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  • 1
    In: Journal of Virology, American Society for Microbiology, Vol. 94, No. 15 ( 2020-07-16)
    Abstract: Currently, there are four seasonal coronaviruses associated with relatively mild respiratory tract disease in humans. However, there is also a plethora of animal coronaviruses which have the potential to cross the species border. This regularly results in the emergence of new viruses in humans. In 2002, severe acute respiratory syndrome coronavirus (SARS-CoV) emerged and rapidly disappeared in May 2003. In 2012, Middle East respiratory syndrome coronavirus (MERS-CoV) was identified as a possible threat to humans, but its pandemic potential so far is minimal, as human-to-human transmission is ineffective. The end of 2019 brought us information about severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emergence, and the virus rapidly spread in 2020, causing an unprecedented pandemic. At present, studies on the virus are carried out using a surrogate system based on the immortalized simian Vero E6 cell line. This model is convenient for diagnostics, but it has serious limitations and does not allow for understanding of the biology and evolution of the virus. Here, we show that fully differentiated human airway epithelium cultures constitute an excellent model to study infection with the novel human coronavirus SARS-CoV-2. We observed efficient replication of the virus in the tissue, with maximal replication at 2 days postinfection. The virus replicated in ciliated cells and was released apically. IMPORTANCE Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged by the end of 2019 and rapidly spread in 2020. At present, it is of utmost importance to understand the biology of the virus, rapidly assess the treatment potential of existing drugs, and develop new active compounds. While some animal models for such studies are under development, most of the research is carried out in Vero E6 cells. Here, we propose fully differentiated human airway epithelium cultures as a model for studies on SARS-CoV-2.
    Type of Medium: Online Resource
    ISSN: 0022-538X , 1098-5514
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2020
    detail.hit.zdb_id: 1495529-5
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  • 2
    In: Journal of Electroanalytical Chemistry, Elsevier BV, Vol. 342, No. 1 ( 1992-2), p. 57-61
    Type of Medium: Online Resource
    ISSN: 1572-6657
    Language: English
    Publisher: Elsevier BV
    Publication Date: 1992
    detail.hit.zdb_id: 1491150-4
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  • 3
    Online Resource
    Online Resource
    MDPI AG ; 2021
    In:  International Journal of Molecular Sciences Vol. 22, No. 2 ( 2021-01-19), p. 960-
    In: International Journal of Molecular Sciences, MDPI AG, Vol. 22, No. 2 ( 2021-01-19), p. 960-
    Abstract: It is well known that living cells interact mechanically with their microenvironment. Many basic cell functions, like migration, proliferation, gene expression, and differentiation, are influenced by external forces exerted on the cell. That is why it is extremely important to study how mechanical properties of the culture substrate influence the cellular molecular regulatory pathways. Optical microscopy is one of the most common experimental method used to visualize and study cellular processes. Confocal microscopy allows to observe changes in the 3D organization of the cytoskeleton in response to a precise mechanical stimulus applied with, for example, a bead trapped with optical tweezers. Optical tweezers-based method (OT) is a microrheological technique which employs a focused laser beam and polystyrene or latex beads to study mechanical properties of biological systems. Latex beads, functionalized with a specific protein, can interact with proteins located on the surface of the cellular membrane. Such interaction can significantly affect the cell’s behavior. In this work, we demonstrate that beads alone, placed on the cell surface, significantly change the architecture of actin, microtubule, and intermediate filaments. We also show that the observed molecular response to such stimulus depends on the duration of the cell–bead interaction. Application of cytoskeletal drugs: cytochalasin D, jasplakinolide, and docetaxel, abrogates remodeling effects of the cytoskeleton. More important, when cells are plated on elastic substrates, which mimic the mechanical properties of physiological cellular environment, we observe formation of novel, “cup-like” structures formed by the microtubule cytoskeleton upon interaction with latex beads. These results provide new insights into the function of the microtubule cytoskeleton. Based on these results, we conclude that rigidity of the substrate significantly affects the cellular processes related to every component of the cytoskeleton, especially their architecture.
