Development and characterization of self-assembling nanoparticles using a bio-inspired amphipathic peptide for gene delivery

The design of a non-viral gene delivery vehicle capable of delivering and releasing a functional nucleic acid cargo intracellularly remains a formidable challenge. For systemic gene therapy to be successful a delivery vehicle is required that protects the nucleic acid cargo from enzymatic degradation, extravasates from the vasculature, traverses the cell membrane, disrupts the endosomal vesicles and unloads the cargo at its destination site, namely the nucleus for the purposes of gene delivery. This manuscript reports the extensive investigation of a novel amphipathic peptide composed of repeating RALA units capable of overcoming the biological barriers to gene delivery both in vitro and in vivo. Our data demonstrates the spontaneous self-assembly of cationic DNA-loaded nanoparticles when the peptide is complexed with pDNA. Nanoparticles were < 100 nm, were stable in the presence of serum and were fusogenic in nature, with increased peptide α-helicity at a lower pH. Nanoparticles proved to be non-cytotoxic, readily traversed the plasma membrane of both cancer and fibroblast cell lines and elicited reporter-gene expression following intravenous delivery in vivo. The results of this study indicate that RALA presents an exciting delivery platform for the systemic delivery of nucleic acid therapeutics.

Instructional Methods For Optimal Learning For An Aerospace Assembly Task: digital animation v pictogram-based instructions

Previous studies on work instruction delivery for complex assembly tasks have shown that the mode and delivery method for the instructions in an engineering context can influence both build time and product quality. The benefits of digital, animated instructional formats when compared to static pictures and text only formats have already been demonstrated. Although pictograms have found applications for relatively straight forward operations and activities, their applicability to relatively complex assembly tasks has yet to be demonstrated. This study compares animated instructions and pictograms for the assembly of an aircraft panel. Based around a series of build experiments, the work records build time as well as the number of media references to measure and compare build efficiency. The number of build errors and the time required to correct them is also recorded. The experiments included five participants completing five builds over five consecutive days for each media type. Results showed that on average the total build time was 13.1% lower for the group using animated instructions. The benefit of animated instructions on build time was most prominent in the first three builds, by build four this benefit had disappeared. There were a similar number of instructional references for the two groups over the five builds but the pictogram users required a lot more references during build 1. There were more errors among the group using pictograms requiring more time for corrections during the build.

Optimizing geometric accuracy of four dimensional CT scans acquired using the wall and couch mounted Varian Real-Time Position Management camera systems

Objective:

The aim of this study was to identify sources of anatomical misrepresentation due to the location of camera mounting, tumour motion velocity and image processing artefacts in order to optimise the 4DCT scan protocol and improve geometrical-temporal accuracy.

A phantom with an imaging insert was driven with a sinusoidal superior-inferior motion of varying amplitude and period for 4DCT scanning. The length of a high density cube within the insert was measured using treatment planning software to determine the accuracy of its spatial representation. Scan parameters were varied including the tube rotation period and the cine time between reconstructed images. A CT image quality phantom was used to measure various image quality signatures under the scan parameters tested.

No significant difference in spatial accuracy was found for 4DCT scans carried out using the wall mounted or couch mounted camera for sinusoidal target motion. Greater spatial accuracy was found for 4DCT scans carried out using a tube rotation speed of 0.5s rather than 1.0s. The reduction in image quality when using a faster rotation speed was not enough to require an increase in patient dose.

4DCT accuracy may be increased by optimising scan parameters, including choosing faster tube rotation speeds. Peak misidentification in the recorded breathing trace leads to spatial artefacts and this risk can be reduced by using a couch mounted infrared camera.

This study explicitly shows that 4DCT scan accuracy is improved by scanning with a faster CT tube rotation speed.

Conventional in vivo irradiation procedures are insufficient to accurately determine tumor responses to non-uniform radiation fields

Purpose: To determine differences in overall tumor responses measured by volumetric assessment and bioluminescence imaging (BLI) following exposure to uniform and non-uniform radiation fields in an ectopic prostate tumor model.

Materials and methods: Bioluminescent human prostate tumor xenografts were established by subcutaneous implantation into male mice. Tumors were irradiated with uniform or non-uniform field configurations using conventional in vivo irradiation procedures performed using a 225 kVp generator with custom lead shielding. Tumor responses were measured using Vernier calipers and by BLI using an in vivo imaging system. Survival was defined as the time to quadroupling of pre-treatment tumor volume. 

Results: The correlation between BLI and tumor volume measurements was found to be different for un-irradiated (R = 0.61), uniformly irradiated (R = 0.34) and partially irradiated (R = 0.30) tumors. Uniformly irradiated tumors resulted in an average tumor growth delay of 60 days with median survival of 75 days, compared to partially irradiated tumors which showed an average growth delay of 24 days and median survival of 38 days. 

Conclusions: Correlation between BLI and tumor volume measurements is lower for partially irradiated tumors than those exposed to uniform dose distributions. The response of partially irradiated tumors suggests non-uniformity in response beyond physical dose distribution within the target volume. Dosimetric uncertainty associated with conventional in vivo irradiation procedures prohibits their ability to accurately determine tumor response to non-uniform radiation fields and stresses the need for image guided small animal radiation research platforms.

