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Sample records for confirm knockout results

  1. Rescue of heart lipoprotein lipase-knockout mice confirms a role for triglyceride in optimal heart metabolism and function.

    Science.gov (United States)

    Khan, Raffay S; Lin, Yan; Hu, Yunying; Son, Ni-Huiping; Bharadwaj, Kalyani G; Palacios, Carla; Chokshi, Aalap; Ji, Ruiping; Yu, Shuiqing; Homma, Sunichi; Schulze, P Christian; Tian, Rong; Goldberg, Ira J

    2013-12-01

    Hearts utilize fatty acids as a primary source of energy. The sources of those lipids include free fatty acids and lipoprotein triglycerides. Deletion of the primary triglyceride-hydrolyzing enzyme lipoprotein lipase (LPL) leads to cardiac dysfunction. Whether heart LPL-knockout (hLPL0) mice are compromised due a deficiency in energetic substrates is unknown. To test whether alternative sources of energy will prevent cardiac dysfunction in hLPL0 mice, two different models were used to supply nonlipid energy. 1) hLPL0 mice were crossed with mice transgenically expressing GLUT1 in cardiomyocytes to increase glucose uptake into the heart; this cross-corrected cardiac dysfunction, reduced cardiac hypertrophy, and increased myocardial ATP. 2) Mice were randomly assigned to a sedentary or training group (swimming) at 3 mo of age, which leads to increased skeletal muscle production of lactate. hLPL0 mice had greater expression of the lactate transporter monocarboxylate transporter-1 (MCT-1) and increased cardiac lactate uptake. Compared with hearts from sedentary hLPL0 mice, hearts from trained hLPL0 mice had adaptive hypertrophy and improved cardiac function. We conclude that defective energy intake and not the reduced uptake of fat-soluble vitamins or cholesterol is responsible for cardiac dysfunction in hLPL0 mice. In addition, our studies suggest that adaptations in cardiac metabolism contribute to the beneficial effects of exercise on the myocardium of patients with heart failure.

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

    Science.gov (United States)

    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.

  3. 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.

  4. Do we need invasive confirmation of cardiac magnetic resonance results?

    Directory of Open Access Journals (Sweden)

    Paweł Siastała

    2017-03-01

    Full Text Available Introduction : Coronary artery revascularization is indicated in patients with documented significant obstruction of coronary blood flow associated with a large area of myocardial ischemia and/or untreatable symptoms. There are a few invasive or noninvasive methods that can provide information about the functional results of coronary artery narrowing. The application of more than one method of ischemia detection in one patient to reevaluate the indications for revascularization is used in case of atypical or no symptoms and/or borderline stenosis. Aim : To evaluate whether the results of cardiac magnetic resonance need to be reconfirmed by the invasive functional method. Material and methods : The hospital database revealed 25 consecutive patients with 29 stenoses who underwent cardiac magnetic resonance (CMR and fractional flow reserve (FFR between the end of 2010 and the end of 2014. The maximal time interval between CMR and FFR was 6 months. None of the patients experienced any clinical events or underwent procedures on coronary arteries between the studies. Results: According to the analysis, the agreement of CMR perfusion with the FFR method was at the level of 89.7%. Assuming that FFR is the gold standard in assessing the severity of stenoses, the sensitivity of CMR perfusion was 90.9%. The percentage of non-severe lesions which were correctly identified in CMR was 88.9%. Conclusions : The study shows that CMR perfusion is a highly sensitive method to detect hemodynamically significant CAD and exclude nonsevere lesions. With FFR as the reference standard, the diagnostic accuracy of MR perfusion to detect ischemic CAD is high.

  5. Do we need invasive confirmation of cardiac magnetic resonance results?

    Science.gov (United States)

    Siastała, Paweł; Kądziela, Jacek; Małek, Łukasz A; Śpiewak, Mateusz; Lech, Katarzyna; Witkowski, Adam

    2017-01-01

    Coronary artery revascularization is indicated in patients with documented significant obstruction of coronary blood flow associated with a large area of myocardial ischemia and/or untreatable symptoms. There are a few invasive or noninvasive methods that can provide information about the functional results of coronary artery narrowing. The application of more than one method of ischemia detection in one patient to reevaluate the indications for revascularization is used in case of atypical or no symptoms and/or borderline stenosis. To evaluate whether the results of cardiac magnetic resonance need to be reconfirmed by the invasive functional method. The hospital database revealed 25 consecutive patients with 29 stenoses who underwent cardiac magnetic resonance (CMR) and fractional flow reserve (FFR) between the end of 2010 and the end of 2014. The maximal time interval between CMR and FFR was 6 months. None of the patients experienced any clinical events or underwent procedures on coronary arteries between the studies. According to the analysis, the agreement of CMR perfusion with the FFR method was at the level of 89.7%. Assuming that FFR is the gold standard in assessing the severity of stenoses, the sensitivity of CMR perfusion was 90.9%. The percentage of non-severe lesions which were correctly identified in CMR was 88.9%. The study shows that CMR perfusion is a highly sensitive method to detect hemodynamically significant CAD and exclude nonsevere lesions. With FFR as the reference standard, the diagnostic accuracy of MR perfusion to detect ischemic CAD is high.

  6. PDHA1 gene knockout in prostate cancer cells results in metabolic reprogramming towards greater glutamine dependence

    Science.gov (United States)

    Li, Yaqing; Li, Xiaoran; Li, Xiaoli; Zhong, Yali; Ji, Yasai; Yu, Dandan; Zhang, Mingzhi; Wen, Jian-Guo; Zhang, Hongquan; Goscinski, Mariusz Adam; Nesland, Jahn M.; Suo, Zhenhe

    2016-01-01

    Alternative pathways of metabolism endowed cancer cells with metabolic stress. Inhibiting the related compensatory pathways might achieve synergistic anticancer results. This study demonstrated that pyruvate dehydrogenase E1α gene knockout (PDHA1 KO) resulted in alterations in tumor cell metabolism by rendering the cells with increased expression of glutaminase1 (GLS1) and glutamate dehydrogenase1 (GLUD1), leading to an increase in glutamine-dependent cell survival. Deprivation of glutamine induced cell growth inhibition, increased reactive oxygen species and decreased ATP production. Pharmacological blockade of the glutaminolysis pathway resulted in massive tumor cells apoptosis and dysfunction of ROS scavenge in the LNCaP PDHA1 KO cells. Further examination of the key glutaminolysis enzymes in human prostate cancer samples also revealed that higher levels of GLS1 and GLUD1 expression were significantly associated with aggressive clinicopathological features and poor clinical outcome. These insights supply evidence that glutaminolysis plays a compensatory role for cell survival upon alternative energy metabolism and targeting the glutamine anaplerosis of energy metabolism via GLS1 and GLUD1 in cancer cells may offer a potential novel therapeutic strategy. PMID:27462778

  7. A Jacob/Nsmf Gene Knockout Results in Hippocampal Dysplasia and Impaired BDNF Signaling in Dendritogenesis.

    Directory of Open Access Journals (Sweden)

    Christina Spilker

    2016-03-01

    Full Text Available Jacob, the protein encoded by the Nsmf gene, is involved in synapto-nuclear signaling and docks an N-Methyl-D-Aspartate receptor (NMDAR-derived signalosome to nuclear target sites like the transcription factor cAMP-response-element-binding protein (CREB. Several reports indicate that mutations in NSMF are related to Kallmann syndrome (KS, a neurodevelopmental disorder characterized by idiopathic hypogonadotropic hypogonadism (IHH associated with anosmia or hyposmia. It has also been reported that a protein knockdown results in migration deficits of Gonadotropin-releasing hormone (GnRH positive neurons from the olfactory bulb to the hypothalamus during early neuronal development. Here we show that mice that are constitutively deficient for the Nsmf gene do not present phenotypic characteristics related to KS. Instead, these mice exhibit hippocampal dysplasia with a reduced number of synapses and simplification of dendrites, reduced hippocampal long-term potentiation (LTP at CA1 synapses and deficits in hippocampus-dependent learning. Brain-derived neurotrophic factor (BDNF activation of CREB-activated gene expression plays a documented role in hippocampal CA1 synapse and dendrite formation. We found that BDNF induces the nuclear translocation of Jacob in an NMDAR-dependent manner in early development, which results in increased phosphorylation of CREB and enhanced CREB-dependent Bdnf gene transcription. Nsmf knockout (ko mice show reduced hippocampal Bdnf mRNA and protein levels as well as reduced pCREB levels during dendritogenesis. Moreover, BDNF application can rescue the morphological deficits in hippocampal pyramidal neurons devoid of Jacob. Taken together, the data suggest that the absence of Jacob in early development interrupts a positive feedback loop between BDNF signaling, subsequent nuclear import of Jacob, activation of CREB and enhanced Bdnf gene transcription, ultimately leading to hippocampal dysplasia.

  8. 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.

  9. [Confirming Indicators of Qualitative Results by Chromatography-mass Spectrometry in Biological Samples].

    Science.gov (United States)

    Liu, S D; Zhang, D M; Zhang, W; Zhang, W F

    2017-04-01

    Because of the exist of complex matrix, the confirming indicators of qualitative results for toxic substances in biological samples by chromatography-mass spectrometry are different from that in non-biological samples. Even in biological samples, the confirming indicators are different in various application areas. This paper reviews the similarities and differences of confirming indicators for the analyte in biological samples by chromatography-mass spectrometry in the field of forensic toxicological analysis and other application areas. These confirming indicators include retention time (RT), relative retention time (RRT), signal to noise (S/N), characteristic ions, relative abundance of characteristic ions, parent ion-daughter ion pair and abundance ratio of ion pair, etc. Copyright© by the Editorial Department of Journal of Forensic Medicine.

  10. Confirmed results of the 248Cm(48Ca,4n)292116 experiment

    International Nuclear Information System (INIS)

    Patin, J B; Moody, K J; Stoyer, M A; Wild, J F; Shaughnessy, D A; Stoyer, N J

    2003-01-01

    The results of a detailed analysis performed on the data obtained in the 248 Cm( 48 Ca,4n) 292 116 reaction is presented. This analysis is independent of the original data analysis performed in Dubna in which three separate decay chains were found. Each decay chain began with an evaporation residue followed by three α decays and ended in a spontaneous fission event, all correlated in time and position. The analysis presented confirms that the three events are present in the data. A summary of the three events will be given as well as a description of the analysis performed

  11. Knockout reactions: experimental aspects

    Energy Technology Data Exchange (ETDEWEB)

    Cortina Gil, D. [Santiago de Compostela Univ. (Spain)

    2007-07-01

    The availability of radioactive beams has given rise to intense activity in the field of direct reactions. The removal of one(two)-nucleon (referred to as nucleon knockout in this text) from a fast exotic projectile has been extensively investigated. This lecture provides a general overview of the experimental results achieved using this technique. The sensitivity of the method to different experimental aspects is illustrated with a few examples. Special attention is given to the application of nucleon-knockout reactions as a general purpose spectroscopic tool. (author)

  12. Knockout reactions: experimental aspects

    International Nuclear Information System (INIS)

    Cortina Gil, D.

    2007-01-01

    The availability of radioactive beams has given rise to intense activity in the field of direct reactions. The removal of one(two)-nucleon (referred to as nucleon knockout in this text) from a fast exotic projectile has been extensively investigated. This lecture provides a general overview of the experimental results achieved using this technique. The sensitivity of the method to different experimental aspects is illustrated with a few examples. Special attention is given to the application of nucleon-knockout reactions as a general purpose spectroscopic tool. (author)

  13. Confirmation of the Z(4430)- resonance and other exotic meson results from the LHCb experiment

    CERN Multimedia

    CERN. Geneva

    2014-01-01

    The LHCb experiment at CERN has recently confirmed the existence of the exotic Z(4430)- state first observed by the Belle experiment in 2008. Its quantum numbers have been measured and the resonant nature of this state has been demonstrated for the first time. As it is charged, the Z(4430)- cannot be classified as a conventional charmonium (ccbar) state, making it a candidate for an exotic resonance composed of four quarks (ccbar udbar). This talk will outline the history of the Z(4430)-, its possible interpretations and describe how the signature of this exotic state can be extracted from the large sample of B0 -> psi(2S) K+pi- decays that LHCb has collected during Run-1 of the LHC. I will also describe recent LHCb results that probe the nature of the exotic X(3872) particle and help to clarify our understanding of the f0(500) and f0(980) scalar mesons that have long thought to be four quark states.

  14. Confirmation of ACRU model results for applications in land use and climate change studies

    Directory of Open Access Journals (Sweden)

    G. P. W. Jewitt

    2010-12-01

    Full Text Available The hydrological responses of a catchment are sensitive to, and strongly coupled to, land use and climate, and changes thereof. The hydrological responses to the impacts of changing land use and climate will be the result of complex interactions, where the change in one may moderate or exacerbate the effects of the other. Further difficulties in assessing these interactions are that dominant drivers of the hydrological system may vary at different spatial and temporal scales. To assess these interactions, a process-based hydrological model, sensitive to land use and climate, and changes thereof, needs to be used. For this purpose the daily time step ACRU model was selected. However, to be able to use a hydrological model such as ACRU with confidence its representation of reality must be confirmed by comparing simulated output against observations across a range of climatic conditions. Comparison of simulated against observed streamflow was undertaken in three climatically diverse South African catchments, ranging from the semi-arid, sub-tropical Luvuvhu catchment, to the winter rainfall Upper Breede catchment and the sub-humid Mgeni catchment. Not only do the climates of the catchments differ, but their primary land uses also vary. In the upper areas of the Mgeni catchment commercial plantation forestry is dominant, while in the middle reaches there are significant areas of commercial plantation sugarcane and urban areas, while the lower reaches are dominated by urban areas. The Luvuvhu catchment has a large proportion of subsistence agriculture and informal residential areas. In the Upper Breede catchment in the Western Cape, commercial orchards and vineyards are the primary land uses. Overall the ACRU model was able to represent the high, low and total flows, with satisfactory Nash-Sutcliffe efficiency indexes obtained for the selected catchments. The study concluded that the ACRU model can be used with confidence to simulate the streamflows

  15. Confirmation of ACRU model results for applications in land use and climate change studies

    Science.gov (United States)

    Warburton, M. L.; Schulze, R. E.; Jewitt, G. P. W.

    2010-12-01

    The hydrological responses of a catchment are sensitive to, and strongly coupled to, land use and climate, and changes thereof. The hydrological responses to the impacts of changing land use and climate will be the result of complex interactions, where the change in one may moderate or exacerbate the effects of the other. Further difficulties in assessing these interactions are that dominant drivers of the hydrological system may vary at different spatial and temporal scales. To assess these interactions, a process-based hydrological model, sensitive to land use and climate, and changes thereof, needs to be used. For this purpose the daily time step ACRU model was selected. However, to be able to use a hydrological model such as ACRU with confidence its representation of reality must be confirmed by comparing simulated output against observations across a range of climatic conditions. Comparison of simulated against observed streamflow was undertaken in three climatically diverse South African catchments, ranging from the semi-arid, sub-tropical Luvuvhu catchment, to the winter rainfall Upper Breede catchment and the sub-humid Mgeni catchment. Not only do the climates of the catchments differ, but their primary land uses also vary. In the upper areas of the Mgeni catchment commercial plantation forestry is dominant, while in the middle reaches there are significant areas of commercial plantation sugarcane and urban areas, while the lower reaches are dominated by urban areas. The Luvuvhu catchment has a large proportion of subsistence agriculture and informal residential areas. In the Upper Breede catchment in the Western Cape, commercial orchards and vineyards are the primary land uses. Overall the ACRU model was able to represent the high, low and total flows, with satisfactory Nash-Sutcliffe efficiency indexes obtained for the selected catchments. The study concluded that the ACRU model can be used with confidence to simulate the streamflows of the three selected

  16. GRAN SASSO/GRENOBLE: Artificial neutrino source confirms solar neutrino result

    International Nuclear Information System (INIS)

    Anon.

    1995-01-01

    In 1992, the Gallex experiment announced the first observation of the neutrinos produced in the primary proton-proton fusion reaction in the core of the Sun, reaction at the origin of the energy production by our star (September 1992, page 1). The Gallex team stressed that the observed neutrino flux was only about two-thirds of the predicted level, confirming the deficit observed by the two pioneering experiments, Ray Davis' chlorine-based detector in the USA and the Kamiokande study in Japan (which are only sensitive to neutrinos from subsidiary solar fusion processes). This deficit demands explanation, and could considerably modify our understanding of how stars shine and/or of neutrino physics. But before drawing conclusions, the Gallex result had to be checked. Gallex, installed in the Italian Gran Sasso underground Laboratory, is a radiochemical experiment using neutrino interactions to transform gallium-71 into germanium-71. The latter is radioactive and decays with a half-life of 11.4 days. Counting the germanium-71 atoms extracted from the target tank measures the neutrino flux to which the detector is exposed. Neutrinos are famous for their reluctance to interact. 65 billion per square centimetre per second on the surface of the Earth produce only one germanium-71 atom in the Gallex target containing 30 tons of gallium. This is at the limit of homeopathy (extracting few atoms of germanium-71 from a solution containing 10 30 atoms) and needs careful checking. Since it is not possible to switch off the Sun, the only recourse was to build an artificial neutrino source more powerful than the Sun as a benchmark. This was done last summer. Last May, 36 kilograms of chromium grains were placed in the Siloe reactor of the French Commissariat à l'énergie atomique, Grenoble. The chromium had been previously enriched to 40% chromium-50 by the Kurchatov Institute in Moscow (natural chromium contains only 4.5% chromium-50). A dedicated core was built for

  17. Lymphocyte signaling: beyond knockouts.

    Science.gov (United States)

    Saveliev, Alexander; Tybulewicz, Victor L J

    2009-04-01

    The analysis of lymphocyte signaling was greatly enhanced by the advent of gene targeting, which allows the selective inactivation of a single gene. Although this gene 'knockout' approach is often informative, in many cases, the phenotype resulting from gene ablation might not provide a complete picture of the function of the corresponding protein. If a protein has multiple functions within a single or several signaling pathways, or stabilizes other proteins in a complex, the phenotypic consequences of a gene knockout may manifest as a combination of several different perturbations. In these cases, gene targeting to 'knock in' subtle point mutations might provide more accurate insight into protein function. However, to be informative, such mutations must be carefully based on structural and biophysical data.

  18. Knockout of the aryl hydrocarbon receptor results in distinct hepatic and renal phenotypes in rats and mice

    Energy Technology Data Exchange (ETDEWEB)

    Harrill, Joshua A. [The Hamner Institute for Health Sciences, Institute for Chemical Safety Sciences, RTP, NC 27709 (United States); Hukkanen, Renee R.; Lawson, Marie; Martin, Greg [The Dow Chemical Company, Midland, MI 48640 (United States); Gilger, Brian [North Carolina State University, College of Veterinary Medicine, Raleigh, NC 27606 (United States); Soldatow, Valerie [University of North Carolina, Department of Environmental Sciences and Engineering, Chapel Hill, NC 27599 (United States); LeCluyse, Edward L. [The Hamner Institute for Health Sciences, Institute for Chemical Safety Sciences, RTP, NC 27709 (United States); Budinsky, Robert A.; Rowlands, J. Craig [The Dow Chemical Company, Midland, MI 48640 (United States); Thomas, Russell S., E-mail: RThomas@thehamner.org [The Hamner Institute for Health Sciences, Institute for Chemical Safety Sciences, RTP, NC 27709 (United States)

    2013-10-15

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor which plays a role in the development of multiple tissues and is activated by a large number of ligands, including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In order to examine the roles of the AHR in both normal biological development and response to environmental chemicals, an AHR knockout (AHR-KO) rat model was created and compared with an existing AHR-KO mouse. AHR-KO rats harboring either 2-bp or 29-bp deletion mutation in exon 2 of the AHR were created on the Sprague–Dawley genetic background using zinc-finger nuclease (ZFN) technology. Rats harboring either mutation type lacked expression of AHR protein in the liver. AHR-KO rats were also insensitive to thymic involution, increased hepatic weight and the induction of AHR-responsive genes (Cyp1a1, Cyp1a2, Cyp1b1, Ahrr) following acute exposure to 25 μg/kg TCDD. AHR-KO rats had lower basal expression of transcripts for these genes and also accumulated ∼ 30–45-fold less TCDD in the liver at 7 days post-exposure. In untreated animals, AHR-KO mice, but not AHR-KO rats, had alterations in serum analytes indicative of compromised hepatic function, patent ductus venosus of the liver and persistent hyaloid arteries in the eye. AHR-KO rats, but not AHR-KO mice, displayed pathological alterations to the urinary tract: bilateral renal dilation (hydronephrosis), secondary medullary tubular and uroepithelial degenerative changes and bilateral ureter dilation (hydroureter). The present data indicate that the AHR may play significantly different roles in tissue development and homeostasis and toxicity across rodent species. - Highlights: • An AHR knockout rat was generated on a Sprague–Dawley outbred background. • AHR-KO rats lack expression of AHR protein. • AHR-KO rats are insensitive to TCDD-mediated effects. • Data suggests difference in the role of AHR in tissue development of rats and mice. • Abnormalities in vascular

  19. Knockout of the aryl hydrocarbon receptor results in distinct hepatic and renal phenotypes in rats and mice

    International Nuclear Information System (INIS)

    Harrill, Joshua A.; Hukkanen, Renee R.; Lawson, Marie; Martin, Greg; Gilger, Brian; Soldatow, Valerie; LeCluyse, Edward L.; Budinsky, Robert A.; Rowlands, J. Craig; Thomas, Russell S.

    2013-01-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor which plays a role in the development of multiple tissues and is activated by a large number of ligands, including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In order to examine the roles of the AHR in both normal biological development and response to environmental chemicals, an AHR knockout (AHR-KO) rat model was created and compared with an existing AHR-KO mouse. AHR-KO rats harboring either 2-bp or 29-bp deletion mutation in exon 2 of the AHR were created on the Sprague–Dawley genetic background using zinc-finger nuclease (ZFN) technology. Rats harboring either mutation type lacked expression of AHR protein in the liver. AHR-KO rats were also insensitive to thymic involution, increased hepatic weight and the induction of AHR-responsive genes (Cyp1a1, Cyp1a2, Cyp1b1, Ahrr) following acute exposure to 25 μg/kg TCDD. AHR-KO rats had lower basal expression of transcripts for these genes and also accumulated ∼ 30–45-fold less TCDD in the liver at 7 days post-exposure. In untreated animals, AHR-KO mice, but not AHR-KO rats, had alterations in serum analytes indicative of compromised hepatic function, patent ductus venosus of the liver and persistent hyaloid arteries in the eye. AHR-KO rats, but not AHR-KO mice, displayed pathological alterations to the urinary tract: bilateral renal dilation (hydronephrosis), secondary medullary tubular and uroepithelial degenerative changes and bilateral ureter dilation (hydroureter). The present data indicate that the AHR may play significantly different roles in tissue development and homeostasis and toxicity across rodent species. - Highlights: • An AHR knockout rat was generated on a Sprague–Dawley outbred background. • AHR-KO rats lack expression of AHR protein. • AHR-KO rats are insensitive to TCDD-mediated effects. • Data suggests difference in the role of AHR in tissue development of rats and mice. • Abnormalities in vascular

  20. Knockout of the alanine racemase gene in Aeromonas hydrophila HBNUAh01 results in cell wall damage and enhanced membrane permeability.

    Science.gov (United States)

    Liu, Dong; Zhang, Lu; Xue, Wen; Wang, Yaping; Ju, Jiansong; Zhao, Baohua

    2015-07-01

    This study focused on the alanine racemase gene (alr-2), which is involved in the synthesis of d-alanine that forms the backbone of the cell wall. A stable alr-2 knockout mutant of Aeromonas hydrophila HBNUAh01 was constructed. When the mutant was supplemented with d-alanine, growth was unaffected; deprivation of d-alanine caused the growth arrest of the starved mutant cells, but not cell lysis. No alanine racemase activity was detected in the culture of the mutant. Additionally, a membrane permeability assay showed increasing damage to the cell wall during d-alanine starvation. No such damage was observed in the wild type during culture. Scanning and transmission electron microscopy analyses revealed deficiencies of the cell envelope and perforation of the cell wall. Leakage of UV-absorbing substances from the mutants was also observed. Thus, the partial viability of the mutants and their independence of d-alanine for growth indicated that inactivation of alr-2 does not impose an auxotrophic requirement for d-alanine. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  1. Central nervous system-specific knockout of steroidogenic factor 1 results in increased anxiety-like behavior.

    Science.gov (United States)

    Zhao, Liping; Kim, Ki Woo; Ikeda, Yayoi; Anderson, Kimberly K; Beck, Laurel; Chase, Stephanie; Tobet, Stuart A; Parker, Keith L

    2008-06-01

    Steroidogenic factor 1 (SF-1) plays key roles in adrenal and gonadal development, expression of pituitary gonadotropins, and development of the ventromedial hypothalamic nucleus (VMH). If kept alive by adrenal transplants, global knockout (KO) mice lacking SF-1 exhibit delayed-onset obesity and decreased locomotor activity. To define specific roles of SF-1 in the VMH, we used the Cre-loxP system to inactivate SF-1 in a central nervous system (CNS)-specific manner. These mice largely recapitulated the VMH structural defect seen in mice lacking SF-1 in all tissues. In multiple behavioral tests, mice with CNS-specific KO of SF-1 had significantly more anxiety-like behavior than wild-type littermates. The CNS-specific SF-1 KO mice had diminished expression or altered distribution in the mediobasal hypothalamus of several genes whose expression has been linked to stress and anxiety-like behavior, including brain-derived neurotrophic factor, the type 2 receptor for CRH (Crhr2), and Ucn 3. Moreover, transfection and EMSAs support a direct role of SF-1 in Crhr2 regulation. These findings reveal important roles of SF-1 in the hypothalamic expression of key regulators of anxiety-like behavior, providing a plausible molecular basis for the behavioral effect of CNS-specific KO of this nuclear receptor.

  2. Non-Smoker Exposure to Secondhand Cannabis Smoke. I. Urine Screening and Confirmation Results

    Science.gov (United States)

    Cone, Edward J.; Bigelow, George E.; Herrmann, Evan S.; Mitchell, John M.; LoDico, Charles; Flegel, Ronald; Vandrey, Ryan

    2015-01-01

    Increased cannabis potency has renewed concerns that secondhand exposure to cannabis smoke can produce positive drug tests. A systematic study was conducted of smoke exposure on drug-free participants. Six experienced cannabis users smoked cannabis cigarettes (5.3% THC in Session 1 and 11.3% THC in Sessions 2 and 3) in a sealed chamber. Six non-smokers were seated with smokers in an alternating manner. Sessions 1 and 2 were conducted with no ventilation and ventilation was employed in Session 3. Non-smoking participant specimens (collected 0–34 h) were analyzed with four immunoassays at different cutoff concentrations (20, 50, 75 and 100 ng/mL) and by GC-MS (LOQ = 0.75 ng/mL). No presumptive positives occurred for non-smokers at 100 and 75 ng/mL; a single positive occurred at 50 ng/mL; and multiple positives occurred at 20 ng/mL. Maximum THCCOOH concentrations by GC-MS for non-smokers ranged from 1.3 to 57.5 ng/mL. THCCOOH concentrations generally increased with THC potency, but room ventilation substantially reduced exposure levels. These results demonstrate that extreme cannabis smoke exposure can produce positive urine tests at commonly utilized cutoff concentrations. However, positive tests are likely to be rare, limited to the hours immediately post-exposure, and occur only under environmental circumstances where exposure is obvious. PMID:25326203

  3. CRISPR/Cas9-mediated knockout of c-REL in HeLa cells results in profound defects of the cell cycle

    Science.gov (United States)

    Ruiz-Perera, Lucia M.; Kadhim, Hussamadin M.; Tertel, Tobias; Henkel, Elena; Hübner, Wolfgang; Huser, Thomas; Kaltschmidt, Barbara; Kaltschmidt, Christian

    2017-01-01

    Cervical cancer is the fourth common cancer in women resulting worldwide in 266,000 deaths per year. Belonging to the carcinomas, new insights into cervical cancer biology may also have great implications for finding new treatment strategies for other kinds of epithelial cancers. Although the transcription factor NF-κB is known as a key player in tumor formation, the relevance of its particular subunits is still underestimated. Here, we applied CRISPR/Cas9n-mediated genome editing to successfully knockout the NF-κB subunit c-REL in HeLa Kyoto cells as a model system for cervical cancers. We successfully generated a homozygous deletion in the c-REL gene, which we validated using sequencing, qPCR, immunocytochemistry, western blot analysis, EMSA and analysis of off-target effects. On the functional level, we observed the deletion of c-REL to result in a significantly decreased cell proliferation in comparison to wildtype (wt) without affecting apoptosis. The impaired proliferative behavior of c-REL-/- cells was accompanied by a strongly decreased amount of the H2B protein as well as a significant delay in the prometaphase of mitosis compared to c-REL+/+ HeLa Kyoto cells. c-REL-/- cells further showed significantly decreased expression levels of c-REL target genes in comparison to wt. In accordance to our proliferation data, we observed the c-REL knockout to result in a significantly increased resistance against the chemotherapeutic agents 5-Fluoro-2’-deoxyuridine (5-FUDR) and cisplatin. In summary, our findings emphasize the importance of c-REL signaling in a cellular model of cervical cancer with direct clinical implications for the development of new treatment strategies. PMID:28767691

  4. The serotonin transporter knockout rat : A review

    NARCIS (Netherlands)

    Olivier, Jocelien; Cools, Alexander; Ellenbroek, Bart A.; Cuppen, E.; Homberg, Judith; Kalueff, Allan V.; LaPorte, Justin L.

    2010-01-01

    This chapter dicusses the most recent data on the serotonin transporter knock-out rat, a unique rat model that has been generated by target-selected N-ethyl-N-nitrosourea (ENU) driven mutagenesis. The knock-out rat is the result of a premature stopcodon in the serotonin transporter gene, and the

  5. Knock-out reactions

    International Nuclear Information System (INIS)

    de Forest, T. Jr.

    1977-01-01

    It is pointed out that the primary motivation for performing high energy single nucleon knock-out reactions is based on the concept of quasi-elastic scattering. The validity of and corrections to the partial wave impulse approximation and kinematical invariance of knock-out reactions and tests of the reaction mechanism are treated. The effect of distortions on the momentum distribution in the effective momentum approximation for given parameters are plotted. 12 references

  6. 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.

    Directory of Open Access Journals (Sweden)

    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

  7. Endochondral fracture healing with external fixation in the Sost knockout mouse results in earlier fibrocartilage callus removal and increased bone volume fraction and strength.

    Science.gov (United States)

    Morse, A; Yu, N Y C; Peacock, L; Mikulec, K; Kramer, I; Kneissel, M; McDonald, M M; Little, D G

    2015-02-01

    Sclerostin deficiency, via genetic knockout or anti-Sclerostin antibody treatment, has been shown to cause increased bone volume, density and strength of calluses following endochondral bone healing. However, there is limited data on the effect of Sclerostin deficiency on the formative early stage of fibrocartilage (non-bony tissue) formation and removal. In this study we extensively investigate the early fibrocartilage callus. Closed tibial fractures were performed on Sost(-/-) mice and age-matched wild type (C57Bl/6J) controls and assessed at multiple early time points (7, 10 and 14days), as well as at 28days post-fracture after bony union. External fixation was utilized, avoiding internal pinning and minimizing differences in stability stiffness, a variable that has confounded previous research in this area. Normal endochondral ossification progressed in wild type and Sost(-/-) mice with equivalent volumes of fibrocartilage formed at early day 7 and day 10 time points, and bony union in both genotypes by day 28. There were no significant differences in rate of bony union; however there were significant increases in fibrocartilage removal from the Sost(-/-) fracture calluses at day 14 suggesting earlier progression of endochondral healing. Earlier bone formation was seen in Sost(-/-) calluses over wild type with greater bone volume at day 10 (221%, p<0.01). The resultant Sost(-/-) united bony calluses at day 28 had increased bone volume fraction compared to wild type calluses (24%, p<0.05), and the strength of the fractured Sost(-/-) tibiae was greater than that that of wild type fractured tibiae. In summary, bony union was not altered by Sclerostin deficiency in externally-fixed closed tibial fractures, but fibrocartilage removal was enhanced and the resultant united bony calluses had increased bone fraction and increased strength. Crown Copyright © 2014. Published by Elsevier Inc. All rights reserved.

  8. Burden of laboratory-confirmed Campylobacter infections in Guatemala 2008–2012: Results from a facility-based surveillance system

    Science.gov (United States)

    Benoit, Stephen R.; Lopez, Beatriz; Arvelo, Wences; Henao, Olga; Parsons, Michele B.; Reyes, Lissette; Moir, Juan Carlos; Lindblade, Kim

    2015-01-01

    Introduction Campylobacteriosis is one of the leading causes of gastroenteritis worldwide. This study describes the epidemiology of laboratory-confirmed Campylobacter diarrheal infections in two facility-based surveillance sites in Guatemala. Methods Clinical, epidemiologic, and laboratory data were collected on patients presenting with acute diarrhea from select healthcare facilities in the departments of Santa Rosa and Quetzaltenango, Guatemala, from January 2008 through August 2012. Stool specimens were cultured for Campylobacter and antimicrobial susceptibility testing was performed on a subset of isolates. Multidrug resistance (MDR) was defined as resistance to ≥3 antimicrobial classes. Results Campylobacter was isolated from 306 (6.0%) of 5137 stool specimens collected. For children Guatemala; antimicrobial resistance was high, and treatment regimens in the ambulatory setting which included metronidazole and trimethoprim-sulfamethoxazole and lacked oral rehydration were sub-optimal. PMID:24534336

  9. Knockout beyond the dripline

    Energy Technology Data Exchange (ETDEWEB)

    Bonaccorso, A. [INFN, Sez. di Pisa, Largo B. Pontecorvo 3, 56127 Pisa (Italy); Charity, R. J. [Department of Chemistry, Washington University, St. Louis, Missouri 63130 (United States); Kumar, R. [Department of Physics, Deenbandhu Chhoturam University of Science and Technology, Murthal, Sonepat-131039 Haryana (India); Salvioni, G. [INFN, Sez. di Pisa, Largo B. Pontecorvo 3, 56127 Pisa, Italy and Dipartimento di Fisica, Università di Pisa, Largo B. Pontecorvo 3, 56127, Pisa (Italy)

    2015-02-24

    In this contribution, we will describe neutron and proton removal from {sup 9}C and {sup 7}Be which are two particularly interesting nuclei entering the nucleo-synthesis pp-chain [1, 2]. Neutron and proton removal reactions have been used in the past twenty years to probe the single-particle structure of exotic nuclei. The core parallel-momentum distribution can give information on the angular momentum and spin of the nucleon initial state while the total removal cross section is sensitive to the asymptotic part of the initial wave function and also to the reaction mechanism. Because knockout is a peripheral reaction from which the Asymptotic Normalization Constant (ANC) of the single-particle wave function can be extracted, it has been used as an indirect method to obtain the rate of reactions like {sup 8}B(p,γ){sup 9}C or {sup 7}Be(p,γ){sup 8}B. Nucleon removal has recently been applied by the HiRA collaboration [3] to situations in which the remaining “core” is beyond the drip line, such as {sup 8}C and {sup 6}Be, unbound by one or more protons, and whose excitation-energy spectrum can be obtained by the invariant-mass method. By gating on the ground-state peak, “core” parallel-momentum distributions and total knockout cross sections have been obtained similar to previous studies with well-bound “cores”. In addition for each projectile, knock out to final bound states has also been obtained in several cases. We will report on the theoretical description and comparison to this experimental data for a few cases for which advances in the accuracy of the transfer-to-the continuum model [4, 5] have been made [6]. These include the use, when available, of “ab-initio” overlaps for the initial state [7] and in particular their ANC values [8]. Also, the construction of a nucleus-target folding potential for the treatment of the core-target S-matrix [9] using for the cores “ab-initio” densities [10] and state-of-the-art n−{sup 9}Be optical

  10. 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.

  11. 49 CFR 655.61 - Action when an employee has a verified positive drug test result or has a confirmed alcohol test...

    Science.gov (United States)

    2010-10-01

    ... drug test result or has a confirmed alcohol test result of 0.04 or greater, or refuses to submit to a... drug test result or has a confirmed alcohol test result of 0.04 or greater, or refuses to submit to a... performing a safety-sensitive function. (3) If an employee refuses to submit to a drug or alcohol test...

  12. 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.

  13. Burden of laboratory-confirmed Campylobacter infections in Guatemala 2008-2012: results from a facility-based surveillance system.

    Science.gov (United States)

    Benoit, Stephen R; Lopez, Beatriz; Arvelo, Wences; Henao, Olga; Parsons, Michele B; Reyes, Lissette; Moir, Juan Carlos; Lindblade, Kim

    2014-03-01

    Campylobacteriosis is one of the leading causes of gastroenteritis worldwide. This study describes the epidemiology of laboratory-confirmed Campylobacter diarrheal infections in two facility-based surveillance sites in Guatemala. Clinical, epidemiologic, and laboratory data were collected on patients presenting with acute diarrhea from select healthcare facilities in the departments of Santa Rosa and Quetzaltenango, Guatemala, from January 2008 through August 2012. Stool specimens were cultured for Campylobacter and antimicrobial susceptibility testing was performed on a subset of isolates. Multidrug resistance (MDR) was defined as resistance to ≥3 antimicrobial classes. Campylobacter was isolated from 306 (6.0%) of 5137 stool specimens collected. For children Guatemala; antimicrobial resistance was high, and treatment regimens in the ambulatory setting which included metronidazole and trimethoprim-sulfamethoxazole and lacked oral rehydration were sub-optimal. Published by Elsevier Ltd.

  14. Confirmed efficacy of etoposide and dexamethasone in HLH treatment: long-term results of the cooperative HLH-2004 study

    Science.gov (United States)

    Bergsten, Elisabet; Horne, AnnaCarin; Aricó, Maurizio; Astigarraga, Itziar; Egeler, R. Maarten; Filipovich, Alexandra H.; Ishii, Eiichi; Janka, Gritta; Ladisch, Stephan; Lehmberg, Kai; McClain, Kenneth L.; Minkov, Milen; Montgomery, Scott; Nanduri, Vasanta; Rosso, Diego

    2017-01-01

    Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening hyperinflammatory syndrome comprising familial/genetic HLH (FHL) and secondary HLH. In the HLH-94 study, with an estimated 5-year probability of survival (pSu) of 54% (95% confidence interval, 48%-60%), systemic therapy included etoposide, dexamethasone, and, from week 9, cyclosporine A (CSA). Hematopoietic stem cell transplantation (HSCT) was indicated in patients with familial/genetic, relapsing, or severe/persistent disease. In HLH-2004, CSA was instead administered upfront, aiming to reduce pre-HSCT mortality and morbidity. From 2004 to 2011, 369 children aged <18 years fulfilled HLH-2004 inclusion criteria (5 of 8 diagnostic criteria, affected siblings, and/or molecular diagnosis in FHL-causative genes). At median follow-up of 5.2 years, 230 of 369 patients (62%) were alive (5-year pSu, 61%; 56%-67%). Five-year pSu in children with (n = 168) and without (n = 201) family history/genetically verified FHL was 59% (52%-67%) and 64% (57%-71%), respectively (familial occurrence [n = 47], 58% [45%-75%]). Comparing with historical data (HLH-94), using HLH-94 inclusion criteria, pre-HSCT mortality was nonsignificantly reduced from 27% to 19% (P = .064 adjusted for age and sex). Time from start of therapy to HSCT was shorter compared with HLH-94 (P = .020 adjusted for age and sex) and reported neurological alterations at HSCT were 22% in HLH-94 and 17% in HLH-2004 (using HLH-94 inclusion criteria). Five-year pSu post-HSCT overall was 66% (verified FHL, 70% [63%-78%]). Additional analyses provided specific suggestions on potential pre-HSCT treatment improvements. HLH-2004 confirms that a majority of patients may be rescued by the etoposide/dexamethasone combination but intensification with CSA upfront, adding corticosteroids to intrathecal therapy, and reduced time to HSCT did not improve outcome significantly. PMID:28935695

  15. Proton knock-out in Hall A

    International Nuclear Information System (INIS)

    Jager, K. de

    2003-01-01

    Proton knock-out is studied in a broad program in Hall A at Jefferson Lab. The first experiment performed in Hall A studied the 16 O(e,e'p) reaction. Since then proton knock-out experiments have studied a variety of aspects of that reaction, from single-nucleon properties to its mechanism, such as final-state interactions and two-body currents, in nuclei from 2 H to 16 O. In this review the accomplishments of this program will be summarized and an outlook given of expected future results. (orig.)

