The soluble granulocyte-macrophage colony-stimulating factor receptor's carboxyl-terminal domain mediates retention of the soluble receptor on the cell surface through interaction with the granulocyte-macrophage colony-stimulating factor receptor beta-subunit

The hematopoietic cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) mediates its activity through binding to cell-surface receptors. The high-affinity GM-CSF receptor (GMR) consists of two transmembrane-anchored subunits: a ligand-specific, low-affinity subunit (GMRalpha); and a signal-transducing beta-subunit (GMRbeta). The human GMRalpha subunit also exists in a soluble isoform (SOLalpha) which antagonizes GM-CSF activity in vitro. Previous studies by us have shown that coexpression of SOLalpha and a mutated GMRbeta in BHK cells results in retention of SOLalpha on the cell surface and the formation of an intermediate affinity binding complex (Kd approximately 300 pM). This paper investigates the mechanism of the retention of SOLalpha on the cell surface. The data demonstrate that SOLalpha is anchored by a direct, ligand-independent interaction with GMRbeta which also occurs when SOLalpha is coexpressed with wild-type GMRbeta. However, SOLalpha and wild-type GMRbeta form a complex which binds GM-CSF with high affinity (Kd = 39 pM), indistinguishable from the binding characteristics of the TMalpha/GMRbeta complex. The experiments further reveal that the interaction between SOLalpha and GMRbeta is abrogated by removal of the unique 16 amino acid carboxyl-terminal domain of SOLalpha. Specific mutation of cysteine 323 in this carboxyl-domain to alanine also eliminates the cell-surface retention of SOLalpha identifying this residue as being necessary for the formation of the SOLalpha/GMRbeta complex.     

DNA melting investigated by differential scanning calorimetry and Raman spectroscopy

Thermal denaturation of the B form of double-stranded DNA has been probed by differential scanning calorimetry (DSC) and Raman spectroscopy of 160 base pair (bp) fragments of calf thymus DNA. The DSC results indicate a median melting temperature Tm = 75.5 degrees C with calorimetric enthalpy change delta Hcal = 6.7 kcal/mol (bp), van't Hoff enthalpy change delta HVH = 50.4 kcal/mol (cooperative unit), and calorimetric entropy change delta Scal = 19.3 cal/deg.mol (bp), at the experimental conditions of 55 mg DNA/ml in 5 mM sodium cacodylate at pH 6.4. The average cooperative melting unit (nmelt) comprises 7.5 bp. The Raman signature of 160 bp DNA is highly sensitive to temperature. Analyses of several conformation-sensitive Raman bands indicate the following ranges for thermodynamic parameters of melting: 43 < delta HVH < 61 kcal/mol (cooperative unit), 75 < Tm < 80 degrees C and 6 < (nmelt) < 9 bp, consistent with the DSC results. The changes observed in specific Raman band frequencies and intensities as a function of temperature reveal that thermal denaturation is accompanied by disruption of Watson-Crick base pairs, unstacking of the bases and disordering of the B form backbone. These three types of structural change are highly correlated throughout the investigated temperature range of 20 to 93 degrees C. Raman bands diagnostic of purine and pyrimidine unstacking, conformational rearrangements in the deoxyribose-phosphate moieties, and changes in environment of phosphate groups have been identified. Among these, bands at 834 cm-1 (due to a localized vibration of the phosphodiester group), 1240 cm-1 (thymine ring) and 1668 cm-1 (carbonyl groups of dT, dG and dC), are shown by comparison with DSC results to be the most reliable quantitative indicators of DNA melting. Conversely, the intensities of Raman marker bands at 786 cm-1 (cytosine ring), 1014 cm-1 (deoxyribose ring) and 1092 cm-1 (phosphate group) are largely invariant to melting and are proposed as appropriate standards for intensity normalizations.     

Hydrogen monitoring requirements in the global technical regulation on hydrogen and fuel cell vehicles

The United Nations Economic Commission for Europe Global Technical Regulation (GTR) Number 13 (Global Technical Regulation on Hydrogen and Fuel Cell Vehicles) is the defining document regulating safety requirements in hydrogen vehicles, and in particular, fuel cell electric vehicles (FCEVs). GTR Number 13 has been formally adopted and will serve as the basis for the national regulatory standards for FCEV safety in North America (led by the United States), Japan, Korea, and the European Union. The GTR defines safety requirements for these vehicles, including specifications on the allowable hydrogen levels in vehicle enclosures during in-use and post-crash conditions and on the allowable hydrogen emissions levels in vehicle exhaust during certain modes of normal operation. However, in order to be incorporated into national regulations, that is, to be legally binding, methods to verify compliance with the specific requirements must exist. In a collaborative program, the Sensor Laboratories at the National Renewable Energy Laboratory in the United States and the Joint Research Centre, Institute for Energy and Transport in the Netherlands have been evaluating and developing analytical methods that can be used to verify compliance with the hydrogen release requirements as specified in the GTR.     

