2,758 research outputs found
LV-pIN-KDEL: a novel lentiviral vector demonstrates the morphology, dynamics and continuity of the endoplasmic reticulum in live neurones.
No full textBACKGROUND
The neuronal endoplasmic reticulum (ER) is an extensive, complex endomembrane system, containing Ca2+ pumps, and Ca2+ channels that permit it to act as a dynamic calcium store. Currently, there is controversy over the continuity of the ER in neurones, how this intersects with calcium signalling and the possibility of physical compartmentalisation. Unfortunately, available probes of ER structure such as vital dyes are limited by their membrane specificity. The introduction of ER-targeted GFP plasmids has been a considerable step forward, but these are difficult to express in neurones through conventional transfection approaches. To circumvent such problems we have engineered a novel ER-targeted GFP construct, termed pIN-KDEL, into a 3rd generation replication-defective, self-inactivating lentiviral vector system capable of mediating gene transduction in diverse dividing and post-mitotic mammalian cells, including neurones.
RESULTS
Following its expression in HEK293 (or COS-7) cells, LV-pIN-KDEL yielded a pattern of fluorescence that co-localised exclusively with the ER marker sec61beta but with no other major organelle. We found no evidence for cytotoxicity and only rarely inclusion body formation. To explore the utility of the probe in resolving the ER in live cells, HEK293 or COS-7 cells were transduced with LV-pIN-KDEL and, after 48 h, imaged directly at intervals from 1 min to several hours. LV-pIN-KDEL fluorescence revealed the endoplasmic reticulum as a tubular lattice structure whose morphology can change markedly within seconds. Although GFP can be phototoxic, the integrity of the cells and ER was retained for several weeks and even after light exposure for periods up to 24 h. Using LV-pIN-KDEL we have imaged the ER in diverse fixed neuronal cultures and, using real-time imaging, found evidence for extensive, dynamic remodelling of the neuronal ER in live hippocampal cultures, brain slices, explants and glia. Finally, through a Fluorescence Loss in Photobleaching (FLIP) approach, continuous irradiation at a single region of interest removed all the fluorescence of LV-pIN-KDEL-transduced nerve cells in explant cultures, thus, providing compelling evidence that in neurons the endoplasmic reticulum is not only dynamic but also continuous.
CONCLUSION
The lentiviral-based ER-targeted reporter, LV-pIN-KDEL, offers considerable advantages over present systems for defining the architecture of the ER, especially in primary cells such as neurones that are notoriously difficult to transfect. Images and continuous photobleaching experiments of LV-pIN-KDEL-transduced neurones demonstrate that the endoplasmic reticulum is a dynamic structure with a single continuous lumen. The introduction of LV-pIN-KDEL is anticipated to greatly facilitate a real-time visualisation of the structural plasticity and continuous nature of the neuronal ER in healthy and diseased brain tissue
An experimental investigation into pin loading effects on fatigue crack growth in Fibre Metal Laminates
No full textAbstractThis paper provides an experimental investigation into the pin loading effects on the crack growth behaviour in Fibre Metal Laminates. The pin loading effects and bypass loading effects are incorporated in two different tested joints. The analysis of the test results shows that pin loading dominates the crack growth only in the vicinity of the pin hole and the superposition method for analysing stress intensity factor in FMLs with pin loading effects can be applied
The Legal Treatment of Muslim Minorities in Italy. Islam and the Neutral State
No full textslam is a growing presence practically everywhere in Europe. In Italy, however, Islam has met a unique model of state neutrality, religious freedom and church and state collaboration. This book gives a detailed description of the legal treatment of Muslims in Italy, contrasting it with other European states and jurisprudence, and with wider global tendencies that characterize the treatment of Islam. Through focusing on a series of case studies, the author argues that the relationship between church and state in Italy, and more broadly in Europe, should be reconsidered both to secure religious freedom and general welfare.