  16. 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

  17. Peptidomic analysis of the neurolysin-knockout mouse brain.

    Science.gov (United States)

    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

  18. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  20. Applications of UT results to confirm defects findings by utilization of relevant metallurgical investigations techniques on gas/condensate pipeline working in wet sour gas environment

    Science.gov (United States)

    El-Azhari, O. A.; Gajam, S. Y.

    2015-03-01

    The gas/condensate pipe line under investigation is a 12 inch diameter, 48 km ASTM, A106 steel pipeline, carrying hydrocarbons containing wet CO2 and H2S.The pipe line had exploded in a region 100m distance from its terminal; after 24 years of service. Hydrogen induced cracking (HIC) and sour gas corrosion were expected due to the presence of wet H2S in the gas analysis. In other areas of pipe line ultrasonic testing was performed to determine whether the pipeline can be re-operated. The results have shown presence of internal planner defects, this was attributed to the existence of either laminations, type II inclusions or some service defects such as HIC and step wise cracking (SWC).Metallurgical investigations were conducted on fractured samples as per NACE standard (TM-0284-84). The obtained results had shown macroscopic cracks in the form of SWC, microstructure of steel had MnS inclusions. Crack sensitivity analyses were calculated and the microhardness testing was conducted. These results had confirmed that the line material was suffering from sour gas deteriorations. This paper correlates the field UT inspection findings with those methods investigated in the laboratory. Based on the results obtained a new HIC resistance material pipeline needs to be selected.

  1. Repository performance confirmation

    International Nuclear Information System (INIS)

    Hansen, Francis D.

    2011-01-01

    Yucca Mountain license application identified a broad suite of monitoring activities. A revision of the plan was expected to winnow the number of activities down to a manageable size. As a result, an objective process for the next stage of performance confirmation planning was developed as an integral part of an overarching long-term testing and monitoring strategy. The Waste Isolation Pilot Plant compliance monitoring program at once reflects its importance to stakeholders while demonstrating adequate understanding of relevant monitoring parameters. The compliance criteria were stated by regulation and are currently monitored as part of the regulatory rule for disposal. At the outset, the screening practice and parameter selection were not predicated on a direct or indirect correlation to system performance metrics, as was the case for Yucca Mountain. Later on, correlation to performance was established, and the Waste Isolation Pilot Plant continues to monitor ten parameters originally identified in the compliance certification documentation. The monitoring program has proven to be effective for the technical intentions and societal or public assurance. The experience with performance confirmation in the license application process for Yucca Mountain helped identify an objective, quantitative methodology for this purpose. Revision of the existing plan would be based on findings of the total system performance assessment. Identification and prioritization of confirmation activities would then derive from performance metrics associated with performance assessment. Given the understanding of repository performance confirmation, as reviewed in this paper, it is evident that the performance confirmation program for the Yucca Mountain project could be readily re-engaged if licensing activities resumed.

  2. Exploring the Great Schism in the Social Sciences: Confirmation Bias and the Interpretation of Results Relating to Biological Influences on Human Behavior and Psychology.

    Science.gov (United States)

    Winking, Jeffrey

    2018-01-01

    The nature-nurture debate is one that biologists often dismiss as a false dichotomy, as all phenotypic traits are the results of complex processes of gene and environment interactions. However, such dismissiveness belies the ongoing debate that is unmistakable throughout the biological and social sciences concerning the role of biological influences in the development of psychological and behavioral traits in humans. Many have proposed that this debate is due to ideologically driven biases in the interpretation of results. Those favoring biological approaches have been accused of a greater willingness to accept biological explanations so as to rationalize or justify the status quo of inequality. Those rejecting biological approaches have been accused of an unwillingness to accept biological explanations so as to attribute inequalities solely to social and institutional factors, ultimately allowing for the possibility of social equality. While it is important to continue to investigate this topic through further research and debate, another approach is to examine the degree to which the allegations of bias are indeed valid. To accomplish this, a convenience sample of individuals with relevant postgraduate degrees was recruited from Mechanical Turk and social media. Participants were asked to rate the inferential power of different research designs and of mock results that varied in the degree to which they supported different ideologies. Results were suggestive that researchers harbor sincere differences of opinion concerning the inferential value of relevant research. There was no suggestion that ideological confirmation biases drive these differences. However, challenges associated with recruiting a large enough sample of experts as well as identifying believable mock scenarios limit the study's inferential scope.

  3. Survey of quasi-free cluster knockout

    International Nuclear Information System (INIS)

    Roos, P.G.; Chant, N.S.

    1975-01-01

    The investigation of quasi-free knockout reactions has been proceeding for many years now, since the first experiments studying (p,2p) reactions on light nuclei. These experiments clearly showed the dominance of quasi-free proton knockout, and have provided information on the proton holes states in nuclei. From very early in the game people extended these studies to the knock-out of clusters, in an attempt to obtain nuclear structure information about clustering in nuclei. These cluster knockout reactions, excluding the nucleon knockout work, are examined. 20 figures, 16 references

  4. 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.

  5. [An attempt to use ultrasonic technique for confirming the diagnosis, planning and observation of long-term treatment results of painful temporo-mandibular joint dysfunction].

    Science.gov (United States)

    Ey-Chmielewska, H

    1998-01-01

    The author presents an attempt of using ultrasonographic technique in diagnosis, planning and observation of treatment results of temporo-mandibular joint pain dysfunctions. Temporo-mandibular joint pain dysfunctions are interchangeably also called temporo-mandibular joint functional disorders. The assessment of pain symptoms in temporo-mandibular joint dysfunctions pain symptoms is principally based on a subjective estimation by the examining practitioner. There is no univocal definition of the disease or a simple index evidencing important symptoms in decision making. Additionally X-ray technique examinations, being hitherto used, in early stages of the disorder do not allow to diagnose it, and are also burdensome to a patient. The aim of this study was to confirm visibility of anatomical elements of the temporo-mandibular joint in an ultrasound examination, assess the mobility of the articular disc before, during and after prosthetic treatment with and without the use of ultrasound technique, and to determine the period of time necessary to obtain a therapeutic effect. The study material consisted of 180 patients, 128 women and 52 men, aged 20 to 60 years, treated by applying prostheses because of temporo-mandibular joint pain dysfunction, in the Department of Prosthetic Dentistry of the Pomeranian Medical Academy. The patients were divided into 2 groups, control and study group. The control group consisted of 90 patients, 63 women and 27 men. In this group prosthetic treatment planning and observation of results was based on a subjective estimation of the practitioner. The study group here comprised 90 patients, 65 women and 25 men, aged 26 to 60 years. In this group prosthetic treatment planning and observation of treatment results were carried on with the use of ultrasound technique. Data from both groups concerning history, results of examinations carried out by ultrasound technique, and the assessment of ultrasound examination were noted on standard

  6. 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.

  7. 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)

  8. 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

  9. [Bipolar obsessive-compulsive disorder: confirmation of results of the "ABC-OCD" survey in 2 populations of patient members versus non-members of an association].

    Science.gov (United States)

    Hantouche, E G; Kochman, F; Demonfaucon, C; Barrot, I; Millet, B; Lancrenon, S; Akiskal, H S

    2002-01-01

    Clinical data are largely focused on depressive comorbidity in OCD. However in practice, treating resistant or severe OCD sufferers revealed many cases who seem to have an authentic OCD with a hidden comorbid bipolar disorder. Most reports had evaluated the OCD comorbidity in unipolar and bipolar mood disorders (Kruger et al., 1995; Chen et Dilsaver, 1995). The only investigation in clinical population focused on the reverse issue was conducted in Pisa. Perugi et al. (1997) have showed in a consecutive series of 315 OCD outpatients, that 15.7% presented a bipolar comorbidity, mostly with BP-II disorder. Further analyses suggested that when comorbidity occurs with bipolar and unipolar depression, it has a differential impact on the clinical picture and course of OCD. The rate of bipolar comorbidity in OCD was analyzed in a recent epidemiological survey undertaken by the French Association of patients suffering from OCD (FA-OCD or AFTOC in French). In a sample of 453 OCD patients, 76% had suffered from a major depression, 11% from bipolar disorder (DSM IV mania or hypomania), 30% from hypomania (cases that obtained a score > or = 10 on the self-rated Angst Hypomania Checklist). According to the score > or = 10 on Self-rated Questionnaire for Cyclothymic Temperament, 50% were classified as cyclothymic. The self-assessment of soft-bipolar dimensions, such as hypomania and cyclothymia was previously validated in a multi-site study in major depression (Hantouche et al., 1998). Further analyses showed that comorbidity with soft bipolarity was characterized by significant interactions with high levels of impulsivity, anger attacks and suicidal behavior. In order to confirm these data, another cohort (n = 175 patients treated by psychiatrists for OCD) was formed and named "PSY-OCD". Comparative analyses between the two populations allowed showing very few demographic and clinical differences. The frequency rate of "bipolar OCD" was equivalent in both populations: BP

  10. Mucosal barrier injury laboratory-confirmed bloodstream infection: results from a field test of a new National Healthcare Safety Network definition.

    Science.gov (United States)

    See, Isaac; Iwamoto, Martha; Allen-Bridson, Kathy; Horan, Teresa; Magill, Shelley S; Thompson, Nicola D

    2013-08-01

    To assess challenges to implementation of a new National Healthcare Safety Network (NHSN) surveillance definition, mucosal barrier injury laboratory-confirmed bloodstream infection (MBI-LCBI). Multicenter field test. Selected locations of acute care hospitals participating in NHSN central line-associated bloodstream infection (CLABSI) surveillance. Hospital staff augmented their CLABSI surveillance for 2 months to incorporate MBI-LCBI: a primary bloodstream infection due to a selected group of organisms in patients with either neutropenia or an allogeneic hematopoietic stem cell transplant with gastrointestinal graft-versus-host disease or diarrhea. Centers for Disease Control and Prevention (CDC) staff reviewed submitted data to verify whether CLABSIs met MBI-LCBI criteria and summarized the descriptive epidemiology of cases reported. Eight cancer, 2 pediatric, and 28 general acute care hospitals including 193 inpatient units (49% oncology/bone marrow transplant [BMT], 21% adult ward, 20% adult critical care, 6% pediatric, 4% step-down) conducted field testing. Among 906 positive blood cultures reviewed, 282 CLABSIs were identified. Of the 103 CLABSIs that also met MBI-LCBI criteria, 100 (97%) were reported from oncology/BMT locations. Agreement between hospital staff and CDC classification of reported CLABSIs as meeting the MBI-LCBI definition was high (90%; κ = 0.82). Most MBI-LCBIs (91%) occurred in patients meeting neutropenia criteria. Some hospitals indicated that their laboratories' methods of reporting cell counts prevented application of neutropenia criteria; revised neutropenia criteria were created using data from field testing. Hospital staff applied the MBI-LCBI definition accurately. Field testing informed modifications for the January 2013 implementation of MBI-LCBI in the NHSN.

  11. The Effect of Preoperative Antimicrobial Prophylaxis on Intraoperative Culture Results in Patients with a Suspected or Confirmed Prosthetic Joint Infection : A Systematic Review

    NARCIS (Netherlands)

    Wouthuyzen-Bakker, Marjan; Benito, Natividad; Soriano, Alex

    Obtaining reliable cultures during revision arthroplasty is important to adequately diagnose and treat a prosthetic joint infection (PJI). The influence of antimicrobial prophylaxis on culture results remains unclear. Since withholding prophylaxis increases the risk for surgical site infections,

  12. Confirmation of high specificity of an automated enzyme immunoassay test for serological diagnosis of syphilis: retrospective evaluation versus results after implementation.

    Science.gov (United States)

    van Dommelen, Laura; Hoebe, Christian J P A; van Tiel, Frank H; Thijs, Carel; Goossens, Valère J; Bruggeman, Cathrien A; van Loo, Inge H M

    2015-03-01

    The optimal algorithm for serological syphilis screening is still a matter of debate. We have previously evaluated the performance of the Bioelisa Syphilis 3.0, using a selection of archived sera, and in this study compare these results with the Bioelisa results after clinical implementation. All Bioelisa Syphilis 3.0 results obtained since clinical implementation were analyzed. Bioelisa-positive or borderline samples were retested using Treponema pallidum particle agglutination, rapid plasma reagin test, fluorescent treponemal antibody-absorption test, and/or immunoblot. On sera sent in together with cerebrospinal fluid, occasionally both the T. pallidum particle agglutination and Bioelisa were performed. The Bioelisa was performed on 14,622 sera. Bioelisa-positive samples, which were not retested by the previously described assays, were withdrawn from the database (n = 36). In 1.3% of the samples (187/14,586), the Bioelisa was positive or borderline and, ultimately, 115 sera were considered true positive (prevalence 0.8%). The specificity of the Bioelisa was 99.5%. Based on the results of all performed diagnostic assays, the specificity of the Bioelisa of 99.5% is very consistent with that found in the initial study (100%; 95% confidence interval was 98.0%-100%). Interpreting (positive) test results is difficult in the absence of a gold standard, especially when the disease prevalence is low. Results should be viewed in the light of the patients' characteristics.

  13. CERN confirms LHC schedule

    CERN Document Server

    2003-01-01

    The CERN Council held its 125th session on 20 June. Highlights of the meeting included confirmation that the LHC is on schedule for a 2007 start-up, and the announcement of a new organizational structure in 2004.

  14. Performance Confirmation Plan

    International Nuclear Information System (INIS)

    Lindner, E.N.

    2000-01-01

    As described, the purpose of the Performance Confirmation Plan is to specify monitoring, testing, and analysis activities for evaluating the accuracy and adequacy of the information used to determine that performance objectives for postclosure will be met. This plan defines a number of specific performance confirmation activities and associated test concepts in support of the MGR that will be implemented to fulfill this purpose. In doing so, the plan defines an approach to identify key factors and processes, predict performance, establish tolerances and test criteria, collect data (through monitoring, testing, and experiments), analyze these data, and recommend appropriate action. The process of defining which factors to address under performance confirmation incorporates input from several areas. In all cases, key performance confirmation factors are those factors which are: (1) important to safety, (2) measurable and predictable, and (3) relevant to the program (i.e., a factor that is affected by construction, emplacement, or is a time-dependent variable). For the present version of the plan, performance confirmation factors important to safety are identified using the principal factors from the RSS (CRWMS M and O 2000a) (which is derived from TSPA analyses) together with other available performance assessment analyses. With this basis, key performance confirmation factors have been identified, and test concepts and test descriptions have been developed in the plan. Other activities are also incorporated into the performance confirmation program outside of these key factors. Additional activities and tests have been incorporated when they are prescribed by requirements and regulations or are necessary to address data needs and model validation requirements relevant to postclosure safety. These other activities have been included with identified factors to construct the overall performance confirmation program

  15. Performance Confirmation Plan

    International Nuclear Information System (INIS)

    Lindner, E.N.

    2000-01-01

    As described, the purpose of the Performance Confirmation Plan is to specify monitoring, testing, and analysis activities for evaluating the accuracy and adequacy of the information used to determine that performance objectives for postclosure will be met. This plan defines a number of specific performance confirmation activities and associated test concepts in support of the MGR that will be implemented to fulfill this purpose. In doing so, the plan defines an approach to identify key factors and processes, predict performance, establish tolerances and test criteria, collect data (through monitoring, testing, and experiments), analyze these data, and recommend appropriate action. The process of defining which factors to address under performance confirmation incorporates input from several areas. In all cases, key performance confirmation factors are those factors which are: (1) important to safety, (2) measurable and predictable, and (3) relevant to the program (i.e., a factor that i s affected by construction, emplacement, or is a time-dependent variable). For the present version of the plan, performance confirmation factors important to safety are identified using the principal factors from the RSS (CRWMS M and O 2000a) (which is derived from TSPA analyses) together with other available performance assessment analyses. With this basis, key performance confirmation factors have been identified, and test concepts and test descriptions have been developed in the plan. Other activities are also incorporated into the performance confirmation program outside of these key factors. Additional activities and tests have been incorporated when they are prescribed by requirements and regulations or are necessary to address data needs and model validation requirements relevant to postclosure safety. These other activities have been included with identified factors to construct the overall performance confirmation program

  16. The Effect of Preoperative Antimicrobial Prophylaxis on Intraoperative Culture Results in Patients with a Suspected or Confirmed Prosthetic Joint Infection: a Systematic Review.

    Science.gov (United States)

    Wouthuyzen-Bakker, Marjan; Benito, Natividad; Soriano, Alex

    2017-09-01

    Obtaining reliable cultures during revision arthroplasty is important to adequately diagnose and treat a prosthetic joint infection (PJI). The influence of antimicrobial prophylaxis on culture results remains unclear. Since withholding prophylaxis increases the risk for surgical site infections, clarification on this topic is critical. A systematic review was performed with the following research question: in patients who undergo revision surgery of a prosthetic joint, does preoperative antimicrobial prophylaxis affect the culture yield of intraoperative samples in comparison with nonpreoperative antimicrobial prophylaxis? Seven articles were included in the final analysis. In most studies, standard diagnostic culture techniques were used. In patients with a PJI, pooled analysis showed a culture yield of 88% (145/165) in the prophylaxis group versus 95% (344/362) in the nonprophylaxis group ( P = 0.004). Subanalysis of patients with chronic PJIs showed positive cultures in 88% (78/89) versus 91% (52/57), respectively ( P = 0.59). In patients with a suspected chronic infection, a maximum difference of 4% in culture yield between the prophylaxis and nonprophylaxis groups was observed. With the use of standard culture techniques, antimicrobial prophylaxis seems to affect cultures in a minority of patients. Along with the known risk of surgical site infections due to inadequate timing of antimicrobial prophylaxis, we discourage the postponement of prophylaxis until tissue samples are obtained in revision surgery. Future studies are necessary to conclude whether the small percentage of false-negative cultures after prophylaxis can be further reduced with the use of more-sensitive culture techniques, like sonication. Copyright © 2017 American Society for Microbiology.

  17. 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

  18. 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.

  19. Plutonian Moon confirmed

    Science.gov (United States)

    In late February, two separate observations confirmed the 1978 discovery by U.S. Naval Observatory scientist James W. Christy of a moon orbiting the planet Pluto. According to the U.S. Naval Observatory, these two observations were needed before the International Astronomical Society (IAS) would officially recognize the discovery.Two types of observations of the moon, which was named Charon after the ferryman in Greek mythology who carried the dead to Pluto's realm, were needed for confirmation: a transit, in which the moon passes in front of Pluto, and an occultation, in which the moon passes behind the planet. These two phenomena occur only during an 8-year period every 124 years that had been calculated to take place during 1984-1985. Both events were observed in late February.

  20. 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.

  1. Single proton knock-out from 24F

    International Nuclear Information System (INIS)

    Thoennessen, M.; Baumann, T.; Brown, B.A.; Enders, J.; Frank, N.H.; Hansen, P.G.; Heckman, P.; Luther, B.A.; Seitz, J.P.; Stolz, A.; Tryggestad, E.

    2004-01-01

    The measurement of the single proton knock-out reaction from 24 F on a 12 C target at 46.7 MeV/nucleon yielded a 23 O ground state population of (6.6+/-1.0) mb. The data were compared to calculations based on the many-body shell model and the eikonal theory. The results are consistent with a [0d5/26]-bar 1s1/2 configuration of 23 O

  2. Planck early results. XXVI. Detection with Planck and confirmation by XMM-Newton of PLCK G266.6-27.3, an exceptionally X-ray luminous and massive galaxy cluster at z ~ 1

    DEFF Research Database (Denmark)

    Lähteenmäki, A.; Poutanen, T.; Natoli, P.

    2011-01-01

    We present first results on PLCKG266.6-27.3, a galaxy cluster candidate detected at a signal-to-noise ratio of 5 in the Planck All Sky survey. An XMM-Newton validation observation has allowed us to confirm that the candidate isa bona fide galaxy cluster. With these X-ray data we measure an accurate...

  3. 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.

  4. Altered Sleep Homeostasis in Rev-erbα Knockout Mice.

    Science.gov (United States)

    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.

  5. 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.

  6. 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.)

  7. 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.

  8. CONFIRMATION OF CIRCUMSTELLAR PHOSPHINE

    Energy Technology Data Exchange (ETDEWEB)

    Agúndez, M.; Cernicharo, J. [Instituto de Ciencia de Materiales de Madrid, CSIC, C/ Sor Juana Inés de la Cruz 3, E-28049 Cantoblanco (Spain); Decin, L. [Sterrenkundig Instituut Anton Pannekoek, University of Amsterdam, Science Park 904, NL-1098 Amsterdam (Netherlands); Encrenaz, P. [LERMA, Observatoire de Paris, 61 Av. de l' Observatoire, F-75014 Paris (France); Teyssier, D. [European Space Astronomy Centre, Urb. Villafranca del Castillo, P.O. Box 50727, E-28080 Madrid (Spain)

    2014-08-01

    Phosphine (PH{sub 3}) was tentatively identified a few years ago in the carbon star envelopes IRC +10216 and CRL 2688 from observations of an emission line at 266.9 GHz attributable to the J = 1-0 rotational transition. We report the detection of the J = 2-1 rotational transition of PH{sub 3} in IRC +10216 using the HIFI instrument on board Herschel, which definitively confirms the identification of PH{sub 3}. Radiative transfer calculations indicate that infrared pumping in excited vibrational states plays an important role in the excitation of PH{sub 3} in the envelope of IRC +10216, and that the observed lines are consistent with phosphine being formed anywhere between the star and 100 R {sub *} from the star, with an abundance of 10{sup –8} relative to H{sub 2}. The detection of PH{sub 3} challenges chemical models, none of which offer a satisfactory formation scenario. Although PH{sub 3} holds just 2% of the total available phosphorus in IRC +10216, it is, together with HCP, one of the major gas phase carriers of phosphorus in the inner circumstellar layers, suggesting that it could also be an important phosphorus species in other astronomical environments. This is the first unambiguous detection of PH{sub 3} outside the solar system, and is a further step toward a better understanding of the chemistry of phosphorus in space.

  9. 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

  10. 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

  11. 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.

  12. TAM receptor knockout mice are susceptible to retinal autoimmune induction.

    Science.gov (United States)

    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.

  13. 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.

  14. Phantom crash confirms models

    International Nuclear Information System (INIS)

    Anon.

    1989-01-01

    To test computer models of how a nuclear reactor's containment building would fare if an airplane crashed into it, the Muto Institute in Tokyo sponsored a 3.2 million dollar project at Sandia National Laboratory to slam an F-4 Phantom jet into a 500 ton concrete wall. The results showed that the computer calculations were accurate

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

    Directory of Open Access Journals (Sweden)

    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

  16. NUCLEON SPIN: Enigma confirmed

    International Nuclear Information System (INIS)

    Anon.

    1994-01-01

    In 1987 the European Muon Collaboration (EMC - June 1988, page 9) reported results from a polarized muon-proton scattering experiment at CERN which puzzled the particle and nuclear physics communities. Contrary to the prediction of the naive quark model, the EMC found that little of the proton spin seemed to be carried by the spins of the quarks. An extensive experimental programme was therefore immediately proposed at CERN, SLAC (Stanford) and DESY (Hamburg) to measure the spin structure function of the neutron and to repeat the proton measurement with improved accuracy

  17. 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.

  18. 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.

  19. Conditional RARα Knockout Mice Reveal Acute Requirement for Retinoic Acid and RARα in Homeostatic Plasticity

    Directory of Open Access Journals (Sweden)

    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

  20. 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.

  1. 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.

  2. 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.

  3. 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.

  4. 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.

  5. Structure around the island of inversion with single-neutron knockout reactions at GANIL

    CERN Document Server

    Fernández-Domínguez, B; Patterson, N; Thomas, J S; Orr, N; Chartier, M; Catford, W; Achouri, N L; Angélique, J-C; Ashwood, N I; Banu, A; Bastin,B; Brown, J; Borcea, R; Franchoo, S; Freer, M; Gaudefroy, L; Laurent, B; Labiche, M; Lemmon, R C; Negoita, F; Paschalis, S; Paul, E S; Petri, M; Roussel-Chomaz, P; Staniou, M; Taylor, M J; Trache, L

    2010-01-01

    The nuclear structure of the 31Mg nucleus has been studied with the singleneutron knockout reaction. We report on the preliminary results of an experiment performed with the EXOGAM array coupled, for the first time, to the SPEG spectrometer at GANIL.We present a provisional result for the inclusive single-neutron knockout cross section of σinc= 90(5) mb. Preliminary exclusive cross sections for the measured bound states, including the ground state, are also presented. Finally, preliminary longitudinal momentum distributions for the ground state and first excited state are also shown. These results are compared to Monte Carlo Shell-Model calculations in the sd-pf region.

  6. Knockout of Vasohibin-1 Gene in Mice Results in Healthy Longevity with Reduced Expression of Insulin Receptor, Insulin Receptor Substrate 1, and Insulin Receptor Substrate 2 in Their White Adipose Tissue

    Directory of Open Access Journals (Sweden)

    Eichi Takeda

    2017-01-01

    Full Text Available Vasohibin-1 (Vash1, originally isolated as an endothelium-derived angiogenesis inhibitor, has a characteristic of promoting stress tolerance in endothelial cells (ECs. We therefore speculated that the lack of the vash1 gene would result in a short lifespan. However, to our surprise, vash1−/− mice lived significantly longer with a milder senescence phenotype than wild-type (WT mice. We sought the cause of this healthy longevity and found that vash1−/− mice exhibited mild insulin resistance along with reduced expression of the insulin receptor (insr, insulin receptor substrate 1 (irs-1, and insulin receptor substrate 2 (irs-2 in their white adipose tissue (WAT but not in their liver or skeletal muscle. The expression of vash1 dominated in the WAT among those 3 organs. Importantly, vash1−/− mice did not develop diabetes even when fed a high-fat diet. These results indicate that the expression of vash1 was required for the normal insulin sensitivity of the WAT and that the target molecules for this activity were insr, irs1, and irs2. The lack of vash1 caused mild insulin resistance without the outbreak of overt diabetes and might contribute to healthy longevity.

  7. 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

  8. Properties of the {sup 7}He ground state from {sup 8}He neutron knockout

    Energy Technology Data Exchange (ETDEWEB)

    Aksyutina, Yu. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, D-64291 Darmstadt (Germany); Fundamental Fysik, Chalmers Tekniska Hoegskola, S-412 96 Goeteborg (Sweden); Johansson, H.T. [Fundamental Fysik, Chalmers Tekniska Hoegskola, S-412 96 Goeteborg (Sweden); Aumann, T.; Boretzky, K. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, D-64291 Darmstadt (Germany); Borge, M.J.G. [Instituto Estructura de la Materia, CSIC, E-28006 Madrid (Spain); Chatillon, A. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, D-64291 Darmstadt (Germany); Chulkov, L.V. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, D-64291 Darmstadt (Germany); Kurchatov Institute, RU-123182 Moscow (Russian Federation); Cortina-Gil, D. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, D-64291 Darmstadt (Germany); University of Santiago de Compostela, 15706 Santiago de Compostela (Spain); Pramanik, U. Datta [Saha Institute of Nuclear Physics, 1/AF Bidhannagar, Kolkata 700064 (India); Emling, H. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, D-64291 Darmstadt (Germany); Forssen, C. [Fundamental Fysik, Chalmers Tekniska Hoegskola, S-412 96 Goeteborg (Sweden); 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, D-64291 Darmstadt (Germany); Jonson, B. [Fundamental Fysik, Chalmers Tekniska Hoegskola, S-412 96 Goeteborg (Sweden)], E-mail: bjn@fy.chalmers.se; Kulessa, R. [Instytut Fizyki, Universytet Jagiellonski, PL-30-059 Krakow (Poland); Langer, C. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, D-64291 Darmstadt (Germany); Lantz, M. [Fundamental Fysik, Chalmers Tekniska Hoegskola, S-412 96 Goeteborg (Sweden); LeBleis, T. [GSI Helmholtzzentrum fuer Schwerionenforschung GmbH, D-64291 Darmstadt (Germany); Lindahl, A.O. [Institutionen foer Fysik, University of Gothenburg, S-412 96 Goeteborg (Sweden)] (and others)

    2009-08-24

    The unbound nucleus {sup 7}He, produced in neutron-knockout reactions with a 240 MeV/u {sup 8}He beam in a liquid-hydrogen target, has been studied in an experiment at the ALADIN-LAND setup at GSI. From an R-matrix analysis the resonance parameters for {sup 7}He as well as the spectroscopic factor for the {sup 6}He(0{sup +}) + n configuration in its ground-state have been obtained. The spectroscopic factor is 0.61 confirming that {sup 7}He is not a pure single-particle state. An analysis of {sup 5}He data from neutron-knockout reactions of {sup 6}He in a carbon target reveals the presence of an s-wave component at low energies in the {alpha}+n relative energy spectrum. A possible low-lying exited state in {sup 7}He observed in neutron knockout data from {sup 8}He in a carbon target and tentatively interpreted as a I{sup {pi}}=1/2{sup -} state, could not be observed in the present experiment. Possible explanations of the shape difference between the {sup 7}He resonance obtained in the two knockout reactions are discussed in terms of target-dependence or different reaction mechanisms at relativistic energies.

  9. Bacteriologically confirmed tuberculosis in children

    International Nuclear Information System (INIS)

    Ozere, I.; Sele, A.; Ozolina, A.

    2005-01-01

    Tuberculosis in children and adults is a growing problem with important public health implications. In Latvia the incidence of tuberculosis (TB) in children up to age 14 has increased from 7,1 per 100000 in 1992 to 28,8 per 100000 in 2003. The diagnosis of TB is confirmed by isolation and identification of M. tuberculosis (MT) from clinical specimen. Confirmation of the disease, however, is difficult in children due to poor bacilli excretion and even under the best circumstances only about 30-40% of pediatric TB cases are proved bacteriologically. Of the 370 pediatric TB cases diagnosed between January 1, 2001 and December 1, 2003 in Latvia, 53 (14,3%) were confirmed bacteriologically. The clinical, radiological, immunological and anamnestic features of confirmed TB can serve as cornerstones for diagnosing of TB, when culture is not available. Objective To evaluate the sensitivity of diagnostic criteria of TB, clinical and radiological manifestation of TB and drug susceptibility of MT isolated also. Methods All consecutive children (53 in total) up to age 14 diagnosed with bacteriologically confirmed TB during 01.01.2001. -01.12.2003. were prospectively evaluated for reasons mentioned above. Results Of the 53 children identified all but one had respiratory tract TB. 17(32,1 %) children were under 4 years of age, 9 (17%) children were 5-9 years old, but 27 (50,9%) patients were 10-14 years old. During evaluation data on TB source case were found in addition in 13 children and total TB contact history was positive in 37 (69,8%) patients. All clinical and radiographical forms of respiratory tract TB were diagnosed. The most common encountered forms were intrathoracic adenopathy in 10 (18,9%) cases and TB pneumonia in 6 (11,3%) cases in children aged 10-14 years. lnthrathoracic adenopathy associated with segmental parenchymal lesion was the most common form in children under 4 years of age -7 (13,2%) cases respectively. Conclusions 1. The clinical and radiological

  10. 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.

  11. 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.

  12. A STAT-1 knockout mouse model for Machupo virus pathogenesis

    Directory of Open Access Journals (Sweden)

    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.

  13. 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.

  14. 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.

  15. 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

  16. 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

  17. 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

  18. 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

  19. 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

  20. Comparison of a newly developed automated and quantitative hepatitis C virus (HCV) core antigen test with the HCV RNA assay for clinical usefulness in confirming anti-HCV results.

    Science.gov (United States)

    Kesli, Recep; Polat, Hakki; Terzi, Yuksel; Kurtoglu, Muhammet Guzel; Uyar, Yavuz

    2011-12-01

    Hepatitis C virus (HCV) is a global health care problem. Diagnosis of HCV infection is mainly based on the detection of anti-HCV antibodies as a screening test with serum samples. Recombinant immunoblot assays are used as supplemental tests and for the final detection and quantification of HCV RNA in confirmatory tests. In this study, we aimed to compare the HCV core antigen test with the HCV RNA assay for confirming anti-HCV results to determine whether the HCV core antigen test may be used as an alternative confirmatory test to the HCV RNA test and to assess the diagnostic values of the total HCV core antigen test by determining the diagnostic specificity and sensitivity rates compared with the HCV RNA test. Sera from a total of 212 treatment-naive patients were analyzed for anti-HCV and HCV core antigen both with the Abbott Architect test and with the molecular HCV RNA assay consisting of a reverse transcription-PCR method as a confirmatory test. The diagnostic sensitivity, specificity, and positive and negative predictive values of the HCV core antigen assay compared to the HCV RNA test were 96.3%, 100%, 100%, and 89.7%, respectively. The levels of HCV core antigen showed a good correlation with those from the HCV RNA quantification (r = 0.907). In conclusion, the Architect HCV antigen assay is highly specific, sensitive, reliable, easy to perform, reproducible, cost-effective, and applicable as a screening, supplemental, and preconfirmatory test for anti-HCV assays used in laboratory procedures for the diagnosis of hepatitis C virus infection.

  1. 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.

  2. 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)

  3. Scrapping of student bursaries confirmed.

    Science.gov (United States)

    Longhurst, Chris

    2016-07-27

    Student bursaries for nurses will be scrapped from next year, the government has confirmed. Undergraduate nursing and midwifery students in England will now face tuition fees and student loans from August 2017.

  4. Selective enrichment of volatiles confirmed

    Science.gov (United States)

    de Pater, Imke

    2018-05-01

    Hydrogen sulfide gas is detected above Uranus's main cloud deck, confirming the prevalence of H2S ice particles as the main cloud component and a strongly unbalanced nitrogen/sulfur ratio in the planet's deep atmosphere.

  5. 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

  6. 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.

  7. Bacterial and Pneumocystis Infections in the Lungs of Gene-Knockout Rabbits with Severe Combined Immunodeficiency

    Directory of Open Access Journals (Sweden)

    Jun Song

    2018-03-01

    Full Text Available Using the CRISPR/Cas9 gene-editing technology, we recently produced a number of rabbits with mutations in immune function genes, including FOXN1, PRKDC, RAG1, RAG2, and IL2RG. Seven founder knockout rabbits (F0 and three male IL2RG null (−/y F1 animals demonstrated severe combined immunodeficiency (SCID, characterized by absence or pronounced hypoplasia of the thymus and splenic white pulp, and absence of immature and mature T and B-lymphocytes in peripheral blood. Complete blood count analysis showed severe leukopenia and lymphocytopenia accompanied by severe neutrophilia. Without prophylactic antibiotics, the SCID rabbits universally succumbed to lung infections following weaning. Pathology examination revealed severe heterophilic bronchopneumonia caused by Bordetella bronchiseptica in several animals, but a consistent feature of lung lesions in all animals was a severe interstitial pneumonia caused by Pneumocystis oryctolagi, as confirmed by histological examination and PCR analysis of Pneumocystis genes. The results of this study suggest that these SCID rabbits could serve as a useful model for human SCID to investigate the disease pathogenesis and the development of gene and drug therapies.

  8. 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.

  9. Bacterial and Pneumocystis Infections in the Lungs of Gene-Knockout Rabbits with Severe Combined Immunodeficiency

    Science.gov (United States)

    Song, Jun; Wang, Guoshun; Hoenerhoff, Mark J.; Ruan, Jinxue; Yang, Dongshan; Zhang, Jifeng; Yang, Jibing; Lester, Patrick A.; Sigler, Robert; Bradley, Michael; Eckley, Samantha; Cornelius, Kelsey; Chen, Kong; Kolls, Jay K.; Peng, Li; Ma, Liang; Chen, Yuqing Eugene; Sun, Fei; Xu, Jie

    2018-01-01

    Using the CRISPR/Cas9 gene-editing technology, we recently produced a number of rabbits with mutations in immune function genes, including FOXN1, PRKDC, RAG1, RAG2, and IL2RG. Seven founder knockout rabbits (F0) and three male IL2RG null (−/y) F1 animals demonstrated severe combined immunodeficiency (SCID), characterized by absence or pronounced hypoplasia of the thymus and splenic white pulp, and absence of immature and mature T and B-lymphocytes in peripheral blood. Complete blood count analysis showed severe leukopenia and lymphocytopenia accompanied by severe neutrophilia. Without prophylactic antibiotics, the SCID rabbits universally succumbed to lung infections following weaning. Pathology examination revealed severe heterophilic bronchopneumonia caused by Bordetella bronchiseptica in several animals, but a consistent feature of lung lesions in all animals was a severe interstitial pneumonia caused by Pneumocystis oryctolagi, as confirmed by histological examination and PCR analysis of Pneumocystis genes. The results of this study suggest that these SCID rabbits could serve as a useful model for human SCID to investigate the disease pathogenesis and the development of gene and drug therapies. PMID:29593714

  10. Abolition of lemniscal barrellette patterning in Prrxl1 knockout mice: Effects upon ingestive behavior.

    Science.gov (United States)

    Bakalar, Dana; Tamaiev, Jonathan; Zeigler, H Philip; Feinstein, Paul

    2015-01-01

    Ingestive behaviors in mice are dependent on orosensory cues transmitted via the trigeminal nerve, as confirmed by transection studies. However, these studies cannot differentiate between deficits caused by the loss of the lemniscal pathway vs. the parallel paralemniscal pathway. The paired-like homeodomain protein Prrxl1 is expressed widely in the brain and spinal cord, including the trigeminal system. A knockout of Prrxl1 abolishes somatotopic barrellette patterning in the lemniscal brainstem nucleus, but not in the parallel paralemniscal nucleus. Null animals are significantly smaller than littermates by postnatal day 5, but reach developmental landmarks at appropriate times, and survive to adulthood on liquid diet. A careful analysis of infant and adult ingestive behavior reveals subtle impairments in suckling, increases in time spent feeding and the duration of feeding bouts, feeding during inappropriate times of the day, and difficulties in the mechanics of feeding. During liquid diet feeding, null mice display abnormal behaviors including extensive use of the paws to move food into the mouth, submerging the snout in the diet, changes in licking, and also have difficulty consuming solid chow pellets. We suggest that our Prrxl1(-/-) animal is a valuable model system for examining the genetic assembly and functional role of trigeminal lemniscal circuits in the normal control of eating in mammals and for understanding feeding abnormalities in humans resulting from the abnormal development of these circuits.

  11. 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

  12. 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.

  13. 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.

  14. 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.

  15. Frequency and predictors of confirmed hypoglycaemia in type 1 and insulin-treated type 2 diabetes mellitus patients in a real-life setting: results from the DIALOG study.

