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9,178,002
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Cytosine methylation associated with repeat-induced point mutation causes epigenetic gene silencing in Neurospora crassa.
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Repeated DNA sequences are frequently mutated during the sexual cycle in Neurospora crassa by a process named repeat-induced point mutation (RIP). RIP is often associated with methylation of cytosine residues in and around the mutated sequences. Here we demonstrate that this methylation can silence a gene located in nearby, unique sequences. A large proportion of strains that had undergone RIP of a linked duplication flanking a single-copy transgene, hph (hygromycin B phosphotransferase), showed partial silencing of hph. These strains were all heavily methylated throughout the single-copy hph sequences and the flanking sequences. Silencing was alleviated by preventing methylation, either by 5-azacytidine (5AC) treatment or by introduction of a mutation (eth-I) known to reduce intracellular levels of S-adenosylmethionine. Silenced strains exhibited spontaneous reactivation of hph at frequencies of 10(4) to 0.5. Reactivated strains, as well as cells that were treated with 5AC, gave rise to cultures that were hypomethylated and partially hygromycin resistant, indicating that some of the original methylation was propagated by a maintenance mechanism. Gene expression levels were found to be variable within a population of clonally related cells, and this variation was correlated with epigenetically propagated differences in methylation patterns.
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9,178,003
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Isolation and characterization of a temperature-sensitive circadian clock mutant of Neurospora crassa.
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A new circadian clock mutant has been isolated in Neurospora crassa. This new mutation, called period-6 (prd-6), has two features novel to known clock mutations. First, the mutation is temperature sensitive. At restrictive temperatures (above 21 degrees) the mutation shortens circadian period length from a wild-type value of 21.5 hr to 18 hr. At permissive temperatures (below 21 degrees) the mutant has a 20.5-hr period length close to that of the wild-type strain. Second, the prd-6 mutation is epistatic to the previously isolated clock mutation period-2 (prd-2). This epistasis is unusual in that the prd-2 prd-6 double mutant strain has an 18-hr period length at both the restrictive and permissive temperatures. That is, the temperature-sensitive aspect of the phenotype of the prd-6 strain is lost in the prd-2 prd-6 double mutant strain. This suggests that the gene products of the prd-2 and prd-6 loci may interact physically and that the presence of a normal prd-2+ protein is required for low temperature to "rescue" the prd-6 mutant phenotype. These results, combined with our recent finding that prd-2 and some alleles of the frq gene show genetic synergy, suggest that it may be possible to establish a more comprehensive model of the Neurospora circadian clock.
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9,178,004
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A putative rhamnogalacturonase required for sexual development of Neurospora crassa.
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In previous work, the asd-I (ascus development) gene of the filamentous fingus Neurospora crassa was identified as a gene expressed preferentially during the sexual cycle and shown to be essential for normal sexual development. The asd-I gene has been sequenced and further characterized. It contains two introns, the first of which is in-frame and inefficiently or differentially spliced. The predicted ASD-I protein has extensive homology with rhamnogalacturonase B of Aspergillus aculeatus, which cleaves the backbone within the ramified hairy regions of pectin. In homozygous asd-I crosses, sexual development is initiated and large numbers of normal-sized asci are formed. Ascospore delineation does not occur, however, and no sexual progeny are produced. As most asd-I asci contain eight nuclei, the two meiotic divisions and subsequent mitotic division typical of normal crosses seem to occur, but the haploid nuclei are not partitioned into ascospores. In wild-type crosses, the ASD-I protein is present in large amounts in croziers and young asci, but it is only faintly detectable in more mature asci containing developing ascospores. Models to explain the possible role of a rhamnogalacturonase in sexual development are presented.
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9,178,005
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Multiple genes encoding pheromones and a pheromone receptor define the B beta 1 mating-type specificity in Schizophyllum commune.
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The genes defining multiple B mating types in the wood-rotting mushroom Schizophyllum commune are predicted to encode multiple pheromones and pheromone receptors. These genes are clustered in each of two recombinable and independently functioning loci, B alpha and B beta. A difference in specificity at either locus between a mated pair of individuals initiates an identical series of events in sexual morphogenesis. The B alpha 1 locus was recently found to contain genes predicted to encode three lipopeptide pheromones and a pheromone receptor with a seven-transmembrane domain. These gene products interact in hetero-specific pairs, the pheromone of one B alpha specificity with the receptor of any one of the other eight B alpha specificities, and are likely to activate a signaling cascade similar to that known for mating in Saccharomyces cerevisiae. We report here that the B beta 1 locus also contains at least three pheromone genes and one pheromone receptor gene, which function similarly to the genes in the B alpha 1 locus, but only within the series of B beta specificities. A comparison of the DNA sequences of the B alpha 1 and B beta 1 loci suggests that each arose from a common ancestral sequence, allowing us to speculate about the evolution of this unique series of regulatory genes.
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9,178,006
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Mechanism of activation of the Caenorhabditis elegans ras homologue let-60 by a novel, temperature-sensitive, gain-of-function mutation.
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The Caenorhabditis elegans let-60 gene encodes a Ras protein that mediates induction of the hermaphrodite vulva. To better understand how mutations constitutively activate Ras and cause unregulated cell division, we have characterized ga89, a temperature-sensitive, gain-of-function mutation in let-60 ras. At 25 degrees, ga89 increases let-60 activity resulting in a multivulva phenotype. At 15 degrees, ga89 decreases let-60 activity resulting in a vulvaless phenotype in let-60(ga89)/Df animals. The ga89 mutation causes a leucine (L) to phenylalanine (F) substitution at amino acid 19, a residue conserved in all Ras proteins. We introduced the L19F change into human H-Ras protein and found that the in vitro GTPase activity of H-Ras became temperature-dependent. Genetic experiments suggest that LET-60 (L19F) interacts with GAP and GNEF, since mutations that decrease GAP and GNEF activity affect the multivulva phenotype of let-60(ga89) animals. These results suggest that the L19F mutation primarily affects the intrinsic rate of GTP hydrolysis by Ras, and that this effect may be sufficient to account for the activated-Ras phenotype caused by let-60(ga89). Our results suggest that a mutation in a human ras gene analogous to ga89 might contribute to oncogenic transformation.
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9,178,007
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The Caenorhabditis elegans spe-5 gene is required for morphogenesis of a sperm-specific organelle and is associated with an inherent cold-sensitive phenotype.
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The nonrandom segregation of organelles to the appropriate compartment during asymmetric cellular division is observed in many developing systems. Caenorhabditis elegans spermatogenesis is an excellent system to address this issue genetically. The proper progression of spermatogenesis requires specialized intracellular organelles, the fibrous body-membranous organelle complexes (FB-MOs). The FB-MOs play a critical role in cytoplasmic partitioning during the asymmetric cellular division associated with sperm meiosis II. Here we show that spe-5 mutants contain defective, vacuolated FB-MOs and usually arrest spermatogenesis at the spermatocyte stage. Occasionally, spe-5 mutants containing defective FB-MOs will form spermatids that are capable of differentiating into functional spermatozoa. These spe-5 spermatids exhibit an incomplete penetrance for tubulin mis-segregation during the second meiotic division. In addition to morphological and FB-MO segregation defects, all six spe-5 mutants are cold-sensitive, exhibiting a more penetrant sterile phenotype at 16 degrees than 25 degrees. This cold sensitivity could be an inherent property of FB-MO morphogenesis.
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9,178,008
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Insertions of hybrid P elements in the yellow gene of Drosophila cause a large variety of mutant phenotypes.
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A series of yellow mutations associated with a great variety of tissue-specific phenotypes were obtained from several highly unstable Drosophila melanogaster strains carrying the gypsy-induced y2 allele. These mutations are caused by insertion of additional DNA sequences of variable size 69 bp upstream of the yellow transcription start site. These sequences are flanked by identical copies of a deleted 1.2-kb P element arranged in the same or inverted orientation. The central part of the inserted element consists of genomic sequences originating from different regions of the X chromosome. The mutant phenotype caused by these chimeric elements depends on the nature of the sequences present either in the P element or in the central part of the insertion, suggesting that these sequences are able to affect expression of the yellow gene. In addition, sequences present in the central region of the insertions strongly modify the effects of the gypsy-bound suppressor of Hairy-wing [su(Hw)] and modifier of mdg4 [mod(mdg4)] proteins on yellow transcription. Analyses of these mutations give new insights into the mechanisms by which su (Hw) and mod(mdg4) affect enhancer function.
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9,178,009
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The CRE-binding protein dCREB-A is required for Drosophila embryonic development.
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We have previously described the cloning of a cyclic AMP response-element (CRE)-binding protein, dCREB-A, in Drosophila melanogaster that is similar to the mammalian CRE-binding protein CREB. dCREB-A is a member of the bZIP family of transcription factors, shows specific binding to the (CRE), and can activate transcription in cell culture. In this report, we describe the gene structure for dCREB-A, protein expression patterns throughout development and the necessary role for this gene in embryogenesis. The 4.5-kb transcript is encoded in six exons that are distributed over 21 kb of DNA. There are seven start sites and no TATA consensus sequences upstream. The dCREB-A protein is expressed in the nuclei of the embryonic salivary gland, proventriculus and stomadeum. Late in embryogenesis, tracheal cell nuclei and specific nuclei within the segments show staining with anti-dCREB-A antibodies. In adult female ovaries, dCREB-A is expressed in the stage 9 through stage 11 follicle cell nuclei. Null mutations of the dCREB-A gene give rise to animals that no longer express dCREB-A protein and die late in embryogenesis before or at hatching. The absolute requirement of dCREB-A for embryogenesis demonstrates a nonredundant function for a CRE-binding protein that will be useful in studying the role of specific signal transduction cascades in development.
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9,178,010
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Genetic and molecular analysis of smooth, a quantitative trait locus affecting bristle number in Drosophila melanogaster.
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A semi-lethal, sterile allele of the smooth locus (2-91.5), sm3, was discovered in an artificial selection line for low abdominal bristle number that had been started from a P-M dysgenic cross. The fitness effects and extremely low bristle number phenotype of the allele could not be separated by recombination from a P-element insertion at cytological location 56E, and precise excision of the P-element at this site was associated with reversion to wild type. The smooth gene was cloned using the P-element insertion as a tag. The gene encodes a 2.6-kb transcript derived from 10 exons and covers a genomic region of at least 80 kb. The Drosophila smooth gene shares substantial sequence identity with a group of RNA binding proteins, with the closest relationship being to the human heterogeneous nuclear ribonucleoprotein L gene. The smooth gene is by definition an abdominal bristle number quantitative trait locus, but further work is required to discern whether naturally occurring allelic variation at this locus is a source of genetic variation for abdominal bristle number in natural populations.
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9,178,011
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Ras1-mediated modulation of Drosophila homeotic function in cell and segment identity.
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Mutations of the Drosophila homeotic proboscipedia gene (pb, the Hox-A2/B2 homologue) provoke dose-sensitive defects. These were used to search for dose-sensitive dominant modifiers of pb function. Two identified interacting genes were the proto-oncogene Ras1 and its functional antagonist Gap1, prominent intermediaries in known signal transduction pathways. Ras1+ is a positive modifier of pb activity both in normal and ectopic cell contexts, while the Ras1-antagonist Gap1 has an opposite effect. A general role for Ras1 in homeotic function is likely, since Ras1+ activity also modulates functions of the homeotic loci Sex combs reduced and Ultrabithorax. Our data suggest that the modulation occurs by a mechanism independent of transcriptional control of the homeotic loci themselves, or of the Ras1/Gap1 genes. Taken together our data support a role for Ras1-mediated cell signaling in the homeotic control of segmental differentiation.
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9,178,012
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Altering developmental trajectories in mice by restricted index selection.
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A restricted index selection experiment on mice was carried out for 1-4 generations on rate of early postnatal development (growth rate from birth to 10 days of age) vs. rate of development much later in ontogeny (growth rate from 28 to 56 days of age). Early rate of development (E) approximates hyperplasia (changes in cell number) and later rate (L) reflects hypertropy (changes in cell size). The selection criteria were as follows; E+LO was selected to increase early body weight gain while holding late body weight gain constant; E-LO was selected to decrease early body gain while holding late gain constant; EOL+ was selected to increase late gain holding early gain constant; and EOL- was selected to decrease late gain holding early gain constant. After 14 generations of selection, significant divergence among lines has occurred and the changes in the growth trajectories are very close to expectation. The genetic and developmental bases of complex traits are discussed as well as the concept of developmental homoplasy.
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9,178,013
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Recombination creates novel L1 (LINE-1) elements in Rattus norvegicus.
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Mammalian L1 (long interspersed repeated DNA. LINE-1) retrotransposons consist of a 5' untranslated region (UTR) with regulatory properties, two protein encoding regions (ORF I, ORF II, which encodes a reverse transcriptase) and a 3' UTR. L1 elements have been evolving in mammals for > 100 million years and this process continues to generate novel L1 subfamilies in modern species. Here we characterized the youngest known subfamily in Rattus norvegicus, L1mlvi2, and unexpectedly found that this element has a dual ancestry. While its 3' UTR shares the same lineage as its nearest chronologically antecedent subfamilies, L13 and L14, its ORF I sequence does not. The L1mlvi2 ORF I was derived from an ancestral ORF I sequence that was the evolutionary precursor of the L13 and L14 ORF I. We suggest that an ancestral ORF I sequence was recruited into the modern L1mlvi2 subfamily by recombination that possibly could have resulted from template strand switching by the reverse transcriptase during L1 replication. This mechanism could also account for some of the structural features of rodent L1 5' UTR and ORF I sequences including one of the more dramatic features of L1 evolution in mammals, namely the repeated acquisition of novel 5' UTRs.
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9,178,014
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Ancestral polymorphism of Mhc class II genes in mice: implications for balancing selection and the mammalian molecular clock.
