li, Jiejie; Li, Liaoliao; Feng, Chenzhuo; Xiong, Lize; Zuo, Zhiyi

2013-01-01

Parasympathetic tone is a dominant neural regulator for basal heart rate. Glutamate transporters (EAAT) via their glutamate uptake functions regulate glutamate neurotransmission in the central nervous system. We showed that EAAT type 3 (EAAT3) knockout mice had a slower heart rate than wild-type mice when they were anesthetized. We design this study to determine whether non-anesthetized EAAT3 knockout mice have a slower heart rate and, if so, what may be the mechanism for this effect. Young a...

Mu-opioid receptor knockout mice show diminished food-anticipatory activity

NARCIS (Netherlands)

Kas, Martien J H; van den Bos, Ruud; Baars, Annemarie M; Lubbers, Marianne; Lesscher, Heidi M B; Hillebrand, Jacquelien J G; Schuller, Alwin G; Pintar, John E; Spruijt, Berry M

We have previously suggested that during or prior to activation of anticipatory behaviour to a coming reward, mu-opioid receptors are activated. To test this hypothesis schedule induced food-anticipatory activity in mu-opioid receptor knockout mice was measured using running wheels. We hypothesized

Pion-induced knock-out reactions

International Nuclear Information System (INIS)

Jain, B.K.; Phatak, S.C.

1977-01-01

A strong absorption model for pion-induced Knock-out reactions is proposed. The distortion of the in-coming and out-going pions has been included by (1) computing pion wave number in nuclear medium (dispersive effect) and (2) excluding the central region of the nucleus where the real pion-absorption is dominant (absorption effect). In order to study the dependence of the (π + π + p) reaction on the off-shell pion-nucleon t-matrix, different off-shell extrapolations are used. The magnitude of the cross-sections seems to be sensitive to the type of off-shell extrapolation; their shapes, however, are similar. The theoretical results are compared with experimental data. The agreement between the theoretical results for separable off-shell extrapolation and the data is good. (author)

A practical theoretical formalism for atomic multielectron processes: direct multiple ionization by a single auger decay or by impact of a single electron or photon

Science.gov (United States)

Liu, Pengfei; Zeng, Jiaolong; Yuan, Jianmin

2018-04-01

Multiple electron processes occur widely in atoms, molecules, clusters, and condensed matters when they are interacting with energetic particles or intense laser fields. Direct multielectron processes (DMEP) are the most complicated among the general multiple electron processes and are the most difficult to describe theoretically. In this work, a unified and accurate theoretical formalism is proposed on the DMEP of atoms including the multiple auger decay and multiple ionization by an impact of a single electron or a single photon based on the atomic collision theory described by a correlated many-body Green's function. Such a practical treatment is made possible by taking consideration of the different coherence features of the atoms (matter waves) in the initial and final states. We first explain how the coherence characteristics of the ejected continuum electrons is largely destructed, by taking the electron impact direct double ionization process as an example. The direct double ionization process is completely different from the single ionization where the complete interference can be maintained. The detailed expressions are obtained for the energy correlations among the continuum electrons and energy resolved differential and integral cross sections according to the separation of knock-out (KO) and shake-off (SO) mechanisms for the electron impact direct double ionization, direct double and triple auger decay, and double and triple photoionization (TPI) processes. Extension to higher order DMEP than triple ionization is straight forward by adding contributions of the following KO and SO processes. The approach is applied to investigate the electron impact double ionization processes of C+, N+, and O+, the direct double and triple auger decay of the K-shell excited states of C+ 1s2{s}22{p}2{}2D and {}2P, and the double and TPI of lithium. Comparisons with the experimental and other theoretical investigations wherever available in the literature show that our

An In Silico Knockout Model for Gastrointestinal Absorption Using a Systems Pharmacology Approach - Development and Application for Ketones.

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Vittal Shivva

Full Text Available Gastrointestinal absorption and disposition of ketones is complex. Recent work describing the pharmacokinetics (PK of d-β-hydroxybutyrate (BHB following oral ingestion of a ketone monoester ((R-3-hydroxybutyl (R-3-hydroxybutyrate found multiple input sites, nonlinear disposition and feedback on endogenous production. In the current work, a human systems pharmacology model for gastrointestinal absorption and subsequent disposition of small molecules (monocarboxylic acids with molecular weight < 200 Da was developed with an application to a ketone monoester. The systems model was developed by collating the information from the literature and knowledge gained from empirical population modelling of the clinical data. In silico knockout variants of this systems model were used to explore the mechanism of gastrointestinal absorption of ketones. The knockouts included active absorption across different regions in the gut and also a passive diffusion knockout, giving 10 gut knockouts in total. Exploration of knockout variants has suggested that there are at least three distinct regions in the gut that contribute to absorption of ketones. Passive diffusion predominates in the proximal gut and active processes contribute to the absorption of ketones in the distal gut. Low doses are predominantly absorbed from the proximal gut by passive diffusion whereas high doses are absorbed across all sites in the gut. This work has provided mechanistic insight into the absorption process of ketones, in the form of unique in silico knockouts that have potential for application with other therapeutics. Future studies on absorption process of ketones are suggested to substantiate findings in this study.

Histidine decarboxylase knockout mice, a genetic model of Tourette syndrome, show repetitive grooming after induced fear.

Science.gov (United States)

Xu, Meiyu; Li, Lina; Ohtsu, Hiroshi; Pittenger, Christopher

2015-05-19

Tics, such as are seen in Tourette syndrome (TS), are common and can cause profound morbidity, but they are poorly understood. Tics are potentiated by psychostimulants, stress, and sleep deprivation. Mutations in the gene histidine decarboxylase (Hdc) have been implicated as a rare genetic cause of TS, and Hdc knockout mice have been validated as a genetic model that recapitulates phenomenological and pathophysiological aspects of the disorder. Tic-like stereotypies in this model have not been observed at baseline but emerge after acute challenge with the psychostimulant d-amphetamine. We tested the ability of an acute stressor to stimulate stereotypies in this model, using tone fear conditioning. Hdc knockout mice acquired conditioned fear normally, as manifested by freezing during the presentation of a tone 48h after it had been paired with a shock. During the 30min following tone presentation, knockout mice showed increased grooming. Heterozygotes exhibited normal freezing and intermediate grooming. These data validate a new paradigm for the examination of tic-like stereotypies in animals without pharmacological challenge and enhance the face validity of the Hdc knockout mouse as a pathophysiologically grounded model of tic disorders. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Accurate Charge Densities from Powder Diffraction

DEFF Research Database (Denmark)

Bindzus, Niels; Wahlberg, Nanna; Becker, Jacob

Synchrotron powder X-ray diffraction has in recent years advanced to a level, where it has become realistic to probe extremely subtle electronic features. Compared to single-crystal diffraction, it may be superior for simple, high-symmetry crystals owing to negligible extinction effects and minimal...... peak overlap. Additionally, it offers the opportunity for collecting data on a single scale. For charge densities studies, the critical task is to recover accurate and bias-free structure factors from the diffraction pattern. This is the focal point of the present study, scrutinizing the performance...

A STAT-1 knockout mouse model for Machupo virus pathogenesis

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Shurtleff Amy C

2011-06-01

Full Text Available Abstract Background Machupo virus (MACV, a member of the Arenaviridae, causes Bolivian hemorrhagic fever, with ~20% lethality in humans. The pathogenesis of MACV infection is poorly understood, and there are no clinically proven treatments for disease. This is due, in part, to a paucity of small animal models for MACV infection in which to discover and explore candidate therapeutics. Methods Mice lacking signal transducer and activator of transcription 1 (STAT-1 were infected with MACV. Lethality, viral replication, metabolic changes, hematology, histopathology, and systemic cytokine expression were analyzed throughout the course of infection. Results We report here that STAT-1 knockout mice succumbed to MACV infection within 7-8 days, and presented some relevant clinical and histopathological manifestations of disease. Furthermore, the model was used to validate the efficacy of ribavirin in protection against infection. Conclusions The STAT-1 knockout mouse model can be a useful small animal model for drug testing and preliminary immunological analysis of lethal MACV infection.

The Expression of TALEN before Fertilization Provides a Rapid Knock-Out Phenotype in Xenopus laevis Founder Embryos.

Science.gov (United States)

Miyamoto, Kei; Suzuki, Ken-Ichi T; Suzuki, Miyuki; Sakane, Yuto; Sakuma, Tetsushi; Herberg, Sarah; Simeone, Angela; Simpson, David; Jullien, Jerome; Yamamoto, Takashi; Gurdon, J B

2015-01-01

Recent advances in genome editing using programmable nucleases have revolutionized gene targeting in various organisms. Successful gene knock-out has been shown in Xenopus, a widely used model organism, although a system enabling less mosaic knock-out in founder embryos (F0) needs to be explored in order to judge phenotypes in the F0 generation. Here, we injected modified highly active transcription activator-like effector nuclease (TALEN) mRNA to oocytes at the germinal vesicle (GV) stage, followed by in vitro maturation and intracytoplasmic sperm injection, to achieve a full knock-out in F0 embryos. Unlike conventional injection methods to fertilized embryos, the injection of TALEN mRNA into GV oocytes allows expression of nucleases before fertilization, enabling them to work from an earlier stage. Using this procedure, most of developed embryos showed full knock-out phenotypes of the pigmentation gene tyrosinase and/or embryonic lethal gene pax6 in the founder generation. In addition, our method permitted a large 1 kb deletion. Thus, we describe nearly complete gene knock-out phenotypes in Xenopus laevis F0 embryos. The presented method will help to accelerate the production of knock-out frogs since we can bypass an extra generation of about 1 year in Xenopus laevis. Meantime, our method provides a unique opportunity to rapidly test the developmental effects of disrupting those genes that do not permit growth to an adult able to reproduce. In addition, the protocol shown here is considerably less invasive than the previously used host transfer since our protocol does not require surgery. The experimental scheme presented is potentially applicable to other organisms such as mammals and fish to resolve common issues of mosaicism in founders.

Methamphetamine-induced changes in the striatal dopamine pathway in μ-opioid receptor knockout mice

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Park Sang Won

2011-11-01

Full Text Available Abstract Background Repeated exposure to methamphetamine (METH can cause not only neurotoxicity but also addiction. Behavioral sensitization is widely used as an animal model for the study of drug addiction. We previously reported that the μ-opioid receptor knockout mice were resistant to METH-induced behavioral sensitization but the mechanism is unknown. Methods The present study determined whether resistance of the μ-opioid receptor (μ-OR knockout mice to behavioral sensitization is due to differential expression of the stimulatory G protein α subunit (Gαs or regulators of G-protein signaling (RGS coupled to the dopamine D1 receptor. Mice received daily intraperitoneal injections of saline or METH (10 mg/kg for 7 consecutive days to induce sensitization. On day 11(following 4 abstinent days, mice were either given a test dose of METH (10 mg/kg for behavioral testing or sacrificed for neurochemical assays without additional METH treatment. Results METH challenge-induced stereotyped behaviors were significantly reduced in the μ-opioid receptor knockout mice when compared with those in wild-type mice. Neurochemical assays indicated that there is a decrease in dopamine D1 receptor ligand binding and an increase in the expression of RGS4 mRNA in the striatum of METH-treated μ-opioid receptor knockout mice but not of METH-treated wild-type mice. METH treatment had no effect on the expression of Gαs and RGS2 mRNA in the striatum of either strain of mice. Conclusions These results indicate that down-regulation of the expression of the dopamine D1 receptor and up-regulation of RGS4 mRNA expression in the striatum may contribute to the reduced response to METH-induced stereotypy behavior in μ-opioid receptor knockout mice. Our results highlight the interactions of the μ-opioid receptor system to METH-induced behavioral responses by influencing the expression of RGS of dopamine D1 receptors.

Proton-induced $\\alpha$-cluster knockout from $^{12}$C

CERN Document Server

Cowley, A A; Förtsch, S V; Buthelezi, E Z; Neveling, R; Smit, F D; Steyn, G F; van Zyl, J J

2010-01-01

Results of a study of the (p, p ) reaction on 12C with polarized incident protons of 100 MeV are reviewed. Experimental cross section and analyzing power distributions are compared with predictions of a distorted wave impulse approximation (DWIA) theory. The theory reproduces the data reasonably well, suggesting that a quasifree knockout mechanism dominates the reaction. Spectroscopic information extracted from the cross section data is in agreement with a shell model prediction.

Two-proton knockout on neutron-rich nuclei

International Nuclear Information System (INIS)

Bazin, D.; Brown, B.A.; Campbell, C.M.; Church, J.A.; Dinca, D.C.; Enders, J.; Gade, A.; Glasmacher, T.; Hansen, P.G.; Mueller, W.F.; Olliver, H.; Perry, B.C.; Sherrill, B.M.; Terry, J.R.; Tostevin, J.A.

2004-01-01

Two-proton knockout reactions on neutron-rich nuclei [Phys. Rev. Lett. 91 (2003) 012501] have been studied in inverse kinematics at intermediate energy. Strong evidence that the two-proton removal from a neutron-rich system proceeds as a direct reaction is presented, together with a preliminary theoretical discussion of the partial cross sections based on eikonal reaction theory and the many-body shell model. They show that this reaction can be used to characterize the wave functions of the projectiles and holds great promise for the study of neutron-rich nuclei

Proteome analysis of a hepatocyte-specific BIRC5 (survivin)-knockout mouse model during liver regeneration.

Science.gov (United States)

Bracht, Thilo; Hagemann, Sascha; Loscha, Marius; Megger, Dominik A; Padden, Juliet; Eisenacher, Martin; Kuhlmann, Katja; Meyer, Helmut E; Baba, Hideo A; Sitek, Barbara

2014-06-06

The Baculoviral IAP repeat-containing protein 5 (BIRC5), also known as inhibitor of apoptosis protein survivin, is a member of the chromosomal passenger complex and a key player in mitosis. To investigate the function of BIRC5 in liver regeneration, we analyzed a hepatocyte-specific BIRC5-knockout mouse model using a quantitative label-free proteomics approach. Here, we present the analyses of the proteome changes in hepatocyte-specific BIRC5-knockout mice compared to wildtype mice, as well as proteome changes during liver regeneration induced by partial hepatectomy in wildtype mice and mice lacking hepatic BIRC5, respectively. The BIRC5-knockout mice showed an extensive overexpression of proteins related to cellular maintenance, organization and protein synthesis. Key regulators of cell growth, transcription and translation MTOR and STAT1/STAT2 were found to be overexpressed. During liver regeneration proteome changes representing a response to the mitotic stimulus were detected in wildtype mice. Mainly proteins corresponding to proliferation, cell cycle and cytokinesis were up-regulated. The hepatocyte-specific BIRC5-knockout mice showed impaired liver regeneration, which had severe consequences on the proteome level. However, several proteins with function in mitosis were found to be up-regulated upon the proliferative stimulus. Our results show that the E3 ubiquitin-protein ligase UHRF1 is strongly up-regulated during liver regeneration independently of BIRC5.

Quantitative changes of main components of erythrocyte membranes which define architectonics of cells under pttg gene knockout

Directory of Open Access Journals (Sweden)

О. P. Kanyuka

2014-04-01

Full Text Available A pttg gene knockout affects the functional state of erythron in mice which could be associated with structural changes in the structure of erythrocyte membranes. The pttg gene knockout causes a significant modification of fatty acids composition of erythrocyte membrane lipids by reducing the content of palmitic acid and increasing of polyunsaturated fatty acids amount by 18%. Analyzing the erythrocyte surface architectonics of mice under pttg gene knockout, it was found that on the background of reduction of the functionally complete biconcave discs population one could observe an increase of the number of transformed cells at different degeneration stages. Researches have shown that in mice with a pttg gene knockout compared with a control group of animals cytoskeletal protein – β-spectrin was reduced by 17.03%. However, there is a reduction of membrane protein band 3 by 33.04%, simultaneously the content of anion transport protein band 4.5 increases by 35.2% and protein band 4.2 by 32.1%. The lectin blot analysis has helped to reveal changes in the structure of the carbohydrate determinants of ery­throcyte membrane glycoproteins under conditions of directed pttg gene inactivation, accompanied by changes in the type of communication, which joins the terminal residue in carbohydrate determinant of glycoproteins. Thus, a significant redistribution of protein and fatty acids contents in erythrocyte membranes that manifested in the increase of the deformed shape of red blood cells is observed under pttg gene knockout.

Less is More: unveiling the functional core of hematopoietic stem cells through knockout mice

Science.gov (United States)

Rossi, Lara; Lin, Kuanyin K.; Boles, Nathan C.; Yang, Liubin; King, Katherine Y.; Jeong, Mira; Mayle, Allison; Goodell, Margaret A.

2012-01-01

Summary Hematopoietic stem cells (HSCs) represent one of the first recognized somatic stem cells. As such, nearly 200 genes have been examined for roles in HSC function in knockout mice. In this review, we compile the majority of these reports to provide a broad overview of the functional modules revealed by these genetic analyses and highlight some key regulatory pathways involved, including cell cycle control, TGF-β signaling, Pten/AKT signaling, Wnt signaling, and cytokine signaling. Finally, we propose recommendations for characterization of HSC function in knockout mice to facilitate cross-study comparisons that would generate a more cohesive picture of HSC biology. In the field of design, the minimalist movement stripped down buildings and objects to their most basic features, a sentiment that architect Ludwig Mies van der Rohe summarized in his motto “less is more”. By depleting HSCs of specific genes, knockout studies transpose the minimalist approach into research biology, providing insights into the essential core of genetic features that is indispensable for a well-functioning hematopoietic system. PMID:22958929

Object recognition impairment in Fmr1 knockout mice is reversed by amphetamine: involvement of dopamine in the medial prefrontal cortex.

Science.gov (United States)

Ventura, R; Pascucci, T; Catania, M V; Musumeci, S A; Puglisi-Allegra, S

2004-09-01

Fragile X syndrome is an X-linked form of mental retardation including, among others, symptoms such as stereotypic behaviour, hyperactivity, hyperarousal, and cognitive deficits. We hypothesized that hyperactivity and/or compromised attentional, cognitive functions may lead to impaired performance in cognitive tasks in Fmr1 knockout mice, the most widely used animal model of fragile X syndrome, and suggested that psychostimulant treatment may improve performance by acting on one or both components. Since hyperactivity and cognitive functions have been suggested to depend on striatal and prefrontal cortex dopaminergic dysfunction, we assessed whether amphetamine produced beneficial, positive effects by acting on dopaminergic corticostriatal systems. Our results show that Fmr1 knockout mice are not able to discriminate between a familiar object and a novel one in the object recognition test, thus showing a clear-cut cognitive impairment that, to date, has been difficult to demonstrate in other cognitive tasks. Amphetamine improved performance of Fmr1 knockout mice, leading to enhanced ability to discriminate novel versus familiar objects, without significantly affecting locomotor activity. In agreement with behavioural data, amphetamine produced a greater increase in dopamine release in the prefrontal cortex of Fmr1 knockout compared with the wild-type mice, while a weak striatal dopaminergic response was observed in Fmr1 knockout mice. Our data support the view that the psychostimulant ameliorates performance in Fmr1 knockout mice by improving merely cognitive functions through its action on prefrontal cortical dopamine, irrespective of its action on motor hyperactivity. These results indicate that prefrontal cortical dopamine plays a major role in cognitive impairments characterizing Fmr1 knockout mice, thus pointing to an important aetiological factor in the fragile X syndrome.

Knock-Outs, Stick-Outs, Cut-Outs: Clipping Paths Separate Objects from Background.

Science.gov (United States)

Wilson, Bradley

1998-01-01

Outlines a six-step process that allows computer operators, using Photoshop software, to create "knock-outs" to precisely define the path that will serve to separate the object from the background. (SR)

Acute secondhand smoke-induced pulmonary inflammation is diminished in RAGE knockout mice.

Science.gov (United States)

Wood, Tyler T; Winden, Duane R; Marlor, Derek R; Wright, Alex J; Jones, Cameron M; Chavarria, Michael; Rogers, Geraldine D; Reynolds, Paul R

2014-11-15

The receptor for advanced glycation end-products (RAGE) has increasingly been demonstrated to be an important modulator of inflammation in cases of pulmonary disease. Published reports involving tobacco smoke exposure have demonstrated increased expression of RAGE, its participation in proinflammatory signaling, and its role in irreversible pulmonary remodeling. The current research evaluated the in vivo effects of short-term secondhand smoke (SHS) exposure in RAGE knockout and control mice compared with identical animals exposed to room air only. Quantitative PCR, immunoblotting, and immunohistochemistry revealed elevated RAGE expression in controls after 4 wk of SHS exposure and an anticipated absence of RAGE expression in RAGE knockout mice regardless of smoke exposure. Ras activation, NF-κB activity, and cytokine elaboration were assessed to characterize the molecular basis of SHS-induced inflammation in the mouse lung. Furthermore, bronchoalveolar lavage fluid was procured from RAGE knockout and control animals for the assessment of inflammatory cells and molecules. As a general theme, inflammation coincident with leukocyte recruitment was induced by SHS exposure and significantly influenced by the availability of RAGE. These data reveal captivating information suggesting a role for RAGE signaling in lungs exposed to SHS. However, ongoing research is still warranted to fully explain roles for RAGE and other receptors in cells coping with involuntary smoke exposure for prolonged periods of time. Copyright © 2014 the American Physiological Society.

A robust statistical estimation (RoSE) algorithm jointly recovers the 3D location and intensity of single molecules accurately and precisely

Science.gov (United States)

Mazidi, Hesam; Nehorai, Arye; Lew, Matthew D.

2018-02-01

In single-molecule (SM) super-resolution microscopy, the complexity of a biological structure, high molecular density, and a low signal-to-background ratio (SBR) may lead to imaging artifacts without a robust localization algorithm. Moreover, engineered point spread functions (PSFs) for 3D imaging pose difficulties due to their intricate features. We develop a Robust Statistical Estimation algorithm, called RoSE, that enables joint estimation of the 3D location and photon counts of SMs accurately and precisely using various PSFs under conditions of high molecular density and low SBR.

Universal statistics of the knockout tournament

Science.gov (United States)

Baek, Seung Ki; Yi, Il Gu; Park, Hye Jin; Kim, Beom Jun

2013-11-01

We study statistics of the knockout tournament, where only the winner of a fixture progresses to the next. We assign a real number called competitiveness to each contestant and find that the resulting distribution of prize money follows a power law with an exponent close to unity if the competitiveness is a stable quantity and a decisive factor to win a match. Otherwise, the distribution is found narrow. The existing observation of power law distributions in various kinds of real sports tournaments therefore suggests that the rules of those games are constructed in such a way that it is possible to understand the games in terms of the contestants' inherent characteristics of competitiveness.

CD1d knockout mice exhibit aggravated contact hypersensitivity responses due to reduced interleukin-10 production predominantly by regulatory B cells

DEFF Research Database (Denmark)

Fjelbye, Jonas; Antvorskov, Julie C; Buschard, Karsten

2015-01-01

.05) and peritoneal cavity (80.8% decrease; P challenge, which suggests an important regulatory and protective role of CD1d-dependent NKT cells in CHS in our model, at least in part via regulation of IL-10 producing B(regs) ....... knockout (CD1d KO) and wild-type (Wt) mice after contact allergen exposure. For induction of CHS, C57BL/6 CD1d KO mice (n = 6) and C57BL/6 Wt mice (n = 6) were sensitised with 1% (w/v) dinitrochlorobenzene (DNCB) or vehicle for three consecutive days and subsequently challenged with a single dose of 0...

K Basins floor sludge retrieval system knockout pot basket fuel burn accident

International Nuclear Information System (INIS)

HUNT, J.W.

1998-01-01

The K Basins Sludge Retrieval System Preliminary Hazard Analysis Report (HNF-2676) identified and categorized a series of potential accidents associated with K Basins Sludge Retrieval System design and operation. The fuel burn accident was of concern with respect to the potential release of contamination resulting from a runaway chemical reaction of the uranium fuel in a knockout pot basket suspended in the air. The unmitigated radiological dose to an offsite receptor from this fuel burn accident is calculated to be much less than the offsite risk evaluation guidelines for anticipated events. However, because of potential radiation exposure to the facility worker, this accident is precluded with a safety significant lifting device that will prevent the monorail hoist from lifting the knockout pot basket out of the K Basin water pool

ARGINASE ENZYMES IN ISOLATED AIRWAYS FROM NORMAL AND NITRIC OXIDE SYNTHASE 2-KNOCKOUT MICE EXPOSED TO OVALBUMIN

Science.gov (United States)

Bratt, Jennifer M.; Franzi, Lisa M.; Linderholm, Angela L.; Last, Michael S.; Kenyon, Nicholas J.; Last, Jerold A.

2009-01-01

Arginase has been suggested to compete with nitric oxide synthase (NOS) for their common substrate, L-arginine. To study the mechanisms underlying this interaction, we compared arginase expression in isolated airways and the consequences of inhibiting arginase activity in vivo with NO production, lung inflammation, and lung function in both C57BL/6 and NOS2 knockout mice undergoing ovalbumin-induced airway inflammation, a mouse model of asthma. Arginases I and II were measured by western blot in isolated airways from sensitized C57BL/6 mice exposed to ovalbumin aerosol. Physiological and biochemical responses---inflammation, lung compliance, airway hyperreactivity, exhaled NO concentration, arginine concentration--were compared with the responses of NOS2 knockout mice. NOS2 knockout mice had increased total cells in lung lavage, decreased lung compliance, and increased airway hyperreactivity. Both arginase I and arginase II were constitutively expressed in the airways of normal C57BL/6 mice. Arginase I was up-regulated approximately 8-fold in the airways of C57BL/6 mice exposed to ovalbumin. Expression of both arginase isoforms were significantly upregulated in NOS2 knockout mice exposed to ovalbumin, with about 40- and 4-fold increases in arginases I and II, respectively. Arginine concentration in isolated airways was not significantly different in any of the groups studied. Inhibition of arginase by systemic treatment of C57BL/6 mice with a competitive inhibitor, Nω-hydroxy-nor-L-arginine (nor-NOHA), significantly decreased the lung inflammatory response to ovalbumin in these animals. We conclude that NOS2 knockout mice are more sensitive to ovalbumin-induced airway inflammation and its sequelae than are C57BL/6 mice, as determined by increased total cells in lung lavage, decreased lung compliance, and increased airway hyperreactivity, and that these findings are strongly correlated with increased expression of both arginase isoforms in the airways of the NOS2

Knockout mouse model for Fxr2: a model for mental retardation

NARCIS (Netherlands)

C.J.M. Bontekoe (Carola); L. Kirkpatrick; C.E. Bakker (Cathy); A.T. Hoogeveen (Andre); R. McAninch; M. Merriweather; B.A. Oostra (Ben); N.C. Cheng (Ngan Ching); K.L. McIlwain; I.M. Nieuwenhuizen (Ingeborg); L.A. Yuva-Paylor; R. Paylor; A. Nellis; R. Willemsen (Rob); Z. Fang; D. Nelson

2002-01-01

textabstractFragile X syndrome is a common form of mental retardation caused by the absence of the FMR1 protein, FMRP. Fmr1 knockout mice exhibit a phenotype with some similarities to humans, such as macro-orchidism and behavioral abnormalities. Two homologs of FMRP have been

Pre-asymptotic behavior of single-particle overlap integrals of non-Borromean two-neutron halos

International Nuclear Information System (INIS)

Timofeyuk, N.K.; Tostevin, J.A.; Blokhintsev, L.D.

2003-01-01

For non-Borromean two-neutron halo nuclei, modifications to the behavior of single-particle overlap integrals will arise due to the correlations of the two interacting nucleons in the halo. An additional contribution to the overlap integral can be obtained using the Feynman diagram approach. This additional term is modeled using a simple local potential model. We show that these modifications may play a role in detailed interpretations of experimental results from single-nucleon knockout, transfer, and other reactions that probe the single-nucleon overlap functions

Pre-Equilibrium Cluster Emission with Pickup and Knockout

International Nuclear Information System (INIS)

Betak, E.

2005-01-01

We present a generalization of the Iwamoto-Harada-Bisplinghoff pre-equilibrium model of light cluster formation and emission, which is enhanced by allowing for possible admixtures of knockout for strongly coupled ejectiles, like α's. The model is able to attain the Weisskopf-Ewing formula for compound-nucleus decay at long-time limit; it keeps the philosophy of pre-equilibrium decay during the equilibration stage and it describes the initial phase of a reaction as direct process(es) expressed using the language of the exciton model

Efficient CRISPR/Cas9-based gene knockout in watermelon.

Science.gov (United States)

Tian, Shouwei; Jiang, Linjian; Gao, Qiang; Zhang, Jie; Zong, Mei; Zhang, Haiying; Ren, Yi; Guo, Shaogui; Gong, Guoyi; Liu, Fan; Xu, Yong

2017-03-01

CRISPR/Cas9 system can precisely edit genomic sequence and effectively create knockout mutations in T0 generation watermelon plants. Genome editing offers great advantage to reveal gene function and generate agronomically important mutations to crops. Recently, RNA-guided genome editing system using the type II clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (Cas9) has been applied to several plant species, achieving successful targeted mutagenesis. Here, we report the genome of watermelon, an important fruit crop, can also be precisely edited by CRISPR/Cas9 system. ClPDS, phytoene desaturase in watermelon, was selected as the target gene because its mutant bears evident albino phenotype. CRISPR/Cas9 system performed genome editing, such as insertions or deletions at the expected position, in transfected watermelon protoplast cells. More importantly, all transgenic watermelon plants harbored ClPDS mutations and showed clear or mosaic albino phenotype, indicating that CRISPR/Cas9 system has technically 100% of genome editing efficiency in transgenic watermelon lines. Furthermore, there were very likely no off-target mutations, indicated by examining regions that were highly homologous to sgRNA sequences. Our results show that CRISPR/Cas9 system is a powerful tool to effectively create knockout mutations in watermelon.

Hematopoiesis in 5-Fluorouracil-Treated Adenosine A(3) Receptor Knock-Out Mice

Czech Academy of Sciences Publication Activity Database

Hofer, Michal; Pospíšil, Milan; Dušek, L.; Hoferová, Zuzana; Komůrková, Denisa

2015-01-01

Roč. 64, č. 2 (2015), s. 255-262 ISSN 0862-8408 Institutional support: RVO:68081707 Keywords : Adenosine A(3) receptor knock-out mice * Hematopoiesis * 5-fluorouracil-induced hematotoxicity Subject RIV: BO - Biophysics Impact factor: 1.643, year: 2015

Sodium homeostasis is preserved in a global 11β-hydroxysteroid dehydrogenase type 1 knockout mouse model

DEFF Research Database (Denmark)

Christensen, Thorbjørn H; Bailey, Matthew A; Kenyon, Christopher J

2015-01-01

hypothesized that loss of renal 11βHSD1 would result in salt wasting and tested this in a knockout mouse model in which 11βHSD1 was deleted in all body tissues. In balance studies, 11βHSD1 deletion had no effect on water, sodium or potassium metabolism; transition to a low-sodium diet did not reveal...... that global deletion of 11βHSD1 in the mouse would give rise to a salt-wasting renal phenotype. What is the main finding and its importance? We subjected a mouse model of global 11βHSD1 deletion to studies of water and electrolyte balance, renal clearance, urinary steroid excretion, renin-angiotensin system...... a natriuretic phenotype. Renal clearance studies demonstrated identical haemodynamic parameters (arterial blood pressure, renal blood flow and glomerular filtration rate) in knockout and wild-type mice, but revealed an augmented kaliuretic response to thiazides in 11βHSD1 knockout animals. There was no effect...

Predicting the names of the best teams after the knock-out phase of a cricket series.

Science.gov (United States)

Lemmer, Hermanus Hofmeyr

2014-01-01

Cricket players' performances can best be judged after a large number of matches had been played. For test or one-day international (ODI) players, career data are normally used to calculate performance measures. These are normally good indicators of future performances, although various factors influence the performance of a player in a specific match. It is often necessary to judge players' performances based on a small number of scores, e.g. to identify the best players after a short series of matches. The challenge then is to use the best available criteria in order to assess performances as accurately and fairly as possible. In the present study the results of the knock-out phase of an International Cricket Council (ICC) World Cup ODI Series are used to predict the names of the best teams by means of a suitably formulated logistic regression model. Despite using very sparse data, the methods used are reasonably successful. It is also shown that if the same technique is applied to career ratings, very good results are obtained.

One at a time: counting single-nanoparticle/electrode collisions for accurate particle sizing by overcoming the instability of gold nanoparticles under electrolytic conditions

International Nuclear Information System (INIS)

Qiu, Danfeng; Wang, Song; Zheng, Yuanqin; Deng, Zhaoxiang

2013-01-01

In response to an increasing demand for understanding electrochemical processes on the nanometer scale, it now becomes possible to monitor electron transfer reactions at the single-nanoparticle level, namely particle collision electrochemistry. This technique has great potential in the development of research tools towards single-particle electrocatalysis and selective and multiplexed particle sizing. However, one existing problem that may discourage these applications is the relatively weak colloidal stability of nanoparticles in an electrolytic solution. Here we report on a facile but efficient way to achieve a good stability of gold nanoparticles in an acidic media so that ‘zero-aggregation’ collisions can be achieved at a carbon ultramicroelectrode. This allows us to obtain anodic dissolution currents from individual nanoparticles in a ‘one particle at a time’ manner, based on which accurate particle sizing with a resolution of 1–2 nm can be achieved. Our work strongly suggests that to maintain a well dispersed nanoparticle solution during a particle impact electrochemical experiment is critically important for accurate particle sizing, as well as other applications that require information to be extracted from individual nanoparticles (not their aggregates). (paper)

P-glycoprotein interaction with risperidone and 9-OH-risperidone studied in vitro, in knock-out mice and in drug-drug interaction experiments

DEFF Research Database (Denmark)

Ejsing, Thomas B.; Pedersen, Anne D.; Linnet, Kristian

2005-01-01

P-glycoprotein, risperidone, nortriptyline, cyclosporine A, drug-drug interaction, blood-brain barrier, knock-out mice......P-glycoprotein, risperidone, nortriptyline, cyclosporine A, drug-drug interaction, blood-brain barrier, knock-out mice...

Differential gene expression in the EphA4 knockout spinal cord and analysis of the inflammatory response following spinal cord injury.

Directory of Open Access Journals (Sweden)

Kathryn M Munro

Full Text Available Mice lacking the axon guidance molecule EphA4 have been shown to exhibit extensive axonal regeneration and functional recovery following spinal cord injury. To assess mechanisms by which EphA4 may modify the response to neural injury a microarray was performed on spinal cord tissue from mice with spinal cord injury and sham injured controls. RNA was purified from spinal cords of adult EphA4 knockout and wild-type mice four days following lumbar spinal cord hemisection or laminectomy only and was hybridised to Affymetrix All-Exon Array 1.0 GeneChips™. While subsequent analyses indicated that several pathways were altered in EphA4 knockout mice, of particular interest was the attenuated expression of a number of inflammatory genes, including Arginase 1, expression of which was lower in injured EphA4 knockout compared to wild-type mice. Immunohistological analyses of different cellular components of the immune response were then performed in injured EphA4 knockout and wildtype spinal cords. While numbers of infiltrating CD3+ T cells were low in the hemisection model, a robust CD11b+ macrophage/microglial response was observed post-injury. There was no difference in the overall number or spread of macrophages/activated microglia in injured EphA4 knockout compared to wild-type spinal cords at 2, 4 or 14 days post-injury, however a lower proportion of Arginase-1 immunoreactive macrophages/activated microglia was observed in EphA4 knockout spinal cords at 4 days post-injury. Subtle alterations in the neuroinflammatory response in injured EphA4 knockout spinal cords may contribute to the regeneration and recovery observed in these mice following injury.

Markerless gene knockout and integration to express heterologous biosynthetic gene clusters in Pseudomonas putida

DEFF Research Database (Denmark)

Choi, Kyeong Rok; Cho, Jae Sung; Cho, In Jin

2018-01-01

Pseudomonas putida has gained much interest among metabolic engineers as a workhorse for producing valuable natural products. While a few gene knockout tools for P. putida have been reported, integration of heterologous genes into the chromosome of P. putida, an essential strategy to develop stable...... plasmid curing systems, generating final strains free of antibiotic markers and plasmids. This markerless recombineering system for efficient gene knockout and integration will expedite metabolic engineering of P. putida, a bacterial host strain of increasing academic and industrial interest....

A Knowledge-Based System for Display and Prediction of O-Glycosylation Network Behaviour in Response to Enzyme Knockouts.

Directory of Open Access Journals (Sweden)

Andrew G McDonald

2016-04-01

Full Text Available O-linked glycosylation is an important post-translational modification of mucin-type protein, changes to which are important biomarkers of cancer. For this study of the enzymes of O-glycosylation, we developed a shorthand notation for representing GalNAc-linked oligosaccharides, a method for their graphical interpretation, and a pattern-matching algorithm that generates networks of enzyme-catalysed reactions. Software for generating glycans from the enzyme activities is presented, and is also available online. The degree distributions of the resulting enzyme-reaction networks were found to be Poisson in nature. Simple graph-theoretic measures were used to characterise the resulting reaction networks. From a study of in-silico single-enzyme knockouts of each of 25 enzymes known to be involved in mucin O-glycan biosynthesis, six of them, β-1,4-galactosyltransferase (β4Gal-T4, four glycosyltransferases and one sulfotransferase, play the dominant role in determining O-glycan heterogeneity. In the absence of β4Gal-T4, all Lewis X, sialyl-Lewis X, Lewis Y and Sda/Cad glycoforms were eliminated, in contrast to knockouts of the N-acetylglucosaminyltransferases, which did not affect the relative abundances of O-glycans expressing these epitopes. A set of 244 experimentally determined mucin-type O-glycans obtained from the literature was used to validate the method, which was able to predict up to 98% of the most common structures obtained from human and engineered CHO cell glycoforms.

Skeletal muscle-specific HMG-CoA reductase knockout mice exhibit rhabdomyolysis: A model for statin-induced myopathy.

Science.gov (United States)

Osaki, Yoshinori; Nakagawa, Yoshimi; Miyahara, Shoko; Iwasaki, Hitoshi; Ishii, Akiko; Matsuzaka, Takashi; Kobayashi, Kazuto; Yatoh, Shigeru; Takahashi, Akimitsu; Yahagi, Naoya; Suzuki, Hiroaki; Sone, Hirohito; Ohashi, Ken; Ishibashi, Shun; Yamada, Nobuhiro; Shimano, Hitoshi

2015-10-23

HMG-CoA reductase (HMGCR) catalyzes the conversion of HMG-CoA to mevalonic acid (MVA); this is the rate-limiting enzyme of the mevalonate pathway that synthesizes cholesterol. Statins, HMGCR inhibitors, are widely used as cholesterol-reducing drugs. However, statin-induced myopathy is the most adverse side effect of statins. To eludicate the mechanisms underlying statin the myotoxicity and HMGCR function in the skeletal muscle, we developed the skeletal muscle-specific HMGCR knockout mice. Knockout mice exhibited postnatal myopathy with elevated serum creatine kinase levels and necrosis. Myopathy in knockout mice was completely rescued by the oral administration of MVA. These results suggest that skeletal muscle toxicity caused by statins is dependent on the deficiencies of HMGCR enzyme activity and downstream metabolites of the mevalonate pathway in skeletal muscles rather than the liver or other organs. Copyright © 2015 Elsevier Inc. All rights reserved.

Generation of a Nrf2 homozygous knockout human embryonic stem cell line using CRISPR/Cas9

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So-Jung Kim

2017-03-01

Full Text Available Nuclear factor erythroid 2-related factor 2 (NFE2L2 or Nrf2 is a well-known transcription factor that regulates the expression of a large number of anti-oxidant genes in mammalian cells (J.H. Kim et al., 2014. Here, we generated a homozygous Nrf2 knockout human embryonic stem cell (hESC line, H9Nrf2KO-A13, using the CRISPR/Cas9 genome editing method. The Nrf2 homozygous knockout H9 cell line maintains pluripotency, differentiation potential into three germ layers, and a normal karyotype.

Haloperidol inhibits the development of atherosclerotic lesions in LDL receptor knockout mice.

Science.gov (United States)

van der Sluis, Ronald J; Nahon, Joya E; Reuwer, Anne Q; Van Eck, Miranda; Hoekstra, Menno

2015-05-01

Antipsychotic drugs have been shown to modulate the expression of ATP-binding cassette transporter A1 (ABCA1), a key factor in the anti-atherogenic reverse cholesterol transport process, in vitro. Here we evaluated the potential of the typical antipsychotic drug haloperidol to modulate the cholesterol efflux function of macrophages in vitro and their susceptibility to atherosclerosis in vivo. Thioglycollate-elicited peritoneal macrophages were used for in vitro studies. Hyperlipidaemic low-density lipoprotein (LDL) receptor knockout mice were implanted with a haloperidol-containing pellet and subsequently fed a Western-type diet for 5 weeks to induce the development of atherosclerotic lesions in vivo. Haloperidol induced a 54% decrease in the mRNA expression of ABCA1 in peritoneal macrophages. This coincided with a 30% decrease in the capacity of macrophages to efflux cholesterol to apolipoprotein A1. Haloperidol treatment stimulated the expression of ABCA1 (+51%) and other genes involved in reverse cholesterol transport, that is, CYP7A1 (+98%) in livers of LDL receptor knockout mice. No change in splenic ABCA1 expression was noted. However, the average size of the atherosclerotic size was significantly smaller (-31%) in the context of a mildly more atherogenic metabolic phenotype upon haloperidol treatment. More importantly, haloperidol markedly lowered MCP-1 expression (-70%) and secretion (-28%) by peritoneal macrophages. Haloperidol treatment lowered the susceptibility of hyperlipidaemic LDL receptor knockout mice to develop atherosclerotic lesions. Our findings suggest that the beneficial effect of haloperidol on atherosclerosis susceptibility can be attributed to its ability to inhibit macrophage chemotaxis. © 2015 The British Pharmacological Society.

Dopamine D2 receptor function is compromised in the brain of the methionine sulfoxide reductase A knockout mouse

OpenAIRE

Oien, Derek B.; Ortiz, Andrea N.; Rittel, Alexander G.; Dobrowsky, Rick T.; Johnson, Michael A.; Levant, Beth; Fowler, Stephen C.; Moskovitz, Jackob

2010-01-01

Previous research suggests that brain oxidative stress and altered rodent locomotor behavior are linked. We observed bio-behavioral changes in methionine sulfoxide reductase A knockout mice associated with abnormal dopamine signaling. Compromised ability of these knockout mice to reduce methionine sulfoxide enhances accumulation of sulfoxides in proteins. We examined the dopamine D2-receptor function and expression, which has an atypical arrangement and quantity of methionine residues. Indeed...

Improving the efficiency of CHO cell line generation using glutamine synthetase gene knockout cells.

Science.gov (United States)

Fan, Lianchun; Kadura, Ibrahim; Krebs, Lara E; Hatfield, Christopher C; Shaw, Margaret M; Frye, Christopher C

2012-04-01

Although Chinese hamster ovary (CHO) cells, with their unique characteristics, have become a major workhorse for the manufacture of therapeutic recombinant proteins, one of the major challenges in CHO cell line generation (CLG) is how to efficiently identify those rare, high-producing clones among a large population of low- and non-productive clones. It is not unusual that several hundred individual clones need to be screened for the identification of a commercial clonal cell line with acceptable productivity and growth profile making the cell line appropriate for commercial application. This inefficiency makes the process of CLG both time consuming and laborious. Currently, there are two main CHO expression systems, dihydrofolate reductase (DHFR)-based methotrexate (MTX) selection and glutamine synthetase (GS)-based methionine sulfoximine (MSX) selection, that have been in wide industrial use. Since selection of recombinant cell lines in the GS-CHO system is based on the balance between the expression of the GS gene introduced by the expression plasmid and the addition of the GS inhibitor, L-MSX, the expression of GS from the endogenous GS gene in parental CHOK1SV cells will likely interfere with the selection process. To study endogenous GS expression's potential impact on selection efficiency, GS-knockout CHOK1SV cell lines were generated using the zinc finger nuclease (ZFN) technology designed to specifically target the endogenous CHO GS gene. The high efficiency (∼2%) of bi-allelic modification on the CHO GS gene supports the unique advantages of the ZFN technology, especially in CHO cells. GS enzyme function disruption was confirmed by the observation of glutamine-dependent growth of all GS-knockout cell lines. Full evaluation of the GS-knockout cell lines in a standard industrial cell culture process was performed. Bulk culture productivity improved two- to three-fold through the use of GS-knockout cells as parent cells. The selection stringency was

Defects in ultrasonic vocalization of cadherin-6 knockout mice.

Directory of Open Access Journals (Sweden)

Ryoko Nakagawa

Full Text Available BACKGROUND: Although some molecules have been identified as responsible for human language disorders, there is still little information about what molecular mechanisms establish the faculty of human language. Since mice, like songbirds, produce complex ultrasonic vocalizations for intraspecific communication in several social contexts, they can be good mammalian models for studying the molecular basis of human language. Having found that cadherins are involved in the vocal development of the Bengalese finch, a songbird, we expected cadherins to also be involved in mouse vocalizations. METHODOLOGY/PRINCIPAL FINDINGS: To examine whether similar molecular mechanisms underlie the vocalizations of songbirds and mammals, we categorized behavioral deficits including vocalization in cadherin-6 knockout mice. Comparing the ultrasonic vocalizations of cadherin-6 knockout mice with those of wild-type controls, we found that the peak frequency and variations of syllables were differed between the mutant and wild-type mice in both pup-isolation and adult-courtship contexts. Vocalizations during male-male aggression behavior, in contrast, did not differ between mutant and wild-type mice. Open-field tests revealed differences in locomotors activity in both heterozygote and homozygote animals and no difference in anxiety behavior. CONCLUSIONS/SIGNIFICANCE: Our results suggest that cadherin-6 plays essential roles in locomotor activity and ultrasonic vocalization. These findings also support the idea that different species share some of the molecular mechanisms underlying vocal behavior.

Relevant feature set estimation with a knock-out strategy and random forests

DEFF Research Database (Denmark)

Ganz, Melanie; Greve, Douglas N; Fischl, Bruce

2015-01-01

unintuitive and difficult to determine. In this article, we propose a novel MVPA method for group analysis of high-dimensional data that overcomes the drawbacks of the current techniques. Our approach explicitly aims to identify all relevant variations using a "knock-out" strategy and the Random Forest...

Claudin-2 knockout by TALEN-mediated gene targeting in MDCK cells: claudin-2 independently determines the leaky property of tight junctions in MDCK cells.

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Shinsaku Tokuda

Full Text Available Tight junctions (TJs regulate the movements of substances through the paracellular pathway, and claudins are major determinants of TJ permeability. Claudin-2 forms high conductive cation pores in TJs. The suppression of claudin-2 expression by RNA interference in Madin-Darby canine kidney (MDCK II cells (a low-resistance strain of MDCK cells was shown to induce a three-fold increase in transepithelial electrical resistance (TER, which, however, was still lower than in high-resistance strains of MDCK cells. Because RNA interference-mediated knockdown is not complete and only reduces gene function, we considered the possibility that the remaining claudin-2 expression in the knockdown study caused the lower TER in claudin-2 knockdown cells. Therefore, we investigated the effects of claudin-2 knockout in MDCK II cells by establishing claudin-2 knockout clones using transcription activator-like effector nucleases (TALENs, a recently developed genome editing method for gene knockout. Surprisingly, claudin-2 knockout increased TER by more than 50-fold in MDCK II cells, and TER values in these cells (3000-4000 Ω·cm2 were comparable to those in the high-resistance strains of MDCK cells. Claudin-2 re-expression restored the TER of claudin-2 knockout cells dependent upon claudin-2 protein levels. In addition, we investigated the localization of claudin-1, -2, -3, -4, and -7 at TJs between control MDCK cells and their respective knockout cells using their TALENs. Claudin-2 and -7 were less efficiently localized at TJs between control and their knockout cells. Our results indicate that claudin-2 independently determines the 'leaky' property of TJs in MDCK II cells and suggest the importance of knockout analysis in cultured cells.

Generation of ERα-floxed and knockout mice using the Cre/LoxP system

International Nuclear Information System (INIS)

Antonson, P.; Omoto, Y.; Humire, P.; Gustafsson, J.-Å.

2012-01-01

Highlights: ► ERα floxed and knockout mice were generated. ► Disruption of the ERα gene results in sterility in both male and female mice. ► ERα −/− mice have ovaries with hemorrhagic follicles and hypoplastic uterus. ► Female ERα −/− mice develop obesity. -- Abstract: Estrogen receptor alpha (ERα) is a nuclear receptor that regulates a range of physiological processes in response to estrogens. In order to study its biological role, we generated a floxed ERα mouse line that can be used to knock out ERα in selected tissues by using the Cre/LoxP system. In this study, we established a new ERα knockout mouse line by crossing the floxed ERα mice with Cre deleter mice. Here we show that genetic disruption of the ERα gene in all tissues results in sterility in both male and female mice. Histological examination of uterus and ovaries revealed a dramatically atrophic uterus and hemorrhagic cysts in the ovary. These results suggest that infertility in female mice is the result of functional defects of the reproductive tract. Moreover, female knockout mice are hyperglycemic, develop obesity and at the age of 4 months the body weight of these mice was more than 20% higher compared to wild type littermates and this difference increased over time. Our results demonstrate that ERα is necessary for reproductive tract development and has important functions as a regulator of metabolism in females.

Preaxial Polydactyly in Sost/Sostdc1 Double Knockouts

Energy Technology Data Exchange (ETDEWEB)

Yee, C M; Collette, N M; Loots, G G

2011-07-29

In the United States, {approx}5% are born with congenital birth defects due to abnormal function of cellular processes and interactions. Sclerosteosis, a rare autosomal recessive disease, causes hyperostosis of the axial and appendicular skeleton, and patients present radial deviation, digit syndactyly, nail dysplasia, and overall high bone mineral density. Sclerosteosis is due to a loss of function of sclerostin (Sost). Sost is a Wnt (abbrev.) antagonist; when mutated, nonfunctional Sost results in hyperactive osteoblast activity which leads to abnormal high bone mass. Previous studies have shown that Sost overexpression in transgenic mice causes reduced bone mineral density and a variety of limb phenotypes ranging from lost, fused, and split phalanges. Consistent with clinical manifestations of Sclerosteosis, Sost knockout mice exhibit increased generalized bone mineral density and syndactyly of the digits. Sostdc1 is a paralog of Sost that has also been described as an antagonist of Wnt signaling, in developing tooth buds. Unlike Sost knockouts, Sostdc1 null mice do not display any limb abnormalities. To determine if Sost and Sostdc1 have redundant functions during limb patterning, we examined Sost; Sostdc1 mice determined that they exhibit a novel preaxial polydactyly phenotype with a low penetrance. LacZ staining, skeletal preparations, and in situ hybridization experiments were used to help characterize this novel phenotype and understand how this phenotype develops. We find Sost and Sostdc1 to have complementary expression patterns during limb development, and the loss of their expression alters the transcription of several key limb regulators, such as Fgf8, Shh and Grem.

Zinc finger nuclease mediated knockout of ADP-dependent glucokinase in cancer cell lines: effects on cell survival and mitochondrial oxidative metabolism.

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Susan Richter

Full Text Available Zinc finger nucleases (ZFN are powerful tools for editing genes in cells. Here we use ZFNs to interrogate the biological function of ADPGK, which encodes an ADP-dependent glucokinase (ADPGK, in human tumour cell lines. The hypothesis we tested is that ADPGK utilises ADP to phosphorylate glucose under conditions where ATP becomes limiting, such as hypoxia. We characterised two ZFN knockout clones in each of two lines (H460 and HCT116. All four clones had frameshift mutations in all alleles at the target site in exon 1 of ADPGK, and were ADPGK-null by immunoblotting. ADPGK knockout had little or no effect on cell proliferation, but compromised the ability of H460 cells to survive siRNA silencing of hexokinase-2 under oxic conditions, with clonogenic survival falling from 21±3% for the parental line to 6.4±0.8% (p = 0.002 and 4.3±0.8% (p = 0.001 for the two knockouts. A similar increased sensitivity to clonogenic cell killing was observed under anoxia. No such changes were found when ADPGK was knocked out in HCT116 cells, for which the parental line was less sensitive than H460 to anoxia and to hexokinase-2 silencing. While knockout of ADPGK in HCT116 cells caused few changes in global gene expression, knockout of ADPGK in H460 cells caused notable up-regulation of mRNAs encoding cell adhesion proteins. Surprisingly, we could discern no consistent effect on glycolysis as measured by glucose consumption or lactate formation under anoxia, or extracellular acidification rate (Seahorse XF analyser under oxic conditions in a variety of media. However, oxygen consumption rates were generally lower in the ADPGK knockouts, in some cases markedly so. Collectively, the results demonstrate that ADPGK can contribute to tumour cell survival under conditions of high glycolytic dependence, but the phenotype resulting from knockout of ADPGK is cell line dependent and appears to be unrelated to priming of glycolysis in these lines.

Zinc finger nuclease mediated knockout of ADP-dependent glucokinase in cancer cell lines: effects on cell survival and mitochondrial oxidative metabolism.

Science.gov (United States)

Richter, Susan; Morrison, Shona; Connor, Tim; Su, Jiechuang; Print, Cristin G; Ronimus, Ron S; McGee, Sean L; Wilson, William R

2013-01-01

Zinc finger nucleases (ZFN) are powerful tools for editing genes in cells. Here we use ZFNs to interrogate the biological function of ADPGK, which encodes an ADP-dependent glucokinase (ADPGK), in human tumour cell lines. The hypothesis we tested is that ADPGK utilises ADP to phosphorylate glucose under conditions where ATP becomes limiting, such as hypoxia. We characterised two ZFN knockout clones in each of two lines (H460 and HCT116). All four clones had frameshift mutations in all alleles at the target site in exon 1 of ADPGK, and were ADPGK-null by immunoblotting. ADPGK knockout had little or no effect on cell proliferation, but compromised the ability of H460 cells to survive siRNA silencing of hexokinase-2 under oxic conditions, with clonogenic survival falling from 21±3% for the parental line to 6.4±0.8% (p = 0.002) and 4.3±0.8% (p = 0.001) for the two knockouts. A similar increased sensitivity to clonogenic cell killing was observed under anoxia. No such changes were found when ADPGK was knocked out in HCT116 cells, for which the parental line was less sensitive than H460 to anoxia and to hexokinase-2 silencing. While knockout of ADPGK in HCT116 cells caused few changes in global gene expression, knockout of ADPGK in H460 cells caused notable up-regulation of mRNAs encoding cell adhesion proteins. Surprisingly, we could discern no consistent effect on glycolysis as measured by glucose consumption or lactate formation under anoxia, or extracellular acidification rate (Seahorse XF analyser) under oxic conditions in a variety of media. However, oxygen consumption rates were generally lower in the ADPGK knockouts, in some cases markedly so. Collectively, the results demonstrate that ADPGK can contribute to tumour cell survival under conditions of high glycolytic dependence, but the phenotype resulting from knockout of ADPGK is cell line dependent and appears to be unrelated to priming of glycolysis in these lines.

Dopamine transporter and vesicular monoamine transporter knockout mice : implications for Parkinson's disease.

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Miller, G W; Wang, Y M; Gainetdinov, R R; Caron, M G

2001-01-01

One of the most valuable methods for understanding the function of a particular protein is the generation of animals that have had the gene encoding for the protein of interest disrupted, commonly known as a "quo;knockout"quo; or null mutant. By incorporating a sequence of DNA (typically encoding antibiotic resistance to aid in the selection of the mutant gene) into embryonic stem cells by homologous recombination, the normal transcription of the gene is effectively blocked (Fig. 1). Since a particular protein is encoded by two copies of a gene, it is necessary to have the gene on both alleles "quo;knocked out."quo; This is performed by cross-breeding animals with one affected allele (heterozygote) to generate offspring that have inherited two mutant alleles (homozygote). This procedure has been used to generate animals lacking either the plasma membrane dopamine transporter (DAT; Fig. 2) or the vesicular monoamine transporter (VMAT2; Fig. 3). Both DAT and VMAT2 are essential for dopamine homeostasis and are thought to participate in the pathogenesis of Parkinson's disease (1-5). Fig. 1. Maps of the targeting vector and the mock construct. The mouse genomic fragment (clone 11) was isolated from a Stratagene 129 SvJ library by standard colony hybridization using a PCR probe from the 5' end of rat cDNA. The restriction site abbreviations are as follows: H, HindIII; N, NotI; Sc, SacI; Sn, SnaI; X, XbaI; and Xh, XhoI. The region between HindIII and SnaI on clone 11 containing the coding sequence from transmembrane domains 3 and 4 of VMAT2 was deleted and replaced with PGK-neo. The 3' fragment of clone 11 was reserved as an external probe for Southern analysis. To facilitate PCR screening of embryonic stem cell clones, a mock construct containing the SnaI/XbaI fragment and part of the Neo cassette was generated as a positive control. pPNT and pGEM4Z were used to construct knockout and mock vectors, respectively. (Reproduced with permission from ref. 1). Fig. 2. DAT and

Myostatin gene knockout mediated by Cas9-D10A nickase in chicken DF1 cells without off-target effect

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Jeong Hyo Lee

2017-05-01

Full Text Available Objective Based on rapid advancement of genetic modification techniques, genomic editing is expected to become the most efficient tool for improvement of economic traits in livestock as well as poultry. In this study, we examined and verified the nickase of mutated CRISPR-associated protein 9 (Cas9 to modulate the specific target gene in chicken DF1 cells. Methods Chicken myostatin which inhibits muscle cell growth and differentiation during myogenesis was targeted to be deleted and mutated by the Cas9-D10A nickase. After co-transfection of the nickase expression vector with green fluorescent gene (GFP gene and targeted multiplex guide RNAs (gRNAs, the GFP-positive cells were sorted out by fluorescence-activated cell sorting procedure. Results Through the genotyping analysis of the knockout cells, the mutant induction efficiency was 100% in the targeted site. Number of the deleted nucleotides ranged from 2 to 39 nucleotide deletion. There was no phenotypic difference between regular cells and knockout cells. However, myostatin protein was not apparently detected in the knockout cells by Western blotting. Additionally, six off-target sites were predicted and analyzed but any non-specific mutation in the off-target sites was not observed. Conclusion The knockout technical platform with the nickase and multiplex gRNAs can be efficiently and stablely applied to functional genomics study in poultry and finally adapted to generate the knockout poultry for agribio industry.

GPR39 (zinc receptor) knockout mice exhibit depression-like behavior and CREB/BDNF down-regulation in the hippocampus.

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Młyniec, Katarzyna; Budziszewska, Bogusława; Holst, Birgitte; Ostachowicz, Beata; Nowak, Gabriel

2014-10-31

Zinc may act as a neurotransmitter in the central nervous system by activation of the GPR39 metabotropic receptors. In the present study, we investigated whether GPR39 knockout would cause depressive-like and/or anxiety-like behavior, as measured by the forced swim test, tail suspension test, and light/dark test. We also investigated whether lack of GPR39 would change levels of cAMP response element-binding protein (CREB),brain-derived neurotrophic factor (BDNF) and tropomyosin related kinase B (TrkB) protein in the hippocampus and frontal cortex of GPR39 knockout mice subjected to the forced swim test, as measured by Western-blot analysis. In this study, GPR39 knockout mice showed an increased immobility time in both the forced swim test and tail suspension test, indicating depressive-like behavior and displayed anxiety-like phenotype. GPR39 knockout mice had lower CREB and BDNF levels in the hippocampus, but not in the frontal cortex, which indicates region specificity for the impaired CREB/BDNF pathway (which is important in antidepressant response) in the absence of GPR39. There were no changes in TrkB protein in either structure. In the present study, we also investigated activity in the hypothalamus-pituitary-adrenal axis under both zinc- and GPR39-deficient conditions. Zinc-deficient mice had higher serum corticosterone levels and lower glucocorticoid receptor levels in the hippocampus and frontal cortex. There were no changes in the GPR39 knockout mice in comparison with the wild-type control mice, which does not support a role of GPR39 in hypothalamus-pituitary-adrenal axis regulation. The results of this study indicate the involvement of the GPR39 Zn(2+)-sensing receptor in the pathophysiology of depression with component of anxiety. © The Author 2015. Published by Oxford University Press on behalf of CINP.

Gene knockout of the KCNJ8-encoded Kir6.1 K(ATP) channel imparts fatal susceptibility to endotoxemia.

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Kane, Garvan C; Lam, Chen-Fuh; O'Cochlain, Fearghas; Hodgson, Denice M; Reyes, Santiago; Liu, Xiao-Ke; Miki, Takashi; Seino, Susumu; Katusic, Zvonimir S; Terzic, Andre

2006-11-01

Sepsis, the systemic inflammatory response to infection, imposes a high demand for bodily adaptation, with the cardiovascular response a key determinant of outcome. The homeostatic elements that secure cardiac tolerance in the setting of the sepsis syndrome are poorly understood. Here, in a model of acute septic shock induced by endotoxin challenge with Escherichia coli lipopolysaccharide (LPS), knockout of the KCNJ8 gene encoding the vascular Kir6.1 K(ATP) channel pore predisposed to an early and profound survival disadvantage. The exaggerated susceptibility provoked by disruption of this stress-responsive sensor of cellular metabolism was linked to progressive deterioration in cardiac activity, ischemic myocardial damage, and contractile dysfunction. Deletion of KCNJ8 blunted the responsiveness of coronary vessels to cytokine- or metabolic-mediated vasodilation necessary to support myocardial perfusion in the wild-type (WT), creating a deficit in adaptive response in the Kir6.1 knockout. Application of a K(ATP) channel opener drug improved survival in the endotoxic WT but had no effect in the Kir6.1 knockout. Restoration of the dilatory capacity of coronary vessels was required to rescue the Kir6.1 knockout phenotype and reverse survival disadvantage in lethal endotoxemia. Thus, the Kir6.1-containing K(ATP) channel, by coupling vasoreactivity with metabolic demand, provides a vital feedback element for cardiovascular tolerance in endotoxic shock.

