Anti-RSV Peptide-Loaded Liposomes for the Inhibition of Respiratory Syncytial Virus

Although respiratory syncytial virus (RSV) is one of the leading causes of acute respiratory tract infection in infants and adults, effective treatment options remain limited. To circumvent this issue, there is a novel approach, namely, the development of multifunctional liposomes for the delivery of anti RSV-peptides. While most of the peptides that are used for loading with the particulate delivery systems are the penetrating peptides, an alternative approach is the development of liposome-peptide systems, which are loaded with an RSV fusion peptide (RF-482), which has been designed to inhibit the RSV fusion and block infection. The results of this work have revealed that the liposomes themselves can serve as potential RSV inhibitors, whilst the anti-RSV-peptide with liposomes can significantly increase the RSV inhibition when compared with the anti-RSV peptide alone

Design and development of novel screen-printed microelectrode and microbiosensor arrays fabricated using ultrafast pulsed laser ablation

© 2016 Elsevier B.V. All rights reserved. A new generic platform for the development of microbiosensors combining screen-printing and ultrafast pulsed laser technologies has been developed, characterised and evaluated. This new platform consists of a layer of screen-printed carbon ink containing the enzyme and mediator, covered with an insulating layer formed from a dielectric screen printed ink. Microholes were drilled through the insulated layer by ultrafast pulsed laser ablation to generate the microbiosensor array. The geometry of the microelectrode array was evaluated by optical microscopy, white light surface profiling and scanning electron microscopy. The electrochemical behaviour of the microelectrode array was characterised by cyclic voltammetry and compared with macroelectrodes. The analytical performance of the microbiosensor array was evaluated with external counter and reference electrodes for hydrogen peroxide and glucose determination showing linearity up to 4 mmol L-1 and 20 mmol L-1 (360 mg dL-1) respectively. The full screen printed three-electrode configuration shows linearity for glucose determination up to 20 mmol L-1 (360 mg dL-1). This study provides a new fabrication method for microelectrode and microbiosensor arrays capable for the first time to retain the activity of the enzymatic system after processing by pulse laser ablation

High potency of lipid conjugated TLR7 agonist requires nanoparticulate or liposomal formulation

Conjugation of small molecule agonists of Toll-like receptor 7 (TLR7) to proteins, lipids, or polymers is known to modulate potency, and the physical form or formulation of these conjugates is likely to have a major effect on their immunostimulatory activity. Here, we studied the effect of formulation on potency of a 1,2-di-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE) conjugated TLR7 agonist (DOPE-TLR7a) alongside assessing physical form using Dynamic Light Scattering (DLS), Nanosight Particle Tracking (NTA) analysis and Small Angle X-ray Scattering (SAXS). A very high potency of DOPE-TLR7a conjugate (EC50 around 9 nM) was observed either when prepared by direct dilution from DMSO or when formulated into 400-700 nm large multilamella liposomes containing dimethyldioctadecylammonium bromide salt (DDA) and DOPE. When prepared by dissolution in DMSO followed by dilution in aqueous culture medium, 93 ± 5 nm nanoparticles were formed. Without dilution from solution in DMSO, no nanoparticles were observed, and no immunostimulatory activity could be detected without this formulation step. SAXS analysis of the conjugate after DMSO dissolution/water dilution revealed a lamellar order with a layer spacing of 68.7 Å, which correlates with arrangement in groups of 3 bilayers. The addition of another immunostimulatory glycolipid, trehalose-6,6-dibehenate (TDB), to DOPE:DDA liposomes gave no further increase in immunostimulatory activity beyond that provided by incorporating DOPE-TLR7a. Given the importance of nanoparticle or liposomal formulation for activity, we conclude a major mechanism for increase in potency when TLR7 agonists are conjugated to macromolecules is through alteration of physical form

Proposed atomic cascade experimental test of symmetric local hidden-variables theories

The proposal is made of replacing the usual auxiliary assumptions needed for the derivation of testable Bell inequalities by an assumption resting only on symmetry considerations. It is shown that reliable atomic cascade tests of symmetric local hidden-variables theories should combine high statistics, cascades of type 0→1→0, and calcite polarizers. The three conditions have been achieved in experiments already performed, but never before simultaneously

Stochastic optics: A local realistic analysis of optical tests of Bell inequalities