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To investigate the evolutionary dynamics at Mhc class II DR genes of mice (genus Mus), we sequenced the peptide binding regions (PBRs) of 41 DRB (= E beta) genes and eight DRA (= E alpha) genes from 15 strains representing eight species. As expected trees of these PBR sequences imply extensive maintenance of ancestral DRB alleles across species. We use a coalescent simulation model to show that the number of interspecific coalescent events (c) observed on these trees was higher than the number expected for neutral genealogies and similar sample sizes and is more consistent with balancing selection that with neutrality. Patterns of ancestral polymorphism in mouse DRB alleles were also used to examine the tempo of synonymous substitution in the PBR of mouse class II genes. Both absolute and relative rate tests on DRA and DRB genes imply increased substitution rates at two- and fourfold degenerate sites of mice and rats relative to primates, and decreased rates for the DRB genes of primates relative to ungulate and carnivore relatives. Thus rates of synonymous substitution at Mhc DR genes in mammals appear to be subject to generation time effects in ways similar to those found at other mammalian genes.
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9,178,015
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Morph-specific proteins in pollen and styles of distylous turnera (Turneraceae).
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We used nondenaturing isoelectric focusing (IEF) in a survey of plants from 11 populations to identify style and pollen proteins unique to the short-styled morph of Turnera scabra, T. sulnitata and T. krapovickasii. Three protein bands [approximately isoelectric points (pls) 6.1, 6.3 and 6.5] were found only in styles and stigmas of short-styled plants while two bands (approximately pls 6.7 and 6.8, M(r) 56 and 59 kD) occur only in pollen of short-styled plants. Some of these bands appear very late in development, within 24 hr before flowering. Two isozyme loci were mapped to an 8.7 cM region spanning the distyly locus. Using these isozyme markers we identified progeny exhibiting recombination adjacent to the distyly locus. No recombinants between the distyly locus and the locus or loci controlling the presence of the short-styled morph-specific proteins were obtained. This suggest that the loci encoding these proteins are either extremely tightly linked to the distyly locus and in complete disequilibrium with the S allele or exhibit morph-limited expression. Crosses to a plant showing an unusual style protein phenotype demonstrated that an additional unlinked locus is required for full expression of the style proteins. The function of the morph-specific proteins is unknown.
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9,178,016
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Insertions of a novel class of transposable elements with a strong target site preference at the r locus of maize.
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The r locus of maize regulates anthocyanin synthesis in various tissues of maize through the production of helix-loop-helix DNA binding proteins capable of inducing expression of structural genes in the anthocyanin biosynthetic pathway. The complex r variant, R-r: standard (R.r), undergoes frequent mutation through a variety of mechanisms including displaced synapsis and crossing over, and intrachromosomal recombination. Here we report a new mechanism for mutation at the R-r complex: insertion of a novel family of transposable elements. Because the elements were first identified in the R-p gene of the R-r complex, they have been named P instability Factor (PIF). Two different PIF elements were cloned and found to have identical sequences at their termini but divergent internal sequences. In addition, the PIF elements showed a marked specificity of insertion sites. Six out of seven PIF-containing derivatives examined had an element inserted at an identical location. Two different members of the PIF element family were identified at this position. The seventh PIF-containing derivative examined had the element inserted at a distinct position within r. Even at this location, however, the element inserted into a conserved target sequence. The timing of PIF excision is unusual. Germinal excision rates can range up to several percent of progeny. Yet somatic sectors are rare, even in lines exhibiting high germinal reversion rates.
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9,178,017
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Recombination and gene flux caused by gene conversion and crossing over in inversion heterokaryotypes.
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A theoretical analysis of the effects of inversions on recombination and gene flux between arrangements caused by gene conversion and crossing over was carried out. Two different mathematical models of recombination were used: the Poisson model (without interference) and the Counting model (with interference). The main results are as follows. (1) Recombination and gene flux are highly site-dependent both inside and outside the inverted regions. (2) Crossing over overwhelms gene conversion as a cause of gene flux in large inversions, while conversion becomes relatively significant in short inversions and in regions around the breakpoints. (3) Under the Counting model the recombination rate between two markers depends strongly on the position of the markers along the inverted segment. Two equally spaced markers in the central part of the inverted segment have less recombination than if they are in a more extreme position. (4) Inversions affect recombination rates in the univerted regions of the chromosome. Recombination increases in the distal segment and decreases in the proximal segment. These results provide an explanation for a number of observations reported in the literature. Because inversions are ubiquitous in the evolutionary history of many Drosophila species, the effects of inversions on recombination are expected to influence DNA variation patterns.
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9,178,018
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A likelihood approach to populations samples of microsatellite alleles.
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This paper presents a likelihood approach to population samples of microsatellite alleles. A Markov chain recursion method previously published by GRIFFITHS and TAVARE is applied to estimate the likelihood function under different models of microsatellite evolution. The method presented can be applied to estimate a fundamental population genetics parameter theta as well as parameters of the mutational model. The new likelihood estimator provides a better estimator of theta in terms of the mean square error than previous approaches. Furthermore, it is demonstrated how the method may easily be applied to test models of microsatellite evolution. In particular it is shown how to compare a one-step model of microsatellite evolution to a multi-step model by a likelihood ratio test.
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9,178,020
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The probability of fixation in populations of changing size.
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The rate of adaptive evolution of a population ultimately depends on the rate of incorporation of beneficial mutations. Even beneficial mutations may, however, be lost from a population since mutant individuals may, by chance, fail to reproduce. In this paper, we calculate the probability of fixation of beneficial mutations that occur in populations of changing size. We examine a number of demographic models, including a population whose size changes once, a population experiencing exponential growth or decline, one that is experiencing logistic growth or decline, and a population that fluctuates in size. The results are based on a branching process model but are shown to be approximate solutions to the diffusion equation describing changes in the probability of fixation over time. Using the diffusion equation, the probability of fixation of deleterious alleles can also be determined for populations that are changing in size. The results developed in this paper can be used to estimate the fixation flux, defined as the rate at which beneficial alleles fix within a population. The fixation flux measures the rate of adaptive evolution of a population and, as we shall see, depends strongly on changes that occur in population size.
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9,178,019
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Conditions for protected inversion polymorphism under supergene selection.
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Conditions for protected inversion polymorphism under the operation of both karyotype and supergene selection in a viability model have been analytically determined. When supergene selection (the effect of recombination in homokaryotypes lowering the mean fitness of their offspring) is acting on gene arrangements and there is no karyotype selection, it is demonstrated that a polymorphic stable equilibrium is reached by the population, which is a function of only the recombination effects in homokaryotypes. Under both supergene and karyotype selection the degree of dominance (h) of karyotype selection is critical to produce a protected inversion polymorphism. In general, the opportunity for protected polymorphism increases as the degree of dominance decreases. For small s values, the conditions for protected polymorphism are r > 2sh and c > 2s(h-1), where r and c are the average loss of viability for offspring of ST/ST and IN/IN homokaryotypes, respectively. These findings suggest that supergene selection may be an important balancing mechanism contributing to the maintenance of inversion polymorphism.
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9,178,021
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Mapping-linked quantitative trait loci using Bayesian analysis and Markov chain Monte Carlo algorithms.
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A Bayesian method for mapping linked quantitative trait loci (QTL) using multiple linked genetic markers is presented. Parameter estimation and hypothesis testing was implemented via Markov chain Monte Carlo (MCMC) algorithms. Parameters included were allele frequencies and substitution effects for two biallelic QTL, map positions of the QTL, and markers, allele frequencies of the markers, and polygenic and residual variances. Missing data were polygenic effects and multi-locus marker-QTL genotypes. Three different MCMC schemes for testing the presence of a single or two linked QTL on the chromosome were compared. The first approach includes a model indicator variable representing two unlinked QTL, affecting the trait, one linked and one unlinked QTL, or both QTL linked with the markers. The second approach incorporates an indicator variable for each QTL into the model for phenotype, allowing or not allowing for a substitution effect of a QTL, on phenotype, and the third approach is based on model determination by reversible jump MCMC. Methods were evaluated empirically by analyzing simulated granddaughter designs. All methods identified correctly a second, linked QTL and did not reject the one-QTL model when there was only a single QTL, and no additional or an unlinked QTL.
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9,178,022
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Time trends in obesity: an epidemiological perspective.
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The average prevalence of obesity (BMI > 30 kg/m2) among European centers participating in the WHO-MONICA study between 1983 and 1986 was about 15% in men and 22% in women Prevalence figures ranged in men from 7% in Gothenburg and 22% in Lithuania and in women from 9% to 45% in the same places. Some monitoring projects or repeated surveys suggest that the prevalence of obesity has been increasing during the past 15 years in some European countries. A closer look at data from The Netherlands suggest that average weight increase in the order of about 1 kilo can be responsible for quite dramatic increases in the prevalence of obesity. This suggest that only small changes in the daily caloric balance may be sufficient to increase the number of obese subjects in populations. In The Netherlands a decrease in energy intake and fat consumption was observed between 1987 and 1993 and smoking rates remained relatively stable. This could imply that reductions in energy expenditure are the main factors responsible for the increase in the prevalence of obesity. Since the increase in the prevalence of obesity seems to occur particularly in younger age-groups, the consequences of the increase in the prevalence of obesity only become apparent many years later. Especially chronic conditions such as arthritis or conditions related to obesity but occurring later in life such as cerebrovascular accidents, chronic heart failure or breast cancer in women. The rising prevalence of non-insulin dependent diabetes mellitus may be one of the first signs of the increasing problem of obesity in European countries.
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9,178,023
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Hormonal influence on lipid-protein interactions in biological membranes. Lactation effects on alkaline phosphatase activity and intestinal brush border membrane properties in rat.
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It is known that prolactin modifies the fluidity of different biological membranes in rats and that the activity of intestinal alkaline phosphatase varies directly with the fluidity of the membranes in which it is found. Our objective was to study the intestinal alkaline phosphatase (IAP) activity, lipid composition and fluidity of the proximal small intestine brush border membranes under the influence of physiological high levels of prolactin, in rats with 15 days of lactating (Dams 15 days) compared with control virgin rats. The phenomenon was corroborated in dams from which the suckling pups had been withdrawn on the tenth day of lactation (Dams 10 days). The results showed a decrease on the IAP activity in dams in lactation with relation to control virgin and dams with withdrawal of pups. We found decreases in total phospholipids contents and fluidity and an increase in the microviscosity lipid membrane in dams with 15 days of lactating compared to virgins. In the same groups there were no differences in total lipids content and no modifications were observed in the quantity of total cholesterol and proteins. These results suggest that the changes produced by lactation could be one of the causes of alteration of brush border membranes properties by modifying the lipid-protein interactions and the alkaline phosphatase activity in the proximal small intestine.
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9,178,024
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The lipolytic effects of thyrotropin and isoprenaline are not affected by growth hormone in infant adipocytes.
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The lipolytic effects of growth hormone (GH) in children are not fully clarified. In this study, no lipolytic effect of GH on isolated adipocytes obtained at surgery on healthy infants aged 2-5 months and children 3-6 years was observed. Furthermore, GH did not enhance isoprenaline-induced lipolysis, as in adults. The TSH-induced lipolysis which is prominent in neonatal adipocytes was not affected by incubation of adipocytes with GH. Assuming that GH alters adipocyte metabolism primarily by increasing the sensitivity, but not the maximum response, of the beta 2-adrenergic receptor population, it follows that GH, in this sense, should be a less important co-actor in children where beta 2-adrenergic receptors are more abundant.
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9,178,025
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Increased in vitro interleukin-12 production by peripheral blood in high-risk IDDM first degree relatives.
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Cytokines secreted by antigen presenting cells, lymphocytes T and pancreatic beta cells are considered as the major mediators in the pathogenesis of IDDM. It has been suggested that cytokines released by macrophages/monocytes could have an initial role in beta-cell damage. The aim of the present study was the estimation of in vitro production of macrophage-derived cytokines: IL-1 beta, TNF-alpha, IL-12 by peripheral blood in high risk IDDM first degree relatives, since it could reflect early events leading to the development of type 1 diabetes in humans. The study was performed in 25 high risk IDDM subjects and 21 age and sex-matched healthy controls. IL-1 beta, TNF-alpha and IL-12 concentrations in supernatants of whole blood cultures with PHA (10 micrograms/ml) were quantified by ELISA. In the ICA positive relatives of IDDM subjects levels of IL-12 were significantly higher as compared with the control group, both at 48 h (p < 0.02) and at 72 h (p < 0.05) of incubation and positively correlated with TNF-alpha and IL-1 beta (R = 0.46, p < 0.02 and R = 0.32, p < 0.05). We did not observe statistical differences in in vitro production of TNF-alpha and IL-1 beta between the study groups. In conclusion we suggest that our findings support the hypothesis, that IL-12 is involved in the pathogenesis of human IDDM. If the involvement of Th1 cells is confirmed in the destruction of islet beta-cells, it is possible that IL-12 antagonists will have a role in the future prevention of insulin dependent diabetes mellitus.
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9,178,026
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Predictors of change in insulin sensitivity during glucocorticoid treatment.
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Glucocorticoid induced alterations in carbohydrate metabolism can result in hyperglycemia. We evaluated changes in carbohydrate metabolism produced by four days of prednisone (20 mg PO TID) measuring insulin sensitivity, basal glucose, basal insulin and first phase insulin release (FPIR). We correlated these measures of carbohydrate metabolism with changes in free fatty acids and lactate levels both of which have been reported to be possible mediators of insulin sensitivity. Insulin sensitivity decreased by 64% (p = 0.002), basal insulin levels increased 50% (p = 0.026), FPIR tripled (p = 0.064) while fasting glucose levels increased significantly but remained normal. Basal FFAs levels increased (p = 0.045) while lactate levels did not change significantly, and neither predicted changes in SI. Basal levels of SI and FPIR were found to be independent predictors of change in insulin sensitivity and together explained 83% of the change in insulin sensitivity produced by short term treatment with prednisone.
