Pressure-mediated oligonucleotide transfection of rat and human cardiovascular tissues

The application of gene therapy to human disease is currently restricted by the relatively low efficiency and potential hazards of methods of oligonucleotide or gene delivery. Antisense or transcription factor decoy oligonucleotides have been shown to be effective at altering gene expression in cell culture expreriments, but their in vivo application is limited by the efficiency of cellular delivery, the intracellular stability of the compounds, and their duration of activity. We report herein the development of a highly efficient method for naked oligodeoxynucleotide (ODN) transfection into cardiovascular tissues by using controlled, nondistending pressure without the use of viral vectors, lipid formulations, or exposure to other adjunctive, potentially hazardous substances. In this study, we have documented the ability of ex vivo, pressure-mediated transfection to achieve nuclear localization of fluorescent (FITC)-labeled ODN in approximately 90% and 50% of cells in intact human saphenous vein and rat myocardium, respectively. We have further documented that pressure-mediated delivery of antisense ODN can functionally inhibit target gene expression in both of these tissues in a sequence-specific manner at the mRNA and protein levels. This oligonucleotide transfection system may represent a safe means of achieving the intraoperative genetic engineering of failure-resistant human bypass grafts and may provide an avenue for the genetic manipultation of cardiac allograft rejection, allograft vasculopathy, or other transplant diseases.

3D numerical modelling of acoustic horns using the method of fundamental solutions

In the present work, a three-dimensional (3D) formulation based on the method of fundamental solutions (MFS) is applied to the study of acoustic horns. The implemented model follows and extends previous works that only considered two-dimensional and axisymmetric horn configurations. The more realistic case of 3D acoustic horns with symmetry regarding two orthogonal planes is addressed. The use of the domain decomposition technique with two interconnected sub-regions along a continuity boundary is proposed, allowing for the computation of the sound pressure generated by an acoustic horn installed on a rigid screen. In order to reduce the model discretization requirements for these cases, Green’s functions derived with the image source methodology are adopted, automatically accounting for the presence of symmetry conditions. A strategy for the calculation of an optimal position of the virtual sources used by the MFS to define the solution is also used, leading to improved reliability and flexibility of the proposed method. The responses obtained by the developed model are compared to reference solutions, computed by well-established models based on the boundary element method. Additionally, numerically calculated acoustic parameters, such as directivity and beamwidth, are compared with those evaluated experimentally.

Refined method of potential enhancement in the equilibria characterization of activated carbon. Comparison with GCMC and DFT

In this paper, we present a technique for equilibria characterization of activated carbon having slit-shaped pores. This method was first developed by Do (Do, D. D. A new method for the characterisation of micro-mesoporous materials. Presented at the International Symposium on New Trends in Colloid and Interface Science, September 24-26, 1998 Chiba, Japan) and applied by his group and other groups for characterization of pore size distribution (PSD) as well as adsorption equilibria determination of a wide range of hydrocarbons. It is refined in this paper and compared with the grand canonical Monte Carlo (GCMG) simulation and density functional theory (DFT). The refined theory results in a good agreement between the pore filling pressure versus pore width and those obtained by GCMG and DFT. Furthermore, our local isotherms are qualitatively in good agreement with those obtained by the GCMC simulations. The main advantage of this method is that it is about 4 orders of magnitude faster than the GCMC simulations, making it suitable for optimization studies and design purposes. Finally, we apply our method and the GCMG in the derivation of the PSD of a commercial activated carbon. It was found that the PSD derived from our method is comparable to that derived from the GCMG simulations.

Corrosion behaviour of a pressure die cast magnesium alloy

High pressure die casting is the most important production method for casting magnesium alloy components, and uniformity of appearance is an important criterion for acceptance of a component by customers. This paper investigates the influence of uniformity in surface appearance of diecast AZ91D plates on their corrosion behaviour. Through immersion, hydrogen collection and weight loss measurements it was found that corrosion is more likely to occur on the areas of the plate that appear to be darker, leading to a non-uniformly corroded surface. Microstructural analysis showed that the non-uniformity in appearance is related to a difference in the morphology and distribution of porosity across the surface of a diecast AZ91D plate. The darker areas of the surface are high in porosity which breaks the continuity of the beta-phase network and provides shortcut paths for corrosion from the surface to the interior of the casting. The brighter shiny areas of the surface are much less porous, with isolated pores being confined by corrosion resistant beta-precipitates thus reducing the corrosion rate.