Generation of knockout rabbits using transcription activator-like effector nucleases

OpenAIRE

Wang, Yu; Fan, Nana; Song, Jun; Zhong, Juan; Guo, Xiaogang; Tian, Weihua; Zhang, Quanjun; Cui, Fenggong; Li, Li; Newsome, Philip N; Frampton, Jon; Esteban, Miguel A; Lai, Liangxue

2014-01-01

Zinc-finger nucleases and transcription activator-like effector nucleases are novel gene-editing platforms contributing to redefine the boundaries of modern biological research. They are composed of a non-specific cleavage domain and a tailor made DNA-binding module, which enables a broad range of genetic modifications by inducing efficient DNA double-strand breaks at desired loci. Among other remarkable uses, these nucleases have been employed to produce gene knockouts in mid-size and large ...

Serotonin Transporter Knockout Rats Show Improved Strategy Set-Shifting and Reduced Latent Inhibition

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Nonkes, Lourens J. P.; van de Vondervoort, Ilse I. G. M.; de Leeuw, Mark J. C.; Wijlaars, Linda P.; Maes, Joseph H. R.; Homberg, Judith R.

2012-01-01

Behavioral flexibility is a cognitive process depending on prefrontal areas allowing adaptive responses to environmental changes. Serotonin transporter knockout (5-HTT[superscript -/-]) rodents show improved reversal learning in addition to orbitofrontal cortex changes. Another form of behavioral flexibility, extradimensional strategy set-shifting…

Systematic screening for skin, hair, and nail abnormalities in a large-scale knockout mouse program.

Directory of Open Access Journals (Sweden)

John P Sundberg

Full Text Available The International Knockout Mouse Consortium was formed in 2007 to inactivate ("knockout" all protein-coding genes in the mouse genome in embryonic stem cells. Production and characterization of these mice, now underway, has generated and phenotyped 3,100 strains with knockout alleles. Skin and adnexa diseases are best defined at the gross clinical level and by histopathology. Representative retired breeders had skin collected from the back, abdomen, eyelids, muzzle, ears, tail, and lower limbs including the nails. To date, 169 novel mutant lines were reviewed and of these, only one was found to have a relatively minor sebaceous gland abnormality associated with follicular dystrophy. The B6N(Cg-Far2tm2b(KOMPWtsi/2J strain, had lesions affecting sebaceous glands with what appeared to be a secondary follicular dystrophy. A second line, B6N(Cg-Ppp1r9btm1.1(KOMPVlcg/J, had follicular dystrophy limited to many but not all mystacial vibrissae in heterozygous but not homozygous mutant mice, suggesting that this was a nonspecific background lesion. We discuss potential reasons for the low frequency of skin and adnexal phenotypes in mice from this project in comparison to those seen in human Mendelian diseases, and suggest alternative approaches to identification of human disease-relevant models.

Accurate prediction of complex free surface flow around a high speed craft using a single-phase level set method

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Broglia, Riccardo; Durante, Danilo

2017-11-01

This paper focuses on the analysis of a challenging free surface flow problem involving a surface vessel moving at high speeds, or planing. The investigation is performed using a general purpose high Reynolds free surface solver developed at CNR-INSEAN. The methodology is based on a second order finite volume discretization of the unsteady Reynolds-averaged Navier-Stokes equations (Di Mascio et al. in A second order Godunov—type scheme for naval hydrodynamics, Kluwer Academic/Plenum Publishers, Dordrecht, pp 253-261, 2001; Proceedings of 16th international offshore and polar engineering conference, San Francisco, CA, USA, 2006; J Mar Sci Technol 14:19-29, 2009); air/water interface dynamics is accurately modeled by a non standard level set approach (Di Mascio et al. in Comput Fluids 36(5):868-886, 2007a), known as the single-phase level set method. In this algorithm the governing equations are solved only in the water phase, whereas the numerical domain in the air phase is used for a suitable extension of the fluid dynamic variables. The level set function is used to track the free surface evolution; dynamic boundary conditions are enforced directly on the interface. This approach allows to accurately predict the evolution of the free surface even in the presence of violent breaking waves phenomena, maintaining the interface sharp, without any need to smear out the fluid properties across the two phases. This paper is aimed at the prediction of the complex free-surface flow field generated by a deep-V planing boat at medium and high Froude numbers (from 0.6 up to 1.2). In the present work, the planing hull is treated as a two-degree-of-freedom rigid object. Flow field is characterized by the presence of thin water sheets, several energetic breaking waves and plungings. The computational results include convergence of the trim angle, sinkage and resistance under grid refinement; high-quality experimental data are used for the purposes of validation, allowing to

Knockout of GAD65 has major impact on synaptic GABA synthesized from astrocyte-derived glutamine

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Walls, Anne Byriel; Eyjolfsson, Elvar M.; Smeland, Olav B.

2011-01-01

γ-Aminobutyric acid (GABA) synthesis from glutamate is catalyzed by glutamate decarboxylase (GAD) of which two isoforms, GAD65 and GAD67, have been identified. The GAD65 has repeatedly been shown to be important during intensified synaptic activity. To specifically elucidate the significance of G...... glutamine both via direct synthesis and via a pathway involving mitochondrial metabolism. Furthermore, a severe neuronal hypometabolism, involving glycolysis and tricarboxylic acid (TCA) cycle activity, was observed in cerebral cortex of GAD65 knockout mice.......65 for maintenance of the highly compartmentalized intracellular and intercellular GABA homeostasis, GAD65 knockout and corresponding wild-type mice were injected with [1-(13)C]glucose and the astrocyte-specific substrate [1,2-(13)C]acetate. Synthesis of GABA from glutamine in the GABAergic synapses...

Simple and Accurate Analytical Solutions of the Electrostatically Actuated Curled Beam Problem

KAUST Repository

Younis, Mohammad I.

2014-08-17

We present analytical solutions of the electrostatically actuated initially deformed cantilever beam problem. We use a continuous Euler-Bernoulli beam model combined with a single-mode Galerkin approximation. We derive simple analytical expressions for two commonly observed deformed beams configurations: the curled and tilted configurations. The derived analytical formulas are validated by comparing their results to experimental data in the literature and numerical results of a multi-mode reduced order model. The derived expressions do not involve any complicated integrals or complex terms and can be conveniently used by designers for quick, yet accurate, estimations. The formulas are found to yield accurate results for most commonly encountered microbeams of initial tip deflections of few microns. For largely deformed beams, we found that these formulas yield less accurate results due to the limitations of the single-mode approximations they are based on. In such cases, multi-mode reduced order models need to be utilized.

Beijing ambient particle exposure accelerates atherosclerosis in ApoE knockout mice.

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Chen, Tian; Jia, Guang; Wei, Yongjie; Li, Jiucun

2013-11-25

Air pollution is associated with significant adverse health effects including increased cardiovascular morbidity and mortality. However research on the cardiovascular effect of "real-world" exposure to ambient particulate matter (PM) in susceptible animal model is very limited. In this study, we aimed to investigate the association between Beijing ambient particle exposure and the atherosclerosis development in the apolipoprotein E knockout mice (ApoE(-/-) mice). Two parallel exposure chambers were used for whole body exposure among ApoE knockout mice. One of the chambers was supplied with untreated ambient air (PM group) and the other chamber was treated with ambient air filtered by high-efficiency particulate air (HEPA) filter (FA group). Twenty mice were divided into two groups and exposed to ambient PM (n=10 for PM group) or filtered air (n=10 for FA group) for two months from January 18th to March 18th, 2010. During the exposure, the mass concentrations of PM2.5 and PM10 in the two chambers were continuously monitored. Additionally, a receptor source apportionment model of chemical mass balance using 19 organic tracers was applied to determine the contributions of sources on the PM2.5 in terms of natural gas, diesel vehicle, gasoline vehicle, coal burning, vegetable debris, biomass burning and cooking. At the end of the two-month exposure, biomarkers of oxidative stress, inflammation and lipid metabolism in bronchoalveolar lavage fluid (BAL) and blood samples were determined and the plaque area on the aortic endothelium was quantified. In the experiment, the concentrations of PM10 and PM2.5 in PM chamber were 99.45μg/m(3) and 61.0μg/m(3) respectively, while PM2.5 in FA chamber was 17.6μg/m(3). Source apportionment analysis by organic tracers showed that gasoline vehicle (39.9%) and coal burning (24.3%) emission were the two major sources contributing to the mass concentration of PM2.5 in Beijing. Among the ApoE knockout mice, the PM group were significantly

Brief Report: Altered Social Behavior in Isolation-Reared "Fmr1" Knockout Mice

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Heitzer, Andrew M.; Roth, Alexandra K.; Nawrocki, Lauren; Wrenn, Craige C.; Valdovinos, Maria G.

2013-01-01

Social behavior abnormalities in Fragile X syndrome (FXS) are characterized by social withdrawal, anxiety, and deficits in social cognition. To assess these deficits, a model of FXS, the "Fmr1" knockout mouse ("Fmr1" KO), has been utilized. This mouse model has a null mutation in the fragile X mental retardation 1 gene ("Fmr1") and displays…

GPR39 (zinc receptor) knockout mice exhibit depression-like behavior and CREB/BDNF down-regulation in the hippocampus

DEFF Research Database (Denmark)

Młyniec, Katarzyna; Budziszewska, Bogusława; Holst, Birgitte

2015-01-01

Background: Zinc may act as a neurotransmitter in the central nervous system by activation of the GPR39 metabotropic receptors. Methods: In the present study, we investigated whether GPR39 knockout would cause depressive-like and/or anxiety-like behavior, as measured by the forced swim test, tail...... investigated activity in the hypothalamus-pituitary-adrenal axis under both zinc- and GPR39-deficient conditions. Zinc-deficient mice had higher serum corticosterone levels and lower glucocorticoid receptor levels in the hippocampus and frontal cortex. Conclusions: There were no changes in the GPR39 knockout...

Physiological roles of CNS muscarinic receptors gained from knockout mice

DEFF Research Database (Denmark)

Thomsen, Morgane; Sørensen, Gunnar; Dencker, Ditte

2017-01-01

receptors modulating neuronal activity and neurotransmitter release in many brain regions, shaping neuronal plasticity, and affecting functions ranging from motor and sensory function to cognitive processes. As gene targeting technology evolves including the use of conditional, cell type specific strains......, knockout mice are likely to continue to provide valuable insights into brain physiology and pathophysiology, and advance the development of new medications for a range of conditions such as Alzheimer's disease, Parkinson's disease, schizophrenia, and addictions, as well as non-opioid analgesics...

CD8 Knockout Mice Are Protected from Challenge by Vaccination with WR201, a Live Attenuated Mutant of Brucella melitensis

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Samuel L. Yingst

2013-01-01

Full Text Available CD8+ T cells have been reported to play an important role in defense against B. abortus infection in mouse models. In the present report, we use CD8 knockout mice to further elucidate the role of these cells in protection from B. melitensis infection. Mice were immunized orally by administration of B. melitensis WR201, a purine auxotrophic attenuated vaccine strain, then challenged intranasally with B. melitensis 16M. In some experiments, persistence of WR201 in the spleens of CD8 knockout mice was slightly longer than that in the spleens of normal mice. However, development of anti-LPS serum antibody, antigen-induced production of γ-interferon (IFN-γ by immune splenic lymphocytes, protection against intranasal challenge, and recovery of nonimmunized animals from intranasal challenge were similar between normal and knockout animals. Further, primary Brucella infection was not exacerbated in perforin knockout and Fas-deficient mice and these animals’ anti-Brucella immune responses were indistinguishable from those of normal mice. These results indicate that CD8+ T cells do not play an essential role as either cytotoxic cells or IFN-γ producers, yet they do participate in a specific immune response to immunization and challenge in this murine model of B. melitensis infection.

On-Line Self-Calibrating Single Crystal Sapphire Optical Sensor Instrumentation for Accurate and Reliable Coal Gasifier Temperature Measurement

Energy Technology Data Exchange (ETDEWEB)

Kristie Cooper; Gary Pickrell; Anbo Wang

2005-11-01

This report summarizes technical progress April-September 2005 on the Phase II program ''On-Line Self-Calibrating Single Crystal Sapphire Optical Sensor Instrumentation for Accurate and Reliable Coal Gasifier Temperature Measurement'', funded by the Federal Energy Technology Center of the U.S. Department of Energy, and performed by the Center for Photonics Technology of the Bradley Department of Electrical and Computer Engineering at Virginia Tech. The outcome of the first phase of this program was the selection of broadband polarimetric differential interferometry (BPDI) for further prototype instrumentation development. This approach is based on the measurement of the optical path difference (OPD) between two orthogonally polarized light beams in a single-crystal sapphire disk. The objective of this program is to bring the sensor technology, which has already been demonstrated in the laboratory, to a level where the sensor can be deployed in the harsh industrial environments and will become commercially viable. Due to the difficulties described on the last report, field testing of the BPDI system has not continued to date. However, we have developed an alternative high temperature sensing solution, which is described in this report. The sensing system will be installed and tested at TECO's Polk Power Station. Following a site visit in June 2005, our efforts have been focused on preparing for that field test, including he design of the sensor mechanical packaging, sensor electronics, the data transfer module, and the necessary software codes to accommodate this application.. We are currently ready to start sensor fabrication.

Prohormone convertase 2 activity is increased in the hippocampus of Wfs1 knockout mice

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Karin eTein

2015-08-01

Full Text Available BackgroundMutations in WFS1 gene cause Wolfram syndrome, which is a rare autosomal recessive disorder, characterized by diabetes insipidus, diabetes mellitus, optic nerve atrophy and deafness (DIDMOAD. The WFS1 gene product wolframin is located in the endoplasmic reticulum. Mice lacking this gene exhibit disturbances in the processing and secretion of peptides, such as vasopressin and insulin. In the brain, high levels of the wolframin protein have been observed in the hippocampus, amygdala and limbic structures. The aim of this study was to investigate the effect of Wfs1 knockout on peptide processing in mouse hippocampus. A peptidomic approach was used to characterize individual peptides in the hippocampus of wild-type and Wfs1 knockout mice. ResultsWe identified 126 peptides in hippocampal extracts and the levels of 10 peptides differed between Wfs1 KO and wild-type mice at P<0.05. The peptide with the largest alteration was little-LEN, which level was 25 times higher in the hippocampus of Wfs1 KO mice compared to wild-type mice. Processing (cleavage of little-LEN from the Pcsk1n gene product proSAAS involves prohormone convertase 2 (PC2. Thus, PC2 activity was measured in extracts prepared from the hippocampus of Wfs1 knockout mice. The activity of PC2 in Wfs1 mutant mice was significantly higher (149.9±2.3%, p<0.0001, n=8 than in wild-type mice (100.0±7.0%, n=8. However, Western blot analysis showed that protein levels of 7B2, proPC2 and PC2 were same in both groups, and so were gene expression levels.ConclusionsProcessing of proSAAS is altered in the hippocampus of Wfs1-KO mice, which is caused by increased activity of PC2. Increased activity of PC2 in Wfs1 knockout mice is not caused by alteration in the levels of PC2 protein. Our results suggest a functional link between Wfs1 and PC2. Thus, the detailed molecular mechanism of the role of Wfs1 in the regulation of PC2 activity needs further investigation.

Probing the structure of unstable nuclei through the recoiled proton tagged knockout reaction

International Nuclear Information System (INIS)

Ye, Y.; Cao, Z.; Jiang, D.

2010-01-01

Recoiled proton tagged knockout reaction experiments were carried-out for 8 He at 82,5 MeV/u in RIKEN and for 6 He at 65 MeV/u in Lanzhou. The very preliminary results for the distinguish of the reaction mechanism are presented and compared to the kinematics calculation. (authors)

β2-Adrenergic Receptor Knockout Mice Exhibit A Diabetic Retinopathy Phenotype

Science.gov (United States)

Jiang, Youde; Zhang, Qiuhua; Liu, Li; Tang, Jie; Kern, Timothy S.; Steinle, Jena J.

2013-01-01

There is considerable evidence from our lab and others for a functional link between β-adrenergic receptor and insulin receptor signaling pathways in retina. Furthermore, we hypothesize that this link may contribute to lesions similar to diabetic retinopathy in that the loss of adrenergic input observed in diabetic retinopathy may disrupt normal anti-apoptotic insulin signaling, leading to retinal cell death. Our studies included assessment of neural retina function (ERG), vascular degeneration, and Müller glial cells (which express only β1 and β2-adrenergic receptor subtypes). In the current study, we produced β2-adrenergic receptor knockout mice to examine this deletion on retinal neurons and vasculature, and to identify specific pathways through which β2-adrenergic receptor modulates insulin signaling. As predicted from our hypothesis, β2-adrenergic receptor knockout mice display certain features similar to diabetic retinopathy. In addition, loss of β2-adrenergic input resulted in an increase in TNFα, a key inhibitor of insulin receptor signaling. Increased TNFα may be associated with insulin-dependent production of the anti-apoptotic factor, Akt. Since the effects occurred in vivo under normal glucose conditions, we postulate that aspects of the diabetic retinopathy phenotype might be triggered by loss of β2-adrenergic receptor signaling. PMID:23894672

A model of knock-out of oxygen by charged particle irradiation of Bi-2212

International Nuclear Information System (INIS)

Bandyopadhyay, S.K.; Sen, Pintu; Barat, P.; Mukherjee, P.; Das, S.K.; Ghosh, B.

1996-01-01

A model of knock-out of oxygen by charged particle (α and proton) irradiation of Bi 2 Sr 2 CaCu 2 O 8+x (Bi-2212) is proposed on the basis of Monte Carlo TRIM calculations. In Bi-2212, the loosely bound excess oxygen is vulnerable to be displaced by particle irradiation. Binding energy and hence, displacement energy of this loosely bound excess oxygen is less compared to that of stoichiometric lattice bound oxygen and other atoms. The displaced or knocked out oxygen goes to pores or intergranular region and generates large pressure inside the sample. Because of porosity of the material, this displaced oxygen diffuses out and there is a net reduction of oxygen content of the sample. The irradiation induced oxygen knock-out is dominant in the bulk where nonionizing energy loss is maximum. (author). 29 refs., 1 fig., 3 tabs

Age- and region-specific imbalances of basal amino acids and monoamine metabolism in limbic regions of female Fmr1 knock-out mice.

Science.gov (United States)

Gruss, Michael; Braun, Katharina

2004-07-01

The Fragile X syndrome, a common form of mental retardation in humans, originates from the loss of expression of the Fragile X mental retardation gene leading to the absence of the encoded Fragile X mental retardation protein 1 (FMRP). A broad pattern of morphological and behavioral abnormalities is well described for affected humans as well as Fmr1 knock-out mice, a transgenic animal model for the human Fragile X syndrome. In the present study, we examined neurochemical differences between female Fmr1 knock-out and wildtype mice with particular focus on neurotransmission. Significant age- and region-specific differences of basal tissue neurotransmitter and metabolite levels measured by high performance liquid chromatography were found. Those differences were more numerous in juvenile animals (postnatal day (PND) 28-31) compared to adults (postnatal day 209-221). In juvenile female knock-out mice, especially aspartate and taurine were increased in cortical regions, striatum, cerebellum, and brainstem. Furthermore, compared to the wildtype animals, the juvenile knock-out mice displayed an increased level of neuronal inhibition in the hippocampus and brainstem reflected by decreased ratios of (aspartate + glutamate)/(taurine + GABA), as well as an increased dopamine (DA) turnover in cortical regions, striatum, and hippocampus. These results provide the first evidence that the lack of FMRP expression in female Fmr1 knock-out mice is accompanied by age-dependent, region-specific alterations in brain amino acids, and monoamine turnover, which might be related to the reported synaptical and behavioural alterations in these animals.

Generation of ER{alpha}-floxed and knockout mice using the Cre/LoxP system

Energy Technology Data Exchange (ETDEWEB)

Antonson, P., E-mail: per.antonson@ki.se [Department of Biosciences and Nutrition, Karolinska Institutet, Novum, SE-141 83 Huddinge (Sweden); Omoto, Y.; Humire, P. [Department of Biosciences and Nutrition, Karolinska Institutet, Novum, SE-141 83 Huddinge (Sweden); Gustafsson, J.-A. [Department of Biosciences and Nutrition, Karolinska Institutet, Novum, SE-141 83 Huddinge (Sweden); Center for Nuclear Receptors and Cell Signaling, Department of Biology and Biochemistry, University of Houston, Houston, TX 77204 (United States)

2012-08-10

Highlights: Black-Right-Pointing-Pointer ER{alpha} floxed and knockout mice were generated. Black-Right-Pointing-Pointer Disruption of the ER{alpha} gene results in sterility in both male and female mice. Black-Right-Pointing-Pointer ER{alpha}{sup -/-} mice have ovaries with hemorrhagic follicles and hypoplastic uterus. Black-Right-Pointing-Pointer Female ER{alpha}{sup -/-} mice develop obesity. -- Abstract: Estrogen receptor alpha (ER{alpha}) is a nuclear receptor that regulates a range of physiological processes in response to estrogens. In order to study its biological role, we generated a floxed ER{alpha} mouse line that can be used to knock out ER{alpha} in selected tissues by using the Cre/LoxP system. In this study, we established a new ER{alpha} knockout mouse line by crossing the floxed ER{alpha} mice with Cre deleter mice. Here we show that genetic disruption of the ER{alpha} gene in all tissues results in sterility in both male and female mice. Histological examination of uterus and ovaries revealed a dramatically atrophic uterus and hemorrhagic cysts in the ovary. These results suggest that infertility in female mice is the result of functional defects of the reproductive tract. Moreover, female knockout mice are hyperglycemic, develop obesity and at the age of 4 months the body weight of these mice was more than 20% higher compared to wild type littermates and this difference increased over time. Our results demonstrate that ER{alpha} is necessary for reproductive tract development and has important functions as a regulator of metabolism in females.

Studies of UCP2 transgenic and knockout mice reveal that liver UCP2 is not essential for the antiobesity effects of fish oil.

Science.gov (United States)

Tsuboyama-Kasaoka, Nobuyo; Sano, Kayo; Shozawa, Chikako; Osaka, Toshimasa; Ezaki, Osamu

2008-03-01

Uncoupling protein 2 (UCP2) is a possible target molecule for energy dissipation. Many dietary fats, including safflower oil and lard, induce obesity in C57BL/6 mice, whereas fish oil does not. Fish oil increases UCP2 expression in hepatocytes and may enhance UCP2 activity by activating the UCP2 molecule or altering the lipid bilayer environment. To examine the role of liver UCP2 in obesity, we created transgenic mice that overexpressed human UCP2 in hepatocytes and examined whether UCP2 transgenic mice showed less obesity when fed a high-fat diet (safflower oil or lard). In addition, we examined whether fish oil had antiobesity effects in UCP2 knockout mice. UCP2 transgenic and wild-type mice fed a high-fat diet (safflower oil or lard) developed obesity to a similar degree. UCP2 knockout and wild-type mice fed fish oil had lower rates of obesity than mice fed safflower oil. Remarkably, safflower oil did not induce obesity in female UCP2 knockout mice, an unexpected phenotype for which we presently have no explanation. However, this unexpected effect was not observed in male UCP2 knockout mice or in UCP2 knockout mice fed a high-lard diet. These data indicate that liver UCP2 is not essential for fish oil-induced decreases in body fat.

Raphe serotonin neuron-specific oxytocin receptor knockout reduces aggression without affecting anxiety-like behavior in male mice only.

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Pagani, J H; Williams Avram, S K; Cui, Z; Song, J; Mezey, É; Senerth, J M; Baumann, M H; Young, W S

2015-02-01

Serotonin and oxytocin influence aggressive and anxiety-like behaviors, though it is unclear how the two may interact. That the oxytocin receptor is expressed in the serotonergic raphe nuclei suggests a mechanism by which the two neurotransmitters may cooperatively influence behavior. We hypothesized that oxytocin acts on raphe neurons to influence serotonergically mediated anxiety-like, aggressive and parental care behaviors. We eliminated expression of the oxytocin receptor in raphe neurons by crossing mice expressing Cre recombinase under control of the serotonin transporter promoter (Slc6a4) with our conditional oxytocin receptor knockout line. The knockout mice generated by this cross are normal across a range of behavioral measures: there are no effects for either sex on locomotion in an open-field, olfactory habituation/dishabituation or, surprisingly, anxiety-like behaviors in the elevated O and plus mazes. There was a profound deficit in male aggression: only one of 11 raphe oxytocin receptor knockouts showed any aggressive behavior, compared to 8 of 11 wildtypes. In contrast, female knockouts displayed no deficits in maternal behavior or aggression. Our results show that oxytocin, via its effects on raphe neurons, is a key regulator of resident-intruder aggression in males but not maternal aggression. Furthermore, this reduction in male aggression is quite different from the effects reported previously after forebrain or total elimination of oxytocin receptors. Finally, we conclude that when constitutively eliminated, oxytocin receptors expressed by serotonin cells do not contribute to baseline anxiety-like behaviors or maternal care. © 2015 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

CRISPR/Cas9-mediated gene knockout is insensitive to target copy number but is dependent on guide RNA potency and Cas9/sgRNA threshold expression level.

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Yuen, Garmen; Khan, Fehad J; Gao, Shaojian; Stommel, Jayne M; Batchelor, Eric; Wu, Xiaolin; Luo, Ji

2017-11-16

CRISPR/Cas9 is a powerful gene editing tool for gene knockout studies and functional genomic screens. Successful implementation of CRISPR often requires Cas9 to elicit efficient target knockout in a population of cells. In this study, we investigated the role of several key factors, including variation in target copy number, inherent potency of sgRNA guides, and expression level of Cas9 and sgRNA, in determining CRISPR knockout efficiency. Using isogenic, clonal cell lines with variable copy numbers of an EGFP transgene, we discovered that CRISPR knockout is relatively insensitive to target copy number, but is highly dependent on the potency of the sgRNA guide sequence. Kinetic analysis revealed that most target mutation occurs between 5 and 10 days following Cas9/sgRNA transduction, while sgRNAs with different potencies differ by their knockout time course and by their terminal-phase knockout efficiency. We showed that prolonged, low level expression of Cas9 and sgRNA often fails to elicit target mutation, particularly if the potency of the sgRNA is also low. Our findings provide new insights into the behavior of CRISPR/Cas9 in mammalian cells that could be used for future improvement of this platform. Published by Oxford University Press on behalf of Nucleic Acids Research 2017.

Adeno-associated virus LPL(S447X) gene therapy in LDL receptor knockout mice

NARCIS (Netherlands)

Rip, Jaap; Sierts, Jeroen A.; Vaessen, Stefan F. C.; Kastelein, John J. P.; Twisk, Jaap; Kuivenhoven, Jan Albert

2007-01-01

BACKGROUND: Overexpression of lipoprotein lipase (LPL) protects against atherosclerosis in genetically engineered mice. We tested whether a gene therapy vector that delivers human (h) LPL(S447X) cDNA to skeletal muscle could induce similar effects. METHODS: LDL receptor knockout (LDLr-/-) mice were

Using an FPGA for Fast Bit Accurate SoC Simulation

NARCIS (Netherlands)

Wolkotte, P.T.; Holzenspies, P.K.F.; Smit, Gerardus Johannes Maria

In this paper we describe a sequential simulation method to simulate large parallel homo- and heterogeneous systems on a single FPGA. The method is applicable for parallel systems were lengthy cycle and bit accurate simulations are required. It is particularly designed for systems that do not fit

Characterization of a Bacillus subtilis surfactin synthetase knockout and antimicrobial activity analysis.

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Liu, Hongxia; Qu, Xiaoxu; Gao, Ling; Zhao, Shengming; Lu, Zhaoxin; Zhang, Chong; Bie, Xiaomei

2016-11-10

Gene knockout is an important approach to improve the production of antimicrobial compounds. B. subtilis PB2-LS10, derived from B. subtilis PB2-L by a surfactin synthetase (srf) genes knockout, exhibits stronger inhibitory action than its parental strain against all tested pathogenic bacteria and fungi. The antimicrobial extracts produced by B. subtilis PB2-L and B. subtilis PB2-LS10 respectively were characterized by the high-resolution LC-ESI-MS. To provide further insight into the distinct antimicrobial activities, we investigated the impact of the srf genes deletion on the growth and gene transcriptional profile of the strains. The mutant strain grew quickly and reached stationary phase 2h earlier than the wild-type. Prominent expression changes in the modified strain involved genes that were essential to metabolic pathways and processes. Genes related to amino acid transport, ATP-binding cassette (ABC) transporters and protein export were up-regulated in strain PB2-LS10. However, amino acid metabolism, carbohydrate metabolism and fatty acid metabolism were repressed. Because of its excellent antimicrobial activity, strain PB2-LS10 has potential for use in food preservation. Copyright © 2016 Elsevier B.V. All rights reserved.

Acute food deprivation reverses morphine-induced locomotion deficits in M5 muscarinic receptor knockout mice.

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Steidl, Stephan; Lee, Esther; Wasserman, David; Yeomans, John S

2013-09-01

Lesions of the pedunculopontine tegmental nucleus (PPT), one of two sources of cholinergic input to the ventral tegmental area (VTA), block conditioned place preference (CPP) for morphine in drug-naïve rats. M5 muscarinic cholinergic receptors, expressed by midbrain dopamine neurons, are critical for the ability of morphine to increase nucleus accumbens dopamine levels and locomotion, and for morphine CPP. This suggests that M5-mediated PPT cholinergic inputs to VTA dopamine neurons critically contribute to morphine-induced dopamine activation, reward and locomotion. In the current study we tested whether food deprivation, which reduces PPT contribution to morphine CPP in rats, could also reduce M5 contributions to morphine-induced locomotion in mice. Acute 18-h food deprivation reversed the phenotypic differences usually seen between non-deprived wild-type and M5 knockout mice. That is, food deprivation increased morphine-induced locomotion in M5 knockout mice but reduced morphine-induced locomotion in wild-type mice. Food deprivation increased saline-induced locomotion equally in wild-type and M5 knockout mice. Based on these findings, we suggest that food deprivation reduces the contribution of M5-mediated PPT cholinergic inputs to the VTA in morphine-induced locomotion and increases the contribution of a PPT-independent pathway. The contributions of cholinergic, dopaminergic and GABAergic neurons to the effects of acute food deprivation are discussed. Copyright © 2013 Elsevier B.V. All rights reserved.

Impairment of Hepcidin Upregulation by Lipopolysaccharide in the Interleukin-6 Knockout Mouse Brain

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Fa-Li Zhang

2017-11-01

Full Text Available To find out whether the Interleukin-6 (IL-6/signal transducer and activator of transcription 3 (STAT3 signaling pathway is involved in the expression of hepcidin in the mouse brain in vivo, we investigated the phosphorylation of STAT3, as well as the expression of hepcidin mRNA, ferroportin 1 (Fpn1 and ferritin light chain (Ft-L proteins in the cortex and hippocampus of LPS-treated wild type (IL-6+/+ and IL-6 knockout (IL-6-/- mice. We demonstrated that IL-6 knockout could significantly reduce the response of hepcidin mRNA, phospho-STAT3, Fpn1 and Ft-L protein expression to LPS treatment, in both the cortex and hippocampus of mice. Also, Stattic, an inhibitor of STAT3, significantly reduced the expression of phospho-STAT3 and hepcidin mRNA in the cortex and hippocampus of the LPS-treated wild type mice. These findings provide in vivo evidence for the involvement of the IL-6/STAT3 signaling pathway in the expression of hepcidin.

RNA-seq reveals transcriptome changes in goats following myostatin gene knockout

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Cai, Bei; Zhou, Shiwei; Zhu, Haijing; Qu, Lei; Wang, Xiaolong

2017-01-01

Myostatin (MSTN) is a powerful negative regulator of skeletal muscle mass in mammalian species that is primarily expressed in skeletal muscles, and mutations of its encoding gene can result in the double-muscling trait. In this study, the CRISPR/Cas9 technique was used to edit MSTN in Shaanbei Cashmere goats and generate knockout animals. RNA sequencing was used to determine and compare the transcriptome profiles of the muscles from three wild-type (WT) goats, three fibroblast growth factor 5 (FGF5) knockout goats (FGF5+/- group) and three goats with disrupted expression of both the FGF5 and MSTN genes (FM+/- group). The sequence reads were obtained using the Illumina HiSeq 2000 system and mapped to the Capra hircus reference genome using TopHat (v2.0.9). In total, 68.93, 62.04 and 66.26 million clean sequencing reads were obtained from the WT, FM+/- and FGF5+/- groups, respectively. There were 201 differentially expressed genes (DEGs) between the WT and FGF5+/- groups, with 86 down- and 115 up-regulated genes in the FGF5+/- group. Between the WT and FM+/- groups, 121 DEGs were identified, including 81 down- and 40 up-regulated genes in the FM+/- group. A total of 198 DEGs were detected between the FGF5+/- group and FM+/- group, with 128 down- and 70 up-regulated genes in the FM+/- group. At the transcriptome level, we found substantial changes in genes involved in fatty acid metabolism and the biosynthesis of unsaturated fatty acids, such as stearoyl-CoA dehydrogenase, 3-hydroxyacyl-CoA dehydratase 2, ELOVL fatty acid elongase 6 and fatty acid synthase, suggesting that the expression levels of these genes may be directly regulated by MSTN and that these genes are likely downstream targets of MSTN with potential roles in lipid metabolism in goats. Moreover, five randomly selected DEGs were further validated with qRT-PCR, and the results were consistent with the transcriptome analysis. The present study provides insight into the unique transcriptome profile of the

Fast and accurate methods for phylogenomic analyses

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Warnow Tandy

2011-10-01

Full Text Available Abstract Background Species phylogenies are not estimated directly, but rather through phylogenetic analyses of different gene datasets. However, true gene trees can differ from the true species tree (and hence from one another due to biological processes such as horizontal gene transfer, incomplete lineage sorting, and gene duplication and loss, so that no single gene tree is a reliable estimate of the species tree. Several methods have been developed to estimate species trees from estimated gene trees, differing according to the specific algorithmic technique used and the biological model used to explain differences between species and gene trees. Relatively little is known about the relative performance of these methods. Results We report on a study evaluating several different methods for estimating species trees from sequence datasets, simulating sequence evolution under a complex model including indels (insertions and deletions, substitutions, and incomplete lineage sorting. The most important finding of our study is that some fast and simple methods are nearly as accurate as the most accurate methods, which employ sophisticated statistical methods and are computationally quite intensive. We also observe that methods that explicitly consider errors in the estimated gene trees produce more accurate trees than methods that assume the estimated gene trees are correct. Conclusions Our study shows that highly accurate estimations of species trees are achievable, even when gene trees differ from each other and from the species tree, and that these estimations can be obtained using fairly simple and computationally tractable methods.

Characterization of Bombyx mori nucleopolyhedrovirus with a knockout of Bm17

OpenAIRE

Shen, Hongxing; Zhou, Yang; Zhang, Wen; Nin, Bin; Wang, Hua; Wang, Xiaochun; Shao, Shihe; Chen, Huiqing; Guo, Zhongjian; Liu, Xiaoyong; Yao, Qin; Chen, Keping

2012-01-01

Open reading frame 17 (Bm17) gene of Bombyx mori nucleopolyhedrovirus is a highly conserved gene in lepidopteran nucleopolyhedroviruses, but its function remains unknown. In this report, transient-expression and superinfection assays indicated that BM17 localized in the nucleus and cytoplasm of infected BmN cells. To determine the role of Bm17 in baculovirus life cycle, we constructed a Bm17 knockout virus and characterized its properties in cells. Analysis of the production and infection of ...

Spectroscopy of 17C via one-neutron knockout reaction

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Kim Sunji

2016-01-01

Full Text Available A spectroscopic study of 17C was performed via the one-neutron knockout reaction of 18C on a carbon target at RIKEN-RIBF. Three unbound states at excitation energies of 2.66(2, 3.16(5, and 3.97(3 MeV (preliminary were observed. The energies are compared with shell-model calculations and existing measurements to deduce their spin-parities. From the comparison, the states at 2.66(2 and 3.97(3 MeV are suggested to be 1/2− and 3/2−, respectively. From its decay property, the state at 3.16(5 MeV is indicated to be 9/2+.

Pharmacological treatment of fragile X syndrome with GABAergic drugs in a knockout mouse model

NARCIS (Netherlands)

Heulens, Inge; D'Hulst, Charlotte; Van Dam, Debby; De Deyn, Peter P.; Kooy, R. Frank

2012-01-01

Molecular and electrophysiological studies have provided evidence for a general downregulation of the GABAergic system in the Fmr1 knockout mouse. GABA(A) receptors are the main inhibitory receptors in the brain and the GABA(A) receptor was proposed as a novel target for treatment of the fragile X

Imaging colon cancer development in mice: IL-6 deficiency prevents adenoma in azoxymethane-treated Smad3 knockouts

Science.gov (United States)

Harpel, Kaitlin; Leung, Sarah; Faith Rice, Photini; Jones, Mykella; Barton, Jennifer K.; Bommireddy, Ramireddy

2016-02-01

The development of colorectal cancer in the azoxymethane-induced mouse model can be observed by using a miniaturized optical coherence tomography (OCT) imaging system. This system is uniquely capable of tracking disease development over time, allowing for the monitoring of morphological changes in the distal colon due to tumor development and the presence of lymphoid aggregates. By using genetically engineered mouse models deficient in Interleukin 6 (IL-6) and Smad family member 3 (Smad3), the role of inflammation on tumor development and the immune system can be elucidated. Smad3 knockout mice develop inflammatory response, wasting, and colitis associated cancer while deficiency of proinflammatory cytokine IL-6 confers resistance to tumorigenesis. We present pilot data showing that the Smad3 knockout group had the highest tumor burden, highest spleen weight, and lowest thymus weight. The IL-6 deficiency in Smad3 knockout mice prevented tumor development, splenomegaly, and thymic atrophy. This finding suggests that agents that inhibit IL-6 (e.g. anti-IL-6 antibody, non-steroidal anti-inflammatory drugs [NSAIDs], etc.) could be used as novel therapeutic agents to prevent disease progression and increase the efficacy of anti-cancer agents. OCT can also be useful for initiating early therapy and assessing the benefit of combination therapy targeting inflammation.

A knockout mutation of a constitutive GPCR in Tetrahymena decreases both G-protein activity and chemoattraction.

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Thomas J Lampert

Full Text Available Although G-protein coupled receptors (GPCRs are a common element in many chemosensory transduction pathways in eukaryotic cells, no GPCR or regulated G-protein activity has yet been shown in any ciliate. To study the possible role for a GPCR in the chemoresponses of the ciliate Tetrahymena, we have generated a number of macronuclear gene knockouts of putative GPCRs found in the Tetrahymena Genome database. One of these knockout mutants, called G6, is a complete knockout of a gene that we call GPCR6 (TTHERM_00925490. Based on sequence comparisons, the Gpcr6p protein belongs to the Rhodopsin Family of GPCRs. Notably, Gpcr6p shares highest amino acid sequence homologies to GPCRs from Paramecium and several plants. One of the phenotypes of the G6 mutant is a decreased responsiveness to the depolarizing ions Ba²⁺ and K⁺, suggesting a decrease in basal excitability (decrease in Ca²⁺ channel activity. The other major phenotype of G6 is a loss of chemoattraction to lysophosphatidic acid (LPA and proteose peptone (PP, two known chemoattractants in Tetrahymena. Using microsomal [³⁵S]GTPγS binding assays, we found that wild-type (CU427 have a prominent basal G-protein activity. This activity is decreased to the same level by pertussis toxin (a G-protein inhibitor, addition of chemoattractants, or the G6 mutant. Since the basal G-protein activity is decreased by the GPCR6 knockout, it is likely that this gene codes for a constitutively active GPCR in Tetrahymena. We propose that chemoattractants like LPA and PP cause attraction in Tetrahymena by decreasing the basal G-protein stimulating activity of Gpcr6p. This leads to decreased excitability in wild-type and longer runs of smooth forward swimming (less interrupted by direction changes towards the attractant. Therefore, these attractants may work as inverse agonists through the constitutively active Gpcr6p coupled to a pertussis-sensitive G-protein.

An FBXO40 knockout generated by CRISPR/Cas9 causes muscle hypertrophy in pigs without detectable pathological effects.

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Zou, Yunlong; Li, Zhiyuan; Zou, Yunjing; Hao, Haiyang; Li, Ning; Li, Qiuyan

2018-04-15

The regulatory function of Fbxo40 has been well characterized in mice. As a key component of the SCF-E3 ubiquitin ligase complex, Fbxo40 induces IRS1 ubiquitination, thus inactivating the IGF1/Akt pathway. The expression of Fbxo40 is restricted to muscle, and mice with an Fbxo40 null mutation exhibit muscle hypertrophy. However, the function of FBXO40 has not been elucidated in pigs, and it is not known whether FBXO40 mutations affect their health. We therefore generated FBXO40 knockout pigs using somatic cell nuclear transfer (SCNT) technology. CRISPR/Cas9 technology was combined with G418 selection, making it possible to generate donor cells at an efficiency of 75.86%. In muscle from FBXO40 knockout pigs, IRS1 levels were higher, and the IGF1/Akt pathway was stimulated. Mutant animals also had approximately 4% more muscle mass compared to WT controls. The knockout pigs developed normally and no pathological changes were found in major organs. These results demonstrate that FBXO40 is a promising candidate gene for improving production traits in agricultural livestock and for developing therapeutic interventions for muscle diseases. Copyright © 2018. Published by Elsevier Inc.

Energy-switching potential energy surface for the water molecule revisited: A highly accurate singled-sheeted form.

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Galvão, B R L; Rodrigues, S P J; Varandas, A J C

2008-07-28

A global ab initio potential energy surface is proposed for the water molecule by energy-switching/merging a highly accurate isotope-dependent local potential function reported by Polyansky et al. [Science 299, 539 (2003)] with a global form of the many-body expansion type suitably adapted to account explicitly for the dynamical correlation and parametrized from extensive accurate multireference configuration interaction energies extrapolated to the complete basis set limit. The new function mimics also the complicated Sigma/Pi crossing that arises at linear geometries of the water molecule.

Effects of alpha-AMPK knockout on exercise-induced gene activation in mouse skeletal muscle

DEFF Research Database (Denmark)

Jørgensen, Sebastian Beck; Wojtaszewski, Jørgen; Viollet, Benoit

2005-01-01

We tested the hypothesis that 5'AMP-activated protein kinase (AMPK) plays an important role in regulating the acute, exercise-induced activation of metabolic genes in skeletal muscle, which were dissected from whole-body a2- and a1-AMPK knockout (KO) and wild-type (WT) mice at rest, after treadmi...

Peptidomic analysis of the neurolysin-knockout mouse brain.

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Castro, Leandro M; Cavalcanti, Diogo M L P; Araujo, Christiane B; Rioli, Vanessa; Icimoto, Marcelo Y; Gozzo, Fábio C; Juliano, Maria; Juliano, Luiz; Oliveira, Vitor; Ferro, Emer S

2014-12-05

A large number of intracellular peptides are constantly produced following protein degradation by the proteasome. A few of these peptides function in cell signaling and regulate protein-protein interactions. Neurolysin (Nln) is a structurally defined and biochemically well-characterized endooligopeptidase, and its subcellular distribution and biological activity in the vertebrate brain have been previously investigated. However, the contribution of Nln to peptide metabolism in vivo is poorly understood. In this study, we used quantitative mass spectrometry to investigate the brain peptidome of Nln-knockout mice. An additional in vitro digestion assay with recombinant Nln was also performed to confirm the identification of the substrates and/or products of Nln. Altogether, the data presented suggest that Nln is a key enzyme in the in vivo degradation of only a few peptides derived from proenkephalin, such as Met-enkephalin and octapeptide. Nln was found to have only a minor contribution to the intracellular peptide metabolism in the entire mouse brain. However, further studies appear necessary to investigate the contribution of Nln to the peptide metabolism in specific areas of the murine brain. Neurolysin was first identified in the synaptic membranes of the rat brain in the middle 80's by Frederic Checler and colleagues. Neurolysin was well characterized biochemically, and its brain distribution has been confirmed by immunohistochemical methods. The neurolysin contribution to the central and peripheral neurotensin-mediated functions in vivo has been delineated through inhibitor-based pharmacological approaches, but its genuine contribution to the physiological inactivation of neuropeptides remains to be firmly established. As a result, the main significance of this work is the first characterization of the brain peptidome of the neurolysin-knockout mouse. This article is part of a Special Issue entitled: Proteomics, mass spectrometry and peptidomics, Cancun 2013

TAM receptor knockout mice are susceptible to retinal autoimmune induction.

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Ye, Fei; Li, Qiutang; Ke, Yan; Lu, Qingjun; Han, Lixia; Kaplan, Henry J; Shao, Hui; Lu, Qingxian

2011-06-16

TAM receptors are expressed mainly by dendritic cells and macrophages in the immune system, and mice lacking TAM receptors develop systemic autoimmune diseases because of inefficient negative control of the cytokine signaling in those cells. This study aims to test the susceptibility of the TAM triple knockout (tko) mice to the retina-specific autoantigen to develop experimental autoimmune uveoretinitis (EAU). TAM tko mice that were or were not immunized with interphotoreceptor retinoid-binding protein (IRBP) peptides were evaluated for retinal infiltration of the macrophages and CD3(+) T cells by immunohistochemistry, spontaneous activation of CD4(+) T cells, and memory T cells by flow cytometry and proliferation of IRBP-specific CD4(+) T cells by [(3)H]thymidine incorporation assay. Ocular inflammation induced by IRBP peptide immunization and specific T cell transfer were observed clinically by funduscopy and confirmed by histology. Tko mice were found to have less naive, but more activated, memory T cells, among which were exhibited high sensitivity to ocular IRBP autoantigens. Immunization with a low dose of IRBP and adoptive transfer of small numbers of IRBP-specific T cells from immunized tko mice caused the infiltration of lymphocytes, including CD3(+) T cells, into the tko retina. Mice without TAM receptor spontaneously develop IRBP-specific CD4(+) T cells and are more susceptible to retinal autoantigen immunization. This TAM knockout mouse line provides an animal model with which to study the role of antigen-presenting cells in the development of T cell-mediated uveitis.

Evaluation of cimi-shield knock-out bed bug eliminator against house fly (Musca domestica) adults.

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Cimi-Shield Knock-Out (CSKO) Bed Bug Eliminator is a green treatment labeled for use against bed bugs, carpet beetles, ants, roaches, fleas, ticks, silverfish, millipedes and centipedes. The active ingredient is soybean oil. If CSKO is formulated according to label instructions and sprayed directly ...

The impact of a community-led program promoting weight loss and healthy living in Aboriginal communities: the New South Wales Knockout Health Challenge

OpenAIRE

Passmore, Erin; Shepherd, Brooke; Milat, Andrew; Maher, Louise; Hennessey, Kiel; Havrlant, Rachael; Maxwell, Michelle; Hodge, Wendy; Christian, Fiona; Richards, Justin; Mitchell, Jo

2017-01-01

Background Aboriginal people in Australia experience significant health burden from chronic disease. There has been limited research to identify effective healthy lifestyle programs to address risk factors for chronic disease among Aboriginal people. Methods The Knockout Health Challenge is a community-led healthy lifestyle program for Aboriginal communities across New South Wales, Australia. An evaluation of the 2013 Knockout Health Challenge was undertaken. Participants’ self-reported physi...

High throughput and accurate serum proteome profiling by integrated sample preparation technology and single-run data independent mass spectrometry analysis.

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Lin, Lin; Zheng, Jiaxin; Yu, Quan; Chen, Wendong; Xing, Jinchun; Chen, Chenxi; Tian, Ruijun

2018-03-01

Mass spectrometry (MS)-based serum proteome analysis is extremely challenging due to its high complexity and dynamic range of protein abundances. Developing high throughput and accurate serum proteomic profiling approach capable of analyzing large cohorts is urgently needed for biomarker discovery. Herein, we report a streamlined workflow for fast and accurate proteomic profiling from 1μL of blood serum. The workflow combined an integrated technique for highly sensitive and reproducible sample preparation and a new data-independent acquisition (DIA)-based MS method. Comparing with standard data dependent acquisition (DDA) approach, the optimized DIA method doubled the number of detected peptides and proteins with better reproducibility. Without protein immunodepletion and prefractionation, the single-run DIA analysis enables quantitative profiling of over 300 proteins with 50min gradient time. The quantified proteins span more than five orders of magnitude of abundance range and contain over 50 FDA-approved disease markers. The workflow allowed us to analyze 20 serum samples per day, with about 358 protein groups per sample being identified. A proof-of-concept study on renal cell carcinoma (RCC) serum samples confirmed the feasibility of the workflow for large scale serum proteomic profiling and disease-related biomarker discovery. Blood serum or plasma is the predominant specimen for clinical proteomic studies while the analysis is extremely challenging for its high complexity. Many efforts had been made in the past for serum proteomics for maximizing protein identifications, whereas few have been concerned with throughput and reproducibility. Here, we establish a rapid, robust and high reproducible DIA-based workflow for streamlined serum proteomic profiling from 1μL serum. The workflow doesn't need protein depletion and pre-fractionation, while still being able to detect disease-relevant proteins accurately. The workflow is promising in clinical application

Leukocytosis and enhanced susceptibility to endotoxemia but not atherosclerosis in adrenalectomized APOE knockout mice.

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Menno Hoekstra

Full Text Available Hyperlipidemic apolipoprotein E (APOE knockout mice show an enhanced level of adrenal-derived anti-inflammatory glucocorticoids. Here we determined in APOE knockout mice the impact of total removal of adrenal function through adrenalectomy (ADX on two inflammation-associated pathologies, endotoxemia and atherosclerosis. ADX mice exhibited 91% decreased corticosterone levels (P<0.001, leukocytosis (WBC count: 10.0 ± 0.4 x 10E9/L vs 6.5 ± 0.5 x 10E9/L; P<0.001 and an increased spleen weight (P<0.01. FACS analysis on blood leukocytes revealed increased B-lymphocyte numbers (55 ± 2% vs 46 ± 1%; P<0.01. T-cell populations in blood appeared to be more immature (CD62L+: 26 ± 2% vs 19 ± 1% for CD4+ T-cells, P<0.001 and 58 ± 7% vs 47 ± 4% for CD8+ T-cells, P<0.05, which coincided with immature CD4/CD8 double positive thymocyte enrichment. Exposure to lipopolysaccharide failed to increase corticosterone levels in ADX mice and was associated with a 3-fold higher (P<0.05 TNF-alpha response. In contrast, the development of initial fatty streak lesions and progression to advanced collagen-containing atherosclerotic lesions was unaffected. Plasma cholesterol levels were decreased by 35% (P<0.001 in ADX mice. This could be attributed to a decrease in pro-atherogenic very-low-density lipoproteins (VLDL as a result of a diminished hepatic VLDL secretion rate (-24%; P<0.05. In conclusion, our studies show that adrenalectomy induces leukocytosis and enhances the susceptibility for endotoxemia in APOE knockout mice. The adrenalectomy-associated rise in white blood cells, however, does not alter atherosclerotic lesion development probably due to the parallel decrease in plasma levels of pro-atherogenic lipoproteins.

Knockout and transgenic mice of Trp53: what have we learned about p53 in breast cancer?

International Nuclear Information System (INIS)

Blackburn, Anneke C; Jerry, D Joseph

2002-01-01

The human p53 tumor suppressor gene TP53 is mutated at a high frequency in sporadic breast cancer, and Li-Fraumeni syndrome patients who carry germline mutations in one TP53 allele have a high incidence of breast cancer. In the 10 years since the first knockout of the mouse p53 tumor suppressor gene (designated Trp53) was published, much has been learned about the contribution of p53 to biology and tumor suppression in the breast through the use of p53 transgenic and knockout mice. The original mice deficient in p53 showed no mammary gland phenotype. However, studies using BALB/c-Trp53-deficient mice have demonstrated a delayed involution phenotype and a mammary tumor phenotype. Together with other studies of mutant p53 transgenes and p53 bitransgenics, a greater understanding has been gained of the role of p53 in involution, of the regulation of p53 activity by hormones, of the effect of mouse strain and modifier genes on tumor phenotype, and of the cooperation between p53 and other oncogenic pathways, chemical carcinogens and hormonal stimulation in mammary tumorigenesis. Both p53 transgenic and knockout mice are important in vivo tools for understanding breast cancer, and are yet to be exploited for developing therapeutic strategies in breast cancer

Knockout of the Gnrh genes in zebrafish: effects on reproduction and potential compensation by reproductive and feeding-related neuropeptides.

Science.gov (United States)

Marvel, Miranda; Spicer, Olivia Smith; Wong, Ten-Tsao; Zmora, Nilli; Zohar, Yonathan

2018-04-04

Gonadotropin-releasing hormone (GnRH) is known as a pivotal upstream regulator of reproduction in vertebrates. However, reproduction is not compromised in the hypophysiotropic Gnrh3 knockout line in zebrafish (gnrh3-/-). In order to determine if Gnrh2, the only other Gnrh isoform in zebrafish brains, is compensating for the loss of Gnrh3, we generated a double Gnrh knockout zebrafish line. Surprisingly, the loss of both Gnrh isoforms resulted in no major impact on reproduction, indicating that a compensatory response, outside of the Gnrh system, was evoked. A plethora of factors acting along the reproductive hypothalamus-pituitary axis were evaluated as possible compensators based on neuroanatomical and differential gene expression studies. In addition, we also examined the involvement of feeding factors in the brain as potential compensators for Gnrh2, which has known anorexigenic effects. We found that the double knockout fish exhibited upregulation of several genes in the brain, specifically gonadotropin-inhibitory hormone (gnih), secretogranin 2 (scg2), tachykinin 3a (tac3a), and pituitary adenylate cyclase-activating peptide 1 (pacap1), and downregulation of agouti-related peptide 1 (agrp1), indicating the compensation occurs outside of Gnrh cells and therefore is a non-cell autonomous response to the loss of Gnrh. While the differential expression of gnih and agrp1 in the double knockout line was confined to the periventricular nucleus and hypothalamus, respectively, the upregulation of scg2 corresponded with a broader neuronal redistribution in the lateral hypothalamus and hindbrain. In conclusion, our results demonstrate the existence of a redundant reproductive regulatory system that comes into play when Gnrh2 and Gnrh3 are lost.

ATP Synthase β-Chain Overexpression in SR-BI Knockout Mice Increases HDL Uptake and Reduces Plasma HDL Level

Directory of Open Access Journals (Sweden)

Kexiu Song

2014-01-01

Full Text Available HDL cholesterol is known to be inversely correlated with cardiovascular disease due to its diverse antiatherogenic functions. SR-BI mediates the selective uptake of HDL-C. SR-BI knockout diminishes but does not completely block the transport of HDL; other receptors may be involved. Ectopic ATP synthase β-chain in hepatocytes has been previously characterized as an apoA-I receptor, triggering HDL internalization. This study was undertaken to identify the overexpression of ectopic ATP synthase β-chain on DIL-HDL uptake in primary hepatocytes in vitro and on plasma HDL levels in SR-BI knockout mice. Human ATP synthase β-chain cDNA was delivered to the mouse liver by adenovirus and GFP adenovirus as control. The adenovirus-mediated overexpression of β-chain was identified at both mRNA and protein levels on mice liver and validated by its increasing of DiL-HDL uptake in primary hepatocytes. In response to hepatic overexpression of β-chain, plasma HDL-C levels and cholesterol were reduced in SR-BI knockout mice, compared with the control. The present data suggest that ATP synthase β-chain can serve as the endocytic receptor of HDL, and its overexpression can reduce plasma HDL-C.

CD44 and Bak expression in IL-6 or TNF-alpha gene knockout mice after whole lung irradiation

International Nuclear Information System (INIS)

Sakai, Minako; Iwakawa, Mayumi; Ohta, Toshie; Tsujii, Hirohiko; Imai, Takashi; Iwakura, Yoichiro

2008-01-01

To understand the molecular mechanisms that underlie radiation pneumonitis, we examined whether knockout of the tumor necrosis factor (TNF) or the interleukin (IL)-6 gene could give mice an inherent resistance to radiation in the acute phase of alveolar damage after thoracic irradiation. The temporal expression of inflammation (CD44) and apoptosis (Bak) markers in lung after thoracic irradiation was measured to determine the degree of alveolar damage. At 4 weeks post-irradiation (10 Gy), small inflammatory foci were observed in all mice, but there were no obvious histological differences between control (C57BL/6JSlc), TNF-alpha knockout (TNF KO), and IL-6 knockout (IL-6 KO) mice. However, immunohistochemical analysis of CD44 and Bak expression over a time course of 2 weeks highlighted significant differences between the three groups. C57BL/6JSlc and TNF KO mice had increased numbers of both CD44-positive and Bak-positive cells after irradiation, while the IL-6 KO mice showed stable levels of CD44 and Bak. In conclusion, the radioresistant status of IL-6 KO mice in the acute phase of alveolar damage after irradiation suggested an important role for IL-6 in radiation pneumonitis. (author)

LRRK2 knockout mice have an intact dopaminergic system but display alterations in exploratory and motor co-ordination behaviors

Science.gov (United States)

2012-01-01

Mutations in the LRRK2 gene are the most common cause of genetic Parkinson’s disease. Although the mechanisms behind the pathogenic effects of LRRK2 mutations are still not clear, data emerging from in vitro and in vivo models suggests roles in regulating neuronal polarity, neurotransmission, membrane and cytoskeletal dynamics and protein degradation. We created mice lacking exon 41 that encodes the activation hinge of the kinase domain of LRRK2. We have performed a comprehensive analysis of these mice up to 20 months of age, including evaluation of dopamine storage, release, uptake and synthesis, behavioral testing, dendritic spine and proliferation/neurogenesis analysis. Our results show that the dopaminergic system was not functionally comprised in LRRK2 knockout mice. However, LRRK2 knockout mice displayed abnormal exploratory activity in the open-field test. Moreover, LRRK2 knockout mice stayed longer than their wild type littermates on the accelerated rod during rotarod testing. Finally, we confirm that loss of LRRK2 caused degeneration in the kidney, accompanied by a progressive enhancement of autophagic activity and accumulation of autofluorescent material, but without evidence of biphasic changes. PMID:22647713

Genetic rescue of glycosylation-deficient Fgf23 in the Galnt3 knockout mouse.

Science.gov (United States)

Ichikawa, Shoji; Gray, Amie K; Padgett, Leah R; Allen, Matthew R; Clinkenbeard, Erica L; Sarpa, Nicole M; White, Kenneth E; Econs, Michael J

2014-10-01

Fibroblast growth factor 23 (FGF23) is a hormone that inhibits renal phosphate reabsorption and 1,25-dihydroxyvitamin D biosynthesis. The FGF23 subtilisin-like proprotein convertase recognition sequence ((176)RHTR(179)↓) is protected by O-glycosylation through ppGalNAc-T3 (GALNT3) activity. Thus, inactivating GALNT3 mutations render FGF23 susceptible to proteolysis, thereby reducing circulating intact hormone levels and leading to hyperphosphatemic familial tumoral calcinosis. To further delineate the role of glycosylation in the Fgf23 function, we generated an inducible FGF23 transgenic mouse expressing human mutant FGF23 (R176Q and R179Q) found in patients with autosomal dominant hypophosphatemic rickets (ADHR) and bred this animal to Galnt3 knockout mice, a model of familial tumoral calcinosis. Due to the low intact Fgf23 level, Galnt3 knockout mice with wild-type Fgf23 alleles were hyperphosphatemic. In contrast, carriers of the mutant FGF23 transgene, regardless of Galnt3 mutation status, had significantly higher serum intact FGF23, resulting in severe hypophosphatemia. Importantly, serum phosphorus and FGF23 were comparable between transgenic mice with or without normal Galnt3 alleles. To determine whether the presence of the ADHR mutation could improve biochemical and skeletal abnormalities in Galnt3-null mice, these mice were also mated to Fgf23 knock-in mice, carrying heterozygous or homozygous R176Q ADHR Fgf23 mutations. The knock-in mice with functional Galnt3 had normal Fgf23 but were slightly hypophosphatemic. The stabilized Fgf23 ADHR allele reversed the Galnt3-null phenotype and normalized total Fgf23, serum phosphorus, and bone Fgf23 mRNA. However, the skeletal phenotype was unaffected. In summary, these data demonstrate that O-glycosylation by ppGaINAc-T3 is only necessary for proper secretion of intact Fgf23 and, once secreted, does not affect Fgf23 function. Furthermore, the more stable Fgf23 ADHR mutant protein could normalize serum phosphorus

Activation of PPARγ Ameliorates Spatial Cognitive Deficits through Restoring Expression of AMPA Receptors in Seipin Knock-Out Mice.

Science.gov (United States)

Zhou, Libin; Chen, Tingting; Li, Guoxi; Wu, Chaoming; Wang, Conghui; Li, Lin; Sha, Sha; Chen, Lei; Liu, George; Chen, Ling

2016-01-27

A characteristic phenotype of congenital generalized lipodystrophy 2 (CGL2) that is caused by loss-of-function of seipin gene is mental retardation. Here, we show that seipin deficiency in hippocampal CA1 pyramidal cells caused the reduction of peroxisome proliferator-activated receptor gamma (PPARγ). Twelve-week-old systemic seipin knock-out mice and neuronal seipin knock-out (seipin-nKO) mice, but not adipose seipin knock-out mice, exhibited spatial cognitive deficits as assessed by the Morris water maze and Y-maze, which were ameliorated by the treatment with the PPARγ agonist rosiglitazone (rosi). In addition, seipin-nKO mice showed the synaptic dysfunction and the impairment of NMDA receptor-dependent LTP in hippocampal CA1 regions. The density of AMPA-induced current (IAMPA) in CA1 pyramidal cells and GluR1/GluR2 expression were significantly reduced in seipin-nKO mice, whereas the NMDA-induced current (INMDA) and NR1/NR2 expression were not altered. Rosi treatment in seipin-nKO mice could correct the decrease in expression and activity of AMPA receptor (AMPAR) and was accompanied by recovered synaptic function and LTP induction. Furthermore, hippocampal ERK2 and CREB phosphorylation in seipin-nKO mice were reduced and this could be rescued by rosi treatment. Rosi treatment in seipin-nKO mice elevated BDNF concentration. The MEK inhibitor U0126 blocked rosi-restored AMPAR expression and LTP induction in seipin-nKO mice, but the Trk family inhibitor K252a did not. These findings indicate that the neuronal seipin deficiency selectively suppresses AMPAR expression through reducing ERK-CREB activities, leading to the impairment of LTP and spatial memory, which can be rescued by PPARγ activation. Congenital generalized lipodystrophy 2 (CGL2), caused by loss-of-function mutation of seipin gene, is characterized by mental retardation. By the generation of systemic or neuronal seipin knock-out mice, the present study provides in vivo evidence that neuronal seipin

Macrophage migration inhibitory factor (MIF) knockout preserves cardiac homeostasis through alleviating Akt-mediated myocardial autophagy suppression in high-fat diet-induced obesity.

