Activation of mitogen-activated protein kinase cascades during priming of human neutrophils by TNF-alpha and GM-CSF

Oral administration of large doses of protein antigen generally induces a state of systemic unresponsiveness currently termed mucosally induced tolerance. In this study, we used human milk protein (HMP) without casein as a multi-protein antigen for the study of mucosally induced tolerance. The HMP utilized in this study mainly contained secretory (S) IgA, lactoferrin (Lf) and alpha-lactalbumin (Lact). When mice were given 1 or 25 mg of HMP orally 3 times or 25 mg orally four consecutive weeks prior to systemic immunization, antigen-specific serum IgG responses to HMP were induced by subsequent parenteral immunization with 100 microg of HMP. Analysis of IgG subclasses revealed that IgG1 followed by IgG2b accounted for the IgG responses noted. When both HMP and ovalbumin (OVA) were fed to mice, tolerance developed to OVA but not to HMP. To further investigate the nature of immune responses seen following oral gavage of HMP, we examined responses to individual protein of HMP. Brisk serum IgG1 and IgG2b responses to both S-IgA and Lf were induced by oral followed by systemic immunization with HMP. Analysis of splenic CD4+ T cells from mice given oral HMP revealed production of Th2- but not Th1-type cytokines. These results show that oral administration of HMP preferentially induces exclusive Th2-type immune responses, which may prevent the development of HMP (S-IgA and Lf)-specific mucosally induced tolerance.     

Assessing information needs and instrument availability for a pressurized water reactor during severe accidents

A five-step methodology was developed to evaluate information needs for nuclear power plants under accident conditions and the availability of plant instrumentation during severe accidents. Step 1 examines the credible accidents and their relationships to plant safety functions. Step 2 determines the information that personnel involved in accident management will need to understand plant behavior. Step 3 determines the capability of the instrumentation to function properly under severe accident conditions. Step 4 determines the conditions expected during the identified severe accidents. Step 5 compares the instrument capabilities and severe accident conditions, to evaluate the availability of the instrumentation to supply needed plant information. This methodology was applied to a pressurized water reactor with a large dry containment and the results are presented. A companion article describes application of the methodology to a boiling water reactor with a Mark I containment.     

Selective excitatory amino acid uptake in glutamatergic nerve terminals and in glia in the rat striatum: quantitative electron microscopic immunocytochemistry of exogenous (D)-aspartate and endogenous glutamate and GABA

To characterize glutamate/aspartate uptake activity in various cellular and subcellular elements in the striatum, rat striatal slices were exposed to 10 and 50 mu M exogenous (D)-aspartate. After fixation with glutaraldehyde/formaldehyde the distribution of (D)-aspartate was analysed by postembedding immunocytochemistry and the ultrastructural distribution was compared with the distributions of endogenous glutamate and GABA. Light microscopically, (D)-aspartate-like immunoreactivity was localized in conspicuous dots along very weakly labelled dendritic profiles and neuron cell bodies. At the electron microscope level gold particles signalling (D)-aspartate occurred at highest density in nerve terminals making asymmetrical contacts with postsynaptic spines (i.e. resembling synapses of cortical afferents). Astrocytic processes also contained gold particles, but at a lower density than nerve endings. In contrast, dendritic spines were only weakly (D)-aspartate-positive. The difference in labelling at 10 and 50 mu M (D)-aspartate was consistent with 'high-affinity' uptake. Neighbouring sections processed with other antibodies showed that the D-aspartate labelling. Occurred in nerve terminals strongly immunoreactive for glutamate, rather than in terminals very weakly glutamate-immunopositive or in nerve endings immunoreactive for GABA. Glutamate labelling of perfusion-fixed striatum confirmed that terminals forming asymmetrical synaptic contacts with spines were enriched with gold particles, suggesting that these terminals use glutamate as a transmitter. This study demonstrates that high-affinity uptake sites for excitatory amino acids in the striatum are most strongly expressed on presumed glutamatergic nerve terminals and on astrocytes.     

The bipartite geminivirus coat protein aids BR1 function in viral movement by affecting the accumulation of viral single-stranded DNA