    Science.gov (United States)

    Cariou, B; Fontaine, P; Eschwege, E; Lièvre, M; Gouet, D; Huet, D; Madani, S; Lavigne, S; Charbonnel, B

    2015-04-01

    DIALOG assessed the prevalence and predictors of hypoglycaemia in patients with type 1 (T1DM) or insulin-treated type 2 diabetes mellitus (T2DM) in a real-life setting. In this observational study, insulin-treated patients (n=3048) completed prospective daily questionnaires reporting the frequency and consequences of severe/confirmed non-severe hypoglycaemia over 30 days. Patients (n=3743) also retrospectively reported severe hypoglycaemia over the preceding year. In this prospective survey, 85.3% and 43.6% of patients with T1DM and T2DM, respectively, reported experiencing at least one confirmed hypoglycaemic event over 30 days, while 13.4% and 6.4%, respectively, reported at least one severe event. Hypoglycaemia frequency increased with longer duration of diabetes and insulin therapy. Strongly predictive factors for hypoglycaemia were previous hypoglycaemia, >2 injections/day, BMIinsulin therapy>10 years. HbA1c level was not predictive of hypoglycaemia in either T1DM or T2DM. The confirmed hypoglycaemia rate was increased in the lowest compared with the highest tertile of HbA1c in T1DM, but not T2DM. At the time of enrolment, physicians reported severe hypoglycaemia in 23.6% and 11.9% of T1DM and T2DM patients, respectively, during the preceding year; the retrospective survey yielded frequencies of 31.5% and 21.7%, respectively. Also, severe hypoglycaemia led to medical complications in 10.7% and 7.8% of events in T1DM and T2DM patients, respectively, over 30 days. Using a unique combined prospective and retrospective approach, the DIALOG study found a relatively high frequency of hypoglycaemia among insulin-treated patients. These findings emphasize the importance of a patient-centred approach for managing diabetes in which hypoglycaemia risk evaluation is critical. ClinicalTrials.gov: NCT01628341. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

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

    Directory of Open Access Journals (Sweden)

    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.

  17. 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.

  18. Opinion dynamics with confirmation bias.

    Directory of Open Access Journals (Sweden)

    Armen E Allahverdyan

    Full Text Available Confirmation bias is the tendency to acquire or evaluate new information in a way that is consistent with one's preexisting beliefs. It is omnipresent in psychology, economics, and even scientific practices. Prior theoretical research of this phenomenon has mainly focused on its economic implications possibly missing its potential connections with broader notions of cognitive science.We formulate a (non-Bayesian model for revising subjective probabilistic opinion of a confirmationally-biased agent in the light of a persuasive opinion. The revision rule ensures that the agent does not react to persuasion that is either far from his current opinion or coincides with it. We demonstrate that the model accounts for the basic phenomenology of the social judgment theory, and allows to study various phenomena such as cognitive dissonance and boomerang effect. The model also displays the order of presentation effect-when consecutively exposed to two opinions, the preference is given to the last opinion (recency or the first opinion (primacy -and relates recency to confirmation bias. Finally, we study the model in the case of repeated persuasion and analyze its convergence properties.The standard Bayesian approach to probabilistic opinion revision is inadequate for describing the observed phenomenology of persuasion process. The simple non-Bayesian model proposed here does agree with this phenomenology and is capable of reproducing a spectrum of effects observed in psychology: primacy-recency phenomenon, boomerang effect and cognitive dissonance. We point out several limitations of the model that should motivate its future development.

  19. Performance Confirmation Data Acquisition System

    International Nuclear Information System (INIS)

    D.W. Markman

    2000-01-01

    The purpose of this analysis is to identify and analyze concepts for the acquisition of data in support of the Performance Confirmation (PC) program at the potential subsurface nuclear waste repository at Yucca Mountain. The scope and primary objectives of this analysis are to: (1) Review the criteria for design as presented in the Performance Confirmation Data Acquisition/Monitoring System Description Document, by way of the Input Transmittal, Performance Confirmation Input Criteria (CRWMS M and O 1999c). (2) Identify and describe existing and potential new trends in data acquisition system software and hardware that would support the PC plan. The data acquisition software and hardware will support the field instruments and equipment that will be installed for the observation and perimeter drift borehole monitoring, and in-situ monitoring within the emplacement drifts. The exhaust air monitoring requirements will be supported by a data communication network interface with the ventilation monitoring system database. (3) Identify the concepts and features that a data acquisition system should have in order to support the PC process and its activities. (4) Based on PC monitoring needs and available technologies, further develop concepts of a potential data acquisition system network in support of the PC program and the Site Recommendation and License Application

  20. 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.

  1. Model confirmation in climate economics

    Science.gov (United States)

    Millner, Antony; McDermott, Thomas K. J.

    2016-01-01

    Benefit–cost integrated assessment models (BC-IAMs) inform climate policy debates by quantifying the trade-offs between alternative greenhouse gas abatement options. They achieve this by coupling simplified models of the climate system to models of the global economy and the costs and benefits of climate policy. Although these models have provided valuable qualitative insights into the sensitivity of policy trade-offs to different ethical and empirical assumptions, they are increasingly being used to inform the selection of policies in the real world. To the extent that BC-IAMs are used as inputs to policy selection, our confidence in their quantitative outputs must depend on the empirical validity of their modeling assumptions. We have a degree of confidence in climate models both because they have been tested on historical data in hindcasting experiments and because the physical principles they are based on have been empirically confirmed in closely related applications. By contrast, the economic components of BC-IAMs often rely on untestable scenarios, or on structural models that are comparatively untested on relevant time scales. Where possible, an approach to model confirmation similar to that used in climate science could help to build confidence in the economic components of BC-IAMs, or focus attention on which components might need refinement for policy applications. We illustrate the potential benefits of model confirmation exercises by performing a long-run hindcasting experiment with one of the leading BC-IAMs. We show that its model of long-run economic growth—one of its most important economic components—had questionable predictive power over the 20th century. PMID:27432964

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

    Directory of Open Access Journals (Sweden)

    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.

  3. 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.

  4. Performance confirmation data acquisition system

    International Nuclear Information System (INIS)

    McAffee, D.A.; Raczka, N.T.

    1997-01-01

    As part of the Viability Assessment (VA) work, this QAP-3-9 document presents and evaluates a comprehensive set of viable concepts for collecting Performance Confirmation (PC) related data. The concepts include: monitoring subsurface repository air temperatures, humidity levels and gaseous emissions via the subsurface ventilation systems, and monitoring the repository geo-technical parameters and rock mass from bore-holes located along the perimeter main drifts and throughout a series of human-rated Observation Drifts to be located in a plane 25 meters above the plane of the emplacement drifts. A key element of this document is the development and analysis of a purposed multi-purpose Remote Inspection Gantry that would provide direct, real-time visual, thermal, and radiological monitoring of conditions inside operational emplacement drifts and close-up observations of in-situ Waste Packages. Preliminary finite-element analyses are presented that indicate the technological feasibility of operating an inspection gantry inside the operational emplacement drifts for short inspection missions lasting 2--3 hours. Overall reliability, availability, and maintainability of the PC data collection concepts are discussed. Preliminary concepts for PC data collection network are also provided

  5. Performance confirmation data acquisition system

    Energy Technology Data Exchange (ETDEWEB)

    McAffee, D.A.; Raczka, N.T. [Yucca Mountain Project, Las Vegas, NV (United States)

    1997-12-31

    As part of the Viability Assessment (VA) work, this QAP-3-9 document presents and evaluates a comprehensive set of viable concepts for collecting Performance Confirmation (PC) related data. The concepts include: monitoring subsurface repository air temperatures, humidity levels and gaseous emissions via the subsurface ventilation systems, and monitoring the repository geo-technical parameters and rock mass from bore-holes located along the perimeter main drifts and throughout a series of human-rated Observation Drifts to be located in a plane 25 meters above the plane of the emplacement drifts. A key element of this document is the development and analysis of a purposed multi-purpose Remote Inspection Gantry that would provide direct, real-time visual, thermal, and radiological monitoring of conditions inside operational emplacement drifts and close-up observations of in-situ Waste Packages. Preliminary finite-element analyses are presented that indicate the technological feasibility of operating an inspection gantry inside the operational emplacement drifts for short inspection missions lasting 2--3 hours. Overall reliability, availability, and maintainability of the PC data collection concepts are discussed. Preliminary concepts for PC data collection network are also provided.

  6. Using Daily Horoscopes To Demonstrate Expectancy Confirmation.

    Science.gov (United States)

    Munro, Geoffrey D.; Munro, James E.

    2000-01-01

    Describes a classroom demonstration that uses daily horoscopes to show the effect that expectation can have on judgment. Addresses the preparation, procedure, and results of the demonstration, and student evaluations. States that the demonstration appears to be effective for teaching students about expectancy confirmation. (CMK)

  7. Characterisation of iunH gene knockout strain from Mycobacterium tuberculosis

    Directory of Open Access Journals (Sweden)

    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

  8. Molecular characterization and development of Sarcocystis speeri sarcocysts in gamma interferon gene knockout mice.

    Science.gov (United States)

    Dubey, J P; Verma, S K; Dunams, D; Calero-Bernal, R; Rosenthal, B M

    2015-11-01

    The North American opossum (Didelphis virginiana) is the definitive host for at least three named species of Sarcocystis: Sarcocystis falcatula, Sarcocystis neurona and Sarcocystis speeri. The South American opossums (Didelphis albiventris, Didelphis marsupialis and Didelphis aurita) are definitive hosts for S. falcatula and S. lindsayi. The sporocysts of these Sarcocystis species are similar morphologically. They are also not easily distinguished genetically because of the difficulties of DNA extraction from sporocysts and availability of distinguishing genetic markers. Some of these species can be distinguished by bioassay; S. neurona and S. speeri are infective to gamma interferon gene knockout (KO) mice, but not to budgerigars (Melopsittacus undulatus); whereas S. falcatula and S. lindsayi are infective to budgerigars but not to KO mice. The natural intermediate host of S. speeri is unknown. In the present study, development of sarcocysts of S. speeri in the KO mice is described. Sarcocysts were first seen at 12 days post-inoculation (p.i.), and they became macroscopic (up to 4 mm long) by 25 days p.i. The structure of the sarcocyst wall did not change from the time bradyzoites had formed at 50-220 days p.i. Sarcocysts contained unique villar protrusions, 'type 38'. The polymerase chain reaction amplifications and sequences analysis of three nuclear loci (18S rRNA, 28S rRNA and ITS1) and two mitochondrial loci (cox1 and cytb) of S. speeri isolate from an Argentinean opossum (D. albiventris) confirmed its membership among species of Sarcocystis and indicated an especially close relationship to another parasite in this genus that employs opossums as its definitive host, S. neurona. These results should be useful in finding natural intermediate host of S. speeri.

  9. 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

  10. 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.

  11. 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.

  12. A New Way to Confirm Planet Candidates

    Science.gov (United States)

    Kohler, Susanna

    2016-05-01

    automated batch processing of a large number of candidates.In a recently published study the results of which were announced yesterday the teamapplied their code to the entire catalog of 7,470 Kepler objects of interest.New Planets and False PositivesThe teams code was able to successfully evaluate the total false-positive probability (FPP) for 7,056 of the objects of interest. Of these, 428 objects previously identified as candidates were found to have FPP of more than 90%, suggesting that they are most likely false positives.Periods and radii of candidate and confirmed planets in the Kepler Objects of Interest catalog. Blue circles have previously been identified as confirmed planets. Candidates (orange) are shaded by false positive probability; more transparent means more likely to be a false positive. [Morton et al. 2016]In contrast, 1,935 candidates were found to have FPP of less than 1%, and were therefore declared validated planets. Of these confirmations, 1,284 were previously unconfirmed, more than doubling Keplers previous catalog of 1,041 confirmed planets. Morton and collaborators believe that 9 of these newly confirmed planets may fall within the habitable zone of their host stars.While the announcement of 1,284 newly confirmed planets is huge, the analysis presented in this study is the real news. The code used is publicly available and can be applied to any transiting exoplanet candidate. This means that this analysis technique can be used to find batches of exoplanets in data from the extended Kepler mission (K2) or from the future TESS and PLATO transit missions.CitationTimothy D. Morton et al 2016 ApJ 822 86. doi:10.3847/0004-637X/822/2/86

  13. 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.

  14. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  16. 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.

  17. Water on the Moon Confirmed

    Science.gov (United States)

    Showstack, Randy

    2009-11-01

    When NASA's Lunar Crater Observation and Sensing Satellite (LCROSS) and a companion rocket purposely slammed into a crater at the Moon's south pole on 9 October, some observers on Earth lamented as anticlimactic the raised plumes of material that were partially blocked by a crater ridge and were difficult to see with backyard telescopes. However, it turns out that the projectiles struck it big. “Indeed, yes, we found water. We didn’t find just a little bit; we found a significant amount,” said Anthony Colaprete, LCROSS principal investigator with the NASA Ames Research Center, Moffett Field, Calif. At a 13 November news briefing, Colaprete lifted a 2-gallon plastic bucket and said preliminary results indicate that instruments detected about a dozen buckets' worth of water in parts of the two plumes, the first generated by the spent Centaur upper stage of the Atlas V launch vehicle at 1131 UTC and the second generated by LCROSS about 4 minutes later. NASA described the two plumes as a high-angle plume of vapor and fine dust and a lower-angle ejecta curtain of heavier material. LCROSS and the Centaur upper stage hit the permanently shadowed Cabeus crater.

  18. 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

  19. 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 ...

  20. 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

  1. Decreased levels of free D-aspartic acid in the forebrain of serine racemase (Srr) knock-out mice.

    Science.gov (United States)

    Horio, Mao; Ishima, Tamaki; Fujita, Yuko; Inoue, Ran; Mori, Hisashi; Hashimoto, Kenji

    2013-05-01

    d-Serine, an endogenous co-agonist of the N-methyl-d-aspartate (NMDA) receptor is synthesized from l-serine by serine racemase (SRR). A previous study of Srr knockout (Srr-KO) mice showed that levels of d-serine in forebrain regions, such as frontal cortex, hippocampus, and striatum, but not cerebellum, of mutant mice are significantly lower than those of wild-type (WT) mice, suggesting that SRR is responsible for d-serine production in the forebrain. In this study, we attempted to determine whether SRR affects the level of other amino acids in brain tissue. We found that tissue levels of d-aspartic acid in the forebrains (frontal cortex, hippocampus and striatum) of Srr-KO mice were significantly lower than in WT mice, whereas levels of d-aspartic acid in the cerebellum were not altered. Levels of d-alanine, l-alanine, l-aspartic acid, taurine, asparagine, arginine, threonine, γ-amino butyric acid (GABA) and methionine, remained the same in frontal cortex, hippocampus, striatum and cerebellum of WT and mutant mice. Furthermore, no differences in d-aspartate oxidase (DDO) activity were detected in the forebrains of WT and Srr-KO mice. These results suggest that SRR and/or d-serine may be involved in the production of d-aspartic acid in mouse forebrains, although further detailed studies will be necessary to confirm this finding. Copyright © 2013 Elsevier Ltd. All rights reserved.

  2. 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

  3. Piroxicam treatment augments bone abnormalities in interleukin-10 knockout mice.

    Science.gov (United States)

    Holgersen, Kristine; Dobie, Ross; Farquharson, Colin; vanʼt Hof, Rob; Ahmed, Syed Faisal; Hansen, Axel Kornerup; Holm, Thomas L

    2015-02-01

    Osteoporosis and fractures are common complications of inflammatory bowel disease. The pathogenesis is multifactorial and has been partly attributed to intestinal inflammation. The aim of this study was to evaluate bone status and assess the association between bone loss and gut inflammation in an experimental colitis model. Colitis was induced in interleukin-10 knockout mice (PAC IL-10 k.o.) by peroral administration of piroxicam for 12 days. The degree of colitis was assessed by clinical, macroscopic, and microscopic evaluation. Trabecular and cortical bone microarchitecture of tibia were determined using micro-computed tomography. Moreover, the serum levels of bone formation and bone resorption biomarkers were measured, and inflammatory protein profiling was performed on colons. PAC IL-10 k.o. mice developed severe colitis, characterized by hyperplasia and focal transmural inflammation, which was consistent with Crohn's disease-like pathology. The gut inflammation was accompanied by a 14% and 12% reduction in trabecular thickness relative to piroxicam-treated wild type and untreated wild type mice, respectively (P < 0.001). The trabecular bone structure was also changed in PAC IL-10 k.o. mice, whereas no differences in cortical bone geometry were observed. The trabecular thickness was inversely correlated with serum levels of CTX (r = -0.93, P = 0.006). Moreover, numerous inflammatory mediators, including RANKL and osteoprotegerin, were significantly increased in the colon of PAC IL-10 k.o. mice. PAC IL-10 k.o. mice develop bone loss and changed trabecular structure, as a result of increased bone resorption. Thus, the PAC IL-10 k.o. model could be a useful experimental model in preclinical research of inflammatory bowel disease-associated bone loss.

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

    Directory of Open Access Journals (Sweden)

    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.

  5. EDF - 2012 full-year results: Strong results and commitments delivered. Confirmation of the solidity of EDF's integrated and diversified business model. Annual results 2012. Electricite de France S.A.: Statutory Auditors' Report on the consolidated financial statements. Management report 2012

    International Nuclear Information System (INIS)

    2013-01-01

    As the world's biggest electricity generator, the EDF Group covers every sector of expertise, from generation to trading and transmission grids. EDF builds on the expertise of its people, its R and D and engineering skills, its experience as a leading industry operator and the attentive support of its customers to deliver competitive solutions that successfully reconcile economic growth with climate protection. This document presents the 2012 annual results, management report and Consolidated financial statements of the Group at 31 December 2012: EBITDA: euro 16.1 billion, +7.7% of which 4.6% organic growth; Net income excluding non-recurring items: euro 4.2 billion, +16.9%; Net income - Group share: euro 3.3 billion, + 5.3%; Allocation in 2013 of the CSPE receivable to dedicated assets, bringing coverage of eligible provisions to 100% starting as early as 2013; Net financial debt/EBITDA: 2.4x2; Proposed dividend of euro 1.25/share for the 20123 financial year, i.e. a pay-out ratio of 55% of net income excluding non-recurring items. 2013: a decisive year: Launch of 'Spark': plan targeting savings of euro 1 billion as soon as 2013; EBITDA: between 0% and 3% in organic growth excluding Edison; Edison: expectation for recurring EBITDA in line with 2012, with fluctuation in results possible in 2013-2014 linked to calendar effect from the re-negotiation of gas supply contracts; Net investments stable at euro 12 billion; Net financial debt/EBITDA: between 2x and 2.5x; Pay-out ratio: between 55% and 65% of net income excluding non-recurring items. Consolidated Financial Statements: Group accounting standards; comparability; significant events and transactions; regulatory events in France; changes in the scope of consolidation; segment reporting; sales; fuel and energy purchases; other external expenses; personnel expenses; taxes other than income taxes; other operating income and expenses; impairment / reversals; other income and expenses; financial result; income taxes

  6. 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.

  7. Acute multi-sgRNA knockdown of KEOPS complex genes reproduces the microcephaly phenotype of the stable knockout zebrafish model.

    Directory of Open Access Journals (Sweden)

    Tilman Jobst-Schwan

    Full Text Available Until recently, morpholino oligonucleotides have been widely employed in zebrafish as an acute and efficient loss-of-function assay. However, off-target effects and reproducibility issues when compared to stable knockout lines have compromised their further use. Here we employed an acute CRISPR/Cas approach using multiple single guide RNAs targeting simultaneously different positions in two exemplar genes (osgep or tprkb to increase the likelihood of generating mutations on both alleles in the injected F0 generation and to achieve a similar effect as morpholinos but with the reproducibility of stable lines. This multi single guide RNA approach resulted in median likelihoods for at least one mutation on each allele of >99% and sgRNA specific insertion/deletion profiles as revealed by deep-sequencing. Immunoblot showed a significant reduction for Osgep and Tprkb proteins. For both genes, the acute multi-sgRNA knockout recapitulated the microcephaly phenotype and reduction in survival that we observed previously in stable knockout lines, though milder in the acute multi-sgRNA knockout. Finally, we quantify the degree of mutagenesis by deep sequencing, and provide a mathematical model to quantitate the chance for a biallelic loss-of-function mutation. Our findings can be generalized to acute and stable CRISPR/Cas targeting for any zebrafish gene of interest.

  8. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  10. Human Genetic Disorders and Knockout Mice Deficient in Glycosaminoglycan

    Directory of Open Access Journals (Sweden)

    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.

  11. 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.

  12. Selective Attention to Visual Stimuli Using Auditory Distractors Is Altered in Alpha-9 Nicotinic Receptor Subunit Knock-Out Mice.

    Science.gov (United States)

    Terreros, Gonzalo; Jorratt, Pascal; Aedo, Cristian; Elgoyhen, Ana Belén; Delano, Paul H

    2016-07-06

    During selective attention, subjects voluntarily focus their cognitive resources on a specific stimulus while ignoring others. Top-down filtering of peripheral sensory responses by higher structures of the brain has been proposed as one of the mechanisms responsible for selective attention. A prerequisite to accomplish top-down modulation of the activity of peripheral structures is the presence of corticofugal pathways. The mammalian auditory efferent system is a unique neural network that originates in the auditory cortex and projects to the cochlear receptor through the olivocochlear bundle, and it has been proposed to function as a top-down filter of peripheral auditory responses during attention to cross-modal stimuli. However, to date, there is no conclusive evidence of the involvement of olivocochlear neurons in selective attention paradigms. Here, we trained wild-type and α-9 nicotinic receptor subunit knock-out (KO) mice, which lack cholinergic transmission between medial olivocochlear neurons and outer hair cells, in a two-choice visual discrimination task and studied the behavioral consequences of adding different types of auditory distractors. In addition, we evaluated the effects of contralateral noise on auditory nerve responses as a measure of the individual strength of the olivocochlear reflex. We demonstrate that KO mice have a reduced olivocochlear reflex strength and perform poorly in a visual selective attention paradigm. These results confirm that an intact medial olivocochlear transmission aids in ignoring auditory distraction during selective attention to visual stimuli. The auditory efferent system is a neural network that originates in the auditory cortex and projects to the cochlear receptor through the olivocochlear system. It has been proposed to function as a top-down filter of peripheral auditory responses during attention to cross-modal stimuli. However, to date, there is no conclusive evidence of the involvement of olivocochlear

  13. 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...

  14. A female Viking warrior confirmed by genomics.

    Science.gov (United States)

    Hedenstierna-Jonson, Charlotte; Kjellström, Anna; Zachrisson, Torun; Krzewińska, Maja; Sobrado, Veronica; Price, Neil; Günther, Torsten; Jakobsson, Mattias; Götherström, Anders; Storå, Jan

    2017-12-01

    The objective of this study has been to confirm the sex and the affinity of an individual buried in a well-furnished warrior grave (Bj 581) in the Viking Age town of Birka, Sweden. Previously, based on the material and historical records, the male sex has been associated with the gender of the warrior and such was the case with Bj 581. An earlier osteological classification of the individual as female was considered controversial in a historical and archaeological context. A genomic confirmation of the biological sex of the individual was considered necessary to solve the issue. Genome-wide sequence data was generated in order to confirm the biological sex, to support skeletal integrity, and to investigate the genetic relationship of the individual to ancient individuals as well as modern-day groups. Additionally, a strontium isotope analysis was conducted to highlight the mobility of the individual. The genomic results revealed the lack of a Y-chromosome and thus a female biological sex, and the mtDNA analyses support a single-individual origin of sampled elements. The genetic affinity is close to present-day North Europeans, and within Sweden to the southern and south-central region. Nevertheless, the Sr values are not conclusive as to whether she was of local or nonlocal origin. The identification of a female Viking warrior provides a unique insight into the Viking society, social constructions, and exceptions to the norm in the Viking time-period. The results call for caution against generalizations regarding social orders in past societies. © 2017 The Authors American Journal of Physical Anthropology Published by Wiley Periodicals, Inc.

  15. 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.

  16. Current issues in electromagnetic knockout reactions

    International Nuclear Information System (INIS)

    Giusti, C.

    2002-01-01

    The role of correlations and two-body currents in one-nucleon emission reactions induced by an electromagnetic probe is discussed. The theoretical framework for cross section calculations is outlined and some results are presented for the exclusive "1"6O(e,e'p) "1"5N and "1"6O(γ, p) "1"5N reactions in relativistic and nonrelativistic models. A reliable and consistent evaluation of different types of correlations is needed to determine the spectroscopic factors. A consistent description of (e,e'p) and (γ, p) data, with the same spectroscopic factors, can be obtained when meson-exchange currents (MEC) are included in the theoretical model. MEC give an important contribution to (γ, p) cross sections and have only a small effect in (e, e'p). In the relativistic model the two-body seagull current affects the calculated cross sections less than in the nonrelativistic calculations.

  17. results

    Directory of Open Access Journals (Sweden)

    Salabura Piotr

    2017-01-01

    Full Text Available HADES experiment at GSI is the only high precision experiment probing nuclear matter in the beam energy range of a few AGeV. Pion, proton and ion beams are used to study rare dielectron and strangeness probes to diagnose properties of strongly interacting matter in this energy regime. Selected results from p + A and A + A collisions are presented and discussed.

  18. Behavioral and electrophysiological characterization of Dyt1 heterozygous knockout mice.

    Science.gov (United States)

    Yokoi, Fumiaki; Chen, Huan-Xin; Dang, Mai Tu; Cheetham, Chad C; Campbell, Susan L; Roper, Steven N; Sweatt, J David; Li, Yuqing

    2015-01-01

    DYT1 dystonia is an inherited movement disorder caused by mutations in DYT1 (TOR1A), which codes for torsinA. Most of the patients have a trinucleotide deletion (ΔGAG) corresponding to a glutamic acid in the C-terminal region (torsinA(ΔE)). Dyt1 ΔGAG heterozygous knock-in (KI) mice, which mimic ΔGAG mutation in the endogenous gene, exhibit motor deficits and deceased frequency of spontaneous excitatory post-synaptic currents (sEPSCs) and normal theta-burst-induced long-term potentiation (LTP) in the hippocampal CA1 region. Although Dyt1 KI mice show decreased hippocampal torsinA levels, it is not clear whether the decreased torsinA level itself affects the synaptic plasticity or torsinA(ΔE) does it. To analyze the effect of partial torsinA loss on motor behaviors and synaptic transmission, Dyt1 heterozygous knock-out (KO) mice were examined as a model of a frame-shift DYT1 mutation in patients. Consistent with Dyt1 KI mice, Dyt1 heterozygous KO mice showed motor deficits in the beam-walking test. Dyt1 heterozygous KO mice showed decreased hippocampal torsinA levels lower than those in Dyt1 KI mice. Reduced sEPSCs and normal miniature excitatory post-synaptic currents (mEPSCs) were also observed in the acute hippocampal brain slices from Dyt1 heterozygous KO mice, suggesting that the partial loss of torsinA function in Dyt1 KI mice causes action potential-dependent neurotransmitter release deficits. On the other hand, Dyt1 heterozygous KO mice showed enhanced hippocampal LTP, normal input-output relations and paired pulse ratios in the extracellular field recordings. The results suggest that maintaining an appropriate torsinA level is important to sustain normal motor performance, synaptic transmission and plasticity. Developing therapeutics to restore a normal torsinA level may help to prevent and treat the symptoms in DYT1 dystonia.

  19. Behavioral and electrophysiological characterization of Dyt1 heterozygous knockout mice.

    Directory of Open Access Journals (Sweden)

    Fumiaki Yokoi

    Full Text Available DYT1 dystonia is an inherited movement disorder caused by mutations in DYT1 (TOR1A, which codes for torsinA. Most of the patients have a trinucleotide deletion (ΔGAG corresponding to a glutamic acid in the C-terminal region (torsinA(ΔE. Dyt1 ΔGAG heterozygous knock-in (KI mice, which mimic ΔGAG mutation in the endogenous gene, exhibit motor deficits and deceased frequency of spontaneous excitatory post-synaptic currents (sEPSCs and normal theta-burst-induced long-term potentiation (LTP in the hippocampal CA1 region. Although Dyt1 KI mice show decreased hippocampal torsinA levels, it is not clear whether the decreased torsinA level itself affects the synaptic plasticity or torsinA(ΔE does it. To analyze the effect of partial torsinA loss on motor behaviors and synaptic transmission, Dyt1 heterozygous knock-out (KO mice were examined as a model of a frame-shift DYT1 mutation in patients. Consistent with Dyt1 KI mice, Dyt1 heterozygous KO mice showed motor deficits in the beam-walking test. Dyt1 heterozygous KO mice showed decreased hippocampal torsinA levels lower than those in Dyt1 KI mice. Reduced sEPSCs and normal miniature excitatory post-synaptic currents (mEPSCs were also observed in the acute hippocampal brain slices from Dyt1 heterozygous KO mice, suggesting that the partial loss of torsinA function in Dyt1 KI mice causes action potential-dependent neurotransmitter release deficits. On the other hand, Dyt1 heterozygous KO mice showed enhanced hippocampal LTP, normal input-output relations and paired pulse ratios in the extracellular field recordings. The results suggest that maintaining an appropriate torsinA level is important to sustain normal motor performance, synaptic transmission and plasticity. Developing therapeutics to restore a normal torsinA level may help to prevent and treat the symptoms in DYT1 dystonia.

  20. Sildenafil restores endothelial function in the apolipoprotein E knockout mouse

    Directory of Open Access Journals (Sweden)

    Balarini Camille M

    2013-01-01

    Full Text Available Abstract Background Atherosclerosis is an inflammatory process of the arterial walls and is initiated by endothelial dysfunction accompanied by an imbalance in the production of reactive oxygen species (ROS and nitric oxide (NO. Sildenafil, a selective phosphodiesterase-5 (PDE5 inhibitor used for erectile dysfunction, exerts its cardiovascular effects by enhancing the effects of NO. The aim of this study was to investigate the influence of sildenafil on endothelial function and atherosclerosis progression in apolipoprotein E knockout (apoE−/− mice. Methods ApoE−/− mice treated with sildenafil (Viagra®, 40 mg/kg/day, for 3 weeks, by oral gavage were compared to the untreated apoE−/− and the wild-type (WT mice. Aortic rings were used to evaluate the relaxation responses to acetylcholine (ACh in all of the groups. In a separate set of experiments, the roles of NO and ROS in the relaxation response to ACh were evaluated by incubating the aortic rings with L-NAME (NO synthase inhibitor or apocynin (NADPH oxidase inhibitor. In addition, the atherosclerotic lesions were quantified and superoxide production was assessed. Results Sildenafil restored the vasodilator response to acetylcholine (ACh in the aortic rings of the apoE−/− mice. Treatment with L-NAME abolished the vasodilator responses to ACh in all three groups of mice and revealed an augmented participation of NO in the endothelium-dependent vasodilation in the sildenafil-treated animals. The normalized endothelial function in sildenafil-treated apoE−/− mice was unaffected by apocynin highlighting the low levels of ROS production in these animals. Moreover, morphological analysis showed that sildenafil treatment caused approximately a 40% decrease in plaque deposition in the aorta. Conclusion This is the first study demonstrating the beneficial effects of chronic treatment with sildenafil on endothelial dysfunction and atherosclerosis in a model of spontaneous

  1. 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

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

    Directory of Open Access Journals (Sweden)

    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.

  3. 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

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Science.gov (United States)

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

    2001-01-01

    VMAT2 expression in wild-type and DAT knockout midbrain. DAT immunoreactivity in wild-type (A) and DAT knockout midbrain (B). VMAT2 immunoreactivity in wild-type (C) and DAT knockout midbrain (D). Robust immunoreactivity was observed in the ventral tegmental area and substantia nigra pars compacta and reticulata in the wild-type brain. Note absence of DAT immunoreactivity and modest reduction of VMAT2 immunoreactivity in the DAT knockout. Fig. 3. Characterization of VMAT2 gene disruption. (A) Southern blot analysis of mouse genomic DNA. The Southern blot was prepared with 15 μg of genomic DNA per lane and probed with a 1.4-kb 3' external genomic fragment. +/+, wild type littermates; +/-, heterozygote; -/-, homozygote. (B) RT-PCR analysis of mouse brain poly(A)+ RNA. For each reverse transcription assay, 0.5 μg of poly(A)+ RNA was used. Equal volumes of cDNA templates were used for each PCR assay. The PCR primers used flank the neomycin cassette for the purpose of detecting potential readthrough of the neomycin DNA. The heterozygote has a reduced amount of transcripts compared with the wild-type littermate; the homozygote is devoid of VMAT2 transcripts. G3PDH was used as internal control. (C) Western blot analysis of wholebrain synaptic vesicles. Samples (25 μg) of vesicles were solubilized and separated by SDS-PAGE, transferred to nitrocellulose, subjected to Western blot analysis with anti-VMAT2-Ct (top) or anti-a-tubulin (bottom) antibodies, and developed with chemiluminescence. Molecular mass markers (kDa) are shown to the left. To confirm equal loading and transfer of proteins, the blots were stripped and reprobed with an antibody to α-tubulin. (Reproduced with permission from ref. 1). The importance of DAT in neuronal function is highlighted in animals in which DAT has been genetically deleted (DAT KO) (3). In the homozygote DAT KO mice, released dopamine remains in the extracellular space up to 300 times longer than normal. As expected, these animals display

  6. 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.

  7. Calibration and Confirmation in Geophysical Models

    Science.gov (United States)

    Werndl, Charlotte

    2016-04-01

    For policy decisions the best geophysical models are needed. To evaluate geophysical models, it is essential that the best available methods for confirmation are used. A hotly debated issue on confirmation in climate science (as well as in philosophy) is the requirement of use-novelty (i.e. that data can only confirm models if they have not already been used before. This talk investigates the issue of use-novelty and double-counting for geophysical models. We will see that the conclusions depend on the framework of confirmation and that it is not clear that use-novelty is a valid requirement and that double-counting is illegitimate.

  8. 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

  9. 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

  10. 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.

  11. 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.)

  12. 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...

  13. Single-step generation of gene knockout-rescue system in pluripotent stem cells by promoter insertion with CRISPR/Cas9.

    Science.gov (United States)

    Matsunaga, Taichi; Yamashita, Jun K

    2014-02-07

    Specific gene knockout and rescue experiments are powerful tools in developmental and stem cell biology. Nevertheless, the experiments require multiple steps of molecular manipulation for gene knockout and subsequent rescue procedures. Here we report an efficient and single step strategy to generate gene knockout-rescue system in pluripotent stem cells by promoter insertion with CRISPR/Cas9 genome editing technology. We inserted a tetracycline-regulated inducible gene promoter (tet-OFF/TRE-CMV) upstream of the endogenous promoter region of vascular endothelial growth factor receptor 2 (VEGFR2/Flk1) gene, an essential gene for endothelial cell (EC) differentiation, in mouse embryonic stem cells (ESCs) with homologous recombination. Both homo- and hetero-inserted clones were efficiently obtained through a simple selection with a drug-resistant gene. The insertion of TRE-CMV promoter disrupted endogenous Flk1 expression, resulting in null mutation in homo-inserted clones. When the inserted TRE-CMV promoter was activated with doxycycline (Dox) depletion, Flk1 expression was sufficiently recovered from the downstream genomic Flk1 gene. Whereas EC differentiation was almost completely perturbed in homo-inserted clones, Flk1 rescue with TRE-CMV promoter activation restored EC appearance, indicating that phenotypic changes in EC differentiation can be successfully reproduced with this knockout-rescue system. Thus, this promoter insertion strategy with CRISPR/Cas9 would be a novel attractive method for knockout-rescue experiments. Copyright © 2014 Elsevier Inc. All rights reserved.

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

    Directory of Open Access Journals (Sweden)

    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.

  15. 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.

  16. Study of 19C by One-Neutron Knockout

    Directory of Open Access Journals (Sweden)

    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.

  17. 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...

  18. 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....

  19. Development of the Performance Confirmation Program at Yucca Mountain, Nevada

    International Nuclear Information System (INIS)

    G.D. LeCain; D. Barr; D. Weaver; R. Snell; S.W. Goodin; F.D. Hansen

    2006-01-01

    The Yucca Mountain Performance Confirmation program consists of tests, monitoring activities, experiments, and analyses to evaluate the adequacy of assumptions, data, and analyses that form the basis of the conceptual and numerical models of flow and transport associated with a proposed radioactive waste repository at Yucca Mountain, Nevada. The Performance Confirmation program uses an eight-stage risk-informed, performance-based approach. Selection of the Performance Confirmation activities (a parameter and a test method) for inclusion in the Performance Confirmation program was done using a risk-informed performance-based decision analysis. The result of this analysis and review was a Performance Confirmation base portfolio that consists of 20 activities. The 20 Performance Confirmation activities include geologic, hydrologic, and construction/engineering testing. Several of the activities were initiated during site characterization and are ongoing. Others activities will commence during construction and/or post emplacement and will continue until repository closure

  20. 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

  1. 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.

  2. 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.

  3. 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

  4. 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.

  5. 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

  6. 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.

  7. 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.

  8. Screening tests for Chlamydia trachomatis or Neisseria gonorrhoeae using the cobas 4800 PCR system do not require a second test to confirm: an audit of patients issued with equivocal results at a sexual health clinic in the Northwest of England, U.K.

    Science.gov (United States)

    Hopkins, Mark J; Smith, Godfrey; Hart, Ian J; Alloba, Fath

    2012-11-01

    To assess the clinical utility of supplementary PCRs following a positive cobas 4800 CT/NG PCR screening test result. Laboratory reports, for Chlamydia trachomatis or Neisseria gonorrhoeae, issued to genitourinary medicine patients between April 2010 and April 2011 were reviewed retrospectively. Positive reports were routinely confirmed by supplementary PCRs and N gonorrhoeae culture. Clinical records of patients with unconfirmed positive (equivocal) reports were retrieved to determine if the infection was confirmed by a second sample obtained at patient recall and the impact of this process on antibiotic management. Over 15 000 patients were tested during the study period. The prevalence of chlamydia and gonorrhoea was 972 (5.75%) and 76 (0.50%), respectively. A further 78 chlamydia and 2 gonorrhoea equivocal reports were issued. Only 56 (72%) patients with an equivocal chlamydia report returned to the clinic, and of these, only 41 (73%) gave a second sample to retest. Positive predictive value (PPV) of the PCR screening test was calculated at 98.0% and 97.5% for detection of chlamydia infection from urine and rectal swabs, respectively. Most patients accepted antibiotic treatment before their infection status had been confirmed. Prevalence of gonorrhoea infection was low but the PPV of the screening PCR in urine specimens remained high (98.75%). Equivocal reports introduce delays to patient management, while the risk of unnecessary antibiotic therapy appears acceptable to most patients. The cobas 4800 CT/NG PCR screening assay can achieve UK testing standards (PPV >90%) for chlamydia, and low prevalence gonorrhoea in urine without supplementary tests. A patient-led confirmation algorithm is proposed.

  9. Magnetic resonance tomography in confirmed multiple sclerosis

    International Nuclear Information System (INIS)

    Uhlenbrock, D.; Dickmann, E.; Beyer, H.K.; Gehlen, W.; Josef-Hospital, Bochum; Knappschafts-Krankenhaus Bochum

    1985-01-01

    The authors report on 21 cases of confirmed multiple sclerosis examined by both CT and magnetic resonance tomography. To safeguard the results, strict criteria were applied in accordance with the suggestions made by neurological work teams. Pathological lesons were seen in 20 patients; the MR image did not reveal anything abnormal in one case. On the average, 10.3 lesions were seen in the MR tomogram, whereas CT images showed on the average only 2.1 foci. The size and number of lesions in the MR tomogram were independent of the duration of the disease, the presented clinical symptoms, or the type of treatment at the time of examination. Evidently the sensitivity of MR tomography is very high in MS patients, but it has not yet been clarified to what extent this applies also to the specificity. Further research is mandatory. First experiences made by us show that lesions of a similar kind can also occur in diseases such as malignant lymphoma involving the brain, in vitamin B 12 deficiency syndrome, or encephalitis, and can become manifest in the MR tomogram. (orig.) [de

  10. Confirmation of Essure placement using transvaginal ultrasound

    NARCIS (Netherlands)

    Veersema, Sebastiaan; Vleugels, Michel; Koks, Caroline; Thurkow, Andreas; van der Vaart, Huub; Brölmann, Hans

    2011-01-01

    To evaluate the protocol for confirmation of satisfactory Essure placement using transvaginal ultrasound. Prospective multicenter cohort study (Canadian Task Force classification II-2). Outpatient departments of 4 teaching hospitals in the Netherlands. Eleven hundred forty-five women who underwent

  11. escherichia coli serotypes confirmed in experimental mammary ...