Stochastic optics may be considered as simply a local realistic interpretation of quantum optics and, in this sense, it is a first step in the reinterpretation of the whole of quantum theory. However, as it is not possible to interpret all the details of quantum theory in a local realistic manner, as shown by Bell’s theorem, minor changes are introduced in the formalism with the consequence that the new theory makes different predictions in some special cases. In stochastic optics, the quantum-operator formalism is simply considered a formal way of dealing with stochastic fields. In particular, the quantum zero point is taken as a real random electromagnetic radiation filling the whole of space. This radiation noise has the same nature as light signals, the only difference being the greater intensity of the latter. We assume that photon detectors have an intensity threshold just above the level of the noise, thus detecting only signals. Transmission of radiation through polarizers follows Malus’s law, but the interplay of signal and noise leads quite naturally to the prediction that the detection probability of some signals is enhanced, which is known to be a necessary condition for the violation of the empirically tested Bell inequalities. In our view, correlated photon pairs are pairs of light signals supercorrelated in polarization, in the sense that, as well as the signal, the accompanying noise is also correlated. Thus stochastic optics allows predictions for the empirical correlations very close, but not identical, to the quantum ones. The theory is applied to the analysis of all experiments designed to test the Bell inequalities by measuring polarization correlations of photon pairs. The predictions agree with quantum optics and experiments within statistical errors, except for the Holt-Pipkin experiment. In this case, the experimental results agree with stochastic optical predictions within two standard deviations while violating quantum optics by four

Development of a rapid in vitro pre-screen for distinguishing effective liposome-adjuvant delivery systems

Abstract: Liposomes are a strong supporting tool in vaccine technology, as they are a versatile system that not only act as antigen delivery systems but also adjuvants that can be highly effective at stimulating both innate and adaptive immune responses. Their ability to induce cell-mediated immunity makes their use in vaccines a useful tool in the development of novel, more effective vaccines against intracellular infections (e.g. HIV, malaria and tuberculosis). Currently, screening of novel liposome formulations uses murine in vivo models which generate data that often correlates poorly with human data. In addition, these models are both high cost and low throughput, making them prohibitive for large scale screening of formulation libraries. This study uses the cationic liposome formulation DDA:TDB (known as cationic adjuvant formulation 01 (CAF01)), as a lead formulation, along with other liposome formulations of known in vivo efficacy to develop an in vitro screening tool for liposome formulation development. THP-1-derived macrophages were the model antigen presenting cell used to assess the ability of the liposome formulations to attract, associate with and activate antigen presenting cells in vitro, crucial steps necessary for an effective immune response to antigen. By using a combination of in vitro functions, the study highlights the potential use of an in vitro screening tool, to predict the in vivo efficacy of novel liposome formulations. CAF01 was predicted as the most effective liposome formulation when assessing all in vitro functions and a measure of in vitro activation was able to predict 80% of the liposome correctly for their ability to induce an in vivo IFN-ү response

Extratropical storm inundation testbed : intermodel comparisons in Scituate, Massachusetts

Author Posting. © American Geophysical Union, 2013. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Journal of Geophysical Research: Oceans 118 (2013): 5054–5073, doi:10.1002/jgrc.20397.The Integrated Ocean Observing System Super-regional Coastal Modeling Testbed had one objective to evaluate the capabilities of three unstructured-grid fully current-wave coupled ocean models (ADCIRC/SWAN, FVCOM/SWAVE, SELFE/WWM) to simulate extratropical storm-induced inundation in the US northeast coastal region. Scituate Harbor (MA) was chosen as the extratropical storm testbed site, and model simulations were made for the 24–27 May 2005 and 17–20 April 2007 (“Patriot's Day Storm”) nor'easters. For the same unstructured mesh, meteorological forcing, and initial/boundary conditions, intermodel comparisons were made for tidal elevation, surface waves, sea surface elevation, coastal inundation, currents, and volume transport. All three models showed similar accuracy in tidal simulation and consistency in dynamic responses to storm winds in experiments conducted without and with wave-current interaction. The three models also showed that wave-current interaction could (1) change the current direction from the along-shelf direction to the onshore direction over the northern shelf, enlarging the onshore water transport and (2) intensify an anticyclonic eddy in the harbor entrance and a cyclonic eddy in the harbor interior, which could increase the water transport toward the northern peninsula and the southern end and thus enhance flooding in those areas. The testbed intermodel comparisons suggest that major differences in the performance of the three models were caused primarily by (1) the inclusion of wave-current interaction, due to the different discrete algorithms used to solve the three wave models and compute water-current interaction, (2) the criterions used for the wet-dry point treatment of the flooding/drying process simulation, and (3) bottom friction parameterizations.This project was supported by NOAA via the U.S.IOOS Office (award: NA10NOS0120063 and NA11NOS0120141) and was managed by the Southeastern Universities Research Association. The Scituate FVCOM setup was supported by the NOAA-funded IOOS NERACOOS program for NECOFS and the MIT Sea Grant College Program through grant 2012-R/RC-127.2014-04-0

Exposure of the basophilic cell line KU812 to liposomes reveals activation profiles associated with potential anaphylactic responses linked to physico-chemical characteristics

