| 1. |
- Klionsky, Daniel J., et al.
(author)
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Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
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In: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
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Research review (peer-reviewed)abstract
- In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.
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| 4. |
- Bouhafs, Chamseddine, et al.
(author)
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Decoupling and ordering of multilayer graphene on C-face 3C-SiC(111)
- 2016
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In: Applied Physics Letters. - : AMER INST PHYSICS. - 0003-6951 .- 1077-3118. ; 109:20
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Journal article (peer-reviewed)abstract
- We show experimentally that few layer graphene (FLG) grown on the carbon terminated surface (C-face) of 3C-SiC(111) is composed of decoupled graphene sheets. Landau level spectroscopy on FLG graphene is performed using the infrared optical Hall effect. We find that Landau level transitions in the FLG exhibit polarization preserving selection rules and the transition energies obey a square-root dependence on the magnetic field strength. These results show that FLG on C-face 3C-SiC(111) behave effectively as a single layer graphene with linearly dispersing bands (Dirac cones) at the graphene K point. We estimate from the Landau level spectroscopy an upper limit of the Fermi energy of about 60 meV in the FLG, which corresponds to a carrier density below 2.5 x 10(11) cm(-2). Low-energy electron diffraction mu-LEED) reveals the presence of azimuthally rotated graphene domains with a typical size of amp;lt;= 200 nm.mu-LEED mapping suggests that the azimuth rotation occurs between adjacent domains within the same sheet rather than vertically in the stack. Published by AIP Publishing.
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| 5. |
- Bouhafs, Chamseddine, et al.
(author)
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Multi-scale investigation of interface properties, stacking order and decoupling of few layer graphene on C-face 4H-SiC
- 2017
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In: Carbon. - : PERGAMON-ELSEVIER SCIENCE LTD. - 0008-6223 .- 1873-3891. ; 116, s. 722-732
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Journal article (peer-reviewed)abstract
- In this work, we report a multi-scale investigation using several nano-, micro and macro-scale techniques of few layer graphene (FLG) sample consisting of large monolayer (ML) and bilayer (BL) areas grown on C-face 4H-SiC (000-1) by high-temperature sublimation. Single 1 x 1 diffraction patterns are observed by micro-low-energy electron diffraction for ML, BL and trilayer graphene with no indication of out-of-plane rotational disorder. A SiOx layer is identified between graphene and SiC by X-ray photoelectron emission spectroscopy and reflectance measurements. The chemical composition of the interface layer changes towards SiO2 and its thickness increases with aging in normal ambient conditions. The formation mechanism of the interface layer is discussed. It is shown by torsion resonance conductive atomic force microscopy that the interface layer causes the formation of non-ideal Schottky contact between ML graphene and SiC. This is attributed to the presence of a large density of interface states. Mid-infrared optical Hall effect measurements revealed Landau-level transitions in FLG that have a square-root dependence on magnetic field, which evidences a stack of decoupled graphene sheets. Contrary to previous works on decoupled C-face graphene, our BL and FLG are composed of ordered decoupled graphene layers without out-of-plane rotation. (C) 2017 Elsevier Ltd. All rights reserved.
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| 6. |
- Hedlund, Petter, et al.