Using postal questionnaires to evaluate physical activity and diet behaviour change: case study exploring implications of valid responder characteristics in interpreting intervention outcomes.

Background: Patient reported outcome measures (PROMs) are used to evaluate lifestyle interventions but littleis known about differences between patients returning valid and invalid responses, or of potential for bias inevaluations. We aimed to examine the characteristics of patients who returned valid responses to lifestylequestionnaires compared to those whose responses were invalid for evaluating lifestyle change. 
Methods: We conducted a secondary data analysis from the SPHERE Study, a trial of an intervention to improveoutcomes for patients with coronary heart disease in primary care. Postal questionnaires were used to assessphysical activity (Godin) and diet (DINE) among study participants at baseline and 18 month follow-up. Three binaryresponse variables were generated for analysis: (1) valid Godin score; (2) valid DINE Fibre score; and (3) validDINE Total Fat score. Multivariate analysis comprised generalised estimating equation regression to examine theassociation of patients’ characteristics with their return of valid responses at both timepoints. 
Results: Overall, 92.1% of participants (832/903) returned questionnaires at both baseline and 18 months. Relativelyfewer valid Godin scores were returned by those who left school aged <15 years (36.5%) than aged 18 and over(50.5%), manual workers (39.5%) than non-manual (49.5%) and those with an elevated cholesterol (>5 mmol)(34.7%) than those with a lower cholesterol (44.4%) but multivariate analysis identified that only school leaving age(p = 0.047) was of statistical significance.Relatively fewer valid DINE scores were returned by manual than non-manual workers (fibre: 80.8% v 86.8%;fat: 71.2% v 80.0%), smokers (fibre: 72.6% v 84.7%; fat: 67.5% v 76.9%), patients with diabetes (fibre: 75.9% v 82.9%;fat: 66.9% v 75.8%) and those with cholesterol >5 mmol (fat: 68.2% v 76.2%) but multivariate analysis showedstatistical significance only for smoking (fibre: p = 0.013; fat: p = 0.045), diabetes (fibre: p = 0.039; fat: p = 0.047), andcholesterol (fat: p = 0.039). 
Conclusions: Our findings illustrate the importance of detailed reporting of research methods, with clearinformation about response rates, respondents and valid outcome data. Outcome measures which are relevant to astudy population should be chosen carefully. The impact of methods of outcome measurement and valid responserates in evaluating healthcare requires further study.

Ultrahigh Brilliance Multi-MeV γ-Ray Beams from Nonlinear Relativistic Thomson Scattering

We report on the generation of a narrow divergence (θγ<2.5mrad), multi-MeV (Emax≈18MeV) and ultrahigh peak brilliance (>1.8×1020photonss-1mm-2mrad-2 0.1% BW) γ-ray beam from the scattering of an ultrarelativistic laser-wakefield accelerated electron beam in the field of a relativistically intense laser (dimensionless amplitude a0≈2). The spectrum of the generated γ-ray beam is measured, with MeV resolution, seamlessly from 6 to 18 MeV, giving clear evidence of the onset of nonlinear relativistic Thomson scattering. To the best of our knowledge, this photon source has the highest peak brilliance in the multi-MeV regime ever reported in the literature.

Ultra-broadband Polarisers Based on Metastable Free-Standing Aligned Carbon Nanotube Membranes

A carbon nanotube free-standing linearly dichroic polariser is developed using solid-state extrusion. Membrane cohesion is experimentally and numerically demonstrated to derive from inter-tube van der Waals interactions in this family of planar metastable morphologies, controlled by the chemical vapour deposition conditions. Ultra-broadband polarisation (400 nm – 2.5 mm) is shown and corroborated by effective medium and full numerical simulations.

Electrical conduction and rheological behaviour of composites of poly(epsilon-caprolactone) and MWCNTs

Two mechanisms of conduction were identified from temperature dependent (120 K-340 K) DC electrical resistivity measurements of composites of poly(c-caprolactone) (PCL) and multi-walled carbon nanotubes (MWCNTs). Activation of variable range hopping (VRH) occurred at lower temperatures than that for temperature fluctuation induced tunneling (TFIT). Experimental data was in good agreement with the VRH model in contrast to the TFIT model, where broadening of tunnel junctions and increasing electrical resistivity at T > T-g is a consequence of a large difference in the coefficients of thermal expansion of PCL and MWCNTs. A numerical model was developed to explain this behavior accounting for a thermal expansion effect by supposing the large increase in electrical resistivity corresponds to the larger relative deformation due to thermal expansion associated with disintegration of the conductive MWCNT network. MWCNTs had a significant nucleating effect on PCL resulting in increased PCL crystallinity and an electrically insulating layer between MWCNTs. The onset of rheological percolation at similar to 0.18 vol% MWCNTs was clearly evident as storage modulus, G' and complex viscosity, vertical bar eta*vertical bar increased by several orders of magnitude. From Cole-Cole and Van Gurp-Palmen plots, and extraction of crossover points (G(c)) from overlaying plots of G' and G '' as a function of frequency, the onset of rheological percolation at 0.18 vol% MWCNTs was confirmed, a similar MWCNT loading to that determined for electrical percolation.