Manufacture of high pressure die-cast radio frequency filter bodies

The manufacture of a radio frequency filter box using high pressure die casting (HPDC) is compared to the traditional high speed machining route. This paper describes an industrial exercise that concluded HPDC to be an economical and appropriate method to produce larger volumes of thin-walled telecommunications components. Modifications to the component design were made to make the component suitable for the HPDC process. Development of the die design through simulation modelling is described. The wrought alloy was replaced by near-eutectic Al-Si die casting alloy that was found to give better temperature stability performance. Apart from the economic benefits, HPDC was found to give lower filter efficiency losses through better surface finish. The effects of HPDC process variables, such as intensification pressure and injection piston velocity, on component quality, particularly porosity levels, were investigated. The pressure was analysed in terms of HPDC machine set pressure and the pressure measured in the die cavity by pressure sensors. Porosity was found to decrease with increased pressure and slightly increase with higher casting velocities.

The effect of positive end-expiratory pressure level on peak expiratory flow during manual hyperinflation

Including positive end-expiratory pressure (PEEP) in the manual resuscitation bag (MRB) may render manual hyperinflation (MHI) ineffective as a secretion maneuver technique in mechanically ventilated patients. In this study we aimed to determine the effect of increased PEEP or decreased compliance on peak expiratory flow rate (PEF) during MHI. A blinded, randomized study was performed on a lung simulator by 10 physiotherapists experienced in MHI and intensive care practice. PEEP levels of 0-15 cm H2O, compliance levels of 0.05 and 0.02 L/cm H2O, and MRB type were randomized. The Mapleson-C MRB generated significantly higher PEF (P < 0.01, d = 2.72) when compared with the Laerdal MRB for all levels of PEEP. In normal compliance (0.05 L/cm H2O) there was a significant decrease in PEF (P < 0.01, d = 1.45) for a PEEP more than 10 cm H2O in the Mapleson-C circuit. The Laerdal MRB at PEEP levels of more than 10 cm H2O did not generate a PEF that is theoretically capable of producing two-phase gas-liquid flow and, consequently, mobilizing pulmonary secretions. If MHI is indicated as a result of mucous plugging, the Mapleson-C MRB may be the most effective method of secretion mobilization.

Development of a non-intrusive heat transfer coefficient gauge and its application to high pressure die casting Effect of the process parameters

A heat transfer coefficient gauge has been built, obeying particular rules in order to ensure the relevance and accuracy of the collected information. The gauge body is made out of the same materials as the die casting die (H13). It is equipped with six thermocouples located at different depths in the body and with a sapphire light pipe. The light pipe is linked to an optic fibre, which is connected to a monochromatic pyrometer. Thermocouples and pyrometer measurements are recorded with a data logger. A high pressure die casting die was instrumented with one such gauge. A set of 150 castings was done and the data recorded. During the casting, some process parameters have been modified such as piston velocity, intensification pressure, delay before switch to the intensification stage, temperature of the alloy, etc.... The data was treated with an inverse method in order to transform temperature measurements into heat flux density and heat transfer coefficient plots. The piston velocity and the initial temperature of the die seem to be the process parameters that have the greatest influence on the heat transfer. (c) 2005 Elsevier B.V. All rights reserved.

New method to determine PSD using supercritical adsorption: Applied to methane adsorption in activated carbon

Adsorption of supercritical fluids is increasingly carried out to determine the micropore size distribution. This is largely motivated by the advances in the use of supercritical adsorption in high energy applications, such as hydrogen and methane storage in porous media. Experimental data are reported as mass excess versus pressure, and when these data are matched against the theoretical mass excess, significant errors could occur if the void volume used in the calculation of the experimental mass excess is incorrectly determined [Malbrunot, P.; Vidal, D.; Vermesse, J.; Chahine, R.; Bose, T. K. Langmuir 1997, 13, 539]. 1 The incorrect value for the void volume leads to a wrong description of the maximum in the plot of mass excess versus pressure as well as the part of the isotherm over the pressure region where the isotherm is decreasing. Because of this uncertainty in the maximum and the decreasing part of the isotherm, we propose a new method in which the problems associated with this are completely avoided. Our method involves only the relationship between the amount that is introduced into the adsorption cell and the equilibrium pressure. This information of direct experimental data has two distinct advantages. The first is that the data is the raw data without any manipulation (i.e., involving further calculations), and the second one is that this relationship always monotonically increases with pressure. We will illustrate this new method with the adsorption data of methane in a commercial sample of activated carbon.