Science.gov (United States)

Xu, X; Ren, J

2015-03-01

Macrophage migration inhibitory factor (MIF) has a role in the development of obesity and diabetes. However, whether MIF has a role in fat diet-induced obesity and associated cardiac anomalies still remains unknown. The aim of this study was to examine the impact of MIF knockout on high-fat diet-induced obesity, obesity-associated cardiac anomalies and the underlying mechanisms involved with a focus on Akt-mediated autophagy. Adult male wild-type (WT) and MIF knockout (MIF(-/-)) mice were placed on 45% high-fat diet for 5 months. Oxygen consumption, CO2 production, respiratory exchange ratio, locomotor activity and heat generation were measured using energy calorimeter. Echocardiographic, cardiomyocyte mechanical and intracellular Ca2+ properties were assessed. Apoptosis was examined using terminal dUTP nick end labeling staining and western blot analysis. Akt signaling pathway and autophagy markers were evaluated. Cardiomyocytes isolated from WT and MIF(-/-) mice were treated with recombinant mouse MIF (rmMIF). High-fat diet feeding elicited increased body weight gain, insulin resistance and caloric disturbance in WT and MIF(-/-) mice. High-fat diet induced unfavorable geometric, contractile and histological changes in the heart, the effects of which were alleviated by MIF knockout. In addition, fat diet-induced cardiac anomalies were associated with Akt activation and autophagy suppression, which were nullified by MIF deficiency. In cardiomyocytes from WT mice, autophagy was inhibited by exogenous rmMIF through Akt activation. In addition, MIF knockout rescued palmitic acid-induced suppression of cardiomyocyte autophagy, the effect of which was nullified by rmMIF. These results indicate that MIF knockout preserved obesity-associated cardiac anomalies without affecting fat diet-induced obesity, probably through restoring myocardial autophagy in an Akt-dependent manner. Our findings provide new insights for the role of MIF in obesity and associated cardiac

An inducible knockout mouse to model the cell-autonomous role of PTEN in initiating endometrial, prostate and thyroid neoplasias

Science.gov (United States)

Mirantes, Cristina; Eritja, Núria; Dosil, Maria Alba; Santacana, Maria; Pallares, Judit; Gatius, Sónia; Bergadà, Laura; Maiques, Oscar; Matias-Guiu, Xavier; Dolcet, Xavier

2013-01-01

SUMMARY PTEN is one of the most frequently mutated tumor suppressor genes in human cancers. The role of PTEN in carcinogenesis has been validated by knockout mouse models. PTEN heterozygous mice develop neoplasms in multiple organs. Unfortunately, the embryonic lethality of biallelic excision of PTEN has inhibited the study of complete PTEN deletion in the development and progression of cancer. By crossing PTEN conditional knockout mice with transgenic mice expressing a tamoxifen-inducible Cre-ERT under the control of a chicken actin promoter, we have generated a tamoxifen-inducible mouse model that allows temporal control of PTEN deletion. Interestingly, administration of a single dose of tamoxifen resulted in PTEN deletion mainly in epithelial cells, but not in stromal, mesenchymal or hematopoietic cells. Using the mT/mG double-fluorescent Cre reporter mice, we demonstrate that epithelial-specific PTEN excision was caused by differential Cre activity among tissues and cells types. Tamoxifen-induced deletion of PTEN resulted in extremely rapid and consistent formation of endometrial in situ adenocarcinoma, prostate intraepithelial neoplasia and thyroid hyperplasia. We also analyzed the role of PTEN ablation in other epithelial cells, such as the tubular cells of the kidney, hepatocytes, colonic epithelial cells or bronchiolar epithelium, but those tissues did not exhibit neoplastic growth. Finally, to validate this model as a tool to assay the efficacy of anti-tumor drugs in PTEN deficiency, we administered the mTOR inhibitor everolimus to mice with induced PTEN deletion. Everolimus dramatically reduced the progression of endometrial proliferations and significantly reduced thyroid hyperplasia. This model could be a valuable tool to study the cell-autonomous mechanisms involved in PTEN-loss-induced carcinogenesis and provides a good platform to study the effect of anti-neoplastic drugs on PTEN-negative tumors. PMID:23471917

Identification of differentially expressed proteins in spontaneous thymic lymphomas from knockout mice with deletion of p53

DEFF Research Database (Denmark)

Honoré, Bent; Buus, Søren; Claësson, Mogens H

2008-01-01

ABSTRACT: BACKGROUND: Knockout mice with a deletion of p53 spontaneously develop thymic lymphomas. Two cell lines (SM5 and SM7), established from two independent tumours, exhibited about fifty to seventy two-fold differentially expressed proteins compared to wild type thymocytes by two-dimensiona......ABSTRACT: BACKGROUND: Knockout mice with a deletion of p53 spontaneously develop thymic lymphomas. Two cell lines (SM5 and SM7), established from two independent tumours, exhibited about fifty to seventy two-fold differentially expressed proteins compared to wild type thymocytes by two...... alpha type 3, transforming acidic coiled-coil containing protein 3, mitochondrial ornithine aminotransferase and epidermal fatty acid binding protein and down-regulation of adenylosuccinate synthetase, tubulin beta-3 chain, a 25 kDa actin fragment, proteasome subunit beta type 9, cofilin-1 and glia...

Wip1 knockout inhibits the proliferation and enhances the migration of bone marrow mesenchymal stem cells

International Nuclear Information System (INIS)

Tang, Yiting; Liu, Lan; Sheng, Ming; Xiong, Kai; Huang, Lei; Gao, Qian; Wei, Jingliang; Wu, Tianwen; Yang, Shulin; Liu, Honglin; Mu, Yulian; Li, Kui

2015-01-01

Mesenchymal stem cells (MSCs), a unique population of multipotent adult progenitor cells originally found in bone marrow (BM), are extremely useful for multifunctional therapeutic approaches. However, the growth arrest and premature senescence of MSCs in vitro prevent the in-depth characterization of these cells. In addition, the regulatory factors involved in MSCs migration remain largely unknown. Given that protein phosphorylation is associated with the processes of MSCs proliferation and migration, we focused on wild-type p53-inducible phosphatase-1 (Wip1), a well-studied modulator of phosphorylation, in this study. Our results showed that Wip1 knockout significantly inhibited MSCs proliferation and induced G2-phase cell-cycle arrest by reducing cyclinB1 expression. Compared with WT-MSCs, Wip1 −/− MSCs displayed premature growth arrest after six passages in culture. Transwell and scratch assays revealed that Wip1 −/− MSCs migrate more effectively than WT-MSCs. Moreover, the enhanced migratory response of Wip1 −/− MSCs may be attributed to increases in the induction of Rac1-GTP activity, the pAKT/AKT ratio, the rearrangement of filamentous-actin (f-actin), and filopodia formation. Based on these results, we then examined the effect of treatment with a PI3K/AKT and Rac1 inhibitor, both of which impaired the migratory activity of MSCs. Therefore, we propose that the PI3K/AKT/Rac1 signaling axis mediates the Wip1 knockout-induced migration of MSCs. Our findings indicate that the principal function of Wip1 in MSCs transformation is the maintenance of proliferative capacity. Nevertheless, knocking out Wip1 increases the migratory capacity of MSCs. This dual effect of Wip1 provides the potential for purposeful routing of MSCs. - Highlights: • Wip1 knockout inhibited MSCs proliferation through reducing cyclinB1 expression. • Wip1 −/− MSCs displayed premature growth arrest in vitro after six passages. • Knocking out Wip1 increases the migratory

Wip1 knockout inhibits the proliferation and enhances the migration of bone marrow mesenchymal stem cells

Energy Technology Data Exchange (ETDEWEB)

Tang, Yiting [College of Animal Science and Technology, Nanjing Agricultural University, Nanjing 210095 (China); State Key Laboratory of Animal Nutrition and Key Laboratory of Farm Animal Genetic Resources and Germplasm Innovation of Ministry of Agriculture, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193 (China); Liu, Lan [State Key Laboratory of Animal Nutrition and Key Laboratory of Farm Animal Genetic Resources and Germplasm Innovation of Ministry of Agriculture, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193 (China); Sheng, Ming [College of Animal Science and Technology, Nanjing Agricultural University, Nanjing 210095 (China); Xiong, Kai [Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, Grønnegårdsvej 7, 1870 Frederiksberg C (Denmark); Huang, Lei; Gao, Qian; Wei, Jingliang; Wu, Tianwen; Yang, Shulin [State Key Laboratory of Animal Nutrition and Key Laboratory of Farm Animal Genetic Resources and Germplasm Innovation of Ministry of Agriculture, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193 (China); Liu, Honglin, E-mail: liuhonglinnjau@163.com [College of Animal Science and Technology, Nanjing Agricultural University, Nanjing 210095 (China); Mu, Yulian, E-mail: muyulian76@iascaas.net.cn [State Key Laboratory of Animal Nutrition and Key Laboratory of Farm Animal Genetic Resources and Germplasm Innovation of Ministry of Agriculture, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193 (China); Li, Kui [State Key Laboratory of Animal Nutrition and Key Laboratory of Farm Animal Genetic Resources and Germplasm Innovation of Ministry of Agriculture, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193 (China)

2015-06-10

Mesenchymal stem cells (MSCs), a unique population of multipotent adult progenitor cells originally found in bone marrow (BM), are extremely useful for multifunctional therapeutic approaches. However, the growth arrest and premature senescence of MSCs in vitro prevent the in-depth characterization of these cells. In addition, the regulatory factors involved in MSCs migration remain largely unknown. Given that protein phosphorylation is associated with the processes of MSCs proliferation and migration, we focused on wild-type p53-inducible phosphatase-1 (Wip1), a well-studied modulator of phosphorylation, in this study. Our results showed that Wip1 knockout significantly inhibited MSCs proliferation and induced G2-phase cell-cycle arrest by reducing cyclinB1 expression. Compared with WT-MSCs, Wip1{sup −/−} MSCs displayed premature growth arrest after six passages in culture. Transwell and scratch assays revealed that Wip1{sup −/−} MSCs migrate more effectively than WT-MSCs. Moreover, the enhanced migratory response of Wip1{sup −/−} MSCs may be attributed to increases in the induction of Rac1-GTP activity, the pAKT/AKT ratio, the rearrangement of filamentous-actin (f-actin), and filopodia formation. Based on these results, we then examined the effect of treatment with a PI3K/AKT and Rac1 inhibitor, both of which impaired the migratory activity of MSCs. Therefore, we propose that the PI3K/AKT/Rac1 signaling axis mediates the Wip1 knockout-induced migration of MSCs. Our findings indicate that the principal function of Wip1 in MSCs transformation is the maintenance of proliferative capacity. Nevertheless, knocking out Wip1 increases the migratory capacity of MSCs. This dual effect of Wip1 provides the potential for purposeful routing of MSCs. - Highlights: • Wip1 knockout inhibited MSCs proliferation through reducing cyclinB1 expression. • Wip1{sup −/−} MSCs displayed premature growth arrest in vitro after six passages. • Knocking out Wip1

RF-knockout Extraction System for the CNAO Synchrotron

CERN Document Server

Carmignani, Nicola; Serio, Mario; Balbinot, Giovanni; Bressi, Erminia; Caldara, Michele; Pullia, Marco; Bosser, Jacques; Venchi, Giuseppe

2010-01-01

The National Centre for Oncological Hadrontherapy (CNAO) is a centre in Italy for the treatment of patients affected by tumours with proton and carbon ions beams accelerated in a synchrotron. The synchrotron extraction method is based on the use of a betatron core. This work aims to verify, through a theoretical study and a simulation, the possibility of using the RF-knockout extraction method exploiting the existing hardware. A simulation program has been written to simulate the extraction system of the synchrotron with the purpose to define the parameters of the radio frequency. Two types of radio frequencies have been compared in order to obtain a constant spill with the minimum ripple: a carrier wave with a frequency and amplitude modulation, and a gaussian narrow band noise modulated in amplitude. Results of the simulation and considerations on the kicker characteristics are presented

Photogrammetry-Based Head Digitization for Rapid and Accurate Localization of EEG Electrodes and MEG Fiducial Markers Using a Single Digital SLR Camera.

Science.gov (United States)

Clausner, Tommy; Dalal, Sarang S; Crespo-García, Maité

2017-01-01

The performance of EEG source reconstruction has benefited from the increasing use of advanced head modeling techniques that take advantage of MRI together with the precise positions of the recording electrodes. The prevailing technique for registering EEG electrode coordinates involves electromagnetic digitization. However, the procedure adds several minutes to experiment preparation and typical digitizers may not be accurate enough for optimal source reconstruction performance (Dalal et al., 2014). Here, we present a rapid, accurate, and cost-effective alternative method to register EEG electrode positions, using a single digital SLR camera, photogrammetry software, and computer vision techniques implemented in our open-source toolbox, janus3D . Our approach uses photogrammetry to construct 3D models from multiple photographs of the participant's head wearing the EEG electrode cap. Electrodes are detected automatically or semi-automatically using a template. The rigid facial features from these photo-based models are then surface-matched to MRI-based head reconstructions to facilitate coregistration to MRI space. This method yields a final electrode coregistration error of 0.8 mm, while a standard technique using an electromagnetic digitizer yielded an error of 6.1 mm. The technique furthermore reduces preparation time, and could be extended to a multi-camera array, which would make the procedure virtually instantaneous. In addition to EEG, the technique could likewise capture the position of the fiducial markers used in magnetoencephalography systems to register head position.

EFFECTS OF PHYSICAL TRAINING ON THE MYOCARDIUM OF FEMALE LDL KNOCKOUT OVARIECTOMIZED MICE

OpenAIRE

Brianezi, Ledimar; Marques, Mara Rubia; Cardoso, Clever Gomes; Miranda, Maria Luiza de Jesus; Fonseca, Fernando Luiz Affonso; Maifrino, Laura Beatriz Mesiano

2017-01-01

ABSTRACT Introduction: The emergence of coronary heart disease increases with menopause, physical inactivity and with dyslipidemia. It is known that physical training promotes the improvement of cardiovascular functions. Objective: The purpose of this study was to investigate the effects of aerobic physical training on the left ventricle in female LDL knockout ovariectomized mice. Methods: Thirty animals were divided into 6 groups (n=5), namely, sedentary non-ovariectomized control; sedentary...

Glutamine synthetase gene knockout-human embryonic kidney 293E cells for stable production of monoclonal antibodies.

Science.gov (United States)

Yu, Da Young; Lee, Sang Yoon; Lee, Gyun Min

2018-05-01

Previously, it was inferred that a high glutamine synthetase (GS) activity in human embryonic kidney (HEK) 293E cells results in elevated resistance to methionine sulfoximine (MSX) and consequently hampers GS-mediated gene amplification and selection by MSX. To overcome this MSX resistance in HEK293E cells, a GS-knockout HEK293E cell line was generated using the CRISPR/Cas9 system to target the endogenous human GS gene. The GS-knockout in the HEK293E cell line (RK8) was confirmed by Western blot analysis of GS and by observation of glutamine-dependent growth. Unlike the wild type HEK293E cells, the RK8 cells were successfully used as host cells to generate a recombinant HEK293E cell line (rHEK293E) producing a monoclonal antibody (mAb). When the RK8 cells were transfected with the GS expression vector containing the mAb gene, rHEK293E cells producing the mAb could be selected in the absence as well as in the presence of MSX. The gene copies and mRNA expression levels of the mAb in rHEK293E cells were also quantified using qRT-PCR. Taken together, the GS-knockout HEK293E cell line can be used as host cells to generate stable rHEK293E cells producing a mAb through GS-mediated gene selection in the absence as well as in the presence of MSX. © 2018 Wiley Periodicals, Inc.

CRISPR/Cas9 mediated chicken Stra8 gene knockout and inhibition of male germ cell differentiation.

Directory of Open Access Journals (Sweden)

Yani Zhang

Full Text Available An efficient genome editing approach had been established to construct the stable transgenic cell lines in the domestic chicken (Gallus gallus domesticus at present. Our objectives were to investigate gene function in the differentiation process of chicken embryonic stem cells (ESCs into spermatogonial stem cells(SSCs. Three guides RNA (gRNAs were designed to knockout the Stra8 gene, and knockout efficiency was evaluated in domestic chicken cells using cleavage activity of in vitro transcription of gRNA, Luciferase-SSA assay, T7 endonuclease I assay(T7E1 and TA clone sequence. In addition, the Cas9/gRNA plasmid was transfected into ESCs to confirm the function of Stra8. SSA assay results showed that luciferase activity of the vector expressing gRNA-1 and gRNA- 2 was higher than that of gRNA-3. TA clone sequencing showed that the knockdown efficiency was 25% (10/40 in DF-1 cells, the knockdown efficiency was 23% (9/40 in chicken ESCs. T7E1 assay indicated that there were cleavage activity for three individuals, and the knockdown efficiency was 12% (3/25. Cell morphology, qRT-PCR, immunostaining and FCS indicated that Cas9/gRNA not only resulted in the knockout of Stra8 gene, but also suggested that the generation of SSCs was blocked by the Stra8 gene knockdown in vitro. Taken together, our results indicate that the CRISPR/Cas9 system could mediate stable Stra8 gene knockdown in domestic chicken's cells and inhibit ECSs differentiation into SSCs.

Medium effects on spin observables of proton knockout reactions

International Nuclear Information System (INIS)

Krein, G.; Maris, T.A.J.; Rodrigues, B.B.; Veit, E.A.

1994-07-01

Medium modifications of the properties of bound nucleons and mesons are investigated by means of medium energy quasi free proton knockout reactions with polarized incident protons. The sensitivity of the spin observables of these reactions to modifications of the nucleon and meson properties is studied using the Bonn one-boson exchange model of the nucleon-nucleon interaction. A method proposed to extract the pp analysing power in medium from the (p, 2 p) asymmetries indicates a reduction of this quantity compared to its free space value. This reduction is linked to modifications of masses and coupling constants of the nucleons and mesons in the nucleus. The implications of these modifications for another spin observable to be measured in the future are discussed. (author). 39 refs, 9 figs

Drop tests of the Three Mile Island knockout canister

International Nuclear Information System (INIS)

Box, W.D.; Aaron, W.S.; Shappert, L.B.; Childress, P.C.; Quinn, G.J.; Smith, J.V.

1987-01-01

A type of Three Mile Island Unit 2 (TMI-2) defueling canister, called a ''knockout'' canister, was subjected to a series of drop tests at the Oak Ridge National Laboratory's Drop Test Facility. These tests confirmed the structural integrity of internal fixed neutron poisons in support of a request for NRC licensing of this type of canister for the shipment of TMI-2 reactor fuel debris to the Idaho National Engineering Laboratory (INEL) for the Core Examination R and D Program. This report presents the data generated and the results obtained from a series of four drop tests that included two drops with the test assembly in the vertical position and two drops with the assembly in the horizontal position

Medium effects on spin observables of proton knockout reactions

Energy Technology Data Exchange (ETDEWEB)

Krein, G [Instituto de Fisica Teorica (IFT), Sao Paulo, SP (Brazil); Maris, T A.J.; Rodrigues, B B; Veit, E A [Rio Grande do Sul Univ., Porto Alegre, RS (Brazil). Inst. de Fisica

1994-07-01

Medium modifications of the properties of bound nucleons and mesons are investigated by means of medium energy quasi free proton knockout reactions with polarized incident protons. The sensitivity of the spin observables of these reactions to modifications of the nucleon and meson properties is studied using the Bonn one-boson exchange model of the nucleon-nucleon interaction. A method proposed to extract the pp analysing power in medium from the (p, 2 p) asymmetries indicates a reduction of this quantity compared to its free space value. This reduction is linked to modifications of masses and coupling constants of the nucleons and mesons in the nucleus. The implications of these modifications for another spin observable to be measured in the future are discussed. (author). 39 refs, 9 figs.

Altered expression and modulation of activity-regulated cytoskeletal associated protein (Arc) in serotonin transporter knockout rats.

NARCIS (Netherlands)

Molteni, R.; Calabrese, F.; Maj, P.F.; Olivier, J.D.A.; Racagni, G.; Ellenbroek, A.A.; Riva, M.A.

2009-01-01

A gene variant in the human serotonin transporter (SERT) can increase the vulnerability to mood disorders. SERT knockout animals show similarities to the human condition and represent an important tool to investigate the mechanisms underlying the pathologic condition in humans. Along this line of

Protein phosphatase 2ACα gene knock-out results in cortical atrophy through activating hippo cascade in neuronal progenitor cells.

Science.gov (United States)

Liu, Bo; Sun, Li-Hua; Huang, Yan-Fei; Guo, Li-Jun; Luo, Li-Shu

2018-02-01

Protein phosphatase 2ACα (PP2ACα), a vital member of the protein phosphatase family, has been studied primarily as a regulator for the development, growth and protein synthesis of a lot of cell types. Dysfunction of PP2ACα protein results in neurodegenerative disease; however, this finding has not been directly confirmed in the mouse model with PP2ACα gene knock-out. Therefore, in this study presented here, we generated the PP2ACα gene knock-out mouse model by the Cre-loxP targeting gene system, with the purpose to directly observe the regulatory role of PP2ACα gene in the development of mouse's cerebral cortex. We observe that knocking-out PP2ACα gene in the central nervous system (CNS) results in cortical neuronal shrinkage, synaptic plasticity impairments, and learning/memory deficits. Further study reveals that PP2ACα gene knock-out initiates Hippo cascade in cortical neuroprogenitor cells (NPCs), which blocks YAP translocation into the nuclei of NPCs. Notably, p73, directly targeted by Hippo cascade, can bind to the promoter of glutaminase2 (GLS2) that plays a dominant role in the enzymatic regulation of glutamate/glutamine cycle. Finally, we find that PP2ACα gene knock-out inhibits the glutamine synthesis through up-regulating the activity of phosphorylated-p73 in cortical NPCs. Taken together, it concludes that PP2ACα critically supports cortical neuronal growth and cognitive function via regulating the signaling transduction of Hippo-p73 cascade. And PP2ACα indirectly modulates the glutamine synthesis of cortical NPCs through targeting p73 that plays a direct transcriptional regulatory role in the gene expression of GLS2. Copyright © 2017 Elsevier Ltd. All rights reserved.

Bone phenotypes of P2 receptor knockout mice

DEFF Research Database (Denmark)

Orriss, Isabel; Syberg, Susanne; Wang, Ning

2011-01-01

The action of extracellular nucleotides is mediated by ionotropic P2X receptors and G-protein coupled P2Y receptors. The human genome contains 7 P2X and 8 P2Y receptor genes. Knockout mice strains are available for most of them. As their phenotypic analysis is progressing, bone abnormalities have...... been observed in an impressive number of these mice: distinct abnormalities in P2X7-/- mice, depending on the gene targeting construct and the genetic background, decreased bone mass in P2Y1-/- mice, increased bone mass in P2Y2-/- mice, decreased bone resorption in P2Y6-/- mice, decreased bone...... formation and bone resorption in P2Y13-/- mice. These findings demonstrate the unexpected importance of extracellular nucleotide signalling in the regulation of bone metabolism via multiple P2 receptors and distinct mechanisms involving both osteoblasts and osteoclasts....

Analysis of knockout mice suggests a role for VGF in the control of fat storage and energy expenditure

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Chakraborty Tandra

2009-10-01

Full Text Available Abstract Background Previous studies of mixed background mice have demonstrated that targeted deletion of Vgf produces a lean, hypermetabolic mouse that is resistant to diet-, lesion-, and genetically-induced obesity. To investigate potential mechanism(s and site(s of action of VGF, a neuronal and endocrine secreted protein and neuropeptide precursor, we further analyzed the metabolic phenotypes of two independent VGF knockout lines on C57Bl6 backgrounds. Results Unlike hyperactive VGF knockout mice on a mixed C57Bl6-129/SvJ background, homozygous mutant mice on a C57Bl6 background were hypermetabolic with similar locomotor activity levels to Vgf+/Vgf+ mice, during day and night cycles, indicating that mechanism(s other than hyperactivity were responsible for their increased energy expenditure. In Vgf-/Vgf- knockout mice, morphological analysis of brown and white adipose tissues (BAT and WAT indicated decreased fat storage in both tissues, and decreased adipocyte perimeter and area in WAT. Changes in gene expression measured by real-time RT-PCR were consistent with increased fatty acid oxidation and uptake in BAT, and increased lipolysis, decreased lipogenesis, and brown adipocyte differentiation in WAT, suggesting that increased sympathetic nervous system activity in Vgf-/Vgf- mice may be associated with or responsible for alterations in energy expenditure and fat storage. In addition, uncoupling protein 1 (UCP1 and UCP2 protein levels, mitochondrial number, and mitochondrial cristae density were upregulated in Vgf-/Vgf- BAT. Using immunohistochemical and histochemical techniques, we detected VGF in nerve fibers innervating BAT and Vgf promoter-driven reporter expression in cervical and thoracic spinal ganglia that project to and innervate the chest wall and tissues including BAT. Moreover, VGF peptide levels were quantified by radioimmunoassay in BAT, and were found to be down-regulated by a high fat diet. Lastly, despite being hypermetabolic

Protease activity of legumain is inhibited by an increase of cystatin E/M in the DJ-1-knockout mouse spleen, cerebrum and heart

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Takuya Yamane

2017-03-01

Full Text Available Legumain (EC 3.4.22.34 is an asparaginyl endopeptidase. Legumain activity has been detected in various mouse tissues including the kidney, spleen and epididymis. Legumain is overexpressed in the majority of human solid tumors and transcription of the legumain gene is regulated by the p53 tumor suppressor in HCT116 cells. The legumain activity is also increased under acid conditions in Alzheimer's disease brains. DJ-1/PARK7, a cancer- and Parkinson's disease-associated protein, works as a coactivator to various transcription factors, including the androgen receptor, p53, PSF, Nrf2, SREBP and RREB1. Recently, we found that legumain expression, activation and cleavage of annexin A2 are regulated by DJ-1 through p53. In this study, we found that the expression levels of legumain mRNA were increased in the cerebrum, kidney, spleen, heart, lung, epididymis, stomach, small intestine and pancreas from DJ-1-knockout mice, although legumain activity levels were decreased in the cerebrum, spleen and heart from DJ-1-knockout mice. Furthermore, we found that cystatin E/M expression was increased in the spleen, cerebrum and heart from DJ-1-knockout mice. These results suggest that reduction of legumain activity is caused by an increase of cystatin E/M expression in the spleen, cerebrum and heart from DJ-1-knockout mice.

Prion protein (PrP) gene-knockout cell lines: insight into functions of the PrP

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Sakudo, Akikazu; Onodera, Takashi

2015-01-01

Elucidation of prion protein (PrP) functions is crucial to fully understand prion diseases. A major approach to studying PrP functions is the use of PrP gene-knockout (Prnp−/−) mice. So far, six types of Prnp−/− mice have been generated, demonstrating the promiscuous functions of PrP. Recently, other PrP family members, such as Doppel and Shadoo, have been found. However, information obtained from comparative studies of structural and functional analyses of these PrP family proteins do not fully reveal PrP functions. Recently, varieties of Prnp−/− cell lines established from Prnp−/− mice have contributed to the analysis of PrP functions. In this mini-review, we focus on Prnp−/− cell lines and summarize currently available Prnp−/− cell lines and their characterizations. In addition, we introduce the recent advances in the methodology of cell line generation with knockout or knockdown of the PrP gene. We also discuss how these cell lines have provided valuable insights into PrP functions and show future perspectives. PMID:25642423

Characterization of the serotonin transporter knockout rat : A selective change in the functioning of the serotonergic system

NARCIS (Netherlands)

Homberg, J. R.; Olivier, J.D.A.; Smits, B. M. G.; Mul, J. D.; Mudde, J.; Verheul, M.; Nieuwenhuizen, O. F. M.; Cools, A. R.; Ronken, E; Cremers, Thomas; Schoffelmeere, A. N. M.; Ellenbroeik, B. A.; Cuppen, E.

2007-01-01

Serotonergic signaling is involved in many neurobiological processes and disturbed 5-HT homeostasis is implicated in a variety of psychiatric and addictive disorders. Here, we describe the functional characterization of the serotonin transporter (SERT) knockout rat model, that is generated by

Characterization of the serotonin transporter knockout rat: a selective change in the functioning of the serotonergic system.

NARCIS (Netherlands)

Homberg, J.R.; Olivier, J.D.A.; Smits, B.M.; Mul, J.D.; Mudde, J.; Verheul, M.; Nieuwenhuizen, O.F.; Cools, A.R.; Ronken, E.; Cremers, T.; Schoffelmeer, A.N.; Ellenbroek, B.A.; Cuppen, E.

2007-01-01

Serotonergic signaling is involved in many neurobiological processes and disturbed 5-HT homeostasis is implicated in a variety of psychiatric and addictive disorders. Here, we describe the functional characterization of the serotonin transporter (SERT) knockout rat model, that is generated by

Accurate phylogenetic tree reconstruction from quartets: a heuristic approach.

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Reaz, Rezwana; Bayzid, Md Shamsuzzoha; Rahman, M Sohel

2014-01-01

Supertree methods construct trees on a set of taxa (species) combining many smaller trees on the overlapping subsets of the entire set of taxa. A 'quartet' is an unrooted tree over 4 taxa, hence the quartet-based supertree methods combine many 4-taxon unrooted trees into a single and coherent tree over the complete set of taxa. Quartet-based phylogeny reconstruction methods have been receiving considerable attentions in the recent years. An accurate and efficient quartet-based method might be competitive with the current best phylogenetic tree reconstruction methods (such as maximum likelihood or Bayesian MCMC analyses), without being as computationally intensive. In this paper, we present a novel and highly accurate quartet-based phylogenetic tree reconstruction method. We performed an extensive experimental study to evaluate the accuracy and scalability of our approach on both simulated and biological datasets.

DEEP WIDEBAND SINGLE POINTINGS AND MOSAICS IN RADIO INTERFEROMETRY: HOW ACCURATELY DO WE RECONSTRUCT INTENSITIES AND SPECTRAL INDICES OF FAINT SOURCES?

Energy Technology Data Exchange (ETDEWEB)

Rau, U.; Bhatnagar, S.; Owen, F. N., E-mail: rurvashi@nrao.edu [National Radio Astronomy Observatory, Socorro, NM-87801 (United States)

2016-11-01

Many deep wideband wide-field radio interferometric surveys are being designed to accurately measure intensities, spectral indices, and polarization properties of faint source populations. In this paper, we compare various wideband imaging methods to evaluate the accuracy to which intensities and spectral indices of sources close to the confusion limit can be reconstructed. We simulated a wideband single-pointing (C-array, L-Band (1–2 GHz)) and 46-pointing mosaic (D-array, C-Band (4–8 GHz)) JVLA observation using a realistic brightness distribution ranging from 1 μ Jy to 100 mJy and time-, frequency-, polarization-, and direction-dependent instrumental effects. The main results from these comparisons are (a) errors in the reconstructed intensities and spectral indices are larger for weaker sources even in the absence of simulated noise, (b) errors are systematically lower for joint reconstruction methods (such as Multi-Term Multi-Frequency-Synthesis (MT-MFS)) along with A-Projection for accurate primary beam correction, and (c) use of MT-MFS for image reconstruction eliminates Clean-bias (which is present otherwise). Auxiliary tests include solutions for deficiencies of data partitioning methods (e.g., the use of masks to remove clean bias and hybrid methods to remove sidelobes from sources left un-deconvolved), the effect of sources not at pixel centers, and the consequences of various other numerical approximations within software implementations. This paper also demonstrates the level of detail at which such simulations must be done in order to reflect reality, enable one to systematically identify specific reasons for every trend that is observed, and to estimate scientifically defensible imaging performance metrics and the associated computational complexity of the algorithms/analysis procedures.

Quasifree knockout of proton pairs from carbon with 640 MeV protons

International Nuclear Information System (INIS)

Komarov, V.I.; Kosarev, G.I.; Netzband, D.; Mueller, H.; Stiehler, T.; Tesch, S.

1980-10-01

The direct nuclear reaction C(p,3p) at 640 MeV has been investigated in an exclusive type of experiment using scintillation counter technique. The measuring conditions have been selected according to the kinematics of quasi-free two-nucleon knockout at large momentum transfer. A phenomenological model is discussed, which is capable of describing qualitatively the dependence of the differential cross section on the opening angle of the forward emitted proton pair as well as on the energy of backward going protons. (author)

An episomal vector-based CRISPR/Cas9 system for highly efficient gene knockout in human pluripotent stem cells.

Science.gov (United States)

Xie, Yifang; Wang, Daqi; Lan, Feng; Wei, Gang; Ni, Ting; Chai, Renjie; Liu, Dong; Hu, Shijun; Li, Mingqing; Li, Dajin; Wang, Hongyan; Wang, Yongming

2017-05-24

Human pluripotent stem cells (hPSCs) represent a unique opportunity for understanding the molecular mechanisms underlying complex traits and diseases. CRISPR/Cas9 is a powerful tool to introduce genetic mutations into the hPSCs for loss-of-function studies. Here, we developed an episomal vector-based CRISPR/Cas9 system, which we called epiCRISPR, for highly efficient gene knockout in hPSCs. The epiCRISPR system enables generation of up to 100% Insertion/Deletion (indel) rates. In addition, the epiCRISPR system enables efficient double-gene knockout and genomic deletion. To minimize off-target cleavage, we combined the episomal vector technology with double-nicking strategy and recent developed high fidelity Cas9. Thus the epiCRISPR system offers a highly efficient platform for genetic analysis in hPSCs.

Comparative effects of chlorpyrifos in wild type and cannabinoid Cb1 receptor knockout mice

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Baireddy, Praveena; Liu, Jing; Hinsdale, Myron; Pope, Carey, E-mail: carey.pope@okstate.edu

2011-11-15

Endocannabinoids (eCBs) modulate neurotransmission by inhibiting the release of a variety of neurotransmitters. The cannabinoid receptor agonist WIN 55.212-2 (WIN) can modulate organophosphorus (OP) anticholinesterase toxicity in rats, presumably by inhibiting acetylcholine (ACh) release. Some OP anticholinesterases also inhibit eCB-degrading enzymes. We studied the effects of the OP insecticide chlorpyrifos (CPF) on cholinergic signs of toxicity, cholinesterase activity and ACh release in tissues from wild type (+/+) and cannabinoid CB1 receptor knockout (-/-) mice. Mice of both genotypes (n = 5-6/treatment group) were challenged with CPF (300 mg/kg, 2 ml/kg in peanut oil, sc) and evaluated for functional and neurochemical changes. Both genotypes exhibited similar cholinergic signs and cholinesterase inhibition (82-95% at 48 h after dosing) in cortex, cerebellum and heart. WIN reduced depolarization-induced ACh release in vitro in hippocampal slices from wild type mice, but had no effect in hippocampal slices from knockouts or in striatal slices from either genotype. Chlorpyrifos oxon (CPO, 100 {mu}M) reduced release in hippocampal slices from both genotypes in vitro, but with a greater reduction in tissues from wild types (21% vs 12%). CPO had no significant in vitro effect on ACh release in striatum. CPF reduced ACh release in hippocampus from both genotypes ex vivo, but reduction was again significantly greater in tissues from wild types (52% vs 36%). In striatum, CPF led to a similar reduction (20-23%) in tissues from both genotypes. Thus, while CB1 deletion in mice had little influence on the expression of acute toxicity following CPF, CPF- or CPO-induced changes in ACh release appeared sensitive to modulation by CB1-mediated eCB signaling in a brain-regional manner. -- Highlights: Black-Right-Pointing-Pointer C57Bl/6 mice showed dose-related cholinergic toxicity following subcutaneous chlorpyrifos exposure. Black-Right-Pointing-Pointer Wild type and

Comparative effects of chlorpyrifos in wild type and cannabinoid Cb1 receptor knockout mice

International Nuclear Information System (INIS)

Baireddy, Praveena; Liu, Jing; Hinsdale, Myron; Pope, Carey

2011-01-01

Endocannabinoids (eCBs) modulate neurotransmission by inhibiting the release of a variety of neurotransmitters. The cannabinoid receptor agonist WIN 55.212-2 (WIN) can modulate organophosphorus (OP) anticholinesterase toxicity in rats, presumably by inhibiting acetylcholine (ACh) release. Some OP anticholinesterases also inhibit eCB-degrading enzymes. We studied the effects of the OP insecticide chlorpyrifos (CPF) on cholinergic signs of toxicity, cholinesterase activity and ACh release in tissues from wild type (+/+) and cannabinoid CB1 receptor knockout (−/−) mice. Mice of both genotypes (n = 5–6/treatment group) were challenged with CPF (300 mg/kg, 2 ml/kg in peanut oil, sc) and evaluated for functional and neurochemical changes. Both genotypes exhibited similar cholinergic signs and cholinesterase inhibition (82–95% at 48 h after dosing) in cortex, cerebellum and heart. WIN reduced depolarization-induced ACh release in vitro in hippocampal slices from wild type mice, but had no effect in hippocampal slices from knockouts or in striatal slices from either genotype. Chlorpyrifos oxon (CPO, 100 μM) reduced release in hippocampal slices from both genotypes in vitro, but with a greater reduction in tissues from wild types (21% vs 12%). CPO had no significant in vitro effect on ACh release in striatum. CPF reduced ACh release in hippocampus from both genotypes ex vivo, but reduction was again significantly greater in tissues from wild types (52% vs 36%). In striatum, CPF led to a similar reduction (20–23%) in tissues from both genotypes. Thus, while CB1 deletion in mice had little influence on the expression of acute toxicity following CPF, CPF- or CPO-induced changes in ACh release appeared sensitive to modulation by CB1-mediated eCB signaling in a brain-regional manner. -- Highlights: ► C57Bl/6 mice showed dose-related cholinergic toxicity following subcutaneous chlorpyrifos exposure. ► Wild type and cannabinoid CB1 receptor knockout littermates

Increased Contextual Fear Conditioning in iNOS Knockout Mice: Additional Evidence for the Involvement of Nitric Oxide in Stress-Related Disorders and Contribution of the Endocannabinoid System

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Gomes, Felipe V.; Silva, Andréia L.; Uliana, Daniela L.; Camargo, Laura H. A.; Guimarães, Francisco S.; Cunha, Fernando Q.; Joca, Sâmia R. L.; Resstel, Leonardo B. M.

2015-01-01

Background: Inducible or neuronal nitric oxide synthase gene deletion increases or decreases anxiety-like behavior in mice, respectively. Since nitric oxide and endocannabinoids interact to modulate defensive behavior, the former effect could involve a compensatory increase in basal brain nitric oxide synthase activity and/or changes in the endocannabinoid system. Thus, we investigated the expression and extinction of contextual fear conditioning of inducible nitric oxide knockout mice and possible involvement of endocannabinoids in these responses. Methods: We evaluated the effects of a preferential neuronal nitric oxide synthase inhibitor, 7-nitroindazol, nitric oxide synthase activity, and mRNA changes of nitrergic and endocannabinoid systems components in the medial prefrontal cortex and hippocampus of wild-type and knockout mice. The effects of URB597, an inhibitor of the fatty acid amide hydrolase enzyme, which metabolizes the endocannabinoid anandamide, WIN55,212-2, a nonselective cannabinoid agonist, and AM281, a selective CB1 antagonist, on contextual fear conditioning were also evaluated. Results: Contextual fear conditioning expression was similar in wild-type and knockout mice, but the latter presented extinction deficits and increased basal nitric oxide synthase activity in the medial prefrontal cortex. 7-Nitroindazol decreased fear expression and facilitated extinction in wild-type and knockout mice. URB597 decreased fear expression in wild-type and facilitated extinction in knockout mice, whereas WIN55,212-2 and AM281 increased it in wild-type mice. Nonconditioned knockout mice showed changes in the mRNA expression of nitrergic and endocannabinoid system components in the medial prefrontal cortex and hippocampus that were modified by fear conditioning. Conclusion: These data reinforce the involvement of the nitric oxide and endocannabinoids (anandamide) in stress-related disorders and point to a deregulation of the endocannabinoid system in

Defining the range of pathogens susceptible to Ifitm3 restriction using a knockout mouse model.

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Aaron R Everitt

Full Text Available The interferon-inducible transmembrane (IFITM family of proteins has been shown to restrict a broad range of viruses in vitro and in vivo by halting progress through the late endosomal pathway. Further, single nucleotide polymorphisms (SNPs in its sequence have been linked with risk of developing severe influenza virus infections in humans. The number of viruses restricted by this host protein has continued to grow since it was first demonstrated as playing an antiviral role; all of which enter cells via the endosomal pathway. We therefore sought to test the limits of antimicrobial restriction by Ifitm3 using a knockout mouse model. We showed that Ifitm3 does not impact on the restriction or pathogenesis of bacterial (Salmonella typhimurium, Citrobacter rodentium, Mycobacterium tuberculosis or protozoan (Plasmodium berghei pathogens, despite in vitro evidence. However, Ifitm3 is capable of restricting respiratory syncytial virus (RSV in vivo either through directly restricting RSV cell infection, or by exerting a previously uncharacterised function controlling disease pathogenesis. This represents the first demonstration of a virus that enters directly through the plasma membrane, without the need for the endosomal pathway, being restricted by the IFITM family; therefore further defining the role of these antiviral proteins.

Brain response to traumatic brain injury in wild-type and interleukin-6 knockout mice: a microarray analysis

DEFF Research Database (Denmark)

Poulsen, Christian Bjørn; Penkowa, Milena; Borup, Rehannah

2005-01-01

Traumatic injury to the brain is one of the leading causes of injury-related death or disability. Brain response to injury is orchestrated by cytokines, such as interleukin (IL)-6, but the full repertoire of responses involved is not well known. We here report the results obtained with microarrays...... in wild-type and IL-6 knockout mice subjected to a cryolesion of the somatosensorial cortex and killed at 0, 1, 4, 8 and 16 days post-lesion. Overall gene expression was analyzed by using Affymetrix genechips/oligonucleotide arrays with approximately 12,400 probe sets corresponding to approximately 10...... in the initial tissue injury and later regeneration of the parenchyma. IL-6 deficiency showed a dramatic effect in the expression of many genes, especially in the 1 day post-lesion timing, which presumably underlies the poor capacity of IL-6 knockout mice to cope with brain damage. The results highlight...

Phenotypic screening of hepatocyte nuclear factor (HNF) 4-γ receptor knockout mice

International Nuclear Information System (INIS)

Gerdin, Anna Karin; Surve, Vikas V.; Joensson, Marie; Bjursell, Mikael; Bjoerkman, Maria; Edenro, Anne; Schuelke, Meint; Saad, Alaa; Bjurstroem, Sivert; Lundgren, Elisabeth Jensen; Snaith, Michael; Fransson-Steen, Ronny; Toernell, Jan; Berg, Anna-Lena; Bohlooly-Y, Mohammad

2006-01-01

Using the mouse as a model organism in pharmaceutical research presents unique advantages as its physiology in many ways resembles the human physiology, it also has a relatively short generation time, low breeding and maintenance costs, and is available in a wide variety of inbred strains. The ability to genetically modify mouse embryonic stem cells to generate mouse models that better mimic human disease is another advantage. In the present study, a comprehensive phenotypic screening protocol is applied to elucidate the phenotype of a novel mouse knockout model of hepatocyte nuclear factor (HNF) 4-γ. HNF4-γ is expressed in the kidneys, gut, pancreas, and testis. First level of the screen is aimed at general health, morphologic appearance, normal cage behaviour, and gross neurological functions. The second level of the screen looks at metabolic characteristics and lung function. The third level of the screen investigates behaviour more in-depth and the fourth level consists of a thorough pathological characterisation, blood chemistry, haematology, and bone marrow analysis. When compared with littermate wild-type mice (HNF4-γ +/+ ), the HNF4-γ knockout (HNF4-γ -/- ) mice had lowered energy expenditure and locomotor activity during night time that resulted in a higher body weight despite having reduced intake of food and water. HNF4-γ -/- mice were less inclined to build nest and were found to spend more time in a passive state during the forced swim test

A Knockout Experiment: Disciplinary Divides and Experimental Skill in Animal Behaviour Genetics.

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Nelson, Nicole C

2015-07-01

In the early 1990s, a set of new techniques for manipulating mouse DNA allowed researchers to 'knock out' specific genes and observe the effects of removing them on a live mouse. In animal behaviour genetics, questions about how to deploy these techniques to study the molecular basis of behaviour became quite controversial, with a number of key methodological issues dissecting the interdisciplinary research field along disciplinary lines. This paper examines debates that took place during the 1990s between a predominately North American group of molecular biologists and animal behaviourists around how to design, conduct, and interpret behavioural knockout experiments. Drawing from and extending Harry Collins's work on how research communities negotiate what counts as a 'well-done experiment,' I argue that the positions practitioners took on questions of experimental skill reflected not only the experimental traditions they were trained in but also their differing ontological and epistemological commitments. Different assumptions about the nature of gene action, eg., were tied to different positions in the knockout mouse debates on how to implement experimental controls. I conclude by showing that examining representations of skill in the context of a community's knowledge commitments sheds light on some of the contradictory ways in which contemporary animal behaviour geneticists talk about their own laboratory work as a highly skilled endeavour that also could be mechanised, as easy to perform and yet difficult to perform well.

The time point of β-catenin knockout in hepatocytes determines their response to xenobiotic activation of the constitutive androstane receptor

International Nuclear Information System (INIS)

Ganzenberg, Katrin; Singh, Yasmin; Braeuning, Albert

2013-01-01

The constitutive androstane receptor (CAR) controls the expression of drug-metabolizing enzymes and regulates hepatocyte proliferation. Studies with transgenic mice with an early postnatal conditional hepatocyte-specific knockout of the β-catenin gene Ctnnb1 revealed that β-catenin deficiency decreases the magnitude of induction of drug-metabolizing enzymes by CAR activators, abrogates zonal differences in the hepatocytes’ susceptibility to these compounds, and impacts on hepatocyte proliferation. These data, however, do not allow distinguishing between effects caused by β-catenin deficiency during postnatal liver development and acute effects of β-catenin deficiency in the adult animal at the time point of CAR activation. Therefore, CAR activation was now studied in a different mouse model allowing for the hepatocyte-specific knockout of β-catenin in adult mice. Treatment of these mice with 3 mg/kg body weight of the model CAR activator 1,4-bis-[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP) confirmed previous findings related to the coordinate regulation of drug metabolism by β-catenin and CAR. More importantly, the present study clarified that the impact of β-catenin signaling on CAR-mediated enzyme induction in the liver is not merely due to developmental defects caused by a postnatal lack of β-catenin, but depends on the presence of β-catenin at the time point of xenobiotic treatment. The study also revealed interesting differences between the two mouse models: hepatic zonation of TCPOBOP-dependent induction of drug-metabolizing enzymes was restored in mice with late knockout of β-catenin, and the strong proliferative response of female mice was exclusively abolished when using animals with a late β-catenin knockout. This suggests a β-catenin-dependent postnatal priming of hepatocytes during postnatal liver development, later affecting the proliferative response of adult animals to CAR-activating xenobiotics

Knockout of the interleukin-36 receptor protects against renal ischemia-reperfusion injury by reduction of proinflammatory cytokines.

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Nishikawa, Hirofumi; Taniguchi, Yoshinori; Matsumoto, Tatsuki; Arima, Naoki; Masaki, Mamoru; Shimamura, Yoshiko; Inoue, Kosuke; Horino, Taro; Fujimoto, Shimpei; Ohko, Kentaro; Komatsu, Toshihiro; Udaka, Keiko; Sano, Shigetoshi; Terada, Yoshio

2018-03-01

IL-36, a newly named member of the IL-1 cytokine family, includes 3 isoforms, IL-36α, IL-36β, and IL-36γ, all of which bind to a heterodimer containing the IL-36 receptor (IL-36R). Little is known about the role of the IL-36 axis in acute kidney injury (AKI) pathogenesis. Therefore, we evaluated IL-36 function in the bilateral renal ischemia-reperfusion injury model of AKI using IL-36R knockout and wild-type mice. IL-36R was found to be expressed in the kidney, mainly in proximal tubules. In IL-36R knockout mice, plasma creatinine, blood urea nitrogen, and IL-6 levels after ischemia-reperfusion injury were significantly lower than those in wild-type mice. Immunohistological analysis revealed mild tubular injury. IL-36α/β/γ levels were increased after ischemia-reperfusion injury, and IL-36α was expressed in lymphocytes and proximal tubular cells, but post-ischemia-reperfusion injury mRNA levels of IL-6 and TNF-α were low in IL-36R knockout mice. In primary cultures of renal tubular epithelial cells, IL-36α treatment upregulated NF-κB activity and Erk phosphorylation. Notably, in patients with AKI, urine IL-36α levels were increased, and IL-36α staining in renal biopsy samples was enhanced. Thus, IL-36α/IL-36R blockage could serve as a potential therapeutic target in AKI. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Genetic background can result in a marked or minimal effect of gene knockout (GPR55 and CB2 receptor in experimental autoimmune encephalomyelitis models of multiple sclerosis.

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Sofia Sisay

Full Text Available Endocannabinoids and some phytocannabinoids bind to CB1 and CB2 cannabinoid receptors, transient receptor potential vanilloid one (TRPV1 receptor and the orphan G protein receptor fifty-five (GPR55. Studies using C57BL/10 and C57BL/6 (Cnr2 (tm1Zim CB2 cannabinoid receptor knockout mice have demonstrated an immune-augmenting effect in experimental autoimmune encephalomyelitis (EAE models of multiple sclerosis. However, other EAE studies in Biozzi ABH mice often failed to show any treatment effect of either CB2 receptor agonism or antagonism on inhibition of T cell autoimmunity. The influence of genetic background on the induction of EAE in endocannabinoid system-related gene knockout mice was examined. It was found that C57BL/6.GPR55 knockout mice developed less severe disease, notably in female mice, following active induction with myelin oligodendrocyte glycoprotein 35-55 peptide. In contrast C57BL/6.CB2 (Cnr2 (Dgen receptor knockout mice developed augmented severity of disease consistent with the genetically and pharmacologically-distinct, Cnr2 (tm1Zim mice. However, when the knockout gene was bred into the ABH mouse background and EAE induced with spinal cord autoantigens the immune-enhancing effect of CB2 receptor deletion was lost. Likewise CB1 receptor and transient receptor potential vanilloid one knockout mice on the ABH background demonstrated no alteration in immune-susceptibility, in terms of disease incidence and severity of EAE, in contrast to that reported in some C57BL/6 mouse studies. Furthermore the immune-modulating influence of GPR55 was marginal on the ABH mouse background. Whilst sedative doses of tetrahydrocannabinol could induce immunosuppression, this was associated with a CB1 receptor rather than a CB2 receptor-mediated effect. These data support the fact that non-psychoactive doses of medicinal cannabis have a marginal influence on the immune response in MS. Importantly, it adds a note of caution for the translational

Proteomic analysis of tissue from α1,3-galactosyltransferase knockout mice reveals that a wide variety of proteins and protein fragments change expression level.

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Louise Thorlacius-Ussing

Full Text Available A barrier in a pig-to-man xenotransplantation is that the Galα1-3Galβ1-4GlcNAc-R carbohydrate (α-Gal epitope expressed on pig endothelial cells reacts with naturally occurring antibodies in the recipient's blood leading to rejection. Deletion of the α1,3-galactosyltransferase gene prevents the synthesis of the α-Gal epitope. Therefore, knockout models of the α1,3-galactosyltransferase gene are widely used to study xenotransplantation. We have performed proteomic studies on liver and pancreas tissues from wild type and α1,3-galactosyltransferase gene knockout mice. The tissues were analyzed by two-dimensional polyacrylamide gel electrophoresis and liquid chromatography-tandem mass spectrometry. The analyses revealed that a wide variety of proteins and protein fragments are differentially expressed suggesting that knockout of the α1,3-galactosyltransferase gene affects the expression of several other genes.

The importance of accurate meteorological input fields and accurate planetary boundary layer parameterizations, tested against ETEX-1

International Nuclear Information System (INIS)

Brandt, J.; Ebel, A.; Elbern, H.; Jakobs, H.; Memmesheimer, M.; Mikkelsen, T.; Thykier-Nielsen, S.; Zlatev, Z.

1997-01-01

Atmospheric transport of air pollutants is, in principle, a well understood process. If information about the state of the atmosphere is given in all details (infinitely accurate information about wind speed, etc.) and infinitely fast computers are available then the advection equation could in principle be solved exactly. This is, however, not the case: discretization of the equations and input data introduces some uncertainties and errors in the results. Therefore many different issues have to be carefully studied in order to diminish these uncertainties and to develop an accurate transport model. Some of these are e.g. the numerical treatment of the transport equation, accuracy of the mean meteorological input fields and parameterizations of sub-grid scale phenomena (as e.g. parameterizations of the 2 nd and higher order turbulence terms in order to reach closure in the perturbation equation). A tracer model for studying transport and dispersion of air pollution caused by a single but strong source is under development. The model simulations from the first ETEX release illustrate the differences caused by using various analyzed fields directly in the tracer model or using a meteorological driver. Also different parameterizations of the mixing height and the vertical exchange are compared. (author)

miR-132/212 knockout mice reveal roles for these miRNAs in regulating cortical synaptic transmission and plasticity.

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Judit Remenyi

Full Text Available miR-132 and miR-212 are two closely related miRNAs encoded in the same intron of a small non-coding gene, which have been suggested to play roles in both immune and neuronal function. We describe here the generation and initial characterisation of a miR-132/212 double knockout mouse. These mice were viable and fertile with no overt adverse phenotype. Analysis of innate immune responses, including TLR-induced cytokine production and IFNβ induction in response to viral infection of primary fibroblasts did not reveal any phenotype in the knockouts. In contrast, the loss of miR-132 and miR-212, while not overtly affecting neuronal morphology, did affect synaptic function. In both hippocampal and neocortical slices miR-132/212 knockout reduced basal synaptic transmission, without affecting paired-pulse facilitation. Hippocampal long-term potentiation (LTP induced by tetanic stimulation was not affected by miR-132/212 deletion, whilst theta burst LTP was enhanced. In contrast, neocortical theta burst-induced LTP was inhibited by loss of miR-132/212. Together these results indicate that miR-132 and/or miR-212 play a significant role in synaptic function, possibly by regulating the number of postsynaptic AMPA receptors under basal conditions and during activity-dependent synaptic plasticity.

Functional categorization of gene expression changes in the cerebellum of a Cln3-knockout mouse model for Batten disease.

Science.gov (United States)

Brooks, Andrew I; Chattopadhyay, Subrata; Mitchison, Hannah M; Nussbaum, Robert L; Pearce, David A

2003-01-01

Juvenile neuronal ceroid lipofuscinosis (JNCL or Batten Disease) is the most common progressive neurodegenerative disorder of childhood. The disease is inherited in an autosomal recessive manner and is the result of mutations in the CLN3 gene. One brain region severely affected in Batten disease is the cerebellum. Using a mouse model for Batten disease which shares pathological similarities to the disease in humans we have used oligonucleotide arrays to profile approximately 19000 mRNAs in the cerebellum. We have identified reproducible changes of twofold or more in the expression of 756 gene products in the cerebellum of 10-week-old Cln3-knockout mice as compared to wild-type controls. We have subsequently divided these genes with altered expression into 14 functional categories. We report a significant alteration in expression of genes associated with neurotransmission, neuronal cell structure and development, immune response and inflammation, and lipid metabolism. An apparent shift in metabolism toward gluconeogenesis is also evident in Cln3-knockout mice. Further experimentation will be necessary to understand the contribution of these changes in expression to a disease state. Detailed analysis of the functional consequences of altered expression of genes in the cerebellum of the Cln3-knockout mice may provide valuable clues in understanding the molecular basis of the pathological mechanisms underlying Batten disease.

Gene knockout and overexpression analysis revealed the role of N-acetylmuramidase in autolysis of Lactobacillus delbrueckii subsp. bulgaricus ljj-6.

Directory of Open Access Journals (Sweden)

Xiao-Yang Pang

Full Text Available Autolysis of lactic acid bacteria (LAB plays a vital role in dairy processing. During cheese making, autolysis of LAB affects cheese flavor development through release of intracellular enzymes and restricts the proliferation of cells in yogurt fermentation and probiotics production. In order to explore the mechanism of autolysis, the gene for the autolytic enzymes of L. bulgaricus, N-acetylmuramidase (mur, was cloned and sequenced (GenBank accession number: KF157911. Mur gene overexpression and gene knockout vectors were constructed based on pMG76e and pUC19 vectors. Recombinant plasmids were transformed into L. bulgaricus ljj-6 by electroporation, then three engineered strains with pMG76e-mur vector and fifteen engineered strains with pUC19-mur::EryBII were screened. The autolysis of the mur knockout strain was significantly lower and autolysis of the mur overexpressed strain was significantly higher compared with that of the wild type strain ljj-6. This result suggested that the mur gene played an important role in autolysis of L. bulgaricus. On the other hand, autolytic activity in a low degree was still observed in the mur knockout strain, which implied that other enzymes but autolysin encoded by mur were also involved in autolysis of L. bulgaricus.

Excited states of virtual clusters in a nucleus and the processes of quasi-elastic cluster knock-out at high energies

International Nuclear Information System (INIS)

Golovanova, N.F.; Il'in, I.M.; Neudatchin, V.G.; Smirnov, Yu.F.; Tchuvil'sky, Yu.M.

1976-01-01

The quasi-elastic knock-out of nucleon clusters from nuclei by an incident high-energy hadron is considered within the framework of the Glauber-Sitenko multiple scattering theory. It is shown that the significant contribution to the cross section for the process comes not only from the hadron elastic scattering by a nonexcited virtual cluster but also from collisions with an excited virtual cluster, accompanied by de-excitation of this cluster. This necessitates modification of the usual theory of quasi-elastic cluster knock-out. First, the angular correlations of the knocked-out cluster and scattered hadron are no longer determined by the momentum distribution of the cluster in the nucleus. They are determined by another form factor F(q) which can be called the modified momentum distribution. Secondly, the meaning and values of the effective numbers of clusters Nsup(eff) have been changed. Thirdly, the characteristics of the processes depend not only on the modulus of momentum q, which the cluster had in the nucleus, but also on its direction relative to an incident beam. A method has been developed for the calculation of the fractional parentage coefficients, which are necessary for the calculation of the cluster knock-out from the p-shell nuclei. (Auth.)

Gene knockout and overexpression analysis revealed the role of N-acetylmuramidase in autolysis of Lactobacillus delbrueckii subsp. bulgaricus ljj-6.

Science.gov (United States)

Pang, Xiao-Yang; Cui, Wen-Ming; Liu, Lu; Zhang, Shu-Wen; Lv, Jia-Ping

2014-01-01

Autolysis of lactic acid bacteria (LAB) plays a vital role in dairy processing. During cheese making, autolysis of LAB affects cheese flavor development through release of intracellular enzymes and restricts the proliferation of cells in yogurt fermentation and probiotics production. In order to explore the mechanism of autolysis, the gene for the autolytic enzymes of L. bulgaricus, N-acetylmuramidase (mur), was cloned and sequenced (GenBank accession number: KF157911). Mur gene overexpression and gene knockout vectors were constructed based on pMG76e and pUC19 vectors. Recombinant plasmids were transformed into L. bulgaricus ljj-6 by electroporation, then three engineered strains with pMG76e-mur vector and fifteen engineered strains with pUC19-mur::EryBII were screened. The autolysis of the mur knockout strain was significantly lower and autolysis of the mur overexpressed strain was significantly higher compared with that of the wild type strain ljj-6. This result suggested that the mur gene played an important role in autolysis of L. bulgaricus. On the other hand, autolytic activity in a low degree was still observed in the mur knockout strain, which implied that other enzymes but autolysin encoded by mur were also involved in autolysis of L. bulgaricus.

Gene knockout of tau expression does not contribute to the pathogenesis of prion disease.

Science.gov (United States)

Lawson, Victoria A; Klemm, Helen M; Welton, Jeremy M; Masters, Colin L; Crouch, Peter; Cappai, Roberto; Ciccotosto, Giuseppe D

2011-11-01

Prion diseases or transmissible spongiform encephalopathies are a group of fatal and transmissible disorders affecting the central nervous system of humans and animals. The principal agent of prion disease transmission and pathogenesis is proposed to be an abnormal protease-resistant isoform of the normal cellular prion protein. The microtubule-associated protein tau is elevated in patients with Creutzfeldt-Jakob disease. To determine whether tau expression contributes to prion disease pathogenesis, tau knockout and control wild-type mice were infected with the M1000 strain of mouse-adapted human prions. Immunohistochemical analysis for total tau expression in prion-infected wild-type mice indicated tau aggregation in the cytoplasm of a subpopulation of neurons in regions associated with spongiform change. Western immunoblot analysis of brain homogenates revealed a decrease in total tau immunoreactivity and epitope-specific changes in tau phosphorylation. No significant difference in incubation period or other disease features were observed between tau knockout and wild-type mice with clinical prion disease. These results demonstrate that, in this model of prion disease, tau does not contribute to the pathogenesis of prion disease and that changes in the tau protein profile observed in mice with clinical prion disease occurs as a consequence of the prion-induced pathogenesis.

Loss of CO2 sensing by the olfactory system of CNGA3 knockout mice

Directory of Open Access Journals (Sweden)

Jinlong HAN, Minmin LUO

2010-12-01

Full Text Available Atmospheric CO2 can signal the presence of food, predators or environmental stress and trigger stereotypical behaviors in both vertebrates and invertebrates. Recent studies have shown that the necklace olfactory system in mice sensitively detects CO2 in the air. Olfactory CO2 neurons are believed to rely on cyclic guanosine monophosphate (cGMP as the key second messenger; however, the specific ion channel underlying CO­2 responses remains unclear. Here we show that CO2-evoked neuronal and behavioral responses require cyclic nucleotide-gated (CNG channels consisting of the CNGA3 subunit. Through Ca2+-imaging, we found that CO2-triggered Ca2+ influx was abolished in necklace olfactory sensory neurons (OSNs of CNGA3-knockout mice. Olfactory detection tests using a Go/No-go paradigm showed that these knockout mice failed to detect 0.5% CO2. Thus, sensitive detection of atmospheric CO2 depends on the function of CNG channels consisting of the CNGA3 subunit in necklace OSNs. These data support the important role of the necklace olfactory system in CO2 sensing and extend our understanding of the signal transduction pathway mediating CO2 detection in mammals [Current Zoology 56 (6: 793–799, 2010].

p21WAF1/Cip1/Sdi1 knockout mice respond to doxorubicin with reduced cardiotoxicity

International Nuclear Information System (INIS)

Terrand, Jerome; Xu, Beibei; Morrissy, Steve; Dinh, Thai Nho; Williams, Stuart; Chen, Qin M.