    African Journals Online (AJOL)

    DJFLEX

    VARIATIONS IN VIRULENCE OF THREE (3) ESCHERICHIA COLI. SEROTYPES CONFIRMED IN ... ows are susceptible to E. coli infection because. E. coli exist in the .... Coli infections in mice: A laboratory animal model for research in.

  12. Experience with confirmation measurement at Los Alamos

    International Nuclear Information System (INIS)

    Marshall, R.S.; Wagner, R.P.; Hsue, F.

    1985-01-01

    Confirmation measurements are used at Los Alamos in support of incoming and outgoing shipment accountibility and for support of both at 235 U and Pu inventories. Statistical data are presented to show the consistency of measurements on items of identical composition and on items measured at two facilitis using similar instruments. A description of confirmation measurement techniques used in support of 235 U and Pu inventories and a discussion on the ability of the measurements to identify items with misstated SNM are given

  13. Experience with confirmation measurement at Los Alamos

    International Nuclear Information System (INIS)

    Marshall, R.S.; Wagner, R.P.

    1985-01-01

    Confirmation measurements are used at Los Alamos in support of incoming and outgoing shipment accountability and for support of both 235 U and Pu inventories. Statistical data are presented to show the consistency of measurements on items of identical composition and on items measured at two facilities using similar instruments. A description of confirmation measurement techniques used in support of 235 U and Pu inventories and a discussion on the ability of the measurements to identify items with misstated SNM are given

  14. 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.

  15. 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.

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

    Science.gov (United States)

    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.

  17. 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

  18. 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.

  19. Spectroscopy of 17C via one-neutron knockout reaction

    Directory of Open Access Journals (Sweden)

    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+.

  20. 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....

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

    Science.gov (United States)

    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.

  2. 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...

  3. 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.

  4. The α-fetoprotein knock-out mouse model suggests that parental behavior is sexually differentiated under the influence of prenatal estradiol

    Science.gov (United States)

    Keller, Matthieu; Pawluski, Jodi L.; Brock, Olivier; Douhard, Quentin; Bakker, Julie

    2010-01-01

    In rodent species, sexual differentiation of the brain for many reproductive processes depends largely on estradiol. This was recently confirmed again by using the α-fetoprotein knockout (AFP-KO) mouse model, which lacks the protective actions of α-fetoprotein against maternal estradiol and as a result represents a good model to determine the contribution of prenatal estradiol to the sexual differentiation of the brain and behavior. Female AFP-KO mice were defeminized and masculinized with regard to their neuroendocrine responses as well as sexual behavior. Since parental behavior is also strongly sexually differentiated in mice, we used the AFP-KO mouse model here to ask whether parental responses are differentiated prenatally under the influence of estradiol. It was found that AFP-KO females showed longer latencies to retrieve pups to the nest and also exhibited lower levels of crouching over the pups in the nest in comparison to WT females. In fact, they resembled males (WT and AFP-KO). Other measures of maternal behavior, for example the incidence of infanticide, tended to be higher in AFP-KO females than in WT females but this increase failed to reach statistical significance. The deficits observed in parental behavior of AFP-KO females could not be explained by any changes in olfactory function, novelty recognition or anxiety. Thus our results suggest that prenatal estradiol defeminizes the parental brain in mice. PMID:20109458

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

    Science.gov (United States)

    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

  6. Nonintrusive irradiated fuel inventory confirmation technique

    International Nuclear Information System (INIS)

    Dowdy, E.J.; Nicholson, N.; Caldwell, J.T.

    1980-01-01

    Successful tests showing correlation between the intensity of the Cerenkov glow surrounding irradiated fuel assemblies in water-filled spent fuel storage ponds and the exposure and cooling times of assemblies have been concluded. Fieldable instruments used in subsequent tests confirmed that such measurements can be made easily and rapidly, without fuel assembly movement or the introduction of apparatus into the storage ponds

  7. Confirmation of Essure placement using transvaginal ultrasound.

    Science.gov (United States)

    Veersema, Sebastiaan; Vleugels, Michel; Koks, Caroline; Thurkow, Andreas; van der Vaart, Huub; Brölmann, Hans

    2011-01-01

    To evaluate the protocol for confirmation of satisfactory Essure placement using transvaginal ultrasound. Prospective multicenter cohort study (Canadian Task Force classification II-2). Outpatient departments of 4 teaching hospitals in the Netherlands. Eleven hundred forty-five women who underwent hysteroscopic sterilization using the Essure device between March 2005 and December 2007. Transvaginal ultrasound examination 12 weeks after uncomplicated successful bilateral placement or as indicated according to the transvaginal ultrasound protocol after 4 weeks, and hysterosalpingography (HSG) at 12 weeks to confirm correct placement of the device after 3 months. The rate of successful placement was 88.4% initially. In 164 women (15%), successful placement was confirmed at HSG according the protocol. In 9 patients (0.84%), incorrect position of the device was observed at HSG. The cumulative pregnancy rate after 18 months was 3.85 per thousand women. Transvaginal ultrasound should be the first diagnostic test used to confirm the adequacy of hysteroscopic Essure sterilization because it is minimally invasive, averts ionizing radiation, and does not decrease the effectiveness of the Essure procedure. Copyright © 2011 AAGL. Published by Elsevier Inc. All rights reserved.

  8. 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...

  9. 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

  10. 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)

  11. 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.

  12. Zika virus infection of adult and fetal STAT2 knock-out hamsters.

    Science.gov (United States)

    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.

  13. 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.

  14. Manned in Situ Confirmation of Lunar Ice

    Science.gov (United States)

    Gerené, S. P. B.; Hummeling, R. W. J.; Ockels, W. J.

    A study is performed to investigate the feasibility of a manned expedition to the Moon using the European Ariane-5 launcher. The primary objective of this lunar mission is to confirm the presence of water at the South-Pole craters. It is believed that these permanently shadowed craters contain water in the form of ice. Secondary objective is to perform lunar surface science and making a first step towards a lunar outpost. Early results show that a minimum of two Ariane-5 launches is required. In this `two Ariane' scenario the first launch will bring a Lunar Landing Vehicle (LLV) into low lunar orbit. The second will launch two astronauts in a Crew Transfer Vehicle into a rendez- vous trajectory with the LLV. Arrived at the Moon, the astronauts will enter the LLV, undock from the CTV and land at the designated site located near the rim of the South-Pole Shackleton crater. The transfer strategy for both spacecraft will be the so-called direct transfer, taking about four days. At arrival the LLV will start mapping the landing site at a ground resolution of one meter. As a consequence of the polar orbit, the CTV has to arrive fourteen days later and surface operations can take about twelve days, accumulating in a total mission-duration of 36 days. 32 days for the CTV and 22 days for the LLV. In case a `two Ariane' flight does not posses sufficient capabilities also a `three Ariane' scenario is developed, in which the LLV is split-up into two stages and launched separately. These two will dock at the Moon forming a descent stage and an ascent stage. The third launch will be a CTV. During surface operations, astronauts will set up a solar power unit, install the sample retrieval system and carry out surface science. Samples of the crater floor will be retrieved by means of a probe or robot guided along a cable suspended over the crater rim. Also, this paper shows the way in which European astronauts can be brought to the Moon for other future missions, like the

  15. 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.

  16. Tensor-based morphometry and stereology reveal brain pathology in the complexin1 knockout mouse.

    Science.gov (United States)

    Kielar, Catherine; Sawiak, Stephen J; Navarro Negredo, Paloma; Tse, Desmond H Y; Morton, A Jennifer

    2012-01-01

    Complexins (Cplxs) are small, soluble, regulatory proteins that bind reversibly to the SNARE complex and modulate synaptic vesicle release. Cplx1 knockout mice (Cplx1(-/-)) have the earliest known onset of ataxia seen in a mouse model, although hitherto no histopathology has been described in these mice. Nevertheless, the profound neurological phenotype displayed by Cplx1(-/-) mutants suggests that significant functional abnormalities must be present in these animals. In this study, MRI was used to automatically detect regions where structural differences were not obvious when using a traditional histological approach. Tensor-based morphometry of Cplx1(-/-) mouse brains showed selective volume loss from the thalamus and cerebellum. Stereological analysis of Cplx1(-/-) and Cplx1(+/+) mice brain slices confirmed the volume loss in the thalamus as well as loss in some lobules of the cerebellum. Finally, stereology was used to show that there was loss of cerebellar granule cells in Cplx1(-/-) mice when compared to Cplx1(+/+) animals. Our study is the first to describe pathological changes in Cplx1(-/-) mouse brain. We suggest that the ataxia in Cplx1(-/-) mice is likely to be due to pathological changes in both cerebellum and thalamus. Reduced levels of Cplx proteins have been reported in brains of patients with neurodegenerative diseases. Therefore, understanding the effects of Cplx depletion in brains from Cplx1(-/-) mice may also shed light on the mechanisms underlying pathophysiology in disorders in which loss of Cplx1 occurs.

  17. Organelle and cellular abnormalities associated with hippocampal heterotopia in neonatal doublecortin knockout mice.

    Directory of Open Access Journals (Sweden)

    Reham Khalaf-Nazzal

    Full Text Available Heterotopic or aberrantly positioned cortical neurons are associated with epilepsy and intellectual disability. Various mouse models exist with forms of heterotopia, but the composition and state of cells developing in heterotopic bands has been little studied. Dcx knockout (KO mice show hippocampal CA3 pyramidal cell lamination abnormalities, appearing from the age of E17.5, and mice suffer from spontaneous epilepsy. The Dcx KO CA3 region is organized in two distinct pyramidal cell layers, resembling a heterotopic situation, and exhibits hyperexcitability. Here, we characterized the abnormally organized cells in postnatal mouse brains. Electron microscopy confirmed that the Dcx KO CA3 layers at postnatal day (P 0 are distinct and separated by an intermediate layer devoid of neuronal somata. We found that organization and cytoplasm content of pyramidal neurons in each layer were altered compared to wild type (WT cells. Less regular nuclei and differences in mitochondria and Golgi apparatuses were identified. Each Dcx KO CA3 layer at P0 contained pyramidal neurons but also other closely apposed cells, displaying different morphologies. Quantitative PCR and immunodetections revealed increased numbers of oligodendrocyte precursor cells (OPCs and interneurons in close proximity to Dcx KO pyramidal cells. Immunohistochemistry experiments also showed that caspase-3 dependent cell death was increased in the CA1 and CA3 regions of Dcx KO hippocampi at P2. Thus, unsuspected ultrastructural abnormalities and cellular heterogeneity may lead to abnormal neuronal function and survival in this model, which together may contribute to the development of hyperexcitability.

  18. Hyperactivity and lack of social discrimination in the adolescent Fmr1 knockout mouse.

    Science.gov (United States)

    Sørensen, Emilie M; Bertelsen, Freja; Weikop, Pia; Skovborg, Maria M; Banke, Tue; Drasbek, Kim R; Scheel-Krüger, Jørgen

    2015-12-01

    The aims of this study were to investigate behaviour relevant to human autism spectrum disorder (ASD) and the fragile X syndrome in adolescent Fmr1 knockout (KO) mice and to evaluate the tissue levels of striatal monoamines. Fmr1 KO mice were evaluated in the open field, marble burying and three-chamber test for the presence of hyperactivity, anxiety, repetitive behaviour, sociability and observation of social novelty compared with wild-type (WT) mice. The Fmr1 KO mice expressed anxiety and hyperactivity in the open field compared with WT mice. This increased level of hyperactivity was confirmed in the three-chamber test. Fmr1 KO mice spent more time with stranger mice compared with the WT. However, after a correction for hyperactivity, their apparent increase in sociability became identical to that of the WT. Furthermore, the Fmr1 KO mice could not differentiate between a familiar or a novel mouse. Monoamines were measured by HPLC: Fmr1 KO mice showed an increase in the striatal dopamine level. We conclude that the fragile X syndrome model seems to be useful for understanding certain aspects of ASD and may have translational interest for studies of social behaviour when hyperactivity coexists in ASD patients.

  19. Tensor-based morphometry and stereology reveal brain pathology in the complexin1 knockout mouse.

    Directory of Open Access Journals (Sweden)

    Catherine Kielar

    Full Text Available Complexins (Cplxs are small, soluble, regulatory proteins that bind reversibly to the SNARE complex and modulate synaptic vesicle release. Cplx1 knockout mice (Cplx1(-/- have the earliest known onset of ataxia seen in a mouse model, although hitherto no histopathology has been described in these mice. Nevertheless, the profound neurological phenotype displayed by Cplx1(-/- mutants suggests that significant functional abnormalities must be present in these animals. In this study, MRI was used to automatically detect regions where structural differences were not obvious when using a traditional histological approach. Tensor-based morphometry of Cplx1(-/- mouse brains showed selective volume loss from the thalamus and cerebellum. Stereological analysis of Cplx1(-/- and Cplx1(+/+ mice brain slices confirmed the volume loss in the thalamus as well as loss in some lobules of the cerebellum. Finally, stereology was used to show that there was loss of cerebellar granule cells in Cplx1(-/- mice when compared to Cplx1(+/+ animals. Our study is the first to describe pathological changes in Cplx1(-/- mouse brain. We suggest that the ataxia in Cplx1(-/- mice is likely to be due to pathological changes in both cerebellum and thalamus. Reduced levels of Cplx proteins have been reported in brains of patients with neurodegenerative diseases. Therefore, understanding the effects of Cplx depletion in brains from Cplx1(-/- mice may also shed light on the mechanisms underlying pathophysiology in disorders in which loss of Cplx1 occurs.

  20. PERFORMANCE CONFIRMATION IN-SITU INSTRUMENTATION

    International Nuclear Information System (INIS)

    N.T. Raczka

    2000-01-01

    The purpose of this document is to identify and analyze the types of in-situ instruments and methods that could be used in support of the data acquisition portion of the Performance Confirmation (PC) program at the potential nuclear waste repository at Yucca Mountain. The PC program will require geomechanical , geophysical, thermal, and hydrologic instrumentation of several kinds. This analysis is being prepared to document the technical issues associated with each type of measurement during the PC period. This analysis utilizes the ''Performance Confirmation Input Criteria'' (CRWMS M andO 1999a) as its starting point. The scope of this analysis is primarily on the period after the start of waste package emplacement and before permanent closure of the repository, a period lasting between 15 and 300 years after last package emplacement (Stroupe 2000, Attachment 1, p. 1). The primary objectives of this analysis are to: (1) Review the design criteria as presented in the ''Performance Confirmation Input Criteria'' (CRWMS M andO 1999a). The scope of this analysis will be limited to the instrumentation related to parameters that require continuous monitoring of the conditions underground. (2) Preliminary identification and listing of the data requirements and parameters as related to the current repository layout in support of PC monitoring. (3) Preliminary identification of methods and instrumentation for the acquisition of the required data. Although the ''Performance Confirmation Input Criteria'' (CRWMS M andO 1999a) defines a broad range of data that must be obtained from a variety of methods, the focus of this analysis is on instrumentation related to the performance of the rock mass and the formation of water in the repository environment, that is obtainable from in-situ observation, testing, and monitoring

  1. Heat Flux Inhibition by Whistlers: Experimental Confirmation

    International Nuclear Information System (INIS)

    Eichler, D.

    2002-01-01

    Heat flux in weakly magnetized collisionless plasma is, according to theoretical predictions, limited by whistler turbulence that is generated by heat flux instabilities near threshold. Observations of solar wind electrons by Gary and coworkers appear to confirm the limit on heat flux as being roughly the product of the magnetic energy density and the electron thermal velocity, in agreement with prediction (Pistinner and Eichler 1998)

  2. 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.

  3. 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

  4. Beijing ambient particle exposure accelerates atherosclerosis in ApoE knockout mice.

    Science.gov (United States)

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

    2013-11-25

    higher than the FA group in terms of serum total cholesterol, low-density lipoprotein, tumor necrosis factor-alpha (TNF-alpha) and C-reactive protein as well as TNF-alpha and interleukin-6 in BAL. Also the total antioxidant capacity and oxidized low-density lipoprotein were significantly different between the two groups. In addition, pathological analysis of aortic arch reveals that the plaques area in the PM group increased significantly compared to the FA group. Our results demonstrated that ambient PM exposure could induce considerable oxidative stress and systemic inflammation in ApoE knockout mice and contribute to the progression of atherosclerosis. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  5. 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.

  6. 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.

  7. Efficient Generation of Myostatin Knock-Out Sheep Using CRISPR/Cas9 Technology and Microinjection into Zygotes.

    Directory of Open Access Journals (Sweden)

    M Crispo

    Full Text Available While CRISPR/Cas9 technology has proven to be a valuable system to generate gene-targeted modified animals in several species, this tool has been scarcely reported in farm animals. Myostatin is encoded by MSTN gene involved in the inhibition of muscle differentiation and growth. We determined the efficiency of the CRISPR/Cas9 system to edit MSTN in sheep and generate knock-out (KO animals with the aim to promote muscle development and body growth. We generated CRISPR/Cas9 mRNAs specific for ovine MSTN and microinjected them into the cytoplasm of ovine zygotes. When embryo development of CRISPR/Cas9 microinjected zygotes (n = 216 was compared with buffer injected embryos (n = 183 and non microinjected embryos (n = 173, cleavage rate was lower for both microinjected groups (P<0.05 and neither was affected by CRISPR/Cas9 content in the injected medium. Embryo development to blastocyst was not affected by microinjection and was similar among the experimental groups. From 20 embryos analyzed by Sanger sequencing, ten were mutant (heterozygous or mosaic; 50% efficiency. To obtain live MSTN KO lambs, 53 blastocysts produced after zygote CRISPR/Cas9 microinjection were transferred to 29 recipient females resulting in 65.5% (19/29 of pregnant ewes and 41.5% (22/53 of newborns. From 22 born lambs analyzed by T7EI and Sanger sequencing, ten showed indel mutations at MSTN gene. Eight showed mutations in both alleles and five of them were homozygous for indels generating out-of frame mutations that resulted in premature stop codons. Western blot analysis of homozygous KO founders confirmed the absence of myostatin, showing heavier body weight than wild type counterparts. In conclusion, our results demonstrate that CRISPR/Cas9 system was a very efficient tool to generate gene KO sheep. This technology is quick and easy to perform and less expensive than previous techniques, and can be applied to obtain genetically modified animal models of interest for

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

    Directory of Open Access Journals (Sweden)

    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.

  9. 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.

  10. 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.

  11. Rearing-environment-dependent hippocampal local field potential differences in wild-type and inositol trisphosphate receptor type 2 knockout mice.

    Science.gov (United States)

    Tanaka, Mika; Wang, Xiaowen; Mikoshiba, Katsuhiko; Hirase, Hajime; Shinohara, Yoshiaki

    2017-10-15

    Mice reared in an enriched environment are demonstrated to have larger hippocampal gamma oscillations than those reared in isolation, thereby confirming previous observations in rats. To test whether astrocytic Ca 2+ surges are involved in this experience-dependent LFP pattern modulation, we used inositol trisphosphate receptor type 2 (IP 3 R2)-knockout (KO) mice, in which IP 3 /Ca 2+ signalling in astrocytes is largely diminished. We found that this experience-dependent gamma power alteration persists in the KO mice. Interestingly, hippocampal ripple events, the synchronized events critical for memory consolidation, are reduced in magnitude and frequency by both isolated rearing and IP 3 R2 deficiency. Rearing in an enriched environment (ENR) is known to enhance cognitive and memory abilities in rodents, whereas social isolation (ISO) induces depression-like behaviour. The hippocampus has been documented to undergo morphological and functional changes depending on these rearing environments. For example, rearing condition during juvenility alters CA1 stratum radiatum gamma oscillation power in rats. In the present study, hippocampal CA1 local field potentials (LFP) were recorded from bilateral CA1 in urethane-anaesthetized mice that were reared in either an ENR or ISO condition. Similar to previous findings in rats, gamma oscillation power during theta states was higher in the ENR group. Ripple events that occur during non-theta periods in the CA1 stratum pyramidale also had longer intervals in ISO mice. Because astrocytic Ca 2+ elevations play a key role in synaptic plasticity, we next tested whether these changes in LFP are also expressed in inositol trisphosphate receptor type 2 (IP 3 R2)-knockout (KO) mice, in which astrocytic Ca 2+ elevations are largely diminished. We found that the gamma power was also higher in IP 3 R2-KO-ENR mice compared to IP 3 R2-KO-ISO mice, suggesting that the rearing-environment-dependent gamma power alteration does not necessarily

  12. Troubleshooting Requests e-mail Confirmation

    CERN Multimedia

    TS Department

    2004-01-01

    In an ongoing effort to improve quality of the repair requests, a new e-mail confirmation automatic system will be implemented starting from the 21st October. All repair requests transmitted to the TCR (72201) or the FM Helpdesk (77777) will be confirmed in an e-mail to the requestor, provided that the latter has a valid e-mail address in the HR database. The e-mail will contain a reference number, a brief description of the problem, the location and a contact where more information can be obtained. A second e-mail will be sent when the processing of the repair request is finished. We hope that this initiative will improve the transparency and quality of our service. Helpdesk Troubleshooting Requests (reminder) We remind you that all the repair requests and other communication concerning the CERN machine buildings have to be transmitted to the TCR via 72201, whereas the ones concerning tertiary buildings are handled directly by the FM helpdesk under the phone number 77777, i.e. problems on systems and equ...

  13. Kepler Confirmation of Multi-Planet Systems

    Science.gov (United States)

    Cochran, W. D.

    2011-10-01

    The NASA Kepler spacecraft has detected 170 candidate multi-planet systems in the first two quarters of data released in February 2011 by Borucki et al. (2011). These systems comprise 115 double candidate systems, 45 triple candidate sys- tems, and 10 systems with 4 or more candidate planets. The architecture and dynamics of these systems were discussed by Lissauer et al. (2011), and a comparison of candidates in single- and multi-planet systems was presented by Latham et al. (2011). Proceeding from "planetary candidate" systems to confirmed and validated multi-planet systems is a difficult process, as most of these systems orbit stars too faint to obtain extremely precise (1ms-1) radial velocity confimation. Here, we discuss in detail the use of transit timing vari- ations (cf. e.g. Holman et al., 2010) to confirm planets near a mean motion resonance. We also discuss extensions to the BLENDER validation (Torres et al., 2004, 2011; Fressin et al., 2011) to validate planets in multi-planet systems. Kepler was competitively selected as the tenth Discovery mission. Funding for the Kepler Mis- sion is provided by NASA's Science Mission Direc- torate. We are deeply grateful for the very hard work of the entire Kepler team.

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

    Directory of Open Access Journals (Sweden)

    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.

  15. 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.

  16. 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

  17. 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

  18. 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)

  19. Molecular Confirmation of Salmonella typhimuriumin Poultry from Kathmandu Valley

    Directory of Open Access Journals (Sweden)

    Sanjeev Kumar Adhikari

    2018-05-01

    Full Text Available A prevalence study was carried to isolate Salmonella typhimurium from blood (n= 50 and gut samples (n=100 of poultry in Kathmandu valley during early 2016. Salmonella typhimurium bacteria isolated in the selective media were biochemically confirmed based on Bergey’s Manual. Two sets of oligonucleotide primers-the genus specific 16S rRNA and the organism specific invA were employed for molecular level confirmation by the Polymerase Chain Reaction (PCR assay. The amplified fragments in 1% agarose gel observed at 406bp and 285bp, respectively confirmed the isolates to be Salmonella typhimurium. Of 150 samples tested, Salmonella typhimurium were isolated from 49 samples, among which nine were from blood (18% and forty from the gut (40%. The present result indicated an alarmingly high level of Salmonella typhimurium, which can result inzoonotic infection in humans owing to increased contact with poultry and consumption of poultry products in the Kathmandu valley.

  20. Maximal Oxygen Consumption is Reduced in Aquaporin-1 Knockout Mice

    Directory of Open Access Journals (Sweden)

    Samer Al-Samir

    2016-08-01

    Full Text Available We have measured maximal oxygen consumption (V’O2,max of mice lacking one or two of the established mouse red-cell CO2 channels AQP1, AQP9 and Rhag. We intended to study whether these proteins, by acting as channels for O2, determine O2 exchange in the lung and in the periphery. We found that V’O2,max as determined by the Helox technique is reduced by ~ 16%, when AQP1 is knocked out, but not when AQP9 or Rhag are lacking. This figure holds for animals respiring normoxic as well as hypoxic gas mixtures. To see whether the reduction of V’O2,max is due to impaired O2 uptake in the lung, we measured carotid arterial O2 saturation (SO2 by pulse oximetry. Neither under normoxic (inspiratory O2 21% nor under hypoxic conditions (11% O2 is there a difference in SO2 between AQP1null and WT mice, suggesting that AQP1 is not critical for O2 uptake in the lung. The fact that the % reduction of V’O2,max is identical in normoxia and hypoxia indicates moreover that the limitation of V’O2,max is not due to an O2 diffusion problem, neither in the lung nor in the periphery. Instead, it appears likely that AQP1null animals exhibit a reduced V’O2,max due to the reduced wall thickness and muscle mass of the left ventricles of their hearts, as reported previously. We conclude that very likely the properties of the hearts of AQP1 knockout mice cause a reduced maximal cardiac output and thus cause a reduced V’O2,max, which constitutes a new phenotype of these mice.

  1. Characterization of Heterogeneous Prostate Tumors in Targeted Pten Knockout Mice.

    Directory of Open Access Journals (Sweden)

    Hanneke Korsten

    Full Text Available Previously, we generated a preclinical mouse prostate tumor model based on PSA-Cre driven inactivation of Pten. In this model homogeneous hyperplastic prostates (4-5m developed at older age (>10m into tumors. Here, we describe the molecular and histological characterization of the tumors in order to better understand the processes that are associated with prostate tumorigenesis in this targeted mouse Pten knockout model. The morphologies of the tumors that developed were very heterogeneous. Different histopathological growth patterns could be identified, including intraductal carcinoma (IDC, adenocarcinoma and undifferentiated carcinoma, all strongly positive for the epithelial cell marker Cytokeratin (CK, and carcinosarcomas, which were negative for CK. IDC pattern was already detected in prostates of 7-8 month old mice, indicating that it could be a precursor stage. At more than 10 months IDC and carcinosarcoma were most frequently observed. Gene expression profiling discriminated essentially two molecular subtypes, denoted tumor class 1 (TC1 and tumor class 2 (TC2. TC1 tumors were characterized by high expression of epithelial markers like Cytokeratin 8 and E-Cadherin whereas TC2 tumors showed high expression of mesenchyme/stroma markers such as Snail and Fibronectin. These molecular subtypes corresponded with histological growth patterns: where TC1 tumors mainly represented adenocarcinoma/intraductal carcinoma, in TC2 tumors carcinosarcoma was the dominant growth pattern. Further molecular characterization of the prostate tumors revealed an increased expression of genes associated with the inflammatory response. Moreover, functional markers for senescence, proliferation, angiogenesis and apoptosis were higher expressed in tumors compared to hyperplasia. The highest expression of proliferation and angiogenesis markers was detected in TC2 tumors. Our data clearly showed that in the genetically well-defined PSA-Cre;Pten-loxP/loxP prostate tumor

  2. Separating genetic and hemodynamic defects in neuropilin 1 knockout embryos.

    Science.gov (United States)

    Jones, Elizabeth A V; Yuan, Li; Breant, Christine; Watts, Ryan J; Eichmann, Anne

    2008-08-01

    Targeted inactivation of genes involved in murine cardiovascular development frequently leads to abnormalities in blood flow. As blood fluid dynamics play a crucial role in shaping vessel morphology, the presence of flow defects generally prohibits the precise assignment of the role of the mutated gene product in the vasculature. In this study, we show how to distinguish between genetic defects caused by targeted inactivation of the neuropilin 1 (Nrp1) receptor and hemodynamic defects occurring in homozygous knockout embryos. Our analysis of a Nrp1 null allele bred onto a C57BL/6 background shows that vessel remodeling defects occur concomitantly with the onset of blood flow and cause death of homozygous mutants at E10.5. Using mouse embryo culture, we establish that hemodynamic defects are already present at E8.5 and continuous circulation is never established in homozygous mutants. The geometry of yolk sac blood vessels is altered and remodeling into yolk sac arteries and veins does not occur. To separate flow-induced deficiencies from those caused by the Nrp1 mutation, we arrested blood flow in cultured wild-type and mutant embryos and followed their vascular development. We find that loss of Nrp1 function rather than flow induces the altered geometry of the capillary plexus. Endothelial cell migration, but not replication, is altered in Nrp1 mutants. Gene expression analysis of endothelial cells isolated from freshly dissected wild-type and mutants and after culture in no-flow conditions showed down-regulation of the arterial marker genes connexin 40 and ephrin B2 related to the loss of Nrp1 function. This method allows genetic defects caused by loss-of-function of a gene important for cardiovascular development to be isolated even in the presence of hemodynamic defects.

  3. 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...

  4. 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

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

    Science.gov (United States)

    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.

  6. 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.

  7. Web-Based Honorarium Confirmation System Prototype

    Science.gov (United States)

    Wisswani, N. W.; Catur Bawa, I. G. N. B.

    2018-01-01

    Improving services in academic environment can be applied by regulating salary payment process for all employees. As a form of control to maintain financial transparency, employees should have information concerning salary payment process. Currently, notification process of committee honorarium will be accepted by the employees in a manual manner. The salary will be received by the employee bank account and to know its details, they should go to the accounting unit to find out further information. Though there are some employees entering the accounting unit, they still find difficulty to obtain information about detailed honor information that they received in their accounts. This can be caused by many data collected and to be managed. Based on this issue, this research will design a prototype of web-based system for accounting unit system in order to provide detailed financial transaction confirmation to employee bank accounts that have been informed through mobile banking system. This prototype will be developed with Waterfall method through testing on final users after it is developed through PHP program with MySQL as DBMS

  8. Confirmation of antibodies against L-tryptophan-like epitope in ...

    African Journals Online (AJOL)

    Rachel Oneya

    2016-09-07

    Sep 7, 2016 ... controls confirming previous results obtained with a lower number of patients in Congo. ... searching trypanosomes and white blood cells count in CSF: 13 in ... The well plate was then filled with 200 µL of diluted (2,000-fold).

  9. Evaluation of histologically confirmed carcinoma of the cervix in ...

    African Journals Online (AJOL)

    Data collected was analysed with SPSS version 20.0 software and presented in tables and charts. Results: Sixty two patients with histological confirmation of ... The commonest histological type of cervical cancer was squamous cell carcinoma accounting for 88.9%. Twenty (44.4%) patients were referred for radiotherapy and ...

  10. Laboratory-confirmed Congenital Rubella Syndrome at the ...

    African Journals Online (AJOL)

    Esem

    ORIGINAL ARTICLE. Laboratory-confirmed Congenital Rubella Syndrome at the University Teaching Hospital in Lusaka,. Zambia-Case Reports. 1,2. 3. 3. 4 ... microcephaly. Rubella Immunoglobulin M (IgM) results were positive. The third case, a girl, was seen at twelve weeks and brought in for slow growth rate. On.

  11. Racial athletic stereotype confirmation in college football recruiting.

    Science.gov (United States)

    Thomas, Grant; Good, Jessica J; Gross, Alexi R

    2015-01-01

    The present study tested real-world racial stereotype use in the context of college athletic recruiting. Stereotype confirmation suggests that observers use stereotypes as hypotheses and interpret relevant evidence in a biased way that confirms their stereotypes. Shifting standards suggest that the evaluative standard to which we hold a target changes as a function of their group membership. We examined whether stereotype confirmation and shifting standards effects would be seen in college football coaches during recruiting. College football coaches evaluated a Black or White player on several attributes and made both zero- and non-zero-sum allocations. Results suggested that coaches used the evidence presented to develop biased subjective evaluations of the players based on race while still maintaining equivalent objective evaluations. Coaches also allocated greater overall resources to the Black recruit than the White recruit.

  12. Impaired Sperm Maturation in Rnase9 Knockout Mice1

    Science.gov (United States)

    Westmuckett, Andrew D.; Nguyen, Edward B.; Herlea-Pana, Oana M.; Alvau, Antonio; Salicioni, Ana M.; Moore, Kevin L.

    2014-01-01

    ABSTRACT Ribonuclease, RNase A family, 9 (RNASE9) is a ribonuclease A superfamily member that is expressed only in the epididymis. It is a small, secreted polypeptide, it lacks ribonuclease activity, and its function(s) is unknown. However, epididymis-specific expression suggests a role in sperm maturation. We generated Rnase9−/− mice to study RNASE9 function in vivo. We confirm that RNASE9 expression is restricted to the epididymis. Within the epididymis, RNASE9 is first detected in midcaput, persists through the distal caput and corpus, and wanes in the cauda. Rnase9−/− mice are born at the expected Mendelian ratio, have normal postnatal growth and development, and have no outwardly apparent phenotype. Spermatogenesis is normal, and Rnase9-null sperm are morphologically normal. Rnase9−/− males have normal fertility in unrestricted mating trials, and fertilization rates in in vitro fertilization assays are indistinguishable from wild-type mice. Visual observations coupled with analyses of sperm velocities shortly after swim out from the corpus shows that motility of Rnase9-null sperm is significantly impaired. However, no differences between wild-type and Rnase9-null sperm are detected by computer-assisted sperm analysis 10–90 min after sperm isolation from the corpus or cauda. Assessment of capacitation-dependent signaling pathways in Rnase9-null sperm showed that, while levels of tyrosine phosphorylation of sperm proteins were normal, there was decreased phosphorylation of protein kinase A substrates upon capacitation compared to wild-type mice. In conclusion, RNASE9 is dispensable for fertility, but the absence of RNASE9 during epididymal transit results in impaired sperm maturation. PMID:24719258

  13. Altered neurocircuitry in the dopamine transporter knockout mouse brain.

    Directory of Open Access Journals (Sweden)

    Xiaowei Zhang

    2010-07-01

    Full Text Available The plasma membrane transporters for the monoamine neurotransmitters dopamine, serotonin, and norepinephrine modulate the dynamics of these monoamine neurotransmitters. Thus, activity of these transporters has significant consequences for monoamine activity throughout the brain and for a number of neurological and psychiatric disorders. Gene knockout (KO mice that reduce or eliminate expression of each of these monoamine transporters have provided a wealth of new information about the function of these proteins at molecular, physiological and behavioral levels. In the present work we use the unique properties of magnetic resonance imaging (MRI to probe the effects of altered dopaminergic dynamics on meso-scale neuronal circuitry and overall brain morphology, since changes at these levels of organization might help to account for some of the extensive pharmacological and behavioral differences observed in dopamine transporter (DAT KO mice. Despite the smaller size of these animals, voxel-wise statistical comparison of high resolution structural MR images indicated little morphological change as a consequence of DAT KO. Likewise, proton magnetic resonance spectra recorded in the striatum indicated no significant changes in detectable metabolite concentrations between DAT KO and wild-type (WT mice. In contrast, alterations in the circuitry from the prefrontal cortex to the mesocortical limbic system, an important brain component intimately tied to function of mesolimbic/mesocortical dopamine reward pathways, were revealed by manganese-enhanced MRI (MEMRI. Analysis of co-registered MEMRI images taken over the 26 hours after introduction of Mn(2+ into the prefrontal cortex indicated that DAT KO mice have a truncated Mn(2+ distribution within this circuitry with little accumulation beyond the thalamus or contralateral to the injection site. By contrast, WT littermates exhibit Mn(2+ transport into more posterior midbrain nuclei and contralateral

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

    Directory of Open Access Journals (Sweden)

    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

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

    Directory of Open Access Journals (Sweden)

    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.

  16. Ethanol self-administration in serotonin transporter knockout mice: unconstrained demand and elasticity.

    Science.gov (United States)

    Lamb, R J; Daws, L C

    2013-10-01

    Low serotonin function is associated with alcoholism, leading to speculation that increasing serotonin function could decrease ethanol consumption. Mice with one or two deletions of the serotonin transporter (SERT) gene have increased extracellular serotonin. To examine the relationship between SERT genotype and motivation for alcohol, we compared ethanol self-administration in mice with zero (knockout, KO), one (HET) or two copies (WT) of the SERT gene. All three genotypes learned to self-administer ethanol. The SSRI, fluvoxamine, decreased responding for ethanol in the HET and WT, but not the KO mice. When tested under a progressive ratio schedule, KO mice had lower breakpoints than HET or WT. As work requirements were increased across sessions, behavioral economic analysis of ethanol self-administration indicated that the decreased breakpoint in KO as compared to HET or WT mice was a result of lower levels of unconstrained demand, rather than differences in elasticity, i.e. the proportional decreases in ethanol earned with increasing work requirements were similar across genotypes. The difference in unconstrained demand was unlikely to result from motor or general motivational factors, as both WT and KO mice responded at high levels for a 50% condensed milk solution. As elasticity is hypothesized to measure essential value, these results indicate that KO value ethanol similarly to WT or HET mice despite having lower break points for ethanol. © 2013 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

  17. 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

  18. 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.

  19. Effects of Eaf2 gene knockout on cataract induced by ultraviolet irradiation in mice

    Directory of Open Access Journals (Sweden)

    Yan-Hua Jiang

    2016-02-01

    Full Text Available AIM:To evaluate the effects of Eaf2 gene knockout on cataract in mice induced by ultraviolet irradiation.METHODS:Fifteen wild type mice were used as the control group, and 10 Eaf2 KO mice were used as the experimental group. The 14-week mice were taken as the research objects in the two groups. So the subgroups were: WT -nonUV, WT -UV, Eaf2 KO-nonUV and Eaf2 KO-UV, a total of 4 groups. Observe the lens of mice in vivo with slit lamp microscope, grade the lens opacity with Lens Opacities Classification System II(LOCSII. Then the mice were sacrificed by breaking the neck, the lens were removed and were observed by dark field microscopy. According to the captured images, the proportion of cataract region was analyzed using Image J software. The data of the two groups were statistically analyzed.RESULTS: The results detected by the two methods were similar. In WT-UV group and Eaf2 KO-UV group, the degree of lens opacity was significantly higher than those of WT-nonUV group and Eaf2 KO-nonUV group. The lens opacity of WT-UV group was significantly higher than that in Eaf2 KO-UV group, and the difference was statistically significant(PCONCLUSION: Ultraviolet radiation can lead to the formation of cataract in mice. Eaf2 protein can promote the formation of cataract in mice caused by ultraviolet.

  20. Confirmed and Potential Sources of Legionella Reviewed

    NARCIS (Netherlands)

    van Heijnsbergen, Eri; Schalk, Johanna A C; Euser, Sjoerd M; Brandsema, Petra S; den Boer, Jeroen W; de Roda Husman, Ana Maria|info:eu-repo/dai/nl/139498281

    2015-01-01

    Legionella bacteria are ubiquitous in natural matrices and man-made systems. However, it is not always clear if these reservoirs can act as source of infection resulting in cases of Legionnaires' disease. This review provides an overview of reservoirs of Legionella reported in the literature, other

  1. 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

  2. Legionella confirmation in cooling tower water

    Science.gov (United States)

    Farhat, Maha; Shaheed, Raja A.; Al-Ali, Haidar H.; Al-Ghamdi, Abdullah S.; Al-Hamaqi, Ghadeer M.; Maan, Hawraa S.; Al-Mahfoodh, Zainab A.; Al-Seba, Hussain Z.

