Immunocytochemical study of alpha 1 and beta 2/3 subunits of GABAA receptors in freehand isolated vestibular Deiters' neurons

Vestibular Deiters' neurons have been isolated from bovine brain by the Hydén's freehand dissection technique and challenged with monoclonal antibodies directed toward the alpha 1 and beta 2/3 subunits of the GABAA receptors. Subsequent challenge with fluorescent secondary antibodies and confocal microscopy allowed the study of the cellular distribution of such subunits. In Deiters' neurons the beta 2/3 subunit displayed a clear presence all along the cell body profile and the initial parts of the dendrites. The alpha 1 subunit was found highly present all over the cell interior except the nuclear profiles. The strong presence inside the cells possibly masked its presence on the plasma membrane. However, in part of the cells studied a distinct presence on the plasma membrane was evident. This subunit was visualized also all along the long dendrites of these neurons. The approach we describe here, involving freehand isolated mature neurons from adult animals, may allow a better characterization of the tridimensional distribution of different types of neuronal GABAA receptors in the respect of the approach with brain slices.     

Occupant deaths in large truck crashes in the United States: 25 years of experience

There is public concern about the magnitude of the problem of large truck crashes in the US. Fatalities in large truck crashes have not declined much; however, more large trucks are driving more miles than ever before while fatalities per mile driven have dropped substantially. This study examined how the public health burden of large truck crashes versus the risk per unit of travel has changed over 25 years.The present study focused on the US vehicle occupants in fatal crashes involving a large truck during 1975-1999. Occupant fatalities per 100000 population, per 10000 licensed drivers, per 10000 registered trucks and per 100 million vehicle-miles of travel (VMT) were calculated to determine trends in occupant deaths in large truck crashes.In 1999, large truck crashes resulted in 3916 occupant deaths in passenger vehicles and 747 in large trucks. Passenger vehicle occupant deaths in large truck crashes per 100000 population have increased somewhat since 1975 (1.28 in 1975 and 1.44 in 1999). There have been appreciable declines in occupant deaths per truck VMT since 1975, but the percentage reduction has been greater for occupants of large trucks (67%) than for passenger vehicle occupants (43%). However, truck drivers are at elevated risk of dying relative to their numbers in the workforce. Overall large truck involvements in fatal crashes per truck VMT decreased more than passenger vehicle involvements per passenger VMT (PVMT; 68% versus 33% decreases for single-vehicle crashes and 43% versus 23% for multiple-vehicle crashes). Large truck involvement in fatal crashes has dropped substantially when measured per unit of travel, but the public health burden of large truck crashes, as measured by deaths per 100000 population, has not improved over time because of the large increase in truck mileage. Research is needed on measures to better protect both occupants of large trucks and passenger vehicle occupants colliding with them.     

Effects of adsorbed heat labile serum proteins and fibrinogen on adhesion and apoptosis of monocytes/macrophages on biomaterials

A previously established human monocyte culture protocol was used to determine the effects of varying adsorbed proteins on monocyte/macrophage adhesion and survival on dimethyl-silane (DM) or RGD modified glass coverslips. Cells were allowed to adhere for 2 h in the absence of protein or in the presence of serum, fibrinogen (Fg), heat inactivated serum (HIS), serum supplemented with Fg or HIS with Fg. Cell adhesion and apoptosis rates were determined on days 0 (2 h), 3, 7 and 10 of culture. The presence of serum alone in the initial culture was sufficient to optimize monocyte/macrophage adhesion and survival rates. Adding Fg to serum did not increase adhesion nor decrease apoptotic rates. No protein or the addition of HIS during the initial incubation period significantly decreased monocyte/macrophage adhesion and survival on both surfaces, however, the addition of Fg to HIS restored adhesion and survival rates to those seen with in the presence of serum alone on RGD surfaces. These studies demonstrate that monocyte/macrophage adhesion and survival on biomaterial surfaces are optimized by adsorbed heat labile serum proteins while adsorbed Fg plays a surface property-dependent role.     