2011-01-01

Doxorubicin (Dox) is an antineoplastic agent that can cause cardiomyopathy in humans and experimental animals. As an inducer of reactive oxygen species and a DNA damaging agent, Dox causes elevated expression of p21 WAF1/Cip1/Sdi1 (p21) gene. Elevated levels of p21 mRNA and p21 protein have been detected in the myocardium of mice following Dox treatment. With chronic treatment of Dox, wild type (WT) animals develop cardiomyopathy evidenced by elongated nuclei, mitochondrial swelling, myofilamental disarray, reduced cardiac output, reduced ejection fraction, reduced left ventricular contractility, and elevated expression of ANF gene. In contrast, p21 knockout (p21KO) mice did not show significant changes in the same parameters in response to Dox treatment. In an effort to understand the mechanism of the resistance against Dox induced cardiomyopathy, we measured levels of antioxidant enzymes and found that p21KO mice did not contain elevated basal or inducible levels of glutathione peroxidase and catalase. Measurements of 6 circulating cytokines indicated elevation of IL-6, IL-12, IFNγ and TNFα in Dox treated WT mice but not p21KO mice. Dox induced elevation of IL-6 mRNA was detected in the myocardium of WT mice but not p21KO mice. While the mechanism of the resistance against Dox induced cardiomyopathy remains unclear, lack of inflammatory response may contribute to the observed cardiac protection in p21KO mice. -- Highlights: ► Doxorubicin induces p21 elevation in the myocardium. ► Doxorubicin causes dilated cardiomyopathy in wild type mice. ► p21 Knockout mice are resistant against doxorubicin induced cardiomyopathy. ► Lack of inflammatory response correlates with the resistance in p21 knockout mice.

DNA barcode data accurately assign higher spider taxa

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Jonathan A. Coddington

2016-07-01

Full Text Available The use of unique DNA sequences as a method for taxonomic identification is no longer fundamentally controversial, even though debate continues on the best markers, methods, and technology to use. Although both existing databanks such as GenBank and BOLD, as well as reference taxonomies, are imperfect, in best case scenarios “barcodes” (whether single or multiple, organelle or nuclear, loci clearly are an increasingly fast and inexpensive method of identification, especially as compared to manual identification of unknowns by increasingly rare expert taxonomists. Because most species on Earth are undescribed, a complete reference database at the species level is impractical in the near term. The question therefore arises whether unidentified species can, using DNA barcodes, be accurately assigned to more inclusive groups such as genera and families—taxonomic ranks of putatively monophyletic groups for which the global inventory is more complete and stable. We used a carefully chosen test library of CO1 sequences from 49 families, 313 genera, and 816 species of spiders to assess the accuracy of genus and family-level assignment. We used BLAST queries of each sequence against the entire library and got the top ten hits. The percent sequence identity was reported from these hits (PIdent, range 75–100%. Accurate assignment of higher taxa (PIdent above which errors totaled less than 5% occurred for genera at PIdent values >95 and families at PIdent values ≥ 91, suggesting these as heuristic thresholds for accurate generic and familial identifications in spiders. Accuracy of identification increases with numbers of species/genus and genera/family in the library; above five genera per family and fifteen species per genus all higher taxon assignments were correct. We propose that using percent sequence identity between conventional barcode sequences may be a feasible and reasonably accurate method to identify animals to family/genus. However

A Generic Simulation Approach for the Fast and Accurate Estimation of the Outage Probability of Single Hop and Multihop FSO Links Subject to Generalized Pointing Errors

KAUST Repository

Ben Issaid, Chaouki; Park, Kihong; Alouini, Mohamed-Slim

2017-01-01

When assessing the performance of the free space optical (FSO) communication systems, the outage probability encountered is generally very small, and thereby the use of nave Monte Carlo simulations becomes prohibitively expensive. To estimate these rare event probabilities, we propose in this work an importance sampling approach which is based on the exponential twisting technique to offer fast and accurate results. In fact, we consider a variety of turbulence regimes, and we investigate the outage probability of FSO communication systems, under a generalized pointing error model based on the Beckmann distribution, for both single and multihop scenarios. Selected numerical simulations are presented to show the accuracy and the efficiency of our approach compared to naive Monte Carlo.

A Generic Simulation Approach for the Fast and Accurate Estimation of the Outage Probability of Single Hop and Multihop FSO Links Subject to Generalized Pointing Errors

KAUST Repository

Ben Issaid, Chaouki

2017-07-28

When assessing the performance of the free space optical (FSO) communication systems, the outage probability encountered is generally very small, and thereby the use of nave Monte Carlo simulations becomes prohibitively expensive. To estimate these rare event probabilities, we propose in this work an importance sampling approach which is based on the exponential twisting technique to offer fast and accurate results. In fact, we consider a variety of turbulence regimes, and we investigate the outage probability of FSO communication systems, under a generalized pointing error model based on the Beckmann distribution, for both single and multihop scenarios. Selected numerical simulations are presented to show the accuracy and the efficiency of our approach compared to naive Monte Carlo.

[New method for analyzing pharmacodynamic material basis of traditional Chinese medicines by using specific knockout technology with monoclonal antibodies].

Science.gov (United States)

Zhao, Yan; Qu, Hui-Hua; Wang, Qing-Guo

2013-09-01

Study on pharmacodynamic material basis of traditional Chinese medicines is one of the key issues for the modernization of traditional Chinese medicine. Having introduced the monoclonal antibody technology into the study on pharmacodynamic material basis of traditional Chinese medicines, the author prepared the immunoaffinity chromatography column by using monoclonal antibodies in active components of traditional Chinese medicines, so as to selectively knock out the component from herbs or traditional Chinese medicine compounds, while preserving all of the other components and keeping their amount and ratio unchanged. A comparative study on pharmacokinetics and pharmacodynamics was made to explicitly reveal the correlation between the component and the main purpose of traditional Chinese medicines and compounds. The analysis on pharmacodynamic material basis of traditional Chinese medicines by using specific knockout technology with monoclonal antibodies is a new method for study pharmacodynamic material basis in line with the characteristics of traditional Chinese medicines. Its results can not only help study material basis from a new perspective, but also help find the modern scientific significance in single herb or among compounds of traditional Chinese medicines.

Effects of major histocompatibility complex class II knockout on mouse bone mechanical properties during development

Science.gov (United States)

Simske, Steven J.; Bateman, Ted A.; Smith, Erin E.; Ferguson, Virginia L.; Chapes, Stephen K.

2002-01-01

We investigated the effect of major histocompatibility complex class II (MHC II) knockout on the development of the mouse peripheral skeleton. These C2D mice had less skeletal development at 8, 12 and 16 weeks of age compared to wild-type C57BL/6J (B6) male mice. The C2D mice had decreased femur mechanical, geometric and compositional measurements compared to wild type mice at each of these ages. C2D femur stiffness (S), peak force in 3-pt bending (Pm), and mineral mass (Min-M) were 74%, 64% and 66%, respectively, of corresponding B6 values at 8 weeks of age. Similar differences were measured at 12 weeks (for which C2D femoral S, Pm and Min-M were 71%, 72% and 73%, respectively, of corresponding B6 values) and at 16 weeks (for which C2D femoral S, Pm and Min-M were 80%, 66% and 61%, respectively, of corresponding B6 values). MHC II knockout delays the development of adult bone properties and is accompanied by lower body mass compared to wild-type controls.

Graft function assessment in mouse models of single- and dual- kidney transplantation.

Science.gov (United States)

Wang, Lei; Wang, Ximing; Jiang, Shan; Wei, Jin; Buggs, Jacentha; Fu, Liying; Zhang, Jie; Liu, Ruisheng

2018-05-23

Animal models of kidney transplantation (KTX) are widely used in studying immune response of hosts to implanted grafts. Additionally, KTX can be used in generating kidney-specific knockout animal models by transplantation of kidneys from donors with global knockout of a gene to wild type recipients or vise verse. Dual kidney transplantation (DKT) provides a more physiological environment for recipients than single kidney transplantation (SKT). However, DKT in mice is rare due to technical challenges. In this study, we successfully performed DKT in mice and compared the hemodynamic response and graft function with SKT. The surgical time, complications and survival rate of DKT were not significantly different from SKT, where survival rates were above 85%. Mice with DKT showed less injury and quicker recovery with lower plasma creatinine (Pcr) and higher GFR than SKT mice (Pcr = 0.34 and 0.17 mg/dl in DKT vs. 0.50 and 0.36 mg/dl in SKT at 1 and 3 days, respectively; GFR = 215 and 131 µl/min for DKT and SKT, respectively). In addition, the DKT exhibited better renal functional reserve and long-term outcome of renal graft function than SKT based on the response to acute volume expansion. In conclusion, we have successfully generated a mouse DKT model. The hemodynamic responses of DKT better mimic physiological situations with less kidney injury and better recovery than SKT because of reduced confounding factors such as single nephron hyperfiltration. We anticipate DKT in mice will provide an additional tool for evaluation of renal significance in physiology and disease.

No further loss of dorsal root ganglion cells after axotomy in p75 neurotrophin receptor knockout mice

DEFF Research Database (Denmark)

Sørensen, B.; Lamm, Trine Tandrup; Koltzenburg, M.

2003-01-01

The role of the p75 neurotrophin receptor for neuronal survival after nerve crush was studied in L5 dorsal root ganglia (DRG) of knockout mice and controls with assumption-free stereological methods. Numbers of neuronal A- and B-cells were obtained using the optical fractionator and optical...

Age-dependent changes of cerebral copper metabolism in Atp7b -/- knockout mouse model of Wilson's disease by [64Cu]CuCl2-PET/CT.

Science.gov (United States)

Xie, Fang; Xi, Yin; Pascual, Juan M; Muzik, Otto; Peng, Fangyu

2017-06-01

Copper is a nutritional metal required for brain development and function. Wilson's disease (WD), or hepatolenticular degeneration, is an inherited human copper metabolism disorder caused by a mutation of the ATP7B gene. Many WD patients present with variable neurological and psychiatric symptoms, which may be related to neurodegeneration secondary to copper metabolism imbalance. The objective of this study was to explore the feasibility and use of copper-64 chloride ([ 64 C]CuCl 2 ) as a tracer for noninvasive assessment of age-dependent changes of cerebral copper metabolism in WD using an Atp7b -/- knockout mouse model of WD and positron emission tomography/computed tomography (PET/CT) imaging. Continuing from our recent study of biodistribution and radiation dosimetry of [ 64 C]CuCl 2 in Atp7b -/- knockout mice, PET quantitative analysis revealed low 64 Cu radioactivity in the brains of Atp7b -/- knockout mice at 7th weeks of age, compared with 64 Cu radioactivity in the brains of age- and gender-matched wild type C57BL/6 mice, at 24 h (h) post intravenous injection of [ 64 C]CuCl 2 as a tracer. Furthermore, age-dependent increase of 64 Cu radioactivity was detected in the brains of Atp7b -/- knockout mice from the 13th to 21th weeks of age, based on the data derived from a longitudinal [ 64 C]CuCl 2 -PET/CT study of Atp7b -/- knockout mice with orally administered [ 64 Cu]CuCl 2 as a tracer. The findings of this study support clinical use of [ 64 Cu]CuCl 2 -PET/CT imaging as a tool for noninvasive assessment of age-dependent changes of cerebral copper metabolism in WD patients presenting with variable neurological and psychiatric symptoms.

Studies of an Androgen-Binding Protein Knockout Corroborate a Role for Salivary ABP in Mouse Communication

Czech Academy of Sciences Publication Activity Database

Chung, A. G.; Belone, P. M.; Vošlajerová Bímová, Barbora; Karn, R. C.; Laukaitis, C. M.

2017-01-01

Roč. 205, č. 4 (2017), s. 1517-1527 ISSN 0016-6731 R&D Projects: GA ČR GA15-13265S Institutional support: RVO:67985904 Keywords : androgen-binding protein * knockout mouse * preference testing Subject RIV: EA - Cell Biology OBOR OECD: Biology (theoretical, mathematical, thermal, cryobiology, biological rhythm), Evolutionary biology Impact factor: 4.556, year: 2016

Histidine Decarboxylase Knockout Mice as a Model of the Pathophysiology of Tourette Syndrome and Related Conditions.

Science.gov (United States)

Pittenger, Christopher

2017-01-01

While the normal functions of histamine (HA) in the central nervous system have gradually come into focus over the past 30 years, the relationship of abnormalities in neurotransmitter HA to human disease has been slower to emerge. New insight came with the 2010 description of a rare nonsense mutation in the biosynthetic enzyme histidine decarboxylase (Hdc) that was associated with Tourette syndrome (TS) and related conditions in a single family pedigree. Subsequent genetic work has provided further support for abnormalities of HA signaling in sporadic TS. As a result of this genetic work, Hdc knockout mice, which were generated more than 15 years ago, have been reexamined as a model of the pathophysiology of TS and related conditions. Parallel work in these KO mice and in human carriers of the Hdc mutation has revealed abnormalities in the basal ganglia system and its modulation by dopamine (DA) and has confirmed the etiologic, face, and predictive validity of the model. The Hdc-KO model thus serves as a unique platform to probe the pathophysiology of TS and related conditions, and to generate specific hypotheses for subsequent testing in humans. This chapter summarizes the development and validation of this model and recent and ongoing work using it to further investigate pathophysiological changes that may contribute to these disorders.

Effects of dopamine D1-like and D2-like antagonists on cocaine discrimination in muscarinic receptor knockout mice.

Science.gov (United States)

Thomsen, Morgane; Caine, Simon Barak

2016-04-05

Muscarinic and dopamine brain systems interact intimately, and muscarinic receptor ligands, like dopamine ligands, can modulate the reinforcing and discriminative stimulus (S(D)) effects of cocaine. To enlighten the dopamine/muscarinic interactions as they pertain to the S(D) effects of cocaine, we evaluated whether muscarinic M1, M2 or M4 receptors are necessary for dopamine D1 and/or D2 antagonist mediated modulation of the S(D) effects of cocaine. Knockout mice lacking M1, M2, or M4 receptors, as well as control wild-type mice and outbred Swiss-Webster mice, were trained to discriminate 10mg/kg cocaine from saline in a food-reinforced drug discrimination procedure. Effects of pretreatments with the dopamine D1 antagonist SCH 23390 and the dopamine D2 antagonist eticlopride were evaluated. In intact mice, both SCH 23390 and eticlopride attenuated the cocaine discriminative stimulus effect, as expected. SCH 23390 similarly attenuated the cocaine discriminative stimulus effect in M1 knockout mice, but not in mice lacking M2 or M4 receptors. The effects of eticlopride were comparable in each knockout strain. These findings demonstrate differences in the way that D1 and D2 antagonists modulate the S(D) effects of cocaine, D1 modulation being at least partially dependent upon activity at the inhibitory M2/M4 muscarinic subtypes, while D2 modulation appeared independent of these systems. Copyright © 2016 Elsevier B.V. All rights reserved.

Adaptations in pre- and postsynaptic 5-HT(1A) receptor function and cocaine supersensitivity in serotonin transporter knockout rats

NARCIS (Netherlands)

Homberg, Judith R; De Boer, Sietse F; Raasø, Halfdan S; Olivier, Jocelien D A; Verheul, Mark; Ronken, Eric; Cools, Alexander R; Ellenbroek, Bart A; Schoffelmeer, Anton N M; Vanderschuren, Louk J M J; De Vries, Taco J; Cuppen, Edwin

2008-01-01

RATIONALE: While individual differences in vulnerability to psychostimulants have been largely attributed to dopaminergic neurotransmission, the role of serotonin is not fully understood. OBJECTIVES: To study the rewarding and motivational properties of cocaine in the serotonin transporter knockout

Accurate estimation of the RMS emittance from single current amplifier data

International Nuclear Information System (INIS)

Stockli, Martin P.; Welton, R.F.; Keller, R.; Letchford, A.P.; Thomae, R.W.; Thomason, J.W.G.

2002-01-01

This paper presents the SCUBEEx rms emittance analysis, a self-consistent, unbiased elliptical exclusion method, which combines traditional data-reduction methods with statistical methods to obtain accurate estimates for the rms emittance. Rather than considering individual data, the method tracks the average current density outside a well-selected, variable boundary to separate the measured beam halo from the background. The average outside current density is assumed to be part of a uniform background and not part of the particle beam. Therefore the average outside current is subtracted from the data before evaluating the rms emittance within the boundary. As the boundary area is increased, the average outside current and the inside rms emittance form plateaus when all data containing part of the particle beam are inside the boundary. These plateaus mark the smallest acceptable exclusion boundary and provide unbiased estimates for the average background and the rms emittance. Small, trendless variations within the plateaus allow for determining the uncertainties of the estimates caused by variations of the measured background outside the smallest acceptable exclusion boundary. The robustness of the method is established with complementary variations of the exclusion boundary. This paper presents a detailed comparison between traditional data reduction methods and SCUBEEx by analyzing two complementary sets of emittance data obtained with a Lawrence Berkeley National Laboratory and an ISIS H - ion source

Knockout silkworms reveal a dispensable role for juvenile hormones in holometabolous life cycle.

Science.gov (United States)

Daimon, Takaaki; Uchibori, Miwa; Nakao, Hajime; Sezutsu, Hideki; Shinoda, Tetsuro

2015-08-04

Insect juvenile hormones (JHs) prevent precocious metamorphosis and allow larvae to undergo multiple rounds of status quo molts. However, the roles of JHs during the embryonic and very early larval stages have not been fully understood. We generated and characterized knockout silkworms (Bombyx mori) with null mutations in JH biosynthesis or JH receptor genes using genome-editing tools. We found that embryonic growth and morphogenesis are largely independent of JHs in Bombyx and that, even in the absence of JHs or JH signaling, pupal characters are not formed in first- or second-instar larvae, and precocious metamorphosis is induced after the second instar at the earliest. We also show by mosaic analysis that a pupal specifier gene broad, which is dramatically up-regulated in the late stage of the last larval instar, is essential for pupal commitment in the epidermis. Importantly, the mRNA expression level of broad, which is thought to be repressed by JHs, remained at very low basal levels during the early larval instars of JH-deficient or JH signaling-deficient knockouts. Therefore, our study suggests that the long-accepted paradigm that JHs maintain the juvenile status throughout larval life should be revised because the larval status can be maintained by a JH-independent mechanism in very early larval instars. We propose that the lack of competence for metamorphosis during the early larval stages may result from the absence of an unidentified broad-inducing factor, i.e., a competence factor.

Increased susceptibility to diet-induced obesity in GPRC6A receptor knockout mice

DEFF Research Database (Denmark)

Clemmensen, Christoffer; Smajilovic, Sanela; Madsen, Andreas N

2013-01-01

locomotor activity. Moreover, diet-induced obese Gprc6a KO mice had increased circulating insulin and leptin levels relative to WT animals, thereby demonstrating that endocrine abnormalities associate with the reported disturbances in energy balance. The phenotype was further accompanied by disruptions...... complications is still elusive. In the present study, we investigated the impact of GPRC6A deficiency in a murine model of diet-induced obesity (DIO). Male Gprc6a knockout (KO) mice and WT littermates were subjected to a high-fat diet (HFD) for 25 weeks and exposed to comprehensive metabolic phenotyping...

Forebrain-specific knockout of B-raf kinase leads to deficits in hippocampal long-term potentiation, learning, and memory.

Science.gov (United States)

Chen, Adele P; Ohno, Masuo; Giese, K Peter; Kühn, Ralf; Chen, Rachel L; Silva, Alcino J

2006-01-01

Raf kinases are downstream effectors of Ras and upstream activators of the MEK-ERK cascade. Ras and MEK-ERK signaling play roles in learning and memory (L&M) and neural plasticity, but the roles of Raf kinases in L&M and plasticity are unclear. Among Raf isoforms, B-raf is preferentially expressed in the brain. To determine whether B-raf has a role in synaptic plasticity and L&M, we used the Cre-LoxP gene targeting system to derive forebrain excitatory neuron B-raf knockout mice. This conditional knockout resulted in deficits in ERK activation and hippocampal long-term potentiation (LTP) and impairments in hippocampus-dependent L&M, including spatial learning and contextual discrimination. Despite the widespread expression of B-raf, this mutation did not disrupt other forms of L&M, such as cued fear conditioning and conditioned taste aversion. Our findings demonstrate that B-raf plays a role in hippocampal ERK activation, synaptic plasticity, and L&M.

Beta2-adrenergic activity modulates vascular tone regulation in lecithin:cholesterol acyltransferase knockout mice

Science.gov (United States)

Manzini, S.; Pinna, C.; Busnelli, M.; Cinquanta, P.; Rigamonti, E.; Ganzetti, G.S.; Dellera, F.; Sala, A.; Calabresi, L.; Franceschini, G.; Parolini, C.; Chiesa, G.

2015-01-01

Lecithin:cholesterol acyltransferase (LCAT) deficiency is associated with hypoalphalipoproteinemia, generally a predisposing factor for premature coronary heart disease. The evidence of accelerated atherosclerosis in LCAT-deficient subjects is however controversial. In this study, the effect of LCAT deficiency on vascular tone and endothelial function was investigated in LCAT knockout mice, which reproduce the human lipoprotein phenotype. Aortas from wild-type (Lcatwt) and LCAT knockout (LcatKO) mice exposed to noradrenaline showed reduced contractility in LcatKO mice (P < 0.005), whereas acetylcholine exposure showed a lower NO-dependent relaxation in LcatKO mice (P < 0.05). Quantitative PCR and Western blotting analyses suggested an adequate eNOS expression in LcatKO mouse aortas. Real-time PCR analysis indicated increased expression of β2-adrenergic receptors vs wild-type mice. Aorta stimulation with noradrenaline in the presence of propranolol, to abolish the β-mediated relaxation, showed the same contractile response in the two mouse lines. Furthermore, propranolol pretreatment of mouse aortas exposed to L-NAME prevented the difference in responses between Lcatwt and LcatKO mice. The results indicate that LCAT deficiency leads to increased β2-adrenergic relaxation and to a consequently decreased NO-mediated vasodilation that can be reversed to guarantee a correct vascular tone. The present study suggests that LCAT deficiency is not associated with an impaired vascular reactivity. PMID:26254103

Beta2-adrenergic activity modulates vascular tone regulation in lecithin:cholesterol acyltransferase knockout mice.

Science.gov (United States)

Manzini, S; Pinna, C; Busnelli, M; Cinquanta, P; Rigamonti, E; Ganzetti, G S; Dellera, F; Sala, A; Calabresi, L; Franceschini, G; Parolini, C; Chiesa, G

2015-11-01

Lecithin:cholesterol acyltransferase (LCAT) deficiency is associated with hypoalphalipoproteinemia, generally a predisposing factor for premature coronary heart disease. The evidence of accelerated atherosclerosis in LCAT-deficient subjects is however controversial. In this study, the effect of LCAT deficiency on vascular tone and endothelial function was investigated in LCAT knockout mice, which reproduce the human lipoprotein phenotype. Aortas from wild-type (Lcat(wt)) and LCAT knockout (Lcat(KO)) mice exposed to noradrenaline showed reduced contractility in Lcat(KO) mice (P<0.005), whereas acetylcholine exposure showed a lower NO-dependent relaxation in Lcat(KO) mice (P<0.05). Quantitative PCR and Western blotting analyses suggested an adequate eNOS expression in Lcat(KO) mouse aortas. Real-time PCR analysis indicated increased expression of β2-adrenergic receptors vs wild-type mice. Aorta stimulation with noradrenaline in the presence of propranolol, to abolish the β-mediated relaxation, showed the same contractile response in the two mouse lines. Furthermore, propranolol pretreatment of mouse aortas exposed to L-NAME prevented the difference in responses between Lcat(wt) and Lcat(KO) mice. The results indicate that LCAT deficiency leads to increased β2-adrenergic relaxation and to a consequently decreased NO-mediated vasodilation that can be reversed to guarantee a correct vascular tone. The present study suggests that LCAT deficiency is not associated with an impaired vascular reactivity. Copyright © 2015. Published by Elsevier Inc.

Characterization of nasal potential difference in cftr knockout and F508del-CFTR mice.

Directory of Open Access Journals (Sweden)

Emilie Lyne Saussereau

Full Text Available BACKGROUND: Treatments designed to correct cystic fibrosis transmembrane conductance regulator (CFTR defects must first be evaluated in preclinical experiments in the mouse model of cystic fibrosis (CF. Mice nasal mucosa mimics the bioelectric defect seen in humans. The use of nasal potential difference (V(TE to assess ionic transport is a powerful test evaluating the restoration of CFTR function. Nasal V(TE in CF mice must be well characterized for correct interpretation. METHODS: We performed V(TE measurements in large-scale studies of two mouse models of CF--B6;129 cftr knockout and FVB F508del-CFTR--and their respective wild-type (WT littermates. We assessed the repeatability of the test for cftr knockout mice and defined cutoff points distinguishing between WT and F508del-CFTR mice. RESULTS: We determined the typical V(TE values for CF and WT mice and demonstrated the existence of residual CFTR activity in F508del-CFTR mice. We characterized intra-animal variability in B6;129 mice and defined the cutoff points for F508del-CFTR chloride secretion rescue. Hyperpolarization of more than -2.15 mV after perfusion with a low-concentration Cl(- solution was considered to indicate a normal response. CONCLUSIONS: These data will make it possible to interpret changes in nasal V(TE in mouse models of CF, in future preclinical studies.

p21{sup WAF1/Cip1/Sdi1} knockout mice respond to doxorubicin with reduced cardiotoxicity

Energy Technology Data Exchange (ETDEWEB)

Terrand, Jerome; Xu, Beibei; Morrissy, Steve; Dinh, Thai Nho [Department of Pharmacology,College of Medicine, University of Arizona, 1501 N. Campbell Ave, Tucson, AZ 85724 (United States); Williams, Stuart [Biomedical Engineering Program, College of Medicine, University of Arizona, 1501 N. Campbell Ave, Tucson, AZ 85724 (United States); Chen, Qin M., E-mail: qchen@email.arizona.edu [Department of Pharmacology,College of Medicine, University of Arizona, 1501 N. Campbell Ave, Tucson, AZ 85724 (United States)

2011-11-15

Doxorubicin (Dox) is an antineoplastic agent that can cause cardiomyopathy in humans and experimental animals. As an inducer of reactive oxygen species and a DNA damaging agent, Dox causes elevated expression of p21{sup WAF1/Cip1/Sdi1} (p21) gene. Elevated levels of p21 mRNA and p21 protein have been detected in the myocardium of mice following Dox treatment. With chronic treatment of Dox, wild type (WT) animals develop cardiomyopathy evidenced by elongated nuclei, mitochondrial swelling, myofilamental disarray, reduced cardiac output, reduced ejection fraction, reduced left ventricular contractility, and elevated expression of ANF gene. In contrast, p21 knockout (p21KO) mice did not show significant changes in the same parameters in response to Dox treatment. In an effort to understand the mechanism of the resistance against Dox induced cardiomyopathy, we measured levels of antioxidant enzymes and found that p21KO mice did not contain elevated basal or inducible levels of glutathione peroxidase and catalase. Measurements of 6 circulating cytokines indicated elevation of IL-6, IL-12, IFN{gamma} and TNF{alpha} in Dox treated WT mice but not p21KO mice. Dox induced elevation of IL-6 mRNA was detected in the myocardium of WT mice but not p21KO mice. While the mechanism of the resistance against Dox induced cardiomyopathy remains unclear, lack of inflammatory response may contribute to the observed cardiac protection in p21KO mice. -- Highlights: Black-Right-Pointing-Pointer Doxorubicin induces p21 elevation in the myocardium. Black-Right-Pointing-Pointer Doxorubicin causes dilated cardiomyopathy in wild type mice. Black-Right-Pointing-Pointer p21 Knockout mice are resistant against doxorubicin induced cardiomyopathy. Black-Right-Pointing-Pointer Lack of inflammatory response correlates with the resistance in p21 knockout mice.

Die Bedeutung von Fetuin-A für Proliferation und Fibrogenese im Knockout-Modell

OpenAIRE

Stojanov, Sabrina

2009-01-01

The human cirrhosis of the liver is a progressive illness with serious clinical results, like portal hypertension and terminal functional loss of the organ. At the end stands the liver transplant or the exitus letalis, not to forget the strongly raised prevalence for the hepatocellular carcinoma. For these reasons it is essential to develop new therapy concepts with views of forecast improvement or even healing. With a knockout model of the mouse we examined the effect of fetuin-A for physiol...

The vasopressin deficient Brattleboro rats: A natural knockout model used in the search for CNS effects of vasopressin

NARCIS (Netherlands)

Bohus, B; de Wied, David; Urban, I.J.A.; Burbach, J.P.H.; De Wied, D.

1999-01-01

Behavioral neuroscience is using mon and more gene knockout techniques to produce animals with a specific deletion. These studies have their precedent in nature. A mutation may result in a limited genetic defect, as seen in the vasopressin (VP) deficiency in the Brattleboro rat. The mutation is in a

Requirement of NMDA receptor reactivation for consolidation and storage of nondeclarative taste memory revealed by inducible NR1 knockout.

Science.gov (United States)

Cui, Zhenzhong; Lindl, Kathryn A; Mei, Bing; Zhang, Shuqing; Tsien, Joe Z

2005-08-01

We employed an inducible, reversible and region-specific gene knockout technique to investigate the requirements for cortical NMDA receptors (NMDAR) during the various stages (acquisition, consolidation and storage, and retrieval) of nondeclarative, hippocampal-independent memory in mice using a conditioned taste aversion memory paradigm. Here we show that temporary knockout of the cortical NMDAR during either the learning or postlearning consolidation stage, but not during the retrieval stage, causes severe performance deficits in the 1-month taste memory retention tests. More importantly, we found that the consolidation and storage of the long-term nondeclarative taste memories requires cortical NMDAR reactivation. Thus, the dynamic engagement of the NMDAR during the postlearning stage leads us to postulate that NMDAR reactivation-mediated synaptic re-entry reinforcement is crucial for overcoming the destabilizing effects intrinsic to synaptic protein turnover and for achieving consolidation and storage of nondeclarative memories in the brain.

Accurate calculation of Green functions on the d-dimensional hypercubic lattice

International Nuclear Information System (INIS)

Loh, Yen Lee

2011-01-01

We write the Green function of the d-dimensional hypercubic lattice in a piecewise form covering the entire real frequency axis. Each piece is a single integral involving modified Bessel functions of the first and second kinds. The smoothness of the integrand allows both real and imaginary parts of the Green function to be computed quickly and accurately for any dimension d and any real frequency, and the computational time scales only linearly with d.

Accurate thickness measurement of graphene

International Nuclear Information System (INIS)

Shearer, Cameron J; Slattery, Ashley D; Stapleton, Andrew J; Shapter, Joseph G; Gibson, Christopher T

2016-01-01

Graphene has emerged as a material with a vast variety of applications. The electronic, optical and mechanical properties of graphene are strongly influenced by the number of layers present in a sample. As a result, the dimensional characterization of graphene films is crucial, especially with the continued development of new synthesis methods and applications. A number of techniques exist to determine the thickness of graphene films including optical contrast, Raman scattering and scanning probe microscopy techniques. Atomic force microscopy (AFM), in particular, is used extensively since it provides three-dimensional images that enable the measurement of the lateral dimensions of graphene films as well as the thickness, and by extension the number of layers present. However, in the literature AFM has proven to be inaccurate with a wide range of measured values for single layer graphene thickness reported (between 0.4 and 1.7 nm). This discrepancy has been attributed to tip-surface interactions, image feedback settings and surface chemistry. In this work, we use standard and carbon nanotube modified AFM probes and a relatively new AFM imaging mode known as PeakForce tapping mode to establish a protocol that will allow users to accurately determine the thickness of graphene films. In particular, the error in measuring the first layer is reduced from 0.1–1.3 nm to 0.1–0.3 nm. Furthermore, in the process we establish that the graphene-substrate adsorbate layer and imaging force, in particular the pressure the tip exerts on the surface, are crucial components in the accurate measurement of graphene using AFM. These findings can be applied to other 2D materials. (paper)

Accurate Multisteps Traffic Flow Prediction Based on SVM

Directory of Open Access Journals (Sweden)

Zhang Mingheng

2013-01-01

Full Text Available Accurate traffic flow prediction is prerequisite and important for realizing intelligent traffic control and guidance, and it is also the objective requirement for intelligent traffic management. Due to the strong nonlinear, stochastic, time-varying characteristics of urban transport system, artificial intelligence methods such as support vector machine (SVM are now receiving more and more attentions in this research field. Compared with the traditional single-step prediction method, the multisteps prediction has the ability that can predict the traffic state trends over a certain period in the future. From the perspective of dynamic decision, it is far important than the current traffic condition obtained. Thus, in this paper, an accurate multi-steps traffic flow prediction model based on SVM was proposed. In which, the input vectors were comprised of actual traffic volume and four different types of input vectors were compared to verify their prediction performance with each other. Finally, the model was verified with actual data in the empirical analysis phase and the test results showed that the proposed SVM model had a good ability for traffic flow prediction and the SVM-HPT model outperformed the other three models for prediction.

A study in male and female 5-HT transporter knockout rats : An animal model for anxiety and depression disorders

NARCIS (Netherlands)

Olivier, J D A; Van Der Hart, M G C; Van Swelm, R P L; Dederen, P J; Homberg, J R; Cremers, T; Deen, P M T; Cuppen, E; Cools, A R; Ellenbroek, B A

2008-01-01

Human studies have shown that a reduction of 5-HT transporter (SERT) increases the vulnerability for anxiety and depression. Moreover, women are more vulnerable to develop depression and anxiety disorders than men. For that reason we hypothesized that homozygous 5-HT transporter knockout rat

A study in male and female 5-HT transporter knockout rats: an animal model for anxiety and depression disorders.

NARCIS (Netherlands)

Olivier, J.; Van Der Hart, M.G.C.; Van Swelm, R.P.L.; Dederen, P.J.; Homberg, J.R.; Cremers, T.; Deen, P.M.T.; Cuppen, E.; Cools, A.R.; Ellenbroek, B.A.

2008-01-01

Human studies have shown that a reduction of 5-HT transporter (SERT) increases the vulnerability for anxiety and depression. Moreover, women are more vulnerable to develop depression and anxiety disorders than men. For that reason we hypothesized that homozygous 5-HT transporter knockout rat

Characterization of Bombyx mori nucleopolyhedrovirus with a knockout of Bm17.

Science.gov (United States)

Shen, Hongxing; Zhou, Yang; Zhang, Wen; Nin, Bin; Wang, Hua; Wang, Xiaochun; Shao, Shihe; Chen, Huiqing; Guo, Zhongjian; Liu, Xiaoyong; Yao, Qin; Chen, Keping

2012-12-01

Open reading frame 17 (Bm17) gene of Bombyx mori nucleopolyhedrovirus is a highly conserved gene in lepidopteran nucleopolyhedroviruses, but its function remains unknown. In this report, transient-expression and superinfection assays indicated that BM17 localized in the nucleus and cytoplasm of infected BmN cells. To determine the role of Bm17 in baculovirus life cycle, we constructed a Bm17 knockout virus and characterized its properties in cells. Analysis of the production and infection of budded virions, the level of viral DNA replication revealed showed that there was no significant difference among the mutant, the control, and the Bm17 repaired virus strains. These results suggest that BM17 is not essential for virus replication in cultured cells.

Knockout of exogenous EGFP gene in porcine somatic cells using zinc-finger nucleases

International Nuclear Information System (INIS)

Watanabe, Masahito; Umeyama, Kazuhiro; Matsunari, Hitomi; Takayanagi, Shuko; Haruyama, Erika; Nakano, Kazuaki; Fujiwara, Tsukasa; Ikezawa, Yuka; Nakauchi, Hiromitsu

2010-01-01

Research highlights: → EGFP gene integrated in porcine somatic cells could be knocked out using the ZFN-KO system. → ZFNs induced targeted mutations in porcine primary cultured cells. → Complete absence of EGFP fluorescence was confirmed in ZFN-treated cells. -- Abstract: Zinc-finger nucleases (ZFNs) are expected as a powerful tool for generating gene knockouts in laboratory and domestic animals. Currently, it is unclear whether this technology can be utilized for knocking-out genes in pigs. Here, we investigated whether knockout (KO) events in which ZFNs recognize and cleave a target sequence occur in porcine primary cultured somatic cells that harbor the exogenous enhanced green fluorescent protein (EGFP) gene. ZFN-encoding mRNA designed to target the EGFP gene was introduced by electroporation into the cell. Using the Surveyor nuclease assay and flow cytometric analysis, we confirmed ZFN-induced cleavage of the target sequence and the disappearance of EGFP fluorescence expression in ZFN-treated cells. In addition, sequence analysis revealed that ZFN-induced mutations such as base substitution, deletion, or insertion were generated in the ZFN cleavage site of EGFP-expression negative cells that were cloned from ZFN-treated cells, thereby showing it was possible to disrupt (i.e., knock out) the function of the EGFP gene in porcine somatic cells. To our knowledge, this study provides the first evidence that the ZFN-KO system can be applied to pigs. These findings may open a new avenue to the creation of gene KO pigs using ZFN-treated cells and somatic cell nuclear transfer.

CRISPR/Cas9-based knockouts reveal that CpRLP1 is a negative regulator of the sex pheromone PR-IP in the Closterium peracerosum-strigosum-littorale complex.

Science.gov (United States)

Kanda, Naho; Ichikawa, Machiko; Ono, Ayaka; Toyoda, Atsushi; Fujiyama, Asao; Abe, Jun; Tsuchikane, Yuki; Nishiyama, Tomoaki; Sekimoto, Hiroyuki

2017-12-19

Heterothallic strains of the Closterium peracerosum-strigosum-littorale (C. psl.) complex have two sexes, mating-type plus (mt + ) and mating-type minus (mt - ). Conjugation between these two sexes is regulated by two sex pheromones, protoplast-release-inducing protein (PR-IP) and PR-IP Inducer, which are produced by mt + and mt - cells, respectively. PR-IP mediates the release of protoplasts from mt - cells during mating. In this study, we examined the mechanism of action of CpRLP1 (receptor-like protein 1), which was previously identified in a cDNA microarray analysis as one of the PR-IP-inducible genes. Using CRISPR/Cas9 technology, we generated CpRLP1 knockout mutants in mt - cells of the C. psl. complex. When the knockout mt - cells were mixed with wild-type mt + cells, conjugation was severely reduced. Many cells released protoplasts without pairing, suggesting a loss of synchronization between the two mating partners. Furthermore, the knockout mutants were hypersensitive to PR-IP. We conclude that CpRLP1 is a negative regulator of PR-IP that regulates the timing of protoplast release in conjugating C. psl. cells. As the first report of successful gene knockout in the class Charophyceae, this study provides a basis for research aimed at understanding the ancestral roles of genes that are indispensable for the development of land plants.

Ground state energy and width of 7He from 8Li proton knockout

International Nuclear Information System (INIS)

Denby, D. H.; DeYoung, P. A.; Hall, C. C.; Baumann, T.; Bazin, D.; Spyrou, A.; Breitbach, E.; Howes, R.; Brown, J.; Frank, N.; Gade, A.; Mosby, S. M.; Peters, W. A.; Thoennessen, M.; Hinnefeld, J.; Hoffman, C. R.; Jenson, R. A.; Luther, B.; Olson, C. W.; Schiller, A.

2008-01-01

The ground state energy and width of 7 He has been measured with the Modular Neutron Array (MoNA) and superconducting dipole Sweeper magnet experimental setup at the National Superconducting Cyclotron Laboratory. 7 He was produced by proton knockout from a secondary 8 Li beam. The measured decay energy spectrum is compared to simulations based on Breit-Wigner line shape with an energy-dependent width for the resonant state. The energy of the ground state is found to be 400(10) keV with a full-width at half-maximum of 125( -15 +40 ) keV

Targeting a single function of the multifunctional matrix metalloprotease MT1-MMP

DEFF Research Database (Denmark)

Ingvarsen, Signe; Porse, Astrid; Erpicum, Charlotte

2013-01-01

and pathological events, has been complicated by the lack of specific inhibitors and the fact that some of the potent MMPs are multifunctional enzymes. These factors have also hampered the setup of therapeutic strategies targeting MMP activity. A tempting target is the membrane-associated MT1-MMP, which has well......-documented importance in matrix degradation but which takes part in more than one pathway in this regard. In this report, we describe the selective targeting of a single function of this enzyme by means of a specific monoclonal antibody against MT1-MMP, raised in an MT1-MMP knock-out mouse. The antibody blocks...... matrix in vitro, as well as in lymphatic vessel sprouting assayed ex vivo. This is the first example of the complete inactivation of a single function of a multifunctional MMP and the use of this strategy to pursue its role....

Bcl-xL knockout attenuates mitochondrial respiration and causes oxidative stress that is compensated by pentose phosphate pathway activity

NARCIS (Netherlands)

Pfeiffer, Annika; Schneider, Julia; Bueno, Diones; Dolga, Amalia; Voss, Timo-Daniel; Lewerenz, Jan; Wüllner, Verena; Methner, Axel

2017-01-01

Bcl-xL is an anti-apoptotic protein that localizes to the outer mitochondrial membrane and influences mitochondrial bioenergetics by controlling Ca2+ influx into mitochondria. Here, we analyzed the effect of mitochondrial Bcl-xL on mitochondrial shape and function in knockout (KO), wild type and

Heterozygous CDKL5 Knockout Female Mice Are a Valuable Animal Model for CDKL5 Disorder

OpenAIRE

Fuchs, Claudia; Gennaccaro, Laura; Trazzi, Stefania; Bastianini, Stefano; Bettini, Simone; Martire, Viviana Lo; Ren, Elisa; Medici, Giorgio; Zoccoli, Giovanna; Rimondini, Roberto; Ciani, Elisabetta

2018-01-01

CDKL5 disorder is a severe neurodevelopmental disorder caused by mutations in the X-linked CDKL5 (cyclin-dependent kinase-like five) gene. CDKL5 disorder primarily affects girls and is characterized by early-onset epileptic seizures, gross motor impairment, intellectual disability, and autistic features. Although all CDKL5 female patients are heterozygous, the most valid disease-related model, the heterozygous female Cdkl5 knockout (Cdkl5 +/−) mouse, has been little characterized. The lack of...

Romk1 Knockout Mice Do Not Produce Bartter Phenotype but Exhibit Impaired K Excretion*

Science.gov (United States)

Dong, Ke; Yan, Qingshang; Lu, Ming; Wan, Laxiang; Hu, Haiyan; Guo, Junhua; Boulpaep, Emile; Wang, WenHui; Giebisch, Gerhard; Hebert, Steven C.; Wang, Tong

2016-01-01

Romk knock-out mice show a similar phenotype to Bartter syndrome of salt wasting and dehydration due to reduced Na-K-2Cl-cotransporter activity. At least three ROMK isoforms have been identified in the kidney; however, unique functions of any of the isoforms in nephron segments are still poorly understood. We have generated a mouse deficient only in Romk1 by selective deletion of the Romk1-specific first exon using an ES cell Cre-LoxP strategy and examined the renal phenotypes, ion transporter expression, ROMK channel activity, and localization under normal and high K intake. Unlike Romk−/− mice, there was no Bartter phenotype with reduced NKCC2 activity and increased NCC expression in Romk1−/− mice. The small conductance K channel (SK) activity showed no difference of channel properties or gating in the collecting tubule between Romk1+/+ and Romk1−/− mice. High K intake increased SK channel number per patch and increased the ROMK channel intensity in the apical membrane of the collecting tubule in Romk1+/+, but such regulation by high K intake was diminished with significant hyperkalemia in Romk1−/− mice. We conclude that 1) animal knockouts of ROMK1 do not produce Bartter phenotype. 2) There is no functional linking of ROMK1 and NKCC2 in the TAL. 3) ROMK1 is critical in response to high K intake-stimulated K+ secretion in the collecting tubule. PMID:26728465

Romk1 Knockout Mice Do Not Produce Bartter Phenotype but Exhibit Impaired K Excretion.

Science.gov (United States)

Dong, Ke; Yan, Qingshang; Lu, Ming; Wan, Laxiang; Hu, Haiyan; Guo, Junhua; Boulpaep, Emile; Wang, WenHui; Giebisch, Gerhard; Hebert, Steven C; Wang, Tong

2016-03-04

Romk knock-out mice show a similar phenotype to Bartter syndrome of salt wasting and dehydration due to reduced Na-K-2Cl-cotransporter activity. At least three ROMK isoforms have been identified in the kidney; however, unique functions of any of the isoforms in nephron segments are still poorly understood. We have generated a mouse deficient only in Romk1 by selective deletion of the Romk1-specific first exon using an ES cell Cre-LoxP strategy and examined the renal phenotypes, ion transporter expression, ROMK channel activity, and localization under normal and high K intake. Unlike Romk(-/-) mice, there was no Bartter phenotype with reduced NKCC2 activity and increased NCC expression in Romk1(-/-) mice. The small conductance K channel (SK) activity showed no difference of channel properties or gating in the collecting tubule between Romk1(+/+) and Romk1(-/-) mice. High K intake increased SK channel number per patch and increased the ROMK channel intensity in the apical membrane of the collecting tubule in Romk1(+/+), but such regulation by high K intake was diminished with significant hyperkalemia in Romk1(-/-) mice. We conclude that 1) animal knockouts of ROMK1 do not produce Bartter phenotype. 2) There is no functional linking of ROMK1 and NKCC2 in the TAL. 3) ROMK1 is critical in response to high K intake-stimulated K(+) secretion in the collecting tubule. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Gamma-ray measurements in the one-neutron knockout of {sup 17}C, {sup 19}N, {sup 21}O and {sup 25}F

Energy Technology Data Exchange (ETDEWEB)

Rodriguez-Tajes, C.; Cortina-Gil, D.; Alvarez-Pol, H.; Benjamim, E.; Benlliure, J.; Caamano, M.; Casarejos, E.; Gascon, M.; Kurtukian, T.; Perez-Loureiro, D. [Universidade de Santiago de Compostela, Departmento de Fisica de Particulas, Santiago de Compostela (Spain); Aumann, T.; Chatillon, A.; Geissel, H.; Nociforo, C.; Prochazka, A.; Simon, H.; Suemmerer, K.; Weick, H.; Winkler, M. [GSI Helmholtzzentrum fuer Schwerionenforschung, Darmstadt (Germany); Borge, M.J.G.; Pascual-Izarra, C.; Perea, A.; Tengblad, O. [CSIC, Instituto de Estructura de la Materia, Madrid (Spain); Chulkov, L.V. [GSI Helmholtzzentrum fuer Schwerionenforschung, Darmstadt (Germany); Kurchatov Institute, Moscow (Russian Federation); Eppinger, K.; Faestermann, T.; Gernhaeuser, R.; Kruecken, R.; Maierbeck, P.; Schwertel, S. [Technische Universitaet Muenchen, Physik Department E12, Garching (Germany); Jonson, B. [Chalmers Tekniska Hoegskola, Fundamental Fysik, Goeteborg (Sweden); CERN, PH Department, Geneve (Switzerland); Kanungo, R. [Saint Mary' s University, Astronomy and Physics Department, Halifax, NS (Canada); Nilsson, T.; Zhukov, M.V. [Chalmers Tekniska Hoegskola, Fundamental Fysik, Goeteborg (Sweden)

2012-07-15

One-neutron knockout reactions in a {sup 9}Be target have been investigated at relativistic energies, near 700 MeV/u, for a set of sd-shell, neutron-rich nuclei. The experiment was performed in the FRS spectrometer, at GSI. {gamma}-ray measurements were carried out by means of the MINIBALL {gamma}-ray spectrometer and allowed the determination of partial cross-sections and branching ratios corresponding to the final states of the emerging knockout fragments. Experimental results are presented for {sup 17}C, {sup 19}N, {sup 21}O and {sup 25}F projectiles. The role of excited states of the N - 1 fragments in the composition of the ground state of these neutron-rich projectiles is outlined in this work. (orig.)

Effects of ketoconazole on the biodistribution and metabolism of [{sup 11}C]loperamide and [{sup 11}C]N-desmethyl-loperamide in wild-type and P-gp knockout mice

Energy Technology Data Exchange (ETDEWEB)

Seneca, Nicholas; Zoghbi, Sami S.; Shetty, H. Umesha; Tuan, Edward; Kannan, Pavitra; Taku, Andrew; Innis, Robert B. [Molecular Imaging Branch, National Institute of Mental Health, National Institutes of Health, Bethesda, MD 20892 (United States); Pike, Victor W. [Molecular Imaging Branch, National Institute of Mental Health, National Institutes of Health, Bethesda, MD 20892 (United States)], E-mail: pikev@mail.nih.gov

2010-04-15

Introduction: [{sup 11}C]Loperamide and [{sup 11}C]N-desmethyl-loperamide ([{sup 11}C]dLop) have been proposed as radiotracers for imaging brain P-glycoprotein (P-gp) function. A major route of [{sup 11}C]loperamide metabolism is N-demethylation to [{sup 11}C]dLop. We aimed to test whether inhibition of CYP3A4 with ketoconazole might reduce the metabolism of [{sup 11}C]loperamide and [{sup 11}C]dLop in mice, and thereby improve the quality of these radiotracers. Methods: Studies were performed in wild-type and P-gp knockout (mdr-1a/b -/-) mice. During each of seven study sessions, one pair of mice, comprising one wild-type and one knockout mouse, was pretreated with ketoconazole (50 mg/kg, ip), while another such pair was left untreated. Mice were sacrificed at 30 min after injection of [{sup 11}C]loperamide or [{sup 11}C]dLop. Whole brain and plasma samples were measured for radioactivity and analyzed with radio-high-performance liquid chromatography. Results: Ketoconazole increased the plasma concentrations of [{sup 11}C]loperamide and its main radiometabolite, [{sup 11}C]dLop, by about twofold in both wild-type and knockout mice, whereas the most polar radiometabolite was decreased threefold. Furthermore, ketoconazole increased the brain concentrations of [{sup 11}C]loperamide and the radiometabolite [{sup 11}C]dLop by about twofold in knockout mice, and decreased the brain concentrations of the major and most polar radiometabolite in wild-type and knockout mice by 82% and 49%, respectively. In contrast, ketoconazole had no effect on plasma and brain distribution of administered [{sup 11}C]dLop and its radiometabolites in either wild-type or knockout mice, except to increase the low plasma [{sup 11}C]dLop concentration. The least polar radiometabolite of [{sup 11}C]dLop was identified with LC-MS{sup n} as the N-hydroxymethyl analog of [{sup 11}C]dLop and this also behaved as a P-gp substrate. Conclusion: In this study, ketoconazole (50 mg/kg, ip) proved

Brain Region-Specific Effects of cGMP-Dependent Kinase II Knockout on AMPA Receptor Trafficking and Animal Behavior

Science.gov (United States)

Kim, Seonil; Pick, Joseph E.; Abera, Sinedu; Khatri, Latika; Ferreira, Danielle D. P.; Sathler, Matheus F.; Morison, Sage L.; Hofmann, Franz; Ziff, Edward B.

2016-01-01

Phosphorylation of GluA1, a subunit of AMPA receptors (AMPARs), is critical for AMPAR synaptic trafficking and control of synaptic transmission. cGMP-dependent protein kinase II (cGKII) mediates this phosphorylation, and cGKII knockout (KO) affects GluA1 phosphorylation and alters animal behavior. Notably, GluA1 phosphorylation in the KO…

Alcoholic fatty liver is enhanced in CYP2A5 knockout mice: The role of the PPARα-FGF21 axis

International Nuclear Information System (INIS)

Chen, Xue; Ward, Stephen C.; Cederbaum, Arthur I.; Xiong, Huabao; Lu, Yongke

2017-01-01

Background & aims: Cytochrome P450 2A5 (CYP2A5) is induced by ethanol, and the ethanol induction of CYP2A5 is regulated by nuclear factor-erythroid 2-related factor 2 (NRF2). Cyp2a5 knockout (Cyp2a5 −/− ) mice develop more severe alcoholic fatty liver than Cyp2a5 +/+ mice. Fibroblast growth factor 21 (FGF21), a PPARα-regulated liver hormone, is involved in hepatic lipid metabolism. Alcoholic and non-alcoholic fatty liver are enhanced in Pparα knockout (Pparα −/− ) mice. This study investigates the relationship between the PPARα-FGF21 axis and the enhanced alcoholic fatty liver in Cyp2a5 −/− mice. Methods: Mice were fed the Lieber-Decarli ethanol diet to induce alcoholic fatty liver. Results: More severe alcoholic fatty liver disease was developed in Cyp2a5 −/− mice than in Cyp2a5 +/+ mice. Basal FGF21 levels were higher in Cyp2a5 −/− mice than in Cyp2a5 +/+ mice, but ethanol did not further increase the elevated FGF21 levels in Cyp2a5 −/− mice while FGF21 was induced by ethanol in Cyp2a5 +/+ mice. Basal levels of serum FGF21 were lower in Pparα −/− mice than in Pparα +/+ mice; ethanol induced FGF21 in Pparα +/+ mice but not in Pparα −/− mice, whereas ethanol induced hypertriglyceridemia in Pparα −/− mice but not in Pparα +/+ mice. Administration of recombinant FGF21 normalized serum FGF21 and triglyceride in Pparα −/− mice. Alcoholic fatty liver was enhanced in liver-specific Fgf21 knockout mice. Pparα and Cyp2a5 double knockout (Pparα −/− /Cyp2a5 −/− ) mice developed more severe alcoholic fatty liver than Pparα +/+ /Cyp2a5 −/− mice. Conclusions: These results suggest that CYP2A5 protects against the development of alcoholic fatty liver disease, and the PPARα-FGF21 axis contributes to the protective effects of CYP2A5 on alcoholic fatty liver disease.

Behavioral analysis of NR2C knockout mouse reveals deficit in acquisition of conditioned fear and working memory.

Science.gov (United States)

Hillman, Brandon G; Gupta, Subhash C; Stairs, Dustin J; Buonanno, Andres; Dravid, Shashank M

2011-05-01

N-methyl-D-aspartate (NMDA) receptors play an important role in excitatory neurotransmission and mediate synaptic plasticity associated with learning and memory. NMDA receptors are composed of two NR1 and two NR2 subunits and the identity of the NR2 subunit confers unique electrophysiologic and pharmacologic properties to the receptor. The precise role of NR2C-containing receptors in vivo is poorly understood. We have performed a battery of behavioral tests on NR2C knockout/nβ-galactosidase knock-in mice and found no difference in spontaneous activity, basal anxiety, forced-swim immobility, novel object recognition, pain sensitivity and reference memory in comparison to wildtype counterparts. However, NR2C knockout mice were found to exhibit deficits in fear acquisition and working memory compared to wildtype mice. Deficit in fear acquisition correlated with lack of fear conditioning-induced plasticity at the thalamo-amygdala synapse. These findings suggest a unique role of NR2C-containing receptors in associative and executive learning representing a novel therapeutic target for deficits in cognition. Copyright © 2011 Elsevier Inc. All rights reserved.

Double blind, placebo controlled trial of two probiotic strains in interleukin 10 knockout mice and mechanistic link with cytokine balance

NARCIS (Netherlands)

McCarthy, J.; O'Mahony, L.; O'Callaghan, L.; Sheil, B.; Vaughan, E.E.; Fitzsimons, N.A.; Fitzgibbon, J.; O'Sullivan, G.C.; Kiely, B.; Collins, J.K.; Shanahan, F.

2003-01-01

Background: Prophylactic efficacy against colitis following lactobacillus consumption in interleukin 10 (IL-10) knockout ( KO) mice has been reported. Whether this applies equally to other probiotic strains is unknown, and the mechanism is unclear. Aims: ( 1) To compare the effect of feeding

RETINOIC ACID INDUCTION OF CLEFT PALATE IN EGF AND TGF-ALPHA KNOCKOUT MICE: STAGE SPECIFIC INFLUENCES OF GROWTH FACTOR EXPRESSION

Science.gov (United States)

ABBOTT, B. D., LEFFLER, K.E. AND BUCKALEW, A.R, Reproductive Toxicology Division, NHEERL, ORD, US EPA, Research Triangle Park, North Carolina. Retinoic acid induction of cleft palate (CP) in EGF and TGF knockout mice: Stage specific influences of growth factor expression.<...

Enhanced genome editing tools for multi-gene deletion knock-out approaches using paired CRISPR sgRNAs in CHO cells

DEFF Research Database (Denmark)

Schmieder, Valerie; Bydlinski, Nina; Strasser, Richard

2017-01-01

(sgRNAs) for full gene deletions. This strategy also enables the targeting of regulatory regions, which would not respond to the conventional frameshift mutations, as shown by deleting the α-1,6-Fucosyltransferase 8 (FUT8) promoter resulting in a functional knock-out. Fut8 also served as model...

A Convenient Cas9-based Conditional Knockout Strategy for Simultaneously Targeting Multiple Genes in Mouse.

Science.gov (United States)

Chen, Jiang; Du, Yinan; He, Xueyan; Huang, Xingxu; Shi, Yun S

2017-03-31

The most powerful way to probe protein function is to characterize the consequence of its deletion. Compared to conventional gene knockout (KO), conditional knockout (cKO) provides an advanced gene targeting strategy with which gene deletion can be performed in a spatially and temporally restricted manner. However, for most species that are amphiploid, the widely used Cre-flox conditional KO (cKO) system would need targeting loci in both alleles to be loxP flanked, which in practice, requires time and labor consuming breeding. This is considerably significant when one is dealing with multiple genes. CRISPR/Cas9 genome modulation system is advantaged in its capability in targeting multiple sites simultaneously. Here we propose a strategy that could achieve conditional KO of multiple genes in mouse with Cre recombinase dependent Cas9 expression. By transgenic construction of loxP-stop-loxP (LSL) controlled Cas9 (LSL-Cas9) together with sgRNAs targeting EGFP, we showed that the fluorescence molecule could be eliminated in a Cre-dependent manner. We further verified the efficacy of this novel strategy to target multiple sites by deleting c-Maf and MafB simultaneously in macrophages specifically. Compared to the traditional Cre-flox cKO strategy, this sgRNAs-LSL-Cas9 cKO system is simpler and faster, and would make conditional manipulation of multiple genes feasible.

Indexed variation graphs for efficient and accurate resistome profiling.

Science.gov (United States)

Rowe, Will P M; Winn, Martyn D

2018-05-14

Antimicrobial resistance remains a major threat to global health. Profiling the collective antimicrobial resistance genes within a metagenome (the "resistome") facilitates greater understanding of antimicrobial resistance gene diversity and dynamics. In turn, this can allow for gene surveillance, individualised treatment of bacterial infections and more sustainable use of antimicrobials. However, resistome profiling can be complicated by high similarity between reference genes, as well as the sheer volume of sequencing data and the complexity of analysis workflows. We have developed an efficient and accurate method for resistome profiling that addresses these complications and improves upon currently available tools. Our method combines a variation graph representation of gene sets with an LSH Forest indexing scheme to allow for fast classification of metagenomic sequence reads using similarity-search queries. Subsequent hierarchical local alignment of classified reads against graph traversals enables accurate reconstruction of full-length gene sequences using a scoring scheme. We provide our implementation, GROOT, and show it to be both faster and more accurate than a current reference-dependent tool for resistome profiling. GROOT runs on a laptop and can process a typical 2 gigabyte metagenome in 2 minutes using a single CPU. Our method is not restricted to resistome profiling and has the potential to improve current metagenomic workflows. GROOT is written in Go and is available at https://github.com/will-rowe/groot (MIT license). will.rowe@stfc.ac.uk. Supplementary data are available at Bioinformatics online.

Molecular acidity: An accurate description with information-theoretic approach in density functional reactivity theory.

Science.gov (United States)

Cao, Xiaofang; Rong, Chunying; Zhong, Aiguo; Lu, Tian; Liu, Shubin

2018-01-15

Molecular acidity is one of the important physiochemical properties of a molecular system, yet its accurate calculation and prediction are still an unresolved problem in the literature. In this work, we propose to make use of the quantities from the information-theoretic (IT) approach in density functional reactivity theory and provide an accurate description of molecular acidity from a completely new perspective. To illustrate our point, five different categories of acidic series, singly and doubly substituted benzoic acids, singly substituted benzenesulfinic acids, benzeneseleninic acids, phenols, and alkyl carboxylic acids, have been thoroughly examined. We show that using IT quantities such as Shannon entropy, Fisher information, Ghosh-Berkowitz-Parr entropy, information gain, Onicescu information energy, and relative Rényi entropy, one is able to simultaneously predict experimental pKa values of these different categories of compounds. Because of the universality of the quantities employed in this work, which are all density dependent, our approach should be general and be applicable to other systems as well. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Delayed liver regeneration after partial hepatectomy in adiponectin knockout mice

International Nuclear Information System (INIS)

Ezaki, Hisao; Yoshida, Yuichi; Saji, Yukiko; Takemura, Takayo; Fukushima, Juichi; Matsumoto, Hitoshi; Kamada, Yoshihiro; Wada, Akira; Igura, Takumi; Kihara, Shinji; Funahashi, Tohru; Shimomura, Iichiro; Tamura, Shinji; Kiso, Shinichi; Hayashi, Norio

2009-01-01

We previously demonstrated that adiponectin has anti-fibrogenic and anti-inflammatory effects in the liver of mouse models of various liver diseases. However, its role in liver regeneration remains unclear. The aim of this study was to determine the role of adiponectin in liver regeneration. We assessed liver regeneration after partial hepatectomy in wild-type (WT) and adiponectin knockout (KO) mice. We analyzed DNA replication and various signaling pathways involved in cell proliferation and metabolism. Adiponectin KO mice exhibited delayed DNA replication and increased lipid accumulation in the regenerating liver. The expression levels of peroxisome proliferator-activated receptor (PPAR) α and carnitine palmitoyltransferase-1 (CPT-1), a key enzyme in mitochondrial fatty acid oxidation, were decreased in adiponectin KO mice, suggesting possible contribution of altered fat metabolism to these phenomena. Collectively, the present results highlight a new role for adiponectin in the process of liver regeneration.