    2018-01-01

    Objectives: To investigate the presence of Legionella spp in cooling tower water. Legionella proliferation in cooling tower water has serious public health implications as it can be transmitted to humans via aerosols and cause Legionnaires’ disease. Methods: Samples of cooling tower water were collected from King Fahd Hospital of the University (KFHU) (Imam Abdulrahman Bin Faisal University, 2015/2016). The water samples were analyzed by a standard Legionella culture method, real-time polymerase chain reaction (RT-PCR), and 16S rRNA next-generation sequencing. In addition, the bacterial community composition was evaluated. Results: All samples were negative by conventional Legionella culture. In contrast, all water samples yielded positive results by real-time PCR (105 to 106 GU/L). The results of 16S rRNA next generation sequencing showed high similarity and reproducibility among the water samples. The majority of sequences were Alpha-, Beta-, and Gamma-proteobacteria, and Legionella was the predominant genus. The hydrogen-oxidizing gram-negative bacterium Hydrogenophaga was present at high abundance, indicating high metabolic activity. Sphingopyxis, which is known for its resistance to antimicrobials and as a pioneer in biofilm formation, was also detected. Conclusion: Our findings indicate that monitoring of Legionella in cooling tower water would be enhanced by use of both conventional culturing and molecular methods. PMID:29436561

  3. 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.

  4. 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.

  5. 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.

  6. CERN Confirms commitment to Open Access

    CERN Multimedia

    2005-01-01

    The CERN Library Information desk.At a meeting on the Wednesday before Easter, the Executive Committee endorsed a policy of open access to all the laboratory's results, as expressed in the document ‘Continuing CERN action on Open Access' (http://cds.cern.ch/record/828991/files/open-2005-006.pdf), released by its Scientific Information Policy Board (SIPB) earlier in the month. "This underlines CERN's commitment to sharing the excitement of fundamental research with as wide an audience as possible", said Guido Altarelli, current SIPB chairman. Open Access to scientific knowledge is today the goal of an increasing component of the worldwide scientific community. It is a concept, made possible by new electronic tools, which would bring enormous benefits to all readers by giving them free access to research results. CERN has implicitly supported such moves from its very beginning. Its Convention (http://cds.cern.ch/record/330625/files/cm-p00046871.pdf), adopted in 1953, requires openness, stipulating that "......

  7. 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.

  8. The Effect of Different Photoperiods in Circadian Rhythms of Per3 Knockout Mice

    Directory of Open Access Journals (Sweden)

    D. S. Pereira

    2014-01-01

    Full Text Available The aim of this study was to analyse the circadian behavioural responses of mice carrying a functional knockout of the Per3 gene (Per3−/− to different light : dark (L : D cycles. Male adult wild-type (WT and Per3−/− mice were kept under 12-hour light : 12-hour dark conditions (12L : 12D and then transferred to either a short or long photoperiod and subsequently released into total darkness. All mice were exposed to both conditions, and behavioural activity data were acquired through running wheel activity and analysed for circadian characteristics during these conditions. We observed that, during the transition from 12L : 12D to 16L : 8D, Per3−/− mice take approximately one additional day to synchronise to the new L : D cycle compared to WT mice. Under these long photoperiod conditions, Per3−/− mice were more active in the light phase. Our results suggest that Per3−/− mice are less sensitive to light. The data presented here provides further evidence that Per3 is involved in the suppression of behavioural activity in direct response to light.

  9. Increased sensitivity of apolipoprotein E knockout mice to copper-induced oxidative injury to the liver.

    Science.gov (United States)

    Chen, Yuan; Li, Bin; Zhao, Ran-ran; Zhang, Hui-feng; Zhen, Chao; Guo, Li

    2015-04-10

    Apolipoprotein E (ApoE) genotypes are related to clinical presentations in patients with Wilson's disease, indicating that ApoE may play an important role in the disease. However, our understanding of the role of ApoE in Wilson's disease is limited. High copper concentration in Wilson's disease induces excessive generation of free oxygen radicals. Meanwhile, ApoE proteins possess antioxidant effects. We therefore determined whether copper-induced oxidative damage differ in the liver of wild-type and ApoE knockout (ApoE(-/-)) mice. Both wild-type and ApoE(-/-) mice were intragastrically administered with 0.2 mL of copper sulfate pentahydrate (200 mg/kg; a total dose of 4 mg/d) or the same volume of saline daily for 12 weeks, respectively. Copper and oxidative stress markers in the liver tissue and in the serum were assessed. Our results showed that, compared with the wild-type mice administered with copper, TBARS as a marker of lipid peroxidation, the expression of oxygenase-1 (HO-1), NAD(P)H dehydrogenase, and quinone 1 (NQO1) significantly increased in the ApoE(-/-) mice administered with copper, meanwhile superoxide dismutase (SOD) activity significantly decreased. Thus, it is concluded that ApoE may protect the liver from copper-induced oxidative damage in Wilson's disease. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. 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.

  11. Proton induced nucleon knockout from 40Ca in the Dirac impulse approximation

    International Nuclear Information System (INIS)

    Maxwell, O.V.; Cooper, E.D.

    1989-12-01

    The (p,2p) reaction on 40 Ca at incident proton energies of 200 and 300 MeV is examined within a Dirac distorted wave impulse approximation. The relativistic Love-Franey t-matrix is evaluated at the nucleon-nucleon laboratory energy (as defined within the plane wave approximation), rather than the nucleon-nucleus laboratory energy. Particular attention is paid to the sensitivity of the calculated cross sections and analyzing powers to the properties of the bound states employed. It is found that the analyzing powers depend very little on the bound state properties, while the cross sections depend significantly only on the rms radii of the bound state wave functions. A major success of the model is its ability to fit the cross section data over a particular range of momentum transfers at both 200 and 300 MeV with the same bound state potential. Outside this momentum transfer range, the predicted cross sections are too low. The calculated analyzing powers agree well with the data at 200 MeV, but disagree with the data at 300 MeV. Values for the rms radii of the 1D 3/2 and 1D 5/2 states in 40 Ca are derived from the requirement that the peak positions of the calculated cross sections at 300 MeV agree with the empirical peak positions. Some preliminary results are also reported for neutron knockout from 40 Ca at 150 MeV

  12. Examination of MARCO activity on dendritic cell phenotype and function using a gene knockout mouse.

    Directory of Open Access Journals (Sweden)

    Hiroshi Komine

    Full Text Available We have reported the upregulation of MARCO, a member of the class A scavenger receptor family, on the surface of murine and human dendritic cells (DCs pulsed with tumor lysates. Exposure of murine tumor lysate-pulsed DCs to an anti-MARCO antibody led to loss of dendritic-like processes and enhanced migratory capacity. In this study, we have further examined the biological and therapeutic implications of MARCO expression by DCs. DCs generated from the bone marrow (bm of MARCO knockout (MARCO⁻/⁻ mice were phenotypically similar to DCs generated from the bm of wild-type mice and produced normal levels of IL-12 and TNF-α when exposed to LPS. MARCO⁻/⁻ DCs demonstrated enhanced migratory capacity in response to CCL-21 in vitro. After subcutaneous injection into mice, MARCO⁻/⁻ TP-DCs migrated more efficiently to the draining lymph node leading to enhanced generation of tumor-specific IFN-γ producing T cells and improved tumor regression and survival in B16 melanoma-bearing mice. These results support targeting MARCO on the surface of DCs to improve trafficking and induction of anti-tumor immunity.

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

    Directory of Open Access Journals (Sweden)

    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.

  14. 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.

  15. 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.

  16. 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.)

  17. NASA Observatory Confirms Black Hole Limits

    Science.gov (United States)

    2005-02-01

    The very largest black holes reach a certain point and then grow no more, according to the best survey to date of black holes made with NASA's Chandra X-ray Observatory. Scientists have also discovered many previously hidden black holes that are well below their weight limit. These new results corroborate recent theoretical work about how black holes and galaxies grow. The biggest black holes, those with at least 100 million times the mass of the Sun, ate voraciously during the early Universe. Nearly all of them ran out of 'food' billions of years ago and went onto a forced starvation diet. Focus on Black Holes in the Chandra Deep Field North Focus on Black Holes in the Chandra Deep Field North On the other hand, black holes between about 10 and 100 million solar masses followed a more controlled eating plan. Because they took smaller portions of their meals of gas and dust, they continue growing today. "Our data show that some supermassive black holes seem to binge, while others prefer to graze", said Amy Barger of the University of Wisconsin in Madison and the University of Hawaii, lead author of the paper describing the results in the latest issue of The Astronomical Journal (Feb 2005). "We now understand better than ever before how supermassive black holes grow." One revelation is that there is a strong connection between the growth of black holes and the birth of stars. Previously, astronomers had done careful studies of the birthrate of stars in galaxies, but didn't know as much about the black holes at their centers. DSS Optical Image of Lockman Hole DSS Optical Image of Lockman Hole "These galaxies lose material into their central black holes at the same time that they make their stars," said Barger. "So whatever mechanism governs star formation in galaxies also governs black hole growth." Astronomers have made an accurate census of both the biggest, active black holes in the distance, and the relatively smaller, calmer ones closer by. Now, for the first

  18. 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.

  19. 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.

  20. Histopathology confirms white-nose syndrome in bats in Europe.

    Science.gov (United States)

    Pikula, Jiri; Bandouchova, Hana; Novotny, Ladislav; Meteyer, Carol U; Zukal, Jan; Irwin, Nancy R; Zima, Jan; Martínková, Natália

    2012-01-01

    White-nose syndrome, associated with the fungal skin infection geomycosis, caused regional population collapse in bats in North America. Our results, based on histopathology, show the presence of white-nose syndrome in Europe. Dermatohistopathology on two bats (Myotis myotis) found dead in March 2010 with geomycosis in the Czech Republic had characteristics resembling Geomyces destructans infection in bats confirmed with white-nose syndrome in US hibernacula. In addition, a live M. myotis, biopsied for histopathology during hibernation in April 2011, had typical fungal infection with cupping erosion and invasion of muzzle skin diagnostic for white-nose syndrome and conidiospores identical to G. destructans that were genetically confirmed as G. destructans.

  1. Histopathology confirms white-nose syndrome in bats in Europe

    Science.gov (United States)

    Pikula, J.; Bandouchova, H.; Novotny, L.; Meteyer, C.U.; Zukal, J.; Irwin, N.R.; Zima, J.; Martinkova, N.

    2012-01-01

    White-nose syndrome, associated with the fungal skin infection geomycosis, caused regional population collapse in bats in North America. Our results, based on histopathology, show the presence of white-nose syndrome in Europe. Dermatohistopathology on two bats (Myotis myotis) found dead in March 2010 with geomycosis in the Czech Republic had characteristics resembling Geomyces destructans infection in bats confirmed with white-nose syndrome in US hibernacula. In addition, a live M. myotis, biopsied for histopathology during hibernation in April 2011, had typical fungal infection with cupping erosion and invasion of muzzle skin diagnostic for white-nose syndrome and conidiospores identical to G. destructans that were genetically confirmed as G. destructans. ?? Wildlife Disease Association 2012.

  2. 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

  3. 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.

  4. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  6. 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.

  7. 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

    Science.gov (United States)

    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

  8. Remyelination in experimentally demyelinated connexin 32 KnockOut mice Remielinização em camundongos KnockOut para conexina 32 desmielinizados experimentalmente

    Directory of Open Access Journals (Sweden)

    Adriano Tony Ramos

    2009-06-01

    Full Text Available The aim of this study was to evaluate the role of connexin 32 (Cx 32 during remyelination of the peripheral nervous system, through a local injection of either 0,1% ethidium bromide solution or saline in the sciatic nerve of Cx 32 knockout mice. Euthanasia was performed ranging from 1, 2, 3, 7, 15, 21 to 30 days after injection. Histochemical, immunohistochemical, immunofluorescence and transmission electron microscopical techniques were used to analyze the development of the lesions. Within the sciatic nerves, Schwann cells initially showed signs of intoxication and rejected their sheaths; after seven days, some thin newly formed myelin sheaths with uneven compactness and redundant loops (tomacula were conspicuous. We concluded that the regeneration of lost myelin sheaths within the PNS followed the pattern already reported for this model in other laboratory species. Therefore, these results suggest that absence of Cx 32 did not interfere with the normal pattern of remyelination in this model in young mice.Este estudo visou avaliar o papel da conexina 32 (Cx 32 durante a remielinização no sistema nervoso periférico. Uma injeção local de 0,1% de solução de brometo de etídio foi realizada no nervo ciático de camundongos deletados para a Cx 32, com eutanásia dos animais aos 1, 2, 3, 7, 15, 21 e 30 dias pós-injeção. Avaliações histoquímicas, imunoistoquímicas, por imunofluorescência e por microscopia eletrônica de transmissão foram utilizadas na análise do desenvolvimento das lesões. Nos nervos ciáticos, células de Schwann mostraram inicialmente sinais de intoxicação e rejeitaram suas bainhas. Após sete dias, observaram-se finas bainhas neoformadas, com compactação desigual e alças redundantes (tomácula. Conclui-se que a regeneração de bainhas de mielina perdidas no SNP seguiu o padrão já relatado deste modelo em outras espécies de laboratório. Portanto, estes resultados sugerem que a ausência da Cx 32 n

  9. Gut microbiota is a key modulator of insulin resistance in TLR 2 knockout mice.

    Directory of Open Access Journals (Sweden)

    Andréa M Caricilli

    2011-12-01

    Full Text Available Environmental factors and host genetics interact to control the gut microbiota, which may have a role in the development of obesity and insulin resistance. TLR2-deficient mice, under germ-free conditions, are protected from diet-induced insulin resistance. It is possible that the presence of gut microbiota could reverse the phenotype of an animal, inducing insulin resistance in an animal genetically determined to have increased insulin sensitivity, such as the TLR2 KO mice. In the present study, we investigated the influence of gut microbiota on metabolic parameters, glucose tolerance, insulin sensitivity, and signaling of TLR2-deficient mice. We investigated the gut microbiota (by metagenomics, the metabolic characteristics, and insulin signaling in TLR2 knockout (KO mice in a non-germ free facility. Results showed that the loss of TLR2 in conventionalized mice results in a phenotype reminiscent of metabolic syndrome, characterized by differences in the gut microbiota, with a 3-fold increase in Firmicutes and a slight increase in Bacteroidetes compared with controls. These changes in gut microbiota were accompanied by an increase in LPS absorption, subclinical inflammation, insulin resistance, glucose intolerance, and later, obesity. In addition, this sequence of events was reproduced in WT mice by microbiota transplantation and was also reversed by antibiotics. At the molecular level the mechanism was unique, with activation of TLR4 associated with ER stress and JNK activation, but no activation of the IKKβ-IκB-NFκB pathway. Our data also showed that in TLR2 KO mice there was a reduction in regulatory T cell in visceral fat, suggesting that this modulation may also contribute to the insulin resistance of these animals. Our results emphasize the role of microbiota in the complex network of molecular and cellular interactions that link genotype to phenotype and have potential implications for common human disorders involving obesity, diabetes

  10. Hippocampal Focal Knockout of CBP Affects Specific Histone Modifications, Long-Term Potentiation, and Long-Term Memory

    Science.gov (United States)

    Barrett, Ruth M; Malvaez, Melissa; Kramar, Eniko; Matheos, Dina P; Arrizon, Abraham; Cabrera, Sara M; Lynch, Gary; Greene, Robert W; Wood, Marcelo A

    2011-01-01

    To identify the role of the histone acetyltransferase (HAT) CREB-binding protein (CBP) in neurons of the CA1 region of the hippocampus during memory formation, we examine the effects of a focal homozygous knockout of CBP on histone modifications, gene expression, synaptic plasticity, and long-term memory. We show that CBP is critical for the in vivo acetylation of lysines on histones H2B, H3, and H4. CBP's homolog p300 was unable to compensate for the loss of CBP. Neurons lacking CBP maintained phosphorylation of the transcription factor CREB, yet failed to activate CREB:CBP-mediated gene expression. Loss of CBP in dorsal CA1 of the hippocampus resulted in selective impairments to long-term potentiation and long-term memory for contextual fear and object recognition. Together, these results suggest a necessary role for specific chromatin modifications, selectively mediated by CBP in the consolidation of memories. PMID:21508930

  11. 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

  12. 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

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

    Science.gov (United States)

    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…

  14. 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...

  15. 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

  16. 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...

  17. 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

  18. 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

  19. 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...

  20. 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.

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

    Science.gov (United States)

    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…

  2. 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.

  3. 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

  4. Male aromatase-knockout mice exhibit normal levels of activity, anxiety and "depressive-like" symptomatology.

    Science.gov (United States)

    Dalla, C; Antoniou, K; Papadopoulou-Daifoti, Z; Balthazart, J; Bakker, J

    2005-09-08

    It is well known that estradiol derived from neural aromatization of testosterone plays a crucial role in the development of the male brain and the display of sexual behaviors in adulthood. It was recently found that male aromatase knockout mice (ArKO) deficient in estradiol due to a mutation in the aromatase gene have general deficits in coital behavior and are sexually less motivated. We wondered whether these behavioral deficits of ArKO males could be related to changes in activity, exploration, anxiety and "depressive-like" symptomatology. ArKO and wild type (WT) males were subjected to open field (OF), elevated plus maze (EPM), and forced swim tests (FST), after being exposed or not to chronic mild stress (CMS). CMS was used to evaluate the impact of chronic stressful procedures and to unveil possible differences between genotypes. There was no effect of genotype on OF, EPM and FST behavioral parameters. WT and ArKO mice exposed to CMS or not exhibited the same behavioral profile during these three types of tests. However, all CMS-exposed mice (ArKO and WT) spent less time in the center of the EPM. Additionally, floating duration measured in the FST increased between two tests in both WT and ArKO mice, though that increase was less prominent in mice previously subjected to CMS than in controls. Therefore, both ArKO and WT males displayed the same behavior and had the same response to CMS however CMS exposure slightly modified the behavior displayed by mice of both genotypes in the FST and EPM paradigms. These results show that ArKO males display normal levels of activity, exploration, anxiety and "depressive-like" symptomatology and thus their deficits in sexual behavior are specific in nature and do not result indirectly from other behavioral changes.

  5. Mechanism of hyperphagia contributing to obesity in brain-derived neurotrophic factor knockout mice.

    Science.gov (United States)

    Fox, E A; Biddinger, J E; Jones, K R; McAdams, J; Worman, A

    2013-01-15

    Global-heterozygous and brain-specific homozygous knockouts (KOs) of brain-derived neurotrophic factor (BDNF) cause late- and early-onset obesity, respectively, both involving hyperphagia. Little is known about the mechanism underlying this hyperphagia or whether BDNF loss from peripheral tissues could contribute to overeating. Since global-homozygous BDNF-KO is perinatal lethal, a BDNF-KO that spared sufficient brainstem BDNF to support normal health was utilized to begin to address these issues. Meal pattern and microstructure analyses suggested overeating of BDNF-KO mice was mediated by deficits in both satiation and satiety that resulted in increased meal size and frequency and implicated a reduction of vagal signaling from the gut to the brain. Meal-induced c-Fos activation in the nucleus of the solitary tract, a more direct measure of vagal afferent signaling, however, was not decreased in BDNF-KO mice, and thus was not consistent with a vagal afferent role. Interestingly though, meal-induced c-Fos activation was increased in the dorsal motor nucleus of the vagus nerve (DMV) of BDNF-KO mice. This could imply that augmentation of vago-vagal digestive reflexes occurred (e.g., accommodation), which would support increased meal size and possibly increased meal number by reducing the increase in intragastric pressure produced by a given amount of ingesta. Additionally, vagal sensory neuron number in BDNF-KO mice was altered in a manner consistent with the increased meal-induced activation of the DMV. These results suggest reduced BDNF causes satiety and satiation deficits that support hyperphagia, possibly involving augmentation of vago-vagal reflexes mediated by central pathways or vagal afferents regulated by BDNF levels. Copyright © 2012 IBRO. Published by Elsevier Ltd. All rights reserved.

  6. Calcium current homeostasis and synaptic deficits in hippocampal neurons from Kelch-like 1 knockout mice

    Directory of Open Access Journals (Sweden)

    Paula Patricia Perissinotti

    2015-01-01

    Full Text Available Kelch-like 1 (KLHL1 is a neuronal actin-binding protein that modulates voltage-gated CaV2.1 (P/Q-type and CaV3.2 (α1H T-type calcium channels; KLHL1 knockdown experiments (KD cause down-regulation of both channel types and altered synaptic properties in cultured rat hippocampal neurons (Perissinotti et al., 2014. Here, we studied the effect of ablation of KLHL1 on calcium channel function and synaptic properties in cultured hippocampal neurons from KLHL1 knockout (KO mice. Western blot data showed the P/Q-type channel α1A subunit was less abundant in KO hippocampus compared to wildtype (WT; and PQ-type calcium currents were smaller in KO neurons than WT during early days in vitro, although this decrease was compensated for at late stages by increases in L-type calcium current. In contrast, T-type currents did not change in culture. However, biophysical properties and western blot analysis revealed a differential contribution of T-type channel isoforms in the KO, with CaV3.2 α1H subunit being down-regulated and CaV3.1 α1G up-regulated. Synapsin I levels were reduced in the KO hippocampus; cultured neurons displayed a concomitant reduction in synapsin I puncta and decreased miniature excitatory postsynaptic current (mEPSC frequency. In summary, genetic ablation of the calcium channel modulator resulted in compensatory mechanisms to maintain calcium current homeostasis in hippocampal KO neurons; however, synaptic alterations resulted in a reduction of excitatory synapse number, causing an imbalance of the excitatory-inhibitory synaptic input ratio favoring inhibition.

  7. Hippocampal gene expression patterns in oxytocin male knockout mice are related to impaired social interaction.

    Science.gov (United States)

    Lazzari, Virginia Meneghini; Zimmermann-Peruzatto, Josi Maria; Agnes, Grasiela; Becker, Roberta Oriques; de Moura, Ana Carolina; Almeida, Silvana; Guedes, Renata Padilha; Giovenardi, Marcia

    2017-11-02

    Social interaction between animals is crucial for the survival and life in groups. It is well demonstrated that oxytocin (OT) and vasopressin (AVP) play critical roles in the regulation of social behaviors in mammals, however, other neurotransmitters and hormones are involved in the brain circuitry related to these behaviors. The present study aimed to investigate the gene expression of neurotransmitter receptors in the brain of OT knockout (OTKO) male mice. In this study, we evaluated the expression levels of the OT receptor (Oxtr), AVP receptors 1a and 1b (Avpr1a; Avpr1b), dopamine receptor 2 (Drd2), and the estrogen receptors alpha and beta (Esr1; Esr2) genes in the hippocampus (HPC), olfactory bulb (OB), hypothalamus (HPT) and prefrontal cortex (PFC). AVP gene (Avp) expression was analyzed in the HPT. Gene expression results were discussed regarding to social interaction and sexual behavior findings. Additionally, we analyzed the influence of OT absence on the Avp mRNA expression levels in the HPT. RNA extraction and cDNAs synthesis followed by quantitative polymerase chain reaction were performed for gene expression determination. Results were calculated with the 2 -ΔΔCt method. Our main finding was that HPC is more susceptible to gene expression changes due to the lack of OT. OTKOs exhibited decreased expression of Drd2 and Avpr1b, but increased expression of Oxtr in the HPC. In the PFC, Esr2 was increased. In the HPT, there was a reduced Avp expression in the OTKO group. No differences were detected in the OB and HPT. Despite these changes in gene expression, sexual behavior was not affected. However, OTKO showed higher social investigation and lower aggressive performance than wild-type mice. Our data highlight the importance of OT for proper gene expression of neurotransmitter receptors related to the regulation of social interaction in male mice. Copyright © 2017. Published by Elsevier B.V.

  8. 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.

  9. Comprehensive behavioral study of mGluR3 knockout mice: implication in schizophrenia related endophenotypes

    Science.gov (United States)

    2014-01-01

    Background We previously performed systematic association studies of glutamate receptor gene family members with schizophrenia, and found positive associations of polymorphisms in the GRM3 (a gene of metabotropic glutamate receptor 3: mGluR3) with the disorder. Physiological roles of GRM3 in brain functions and its functional roles in the pathogenesis of schizophrenia remain to be resolved. Results We generated mGluR3 knockout (KO) mice and conducted comprehensive behavioral analyses. KO mice showed hyperactivity in the open field, light/dark transition, and 24-hour home cage monitoring tests, impaired reference memory for stressful events in the Porsolt forced swim test, impaired contextual memory in cued and contextual fear conditioning test, and impaired working memory in the T-Maze forced alternation task test. Hyperactivity and impaired working memory are known as endophenotypes of schizophrenia. We examined long-term synaptic plasticity by assessing long-term potentiation (LTP) in the CA1 region in the hippocampi of KO and wild-type (WT) mice. We observed no differences in the amplitude of LTP between the two genotypes, suggesting that mGluR3 is not essential for LTP in the CA1 region of the mouse hippocampus. As hyperactivity is typically associated with increased dopaminergic transmission, we performed in vivo microdialysis measurements of extracellular dopamine in the nucleus accumbens of KO and WT mice. We observed enhancements in the methamphetamine (MAP)-induced release of dopamine in KO mice. Conclusions These results demonstrate that a disturbance in the glutamate-dopamine interaction may be involved in the pathophysiology of schizophrenia-like behavior, such as hyperactivity in mGluR3 KO mice. PMID:24758191

  10. A conditioned aversion study of sucrose and SC45647 taste in TRPM5 knockout mice.

    Science.gov (United States)

    Eddy, Meghan C; Eschle, Benjamin K; Peterson, Darlene; Lauras, Nathan; Margolskee, Robert F; Delay, Eugene R

    2012-06-01

    Previously, published studies have reported mixed results regarding the role of the TRPM5 cation channel in signaling sweet taste by taste sensory cells. Some studies have reported a complete loss of sweet taste preference in TRPM5 knockout (KO) mice, whereas others have reported only a partial loss of sweet taste preference. This study reports the results of conditioned aversion studies designed to motivate wild-type (WT) and KO mice to respond to sweet substances. In conditioned taste aversion experiments, WT mice showed nearly complete LiCl-induced response suppression to sucrose and SC45647. In contrast, TRPM5 KO mice showed a much smaller conditioned aversion to either sweet substance, suggesting a compromised, but not absent, ability to detect sweet taste. A subsequent conditioned flavor aversion experiment was conducted to determine if TRPM5 KO mice were impaired in their ability to learn a conditioned aversion. In this experiment, KO and WT mice were conditioned to a mixture of SC45647 and amyl acetate (an odor cue). Although WT mice avoided both components of the stimulus mixture, they avoided SC45647 more than the odor cue. The KO mice also avoided both stimuli, but they avoided the odor component more than SC45647, suggesting that while the KO mice are capable of learning an aversion, to them the odor cue was more salient than the taste cue. Collectively, these findings suggest the TRPM5 KO mice have some residual ability to detect SC45647 and sucrose, and, like bitter, there may be a TRPM5-independent transduction pathway for detecting these substances.

  11. 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.

  12. 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.

  13. Use of urinary pregnanediol 3-glucuronide to confirm ovulation.

    Science.gov (United States)

    Ecochard, R; Leiva, R; Bouchard, T; Boehringer, H; Direito, A; Mariani, A; Fehring, R

    2013-10-01

    Urinary hormonal markers may assist in increasing the efficacy of Fertility Awareness Based Methods (FABM). This study uses urinary pregnanediol-3a-glucuronide (PDG) testing to more accurately identify the infertile phase of the menstrual cycle in the setting of FABM. Secondary analysis of an observational and simulation study, multicentre, European study. The study includes 107 women and tracks daily first morning urine (FMU), observed the changes in cervical mucus discharge, and ultrasonography to identify the day of ovulation over 326 menstrual cycles. The following three scenarios were tested: (A) use of the daily pregnandiol-3a-glucuronide (PDG) test alone; (B) use of the PDG test after the first positive urine luteinizing hormone (LH) kit result; (C) use of the PDG test after the disappearance of fertile type mucus. Two models were used: (1) one day of PDG positivity; or (2) waiting for three days of PDG positivity before declaring infertility. After the first positivity of a LH test or the end of fertile mucus, three consecutive days of PDG testing over a threshold of 5μg/mL resulted in a 100% specificity for ovulation confirmation. They were respectively associated an identification of an average of 6.1 and 7.6 recognized infertile days. The results demonstrate a clinical scenario with 100% specificity for ovulation confirmation and provide the theoretical background for a future development of a competitive lateral flow assay for the detection of PDG in the urine. Copyright © 2013 Elsevier Inc. All rights reserved.

  14. 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...

  15. 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

    significant recovery from spared-nerve injury-induced mechanical hypersensitivity are two novel phenotypes which characterize the global CB1R-/- mice. An increase in transient receptor potential channel of melastatin 8 channel function in DRG neurons may underlie the cold phenotype. Recovery of mechanical thresholds in the CB1R knockouts was independent of motor function. These results indicate that CB1R expression contributes to the development of persistent mechanical hypersensitivity, protects against the development of robust cold allodynia but is not involved in motor impairment following spared-nerve injury in mice. © The Author(s) 2016.

  16. 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

  17. Minimal Effects of Age and Exposure to a Noisy Environment on Hearing in Alpha9 Nicotinic Receptor Knockout Mice

    Directory of Open Access Journals (Sweden)

    Amanda M. Lauer

    2017-06-01

    Full Text Available Studies have suggested a role of weakened medial olivocochlear (OC efferent feedback in accelerated hearing loss and increased susceptibility to noise. The present study investigated the progression of hearing loss with age and exposure to a noisy environment in medial OC-deficient mice. Alpha9 nicotinic acetylcholine receptor knockout (α9KO and wild types were screened for hearing loss using auditory brainstem responses. α9KO mice housed in a quiet environment did not show increased hearing loss compared to wild types in young adulthood and middle age. Challenging the medial OC system by housing in a noisy environment did not increase hearing loss in α9KO mice compared to wild types. ABR wave 1 amplitudes also did not show differences between α9KO mice and wild types. These data suggest that deficient medial OC feedback does not result in early onset of hearing loss.

  18. Gamma aminobutyric acid transporter subtype 1 gene knockout mice: a new model for attention deficit/hyperactivity disorder

    Institute of Scientific and Technical Information of China (English)

    Ping Yang; Guoqiang Cai; Youqing Cai; Jian Fei; Guoxiang Liu

    2013-01-01

    Attention deficit/hyperactivity disorder (ADHD) is characterized by hyperactivity,impaired sustained attention,impulsivity,and is usually accompanied by varying degrees of learning difficulties and lack of motor coordination.However,the pathophysiology and etiology of ADHD remain inconclusive so far.Our previous studies have demonstrated that the gamma aminobutyric acid transporter subtype 1 (GAT1) gene knockout (ko) mouse (gat1-/-)is hyperactive and exhibited impaired memory performance in the Morris water maze.In the current study,we found that the gat1-/-mice showed low levels of attentional focusing and increased impulsivity.In addition,the gat1-/-mice displayed ataxia characterized by defects in motor coordination and balance skills.The hyperactivity in the ko mice was reduced by both methylphenidate and amphetamine.Collectively,these results suggest that GAT1 ko mouse is a new animal model for ADHD studying and GAT1 may be a new target to treat ADHD.

  19. A homozygous Keap1-knockout human embryonic stem cell line generated using CRISPR/Cas9 mediates gene targeting

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    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.

  20. 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.

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

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    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

  2. Enhanced glucose tolerance in pancreatic-derived factor (PANDER knockout C57BL/6 mice

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    Shari L. Moak

    2014-11-01

    Full Text Available Pancreatic-derived factor (PANDER; also known as FAM3B is a uniquely structured protein strongly expressed within and secreted from the endocrine pancreas. PANDER has been hypothesized to regulate fasting and fed glucose homeostasis, hepatic lipogenesis and insulin signaling, and to serve a potential role in the onset or progression of type 2 diabetes (T2D. Despite having potentially pivotal pleiotropic roles in glycemic regulation and T2D, there has been limited generation of stable animal models for the investigation of PANDER function, and there are no models on well-established genetic murine backgrounds for T2D. Our aim was to generate an enhanced murine model to further elucidate the biological function of PANDER. Therefore, a pure-bred PANDER knockout C57BL/6 (PANKO-C57 model was created and phenotypically characterized with respect to glycemic regulation and hepatic insulin signaling. The PANKO-C57 model exhibited an enhanced metabolic phenotype, particularly with regard to enhanced glucose tolerance. Male PANKO-C57 mice displayed decreased fasting plasma insulin and C-peptide levels, whereas leptin levels were increased as compared with matched C57BL/6J wild-type mice. Despite similar peripheral insulin sensitivity between both groups, hepatic insulin signaling was significantly increased during fasting conditions, as demonstrated by increased phosphorylation of hepatic PKB/Akt and AMPK, along with mature SREBP-1 expression. Insulin stimulation of PANKO-C57 mice resulted in increased hepatic triglyceride and glycogen content as compared with wild-type C57BL/6 mice. In summary, the PANKO-C57 mouse represents a suitable model for the investigation of PANDER in multiple metabolic states and provides an additional tool to elucidate the biological function and potential role in T2D.

  3. Highly efficient generation of GGTA1 biallelic knockout inbred mini-pigs with TALENs.

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    Jige Xin

    Full Text Available Inbred mini-pigs are ideal organ donors for future human xenotransplantations because of their clear genetic background, high homozygosity, and high inbreeding endurance. In this study, we chose fibroblast cells from a highly inbred pig line called Banna mini-pig inbred line (BMI as donor nuclei for nuclear transfer, combining with transcription activator-like effector nucleases (TALENs and successfully generated α-1,3-galactosyltransferase (GGTA1 gene biallelic knockout (KO pigs. To validate the efficiency of TALEN vectors, in vitro-transcribed TALEN mRNAs were microinjected into one-cell stage parthenogenetically activated porcine embryos. The efficiency of indel mutations at the GGTA1-targeting loci was as high as 73.1% (19/26 among the parthenogenetic blastocysts. TALENs were co-transfected into porcine fetal fibroblasts of BMI with a plasmid containing neomycin gene. The targeting efficiency reached 89.5% (187/209 among the survived cell clones after a 10 d selection. More remarkably 27.8% (58/209 of colonies were biallelic KO. Five fibroblast cell lines with biallelic KO were chosen as nuclear donors for somatic cell nuclear transfer (SCNT. Three miniature piglets with biallelic mutations of the GGTA1 gene were achieved. Gal epitopes on the surface of cells from all the three biallelic KO piglets were completely absent. The fibroblasts from the GGTA1 null piglets were more resistant to lysis by pooled complement-preserved normal human serum than those from wild-type pigs. These results indicate that a combination of TALENs technology with SCNT can generate biallelic KO pigs directly with high efficiency. The GGTA1 null piglets with inbred features created in this study can provide a new organ source for xenotransplantation research.

  4. Altered Morphology and Function of the Lacrimal Functional Unit in Protein Kinase Cα Knockout Mice

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    Chen, Zhuo; Li, Zhijie; Basti, Surendra; Farley, William J.

    2010-01-01

    Purpose. Protein kinase C (PKC) α plays a major role in the parasympathetic neural stimulation of lacrimal gland (LG) secretion. It also has been reported to have antiapoptotic properties and to promote cell survival. Therefore, the hypothesis for the present study was that PKCα knockout (−/−) mice have impaired ocular surface–lacrimal gland signaling, rendering them susceptible to desiccating stress and impaired corneal epithelial wound healing. In this study, the lacrimal function unit (LFU) and the stressed wound-healing response were examined in PKCα−/− mice. Methods. In PKCα+/+ control mice and PKCα−/− mice, tear production, osmolarity, and clearance rate were evaluated before and after experimental desiccating stress. Histology and immunofluorescent staining of PKC and epidermal growth factor were performed in tissues of the LFU. Cornified envelope (CE) precursor protein expression and cell proliferation were evaluated. The time course of healing and degree of neutrophil infiltration was evaluated after corneal epithelial wounding. Results. Compared with the PKCα+/+ mice, the PKCα−/− mice were noted to have significantly increased lacrimal gland weight, with enlarged, carbohydrate-rich, PAS-positive acinar cells; increased corneal epithelia permeability, with reduced CE expression; and larger conjunctival epithelial goblet cells. The PKCα−/− mice showed more rapid corneal epithelial healing, with less neutrophil infiltration and fewer proliferating cells than did the PKCα+/+ mice. Conclusions. The PKCα−/− mice showed lower tear production, which appeared to be caused by impaired secretion by the LG and conjunctival goblet cells. Despite their altered tear dynamics, the PKCα−/− mice demonstrated more rapid corneal epithelial wound healing, perhaps due to decreased neutrophil infiltration. PMID:20505191

  5. Knockout Serum Replacement Promotes Cell Survival by Preventing BIM from Inducing Mitochondrial Cytochrome C Release.

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    Yuki Ishii

    Full Text Available Knockout serum replacement (KOSR is a nutrient supplement commonly used to replace serum for culturing stem cells. We show here that KOSR has pro-survival activity in chronic myelogenous leukemia (CML cells transformed by the BCR-ABL oncogene. Inhibitors of BCR-ABL tyrosine kinase kill CML cells by stimulating pro-apoptotic BIM and inhibiting anti-apoptotic BCL2, BCLxL and MCL1. We found that KOSR protects CML cells from killing by BCR-ABL inhibitors--imatinib, dasatinib and nilotinib. The protective effect of KOSR is reversible and not due to the selective outgrowth of drug-resistant clones. In KOSR-protected CML cells, imatinib still inhibited the BCR-ABL tyrosine kinase, reduced the phosphorylation of STAT, ERK and AKT, down-regulated BCL2, BCLxL, MCL1 and up-regulated BIM. However, these pro-apoptotic alterations failed to cause cytochrome c release from the mitochondria. With mitochondria isolated from KOSR-cultured CML cells, we showed that addition of recombinant BIM protein also failed to cause cytochrome c release. Besides the kinase inhibitors, KOSR could protect cells from menadione, an inducer of oxidative stress, but it did not protect cells from DNA damaging agents. Switching from serum to KOSR caused a transient increase in reactive oxygen species and AKT phosphorylation in CML cells that were protected by KOSR but not in those that were not protected by this nutrient supplement. Treatment of KOSR-cultured cells with the PH-domain inhibitor MK2206 blocked AKT phosphorylation, abrogated the formation of BIM-resistant mitochondria and stimulated cell death. These results show that KOSR has cell-context dependent pro-survival activity that is linked to AKT activation and the inhibition of BIM-induced cytochrome c release from the mitochondria.