The movement of bipartite geminiviruses such as squash leaf curl virus (SqLCV) requires the cooperative interaction of two essential virus-encoded movement proteins, BR1 and BL1. While the viral coat protein AR1 is not essential for systemic infection, genetic studies demonstrate that its presence masks the defective phenotype of certain BR1 missense mutants, thus suggesting that coat protein does interact with the viral movement pathway. To further examine the mechanism of this interaction, we have constructed alanine-scanning mutants of AR1 and studied them for the ability to mask the infectivity defects of appropriate BR1 mutants, for the ability to target to the nucleus and to bind viral single-stranded DNA (ssDNA) and multimerize, and for effects on the accumulation of replicated viral ssDNA. We identified a specific region of AR1 required for masking of appropriate BR1 mutants and showed that this same region of AR1 was also important for ssDNA binding and the accumulation of viral replicated ssDNA. This region of AR1 also overlapped that involved in multimerization of the coat protein. We also found that the accumulation in protoplasts of single-stranded forms of a recombinant plasmid that included the SqLCV replication origin but was too large to be encapsidated was dependent on the presence of AR1 but did not appear to require encapsidation. These findings extend our model for SqLCV movement, demonstrating that coat protein affects viral movement through its ability to induce the accumulation of replicated viral ssDNA genomes. They further suggested that encapsidation was not required for the AR1-dependent accumulation of viral ssDNA.     

On the future of the speciality of Obstetrics and Gynecology. A Dutch view

Between 4 and 15 ng/ml serum prostate-specific antigen (PSA) has a low specificity for prostate cancer (PCa). One accepted method to enhance this specificity is transrectal ultrasonography (TRUS)-measured PSA-density (PSA-D). We compared this method with a new alternative, transition zone PSA (PSA-TZ). We measured total and transition zone prostatic volumes by TRUS and calculated PSA-D and PSA-TZ in 59 patients with suspicion of PCa and PSA between 4 and 15 ng/ml. All patients then had sextant biopsies of the prostate, 30 were positive for PCa and 29 showed benign tissue. With a cut-off value of 0.35, PSA-TZ had a positive predicted value of 77% for PCa, whereas PSA-D, with a cut-off value of 0.12, had a positive predicted value of 55%. Our data suggest PSA-TZ to be more reliable for avoiding unnecessary biopsies in patients with PCa suspicion and serum PSA below 15 ng/ml. PSA-TZ, calculated by TRUS, enhances the specificity of PSA for needle biopsy diagnosis of PCa.     

The prevalence of foot ulceration and its correlates in type 2 diabetic patients: a population-based study

The prevalence of peripheral neuropathy, peripheral vascular disease, and foot ulceration in Type 2 diabetic patients in the community were determined in a community-based study. Eight hundred and eleven subjects (404 male, 407 female, mean age 65.4 (range 34-90) years, diabetes duration 7.4 (0-50) years) from 37 general practices in three UK cities were studied. Neuropathy was diagnosed clinically using modified neuropathy disability scores which were ascertained using structured interviews and clinical examinations by one observer in each city. Peripheral vascular disease was diagnosed if a history of revascularization was present or > or = 2 foot pulses were absent. History of current or previous foot ulceration was recorded. The prevalence of neuropathy was 41.6% (95% confidence limits 38.3-44.9%) and the prevalence of PVD, 11% (9.1-13.7%). Forty-eight percent of neuropathic patients reported significant neuropathic symptoms. Forty-three patients (5.3% (3.8-6.8%)) had current or past foot ulcers; 20 of these were pure neuropathic ulcers, 13 neuroischaemic, 5 pure vascular, and 5 were unclassified. Multiple logistic regression showed history of amputation, neuropathy disability score, and peripheral vascular disease to be significantly associated with foot ulceration after adjusting for age and diabetes duration. A substantial proportion of Type 2 diabetic patients, often elderly patients who do not attend hospitals, suffered from peripheral neuropathy and peripheral vascular disease. These patients are at risk of foot ulceration and may benefit from preventive footcare.     

The plant cDNA LCT1 mediates the uptake of calcium and cadmium in yeast

Nonessential metal ions such as cadmium are most likely transported across plant membranes via transporters for essential cations. To identify possible pathways for Cd2+ transport we tested putative plant cation transporters for Cd2+ uptake activity by expressing cDNAs in Saccharomyces cerevisiae and found that expression of one clone, LCT1, renders the growth of yeast more sensitive to cadmium. Ion flux assays showed that Cd2+ sensitivity is correlated with an increase in Cd2+ uptake. LCT1-dependent Cd2+ uptake is saturable, lies in the high-affinity range (apparent KM for Cd2+ = 33 microM) and is sensitive to block by La3+ and Ca2+. Growth assays demonstrated a sensitivity of LCT1-expressing yeast cells to extracellular millimolar Ca2+ concentrations. LCT1-dependent increase in Ca2+ uptake correlated with the observed phenotype. Furthermore, LCT1 complements a yeast disruption mutant in the MID1 gene, a non-LCT1-homologous yeast gene encoding a membrane Ca2+ influx system required for recovery from the mating response. We conclude that LCT1 mediates the uptake of Ca2+ and Cd2+ in yeast and may therefore represent a first plant cDNA encoding a plant Ca2+ uptake or an organellar Ca2+ transport pathway in plants and may contribute to transport of the toxic metal Cd2+ across plant membranes.