(author)
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Erectile dysfunction in cyclic CMP-dependent kinase I-deficient mice
- 2000
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In: Proceedings of the National Academy of Sciences of the United States of America. - : National Academy of Sciences; 1999. - 0027-8424 .- 1091-6490. ; 97:5, s. 2349-2354
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Journal article (peer-reviewed)abstract
- The generation of nitric oxide (NO) in penile erectile tissue and the subsequent elevation of cyclic GMP (cGMP) levels are important for normal penile erection. Current treatments of erectile dysfunction elevate either cGMP levels by blocking cGMP degrading phosphodiesterase 5 or cyclic AMP (cAMP) levels by intrapenile injection of prostaglandin E1. The molecular target or targets of cGMP in erectile tissue and the role of cAMP for normal penile erection are not known. Herein, we report that mice lacking cGMP-dependent kinase I (cGKI) have a very low ability to reproduce and that their corpora cavernosa fail to relax on activation of the NO/cGMP signaling cascade. Elevation of cAMP by forskolin, however, induces similar relaxation in normal and cGKI-null corpus cavernosum. In addition, sperm derived from cGKI-null mice is normal, can undergo acrosomal reactions, and can efficiently fertilize eggs. Altogether, these data identify cGKI as the downstream target of cGMP in erectile tissue and provide evidence that cAMP signaling cannot compensate for the absence of the cGMP/cGKI signaling cascade in vivo.Penile erection is produced by an increased blood flow to the corpus cavernosum (CC), made possible by opening of penile resistance vessels (helicine arteries), relaxation of the CC cells, and occlusion of the venous outflow (1). The erectile response in several animal models depends on nitric oxide (NO), produced by nerves as well as vascular endothelium (1, 2–4). NO activates soluble guanylate cyclase, which leads to the production of cyclic GMP (cGMP). cGMP signals via three different receptors in eukaryotic cells, including ion channels, phosphodiesterases, and protein kinases (5). At present, however, the molecular targets that are activated by cGMP and finally execute the relaxation of penile smooth muscle are not known. In addition, two different cGMP-dependent protein kinases (cGKI and cGKII) have been identified in mammals (6, 7). cGKII is expressed in the small intestine, brain, and cartilage (8–10), whereas high levels of cGKI are found in vascular and intestinal smooth muscle, platelets, Purkinje cells of the cerebellum, and CC cells (11, 12). Inactivation of cGKI in mice abolished both NO/cGMP-dependent relaxation of vascular and intestinal smooth muscle and inhibition of platelet aggregation, causing hypertension, intestinal dysmotility, and abnormal hemostasis (13). In the present study, we investigated the function of cGKI in erectile tissue and the capability of cGKI-deficient mice to reproduce. Furthermore, we analyzed whether a cross-talk exists between the cGMP and cyclic AMP (cAMP) signaling cascades in smooth muscle (5, 14), i.e., whether cAMP can cause relaxation via cGKI.
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| 7. |
- Peev, D., et al.
(author)
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Anisotropic contrast optical microscope
- 2016
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In: Review of Scientific Instruments. - : AMER INST PHYSICS. - 0034-6748 .- 1089-7623. ; 87:11
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Journal article (peer-reviewed)abstract
- An optical microscope is described that reveals contrast in the Mueller matrix images of a thin, transparent, or semi-transparent specimen located within an anisotropic object plane (anisotropic filter). The specimen changes the anisotropy of the filter and thereby produces contrast within the Mueller matrix images. Here we use an anisotropic filter composed of a semi-transparent, nanostructured thin film with sub-wavelength thickness placed within the object plane. The sample is illuminated as in common optical microscopy but the light is modulated in its polarization using combinations of linear polarizers and phase plate (compensator) to control and analyze the state of polarization. Direct generalized ellipsometry data analysis approaches permit extraction of fundamental Mueller matrix object plane images dispensing with the need of Fourier expansion methods. Generalized ellipsometry model approaches are used for quantitative image analyses. These images are obtained from sets of multiple images obtained under various polarizer, analyzer, and compensator settings. Up to 16 independent Mueller matrix images can be obtained, while our current setup is limited to 11 images normalized by the unpolarized intensity. We demonstrate the anisotropic contrast optical microscope by measuring lithographically defined micro-patterned anisotropic filters, and we quantify the adsorption of an organic self-assembled monolayer film onto the anisotropic filter. Comparison with an isotropic glass slide demonstrates the image enhancement obtained by our method over microscopy without the use of an anisotropic filter. In our current instrument, we estimate the limit of detection for organic volumetric mass within the object plane of approximate to 49 fg within approximate to 7 x 7 mu m(2) object surface area. Compared to a quartz crystal microbalance with dissipation instrumentation, where contemporary limits require a total load of approximate to 500 pg for detection, the instrumentation demonstrated here improves sensitivity to a total mass required for detection by 4 orders of magnitude. We detail the design and operation principles of the anisotropic contrast optical microscope, and we present further applications to the detection of nanoparticles, to novel approaches for imaging chromatography and to new contrast modalities for observations on living cells. Published by AIP Publishing.
