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Biomedical subjects

A Butler

Publications and source records attributed to A Butler.

At least 19 recordsLinked to original sources
PubMed1997

Thyroid hormone-mediated enhancement of heterodimer formation between thyroid hormone receptor beta and retinoid X receptor.

A subset of nuclear receptors, including those for thyroid hormone (TR), retinoic acid, vitamin D3, and eicosanoids, can form heterodimers with the retinoid X receptor (RXR) on DNA regulatory elements in the absence of their cognate ligands. In a mammalian two-hybrid assay, we have found that recruitment of a VP16-RXR chimera by a Gal4-TRbeta ligand-binding domain fusion is enhanced up to 50-fold by thyroid hormone (T3). This was also observed with a mutant fusion, Gal4-TR(L454A), lacking ligand-inducible activation function (AF-2) and unable to interact with putative coactivators, suggesting that the AF-2 activity of TR or intermediary cofactors is not involved in this effect. The wild-type and mutant Gal4-TR fusions also exhibited hormone-dependent recruitment of RXR in yeast. Hormone-dependent recruitment of RXR was also evident with another Gal4-TR mutant, AHTm, which does not interact with the nuclear receptor corepressor N-CoR, suggesting that ligand-enhanced dimerization is not a result of T3-induced corepressor release. Finally, we have shown that the interaction between RXR and TR is augmented by T3 in vitro, arguing against altered expression of either partner in vivo mediating this effect. We propose that ligand-dependent heterodimerization of TR and RXR in solution may provide a further level of control in nuclear receptor signaling.

Cell Nucleus↗
PubMed1997

Behavioral defects in C. elegans egl-36 mutants result from potassium channels shifted in voltage-dependence of activation.

Mutations in the C. elegans egl-36 gene result in defective excitation of egg-laying and enteric muscles. Dominant gain-of-function alleles inhibit enteric and egg-laying muscle contraction, whereas a putative null mutation has no observed phenotype. egl-36 encodes a Shaw-type (Kv3) voltage-dependent potassium channel subunit. In Xenopus oocytes, wild-type egl-36 expresses noninactivating channels with slow activation kinetics. One gain-of-function mutation causes a single amino acid substitution in S6, and the other causes a substitution in the cytoplasmic amino terminal domain. Both mutant alleles produce channels dramatically shifted in their midpoints of activation toward hyperpolarized voltages. An egl-36::gfp fusion is expressed in egg-laying muscles and in a pair of enteric muscle motor neurons. The mutant egl-36 phenotypes can thus be explained by expression in these cells of potassium channels that are inappropriately opened at hyperpolarized potentials, causing decreased excitability due to increased potassium conductance.

Animals↗
PubMed1997

Structure of the mouse serum amyloid A 5 (Saa5) gene: relationship to other members of the serum amyloid A family.

The mouse and human genes which encode the serum amyloid A (SAA) proteins have been extensively characterized. In the present study, the authors report the sequencing of the constitutively expressed mouse Saa5 gene, including approximately 2 kilobases of promoter sequence which extends into the sequence of the adjacent Saa4 pseudogene. The Saa5 gene was thus determined to have a 4 exon/3 intron structure, with a B2 repeat located approximately 450 base pairs upstream, and a microsatellite contained within intron 2. Analyses of Saa5 intronic and promoter sequences, by comparison with representative members of the mouse Saa and human SAA families, provided evidence which strongly suggests that mouse Saa5 and the constitutively expressed human SAA4 gene are evolutionary and functional homologues. Furthermore, putative transcription motifs were identified within the Saa5 and SAA4 promoters which may account, in part, for the constitutive expression and unique induction profiles of these genes.

Animals↗
PubMed1997

Sequence-specific epigenetic effects of the maternal somatic genome on developmental rearrangements of the zygotic genome in Paramecium primaurelia.

In ciliates, the germ line genome is extensively rearranged during the development of the somatic macronucleus from a mitotic product of the zygotic nucleus. Germ line chromosomes are fragmented in specific regions, and a large number of internal sequence elements are eliminated. It was previously shown that transformation of the vegetative macronucleus of Paramecium primaurelia with a plasmid containing a subtelomeric surface antigen gene can affect the processing of the homologous germ line genomic region during development of a new macronucleus in sexual progeny of transformed clones. The gene and telomere-proximal flanking sequences are deleted from the new macronuclear genome, although the germ line genome remains wild type. Here we show that plasmids containing nonoverlapping segments of the same genomic region are able to induce similar terminal deletions; the locations of deletion end points depend on the particular sequence used. Transformation of the maternal macronucleus with a sequence internal to a macronuclear chromosome also causes the occurrence of internal deletions between short direct repeats composed of alternating thymines and adenines. The epigenetic influence of maternal macronuclear sequences on developmental rearrangements of the zygotic genome thus appears to be both sequence specific and general, suggesting that this trans-nucleus effect is mediated by pairing of homologous sequences.