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9,178,027
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Cholesterol 7 alpha-hydroxylase activity in hypothyroidism and hyperthyroidism in humans.
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Alterations of serum cholesterol levels are well recognized findings in hypothyroidism and hyperthyroidism. It remains unclear, whether thyroid hormones may affect serum concentrations of cholesterol through changes in the activity of cholesterol 7 alpha-hydroxylase, the rate-limiting enzyme in the catabolic conversion of cholesterol to bile acids. We determined serum concentrations of the bile acid precursor 7 alpha-hydroxy-4-cholesten-3-one, which reflects cholesterol 7 alpha-hydroxylase activity in the liver, in 19 patients with hypothyroidism and in 10 patients with hyperthyroidism before and after treatment, respectively. In patients with hypothyroidism, serum concentrations of cholesterol and LDL-cholesterol decreased by 33% (p < 0.0005) and 39% (p < 0.0005), respectively, after replacement therapy with thyroid hormones. In contrast, serum concentrations of 7 alpha-hydroxy-4-cholesten-3-one (21.7 +/- 15.8 ng/ml vs 24.5 +/- 18.1 ng/ml before treatment, n.s.) as well as serum HDL-cholesterol were unchanged during substitution therapy. In patients with hyperthyroidism, serum concentrations of cholesterol and LDL-cholesterol increased by 27% (p < 0.01) and 39% (p < 0.01) after antithyroid treatment, respectively. Again, serum concentrations of 7 alpha-hydroxy-4-cholesten-3-one did not change significantly during treatment (15.8 +/- 12.6 ng/ml vs 14.7 +/- 8.1 ng/ml before treatment, n.s.). These findings indicate that in humans, thyroid hormones influence serum lipid concentrations by other mechanisms than by affecting the activity of cholesterol 7 alpha-hydroxylase.
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9,178,028
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Repeated administration of growth hormone-releasing hormone with or without previous administration of pyridostigmine in insulin-dependent diabetes mellitus.
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In insulin dependent diabetes mellitus (IDDM) either elevated growth hormone (GH) levels or abnormal responses to specific as well as unspecific stimuli have been reported. As hyperglycemia is known to blunt GH response to various stimuli, a normal GH response to GHRH in presence of hyperglycemia should also be considered inappropriate. To investigate the mechanism underlying this inappropriate GH response, in 9 patients with IDDM, selected for normal GH response to GHRH, we studied the GH response to two consecutive GHRH boluses (1 microgram/kg), the second of which preceded 30 min before by pyridostigmine (120 mg p.o.). Seven age matched normal volunteers were evaluated as control group. Basal plasma glucose and serum GH levels were significantly higher in patients with IDDM than in normal subjects (184.4 +/- 9.6 vs 86.2 +/- 4.4 mg/dl, p < 0.01 and 2.4 +/- 1.0 vs 1.0 +/- 0.4 microgram/l, p < 0.01 respectively). Both in normal subjects and in patients with IDDM the GH response to the second consecutive GHRH administration was lower than that of the first GHRH bolus (delta AUC: 82.5 +/- 28.3 vs 401.1 +/- 131.2 micrograms/l/h, p < 0.05 and 77.2 +/- 30.4 vs 336.8 +/- 60.0 p < 0.02, respectively). Pyridostigmine was able to restore the blunted GH responsiveness to the second GHRH administration in both groups, but this response was found higher in normal than in diabetic subjects (delta AUC: 1250.8 +/- 136.2 vs 527.5 +/- 147.6, p < 0.01). Since the GH-releasing effect of PD is likely to be mediated by the inhibition of hypothalamic somatostatin release, our results suggest that there is also an impaired somatostatin tone in hyperglycemic type 1 diabetic patients with normal GH response to GHRH.
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9,178,029
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Immunoreactive growth hormone-releasing hormone (IR-GHRH) in the feto-placental circulation and differential effects of L-dopa, L-arginine and somatostatin-14 on the plasma levels of IR-GHRH in normal adults.
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The relation of the physiological releases of growth hormone-releasing hormone (GHRH) and growth hormone (GH) into the circulation in various conditions was investigated using a sensitive and specific radioimmunoassay for plasma GHRH. The mean fasting plasma level of immunoreactive (IR)-GHRH in 72 normal adults was 10.3 +/- 0.5 (mean +/- SEM) pg/ml and there was no significant sex difference in the level. The concentrations of IR-GHRH in plasma from the umbilical artery and umbilical vein were 107.3 +/- 20.5 pg/ml and 33.6 +/- 3.8 pg/ml, respectively, and a marked arterio-venous gradient was observed in all 12 individuals examined. The plasma level of IR-GHRH in the maternal vein was significantly lower than that in the cord blood, but was similar to that in non-pregnant women. In normal adults, although there was no apparent fluctuation in the level of plasma IR-GHRH or of plasma GH during bed rest, a significant increase of plasma IR-GHRH was detected followed by, or synchronized with the surge of plasma GH after oral administration of L-dopa. In contrast, on L-arginine infusion, no proportional elevation of plasma IR-GHRH with increase in plasma GH was observed. During and after intravenous infusion of somatostatin, the circulating IR-GHRH level did not increase, but on stopping the infusion there was an immediate and marked rebound surge of GH. We conclude that 1) the elevated IR-GHRH in the cord blood plasma originates from the fetus and may have a primary role in enhancing secretion of GH which promotes growth in early human life, and 2) the participations of GHRH in the mechanisms of GH secretion seen after administrations of L-dopa, L-arginine and somatostatin are different.
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9,178,033
|
Behavioral symptoms in adult rats after postnatal L-nitro-arginine.
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We addressed experimentally the suggestion by Gally et al. [Gally J. A., Read Montague P., Reeke G. N. Jr and Edelman G. M. (1990) Proc. Natl Acad. Sci. U.S.A. 87, 3547-3551] that nitric oxide may play a role in the use-dependent modification of synaptic efficacy in the developing nervous system. In a preliminary control experiment, we treated rat pups from postnatal day 8 to postnatal day 22 with a nitric oxide synthase blocker (L-nitro-arginine) and compared their growth curves and brain weights to those of saline injected control pubs. No significant differences were found after the 14 days of nitric oxide synthase inhibition. In the subsequent experiment, we inhibited nitric oxide synthesis in rat pups from postnatal day 8 to day 29 and assessed their place learning ability and open field behavior as adults. We found an increased speed of habituation of locomotion in an open field in 5-month-old rats that had been treated postnatally with a nitric oxide synthase blocker. There was no difference between treated and non-treated rats with respect to place learning in a water maze. We conclude that perturbation of nitric oxide production during early postnatal development does not preclude normal learning and memory function in the adult.
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9,178,034
|
Weaver mutant mouse cerebellar granule cells respond normally to chronic depolarization.
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We studied the effects of chronic K(+)-induced membrane depolarization and treatment with N-methyl-D-aspartate (NMDA) on cerebellar granule cells (CGCs) from weaver mutant mice and non-weaver litter-mates. The weaver mutation is a Gly-to-Ser substitution in a conserved region of the Girk2 G protein-coupled inward rectifying potassium channel [Patil N., Cox D. R., Bhat D., Faham M., Myers R. M. and Peterson A. S. (1995) Nature Genet. 11, 126-129] which induces early death of CGCs. The biochemical differentiation of CGCs was estimated as the rate of 2-deoxy-D-glucose accumulation and the expression of neural cell adhesion molecule (NCAM). High (25 mM) K+ ion concentration or treatment with NMDA greatly promoted the biochemical differentiation of both weaver mutant and non-weaver litter-mate mouse CGCs. In contrast to the marked effect on biochemical differentiation in both weaver and non-weaver mice CGSs, chronic high K+ treatment only had limited effect on survival. The survival of weaver mutant mouse CGCs in medium containing 5 mM K+ ions was very low, only 20% of the plated cells surviving at 7 days after plating, as opposed to the 50% for non-weaver CGCs. Chronic high K+ treatment improved the relative survival of weaver mutant mouse CGCs 1.6 2.2-fold and that of non-weaver CGCs 1.2-1.4-fold; the same number of CGCs (about 20% of the plated cells) were rescued by high K+ in both types of culture. The findings indicate that, in culture weaver mutant mouse, CGCs have a normal response to membrane depolarization and that the normal function of the Girk2 potassium channel is not critical for the survival of differentiated CGCs.
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9,178,035
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Enhanced GABA release in cell-damaging conditions in the adult and developing mouse hippocampus.
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The release of [3H]GABA from hippocampal slices from adult (3-month-old) and developing (7-day-old) mice was studied in cell-damaging conditions in vitro using a superfusion system. Cell damage was induced by modified superfusion media, including hypoxia, hypoglycemia, ischemia, the presence of Free radicals and oxidative stress. The basal release of GABA from the immature and mature hippocampus was generally markedly increased in all cell-damaging conditions. In 7-day-old mice the release was enhanced most in the presence of free radicals. 1.0 mM NaCN and ischemia, whereas in the adults 1.0 mM NaCN provoked the largest release of GABA, followed by ischemia and free radical-containing media. Potassium stimulation (50 mM K+) was still able to potentiate the release in all cell-damaging conditions in both age groups. It was shown by superfusing the slices in Ca- and Na-free media that ischemia-induced GABA release was Ca-independent, occurring by a reversed operation of Na-dependent cell membrane carriers in both adult and developing hippocampus. Glutamate and its receptor agonists, N-methyl-D-aspartate (NMDA), kainate and 2-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), potentiated GABA release only in the immature hippocampus by a receptor-mediated mechanism. The enhancement by kainate and AMPA receptors also operated under ischemic conditions. The massive amount of GABA released simultaneously with excitatory amino acids in the mature and immature hippocampus may be an important protective mechanism against excitotoxicity, counteracting harmful effects that lead to neuronal death. The GABA release induced by activation of presynaptic glutamate receptors may contribute particularly to the maintenance of homeostasis in the hippocampus upon impending hyperexcitation.
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9,178,036
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Induction of c-fos mRNA by lead in PC 12 cells.
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Addition of lead acetate to PC 12 pheochromocytoma cells elicits induction of c-fos, an immediate early response gene. Induction of c-fos was concentration- and time-dependent: the lowest concentration of lead acetate tested that induced c-fos was 10 microM; induction was observed after a 30 min incubation and remained high after 90 min. Treatment with lead acetate and cycloheximide superinduced c-fos mRNA. Actinomycin D, an inhibitor of mRNA transcription, decreased the level of c-fos mRNA induced by lead acetate by almost 80%. Cadmium chloride and zinc chloride did not induce c-fos mRNA. Since the c-fos gene encodes a transcription factor, Pb2+ has the potential to deregulate the expression of other genes.
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9,178,038
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Localisation and expression of metallothionein immunoreactivity in the developing sheep brain.
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Metallothioneins are small cysteine-rich proteins that bind heavy metals. In higher mammals there are complex families of metallothionein isoforms, which are well characterised at the DNA level but less so in terms of their cellular expression and function. In particular, little is known about the localisation of metallothionein in the developing mammalian brain. In this study using sheep fetuses, we have shown that metallothionein 1 and 2 isoform expression undergoes shifts in regional and cellular localisation during development of the brain. Metallothionein 1 and 2 expression is first detected by embryonic days E72 E73 (gestation is 150 days) at the mRNA level and the metallothionein protein is observed in cells of the proliferating ventricular zones. Subsequent expression is detected in radial glial cells, oligodendrocytes and astrocytes in several regions of the brain, most notably the cerebral cortex. In the adult brain, metallothionein is expressed in astrocytes but not in oligodendrocytes. Double-labelling immunohistochemistry using the glial fibrillary acidic protein (GFAP) an astrocyte marker, and metallothionein revealed that although there is an overlap in the profiles of the two proteins, there is no simple correlation in their expression. These observations are consistent with metallothionein, under physiological conditions, being regulated mainly by intracellular factors.
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9,178,037
|
A comparative study of the distribution of alpha- and gamma-enolase subunits in cultured rat neural cells and fibroblasts.
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We report the presence and distribution of alpha (ubiquitous) and gamma (neuron-specific) subunits of the dimeric glycolytic enzyme enolase (2-phospho-D-glycerate hydrolase) in cultured neural cells. The gamma gamma enolase is found in vivo at high levels only in neurons and neuroendocrine cells. Neuronal cells in culture also contain relatively high levels of alpha gamma and gamma gamma enolase. Here we show, by enzymatic and immunological techniques, that the gamma subunit also is expressed in cultured rat astrocytes and meningeal fibroblasts and, as we previously reported, in oligodendrocytes. Both neuron-specific isoforms alpha gamma and gamma gamma are expressed in all these cells, but the alpha alpha isoform accounts for the major part of total enolase activity. The sum of alpha gamma and gamma gamma enolase activities increases with time in culture. i.e. maturation processes, reaching the highest level in oligodendrocytes (40% of total enolase activity) and 15 and 10% of total enzymatic activity in astrocytes and fibroblasts, respectively. The gamma enolase transcripts were found not only in cultured neuronal cells but also in cultured oligodendrocytes astrocytes, and meningeal fibroblasts. Our data indicate that neuron-specific enolase should be used with caution as a specific marker for neuronal cell differentiation.
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9,178,039
|
Properties of developing lateral geniculate neurones in the mouse.