Working on the concepts of religious freedom, state neutrality, and relationship between church and state, Andrea Pin develops a theoretical framework that combines the state level with the supranational level in the form of the European Convention of Human Rights, which ultimately shapes a unitary but flexible understanding of pluralism. This approach should better accommodate not just Muslims’ needs, but religious needs in general in Italy and elsewhere
Design of pin connections between steel members
No full textThe design of pin connections between steel members has been established for many years in design codes. However, recently, in the scope of the revision of Eurocode 3, Part 1–8 (EN 1993–1-8), questions were raised concerning the safety of the corresponding design verifications. This paper identifies two main aspects that require revision, namely: (i) the possibility to design a pin as a bolt in shear and (ii) the verification of the resistance of the pin itself. Based on a thorough literature review, experimental tests and a parametric study, a new proposal submitted to CEN as an amendment to the code, is presented to solve these two identified issues.Green Open Access added to TU Delft Institutional Repository 'You share, we take care!' - Taverne project https://www.openaccess.nl/en/you-share-we-take-care Otherwise as indicated in the copyright section: the publisher is the copyright holder of this work and the author uses the Dutch legislation to make this work public.Steel & Composite Structure
Real Exchange Rate, Foreign Trade and Employment: Evidence from China
No full textCoordination of macro-economic development and employment is an essential issue for China's social development, which largely depends on economic expansion, as well as integration into the global market to create jobs. Through the literature review and empirical test, this paper analyses the relationship between macro-economic policy and employment, and discusses the impact of real exchange rate and foreign trade on employment. The research indicates that a stable and competitive exchange rate policy plays an indispensable role in employment promotion, more effective than monetary and fiscal policies, while the export growth also plays a positive role in employment promotion.China, employment, foreign trade, real exchange rate
Pin-pin resonance as a reference in determining ballasted railway track vibration behaviour
No full textPin-pin resonance is one of the most significant preferred vibration modes of beams, which are supported at equal distances, such as rails at sleepers in railway track structures do. Pin-pin resonance is a vibration that appears in one basic (first) mode and several higher modes, however the basic mode will have the highest amplitudes. In operational conditions of railways, pin-pin resonance only partly influences wheel-rail contact of the train while the speed dependent sleeper-passing frequency is more important. Among other track resonances, pin-pin resonance plays an important role in noise and vibration radiation of the rails and can be used as a meaningful instrument in track system dynamics recognition and optimization. However, existing simple analytic approximations are not sufficiently reliable to perform these recognitions."This will be shown in this paper by means of new tests and simulations that are based on improved models and methods but restricted to ballasted track structures.Railway EngineeringCivil Engineering and Geoscience
Quantum Pin Codes
No full textarXiv: 1906.11394We introduce quantum pin codes: a class of quantum CSS codes. Quantum pin codes are a vast generalization of quantum color codes and Reed-Muller codes. A lot of the structure and properties of color codes carries over to pin codes. Pin codes have gauge operators, an unfolding procedure and their stabilizers form multi-orthogonal spaces. This last feature makes them interesting for devising magic-state distillation protocols. We study examples of these codes and their properties
Effect of Laser Annealing on amorphous Silicon Carbide Films Prepared by PECVD
No full textIn order to improve the performance of PECVD SiC in MEMS, pulse excimer laser annealing process was introduced in this paper. The effect was comprehensively investigated in several respects, including morphology, mechanical properties and residual stress elimination. The study revealed that: after laser annealing with adequate energy density, amorphous SiC (a-SiC) transformed into polycrystalline SiC, and larger crystal grains were formed with higher energy density; the process effectively enhanced the films' mechanical property, a moderate energy density about 90mJ/cm2 resulted in comparatively greatest Young's modulus and hardness; the stress indicating structure exhibited that the annealing eliminated the compressive stress of films as deposited effectively. Therefore, pulse excimer laser annealing process is a promising method to improve the performance of a-SiC films prepared by PECVD, and makes the material more applicable in many MEMS fields.Engineering, Electrical & ElectronicNanoscience & NanotechnologyEICPCI-S(ISTP)
Numerical investigation of bubble dynamics and flow boiling heat transfer in cylindrical micro-pin-fin heat exchangers
No full textMicro-pin-fin evaporators are a promising alternative to multi-microchannel heat sinks for two-phase cooling of high power-density devices. Within pin-fin evaporators, the refrigerant flows through arrays of obstacles in cross-flow and is not restricted by the walls of a channel. The dynamics of bubbles generated upon flow boiling and the associated heat transfer mechanisms are expected to be substantially different from those pertinent to microchannels; however, the fundamental aspects of two-phase flows evolving through micro-pin-fin arrays are still little understood. This article presents a systematic analysis of flow boiling within a micro-pin-fin evaporator, encompassing bubble, thin-film dynamics and heat transfer. The flow is studied by means of numerical simulations, performed using a customised boiling solver in OpenFOAM v2106, which adopts the built-in geometric Volume of Fluid method to capture the liquid–vapour interface dynamics. The numerical model of the evaporator includes in-line arrays of pin-fins of diameter of 50μm and height of 100μm, streamwise pitch of 91.7μm and cross-stream pitch of 150μm. The fluid utilised is refrigerant R236fa at a saturation temperature of 30 °C. The range of operating conditions simulated includes values of mass flux G=500–2000kg/(m2s), heat flux q=200kW/m2, and inlet subcooling ΔTsub=0–5K. This study shows that bubbles nucleated in a pin-fin evaporator tend to travel along the channels formed in between the pin-fin lines. Bubbles grow due to liquid evaporation and elongate in the direction of the flow, leaving thin liquid films that partially cover the pin-fins surface. The main contributions to heat transfer arise from the evaporation of this thin liquid film and from a cross-stream convective motion induced by the bubbles in the gap between the cylinders, which displace the hot fluid otherwise stagnant in the cylinders wakes. When the mass flow rate is increased, bubbles depart earlier from the nucleation sites and grow more slowly, which results in a reduction of the two-phase heat transfer. Higher inlet subcooling yields lower two-phase heat transfer coefficients because condensation becomes important when bubbles depart from the hot pin-fin surfaces and reach highly subcooled regions, thus reducing the two-phase heat transfer
Pin/hole joint construction and laminate therefor
No full textA pin/hole joint construction in a laminate (1) comprises layers (9) of fibres and resin bonded to one another. At least one hole (2) extends transversely completely through the layers in the laminate, in which hole a pin (3) is accommodated that bears against the inner surface of the hole for transferring forces between the laminate and the pin. In the laminate a first region (13) is defined in which the layers are directly on top of one another and a second region (14) in which at least one reinforcing layer (5) is provided that is bonded to the adjacent layers of fibres and resin, in which second region the at least one hole is located. All layers of fibres and resin are continued into the second region.Aerospace Engineerin
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