Shorter duration of non-rapid eye movement sleep slow waves in EphA4 knockout mice.

Science.gov (United States)

Freyburger, Marlène; Poirier, Gaétan; Carrier, Julie; Mongrain, Valérie

2017-10-01

Slow waves occurring during non-rapid eye movement sleep have been associated with neurobehavioural performance and memory. In addition, the duration of previous wakefulness and sleep impacts characteristics of these slow waves. However, molecular mechanisms regulating the dynamics of slow-wave characteristics remain poorly understood. The EphA4 receptor regulates glutamatergic transmission and synaptic plasticity, which have both been linked to sleep slow waves. To investigate if EphA4 regulates slow-wave characteristics during non-rapid eye movement sleep, we compared individual parameters of slow waves between EphA4 knockout mice and wild-type littermates under baseline conditions and after a 6-h sleep deprivation. We observed that, compared with wild-type mice, knockout mice display a shorter duration of positive and negative phases of slow waves under baseline conditions and after sleep deprivation. However, the mutation did not change slow-wave density, amplitude and slope, and did not affect the sleep deprivation-dependent changes in slow-wave characteristics, suggesting that EphA4 is not involved in the response to elevated sleep pressure. Our present findings suggest a role for EphA4 in shaping cortical oscillations during sleep that is independent from sleep need. © 2017 European Sleep Research Society.

Accurate Prediction of Motor Failures by Application of Multi CBM Tools: A Case Study

Science.gov (United States)

Dutta, Rana; Singh, Veerendra Pratap; Dwivedi, Jai Prakash

2018-02-01

Motor failures are very difficult to predict accurately with a single condition-monitoring tool as both electrical and the mechanical systems are closely related. Electrical problem, like phase unbalance, stator winding insulation failures can, at times, lead to vibration problem and at the same time mechanical failures like bearing failure, leads to rotor eccentricity. In this case study of a 550 kW blower motor it has been shown that a rotor bar crack was detected by current signature analysis and vibration monitoring confirmed the same. In later months in a similar motor vibration monitoring predicted bearing failure and current signature analysis confirmed the same. In both the cases, after dismantling the motor, the predictions were found to be accurate. In this paper we will be discussing the accurate predictions of motor failures through use of multi condition monitoring tools with two case studies.

Sub-micrometre accurate free-form optics by three-dimensional printing on single-mode fibres

Science.gov (United States)

Gissibl, Timo; Thiele, Simon; Herkommer, Alois; Giessen, Harald

2016-01-01

Micro-optics are widely used in numerous applications, such as beam shaping, collimation, focusing and imaging. We use femtosecond 3D printing to manufacture free-form micro-optical elements. Our method gives sub-micrometre accuracy so that direct manufacturing even on single-mode fibres is possible. We demonstrate the potential of our method by writing different collimation optics, toric lenses, free-form surfaces with polynomials of up to 10th order for intensity beam shaping, as well as chiral photonic crystals for circular polarization filtering, all aligned onto the core of the single-mode fibres. We determine the accuracy of our optics by analysing the output patterns as well as interferometrically characterizing the surfaces. We find excellent agreement with numerical calculations. 3D printing of microoptics can achieve sufficient performance that will allow for rapid prototyping and production of beam-shaping and imaging devices. PMID:27339700

Prion protein (PrP) gene-knockout cell lines: insight into functions of the PrP

OpenAIRE

Sakudo, Akikazu; Onodera, Takashi

2015-01-01

Elucidation of prion protein (PrP) functions is crucial to fully understand prion diseases. A major approach to studying PrP functions is the use of PrP gene-knockout (Prnp ?/?) mice. So far, six types of Prnp ?/? mice have been generated, demonstrating the promiscuous functions of PrP. Recently, other PrP family members, such as Doppel and Shadoo, have been found. However, information obtained from comparative studies of structural and functional analyses of these PrP family proteins do not ...

On a calculation of nucleon knock-out cross sections in a collision of relativistic nuclei

International Nuclear Information System (INIS)

Goryachev, B.I.; Lin'kova, N.V.

1985-01-01

It is shown that in the framework of the two-stage model one can obtain knock-out cross sections of the given number of nucleons from the nucleus-target at a certain number of nucleons knocked out from the nucleus-projectile. The first stage is considered as a fast process of nucleon collisions of interacting nuclei which is completed with knock out of one or several nucleons. The second stage-comparatively slow - is related to de-excitation of nuclei-fragments

GABA(A)-benzodiazepine receptor complex sensitivity in 5-HT(1A) receptor knockout mice on a 129/Sv background.

NARCIS (Netherlands)

Pattij, T.; Groenink, L.; Oosting, R.S.; Gugten, J. van der; Maes, R.A.A.; Olivier, B.

2002-01-01

Previous studies in 5-HT(1A) receptor knockout (1AKO) mice on a mixed Swiss Websterx129/Sv (SWx129/Sv) and a pure 129/Sv genetic background suggest a differential gamma-aminobutyric acid (GABA(A))-benzodiazepine receptor complex sensitivity in both strains, independent from the anxious phenotype. To

Histone deacetylase 6 inhibition reduces cysts by decreasing cAMP and Ca2+ in knock-out mouse models of polycystic kidney disease.

Science.gov (United States)

Yanda, Murali K; Liu, Qiangni; Cebotaru, Valeriu; Guggino, William B; Cebotaru, Liudmila

2017-10-27

Autosomal dominant polycystic kidney disease (ADPKD) is associated with progressive enlargement of multiple renal cysts, often leading to renal failure that cannot be prevented by a current treatment. Two proteins encoded by two genes are associated with ADPKD: PC1 ( pkd1 ), primarily a signaling molecule, and PC2 ( pkd2 ), a Ca 2+ channel. Dysregulation of cAMP signaling is central to ADPKD, but the molecular mechanism is unresolved. Here, we studied the role of histone deacetylase 6 (HDAC6) in regulating cyst growth to test the possibility that inhibiting HDAC6 might help manage ADPKD. Chemical inhibition of HDAC6 reduced cyst growth in PC1-knock-out mice. In proximal tubule-derived, PC1-knock-out cells, adenylyl cyclase 6 and 3 (AC6 and -3) are both expressed. AC6 protein expression was higher in cells lacking PC1, compared with control cells containing PC1. Intracellular Ca 2+ was higher in PC1-knock-out cells than in control cells. HDAC inhibition caused a drop in intracellular Ca 2+ and increased ATP-simulated Ca 2+ release. HDAC6 inhibition reduced the release of Ca 2+ from the endoplasmic reticulum induced by thapsigargin, an inhibitor of endoplasmic reticulum Ca 2+ -ATPase. HDAC6 inhibition and treatment of cells with the intracellular Ca 2+ chelator 1,2-bis(2-aminophenoxy)ethane- N , N , N ', N '-tetraacetic acid tetrakis(acetoxymethyl ester) reduced cAMP levels in PC1-knock-out cells. Finally, the calmodulin inhibitors W-7 and W-13 reduced cAMP levels, and W-7 reduced cyst growth, suggesting that AC3 is involved in cyst growth regulated by HDAC6. We conclude that HDAC6 inhibition reduces cell growth primarily by reducing intracellular cAMP and Ca 2+ levels. Our results provide potential therapeutic targets that may be useful as treatments for ADPKD. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Cortical Gene Expression After a Conditional Knockout of 67 kDa Glutamic Acid Decarboxylase in Parvalbumin Neurons.

Science.gov (United States)

Georgiev, Danko; Yoshihara, Toru; Kawabata, Rika; Matsubara, Takurou; Tsubomoto, Makoto; Minabe, Yoshio; Lewis, David A; Hashimoto, Takanori

2016-07-01

In the cortex of subjects with schizophrenia, expression of glutamic acid decarboxylase 67 (GAD67), the enzyme primarily responsible for cortical GABA synthesis, is reduced in the subset of GABA neurons that express parvalbumin (PV). This GAD67 deficit is accompanied by lower cortical levels of other GABA-associated transcripts, including GABA transporter-1, PV, brain-derived neurotrophic factor (BDNF), tropomyosin receptor kinase B, somatostatin, GABAA receptor α1 subunit, and KCNS3 potassium channel subunit mRNAs. In contrast, messenger RNA (mRNA) levels for glutamic acid decarboxylase 65 (GAD65), another enzyme for GABA synthesis, are not altered. We tested the hypothesis that this pattern of GABA-associated transcript levels is secondary to the GAD67 deficit in PV neurons by analyzing cortical levels of these GABA-associated mRNAs in mice with a PV neuron-specific GAD67 knockout. Using in situ hybridization, we found that none of the examined GABA-associated transcripts had lower cortical expression in the knockout mice. In contrast, PV, BDNF, KCNS3, and GAD65 mRNA levels were higher in the homozygous mice. In addition, our behavioral test battery failed to detect a change in sensorimotor gating or working memory, although the homozygous mice exhibited increased spontaneous activities. These findings suggest that reduced GAD67 expression in PV neurons is not an upstream cause of the lower levels of GABA-associated transcripts, or of the characteristic behaviors, in schizophrenia. In PV neuron-specific GAD67 knockout mice, increased levels of PV, BDNF, and KCNS3 mRNAs might be the consequence of increased neuronal activity secondary to lower GABA synthesis, whereas increased GAD65 mRNA might represent a compensatory response to increase GABA synthesis. © The Author 2016. Published by Oxford University Press on behalf of the Maryland Psychiatric Research Center. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Accurate geometrical optics model for single-lens stereovision system using a prism.

Science.gov (United States)

Cui, Xiaoyu; Lim, Kah Bin; Guo, Qiyong; Wang, DaoLei

2012-09-01

In this paper, we proposed a new method for analyzing the image formation of a prism. The prism was considered as a single optical system composed of some planes. By analyzing each plane individually and then combining them together, we derived a transformation matrix which can express the relationship between an object point and its image by the refraction of a prism. We also explained how to use this matrix for epipolar geometry and three-dimensional point reconstruction. Our method is based on optical geometry and could be used in a multiocular prism. Experimentation results are presented to prove the accuracy of our method is better than former researchers' and is comparable with that of the multicamera stereovision system.

Hydrogen atoms can be located accurately and precisely by x-ray crystallography.

Science.gov (United States)

Woińska, Magdalena; Grabowsky, Simon; Dominiak, Paulina M; Woźniak, Krzysztof; Jayatilaka, Dylan

2016-05-01

Precise and accurate structural information on hydrogen atoms is crucial to the study of energies of interactions important for crystal engineering, materials science, medicine, and pharmacy, and to the estimation of physical and chemical properties in solids. However, hydrogen atoms only scatter x-radiation weakly, so x-rays have not been used routinely to locate them accurately. Textbooks and teaching classes still emphasize that hydrogen atoms cannot be located with x-rays close to heavy elements; instead, neutron diffraction is needed. We show that, contrary to widespread expectation, hydrogen atoms can be located very accurately using x-ray diffraction, yielding bond lengths involving hydrogen atoms (A-H) that are in agreement with results from neutron diffraction mostly within a single standard deviation. The precision of the determination is also comparable between x-ray and neutron diffraction results. This has been achieved at resolutions as low as 0.8 Å using Hirshfeld atom refinement (HAR). We have applied HAR to 81 crystal structures of organic molecules and compared the A-H bond lengths with those from neutron measurements for A-H bonds sorted into bonds of the same class. We further show in a selection of inorganic compounds that hydrogen atoms can be located in bridging positions and close to heavy transition metals accurately and precisely. We anticipate that, in the future, conventional x-radiation sources at in-house diffractometers can be used routinely for locating hydrogen atoms in small molecules accurately instead of large-scale facilities such as spallation sources or nuclear reactors.

13C(α,n)16O reaction as the knock-out exchange process

International Nuclear Information System (INIS)

Kim, G.; Khajdarov, R.R.; Zaparov, Eh.A.

2000-01-01

S-factor for the 13 C(α,n) 16 O reaction is studied. In the framework of the simple phenomenological model this reaction is analysed as neutron knocked-out by α-particle exchange process. The analysis demonstrates the importance of taking into account 2p-state in 13 C. The 13 C(α,n) 16 O cross section is considered both as the knock-out exchange process and as it's combination with process through a compound nucleus. It was shown that for E α s value extrapolated to low energies is found to be noticeably larger that of R-matrix analysis. Different ways of improving the proposed model are discussed. (author)

Aggravated brain damage after hypoxic ischemia in immature adenosine A2A knockout mice.

Science.gov (United States)

Adén, Ulrika; Halldner, Linda; Lagercrantz, Hugo; Dalmau, Ishar; Ledent, Catherine; Fredholm, Bertil B

2003-03-01

Cerebral hypoxic ischemia (HI) is an important cause of brain injury in the newborn infant. Adenosine is believed to protect against HI brain damage. However, the roles of the different adenosine receptors are unclear, particularly in young animals. We examined the role of adenosine A2A receptors (A2AR) using 7-day-old A2A knockout (A2AR(-/-)) mice in a model of HI. HI was induced in 7-day-old CD1 mice by exposure to 8% oxygen for 30 minutes after occlusion of the left common carotid artery. The resulting unilateral focal lesion was evaluated with the use of histopathological scoring and measurements of residual brain areas at 5 days, 3 weeks, and 3 months after HI. Behavioral evaluation of brain injury by locomotor activity, rotarod, and beam-walking test was made 3 weeks and 3 months after HI. Cortical cerebral blood flow, assessed by laser-Doppler flowmetry, and rectal temperature were measured during HI. Reduction in cortical cerebral blood flow during HI and rectal temperature did not differ between wild-type (A2AR(+/+)) and knockout mice. In the A2AR(-/-) animals, brain injury was aggravated compared with wild-type mice. The A2AR(-/-) mice subjected to HI displayed increased forward locomotion and impaired rotarod performance in adulthood compared with A2AR(+/+) mice subjected to HI, whereas beam-walking performance was similarly defective in both groups. These results suggest that, in contrast to the situation in adult animals, A2AR play an important protective role in neonatal HI brain injury.

Fast and accurate three-dimensional point spread function computation for fluorescence microscopy.

Science.gov (United States)

Li, Jizhou; Xue, Feng; Blu, Thierry

2017-06-01

The point spread function (PSF) plays a fundamental role in fluorescence microscopy. A realistic and accurately calculated PSF model can significantly improve the performance in 3D deconvolution microscopy and also the localization accuracy in single-molecule microscopy. In this work, we propose a fast and accurate approximation of the Gibson-Lanni model, which has been shown to represent the PSF suitably under a variety of imaging conditions. We express the Kirchhoff's integral in this model as a linear combination of rescaled Bessel functions, thus providing an integral-free way for the calculation. The explicit approximation error in terms of parameters is given numerically. Experiments demonstrate that the proposed approach results in a significantly smaller computational time compared with current state-of-the-art techniques to achieve the same accuracy. This approach can also be extended to other microscopy PSF models.

[Effects of aquaporin-4 gene knockout on behavior changes and cerebral morphology during aging in mice].

Science.gov (United States)

Su, Shengan; Lu, Yunbi; Zhang, Weiping

2013-05-01

To investigate the effects of aquaporin-4 (AQP4) gene knockout on the behavior changes and cerebral morphology during aging in mice,and to compare that of young and aged mice between AQP4 knockout mice (AQP4(-/-)) and wild type mice (AQP4(+/+)). Fifty-eight CD-1 mice were divided into four groups: young (2-3 months old) AQP4(-/-), aged (17-19 months old) AQP4(-/-), young AQP4(+/+) and aged AQP4(+/+). The activity levels and exploring behavior of mice were tested in open field. The neurons were stained with toluidine blue and NeuN, the astrocytes and microglia were stained with GFAP and Iba-1, respectively. The morphological changes of neuron, astrocyte and microglia were then analyzed. Compared with young mice, the total walking distance in open field of aged AQP4(+/+) mice and aged AQP4(-/-) mice decreased 41.2% and 44.1%, respectively (Ptime in the central area of open field. The density of neuron in cortex of aged AQP4(+/+) mice and aged AQP4(-/-) mice decreased 19.6% and 15.8%, respectively (P<0.05), while there was no difference in the thickness of neuron cell body in hippocampus CA1 region. The density of astrocyte in hippocampus CA3 region of aged AQP4(+/+) mice and aged AQP4(-/-) mice increased 57.7% and 64.3%, respectively (P<0.001), while there was no difference in the area of astrocyte. The area of microglia in hippocampus CA3 region of aged AQP4(+/+) mice and aged AQP4(-/-) mice increased 46.9% and 52.0%, respectively (P<0.01), while there was no difference in the density of microglia. Compared with AQP4(+/+) mice, the young and aged AQP4(-/-) mice showed smaller area of astrocyte in hippocampus CA3 region, reduced 18.0% in young mice and 23.6% in aged mice. There was no difference between AQP4(+/+) mice and AQP4(-/-) mice for other observed indexes. AQP4 may be involved in change of astrocyte and astrocyte-related behaviors during aging. AQP4 gene knockout may have limited effects on the change of neuron, microglia and most neuronal behaviors in aging

Momentum profile analysis in one-neutron knockout from Borromean nuclei

Energy Technology Data Exchange (ETDEWEB)

Aksyutina, Yu. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, ExtreMe Matter Institute, EMMI, D-64291 Darmstadt (Germany); Aumann, T. [Institut fuer Kernphysik, Technische Universitaet, D-64289 Darmstadt (Germany); Boretzky, K. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, ExtreMe Matter Institute, EMMI, D-64291 Darmstadt (Germany); Borge, M.J.G. [Instituto Estructura de la Materia, CSIC, E-28006 Madrid (Spain); Caesar, C.; Chatillon, A. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, ExtreMe Matter Institute, EMMI, D-64291 Darmstadt (Germany); Chulkov, L.V. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, ExtreMe Matter Institute, EMMI, D-64291 Darmstadt (Germany); Kurchatov Institute, RU-123182 Moscow (Russian Federation); Cortina-Gil, D. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, ExtreMe Matter Institute, EMMI, D-64291 Darmstadt (Germany); University of Santiago de Compostela, 15706 Santiago de Compostela (Spain); Datta Pramanik, U. [Saha Institute of Nuclear Physics, 1/AF Bidhannagar, Kolkata 700064 (India); Emling, H. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, ExtreMe Matter Institute, EMMI, D-64291 Darmstadt (Germany); Fynbo, H.O.U. [Department of Physics and Astronomy, University of Aarhus, DK-8000 Aarhus C (Denmark); Geissel, H.; Ickert, G. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, ExtreMe Matter Institute, EMMI, D-64291 Darmstadt (Germany); Johansson, H.T. [Fundamental Fysik, Chalmers Tekniska Hoegskola, S-412 96 Goeteborg (Sweden); and others

2013-01-29

One-neutron knockout reactions from Borromean nuclei are analyzed using a profile function analysis technique. The profile function, which is derived as the square root of the variance of the measured fragment + neutron momenta as a function of their relative energy, is shown to be very sensitive to the angular momentum of the knocked out neutron. Three cases are studied here: {sup 7}He, where the profile function analysis shows a presence of (s{sub 1/2}){sup 2} component in the {sup 8}He ground-state wave-function, {sup 10}Li, where the presence of a 11(2)% d-wave contribution to the relative energy spectrum above 1.5 MeV is found and, finally, the presence of a major s contribution around 0.5 MeV relative energy in the {sup 13}Be case and that the observed decay to the {sup 12}Be 2{sup +} state originates in a d state in {sup 13}Be.

Smooth muscle cells in atherosclerosis originate from the local vessel wall and not circulating progenitor cells in ApoE knockout mice

DEFF Research Database (Denmark)

Bentzon, Jacob Fog; Weile, Charlotte; Sondergaard, Claus S

2006-01-01

Recent studies of bone marrow (BM)-transplanted apoE knockout (apoE-/-) mice have concluded that a substantial fraction of smooth muscle cells (SMCs) in atherosclerosis arise from circulating progenitor cells of hematopoietic origin. This pathway, however, remains controversial. In the present st...

Simple, fast and accurate two-diode model for photovoltaic modules

Energy Technology Data Exchange (ETDEWEB)

Ishaque, Kashif; Salam, Zainal; Taheri, Hamed [Faculty of Electrical Engineering, Universiti Teknologi Malaysia, UTM 81310, Skudai, Johor Bahru (Malaysia)

2011-02-15

This paper proposes an improved modeling approach for the two-diode model of photovoltaic (PV) module. The main contribution of this work is the simplification of the current equation, in which only four parameters are required, compared to six or more in the previously developed two-diode models. Furthermore the values of the series and parallel resistances are computed using a simple and fast iterative method. To validate the accuracy of the proposed model, six PV modules of different types (multi-crystalline, mono-crystalline and thin-film) from various manufacturers are tested. The performance of the model is evaluated against the popular single diode models. It is found that the proposed model is superior when subjected to irradiance and temperature variations. In particular the model matches very accurately for all important points of the I-V curves, i.e. the peak power, short-circuit current and open circuit voltage. The modeling method is useful for PV power converter designers and circuit simulator developers who require simple, fast yet accurate model for the PV module. (author)

Obese Neuronal PPARγ Knockout Mice Are Leptin Sensitive but Show Impaired Glucose Tolerance and Fertility.

Science.gov (United States)

Fernandez, Marina O; Sharma, Shweta; Kim, Sun; Rickert, Emily; Hsueh, Katherine; Hwang, Vicky; Olefsky, Jerrold M; Webster, Nicholas J G

2017-01-01

The peroxisome-proliferator activated receptor γ (PPARγ) is expressed in the hypothalamus in areas involved in energy homeostasis and glucose metabolism. In this study, we created a deletion of PPARγ brain-knockout (BKO) in mature neurons in female mice to investigate its involvement in metabolism and reproduction. We observed that there was no difference in age at puberty onset between female BKOs and littermate controls, but the BKOs gave smaller litters when mated and fewer oocytes when ovulated. The female BKO mice had regular cycles but showed an increase in the number of cycles with prolonged estrus. The mice also had increased luteinizing hormone (LH) levels during the LH surge and histological examination showed hemorrhagic corpora lutea. The mice were challenged with a 60% high-fat diet (HFD). Metabolically, the female BKO mice showed normal body weight, glucose and insulin tolerance, and leptin levels but were protected from obesity-induced leptin resistance. The neuronal knockout also prevented the reduction in estrous cycles due to the HFD. Examination of ovarian histology showed a decrease in the number of primary and secondary follicles in both genotypes due to the HFD, but the BKO ovaries showed an increase in the number of hemorrhagic follicles. In summary, our results show that neuronal PPARγ is required for optimal female fertility but is also involved in the adverse effects of diet-induced obesity by creating leptin resistance potentially through induction of the repressor Socs3. Copyright © 2017 by the Endocrine Society.

A single-aliquot OSL protocol using bracketing regenerative doses to accurately determine equivalent doses in quartz

CERN Document Server

Folz, E

1999-01-01

In most cases, sediments show inherent heterogeneity in their luminescence behaviours and bleaching histories, and identical aliquots are not available: single-aliquot determination of the equivalent dose (ED) is then the approach of choice and the advantages of using regenerative protocols are outlined. Experiments on five laboratory bleached and dosed quartz samples, following the protocol described by Murray and Roberts (1998. Measurement of the equivalent dose in quartz using a regenerative-dose single aliquot protocol. Radiation Measurements 27, 171-184), showed the hazards of using a single regeneration dose: a 10% variation in the regenerative dose yielded some equivalent dose estimates that differed from the expected value by more than 5%. A protocol is proposed that allows the use of different regenerative doses to bracket the estimated equivalent dose. The measured ED is found to be in excellent agreement with the known value when the main regeneration dose is within 10% of the true equivalent dose.

A single-aliquot OSL protocol using bracketing regenerative doses to accurately determine equivalent doses in quartz

International Nuclear Information System (INIS)

Folz, Elise; Mercier, Norbert

1999-01-01

In most cases, sediments show inherent heterogeneity in their luminescence behaviours and bleaching histories, and identical aliquots are not available: single-aliquot determination of the equivalent dose (ED) is then the approach of choice and the advantages of using regenerative protocols are outlined. Experiments on five laboratory bleached and dosed quartz samples, following the protocol described by Murray and Roberts (1998. Measurement of the equivalent dose in quartz using a regenerative-dose single aliquot protocol. Radiation Measurements 27, 171-184), showed the hazards of using a single regeneration dose: a 10% variation in the regenerative dose yielded some equivalent dose estimates that differed from the expected value by more than 5%. A protocol is proposed that allows the use of different regenerative doses to bracket the estimated equivalent dose. The measured ED is found to be in excellent agreement with the known value when the main regeneration dose is within 10% of the true equivalent dose

The testosterone-dependent and independent transcriptional networks in the hypothalamus of Gpr54 and Kiss1 knockout male mice are not fully equivalent

Directory of Open Access Journals (Sweden)

Sutcliffe Margaret

2011-04-01

Full Text Available Abstract Background Humans and mice with loss of function mutations in GPR54 (KISS1R or kisspeptin do not progress through puberty, caused by a failure to release GnRH. The transcriptional networks regulated by these proteins in the hypothalamus have yet to be explored by genome-wide methods. Results We show here, using 1 million exon mouse arrays (Exon 1.0 Affymetrix and quantitative polymerase chain reaction (QPCR validation to analyse microdissected hypothalamic tissue from Gpr54 and Kiss1 knockout mice, the extent of transcriptional regulation in the hypothalamus. The sensitivity to detect important transcript differences in microdissected RNA was confirmed by the observation of counter-regulation of Kiss1 expression in Gpr54 knockouts and confirmed by immunohistochemistry (IHC. Since Gpr54 and Kiss1 knockout animals are effectively pre-pubertal with low testosterone (T levels, we also determined which of the validated transcripts were T-responsive and which varied according to genotype alone. We observed four types of transcriptional regulation (i genotype only dependent regulation, (ii T only dependent regulation, (iii genotype and T-dependent regulation with interaction between these variables, (iv genotype and T-dependent regulation with no interaction between these variables. The results implicate for the first time several transcription factors (e.g. Npas4, Esr2, proteases (Klk1b22, and the orphan 10-transmembrane transporter TMEM144 in the biology of GPR54/kisspeptin function in the hypothalamus. We show for the neuronal activity regulated transcription factor NPAS4, that distinct protein over-expression is seen in the hypothalamus and hippocampus in Gpr54 knockout mice. This links for the first time the hypothalamic-gonadal axis with this important regulator of inhibitory synapse formation. Similarly we confirm TMEM144 up-regulation in the hypothalamus by RNA in situ hybridization and western blot. Conclusions Taken together, global

Accurate RNA consensus sequencing for high-fidelity detection of transcriptional mutagenesis-induced epimutations.

Science.gov (United States)

Reid-Bayliss, Kate S; Loeb, Lawrence A

2017-08-29

Transcriptional mutagenesis (TM) due to misincorporation during RNA transcription can result in mutant RNAs, or epimutations, that generate proteins with altered properties. TM has long been hypothesized to play a role in aging, cancer, and viral and bacterial evolution. However, inadequate methodologies have limited progress in elucidating a causal association. We present a high-throughput, highly accurate RNA sequencing method to measure epimutations with single-molecule sensitivity. Accurate RNA consensus sequencing (ARC-seq) uniquely combines RNA barcoding and generation of multiple cDNA copies per RNA molecule to eliminate errors introduced during cDNA synthesis, PCR, and sequencing. The stringency of ARC-seq can be scaled to accommodate the quality of input RNAs. We apply ARC-seq to directly assess transcriptome-wide epimutations resulting from RNA polymerase mutants and oxidative stress.

Physical mechanisms of single-event effects in advanced microelectronics

Energy Technology Data Exchange (ETDEWEB)

Schrimpf, Ronald D. [Electrical Engineering and Computer Science, Vanderbilt University, 5635 Stevenson Center, Nashville, TN 37235 (United States)]. E-mail: ron.schrimpf@vanderbilt.edu; Weller, Robert A. [Electrical Engineering and Computer Science, Vanderbilt University, 5635 Stevenson Center, Nashville, TN 37235 (United States); Mendenhall, Marcus H. [Free Electron Laser Center, Vanderbilt University, Station B 351816, Nashville, TN 37235 (United States); Reed, Robert A. [Electrical Engineering and Computer Science, Vanderbilt University, 5635 Stevenson Center, Nashville, TN 37235 (United States); Massengill, Lloyd W. [Electrical Engineering and Computer Science, Vanderbilt University, 5635 Stevenson Center, Nashville, TN 37235 (United States)

2007-08-15

The single-event error rate in advanced semiconductor technologies can be estimated more accurately than conventional methods by using simulation based on accurate descriptions of a large number of individual particle interactions. The results can be used to select the ion types and energies for accelerator testing and to identify situations in which nuclear reactions will contribute to the error rate.

Physical mechanisms of single-event effects in advanced microelectronics

International Nuclear Information System (INIS)

Schrimpf, Ronald D.; Weller, Robert A.; Mendenhall, Marcus H.; Reed, Robert A.; Massengill, Lloyd W.

2007-01-01

The single-event error rate in advanced semiconductor technologies can be estimated more accurately than conventional methods by using simulation based on accurate descriptions of a large number of individual particle interactions. The results can be used to select the ion types and energies for accelerator testing and to identify situations in which nuclear reactions will contribute to the error rate

Dual gene activation and knockout screen reveals directional dependencies in genetic networks. | Office of Cancer Genomics

Science.gov (United States)

Understanding the direction of information flow is essential for characterizing how genetic networks affect phenotypes. However, methods to find genetic interactions largely fail to reveal directional dependencies. We combine two orthogonal Cas9 proteins from Streptococcus pyogenes and Staphylococcus aureus to carry out a dual screen in which one gene is activated while a second gene is deleted in the same cell. We analyze the quantitative effects of activation and knockout to calculate genetic interaction and directionality scores for each gene pair.

PKD1 Mono-Allelic Knockout Is Sufficient to Trigger Renal Cystogenesis in a Mini-Pig Model

OpenAIRE

He, Jin; Li, Qiuyan; Fang, Suyun; Guo, Ying; Liu, Tongxin; Ye, Jianhua; Yu, Zhengquan; Zhang, Ran; Zhao, Yaofeng; Hu, Xiaoxiang; Bai, Xueyuan; Chen, Xiangmei; Li, Ning

2015-01-01

PKD1 and PKD2 mutations could lead to autosomal dominant polycystic kidney disease (ADPKD), which afflicts millions of people worldwide. Due to the marked differences in the lifespan, size, anatomy, and physiology from humans, rodent ADPKD models cannot fully mimic the disease. To obtain a large animal model that recapitulates the disease, we constructed a mini-pig model by mono-allelic knockout (KO) of PKD1 using zinc finger nuclease. The mono-allelic KO pigs had lower PKD1 expression than t...

Efficient gene knock-out and knock-in with transgenic Cas9 in Drosophila.

Science.gov (United States)

Xue, Zhaoyu; Ren, Mengda; Wu, Menghua; Dai, Junbiao; Rong, Yikang S; Gao, Guanjun

2014-03-21

Bacterial Cas9 nuclease induces site-specific DNA breaks using small gRNA as guides. Cas9 has been successfully introduced into Drosophila for genome editing. Here, we improve the versatility of this method by developing a transgenic system that expresses Cas9 in the Drosophila germline. Using this system, we induced inheritable knock-out mutations by injecting only the gRNA into embryos, achieved highly efficient mutagenesis by expressing gRNA from the promoter of a novel non-coding RNA gene, and recovered homologous recombination-based knock-in of a fluorescent marker at a rate of 4.5% by co-injecting gRNA with a circular DNA donor. Copyright © 2014 Xue et al.

Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection.

Science.gov (United States)

Baba, Tomoya; Ara, Takeshi; Hasegawa, Miki; Takai, Yuki; Okumura, Yoshiko; Baba, Miki; Datsenko, Kirill A; Tomita, Masaru; Wanner, Barry L; Mori, Hirotada

2006-01-01

We have systematically made a set of precisely defined, single-gene deletions of all nonessential genes in Escherichia coli K-12. Open-reading frame coding regions were replaced with a kanamycin cassette flanked by FLP recognition target sites by using a one-step method for inactivation of chromosomal genes and primers designed to create in-frame deletions upon excision of the resistance cassette. Of 4288 genes targeted, mutants were obtained for 3985. To alleviate problems encountered in high-throughput studies, two independent mutants were saved for every deleted gene. These mutants-the 'Keio collection'-provide a new resource not only for systematic analyses of unknown gene functions and gene regulatory networks but also for genome-wide testing of mutational effects in a common strain background, E. coli K-12 BW25113. We were unable to disrupt 303 genes, including 37 of unknown function, which are candidates for essential genes. Distribution is being handled via GenoBase (http://ecoli.aist-nara.ac.jp/).

Development of Murine Cyp3a Knockout Chimeric Mice with Humanized Liver.

Science.gov (United States)

Kato, Kota; Ohbuchi, Masato; Hamamura, Satoko; Ohshita, Hiroki; Kazuki, Yasuhiro; Oshimura, Mitsuo; Sato, Koya; Nakada, Naoyuki; Kawamura, Akio; Usui, Takashi; Kamimura, Hidetaka; Tateno, Chise

2015-08-01

We developed murine CYP3A knockout ko chimeric mice with humanized liver expressing human P450S similar to those in humans and whose livers and small intestines do not express murine CYP3A this: approach may overcome effects of residual mouse metabolic enzymes like Cyp3a in conventional chimeric mice with humanized liver, such as PXB-mice [urokinase plasminogen activator/severe combined immunodeficiency (uPA/SCID) mice repopulated with over 70% human hepatocytes] to improve the prediction of drug metabolism and pharmacokinetics in humans. After human hepatocytes were transplanted into Cyp3a KO/uPA/SCID host mice, human albumin levels logarithmically increased until approximately 60 days after transplantation, findings similar to those in PXB-mice. Quantitative real-time-polymerase chain reaction analyses showed that hepatic human P450s, UGTs, SULTs, and transporters mRNA expression levels in Cyp3a KO chimeric mice were also similar to those in PXB-mice and confirmed the absence of Cyp3a11 mRNA expression in mouse liver and intestine. Findings for midazolam and triazolam metabolic activities in liver microsomes were comparable between Cyp3a KO chimeric mice and PXB-mice. In contrast, these activities in the intestine of Cyp3a KO chimeric mice were attenuated compared with PXB-mice. Owing to the knockout of murine Cyp3a, hepatic Cyp2b10 and 2c55 mRNA levels in Cyp3a KO/uPA/SCID mice (without hepatocyte transplants) were 8.4- and 61-fold upregulated compared with PXB-mice, respectively. However, human hepatocyte transplantation successfully restored Cyp2b10 level nearly fully and Cyp2c55 level partly (still 13-fold upregulated) compared with those in PXB-mice. Intestinal Cyp2b10 and 2c55 were also repressed by human hepatocyte transplantation in Cyp3a KO chimeric mice. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

Sarcocystis pantherophis, n. sp. from eastern rat snakes (Pantherophis alleghaniensis) definitive hosts and interferongamma gene knockout mice as experimental intermediate hosts

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Here we report a new species, Sarcocystis pantherophisi with the Eastern rat snake (Pantherophis alleghaniensis) as natural definitive host and the interferon gamma gene knockout (KO) mouse as the experimental intermediate host. Sporocysts (n=15) from intestinal contents of the snake were 17.3 x 10....

RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome

Directory of Open Access Journals (Sweden)

Dewey Colin N

2011-08-01

Full Text Available Abstract Background RNA-Seq is revolutionizing the way transcript abundances are measured. A key challenge in transcript quantification from RNA-Seq data is the handling of reads that map to multiple genes or isoforms. This issue is particularly important for quantification with de novo transcriptome assemblies in the absence of sequenced genomes, as it is difficult to determine which transcripts are isoforms of the same gene. A second significant issue is the design of RNA-Seq experiments, in terms of the number of reads, read length, and whether reads come from one or both ends of cDNA fragments. Results We present RSEM, an user-friendly software package for quantifying gene and isoform abundances from single-end or paired-end RNA-Seq data. RSEM outputs abundance estimates, 95% credibility intervals, and visualization files and can also simulate RNA-Seq data. In contrast to other existing tools, the software does not require a reference genome. Thus, in combination with a de novo transcriptome assembler, RSEM enables accurate transcript quantification for species without sequenced genomes. On simulated and real data sets, RSEM has superior or comparable performance to quantification methods that rely on a reference genome. Taking advantage of RSEM's ability to effectively use ambiguously-mapping reads, we show that accurate gene-level abundance estimates are best obtained with large numbers of short single-end reads. On the other hand, estimates of the relative frequencies of isoforms within single genes may be improved through the use of paired-end reads, depending on the number of possible splice forms for each gene. Conclusions RSEM is an accurate and user-friendly software tool for quantifying transcript abundances from RNA-Seq data. As it does not rely on the existence of a reference genome, it is particularly useful for quantification with de novo transcriptome assemblies. In addition, RSEM has enabled valuable guidance for cost

Progressive hearing loss and degeneration of hair cell stereocilia in taperin gene knockout mice

International Nuclear Information System (INIS)

Chen, Mo; Wang, Qin; Zhu, Gang-Hua; Hu, Peng; Zhou, Yuan; Wang, Tian; Lai, Ruo-Sha; Xiao, Zi-An; Xie, Ding-Hua

2016-01-01

The TPRN gene encodes taperin, which is prominently present at the taper region of hair cell stereocilia. Mutations in TPRN have been reported to cause autosomal recessive nonsyndromic deafness 79(DFNB 79). To investigate the role of taperin in pathogenesis of hearing loss, we generated TPRN knockout mice using TALEN technique. Sanger sequencing confirmed an 11 bp deletion at nucleotide 177–187 in exon 1 of TPRN, which results in a truncated form of taperin protein. Heterozygous TPRN +/− mice showed apparently normal auditory phenotypes to their wide-type (WT) littermates. Homozygous TPRN −/− mice exhibited progressive sensorineural hearing loss as reflected by auditory brainstem response to both click and tone burst stimuli at postnatal days 15 (P15), 30 (P30), and 60 (P60). Alex Fluor-594 phalloidin labeling showed no obvious difference in hair cell numbers in the cochlea between TPRN −/− mice and WT mice under light microscope. However, scanning electronic microscopy revealed progressive degeneration of inner hair cell stereocilia, from apparently normal at postnatal days 3 (P3) to scattered absence at P15 and further to substantial loss at P30. The outer hair cell stereocilia also showed progressive degeneration, though much less severe, Collectively, we conclude that taperin plays an important role in maintenance of hair cell stereocilia. Establishment of TPRN knockout mice enables further investigation into the function of this gene. - Highlights: • TPRN −/− mice were generated using TALEN technique. • TPRN −/− mice presented progressive hearing loss. • WT and TPRN −/− mice showed no difference in hair cell numbers. • TPRN −/− mice showed progressive degeneration of hair cell stereocilia.

Evaluation of organ-specific glucose metabolism by 18F-FDG in insulin receptor substrate-1 (IRS-1) knockout mice as a model of insulin resistance

International Nuclear Information System (INIS)

Cheng, Chao; Nakamura, Akinobu; Minamimoto, Ryogo; Shinoda, Kazuaki; Tateishi, Ukihide; Terauchi, Yasuo; Inoue, Tomio; Goto, Atsuhi; Kadowaki, Takashi

2011-01-01

Insulin resistance (IR) is a physiological condition in which the body produces insulin but does not result in a sufficient biological effect. Insulin resistance is usually asymptomatic but is associated with health problems and is a factor in the metabolic syndrome. The aim of the present study is to clarify organ-specific insulin resistance in normal daily conditions using [ 18 F]-2-fluoro-2-deoxy-D-glucose ([ 18 F]-FDG). The biodistribution of [ 18 F]-FDG was examined in insulin receptor substrate-1 (IRS-1) knockout mice, an animal model of skeletal muscle insulin resistance, and C57BL/6J (wild-type) mice with and without insulin loading. Mice received 0.5 MBq of [ 18 F]-FDG injected into the tail vein, immediately followed by nothing (control cohorts) or an intraperitoneal injection of 1.5 mU/g body weight of human insulin as an insulin loading test. Blood glucose concentrations for all of the experimental animals were assessed at 0, 20, 40, and 60 min post-injection. The mice were subsequently killed, and tissue was collected for evaluation of [ 18 F]-FDG biodistribution. The radioactivity of each organ was measured using a gamma counter. In the absence of insulin, the blood glucose concentrations of wild-type mice (132±26 mg/dl) and IRS-1 knockout mice (134±18 mg/dl) were not significantly different. Blood glucose concentrations decreased following insulin administration, with lower concentrations in wild-type mice than in knockout mice at 20, 40, and 60 min. A statistically significant difference in [ 18 F]-FDG uptake between wild-type mice and IRS-1 knockout mice was confirmed in the heart, abdominal muscle, and femoral muscle. With insulin loading, [ 18 F]-FDG uptake in the heart, back muscle, and abdominal muscle was significantly increased compared to without insulin loading in both wild-type mice and knockout mice. Our results showed that IR significantly affected [ 18 F]-FDG uptake in the heart in normal daily conditions. IR was associated with

Direct detection of single-nucleotide polymorphisms in bacterial DNA by SNPtrap

DEFF Research Database (Denmark)

Grønlund, Hugo Ahlm; Moen, Birgitte; Hoorfar, Jeffrey

2011-01-01

A major challenge with single-nucleotide polymorphism (SNP) fingerprinting of bacteria and higher organisms is the combination of genome-wide screenings with the potential of multiplexing and accurate SNP detection. Single-nucleotide extension by the minisequencing principle represents a technolo...

Polyomic profiling reveals significant hepatic metabolic alterations in glucagon-receptor (GCGR knockout mice: implications on anti-glucagon therapies for diabetes

Directory of Open Access Journals (Sweden)

Molloy Mark P

2011-06-01

Full Text Available Abstract Background Glucagon is an important hormone in the regulation of glucose homeostasis, particularly in the maintenance of euglycemia and prevention of hypoglycemia. In type 2 Diabetes Mellitus (T2DM, glucagon levels are elevated in both the fasted and postprandial states, which contributes to inappropriate hyperglycemia through excessive hepatic glucose production. Efforts to discover and evaluate glucagon receptor antagonists for the treatment of T2DM have been ongoing for approximately two decades, with the challenge being to identify an agent with appropriate pharmaceutical properties and efficacy relative to potential side effects. We sought to determine the hepatic & systemic consequence of full glucagon receptor antagonism through the study of the glucagon receptor knock-out mouse (Gcgr-/- compared to wild-type littermates. Results Liver transcriptomics was performed using Affymetric expression array profiling, and liver proteomics was performed by iTRAQ global protein analysis. To complement the transcriptomic and proteomic analyses, we also conducted metabolite profiling (~200 analytes using mass spectrometry in plasma. Overall, there was excellent concordance (R = 0.88 for changes associated with receptor knock-out between the transcript and protein analysis. Pathway analysis tools were used to map the metabolic processes in liver altered by glucagon receptor ablation, the most notable being significant down-regulation of gluconeogenesis, amino acid catabolism, and fatty acid oxidation processes, with significant up-regulation of glycolysis, fatty acid synthesis, and cholesterol biosynthetic processes. These changes at the level of the liver were manifested through an altered plasma metabolite profile in the receptor knock-out mice, e.g. decreased glucose and glucose-derived metabolites, and increased amino acids, cholesterol, and bile acid levels. Conclusions In sum, the results of this study suggest that the complete ablation

Structure of the unbound nucleus 13Be: One-neutron knockout reaction data from 14Be analyzed in a holistic approach

DEFF Research Database (Denmark)

Aksyutina, Yu; Aumann, T.; Boretzky, K.

2013-01-01

At the ALADIN-LAND setup at GSI the unbound nucleus 13Be has been produced in one-neutron knockout reactions from a 304 MeV/nucleon relativistic beam of 14Be ions impinging on a liquid hydrogen target. An analysis of the data including all available information about 13Be, and in particular recen...

Global gene expression profiles of MT knockout and wild-type mice in the condition of doxorubicin-induced cardiomyopathy.

Science.gov (United States)

Shuai, Yi; Guo, Jun; Dong, Yansheng; Zhong, Weijian; Xiao, Ping; Zhou, Tong; Zhang, Lishi; Peng, Shuangqing

2011-01-15

Increasing evidence from in vivo and in vitro studies has indicated that MT exerts protective effects against DOX-induced cardiotoxicity; however the underlying precise mechanisms still remain an enigma. Therefore, the present study was designed using MT knockout mice in concert with genomic approaches to explore the possible molecular and cellular mechanisms in terms of the genetic network changes. MT-I/II null (MT⁻/⁻) mice and corresponding wild-type mice (MT+/+) were administrated with a single dose of DOX (15 mg/kg, i.p.) or equal volume of saline. Animals were sacrificed on the 4th day after DOX administration and samples were collected for further analyses. Global gene expression profiles of cardiac mRNA from two genotype mice revealed that 381 characteristically MT-responsive genes were identified between MT+/+ mice and MT⁻/⁻ mice in response to DOX, including fos, ucp3, car3, atf3, map3k6, etc. Functional analysis implied MAPK signaling pathway, p53 signaling pathway, Jak-STAT signaling pathway, PPAR signaling pathway, Wnt signaling pathway, etc. might be involved to mediate the protection of DOX cardiomyopathy by MT. Results from the present study not only validated the previously reported possible mechanisms of MT protection against DOX toxicity, but also provided new clues into the molecular mechanisms involved in this process. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

PrPC displays an essential protective role from oxidative stress in an astrocyte cell line derived from PrPC knockout mice

International Nuclear Information System (INIS)

Bertuchi, Fernanda R.; Bourgeon, Dominique M.G.; Landemberger, Michele C.; Martins, Vilma R.; Cerchiaro, Giselle

2012-01-01

Highlights: ► PrP C in solution acts as a radical scavenger. ► PrP C reduces hydrogen peroxide toxicity in astrocytes. ► Increase in ROS disrupted the cell cycle in the PrP C -knockout astrocytes. ► PrP C prevents the cell death independently of an SOD-like activity. -- Abstract: The PrP C protein, which is especially present in the cellular membrane of nervous system cells, has been extensively studied for its controversial antioxidant activity. In this study, we elucidated the free radical scavenger activity of purified murine PrP C in solution and its participation as a cell protector in astrocytes that were subjected to treatment with an oxidant. In vitro and using an EPR spin-trapping technique, we observed that PrP C decreased the oxidation of the DMPO trap in a Fenton reaction system (Cu 2+ /ascorbate/H 2 O 2 ), which was demonstrated by approximately 70% less DMPO/OH · . In cultured PrP C -knockout astrocytes from mice, the absence of PrP C caused an increase in intracellular ROS (reactive oxygen species) generation during the first 3 h of H 2 O 2 treatment. This rapid increase in ROS disrupted the cell cycle in the PrP C -knockout astrocytes, which increased the population of cells in the sub-G1 phase when compared with cultured wild-type astrocytes. We conclude that PrP C in solution acts as a radical scavenger, and in astrocytes, it is essential for protection from oxidative stress caused by an external chemical agent, which is a likely condition in human neurodegenerative CNS disorders and pathological conditions such as ischemia.

Effects of long- and short-term darbepoetin-α treatment on oxidative stress, inflammation and endothelial injury in ApoE knockout mice.

Science.gov (United States)

Özdemir, Evrim Dursun; Hanikoglu, Aysegul; Cort, Aysegul; Ozben, Beste; Suleymanlar, Gultekin; Ozben, Tomris

2017-07-01

Atherosclerosis and atherosclerosis-related complications are the main cause of death in the world. Vascular injury in response to inflammation and enhanced oxidant stress promotes endothelial dysfunction and leads to atherosclerotic lesions. Low-dose treatment with darbepoetin-α may be a potential therapeutic tool for endothelial injury and atherosclerosis. In order to study the effect of darbepoetin-α on endothelial injury and atherosclerosis, we used ApoE-/- mice as the atherosclerotic mice model. We monitored atherosclerosis and plaque formation histochemically in ApoE knockout mice at early and late stages of atherosclerosis. Darbepoetin-α was injected intraperitoneally at a dose of 0.1 μg/kg to ApoE-/- mice. The results of 2 ApoE-/- mice groups injected with darbepoetin-α (early and late stages of atherosclerosis) were compared to the results of the corresponding saline injected ApoE-/- mice groups and the control (C57BL/6) mice. Lipid profile (total cholesterol, triglyceride), inflammation (CRP, IL-6, histamine), endothelial injury (ICAM-1, selectin) and oxidative stress markers (lipid peroxidation, protein oxidation) were significantly increased in 4 atherosclerotic groups compared to the control group. Short-term darbepoetin-α had no marked effects on indicators of inflammation and endothelial injury in the ApoE knockout mice groups compared to the ApoE knockout mice not treated with darbepoetin-α, however, darbepoetin-α significantly decreased 8-isoprostane and protein carbonyl content. Long term darbepoetin-α treatment reduced oxidative stress in ApoE-/- mice. This study contributes to understanding and elucidating the biochemical changes occurring during early and late stages of atherosclerosis development regarding lipid profile, inflammation, endothelial injury and oxidative stress markers.

Accurate artificial boundary conditions for the semi-discretized linear Schrödinger and heat equations on rectangular domains

Science.gov (United States)

Ji, Songsong; Yang, Yibo; Pang, Gang; Antoine, Xavier

2018-01-01

The aim of this paper is to design some accurate artificial boundary conditions for the semi-discretized linear Schrödinger and heat equations in rectangular domains. The Laplace transform in time and discrete Fourier transform in space are applied to get Green's functions of the semi-discretized equations in unbounded domains with single-source. An algorithm is given to compute these Green's functions accurately through some recurrence relations. Furthermore, the finite-difference method is used to discretize the reduced problem with accurate boundary conditions. Numerical simulations are presented to illustrate the accuracy of our method in the case of the linear Schrödinger and heat equations. It is shown that the reflection at the corners is correctly eliminated.

Fast and accurate methods for the performance testing of highly-efficient c-Si photovoltaic modules using a 10 ms single-pulse solar simulator and customized voltage profiles

International Nuclear Information System (INIS)

Virtuani, A; Rigamonti, G; Friesen, G; Chianese, D; Beljean, P

2012-01-01

Performance testing of highly efficient, highly capacitive c-Si modules with pulsed solar simulators requires particular care. These devices in fact usually require a steady-state solar simulator or pulse durations longer than 100–200 ms in order to avoid measurement artifacts. The aim of this work was to validate an alternative method for the testing of highly capacitive c-Si modules using a 10 ms single pulse solar simulator. Our approach attempts to reconstruct a quasi-steady-state I–V (current–voltage) curve of a highly capacitive device during one single 10 ms flash by applying customized voltage profiles–-in place of a conventional V ramp—to the terminals of the device under test. The most promising results were obtained by using V profiles which we name ‘dragon-back’ (DB) profiles. When compared to the reference I–V measurement (obtained by using a multi-flash approach with approximately 20 flashes), the DB V profile method provides excellent results with differences in the estimation of P max (as well as of I sc , V oc and FF) below ±0.5%. For the testing of highly capacitive devices the method is accurate, fast (two flashes—possibly one—required), cost-effective and has proven its validity with several technologies making it particularly interesting for in-line testing. (paper)

Benchmarking density-functional-theory calculations of rotational g tensors and magnetizabilities using accurate coupled-cluster calculations.

Science.gov (United States)

Lutnaes, Ola B; Teale, Andrew M; Helgaker, Trygve; Tozer, David J; Ruud, Kenneth; Gauss, Jürgen

2009-10-14

An accurate set of benchmark rotational g tensors and magnetizabilities are calculated using coupled-cluster singles-doubles (CCSD) theory and coupled-cluster single-doubles-perturbative-triples [CCSD(T)] theory, in a variety of basis sets consisting of (rotational) London atomic orbitals. The accuracy of the results obtained is established for the rotational g tensors by careful comparison with experimental data, taking into account zero-point vibrational corrections. After an analysis of the basis sets employed, extrapolation techniques are used to provide estimates of the basis-set-limit quantities, thereby establishing an accurate benchmark data set. The utility of the data set is demonstrated by examining a wide variety of density functionals for the calculation of these properties. None of the density-functional methods are competitive with the CCSD or CCSD(T) methods. The need for a careful consideration of vibrational effects is clearly illustrated. Finally, the pure coupled-cluster results are compared with the results of density-functional calculations constrained to give the same electronic density. The importance of current dependence in exchange-correlation functionals is discussed in light of this comparison.

Skeletal Muscle Fibre-Specific Knockout of p53 Does Not Reduce Mitochondrial Content or Enzyme Activity

Directory of Open Access Journals (Sweden)

Ben Stocks

2017-12-01

Full Text Available Tumour protein 53 (p53 has been implicated in the regulation of mitochondrial biogenesis in skeletal muscle, with whole-body p53 knockout mice displaying impairments in basal mitochondrial content, respiratory capacity, and enzyme activity. This study aimed to determine the effect of skeletal muscle-specific loss of p53 on mitochondrial content and enzyme activity. Mitochondrial protein content, enzyme activity and mRNA profiles were assessed in skeletal muscle of 8-week-old male muscle fibre-specific p53 knockout mice (p53 mKO and floxed littermate controls (WT under basal conditions. p53 mKO and WT mice displayed similar content of electron transport chain proteins I-V and citrate synthase enzyme activity in skeletal muscle. In addition, the content of proteins regulating mitochondrial morphology (MFN2, mitofillin, OPA1, DRP1, FIS1, fatty acid metabolism (β-HAD, ACADM, ACADL, ACADVL, carbohydrate metabolism (HKII, PDH, energy sensing (AMPKα2, AMPKβ2, and gene transcription (NRF1, PGC-1α, and TFAM were comparable in p53 mKO and WT mice (p > 0.05. Furthermore, p53 mKO mice exhibited normal mRNA profiles of targeted mitochondrial, metabolic and transcriptional proteins (p > 0.05. Thus, it appears that p53 expression in skeletal muscle fibres is not required to develop or maintain mitochondrial protein content or enzyme function in skeletal muscle under basal conditions.

Towards accurate emergency response behavior

International Nuclear Information System (INIS)

Sargent, T.O.

1981-01-01

Nuclear reactor operator emergency response behavior has persisted as a training problem through lack of information. The industry needs an accurate definition of operator behavior in adverse stress conditions, and training methods which will produce the desired behavior. Newly assembled information from fifty years of research into human behavior in both high and low stress provides a more accurate definition of appropriate operator response, and supports training methods which will produce the needed control room behavior. The research indicates that operator response in emergencies is divided into two modes, conditioned behavior and knowledge based behavior. Methods which assure accurate conditioned behavior, and provide for the recovery of knowledge based behavior, are described in detail

Paired immunoglobulin-like receptor B knockout does not enhance axonal regeneration or locomotor recovery after spinal cord injury.

Science.gov (United States)

Nakamura, Yuka; Fujita, Yuki; Ueno, Masaki; Takai, Toshiyuki; Yamashita, Toshihide

2011-01-21

Myelin components that inhibit axonal regeneration are believed to contribute significantly to the lack of axonal regeneration noted in the adult central nervous system. Three proteins found in myelin, Nogo, myelin-associated glycoprotein, and oligodendrocyte-myelin glycoprotein, inhibit neurite outgrowth in vitro. All of these proteins interact with the same receptors, namely, the Nogo receptor (NgR) and paired immunoglobulin-like receptor B (PIR-B). As per previous reports, corticospinal tract (CST) regeneration is not enhanced in NgR-knock-out mice after spinal cord injury. Therefore, we assessed CST regeneration in PIR-B-knock-out mice. We found that hindlimb motor function, as assessed using the Basso mouse scale, footprint test, inclined plane test, and beam walking test, did not differ between the PIR-B-knock-out and wild-type mice after dorsal hemisection of the spinal cord. Further, tracing of the CST fibers after injury did not reveal enhanced axonal regeneration or sprouting in the CST of the PIR-B-knock-out mice. Systemic administration of NEP1-40, a NgR antagonist, to PIR-B knock-out mice did not enhance the regenerative response. These results indicate that PIR-B knock-out is not sufficient to induce extensive axonal regeneration after spinal cord injury.

Investigation on the Metabolic Regulation of pgi gene knockout Escherichia coli by Enzyme Activities and Intracellular Metabolite Concentrations

Directory of Open Access Journals (Sweden)

Nor ‘Aini, A. R.

2006-01-01

Full Text Available An integrated analysis of the cell growth characteristics, enzyme activities, intracellular metabolite concentrations was made to investigate the metabolic regulation of pgi gene knockout Escherichia coli based on batch culture and continuous culture which was performed at the dilution rate of 0.2h-1. The enzymatic study identified that pathways of pentose phosphate, ED pathway and glyoxylate shunt were all active in pgi mutant. The glycolysis enzymes i.e glyceraldehyde-3-phosphate dehydrogenase, fructose diphosphatase, pyruvate kinase, triose phosphate isomerase were down regulated implying that the inactivation of pgi gene reduced the carbon flux through glycolytic pathway. Meanwhile, the pentose phosphate pathway was active as a major route for intermediary carbohydrate metabolism instead of glycolysis. The pentose phosphate pathway generates most of the major reducing co-factor NADPH as shown by the increased of NADPH/NADP+ ratio in the mutant when compared with the parent strain. The fermentative enzymes such as acetate kinase and lactate dehydrogenase were down regulated in the mutant. Knockout of pgi gene results in the significant increase in the intracellular concentration of glucose-6-phosphate and decrease in the concentration of oxaloacetate. The slow growth rate of the mutant was assumed to be affected by the accumulation of glucose-6-phosphate and imbalance of NADPH reoxidation.

Knockout of Tmem70 alters biogenesis of ATP synthase and leads to embryonal lethality in mice

Czech Academy of Sciences Publication Activity Database

Vrbacký, Marek; Kovalčíková, Jana; Chawengsaksophak, Kallayanee; Beck, Inken; Mráček, Tomáš; Nůsková, Hana; Sedmera, David; Papoušek, František; Kolář, František; Sobol, Margaryta; Hozák, Pavel; Sedláček, Radislav; Houštěk, Josef

2016-01-01

Roč. 25, č. 21 (2016), s. 4674-4685 ISSN 0964-6906 R&D Projects: GA ČR(CZ) GB14-36804G; GA MŠk(CZ) LL1204; GA MŠk(CZ) ED1.1.00/02.0109; GA TA ČR(CZ) TE01020118; GA MŠk(CZ) LM2015062; GA MZd(CZ) NV16-33018A; GA MŠk(CZ) LM2015040 Institutional support: RVO:67985823 ; RVO:68378050 Keywords : mouse knockout * mitochondria * ATP synthase * TMEM70 * biogenesis * mitochondrial diseases Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.340, year: 2016

Increased radiosensitivity and radiation-induced apoptosis in SRC-3 knockout mice

International Nuclear Information System (INIS)

Jin Jie; Wang Yu; Xu Yang; Chen Shilei; Wang Junping; Ran Xinze; Su Yongping; Wang Jin

2014-01-01

Steroid receptor coactivator-3 (SRC-3), a multifunctional transcriptional coactivator, plays an important role in regulation of cell apoptosis in chemoresistant cancer cells. However, its role in radiation-induced apoptosis in hematopoietic cells is still unclear. In this study, we used SRC-3 knockout (SRC-3 -/- ) mice to assess the role of SRC-3 in radiation-induced hematopoietic injury in vivo. After a range of doses of irradiation, SRC-3 -/- mice exhibited lower counts of peripheral blood cells and bone marrow (BM) mononuclear cells and excessive BM depression, which resulted in a significantly higher mortality compared with wildtype mice. Moreover, BM mononuclear cells obtained from SRC-3 -/- mice showed a remarkable increase in radiation-induced apoptosis. Collectively, our data demonstrate that SRC-3 plays a role in radiation-induced apoptosis of BM hematopoietic cells. Regulation of SRC-3 might influence the radiosensitivity of hematopoietic cells, which highlights a potential therapeutic target for radiation-induced hematopoietic injury. (author)

Metabonomic study of the biochemical profiles of heterozygous myostatin knockout swine

Directory of Open Access Journals (Sweden)

Jianxiang XU,Dengke PAN,Jie ZHAO,Jianwu WANG,Xiaohong HE,Yuehui MA,Ning LI

2015-03-01

Full Text Available Myostatin is a transforming growth factor-β family member that normally acts to limit skeletal muscle growth. Myostatin gene (MSTN knockout (KO mice show possible effects for the prevention or treatment of metabolic disorders such as obesity and type 2 diabetes. We applied chromatography and mass spectrometry based metabonomics to assess system-wide metabolic response of heterozygous MSTN KO (MSTN+/- swine. Most of the metabolic data for MSTN+/- swine were similar to the data for wild type (WT control swine. There were, however, metabolic changes related to fatty acid metabolism, glucose utilization, lipid metabolism, as well as BCAA catabolism caused by monoallelic MSTN depletion.The statistical analyses suggested that: (1 most metabolic changes were not significant in MSTN+/- swine compared to WT swine; (2 only a few metabolic properties were significantly different between KO and WT swine, especially for lipid metabolism. Significantly, these minor changes were most evident in female KO swine and suggested differences in gender sensitivity to myostatin.

Increased amphetamine-induced locomotor activity, sensitization, and accumbal dopamine release in M5 muscarinic receptor knockout mice

DEFF Research Database (Denmark)

Schmidt, Lene S; Miller, Anthony D; Lester, Deranda B

2010-01-01

showed that M(5) receptor knockout (M (5) (-/-) ) mice are less sensitive to the reinforcing properties of addictive drugs. MATERIALS AND METHODS: Here, we investigate the role of M(5) receptors in the effects of amphetamine and cocaine on locomotor activity, locomotor sensitization, and dopamine release......-induced hyperactivity and dopamine release as well as amphetamine sensitization are enhanced in mice lacking the M(5) receptor. These results support the concept that the M(5) receptor modulates effects of addictive drugs....