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This study describes the properties of neurones recorded in vitro from the dorsal lateral geniculate nucleus (dLGN) of the mouse between developmental stages E16 and P36 and represents the first systematic study of the development of rodent thalamic neurones. The results demonstrate that thalamo-cortical neurones in the mouse dLGN undergo a series of important changes as they mature. Prenatally recorded cells had low resting potentials and could not generate action potentials but as they mature, mouse dLGN neurones become more polarised and show an increase in membrane time constant and spike threshold, while action potentials increase in amplitude and decrease in width. The low-threshold spike (LTS) complex appears at the time of birth, but does not show properties typical of adult cells until at least the third postnatal week. Immature action potentials are primarily sodium-dependent but gain a significant calcium component in the second postnatal week, which is associated with a supra-threshold oscillation of the membrane potential. The electrical activity during this critical period is strongly influenced by the interaction of powerful inward and outward rectification with calcium conductances which determines the appearance of voltage responses to intracellular current injection. The membrane potential in recordings from neurones during the first postnatal week was dominated by intense TTX-sensitive depolarising synaptic-like events which attained amplitudes of 60 mV in several neurones at stages P5-P8. These changes are discussed in relationship to the formation of appropriate connections in the developing visual system.
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9,178,040
|
Development of vulnerability to hypoxic damage in in vitro hippocampal neurons.
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We investigated the relationship between sensitivity to hypoxia and culture age in in vitro hippocampal neurons. Hypoxia was induced by 24 hr incubation in an oxygen-free environment. Up to 6 days in vitro (DIV) mortality was very low or negligible, with few exceptions. Starting at 7 DIV, significant mortality began to be observed; in the age range 7 10 DIV, mortality of 50% or more was observed in five out of 11 experiments (45%) and average mortality was 51 +/- 15% (mean +/- standard deviation, N = 11). In older (12 18 DIV) cultures, mortality of 50% or more was the rule (13 out of 13 experiments) and average mortality was 83 +/- 16% (mean +/- standard deviation, N = 13). The data could be fitted by a sigmoid line (r = 0.87, P < 10(-6) in which 50% mortality corresponds to 8.6 DIV. The N-methyl-D-aspartate antagonist amino-phosphono-valerate and the nitric oxide synthase inhibitor nitroarginine both provided protection. Degree of protection was comparable for the two compounds, but was not observed in cultures younger than approximately 7 DIV. By contrast exogenous creatine was not protective, at variance with findings from other models. The data represent the first description of how sensitivity to hypoxic damage varies during the lifetime of an in vitro neuronal hippocampal culture. Moreover, they suggest the hypothesis that some maturational changes occurring at 79 days in vitro may make previously resistant in vitro neurons significantly sensitive to hypoxic damage, and that at least some of these changes may reflect the development of N-methyl-D-aspartate-mediated glutamatergic transmission.
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9,178,041
|
Implantation of cultured human leptomeningeal cells into rat brain.
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Since previous studies have shown that cells cultured from human leptomeninges can express neuronal and glial antigens under appropriate culture conditions [DeGiorgio L. A. et al. (1994) J. Neurol. Sci. 124, 141 148; Bernstein J. J. et al. (1996) Int. J. Derl Neurosci. 14(5), 681 687], we have studied the developmental characteristics of these cells further by grafting them into young adult rat brains. Cells were labeled in culture with Fast Blue and were identified unequivocally by hybridization with nick-translated human DNA. Intensely Fast Blue positive human leptomeningeal cells were concentrated in the implant pocket and adjacent rat leptomeninges al one and two weeks postimplant. Human and rat leptomeningeal cells were similar morphologically and were equally immunopositive for vimentin and fibronectin. Implanted human cells did not express the neuronal and glial proteins they had in vitro. Cells which hybridized with human DNA corresponded to the intensely Fast Blue positive cells. Small groups of human DNA hybridizing cells were also observed in the choroid plexus. Less intensely Fast Blue positive neurons and glia were found in the brain but these hybridized with rat DNA. A minority of human leptomeningeal cells implanted into rat brain are subsequently found in host leptomeninges where they demonstrate properties characteristic of leptomeningeal fibroblasts. Small numbers of implanted cells can survive for two weeks.
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9,178,042
|
Developmentally regulated release of intraretinal neurotrophic factors in vitro.
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The effects of conditioned media either from aggregates or from explants of embryonic chick retinae and of recombinant neurotrophins were tested upon the survival in vitro of ganglion cells in dissociated cell cultures from the retina of newborn rats. Ganglion cells were identified by the detection of retrogradely transported horseradish peroxidase injected bilaterally into the superior colliculus. Conditioned media increased significantly the survival of ganglion cells after 2 days in culture, at a wide range of plating densities, and had no effect upon adhesion of rat retinal cells. Media conditioned by cell ensembles from chick retinae from embryonic day 8 (E8) to E16 had neurotrophic effects. Release of neurotrophic activity peaked at E10 E12, irrespective of the numbers of cells or total concentration of protein in the conditioned media. The active molecules were non-dialyzable and were released either in the presence or in the absence of fetal calf serum. The neurotrophic activity was abolished by trypsinization, and recovered by salting-out with 25 75% ammonium sulfate. NT-4, BDNF and, to a lesser extent, NT-3, increased the survival of ganglion cells in our assay, while NGF had no effect. The data show that chick retinal cells release soluble trophic proteins according to a developmentally regulated pattern. These neurotrophic factors may be involved in local competitive interactions that help control naturally occurring neuron death among ganglion cells of the vertebrate retina.
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9,178,043
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Postnatal changes of brain monoamine levels in prenatally malnourished and control rats.
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The effects of age and prenatal protein malnutrition (6% casein diet) on the concentration of monoamine neurotransmitters and their metabolites and precursors in the hippocampal formation, striatum, brain stem and cerebral cortex were investigated in 1-, 15-, 30-, 45-, 90- and 220-day-old rats. Concentrations of all neurotransmitters, i.e. dopamine, norepinephrine and serotonin changed significantly during the development. However, two main patterns were recognized. Serotonin in all areas, and dopamine in the striatum, increased from birth to day 45, and declined significantly in 90-day-old rats. In contrast, norepinephrine in all areas, and dopamine in areas other than the striatum, showed the lowest levels in 30-day-old rats, with levels increasing gradually after this age. Concentrations of metabolites paralleled changes in corresponding neurotransmitter levels. Prenatal protein malnutrition did not significantly affect any neurotransmitter concentrations with the exception of increased tryptophan levels (181%) in the hippocampal formation of newborn rats and decreased tyrosine levels (59%) in the striatum of day 30 rats. The results indicate that the monoamine transmitter content varied dynamically throughout postnatal life; however, they seem to counteract the insult from prenatal protein malnutrition after postnatal nutritional rehabilitation.
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9,178,046
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Psychiatry of the elderly: a consensus statement.
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This consensus statement, published by The Division of Mental Health and Prevention of Substance Abuse of the World Health Organisation, was co-sponsored by WHO and the Geriatric Section of the World Psychiatric Association, under the Section presidency of Professor J. Wertheimer. The final statement was prepared by an inter-disciplinary group representing the principal international association, at a meeting in Lausanne, 5-7 February 1996, chaired by Professor H. Hafner. Professor Cornelius Katona and Dr. Nori Graham were co-rapporteurs. A full list of participants is available from Professor Wertheimer.
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9,178,047
|
The Center for Epidemiological Studies-Depression (CES-D) Scale: assessment of depression in the medically ill elderly.
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This study examines the use of the Center for Epidemiological Studies-Depression Scale (CES-D) in a sample of elderly, medically ill inpatients. Seventy-six individuals completed the CES-D and a psychiatric interview, from which DSM-III-R diagnoses of depression were obtained. Analyses of sensitivity and specificity indicated that use of an alternative scoring method which more closely approximates current diagnostic criteria for depression may improve the predictive power of the test. Employment of stringent cut-scores was not supported, as sensitivity was compromised. Item analyses demonstrated that seven of the CES-D items failed to discriminate major, minor and nondepressed patients, and that several of these items tapped somatic symptoms. These findings suggest that the validity of the CES-D may be compromised when used with elderly medical patients. and modifications for its use are recommended.
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9,178,048
|
Sexual relationships in married dementia sufferers.
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To determine the proportion of couples, one of whom suffers from dementia, continuing with a sexual relationship, their level of satisfaction with their sexual relationship and the associations of remaining sexually active.
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9,178,050
|
Differentiating between demented and psychiatric patients with the Dutch version of the IQCODE.
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This study presents psychometric data on the Dutch version of the Informant Questionnaire on Cognitive Decline in the Elderly (IQCODE-N). We focused on differentiating between inpatients with dementia or a mood/psychotic disorder. A cutoff point of 3.9 on the IQCODE-N correctly predicted group membership in 87.8% and 78.8% respectively. Internal consistency and principal component analysis suggest that the IQCODE-N measures a unidimensional construct. Correlations with a Dementia Screening Test (r = 0.62), nurses' observations on memory disorders (r = 0.71) and a global assessment of severity of dementia (r = 0.72) support the validity of the informant ratings. Patients' age and level of education did not confound informant ratings. Only a small fraction of the informants could not complete the IQCODE-N, which makes it an easily applicable rating scale for cognitive decline.
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9,178,049
|
Health-related quality of life in older patients with schizophrenia and other psychoses: relationships among psychosocial and psychiatric factors.
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Few multivariate studies relating psychosocial factors to symptoms of psychosis among older patients exist. We assessed environmental stressors, satisfaction with emotional support, coping responses and psychiatric symptoms, and sought to relate these factors to quality of well-being among older patients with schizophrenia and other psychoses.
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9,178,051
|
Stress of caregivers of dementia patients in the Singapore Chinese family.
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In a study of 50 family caregivers of elderly Chinese patients with dementia in Singapore, 28 (56%) scored five points or more on the 28-item General Health Questionnaire (GHQ). The GHQ scores correlated significantly with duration of care; presence of delusion, hallucination, depression, insomnia, incontinence and agitation; and the total score of the Behavioural Pathology in Alzheimer's Disease Rating Scale. On multiple regression analysis the only variables to achieve a significant relationship with the GHQ scores were duration of care, depression and the total behavioural score.
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9,178,052
|
Carers' knowledge of dementia and their expressed concerns.
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The authors wished to determine how much carers from different settings caring for patients with dementia knew about the disorder and elicit their main concerns about the disease.
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9,178,054
|
Coroner's verdicts in the elderly: a suicide or an open verdict?
|
To identify variables, other than legal definitions, that may have influenced North Cheshire's Coroner in returning a verdict of 'suicide' or an 'open verdict' on unexpected deaths of the elderly.
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9,178,053
|
Characteristics of elderly psychiatric patients retained in a state hospital during downsizing: a prospective study with replication.
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This study examined the clinical characteristics of elderly inpatients associated with retention in a large state hospital during a period of rapid reduction in the inpatient census.
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9,178,055
|
Prevalence and correlates of aggressive behaviours occurring in patients with Alzheimer's disease.
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To determine the prevalence and clinical correlates of verbal and physical aggression occurring in Alzheimer's disease sufferers.
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9,178,056
|
Behaviour therapy for obsessive compulsive disorder in a 78-year-old woman.
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To describe a case of late onset obsessive compulsive disorder (OCD) and determine the impact of a behavioural intervention on OCD symptoms.
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9,178,071
|
B-cell activation versus tolerance--the central role of immunoglobulin receptor engagement and T-cell help.
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The critical variables responsible for the decision between activation and tolerance in the B-cell compartment were studied in the well-characterised hen egg lysozyme:antilysozyme transgenic model. Mature B-cells exposed to a low antigenic signal, which resulted in a receptor occupancy of between 5% and 25%, remained 'indifferent' (i.e. were neither activated nor tolerant), had a normal lifespan and recirculated through B-cell follicles. When receptor occupancy reached a critical threshold of approximately 50%, the B-cells became activated and moved to the outer T-cell zones where they died. This threshold appeared to be lower in the case of immature B-cells. On addition of T-cell help, however, the B-cells were rescued, proliferated and secreted antibody. Thus the outcome of the interaction between B-cells and antigen (self or foreign) is determined largely by the degree of antigen receptor engagement and availability of T-cell help.
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9,178,070
|
CD40: a pivotal receptor in the determination of life/death decisions in B lymphocytes.
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CD40 is a 48 kDa glycoprotein predominantly expressed on B cells in both mouse and man, which interacts with a counterligand (CD40L), expressed on activated CD4+ T cells. CD40/CD40L interactions are now known to be essential for the initiation of antibody responses to T-dependent antigens. In this review we discuss the immunobiology of CD40, with a special emphasis on our own studies in the mouse. These have focused on signal transduction via CD40, the role of cytokines (both T cell-derived and B cell-derived) in CD40-mediated B cell activation, and the role of CD40 in protecting B cells from apoptotic cell death. The available data indicate clearly that this protein is a pivotal receptor on B cells, both for the delivery of activating signals and for promoting B cell survival.
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9,178,073
|
The role of T cells in the regulation of B cell tolerance.
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The study of conventional models of B cell tolerance has suggested that self-tolerance is imposed on B cells at an early stage in their development due to a peculiar sensitivity of immature B cells to tolerance induction. While this concept accounts for some aspects of central B cell tolerance, it is inconsistent with recent reports of tolerance induction in mature splenic B cells from immunoglobulin transgenic mice. We present an alternative model, the hierarchical model (Aust. N. Z. J. Med. 25, 761-767, 1995), in which regulation of naive B cell reactivity is a function of antigen signal strength and availability of T cell help, but is independent of B cell maturation stage. In turn, the development of tolerance or memory in the T cell compartment is dependent on a combination of antigen-MHC recognition by T cells and antigen-nonspecific signalling by antigen-presenting cells. Using a transgenic model of T-B collaboration, we have shown that both immature and mature self-reactive B cells can be rescued and induced to secrete auto-antibody if the B cell determinant is linked to a carrier protein bearing a foreign T cell determinant.