DETERMINING THE BANDWIDTH OF A KERNEL SPECTRUM ESTIMATE

Abstract. A cross-validated form of Whittle's frequency domain approximation to the likelihood function of a stationary Gaussian process is described, and proposed as a criterion for choosing the bandwidth in a kernel spectrum estimate. The criterion is shown to be equivalent, in large samples, to the mean integrated squared error. The statistical properties of the spectrum estimate whose bandwidth maximizes the criterion have been explored in a limited simulation.     

Effects of ethanol on gene expression in rat bone: transient dose-dependent changes in mRNA levels for matrix proteins, skeletal growth factors, and cytokines are followed by reductions in bone formation

Several studies were performed in female rats to determine dose and time course changes in mRNA levels for matrix proteins in bone after a single administration of ethanol. As expected, dose-dependent transient increases in blood ethanol were measured. Additionally, there was mild hypocalcemia with no change in immunoreactive parathyroid hormone. Coordinated dose-dependent increases in mRNA for type 1 collagen, osteonectin, and osteocalcin were noted in the proximal tibial metaphysis 6 hr after ethanol was given, with the peak values occurring at a dose of 1.2 g/kg (0.4 ml). Similar increases in mRNA levels for matrix proteins were noted in lumbar vertebrae after ethanol treatment. The changes were specific for bone; ethanol had no effect on mRNA levels for matrix proteins in the uterus or liver, although the mRNA concentrations tended to be reduced in uterus. Message levels for several cytokines implicated in the regulation of bone turnover were also assayed; mRNA levels for transforming growth factor-beta1, transforming growth factor-beta2, interferon-gamma, and interleukin-6 were unchanged at doses ranging from 0.14 to 1.7 g/kg. At the highest dose of ethanol, the mRNA level for tumor necrosis factor-alpha was elevated while the level for insulin-like growth factor-1 was reduced. The time course effects of ethanol (0.4 ml dose) were determined in a separate experiment. Ethanol resulted in a transient increase in mRNA levels for the three bone matrix proteins assayed. However, matrix protein synthesis, as determined by incorporation of 3H-proline into the proximal tibial metaphysis, was not changed after 6 hr. The changes in mRNA levels for the matrix proteins were preceded by brief, transient decreases in mRNA levels for interleukin-1beta, interferon-gamma, and migration inhibitory factor, and followed by a more prolonged decrease in the mRNA level for insulin-like growth factor-1. A subsequent study was performed to determine the effects of repetitive daily treatment with ethanol on rat bone. After 7 days, there were highly significant decreases in the mRNA level for type 1 collagen, as well as decreased bone formation. These results suggest that ethanol may alter bone metabolism by disturbing signal transduction pathways that regulate the expression of genes for bone matrix proteins, skeletal growth factors, and cytokines.     

Surveillance and control of aflatoxin contamination of dried figs and fig paste imported into the United Kingdom.

Samples of dried figs and fig pastes from Turkey supplied voluntarily by UK importers and retailers during the period November 1988 to January 1989 showed both a high incidence and high levels of contamination with aflatoxins. In the samples tested, 24% had total aflatoxin concentrations above 10 micrograms/kg, with the highest level being 165 micrograms/kg. More rigorous monitoring of the 1989 fig harvest was undertaken on bulk consignments for all figs from Turkey entering the UK. For whole dried figs 20 kg samples were taken (as 20 sub-samples), and for fig paste 5 kg samples were taken (again as 20 sub-samples). Figs were minced, blended with water and mixed prior to sub-sampling for analysis. Analysis was by immunoaffinity column clean-up with HPLC determination of aflatoxins with fluorescence detection. Examination showed that 11% of 112 consignments of fig paste and 9% of 93 consignments of whole dried figs were contaminated with total alfatoxin concentrations above 10 micrograms/kg, with the highest level of contamination being 40 micrograms/kg. As a result of this surveillance programme 14 consignments of figs were refused entry into the United Kingdom.     

Impacts of climate change on the fate and behaviour of pesticides in surface and groundwater--A UK perspective

Over the last two decades significant effort has been dedicated to understanding the fate and transport of pesticides in surface water and groundwater and to use this understanding in the development of environmental policy and regulation. However, there have been few studies that have investigated the relationships between pesticides and climate change, and where this work has been undertaken it has principally been in relation to the impacts of climate change on agricultural production rather than in the context of environmental protection. This study addresses that gap by reviewing how climate change may impact the fate and transport of pesticides in surface and groundwaters as a pre-cursor to quantitative studies. In order to structure the review, we have adopted a source-pathway-receptor approach where climate sensitivities of pesticide source terms, environmental pathways and receptors are reviewed. The main climate drivers for changing pesticide fate and behaviour are thought to be changes in rainfall seasonality and intensity and increased temperatures, but the effect of climate change on pesticide fate and transport is likely to be very variable and difficult to predict. In the long-term, indirect impacts, such as land-use change driven by changes in climate, may have a more significant effect on pesticides in surface and groundwaters than the direct impacts of climate change on pesticide fate and transport. The review focuses on climate change scenarios and case studies from the UK; however, the general conclusions can be applied more widely.