331 research outputs found

    Evaluation of Intra-Host Variants of the Entire Hepatitis B Virus Genome

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    Genetic analysis of hepatitis B virus (HBV) frequently involves study of intra-host variants, identification of which is commonly achieved using short regions of the HBV genome. However, the use of short sequences significantly limits evaluation of genetic relatedness among HBV strains. Although analysis of HBV complete genomes using genetic cloning has been developed, its application is highly labor intensive and practiced only infrequently. We describe here a novel approach to whole genome (WG) HBV quasispecies analysis based on end-point, limiting-dilution real-time PCR (EPLD-PCR) for amplification of single HBV genome variants, and their subsequent sequencing. EPLD-PCR was used to analyze WG quasispecies from serum samples of patients (n = 38) infected with HBV genotypes A, B, C, D, E and G. Phylogenetic analysis of the EPLD-isolated HBV-WG quasispecies showed the presence of mixed genotypes, recombinant variants and sub-populations of the virus. A critical observation was that HBV-WG consensus sequences obtained by direct sequencing of PCR fragments without EPLD are genetically close, but not always identical to the major HBV variants in the intra-host population, thus indicating that consensus sequences should be judiciously used in genetic analysis. Sequence-based studies of HBV WG quasispecies should afford a more accurate assessment of HBV evolution in various clinical and epidemiological settings

    Cannabinoid type 1 receptor-containing axons innervate NPY/AgRP neurons in the mouse arcuate nucleus

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    Objectives: Phytocannabinoids, such as THC and endocannabinoids, are well known to promote feeding behavior and to control energy metabolism through cannabinoid type 1 receptors (CB1R). However, the underlying mechanisms are not fully understood. Generally, cannabinoid-conducted retrograde dis-inhibition of hunger-promoting neurons has been suggested to promote food intake, but so far it has not been demonstrated due to technical limitations. Methods: We applied immunohistochemical labeling of CB1R for light microscopy and electron microscopy combined with three-dimensional reconstruction from serial sections in CB1R-expressing and CB1R-null mice, which served as a negative control. Hunger-promoting neurons expressing Agouti-related protein and neuropeptide Y (AgRP/NPY) in the hypothalamic arcuate nucleus were identified in NPY-GFP and NPY-hrGFP mice. Results: Using three-dimensional reconstruction from serial sections we demonstrated numerous discontinuous segments of anti-CB1R labeling in the synaptic boutons and axonal shafts in the arcuate nucleus. We observed CB1R in the symmetric, presumed GABAergic, synaptic boutons innervating AgRP/NPY neurons. We also detected CB1R-containing axons producing symmetric and asymmetric synapses onto AgRP/NPY-negative neurons. Furthermore, we identified CB1R in close apposition to the endocannabinoid (2-arachidonoylglycerol)-synthesizing enzyme diacylglycerol lipase-alpha at AgRP/NPY neurons. Conclusions: Our immunohistochemical and ultrastructural study demonstrates the morphological substrate for cannabinoid-conducted feeding behavior via retrograde dis-inhibition of hunger-promoting AgRP/NPY neurons. Keywords: Hypothalamus, Arcuate nucleus, Agouti-related protein, Neuropeptide Y, Electron microscopy, 3D reconstructio

    Cannabinoid Type 1 Receptor is Undetectable in Rodent and Primate Cerebral Neural Stem Cells but Participates in Radial Neuronal Migration