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9,178,072
|
Apoptosis and the B cell response to antigen.
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The primary B cell response to T cell dependent antigens comprises two pathways of differentiation; one resulting in formation of foci of antibody forming cells in the extrafollicular regions of secondary lymphoid organs and the other giving rise to germinal centers within the follicles. Foci of antibody forming cells are detectable for only a limited time, before involuting due to apoptosis of the plasma cells. Similarly in the germinal center, regulation of cell number, selection of high affinity variants generated by somatic hypermutation, and the resolution of the germinal center itself all involve the death of unwanted B cells. In this review we describe recent experiments which have allowed determination of the role of certain forms of apoptosis in the B cell response to antigen.
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9,178,074
|
The importance of efficacy and partial agonism in evaluating models of B lymphocyte activation.
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Immunologists have developed a range of in vitro techniques for probing the receptor mediated response of cells comprising the immune system. An important and ubiquitous method is the use of antibodies in either soluble or aggregated form to engage cell surface receptors and transmit a signal. Models of cell and molecular interactions, derived from the use of these antibodies, form the basis of our efforts to understand and explain the corresponding in vivo systems. However, interpreting in vitro experiments and distinguishing between alternative models is difficult. This complexity is illustrated here using B cell stimulation by surface immunoglobulin and CD40. The fluorescent cell labelling dye carboxyfluorescein, diacetate, succinimidyl ester (CFSE) is used to show that many anti-Ig and CD40 stimulatory agents, used to assess the role of B cells and lymphokines, are partial agonists. By modelling each step in B cell signalling, activation and division it is possible to show that small changes in signal contributed by a second receptor can generate numerous distinct dose response curves that are highly dependent on the "efficacy" of signal transmission by the primary ligand and the number of cell divisions taken in culture. Differences in dose response curves become particularly striking if the primary activating stimulus is a partial agonist. Although exemplified here with B cell stimulation the conclusions are applicable to other in vitro activation systems and suggest ways to improve both the design and interpretation of in vitro experiments.
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9,178,077
|
Association between a history of childhood sexual abuse and subsequent, adolescent psychoactive substance use disorder in a sample of HIV seropositive men.
|
Examine whether an association exists between having a sexual abuse history and having a psychoactive substance use disorder (PSUD) history in a population of HIV seropositive men.
|
9,178,078
|
Weekly and seasonal variation in sexual behaviors among adolescent women with sexually transmitted diseases.
|
The objective of this research is to describe aspects of the organization of adolescent sexual behavior in order to understand factors associated with risk for sexually transmitted diseases (STD).
|
9,178,079
|
Effect of parental mental health status on adolescents' dietary behaviors.
|
The purpose of this study was to explore whether adolescents of substance-abusing and depressed parents were more likely to have poor dietary behaviors than those in the health comparison families.
|
9,178,080
|
Developmental and food profiles of infants born to adolescent and adult mothers.
|
To compare developmental markers and dietary intake of infants born to lower socioeconomic adolescent and adult mothers.
|
9,178,081
|
Lactational performance of adolescent mothers shows preliminary differences from that of adult women.
|
The purposes of this study were to characterize milk production, milk composition, and the lactational behavior of adolescent mothers, and to compare their lactational performance with that of adult females.
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9,178,082
|
Covariations of eating behaviors with other health-related behaviors among adolescents.
|
The study objectives are: (1) to examine and compare patterns of covariation of a wide range of health behaviors among adolescent boys and girls; (2) to determine whether eating behaviors are part of a larger construct of health-related behaviors and to identify the behaviors with which they share underlying similarities; and (3) to determine whether youth engaging in other health-compromising behaviors are at risk for unhealthy eating.
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9,178,083
|
Black-white differences in body size perceptions and weight management practices among adolescent females.
|
This study compares body size perceptions and weight management practices of black and white adolescent females.
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9,178,084
|
Use of body mass index to monitor treatment of obese adolescents.
|
Absolute weight loss may not always be the best measure of adherence and response to therapy in obese adolescents if weight gain owing to linear growth is not considered. We wished to compare short-term absolute weight and height changes with changes in body mass index (BMI) in a group of severely obese adolescents to determine the most meaningful measure of treatment response.
|
9,178,085
|
NMR study of the peptide present in the principal neutralizing determinant (PND) of HIV-1 envelope glycoprotein gp120.
|
The peptide sequence Gly-Pro-Gly-Arg-Ala-Phe (GPGRAF) is present in many principal neutralizing determinants (PND) of the human immunodeficiency virus type-1 (HIV-1). It has been shown that peptides from the PND sequence contain a significant beta turn in the conserved Gly-Pro-Gly-Arg sequence. In order to find out whether or not the smaller subunits also contain this turn, we have studied the NMR of a hexapeptide [GPGPRAF, peptide (I)], a heptapeptide Gly-Pro-Gly-Arg-Ala-Phe-Cys [GPGRAFC, peptide (II)] and a dodecapeptide [GPGRAFGPGRAF, peptide (III)], retaining the side chain protecting groups. Although the majority of conformations for these peptides are disordered, there is a considerable propensity of structures with beta turn in the GPGR sequence. While peptide (I) and peptide (III) seem to have both type I and type II beta turn conformations, peptide (II) shows a propensity of only type II beta turn. The nascent structures obtained in these peptides may get stabilized as the receptor binding conformation in the presence of the receptors, thus playing a significant role in vaccine development against HIV.
|
9,178,086
|
Simultaneous chromatographic analysis of eight fat-soluble vitamins in plasma.
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A high-performance liquid chromatographic (HPLC) method for the simultaneous analysis of eight fat-soluble vitamins including A acid, retinol and retinal, vitamins D2 and D3 together with menadione (vitamin K3) is described. The eight vitamins extracted from plasma were analyzed by reversed-phase HPLC with UV detection at 245 nm. The mobile phase was composed of methanol and ethanol (85:15, v/v) with 0.1% triethylamine. The extraction efficiency of different solvents, namely, dichloromethane, benzene, methanol, ethanol, chloroform and hexane, with or without the addition of detergents was studied. There is no single solvent system that could extract all eight vitamins in one step. A recovery study of the vitamins was performed using rabbit plasma. An average recovery for individual vitamins was about 40% or more with the described extraction methods. The absorbance response was linear at the nanogram level. The present method may be broadly applied with alternative extraction methods for the eight vitamins from different sources.
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9,178,088
|
Large-scale purification and long-term stability of human butyrylcholinesterase: a potential bioscavenger drug.
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Butyrylcholinesterase from human plasma (HuBChE) is a potential drug candidate for detoxification of certain harmful chemicals that contain carboxylic or phosphoric acid ester bonds. Large quantities of purified HuBChE, displaying a high stability upon long-term storage, are required for the evaluation of its therapeutic capacity and its pharmaceutical properties. Several modifications of a previously reported procedure enabled us to purify the enzyme > 15,000-fold from pools of up to 100 1 of human plasma. The three-step procedure is based on precipitation of plasma proteins by ammonium sulfate (step I) and batch adsorption of HuBChE on procainamide-Sepharose 4B gel (step II). Ammonium sulfate was also employed in the third stage to fractionate the final product from procainamide-containing HuBChE solution. The overall yield (63%) of electrophoretically pure enzyme was significantly higher than that previously reported (34%) for the purification of HuBChE from 12.5 1 of plasma or from 5 kg of Cohn fraction IV-4. Purified HuBChE was stored at 5 degrees C in 10 mM phosphate buffer (pH 7.4) containing 1 mM EDTA and 0.02% NaN3. The specific activity, protein migration on gel electrophoresis, thermostability at 54 degrees C and the mean residence time in the circulation of mice remained essentially constant for at least 46 months. The modifications introduced can provide large quantities of purified enzyme that maintains its activity and bioavailability properties for several years.
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9,178,087
|
In vivo study of halothane hepatotoxicity in the rat using magnetic resonance imaging and 31P spectroscopy.
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Using magnetic resonance imaging (MRI) and spectroscopy (MRS), in vivo halothane hepatotoxicity was assessed in male Wistar rats. With 1.5% halothane in 100 or 20% O2, an edematous region, characterized by increased intensity on T2 weighted images and an increase in regional tissue water content (rho water), was seen proximal to the hepatic portal vein in the liver. Both spin-lattice relaxation (T1) and spin-spin relaxation (T2) increased in this region, relative to distal regions of the liver. Similarly, a high signal intensity on proton density weighted images was observed in this area. As halothane anaesthesia progressed, a decrease in the adenosine triphosphate-inorganic phosphate ratio (ATP/Pi) and an increase in the phosphomonoester-phosphodiester (PME/PDE) ratio was detected in the liver. In addition, intracellular pH decreased and intracellular free magnesium concentration [Mg2+] increased with time of exposure. Excessive vacuolation, ribosomal disappearance from rough endoplasmic reticulum, mitochondrial swelling and fragmentation of smooth endoplasmic reticulum were observed by transmission electron microscopy (TEM) in samples from the edematous region of the liver.
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9,178,090
|
Quantification of total RNA by ethidium bromide fluorescence may not accurately reflect the RNA mass.
|
The fluorescent signal from purified large and small ribosomal RNA subunits stained with ethidium bromide was quantified using agarose gels and image analysis. A significantly smaller fluorescent signal was observed for 23 S rRNA compared to 16 S rRNA when normalized to mass. Therefore, small changes in amounts of 23 S or 16 S rRNA within a sample can have a profound impact on observed bulk fluorescence. A comparison of 23 S/16 S rRNA fluorescence ratios for 4 marine bacteria grown in batch culture indicates that the lower fluorescence of 23 S rRNA is widespread. The 23 S/16 S rRNA fluorescence ratios also suggest that intracellular concentrations of ribosomal subunits do not always adhere to a stoichiometry of 2.0.
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9,178,089
|
Determination of cell surface charge by photometric titration.
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A colloid titration method has been frequently used to determine the number of charged residues at the cell surface. Here we present a new version of this technique, based on photometric measurements of a metachromatic shift in the maximum absorption of toluidine blue as it binds to the cell surface. The major improvements are: (1) simplified methodology and (2) increased precision of equivalence point determination. The data are analyzed using Gran's theory, which allows measurements to be taken at regular intervals instead of being concentrated around the equivalence titration point. We used this method to characterize the cell surface charge of three populations of rat mast cells: (1) peritoneal mast cells (PMC), (2) bone marrow-derived mast cells (BMMC) and (3) a rat cultured mast cell line (RCMC). Our results indicate that PMC have (4.23 +/- 0.59) x 10(8), while BMMC (8.58 +/- 0.26) x 10(7) negatively charged residues per cell. The results for RCMC were similar to those for BMMC. Taking into account the size differences between PMC and BMMC, the average charge density of PMC was also significantly higher than that of BMMC. The differences in cell surface charge were analyzed in the light of different sensitivities of mast cells to polycationic secretagogues.
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9,178,091
|
Polyphenol oxidase activity staining in polyacrylamide electrophoresis gels.
|
An analytical method allowing the detection of polyphenol oxidase activity on polyacrylamide gel electrophoresis (PAGE) is described. The method is rapid, sensitive and specific and is based on a coupling reaction between 4-tert-butyl-o-benzoquinone and the aromatic amine, 4-amino-N,N-diethylaniline sulphate. Catecholase activity of polyphenol oxidase appears as blue stained bands on a colourless background.
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9,178,094
|
Influence of polyethylene glycol graftings on the in vitro degradation and calcification of bovine pericardium.
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Calcification is a frequent cause of the clinical failure of bio-prosthetic heart valves fabricated from glutaraldehyde pretreated bovine pericardium (GATBP). This article reports on various chemical techniques for grafting polyethylene glycol (PEG) on bovine pericardium, their biostability, and calcification. The process of calcification profile was studied by in vitro experiments via the incubation of pericardial samples in a metastable solution of calcium phosphate. The calcification profile of PEG-modified bovine pericardium through glutaraldehyde linkages was significantly reduced compared to other methods of grafting. The mechanical property of these PEG-modified tissues after enzyme (collagenase) digestion and calcification were also investigated. PEG grafting of BP via glutaraldehyde or hexamethylene diisocyanate had shown better mechanical stability compared to other grafting methods used. In conclusion, it seems that the surface modification of bovine pericardium through high molecular weight PEGs via glutaraldehyde linkages may provide new ways of controlling tissue biodegradation and calcification.
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9,178,095
|
Genetic and biochemical analysis of the transmembrane domain of Arabidopsis 3-hydroxy-3-methylglutaryl coenzyme A reductase.
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We have examined the amino terminal membrane anchoring domain of Arabidopsis thaliana 3-hydroxy-3-methylglutaryl coenzyme A reductase (Hmg1p), a key enzyme of the isoprenoid biosynthetic pathway. Using both in vitro and in vivo approaches, we have analyzed a series of recombinant derivatives to identify key structural elements which play a role in defining Hmg1p transmembrane topology. Based on our results, we have proposed a topological model for Hmg1p in which the enzyme spans the lipid bilayer twice. We have shown the two transmembrane segments, designated TMS1 and TMS2, to be structurally and functionally inequivalent in their ability to direct the targeting and orientation of reporter proteins. Furthermore, we provide evidence indicating both the extreme amino terminal end and carboxyl terminal domain of the protein reside in the cytosol. This model therefore provides a key basis for the future examination of the role of the transmembrane domain in the targeting and regulation of Hmg1p in plant cells.
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9,178,096
|
Differentiation of human trophoblast cells in vitro is inhibited by dimethylsulfoxide.