  6. Tissue inhibitor of metalloproteinase-3 knockout mice exhibit enhanced energy expenditure through thermogenesis.

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    Yohsuke Hanaoka

    Full Text Available Tissue inhibitors of metalloproteinases (TIMPs regulate matrix metalloproteinase activity and maintain extracellular matrix homeostasis. Although TIMP-3 has multiple functions (e.g., apoptosis, inhibition of VEGF binding to VEGF receptor, and inhibition of TNFα converting enzyme, its roles in thermogenesis and metabolism, which influence energy expenditure and can lead to the development of metabolic disorders when dysregulated, are poorly understood. This study aimed to determine whether TIMP-3 is implicated in metabolism by analyzing TIMP-3 knockout (KO mice. TIMP-3 KO mice had higher body temperature, oxygen consumption, and carbon dioxide production than wild-type (WT mice, although there were no differences in food intake and locomotor activity. These results suggest that metabolism is enhanced in TIMP-3 KO mice. Real-time PCR analysis showed that the expression of PPAR-δ, UCP-2, NRF-1 and NRF-2 in soleus muscle, and PGC-1α and UCP-2 in gastrocnemius muscle, was higher in TIMP-3 KO mice than in WT mice, suggesting that TIMP-3 deficiency may increase mitochondrial activity. When exposed to cold for 8 hours to induce thermogenesis, TIMP-3 KO mice had a higher body temperature than WT mice. In the treadmill test, oxygen consumption and carbon dioxide production were higher in TIMP-3 KO mice both before and after starting exercise, and the difference was more pronounced after starting exercise. Our findings suggest that TIMP-3 KO mice exhibit enhanced metabolism, as reflected by a higher body temperature than WT mice, possibly due to increased mitochondrial activity. Given that TIMP-3 deficiency increases energy expenditure, TIMP-3 may present a novel therapeutic target for preventing metabolic disorders.

  7. Altered social behaviours in neurexin 1α knockout mice resemble core symptoms in neurodevelopmental disorders.

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    Hannah Mary Grayton

    Full Text Available BACKGROUND: Copy number variants have emerged as an important genomic cause of common, complex neurodevelopmental disorders. These usually change copy number of multiple genes, but deletions at 2p16.3, which have been associated with autism, schizophrenia and mental retardation, affect only the neurexin 1 gene, usually the alpha isoform. Previous analyses of neurexin 1α (Nrxn1α knockout (KO mouse as a model of these disorders have revealed impairments in synaptic transmission but failed to reveal defects in social behaviour, one of the core symptoms of autism. METHODS: We performed a detailed investigation of the behavioural effects of Nrxn1α deletion in mice bred onto a pure genetic background (C57BL/6J to gain a better understanding of its role in neurodevelopmental disorders. Wildtype, heterozygote and homozygote Nrxn1α KO male and female mice were tested in a battery of behavioural tests (n = 9-16 per genotype, per sex. RESULTS: In homozygous Nrxn1α KO mice, we observed altered social approach, reduced social investigation, and reduced locomotor activity in novel environments. In addition, male Nrxn1α KO mice demonstrated an increase in aggressive behaviours. CONCLUSIONS: These are the first experimental data that associate a deletion of Nrxn1α with alterations of social behaviour in mice. Since this represents one of the core symptom domains affected in autism spectrum disorders and schizophrenia in humans, our findings suggest that deletions within NRXN1 found in patients may be responsible for the impairments seen in social behaviours, and that the Nrxn1α KO mice are a useful model of human neurodevelopmental disorder.

  8. Experimental evidence for the involvement of PDLIM5 in mood disorders in hetero knockout mice.

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    Yasue Horiuchi

    Full Text Available BACKGROUND: Reports indicate that PDLIM5 is involved in mood disorders. The PDLIM5 (PDZ and LIM domain 5 gene has been genetically associated with mood disorders; it's expression is upregulated in the postmortem brains of patients with bipolar disorder and downregulated in the peripheral lymphocytes of patients with major depression. Acute and chronic methamphetamine (METH administration may model mania and the evolution of mania into psychotic mania or schizophrenia-like behavioral changes, respectively. METHODS: To address whether the downregulation of PDLIM5 protects against manic symptoms and cause susceptibility to depressive symptoms, we evaluated the effects of reduced Pdlim5 levels on acute and chronic METH-induced locomotor hyperactivity, prepulse inhibition, and forced swimming by using Pdlim5 hetero knockout (KO mice. RESULTS: The homozygous KO of Pdlim5 is embryonic lethal. The effects of METH administration on locomotor hyperactivity and the impairment of prepulse inhibition were lower in Pdlim5 hetero KO mice than in wild-type mice. The transient inhibition of PDLIM5 (achieved by blocking the translocation of protein kinase C epsilon before the METH challenge had a similar effect on behavior. Pdlim5 hetero KO mice showed increased immobility time in the forced swimming test, which was diminished after the chronic administration of imipramine. Chronic METH treatment increased, whereas chronic haloperidol treatment decreased, Pdlim5 mRNA levels in the prefrontal cortex. Imipramine increased Pdlim5 mRNA levels in the hippocampus. CONCLUSION: These findings are partially compatible with reported observations in humans, indicating that PDLIM5 is involved in psychiatric disorders, including mood disorders.

  9. Metabolomic profiles of arsenic (+3 oxidation state) methyltransferase knockout mice: Effect of sex and arsenic exposure

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    Huang, Madelyn C.; Douillet, Christelle; Su, Mingming; Zhou, Kejun; Wu, Tao; Chen, Wenlian; Galanko, Joseph A.; Drobná, Zuzana; Saunders, R. Jesse; Martin, Elizabeth; Fry, Rebecca C.; Jia, Wei; Stýblo, Miroslav

    2016-01-01

    Arsenic (+3 oxidation state) methyltransferase (As3mt) is the key enzyme in the pathway for methylation of inorganic arsenic (iAs). Altered As3mt expression and AS3MT polymorphism have been linked to changes in iAs metabolism and in susceptibility to iAs toxicity in laboratory models and in humans. As3mt-knockout mice have been used to study the association between iAs metabolism and adverse effects of iAs exposure. However, little is known about systemic changes in metabolism of these mice and how these changes lead to their increased susceptibility to iAs toxicity. Here, we compared plasma and urinary metabolomes of male and female wild-type (WT) and As3mt-KO (KO) C57BL6 mice and examined metabolomic shifts associated with iAs exposure in drinking water. Surprisingly, exposure to 1 ppm As elicited only small changes in the metabolite profiles of either WT or KO mice. In contrast, comparisons of KO mice with WT mice revealed significant differences in plasma and urinary metabolites associated with lipid (phosphatidylcholines, cytidine, acyl-carnitine), amino acid (hippuric acid, acetylglycine, urea), and carbohydrate (L-sorbose, galactonic acid, gluconic acid) metabolism. Notably, most of these differences were sex-specific. Sex-specific differences were also found between WT and KO mice in plasma triglyceride and lipoprotein cholesterol levels. Some of the differentially changed metabolites (phosphatidylcholines, carnosine, and sarcosine) are substrates or products of reactions catalyzed by other methyltransferases. These results suggest that As3mt KO alters major metabolic pathways in a sex-specific manner, independent of iAs treatment, and that As3mt may be involved in other cellular processes beyond iAs methylation. PMID:26883664

  10. Impact of food restriction and cocaine on locomotion in ghrelin- and ghrelin-receptor knockout mice.

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    Clifford, Shane; Zeckler, Rosie Albarran; Buckman, Sam; Thompson, Jeff; Hart, Nigel; Wellman, Paul J; Smith, Roy G

    2011-07-01

    Food restriction (FR) augments the behavioral and reinforcing effects of psychomotor stimulants such as cocaine or amphetamine; effects that may be related to the capacity of FR to increase plasma levels of ghrelin (GHR), a 28-amino acid orexigenenic peptide linked to activation of brain dopamine systems. The present study used wild-type (WT) mice or mutant mice sustaining knockout of either GHR [GHR((-/-)) ] or of the growth hormone secretagogue receptor [GHS-R((-/-)) ] and subjected to FR or not to evaluate the role of GHR and GHS-R in cocaine-stimulated locomotion. WT, GHR((-/-)) , and GHS-R((-/-)) mice were either restricted to 60% of baseline caloric intake or allowed to free-feed (FF). Mice were treated with 0, 1.25, 2.5 and 5.0 mg/kg cocaine on separate test days (in random dose order) and forward locomotion was recorded on each drug day for 45 minutes after drug dosing. Food (and water) was available immediately after (but not during) each activity test. For FF mice, there was no interaction between cocaine and GHR status on locomotion. FR-WT mice treated with saline exhibited significant increases in anticipatory locomotion (relative to FF-WT mice), whereas FR-GHS-R((-/-)) mice did not. Cocaine significantly increased locomotion in FR-GHR((-/-)) and FR-GHS-R((-/-)) mice to the levels noted in FR-WT mice. These results suggest that GHS-R activity, but not GHR activity, is required for FR to augment food-associated anticipatory locomotion, but do not support the contention that GHR pathways are required for the capacity of FR to augment the acute effect of cocaine on locomotion. © 2010 The Authors, Addiction Biology © 2010 Society for the Study of Addiction.

  11. Decreased Neointimal Extracellular Matrix Formation in RAGE-Knockout Mice After Microvascular Denudation

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    Groezinger, Gerd, E-mail: gerd.groezinger@med.uni-tuebingen.de; Schmehl, Joerg, E-mail: joerg.schmehl@med.uni-tuebingen.de; Bantleon, Ruediger, E-mail: ruediger.bantleon@med.uni-tuebingen.de; Kehlbach, Rainer, E-mail: rainer.kehlbach@uni-tuebingen.de [University of Tuebingen, Department of Diagnostic and Interventional Radiology (Germany); Mehra, Tarun, E-mail: tarun.mehra@med.uni-tuebingen.de [University of Tuebingen, Department of Dermatology (Germany); Claussen, Claus, E-mail: gerd.groezinger@med.uni-tuebingen.de; Wiesinger, Benjamin, E-mail: benjamin.wiesinger@med.uni-tuebingen.de [University of Tuebingen, Department of Diagnostic and Interventional Radiology (Germany)

    2012-12-15

    Purpose: To evaluate in vivo the role of RAGE (receptor for advanced glycated end products) in the development of restenosis and neointimal proliferation in RAGE-deficient knockout (KO) mice compared with wild-type (WT) mice in an animal model. Materials and Methods: Sixteen WT and 15 RAGE-deficient mice underwent microvascular denudation of the common femoral artery under general anaesthesia. Contralateral arteries underwent a sham operation and served as controls. Four weeks after the intervention, all animals were killed, and paraformaldehyde-fixed specimens of the femoral artery were analysed with different stains (hematoxylin and eosin and Elastica van Gieson) and several different types of immunostaining (proliferating cell nuclear antigen, {alpha}-actin, collagen, von Willebrand factor, RAGE). Luminal area, area of the neointima, and area of the media were measured in all specimens. In addition, colony-formation assays were performed, and collagen production by WT smooth muscle cells (SMCs) and RAGE-KO SMCs was determined. For statistical analysis, P < 0.05 was considered statistically significant. Results: Four weeks after denudation, WT mice showed a 49.6% loss of luminal area compared with 14.9% loss of luminal area in RAGE-deficient mice (sham = 0% loss) (P < 0.001). The neointima was 18.2 (*1000 {mu}m{sup 2} [n = 15) in the WT group compared with only 8.4 (*1000 {mu}m{sup 2} [n = 16]) in the RAGE-KO group. RAGE-KO SMCs showed significantly decreased proliferation activity and production of extracellular matrix protein. Conclusion: RAGE may be shown to play a considerable role in the formation of neointima leading to restenosis after vascular injury.

  12. Comprehensive behavioral analysis of pituitary adenylate cyclase-activating polypeptide (PACAP knockout mice

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    Satoko eHattori

    2012-10-01

    Full Text Available Pituitary adenylate cyclase-activating polypeptide (PACAP is a neuropeptide acting as a neurotransmitter, neuromodulator, or neurotrophic factor. PACAP is widely expressed throughout the brain and exerts its functions through the PACAP-specific receptor (PAC1. Recent studies reveal that genetic variants of the PACAP and PAC1 genes are associated with mental disorders, and several behavioral abnormalities of PACAP knockout (KO mice are reported. However, an insufficient number of backcrosses was made using PACAP KO mice on the C57BL/6J background due to their postnatal mortality. To elucidate the effects of PACAP on neuropsychiatric function, the PACAP gene was knocked out in F1 hybrid mice (C57BL/6J x 129SvEv for appropriate control of the genetic background. The PACAP KO mice were then subjected to a behavioral test battery. PACAP deficiency had no significant effects on neurological screen. As shown previously, the mice exhibited significantly increased locomotor activity in a novel environment and abnormal anxiety-like behavior, while no obvious differences between genotypes were shown in home cage activity. In contrast to previous reports, the PACAP KO mice showed normal prepulse inhibition and slightly decreased depression-like behavior. Previous study demonstrates that the social interaction in a resident-intruder test was decreased in PACAP KO mice. On the other hand, we showed that PACAP KO mice exhibited increased social interaction in Crawley’s three-chamber social approach test, although PACAP KO had no significant impact on social interaction in a home cage. PACAP KO mice also exhibited mild performance deficit in working memory in an eight-arm radial maze and the T-maze, while they did not show any significant abnormalities in the left-right discrimination task in the T-maze. These results suggest that PACAP has an important role in the regulation of locomotor activity, social behavior, anxiety-like behavior and, potentially

  13. Single-neutron knockout from 20C and the structure of 19C

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    J.W. Hwang

    2017-06-01

    Full Text Available The low-lying unbound level structure of the halo nucleus 19C has been investigated using single-neutron knockout from 20C on a carbon target at 280 MeV/nucleon. The invariant mass spectrum, derived from the momenta of the forward going beam velocity 18C fragment and neutrons, was found to be dominated by a very narrow near threshold (Erel=0.036(1 MeV peak. Two less strongly populated resonance-like features were also observed at Erel=0.84(4 and 2.31(3 MeV, both of which exhibit characteristics consistent with neutron p-shell hole states. Comparisons of the energies, measured cross sections and parallel momentum distributions to the results of shell-model and eikonal reaction calculations lead to spin-parity assignments of 5/21+ and 1/21− for the levels at Ex=0.62(9 and 2.89(10 MeV with Sn=0.58(9 MeV. Spectroscopic factors were also deduced and found to be in reasonable accord with shell-model calculations. The valence neutron configuration of the 20C ground state is thus seen to include, in addition to the known 1s1/22 component, a significant 0d5/22 contribution. The level scheme of 19C, including significantly the 1/21− cross-shell state, is well accounted for by the YSOX shell-model interaction developed from the monopole-based universal interaction.

  14. Somatostatin receptor 1 and 5 double knockout mice mimic neurochemical changes of Huntington's disease transgenic mice.

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    Padmesh S Rajput

    Full Text Available Selective degeneration of medium spiny neurons and preservation of medium sized aspiny interneurons in striatum has been implicated in excitotoxicity and pathophysiology of Huntington's disease (HD. However, the molecular mechanism for the selective sparing of medium sized aspiny neurons and vulnerability of projection neurons is still elusive. The pathological characteristic of HD is an extensive reduction of the striatal mass, affecting caudate putamen. Somatostatin (SST positive neurons are selectively spared in HD and Quinolinic acid/N-methyl-D-aspartic acid induced excitotoxicity, mimic the model of HD. SST plays neuroprotective role in excitotoxicity and the biological effects of SST are mediated by five somatostatin receptor subtypes (SSTR1-5.To delineate subtype selective biological responses we have here investigated changes in SSTR1 and 5 double knockout mice brain and compared with HD transgenic mouse model (R6/2. Our study revealed significant loss of dopamine and cAMP regulated phosphoprotein of 32 kDa (DARPP-32 and comparable changes in SST, N-methyl-D-aspartic acid receptors subtypes, calbindin and brain nitric oxide synthase expression as well as in key signaling proteins including calpain, phospho-extracellular-signal-regulated kinases1/2, synapsin-IIa, protein kinase C-α and calcineurin in SSTR1/5(-/- and R6/2 mice. Conversely, the expression of somatostatin receptor subtypes, enkephalin and phosphatidylinositol 3-kinases were strain specific. SSTR1/5 appears to be important in regulating NMDARs, DARPP-32 and signaling molecules in similar fashion as seen in HD transgenic mice.This is the first comprehensive description of disease related changes upon ablation of G- protein coupled receptor gene. Our results indicate that SST and SSTRs might play an important role in regulation of neurodegeneration and targeting this pathway can provide a novel insight in understanding the pathophysiology of Huntington's disease.

  15. Host resistance of CD18 knockout mice against systemic infection with Listeria monocytogenes

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    Wu, Huaizhu; Prince, Joseph E.; Brayton, Cory F.; Shah, Chirayu; Zeve, Daniel; Gregory, Stephen H.; Smith, C. Wayne; Ballantyne, Christie M.

    2003-01-01

    Mice with targeted mutations of CD18, the common beta2 subunit of CD11/CD18 integrins, have leukocytosis, impaired transendothelial neutrophil emigration, and reduced host defense to Streptococcus pneumoniae, a gram-positive extracellular bacterium. Previous studies using blocking monoclonal antibodies suggested roles for CD18 and CD11b in hepatic neutrophil recruitment and host innate response to Listeria monocytogenes, a gram-positive intracellular bacterium. We induced systemic listeriosis in CD18 knockout (CD18-ko) and wild-type (WT) mice by tail vein injection with Listeria. By 14 days postinjection (dpi), 8 of 10 WT mice died, compared with 2 of 10 CD18-ko mice (P Listeria organisms in livers and spleens were similar in both groups at 20 min postinfection. By 3, 5, and 7 dpi, however, numbers of Listeria organisms were significantly lower in livers and spleens of CD18-ko mice than in WT mice. Histopathology showed that following Listeria infection, CD18-ko mice had milder inflammatory and necrotizing lesions in both spleens and livers than did WT mice. Cytokine assays indicated that baseline interleukin-1beta and granulocyte colony-stimulating factor (G-CSF) levels were higher in CD18-ko mice than in WT mice and that CD18-ko splenocytes produced higher levels of interleukin-1beta and G-CSF than WT splenocytes under the same amount of Listeria stimulation. These findings show that CD18 is not an absolute requirement for antilisterial innate immunity or hepatic neutrophil recruitment. We propose that the absence of CD18 in the mice results in the priming of innate immunity, as evidenced by elevated cytokine expression, and neutrophilic leukocytosis, which augments antilisterial defense.

  16. Large-conductance Ca2+-activated K+ channel β1-subunit knockout mice are not hypertensive

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    Garver, Hannah; Galligan, James J.; Fink, Gregory D.

    2011-01-01

    Large-conductance Ca2+-activated K+ (BK) channels are composed of pore-forming α-subunits and accessory β1-subunits that modulate Ca2+ sensitivity. BK channels regulate arterial myogenic tone and renal Na+ clearance/K+ reabsorption. Previous studies using indirect or short-term blood pressure measurements found that BK channel β1-subunit knockout (BK β1-KO) mice were hypertensive. We evaluated 24-h mean arterial pressure (MAP) and heart rate in BK β1-KO mice using radiotelemetry. BK β1-KO mice did not have a higher 24-h average MAP when compared with wild-type (WT) mice, although MAP was ∼10 mmHg higher at night. The dose-dependent peak declines in MAP by nifedipine were only slightly larger in BK β1-KO mice. In BK β1-KO mice, giving 1% NaCl to mice to drink for 7 days caused a transient (5 days) elevation of MAP (∼5 mmHg); MAP returned to pre-saline levels by day 6. BK β1-KO mesenteric arteries in vitro demonstrated diminished contractile responses to paxilline, increased reactivity to Bay K 8644 and norepinephrine (NE), and maintained relaxation to isoproterenol. Paxilline and Bay K 8644 did not constrict WT or BK β1-KO mesenteric veins (MV). BK β1-subunits are not expressed in MV. The results indicate that BK β1-KO mice are not hypertensive on normal or high-salt intake. BK channel deficiency increases arterial reactivity to NE and L-type Ca2+ channel function in vitro, but the L-type Ca2+ channel modulation of MAP is not altered in BK β1-KO mice. BK and L-type Ca2+ channels do not modulate murine venous tone. It appears that selective loss of BK channel function in arteries only is not sufficient to cause sustained hypertension. PMID:21131476

  17. Altered Social Behaviours in Neurexin 1α Knockout Mice Resemble Core Symptoms in Neurodevelopmental Disorders

    Science.gov (United States)

    Grayton, Hannah Mary; Missler, Markus

    2013-01-01

    Background Copy number variants have emerged as an important genomic cause of common, complex neurodevelopmental disorders. These usually change copy number of multiple genes, but deletions at 2p16.3, which have been associated with autism, schizophrenia and mental retardation, affect only the neurexin 1 gene, usually the alpha isoform. Previous analyses of neurexin 1α (Nrxn1α) knockout (KO) mouse as a model of these disorders have revealed impairments in synaptic transmission but failed to reveal defects in social behaviour, one of the core symptoms of autism. Methods We performed a detailed investigation of the behavioural effects of Nrxn1α deletion in mice bred onto a pure genetic background (C57BL/6J) to gain a better understanding of its role in neurodevelopmental disorders. Wildtype, heterozygote and homozygote Nrxn1α KO male and female mice were tested in a battery of behavioural tests (n = 9–16 per genotype, per sex). Results In homozygous Nrxn1α KO mice, we observed altered social approach, reduced social investigation, and reduced locomotor activity in novel environments. In addition, male Nrxn1α KO mice demonstrated an increase in aggressive behaviours. Conclusions These are the first experimental data that associate a deletion of Nrxn1α with alterations of social behaviour in mice. Since this represents one of the core symptom domains affected in autism spectrum disorders and schizophrenia in humans, our findings suggest that deletions within NRXN1 found in patients may be responsible for the impairments seen in social behaviours, and that the Nrxn1α KO mice are a useful model of human neurodevelopmental disorder. PMID:23840597

  18. Reduced emotional and corticosterone responses to stress in μ-opioid receptor knockout mice

    Science.gov (United States)

    Ide, Soichiro; Sora, Ichiro; Ikeda, Kazutaka; Minami, Masabumi; Uhl, George R.; Ishihara, Kumatoshi

    2014-01-01

    The detailed mechanisms of emotional modulation in the nervous system by opioids remain to be elucidated, although the opioid system is well known to play important roles in the mechanisms of analgesia and drug dependence. In the present study, we conducted behavioral tests of anxiety and depression and measured corticosterone concentrations in both male and female μ-opioid receptor knockout (MOP-KO) mice to reveal the involvement of μ-opioid receptors in stress-induced emotional responses. MOP-KO mice entered more and spent more time in the open arms of the elevated plus maze compared with wild-type mice. MOP-KO mice also displayed significantly decreased immobility in a 15 min tail-suspension test compared with wild-type mice. Similarly, MOP-KO mice exhibited significantly decreased immobility on days 2, 3, and 4 in a 6 min forced swim test conducted for 5 consecutive days. The increase in plasma corticosterone concentration induced by tail-suspension, repeated forced swim, or restraint stress was reduced in MOP-KO mice compared with wild-type mice. Corticosterone levels were not different between wild-type and MOP-KO mice before stress exposure. In contrast, although female mice tended to exhibit fewer anxiety-like responses in the tail-suspension test in both genotypes, no significant gender differences were observed in stress-induced emotional responses. These results suggest that MOPs play an important facilitatory role in emotional responses to stress, including anxiety- and depression-like behavior and corticosterone levels. PMID:19596019

  19. Ghrelin knockout mice show decreased voluntary alcohol consumption and reduced ethanol-induced conditioned place preference.

    Science.gov (United States)

    Bahi, Amine; Tolle, Virginie; Fehrentz, Jean-Alain; Brunel, Luc; Martinez, Jean; Tomasetto, Catherine-Laure; Karam, Sherif M

    2013-05-01

    Recent work suggests that stomach-derived hormone ghrelin receptor (GHS-R1A) antagonism may reduce motivational aspects of ethanol intake. In the current study we hypothesized that the endogenous GHS-R1A agonist ghrelin modulates alcohol reward mechanisms. For this purpose ethanol-induced conditioned place preference (CPP), ethanol-induced locomotor stimulation and voluntary ethanol consumption in a two-bottle choice drinking paradigm were examined under conditions where ghrelin and its receptor were blocked, either using ghrelin knockout (KO) mice or the specific ghrelin receptor (GHS-R1A) antagonist "JMV2959". We showed that ghrelin KO mice displayed lower ethanol-induced CPP than their wild-type (WT) littermates. Consistently, when injected during CPP-acquisition, JMV2959 reduced CPP-expression in C57BL/6 mice. In addition, ethanol-induced locomotor stimulation was lower in ghrelin KO mice. Moreover, GHS-R1A blockade, using JMV2959, reduced alcohol-stimulated locomotion only in WT but not in ghrelin KO mice. When alcohol consumption and preference were assessed using the two-bottle choice test, both genetic deletion of ghrelin and pharmacological antagonism of the GHS-R1A (JMV2959) reduced voluntary alcohol consumption and preference. Finally, JMV2959-induced reduction of alcohol intake was only observed in WT but not in ghrelin KO mice. Taken together, these results suggest that ghrelin neurotransmission is necessary for the stimulatory effect of ethanol to occur, whereas lack of ghrelin leads to changes that reduce the voluntary intake as well as conditioned reward by ethanol. Our findings reveal a major, novel role for ghrelin in mediating ethanol behavior, and add to growing evidence that ghrelin is a key mediator of the effects of multiple abused drugs. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. 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

  1. 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.

  2. Contribution of PPARγ in modulation of acrolein-induced inflammatory signaling in gp91phox knock-out mice.

    Science.gov (United States)

    Yousefipour, Zivar; Chug, Neha; Marek, Katarzyna; Nesbary, Alicia; Mathew, Joseph; Ranganna, Kasturi; Newaz, Mohammad A

    2017-08-01

    Oxidative stress and inflammation are major contributors to acrolein toxicity. Peroxisome proliferator activated receptor gamma (PPARγ) has antioxidant and anti-inflammatory effects. We investigated the contribution of PPARγ ligand GW1929 to the attenuation of oxidative stress in acrolein-induced insult. Male gp91 phox knock-out (KO) mice were treated with acrolein (0.5 mg·(kg body mass) -1 by intraperitoneal injection for 7 days) with or without GW1929 (GW; 0.5 mg·(kg body mass) -1 ·day -1 , orally, for 10 days). The livers were processed for further analyses. Acrolein significantly increased 8-isoprostane and reduced PPARγ activity (P acrolein-treated WT mice, and was reduced by GW1929 (by 65%). KO mice exhibited higher xanthine oxidase (XO). Acrolein increased XO and COX in WT mice and XO in KO mice. GW1929 significantly reduced COX in WT and KO mice and reduced XO in KO mice. Acrolein significantly reduced the total antioxidant status in WT and KO mice (P acrolein-treated WT mice. GW1929 reduced NF-κB levels (by 51%) in KO mice. Acrolein increased CD36 in KO mice (by 43%), which was blunted with GW1929. Data confirms that the generation of free radicals by acrolein is mainly through NAD(P)H, but other oxygenates play a role too. GW1929 may alleviate the toxicity of acrolein by attenuating NF-κB, COX, and CD36.

  3. Analytical approach for confirming the achievement of LMFBR reliability goals

    International Nuclear Information System (INIS)

    Ingram, G.E.; Elerath, J.G.; Wood, A.P.

    1981-01-01

    The approach, recommended by GE-ARSD, for confirming the achievement of LMFBR reliability goals relies upon a comprehensive understanding of the physical and operational characteristics of the system and the environments to which the system will be subjected during its operational life. This kind of understanding is required for an approach based on system hardware testing or analyses, as recommended in this report. However, for a system as complex and expensive as the LMFBR, an approach which relies primarily on system hardware testing would be prohibitive both in cost and time to obtain the required system reliability test information. By using an analytical approach, results of tests (reliability and functional) at a low level within the specific system of interest, as well as results from other similar systems can be used to form the data base for confirming the achievement of the system reliability goals. This data, along with information relating to the design characteristics and operating environments of the specific system, will be used in the assessment of the system's reliability

  4. Confirmation test of powder mixing process in J-MOX

    International Nuclear Information System (INIS)

    Ota, Hiroshi; Osaka, Shuichi; Kurita, Ichiro

    2009-01-01

    Japan Nuclear Fuel Ltd. (hereafter, JNFL) MOX Fuel Fabrication Plant (hereafter, J-MOX) is what fabricates MOX fuel for domestic light water power plants. Development of design concept of J-MOX was started mid 90's and the frame of J-MOX process was clarified around 2000 including adoption of MIMAS process as apart of J-MOX powder process. JNFL requires to take an answer to any technical question that has not been clarified ever before by world's MOX and/or Uranium fabricators before it commissions equipment procurement. J-MOX is to be constructed adjacent to the Rokkasho Reprocessing Plant (RRP) and to utilize MH-MOX powder recovered at RRP. The combination of the MIMAS process and the MH-MOX powder is what has never tried in the world. Therefore JNFL started a series of confirmation tests of which the most important is the powder test to confirm the applicability of MH-MOX powder to the MIMAS process. The MH-MOX powder, consisting of 50% plutonium oxide and 50% uranium oxide, originates JAEA development utilizing microwave heating (MH) technology. The powder test started with laboratory scale small equipment utilizing both uranium and the MOX powder in 2000, left a solution to tough problem such as powder adhesion onto equipment, and then was followed by a large scale equipment test again with uranium and the MOX powder. For the MOX test, actual size equipment within glovebox was manufactured and installed in JAEA plutonium fuel center in 2005, and based on results taken so far an understanding that the MIMAS equipment, with the MH-MOX powder, can present almost same quality MOX pellet as what is introduced as fabricated in Europe was developed. The test was finished at the end of Japanese fiscal year (JFY) 2007, and it was confirmed that the MOX pellets fabricated in this test were almost satisfied with the targeted specifications set for domestic LWR MOX fuels. (author)

  5. 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.)

  6. 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.

  7. 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.

  8. 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.

  9. 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

  10. Isotopic Generation and Confirmation of the PWR Application Model?

    International Nuclear Information System (INIS)

    L.B. Wimmer

    2003-01-01

    The objective of this calculation is to establish an isotopic database to represent commercial spent nuclear fuel (CSNF) from pressurized water reactors (PWRs) in criticality analyses performed for the proposed Monitored Geologic Repository at Yucca Mountain, Nevada. Confirmation of the conservatism with respect to criticality in the isotopic concentration values represented by this isotopic database is performed as described in Section 3.5.3.1.2 of the ''Disposal Criticality Analysis Methodology Topical Report'' (YMP 2000). The isotopic database consists of the set of 14 actinides and 15 fission products presented in Section 3.5.2.1.1 of YMP 2000 for use in CSNF burnup credit. This set of 29 isotopes is referred to as the principal isotopes. The oxygen isotope from the UO 2 fuel is also included in the database. The isotopic database covers enrichments of 235 U ranging from 1.5 to 5.5 weight percent (wt%) and burnups ranging from approximately zero to 75 GWd per metric ton of uranium (mtU). The choice of fuel assembly and operating history values used in generating the isotopic database are provided is Section 5. Tables of isotopic concentrations for the 29 principal isotopes (plus oxygen) as a function of enrichment and burnup are provided in Section 6.1. Results of the confirmation of the conservatism with respect to criticality in the isotopic concentration values are provided in Section 6.2

  11. Carcinogenic effects of riddelliine on P53 knockout mice

    Science.gov (United States)

    Riddelliine is a pyrrolizidine alkaloid found in Senecio riddellii and several other Senecio spp. Pyrrolizidine alkaloids are a group of over 600 toxins, found in more than 6,000 plants worldwide. As a result they are likely the most economically significant plant toxin in the world, affecting a wi...

  12. Change in the balance of excitatory and inhibitory midline fiber crossing as an explanation for the hopping phenotype in EphA4 knockout mice

    DEFF Research Database (Denmark)

    Restrepo, Carlos E.; Margaryan, Gayane; Borgius, Lotta

    2011-01-01

    Neuronal networks in the spinal cord termed central pattern generators (CPGs) are responsible for the generation of rhythmic movements, such as walking. The axon guidance molecule EphA4 has been suggested to play a role in the configuration of spinal CPG networks in mammals. In EphA4 knockout (Eph...... compared with EphA4 lacZ/+ mice. These results show that the hopping phenotype is the result of a change in the balance between excitatory and inhibitory signals across the midline and that EphA4-positive neurons play an essential role in the mammalian CPG....

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

    DEFF Research Database (Denmark)

    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...

  14. 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.

  15. 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.

  16. Single-Step Generation of Conditional Knockout Mouse Embryonic Stem Cells

    Directory of Open Access Journals (Sweden)

    Matyas Flemr

    2015-07-01

    Full Text Available Induction of double-strand DNA breaks (DSBs by engineered nucleases, such as CRISPR/Cas9 or transcription activator-like effector nucleases (TALENs, stimulates knockin of exogenous DNA fragments via homologous recombination (HR. However, the knockin efficiencies reported so far have not allowed more complex in vitro genome modifications such as, for instance, simultaneous integration of a DNA fragment at two distinct genomic sites. We developed a reporter system to enrich for cells with engineered nuclease-assisted HR events. Using this system in mouse embryonic stem cells (mESCs, we achieve single-step biallelic and seamless integration of two loxP sites for Cre recombinase-mediated inducible gene knockout, as well as biallelic endogenous gene tagging with high efficiency. Our approach reduces the time and resources required for conditional knockout mESC generation dramatically.

  17. 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.

  18. 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.

  19. 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

  20. Initial Results of Peripheral-Blood Stem-Cell Mobilization, Collection, Cryopreservation, and Engraftment After Autologous Transplantation Confirm That the Capacity-Building Approach Offers Good Chances of Success in Critical Contexts: A Kurdish-Italian Cooperative Project at the Hiwa Cancer Hospital, Sulaymaniyah

    Directory of Open Access Journals (Sweden)

    Ignazio Majolino

    2017-12-01

    Full Text Available Introduction: At Hiwa Cancer Hospital (Sulaymaniyah, Iraqi Kurdistan after the center was started by a cooperative project in June 2016, autologous transplantation was developed. Patients and Methods: To develop the project, the capacity-building approach was adopted, with on-site training and coaching of personnel, educational meetings, lectures, on-the-job training, and the implementation of quality management planning. Results: Here, we report initial results of peripheral-blood stem-cell mobilization and collection of the first 27 patients (age 12 to 61 years; 19 males and 8 females; multiple myeloma, n = 10; plasma cell leukemia, n = 1; Hodgkin lymphoma, n = 12; non-Hodgkin lymphoma, n = 3; and acute myeloid leukemia, n = 1. Only three (11.5% of 26 patients experienced a failure of mobilization. A median of 6.1 × 106/kg CD34-positive cells per patient were collected (range, 2.4 to 20.8, with two apheretic runs. Twenty-four patients underwent autologous transplantation. All but one transplantation engrafted fully and steadily, with 0.5 and 1.0 × 109/L polymorphonucleates on day 10.5 (range, 8 to 12 and day 11 (range, 9 to 15, respectively, and with 20 and 50 × 109/L platelets on day 13 (range, 10 to 17 and day 17 (range, 2 to 44, respectively. More than 95% of patients are projected to survive 1 year after autograft. Conclusion: These data are the result of an Italian effort to establish in Iraqi Kurdistan a leading center for hemopoietic stem-cell transplantation. The capacity building approach was used, with on-site training and coaching as instruments for the development of provider ability and problem solving. With future limitations for immigration, this method will be helpful, especially in the field of high-technology medicine.

  1. 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...

  2. 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...

  3. 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 ...

  4. 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.

  5. 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 ...

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

    Science.gov (United States)

    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.

  7. 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.

  8. 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.

  9. 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

  10. Deficits of learning and memory in Hemojuvelin knockout mice.

    Science.gov (United States)

    Li, Jinglong; Zhang, Peng; Liu, Hongju; Ren, Wei; Song, Jinjing; Rao, Elizabeth; Takahashi, Eiki; Zhou, Ying; Li, Weidong; Chen, Xiaoping

    2015-10-01

    Iron is involved in various physiological processes of the human body to maintain normal functions. Abnormal iron accumulation in brain has been reported as a pathogenesis of several neurodegenerative disorders and cognitive impairments. Hemojuvelin (HVJ) is a membrane-bound and soluble protein in mammals that is responsible for the iron overload condition known as juvenile hemochromatosis. Although iron accumulation in brain has been related to neurodegenerative diseases, it remains unknown the effect of mutation of HVJ gene on cognitive performance. In our studies, HJV(-/-) mice showed deficits in novel object recognition and Morris water maze tests. Furthermore, the expression ration of apoptotic marker Bax and anti-apoptotic marker Bcl-2 in the hippocampus and prefrontal cortex showed higher levels in HJV(-/-) mice. Our results suggested that deletion of HJV gene could increase apoptosis in brain which might contribute to learning and memory deficits in mutant mice. These results indicated that HJV(-/-) mice would be a useful model to study cognitive impairment induced by iron overload in brain.

  11. 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

  12. 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

  13. Tetranectin Knockout Mice Develop Features of Parkinson Disease

    Directory of Open Access Journals (Sweden)

    Er-song Wang

    2014-07-01

    Full Text Available Background/Aims: Aggregation of insoluble α-synuclein to form Lewy bodies (LBs may contribute to the selective loss of midbrain dopaminergic neurons in Parkinson disease (PD. Lack of robust animal models has impeded elucidation of the molecular mechanisms of LB formation and other critical aspects of PD pathogenesis. Methods: We established a mouse model with targeted deletion of the plasminogen-binding protein tetranectin (TN gene (TN-/- and measured the behavioral and histopathological features of PD. Results: Aged (15-to 20-month-old TN-/- mice displayed motor deficits resembling PD symptoms, including limb rigidity and both slower ambulation (bradykinesia and reduced rearing activity in the open field. In addition, these mice exhibited more numerous α-synuclein-positive LB-like inclusions within the substantia nigra pars compacta (SNc and reduced numbers of SNc dopaminergic neurons than age-matched wild type (WT mice. These pathological changes were also accompanied by loss of dopamine terminals in the dorsal striatum. Conclusion: The TN-/- mouse exhibits several key features of PD and so may be a valuable model for studying LB formation and testing candidate neuroprotective therapies for PD and other synucleinopathies.

  14. 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...

  15. XX males SRY negative: a confirmed cause of infertility.

    Science.gov (United States)

    Vetro, Annalisa; Ciccone, Roberto; Giorda, Roberto; Patricelli, Maria Grazia; Della Mina, Erika; Forlino, Antonella; Zuffardi, Orsetta

    2011-10-01

    SOX9 is a widely expressed transcription factor playing several relevant functions during development and essential for testes differentiation. It is considered to be the direct target gene of the protein encoded by SRY and its overexpression in an XX murine gonad can lead to male development in the absence of Sry. Recently, a family was reported with a 178 kb duplication in the gene desert region ending about 500 kb upstream of SOX9 in which 46,XY duplicated persons were completely normal and fertile whereas the 46,XX ones were males who came to clinical attention because of infertility. We report a family with two azoospermic brothers, both 46,XX, SRY negative, having a 96 kb triplication 500 kb upstream of SOX9. Both subjects have been analyzed trough oligonucleotide array-CGH and the triplication was confirmed and characterised through qPCR, defining the minimal region of amplification upstream of SOX9 associated with 46,XX infertile males, SRY negative. Our results confirm that even in absence of SRY, complete male differentiation may occur, possibly driven by overexpression of SOX9 in the gonadal ridge, as a consequence of the amplification of a gene desert region. We hypothesize that this region contains gonadal specific long-range regulation elements whose alteration may impair the normal sex development. Our data show that normal XX males, with alteration in copy number or, possibly, in the critical sequence upstream to SOX9 are a new category of infertility inherited in a dominant way with expression limited to the XX background.

