Fluorescence Correlation Spectroscopy Reveals Efficient Cytosolic Delivery of Protein Cargo by Cell-Permeant Miniature Proteins.

New methods for delivering proteins into the cytosol of mammalian cells are being reported at a rapid pace. Differentiating between these methods in a quantitative manner is difficult, however, as most assays for evaluating cytosolic protein delivery are qualitative and indirect and thus often misleading. Here we make use of fluorescence correlation spectroscopy (FCS) to determine with precision and accuracy the relative efficiencies with which seven different previously reported "cell-penetrating peptides" (CPPs) transport a model protein cargo-the self-labeling enzyme SNAP-tag-beyond endosomal membranes and into the cytosol. Using FCS, we discovered that the miniature protein ZF5.3 is an exceptional vehicle for delivering SNAP-tag to the cytosol. When delivered by ZF5.3, SNAP-tag can achieve a cytosolic concentration as high as 250 nM, generally at least 2-fold and as much as 6-fold higher than any other CPP evaluated. Additionally, we show that ZF5.3 can be fused to a second enzyme cargo-the engineered peroxidase APEX2-and reliably delivers the active enzyme to the cell interior. As FCS allows one to realistically assess the relative merits of protein transduction domains, we anticipate that it will greatly accelerate the identification, evaluation, and optimization of strategies to deliver large, intact proteins to intracellular locales

RentWise: An Updated Tool for Meeting the Needs of Renters

Housing education is critical in the context of today’s economy. This article details the current need for rental education for both consumers and practitioners. We discuss the importance of housing education resources for Extension educators across the United States, describe the updated curriculum RentWise, and provide information related to accessing the curriculum

Controlling aggregation of cholesterol-modified DNA nanostructures.

DNA nanotechnology allows for the design of programmable DNA-built nanodevices which controllably interact with biological membranes and even mimic the function of natural membrane proteins. Hydrophobic modifications, covalently linked to the DNA, are essential for targeted interfacing of DNA nanostructures with lipid membranes. However, these hydrophobic tags typically induce undesired aggregation eliminating structural control, the primary advantage of DNA nanotechnology. Here, we study the aggregation of cholesterol-modified DNA nanostructures using a combined approach of non-denaturing polyacrylamide gel electrophoresis, dynamic light scattering, confocal microscopy and atomistic molecular dynamics simulations. We show that the aggregation of cholesterol-tagged ssDNA is sequence-dependent, while for assembled DNA constructs, the number and position of the cholesterol tags are the dominating factors. Molecular dynamics simulations of cholesterol-modified ssDNA reveal that the nucleotides wrap around the hydrophobic moiety, shielding it from the environment. Utilizing this behavior, we demonstrate experimentally that the aggregation of cholesterol-modified DNA nanostructures can be controlled by the length of ssDNA overhangs positioned adjacent to the cholesterol. Our easy-to-implement method for tuning cholesterol-mediated aggregation allows for increased control and a closer structure-function relationship of membrane-interfacing DNA constructs - a fundamental prerequisite for employing DNA nanodevices in research and biomedicine.This work was supported by: European Research Council (ERC) consolidator grant [DesignerPores 647144 to U.F.K.]. Winton Programme for the Physics of Sustainability; Gates Cambridge; Oppenheimer Ph.D. studentship; and the European Union's Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement No 792270 [to K.G.]. Engineering and Physical Sciences Research Council (EPSRC); and the Cambridge Trust Vice Chancellor’s Award [to A.O.]. National Science Foundation (USA) [DMR-1827346]; National Institutes of Health [P41-GM104601]; the supercomputer time provided through XSEDE allocation grant [MCA05S028]; and the Blue Waters petascale supercomputer system (UIUC) [to A.A. and H.J.]. Winton Programme for the Physics of Sustainability; and Engineering and Physical Sciences Research Council (EPSRC) [to D.S.]. The FEI Tecnai G2-Spirit was funded by the Wellcome Trust [090932/Z/09/Z]. Funding for open access charge: ERC [DesignerPores 647144]