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Dimethyl sulfoxide (DMSO) exerts a number of biological effects, the most frequently cited being induction of cell differentiation. The compound also increases invasiveness and metastatic potential. In contrast to the many reports of DMSO-induced cell differentiation, we report here that DMSO inhibits the morphological differentiation of human cytotrophoblast cells to syncytiotrophoblast, as revealed by immunofluorescence staining for desmosomal protein and nuclei. Cytotrophoblast cells treated with DMSO under differentiation-inducing conditions remained mononucleated with intense desmosomals staining. The effect was dose dependent, with a maximal effect seen at 1.5% DMSO. Concentrations of < or = 0.5% had no effect and concentrations > 2% were cytotoxic. In addition to these morphological changes, DMSO inhibited secretion of human chorionic gonadotropin in a dose-dependent manner. At a concentration of 1.5%, DMSO inhibited secretion by 70%. If cytotrophoblast cells were cultured in the presence of DMSO and then switched to DMSO-free medium, they proceeded to differentiate normally. While the precise mechanism of action remains unknown, judicious use of DMSO may be a useful tool for studying and manipulating the differentiation of human trophoblast cells in vitro. The findings also indicate that care should be used in interpreting results obtained using DMSO as a carrier in drug and inhibitor studies.
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9,178,097
|
Autoregulation of actin synthesis responds to monomeric actin levels.
|
Regulation of the assembly and expression of actin is of major importance in diverse cellular functions such as motility and adhesion and in defining cellular and tissue architecture. These biological processes are controlled by changing the balance between polymerized (F) and soluble (G) actin. Previous studies have indicated the existence of an autoregulatory pathway that links the state of assembly and expression of actin, resulting in the reduction of actin synthesis after actin filaments are depolymerized. We have employed the marine toxins swinholide A and latrunculin A, both disrupting the organization of the actin-cytoskeleton, to determine whether this autoregulatory response is activated by a decrease in the level of polymerized actin or by an increase in monomeric actin concentrations in the cell. We showed that in cells treated with swinholide A the level of filamentous actin is decreased, and using a reversible cross-linking reagent, we found that actin dimers are formed. Latrunculin A also disassembled actin filaments, but produced monomeric actin, followed by a reduction in actin and vinculin expression, while swinholide A treatment elevated the synthesis of these proteins. In cells treated with both latrunculin A and swinholide A, dimeric actin was formed, and actin and vinculin synthesis were higher than in control cells. These results suggest that the substrate that confers an autoregulated reduction in actin expression is monomeric actin, and when its level is decreased by dimeric actin formation, actin synthesis is increased.
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9,178,098
|
Simultaneous activity of two different mechanisms of folate transport in ovarian carcinoma cell lines.
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We investigated whether the folate receptor alpha-isoform (FR alpha), which is overexpressed on ovarian carcinoma cells, is functionally active in internalizing the physiological form et folate, 5-methyl tetrahydrofolate (THF). Six ovarian tumor cell lines, expressing different levels of FR alpha (COR > > OVCAR3 > IGROV1 > OVCAR4 > SKOV3 > OVCAR5), were maintained in folate-depleted medium and internalization of 10 nM evaluated as acid-resistant radioactivity at 0 degree and 37 degrees C. The amount of 5-methyl[1H]THF present in this fraction was not strictly related to the number of membrane receptors, since even cell lines with low FR alpha expression, e.g., OVCAR4, showed efficient internalization. Time-course studies indicated that, whereas no uptake was detected at 0 degree C, at 37 degrees C the internalized fraction showed a slow and constant increase, until 4 h. At this time the internalized radioactivity represented < 50% of the total bound in COR, OVCAR3 and IGROV1 cells, whereas the other cell lines tested internalized fourfold more folate than their surface binding capacity. The incubation in the presence of a concentration (50 nM) of 5-methyl[3H]THF, which best ensures receptors saturation on cells with highest FR levels (COR and OVCAR3), had slight effect on surface binding of all the tested cell lines, including IGROV1 and SKOV3. In contrast, the increase of the uptake was more pronounced, particularly in SKOV3 cells. These results, together with the accumulation curves of folic acid (FA) and 5-methylTHF at 37 degrees C, suggested the presence of a molecule on ovarian carcinoma cells with high affinity for reduced folates, possibly a reduced folate carrier (RFC). Measurement of radioactivity present in the supernatant of IGROV1 and SKOV3 cells, subjected to hypotonic lysis and cell fractionation, further indicated that 5-methyl[3H]THF was translocated to the cytosol and, despite differences in membrane levels of FR alpha expression this internalized fraction was similar in both cell lines. Inhibition experiments to selectively block FR alpha or RFC activity showed a differential sensitivity of the two pathways depending on the cell line examined. Internalization was more consistently inhibited on IGROV1 than on SKOV3 cells by treatments that disrupt FR alpha activity, e.g., incubation with excess FA and phosphatidylinositol specific phospholipase C, whereas Probenecid, which preferentially inhibits the carrier-mediated pathway, showed a strong inhibitory effect on both cell lines. These findings suggest that the internalization of 5-methylTHF in these tumor cells depends not only on the level of overexpressed FR alpha, but another transport route, with features characteristic for RFC, is functional and participates in folate uptake.
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9,178,099
|
Dexamethasone induces rapid actin assembly in human endometrial cells without affecting its synthesis.
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Dexamethasone exerts a stimulatory effect of rapid-onset on the polymerization of actin. This has been documented in human endometrial adenocarcinoma Ishikawa cells, resulting in an acute, dose-dependent decrease in the G/total-actin ratio. In the present study we completely characterized this fast and apparently nongenomic effect of dexamethasone on actin assembly. We followed the morphological alterations of actin cytoskeleton and measured the time-dependent dynamics of actin polymerization both by ruling out any changes of total actin in the cells and by measuring its transcript. Rapid changes in actin polymerization were accurately measured using a highly sensitive and quantitative rhodamine-phalloidin fluorimetric assay. Ishikawa cells, exposed to 0.1 microM dexamethasone for various time periods up to 24 h, showed a highly significant, rapid, and transient increase in the polymerization of actin starting within 15 min of dexamethasone exposure and lasting 2 h. Treated cells showed a significant (1.79-fold) enhancement of the fluorescent signal compared to untreated cells at 15 min. This value decreased continuously in a time-dependent manner, reaching control levels after 120 min and remained so for the next 24 h. Confocal laser scanning microscopy studies confirmed these findings. Intensive coloration of microfilaments over several scanning sections suggested an enhanced degree of actin polymerization in cells preincubated for 15 min with 0.1 microM dexamethasone. Moreover, actin filaments were more resistant to cytochalasin B. Additionally, quantitative immunoblot analysis showed that the content of total cellular actin remained the same during this period, suggesting that the biosynthesis of actin was unaffected. Northern blot analysis showed that the concentration of the actin transcript was also unaffected. Our data suggest that glucocorticoids induce a fast and self-limited polymerization of actin in human endometrial cells without affecting its synthesis. These findings strengthen the hypothesis that glucocorticoids exert rapid, nongenomic cellular effects and that the actin-based cytoskeleton is an integral part of this pathway, playing an essential role in receiving and mediating steroid signals for the modulation of cellular responses.
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9,178,100
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Transforming growth factor-beta 1 regulation of bone sialoprotein gene transcription: identification of a TGF-beta activation element in the rat BSP gene promoter.
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Transforming growth factor-beta (TGF-beta) increases steady-state mRNA levels of several extracellular matrix proteins in mineralized connective tissues. Bone sialoprotein (BSP) is a major constituent of the bone matrix, thought to initiate and regulate the formation of mineral crystals. To determine the molecular pathways of TGF-beta 1 regulation of bone proteins, we have analyzed the effects of the TGF-beta 1 on the expression of the BSP in the rat osteosarcoma cell line (ROS 17/2.8). TGF-beta 1 at 1 ng/ml, increased BSP mRNA levels in ROS 17/2.8 cells approximately 8-fold: the stimulation was first evident at 3 hr, reached maximal levels at 12 hr and slowly declined thereafter. Since the stability of the BSP mRNA was not significantly affected by TGF-beta 1, and nuclear "run-on" transcription analyses revealed only a approximately 2-fold increase in the transcription of the BSP gene, most of the increase in BSP mRNA appeared to involve a nuclear post-transcriptional mechanism. Moreover, the effects of TGF-beta 1 were indirect, since the increase in BSP mRNA was abrogated by cycloheximide (28 micrograms/ml). To identify the site of transcriptional regulation by TGF-beta 1, transient transfection analyses were performed using BSP gene promoter constructs linked to a luciferase reporter gene. Constructs that included nt -801 to -426 of the promoter sequence were found to enhance transcriptional activity approximately 1.8-fold in cells treated with TGF-beta 1. Within this sequence, approximately 500 nt upstream of the transcription start site, a putative TGF-beta activation element (TAE) was identified that contained the 5'-portion of the nuclear factor-1 (NF-1) canonical sequence (TTGGC) overlapping a consensus sequence for activator protein-2 (AP-2). The functionality of the TAE was shown by an increased binding of a nuclear protein from TGF-beta 1 stimulated cells in gel mobility shift assays and from the attenuation of TGF-beta 1-induced luciferase activity when cells were co-transfected with a double-stranded TAE oligonucleotide. Competition gel mobility shift analyses revealed that the nuclear protein that binds to the TAE has similar properties to, but is distinct from, NF-1 nuclear protein. These studies have therefore identified a TGF-beta activation element (TAE) in the rat BSP gene promoter that mediates the stimulatory effects of TGF-beta 1 on BSP gene transcription.
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9,178,101
|
Reduced drug accumulation and multidrug resistance in human breast cancer cells without associated P-glycoprotein or MRP overexpression.
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MCF-7 human breast cancer cells selected in Adriamycin in the presence of verapamil developed a multidrug resistant phenotype, which was characterized by as much as 100,000-fold resistance to mitoxantrone, 667-fold resistance to daunorubicin, and 600-fold resistance to doxorubicin. Immunoblot and PCR analyses demonstrated no increase in MDR-1 or MRP expression in resistant cells, relative to parental cells. This phenotype is similar to one previously described in mitoxantrone-selected cells. The cells, designated MCF-7 AdVp, displayed a slower growth rate without alteration in topoisomerase II alpha level or activity. Increased efflux and reduced accumulation of daunomycin and rhodamine were observed when compared to parental cells. Depletion of ATP resulted in complete abrogation of efflux of both daunomycin and rhodamine. No apparent alterations in subcellular daunorubicin distribution were observed by confocal microscopy. No differences were noted in intracellular pH. Molecular cloning studies using DNA differential display identified increased expression of the alpha subunit of the amiloride-sensitive sodium channel in resistant cells. Quantitative PCR studies demonstrated an eightfold overexpression of the alpha subunit of the Na+ channel in the resistant subline. This channel may be linked to the mechanism of drug resistance in the AdVp cells. The results presented here support the hypothesis that a novel energy-dependent protein is responsible for the efflux in the AdVp cells. Further identification awaits molecular cloning studies.
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9,178,102
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Identification of Rad's effector-binding domain, intracellular localization, and analysis of expression in Pima Indians.
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In order to characterize the endogenous gene product for rad (ras-related protein associated with diabetes), we prepared antibodies to synthetic peptides that correspond to amino acids (109-121, 178-195, 254-271) within the protein. These antibodies were used to analyze the expression, structure, and function of rad. Western analysis with these antibodies revealed that rad was a 46 kDa protein which was expressed during myotube formation. Further, immunolocalization studies showed that rad localized to thin filamentous regions in skeletal muscle. Interestingly, when muscle biopsies from diabetic and control Pima Indians were compared, no differences in rad protein or mRNA expression were observed. Similarly, no differences were observed in protein expression in diabetic and control Zucker diabetic fatty (ZDF) rats. Functional analysis of muscle rad revealed that its GTP-binding activity was inhibited by the addition of N-ethylmaliemide, GTP, GTP gamma S, and GDP beta S but not ATP or dithiothreitol. Moreover, cytosol-dependent rad-GTPase activity was stimulated by the peptide corresponding to amino acids 109-121. Antibodies corresponding to this epitope inhibited cytosol-dependent rad-GTPase activity. Taken together, the results indicate that 1) rad is a 46 kDa GTP-binding protein localized to thin filaments in muscle and its expression increases during myoblast fusion, 2) expression of rad in Pima Indians and ZDF rats does not correlate with diabetes, and 3) the amino acids (109-121) may be involved in regulating rad-GTPase activity, perhaps by interacting with a cytosolic factor(s) regulating nucleotide exchange and/or hydrolysis.
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9,178,103
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Alteration of the kinetics of type I procollagen synthesis in human osteosarcoma cells by 1,25-dihydroxyvitamin D3.
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The kinetics of type I procollagen synthesis in a human osteosarcoma cell line, MG 63, were investigated after treatment with 1,25-dihydroxyvitamin D3 (1,25-(OH)2 D3), a hormonal inducer of phenotypic differentiation. Pulse label and chase experiments demonstrated greatly enhanced production and more rapid reduction of intracellular procollagen molecules in the 1,25-(OH)2 D3-treated cells as compared to the nontreated case. After a chase for 1 h, labeled procollagen was reduced by nine-tenths in 1,25-(OH)2 D3-treated cells, while half of the radioactivity still remained in nontreated cells. The expression rate of type I collagen, which was examined by pulse label experiment, was elevated in association with an increase in the mRNA coding for the type I collagen alpha 1 chain by 1,25-(OH)2 D3 treatment. However, the amount of intracellular procollagen present after 4 h continuous labeling was almost the same, independent of the 1,25-(OH)2 D3 treatment. Thus, we conclude that strage of the molecule was not affected. The results therefore suggest an increase in both the synthesis and secretion of type I collagen. The 1,25-(OH)2 D3 treatment was also found to induce the alpha subunit of prolyl 4-hydroxylase and to be associated with an elevated level of hydroxyproline in the procollagen. Moreover, gelatinase B-resistant procollagen molecules, indicative of intracellular procollagen molecules in the stable triple helical form, were detected only in the 1,25-(OH)2 D3-treated cells. These data suggest more efficient proline hydroxylation is involved in rapid secretion of procollagen after hormone administration. The present evidence points to posttranslational control of procollagen synthesis.