BSN723T Prevents Atherosclerosis and Weight Gain in ApoE Knockout Mice Fed a Western Diet

OpenAIRE

Williams, Jarrod; Ensor, Charles; Gardner, Scott; Smith, Rebecca; Lodder, Robert

2015-01-01

Objective This study tests the hypothesis that BSN723T can prevent the development of hyperlipidemia and atherosclerosis in ApoE-/- knockout mice fed a Western (high fat, high cholesterol, and high sucrose) diet. BSN723T is a combination drug therapy consisting of D-tagatose and dihydromyricetin (BSN723). Background D-tagatose has an antihyperglycemic effect in animal and human studies and shows promise as a treatment for type 2 diabetes and obesity. Many claims regarding BSN723's pharmacolog...

Comparison of the Tastes of L-Alanine and Monosodium Glutamate in C57BL/6J Wild Type and T1r3 Knockout Mice.

Science.gov (United States)

Eddy, Meghan C; Eschle, Benjamin K; Delay, Eugene R

2017-09-01

Previous research showed that L-alanine and monosodium L-glutamate elicit similar taste sensations in rats. This study reports the results of behavioral experiments designed to compare the taste capacity of C57BL/6J wild type and T1r3- mice for these 2 amino acids. In conditioned taste aversion (CTA) experiments, wild-type mice exhibited greater sensitivity than knockout mice for both L-amino acids, although knockout mice were clearly able to detect both amino acids at 50 mM and higher concentrations. Generalization of CTA between L-alanine and L-glutamate was bidirectionally equivalent for both mouse genotypes, indicating that both substances elicited similar tastes in both genotypes. This was verified by the discrimination experiments in which both mouse genotypes performed at or near chance levels at 75 and 150 mM. Above 150 mM, discrimination performance improved, suggesting the taste qualities of the 2 L-amino acids are not identical. No differences between knockout and wild-type mice in discrimination ability were detected. These results indicate that while the T1r3 receptor is important for tasting L-alanine and L-glutamate, other receptors are also important for tasting these amino acids. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Morphological and genetic characterization of group I Clostridium botulinum type B strain 111 and the transcriptional regulator spoIIID gene knockout mutant in sporulation.

Science.gov (United States)

Hosomi, Koji; Kuwana, Ritsuko; Takamatsu, Hiromu; Kohda, Tomoko; Kozaki, Shunji; Mukamoto, Masafumi

2015-06-01

Clostridium botulinum is a heat-resistant spore-forming bacterium that causes the serious paralytic illness botulism. Heat-resistant spores may cause food sanitation hazards and sporulation plays a central role in the survival of C. botulinum. We observed morphological changes and investigated the role of the transcriptional regulator SpoIIID in the sporulation of C. botulinum type B strain 111 in order to elucidate the molecular mechanism in C. botulinum. C. botulinum type B formed heat-resistant spores through successive morphological changes corresponding to those of Bacillus subtilis, a spore-forming model organism. An analysis of the spoIIID gene knockout mutant revealed that the transcriptional regulator SpoIIID contributed to heat-resistant spore formation by C. botulinum type B and activated the transcription of the sigK gene later during sporulation. Transcription of the spoIIID gene, which differed from that in B. subtilis and Clostridium difficile, was observed in the sigE gene knockout mutant of C. botulinum type B. An analysis of the sigF gene knockout mutant showed that the sporulation-specific sigma factor SigF was essential for transcription of the spoIIID gene in C. botulinum type B. These results suggest that the regulation of sporulation in C. botulinum is not similar to that in B. subtilis and other clostridia. Copyright © 2015 Elsevier Ltd. All rights reserved.

Spectrally accurate contour dynamics

International Nuclear Information System (INIS)

Van Buskirk, R.D.; Marcus, P.S.

1994-01-01

We present an exponentially accurate boundary integral method for calculation the equilibria and dynamics of piece-wise constant distributions of potential vorticity. The method represents contours of potential vorticity as a spectral sum and solves the Biot-Savart equation for the velocity by spectrally evaluating a desingularized contour integral. We use the technique in both an initial-value code and a newton continuation method. Our methods are tested by comparing the numerical solutions with known analytic results, and it is shown that for the same amount of computational work our spectral methods are more accurate than other contour dynamics methods currently in use

STAT2 Knockout Syrian Hamsters Support Enhanced Replication and Pathogenicity of Human Adenovirus, Revealing an Important Role of Type I Interferon Response in Viral Control.

Directory of Open Access Journals (Sweden)

Karoly Toth

2015-08-01

Full Text Available Human adenoviruses have been studied extensively in cell culture and have been a model for studies in molecular, cellular, and medical biology. However, much less is known about adenovirus replication and pathogenesis in vivo in a permissive host because of the lack of an adequate animal model. Presently, the most frequently used permissive immunocompetent animal model for human adenovirus infection is the Syrian hamster. Species C human adenoviruses replicate in these animals and cause pathology that is similar to that seen with humans. Here, we report findings with a new Syrian hamster strain in which the STAT2 gene was functionally knocked out by site-specific gene targeting. Adenovirus-infected STAT2 knockout hamsters demonstrated an accentuated pathology compared to the wild-type control animals, and the virus load in the organs of STAT2 knockout animals was 100- to 1000-fold higher than that in wild-type hamsters. Notably, the adaptive immune response to adenovirus is not adversely affected in STAT2 knockout hamsters, and surviving hamsters cleared the infection by 7 to 10 days post challenge. We show that the Type I interferon pathway is disrupted in these hamsters, revealing the critical role of interferon-stimulated genes in controlling adenovirus infection. This is the first study to report findings with a genetically modified Syrian hamster infected with a virus. Further, this is the first study to show that the Type I interferon pathway plays a role in inhibiting human adenovirus replication in a permissive animal model. Besides providing an insight into adenovirus infection in humans, our results are also interesting from the perspective of the animal model: STAT2 knockout Syrian hamster may also be an important animal model for studying other viral infections, including Ebola-, hanta-, and dengue viruses, where Type I interferon-mediated innate immunity prevents wild type hamsters from being effectively infected to be used as

Common arterial trunk and ventricular non-compaction in Lrp2 knockout mice indicate a crucial role of LRP2 in cardiac development

NARCIS (Netherlands)

T. Baardman (Taco); M.V. Zwier (Mathijs V.); L.J. Wisse (Lambertus); A.C. Gittenberger-De Groot (Adriana); W.S. Kerstjens-Frederikse (Wilhelmina); R.M.W. Hofstra (Robert); A. Jurdzinski (Angelika); B.P. Hierck (Beerend); M.R.M. Jongbloed (Monique); R.M.F. Berger (Rolf); T. Plösch (Torsten); M.C. DeRuiter (Marco)

2016-01-01

textabstractLipoprotein-related receptor protein 2 (LRP2) is important for development of the embryonic neural crest and brain in both mice and humans. Although a role in cardiovascular development can be expected, the hearts of Lrp2 knockout (KO) mice have not yet been investigated. We studied the

Common arterial trunk and ventricular non-compaction in Lrp2 knockout mice indicate a crucial role of LRP2 in cardiac development

NARCIS (Netherlands)

Baardman, Maria E.; Zwier, Mathijs V.; Wisse, Lambertus J.; Gittenberger-de Groot, Adriana C.; Kerstjens-Frederikse, Wilhelmina S.; Hofstra, Robert M. W.; Jurdzinski, Angelika; Hierck, Beerend P.; Jongbloed, Monique R. M.; Berger, Rolf M. F.; Plosch, Torsten; DeRuiter, Marco C.

2016-01-01

Lipoprotein-related receptor protein 2 (LRP2) is important for development of the embryonic neural crest and brain in both mice and humans. Although a role in cardiovascular development can be expected, the hearts of Lrp2 knockout (KO) mice have not yet been investigated. We studied the

Bone growth and turnover in progesterone receptor knockout mice.

Energy Technology Data Exchange (ETDEWEB)

Rickard, David J.; Iwaniec, Urszula T.; Evans, Glenda; Hefferan, Theresa E.; Hunter, Jaime C.; Waters, Katrina M.; Lydon, John P.; O' Malley, Bert W.; Khosla, Sundeep; Spelsberg, Thomas C.; Turner, Russell T.

2008-05-01

The role of progesterone receptor (PR) signaling in skeletal metabolism is controversial. To address whether signaling through the PR is necessary for normal bone growth and turnover, we performed histomorphometric and mCT analyses of bone from homozygous female PR knockout (PRKO) mice at 6, 12, and 26 weeks of age. These mice possess a null mutation of the PR locus, which blocks the gene expression of A and B isoforms of PR. Body weight gain, uterine weight gain and tibia longitudinal bone growth was normal in PRKO mice. In contrast, total and cortical bone mass were increased in long bones of post-pubertal (12 and 26-week-old) PRKO mice, whereas cancellous bone mass was normal in the tibia but increased in the humerus. The striking 57% decrease in cancellous bone from the proximal tibia metaphysis which occurred between 6 and 26 weeks in WT mice was abolished in PRKO mice. The improved bone balance in aging PRKO mice was associated with elevated bone formation and a tendency toward reduced osteoclast perimeter. Taken together, these findings suggest that PR signaling in mice attenuates the accumulation of cortical bone mass during adolescence and is required for early age-related loss of cancellous bone.

Analyzing AbrB-Knockout Effects through Genome and Transcriptome Sequencing of Bacillus licheniformis DW2

Science.gov (United States)

Shu, Cheng-Cheng; Wang, Dong; Guo, Jing; Song, Jia-Ming; Chen, Shou-Wen; Chen, Ling-Ling; Gao, Jun-Xiang

2018-01-01

As an industrial bacterium, Bacillus licheniformis DW2 produces bacitracin which is an important antibiotic for many pathogenic microorganisms. Our previous study showed AbrB-knockout could significantly increase the production of bacitracin. Accordingly, it was meaningful to understand its genome features, expression differences between wild and AbrB-knockout (ΔAbrB) strains, and the regulation of bacitracin biosynthesis. Here, we sequenced, de novo assembled and annotated its genome, and also sequenced the transcriptomes in three growth phases. The genome of DW2 contained a DNA molecule of 4,468,952 bp with 45.93% GC content and 4,717 protein coding genes. The transcriptome reads were mapped to the assembled genome, and obtained 4,102∼4,536 expressed genes from different samples. We investigated transcription changes in B. licheniformis DW2 and showed that ΔAbrB caused hundreds of genes up-regulation and down-regulation in different growth phases. We identified a complete bacitracin synthetase gene cluster, including the location and length of bacABC, bcrABC, and bacT, as well as their arrangement. The gene cluster bcrABC were significantly up-regulated in ΔAbrB strain, which supported the hypothesis in previous study of bcrABC transporting bacitracin out of the cell to avoid self-intoxication, and was consistent with the previous experimental result that ΔAbrB could yield more bacitracin. This study provided a high quality reference genome for B. licheniformis DW2, and the transcriptome data depicted global alterations across two strains and three phases offered an understanding of AbrB regulation and bacitracin biosynthesis through gene expression. PMID:29599755

Zadoff-Chu coded ultrasonic signal for accurate range estimation

KAUST Repository

AlSharif, Mohammed H.

2017-11-02

This paper presents a new adaptation of Zadoff-Chu sequences for the purpose of range estimation and movement tracking. The proposed method uses Zadoff-Chu sequences utilizing a wideband ultrasonic signal to estimate the range between two devices with very high accuracy and high update rate. This range estimation method is based on time of flight (TOF) estimation using cyclic cross correlation. The system was experimentally evaluated under different noise levels and multi-user interference scenarios. For a single user, the results show less than 7 mm error for 90% of range estimates in a typical indoor environment. Under the interference from three other users, the 90% error was less than 25 mm. The system provides high estimation update rate allowing accurate tracking of objects moving with high speed.

Zadoff-Chu coded ultrasonic signal for accurate range estimation

KAUST Repository

AlSharif, Mohammed H.; Saad, Mohamed; Siala, Mohamed; Ballal, Tarig; Boujemaa, Hatem; Al-Naffouri, Tareq Y.

2017-01-01

This paper presents a new adaptation of Zadoff-Chu sequences for the purpose of range estimation and movement tracking. The proposed method uses Zadoff-Chu sequences utilizing a wideband ultrasonic signal to estimate the range between two devices with very high accuracy and high update rate. This range estimation method is based on time of flight (TOF) estimation using cyclic cross correlation. The system was experimentally evaluated under different noise levels and multi-user interference scenarios. For a single user, the results show less than 7 mm error for 90% of range estimates in a typical indoor environment. Under the interference from three other users, the 90% error was less than 25 mm. The system provides high estimation update rate allowing accurate tracking of objects moving with high speed.

Germinated Brown Rice Attenuates Atherosclerosis and Vascular Inflammation in Low-Density Lipoprotein Receptor-Knockout Mice.

Science.gov (United States)

Zhao, Ruozhi; Ghazzawi, Nora; Wu, Jiansu; Le, Khuong; Li, Chunyang; Moghadasian, Mohammed H; Siow, Yaw L; Apea-Bah, Franklin B; Beta, Trust; Yin, Zhengfeng; Shen, Garry X

2018-05-02

The present study investigates the impact of germinated brown rice (GBR) on atherosclerosis and the underlying mechanism in low-density lipoprotein receptor-knockout (LDLr-KO) mice. The intensity of atherosclerosis in aortas of LDLr-KO mice receiving diet supplemented with 60% GBR (weight/weight) was significantly less than that in mice fed with 60% white rice (WR) or control diet ( p mice fed with WR diet was significantly more than that from mice receiving the control diet ( p mice in comparison to the WR diet ( p mice compared to WR. The anti-atherosclerotic effect of GBR in LDLr-KO mice at least in part results from its anti-inflammatory activity.

Knockout of a P-glycoprotein gene increases susceptibility to abamectin and emamectin benzoate in Spodoptera exigua.

Science.gov (United States)

Zuo, Y-Y; Huang, J-L; Wang, J; Feng, Y; Han, T-T; Wu, Y-D; Yang, Y-H

2018-02-01

P-glycoprotein [P-gp or the ATP-binding cassette transporter B1 (ABCB1)] is an important participant in multidrug resistance of cancer cells, yet the precise function of this arthropod transporter is unknown. The aim of this study was to determine the importance of P-gp for susceptibility to insecticides in the beet armyworm (Spodoptera exigua) using clustered regularly interspaced short palindromic repeats/CRISPR-associated 9 (CRISPR/Cas9) gene-editing technology. We cloned an open reading frame (ORF) encoding the S. exigua P-gp protein (SeP-gp) predicted to display structural characteristics common to P-gp and other insect ABCB1 transporters. A knockout line with a frame shift deletion of four nucleotides in the SeP-gp ORF was established using the CRISPR/Cas9 gene-editing system to test its potential role in determining susceptibility to chemical insecticides or insecticidal proteins from the bacterium Bacillus thuringiensis (Bt). Results from comparative bioassays demonstrate that knockout of SeP-gp significantly increases susceptibility of S. exigua by around threefold to abamectin and emamectin benzoate (EB), but not to spinosad, chlorfenapyr, beta-cypermethrin, carbosulfan indoxacarb, chlorpyrifos, phoxim, diafenthiuron, chlorfluazuron, chlorantraniliprole or two Bt toxins (Cry1Ca and Cry1Fa). Our data support an important role for SeP-gp in susceptibility of S. exigua to abamectin and EB and imply that overexpression of SeP-gp may contribute to abamectin and EB resistance in S. exigua. © 2017 The Royal Entomological Society.

Management Of Hanford KW Basin Knockout Pot Sludge As Spent Nuclear Fuel

International Nuclear Information System (INIS)

Raymond, R. E.; Evans, K. M.

2012-01-01

CH2M Hill Plateau Remediation Company (CHPRC) and AREVA Federal Services, LLC (AFS) have been working collaboratively to develop and deploy technologies to remove, transport, and interim store remote-handled sludge from the 10S-K West Reactor Fuel Storage Basin on the U.S. Department of Energy (DOE) Hanford Site near Richland, WA, USA. Two disposal paths exist for the different types of sludge found in the K West (KW) Basin. One path is to be managed as Spent Nuclear Fuel (SNF) with eventual disposal at an SNF at a yet to be licensed repository. The second path will be disposed as remote-handled transuranic (RH-TRU) waste at the Waste Isolation Pilot Plant (WIPP) in Carlsbad, NM. This paper describes the systems developed and executed by the Knockout Pot (KOP) Disposition Subproject for processing and interim storage of the sludge managed as SNF, (i.e., KOP material)

Simple Meets Single: The Application of CRISPR/Cas9 in Haploid Embryonic Stem Cells

Directory of Open Access Journals (Sweden)

Zixi Yin

2017-01-01

Full Text Available The CRISPR/Cas9 system provides a powerful method for the genetic manipulation of the mammalian genome, allowing knockout of individual genes as well as the generation of genome-wide knockout cell libraries for genetic screening. However, the diploid status of most mammalian cells restricts the application of CRISPR/Cas9 in genetic screening. Mammalian haploid embryonic stem cells (haESCs have only one set of chromosomes per cell, avoiding the issue of heterozygous recessive mutations in diploid cells. Thus, the combination of haESCs and CRISPR/Cas9 facilitates the generation of genome-wide knockout cell libraries for genetic screening. Here, we review recent progress in CRISPR/Cas9 and haPSCs and discuss their applications in genetic screening.

Accurate and robust brain image alignment using boundary-based registration.

Science.gov (United States)

Greve, Douglas N; Fischl, Bruce

2009-10-15

The fine spatial scales of the structures in the human brain represent an enormous challenge to the successful integration of information from different images for both within- and between-subject analysis. While many algorithms to register image pairs from the same subject exist, visual inspection shows that their accuracy and robustness to be suspect, particularly when there are strong intensity gradients and/or only part of the brain is imaged. This paper introduces a new algorithm called Boundary-Based Registration, or BBR. The novelty of BBR is that it treats the two images very differently. The reference image must be of sufficient resolution and quality to extract surfaces that separate tissue types. The input image is then aligned to the reference by maximizing the intensity gradient across tissue boundaries. Several lower quality images can be aligned through their alignment with the reference. Visual inspection and fMRI results show that BBR is more accurate than correlation ratio or normalized mutual information and is considerably more robust to even strong intensity inhomogeneities. BBR also excels at aligning partial-brain images to whole-brain images, a domain in which existing registration algorithms frequently fail. Even in the limit of registering a single slice, we show the BBR results to be robust and accurate.

Accurate measurement of junctional conductance between electrically coupled cells with dual whole-cell voltage-clamp under conditions of high series resistance.

Science.gov (United States)

Hartveit, Espen; Veruki, Margaret Lin

2010-03-15

Accurate measurement of the junctional conductance (G(j)) between electrically coupled cells can provide important information about the functional properties of coupling. With the development of tight-seal, whole-cell recording, it became possible to use dual, single-electrode voltage-clamp recording from pairs of small cells to measure G(j). Experiments that require reduced perturbation of the intracellular environment can be performed with high-resistance pipettes or the perforated-patch technique, but an accompanying increase in series resistance (R(s)) compromises voltage-clamp control and reduces the accuracy of G(j) measurements. Here, we present a detailed analysis of methodologies available for accurate determination of steady-state G(j) and related parameters under conditions of high R(s), using continuous or discontinuous single-electrode voltage-clamp (CSEVC or DSEVC) amplifiers to quantify the parameters of different equivalent electrical circuit model cells. Both types of amplifiers can provide accurate measurements of G(j), with errors less than 5% for a wide range of R(s) and G(j) values. However, CSEVC amplifiers need to be combined with R(s)-compensation or mathematical correction for the effects of nonzero R(s) and finite membrane resistance (R(m)). R(s)-compensation is difficult for higher values of R(s) and leads to instability that can damage the recorded cells. Mathematical correction for R(s) and R(m) yields highly accurate results, but depends on accurate estimates of R(s) throughout an experiment. DSEVC amplifiers display very accurate measurements over a larger range of R(s) values than CSEVC amplifiers and have the advantage that knowledge of R(s) is unnecessary, suggesting that they are preferable for long-duration experiments and/or recordings with high R(s). Copyright (c) 2009 Elsevier B.V. All rights reserved.

Accurate Sample Time Reconstruction of Inertial FIFO Data

Directory of Open Access Journals (Sweden)

Sebastian Stieber

2017-12-01

Full Text Available In the context of modern cyber-physical systems, the accuracy of underlying sensor data plays an increasingly important role in sensor data fusion and feature extraction. The raw events of multiple sensors have to be aligned in time to enable high quality sensor fusion results. However, the growing number of simultaneously connected sensor devices make the energy saving data acquisition and processing more and more difficult. Hence, most of the modern sensors offer a first-in-first-out (FIFO interface to store multiple data samples and to relax timing constraints, when handling multiple sensor devices. However, using the FIFO interface increases the negative influence of individual clock drifts—introduced by fabrication inaccuracies, temperature changes and wear-out effects—onto the sampling data reconstruction. Furthermore, additional timing offset errors due to communication and software latencies increases with a growing number of sensor devices. In this article, we present an approach for an accurate sample time reconstruction independent of the actual clock drift with the help of an internal sensor timer. Such timers are already available in modern sensors, manufactured in micro-electromechanical systems (MEMS technology. The presented approach focuses on calculating accurate time stamps using the sensor FIFO interface in a forward-only processing manner as a robust and energy saving solution. The proposed algorithm is able to lower the overall standard deviation of reconstructed sampling periods below 40 μ s, while run-time savings of up to 42% are achieved, compared to single sample acquisition.

Accurate Evaluation of Quantum Integrals

Science.gov (United States)

Galant, D. C.; Goorvitch, D.; Witteborn, Fred C. (Technical Monitor)

1995-01-01

Combining an appropriate finite difference method with Richardson's extrapolation results in a simple, highly accurate numerical method for solving a Schrodinger's equation. Important results are that error estimates are provided, and that one can extrapolate expectation values rather than the wavefunctions to obtain highly accurate expectation values. We discuss the eigenvalues, the error growth in repeated Richardson's extrapolation, and show that the expectation values calculated on a crude mesh can be extrapolated to obtain expectation values of high accuracy.

MicroRNA-155 knockout mice are susceptible to Mycobacterium tuberculosis infection.

Science.gov (United States)

Iwai, Hiroki; Funatogawa, Keiji; Matsumura, Kazunori; Kato-Miyazawa, Masako; Kirikae, Fumiko; Kiga, Kotaro; Sasakawa, Chihiro; Miyoshi-Akiyama, Tohru; Kirikae, Teruo

2015-05-01

MicroRNAs (miRNAs) are short, conserved, non-coding RNA molecules that repress translation, followed by the decay of miRNA-targeted mRNAs that encode molecules involved in cell differentiation, development, immunity and apoptosis. At least six miRNAs, including microRNA-155 (miR-155), were up-regulated when born marrow-derived macrophages from C57BL/6 mice were infected with Mycobacterium tuberculosis Erdman. C57BL/6 mice intravenously infected with Erdman showed up-regulation of miR-155 in livers and lungs. Following infection, miR-155-deficient C57BL/6 mice died significantly earlier and had significantly higher numbers of CFU in lungs than wild-type mice. Moreover, fewer CD4(+) T cells, but higher numbers of monocytes and neutrophils, were present in the lungs of Erdman-infected miR-155 knockout (miR-155(-/-)) than of wild-type mice. These findings indicated that miR-155 plays a critical role in immune responses to M. tuberculosis. Copyright © 2015 Elsevier Ltd. All rights reserved.

Single ionization of Ne, Ar and Kr by proton impact: Single differential distributions in energy and angle

Energy Technology Data Exchange (ETDEWEB)

Otranto, S [CONICET and Dto. de Fisica, Universidad Nacional del Sur, 8000 BahIa Blanca (Argentina); Miraglia, J E [Instituto de AstronomIa y Fisica del Espacio, CONICET and Universidad de Buenos Aires C1428EGA (Argentina); Olson, R E, E-mail: sotranto@uns.edu.a [Physics Department, Missouri University of Science and Technology, Rolla MO 65409 (United States)

2009-11-01

In this work we present a theoretical study of singly differential cross sections in energy and angle for the single ionization of neon, argon and krypton by proton impact. Theoretical results obtained by means of the Continuum Distorted Wave-Eikonal initial state (CDW-EIS) model are compared to those provided by the First Born Approximation (FBA) and the classical trajectory Monte Carlo (CTMC) method as well as to experimental data from several laboratories. We note in particular for argon, that the CDW-EIS model does not reproduce the experimental data as accurately as expected, while the CTMC instead is in very good agreement.

Highly accurate potential calculations for cylindrically symmetric geometries using multi-region FDM: A review

Energy Technology Data Exchange (ETDEWEB)

Edwards, David, E-mail: dej@kingcon.com [IJL Research Center, Newark, VT 05871 (United States)

2011-07-21

This paper is a review of multi-region FDM, a numerical technique for accurately determining electrostatic potentials in cylindrically symmetric geometries. Multi-region FDM can be thought of as the union of various individual elements: a single region FDM process: a method for algorithmic development; a method for auto creating a multi-region structure; the process for the relaxation of multi-region structures. Each element will be briefly described along with its integration into the multi-region relaxation process itself.

Remodeling of repolarization and arrhythmia susceptibility in a myosin-binding protein C knockout mouse model.

Science.gov (United States)

Toib, Amir; Zhang, Chen; Borghetti, Giulia; Zhang, Xiaoxiao; Wallner, Markus; Yang, Yijun; Troupes, Constantine D; Kubo, Hajime; Sharp, Thomas E; Feldsott, Eric; Berretta, Remus M; Zalavadia, Neil; Trappanese, Danielle M; Harper, Shavonn; Gross, Polina; Chen, Xiongwen; Mohsin, Sadia; Houser, Steven R

2017-09-01

Hypertrophic cardiomyopathy (HCM) is one of the most common genetic cardiac diseases and among the leading causes of sudden cardiac death (SCD) in the young. The cellular mechanisms leading to SCD in HCM are not well known. Prolongation of the action potential (AP) duration (APD) is a common feature predisposing hypertrophied hearts to SCD. Previous studies have explored the roles of inward Na + and Ca 2+ in the development of HCM, but the role of repolarizing K + currents has not been defined. The objective of this study was to characterize the arrhythmogenic phenotype and cellular electrophysiological properties of mice with HCM, induced by myosin-binding protein C (MyBPC) knockout (KO), and to test the hypothesis that remodeling of repolarizing K + currents causes APD prolongation in MyBPC KO myocytes. We demonstrated that MyBPC KO mice developed severe hypertrophy and cardiac dysfunction compared with wild-type (WT) control mice. Telemetric electrocardiographic recordings of awake mice revealed prolongation of the corrected QT interval in the KO compared with WT control mice, with overt ventricular arrhythmias. Whole cell current- and voltage-clamp experiments comparing KO with WT mice demonstrated ventricular myocyte hypertrophy, AP prolongation, and decreased repolarizing K + currents. Quantitative RT-PCR analysis revealed decreased mRNA levels of several key K + channel subunits. In conclusion, decrease in repolarizing K + currents in MyBPC KO ventricular myocytes contributes to AP and corrected QT interval prolongation and could account for the arrhythmia susceptibility. NEW & NOTEWORTHY Ventricular myocytes isolated from the myosin-binding protein C knockout hypertrophic cardiomyopathy mouse model demonstrate decreased repolarizing K + currents and action potential and QT interval prolongation, linking cellular repolarization abnormalities with arrhythmia susceptibility and the risk for sudden cardiac death in hypertrophic cardiomyopathy. Copyright © 2017

Conditional RARα Knockout Mice Reveal Acute Requirement for Retinoic Acid and RARα in Homeostatic Plasticity

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Federica eSarti

2012-02-01

Full Text Available All-trans retinoic acid (RA plays important roles in brain development through regulating gene transcription. Recently, a novel postdevelopmental role of RA in mature brain was proposed. Specifically, RA rapidly enhanced excitatory synaptic transmission independent of transcriptional regulation. RA synthesis was induced when excitatory synaptic transmission was chronically blocked, and RA then activated dendritic protein synthesis and synaptic insertion of homomeric GluA1 AMPA receptors, thereby compensating for the loss of neuronal activity in a homeostatic fashion. This action of RA was suggested to be mediated by its canonical receptor RARα but no genetic evidence was available. Thus, we here tested the fundamental requirement of RARα in homeostatic plasticity using conditional RARα knockout mice, and additionally performed a structure-function analysis of RARα. We show that acutely deleting RARα in neurons eliminated RA’s effect on excitatory synaptic transmission, and inhibited activity blockade-induced homeostatic synaptic plasticity. By expressing various RARα rescue constructs in RARα knockout neurons, we found that the DNA-binding domain of RARα was dispensable for its role in regulating synaptic strength, further supporting the notion that RA and RARα act in a non-transcriptional manner in this context. By contrast, the ligand-binding domain (LBD and the mRNA-binding domain (F-domain are both necessary and sufficient for the function of RARα in homeostatic plasticity. Furthermore, we found that homeostatic regulation performed by the LBD/F domains leads to insertion of calcium-permeable AMPA receptors. Our results confirm with unequivocal genetic approaches that RA and RARα perform essential non-transcriptional functions in regulating synaptic strength, and establish a functional link between the various domains of RARα and their involvement in regulating protein synthesis and excitatory synaptic transmission during

Approaching system equilibrium with accurate or not accurate feedback information in a two-route system

Science.gov (United States)

Zhao, Xiao-mei; Xie, Dong-fan; Li, Qi

2015-02-01

With the development of intelligent transport system, advanced information feedback strategies have been developed to reduce traffic congestion and enhance the capacity. However, previous strategies provide accurate information to travelers and our simulation results show that accurate information brings negative effects, especially in delay case. Because travelers prefer to the best condition route with accurate information, and delayed information cannot reflect current traffic condition but past. Then travelers make wrong routing decisions, causing the decrease of the capacity and the increase of oscillations and the system deviating from the equilibrium. To avoid the negative effect, bounded rationality is taken into account by introducing a boundedly rational threshold BR. When difference between two routes is less than the BR, routes have equal probability to be chosen. The bounded rationality is helpful to improve the efficiency in terms of capacity, oscillation and the gap deviating from the system equilibrium.

Impairment in extinction of cued fear memory in syntenin-1 knockout mice.

Science.gov (United States)

Talukdar, Gourango; Inoue, Ran; Yoshida, Tomoyuki; Mori, Hisashi

2018-03-01

Syntenin-1 is a PDZ domain-containing intracellular scaffold protein involved in exosome production, synapse formation, and synaptic plasticity. We tested whether syntenin-1 can regulate learning and memory through its effects on synaptic plasticity. Specifically, we investigated the role of syntenin-1 in contextual and cued fear conditioning and extinction of conditioned fear using syntenin-1 knockout (KO) mice. Genetic disruption of syntenin-1 had little effect on contextual and cued fear memory. However, syntenin-1 KO mice exhibited selective impairment in cued fear extinction retention. This extinction retention deficit in syntenin-1 KO mice was associated with reduced c-Fos-positive neurons in the basolateral amygdala (BLA) and infralimbic cortex (IL) after extinction training and increased c-Fos-positive neurons in the BLA after an extinction retention test. Our results suggest that syntenin-1 plays an important role in extinction of cued fear memory by modulating neuronal activity in the BLA and IL. Copyright © 2018 Elsevier Inc. All rights reserved.

Zika virus infection of adult and fetal STAT2 knock-out hamsters.

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Siddharthan, Venkatraman; Van Wettere, Arnaud J; Li, Rong; Miao, Jinxin; Wang, Zhongde; Morrey, John D; Julander, Justin G

2017-07-01

Zika virus (ZIKV) infection was investigated in adult and fetal STAT2 knock-out (KO) hamsters. Subcutaneous injection of ZIKV of adults resulted in morbidity, mortality, and infection of the uterus, placenta, brain, spinal cord, and testicles, thus providing an opportunity to evaluate congenital ZIKV infection in a second rodent species besides mice. ZIKV-infected cells with morphologies of Sertoli cells and spermatogonia were observed in the testes, which may have implications for sexual transmission and male sterility. Neonates exposed as fetuses to ZIKV at 8 days post-coitus were not smaller than controls. Nevertheless, infectious virus and ZIKV RNA was detected in some, but not all, placentas and fetal brains of KO hamsters. STAT2 KO hamsters may be useful for addressing sexual transmission, pathogenesis, routes of fetal infection, and neurological disease outcomes, and may also be used in antiviral or vaccine studies to identify intervention strategies. Copyright © 2017. Published by Elsevier Inc.

Modeling single cell antibody excretion on a biosensor

NARCIS (Netherlands)

Stojanovic, Ivan; Baumgartner, W.; van der Velden, T.J.G.; Terstappen, Leonardus Wendelinus Mathias Marie; Schasfoort, Richardus B.M.

2016-01-01

We simulated, using Comsol Multiphysics, the excretion of antibodies by single hybridoma cells and their subsequent binding on a surface plasmon resonance imaging (SPRi) sensor. The purpose was to confirm that SPRi is suitable to accurately quantify antibody (anti-EpCAM) excretion. The model showed

Cancer resistance of SR/CR mice in the genetic knockout backgrounds of leukocyte effector mechanisms: determinations for functional requirements.

Science.gov (United States)

Sanders, Anne M; Stehle, John R; Blanks, Michael J; Riedlinger, Gregory; Kim-Shapiro, Jung W; Monjazeb, Arta M; Adams, Jonathan M; Willingham, Mark C; Cui, Zheng

2010-03-31

Spontaneous Regression/Complete Resistant (SR/CR) mice are a colony of cancer-resistant mice that can detect and rapidly destroy malignant cells with innate cellular immunity, predominately mediated by granulocytes. Our previous studies suggest that several effector mechanisms, such as perforin, granzymes, or complements, may be involved in the killing of cancer cells. However, none of these effector mechanisms is known as critical for granulocytes. Additionally, it is unclear which effector mechanisms are required for the cancer killing activity of specific leukocyte populations and the survival of SR/CR mice against the challenges of lethal cancer cells. We hypothesized that if any of these effector mechanisms was required for the resistance to cancer cells, its functional knockout in SR/CR mice should render them sensitive to cancer challenges. This was tested by cross breeding SR/CR mice into the individual genetic knockout backgrounds of perforin (Prf-/-), superoxide (Cybb-/), or inducible nitric oxide (Nos2-/). SR/CR mice were bred into individual Prf-/-, Cybb-/-, or Nos2-/- genetic backgrounds and then challenged with sarcoma 180 (S180). Their overall survival was compared to controls. The cancer killing efficiency of purified populations of macrophages and neutrophils from these immunodeficient mice was also examined. When these genetically engineered mice were challenged with cancer cells, the knockout backgrounds of Prf-/-, Cybb-/-, or Nos2-/- did not completely abolish the SR/CR cancer resistant phenotype. However, the Nos2-/- background did appear to weaken the resistance. Incidentally, it was also observed that the male mice in these immunocompromised backgrounds tended to be less cancer-resistant than SR/CR controls. Despite the previously known roles of perforin, superoxide or nitric oxide in the effector mechanisms of innate immune responses, these effector mechanisms were not required for cancer-resistance in SR/CR mice. The resistance was

Drop tests of the Three Mile Island knockout canister

International Nuclear Information System (INIS)

Box, W.D.; Aaron, W.S.; Shappert, L.B.; Childress, P.C.; Quinn, G.J.; Smith, J.V.

1986-09-01

A type of Three Mile Island Unit 2 (TMI-2) defueling canister, called a ''knockout'' canister, was subjected to a series of drop tests at the Oak Ridge National Laboratory's Drop Test Facility. These tests were designed to confirm the structural integrity of internal fixed neutron poisons in support of a request for NRC licensing of this type of canister for the shipment of TMI-2 reactor fuel debris to the Idaho National Engineering Laboratory (INEL) for the Core Examination R and D Program. Work conducted at the Oak Ridge National Laboratory included (1) precise physical measurements of the internal poison rod configuration before assembly, (2) canister assembly and welding, (3) nondestructive examination (an initial hydrostatic pressure test and an x-ray profile of the internals before and after each drop test), (4) addition of a simulated fuel load, (5) instrumentation of the canister for each drop test, (6) fabrication of a cask simulation vessel with a developed and tested foam impact limiter, (7) use of refrigeration facilities to cool the canister to well below freezing prior to three of the drops, (8) recording the drop test with still, high-speed, and normal-speed photography, (9) recording the accelerometer measurements during impact, (10) disassembly and post-test examination with precise physical measurements, and (11) preparation of the final report

Role of interferon-gamma in the pathogenesis of LCMV-induced meningitis: unimpaired leucocyte recruitment, but deficient macrophage activation in interferon-gamma knock-out mice

DEFF Research Database (Denmark)

Nansen, A; Christensen, Jan Pravsgaard; Röpke, C

1998-01-01

, a viral peptide could also elicit a T cell mediated inflammatory response in virus-primed IFN-gamma knock-out mice, indicating that redundancy of this cytokine as a proinflammatory mediator is not restricted to inflammatory reactions triggered by an active infection. Thus, T cell mediated inflammation may...

Studies of an Androgen-Binding Protein Knockout Corroborate a Role for Salivary ABP in Mouse Communication.

Science.gov (United States)

Chung, Amanda G; Belone, Phillip M; Bímová, Barbora Vošlajerová; Karn, Robert C; Laukaitis, Christina M

2017-04-01

The house mouse Androgen-binding protein ( Abp ) gene family is comprised of 64 paralogs, 30 Abpa and 34 Abpbg , encoding the alpha (ABPA) and beta-gamma (ABPBG) protein subunits that are disulfide-bridged to form dimers in secretions. Only 14 Abp genes are expressed in distinct patterns in the lacrimal (11) and submandibular glands (3). We created a knockout mouse line lacking two of the three genes expressed in submandibular glands, Abpa27 and Abpbg27 , by replacing them with the neomycin resistance gene. The knockout genotype (-/-) showed no Abpa27 or Abpbg27 transcripts in submandibular gland complementary DNA (cDNA) libraries and there was a concomitant lack of protein expression of ABPA27 and ABPBG27 in the -/- genotype saliva, shown by elimination of these two proteins from the saliva proteome and the loss of cross-reactive material in the acinar cells of the submandibular glands. We also observed a decrease in BG26 protein in the -/- animals, suggesting monomer instability. Overall, we observed no major phenotypic changes in the -/- genotype, compared with their +/+ and +/- siblings raised in a laboratory setting, including normal growth curves, tissue histology, fecundity, and longevity. The only difference is that male and female C57BL/6 mice preferred saliva of the opposite sex containing ABP statistically significantly more than saliva of the opposite sex without ABP in a Y-maze test. These results show for the first time that mice can sense the presence of ABP between saliva targets with and without ABPs, and that they spend more time investigating the target containing ABP. Copyright © 2017 by the Genetics Society of America.

Accurate modeling and evaluation of microstructures in complex materials

Science.gov (United States)

Tahmasebi, Pejman

2018-02-01

Accurate characterization of heterogeneous materials is of great importance for different fields of science and engineering. Such a goal can be achieved through imaging. Acquiring three- or two-dimensional images under different conditions is not, however, always plausible. On the other hand, accurate characterization of complex and multiphase materials requires various digital images (I) under different conditions. An ensemble method is presented that can take one single (or a set of) I(s) and stochastically produce several similar models of the given disordered material. The method is based on a successive calculating of a conditional probability by which the initial stochastic models are produced. Then, a graph formulation is utilized for removing unrealistic structures. A distance transform function for the Is with highly connected microstructure and long-range features is considered which results in a new I that is more informative. Reproduction of the I is also considered through a histogram matching approach in an iterative framework. Such an iterative algorithm avoids reproduction of unrealistic structures. Furthermore, a multiscale approach, based on pyramid representation of the large Is, is presented that can produce materials with millions of pixels in a matter of seconds. Finally, the nonstationary systems—those for which the distribution of data varies spatially—are studied using two different methods. The method is tested on several complex and large examples of microstructures. The produced results are all in excellent agreement with the utilized Is and the similarities are quantified using various correlation functions.

Female preproenkephalin-knockout mice display altered emotional responses

Science.gov (United States)

Ragnauth, A.; Schuller, A.; Morgan, M.; Chan, J.; Ogawa, S.; Pintar, J.; Bodnar, R. J.; Pfaff, D. W.

2001-01-01

The endogenous opioid system has been implicated in sexual behavior, palatable intake, fear, and anxiety. The present study examined whether ovariectomized female transgenic preproenkephalin-knockout (PPEKO) mice and their wild-type and heterozygous controls displayed alterations in fear and anxiety paradigms, sucrose intake, and lordotic behavior. To examine stability of responding, three squads of the genotypes were tested across seasons over a 20-month period. In a fear-conditioning paradigm, PPEKO mice significantly increased freezing to both fear and fear + shock stimuli relative to controls. In the open field, PPEKO mice spent significantly less time and traversed significantly less distance in the center of an open field than wild-type controls. Further, PPEKO mice spent significantly less time and tended to be less active on the light side of a dark–light chamber than controls, indicating that deletion of the enkephalin gene resulted in exaggerated responses to fear or anxiety-provoking environments. These selective deficits were observed consistently across testing squads spanning 20 months and different seasons. In contrast, PPEKO mice failed to differ from corresponding controls in sucrose, chow, or water intake across a range (0.0001–20%) of sucrose concentrations and failed to differ in either lordotic or female approach to male behaviors when primed with estradiol and progesterone, thereby arguing strongly for the selectivity of a fear and anxiety deficit which was not caused by generalized and nonspecific debilitation. These transgenic data strongly suggest that opioids, and particularly enkephalin gene products, are acting naturally to inhibit fear and anxiety. PMID:11172058

Accurate determination of rates from non-uniformly sampled relaxation data

Energy Technology Data Exchange (ETDEWEB)

Stetz, Matthew A.; Wand, A. Joshua, E-mail: wand@upenn.edu [University of Pennsylvania Perelman School of Medicine, Johnson Research Foundation and Department of Biochemistry and Biophysics (United States)

2016-08-15

The application of non-uniform sampling (NUS) to relaxation experiments traditionally used to characterize the fast internal motion of proteins is quantitatively examined. Experimentally acquired Poisson-gap sampled data reconstructed with iterative soft thresholding are compared to regular sequentially sampled (RSS) data. Using ubiquitin as a model system, it is shown that 25 % sampling is sufficient for the determination of quantitatively accurate relaxation rates. When the sampling density is fixed at 25 %, the accuracy of rates is shown to increase sharply with the total number of sampled points until eventually converging near the inherent reproducibility of the experiment. Perhaps contrary to some expectations, it is found that accurate peak height reconstruction is not required for the determination of accurate rates. Instead, inaccuracies in rates arise from inconsistencies in reconstruction across the relaxation series that primarily manifest as a non-linearity in the recovered peak height. This indicates that the performance of an NUS relaxation experiment cannot be predicted from comparison of peak heights using a single RSS reference spectrum. The generality of these findings was assessed using three alternative reconstruction algorithms, eight different relaxation measurements, and three additional proteins that exhibit varying degrees of spectral complexity. From these data, it is revealed that non-linearity in peak height reconstruction across the relaxation series is strongly correlated with errors in NUS-derived relaxation rates. Importantly, it is shown that this correlation can be exploited to reliably predict the performance of an NUS-relaxation experiment by using three or more RSS reference planes from the relaxation series. The RSS reference time points can also serve to provide estimates of the uncertainty of the sampled intensity, which for a typical relaxation times series incurs no penalty in total acquisition time.

Comparative proteomics analysis of apoptotic Spodoptera frugiperda cells during p35 knockout Autographa californica multiple nucleopolyhedrovirus infection.

Science.gov (United States)

Yu, Qian; Xiong, Youhua; Liu, Jianliang; Wang, Qin; Qiu, Yuanxin; Wen, Dongling

2016-06-01

Infection with Autographa californica multiple nucleopolyhedrovirus (AcMNPV) mutants lacking a functional p35 gene can induce host cell apoptosis, which provides the possibility to use the potential of these viruses in the biological control of pest insects. Nonetheless, the proteomics or the protein changes of Spodoptera frugiperda (Sf9) cells infected with p35 knockout AcMNPV have not yet been studied. To further improve the use of AcMNPV, we set out to analyze the protein composition and protein changes of Sf9 cells of different infection stages by isobaric tag for relative and absolute quantification (iTRAQ) techniques. A total of 4004 sf9 proteins were identified by iTRAQ. After comparation of the significantly expressed 483 proteins from p35koAcMNPV-infected Sf9 cells and the significantly expressed 413 proteins from wtAcMNPV-infected Sf9 cells, we found that 226 proteins were specific to p35koAcMNPV-infected Sf9 cells. The 226 proteins were categorized according to GO classification for insects and were categorized into: biological processes, molecular functions and cellular components. Of interest, the most up-regulated proteins related to Epstein-Barr virus infection, RNA transport, Calcium signaling pathway, cGMP-PKG signaling pathway, oxidative phosphorylation and N-Glycan biosynthesis. Determination of the protein changes in p35 knockout AcMNPV-infected Sf9 cells would facilitate the better use of this virus-host cell interaction in pest insect control and other related fields. Copyright © 2016 Elsevier Inc. All rights reserved.

Muscle Glycogen Remodeling and Glycogen Phosphate Metabolism following Exhaustive Exercise of Wild Type and Laforin Knockout Mice*

Science.gov (United States)

Irimia, Jose M.; Tagliabracci, Vincent S.; Meyer, Catalina M.; Segvich, Dyann M.; DePaoli-Roach, Anna A.; Roach, Peter J.

2015-01-01

Glycogen, the repository of glucose in many cell types, contains small amounts of covalent phosphate, of uncertain function and poorly understood metabolism. Loss-of-function mutations in the laforin gene cause the fatal neurodegenerative disorder, Lafora disease, characterized by increased glycogen phosphorylation and the formation of abnormal deposits of glycogen-like material called Lafora bodies. It is generally accepted that the phosphate is removed by the laforin phosphatase. To study the dynamics of skeletal muscle glycogen phosphorylation in vivo under physiological conditions, mice were subjected to glycogen-depleting exercise and then monitored while they resynthesized glycogen. Depletion of glycogen by exercise was associated with a substantial reduction in total glycogen phosphate and the newly resynthesized glycogen was less branched and less phosphorylated. Branching returned to normal on a time frame of days, whereas phosphorylation remained suppressed over a longer period of time. We observed no change in markers of autophagy. Exercise of 3-month-old laforin knock-out mice caused a similar depletion of glycogen but no loss of glycogen phosphate. Furthermore, remodeling of glycogen to restore the basal branching pattern was delayed in the knock-out animals. From these results, we infer that 1) laforin is responsible for glycogen dephosphorylation during exercise and acts during the cytosolic degradation of glycogen, 2) excess glycogen phosphorylation in the absence of laforin delays the normal remodeling of the branching structure, and 3) the accumulation of glycogen phosphate is a relatively slow process involving multiple cycles of glycogen synthesis-degradation, consistent with the slow onset of the symptoms of Lafora disease. PMID:26216881

Enhanced brain disposition and effects of Δ9-tetrahydrocannabinol in P-glycoprotein and breast cancer resistance protein knockout mice.

Directory of Open Access Journals (Sweden)

Adena S Spiro

Full Text Available The ABC transporters P-glycoprotein (P-gp, Abcb1 and breast cancer resistance protein (Bcrp, Abcg2 regulate the CNS disposition of many drugs. The main psychoactive constituent of cannabis Δ(9-tetrahydrocannabinol (THC has affinity for P-gp and Bcrp, however it is unknown whether these transporters modulate the brain accumulation of THC and its functional effects on the CNS. Here we aim to show that mice devoid of Abcb1 and Abcg2 retain higher brain THC levels and are more sensitive to cannabinoid-induced hypothermia than wild-type (WT mice. Abcb1a/b (-/-, Abcg2 (-/- and wild-type (WT mice were injected with THC before brain and blood were collected and THC concentrations determined. Another cohort of mice was examined for THC-induced hypothermia by measuring rectal body temperature. Brain THC concentrations were higher in both Abcb1a/b (-/- and Abcg2 (-/- mice than WT mice. ABC transporter knockout mice exhibited delayed elimination of THC from the brain with the effect being more prominent in Abcg2 (-/- mice. ABC transporter knockout mice were more sensitive to THC-induced hypothermia compared to WT mice. These results show P-gp and Bcrp prolong the brain disposition and hypothermic effects of THC and offer a novel mechanism for both genetic vulnerability to the psychoactive effects of cannabis and drug interactions between CNS therapies and cannabis.

A tubulin alpha 8 mouse knockout model indicates a likely role in spermatogenesis but not in brain development.

Directory of Open Access Journals (Sweden)

Christine P Diggle

Full Text Available Tubulin alpha 8 (Tuba8 is the most divergent member of the highly conserved alpha tubulin family, and uniquely lacks two key post-translational modification sites. It is abundantly expressed in testis and muscle, with lower levels in the brain. We previously identified homozygous hypomorphic TUBA8 mutations in human subjects with a polymicrogyria (PMG syndrome, suggesting its involvement in development of the cerebral cortex. We have now generated and characterized a Tuba8 knockout mouse model. Homozygous mice were confirmed to lack Tuba8 protein in the testis, but did not display PMG and appeared to be neurologically normal. In response to this finding, we re-analyzed the human PMG subjects using whole exome sequencing. This resulted in identification of an additional homozygous loss-of-function mutation in SNAP29, suggesting that SNAP29 deficiency, rather than TUBA8 deficiency, may underlie most or all of the neurodevelopmental anomalies in these subjects. Nonetheless, in the mouse brain, Tuba8 specifically localised to the cerebellar Purkinje cells, suggesting that the human mutations may affect or modify motor control. In the testis, Tuba8 localisation was cell-type specific. It was restricted to spermiogenesis with a strong acrosomal localization that was gradually replaced by cytoplasmic distribution and was absent from spermatozoa. Although the knockout mice were fertile, the localisation pattern indicated that Tuba8 may have a role in spermatid development during spermatogenesis, rather than as a component of the mature microtubule-rich flagellum itself.

An accurate solver for forward and inverse transport

International Nuclear Information System (INIS)

Monard, Francois; Bal, Guillaume

2010-01-01

This paper presents a robust and accurate way to solve steady-state linear transport (radiative transfer) equations numerically. Our main objective is to address the inverse transport problem, in which the optical parameters of a domain of interest are reconstructed from measurements performed at the domain's boundary. This inverse problem has important applications in medical and geophysical imaging, and more generally in any field involving high frequency waves or particles propagating in scattering environments. Stable solutions of the inverse transport problem require that the singularities of the measurement operator, which maps the optical parameters to the available measurements, be captured with sufficient accuracy. This in turn requires that the free propagation of particles be calculated with care, which is a difficult problem on a Cartesian grid. A standard discrete ordinates method is used for the direction of propagation of the particles. Our methodology to address spatial discretization is based on rotating the computational domain so that each direction of propagation is always aligned with one of the grid axes. Rotations are performed in the Fourier domain to achieve spectral accuracy. The numerical dispersion of the propagating particles is therefore minimal. As a result, the ballistic and single scattering components of the transport solution are calculated robustly and accurately. Physical blurring effects, such as small angular diffusion, are also incorporated into the numerical tool. Forward and inverse calculations performed in a two-dimensional setting exemplify the capabilities of the method. Although the methodology might not be the fastest way to solve transport equations, its physical accuracy provides us with a numerical tool to assess what can and cannot be reconstructed in inverse transport theory.

visnormsc: A Graphical User Interface to Normalize Single-cell RNA Sequencing Data.

Science.gov (United States)

Tang, Lijun; Zhou, Nan

2017-12-26

Single-cell RNA sequencing (RNA-seq) allows the analysis of gene expression with high resolution. The intrinsic defects of this promising technology imports technical noise into the single-cell RNA-seq data, increasing the difficulty of accurate downstream inference. Normalization is a crucial step in single-cell RNA-seq data pre-processing. SCnorm is an accurate and efficient method that can be used for this purpose. An R implementation of this method is currently available. On one hand, the R package possesses many excellent features from R. On the other hand, R programming ability is required, which prevents the biologists who lack the skills from learning to use it quickly. To make this method more user-friendly, we developed a graphical user interface, visnormsc, for normalization of single-cell RNA-seq data. It is implemented in Python and is freely available at https://github.com/solo7773/visnormsc . Although visnormsc is based on the existing method, it contributes to this field by offering a user-friendly alternative. The out-of-the-box and cross-platform features make visnormsc easy to learn and to use. It is expected to serve biologists by simplifying single-cell RNA-seq normalization.

Age-Related Hearing Loss in Mn-SOD Heterozygous Knockout Mice

Directory of Open Access Journals (Sweden)

Makoto Kinoshita

2013-01-01

Full Text Available Age-related hearing loss (AHL reduces the quality of life for many elderly individuals. Manganese superoxide dismutase (Mn-SOD, one of the antioxidant enzymes acting within the mitochondria, plays a crucial role in scavenging reactive oxygen species (ROS. To determine whether reduction in Mn-SOD accelerates AHL, we evaluated auditory function in Mn-SOD heterozygous knockout (HET mice and their littermate wild-type (WT C57BL/6 mice by means of auditory brainstem response (ABR. Mean ABR thresholds were significantly increased at 16 months when compared to those at 4 months in both WT and HET mice, but they did not significantly differ between them at either age. The extent of hair cell loss, spiral ganglion cell density, and thickness of the stria vascularis also did not differ between WT and HET mice at either age. At 16 months, immunoreactivity of 8-hydroxydeoxyguanosine was significantly greater in the SGC and SV in HET mice compared to WT mice, but that of 4-hydroxynonenal did not differ between them. These findings suggest that, although decrease of Mn-SOD by half may increase oxidative stress in the cochlea to some extent, it may not be sufficient to accelerate age-related cochlear damage under physiological aging process.

Anti-Atherosclerotic Action of Agmatine in ApoE-Knockout Mice.

Science.gov (United States)

Wiśniewska, Anna; Olszanecki, Rafał; Totoń-Żurańska, Justyna; Kuś, Katarzyna; Stachowicz, Aneta; Suski, Maciej; Gębska, Anna; Gajda, Mariusz; Jawień, Jacek; Korbut, Ryszard

2017-08-04

Atherosclerosis is an inflammatory disease in which dysfunction of mitochondria play an important role, and disorders of lipid management intensify this process. Agmatine, an endogenous polyamine formed by decarboxylation of arginine, exerts a protective effect on mitochondria and modulates fatty acid metabolism. We investigated the effect of exogenous agmatine on the development of atherosclerosis and changes in lipid profile in apolipoprotein E knockout (apoE-/-) mice. Agmatine caused an approximate 40% decrease of atherosclerotic lesions, as estimated by en face and cross-section methods with an influence on macrophage but not on smooth muscle content in the plaques. Agmatine treatment did not changed gelatinase activity within the plaque area. What is more, the action of agmatine was associated with an increase in the number of high density lipoproteins (HDL) in blood. Real-Time PCR analysis showed that agmatine modulates liver mRNA levels of many factors involved in oxidation of fatty acid and cholesterol biosynthesis. Two-dimensional electrophoresis coupled with mass spectrometry identified 27 differentially expressed mitochondrial proteins upon agmatine treatment in the liver of apoE-/- mice, mostly proteins related to metabolism and apoptosis. In conclusion, prolonged administration of agmatine inhibits atherosclerosis in apoE-/- mice; however, the exact mechanisms linking observed changes and elevations of HDL plasma require further investigation.

AMS measurement of C-14 concentration in a single-year ring of a 2500-yr-old tree

International Nuclear Information System (INIS)

Sakurai, H.; Gandou, T.; Kato, W.; Sawaki, Y.; Matsumoto, T.; Aoki, T.; Matsuzaki, H.; Gunji, S.; Tokanai, F.

2004-01-01

The 14 C concentration in rings of an old tree that date back approximately 2500 yr has been measured at single-year intervals with a highly accurate liquid scintillation counter (LSC) (0.2%) to investigate the 11-yr periodicity of solar activity. To investigate the applicability of AMS to accurate 14 C measurement, 16 graphite samples produced from the cellulose of a single-year tree ring of a 2500-yr-old cedar were measured with the micro analysis laboratory tandem (MALT) accelerator, at The University of Tokyo, and the results were compared with the 14 C age determined using LSC. The average 14 C age of the single-year tree ring calculated from 16 measurements was 2496 ± 23 yr BP, corresponding to the statistical accuracy of 0.26%. This was consistent with the age of 2514 ± 23 yr BP determined using LSC within the acceptable error range, which indicates that accelerator mass spectrometry (AMS) is applicable for accurate 14 C measurement using multi-graphite for the same single-year tree ring

EMHP: an accurate automated hole masking algorithm for single-particle cryo-EM image processing.

Science.gov (United States)

Berndsen, Zachary; Bowman, Charles; Jang, Haerin; Ward, Andrew B

2017-12-01

The Electron Microscopy Hole Punch (EMHP) is a streamlined suite of tools for quick assessment, sorting and hole masking of electron micrographs. With recent advances in single-particle electron cryo-microscopy (cryo-EM) data processing allowing for the rapid determination of protein structures using a smaller computational footprint, we saw the need for a fast and simple tool for data pre-processing that could run independent of existing high-performance computing (HPC) infrastructures. EMHP provides a data preprocessing platform in a small package that requires minimal python dependencies to function. https://www.bitbucket.org/chazbot/emhp Apache 2.0 License. bowman@scripps.edu. Supplementary data are available at Bioinformatics online. © The Author(s) 2017. Published by Oxford University Press.

Can an electronic device with a single cuff be accurate in a wide range of arm size? Validation of the Visomat Comfort 20/40 device for home blood pressure monitoring.

Science.gov (United States)

Stergiou, G S; Tzamouranis, D; Nasothimiou, E G; Protogerou, A D

2008-11-01

An appropriate cuff according to the individual's arm circumference is recommended with all blood pressure (BP) monitors. An electronic device for home monitoring has been developed (Visomat Comfort 20/40) that estimates the individual's arm circumference by measuring the cuff filing volume and makes an adjustment of measured BP taking into account the estimated arm circumference. Thus the manufacturer recommends the use of a single cuff for arm circumference 23-43 cm. The device accuracy was assessed using the European Society of Hypertension International Protocol. Simultaneous BP measurements were obtained in 33 adults by two observers (connected mercury sphygmomanometers) four times, sequentially with three measurements taken using the tested device. Absolute device-observer BP differences were classified into difference differences differences difference (systolic/diastolic) was 3.7 +/- 5.6/-1.5 +/- 4.7 mm Hg (4.7 +/- 4.9/ - 1.7 +/- 4.3 in arm circumference 23-29 cm [39 readings] and 3.1 +/- 5.9/-1.4 +/- 5.0 in arm 30-34 cm [60 readings], P=NS). In conclusion, the device fulfils the International Protocol requirements and can be recommended for clinical use. Interestingly, the device was accurate using a single cuff in a wide range of arm circumference (23-34 cm). This study provides no information about the device accuracy in larger arms.

Highly Accurate Classification of Watson-Crick Basepairs on Termini of Single DNA Molecules

Science.gov (United States)

Winters-Hilt, Stephen; Vercoutere, Wenonah; DeGuzman, Veronica S.; Deamer, David; Akeson, Mark; Haussler, David

2003-01-01

We introduce a computational method for classification of individual DNA molecules measured by an α-hemolysin channel detector. We show classification with better than 99% accuracy for DNA hairpin molecules that differ only in their terminal Watson-Crick basepairs. Signal classification was done in silico to establish performance metrics (i.e., where train and test data were of known type, via single-species data files). It was then performed in solution to assay real mixtures of DNA hairpins. Hidden Markov Models (HMMs) were used with Expectation/Maximization for denoising and for associating a feature vector with the ionic current blockade of the DNA molecule. Support Vector Machines (SVMs) were used as discriminators, and were the focus of off-line training. A multiclass SVM architecture was designed to place less discriminatory load on weaker discriminators, and novel SVM kernels were used to boost discrimination strength. The tuning on HMMs and SVMs enabled biophysical analysis of the captured molecule states and state transitions; structure revealed in the biophysical analysis was used for better feature selection. PMID:12547778

Spectrally accurate initial data in numerical relativity

Science.gov (United States)

Battista, Nicholas A.

Einstein's theory of general relativity has radically altered the way in which we perceive the universe. His breakthrough was to realize that the fabric of space is deformable in the presence of mass, and that space and time are linked into a continuum. Much evidence has been gathered in support of general relativity over the decades. Some of the indirect evidence for GR includes the phenomenon of gravitational lensing, the anomalous perihelion of mercury, and the gravitational redshift. One of the most striking predictions of GR, that has not yet been confirmed, is the existence of gravitational waves. The primary source of gravitational waves in the universe is thought to be produced during the merger of binary black hole systems, or by binary neutron stars. The starting point for computer simulations of black hole mergers requires highly accurate initial data for the space-time metric and for the curvature. The equations describing the initial space-time around the black hole(s) are non-linear, elliptic partial differential equations (PDE). We will discuss how to use a pseudo-spectral (collocation) method to calculate the initial puncture data corresponding to single black hole and binary black hole systems.

Recording the dynamic endocytosis of single gold nanoparticles by AFM-based force tracing.

Science.gov (United States)

Ding, Bohua; Tian, Yongmei; Pan, Yangang; Shan, Yuping; Cai, Mingjun; Xu, Haijiao; Sun, Yingchun; Wang, Hongda

2015-05-07

We utilized force tracing to directly record the endocytosis of single gold nanoparticles (Au NPs) with different sizes, revealing the size-dependent endocytosis dynamics and the crucial role of membrane cholesterol. The force, duration and velocity of Au NP invagination are accurately determined at the single-particle and microsecond level unprecedentedly.

Electron scattering and nuclear structure

International Nuclear Information System (INIS)

Frois, B.

1987-01-01

The search for the appropriate degrees of freedom to describe nuclei is the central focus of nuclear physics today. Therefore the authors explore in this review their current understanding of nuclear structure as defined by electromagnetic data. The precision of the electromagnetic probe allows us to define accurately the limits of present theoretical descriptions. The authors review here a broad range of subjects that have been addressed by recent experiments, from the study of meson exchange currents and single-particle distributions to collective excitations in heavy nuclei. However, they do not discuss elastic magnetic scattering, inelastic excitation of discrete states, or single-nucleon knockout reactions since these reactions were recently reviewed. The principal aim of this review is to offer a fresh perspective on nuclear structure, based on the new generation of electron scattering data presented here and in the above-mentioned articles

Inflammation in Lafora Disease: Evolution with Disease Progression in Laforin and Malin Knock-out Mouse Models.