Coercive Nuclear Campaigns in the 21st Century: Understanding Adversary Incentives and Options for Nuclear Escalation

FY 2012-2013. Project Leads: Lieber, Keir A. and Press, Daryl G.The project leads will examine why and how regional powers armed with nuclear weapons may employ them coercively during a conventional war. More specifically, a set of research questions will provide insight on the paths of nuclear escalation, weak state strategies of coercive escalation, and the type of targets regional powers may strike to pursue escalatory strategies.NAApproved for public release; distribution is unlimited

MRI T2 and T1ρ relaxation in patients at risk for knee osteoarthritis: A systematic review and meta-analysis

© 2019 The Author(s). Background: Magnetic resonance imaging (MRI) T2 and T1ρ relaxation are increasingly being proposed as imaging biomarkers potentially capable of detecting biochemical changes in articular cartilage before structural changes are evident. We aimed to: 1) summarize MRI methods of published studies investigating T2 and T1ρ relaxation time in participants at risk for but without radiographic knee OA; and 2) compare T2 and T1ρ relaxation between participants at-risk for knee OA and healthy controls. Methods: We conducted a systematic review of studies reporting T2 and T1ρ relaxation data that included both participants at risk for knee OA and healthy controls. Participant characteristics, MRI methodology, and T1ρ and T2 relaxation data were extracted. Standardized mean differences (SMDs) were calculated within each study. Pooled effect sizes were then calculated for six commonly segmented knee compartments. Results: 55 articles met eligibility criteria. There was considerable variability between scanners, coils, software, scanning protocols, pulse sequences, and post-processing. Moderate risk of bias due to lack of blinding was common. Pooled effect sizes indicated participants at risk for knee OA had lengthened T2 relaxation time in all compartments (SMDs from 0.33 to 0.74; p \u3c 0.01) and lengthened T1ρ relaxation time in the femoral compartments (SMD from 0.35 to 0.40; p \u3c 0.001). Conclusions: T2 and T1ρ relaxation distinguish participants at risk for knee OA from healthy controls. Greater standardization of MRI methods is both warranted and required for progress towards biomarker validation

Travelling for Umrah:destination attributes, destination image, and post-travel intentions

This paper examines the links between cosmopolitanism, self-identity, and a desire for social interaction perceived destination image and behavioural intentions. A model tested using a sample of 538 Iranian visitors to Mecca for the purpose of Umrah. The result from the structural model suggests that destination attributes influence perceived destination image. Further, such tourists are likely to revisit or recommend Islamic destinations if their experience matches their perceived image of the destination. This implies that, while the religious characteristics of the destination remain important, destination managers cannot disregard the tangential, non-religious attributes of a destination which are crucial in order to satisfy more conventional tourist desires. As such, this study suggests that those managing religious travel destinations should endeavour to foster a welcoming image, where experience, interaction and tolerance are at the forefront of the destination’s offering

Effectiveness of Stroke Early Supported Discharge: Analysis From a National Stroke Registry

BackgroundImplementation of stroke early supported discharge (ESD) services has been recommended in many countries’ clinical guidelines, based on clinical trial evidence. This is the first observational study to investigate the effectiveness of ESD service models operating in real-world conditions, at scale.Methods AND RESULTSUsing historical prospective data from the United Kingdom Sentinel Stroke National Audit Programme (January 1, 2016–December 31, 2016), measures of ESD effectiveness were “days to ESD” (number of days from hospital discharge to first ESD contact; n=6222), “rehabilitation intensity” (total number of treatment days/total days with ESD; n=5891), and stroke survivor outcome (modified Rankin scale at ESD discharge; n=6222). ESD service models (derived from Sentinel Stroke National Audit Programme postacute organizational audit data) were categorized with a 17-item score, reflecting adoption of ESD consensus core components (evidence-based criteria). Multilevel modeling analysis was undertaken as patients were clustered within ESD teams across the Midlands, East, and North of England (n=31). A variety of ESD service models had been adopted, as reflected by variability in the ESD consensus score. Controlling for patient characteristics and Sentinel Stroke National Audit Programme hospital score, a 1-unit increase in ESD consensus score was significantly associated with a more responsive ESD service (reduced odds of patient being seen after ≥1 day of 29% [95% CI, 1%–49%] and increased treatment intensity by 2% [95% CI, 0.3%–4%]). There was no association with stroke survivor outcome measured by the modified Rankin Scale.ConclusionsThis study has shown that adopting defined core components of ESD is associated with providing a more responsive and intensive ESD service. This shows that adherence to evidence-based criteria is likely to result in a more effective ESD service as defined by process measures