A novel method for the production of aluminium nitride

A method has been developed to produce thick (> 400 mu m) AlN surface layers oil aluminium plates at 540 degrees C, under nitrogen at atmospheric pressure. A critical element of the process is the use of Mg powder placed in close proximity to the Al plate surface. The Mg reduces/disrupts the natural, protective oxide film on the Al surface. The nitride layers form through two distinct modes, one growing outward from the Al plate surface and the other growing into the Al. (c) 2006 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Field observations of instantaneous water slopes and horizontal pressure gradients in the swash-zone

Field observations of instantaneous water surface slopes in the swash zone are presented. For free-surface flows with a hydrostatic pressure distribution the surface slope is equivalent to the horizontal pressure gradient. Observations were made using a novel technique which in its simplest form consists of a horizontal stringline extending seaward from the beach face. Visual observation, still photography or video photography is then sufficient to determine the surface slope where the free-surface cuts the line or between reference points in the image. The method resolves the mean surface gradient over a cross-shore distance of 5 m or more to within +/- 0.001, or 1/20th -1/100th of typical beach gradients. In addition, at selected points and at any instant in time during the swash cycle, the water surface slope can be determined exactly to be dipping either seaward or landward. Close to the location of bore collapse landward dipping water surface slopes of order 0.05-0.1 occur over a very small region (order 0.5 m) at the blunt or convex leading edge of the swash. In the middle and upper swash the water surface slope at this leading edge is usually very close to horizontal or slightly seaward. Behind the leading edge, the water surface slope was observed to be very close to horizontal or dipping seaward at all times throughout the swash uprush. During the backwash the water surface slope was observed to be always dipping seaward, approaching the beach slope, and remained seaward until a new uprush edge or incident bore passed any particular cross-shore location of interest. The observations strongly Suggest that the swash boundary layer is subject to an adverse pressure gradient during uprush and a favourable pressure gradient during the backwash. Furthermore, assuming Euler's equations are a good approximation in the swash, the observations also show that the total fluid acceleration is negative (offshore) for almost the whole of the uprush and for the entire backwash. The observations are contrary to recent work suggesting significant shoreward directed accelerations and pressure gradients occur in the swash (i.e., delta u/delta t > 0 similar to delta p/delta x < 0), but consistent with analytical and numerical solutions for swash uprush and backwash. The results have important implications for sediment transport modelling in the swash zone.

Measurement and stability of FTY720 in human whole blood by high-performance liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry

We report here a validated method for the quantification of a new immunosuppressant drug FTY720, using HPLC-tandem mass spectrometry. Whole blood samples (500 mu l) were subjected to liquid-liquid extraction, in the presence of an internal standard (Y-32919). Mass spectrometric detection was by selected reaction monitoring with an atmospheric pressure chemical ionization source in positive ionization mode (FTY720: m/z 308.3 -> 255.3). The assay was linear from 0.2 to 25 mu g/l (r(2) > 0.997, n = 5). The inter- and intra-day analytical recovery and imprecision for quality control samples (0.5, 7 and 15 mu g/l) were 95.8-103.2 and < 5.5%, respectively. At the lower limit of quantification (0.2 mu g/l) the interand intra-day analytical recovery was 99.0-102.8% with imprecision of < 7.6% (n = 5). The assay had a mean relative recovery of 100.5 +/- 5.8% (n = 15). Extracted samples were stable for 16 h. IFTY720 quality control samples were stable at room temperature for 16 h at 4 degrees C for at least 8 days and when taken through at least three freeze-thaw cycles. In conclusion, the method described displays analytical performance characteristics that are suitable for pharmacokinetic studies in humans. (c) 2006 Elsevier B.V. All rights reserved.