    Type of Medium: Online Resource
    ISSN: 1422-0067
    Language: English
    Publisher: MDPI AG
    Publication Date: 2021
    detail.hit.zdb_id: 2019364-6
    SSG: 12
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  • 4
    Online Resource
    Online Resource
    Walter de Gruyter GmbH ; 2020
    In:  Bio-Algorithms and Med-Systems Vol. 16, No. 3 ( 2020-09-21)
    In: Bio-Algorithms and Med-Systems, Walter de Gruyter GmbH, Vol. 16, No. 3 ( 2020-09-21)
    Abstract: Deconvolution microscopy is a very useful, software-based technique allowing to deblur microscopy images and increase both lateral and axial resolutions. It can be used along with many of fluorescence microscopy imaging techniques. By increasing axial resolution, it also enables three-dimensional imaging using a basic wide-field fluorescence microscope. Unfortunately, commercially available deconvolution software is expensive, while freely available programs have limited capabilities of a batch file processing. In this work we present BatchDeconvolution , a Fiji plugin that bridges two programs that we used subsequently in an image deconvolution pipeline: PSF Generator and DeconvolutionLab2 , both from Biomedical Imaging Group, EPFL. Our software provides a simple way to perform a batch processing of multiple microscopy files with minimal working time required from the user.
    Type of Medium: Online Resource
    ISSN: 1896-530X , 1895-9091
    Language: English
    Publisher: Walter de Gruyter GmbH
    Publication Date: 2020
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  • 5
    In: Scientific Reports, Springer Science and Business Media LLC, Vol. 9, No. 1 ( 2019-07-31)
    Abstract: Moderate intravascular hemolysis is a common condition in newborns. It is followed by the accumulation of bilirubin, which is a secondary product of the activity of heme oxygenase-1, an enzyme that catalyzes the breakdown of heme released from disrupted erythrocytes and taken up by hepatic macrophages. Although these cells are a major site of enzymatic heme breakdown in adults, we show here that epithelial cells of proximal tubules in the kidneys perform the functions of both heme uptake and catabolism in mouse neonates. A time-course study examining mouse pups during the neonatal period showed a gradual recovery from hemolysis, and concomitant decreases in the expression of heme-related genes and non-heme iron transporters in the proximal tubules. By adjusting the expression of iron-handling proteins in response to the disappearance of hemolysis in mouse neonates, the kidneys may play a role in the detoxification of iron and contribute to its recirculation from the primary urine to the blood.
    Type of Medium: Online Resource
    ISSN: 2045-2322
    Language: English
    Publisher: Springer Science and Business Media LLC
    Publication Date: 2019
    detail.hit.zdb_id: 2615211-3
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  • 6
    Online Resource
    Online Resource
    Elsevier BV ; 2023
    In:  MethodsX Vol. 10 ( 2023), p. 102107-
    In: MethodsX, Elsevier BV, Vol. 10 ( 2023), p. 102107-
    Type of Medium: Online Resource
    ISSN: 2215-0161
    Language: English
    Publisher: Elsevier BV
    Publication Date: 2023
    detail.hit.zdb_id: 2830212-6
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  • 7
    In: European Journal of Neuroscience, Wiley
    Abstract: Drug seeking is associated with the ventral tegmental area (VTA) dopaminergic (DA) activity. Previously, we have shown that brief optogenetic inhibition of VTA DA neurons with 1 s pulses delivered every 9 s attenuates cocaine seeking under extinction conditions in rats without producing overt signs of dysphoria or locomotor sedation. Whether recruitment of neuronal pathways inhibiting VTA neuronal activity would suppress drug seeking remains unknown. Here, we asked if optogenetic stimulation of the lateral habenula (LHb) efferents in the rostromedial tegmental nucleus (RMTg) as well as RMTg efferents in VTA would reduce drug seeking. To investigate this, we measured how recruitment of elements of this inhibitory pathway affects cocaine seeking in male rats under extinction conditions. The effectiveness of brief optogenetic manipulations was confirmed electrophysiologically at the level of electrical activity of VTA DA neurons. Real‐time conditioned place aversion (RT‐CPA) and open field tests were performed to control for potential dysphoric/sedating effects of brief optogenetic stimulation of LHb‐RMTg‐VTA circuitry. Optogenetic stimulation of either RMTg or LHb inhibited VTA DAergic neuron firing, whereas similar stimulation of RMTg efferents in VTA or LHb efferents in RMTg reduced cocaine seeking under extinction conditions. Moreover, stimulation of LHb‐RMTg efferents produced an effect that was maintained 24 h later, during cocaine seeking test without stimulation. This effect was specific, as brief optogenetic stimulation did not affect locomotor activity and was not aversive. Our results indicate that defined inhibitory pathways can be recruited to inhibit cocaine seeking, providing potential new targets for non‐pharmacological treatment of drug craving.