  16. Neuroimaging findings in children with retinopathy-confirmed cerebral malaria

    Energy Technology Data Exchange (ETDEWEB)

    Potchen, Michael J. [Michigan State University, Department of Radiology, 184 Radiology Building, East Lansing, MI 48824-1303 (United States)], E-mail: mjp@rad.msu.edu; Birbeck, Gretchen L. [Michigan State University, International Neurologic and Psychiatric Epidemiology Program, 324 West Fee Hall, East Lansing, MI 48824 (United States)], E-mail: Gretchen.Birbeck@ht.msu.edu; DeMarco, J. Kevin [Michigan State University, Department of Radiology, 184 Radiology Building, East Lansing, MI 48824-1303 (United States)], E-mail: jkd@rad.msu.edu; Kampondeni, Sam D. [University of Malawi, Department of Radiology, Queen Elizabeth Central Hospital, Blantyre (Malawi)], E-mail: kamponde@msu.edu; Beare, Nicholas [St. Paul' s Eye Unit, Royal Liverpool University Hospital, Prescot Street, Liverpool L7 8XP (United Kingdom)], E-mail: nbeare@btinternet.com; Molyneux, Malcolm E. [Malawi-Liverpool-Wellcome Trust Clinical Research Programme, College of Medicine (Malawi); School of Tropical Medicine, University of Liverpool, Liverpool (United Kingdom)], E-mail: mmolyneux999@google.com; Taylor, Terrie E. [Michigan State University, College of Osteopathic Medicine, B309-B West Fee Hall, East Lansing, MI 48824 (United States); University of Malawi, College of Medicine, Blantyre Malaria Project, Blantyre (Malawi)], E-mail: taylort@msu.edu

    2010-04-15

    Purpose: To describe brain CT findings in retinopathy-confirmed, paediatric cerebral malaria. Materials and methods: In this outcomes study of paediatric cerebral malaria, a subset of children with protracted coma during initial presentation was scanned acutely. Survivors experiencing adverse neurological outcomes also underwent a head CT. All children had ophthalmological examination to confirm the presence of the retinopathy specific for cerebral malaria. Independent interpretation of CT images was provided by two neuroradiologists. Results: Acute brain CT findings in three children included diffuse oedema with obstructive hydrocephalus (2), acute cerebral infarctions in multiple large vessel distributions with secondary oedema and herniation (1), and oedema of thalamic grey matter (1). One child who was reportedly normal prior to admission had parenchymal atrophy suggestive of pre-existing CNS injury. Among 56 survivors (9-84 months old), 15 had adverse neurologic outcomes-11/15 had a follow-up head CT, 3/15 died and 1/15 refused CT. Follow-up head CTs obtained 7-18 months after the acute infection revealed focal and multifocal lobar atrophy correlating to regions affected by focal seizures during the acute infection (5/11). Other findings were communicating hydrocephalus (2/11), vermian atrophy (1/11) and normal studies (3/11). Conclusions: The identification of pre-existing imaging abnormalities in acute cerebral malaria suggests that population-based studies are required to establish the rate and nature of incidental imaging abnormalities in Malawi. Children with focal seizures during acute cerebral malaria developed focal cortical atrophy in these regions at follow-up. Longitudinal studies are needed to further elucidate mechanisms of CNS injury and death in this common fatal disease.

  17. Neuroimaging findings in children with retinopathy-confirmed cerebral malaria

    International Nuclear Information System (INIS)

    Potchen, Michael J.; Birbeck, Gretchen L.; DeMarco, J. Kevin; Kampondeni, Sam D.; Beare, Nicholas; Molyneux, Malcolm E.; Taylor, Terrie E.

    2010-01-01

    Purpose: To describe brain CT findings in retinopathy-confirmed, paediatric cerebral malaria. Materials and methods: In this outcomes study of paediatric cerebral malaria, a subset of children with protracted coma during initial presentation was scanned acutely. Survivors experiencing adverse neurological outcomes also underwent a head CT. All children had ophthalmological examination to confirm the presence of the retinopathy specific for cerebral malaria. Independent interpretation of CT images was provided by two neuroradiologists. Results: Acute brain CT findings in three children included diffuse oedema with obstructive hydrocephalus (2), acute cerebral infarctions in multiple large vessel distributions with secondary oedema and herniation (1), and oedema of thalamic grey matter (1). One child who was reportedly normal prior to admission had parenchymal atrophy suggestive of pre-existing CNS injury. Among 56 survivors (9-84 months old), 15 had adverse neurologic outcomes-11/15 had a follow-up head CT, 3/15 died and 1/15 refused CT. Follow-up head CTs obtained 7-18 months after the acute infection revealed focal and multifocal lobar atrophy correlating to regions affected by focal seizures during the acute infection (5/11). Other findings were communicating hydrocephalus (2/11), vermian atrophy (1/11) and normal studies (3/11). Conclusions: The identification of pre-existing imaging abnormalities in acute cerebral malaria suggests that population-based studies are required to establish the rate and nature of incidental imaging abnormalities in Malawi. Children with focal seizures during acute cerebral malaria developed focal cortical atrophy in these regions at follow-up. Longitudinal studies are needed to further elucidate mechanisms of CNS injury and death in this common fatal disease.

  18. Confirmation of identity and detection limit in neutron activation analysis

    International Nuclear Information System (INIS)

    Yustina Tri Handayani; Slamet Wiyuniati; Tulisna

    2010-01-01

    Neutron Activation Analysis (NAA) based on neutron capture by nuclides. Of the various possibilities of radionuclides that occur, radionuclides and gamma radiation which provides the identity of the element were analyzed and the best sensitivity should be determined. Confirmation for elements in sediment samples was done theoretically and experimentally. The result of confirmation shows that Al, V, Cr K, Na, Ca and Zn were analyzed based on radionuclides of Al-28, V-52, Cr-51 , K-42, Na-24, Ca-48, Zn-65. Elements of Mg, Mn, Fe, Co were analyzed based on radionuclides of Mg-27, Mn-56, Fe-59, Co-60 through peak which the highest value of combined probability of radiation emission and efficiency. Cu can be analyzed through Cu-64 or Cu-66, but the second is more sensitive. Detection limit is determined at a certain measurement conditions carried out by a laboratory. Detection limit in the NAA is determined based on the Compton continue area by Curie method. The detection limit of Al, V, Ca, Mg, Mn, As, K, Na, Mg, Ce, Co, Cr, Fe, La, Sc, and Zn in sediment samples are 240, 27, 4750, 2600, 21, 3.3 , 75, 1.4, 1.8, 0.5, 2.7, 29, 1, 0.05, and 37 ppm. Analysis of Cu in sediments which concentrations of 98.6 ppm, Cu-66 is not detected. Tests using pure standard solutions of Cu obtained detection limit of 0.12 µg, or 7.9 ppm in samples of 15 mg. In general, the detection limit obtained was higher than the detection limit of the reference, it was caused by the differences in the sample matrix and analytical conditions. (author)

  19. Confirmation of a blocked amino terminus of sulfhydryl oxidase

    International Nuclear Information System (INIS)

    Janolino, V.G.; Morrison-Rowe, S.J.; Swaisgood, H.E.

    1990-01-01

    The isolation of sulfhydryl oxidase from bovine milk in a suitably pure form for sequencing was carried out by transient covalent affinity chromatography of diafiltered whey using cysteinylsuccinamidopropyl-glass as matrix. The glutathione-eluted proteins were separated by SDS-PAGE. By radiolabeling the affinity chromatography-purified enzyme with [ 14 C]iodoacetate before subjecting to SDS-PAGE, the sulfhydryl oxidase band was identified, because sulfhydryl oxidase is known to be inactivated by alkylation of one sulfhydryl group per mole. The results confirmed that sulfhydryl oxidase corresponds to the 85 (± 5)-kDa band observed on SDS-PAGE. The protein band corresponding to radiolabeled sulfhydryl oxidase was recovered from SDS-PAGE gels by electrophoretic elution and by electroblotting on polyvinylidene difluoride membrane and subjected to gas phase sequencing. Precautions were taken during electrophoretic elution to prevent reactions that result in N-terminal blocking. Both methods of protein recovery yielded negative results when subjected to sequence analysis indicating that the N-terminus of sulfhydryl oxidase is blocked

  20. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  3. Poliomyelitis in Osun State, Nigeria: Two Confirmed Cases After 6 ...

    African Journals Online (AJOL)

    The Clinico-epidemological characteristics of two confirmed cases of poliomyelitis detected by Acute Flaccid Paralysis (AFP) surveillance in Osun State of Nigeria after almost 6 years of the last confirmed case in the State was reported to provide information for formulating possible aetiological hypothesis and to adequately ...

  4. The role of laboratory confirmations and molecular epidemiology in ...

    African Journals Online (AJOL)

    This review reports on the role of laboratory confirmation and molecular epidemiology in global eradication of measles. The role of laboratory confirmation and molecular epidemiology in defining the origins of measles outbreaks cannot be overemphasized. New serological tests based on recombinant proteins detect only a ...

  5. Piper betle induces phase I & II genes through Nrf2/ARE signaling pathway in mouse embryonic fibroblasts derived from wild type and Nrf2 knockout cells.

    Science.gov (United States)

    Wan Hasan, Wan Nuraini; Kwak, Mi-Kyoung; Makpol, Suzana; Wan Ngah, Wan Zurinah; Mohd Yusof, Yasmin Anum

    2014-02-23

    Nuclear factor-erythroid 2 p45 related factor 2 (Nrf2) is a primary transcription factor, protecting cells from oxidative stress by regulating a number of antioxidants and phase II detoxifying enzymes. Dietary components such as sulforaphane in broccoli and quercetin in onions have been shown to be inducers of Nrf2. Piper betle (PB) grows well in tropical climate and the leaves are used in a number of traditional remedies for the treatment of stomach ailments and infections among Asians. The aim of this study was to elucidate the effect of Piper betle (PB) leaves extract in Nrf2 signaling pathway by using 2 types of cells; mouse embryonic fibroblasts (MEFs) derived from wild-type (WT) and Nrf2 knockout (N0) mice. WT and N0 cells were treated with 5 and 10 μg/ml of PB for 10 and 12-h for the determination of nuclear translocation of Nrf2 protein. Luciferase reporter gene activity was performed to evaluate the antioxidant response element (ARE)-induction by PB. Real-time PCR and Western blot were conducted on both WT and N0 cells after PB treatment for the determination of antioxidant enzymes [superoxide dismutase (SOD1) and heme-oxygenase (HO-1)], phase I oxidoreductase enzymes [ quinone oxidoreductase (NQO1)] and phase II detoxifying enzyme [glutathione S-transferase (GST)]. Nuclear translocation of Nrf2 by PB in WT cells was better after 10 h incubation compared to 12 h. Real time PCR and Western blot analysis showed increased expressions of Nrf2, NQO1 and GSTA1 genes with corresponding increases in glutathione, NQO1 and HO-1 proteins in WT cells. Reporter gene ARE was stimulated by PB as shown by ARE/luciferase assay. Interestingly, PB induced SOD1 gene and protein expressions in N0 cells but not in WT cells. The results of this study confirmed that PB activated Nrf2-ARE signaling pathway which subsequently induced some phase I oxidoreductase, phase II detoxifying and antioxidant genes expression via ARE reporter gene involved in the Nrf2 pathway with the

  6. An alternative method to achieve metrological confirmation in measurement process

    Science.gov (United States)

    Villeta, M.; Rubio, E. M.; Sanz, A.; Sevilla, L.

    2012-04-01

    Metrological confirmation process must be designed and implemented to ensure that metrological characteristics of the measurement system meet metrological requirements of the measurement process. The aim of this paper is to present an alternative method to the traditional metrological requirements about the relationship between tolerance and measurement uncertainty, to develop such confirmation processes. The proposed way to metrological confirmation considers a given inspection task of the measurement process into the manufacturing system, and it is based on the Index of Contamination of the Capability, ICC. Metrological confirmation process is then developed taking into account the producer risks and economic considerations on this index. As a consequence, depending on the capability of the manufacturing process, the measurement system will be or will not be in adequate state of metrological confirmation for the measurement process.

  7. Computer assisted radionuclide angiography to confirm reversible ischemic cerebral dysfunction

    International Nuclear Information System (INIS)

    Buell, U.; Lanksch, W.; Tosch, U.; Kleinhans, E.; Steinhoff, H.

    1982-01-01

    Computer assisted radionuclide angiography (CARNA) was employed in patients with transient ischemic attack (TIA) or prolonged reversible ischemic neurologic deficit (PRIND) to establish the sensitivity of CARNA in detecting and quantifying changes of cerebral perfusion in such selected patients. Moreover, results of CARNA were compared with findings of cranial radiographic angiography (RGA) to obtain data on combined sensitivities of these methods. CARNA may be the preferred noninvasive procedure employed because it detects and quantifies the vascular supply disorder in patients with TIA and PRIND. If no computer assistance is used to evaluate cranial radionuclide angiography, results are considerable less accurate. Specifity of CARNA is 84.6%. If CARNA is negative (25.2% in TIA; 12.7% in PRIND), a further method must be employed to confirm the cranial vascular origin of the attack. This may be RGA in TIA and transmission computed axial tomography (T-CAT) T-CAT in PRIND. This diagnos - tic sequence lead to 92.4% true positive in TIA and to 93.2% true positives in PRIND

  8. 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.

  9. 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.

  10. Managing a duopolistic water market with confirmed proposals. An experiment

    Directory of Open Access Journals (Sweden)

    García-Gallego, Aurora

    2012-03-01

    Full Text Available We report results from experimental water markets in which owners of two different sources of water supply water to households and farmers. The final water quality consumed by each type of consumer is determined through mixing of qualities from two different resources. We compare the standard duopolistic market structure with an alternative market clearing mechanism inspired by games with confirmed strategies (which have been shown to yield collusive outcomes. As in the static case, complex dynamic markets operating under a confirmed proposals protocol yield less efficient outcomes because coordination among independent suppliers has the usual effects of restricting output and increasing prices to the users. Our results suggest that, when market mechanisms are used to allocate water to its users, the rule of thumb used by competition authorities can also serve as a guide towards water market regulation.

    Se presentan resultados de un experimento con mercados acuíferos en el que los propietarios de agua de distinta calidad la ofrecen a hogares y agricultores. La calidad finalmente consumida por cada tipo de consumidor se determina a partir de una mezcla de las dos calidades. Se compara el duopolio estándar con una forma alternativa de cerrar el mercado que está inspirada en los juegos con propuestas confirmadas, que consiguen resultados relativamente más colusivos. Como en el caso estático, los mercados dinámicos y complejos que operan bajo un protocolo de propuestas confirmadas son menos eficientes porque la coordinación entre oferentes independientes tiene los efectos de restringir el output y de provocar un crecimiento de los precios. Nuestros resultados sugieren que cuando los mecanismos de mercado se utilizan para distribuir el agua a sus usuarios, la regla utilizada por parte de las autoridades de la competencia puede servir también como guía para la regulación de los mercados acuíferos.

  11. Culture confirmation of tuberculosis cases in Birmingham, UK.

    Science.gov (United States)

    Hayer, Kalbir S; Sitch, Alice J; Dedicoat, Martin; Wood, Annette L

    2013-10-01

    The proportion of culture-confirmed tuberculosis (TB) cases in Birmingham had gradually decreased to less than 65% in 2008. Reasons for this were unclear, therefore this study assessed diagnostic methods used for confirming TB and reviewed factors involved in positive culture. A cross-sectional study was carried out. A list of notified TB cases for Birmingham in those aged 16 y and over in 2009 was collated. Where no positive culture was recorded, further data were collected from hospital databases and case notes. Of 449 TB cases, 419 (93%) had samples taken for culture testing. Of all cases, 309 (69%) were confirmed by culture testing; of those receiving culture testing, 73% were confirmed. Pulmonary TB was identified as a predictor of positive culture in both the unadjusted and adjusted analyses: odds ratio (OR) 2.05, 95% confidence interval (CI) 1.32-3.19, and OR 2.32, 95% CI 1.29-4.17, respectively. Gender, age, ethnicity, UK born, and treatment delay were not significantly associated with positive culture. Of 140 cases not confirmed by culture, 129 (92%) had their diagnosis supported by at least one other test. The vast majority of TB cases had microbiological specimens taken to help confirm the disease. Furthermore, culture confirmation rates in Birmingham were meeting national targets in 2009. However culture confirmation rates were significantly lower in extrapulmonary TB, therefore further work is suggested in this group. The role of other investigations (e.g. interferon-gamma release assay (IGRA), Mantoux) is unclear. Further collaboration between clinicians, histopathologists, and microbiologists is advised to ensure samples are sent appropriately and culture confirmation is optimized.

  12. Anaplasma phagocytophilum in Danish sheep: confirmation by DNA sequencing

    Directory of Open Access Journals (Sweden)

    Thamsborg Stig M

    2009-12-01

    Full Text Available Abstract Background The presence of Anaplasma phagocytophilum, an Ixodes ricinus transmitted bacterium, was investigated in two flocks of Danish grazing lambs. Direct PCR detection was performed on DNA extracted from blood and serum with subsequent confirmation by DNA sequencing. Methods 31 samples obtained from clinically normal lambs in 2000 from Fussingø, Jutland and 12 samples from ten lambs and two ewes from a clinical outbreak at Feddet, Zealand in 2006 were included in the study. Some of the animals from Feddet had shown clinical signs of polyarthritis and general unthriftiness prior to sampling. DNA extraction was optimized from blood and serum and detection achieved by a 16S rRNA targeted PCR with verification of the product by DNA sequencing. Results Five DNA extracts were found positive by PCR, including two samples from 2000 and three from 2006. For both series of samples the product was verified as A. phagocytophilum by DNA sequencing. Conclusions A. phagocytophilum was detected by molecular methods for the first time in Danish grazing lambs during the two seasons investigated (2000 and 2006.

  13. CONFIRMATION OF FAINT DWARF GALAXIES IN THE M81 GROUP

    Energy Technology Data Exchange (ETDEWEB)

    Chiboucas, Kristin [Gemini Observatory, 670 North A' ohoku Pl, Hilo, HI 96720 (United States); Jacobs, Bradley A.; Tully, R. Brent [Institute for Astronomy, University of Hawaii, 2680 Woodlawn Drive, Honolulu, HI 96821 (United States); Karachentsev, Igor D., E-mail: kchibouc@gemini.edu, E-mail: bjacobs@ifa.hawaii.edu, E-mail: tully@ifa.hawaii.edu, E-mail: ikar@luna.sao.ru [Special Astrophysical Observatory (SAO), Russian Academy of Sciences, Nizhnij Arkhyz, Karachai-Cherkessian Republic 369167 (Russian Federation)

    2013-11-01

    We have followed up on the results of a 65 deg{sup 2} CFHT/MegaCam imaging survey of the nearby M81 Group searching for faint and ultra-faint dwarf galaxies. The original survey turned up 22 faint candidate dwarf members. Based on two-color HST ACS/WFC and WFPC2 photometry, we now confirm 14 of these as dwarf galaxy members of the group. Distances and stellar population characteristics are discussed for each. To a completeness limit of M{sub r{sup '}}= -10, we find a galaxy luminosity function slope of –1.27 ± 0.04 for the M81 Group. In this region, there are now 36 M81 Group members known, including 4 blue compact dwarfs; 8 other late types including the interacting giants M81, NGC 3077, and M82; 19 early type dwarfs; and at least 5 potential tidal dwarf galaxies. We find that the dSph galaxies in M81 appear to lie in a flattened distribution, similar to that found for the Milky Way and M31. One of the newly discovered dSph galaxies has properties similar to the ultra-faint dwarfs being found in the Local Group with a size R{sub e} ∼ 100 pc and total magnitude estimates M{sub r{sup '}}= -6.8 and M{sub I} ∼ –9.1.

  14. Pathological Confirmation of Optic Neuropathy in Familial Dysautonomia.

    Science.gov (United States)

    Mendoza-Santiesteban, Carlos E; Palma, Jose-Alberto; Hedges, Thomas R; Laver, Nora V; Farhat, Nada; Norcliffe-Kaufmann, Lucy; Kaufmann, Horacio

    2017-03-01

    Clinical data suggest that optic neuropathy and retinal ganglion cell loss are the main cause of visual decline in patients with familial dysautonomia, but this has not previously been confirmed by pathological analyses. We studied retinas and optic nerves in 6 eyes from 3 affected patients obtained at autopsy. Analyses included routine neurohistology and immunohistochemistry for neurofilaments, cytochrome c oxidase (COX), and melanopsin-containing ganglion cells. We observed profound axon loss in the temporal portions of optic nerves with relative preservation in the nasal portions; this correlated with clinical and optical coherence tomography findings in 1 patient. Retinal ganglion cell layers were markedly reduced in the central retina, whereas melanopsin-containing ganglion cells were relatively spared. COX staining was reduced in the temporal portions of the optic nerve indicating reduced mitochondrial density. Axonal swelling with degenerating lysosomes and mitochondria were observed by electron microscopy. These findings support the concept that there is a specific optic neuropathy and retinopathy in patients with familial dysautonomia similar to that seen in other optic neuropathies with mitochondrial dysfunction. This raises the possibility that defective expression of the IkB kinase complex-associated protein (IKAP) resulting from mutations in IKBKAP affects mitochondrial function in the metabolism-dependent retinal parvocellular ganglion cells in this condition. © 2017 American Association of Neuropathologists, Inc. All rights reserved.

  15. Confirmation of Faint Dwarf Galaxies in the M81 Group

    Science.gov (United States)

    Chiboucas, Kristin; Jacobs, Bradley A.; Tully, R. Brent; Karachentsev, Igor D.

    2013-11-01

    We have followed up on the results of a 65 deg2 CFHT/MegaCam imaging survey of the nearby M81 Group searching for faint and ultra-faint dwarf galaxies. The original survey turned up 22 faint candidate dwarf members. Based on two-color HST ACS/WFC and WFPC2 photometry, we now confirm 14 of these as dwarf galaxy members of the group. Distances and stellar population characteristics are discussed for each. To a completeness limit of M_{r^{\\prime }} = -10, we find a galaxy luminosity function slope of -1.27 ± 0.04 for the M81 Group. In this region, there are now 36 M81 Group members known, including 4 blue compact dwarfs; 8 other late types including the interacting giants M81, NGC 3077, and M82; 19 early type dwarfs; and at least 5 potential tidal dwarf galaxies. We find that the dSph galaxies in M81 appear to lie in a flattened distribution, similar to that found for the Milky Way and M31. One of the newly discovered dSph galaxies has properties similar to the ultra-faint dwarfs being found in the Local Group with a size Re ~ 100 pc and total magnitude estimates M_{r^{\\prime }} = -6.8 and MI ~ -9.1.

  16. Consumer satisfaction and confirmation of habits of comprehension

    DEFF Research Database (Denmark)

    Sørensen, Bent; Andersen, Christian; Andersen, Morten Purup

    2014-01-01

    the formation of consumer satisfaction; the perspective is that of the confirmation paradigm within advertisement research. Inductive advertisements support cognitive habit formation through confirmation, and the confirmation paradigm explains exactly consumer satisfaction with reference to confirmation. Hence......The purpose of this article is twofold: First, within a Peircean framework it shall be demonstrated how there is a relation between the compositional structure of certain types of print advertisements and their bringing about inductive comprehension, and how the consumer can be understood...... as a bundle of habits. It is the assumption that advertising that supports an inductive effect particularly appeals to the cognitive tendency of habit formation in the consumer. Second, it is asked whether advertisements that predominantly invite inductive processes of comprehension also influence...

  17. Detection and confirmation of toxigenic Vibrio cholerae O1 in ...

    African Journals Online (AJOL)

    2013-08-20

    Aug 20, 2013 ... mental water samples, as well as for confirmation of clinical isolates. Keywords: ... Monitoring the presence of V. cholerae in drinking water sources ... have several advantages: they are rapid, sensitive, highly selective and do ...

  18. First confirmed record of Elodea canadensis Michx. (Hydrocharitaceae in Greece

    Directory of Open Access Journals (Sweden)

    Poulis Georgios

    2017-12-01

    Full Text Available The paper confirms the presence of Elodea canadensis Michx. in Greece and outlines the history of contradictory relevant reports. This is also the first report of the species′ presence in the transboundary lake Great Prespa.

  19. CERN confirms goal of 2007 start-up for LHC

    CERN Document Server

    2005-01-01

    Speaking at the 131st session of CERN Council on 17 December 2004, the Director-General, Robert Aymar, confirmed that the top priority is to maintain the goal of starting up the Large Hadron Collider (LHC) in 2007.

  20. NIH study confirms risk factors for male breast cancer

    Science.gov (United States)

    Pooled data from studies of about 2,400 men with breast cancer and 52,000 men without breast cancer confirmed that risk factors for male breast cancer include obesity, a rare genetic condition called Klinefelter syndrome, and gynecomastia.

  1. Scientists confirm delay in testing new CERN particle accelerator

    CERN Multimedia

    2007-01-01

    "Scientists seeking to uncover the secrets of the universe will have to wait a little longer after the CERN laboratory inswitzerland on Monday confirmed a delay in tests of a massive new particle accelerator." (1 page)

  2. Confirmation of the absolute configuration of (−)-aurantioclavine

    KAUST Repository

    Behenna, Douglas C.; Krishnan, Shyam; Stoltz, Brian M.

    2011-01-01

    We confirm our previous assignment of the absolute configuration of (-)-aurantioclavine as 7R by crystallographically characterizing an advanced 3-bromoindole intermediate reported in our previous synthesis. This analysis also provides additional

  3. Generation of beta-lactoglobulin knock-out goats using CRISPR/Cas9.

    Directory of Open Access Journals (Sweden)

    Wenjun Zhou

    Full Text Available Goat's milk, considered a substitute for cow's milk, has a high nutritional value. However, goat's milk contains various allergens, predominantly β-lactoglobulin (BLG. In this study, we employed the CRISPR/Cas9 system to target the BLG locus in goat fibroblasts for sgRNA optimization and generate BLG knock-out goats through co-injection of Cas9 mRNA and small guide RNAs (sgRNAs into goat embryos at the one-cell stage. We firstly tested sgRNA editing efficiencies in goat fibroblast cells, and approximately 8.00%-9.09% of the cells were modified in single sgRNA-guided targeting experiment. Among the kids, the genome-targeting efficiencies of single sgRNA were 12.5% (10 ng/μL sg1 and 0% (10 ng/μL sg2 and efficiencies of dual sgRNAs were 25.0% (25 ng/μL sg2+sg3 group and 28.6% (50 ng/μL sg2+sg3 group. Relative expression of BLG in BLG knock-out goat mammary glands significantly (p < 0.01 decreased as well as other milk protein coding genes, such as CSN1S1, CSN1S2, CSN2, CSN3 and LALBA (p < 0.05. As expected, BLG protein had been abolished in the milk of the BLG knock-out goat. In addition, most of the targeted kids were chimeric (3/4, and their various body tissues were edited simultaneously. Our study thus provides a basis for optimizing the quality of goat milk, which can be applied to biomedical and agricultural research.

  4. 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.

  5. 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.

  6. Marginal level dystrophin expression improves clinical outcome in a strain of dystrophin/utrophin double knockout mice.

    Directory of Open Access Journals (Sweden)

    Dejia Li

    2010-12-01

    Full Text Available Inactivation of all utrophin isoforms in dystrophin-deficient mdx mice results in a strain of utrophin knockout mdx (uko/mdx mice. Uko/mdx mice display severe clinical symptoms and die prematurely as in Duchenne muscular dystrophy (DMD patients. Here we tested the hypothesis that marginal level dystrophin expression may improve the clinical outcome of uko/mdx mice. It is well established that mdx3cv (3cv mice express a near-full length dystrophin protein at ∼5% of the normal level. We crossed utrophin-null mutation to the 3cv background. The resulting uko/3cv mice expressed the same level of dystrophin as 3cv mice but utrophin expression was completely eliminated. Surprisingly, uko/3cv mice showed a much milder phenotype. Compared to uko/mdx mice, uko/3cv mice had significantly higher body weight and stronger specific muscle force. Most importantly, uko/3cv outlived uko/mdx mice by several folds. Our results suggest that a threshold level dystrophin expression may provide vital clinical support in a severely affected DMD mouse model. This finding may hold clinical implications in developing novel DMD therapies.

  7. 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

  8. 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

  9. 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)

  10. 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...

  11. 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

  12. 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

  13. 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

  14. 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.

  15. Lentivirus-ABCG1 instillation reduces lipid accumulation and improves lung compliance in GM-CSF knock-out mice

    Energy Technology Data Exchange (ETDEWEB)

    Malur, Anagha; Huizar, Isham [Program in Lung Cell Biology and Translational Research, Division of Pulmonary, Critical Care and Sleep Medicine, East Carolina University, Greenville, NC (United States); Wells, Greg [Department of Microbiology and Immunology, East Carolina University, Greenville, NC (United States); Barna, Barbara P. [Program in Lung Cell Biology and Translational Research, Division of Pulmonary, Critical Care and Sleep Medicine, East Carolina University, Greenville, NC (United States); Malur, Achut G. [Department of Microbiology and Immunology, East Carolina University, Greenville, NC (United States); Thomassen, Mary Jane, E-mail: thomassenm@ecu.edu [Program in Lung Cell Biology and Translational Research, Division of Pulmonary, Critical Care and Sleep Medicine, East Carolina University, Greenville, NC (United States); Department of Microbiology and Immunology, East Carolina University, Greenville, NC (United States)

    2011-11-18

    Highlights: Black-Right-Pointing-Pointer Lentivirus-ABCG1 reduces lipid accumulation in lungs of GM-CSF knock-out mice. Black-Right-Pointing-Pointer Up-regulation of ABCG1 improves lung function. Black-Right-Pointing-Pointer Upregulation of ABCG1 improves surfactant metabolism. -- Abstract: We have shown decreased expression of the nuclear transcription factor, peroxisome proliferator-activated receptor-gamma (PPAR{gamma}) and the PPAR{gamma}-regulated ATP-binding cassette transporter G1 (ABCG1) in alveolar macrophages from patients with pulmonary alveolar proteinosis (PAP). PAP patients also exhibit neutralizing antibodies to granulocyte-macrophage colony stimulating factor (GM-CSF), an upregulator of PPAR{gamma}. In association with functional GM-CSF deficiency, PAP lung is characterized by surfactant-filled alveolar spaces and lipid-filled alveolar macrophages. Similar pathology characterizes GM-CSF knock-out (KO) mice. We reported previously that intratracheal instillation of a lentivirus (lenti)-PPAR{gamma} plasmid into GM-CSF KO animals elevated ABCG1 and reduced alveolar macrophage lipid accumulation. Here, we hypothesized that instillation of lenti-ABCG1 might be sufficient to decrease lipid accumulation and improve pulmonary function in GM-CSF KO mice. Animals received intratracheal instillation of lenti-ABCG1 or control lenti-enhanced Green Fluorescent Protein (eGFP) plasmids and alveolar macrophages were harvested 10 days later. Alveolar macrophage transduction efficiency was 79% as shown by lenti-eGFP fluorescence. Quantitative PCR analyses indicated a threefold (p = 0.0005) increase in ABCG1 expression with no change of PPAR{gamma} or ABCA1 in alveolar macrophages of lenti-ABCG1 treated mice. ABCG1 was unchanged in control lenti-eGFP and PBS-instilled groups. Oil Red O staining detected reduced intracellular neutral lipid in alveolar macrophages from lenti-ABCG1 treated mice. Extracellular cholesterol and phospholipids were also decreased as shown by

  16. 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

  17. 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.

  18. Lentivirus-ABCG1 instillation reduces lipid accumulation and improves lung compliance in GM-CSF knock-out mice

    International Nuclear Information System (INIS)

    Malur, Anagha; Huizar, Isham; Wells, Greg; Barna, Barbara P.; Malur, Achut G.; Thomassen, Mary Jane

    2011-01-01

    Highlights: ► Lentivirus-ABCG1 reduces lipid accumulation in lungs of GM-CSF knock-out mice. ► Up-regulation of ABCG1 improves lung function. ► Upregulation of ABCG1 improves surfactant metabolism. -- Abstract: We have shown decreased expression of the nuclear transcription factor, peroxisome proliferator-activated receptor-gamma (PPARγ) and the PPARγ-regulated ATP-binding cassette transporter G1 (ABCG1) in alveolar macrophages from patients with pulmonary alveolar proteinosis (PAP). PAP patients also exhibit neutralizing antibodies to granulocyte–macrophage colony stimulating factor (GM-CSF), an upregulator of PPARγ. In association with functional GM-CSF deficiency, PAP lung is characterized by surfactant-filled alveolar spaces and lipid-filled alveolar macrophages. Similar pathology characterizes GM-CSF knock-out (KO) mice. We reported previously that intratracheal instillation of a lentivirus (lenti)-PPARγ plasmid into GM-CSF KO animals elevated ABCG1 and reduced alveolar macrophage lipid accumulation. Here, we hypothesized that instillation of lenti-ABCG1 might be sufficient to decrease lipid accumulation and improve pulmonary function in GM-CSF KO mice. Animals received intratracheal instillation of lenti-ABCG1 or control lenti-enhanced Green Fluorescent Protein (eGFP) plasmids and alveolar macrophages were harvested 10 days later. Alveolar macrophage transduction efficiency was 79% as shown by lenti-eGFP fluorescence. Quantitative PCR analyses indicated a threefold (p = 0.0005) increase in ABCG1 expression with no change of PPARγ or ABCA1 in alveolar macrophages of lenti-ABCG1 treated mice. ABCG1 was unchanged in control lenti-eGFP and PBS-instilled groups. Oil Red O staining detected reduced intracellular neutral lipid in alveolar macrophages from lenti-ABCG1 treated mice. Extracellular cholesterol and phospholipids were also decreased as shown by analysis of bronchoalveolar lavage fluid. Lung compliance was diminished in untreated GMCSF KO mice

  19. Generation of a Tph2 Conditional Knockout Mouse Line for Time- and Tissue-Specific Depletion of Brain Serotonin

    Science.gov (United States)

    Migliarini, Sara; Pacini, Giulia; Pasqualetti, Massimo

    2015-01-01

    Serotonin has been gaining increasing attention during the last two decades due to the dual function of this monoamine as key regulator during critical developmental events and as neurotransmitter. Importantly, unbalanced serotonergic levels during critical temporal phases might contribute to the onset of neuropsychiatric disorders, such as schizophrenia and autism. Despite increasing evidences from both animal models and human genetic studies have underpinned the importance of serotonin homeostasis maintenance during central nervous system development and adulthood, the precise role of this molecule in time-specific activities is only beginning to be elucidated. Serotonin synthesis is a 2-step process, the first step of which is mediated by the rate-limiting activity of Tph enzymes, belonging to the family of aromatic amino acid hydroxylases and existing in two isoforms, Tph1 and Tph2, responsible for the production of peripheral and brain serotonin, respectively. In the present study, we generated and validated a conditional knockout mouse line, Tph2 flox/flox, in which brain serotonin can be effectively ablated with time specificity. We demonstrated that the Cre-mediated excision of the third exon of Tph2 gene results in the production of a Tph2 null allele in which we observed the near-complete loss of brain serotonin, as well as the growth defects and perinatal lethality observed in serotonin conventional knockouts. We also revealed that in mice harbouring the Tph2 null allele, but not in wild-types, two distinct Tph2 mRNA isoforms are present, namely Tph2Δ3 and Tph2Δ3Δ4, with the latter showing an in-frame deletion of amino acids 84–178 and coding a protein that could potentially retain non-negligible enzymatic activity. As we could not detect Tph1 expression in the raphe, we made the hypothesis that the Tph2Δ3Δ4 isoform can be at the origin of the residual, sub-threshold amount of serotonin detected in the brain of Tph2 null/null mice. Finally, we set

  20. 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

  1. Suppression or knockout of SaF/SaM overcomes the Sa-mediated hybrid male sterility in rice

    Institute of Scientific and Technical Information of China (English)

    Yongyao Xie; Baixiao Niu; Yunming Long; Gousi Li; Jintao Tang; Yaling Zhang; Ding Ren; Yao-Guang Liu; Letian Chen

    2017-01-01

    Hybrids between the indica and japonica subspecies of rice (Oryza sativa) are usually sterile, which hinders utilization of heterosis in the inter-subspecific hybrid breeding. The complex locus Sa comprises two adjacently located genes, SaF and SaM, which interact to cause abortion of pollen grains carrying the japonica allele in japonica-indica hybrids. Here we showed that silencing of SaF or SaM by RNA interference restored male fertility in indica-japonica hybrids with heterozygous Sa. We further used clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9-based genome editing to knockout the SaF and SaM alleles, respectively, of an indica rice line to create hybrid-compatible lines. The resultant artificial neutral alleles did not affect pollen viability and other agricultural traits, but did break down the reproductive barrier in the hybrids. We found that some rice lines have natural neutral allele Sa-n, which was compatible with the typical japonica or indica Sa alleles in hybrids. Our results demonstrate that SaF and SaM are required for hybrid male sterility, but are not essential for pollen development. This study provides effective approaches for the generation of hybrid-compatible lines by knocking out the Sa locus or using the natural Sa-n allele to overcome hybrid male sterility in rice breeding.

  2. An extremely high dietary iodide supply forestalls severe hypothyroidism in Na+/I- symporter (NIS) knockout mice.

    Science.gov (United States)

    Ferrandino, Giuseppe; Kaspari, Rachel R; Reyna-Neyra, Andrea; Boutagy, Nabil E; Sinusas, Albert J; Carrasco, Nancy

    2017-07-13

    The sodium/iodide symporter (NIS) mediates active iodide (I - ) accumulation in the thyroid, the first step in thyroid hormone (TH) biosynthesis. Mutations in the SLC5A5 gene encoding NIS that result in a non-functional protein lead to congenital hypothyroidism due to I - transport defect (ITD). ITD is a rare autosomal disorder that, if not treated promptly in infancy, can cause mental retardation, as the TH decrease results in improper development of the nervous system. However, in some patients, hypothyroidism has been ameliorated by unusually large amounts of dietary I - . Here we report the first NIS knockout (KO) mouse model, obtained by targeting exons 6 and 7 of the Slc5a5 gene. In NIS KO mice, in the thyroid, stomach, and salivary gland, NIS is absent, and hence there is no active accumulation of the NIS substrate pertechnetate ( 99m TcO 4 - ). NIS KO mice showed undetectable serum T 4 and very low serum T 3 levels when fed a diet supplying the minimum I - requirement for rodents. These hypothyroid mice displayed oxidative stress in the thyroid, but not in the brown adipose tissue or liver. Feeding the mice a high-I - diet partially rescued TH biosynthesis, demonstrating that, at high I - concentrations, I - enters the thyroid through routes other than NIS.

  3. Whole body analysis of the knockout gene mouse model for cystic fibrosis using thermal and fast neutron activation analysis

    International Nuclear Information System (INIS)

    Mason, M.M.; Morris, J.S.; Derenzy, B.A.; Spate, V.L.; Horsman, T.L.; Baskett, C.K.; Nichols, T.A.; Colbert, J.W.; Clarke, L.L.; Gawenis, L.R.; Hillman, L.S.