Measurements of polarimetric sensitivity to hydrostatic pressure, strain and temperature in birefringent dual-core microstructured polymer fiber

We experimentally characterized a birefringent microstructured polymer fiber of specific construction, which allows for single mode propagation in two cores separated by a pair of large holes. The fiber exhibits high birefringence in each of the cores as well as relatively weak coupling between the cores. Spectral dependence of the group and the phase modal birefringence was measured using an interferometric method. We have also measured the sensing characteristics of the fiber such as polarimetric sensitivity to hydrostatic pressure, strain and temperature. Moreover, we have studied the effect of hydrostatic pressure and strain on coupling between the cores.

Analysis of the intraocular pressure pulse

This thesis was concerned with investigating methods of improving the IOP pulse’s potential as a measure of clinical utility. There were three principal sections to the work. 1. Optimisation of measurement and analysis of the IOP pulse. A literature review, covering the years 1960 – 2002 and other relevant scientific publications, provided a knowledge base on the IOP pulse. Initial studies investigated suitable instrumentation and measurement techniques. Fourier transformation was identified as a promising method of analysing the IOP pulse and this technique was developed. 2. Investigation of ocular and systemic variables that affect IOP pulse measurements In order to recognise clinically important changes in IOP pulse measurement, studies were performed to identify influencing factors. Fourier analysis was tested against traditional parameters in order to assess its ability to detect differences in IOP pulse. In addition, it had been speculated that the waveform components of the IOP pulse contained vascular characteristic analogous to those components found in arterial pulse waves. Validation studies to test this hypothesis were attempted. 3. The nature of the intraocular pressure pulse in health and disease and its relation to systemic cardiovascular variables. Fourier analysis and traditional parameters were applied to the IOP pulse measurements taken on diseased and healthy eyes. Only the derived parameter, pulsatile ocular blood flow (POBF) detected differences in diseased groups. The use of an ocular pressure-volume relationship may have improved the POBF measure’s variance in comparison to the measurement of the pulse’s amplitude or Fourier components. Finally, the importance of the driving force of pulsatile blood flow, the arterial pressure pulse, is highlighted. A method of combining the measurements of pulsatile blood flow and pulsatile blood pressure to create a measure of ocular vascular impedance is described along with its advantages for future studies.

Three-dimensional fluid pressure mapping in porous media using magnetic resonance imaging with gas-filled liposomes

This paper presents and demonstrates a method for using magnetic resonance imaging to measure local pressure of a fluid saturating a porous medium. The method is tested both in a static system of packed silica gel and in saturated sintered glass cylinders experiencing fluid flow. The fluid used contains 3% gas in the form of 3-μm average diameter gas filled 1,2-distearoyl-sn-glycero-3-phosphocholine (C18:0, MW: 790.16) liposomes suspended in 5% glycerol and 0.5% Methyl cellulose with water. Preliminary studies at 2.35 T demonstrate relative magnetic resonance signal changes of 20% per bar in bulk fluid for an echo time TE=40 ms, and 6-10% in consolidated porous media for TE=10 ms, over the range 0.8-1.8 bar for a spatial resolution of 0.1 mm3 and a temporal resolution of 30 s. The stability of this solution with relation to applied pressure and methods for improving sensitivity are discussed. © 2007 Elsevier Inc. All rights reserved.

MRI pressure and stress measurement in novel homogeneous soft solids

This paper presents MRI measurements of a novel semi solid MR contrast agent to pressure. The agent is comprised of potassium chloride cross linked carageenan gum at a concentration of 2% w/v, with micron size lipid coated bubbles of air at a concentration of 3% v/v. The choice for an optimum suspending medium, the methods of production and the preliminary MRI results are presented herein. The carageenan gum is shown to be ideally elastic for compressions relating to volume changes less than 15%, in contrast to the inelastic gellan gum also tested. Although slightly lower than that of gellan gum, carageenan has a water diffusion coefficient of 1.72×10-9 m2.s-1 indicating its suitability to this purpose. RARE imaging is performed whilst simultaneously compressing test and control samples and a maximum sensitivity of 1.6% MR signal change per % volume change is found which is shown to be independent of proton density variations due to the presence of microbubbles and compression. This contrast agent could prove useful for numerous applications, and particularly in chemical engineering. More generally the method allows the user to non-invasively image with MRI any process that causes, within the solid, local changes either in bubble size or bubble shape. © 2008 American Institute of Physics.