Lipidic nanoparticles (LNP), particularly liposomes, have been proven to be a successful and versatile platform for intracellular drug delivery for decades. Whilst primarily developed for small molecule delivery, liposomes have recently undergone a renaissance due to their success in vaccination strategies, delivering nucleic acids, in the COVID-19 pandemic. As such, liposomes are increasingly being investigated for the delivery of nucleic acids, beyond mRNA, as non-viral gene delivery vectors. Although not generally considered toxic, liposomes are increasingly shown to not be immunologically inert, which may have advantages in vaccine applications but may limit their use in other conditions where immunological responses may lead to adverse events, particularly those associated with complement activation. We sought to assess a small panel of liposomes varying in a number of physico-chemical characteristics associated with complement activation and inflammatory responses, and examine how basophil-like cells may respond to them. Basophils, as well as other cell types, are involved in the anaphylactic responses to liposomes but are difficult to isolate in sufficient numbers to conduct large scale analysis. Here, we report the use of the human KU812 cell line as a surrogate for primary basophils. Multiple phenotypic markers of activation were assessed, as well as the release of histamine and inflammasome activity within the cells. We found that larger liposomes were more likely to result in KU812 activation, and that non-PEGylated liposomes were potent stimulators of inflammasome activity (four-fold greater IL-1β secretion than untreated controls), and a lower ratio of cholesterol to lipid was also associated with greater IL-1β secretion ([Cholesterol:DSPC ratio] 1:10; 0.35 pg/mL IL-1β vs. 5:10; 0.1 pg/mL). Additionally, PEGylation appeared to be associated with direct KU812 activation. These results suggest possible mechanisms related to the consequences of complement activation that may be underpinned by basophilic cells, in addition to other immune cell types. Investigation of the mechanisms behind these responses, and their impact on use in vivo, are now warranted

Wave-driven inner-shelf motions on the Oregon coast

Author Posting. © American Meteorological Society, 2009. This article is posted here by permission of American Meteorological Society for personal use, not for redistribution. The definitive version was published in Journal of Physical Oceanography 39 (2009): 2942-2956, doi:10.1175/2009JPO4041.1.Recent work by S. Lentz et al. documents offshore transport in the inner shelf due to a wave-driven return flow associated with the Hasselmann wave stress (the Stokes–Coriolis force). This analysis is extended using observations from the central Oregon coast to identify the wave-driven return flow present and quantify the potential bias of wind-driven across-shelf exchange by unresolved wave-driven circulation. Using acoustic Doppler current profiler (ADCP) measurements at six stations, each in water depths of 13–15 m, observed depth-averaged, across-shelf velocities were generally correlated with theoretical estimates of the proposed return flow. During times of minimal wind forcing, across-shelf velocity profiles were vertically sheared, with stronger velocities near the top of the measured portion of the water column, and increased in magnitude with increasing significant wave height, consistent with circulation due to the Hasselmann wave stress. Yet velocity magnitudes and vertical shears were stronger than that predicted by linear wave theory, and more similar to the stratified “summer” velocity profiles described by S. Lentz et al. Additionally, substantial temporal and spatial variability of the wave-driven return flow was found, potentially due to changing wind and wave conditions as well as local bathymetric variability. Despite the wave-driven circulation found, subtracting estimates of the return flow from the observed across-shelf velocity had no significant effect on estimates of the across-shelf exchange due to along-shelf wind forcing at these water depths along the Oregon coast during summer.This work was performed with the Partnership for Interdisciplinary Studies of Coastal Oceans (PISCO), funded primarily by the Gordon and Betty Moore Foundation and David and Lucile Packard Foundation. SL acknowledges support from NSF Ocean Science Grant #OCE-0548961. AK acknowledges support from the WHOI Coastal Ocean Institute Fellowship

Using microfluidics for scalable manufacturing of nanomedicines from bench to GMP : a case study using protein-loaded liposomes

Nanomedicines are well recognised for their ability to improve therapeutic outcomes. Yet, due to their complexity, nanomedicines are challenging and costly to produce using traditional manufacturing methods. For nanomedicines to be widely exploited, new manufacturing technologies must be adopted to reduce development costs and provide a consistent product. Within this study, we investigate microfluidic manufacture of nanomedicines. Using protein-loaded liposomes as a case study, we manufacture liposomes with tightly defined physico-chemical attributes (size, PDI, protein loading and release) from small-scale (1 mL) through to GMP volume production (200 mL/min). To achieve this, we investigate two different laminar flow microfluidic cartridge designs (based on a staggered herringbone design and a novel toroidal mixer design); for the first time we demonstrate the use of a new microfluidic cartridge design which delivers seamless scale-up production from bench-scale (12 mL/min) through GMP production requirements of over 20 L/h using the same standardised normal operating parameters. We also outline the application of tangential flow filtration for down-stream processing and high product yield. This work confirms that defined liposome products can be manufactured rapidly and reproducibly using a scale-independent production process, thereby de-risking the journey from bench to approved product