Science.gov (United States)

López-González, Irene; Viana, Rosa; Sanz, Pascual; Ferrer, Isidre

2017-07-01

Lafora progressive myoclonus epilepsy (Lafora disease, LD) is a fatal rare autosomal recessive neurodegenerative disorder characterized by the accumulation of insoluble ubiquitinated polyglucosan inclusions in the cytoplasm of neurons, which is most commonly associated with mutations in two genes: EPM2A, encoding the glucan phosphatase laforin, and EPM2B, encoding the E3-ubiquitin ligase malin. The present study analyzes possible inflammatory responses in the mouse lines Epm2a -/- (laforin knock-out) and Epm2b -/- (malin knock-out) with disease progression. Increased numbers of reactive astrocytes (expressing the GFAP marker) and microglia (expressing the Iba1 marker) together with increased expression of genes encoding cytokines and mediators of the inflammatory response occur in both mouse lines although with marked genotype differences. C3ar1 and CxCl10 messenger RNAs (mRNAs) are significantly increased in Epm2a -/- mice aged 12 months when compared with age-matched controls, whereas C3ar1, C4b, Ccl4, CxCl10, Il1b, Il6, Tnfα, and Il10ra mRNAs are significantly upregulated in Epm2b -/- at the same age. This is accompanied by increased protein levels of IL1-β, IL6, TNFα, and Cox2 particularly in Epm2b -/- mice. The severity of inflammatory changes correlates with more severe clinical symptoms previously described in Epm2b -/- mice. These findings show for the first time increased innate inflammatory responses in a neurodegenerative disease with polyglucosan intraneuronal deposits which increase with disease progression, in a way similar to what is seen in neurodegenerative diseases with abnormal protein aggregates. These findings also point to the possibility of using anti-inflammatory agents to mitigate the degenerative process in LD.

Remarks on the observability of single beacon underwater navigation

DEFF Research Database (Denmark)

Jouffroy, Jerome; Ross, Andrew

This paper contributes a simple and intuitive result in the analysis of underwater navigation using a single ranging beacon. This analysis should help with the design of small and lightweight underwater vehicles by reducing the amount of instrumentation required for accurate navigation. The concept...

Highly accurate fluorogenic DNA sequencing with information theory-based error correction.

Science.gov (United States)

Chen, Zitian; Zhou, Wenxiong; Qiao, Shuo; Kang, Li; Duan, Haifeng; Xie, X Sunney; Huang, Yanyi

2017-12-01

Eliminating errors in next-generation DNA sequencing has proved challenging. Here we present error-correction code (ECC) sequencing, a method to greatly improve sequencing accuracy by combining fluorogenic sequencing-by-synthesis (SBS) with an information theory-based error-correction algorithm. ECC embeds redundancy in sequencing reads by creating three orthogonal degenerate sequences, generated by alternate dual-base reactions. This is similar to encoding and decoding strategies that have proved effective in detecting and correcting errors in information communication and storage. We show that, when combined with a fluorogenic SBS chemistry with raw accuracy of 98.1%, ECC sequencing provides single-end, error-free sequences up to 200 bp. ECC approaches should enable accurate identification of extremely rare genomic variations in various applications in biology and medicine.

Functional characterization of Pol III U6 promoters for gene knockdown and knockout in Plutella xylostella.

Science.gov (United States)

Huang, Yuping; Wang, Yajun; Zeng, Baosheng; Liu, Zhaoxia; Xu, Xuejiao; Meng, Qian; Huang, Yongping; Yang, Guang; Vasseur, Liette; Gurr, Geoff M; You, Minsheng

2017-10-01

RNA polymerase type III (Pol-III) promoters such as U6 are commonly used to express small RNAs, including short hairpin RNAs (shRNAs) and single guide RNAs (sgRNAs). Functional U6 promoters are widely used in CRISPR systems, and their characterization can facilitate genome editing of non-model organisms. In the present study, six U6 small nuclear RNA (snRNA) promoters containing two conserved elements of a proximal sequence element (PSEA) and a TATA box, were identified and characterized in the diamondback moth (Plutella xylostella) genome. Relative efficiency of the U6 promoters to express shRNA induced EGFP knockdown was tested in a P. xylostella cell line, revealing that the PxU6:3 promoter had the strongest expression effect. Further work with the PxU6:3 promoter showed its efficacy in EGFP knockout using CRISPR/Cas9 system in the cells. The expression plasmids with versatile Pxabd-A gene specific sgRNA driven by the PxU6:3 promoter, combined with Cas9 mRNA, could induce mutagenesis at specific genomic loci in vivo. The phenotypes induced by sgRNA expression plasmids were similar to those done in vitro transcription sgRNAs. A plasmid with two tandem arranged PxU6:3:sgRNA expression cassettes targeting Pxabd-A loci was generated, which caused a 28,856 bp fragment deletion, suggesting that the multi-sgRNA expression plasmid can be used for multi-targeting. Our work indicates that U6 snRNA promoters can be used for functional studies of genes with the approach of reverse genetics in P. xylostella. These essential promoters also provide valuable potential for CRISPR-derived gene drive as a tactic for population control in this globally significant pest. Copyright © 2017 Elsevier Ltd. All rights reserved.

Action potentials and ion conductances in wild-type and CALHM1-knockout type II taste cells

Science.gov (United States)

Saung, Wint Thu; Foskett, J. Kevin

2017-01-01

Taste bud type II cells fire action potentials in response to tastants, triggering nonvesicular ATP release to gustatory neurons via voltage-gated CALHM1-associated ion channels. Whereas CALHM1 regulates mouse cortical neuron excitability, its roles in regulating type II cell excitability are unknown. In this study, we compared membrane conductances and action potentials in single identified TRPM5-GFP-expressing circumvallate papillae type II cells acutely isolated from wild-type (WT) and Calhm1 knockout (KO) mice. The activation kinetics of large voltage-gated outward currents were accelerated in cells from Calhm1 KO mice, and their associated nonselective tail currents, previously shown to be highly correlated with ATP release, were completely absent in Calhm1 KO cells, suggesting that CALHM1 contributes to all of these currents. Calhm1 deletion did not significantly alter resting membrane potential or input resistance, the amplitudes and kinetics of Na+ currents either estimated from action potentials or recorded from steady-state voltage pulses, or action potential threshold, overshoot peak, afterhyperpolarization, and firing frequency. However, Calhm1 deletion reduced the half-widths of action potentials and accelerated the deactivation kinetics of transient outward currents, suggesting that the CALHM1-associated conductance becomes activated during the repolarization phase of action potentials. NEW & NOTEWORTHY CALHM1 is an essential ion channel component of the ATP neurotransmitter release mechanism in type II taste bud cells. Its contribution to type II cell resting membrane properties and excitability is unknown. Nonselective voltage-gated currents, previously associated with ATP release, were absent in cells lacking CALHM1. Calhm1 deletion was without effects on resting membrane properties or voltage-gated Na+ and K+ channels but contributed modestly to the kinetics of action potentials. PMID:28202574

An accurate and portable solid state neutron rem meter

Energy Technology Data Exchange (ETDEWEB)

Oakes, T.M. [Nuclear Science and Engineering Institute, University of Missouri, Columbia, MO (United States); Bellinger, S.L. [Department of Mechanical and Nuclear Engineering, Kansas State University, Manhattan, KS (United States); Miller, W.H. [Nuclear Science and Engineering Institute, University of Missouri, Columbia, MO (United States); Missouri University Research Reactor, Columbia, MO (United States); Myers, E.R. [Department of Physics, University of Missouri, Kansas City, MO (United States); Fronk, R.G.; Cooper, B.W [Department of Mechanical and Nuclear Engineering, Kansas State University, Manhattan, KS (United States); Sobering, T.J. [Electronics Design Laboratory, Kansas State University, KS (United States); Scott, P.R. [Department of Physics, University of Missouri, Kansas City, MO (United States); Ugorowski, P.; McGregor, D.S; Shultis, J.K. [Department of Mechanical and Nuclear Engineering, Kansas State University, Manhattan, KS (United States); Caruso, A.N., E-mail: carusoan@umkc.edu [Department of Physics, University of Missouri, Kansas City, MO (United States)

2013-08-11

Accurately resolving the ambient neutron dose equivalent spanning the thermal to 15 MeV energy range with a single configuration and lightweight instrument is desirable. This paper presents the design of a portable, high intrinsic efficiency, and accurate neutron rem meter whose energy-dependent response is electronically adjusted to a chosen neutron dose equivalent standard. The instrument may be classified as a moderating type neutron spectrometer, based on an adaptation to the classical Bonner sphere and position sensitive long counter, which, simultaneously counts thermalized neutrons by high thermal efficiency solid state neutron detectors. The use of multiple detectors and moderator arranged along an axis of symmetry (e.g., long axis of a cylinder) with known neutron-slowing properties allows for the construction of a linear combination of responses that approximate the ambient neutron dose equivalent. Variations on the detector configuration are investigated via Monte Carlo N-Particle simulations to minimize the total instrument mass while maintaining acceptable response accuracy—a dose error less than 15% for bare {sup 252}Cf, bare AmBe, an epi-thermal and mixed monoenergetic sources is found at less than 4.5 kg moderator mass in all studied cases. A comparison of the energy dependent dose equivalent response and resultant energy dependent dose equivalent error of the present dosimeter to commercially-available portable rem meters and the prior art are presented. Finally, the present design is assessed by comparison of the simulated output resulting from applications of several known neutron sources and dose rates.

Accurate, fully-automated NMR spectral profiling for metabolomics.

Directory of Open Access Journals (Sweden)

Siamak Ravanbakhsh

Full Text Available Many diseases cause significant changes to the concentrations of small molecules (a.k.a. metabolites that appear in a person's biofluids, which means such diseases can often be readily detected from a person's "metabolic profile"-i.e., the list of concentrations of those metabolites. This information can be extracted from a biofluids Nuclear Magnetic Resonance (NMR spectrum. However, due to its complexity, NMR spectral profiling has remained manual, resulting in slow, expensive and error-prone procedures that have hindered clinical and industrial adoption of metabolomics via NMR. This paper presents a system, BAYESIL, which can quickly, accurately, and autonomously produce a person's metabolic profile. Given a 1D 1H NMR spectrum of a complex biofluid (specifically serum or cerebrospinal fluid, BAYESIL can automatically determine the metabolic profile. This requires first performing several spectral processing steps, then matching the resulting spectrum against a reference compound library, which contains the "signatures" of each relevant metabolite. BAYESIL views spectral matching as an inference problem within a probabilistic graphical model that rapidly approximates the most probable metabolic profile. Our extensive studies on a diverse set of complex mixtures including real biological samples (serum and CSF, defined mixtures and realistic computer generated spectra; involving > 50 compounds, show that BAYESIL can autonomously find the concentration of NMR-detectable metabolites accurately (~ 90% correct identification and ~ 10% quantification error, in less than 5 minutes on a single CPU. These results demonstrate that BAYESIL is the first fully-automatic publicly-accessible system that provides quantitative NMR spectral profiling effectively-with an accuracy on these biofluids that meets or exceeds the performance of trained experts. We anticipate this tool will usher in high-throughput metabolomics and enable a wealth of new applications of

Accurate path integration in continuous attractor network models of grid cells.

Science.gov (United States)

Burak, Yoram; Fiete, Ila R

2009-02-01

Grid cells in the rat entorhinal cortex display strikingly regular firing responses to the animal's position in 2-D space and have been hypothesized to form the neural substrate for dead-reckoning. However, errors accumulate rapidly when velocity inputs are integrated in existing models of grid cell activity. To produce grid-cell-like responses, these models would require frequent resets triggered by external sensory cues. Such inadequacies, shared by various models, cast doubt on the dead-reckoning potential of the grid cell system. Here we focus on the question of accurate path integration, specifically in continuous attractor models of grid cell activity. We show, in contrast to previous models, that continuous attractor models can generate regular triangular grid responses, based on inputs that encode only the rat's velocity and heading direction. We consider the role of the network boundary in the integration performance of the network and show that both periodic and aperiodic networks are capable of accurate path integration, despite important differences in their attractor manifolds. We quantify the rate at which errors in the velocity integration accumulate as a function of network size and intrinsic noise within the network. With a plausible range of parameters and the inclusion of spike variability, our model networks can accurately integrate velocity inputs over a maximum of approximately 10-100 meters and approximately 1-10 minutes. These findings form a proof-of-concept that continuous attractor dynamics may underlie velocity integration in the dorsolateral medial entorhinal cortex. The simulations also generate pertinent upper bounds on the accuracy of integration that may be achieved by continuous attractor dynamics in the grid cell network. We suggest experiments to test the continuous attractor model and differentiate it from models in which single cells establish their responses independently of each other.

Spectrally resolved single-molecule electrometry

Science.gov (United States)

Ruggeri, F.; Krishnan, M.

2018-03-01

Escape-time electrometry is a recently developed experimental technique that offers the ability to measure the effective electrical charge of a single biomolecule in solution with sub-elementary charge precision. The approach relies on measuring the average escape-time of a single charged macromolecule or molecular species transiently confined in an electrostatic fluidic trap. Comparing the experiments with the predictions of a mean-field model of molecular electrostatics, we have found that the measured effective charge even reports on molecular conformation, e.g., folded or disordered state, and non-uniform charge distribution in disordered proteins or polyelectrolytes. Here we demonstrate the ability to use the spectral dimension to distinguish minute differences in electrical charge between individual molecules or molecular species in a single simultaneous measurement, under identical experimental conditions. Using one spectral channel for referenced measurement, this kind of photophysical distinguishability essentially eliminates the need for accurate knowledge of key experimental parameters, otherwise obtained through intensive characterization of the experimental setup. As examples, we demonstrate the ability to detect small differences (˜5%) in the length of double-stranded DNA fragments as well as single amino acid exchange in an intrinsically disordered protein, prothymosin α.

Predictive validity and immune cell involvement in the pathogenesis of piroxicam-accelerated colitis in interleukin-10 knockout mice

DEFF Research Database (Denmark)

Holgersen, Kristine; Kvist, Peter Helding; Hansen, Axel Jacob Kornerup

2014-01-01

Piroxicam administration is a method for induction of enterocolitis in interleukin-10 knockout (IL-10 k.o.) mice. The piroxicam-accelerated colitis (PAC) IL-10 k.o. model combines a dysregulated immune response against the gut microbiota with a decreased mucosal integrity. The predictive validity...... and pathogenic mechanisms of the model have not been thoroughly investigated. In this study, IL-10 k.o. mice received piroxicam in the chow, and model qualification was performed by examining the efficacy of prophylactic anti-IL-12/23p40 monoclonal antibody (mAb), anti-TNFαmAb, cyclosporine A (CsA) and oral...

Cannabinoid 1 receptor knockout mice display cold allodynia, but enhanced recovery from spared-nerve injury-induced mechanical hypersensitivity.

Science.gov (United States)

Sideris, Alexandra; Piskoun, Boris; Russo, Lori; Norcini, Monica; Blanck, Thomas; Recio-Pinto, Esperanza

2016-01-01

The function of the Cannabinoid 1 receptor (CB1R) in the development of neuropathic pain is not clear. Mounting evidence suggest that CB1R expression and activation may contribute to pain. Cannabinoid 1 receptor knockout mice (CB1R-/-) generated on a C57Bl/6 background exhibit hypoalgesia in the hotplate assay and formalin test. These findings suggest that Cannabinoid 1 receptor expression mediates the responses to at least some types of painful stimuli. By using this mouse line, we sought to determine if the lack of Cannabinoid 1 receptor unveils a general hypoalgesic phenotype, including protection against the development of neuropathic pain. The acetone test was used to measure cold sensitivity, the electronic von Frey was used to measure mechanical thresholds before and after spared-nerve injury, and analysis of footprint patterns was conducted to determine if motor function is differentially affected after nerve-injury in mice with varying levels of Cannabinoid 1 receptor. At baseline, CB1R-/- mice were hypersensitive in the acetone test, and this phenotype was maintained after spared-nerve injury. Using calcium imaging of lumbar dorsal root ganglion (DRG) cultures, a higher percentage of neurons isolated from CB1R-/- mice were menthol sensitive relative to DRG isolated from wild-type (CB1R+/+) mice. Baseline mechanical thresholds did not differ among genotypes, and mechanical hypersensitivity developed similarly in the first two weeks following spared-nerve injury (SNI). At two weeks post-SNI, CB1R-/- mice recovered significantly from mechanical hypersensitivity, while the CB1R+/+ mice did not. Heterozygous knockouts (CB1R+/-) transiently developed cold allodynia only after injury, but recovered mechanical thresholds to a similar extent as the CB1R-/- mice. Sciatic functional indices, which reflect overall nerve health, and alternation coefficients, which indicate uniformity of strides, were not significantly different among genotypes. Cold allodynia and

Single-sided natural ventilation through a centre-pivot roof window

DEFF Research Database (Denmark)

Iqbal, Ahsan; Nielsen, Peter V.; Gunner, Amalie

2014-01-01

The characteristics of centre pivot roof windows for wind driven single-sided ventilation has not been studied before. These types of windows are dominating roof windows in Europe. Knowledge of flow characteristics of this kind of window is essential for accurate designing of natural ventilation...... systems. In this study, numerical methods were used to characterise a centre-pivot roof window for wind-driven single-sided ventilation. A 1:20 scale model house of the Energy Flex House (Denmark) was used in this study. The roof slope was 36o. It was found that the single-sided ventilation through...

Model independent approach to the single photoelectron calibration of photomultiplier tubes

Energy Technology Data Exchange (ETDEWEB)

Saldanha, R.; Grandi, L.; Guardincerri, Y.; Wester, T.

2017-08-01

The accurate calibration of photomultiplier tubes is critical in a wide variety of applications in which it is necessary to know the absolute number of detected photons or precisely determine the resolution of the signal. Conventional calibration methods rely on fitting the photomultiplier response to a low intensity light source with analytical approximations to the single photoelectron distribution, often leading to biased estimates due to the inability to accurately model the full distribution, especially at low charge values. In this paper we present a simple statistical method to extract the relevant single photoelectron calibration parameters without making any assumptions about the underlying single photoelectron distribution. We illustrate the use of this method through the calibration of a Hamamatsu R11410 photomultiplier tube and study the accuracy and precision of the method using Monte Carlo simulations. The method is found to have significantly reduced bias compared to conventional methods and works under a wide range of light intensities, making it suitable for simultaneously calibrating large arrays of photomultiplier tubes.

An accurate bound on tensor-to-scalar ratio and the scale of inflation

International Nuclear Information System (INIS)

Choudhury, Sayantan; Mazumdar, Anupam

2014-01-01

In this paper we provide an accurate bound on primordial gravitational waves, i.e. tensor-to-scalar ratio (r) for a general class of single-field models of inflation where inflation occurs always below the Planck scale, and the field displacement during inflation remains sub-Planckian. If inflation has to make connection with the real particle physics framework then it must be explained within an effective field theory description where it can be trustable below the UV cut-off of the scale of gravity. We provide an analytical estimation and estimate the largest possible r, i.e. r⩽0.12, for the field displacement less than the Planck cut-off

Improving Genetic Evaluation of Litter Size Using a Single-step Model

DEFF Research Database (Denmark)

Guo, Xiangyu; Christensen, Ole Fredslund; Ostersen, Tage

A recently developed single-step method allows genetic evaluation based on information from phenotypes, pedigree and markers simultaneously. This paper compared reliabilities of predicted breeding values obtained from single-step method and the traditional pedigree-based method for two litter size...... traits, total number of piglets born (TNB), and litter size at five days after birth (Ls 5) in Danish Landrace and Yorkshire pigs. The results showed that the single-step method combining phenotypic and genotypic information provided more accurate predictions than the pedigree-based method, not only...

Effects of stimulation of soluble guanylate cyclase on diabetic nephropathy in diabetic eNOS knockout mice on top of angiotensin II receptor blockade.

Directory of Open Access Journals (Sweden)

Ina M Ott

Full Text Available The prevalence of diabetes mellitus and its complications, such as diabetic nephropathy (DN, is rising worldwide and prevention and treatment are therefore becoming increasingly important. Therapy of DN is particularly important for patients who do not adequately respond to angiotensin receptor blocker (ARB treatment. Novel approaches include the stimulation of soluble guanylate cyclase (sGC as it is reported to have beneficial effects on cardiac and renal damage. We aimed to investigate the effects of the sGC stimulator riociguat and ARB telmisartan on kidney function and structure in a hypertensive model of diabetic nephropathy. Seventy-six diabetic male eNOS knockout C57BL/6J mice were randomly divided after having received streptozotocin: telmisartan (1 mg/kg/d, riociguat (3 mg/kg/d, riociguat+telmisartan (3+1 mg/kg/d, and vehicle. Fourteen mice were used as non-diabetic controls. Treatment duration was 11 weeks. Glucose concentrations were increased and similar in all diabetic groups. Telmisartan insignificantly reduced blood pressure by 5.9 mmHg compared with diabetic controls (111.2±2.3 mmHg vs. 117.1±2.2 mmHg; p = 0.071. Treatment with riociguat both alone and in combination with telmisartan led to a significant reduction of blood pressure towards diabetic vehicle (105.2±2.5 mmHg and 105.0±3.2 mmHg, respectively, vs. 117.1±2.2 mmHg. Combined treatment also significantly decreased albuminuria compared with diabetic controls (47.3±9.6 µg/24 h vs. 170.8±34.2 µg/24 h; p = 0.002 reaching levels similar to those of non-diabetic controls (34.4±10.6 µg/24 h, whereas the reduction by single treatment with either telmisartan (97.8±26.4 µg/24 h or riociguat (97.1±15.7 µg/24 h was not statistically significant. The combination treatment led to a significant (p<0.01 decrease of tissue immunoreactivity of malondialdehyde, as consequence of reduced oxidative stress. In conclusion, stimulation of sGC significantly reduced urinary

Ground Vibration Attenuation Measurement using Triaxial and Single Axis Accelerometers

Science.gov (United States)

Mohammad, A. H.; Yusoff, N. A.; Madun, A.; Tajudin, S. A. A.; Zahari, M. N. H.; Chik, T. N. T.; Rahman, N. A.; Annuar, Y. M. N.

2018-04-01

Peak Particle Velocity is one of the important term to show the level of the vibration amplitude especially traveling wave by distance. Vibration measurement using triaxial accelerometer is needed to obtain accurate value of PPV however limited by the size and the available channel of the data acquisition module for detailed measurement. In this paper, an attempt to estimate accurate PPV has been made by using only a triaxial accelerometer together with multiple single axis accelerometer for the ground vibration measurement. A field test was conducted on soft ground using nine single axis accelerometers and a triaxial accelerometer installed at nine receiver location R1 to R9. Based from the obtained result, the method shows convincing similarity between actual PPV with the calculated PPV with error ratio 0.97. With the design method, vibration measurement equipment size can be reduced with fewer channel required.

Establishment of pten knockout medaka with transcription activator-like effector nucleases (TALENs as a model of PTEN deficiency disease.

Directory of Open Access Journals (Sweden)

Yuriko Matsuzaki

Full Text Available Phosphatase and tensin homolog (PTEN is a lipid and protein phosphatase that antagonizes signaling by the phosphatidylinositol 3-kinase (PI3K-AKT signaling pathway. The PTEN gene is a major tumor suppressor, with mutations of this gene occurring frequently in tumors of humans and mice. We have now developed mutant medaka deficient in PTEN with the use of transcription activator-like effector nuclease (TALEN technology. Medaka possesses two pten genes, ptena and ptenb, similar to zebrafish. We established 16 ptena mutant lines and two ptenb mutant lines. Homozygous single pten mutants were found to be viable and fertile. In contrast, pten double-knockout (dko embryos manifested severe abnormalities in vasculogenesis, eye size, and tail development at 72 hours post fertilization(hpf and died before hatching. Immunoblot analysis revealed that the ratio of phosphorylated to total forms of AKT (pAKT/AKT in pten dko embryos was four times that in wild-type embryos, indicative of up-regulation of signaling by the PI3K-AKT pathway. Treatment of pten dko embryos with the PI3K inhibitor LY294002 reduced the pAKT/AKT ratio by about one-half and partially rescued the defect in vasculogenesis. Additional inhibitors of the PI3K-AKT pathway, including rapamycin and N-α-tosyl-L-phenylalanyl chloromethyl ketone, also partially restored vasculogenesis in the dko embryos. Our model system thus allows pten dko embryos to be readily distinguished from wild-type embryos at an early stage of development and is suitable for the screening of drugs able to compensate for PTEN deficiency.

Split dose recovery studies using homologous recombination deficient gene knockout chicken B lymphocyte cells

International Nuclear Information System (INIS)

Rao, B.S.S.; Tano, Kaori; Utsumi, Hiroshi; Takeda, Shunichi

2007-01-01

To understand the role of proteins involved in double strand breaks (DSB) repair modulating sublethal damage (SLD) recovery, chicken B lymphoma (DT 40) cell lines either proficient or deficient in RAD52, XRCC2, XRCC3, RAD51C and RAD51D were subjected to fractionated irradiation and their survival curves charted. Survival curves of both WT DT40 and RAD52 -/- cells had a big shoulder while all the other cells exhibited small shoulders. However, at the higher doses of radiation, RAD51C -/- cells displayed hypersensitivity comparable to the data obtained for the homologous recombination deficient RAD54 -/- cells. Repair of SLD was measured as an increase in survival after a split dose irradiation with an interval of incubation between the radiation doses. All the cell lines (parental DT40 and genetic knockout cell lines viz., RAD52 -/- , XRCC2 -/- XRCC3 -/- RAD51C -/- and RAD51D -/- ) used in this study demonstrated a typical split-dose recovery capacity with a specific peak, which varied depending on the cell type. The maximum survival of WT DT40 and RAD52 -/- was reached at about 1-2 hours after the first dose of radiation and then decreased to a minimum thereafter (5 h). The increase in the survival peaked once again by about 8 hours. The survival trends observed in XRCC2 -/- , XRCC3 -/- , RAD51C -/- and RAD51D -/- knockout cells were also similar, except for the difference in the initial delay of a peak survival for RAD51D -/- and lower survival ratios. The second phase of increase in the survival in these cell lines was much slower in XRCC2 -/- , XRCC3 -/- , RAD51C -/- nd RAD51D -/- and further delayed when compared with that of RAD52 -/- and parental DT40 cells suggesting a dependence on their cell cycle kinetics. This study demonstrates that the participation of RAD52, XRCC2, XRCC3, RAD51C and RAD51D in the DSB repair via homologous recombination is of less importance in comparison to RAD54, as RAD54 deficient cells demonstrated complete absence of SLD recovery

Exposure to diesel exhaust up-regulates iNOS expression in ApoE knockout mice

International Nuclear Information System (INIS)

Bai Ni; Kido, Takashi; Kavanagh, Terrance J.; Kaufman, Joel D.; Rosenfeld, Michael E.; Breemen, Cornelis van; Eeden, Stephan F. van

2011-01-01

Traffic related particulate matter air pollution is a risk factor for cardiovascular events; however, the biological mechanisms are unclear. We hypothesize that diesel exhaust (DE) inhalation induces up-regulation of inducible nitric oxide synthase (iNOS), which is known to contribute to vascular dysfunction, progression of atherosclerosis and ultimately cardiovascular morbidity and mortality. Methods: ApoE knockout mice (30-week) were exposed to DE (at 200 μg/m 3 of particulate matter) or filtered-air (control) for 7 weeks (6 h/day, 5 days/week). iNOS expression in the blood vessels and heart was evaluated by immunohistochemistry and western blotting analysis. To examine iNOS activity, thoracic aortae were mounted in a wire myograph, and vasoconstriction stimulated by phenylephrine (PE) was measured with and without the presence of the specific inhibitor for iNOS (1400 W). NF-κB (p65) activity was examined by ELISA. The mRNA expression of iNOS and NF-κB (p65) was determined by real-time PCR. Results: DE exposure significantly enhanced iNOS expression in the thoracic aorta (4-fold) and heart (1.5 fold). DE exposure significantly attenuated PE-stimulated vasoconstriction by ∼ 20%, which was partly reversed by 1400 W. The mRNA expression of iNOS and NF-κB was significantly augmented after DE exposure. NF-κB activity was enhanced 2-fold after DE inhalation, and the augmented NF-κB activity was positively correlated with iNOS expression (R 2 = 0.5998). Conclusions: We show that exposure to DE increases iNOS expression and activity possibly via NF-κB-mediated pathway. We suspect that DE exposure-caused up-regulation of iNOS contributes to vascular dysfunction and atherogenesis, which could ultimately lead to urban air pollution-associated cardiovascular morbidity and mortality. - Highlights: → Exposed ApoE knockout mice (30-week) to diesel exhaust (DE) for 7 weeks. → Examine iNOS expression and activity in the blood vessels and heart. → DE exposure

Autism-related behavioral abnormalities in synapsin knockout mice.

Science.gov (United States)

Greco, Barbara; Managò, Francesca; Tucci, Valter; Kao, Hung-Teh; Valtorta, Flavia; Benfenati, Fabio

2013-08-15

Several synaptic genes predisposing to autism-spectrum disorder (ASD) have been identified. Nonsense and missense mutations in the SYN1 gene encoding for Synapsin I have been identified in families segregating for idiopathic epilepsy and ASD and genetic mapping analyses have identified variations in the SYN2 gene as significantly contributing to epilepsy predisposition. Synapsins (Syn I/II/III) are a multigene family of synaptic vesicle-associated phosphoproteins playing multiple roles in synaptic development, transmission and plasticity. Lack of SynI and/or SynII triggers a strong epileptic phenotype in mice associated with mild cognitive impairments that are also present in the non-epileptic SynIII(-/-) mice. SynII(-/-) and SynIII(-/-) mice also display schizophrenia-like traits, suggesting that Syns could be involved in the regulation of social behavior. Here, we studied social interaction and novelty, social recognition and social dominance, social transmission of food preference and social memory in groups of male SynI(-/-), SynII(-/-) and SynIII(-/-) mice before and after the appearance of the epileptic phenotype and compared their performances with control mice. We found that deletion of Syn isoforms widely impairs social behaviors and repetitive behaviors, resulting in ASD-related phenotypes. SynI or SynIII deletion altered social behavior, whereas SynII deletion extensively impaired various aspects of social behavior and memory, altered exploration of a novel environment and increased self-grooming. Social impairments of SynI(-/-) and SynII(-/-) mice were evident also before the onset of seizures. The results demonstrate an involvement of Syns in generation of the behavioral traits of ASD and identify Syn knockout mice as a useful experimental model of ASD and epilepsy. Copyright © 2013 Elsevier B.V. All rights reserved.

Increased anxiety-related behaviour in Hint1 knockout mice.

Science.gov (United States)

Varadarajulu, Jeeva; Lebar, Maria; Krishnamoorthy, Gurumoorthy; Habelt, Sonja; Lu, Jia; Bernard Weinstein, I; Li, Haiyang; Holsboer, Florian; Turck, Christoph W; Touma, Chadi

2011-07-07

Several reports have implicated a role for the histidine triad nucleotide-binding protein-1 (Hint1) in psychiatric disorders. We have studied the emotional behaviour of male Hint1 knockout (Hint1 KO) mice in a battery of tests and performed biochemical analyses on brain tissue. The behavioural analysis revealed that Hint1 KO mice exhibit an increased emotionality phenotype compared to wildtype (WT) mice, while no significant differences in locomotion or general exploratory activity were noted. In the elevated plus-maze (EPM) test, the Hint1 KO animals entered the open arms of the apparatus less often than WT littermates. Similarly, in the dark-light box test, Hint1 KO mice spent less time in the lit compartment and the number of entries were reduced, which further confirmed an increased anxiety-related behaviour. Moreover, the Hint1 KO animals showed significantly more struggling and less floating behaviour in the forced swim test (FST), indicating an increased emotional arousal in aversive situations. Hint1 is known as a protein kinase C (PKC) interacting protein. Western blot analysis showed that PKCγ expression was elevated in Hint1 KO compared to WT mice. Interestingly, PKCγ mRNA levels of the two groups did not show a significant difference, implying a post-transcriptional PKCγ regulation. In addition, PKC enzymatic activity was increased in Hint1 KO compared to WT mice. In summary, our results indicate a role for Hint1 and PKCγ in modulating anxiety-related and stress-coping behaviour in mice. Copyright © 2011 Elsevier B.V. All rights reserved.

Accurate Extraction of Charge Carrier Mobility in 4-Probe Field-Effect Transistors

KAUST Repository

Choi, Hyun Ho; Rodionov, Yaroslav I.; Paterson, Alexandra F.; Panidi, Julianna; Saranin, Danila; Kharlamov, Nikolai; Didenko, Sergei I.; Anthopoulos, Thomas D.; Cho, Kilwon; Podzorov, Vitaly

2018-01-01

Charge carrier mobility is an important characteristic of organic field-effect transistors (OFETs) and other semiconductor devices. However, accurate mobility determination in FETs is frequently compromised by issues related to Schottky-barrier contact resistance, that can be efficiently addressed by measurements in 4-probe/Hall-bar contact geometry. Here, it is shown that this technique, widely used in materials science, can still lead to significant mobility overestimation due to longitudinal channel shunting caused by voltage probes in 4-probe structures. This effect is investigated numerically and experimentally in specially designed multiterminal OFETs based on optimized novel organic-semiconductor blends and bulk single crystals. Numerical simulations reveal that 4-probe FETs with long but narrow channels and wide voltage probes are especially prone to channel shunting, that can lead to mobilities overestimated by as much as 350%. In addition, the first Hall effect measurements in blended OFETs are reported and how Hall mobility can be affected by channel shunting is shown. As a solution to this problem, a numerical correction factor is introduced that can be used to obtain much more accurate experimental mobilities. This methodology is relevant to characterization of a variety of materials, including organic semiconductors, inorganic oxides, monolayer materials, as well as carbon nanotube and semiconductor nanocrystal arrays.

Accurate alpha sticking fractions from improved calculations relevant for muon catalyzed fusion

International Nuclear Information System (INIS)

Szalewicz, K.

1990-05-01

Recent experiments have shown that under proper conditions a single muon may catalyze almost two hundred fusions in its lifetime. This process proceeds through formation of muonic molecular ions. Properties of these ions are central to the understanding of the phenomenon. Our work included the most accurate calculations of the energy levels and Coulombic sticking fractions for tdμ and other muonic molecular ions, calculations of Auger transition rates, calculations of corrections to the energy levels due to interactions with the most molecule, and calculation of the reactivation of muons from α particles. The majority of our effort has been devoted to the theory and computation of the influence of the strong nuclear forces on fusion rates and sticking fractions. We have calculated fusion rates for tdμ including the effects of nuclear forces on the molecular wave functions. We have also shown that these results can be reproduced to almost four digit accuracy by using a very simple quasifactorizable expression which does not require modifications of the molecular wave functions. Our sticking fractions are more accurate than any other theoretical values. We have used a more sophisticated theory than any other work and our numerical calculations have converged to at least three significant digits

Accurate Extraction of Charge Carrier Mobility in 4-Probe Field-Effect Transistors

KAUST Repository

Choi, Hyun Ho

2018-04-30

Charge carrier mobility is an important characteristic of organic field-effect transistors (OFETs) and other semiconductor devices. However, accurate mobility determination in FETs is frequently compromised by issues related to Schottky-barrier contact resistance, that can be efficiently addressed by measurements in 4-probe/Hall-bar contact geometry. Here, it is shown that this technique, widely used in materials science, can still lead to significant mobility overestimation due to longitudinal channel shunting caused by voltage probes in 4-probe structures. This effect is investigated numerically and experimentally in specially designed multiterminal OFETs based on optimized novel organic-semiconductor blends and bulk single crystals. Numerical simulations reveal that 4-probe FETs with long but narrow channels and wide voltage probes are especially prone to channel shunting, that can lead to mobilities overestimated by as much as 350%. In addition, the first Hall effect measurements in blended OFETs are reported and how Hall mobility can be affected by channel shunting is shown. As a solution to this problem, a numerical correction factor is introduced that can be used to obtain much more accurate experimental mobilities. This methodology is relevant to characterization of a variety of materials, including organic semiconductors, inorganic oxides, monolayer materials, as well as carbon nanotube and semiconductor nanocrystal arrays.

Analysis of different multiplicities and their interference in quasi-elastic cluster knock-out by fast hadrons

International Nuclear Information System (INIS)

Golovanova, N.F.; Ibraeva, E.T.; Neudatchin, V.G.

1978-01-01

Different multiplicities and their interference in hadron scattering have been investigated on the basis of a new dynamic approach to quasi-elastic knock-out of nucleon clusters by fast hadrons from light nuclei. It is shown that in the region of momentum transfer values p, where scattering multiplicities less than b are predominant, the effective numbers and form factors determined in Refs. 1) -- 3) no longer act as pure structural nuclear factors (b means the number of nucleons in the knocked-out cluster). These characteristics are significantly dependent on the process dynamics. Only in the region of values p, where the maximum hadron scattering multiplicity b is realized, the effective numbers and form factors do assume the purely structural meaning. (auth.)

Pharmacokinetics of dietary cancer chemopreventive compound dibenzoylmethane in rats and the impact of nanoemulsion and genetic knockout of Nrf2 on its disposition.

Science.gov (United States)

Lin, Wen; Hong, Jin-Liern; Shen, Guoxiang; Wu, Rachel T; Wang, Yuwen; Huang, Mou-Tuan; Newmark, Harold L; Huang, Qingrong; Khor, Tin Oo; Heimbach, Tycho; Kong, Ah-Ng

2011-03-01

The pharmacokinetic disposition of a dietary cancer chemopreventive compound dibenzoylmethane (DBM) was studied in male Sprague-Dawley rats after intravenous (i.v.) and oral (p.o.) administrations. Following a single i.v. bolus dose, the mean plasma clearance (CL) of DBM was low compared with the hepatic blood flow. DBM displayed a high volume of distribution (Vss). The elimination terminal t1/2 was long. The mean CL, Vss and AUC0-∞/dose were similar between the i.v. 10 and 10 mg/kg doses. After single oral doses (10, 50 and 250 mg/kg), the absolute oral bioavailability (F*) of DBM was 7.4%-13.6%. The increase in AUC was not proportional to the oral doses, suggesting non-linearity. In silico prediction of oral absorption also demonstrated low DBM absorption in vivo. An oil-in-water nanoemulsion containing DBM was formulated to potentially overcome the low F* due to poor water solubility of DBM, with enhanced oral absorption. Finally, to examine the role of Nrf2 on the pharmacokinetics of DBM, since DBM activates the Nrf2-dependent detoxification pathways, Nrf2 wild-type (+/+) mice and Nrf2 knockout (-/-) mice were utilized. There was an increased systemic plasma exposure of DBM in Nrf2 (-/-) mice, suggesting that the Nrf2 genotype could also play a role in the pharmacokinetic disposition of DBM. Taken together, the results show that DBM has low oral bioavailability which could be due in part to poor water solubility and this could be overcome by a nanotechnology-based drug delivery system and furthermore the Nrf2 genotype could also play a role in the pharmacokinetics of DBM. Copyright © 2010 John Wiley & Sons, Ltd.

Mouse nuclear myosin I knock-out shows interchangeability and redundancy of myosin isoforms in the cell nucleus.

Science.gov (United States)

Venit, Tomáš; Dzijak, Rastislav; Kalendová, Alžběta; Kahle, Michal; Rohožková, Jana; Schmidt, Volker; Rülicke, Thomas; Rathkolb, Birgit; Hans, Wolfgang; Bohla, Alexander; Eickelberg, Oliver; Stoeger, Tobias; Wolf, Eckhard; Yildirim, Ali Önder; Gailus-Durner, Valérie; Fuchs, Helmut; de Angelis, Martin Hrabě; Hozák, Pavel

2013-01-01

Nuclear myosin I (NM1) is a nuclear isoform of the well-known "cytoplasmic" Myosin 1c protein (Myo1c). Located on the 11(th) chromosome in mice, NM1 results from an alternative start of transcription of the Myo1c gene adding an extra 16 amino acids at the N-terminus. Previous studies revealed its roles in RNA Polymerase I and RNA Polymerase II transcription, chromatin remodeling, and chromosomal movements. Its nuclear localization signal is localized in the middle of the molecule and therefore directs both Myosin 1c isoforms to the nucleus. In order to trace specific functions of the NM1 isoform, we generated mice lacking the NM1 start codon without affecting the cytoplasmic Myo1c protein. Mutant mice were analyzed in a comprehensive phenotypic screen in cooperation with the German Mouse Clinic. Strikingly, no obvious phenotype related to previously described functions has been observed. However, we found minor changes in bone mineral density and the number and size of red blood cells in knock-out mice, which are most probably not related to previously described functions of NM1 in the nucleus. In Myo1c/NM1 depleted U2OS cells, the level of Pol I transcription was restored by overexpression of shRNA-resistant mouse Myo1c. Moreover, we found Myo1c interacting with Pol II. The ratio between Myo1c and NM1 proteins were similar in the nucleus and deletion of NM1 did not cause any compensatory overexpression of Myo1c protein. We observed that Myo1c can replace NM1 in its nuclear functions. Amount of both proteins is nearly equal and NM1 knock-out does not cause any compensatory overexpression of Myo1c. We therefore suggest that both isoforms can substitute each other in nuclear processes.

Behavioral characterization of CD36 knockout mice with SHIRPA primary screen.

Science.gov (United States)

Zhang, Shuxiao; Wang, Wei; Li, Juan; Cheng, Ke; Zhou, Jingjing; Zhu, Dan; Yang, Deyu; Liang, Zihong; Fang, Liang; Liao, Li; Xie, Peng

2016-02-15

CD36 is a member of the class B scavenger receptor family of cell surface proteins, which plays a major role in fatty acid, glucose and lipid metabolism. Besides, CD36 functions as a microglial surface receptor for amyloid beta peptide. Regarding this, we suggest CD36 might also contribute to neuropsychiatric disease. The aim of this study was to achieve a behavioral phenotype of CD36 knockout (CD36(-/-)) mice. We characterized the behavior of CD36(-/-) mice and C57BL/6J mice by subjecting them to a series of tests, which include SHIRPA primary behavioral screen test, 1% sucrose preference test, elevated plus-maze test, open-field test and forced swimming test. The results showed that CD36(-/-) mice traversed more squares, emitted more defecation, exhibited higher tail elevation and had more aggressive behaviors than C57BL/6J mice. The CD36(-/-) mice spent more time and traveled longer distance in periphery zone in the open-field test. Meanwhile, the numbers that CD36(-/-) mice entered in the open arms of elevated plus-maze were reduced. These findings suggest that CD36(-/-) mice present an anxious phenotype and might be involved in neuropsychiatric disorders. Copyright © 2015. Published by Elsevier B.V.

Exacerbation of spontaneous autoimmune nephritis following regulatory T cell depletion in B cell lymphoma 2-interacting mediator knock-out mice.

Science.gov (United States)

Wang, Y M; Zhang, G Y; Wang, Y; Hu, M; Zhou, J J; Sawyer, A; Cao, Q; Wang, Y; Zheng, G; Lee, V W S; Harris, D C H; Alexander, S I

2017-05-01

Regulatory T cells (T regs ) have been recognized as central mediators for maintaining peripheral tolerance and limiting autoimmune diseases. The loss of T regs or their function has been associated with exacerbation of autoimmune disease. However, the temporary loss of T regs in the chronic spontaneous disease model has not been investigated. In this study, we evaluated the role of T regs in a novel chronic spontaneous glomerulonephritis model of B cell lymphoma 2-interacting mediator (Bim) knock-out mice by transient depleting T regs . Bim is a pro-apoptotic member of the B cell lymphoma 2 (Bcl-2) family. Bim knock-out (Bim -/- ) mice fail to delete autoreactive T cells in thymus, leading to chronic spontaneous autoimmune kidney disease. We found that T reg depletion in Bim -/- mice exacerbated the kidney injury with increased proteinuria, impaired kidney function, weight loss and greater histological injury compared with wild-type mice. There was a significant increase in interstitial infiltrate of inflammatory cells, antibody deposition and tubular damage. Furthermore, the serum levels of cytokines interleukin (IL)-2, IL-4, IL-6, IL-10, IL-17α, interferon (IFN)-γ and tumour necrosis factor (TNF)-α were increased significantly after T reg depletion in Bim -/- mice. This study demonstrates that transient depletion of T regs leads to enhanced self-reactive T effector cell function followed by exacerbation of kidney disease in the chronic spontaneous kidney disease model of Bim-deficient mice. © 2017 British Society for Immunology.

Calibration of reconstruction parameters in atom probe tomography using a single crystallographic orientation

International Nuclear Information System (INIS)

Suram, Santosh K.; Rajan, Krishna

2013-01-01

The purpose of this work is to develop a methodology to estimate the APT reconstruction parameters when limited crystallographic information is available. Reliable spatial scaling of APT data currently requires identification of multiple crystallographic poles from the field desorption image for estimating the reconstruction parameters. This requirement limits the capacity of accurately reconstructing APT data for certain complex systems, such as highly alloyed systems and nanostructured materials wherein more than one pole is usually not observed within one grain. To overcome this limitation, we develop a quantitative methodology for calibrating the reconstruction parameters in an APT dataset by ensuring accurate inter-planar spacing and optimizing the curvature correction for the atomic planes corresponding to a single crystallographic orientation. We validate our approach on an aluminum dataset and further illustrate its capabilities by computing geometric reconstruction parameters for W and Al–Mg–Sc datasets. - Highlights: ► Quantitative approach is developed to accurately reconstruct APT data. ► Curvature of atomic planes in APT data is used to calibrate the reconstruction. ► APT reconstruction parameters are determined from a single crystallographic axis. ► Quantitative approach is demonstrated on W, Al and Al–Mg–Sc systems. ► Accurate APT reconstruction of complex materials is now possible

A single source microwave photonic filter using a novel single-mode fiber to multimode fiber coupling technique.

Science.gov (United States)

Chang, John; Fok, Mable P; Meister, James; Prucnal, Paul R

2013-03-11

In this paper we present a fully tunable and reconfigurable single-laser multi-tap microwave photonic FIR filter that utilizes a special SM-to-MM combiner to sum the taps. The filter requires only a single laser source for all the taps and a passive component, a SM-to-MM combiner, for incoherent summing of signal. The SM-to-MM combiner does not produce optical interference during signal merging and is phase-insensitive. We experimentally demonstrate an eight-tap filter with both positive and negative programmable coefficients with excellent correspondence between predicted and measured values. The magnitude response shows a clean and accurate function across the entire bandwidth, and proves successful operation of the FIR filter using a SM-to-MM combiner.

Real-time single-molecule observation of rolling-circle DNA replication

NARCIS (Netherlands)

Tanner, Nathan A.; Loparo, Joseph J.; Hamdan, Samir M.; Jergic, Slobodan; Dixon, Nicholas E.; Oijen, Antoine M. van

2009-01-01

We present a simple technique for visualizing replication of individual DNA molecules in real time. By attaching a rolling-circle substrate to a TIRF microscope-mounted flow chamber, we are able to monitor the progression of single-DNA synthesis events and accurately measure rates and processivities

Development of a keV single-ion-implanter for nanofabrication

International Nuclear Information System (INIS)

Yang, C.; Jamieson, D.N.; Hopf, T.; Tamanyan, G.; Spizziri, P.; Pakes, C.; Andresen, S.E.; Hudson, F.; Gauja, E.; Dzurak, A.; Clark, R.G.

2005-01-01

Traditional methods of doping semiconductors have a difficulty meeting the demand for high precision doping due to large statistical fluctuations in the numbers of dopant atoms introduced in the ever shrinking volume in micro- and nano-electronics devices, especially when the fabrication process approaches the nanometre scale. The statistical fluctuations in doping semiconductors for the fabrication of devices with a very small feature size may lead to inconsistent and unreliable performance. This paper describes the adaptation of a commercial ion implanter into a single-ion-implantation system for the accurate delivery of dopants into a nanometre or micrometre area in a silicon substrate. All the implanted ions can be accurately counted with near 100% certainty through online detection using the silicon substrate itself as an ion detector. A variety of ion species including B + , N + , P + at the energy range of 10-15 keV can be delivered in the single ion implantation system. (author). 6 refs., 6 figs

Elastic Properties of Nucleic Acids by Single-Molecule Force Spectroscopy.

Science.gov (United States)

Camunas-Soler, Joan; Ribezzi-Crivellari, Marco; Ritort, Felix

2016-07-05

We review the current knowledge on the use of single-molecule force spectroscopy techniques to extrapolate the elastic properties of nucleic acids. We emphasize the lesser-known elastic properties of single-stranded DNA. We discuss the importance of accurately determining the elastic response in pulling experiments, and we review the simplest models used to rationalize the experimental data as well as the experimental approaches used to pull single-stranded DNA. Applications used to investigate DNA conformational transitions and secondary structure formation are also highlighted. Finally, we provide an overview of the effects of salt and temperature and briefly discuss the effects of contour length and sequence dependence.

Hydrogen sulfide detection based on reflection: from a poison test approach of ancient China to single-cell accurate localization.

Science.gov (United States)

Kong, Hao; Ma, Zhuoran; Wang, Song; Gong, Xiaoyun; Zhang, Sichun; Zhang, Xinrong

2014-08-05

With the inspiration of an ancient Chinese poison test approach, we report a rapid hydrogen sulfide detection strategy in specific areas of live cells using silver needles with good spatial resolution of 2 × 2 μm(2). Besides the accurate-localization ability, this reflection-based strategy also has attractive merits of convenience and robust response when free pretreatment and short detection time are concerned. The success of endogenous H2S level evaluation in cellular cytoplasm and nuclear of human A549 cells promises the application potential of our strategy in scientific research and medical diagnosis.

The effect of insulin deficiency on tau and neurofilament in the insulin knockout mouse

International Nuclear Information System (INIS)

Schechter, Ruben; Beju, Delia; Miller, Kenneth E.

2005-01-01

Complications of diabetes mellitus within the nervous system are peripheral and central neuropathy. In peripheral neuropathy, defects in neurofilament and microtubules have been demonstrated. In this study, we examined the effects of insulin deficiency within the brain in insulin knockout mice (I(-/-)). The I(-/-) exhibited hyperphosphorylation of tau, at threonine 231, and neurofilament. In addition, we showed hyperphosphorylation of c-Jun N-terminal kinase (JNK) and glycogen synthase kinase 3 β (GSK-3 β) at serine 9. Extracellular signal-regulated kinase 1 (ERK 1) showed decrease in phosphorylation, whereas ERK 2 showed no changes. Ultrastructural examination demonstrated swollen mitochondria, endoplasmic reticulum, and Golgi apparatus, and dispersion of the nuclear chromatin. Microtubules showed decrease in the number of intermicrotubule bridges and neurofilament presented as bunches. Thus, lack of insulin brain stimulation induces JNK hyperphosphorylation followed by hyperphosphorylation of tau and neurofilament, and ultrastructural cellular damage, that over time may induce decrease in cognition and learning disabilities

The effect of insulin deficiency on tau and neurofilament in the insulin knockout mouse

Energy Technology Data Exchange (ETDEWEB)

Schechter, Ruben [William K. Warren Medical Research Institute, University of Oklahoma Medical Health Science Center, Tulsa, OK 74107 (United States); Department of Anatomy and Cell Biology, Oklahoma State University Center for Health Science, Tulsa, OK 74107 (United States); schechter@okstate edu, E-mail: ruben; Beju, Delia [William K. Warren Medical Research Institute, University of Oklahoma Medical Health Science Center, Tulsa, OK 74107 (United States); Department of Anatomy and Cell Biology, Oklahoma State University Center for Health Science, Tulsa, OK 74107 (United States); Miller, Kenneth E [Department of Anatomy and Cell Biology, Oklahoma State University Center for Health Science, Tulsa, OK 74107 (United States)

2005-09-09

Complications of diabetes mellitus within the nervous system are peripheral and central neuropathy. In peripheral neuropathy, defects in neurofilament and microtubules have been demonstrated. In this study, we examined the effects of insulin deficiency within the brain in insulin knockout mice (I(-/-)). The I(-/-) exhibited hyperphosphorylation of tau, at threonine 231, and neurofilament. In addition, we showed hyperphosphorylation of c-Jun N-terminal kinase (JNK) and glycogen synthase kinase 3 {beta} (GSK-3 {beta}) at serine 9. Extracellular signal-regulated kinase 1 (ERK 1) showed decrease in phosphorylation, whereas ERK 2 showed no changes. Ultrastructural examination demonstrated swollen mitochondria, endoplasmic reticulum, and Golgi apparatus, and dispersion of the nuclear chromatin. Microtubules showed decrease in the number of intermicrotubule bridges and neurofilament presented as bunches. Thus, lack of insulin brain stimulation induces JNK hyperphosphorylation followed by hyperphosphorylation of tau and neurofilament, and ultrastructural cellular damage, that over time may induce decrease in cognition and learning disabilities.

Disease Model Discovery from 3,328 Gene Knockouts by The International Mouse Phenotyping Consortium

Science.gov (United States)

Meehan, Terrence F.; Conte, Nathalie; West, David B.; Jacobsen, Julius O.; Mason, Jeremy; Warren, Jonathan; Chen, Chao-Kung; Tudose, Ilinca; Relac, Mike; Matthews, Peter; Karp, Natasha; Santos, Luis; Fiegel, Tanja; Ring, Natalie; Westerberg, Henrik; Greenaway, Simon; Sneddon, Duncan; Morgan, Hugh; Codner, Gemma F; Stewart, Michelle E; Brown, James; Horner, Neil; Haendel, Melissa; Washington, Nicole; Mungall, Christopher J.; Reynolds, Corey L; Gallegos, Juan; Gailus-Durner, Valerie; Sorg, Tania; Pavlovic, Guillaume; Bower, Lynette R; Moore, Mark; Morse, Iva; Gao, Xiang; Tocchini-Valentini, Glauco P; Obata, Yuichi; Cho, Soo Young; Seong, Je Kyung; Seavitt, John; Beaudet, Arthur L.; Dickinson, Mary E.; Herault, Yann; Wurst, Wolfgang; de Angelis, Martin Hrabe; Lloyd, K.C. Kent; Flenniken, Ann M; Nutter, Lauryl MJ; Newbigging, Susan; McKerlie, Colin; Justice, Monica J.; Murray, Stephen A.; Svenson, Karen L.; Braun, Robert E.; White, Jacqueline K.; Bradley, Allan; Flicek, Paul; Wells, Sara; Skarnes, William C.; Adams, David J.; Parkinson, Helen; Mallon, Ann-Marie; Brown, Steve D.M.; Smedley, Damian

2017-01-01

Although next generation sequencing has revolutionised the ability to associate variants with human diseases, diagnostic rates and development of new therapies are still limited by our lack of knowledge of function and pathobiological mechanism for most genes. To address this challenge, the International Mouse Phenotyping Consortium (IMPC) is creating a genome- and phenome-wide catalogue of gene function by characterizing new knockout mouse strains across diverse biological systems through a broad set of standardised phenotyping tests, with all mice made readily available to the biomedical community. Analysing the first 3328 genes reveals models for 360 diseases including the first for type C Bernard-Soulier, Bardet-Biedl-5 and Gordon Holmes syndromes. 90% of our phenotype annotations are novel, providing the first functional evidence for 1092 genes and candidates in unsolved diseases such as Arrhythmogenic Right Ventricular Dysplasia 3. Finally, we describe our role in variant functional validation with the 100,000 Genomes and other projects. PMID:28650483

Chemical Transport Knockout for Oxidized Vitamin C, Dehydroascorbic Acid, Reveals Its Functions in vivo

Directory of Open Access Journals (Sweden)

Hongbin Tu

2017-09-01

Full Text Available Despite its transport by glucose transporters (GLUTs in vitro, it is unknown whether dehydroascorbic acid (oxidized vitamin C, DHA has any in vivo function. To investigate, we created a chemical transport knockout model using the vitamin C analog 6-bromo-ascorbate. This analog is transported on sodium-dependent vitamin C transporters but its oxidized form, 6-bromo-dehydroascorbic acid, is not transported by GLUTs. Mice (gulo−/− unable to synthesize ascorbate (vitamin C were raised on 6-bromo-ascorbate. Despite normal survival, centrifugation of blood produced hemolysis secondary to near absence of red blood cell (RBC ascorbate/6-bromo-ascorbate. Key findings with clinical implications were that RBCs in vitro transported dehydroascorbic acid but not bromo-dehydroascorbic acid; RBC ascorbate in vivo was obtained only via DHA transport; ascorbate via DHA transport in vivo was necessary for RBC structural integrity; and internal RBC ascorbate was essential to maintain ascorbate plasma concentrations in vitro/in vivo.

Interactions of the opioid and cannabinoid systems in reward: Insights from knockout studies

Directory of Open Access Journals (Sweden)

Katia eBefort

2015-02-01

Full Text Available The opioid system consists of three receptors, mu, delta, and kappa, which are activated by endogenous opioid peptides (enkephalins, endorphins and dynorphins. The endogenous cannabinoid system comprises lipid neuromodulators (endocannabinoids, enzymes for their synthesis and their degradation and two well-characterized receptors, cannabinoid receptors CB1 and CB2. These systems play a major role in the control of pain as well as in mood regulation, reward processing and the development of addiction. Both opioid and cannabinoid receptors are coupled to G proteins and are expressed throughout the brain reinforcement circuitry. Extending classical pharmacology, research using genetically modified mice has provided important progress in the identification of the specific contribution of each component of these endogenous systems in vivo on reward process. This review will summarize available genetic tools and our present knowledge on the consequences of gene knockout on reinforced behaviors in both systems, with a focus on their potential interactions. A better understanding of opioid-cannabinoid interactions may provide novel strategies for therapies in addicted individuals.

Global analysis of gene expression in the developing brain of Gtf2ird1 knockout mice.

Directory of Open Access Journals (Sweden)

Jennifer O'Leary

Full Text Available Williams-Beuren Syndrome (WBS is a neurodevelopmental disorder caused by a hemizygous deletion of a 1.5 Mb region on chromosome 7q11.23 encompassing 26 genes. One of these genes, GTF2IRD1, codes for a putative transcription factor that is expressed throughout the brain during development. Genotype-phenotype studies in patients with atypical deletions of 7q11.23 implicate this gene in the neurological features of WBS, and Gtf2ird1 knockout mice show reduced innate fear and increased sociability, consistent with features of WBS. Multiple studies have identified in vitro target genes of GTF2IRD1, but we sought to identify in vivo targets in the mouse brain.We performed the first in vivo microarray screen for transcriptional targets of Gtf2ird1 in brain tissue from Gtf2ird1 knockout and wildtype mice at embryonic day 15.5 and at birth. Changes in gene expression in the mutant mice were moderate (0.5 to 2.5 fold and of candidate genes with altered expression verified using real-time PCR, most were located on chromosome 5, within 10 Mb of Gtf2ird1. siRNA knock-down of Gtf2ird1 in two mouse neuronal cell lines failed to identify changes in expression of any of the genes identified from the microarray and subsequent analysis showed that differences in expression of genes on chromosome 5 were the result of retention of that chromosome region from the targeted embryonic stem cell line, and so were dependent upon strain rather than Gtf2ird1 genotype. In addition, specific analysis of genes previously identified as direct in vitro targets of GTF2IRD1 failed to show altered expression.We have been unable to identify any in vivo neuronal targets of GTF2IRD1 through genome-wide expression analysis, despite widespread and robust expression of this protein in the developing rodent brain.

Arterial injury promotes medial chondrogenesis in Sm22 knockout mice.

Science.gov (United States)

Shen, Jianbin; Yang, Maozhou; Jiang, Hong; Ju, Donghong; Zheng, Jian-Pu; Xu, Zhonghui; Liao, Tang-Dong; Li, Li

2011-04-01

Expression of SM22 (also known as SM22alpha and transgelin), a vascular smooth muscle cells (VSMCs) marker, is down-regulated in arterial diseases involving medial osteochondrogenesis. We investigated the effect of SM22 deficiency in a mouse artery injury model to determine the role of SM22 in arterial chondrogenesis. Sm22 knockout (Sm22(-/-)) mice developed prominent medial chondrogenesis 2 weeks after carotid denudation as evidenced by the enhanced expression of chondrogenic markers including type II collagen, aggrecan, osteopontin, bone morphogenetic protein 2, and SRY-box containing gene 9 (SOX9). This was concomitant with suppression of VSMC key transcription factor myocardin and of VSMC markers such as SM α-actin and myosin heavy chain. The conversion tendency from myogenesis to chondrogenesis was also observed in primary Sm22(-/-) VSMCs and in a VSMC line after Sm22 knockdown: SM22 deficiency altered VSMC morphology with compromised stress fibre formation and increased actin dynamics. Meanwhile, the expression level of Sox9 mRNA was up-regulated while the mRNA levels of myocardin and VSMC markers were down-regulated, indicating a pro-chondrogenic transcriptional switch in SM22-deficient VSMCs. Furthermore, the increased expression of SOX9 was mediated by enhanced reactive oxygen species production and nuclear factor-κB pathway activation. These findings suggest that disruption of SM22 alters the actin cytoskeleton and promotes chondrogenic conversion of VSMCs.

Genotypic differences in intruder-evoked immediate early gene activation in male, but not female, vasopressin 1b receptor knockout mice.

Science.gov (United States)

Witchey, Shannah K; Stevenson, Erica L; Caldwell, Heather K

2016-11-24

The neuropeptide arginine vasopressin (Avp) modulates social behaviors via its two centrally expressed receptors, the Avp 1a receptor and the Avp 1b receptor (Avpr1b). Recent work suggests that, at least in mice, Avp signaling through Avpr1b within the CA2 region of the hippocampus is critical for normal aggressive behaviors and social recognition memory. However, this brain area is just one part of a larger neural circuit that is likely to be impacted in Avpr1b knockout (-/-) mice. To identify other brain areas that are affected by altered Avpr1b signaling, genotypic differences in immediate early gene activation, i.e. c-FOS and early growth response factor 1 (EGR-1), were quantified using immunocytochemistry following a single exposure to an intruder. In females, no genotypic differences in intruder-evoked c-FOS or EGR-1 immunoreactivity were observed in any of the brain areas measured. In males, while there were no intruder-evoked genotypic differences in c-FOS immunoreactivity, genotypic differences were observed in EGR-1 immunoreactivity within the ventral bed nucleus of the stria terminalis and the anterior hypothalamus; with Avpr1b -/- males having less EGR-1 immunoreactivity in these regions than controls. These data are the first to identify specific brain areas that may be a part of a neural circuit that includes Avpr1b-expressing cells in the CA2 region of the hippocampus. It is thought that this circuit, when working properly, plays a role in how an animal evaluates its social context.