Effectiveness of stroke early supported discharge

Background Implementation of stroke early supported discharge (ESD) services has been recommended in many countries’ clinical guidelines, based on clinical trial evidence. This is the first observational study to investigate the effectiveness of ESD service models operating in real-world conditions, at scale. Methods AND RESULTS Using historical prospective data from the United Kingdom Sentinel Stroke National Audit Programme (January 1, 2016–December 31, 2016), measures of ESD effectiveness were “days to ESD” (number of days from hospital discharge to first ESD contact; n=6222), “rehabilitation intensity” (total number of treatment days/total days with ESD; n=5891), and stroke survivor outcome (modified Rankin scale at ESD discharge; n=6222). ESD service models (derived from Sentinel Stroke National Audit Programme postacute organizational audit data) were categorized with a 17-item score, reflecting adoption of ESD consensus core components (evidence-based criteria). Multilevel modeling analysis was undertaken as patients were clustered within ESD teams across the Midlands, East, and North of England (n=31). A variety of ESD service models had been adopted, as reflected by variability in the ESD consensus score. Controlling for patient characteristics and Sentinel Stroke National Audit Programme hospital score, a 1-unit increase in ESD consensus score was significantly associated with a more responsive ESD service (reduced odds of patient being seen after ≥1 day of 29% [95% CI, 1%–49%] and increased treatment intensity by 2% [95% CI, 0.3%–4%]). There was no association with stroke survivor outcome measured by the modified Rankin Scale. Conclusions This study has shown that adopting defined core components of ESD is associated with providing a more responsive and intensive ESD service. This shows that adherence to evidence-based criteria is likely to result in a more effective ESD service as defined by process measures. REGISTRATION: URL: http://www.isrctn.com/ ; Unique identifier: ISRCTN15568163

Incorporation by coordination and release of the iron chelator drug deferiprone from zinc-based metal–organic frameworks

A series of new zinc-based metal–organic framework materials has been prepared in which deferiprone is incorporated as a chelating ligand on infinite or tri-zinc secondary building units following deprotonation. Deferiprone is immediately released from the MOFs on treatments with 1 N hydrochloric acid or buffer, but slow release is observed in ethanoic acid

Modifying Polyester Surfaces with Incompatible Polymer Additives

Surface modification of amorphous PET in incompatible blends is demonstrated using fluorocarbon end-functional polystyrenes. Contact angles with water and decane were consistent with high levels of surface fluorocarbon, even for spin-cast films with no further processing required. Hydrophobicity and lipophobicity were further increased by annealing above the glass transition temperature. High resolution depth profiling using complementary ion beam analysis and specular neutron reflectometry has enabled accurate characterisation of the composition profile of the additive including the minimum in additive concentration found just below the surface enriched layer. This analysis quantified the very low compatibility between the modifying polymer and the amorphous PET and was consistent with the highly segregated nature of the adsorbing species and its sharp interface with the subphase. For these incompatible polymer blends, surfaces enriched with the surface active polymer could coexist at equilibrium with extremely low (∼0.4%) bulk loadings of the additive. This suggests that for thicker films at even lower additive concentrations than the minimum 1% that we studied, it may be possible to achieve efficient surface modification. However, at this concentration, the efficiency of surface modification is limited by the processing conditions. Finally we note that in higher loadings of surface active additive there is clear evidence for lateral phase separation into patterned domains of differing composition. The enhancement in surface properties is due to local reorganisation rather than bulk redistribution of the components within the film, as the composition versus depth distributions of the polymer blend components was observed to be relatively unaffected by annealing