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9,178,104
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Partial isolation from intact cells of a cell surface-exposed lysophosphatidylinositol-phospholipase C.
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A novel cell surface phosphoinositide-cleaving phospholipase C (ecto-PLC) activity was isolated from cultured cells by exploiting its presumed external exposure. Biotinylation of intact cells followed by solubilization of the biotinylated proteins from a membrane fraction and recovery onto immobilized-avidin beads, allowed assay of this cell surface enzyme activity apart from the background of the substantial family of intracellular PLCs. Several cell lines of differing ecto-PLC expression were examined as well as cells stably transfected to overexpress the glycosylphosphatidylinositol (GP) anchored protein human placental alkaline phosphatase (PLAP) as a cell surface enzyme marker. The resulting bead preparations from ecto-PLC positive cells possessed calcium-dependent PLC activity with preference for lysophosphatidylinositol (lysaPI) rather than phosphatidylinositol (PI). The function of ecto-PLC of intact cells evidently is not to release GPI-anchored proteins at the cell surface, as no detectable Ca(2+)-dependent release of overexpressed PLAP from ecto-PLC-positive cells was observed. To investigate the cell surface linkage of the ecto-PLC itself, intact cells were treated with bacterial PI-PLC to cleave simple GPI anchors, but no decrease in ecto-PLC activity was observed. High ionic strength washes of biotinylated membranes prior to the generation of bead preparations did not substantially reduce the lysoPI-PLC activity. The results verify that the ecto-PLC is truly cell surface-exposed, and unlike other members of the PLC family that are thought to be peripheral membrane proteins, this novel lysoPI-PLC is most likely a true membrane protein.
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9,178,105
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Parathyroid hormone uses both adenylate cyclase and protein kinase C to regulate acid production in osteoclasts.
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Osteoclasts, isolated from the endosteum of 2.5- to 3-week-old chickens, were treated with acridine orange, a hydrogen ion concentration-sensitive fluorescent dye, in order to monitor changes in acid production. The adenylate cyclase inhibitor, alloxan, blocked parathyroid hormone (PTH)-stimulated acid production. Dibutyryl cyclic adenosine monophosphate, a membrane-permeant form of cyclic adenosine monophosphate, mimicked the PTH effect. Bisindolylmaleimide, a specific inhibitor of protein kinase C (PKC), blocked the initial stimulation (15, 30, and 60 min) of acid production by PTH but had no effect on long-term stimulation (120 min). Confocal microscopy of osteoclasts stained with fluorescein-conjugated bisindolylmateimide revealed a shift in location of PKC from the cytoplasm to the plasma membrane region after treatment with parathyroid hormone. The results of these studies support the hypothesis that PTH regulation of acid production in osteoclasts involves both adenylate cyclase and PKC as effectors.
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9,178,106
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Heparan sulfate-binding peptide promotes the deposition of proteoglycans in the extracellular matrix.
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A synthetic peptide, which was shown to bind extracellular matrix heparan sulfate chains with a high degree of affinity and specificity [Colburn et al. (1996): Arch Biochem Biophys 325:129-138], has now been found to promote the transfer and the deposition of endothelial cell surface proteoglycans in the extracellular matrix. The peptide also induces preferential binding of extracellular matrix heparan sulfate proteoglycans, which have been added to the supernatant growth medium, and the requirement for its presence is stringent in that only a negligible amount of proteoglycans are bound to the cell layer in the absence of the peptide. In addition, antibodies directed against the peptide detect the accumulation of the peptide in the matrix compartment where the peptide is found associated with the proteoglycans transferred from the cell surface. The sequence of events induced by the peptide appears to be an extension of a naturally occurring process since proteoglycans with properties similar to those of the species ordinarily present in the extracellular matrix have been observed to transfer from the cell surface to the matrix during a pulse-chase experiment. We suggest that formation of the complex peptide-proteoglycan with consequent displacement of the proteoglycan from its anchorage on the cell initiates the process of transfer of the heparan sulfate-bound peptide from the cell surface to the extracellular matrix.
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9,178,107
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Reactive oxygen species and antioxidants in inflammatory diseases.
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This paper aims to review the rôle of free radical-induced tissue damage and antioxidant defence mechanisms in inflammatory diseases that involve pathogenic processes similar to the periodontal diseases. There is a clearly defined and substantial role for free radicals or reactive oxygen species (ROS) in periodontitis, but little research has been performed in this area. This paper reviews the considerable data available relating ROS activity and antioxidant defence to inflammatory diseases and attempts to draw parallels with periodontitis, in an effort to stimulate more periodontal research in this important area. The recent discovery of the transcription factor nuclear factor kappa B (NF-kappa B) is reviewed and several potential pathways for cytokine-induced periodontal tissue damage, mediated by NF-kappa B1 are discussed. Emphasis is placed on cytokines that have been studied in periodontitis, principally TNF-alpha, IL-1, IL-6, IL-8 and beta-interferon. The link between cellular production of such important mediators of inflammation and the antioxidant (AO) thiols, cysteine and reduced glutathione (GSH), is discussed and it is hypothesised that NF-kappa B antagonists may offer important therapeutic benefits.
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9,178,108
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Cytokines modulate routes of collagen breakdown. Review with special emphasis on mechanisms of collagen degradation in the periodontium and the burst hypothesis of periodontal disease progression.
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In this paper, we review recent work on collagen degradation, 2 main routes of breakdown are described and their relevance during healthy and inflammatory conditions of the periodontium is discussed. Special attention is paid to the possible role of cytokines, in particular interleukin 1 (IL-1) and transforming growth factor beta (TGF-beta), on the modulation of collagen phagocytosis and metalloproteinase production. IL-1 has been shown to have a dual function in collagen digestion. It inhibits the intracellular phagocytic pathway, but at the same time, it strongly promotes extracellular digestion by inducing the release of collagenolytic enzymes like collagenase. TGF-beta has an opposite effect on both pathways and antagonizes IL-1. Collagenase is released in an inactive form, and a considerable fraction of the proenzyme may become incorporated in the extracellular matrix. This reservoir of latent enzyme can be activated (for instance by plasmin), leading to a sudden and extensive breakdown of the collagenous fibre meshwork. It is suggested that this phenomenon may also take place during progressive periodontitis and could explain an episodic nature of collagenolysis, clinically resulting in bursts of attachment loss (burst hypothesis).
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9,178,109
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Marginal bone level buccal to mandibular molars in digital radiographs from charge-coupled device and storage phosphor systems. An in vitro study.
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The aim of this in vitro study was to compare bone loss measurements in the furcation area of mandibular molars in digital radiographs from a CCD-(Sens-A-Ray) and a storage phosphor (Digora) system, 10 1st and 7 2nd mandibular molars, with intact lingual but reduced buccal bone with furcation involvements, were used. Radiographs were first taken with lead markers at the most apical part of the buccal bone and at the cemento-enamel junction. These radiographs were used to establish a radiographically true distance between the CEJ and the buccal bone level. The lead marker at the CEJ served as a reference point for the observers' subsequent estimates of the extent of bone removed, which were made in radiographs without lead markers at the bone level. 1 exposure (400 ms) for the CCD- and 5 (160 ms, 200 ms, 250 ms, 320 ms, 400 ms) for the storage phosphor system were used. Measurements were made in unprocessed (original) and processed images (contrast enhanced and/or high pass filtered). The results showed underestimation of bone loss, but smaller than previously reported for film radiographs. No significant difference was found between the 2 systems when compared at the same exposure. Nor were any significant differences found between unprocessed and processed images or between storage phosphor images from different exposures. We conclude that digital radiographs are comparable to film based radiographs for measurement of buccal bone loss but that lower exposures can be used, especially with the storage phosphor system.
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9,178,110
|
Clearance of sodium lauryl sulphate from the oral cavity.
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Sodium lauryl sulphate (SLS) is used in toothpaste and mouth rinses as an emulsifying and surface cleaning agent. SLS has been implicated in an increased incidence of oral irritation in subjects predisposed to recurrent aphthous stomatitis (RAU). Hence, the purpose of this study was to determine the levels of SLS found in the oral cavity following rinsing with an SLS containing mouth rinse and brushing with a SLS containing dentifrice. An analytical method to separate SLS from saliva and other complex systems was developed. The method used high performance liquid chromatography (HPLC) and detection performed using conductivity measurements. Standard curves with known concentrations showed a detection limit of less than 0.4 ug SLS/ml of fluid. 2 clinical studies were conducted to determine the amount of SLS retained in the mouth by a healthy population after rinsing or brushing with commercially available products. The results showed, after rinsing, that 96% of the available SLS from the rinse was recovered in the collected samples within 2 min. Similarly, after brushing, 86% of the SLS contained within the toothpaste was recovered from the collected samples within the first 10 min. These results showed that the amount of SLS retained in the oral cavity was minimal and the contact time between SLS and the oral cavity was very short. A 2nd study was conducted to measure the amount of SLS retained in the mouth by a population susceptible to RAU. After rinsing, 97% of the available SLS was recovered within the first 2 min. Following brushing, 89% of the SLS in the dentifrice was recovered within the first 10 min. These results were comparable to those determined by the study involving the healthy population.
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9,178,111
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In vitro effect of the Sensonic toothbrush on Treponema denticola.
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The purpose of this in vitro study was to compare the effects of the Sensonic. Oral-B Braun mechanical and Oral-B manual toothbrushes upon the morphology and cellular integrity of Treponema denticola. This spirochete was chosen because of its frequent isolation from active lesions of inflammatory periodontal disease and its pathogenic potential. T. denticola, strain ATCC 33421, was grown in an anaerobic nitrogen rich atmosphere in enhanced 1186 mycoplasma broth. 160, 5-ml aliquots of cultured microbes were assigned to 1 of 3 brushing treatment groups and a control group. Samples were further divided into 4 groups of 10 samples each and assigned to one of 4 brushing exposure times: 15, 30, 45, and 60 seconds. After treatment, 0.2 ml of each sample was applied to a millipore filter and examined by SEM. Intact microbes were counted from 10 non-overlapping fields at 4500x. Remaining treated samples were pelleted and examined by TEM. A statistically significant reduction of intact microbes for the Sensonic treatment group at each exposure time was found when compared to Oral-B Braun, Oral-B manual, and non-treated controls. TEM examination of Sensonic treated samples revealed separation of the outer membrane at lower exposure times and only cellular debris after exposures of 45 and 60 s. These results suggest that exposure to the sonic frequency generated by the Sensonic toothbrush is capable of severely disrupting the structural integrity of T. denticola.
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9,178,113
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Self-concept and dental health behaviours in adolescents.
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The purpose of this investigation was to examine the relation between some dental health behaviours and 2 measures of self-concept in adolescents. Data from a survey of 41142, 12-16-year-old children from 244 secondary schools throughout England were analysed to obtain information about their frequencies of toothbrushing, use of dental floss and dental attendance, and whether they recalled advice about toothbrushing, in relation to self-esteem and health locus of control (HLOC). Subjects completed a questionnaire, anonymously, in school class. The results showed a significant positive correlation (Spearman) between the frequencies of flossing and toothbrushing, in both sexes, and between social group and toothbrushing frequency, where brushing frequency increased as socio-economic status improved. Some association between use of floss and social group emerged, but this was smaller and less consistent than that observed with toothbrushing brushing frequency. Self-esteem was positively correlated with toothbrushing frequency at ages 12-15 years, while HLOC showed correlations at some ages but not others. Use of dental floss showed no relation to self-concept. Subjects with more favourable self-concept were more likely to make more frequent dental visits than those with a poorer self-view. There was a strong and consistent correlation between recalled advice about toothbrushing and lower self- esteem and external locus of control. The results are in agreement with our earlier reports and suggest that self-concept may play a significant role in mediating changes in dental health behaviour.
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9,178,112
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The effect of SRP on the clinical and microbiological parameters of periodontal diseases.
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The purpose of the present investigation was to examine the effect of SRP on clinical and microbiological parameters in 57 subjects with adult periodontitis (mean age 47 +/- 11 years). Subjects were monitored clinically and microbiologically prior to and 3, 6 and 9 months after full-mouth SRP under local anaesthesia. Clinical assessments of plaque, redness, suppuration, BOP, pocket depth and attachment level were made at 6 sites per tooth. The means of duplicate attachment level measurements taken at each visit were used to assess change between visits. Clinical data were averaged within each subject and then averaged across subjects for each visit. Subgingival plaque samples were taken from the mesial aspect of each tooth and the presence and levels of 40 subgingival taxa were determined using whole genomic DNA probes and checkerboard DNA-DNA hybridization. The mean levels and % of sites colonized by each species (prevalence) was computed for each subject at each visit. Differences in clinical and microbiological parameters before and after SRP were sought using the Wilcoxon signed ranks test or the Quade test for more than 2 visits. Overall, there was a mean gain in attachment level of 0.11 +/- 0.23 mm (range -0.53 to 0.64 mm) 3 months post-therapy. There was a significant decrease in the % of sites exhibiting gingival redness (68 to 57%) and BOP (58 to 52%) as well as a mean (+/-SEM) pocket depth (3.3 +/- 0.06 to 3.1 +/- 0.05 mm). Sites with pre-therapy pocket depths of < 4 mm showed a non-significant increase in pocket depth and attachment level, 4.6 mm pockets showed a significant decrease in pocket depth and a non-significant gain in attachment post-therapy, while > 6 mm pockets showed a significant decrease in pocket depth and attachment level measurements post-therapy. Significant clinical improvements were seen in subjects who had never smoked or were past smokers but not in current smokers. Mean prevalences and levels of P. gingivalis, T. denticola and B. forsythus were significantly reduced after SRP, while A. viscosus showed a significant increase in mean levels. The mean decrease in prevalence of P. gingivalis was similar at all pocket depth categories, while B. forsythus decreased more at shallow and intermediate pockets and A. viscosus increased most at deep sites. P. gingivalis. B. forsythus and T. denticola were equally prevalent among current, past and never smokers pre-therapy, decreased significantly post-SRP in never and past smokers but increased in current smokers. Clinical improvement post-SRP was accompanied by a modest change in the subgingival microbiota, primarily a reduction in P. gingivalis, B. forsythus and T. denticola, suggesting potential targets for therapy and indicating that radical alterations in the subgingival microbiota may not be necessary or desirable in many patients.