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    Cannabinoid type 1 receptor (CB1R) is expressed and participates in several aspects of cerebral cortex embryonic development as demonstrated with whole-transcriptome mRNA sequencing and other contemporary methods. However, the cellular location of CB1R, which helps to specify molecular mechanisms, remains to be documented. Using three-dimensional (3D) electron microscopic reconstruction, we examined CB1R immunolabeling in proliferating neural stem cells (NSCs) and migrating neurons in the embryonic mouse (Mus musculus) and rhesus macaque (Macaca mulatta) cerebral cortex. We found that the mitotic and postmitotic ventricular and subventricular zone (VZ and SVZ) cells are immunonegative in both species while radially migrating neurons in the intermediate zone (IZ) and cortical plate (CP) contain CB1R-positive intracellular vesicles. CB1R immunolabeling was more numerous and more extensive in monkeys compared to mice. In CB1R-knock out mice, projection neurons in the IZ show migration abnormalities such as an increased number of lateral processes. Thus, in radially migrating neurons CB1R provides a molecular substrate for the regulation of cell movement. Undetectable level of CB1R in VZ/SVZ cells indicates that previously suggested direct CB1R-transmitted regulation of cellular proliferation and fate determination demands rigorous re-examination. More abundant CB1R expression in monkey compared to mouse suggests that therapeutic or recreational cannabis use may be more distressing for immature primate neurons than inferred from experiments with rodents

    Galileo E5 AltBOC Signals: Application for Single-Frequency Total Electron Content Estimations

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    Global navigation satellite system signals are known to be an efficient tool to monitor the Earth ionosphere. We suggest Galileo E5 AltBOC phase and pseudorange observables—a single-frequency combination—to estimate the ionospheric total electron content (TEC). We performed a one-month campaign in September 2020 to compare the noise level for different TEC estimations based on single-frequency and dual-frequency data. Unlike GPS, GLONASS, or Galileo E5a and E5b single-frequency TEC estimations (involving signals with binary and quadrature phase-shift keying, such as BPSK and QPSK, or binary offset carrier (BOC) modulation), an extra wideband Galileo E5 AltBOC signal provided the smallest noise level, comparable to that of dual-frequency GPS. For elevation higher than 60 degrees, the 100 s root-mean-square (RMS) of TEC, an estimated TEC noise proxy, was as follows for different signals: ~0.05 TECU for Galileo E5 AltBOC, 0.09 TECU for GPS L5, ~0.1TECU for Galileo E5a/E5b BPSK, and 0.85 TECU for Galileo E1 CBOC. Dual-frequency phase combinations provided RMS values of 0.03 TECU for Galileo E1/E5, 0.03 and 0.07 TECU for GPS L1/L2 and L1/L5. At low elevations, E5 AltBOC provided at least twice less single-frequency TEC noise as compared with data obtained from E5a or E5b. The short dataset of our study could limit the obtained estimates; however, we expect that the AltBOC single-frequency TEC will still surpass the BPSK analogue in noise parameters when the solar cycle evolves and geomagnetic activity increases. Therefore, AltBOC signals could advance geoscience

    Siberian Snakes, Figure-8 and Spin Transparency Techniques for High Precision Experiments with Polarized Hadron Beams in Colliders

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    We present a review of the possibilities to conduct experiments of high efficiency in the nuclear and high energy physics with spin-polarized beams in a collider complex, configuration of which includes Siberian snakes or figure-8 collider ring. Special attention is given to the recently elicited advantageous possibility to conduct high precision experiments in a regime of the spin transparency (ST) when the design global spin tune is close to zero. In this regime, the polarization control is realized by use of spin navigators (SN), which are compact special insertions of magnets dedicated to a high flexibility spin manipulation including frequent spin flips

    Siberian Snakes, Figure-8 and Spin Transparency Techniques for High Precision Experiments with Polarized Hadron Beams in Colliders

    No full text
    We present a review of the possibilities to conduct experiments of high efficiency in the nuclear and high energy physics with spin-polarized beams in a collider complex, configuration of which includes Siberian snakes or figure-8 collider ring. Special attention is given to the recently elicited advantageous possibility to conduct high precision experiments in a regime of the spin transparency (ST) when the design global spin tune is close to zero. In this regime, the polarization control is realized by use of spin navigators (SN), which are compact special insertions of magnets dedicated to a high flexibility spin manipulation including frequent spin flips
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