Characterisation of SiOxCyHz thin films deposited by low-temperature PECVD

SiO_xC_yH_z thin films were deposited from hexamethyldisiloxane (HMDSO)/O₂ mixtures in a parallel plate, capacitively coupled, RF plasma reactor. Polyethylene terephthalate (PET), Si(1 0 0) wafers and KBr tablets were chosen as substrates. Effect of HMDSO/O₂ ratio, total treatment pressure and power input on the properties of the deposited films were investigated. The structure and bondings were studied by means of Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. Wettability characteristics of the deposited thin films were investigated by means of water droplet contact angle measurements. Surface morphology was investigated with atomic force microscopy. Barrier properties of the SiO_xC_yH_z thin films were investigated by measuring the water vapour transmission rate of the coated PET substrates. Correlations between the characteristics of the deposited film and their barrier properties were discussed.     

High-Throughput Analysis of Algal Crude Oils Using High Resolution Mass Spectrometry

Lipid analysis often needs to be specifically optimized for each class of compounds due to its wide variety of chemical and physical properties. It becomes a serious bottleneck in the development of algae-based next generation biofuels when high-throughput analysis becomes essential for the optimization of various process conditions. We propose a high-resolution mass spectrometry-based high-throughput assay as a ‘quick-and-dirty’ protocol to monitor various lipid classes in algal crude oils. Atmospheric pressure chemical ionization was determined to be most effective for this purpose to cover a wide range of lipid classes. With an autosampler-LC pump set-up, we could analyze algal crude samples every one and half minutes, monitoring several lipid species such as TAG, DAG, squalene, sterols, and chlorophyll a. High-mass resolution and high-mass accuracy of the orbitrap mass analyzer provides confidence in the identification of these lipid compounds. MS/MS and MS3 analysis could be performed in parallel for further structural information, as demonstrated for TAG and DAG. This high-throughput method was successfully demonstrated for semi-quantitative analysis of algal oils after treatment with various nanoparticles.     

Quantitative detection of Listeria monocytogenes in raw milk and soft cheeses: Culture-independent versus liquid- and solid-based culture-dependent real time PCR approaches

A culture-independent and two culture-dependent real time (rt-) PCR approaches were developed to quantitatively identify Listeria monocytogenes in raw milk and soft cheeses. The optimised rt-PCR revealed 100% inclusivity and exclusivity. DNA- and cell-based standard curves showed a good linearity of response (R² ≥ 0.987 and R² ≥ 0.998, respectively) for five orders of magnitude (39 × 10⁵ – 3 × 10⁰ genome equivalents and 10⁶–10¹ CFU equivalents, respectively) with about 100% relative accuracy and inter-assay variability ≤0.90%. Up to 1 genome equivalent/and 10 CFU/reaction were quantified in the DNA and cell standard curves, respectively. The rt-PCR was then combined with a liquid- (MPN technique) or a solid- (ALOA and PALCAM) based enumeration. The diagnostic sensitivity of the different approaches was investigated in artificially contaminated raw milk and soft cheeses. The rt-PCR culture-independent approach performed well in raw milk and (with a lower sensitivity) in stracchino cheese-based standard curves. MPN/rt-PCR was the best approach to enumerate low levels of L. monocytogenes in raw milk and stracchino cheese, while the ALOA-based rt-PCR quantification was more effective than the PALCAM-based. These performances were confirmed when 23 real samples of raw milk and soft cheeses by both the rt-PCR approaches were assayed.     

Phase transformation and microstructural evolution after heat treatment of a terbium-doped lithium–aluminum phosphate glass

The crystallization kinetics and phase transformation of a transparent Tb³⁺-doped lithium–aluminum phosphate glass, prepared by melt quenching, were investigated. The energy associated to the glass transition and the crystallization parameters (activation energy for crystallization and Avrami exponent) were evaluated by different methods using the experimental data obtained by differential thermal analysis performed at different heating rates. Using an isoconversional method to determine the change of the activation energy for crystallization with the fraction of crystallization, it was verified that with the increase in the fraction of crystallization from 0.1 to 0.9, the value of the activation energy decreased slightly from ~370 to ~310 kJ mol^?1 and that the Avrami exponent varied from 0.8 to 1, suggesting a surface crystal growth mechanism. Observation of the microstructural evolution of heat-treated glass samples confirmed a surface crystallization process revealing spherulitic crystals constituted mainly by aluminum metaphosphate.