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9,178,114
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In vitro accuracy and reproducibility of automated and conventional periodontal probes.
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The aim of this study was to investigate the accuracy and reproducibility of experienced and inexperienced examiners using 3 automated periodontal probes (Florida Pocket Probe, Florida Disk Probe, Peri Probe) in comparison with 3 conventional periodontal probes (Marquis, Williams and EN-15 probes). Test blocks of aluminium had 30 holes of diameter 1.10 mm and depths ranging from 2.75 to 10.0 mm. machined with a tolerance of +/- 0.01 mm. 8 experienced examiners and 8 inexperienced examiners were selected to perform duplicate measurements on the blocks over 6 visits using each of the 6 probes. 1 automated and 1 conventional probe were used at each examination. The % accuracy and reproducibility for each of the duplicate measurements was calculated and analysed using Friedman 2-way analysis of variance and the Wilcoxon matched pairs test. On average, all probes showed high reproducibility, with the Florida Disk Probe, the Florida Pocket Probe and the Williams probe ranked best and the other 3 probes were less reproducible. On average, all probes showed a high degree of accuracy, automated probes were ranked best and were significantly better than conventional probes. Experience had little effect on reproducibility, with only the Peri Probe showing significant differences at the 5% level between the groups. Experience appeared to be more important for accuracy, as experienced examiners were more accurate than inexperienced examiners, with significant differences at the 5% level for the EN-15, Florida Disk Probe and Peri Probe. However, inexperienced examiners were significantly more accurate using the Williams probe. This in vitro study has shown that automated probes offer increased accuracy over conventional probes and the Florida Pocket and disk probes compare well with conventional probes for reproducibility.
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9,178,115
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Protein degradation by Prevotella intermedia and Actinomyces meyeri supports the growth of non-protein-cleaving oral bacteria in serum.
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The proteolytic activities of oral bacteria are thought to play an important role in the aetiology of dental abscesses. Bacteria-derived proteases may contribute to tissue destruction, and are likely to impair host defence by degrading immunoglobulins and complement. Degraded periodontal tissue and tissue fluid are likely to constitute essential sources of nutrients in the abscess. Tissue fluid, which is derived from serum, is rich in protein and poor in carbohydrate, suggesting that breakdown of protein and fermentation of amino acids is a crucial step to generate energy for growth of the microflora. The number of oral bacterial species that perform hydrolytic cleavage of protein into polypeptides, the first step in protein degradation, is relatively small compared to the large majority of peptidase-producing species. In this study, we therefore investigated the growth-promoting effect of proteinase-producing species like Prevotella intermedia and Actinomyces meyeri on the growth of some non-proteinase producing bacteria in mixed cultures. We used serum as a substitute for the supposed natural substrate of the abscess microflora. The breakdown of serum proteins was investigated using capillary electrophoresis. Poor growth was found in mono- and mixed cultures of non-proteinase producing species Eubacterium lentum, Fusobacterium nucleatum. Peptostreptococcus micros, and Streptococcus intermedius. The presence of P. intermedia in mixed cultures strongly enhanced growth of these 4 species, according to the hypothesis that the growth of the mixed cultures was peptide-limited. The enhanced growth of P. intermedia in pronase-digested serum indicated peptide-limited growth of this organism in serum, despite its production of proteinase. We found that growth of monocultures of Actinomyces meyeri was poor. In contrast, A. meyeri grew well in mixed cultures and its presence stimulated growth of F. nucleatum and P. micros, suggesting a synergistic relationship. The growth of mono- and mixed cultures was investigated using one representative strain of each species. Thus, there is a small risk of having selected unique strains. Proteinase inhibitors reduced the growth of Porphyromonas gingivalis, Prevotella nigrescens, and P. intermedia in trypticase peptone-yeast extract medium with, and without, IgG. Our study indicated that proteinase-producing organisms play a key role in mixed cultures of oral bacteria in human serum by providing polypeptides for growth. This may explain their association with dental abscesses.
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9,178,116
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The impact of the closing of three Massachusetts public chronic disease hospitals: a multidimensional perspective.
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The closing of three public chronic disease hospitals in Massachusetts in 1991 as a cost-cutting measure sparked renewed attention to the consequences of relocation. Massachusetts officials faithfully carried out a series of measures to assure that patients would be transferred to facilities providing high quality care and that the relocation process would be highly sensitive to patient needs. A survey of family representatives revealed that both the relocation process and the outcome tended to be perceived positively. Quasi-experimental studies of health and survival outcomes, however, provided less favorable results. On two of three measures of health change, relocation was found to have no effect. However, relocation was found to increase the likelihood of incontinence. For patients at the hospital with the greatest concentration of older patients, relocation lead to heightened mortality rates. Also disappointing for State officials was the fact that the anticipated cost savings were less than anticipated. The findings point to the need for renewed efforts to understand the circumstances when relocation places institutionalized older people at serious risk, more careful cost estimates of the savings to be achieved through proposed cost-saving policy changes, and more carefully formulated policy guidelines for relocation of the institutionalized elderly that balance the risks associated with relocation against other public policy objectives.
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9,178,118
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Survey of condom-related beliefs, behaviors, and perceived social norms in Mexican migrant laborers.
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This study reports findings from a survey of condom-related beliefs, behaviors, and perceived social norms in Mexican migrant laborers that live and work in the United States for extended periods of time. Snowball sampling was used to recruit 501 Mexican migrants from five "sending towns" in Jalisco, Mexico, with historically high rates of out-migration to the United States. Results showed that subjects reported few negative beliefs about condom use and high efficacy to use condoms in challenging sexual situations but social norms sanctioning condoms were limited. Results also revealed mixed knowledge of HIV transmission, poor knowledge of condom use, and higher condom use with occasional versus regular sex partners. Forty-four percent of male migrants reported sex with prostitutes while in the U.S., with married men reporting less condoms use with prostitutes than single men. It was concluded that condom promotion efforts with Mexican migrants should concentrate on men to encourage consistent use with occasional sex partners, including prostitutes. AIDS prevention education should be provided with sensitivity to the language needs, limited education, and extreme social and geographic marginality of this highly underresearched Latino population.
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9,178,117
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Skin cancer prevention: a time for action.
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Skin cancer is the most common malignancy in the United States accounting for more than 840,000 cases and 9,400 deaths annually. It is estimated that 90% of non-melanoma skin cancers and much of melanoma incidence can be attributed to sun exposure. The evidence suggests that regular use of sunscreen (Sun Protective Factor (SPF) of 15 or higher), wearing protective, tightly woven clothing and wide brimmed hats, and avoiding sun exposure when the ultraviolet rays are strongest (between 11:00 a.m. and 3:00 p.m.) can dramatically reduce the risk of skin cancer. Interventions to promote sun-protection behaviors that target children and adults are necessary to reduce the growing incidence rate of skin cancer in the United States.
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9,178,119
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An examination of differential follow-up rates in cervical cancer screening.
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The purpose of this study was to test the hypothesis that follow-up rates for women with abnormal cervical cancer screening results vary by age, ethnicity, and initial screening results in California's Breast and Cervical Cancer Control Program. The sample consisted of women in the screening program who received an abnormal cervical screening result (N = 1.738). Bivariate and logistic regression analyses were utilized to examine variables that account for differences in follow-up rates among these women. Bivariate analysis showed significant differences by age, race/ethnicity, initial screening results, and urban/rural residence. In logistic regression analysis, these variables also retained significance. Severity of diagnosis was a highly significant predictor of follow-up. Women of color, older women, and women with less severe diagnoses should be targeted as groups needing assistance in adhering to follow-up recommendations.
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9,178,120
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Overweight and obesity in Saudi Arabian adult population, role of socio-demographic variables.
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The objectives of this Community-based National Epidemiological Household Survey, conducted between 1990-1993, were to estimate the prevalence of overweight and obesity in Saudi Arabia and to examine its association with the socio-demographic characteristics of the adult population. A sample of Saudis 20 years and over was selected using a multistage stratified cluster sampling technique with probability proportionate to size. The selected subjects were requested to visit primary health care centers in their localities. Physicians in these clinics took measurements of heights and weights and collected other relevant data. Obesity was measured by the Body Mass Index, using the Quetelet Index. The results showed the sample of 10,651 subjects of which 50.8% were males, had a mean age of 35.8 years (SD = 14.27 years). The prevalence of overweight was 31.2% (95% confidence interval: 30.3%, 32.1%); 33.1% for males and 29.4% for females. For obesity, the overall prevalence was 22.1%; males 17.8% and females 26.6%. The study design suggested that these estimates could be closer to the true values. The multiple logistic regression analysis showed that age, residential area, region, income, gender, and education are statistically significant predictors of obesity. The prevalence of obesity was higher in females than males, lower in subjects living in rural areas with traditional lifestyles than those in more urbanized environments, and increased with increasing age. The observed prevalence and pattern of overweight and obesity with age and gender is similar to those observed in the Arab community and some Western nations. There is a need for increased physical activity and better nutrition education programs to reduce the extent of obesity and to prevent the serious health consequences, especially, in the middle age group.
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9,178,122
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Effects of monensin on the metabolism of periparturient dairy cows.
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The effects of monensin on plasma concentrations and changes in plasma concentrations of energy metabolites and minerals over time were investigated using 24 multiparous Holstein cows. Cows were paired according to farm, predicted date of calving, and body condition score and were randomly allocated to two groups. Treated cows were given a ruminal bolus containing 32 g of monensin at 50 +/- 7 d before predicted calving. Treated cows had lower plasma concentrations of glucose, free fatty acid (FFA), and beta-hydroxybutyrate (BHBA) than did control cows before calving, indicating that monensin influenced energy metabolism. However, no significant differences in plasma concentrations of glucose, FFA, and BHBA were found between groups after calving. Plasma BHBA concentrations increased more before calving in control cows, and plasma FFA and urea concentrations increased significantly before calving in all cows. No significant differences in body weight, plasma concentrations of urea, or whole blood concentrations of glutathione peroxidase were detected between groups before or after calving. Plasma ceruloplasmin activity did not differ between groups before calving, but was significantly higher in treated cows after calving. Plasma concentrations of Ca did not significantly differ between groups before or after calving. Monensin altered both energy and mineral metabolism and has the potential to improve the health and production of dairy cows.
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9,178,121
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Bovine somatotropin dose titration in lactating dairy ewes. 2. Dose determination and factors affecting the response.
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Seventy-four lactating dairy ewes were injected with recombinant bovine somatotropin (bst; sometribove) in a sustained-release formulation. Ewes received 0, 80, 160, or 240 mg of bST/14 d from wk 3 to 8 of lactation (part 1) and 0, 80, or 160 mg of bST/14 d from wk 11 to 18 of lactation (part 2). The optimal dose of bST was studied as well as the factors (lactation stage, lactation number, initial milk production, body weight, and body condition) possibly affecting the increase in milk production following bST injection. Using a quadratic regression model, the maximum theoretical dose was determined to be 181 mg of bST/14 d during the first part of lactation. During the second part of lactation, 143 mg of bST/14 d was the maximum theoretical dose. The increment of milk production did not vary with lactation number, but first lactation ewes, in general, responded better than did multiparous ewes. Relative to initial milk production, improvement was greatest for ewes with average milk production (1500 ml/d) that received a dose of 192.3 mg of bST/14 d during the first part of lactation; improvement was also measured from the highest producers (2000 ml/d) during the second part of lactation. The best response was obtained from ewes with average body condition (score 3 on a five-point scale where 1 = thin to 5 = fat) and a dose of 200 mg of bST/14 d during the first part of lactation; during the second part of lactation, body condition score had no effect. Body weight had no effect on the increment of production at any time.
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9,178,123
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Composition of colostrum from dairy heifers exposed to high air temperatures during late pregnancy and the early postpartum period.
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This study examined the effects of heat stress on composition of colostrum from primiparous cows during late pregnancy and the early postpartum period. Two groups of 6 Holstein heifers were utilized. During the last 3 wk of pregnancy and during the first 36 h after calving, one group was exposed to thermal comfort (temperature-humidity index = 65); the other group was exposed to high air temperatures (temperature-humidity index = 82 from 0900 to 2000 h and temperature-humidity index = 76 from 2100 to 0800 h). Heifers under heat stress had higher rectal temperatures and respiratory rates; lower plasma triiodothyronine and glucose; higher plasma nonesterified fatty acids and beta-hydroxybutyrate; lower intakes of dry matter, net energy for lactation, and crude protein; higher water intakes; and lower body condition scores. The decline of plasma immunoglobulins (Ig) over the final 2 wk of pregnancy was less pronounced for heifers under heat stress. For the first four milkings, colostrum of cows exposed to high air temperatures had lower mean concentrations of IgG and IgA; lower mean percentages of total protein, casein, lactalbumin, fat, and lactose; lower contents (grams per liter) of short- and medium-chain fatty acids; lower energy; lower titratable acidity; and higher pH. Thus, high air temperatures during late pregnancy and the early postpartum period markedly affected the composition of colostrum from primiparous dairy cows.
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