    Type of Medium: Online Resource
    ISSN: 0953-816X , 1460-9568
    Language: English
    Publisher: Wiley
    Publication Date: 2022
    detail.hit.zdb_id: 2005178-5
    SSG: 12
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  • 8
    Online Resource
    Online Resource
    American Society for Cell Biology (ASCB) ; 2004
    In:  Molecular Biology of the Cell Vol. 15, No. 7 ( 2004-07), p. 3497-3508
    In: Molecular Biology of the Cell, American Society for Cell Biology (ASCB), Vol. 15, No. 7 ( 2004-07), p. 3497-3508
    Abstract: To study the dynamics of stress fiber components in cultured fibroblasts, we expressed α-actinin and the myosin II regulatory myosin light chain (MLC) as fusion proteins with green fluorescent protein. Myosin activation was stimulated by treatment with calyculin A, a serine/threonine phosphatase inhibitor that elevates MLC phosphorylation, or with LPA, another agent that ultimately stimulates phosphorylation of MLC via a RhoA-mediated pathway. The resulting contraction caused stress fiber shortening and allowed observation of changes in the spacing of stress fiber components. We have observed that stress fibers, unlike muscle myofibrils, do not contract uniformly along their lengths. Although peripheral regions shortened, more central regions stretched. We detected higher levels of MLC and phosphorylated MLC in the peripheral region of stress fibers. Fluorescence recovery after photobleaching revealed more rapid exchange of myosin and α-actinin in the middle of stress fibers, compared with the periphery. Surprisingly, the widths of the myosin and α-actinin bands in stress fibers also varied in different regions. In the periphery, the banding patterns for both proteins were shorter, whereas in central regions, where stretching occurred, the bands were wider.
    Type of Medium: Online Resource
    ISSN: 1059-1524 , 1939-4586
    Language: English
    Publisher: American Society for Cell Biology (ASCB)
    Publication Date: 2004
    detail.hit.zdb_id: 1474922-1
    SSG: 12
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  • 9
    Online Resource
    Online Resource
    Springer Science and Business Media LLC ; 2018
    In:  Scientific Reports Vol. 8, No. 1 ( 2018-04-13)
    In: Scientific Reports, Springer Science and Business Media LLC, Vol. 8, No. 1 ( 2018-04-13)
    Abstract: APOBEC3 family members are cytidine deaminases with roles in intrinsic responses to infection by retroviruses and retrotransposons, and in the control of other DNA viruses, such as herpesviruses, parvoviruses and hepatitis B virus. Although effects of APOBEC3 members on viral DNA have been demonstrated, it is not known whether they edit RNA genomes through cytidine deamination. Here, we investigated APOBEC3-mediated restriction of Coronaviridae . In experiments in vitro , three human APOBEC3 proteins (A3C, A3F and A3H) inhibited HCoV-NL63 infection and limited production of progeny virus, but did not cause hypermutation of the coronaviral genome. APOBEC3-mediated restriction was partially dependent on enzyme activity, and was reduced by the use of enzymatically inactive APOBEC3. Moreover, APOBEC3 proteins bound to the coronaviral nucleoprotein, and this interaction also affected viral replication. Although the precise molecular mechanism of deaminase-dependent inhibition of coronavirus replication remains elusive, our results further our understanding of APOBEC-mediated restriction of RNA virus infections.
    Type of Medium: Online Resource
    ISSN: 2045-2322
    Language: English
    Publisher: Springer Science and Business Media LLC
    Publication Date: 2018
    detail.hit.zdb_id: 2615211-3
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  • 10
    In: Scientific Reports, Springer Science and Business Media LLC, Vol. 13, No. 1 ( 2023-07-07)
    Abstract: The examination of morphology and migration of cells plays substantial role in understanding the cellular behaviour, being described by plethora of quantitative parameters and models. These descriptions, however, treat cell migration and morphology as independent properties of temporal cell state, while not taking into account their strong interdependence in adherent cells. Here we present the new and simple mathematical parameter called signed morphomigrational angle ( sMM angle ) that links cell geometry with translocation of cell centroid, considering them as one morphomigrational behaviour. The sMM angle combined with pre-existing quantitative parameters enabled us to build a new tool called morphomigrational description , used to assign the numerical values to several cellular behaviours. Thus, the cellular activities that until now were characterized using verbal description or by complex mathematical models, are described here by a set of numbers. Our tool can be further used in automatic analysis of cell populations as well as in studies focused on cellular response to environmental directional signals.
    Type of Medium: Online Resource
    ISSN: 2045-2322
    Language: English
    Publisher: Springer Science and Business Media LLC
    Publication Date: 2023
    detail.hit.zdb_id: 2615211-3
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