Animals↗
PubMed1996

Mapping genes within a YAC by computer-assisted interpretation of partial restriction digestions.

Partial restriction digestion is used to map restriction sites and the location of genes within yeast artificial chromosomes (YACs). Locus-specific probes are hybridised to the partially digested YAC DNA and the fragments to which they hybridise are compared with the pattern of partial digestion products that include each map region. A least squares criterion is presented which allows for error in fragment length determination. This rapidly defines the most likely location of a marker within the restriction map and permits the combination of results from digestions with different restriction enzymes. Approximate confidence intervals may be assigned to gene locations, and tests of goodness-of-fit of the data may be performed. Since the number of erroneously matched fragments increases in proportion to the square of the number of sites, denser maps are not necessarily more informative. Simulations indicate that the optimal number of internal restriction sites given typical experimental error (1% of YAC length) is about five sites; the associated broad support interval (on average one third of YAC length) may be reduced by combining results from different enzyme digestions. Application of a computer implementation of this model to experimental data showed that the model fitted well, and estimates of location were found to be consistent with other evidence.

Animals↗
PubMed1996

A gene map of the human genome.

The human genome is thought to harbor 50,000 to 100,000 genes, of which about half have been sampled to date in the form of expressed sequence tags. An international consortium was organized to develop and map gene-based sequence tagged site markers on a set of two radiation hybrid panels and a yeast artificial chromosome library. More than 16,000 human genes have been mapped relative to a framework map that contains about 1000 polymorphic genetic markers. The gene map unifies the existing genetic and physical maps with the nucleotide and protein sequence databases in a fashion that should speed the discovery of genes underlying inherited human disease. The integrated resource is available through a site on the World Wide Web at http://www.ncbi.nlm.nih.gov/SCIENCE96/.

Amino Acid Sequence↗
PubMed1996

Inactivation of vanadium bromoperoxidase: formation of 2-oxohistidine.

The basis of the irreversible inactivation of the vanadium bromoperoxidase (V-BrPO) isolated from the marine alga Ascophyllum nodosum under turnover conditions at low pH (i.e., 15 to 100 mM H2O2, 0.1 KBr, ca. 15 nM V-BrPO in 0.1 M citrate, pH 4) has been investigated. Inactivation under these conditions was found to produce 2-oxohistidine as identified by HPLC using electrochemical detection. Formation of 2-oxohistidine requires all the components of turnover (i.e., bromide, hydrogen peroxide, and V-BrPO) as well as low pH; inactivation does not occur nor is significant 2-oxohistidine formed in the presence of hydrogen peroxide alone. The oxidation of histidine did not occur by singlet oxygen generated by V-BrPO, because neither 2-oxohistidine nor inactivation occur under the conditions in which singlet oxygen is produced quantitatively by V-BrPO. The addition of aqueous bromine to N alpha-benzoylhistidine at low pH formed N alpha-benzoyl-2-oxohistidine. cis-Dioxovanadium(V) (VO2+) in strong acid and MoO(O2)2(ox)2- (ox2- is oxalate) at pH 5, both of which are functional mimics of V-BrPO by oxidizing bromide by hydrogen peroxide, catalyzed the oxidation of N alpha-benzoylhistidine to N alpha-benzoyl-2-oxohistidine. Furthermore, when hypobromite was added to N alpha-benzoylhistidine in the presence of hydrogen peroxide at neutral pH, conditions under which HOBr would react first with H2O2 to produce singlet oxygen, no N alpha-benzoyl-2-oxohistidine was formed. Thus the oxidation of histidine in V-BrPO is proposed to occur via oxidized bromine species. Irreversible inactivation V-BrPO was also found to be accompanied by release of vanadium.

Bromates↗
PubMed1996

Mapping of the mouse serum amyloid A gene cluster by long-range polymerase chain reaction.

The present study defines the organization of the mouse serum amyloid A (Saa) gene cluster on chromosome 7. A polymerase chain reaction (PCR)-based strategy was used successfully to generate a complete map of the mouse Saa genes, defining a linkage group of 3'-Saa2-5'/5'-Saa1-3'/5'-Saa4-3'/5'-Saa5-3'/5'-+ ++Saa3-3', with a maximum size of 45 kilobases (kb). This contrasts with the 150 kb human SAA gene cluster, which has been previously defined. The tight linkage of both mouse Saas and human SAAs is of potential functional significance, since the genes that encode the acute phase serum amyloid A proteins are known to exhibit co-ordinate transcriptional regulation. The present results thus suggest that selective pressure may exist which maintains the co-ordinately transcribed Saa genes in close physical proximity. This study, furthermore, demonstrates the utility of a novel PCR-based approach for fine mapping of tightly clustered linkage groups. The strategy used possesses a number of advantages over previously described techniques, such as long-range restriction mapping, since it facilitates the concurrent determination of not only precise relative map positions, but also the relative transcriptional orientations of assayed paired loci. Although presently limited in resolution to genes not more than 27 kb apart, future technical advances are likely to extend the applicability of this approach in mapping experiments to less tightly linked clusters of genes.