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| 8. |
- Ravasi, Timothy, et al.
(author)
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An Atlas of Combinatorial Transcriptional Regulation in Mouse and Man
- 2010
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In: CELL. - : Elsevier BV. - 0092-8674 .- 1097-4172. ; 140:5, s. 744-752
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Journal article (peer-reviewed)abstract
- Combinatorial interactions among transcription factors are critical to directing tissue-specific gene expression. To build a global atlas of these combinations, we have screened for physical interactions among the majority of human and mouse DNA-binding transcription factors (TFs). The complete networks contain 762 human and 877 mouse interactions. Analysis of the networks reveals that highly connected TFs are broadly expressed across tissues, and that roughly half of the measured interactions are conserved between mouse and human. The data highlight the importance of TF combinations for determining cell fate, and they lead to the identification of a SMAD3/FLI1 complex expressed during development of immunity. The availability of large TF combinatorial networks in both human and mouse will provide many opportunities to study gene regulation, tissue differentiation, and mammalian evolution.
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| 9. |
- Wittchen, Hans-Ulrich, et al.
(author)
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The need for a behavioural science focus in research on mental health and mental disorders
- 2014
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In: International Journal of Methods in Psychiatric Research. - : Wiley-Blackwell. - 1049-8931 .- 1557-0657. ; 23, s. 28-40
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Journal article (peer-reviewed)abstract
- Psychology as a science offers an enormous diversity of theories, principles, and methodological approaches to understand mental health, abnormal functions and behaviours and mental disorders. A selected overview of the scope, current topics as well as strength and gaps in Psychological Science may help to depict the advances needed to inform future research agendas specifically on mental health and mental disorders. From an integrative psychological perspective, most maladaptive health behaviours and mental disorders can be conceptualized as the result of developmental dysfunctions of psychological functions and processes as well as neurobiological and genetic processes that interact with the environment. The paper presents and discusses an integrative translational model, linking basic and experimental research with clinical research as well as population-based prospective-longitudinal studies. This model provides a conceptual framework to identify how individual vulnerabilities interact with environment over time, and promote critical behaviours that might act as proximal risk factors for ill-health and mental disorders. Within the models framework, such improved knowledge is also expected to better delineate targeted preventive and therapeutic interventions that prevent further escalation in early stages before the full disorder and further complications thereof develop. In contrast to conventional personalized medicine that typically targets individual (genetic) variation of patients who already have developed a disease to improve medical treatment, the proposed framework model, linked to a concerted funding programme of the Science of Behaviour Change, carries the promise of improved diagnosis, treatment and prevention of health-risk behaviour constellations as well as mental disorders.
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| 10. |
- Boosalis, A., et al.
(author)
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Visible to vacuum ultraviolet dielectric functions of epitaxial graphene on 3C and 4H SiC polytypes determined by spectroscopic ellipsometry
- 2012
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In: Applied Physics Letters. - : American Institute of Physics (AIP). - 0003-6951 .- 1077-3118. ; 101:1
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Journal article (peer-reviewed)abstract
- Spectroscopic ellipsometry measurements in the visible to vacuum-ultraviolet spectra (3.5-9.5 eV) are performed to determine the dielectric function of epitaxial graphene on SiC polytypes, including 4H (C-face and Si-face) and 3C SiC (Si-face). The model dielectric function of graphene is composed of two harmonic oscillators and allows the determination of graphene quality, morphology, and strain. A characteristic van Hove singularity at 4.5 eV is present in the dielectric function of all samples, in agreement with observations on exfoliated as well as chemical vapor deposited graphene in the visible range. Model dielectric function analysis suggests that none of our graphene layers experience a significant degree of strain. Graphene grown on the Si-face of 4H SiC exhibits a dielectric function most similar to theoretical predictions for graphene. The carbon buffer layer common for graphene on Si-faces is found to increase polarizability of graphene in the investigated spectrum.
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