    1998-01-01

    A genetically engineered 'knockout gene' mouse model for human cystic fibrosis (CF) has been utilized to study bone mineralization. In CF, the so-called cystic fibrosis transmembrane conductance regulator (CFTR) protein, a chloride ion channel, is either absent or defective. To produce the animal model the murine CFTR gene has been inactivated producing CF symptoms in the homozygotic progeny. CF results in abnormal intestinal absorption of minerals and nutrients which presumably results in substandard bone mineralization. The objective of this study was to determine the feasibility of using whole-body thermal and fast neutron activation analysis to determine mineral and trace-element differences between homozygote controls (+/+) and CF (-/-), murine siblings. Gender-matched juvenile +/+ and -/- litter mates were lyophilized and placed in a BN capsule to reduce thermal-neutron activation and irradiated for 10 seconds at φ fast ∼ 1 x 10 13 n x cm -2 x s -1 using the MURR pneumatic-tube facility. Phosphorus was measured via the 31 P 15 (n,α) 28 Al 13 reaction. After several days decay, the whole-body specimens were re-irradiated in the same facility, but without thermal-neutron shielding, for 5 seconds and the gamma-ray spectrum was recorded at two different decay periods allowing measurement of 77m Se, 24 Na, 27m g, 38 Cl, 42k , 49 Ca, 56 Mn, 66 Cu and 80 Br from the corresponding radiative-capture reactions. (author)

  4. Rapid-throughput skeletal phenotyping of 100 knockout mice identifies 9 new genes that determine bone strength.

    Directory of Open Access Journals (Sweden)

    J H Duncan Bassett

    Full Text Available Osteoporosis is a common polygenic disease and global healthcare priority but its genetic basis remains largely unknown. We report a high-throughput multi-parameter phenotype screen to identify functionally significant skeletal phenotypes in mice generated by the Wellcome Trust Sanger Institute Mouse Genetics Project and discover novel genes that may be involved in the pathogenesis of osteoporosis. The integrated use of primary phenotype data with quantitative x-ray microradiography, micro-computed tomography, statistical approaches and biomechanical testing in 100 unselected knockout mouse strains identified nine new genetic determinants of bone mass and strength. These nine new genes include five whose deletion results in low bone mass and four whose deletion results in high bone mass. None of the nine genes have been implicated previously in skeletal disorders and detailed analysis of the biomechanical consequences of their deletion revealed a novel functional classification of bone structure and strength. The organ-specific and disease-focused strategy described in this study can be applied to any biological system or tractable polygenic disease, thus providing a general basis to define gene function in a system-specific manner. Application of the approach to diseases affecting other physiological systems will help to realize the full potential of the International Mouse Phenotyping Consortium.

  5. 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.

  6. Suppression or knockout of SaF/SaM overcomes the Sa-mediated hybrid male sterility in rice.

    Science.gov (United States)

    Xie, Yongyao; Niu, Baixiao; Long, Yunming; Li, Gousi; Tang, Jintao; Zhang, Yaling; Ren, Ding; Liu, Yao-Guang; Chen, Letian

    2017-09-01

    Hybrids between the indica and japonica subspecies of rice (Oryza sativa) are usually sterile, which hinders utilization of heterosis in the inter-subspecific hybrid breeding. The complex locus Sa comprises two adjacently located genes, SaF and SaM, which interact to cause abortion of pollen grains carrying the japonica allele in japonica-indica hybrids. Here we showed that silencing of SaF or SaM by RNA interference restored male fertility in indica-japonica hybrids with heterozygous Sa. We further used clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9-based genome editing to knockout the SaF and SaM alleles, respectively, of an indica rice line to create hybrid-compatible lines. The resultant artificial neutral alleles did not affect pollen viability and other agricultural traits, but did break down the reproductive barrier in the hybrids. We found that some rice lines have natural neutral allele Sa-n, which was compatible with the typical japonica or indica Sa alleles in hybrids. Our results demonstrate that SaF and SaM are required for hybrid male sterility, but are not essential for pollen development. This study provides effective approaches for the generation of hybrid-compatible lines by knocking out the Sa locus or using the natural Sa-n allele to overcome hybrid male sterility in rice breeding. © 2017 The Authors. Bioelectromagnetics published by Wiley Periodicals, Inc. © 2017 Institute of Botany, Chinese Academy of Sciences.

  7. 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

  8. 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.

  9. Design of a multiplex PCR assay for the simultaneous detection and confirmation of Neisseria gonorrhoeae.

    LENUS (Irish Health Repository)

    O'Callaghan, Isabelle

    2010-05-01

    To improve the detection of Neisseria gonorrhoeae by designing a multiplex PCR assay using two N gonorrhoeae-specific genes as targets, thereby providing detection and confirmation of a positive result simultaneously.

  10. Histologically confirmed intracranial tumors managed at Enugu, Nigeria

    Directory of Open Access Journals (Sweden)

    Chika Anele Ndubuisi

    2017-01-01

    Full Text Available Background: There is controversy about the global distribution of intracranial tumors (ICTs. The previous reports from Africa suggested low frequency and different pattern of distribution of brain tumors from what obtains in other continents. The limitations at that time, including paucity of diagnostic facilities and personnel, have improved. Objective: The objective of this study is to analyze the current trend and distribution of histology confirmed brain tumors managed in Enugu, in a decade. Methods: A retrospective analysis of ICTs managed between 2006 and 2015 at Memfys Hospital, Enugu. Only cases with conclusive histology report were analyzed. The World Health Organization ICT classification was used. Results: This study reviewed 252 patients out of 612 neuroimaging diagnosed brain tumors. Mean age was 42.8 years and male-to-female ratio was 1.2:1.0. Annual frequency increased from 11 in 2006 to 55 in 2015. Metastatic brain tumors accounted for 5.6%, and infratentorial tumors represented 16.3%. Frequency of the common primary tumors were meningioma (32.9%, glioma (23.8%, pituitary adenomas (13.5%, and craniopharyngioma (7.5% (P = 0.001. Vestibular schwannoma accounted for 1.2%. Meningioma did not have gender difference (P = 0.714. Medulloblastoma, glioma, and craniopharyngioma were the most common pediatric tumors. About 8.7% presented unconscious (P < 0.001. There was no significant difference between radiology and histology diagnosis (P = 0.932. Conclusion: Meningioma is the most frequent tumor with increasing male incidence, but the frequency of glioma is increasing. Metastasis, acoustic schwannoma, lymphoma, and germ cell tumors seem to be uncommon. Late presentation is the rule.

  11. Physics of mind: Experimental confirmations of theoretical predictions.

    Science.gov (United States)

    Schoeller, Félix; Perlovsky, Leonid; Arseniev, Dmitry

    2018-02-02

    What is common among Newtonian mechanics, statistical physics, thermodynamics, quantum physics, the theory of relativity, astrophysics and the theory of superstrings? All these areas of physics have in common a methodology, which is discussed in the first few lines of the review. Is a physics of the mind possible? Is it possible to describe how a mind adapts in real time to changes in the physical world through a theory based on a few basic laws? From perception and elementary cognition to emotions and abstract ideas allowing high-level cognition and executive functioning, at nearly all levels of study, the mind shows variability and uncertainties. Is it possible to turn psychology and neuroscience into so-called "hard" sciences? This review discusses several established first principles for the description of mind and their mathematical formulations. A mathematical model of mind is derived from these principles. This model includes mechanisms of instincts, emotions, behavior, cognition, concepts, language, intuitions, and imagination. We clarify fundamental notions such as the opposition between the conscious and the unconscious, the knowledge instinct and aesthetic emotions, as well as humans' universal abilities for symbols and meaning. In particular, the review discusses in length evolutionary and cognitive functions of aesthetic emotions and musical emotions. Several theoretical predictions are derived from the model, some of which have been experimentally confirmed. These empirical results are summarized and we introduce new theoretical developments. Several unsolved theoretical problems are proposed, as well as new experimental challenges for future research. Copyright © 2017. Published by Elsevier B.V.

  12. Genetic Confirmation of Mungbean (Vigna radiata) and Mashbean (Vigna mungo) Interspecific Recombinants using Molecular Markers.

    Science.gov (United States)

    Abbas, Ghulam; Hameed, Amjad; Rizwan, Muhammad; Ahsan, Muhammad; Asghar, Muhammad J; Iqbal, Nayyer

    2015-01-01

    Molecular confirmation of interspecific recombinants is essential to overcome the issues like self-pollination, environmental influence, and inadequacy of morphological characteristics during interspecific hybridization. The present study was conducted for genetic confirmation of mungbean (female) and mashbean (male) interspecific crosses using molecular markers. Initially, polymorphic random amplified polymorphic DNA (RAPD), universal rice primers (URP), and simple sequence repeats (SSR) markers differentiating parent genotypes were identified. Recombination in hybrids was confirmed using these polymorphic DNA markers. The NM 2006 × Mash 88 was most successful interspecific cross. Most of true recombinants confirmed by molecular markers were from this cross combination. SSR markers were efficient in detecting genetic variability and recombination with reference to specific chromosomes and particular loci. SSR (RIS) and RAPD identified variability dispersed throughout the genome. In conclusion, DNA based marker assisted selection (MAS) efficiently confirmed the interspecific recombinants. The results provided evidence that MAS can enhance the authenticity of selection in mungbean improvement program.

  13. 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.

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

    Science.gov (United States)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  16. 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.

  17. 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].

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

    Science.gov (United States)

    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.

  19. 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

  20. 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.

  1. Confirmational study: a positive-based thumb and finger sucking elimination program.

    Science.gov (United States)

    Green, Shari E

    2010-11-01

    This article emphasizes the critical need for information specifically regarding the topic of retained sucking behaviors. The study aimed to confirm results provided by Van Norman of 723 subjects in 1997. Parent surveys were collected on 441 subjects who received an orofacial myofunctional treatment program provided by one certified orofacial myologist. Results of this study do confirm that retained digit sucking behavior may be addressed successfully and expediently by a program based on positive behavior modification techniques.

  2. Eric Besson: the financial advantage of nuclear energy is confirmed

    International Nuclear Information System (INIS)

    Anon.

    2012-01-01

    The French minister of energy, E. Besson said that the study of the Court of Auditors on the real costs of nuclear energy confirmed the competitiveness of nuclear power. The Court of Auditors confirmed also that public expenditures in favor of nuclear energy are balanced by the gain through the tax on nuclear facilities. The Court of Auditors confirms also that dismantlement charges and charges for the management of radioactive wastes are included in the present costs of nuclear energy at an adequate level with today's knowledge. The total cost of nuclear energy is very competitive, it ranges form 32.5 euros/MWh to 49.5 euros/MWh according to the cost accounting method used. One of major parameters for cost elaboration is the knowledge of the lengths of the operating life of the power plant. The longer the extension is, the lower is the investment cost. (A.C.)

  3. 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.

  4. Production of a mouse strain with impaired glucose tolerance by systemic heterozygous knockout of the glucokinase gene and its feasibility as a prediabetes model

    Science.gov (United States)

    SAITO, Mikako; KANEDA, Asako; SUGIYAMA, Tae; IIDA, Ryousuke; OTOKUNI, Keiko; KABURAGI, Misako; MATSUOKA, Hideaki

    2015-01-01

    Exon II of glucokinase (Gk) was deleted to produce a systemic heterozygous Gk knockout (Gk+/−) mouse. The relative expression levels of Gk in the heart, lung, liver, stomach, and pancreas in Gk+/− mice ranged from 0.41–0.68 versus that in wild (Gk+/+) mice. On the other hand, its expression levels in the brain, adipose tissue, and muscle ranged from 0.95–1.03, and its expression levels in the spleen and kidney were nearly zero. Gk knockout caused no remarkable off-target effect on the expression of 7 diabetes causing genes (Shp, Hnf1a, Hnf1b, Irs1, Irs2, Kir6.2, and Pdx1) in 10 organs. The glucose tolerance test was conducted to determine the blood glucose concentrations just after fasting for 24 h (FBG) and at 2 h after high-glucose application (GTT2h). The FBG-GTT2h plots obtained with the wild strain fed the control diet (CD), Gk+/− strain fed the CD, and Gk+/− strain fed the HFD were distributed in separate areas in the FBG-GTT2h diagram. The respective areas could be defined as the normal state, prediabetes state, and diabetes state, respectively. Based on the results, the criteria for prediabetes could be defined for the Gk+/− strain developed in this study. PMID:25765873

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

    Directory of Open Access Journals (Sweden)

    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.

  6. 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

  7. Thumb rule of visual angle: a new confirmation.

    Science.gov (United States)

    Groot, C; Ortega, F; Beltran, F S

    1994-02-01

    The classical thumb rule of visual angle was reexamined. Hence, the visual angle was measured as a function of a thumb's width and the distance between eye and thumb. The measurement of a thumb's width when held at arm's length was taken on 67 second-year students of psychology. The visual angle was about 2 degrees as R. P. O'Shea confirmed in 1991. Also, we confirmed a linear relationship between the size of a thumb's width at arm's length and the visual angle.

  8. 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.

  9. Regional registration of [6-14C]glucose metabolism during brain activation of α-syntrophin knockout mice

    Science.gov (United States)

    Cruz, Nancy F.; Ball, Kelly K.; Froehner, Stanley C.; Adams, Marvin E.; Dienel, Gerald A.

    2013-01-01

    α-Syntrophin is a component of the dystrophin scaffold-protein complex that serves as an adaptor for recruitment of key proteins to the cytoplasmic side of plasma membranes. α-Syntrophin knockout (KO) causes loss of the polarized localization of aquaporin4 (AQP4) at astrocytic endfeet and interferes with water and K+ homeostasis. During brain activation, release of ions and metabolites from endfeet is anticipated to increase perivascular fluid osmolarity, AQP4-mediated osmotic water flow from endfeet, and metabolite washout from brain. This study tests the hypothesis that reduced levels of endfoot AQP4 increase retention of [14C]metabolites during sensory stimulation. Conscious KO and wildtype mice were pulse-labeled with [6-14C]glucose during unilateral acoustic stimulation or bilateral acoustic plus whisker stimulation, and label retention was assayed by computer-assisted brain imaging or analysis of [14C]metabolites in extracts, respectively. High-resolution autoradiographic assays detected a 17% side-to-side difference (P<0.05) in inferior colliculus of KO mice, not wildtype mice. However, there were no labeling differences between KO and wildtype mice for five major HPLC fractions from four dissected regions, presumably due to insufficient anatomical resolution. The results suggest a role for AQP4-mediated water flow in support of washout of metabolites, and underscore the need for greater understanding of astrocytic water and metabolite fluxes. PMID:23346911

  10. Studies on functional roles of the histaminergic neuron system by using pharmacological agents, knockout mice and positron emission tomography

    International Nuclear Information System (INIS)

    Watanabe, Takehiko; Yanai, Kazuhiko

    2001-01-01

    Since one of us, Takehiko Watanabe (TW), elucidated the location and distribution of the histaminergic neuron system in the brain with antibody raised against L-histidine decarboxylase (a histamine-forming enzyme, HDC) as a marker in 1984 and came to Tohoku University School of Medicine in Sendai, we have been collaborating on the functions of this neuron system by using pharmacological agents, knockout mice of the histamine-related genes, and, in some cases, positron emission tomography (PET). Many of our graduate students and colleagues have been actively involved in histamine research since 1985. Our extensive studies have clarified some of the functions of histamine neurons using methods from molecular techniques to non-invasive human PET imaging. Histamine neurons are involved in many brain functions, such as spontaneous locomotion, arousal in wake-sleep cycle, appetite control, seizures, learning and memory, aggressive behavior and emotion. Particularly, the histaminergic neuron system is one of the most important neuron systems to maintain and stimulate wakefulness. Histamine also functions as a biprotection system against various noxious and unfavorable stimuli (for examples, convulsion, nociception, drug sensitization, ischemic lesions, and stress). Although activators of histamine neurons have not been clinically available until now, we would like to point out that the activation of the histaminergic neuron system is important to maintain mental health. Here, we summarize the newly-discovered functions of histamine neurons mainly on the basis of results from our research groups. (author)

  11. Curcumin Protects against Atherosclerosis in Apolipoprotein E-Knockout Mice by Inhibiting Toll-like Receptor 4 Expression.

    Science.gov (United States)

    Zhang, Shanshan; Zou, Jun; Li, Peiyang; Zheng, Xiumei; Feng, Dan

    2018-01-17

    Toll-like receptor 4 (TLR4) has been reported to play a critical role in the pathogenesis of atherosclerosis, the current study aimed to investigate whether curcumin suppresses atherosclerosis development in ApoE-knockout (ApoE -/- ) mice by inhibiting TLR4 expression. ApoE -/- mice were fed a high-fat diet supplemented with or without curcumin (0.1% w/w) for 16 weeks. Curcumin supplementation significantly reduced TLR4 expression and macrophage infiltration in atherosclerotic plaques. Curcumin also reduced aortic interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) expression, nuclear factor-κB (NF-κB) activity, and plasma IL-1β, TNF-α, soluble VCAM-1 and ICAM-1 levels. In addition, aortic sinus sections revealed that curcumin treatment reduced the extent of atherosclerotic lesions and inhibited atherosclerosis development. In vitro, curcumin inhibited NF-κB activation in macrophages and reduced TLR4 expression induced by lipopolysaccharide. Our results indicate that curcumin protects against atherosclerosis at least partially by inhibiting TLR4 expression and its related inflammatory reaction.

  12. A muscle-specific knockout implicates nuclear receptor coactivator MED1 in the regulation of glucose and energy metabolism.

    Science.gov (United States)

    Chen, Wei; Zhang, Xiaoting; Birsoy, Kivanc; Roeder, Robert G

    2010-06-01

    As conventional transcriptional factors that are activated in diverse signaling pathways, nuclear receptors play important roles in many physiological processes that include energy homeostasis. The MED1 subunit of the Mediator coactivator complex plays a broad role in nuclear receptor-mediated transcription by anchoring the Mediator complex to diverse promoter-bound nuclear receptors. Given the significant role of skeletal muscle, in part through the action of nuclear receptors, in glucose and fatty acid metabolism, we generated skeletal muscle-specific Med1 knockout mice. Importantly, these mice show enhanced insulin sensitivity and improved glucose tolerance as well as resistance to high-fat diet-induced obesity. Furthermore, the white muscle of these mice exhibits increased mitochondrial density and expression of genes specific to type I and type IIA fibers, indicating a fast-to-slow fiber switch, as well as markedly increased expression of the brown adipose tissue-specific UCP-1 and Cidea genes that are involved in respiratory uncoupling. These dramatic results implicate MED1 as a powerful suppressor in skeletal muscle of genetic programs implicated in energy expenditure and raise the significant possibility of therapeutical approaches for metabolic syndromes and muscle diseases through modulation of MED1-nuclear receptor interactions.

  13. Advanced age-related denervation and fiber-type grouping in skeletal muscle of SOD1 knockout mice.

    Science.gov (United States)

    Kostrominova, Tatiana Y

    2010-11-30

    In this study skeletal muscles from 1.5- and 10-month-old Cu/Zn superoxide dismutase (SOD1) homozygous knockout (JLSod1(-/-)) mice obtained from The Jackson Laboratory (C57Bl6/129SvEv background) were compared with muscles from age- and sex-matched heterozygous (JLSod1(+/-)) littermates. The results of this study were compared with previously published data on two different strains of Sod1(-/-) mice: one from Dr. Epstein's laboratory (ELSod1(-/-); C57Bl6 background) and the other from Cephalon, Inc. (CSod1(-/-); 129/CD-1 background). Grouping of succinate dehydrogenase-positive fibers characterized muscles of Sod1(-/-) mice from all three strains. The 10-month-old Sod1(-/-)C and JL mice displayed pronounced denervation of the gastrocnemius muscle, whereas the ELSod1(-/-) mice displayed a small degree of denervation at this age, but developed accelerated age-related denervation later on. Denervation markers were up-regulated in skeletal muscle of 10-month-old JLSod1(-/-) mice. This study is the first to show that metallothionein mRNA and protein expression was up-regulated in the skeletal muscle of 10-month-old JLSod1(-/-) mice and was mostly localized to the small atrophic muscle fibers. In conclusion, all three strains of Sod1(-/-) mice develop accelerated age-related muscle denervation, but the genetic background has significant influence on the progress of denervation. Copyright © 2010 Elsevier Inc. All rights reserved.

  14. [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.

  15. Cellular and Molecular Mechanisms of Anterior Chamber-Associated Immune Deviation (ACAID: What We Have Learned from Knockout Mice

    Directory of Open Access Journals (Sweden)

    Julie Vendomèle

    2017-11-01

    Full Text Available Anterior chamber-associated immune deviation (ACAID is a well-known phenomenon that can occur after an antigen is introduced without any danger signal into the anterior chamber of a murine eye. It is reported to lead to an antigen-specific immune deviation throughout the body. Despite the relatively little evidence of this phenomenon in humans, it has been suggested as a potential prophylactic strategy in allograft rejections and in several autoimmune diseases. Cellular and molecular mechanisms of ACAID have been explored in different murine models mainly as proofs of concept, first by direct analyses of immune components in normal immunocompetent settings and by cell transfer experiments. Later, use of knockout (KO mice has helped considerably to decipher ACAID mechanisms. However, several factors raise questions about the reliability and validity of studies using KO murine models. This mini-review summarizes results obtained with KO mice and discusses their advantages, their potential weaknesses, and their potential methods for further progress.

  16. Studies on functional roles of the histaminergic neuron system by using pharmacological agents, knockout mice and positron emission tomography

    Energy Technology Data Exchange (ETDEWEB)

    Watanabe, Takehiko; Yanai, Kazuhiko [Tohoku Univ., Sendai (Japan). Graduate School of Medicine

    2001-12-01

    Since one of us, Takehiko Watanabe (TW), elucidated the location and distribution of the histaminergic neuron system in the brain with antibody raised against L-histidine decarboxylase (a histamine-forming enzyme, HDC) as a marker in 1984 and came to Tohoku University School of Medicine in Sendai, we have been collaborating on the functions of this neuron system by using pharmacological agents, knockout mice of the histamine-related genes, and, in some cases, positron emission tomography (PET). Many of our graduate students and colleagues have been actively involved in histamine research since 1985. Our extensive studies have clarified some of the functions of histamine neurons using methods from molecular techniques to non-invasive human PET imaging. Histamine neurons are involved in many brain functions, such as spontaneous locomotion, arousal in wake-sleep cycle, appetite control, seizures, learning and memory, aggressive behavior and emotion. Particularly, the histaminergic neuron system is one of the most important neuron systems to maintain and stimulate wakefulness. Histamine also functions as a biprotection system against various noxious and unfavorable stimuli (for examples, convulsion, nociception, drug sensitization, ischemic lesions, and stress). Although activators of histamine neurons have not been clinically available until now, we would like to point out that the activation of the histaminergic neuron system is important to maintain mental health. Here, we summarize the newly-discovered functions of histamine neurons mainly on the basis of results from our research groups. (author)

  17. 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.

  18. Reconstructing gene regulatory networks from knock-out data using Gaussian Noise Model and Pearson Correlation Coefficient.

    Science.gov (United States)

    Mohamed Salleh, Faridah Hani; Arif, Shereena Mohd; Zainudin, Suhaila; Firdaus-Raih, Mohd

    2015-12-01

    A gene regulatory network (GRN) is a large and complex network consisting of interacting elements that, over time, affect each other's state. The dynamics of complex gene regulatory processes are difficult to understand using intuitive approaches alone. To overcome this problem, we propose an algorithm for inferring the regulatory interactions from knock-out data using a Gaussian model combines with Pearson Correlation Coefficient (PCC). There are several problems relating to GRN construction that have been outlined in this paper. We demonstrated the ability of our proposed method to (1) predict the presence of regulatory interactions between genes, (2) their directionality and (3) their states (activation or suppression). The algorithm was applied to network sizes of 10 and 50 genes from DREAM3 datasets and network sizes of 10 from DREAM4 datasets. The predicted networks were evaluated based on AUROC and AUPR. We discovered that high false positive values were generated by our GRN prediction methods because the indirect regulations have been wrongly predicted as true relationships. We achieved satisfactory results as the majority of sub-networks achieved AUROC values above 0.5. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Knockout of Murine Mamld1 Impairs Testicular Growth and Daily Sperm Production but Permits Normal Postnatal Androgen Production and Fertility.

    Science.gov (United States)

    Miyado, Mami; Yoshida, Kaoru; Miyado, Kenji; Katsumi, Momori; Saito, Kazuki; Nakamura, Shigeru; Ogata, Tsutomu; Fukami, Maki

    2017-06-19

    MAMLD1 has been implicated in testicular function in both human and mouse fetuses. Although three patients with MAMLD1 mutations were reported to have hypergonadotropic hypogonadism in their teens, the functional significance of MAMLD1 in the postnatal testis remains unclear. Here, we analyzed the phenotype of Mamld1 knockout (KO) male mice at reproductive ages. The reproductive organs of KO male mice were morphologically unremarkable, except for relatively small testes. Seminiferous tubule size and number of proliferating spermatogonia/spermatocytes were reduced in the KO testis. Daily sperm production of KO mice was mildly attenuated, whereas total sperm counts in epididymal semen remained normal. Sperm motility and morphology, as well as androgen levels in serum and testicular tissues and the number of pups born from cross-mated wildtype (WT) female mice, were comparable between WT and KO male mice. These results indicate that MAMLD1 contributes to the maintenance of postnatal testicular growth and daily sperm production but is dispensable for androgen biosynthesis and fertility. MAMLD1 likely plays supporting roles in multiple and continuous steps of male reproduction.

  20. Knockout of Murine Mamld1 Impairs Testicular Growth and Daily Sperm Production but Permits Normal Postnatal Androgen Production and Fertility

    Directory of Open Access Journals (Sweden)

    Mami Miyado

    2017-06-01

    Full Text Available MAMLD1 has been implicated in testicular function in both human and mouse fetuses. Although three patients with MAMLD1 mutations were reported to have hypergonadotropic hypogonadism in their teens, the functional significance of MAMLD1 in the postnatal testis remains unclear. Here, we analyzed the phenotype of Mamld1 knockout (KO male mice at reproductive ages. The reproductive organs of KO male mice were morphologically unremarkable, except for relatively small testes. Seminiferous tubule size and number of proliferating spermatogonia/spermatocytes were reduced in the KO testis. Daily sperm production of KO mice was mildly attenuated, whereas total sperm counts in epididymal semen remained normal. Sperm motility and morphology, as well as androgen levels in serum and testicular tissues and the number of pups born from cross-mated wildtype (WT female mice, were comparable between WT and KO male mice. These results indicate that MAMLD1 contributes to the maintenance of postnatal testicular growth and daily sperm production but is dispensable for androgen biosynthesis and fertility. MAMLD1 likely plays supporting roles in multiple and continuous steps of male reproduction.

  1. Kidneys From α1,3-Galactosyltransferase Knockout/Human Heme Oxygenase-1/Human A20 Transgenic Pigs Are Protected From Rejection During Ex Vivo Perfusion With Human Blood.

    Science.gov (United States)

    Ahrens, Hellen E; Petersen, Björn; Ramackers, Wolf; Petkov, Stoyan; Herrmann, Doris; Hauschild-Quintern, Janet; Lucas-Hahn, Andrea; Hassel, Petra; Ziegler, Maren; Baars, Wiebke; Bergmann, Sabine; Schwinzer, Reinhard; Winkler, Michael; Niemann, Heiner

    2015-07-01

    Multiple modifications of the porcine genome are required to prevent rejection after pig-to-primate xenotransplantation. Here, we produced pigs with a knockout of the α1,3-galactosyltransferase gene (GGTA1-KO) combined with transgenic expression of the human anti-apoptotic/anti-inflammatory molecules heme oxygenase-1 and A20, and investigated their xenoprotective properties. The GGTA1-KO/human heme oxygenase-1 (hHO-1)/human A20 (hA20) transgenic pigs were produced in a stepwise approach using zinc finger nuclease vectors targeting the GGTA1 gene and a Sleeping Beauty vector coding for hA20. Two piglets were analyzed by quantitative reverse-transcription polymerase chain reaction, flow cytometry, and sequencing. The biological function of the genetic modifications was tested in a (51)Chromium release assay and by ex vivo kidney perfusions with human blood. Disruption of the GGTA1 gene by deletion of few basepairs was demonstrated in GGTA1-KO/hHO-1/hA20 transgenic pigs. The hHO-1 and hA20 mRNA expression was confirmed by quantitative reverse-transcription polymerase chain reaction. Ex vivo perfusion of 2 transgenic kidneys was feasible for the maximum experimental time of 240 minutes without symptoms of rejection. Results indicate that GGTA1-KO/hHO-1/hA20 transgenic pigs are a promising model to alleviate rejection and ischemia-reperfusion damage in porcine xenografts and could serve as a background for further genetic modifications toward the production of a donor pig that is clinically relevant for xenotransplantation.

  2. PrP Knockout Cells Expressing Transmembrane PrP Resist Prion Infection.

    Science.gov (United States)

    Marshall, Karen E; Hughson, Andrew; Vascellari, Sarah; Priola, Suzette A; Sakudo, Akikazu; Onodera, Takashi; Baron, Gerald S

    2017-01-15

    Glycosylphosphatidylinositol (GPI) anchoring of the prion protein (PrP C ) influences PrP C misfolding into the disease-associated isoform, PrP res , as well as prion propagation and infectivity. GPI proteins are found in cholesterol- and sphingolipid-rich membrane regions called rafts. Exchanging the GPI anchor for a nonraft transmembrane sequence redirects PrP C away from rafts. Previous studies showed that nonraft transmembrane PrP C variants resist conversion to PrP res when transfected into scrapie-infected N2a neuroblastoma cells, likely due to segregation of transmembrane PrP C and GPI-anchored PrP res in distinct membrane environments. Thus, it remained unclear whether transmembrane PrP C might convert to PrP res if seeded by an exogenous source of PrP res not associated with host cell rafts and without the potential influence of endogenous expression of GPI-anchored PrP C To further explore these questions, constructs containing either a C-terminal wild-type GPI anchor signal sequence or a nonraft transmembrane sequence containing a flexible linker were expressed in a cell line derived from PrP knockout hippocampal neurons, NpL2. NpL2 cells have physiological similarities to primary neurons, representing a novel and advantageous model for studying transmissible spongiform encephalopathy (TSE) infection. Cells were infected with inocula from multiple prion strains and in different biochemical states (i.e., membrane bound as in brain microsomes from wild-type mice or purified GPI-anchorless amyloid fibrils). Only GPI-anchored PrP C supported persistent PrP res propagation. Our data provide strong evidence that in cell culture GPI anchor-directed membrane association of PrP C is required for persistent PrP res propagation, implicating raft microdomains as a location for conversion. Mechanisms of prion propagation, and what makes them transmissible, are poorly understood. Glycosylphosphatidylinositol (GPI) membrane anchoring of the prion protein (PrP C

  3. Confirming psychogenic nonepileptic seizures with video-EEG: sex matters.

    Science.gov (United States)

    Noe, Katherine H; Grade, Madeline; Stonnington, Cynthia M; Driver-Dunckley, Erika; Locke, Dona E C

    2012-03-01

    The influence of gender on psychogenic nonepileptic seizures (PNES) diagnosis was examined retrospectively in 439 subjects undergoing video-EEG (vEEG) for spell classification, of whom 142 women and 42 men had confirmed PNES. The epileptologist's predicted diagnosis was correct in 72% overall. Confirmed epilepsy was correctly predicted in 94% men and 88% women. In contrast, confirmed PNES was accurately predicted in 86% women versus 61% men (p=0.003). Sex-based differences in likelihood of an indeterminate admission were not observed for predicted epilepsy or physiologic events, but were for predicted PNES (39% men, 12% women, p=0.0002). More frequent failure to record spells in men than women with predicted PNES was not explained by spell frequency, duration of monitoring, age, medication use, or personality profile. PNES are not only less common in men, but also more challenging to recognize in the clinic, and even when suspected more difficult to confirm with vEEG. Copyright © 2012 Elsevier Inc. All rights reserved.

  4. Confirmation of the absolute configuration of (−)-aurantioclavine

    KAUST Repository

    Behenna, Douglas C.

    2011-04-01

    We confirm our previous assignment of the absolute configuration of (-)-aurantioclavine as 7R by crystallographically characterizing an advanced 3-bromoindole intermediate reported in our previous synthesis. This analysis also provides additional support for our model of enantioinduction in the palladium(II)-catalyzed oxidative kinetic resolution of secondary alcohols. © 2010 Elsevier Ltd. All rights reserved.

  5. Exploratory laparotomy in the management of confirmed necrotizing ...

    African Journals Online (AJOL)

    confirmed NEC who undergo laparotomy remain high in infants despite optimal medical and surgical care. Ann. Pediatr Surg 11:123–126 c 2015 Annals of Pediatric. Surgery. ... Radcliffe Hospital, Headley Way, Headington, Oxford OX3 9DU, UK e-mail: ... The institutional review board of the John Radcliffe. Hospital, Oxford ...

  6. 18 CFR 300.10 - Application for confirmation and approval.

    Science.gov (United States)

    2010-04-01

    ... REGULATORY COMMISSION, DEPARTMENT OF ENERGY REGULATIONS FOR FEDERAL POWER MARKETING ADMINISTRATIONS CONFIRMATION AND APPROVAL OF THE RATES OF FEDERAL POWER MARKETING ADMINISTRATIONS Filing Requirements § 300.10... with applicable laws and that it is the lowest possible rate consistent with sound business principles...

  7. Theory-Led Confirmation Bias and Experimental Persona

    Science.gov (United States)

    Allen, Michael

    2011-01-01

    Questionnaire and interview findings from a survey of three Year 8 (ages 12-13 years) science practical lessons (n = 52) demonstrate how pupils' data collection and inference making were sometimes biased by desires to confirm a personal theory. A variety of behaviours are described where learners knowingly rejected anomalies, manipulated…

  8. Factors associated with child sexual abuse confirmation at forensic examinations

    Directory of Open Access Journals (Sweden)

    Welington dos Santos Silva

    Full Text Available Abstract The aim of this study is identify potential factors associated with child sexual abuse confirmation at forensic examinations. The forensic files of children under 12 years of age reporting sexual abuse at the Nina Rodrigues Institute of Forensic Medicine in Salvador, Bahia, Brazil between January 2008 and December 2009 were reviewed. A multivariate analysis was conducted to identify factors associated with finding evidence of sexual abuse in forensic examinations. The proportion of cases confirmed by the forensic physician based on material evidence was 10.4%. Adjusted analysis showed that the variables place of birth, type of abuse reported, family relationship between the child and the perpetrator, and the interval between the reported abuse and the forensic examination were not independently associated with finding forensic evidence of sexual abuse. A report of penetration was associated with a five-fold greater likelihood of confirmation, while the victim being 10-11 years of age was associated with a two-fold of abuse confirmation than younger children. These findings should be taken into consideration when drawing up guidelines for the multidisciplinary evaluation of children suspected of being victims of sexual abuse and in deciding whether to refer the child for forensic examination.

  9. Psychotherapy of Depression: A Self-Confirmation Model.

    Science.gov (United States)

    Andrews, John D. W.

    1989-01-01

    Concepts of self-confirmation, interpersonal diagnosis, and prototype construction are used to integrate research and clinical findings concerning depression. Various theoretical accounts and bodies of data that fit within this integrative conceptual framework are examined, and implications for psychotherapy are discussed. (SLD)

  10. Confirmation of radiation pressure effects in laser--plasma interactions

    International Nuclear Information System (INIS)

    Attwood, D.T.; Sweeney, D.W.; Auerbach, J.M.; Lee, P.H.Y.

    1977-10-01

    Interferometric data resolved in 1μm and 15 psec confirms the dominant role of radiation pressure during high intensity laser-plasma interactions. Specifically observed manifestations include electron density profiles steepened to 1 μm scale length, clearly defined upper and lower density shelves, and small and large scale deformation of transverse isodensity surfaces

  11. Confirmation of antibodies against L-tryptophan-like epitope in ...

    African Journals Online (AJOL)

    Confirmation of antibodies against L-tryptophan-like epitope in human African trypanosomosis serological diagnostic. ... number of patients in Congo. A diagnostic test based on this synthetic epitope, especially in combination with other tests, might improve the HAT diagnostic test in field conditions. Key words: Tryptophan ...

  12. 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.

  13. 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

  14. [Epidemiological investigation on confirmed cases of schistosomiasis in Hubei Province].

    Science.gov (United States)

    Yan-Yan, Chen; Shun-Xiang, Cai; Guo, Li; Ying, Xiao; Xiao-Wei, Shan; Juan, Zhang; Jian-Bing, Liu

    2016-05-10

    To grasp the distribution and epidemiology of confirmed cases of schistosomiasis in Hubei Province, so as to provide the evidence for promoting the prevention and control work. The confirmed cases of schistosomiasis in Hubei Province from 2010 to 2014 were epidemiologically investigated, and the prevalence characteristics and main influencing factors were analyzed. A total of 10 102 confirmed cases from 2010 to 2014 were surveyed. There were 1 062 local infected patients, accounting for 10.51% and including 354 repeated infections and 17 newly infected. There were 290 foreigninfected patients, accounting for 2.87%, with 206 repeated infection cases and 84 newly infected. There were 8 750 historical patients, including 2 229 patients who leaked the former schistosomiasis investigations, accounting for 22.06%; 570 patients missed treatment, accounting for 5.64%; 3 640 patients were treated with non-standard therapy, accounting for 36.03%; 2 311 patients were treated with poor medication efficacy, accounting for 22.88%. The multivariate non-conditional Logistic regression, targeting at confirmed cases in 2014, showed that, for the leaking investigations, the potential risk factors included the age, educational level, and latrine renovation ( b >0, OR >1), the protective factors were the times of previous treatment, cattle feeding in villager team, and Oncomelania hupensis snails in surroundings ( b <0, OR <1); for the treatment-missing, the age, educational level, snails in the surroundings of residence were risk factors ( b <0, OR <1); for the substandard treatment, the risk factors included the occupation and snails in the surroundings of residence ( b >0, OR >1), and the educational level and snails in the own field were protective factors ( b <0, OR <1). The epidemiological investigation on the confirmed cases of schistosomiasis could grasp the epidemic factors so as to improve the management and carry out the scientific control.

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

    Directory of Open Access Journals (Sweden)

    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.

  16. 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....

  17. 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

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

    Energy Technology Data Exchange (ETDEWEB)

    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

  19. [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

  20. Knockout of the alanine racemase gene in Lactobacillus plantarum results in septation defects and cell wall perforation

    NARCIS (Netherlands)

    Palumbo, E.; Favier, C.F.; Deghorain, M.; Cocconcelli, P.S.; Grangette, C.; Mercenier, A.M.E.; Vaughan, E.E.; Hols, P.

    2004-01-01

    A stable mutant of Lactobacillus plantarum deficient in alanine racemase (Alr) was constructed by two successive homologous recombination steps. When the mutant was supplemented with D-alanine, growth and viability were unaffected. Surprisingly, deprivation Of D-alanine during exponential growth did

  1. Caldwell Ranch Exploration and Confirmation Project, Northwest Geysers, CA

    Energy Technology Data Exchange (ETDEWEB)

    Walters, Mark A.

    2013-04-25

    The purpose of the Caldwell Ranch Exploration and Confirmation Project was to drill, test, and confirm the present economic viability of the undeveloped geothermal reservoir in the 870 acre Caldwell Ranch area of the Northwest Geysers that included the CCPA No.1 steam field. All of the drilling, logging, and sampling challenges were met. Three abandoned wells, Prati 5, Prati 14 and Prati 38 were re-opened and recompleted to nominal depths of 10,000 feet in 2010. Two of the wells required sidetracking. The flow tests indicated Prati 5 Sidetrack 1 (P-5 St1), Prati 14 (P-14) and Prati 38 Sidetrack 2 (P-38 St2) were collectively capable of initially producing an equivalent of 12 megawatts (MWe) of steam using a conversion rate of 19,000 pounds of steam/hour

  2. Confirmation tests of PWR surveillance capsule shipping container

    International Nuclear Information System (INIS)

    Tomita, N.; Ue, K.; Ohashi, M.; Asada, K.; Yoneda, Y.

    1980-01-01

    Mitsubishi Heavy Industries, Ltd. carried out the confirmation tests to confirm the reliability of the PWR surveillance capsule shipping container and to collect cask design data using a 10-ton weight full scale model at Kobe Shipyard and Engine Works. This report presents the outline of these tests. The B Type container was a cylinder 3289 mm long, 1080 mm in diameter and designed in accordance with the new modified Japanese regulations similar to IAEA regulation. These tests consist of four 9 m drop tests, two 1 m puncture tests, a fire test and an immersion test. In conclusion, safetyness of this container has been proved and various technical data for cask design were also collected through these tests. (author)

  3. Confirmation of the decontamination ability using the d