Mixture models for single-cell assays with applications to vaccine studies

OpenAIRE

Finak, Greg; McDavid, Andrew; Chattopadhyay, Pratip; Dominguez, Maria; De Rosa, Steve; Roederer, Mario; Gottardo, Raphael

2013-01-01

Blood and tissue are composed of many functionally distinct cell subsets. In immunological studies, these can be measured accurately only using single-cell assays. The characterization of these small cell subsets is crucial to decipher system-level biological changes. For this reason, an increasing number of studies rely on assays that provide single-cell measurements of multiple genes and proteins from bulk cell samples. A common problem in the analysis of such data is to identify biomarkers...

Single-photon double and triple ionization of acetaldehyde (ethanal) studied by multi-electron coincidence spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Zagorodskikh, S. [Department of Physics and Astronomy, Uppsala University, Box 516, SE-751 20 Uppsala (Sweden); Department of Physics, University of Gothenburg, Origovägen 6B, SE-412 96 Gothenburg (Sweden); Zhaunerchyk, V. [Department of Physics, University of Gothenburg, Origovägen 6B, SE-412 96 Gothenburg (Sweden); Department of Physics and Astronomy, Uppsala University, Box 516, SE-751 20 Uppsala (Sweden); Mucke, M. [Department of Physics and Astronomy, Uppsala University, Box 516, SE-751 20 Uppsala (Sweden); Eland, J.H.D. [Department of Chemistry, Physical and Theoretical Chemistry Laboratory, Oxford University, South Parks Road, Oxford OX1 3QZ (United Kingdom); Department of Physics, University of Gothenburg, Origovägen 6B, SE-412 96 Gothenburg (Sweden); Department of Physics and Astronomy, Uppsala University, Box 516, SE-751 20 Uppsala (Sweden); Squibb, R.J. [Department of Physics, University of Gothenburg, Origovägen 6B, SE-412 96 Gothenburg (Sweden); Department of Physics and Astronomy, Uppsala University, Box 516, SE-751 20 Uppsala (Sweden); Karlsson, L. [Department of Physics and Astronomy, Uppsala University, Box 516, SE-751 20 Uppsala (Sweden); Linusson, P. [Department of Physics, Stockholm University, AlbaNova University Center, SE-106 91 Stockholm (Sweden); Feifel, R., E-mail: raimund.feifel@gu.se [Department of Physics, University of Gothenburg, Origovägen 6B, SE-412 96 Gothenburg (Sweden); Department of Physics and Astronomy, Uppsala University, Box 516, SE-751 20 Uppsala (Sweden)

2015-12-16

Highlights: • The first ever valence double ionization spectrum of acetaldehyde is reported. • The first ever site-selectively extracted Auger spectra of acetaldehyde are reported. • The first ever Auger spectra of acetaldehyde involving shake-up states are reported. • The first ever triple ionization spectra of acetaldehyde are reported. • The first ever energy sharing of electron pairs emitted by acetaldehyde is presented. - Abstract: Single-photon multiple ionization processes of acetaldehyde (ethanal) have been experimentally investigated by utilizing a multi-particle coincidence technique based on the time-of-flight magnetic bottle principle, in combination with either a synchrotron radiation source or a pulsed helium discharge lamp. The processes investigated include double and triple ionization in the valence region as well as single and double Auger decay of core-ionized acetaldehyde. The latter are studied site-selectively for chemically different carbon core vacancies, scrutinizing early theoretical predictions specifically made for the case of acetaldehyde. Moreover, Auger processes in shake-up and core-valence ionized states are investigated. In the cases where the processes involve simultaneous emission of two electrons, the distributions of the energy sharing are presented, emphasizing either the knock-out or shake-off mechanism.

Accurate detection of carcinoma cells by use of a cell microarray chip.

Directory of Open Access Journals (Sweden)

Shohei Yamamura

Full Text Available BACKGROUND: Accurate detection and analysis of circulating tumor cells plays an important role in the diagnosis and treatment of metastatic cancer treatment. METHODS AND FINDINGS: A cell microarray chip was used to detect spiked carcinoma cells among leukocytes. The chip, with 20,944 microchambers (105 µm width and 50 µm depth, was made from polystyrene; and the formation of monolayers of leukocytes in the microchambers was observed. Cultured human T lymphoblastoid leukemia (CCRF-CEM cells were used to examine the potential of the cell microarray chip for the detection of spiked carcinoma cells. A T lymphoblastoid leukemia suspension was dispersed on the chip surface, followed by 15 min standing to allow the leukocytes to settle down into the microchambers. Approximately 29 leukocytes were found in each microchamber when about 600,000 leukocytes in total were dispersed onto a cell microarray chip. Similarly, when leukocytes isolated from human whole blood were used, approximately 89 leukocytes entered each microchamber when about 1,800,000 leukocytes in total were placed onto the cell microarray chip. After washing the chip surface, PE-labeled anti-cytokeratin monoclonal antibody and APC-labeled anti-CD326 (EpCAM monoclonal antibody solution were dispersed onto the chip surface and allowed to react for 15 min; and then a microarray scanner was employed to detect any fluorescence-positive cells within 20 min. In the experiments using spiked carcinoma cells (NCI-H1650, 0.01 to 0.0001%, accurate detection of carcinoma cells was achieved with PE-labeled anti-cytokeratin monoclonal antibody. Furthermore, verification of carcinoma cells in the microchambers was performed by double staining with the above monoclonal antibodies. CONCLUSION: The potential application of the cell microarray chip for the detection of CTCs was shown, thus demonstrating accurate detection by double staining for cytokeratin and EpCAM at the single carcinoma cell level.

A facile electrode preparation method for accurate electrochemical measurements of double-side-coated electrode from commercial Li-ion batteries

Science.gov (United States)

Zhou, Ge; Wang, Qiyu; Wang, Shuo; Ling, Shigang; Zheng, Jieyun; Yu, Xiqian; Li, Hong

2018-04-01

The post mortem electrochemical analysis, including charge-discharge and electrochemical impedance spectroscopy (EIS) measurements, are critical steps for revealing the failure mechanisms of commercial lithium-ion batteries (LIBs). These post measurements usually require the reassembling of coin-cell with electrode which is often double-side-coated in commercial LIBs. It is difficult to use such double-side-coated electrode to perform accurate electrochemical measurements because the back side of the electrode is coated with active materials, rather than single-side-coated electrode that is often used in coin-cell measurements. In this study, we report a facile tape-covering sample preparation method, which can effectively suppress the influence of back side of the double-side-coated electrodes on capacity and EIS measurements in coin-cells. By tape-covering the unwanted side, the areal capacity of the desired investigated side of the electrode has been accurately measured with an experimental error of about 0.5% at various current densities, and accurate EIS measurements and analysis have been conducted as well.

Phosphocreatine kinetics at the onset of contractions in skeletal muscle of MM creatine kinase knockout mice

Science.gov (United States)

Roman, Brian B.; Meyer, Ronald A.; Wiseman, Robert W.

2002-01-01

Phosphocreatine (PCr) depletion during isometric twitch stimulation at 5 Hz was measured by (31)P-NMR spectroscopy in gastrocnemius muscles of pentobarbital-anesthetized MM creatine kinase knockout (MMKO) vs. wild-type C57B (WT) mice. PCr depletion after 2 s of stimulation, estimated from the difference between spectra gated to times 200 ms and 140 s after 2-s bursts of contractions, was 2.2 +/- 0.6% of initial PCr in MMKO muscle vs. 9.7 +/- 1.6% in WT muscles (mean +/- SE, n = 7, P muscle after 2 s only if ADP-stimulated oxidative phosphorylation was included in the model. Taken together, the results suggest that cytoplasmic ADP more rapidly increases and oxidative phosphorylation is more rapidly activated at the onset of contractions in MMKO compared with WT muscles.

Fast and accurate focusing analysis of large photon sieve using pinhole ring diffraction model.

Science.gov (United States)

Liu, Tao; Zhang, Xin; Wang, Lingjie; Wu, Yanxiong; Zhang, Jizhen; Qu, Hemeng

2015-06-10

In this paper, we developed a pinhole ring diffraction model for the focusing analysis of a large photon sieve. Instead of analyzing individual pinholes, we discuss the focusing of all of the pinholes in a single ring. An explicit equation for the diffracted field of individual pinhole ring has been proposed. We investigated the validity range of this generalized model and analytically describe the sufficient conditions for the validity of this pinhole ring diffraction model. A practical example and investigation reveals the high accuracy of the pinhole ring diffraction model. This simulation method could be used for fast and accurate focusing analysis of a large photon sieve.

Fingerprint-Inspired Flexible Tactile Sensor for Accurately Discerning Surface Texture.

Science.gov (United States)

Cao, Yudong; Li, Tie; Gu, Yang; Luo, Hui; Wang, Shuqi; Zhang, Ting

2018-04-01

Inspired by the epidermal-dermal and outer microstructures of the human fingerprint, a novel flexible sensor device is designed to improve haptic perception and surface texture recognition, which is consisted of single-walled carbon nanotubes, polyethylene, and polydimethylsiloxane with interlocked and outer micropyramid arrays. The sensor shows high pressure sensitivity (-3.26 kPa -1 in the pressure range of 0-300 Pa), and it can detect the shear force changes induced by the dynamic interaction between the outer micropyramid structure on the sensor and the tested material surface, and the minimum dimension of the microstripe that can be discerned is as low as 15 µm × 15 µm (interval × width). To demonstrate the texture discrimination capability, the sensors are tested for accurately discerning various surface textures, such as the textures of different fabrics, Braille characters, the inverted pyramid patterns, which will have great potential in robot skins and haptic perception, etc. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Spatial reconstruction of single-cell gene expression

Science.gov (United States)

Satija, Rahul; Farrell, Jeffrey A.; Gennert, David; Schier, Alexander F.; Regev, Aviv

2015-01-01

Spatial localization is a key determinant of cellular fate and behavior, but spatial RNA assays traditionally rely on staining for a limited number of RNA species. In contrast, single-cell RNA-seq allows for deep profiling of cellular gene expression, but established methods separate cells from their native spatial context. Here we present Seurat, a computational strategy to infer cellular localization by integrating single-cell RNA-seq data with in situ RNA patterns. We applied Seurat to spatially map 851 single cells from dissociated zebrafish (Danio rerio) embryos, inferring a transcriptome-wide map of spatial patterning. We confirmed Seurat’s accuracy using several experimental approaches, and used it to identify a set of archetypal expression patterns and spatial markers. Additionally, Seurat correctly localizes rare subpopulations, accurately mapping both spatially restricted and scattered groups. Seurat will be applicable to mapping cellular localization within complex patterned tissues in diverse systems. PMID:25867923

Accurate and Simple Calibration of DLP Projector Systems

DEFF Research Database (Denmark)

Wilm, Jakob; Olesen, Oline Vinter; Larsen, Rasmus

2014-01-01

does not rely on an initial camera calibration, and so does not carry over the error into projector calibration. A radial interpolation scheme is used to convert features coordinates into projector space, thereby allowing for a very accurate procedure. This allows for highly accurate determination...

Apolipoprotein E-knockout mice on high-fat diet show autoimmune injury on kidney and aorta

Energy Technology Data Exchange (ETDEWEB)

Wang, Yuehai [Cardiovascular Department, Liaocheng People’s Hospital of Shandong University, Liaocheng, Shandong 252000 (China); Cardiovascular Department, The Second Clinical Medical College of Fujian Medical University, Quanzhou, Fujian 362000 (China); Lu, Huixia [The Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and Chinese Ministry of Health, Shandong University Qilu Hospital, Jinan, Shandong 250012 (China); Huang, Ziyang, E-mail: huangziyang666@126.com [Cardiovascular Department, The Second Clinical Medical College of Fujian Medical University, Quanzhou, Fujian 362000 (China); Lin, Huili [Cardiovascular Department, The Second Clinical Medical College of Fujian Medical University, Quanzhou, Fujian 362000 (China); Lei, Zhenmin [Department of OB/GYN, University of Louisville School of Medicine, Louisville, KY 40292 (United States); Chen, Xiaoqing [Department of Rheumatism and Immunology, The Second Clinical Medical College of Fujian Medical University, Quanzhou, Fujian 362000 (China); Tang, Mengxiong; Gao, Fei; Dong, Mei [The Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and Chinese Ministry of Health, Shandong University Qilu Hospital, Jinan, Shandong 250012 (China); Li, Rongda [Department of Rheumatism and Immunology, The Second Clinical Medical College of Fujian Medical University, Quanzhou, Fujian 362000 (China); Lin, Ling, E-mail: qzlinl@163.com [Department of Rheumatism and Immunology, The Second Clinical Medical College of Fujian Medical University, Quanzhou, Fujian 362000 (China)

2014-07-18

Highlights: • Titers of ANA and anti-dsDNA antibodies were similar in ApoE{sup −/−} and Fas{sup −/−} mice. • The spleen weights and glomerular areas were similar in ApoE{sup −/−} and Fas{sup −/−} mice. • Expressions of IgG and C3 in glomeruli were similar in ApoE{sup −/−} and Fas{sup −/−} mice. • IgG, C3 and macrophage infiltration in aortic plaques were found in ApoE{sup −/−} mice. - Abstract: Background: Apolipoprotein E-knockout (ApoE{sup −/−}) mice is a classic model of atherosclerosis. We have found that ApoE{sup −/−} mice showed splenomegaly, higher titers of serum anti-nuclear antibody (ANA) and anti-dsDNA antibody compared with C57B6/L (B6) mice. However, whether ApoE{sup −/−} mice show autoimmune injury remains unclear. Methods and results: Six females and six males in each group, ApoE{sup −/−}, Fas{sup −/−} and B6 mice, were used in this study. The titers of serum ANA, anti-dsDNA antibody and creatinine and urine protein were measured by ELISA after 4 months of high-fat diet. The spleen weight and the glomerular area were determined. The expressions of IgG, C3 and macrophage in kidney and atherosclerotic plaque were detected by immunostaining followed by morphometric analysis. Similar to the characteristics of Fas{sup −/−} mice, a model of systemic lupus erythematosus (SLE), ApoE{sup −/−} mice, especially female, displayed significant increases of spleen weight and glomerular area when compared to B6 mice. Also, elevated titers of serum ANA, anti-dsDNA antibody and creatinine and urine protein. Moreover, the expressions of IgG, C3 and macrophage in glomeruli and aortic plaques were found in ApoE{sup −/−} mice. In addition, the IgG and C3 expressions in glomeruli and plaques significantly increased (or a trend of increase) in female ApoE{sup −/−} mice compared with males. Conclusions: Apolipoprotein E-knockout mice on high-fat diet show autoimmune injury on kidney and aorta.

Apolipoprotein E-knockout mice on high-fat diet show autoimmune injury on kidney and aorta

International Nuclear Information System (INIS)

Wang, Yuehai; Lu, Huixia; Huang, Ziyang; Lin, Huili; Lei, Zhenmin; Chen, Xiaoqing; Tang, Mengxiong; Gao, Fei; Dong, Mei; Li, Rongda; Lin, Ling

2014-01-01

Highlights: • Titers of ANA and anti-dsDNA antibodies were similar in ApoE −/− and Fas −/− mice. • The spleen weights and glomerular areas were similar in ApoE −/− and Fas −/− mice. • Expressions of IgG and C3 in glomeruli were similar in ApoE −/− and Fas −/− mice. • IgG, C3 and macrophage infiltration in aortic plaques were found in ApoE −/− mice. - Abstract: Background: Apolipoprotein E-knockout (ApoE −/− ) mice is a classic model of atherosclerosis. We have found that ApoE −/− mice showed splenomegaly, higher titers of serum anti-nuclear antibody (ANA) and anti-dsDNA antibody compared with C57B6/L (B6) mice. However, whether ApoE −/− mice show autoimmune injury remains unclear. Methods and results: Six females and six males in each group, ApoE −/− , Fas −/− and B6 mice, were used in this study. The titers of serum ANA, anti-dsDNA antibody and creatinine and urine protein were measured by ELISA after 4 months of high-fat diet. The spleen weight and the glomerular area were determined. The expressions of IgG, C3 and macrophage in kidney and atherosclerotic plaque were detected by immunostaining followed by morphometric analysis. Similar to the characteristics of Fas −/− mice, a model of systemic lupus erythematosus (SLE), ApoE −/− mice, especially female, displayed significant increases of spleen weight and glomerular area when compared to B6 mice. Also, elevated titers of serum ANA, anti-dsDNA antibody and creatinine and urine protein. Moreover, the expressions of IgG, C3 and macrophage in glomeruli and aortic plaques were found in ApoE −/− mice. In addition, the IgG and C3 expressions in glomeruli and plaques significantly increased (or a trend of increase) in female ApoE −/− mice compared with males. Conclusions: Apolipoprotein E-knockout mice on high-fat diet show autoimmune injury on kidney and aorta

Improved single particle potential for transport model simulations of nuclear reactions induced by rare isotope beams

International Nuclear Information System (INIS)

Xu Chang; Li Baoan

2010-01-01

Taking into account more accurately the isospin dependence of nucleon-nucleon interactions in the in-medium many-body force term of the Gogny effective interaction, new expressions for the single-nucleon potential and the symmetry energy are derived. Effects of both the spin (isospin) and the density dependence of nuclear effective interactions on the symmetry potential and the symmetry energy are examined. It is shown that they both play a crucial role in determining the symmetry potential and the symmetry energy at suprasaturation densities. The improved single-nucleon potential will be useful for more accurate simulation of nuclear reactions induced by rare-isotope beams within transport models.

Diacylglycerol lipase a knockout mice demonstrate metabolic and behavioral phenotypes similar to those of cannabinoid receptor 1 knockout mice

Directory of Open Access Journals (Sweden)

David R Powell

2015-06-01

Full Text Available After creating >4650 knockouts (KOs of independent mouse genes, we screened them by high-throughput phenotyping and found that cannabinoid receptor 1 (Cnr1 KO mice had the same lean phenotype published by others. We asked if our KOs of DAG lipase a or b (Dagla or Daglb, which catalyze biosynthesis of the endocannabinoid (EC 2-Arachidonoylglycerol (2-AG, or Napepld, which catalyzes biosynthesis of the EC anandamide, shared the lean phenotype of Cnr1 KO mice. We found that Dagla KO mice, but not Daglb or Napepld KO mice, were among the leanest of 3651 chow-fed KO lines screened. In confirmatory studies, chow- or high fat diet-fed Dagla and Cnr1 KO mice were leaner than wild type (WT littermates; when data from multiple cohorts of adult mice were combined, body fat was 47% and 45% lower in Dagla and Cnr1 KO mice, respectively, relative to WT values. In contrast, neither Daglb nor Napepld KO mice were lean. Weanling Dagla KO mice ate less than WT mice and had body weight similar to pair-fed WT mice, and adult Dagla KO mice had normal activity and VO2 levels, similar to Cnr1 KO mice. Our Dagla and Cnr1 KO mice also had low fasting insulin, triglyceride and total cholesterol levels, and after a glucose challenge had normal glucose but very low insulin levels. Dagla and Cnr1 KO mice also showed similar responses to a battery of behavioral tests. These data suggest: 1 the lean phenotype of young Dagla and Cnr1 KO mice is mainly due to hypophagia; 2 in pathways where ECs signal through Cnr1 to regulate food intake and other metabolic and behavioral phenotypes observed in Cnr1 KO mice, Dagla alone provides the 2-AG that serves as the EC signal; and 3 small molecule Dagla inhibitors with a pharmacokinetic profile similar to that of Cnr1 inverse agonists are likely to mirror the ability of these Cnr1 inverse agonists to lower body weight and improve glycemic control in obese patients with type 2 diabetes, but may also induce undesirable neuropsychiatric

An accurate, fast, and scalable solver for high-frequency wave propagation

Science.gov (United States)

Zepeda-Núñez, L.; Taus, M.; Hewett, R.; Demanet, L.

2017-12-01

In many science and engineering applications, solving time-harmonic high-frequency wave propagation problems quickly and accurately is of paramount importance. For example, in geophysics, particularly in oil exploration, such problems can be the forward problem in an iterative process for solving the inverse problem of subsurface inversion. It is important to solve these wave propagation problems accurately in order to efficiently obtain meaningful solutions of the inverse problems: low order forward modeling can hinder convergence. Additionally, due to the volume of data and the iterative nature of most optimization algorithms, the forward problem must be solved many times. Therefore, a fast solver is necessary to make solving the inverse problem feasible. For time-harmonic high-frequency wave propagation, obtaining both speed and accuracy is historically challenging. Recently, there have been many advances in the development of fast solvers for such problems, including methods which have linear complexity with respect to the number of degrees of freedom. While most methods scale optimally only in the context of low-order discretizations and smooth wave speed distributions, the method of polarized traces has been shown to retain optimal scaling for high-order discretizations, such as hybridizable discontinuous Galerkin methods and for highly heterogeneous (and even discontinuous) wave speeds. The resulting fast and accurate solver is consequently highly attractive for geophysical applications. To date, this method relies on a layered domain decomposition together with a preconditioner applied in a sweeping fashion, which has limited straight-forward parallelization. In this work, we introduce a new version of the method of polarized traces which reveals more parallel structure than previous versions while preserving all of its other advantages. We achieve this by further decomposing each layer and applying the preconditioner to these new components separately and

Effects of PPARs agonists on cardiac metabolism in littermate and cardiomyocyte-specific PPAR-γ-knockout (CM-PGKO mice.

Directory of Open Access Journals (Sweden)

Michelangela Barbieri

Full Text Available Understanding the molecular regulatory mechanisms controlling for myocardial lipid metabolism is of critical importance for the development of new therapeutic strategies for heart diseases. The role of PPARγ and thiazolidinediones in regulation of myocardial lipid metabolism is controversial. The aim of our study was to assess the role of PPARγ on myocardial lipid metabolism and function and differentiate local/from systemic actions of PPARs agonists using cardiomyocyte-specific PPARγ -knockout (CM-PGKO mice. To this aim, the effect of PPARγ, PPARγ/PPARα and PPARα agonists on cardiac function, intra-myocyte lipid accumulation and myocardial expression profile of genes and proteins, affecting lipid oxidation, uptake, synthesis, and storage (CD36, CPT1MIIA, AOX, FAS, SREBP1-c and ADPR was evaluated in cardiomyocyte-specific PPARγ-knockout (CM-PGKO and littermate control mice undergoing standard and high fat diet (HFD. At baseline, protein levels and mRNA expression of genes involved in lipid uptake, oxidation, synthesis, and accumulation of CM-PGKO mice were not significantly different from those of their littermate controls. At baseline, no difference in myocardial lipid content was found between CM-PGKO and littermate controls. In standard condition, pioglitazone and rosiglitazone do not affect myocardial metabolism while, fenofibrate treatment significantly increased CD36 and CPT1MIIA gene expression. In both CM-PGKO and control mice submitted to HFD, six weeks of treatment with rosiglitazone, fenofibrate and pioglitazone lowered myocardial lipid accumulation shifting myocardial substrate utilization towards greater contribution of glucose. In conclusion, at baseline, PPARγ does not play a crucial role in regulating cardiac metabolism in mice, probably due to its low myocardial expression. PPARs agonists, indirectly protect myocardium from lipotoxic damage likely reducing fatty acids delivery to the heart through the actions on adipose

Robust and Accurate Anomaly Detection in ECG Artifacts Using Time Series Motif Discovery

Science.gov (United States)

Sivaraks, Haemwaan

2015-01-01

Electrocardiogram (ECG) anomaly detection is an important technique for detecting dissimilar heartbeats which helps identify abnormal ECGs before the diagnosis process. Currently available ECG anomaly detection methods, ranging from academic research to commercial ECG machines, still suffer from a high false alarm rate because these methods are not able to differentiate ECG artifacts from real ECG signal, especially, in ECG artifacts that are similar to ECG signals in terms of shape and/or frequency. The problem leads to high vigilance for physicians and misinterpretation risk for nonspecialists. Therefore, this work proposes a novel anomaly detection technique that is highly robust and accurate in the presence of ECG artifacts which can effectively reduce the false alarm rate. Expert knowledge from cardiologists and motif discovery technique is utilized in our design. In addition, every step of the algorithm conforms to the interpretation of cardiologists. Our method can be utilized to both single-lead ECGs and multilead ECGs. Our experiment results on real ECG datasets are interpreted and evaluated by cardiologists. Our proposed algorithm can mostly achieve 100% of accuracy on detection (AoD), sensitivity, specificity, and positive predictive value with 0% false alarm rate. The results demonstrate that our proposed method is highly accurate and robust to artifacts, compared with competitive anomaly detection methods. PMID:25688284

Single Nucleobase Identification Using Biophysical Signatures from Nanoelectronic Quantum Tunneling.

Science.gov (United States)

Korshoj, Lee E; Afsari, Sepideh; Khan, Sajida; Chatterjee, Anushree; Nagpal, Prashant

2017-03-01

Nanoelectronic DNA sequencing can provide an important alternative to sequencing-by-synthesis by reducing sample preparation time, cost, and complexity as a high-throughput next-generation technique with accurate single-molecule identification. However, sample noise and signature overlap continue to prevent high-resolution and accurate sequencing results. Probing the molecular orbitals of chemically distinct DNA nucleobases offers a path for facile sequence identification, but molecular entropy (from nucleotide conformations) makes such identification difficult when relying only on the energies of lowest-unoccupied and highest-occupied molecular orbitals (LUMO and HOMO). Here, nine biophysical parameters are developed to better characterize molecular orbitals of individual nucleobases, intended for single-molecule DNA sequencing using quantum tunneling of charges. For this analysis, theoretical models for quantum tunneling are combined with transition voltage spectroscopy to obtain measurable parameters unique to the molecule within an electronic junction. Scanning tunneling spectroscopy is then used to measure these nine biophysical parameters for DNA nucleotides, and a modified machine learning algorithm identified nucleobases. The new parameters significantly improve base calling over merely using LUMO and HOMO frontier orbital energies. Furthermore, high accuracies for identifying DNA nucleobases were observed at different pH conditions. These results have significant implications for developing a robust and accurate high-throughput nanoelectronic DNA sequencing technique. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

When Is Network Lasso Accurate?

Directory of Open Access Journals (Sweden)

Alexander Jung

2018-01-01

Full Text Available The “least absolute shrinkage and selection operator” (Lasso method has been adapted recently for network-structured datasets. In particular, this network Lasso method allows to learn graph signals from a small number of noisy signal samples by using the total variation of a graph signal for regularization. While efficient and scalable implementations of the network Lasso are available, only little is known about the conditions on the underlying network structure which ensure network Lasso to be accurate. By leveraging concepts of compressed sensing, we address this gap and derive precise conditions on the underlying network topology and sampling set which guarantee the network Lasso for a particular loss function to deliver an accurate estimate of the entire underlying graph signal. We also quantify the error incurred by network Lasso in terms of two constants which reflect the connectivity of the sampled nodes.

The FKBP5 Gene Affects Alcohol Drinking in Knockout Mice and Is Implicated in Alcohol Drinking in Humans

Directory of Open Access Journals (Sweden)

Bin Qiu

2016-08-01

Full Text Available FKBP5 encodes FK506-binding protein 5, a glucocorticoid receptor (GR-binding protein implicated in various psychiatric disorders and alcohol withdrawal severity. The purpose of this study is to characterize alcohol preference and related phenotypes in Fkbp5 knockout (KO mice and to examine the role of FKBP5 in human alcohol consumption. The following experiments were performed to characterize Fkpb5 KO mice. (1 Fkbp5 KO and wild-type (WT EtOH consumption was tested using a two-bottle choice paradigm; (2 The EtOH elimination rate was measured after intraperitoneal (IP injection of 2.0 g/kg EtOH; (3 Blood alcohol concentration (BAC was measured after 3 h limited access of alcohol; (4 Brain region expression of Fkbp5 was identified using LacZ staining; (5 Baseline corticosterone (CORT was assessed. Additionally, two SNPs, rs1360780 (C/T and rs3800373 (T/G, were selected to study the association of FKBP5 with alcohol consumption in humans. Participants were college students (n = 1162 from 21–26 years of age with Chinese, Korean or Caucasian ethnicity. The results, compared to WT mice, for KO mice exhibited an increase in alcohol consumption that was not due to differences in taste sensitivity or alcohol metabolism. Higher BAC was found in KO mice after 3 h of EtOH access. Fkbp5 was highly expressed in brain regions involved in the regulation of the stress response, such as the hippocampus, amygdala, dorsal raphe and locus coeruleus. Both genotypes exhibited similar basal levels of plasma corticosterone (CORT. Finally, single nucleotide polymorphisms (SNPs in FKBP5 were found to be associated with alcohol drinking in humans. These results suggest that the association between FKBP5 and alcohol consumption is conserved in both mice and humans.

Generation of Newly Discovered Resistance Gene mcr-1 Knockout in Escherichia coli Using the CRISPR/Cas9 System.

Science.gov (United States)

Sun, Lichang; He, Tao; Zhang, Lili; Pang, Maoda; Zhang, Qiaoyan; Zhou, Yan; Bao, Hongduo; Wang, Ran

2017-07-28

The mcr-1 gene is a new "superbug" gene discoverd in China in 2016 that makes bacteria highly resistant to the last-resort class of antibiotics. The mcr-1 gene raised serious concern about its possible global dissemination and spread. Here, we report a potential anti-resistant strategy using the CRISPR/Cas9-mediated approach that can efficiently induce mcr-1 gene knockout in Escherichia coli . Our findings suggested that using the CRISPR/Cas9 system to knock out the resistance gene mcr-1 might be a potential anti-resistant strategy. Bovine myeloid antimicrobial peptide-27 could help deliver plasmid pCas::mcr targeting specific DNA sequences of the mcr-1 gene into microbial populations.

A homozygous Keap1-knockout human embryonic stem cell line generated using CRISPR/Cas9 mediates gene targeting

Directory of Open Access Journals (Sweden)

So-Jung Kim

2017-03-01

Full Text Available Kelch-like ECH-associated protein 1 (keap1 is a cysteine-rich protein that interacts with transcription factor Nrf2 in a redox-sensitive manner, leading to the degradation of Nrf2 (Kim et al., 2014a. Disruption of Keap1 results in the induction of Nrf2-related signaling pathways involving the expression of a set of anti-oxidant and anti-inflammatory genes. We generated biallelic mutants of the Keap1 gene using a CRISPR-Cas9 genome editing method in the H9 human embryonic stem cell (hESC. The Keap1 homozygous-knockout H9 cell line retained normal morphology, gene expression, and in vivo differentiation potential.

Controlling single-molecule junction conductance by molecular interactions

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Kitaguchi, Y.; Habuka, S.; Okuyama, H.; Hatta, S.; Aruga, T.; Frederiksen, T.; Paulsson, M.; Ueba, H.

2015-01-01

For the rational design of single-molecular electronic devices, it is essential to understand environmental effects on the electronic properties of a working molecule. Here we investigate the impact of molecular interactions on the single-molecule conductance by accurately positioning individual molecules on the electrode. To achieve reproducible and precise conductivity measurements, we utilize relatively weak π-bonding between a phenoxy molecule and a STM-tip to form and cleave one contact to the molecule. The anchoring to the other electrode is kept stable using a chalcogen atom with strong bonding to a Cu(110) substrate. These non-destructive measurements permit us to investigate the variation in single-molecule conductance under different but controlled environmental conditions. Combined with density functional theory calculations, we clarify the role of the electrostatic field in the environmental effect that influences the molecular level alignment. PMID:26135251

The nature and nurture of cell heterogeneity: accounting for macrophage gene-environment interactions with single-cell RNA-Seq.

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Wills, Quin F; Mellado-Gomez, Esther; Nolan, Rory; Warner, Damien; Sharma, Eshita; Broxholme, John; Wright, Benjamin; Lockstone, Helen; James, William; Lynch, Mark; Gonzales, Michael; West, Jay; Leyrat, Anne; Padilla-Parra, Sergi; Filippi, Sarah; Holmes, Chris; Moore, Michael D; Bowden, Rory

2017-01-07

Single-cell RNA-Seq can be a valuable and unbiased tool to dissect cellular heterogeneity, despite the transcriptome's limitations in describing higher functional phenotypes and protein events. Perhaps the most important shortfall with transcriptomic 'snapshots' of cell populations is that they risk being descriptive, only cataloging heterogeneity at one point in time, and without microenvironmental context. Studying the genetic ('nature') and environmental ('nurture') modifiers of heterogeneity, and how cell population dynamics unfold over time in response to these modifiers is key when studying highly plastic cells such as macrophages. We introduce the programmable Polaris™ microfluidic lab-on-chip for single-cell sequencing, which performs live-cell imaging while controlling for the culture microenvironment of each cell. Using gene-edited macrophages we demonstrate how previously unappreciated knockout effects of SAMHD1, such as an altered oxidative stress response, have a large paracrine signaling component. Furthermore, we demonstrate single-cell pathway enrichments for cell cycle arrest and APOBEC3G degradation, both associated with the oxidative stress response and altered proteostasis. Interestingly, SAMHD1 and APOBEC3G are both HIV-1 inhibitors ('restriction factors'), with no known co-regulation. As single-cell methods continue to mature, so will the ability to move beyond simple 'snapshots' of cell populations towards studying the determinants of population dynamics. By combining single-cell culture, live-cell imaging, and single-cell sequencing, we have demonstrated the ability to study cell phenotypes and microenvironmental influences. It's these microenvironmental components - ignored by standard single-cell workflows - that likely determine how macrophages, for example, react to inflammation and form treatment resistant HIV reservoirs.

Parkinsonian rest tremor can be detected accurately based on neuronal oscillations recorded from the subthalamic nucleus.

Science.gov (United States)

Hirschmann, J; Schoffelen, J M; Schnitzler, A; van Gerven, M A J

2017-10-01

To investigate the possibility of tremor detection based on deep brain activity. We re-analyzed recordings of local field potentials (LFPs) from the subthalamic nucleus in 10 PD patients (12 body sides) with spontaneously fluctuating rest tremor. Power in several frequency bands was estimated and used as input to Hidden Markov Models (HMMs) which classified short data segments as either tremor-free rest or rest tremor. HMMs were compared to direct threshold application to individual power features. Applying a threshold directly to band-limited power was insufficient for tremor detection (mean area under the curve [AUC] of receiver operating characteristic: 0.64, STD: 0.19). Multi-feature HMMs, in contrast, allowed for accurate detection (mean AUC: 0.82, STD: 0.15), using four power features obtained from a single contact pair. Within-patient training yielded better accuracy than across-patient training (0.84vs. 0.78, p=0.03), yet tremor could often be detected accurately with either approach. High frequency oscillations (>200Hz) were the best performing individual feature. LFP-based markers of tremor are robust enough to allow for accurate tremor detection in short data segments, provided that appropriate statistical models are used. LFP-based markers of tremor could be useful control signals for closed-loop deep brain stimulation. Copyright © 2017 International Federation of Clinical Neurophysiology. Published by Elsevier B.V. All rights reserved.

Single-atom reversible recording at room temperature

DEFF Research Database (Denmark)

Quaade, Ulrich; Stokbro, Kurt; Lin, Rong

2001-01-01

investigate two important aspects of using this single-atom switch as a memory device. First, the switching is electron stimulated, and through detailed modelling the switching probability per electron is accurately deduced. Second, we have investigated the possibilities for desorbing single hydrogen atoms...... to construct ordered arrays of switches to manufacture a memory device. Two desorption mechanisms have been considered: the well known electron-induced desorption at negative sample bias and a novel mechanism probably involving elastic deformation of the tip. For both mechanisms mechanical stability of the STM...... is of crucial importance. With our equipment it was possible to create a row of four switches in a controlled way.(Some figures in this article are in colour only in the electronic version)....

Antibodies directed against monomorphic and evolutionary conserved self epitopes may be generated in 'knock-out' mice. Development of monoclonal antibodies directed against monomorphic MHC class I determinants

DEFF Research Database (Denmark)

Claesson, M H; Endel, B; Ulrik, J

1994-01-01

Beta-2 microglobulin (beta 2m) gene 'knock-out' mice (C1D) were primed with purified H-2Kb and H-2Db molecules and spleen cells from immunized mice were used to generate monoclonal antibody secreting B-cell hybridomas. Approximately 0.2% of the Ig-secreting primary microcultures contained H-2b...

A Single Test Combining Blood Markers and Elastography is More Accurate Than Other Fibrosis Tests in the Main Causes of Chronic Liver Diseases.

Science.gov (United States)

Ducancelle, Alexandra; Leroy, Vincent; Vergniol, Julien; Sturm, Nathalie; Le Bail, Brigitte; Zarski, Jean Pierre; Nguyen Khac, Eric; Salmon, Dominique; de Ledinghen, Victor; Calès, Paul

2017-08-01

International guidelines suggest combining a blood test and liver stiffness measurement (LSM) to stage liver fibrosis in chronic hepatitis C (CHC) and non-alcoholic fatty liver disease (NAFLD). Therefore, we compared the accuracies of these tests between the main etiologies of chronic liver diseases. Overall, 1968 patients were included in 5 etiologies: CHC: 698, chronic hepatitis B: 152, human immunodeficiency virus/CHC: 628, NAFLD: 225, and alcoholic liver disease (ALD): 265. Sixteen tests [13 blood tests, LSM (Fibroscan), 2 combined: FibroMeters] were evaluated. References were Metavir staging and CHC etiology. Accuracy was evaluated mainly with the Obuchowski index (OI) and accessorily with area under the receiver operating characteristics (F≥2, F≥3, cirrhosis). OIs in CHC were: FibroMeters: 0.812, FibroMeters: 0.785 to 0.797, Fibrotest: 0.762, CirrhoMeters: 0.756 to 0.771, LSM: 0.754, Hepascore: 0.752, FibroMeter: 0.750, aspartate aminotransferase platelet ratio index: 0.742, Fib-4: 0.741. In other etiologies, most tests had nonsignificant changes in OIs. In NAFLD, CHC-specific tests were more accurate than NAFLD-specific tests. The combined FibroMeters had significantly higher accuracy than their 2 constitutive tests (FibroMeters and LSM) in at least 1 diagnostic target in all etiologies, except in ALD where LSM had the highest OI, and in 3 diagnostic targets (OIs and 2 area under the receiver operating characteristics) in CHC and NAFLD. Some tests developed in CHC outperformed other tests in their specific etiologies. Tests combining blood markers and LSM outperformed single tests, validating recent guidelines and extending them to main etiologies. Noninvasive fibrosis evaluation can thus be simplified in the main etiologies by using a unique test: either LSM alone, especially in ALD, or preferably combined to blood markers.

High-Throughput Accurate Single-Cell Screening of Euglena gracilis with Fluorescence-Assisted Optofluidic Time-Stretch Microscopy.

Directory of Open Access Journals (Sweden)

Baoshan Guo

Full Text Available The development of reliable, sustainable, and economical sources of alternative fuels is an important, but challenging goal for the world. As an alternative to liquid fossil fuels, algal biofuel is expected to play a key role in alleviating global warming since algae absorb atmospheric CO2 via photosynthesis. Among various algae for fuel production, Euglena gracilis is an attractive microalgal species as it is known to produce wax ester (good for biodiesel and aviation fuel within lipid droplets. To date, while there exist many techniques for inducing microalgal cells to produce and accumulate lipid with high efficiency, few analytical methods are available for characterizing a population of such lipid-accumulated microalgae including E. gracilis with high throughout, high accuracy, and single-cell resolution simultaneously. Here we demonstrate high-throughput, high-accuracy, single-cell screening of E. gracilis with fluorescence-assisted optofluidic time-stretch microscopy-a method that combines the strengths of microfluidic cell focusing, optical time-stretch microscopy, and fluorescence detection used in conventional flow cytometry. Specifically, our fluorescence-assisted optofluidic time-stretch microscope consists of an optical time-stretch microscope and a fluorescence analyzer on top of a hydrodynamically focusing microfluidic device and can detect fluorescence from every E. gracilis cell in a population and simultaneously obtain its image with a high throughput of 10,000 cells/s. With the multi-dimensional information acquired by the system, we classify nitrogen-sufficient (ordinary and nitrogen-deficient (lipid-accumulated E. gracilis cells with a low false positive rate of 1.0%. This method holds promise for evaluating cultivation techniques and selective breeding for microalgae-based biofuel production.

Assessing the measurement of aerosol single scattering albedo by Cavity Attenuated Phase-Shift Single Scattering Monitor (CAPS PMssa)

Science.gov (United States)

Perim de Faria, Julia; Bundke, Ulrich; Onasch, Timothy B.; Freedman, Andrew; Petzold, Andreas

2016-04-01

The necessity to quantify the direct impact of aerosol particles on climate forcing is already well known; assessing this impact requires continuous and systematic measurements of the aerosol optical properties. Two of the main parameters that need to be accurately measured are the aerosol optical depth and single scattering albedo (SSA, defined as the ratio of particulate scattering to extinction). The measurement of single scattering albedo commonly involves the measurement of two optical parameters, the scattering and the absorption coefficients. Although there are well established technologies to measure both of these parameters, the use of two separate instruments with different principles and uncertainties represents potential sources of significant errors and biases. Based on the recently developed cavity attenuated phase shift particle extinction monitor (CAPS PM_{ex) instrument, the CAPS PM_{ssa instrument combines the CAPS technology to measure particle extinction with an integrating sphere capable of simultaneously measuring the scattering coefficient of the same sample. The scattering channel is calibrated to the extinction channel, such that the accuracy of the single scattering albedo measurement is only a function of the accuracy of the extinction measurement and the nephelometer truncation losses. This gives the instrument an accurate and direct measurement of the single scattering albedo. In this study, we assess the measurements of both the extinction and scattering channels of the CAPS PM_{ssa through intercomparisons with Mie theory, as a fundamental comparison, and with proven technologies, such as integrating nephelometers and filter-based absorption monitors. For comparison, we use two nephelometers, a TSI 3563 and an Aurora 4000, and two measurements of the absorption coefficient, using a Particulate Soot Absorption Photometer (PSAP) and a Multi Angle Absorption Photometer (MAAP). We also assess the indirect absorption coefficient

Accurate Measurements of Aircraft Engine Soot Emissions Using a CAPS PMssa Monitor

Science.gov (United States)

Onasch, Timothy; Thompson, Kevin; Renbaum-Wolff, Lindsay; Smallwood, Greg; Make-Lye, Richard; Freedman, Andrew

2016-04-01

We present results of aircraft engine soot emissions measurements during the VARIAnT2 campaign using CAPS PMssa monitors. VARIAnT2, an aircraft engine non-volatile particulate matter (nvPM) emissions field campaign, was focused on understanding the variability in nvPM mass measurements using different measurement techniques and accounting for possible nvPM sampling system losses. The CAPS PMssa monitor accurately measures both the optical extinction and scattering (and thus single scattering albedo and absorption) of an extracted sample using the same sample volume for both measurements with a time resolution of 1 second and sensitivity of better than 1 Mm-1. Absorption is obtained by subtracting the scattering signal from the total extinction. Given that the single scattering albedo of the particulates emitted from the aircraft engine measured at both 630 and 660 nm was on the order of 0.1, any inaccuracy in the scattering measurement has little impact on the accuracy of the ddetermined absorption coefficient. The absorption is converted into nvPM mass using a documented Mass Absorption Coefficient (MAC). Results of soot emission indices (mass soot emitted per mass of fuel consumed) for a turbojet engine as a function of engine power will be presented and compared to results obtained using an EC/OC monitor.

Pregnenolone rescues schizophrenia-like behavior in dopamine transporter knockout mice.

Directory of Open Access Journals (Sweden)

Peiyan Wong

Full Text Available Pregnenolone belongs to a class of endogenous neurosteroids in the central nervous system (CNS, which has been suggested to enhance cognitive functions through GABA(A receptor signaling by its metabolites. It has been shown that the level of pregnenolone is altered in certain brain areas of schizophrenic patients, and clozapine enhances pregnenolone in the CNS in rats, suggesting that pregnenolone could be used to treat certain symptoms of schizophrenia. In addition, early phase proof-of-concept clinical trials have indicated that pregnenolone is effective in reducing the negative symptoms and cognitive deficits of schizophrenia patients. Here, we evaluate the actions of pregnenolone on a mouse model for schizophrenia, the dopamine transporter knockout mouse (DAT KO. DAT KO mice mirror certain symptoms evident in patients with schizophrenia, such as the psychomotor agitation, stereotypy, deficits of prepulse inhibition and cognitive impairments. Following acute treatment, pregnenolone was found to reduce the hyperlocomotion, stereotypic bouts and pre-pulse inhibition (PPI deficits in DAT KO mice in a dose-dependent manner. At 60 mg/kg of pregnenolone, there were no significant differences in locomotor activities and stereotypy between wild-type and DAT KO mice. Similarly, acute treatment of 60 mg/kg of pregnenolone fully rescued PPI deficits of DAT KO mice. Following chronic treatment with pregnenolone at 60 mg/kg, the cognitive deficits of DAT KO mice were rescued in the paradigms of novel object recognition test and social transmission of food preference test. Pregnenolone thus holds promise as a therapeutic candidate in schizophrenia.

Altered Sleep Homeostasis in Rev-erbα Knockout Mice.

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Mang, Géraldine M; La Spada, Francesco; Emmenegger, Yann; Chappuis, Sylvie; Ripperger, Jürgen A; Albrecht, Urs; Franken, Paul

2016-03-01

The nuclear receptor REV-ERBα is a potent, constitutive transcriptional repressor critical for the regulation of key circadian and metabolic genes. Recently, REV-ERBα's involvement in learning, neurogenesis, mood, and dopamine turnover was demonstrated suggesting a specific role in central nervous system functioning. We have previously shown that the brain expression of several core clock genes, including Rev-erbα, is modulated by sleep loss. We here test the consequences of a loss of REV-ERBα on the homeostatic regulation of sleep. EEG/EMG signals were recorded in Rev-erbα knockout (KO) mice and their wild type (WT) littermates during baseline, sleep deprivation, and recovery. Cortical gene expression measurements after sleep deprivation were contrasted to baseline. Although baseline sleep/wake duration was remarkably similar, KO mice showed an advance of the sleep/wake distribution relative to the light-dark cycle. After sleep onset in baseline and after sleep deprivation, both EEG delta power (1-4 Hz) and sleep consolidation were reduced in KO mice indicating a slower increase of homeostatic sleep need during wakefulness. This slower increase might relate to the smaller increase in theta and gamma power observed in the waking EEG prior to sleep onset under both conditions. Indeed, the increased theta activity during wakefulness predicted delta power in subsequent NREM sleep. Lack of Rev-erbα increased Bmal1, Npas2, Clock, and Fabp7 expression, confirming the direct regulation of these genes by REV-ERBα also in the brain. Our results add further proof to the notion that clock genes are involved in sleep homeostasis. Because accumulating evidence directly links REV-ERBα to dopamine signaling the altered homeostatic regulation of sleep reported here are discussed in that context. © 2016 Associated Professional Sleep Societies, LLC.

Accurate recapture identification for genetic mark–recapture studies with error-tolerant likelihood-based match calling and sample clustering

Science.gov (United States)

Sethi, Suresh; Linden, Daniel; Wenburg, John; Lewis, Cara; Lemons, Patrick R.; Fuller, Angela K.; Hare, Matthew P.

2016-01-01

Error-tolerant likelihood-based match calling presents a promising technique to accurately identify recapture events in genetic mark–recapture studies by combining probabilities of latent genotypes and probabilities of observed genotypes, which may contain genotyping errors. Combined with clustering algorithms to group samples into sets of recaptures based upon pairwise match calls, these tools can be used to reconstruct accurate capture histories for mark–recapture modelling. Here, we assess the performance of a recently introduced error-tolerant likelihood-based match-calling model and sample clustering algorithm for genetic mark–recapture studies. We assessed both biallelic (i.e. single nucleotide polymorphisms; SNP) and multiallelic (i.e. microsatellite; MSAT) markers using a combination of simulation analyses and case study data on Pacific walrus (Odobenus rosmarus divergens) and fishers (Pekania pennanti). A novel two-stage clustering approach is demonstrated for genetic mark–recapture applications. First, repeat captures within a sampling occasion are identified. Subsequently, recaptures across sampling occasions are identified. The likelihood-based matching protocol performed well in simulation trials, demonstrating utility for use in a wide range of genetic mark–recapture studies. Moderately sized SNP (64+) and MSAT (10–15) panels produced accurate match calls for recaptures and accurate non-match calls for samples from closely related individuals in the face of low to moderate genotyping error. Furthermore, matching performance remained stable or increased as the number of genetic markers increased, genotyping error notwithstanding.

Assessment of the Tensile Properties for Single Fibers

Science.gov (United States)

2018-02-01

release; distribution is unlimited. 24 7. Conclusions A method for accurately characterizing the tensile material properties of single fibers...subject to any penalty for failing to comply with a collection of information if it does not display a currently valid OMB control number. PLEASE DO NOT...10. SPONSOR/MONITOR’S ACRONYM(S) 11. SPONSOR/MONITOR’S REPORT NUMBER(S) 12. DISTRIBUTION/ AVAILABILITY STATEMENT 13. SUPPLEMENTARY NOTES

A new, accurate predictive model for incident hypertension

DEFF Research Database (Denmark)

Völzke, Henry; Fung, Glenn; Ittermann, Till

2013-01-01

Data mining represents an alternative approach to identify new predictors of multifactorial diseases. This work aimed at building an accurate predictive model for incident hypertension using data mining procedures.......Data mining represents an alternative approach to identify new predictors of multifactorial diseases. This work aimed at building an accurate predictive model for incident hypertension using data mining procedures....

Accurate quantum yields by laser gain vs absorption spectroscopy - Investigation of Br/Br(asterisk) channels in photofragmentation of Br2 and IBr

Science.gov (United States)

Haugen, H. K.; Weitz, E.; Leone, S. R.

1985-01-01

Various techniques have been used to study photodissociation dynamics of the halogens and interhalogens. The quantum yields obtained by these techniques differ widely. The present investigation is concerned with a qualitatively new approach for obtaining highly accurate quantum yields for electronically excited states. This approach makes it possible to obtain an accuracy of 1 percent to 3 percent. It is shown that measurement of the initial transient gain/absorption vs the final absorption in a single time-resolved signal is a very accurate technique in the study of absolute branching fractions in photodissociation. The new technique is found to be insensitive to pulse and probe laser characteristics, molecular absorption cross sections, and absolute precursor density.

Attenuated stress response to acute restraint and forced swimming stress in arginine vasopressin 1b receptor subtype (Avpr1b) receptor knockout mice and wild-type mice treated with a novel Avpr1b receptor antagonist.

Science.gov (United States)

Roper, J A; Craighead, M; O'Carroll, A-M; Lolait, S J

2010-11-01

Arginine vasopressin (AVP) synthesised in the parvocellular region of the hypothalamic paraventricular nucleus and released into the pituitary portal vessels acts on the 1b receptor subtype (Avpr1b) present in anterior pituitary corticotrophs to modulate the release of adrenocorticotrophic hormone (ACTH). Corticotrophin-releasing hormone is considered the major drive behind ACTH release; however, its action is augmented synergistically by AVP. To determine the extent of vasopressinergic influence in the hypothalamic-pituitary-adrenal axis response to restraint and forced swimming stress, we compared the stress hormone levels [plasma ACTH in both stressors and corticosterone (CORT) in restraint stress only] following acute stress in mutant Avpr1b knockout (KO) mice compared to their wild-type controls following the administration of a novel Avpr1b antagonist. Restraint and forced swimming stress-induced increases in plasma ACTH were significantly diminished in mice lacking a functional Avpr1b and in wild-type mice that had been pre-treated with Avpr1b antagonist. A corresponding decrease in plasma CORT levels was also observed in acute restraint-stressed knockout male mice, and in Avpr1b-antagonist-treated male wild-type mice. By contrast, plasma CORT levels were not reduced in acutely restraint-stressed female knockout animals, or in female wild-type animals pre-treated with Avpr1b antagonist. These results demonstrate that pharmacological antagonism or inactivation of Avpr1b causes a reduction in the hypothalamic-pituitary-adrenal (HPA) axis response, particularly ACTH, to acute restraint and forced swimming stress, and show that Avpr1b knockout mice constitute a model by which to study the contribution of Avpr1b to the HPA axis response to acute stressors. © 2010 The Authors. Journal of Neuroendocrinology © 2010 Blackwell Publishing Ltd.

Single-molecule dataset (SMD): a generalized storage format for raw and processed single-molecule data.

Science.gov (United States)

Greenfeld, Max; van de Meent, Jan-Willem; Pavlichin, Dmitri S; Mabuchi, Hideo; Wiggins, Chris H; Gonzalez, Ruben L; Herschlag, Daniel

2015-01-16

Single-molecule techniques have emerged as incisive approaches for addressing a wide range of questions arising in contemporary biological research [Trends Biochem Sci 38:30-37, 2013; Nat Rev Genet 14:9-22, 2013; Curr Opin Struct Biol 2014, 28C:112-121; Annu Rev Biophys 43:19-39, 2014]. The analysis and interpretation of raw single-molecule data benefits greatly from the ongoing development of sophisticated statistical analysis tools that enable accurate inference at the low signal-to-noise ratios frequently associated with these measurements. While a number of groups have released analysis toolkits as open source software [J Phys Chem B 114:5386-5403, 2010; Biophys J 79:1915-1927, 2000; Biophys J 91:1941-1951, 2006; Biophys J 79:1928-1944, 2000; Biophys J 86:4015-4029, 2004; Biophys J 97:3196-3205, 2009; PLoS One 7:e30024, 2012; BMC Bioinformatics 288 11(8):S2, 2010; Biophys J 106:1327-1337, 2014; Proc Int Conf Mach Learn 28:361-369, 2013], it remains difficult to compare analysis for experiments performed in different labs due to a lack of standardization. Here we propose a standardized single-molecule dataset (SMD) file format. SMD is designed to accommodate a wide variety of computer programming languages, single-molecule techniques, and analysis strategies. To facilitate adoption of this format we have made two existing data analysis packages that are used for single-molecule analysis compatible with this format. Adoption of a common, standard data file format for sharing raw single-molecule data and analysis outcomes is a critical step for the emerging and powerful single-molecule field, which will benefit both sophisticated users and non-specialists by allowing standardized, transparent, and reproducible analysis practices.

Sequence Dependent Interactions Between DNA and Single-Walled Carbon Nanotubes

Science.gov (United States)

Roxbury, Daniel

It is known that single-stranded DNA adopts a helical wrap around a single-walled carbon nanotube (SWCNT), forming a water-dispersible hybrid molecule. The ability to sort mixtures of SWCNTs based on chirality (electronic species) has recently been demonstrated using special short DNA sequences that recognize certain matching SWCNTs of specific chirality. This thesis investigates the intricacies of DNA-SWCNT sequence-specific interactions through both experimental and molecular simulation studies. The DNA-SWCNT binding strengths were experimentally quantified by studying the kinetics of DNA replacement by a surfactant on the surface of particular SWCNTs. Recognition ability was found to correlate strongly with measured binding strength, e.g. DNA sequence (TAT)4 was found to bind 20 times stronger to the (6,5)-SWCNT than sequence (TAT)4T. Next, using replica exchange molecular dynamics (REMD) simulations, equilibrium structures formed by (a) single-strands and (b) multiple-strands of 12-mer oligonucleotides adsorbed on various SWCNTs were explored. A number of structural motifs were discovered in which the DNA strand wraps around the SWCNT and 'stitches' to itself via hydrogen bonding. Great variability among equilibrium structures was observed and shown to be directly influenced by DNA sequence and SWCNT type. For example, the (6,5)-SWCNT DNA recognition sequence, (TAT)4, was found to wrap in a tight single-stranded right-handed helical conformation. In contrast, DNA sequence T12 forms a beta-barrel left-handed structure on the same SWCNT. These are the first theoretical indications that DNA-based SWCNT selectivity can arise on a molecular level. In a biomedical collaboration with the Mayo Clinic, pathways for DNA-SWCNT internalization into healthy human endothelial cells were explored. Through absorbance spectroscopy, TEM imaging, and confocal fluorescence microscopy, we showed that intracellular concentrations of SWCNTs far exceeded those of the incubation

[The NIR spectra based variety discrimination for single soybean seed].

Science.gov (United States)

Zhu, Da-Zhou; Wang, Kun; Zhou, Guang-Hua; Hou, Rui-Feng; Wang, Cheng

2010-12-01

With the development of soybean producing and processing, the quality breeding becomes more and more important for soybean breeders. Traditional sampling detection methods for soybean quality need to destroy the seed, and does not satisfy the requirement of earlier generation materials sieving for breeding. Near infrared (NIR) spectroscopy has been widely used for soybean quality detection. However, all these applications were referred to mass samples, and they were not suitable for little or single seed detection in breeding procedure. In the present study, the acousto--optic tunable filter (AOTF) NIR spectroscopy was used to measure the single soybean seed. Two varieties of soybean were measured, which contained 60 KENJIANDOU43 seeds and 60 ZHONGHUANG13 seeds. The results showed that NIR spectra combined with soft independent modeling of class analogy (SIMCA) could accurately discriminate the soybean varieties. The classification accuracy for KENJIANDOU43 seeds and ZHONGHUANG13 was 100%. The spectra of single soybean seed were measured at different positions, and it showed that the seed shape has significant influence on the measurement of spectra, therefore, the key point for single seed measurement was how to accurately acquire the spectra and keep their representativeness. The spectra for soybeans with glossy surface had high repeatability, while the spectra of seeds with external defects had significant difference for several measurements. For the fast sieving of earlier generation materials in breeding, one could firstly eliminate the seeds with external defects, then apply NIR spectra for internal quality detection, and in this way the influence of seed shape and external defects could be reduced.

Chronic minocycline treatment improves social recognition memory in adult male Fmr1 knockout mice.

Science.gov (United States)

Yau, Suk Yu; Chiu, Christine; Vetrici, Mariana; Christie, Brian R

2016-10-01

Fragile X syndrome (FXS) is caused by a mutation in the Fmr1 gene that leads to silencing of the gene and a loss of its gene product, Fragile X mental retardation protein (FMRP). Some of the key behavioral phenotypes for FXS include abnormal social anxiety and sociability. Here we show that Fmr1 knock-out (KO) mice exhibit impaired social recognition when presented with a novel mouse, and they display normal social interactions in other sociability tests. Administering minocycline to Fmr1 KO mice throughout critical stages of neural development improved social recognition memory in the novel mouse recognition task. To determine if synaptic changes in the prefrontal cortex (PFC) could have played a role in this improvement, we examined PSD-95, a member of the membrane-associated guanylate kinase family, and signaling molecules (ERK1/2, and Akt) linked to synaptic plasticity in the PFC. Our analyses indicated that while minocycline treatment can enhance behavioral performance, it does not enhance expression of PSD-95, ERK1/2 or Akt in the PFC. Copyright © 2016 Elsevier B.V. All rights reserved.

Characterization of 3-Dimensional PET Systems for Accurate Quantification of Myocardial Blood Flow.

Science.gov (United States)

Renaud, Jennifer M; Yip, Kathy; Guimond, Jean; Trottier, Mikaël; Pibarot, Philippe; Turcotte, Eric; Maguire, Conor; Lalonde, Lucille; Gulenchyn, Karen; Farncombe, Troy; Wisenberg, Gerald; Moody, Jonathan; Lee, Benjamin; Port, Steven C; Turkington, Timothy G; Beanlands, Rob S; deKemp, Robert A

2017-01-01

Three-dimensional (3D) mode imaging is the current standard for PET/CT systems. Dynamic imaging for quantification of myocardial blood flow with short-lived tracers, such as 82 Rb-chloride, requires accuracy to be maintained over a wide range of isotope activities and scanner counting rates. We proposed new performance standard measurements to characterize the dynamic range of PET systems for accurate quantitative imaging. 82 Rb or 13 N-ammonia (1,100-3,000 MBq) was injected into the heart wall insert of an anthropomorphic torso phantom. A decaying isotope scan was obtained over 5 half-lives on 9 different 3D PET/CT systems and 1 3D/2-dimensional PET-only system. Dynamic images (28 × 15 s) were reconstructed using iterative algorithms with all corrections enabled. Dynamic range was defined as the maximum activity in the myocardial wall with less than 10% bias, from which corresponding dead-time, counting rates, and/or injected activity limits were established for each scanner. Scatter correction residual bias was estimated as the maximum cavity blood-to-myocardium activity ratio. Image quality was assessed via the coefficient of variation measuring nonuniformity of the left ventricular myocardium activity distribution. Maximum recommended injected activity/body weight, peak dead-time correction factor, counting rates, and residual scatter bias for accurate cardiac myocardial blood flow imaging were 3-14 MBq/kg, 1.5-4.0, 22-64 Mcps singles and 4-14 Mcps prompt coincidence counting rates, and 2%-10% on the investigated scanners. Nonuniformity of the myocardial activity distribution varied from 3% to 16%. Accurate dynamic imaging is possible on the 10 3D PET systems if the maximum injected MBq/kg values are respected to limit peak dead-time losses during the bolus first-pass transit. © 2017 by the Society of Nuclear Medicine and Molecular Imaging.

Accurately localizing the thyroid tissue in mature cystic teratoma of ovary by single-photon emission computerized tomography/computerized tomography

International Nuclear Information System (INIS)

Demir, Yusuf; Üçler, Rıfkı; Alkiş, İsmet; Bulut, Gülay

2015-01-01

A 30-year-old woman with hyperthyroidism was admitted to hospital. Although increased thyroid function was found, the gland was normal in ultrasonography (USG). Additionally, thyroid iodine uptake and Tc-99m pertechnetate scintigraphy was normal. Abdomen USG detected a cystic pelvic mass in left ovary. A whole-body scan was performed 48 hours after oral ingestion of 29.6 MBq (0.8 mCi) I-131 (iodine-131) revealed a round structure located to the left lower abdomen. Iodine uptake was detected in this cyst which was compatible with functional thyroid tissue demonstrated by SPECT/CT. The patient was underwent surgical operation and histopathology confirmed mature cystic teratoma. Accurate localization and depiction of thyroid tissue in ovary mass was provided with SPECT/CT