Animals↗
PubMed1996

The messenger RNAs for the N-methyl-D-aspartate receptor subunits show region-specific expression of different subunit composition in the human brain.

The expression of the messenger RNAs encoding N-methyl-D-aspartate receptor subunits in neurologically normal post-mortem human brain was studied by in situ hybridization. In the caudate, putamen and nucleus accumbens strong hybridization signals were observed for N-methyl-D-aspartate R1-1 messenger RNA but much weaker signals for N-methyl-D-aspartate R1-3 and N-methyl-D-aspartate R1-4, N-Methyl-D-aspartate R1-2 was not detectable. N-methyl-D-aspartate R2B was the only N-methyl-D-aspartate R2 subunit detected in these nuclei. In the hippocampus the messenger RNAs for both N-methyl-D-aspartate R1-1 and N-methyl-D-aspartate R1-4 were strongly expressed in the dentate gyrus, CA3-CA1 pyramidal cells, subiculum, entorhinal cortex and perirhinal cortex. Much lower expression was seen for N-methyl-D-aspartate R1-2 and N-methyl-D-aspartate R1-3. The messenger RNAs for both N-methyl-D-aspartate R2A and N-methyl-D-aspartate R2B, but not N-methyl-D-aspartate R2C, subunits were expressed in the hippocampus. In the temporal cortex all N-methyl-D-aspartate RI isoforms were expressed (N-methyl-D-aspartate R1-1 and N-methyl-D-aspartate R1-4 being the most abundant) and N-methyl-D-aspartate R2A and N-methyl-D-aspartate R2B but not N-methyl-D-aspartate R2C were also moderately expressed. In the brain stem N-methyl-D-aspartate R1-4 was strongly expressed in various nuclei including the locus coeruleus, nucleus centralis superior and deep pontine nuclei. Only weak expression was seen for N-methyl-D-aspartate RI-1 and N-methyl-D-aspartate R1-3 but not N-methyl-D-aspartate RI-2; of the N-methyl-D-aspartate R2 subunits only N-methyl-D-aspartate R2C was found to be expressed in these nuclei. In the cerebellum all the N-methyl-D-aspartate I isoforms were expressed (mostly N-methyl-D-aspartate R1-4) in the Purkinje layer which also expressed N-methyl-D-aspartate R2A and N-methyl-D-aspartate R2C. In the molecular layer cells were found expressing N-methyl-D-aspartate R1-4 and N-methyl-D-aspartate R2B and cells in the granule layer were found to express N-methyl-D-aspartate R1-1, N-methyl-D-aspartate R1-3 and N-methyl-D-aspartate R1-4 and N-methyl-D-aspartate R2C only. Preliminary studies indicated that the messenger RNA for the N-methyl-D-aspartate R2D subunit was not expressed in the above areas of brain. These results give the first demonstration of the distribution of N-methyl-D-aspartate receptor subunit messenger RNAs in the human brain. The region-specific expression of subunit combinations suggests a heterogeneity of N-methyl-D-aspartate receptors with diverse physiological/pathophysiological roles and provides a rationale for the development of discriminatory N-methyl-D-aspartate receptor antagonists to target selective neuronal populations.

Aged↗
PubMed1996

Interleukins-6 and -8 levels in sera and cervical mucus of fertile, idiopathic infertile, and immunoinfertile women: implication in infertility.

PROBLEM: The purpose of this study was to investigate the levels of interleukin-6 and -8 (IL-6 and IL-8) in sera and cervical mucus of infertile (idiopathic and immunoinfertile) women and to compare the levels with those in age-matched normally cycling fertile women. METHODS: Levels of IL-6 and IL-8 were measured in the sera and cervical mucus of fertile and infertile women by the enzyme-linked immunosorbent assay (ELISA). A non-parametric Mann-Whitney test was used to evaluate significance between the means. Linear regression analysis of IL-6 and IL-8 concentrations in serum versus cervical mucus and with antisperm antibody titers was performed by condition as well as all groups together. P < 0.05 was considered significant. RESULTS: Both IL-6 and IL-8 were detected in sera and cervical mucus of fertile as well as infertile groups. Although serum levels of IL-6 and IL-8 were significantly different between fertile and infertile groups, the differences were more pronounced in the cervical mucus samples. Cervical mucus of idiopathic and immunoinfertile women demonstrated significantly (P = 0.002 to < 0.0001) greater concentrations of IL-6 and IL-8 compared to fertile controls. In general, there was no significant correlation between the serum and cervical mucus IL-6 and IL-8 levels, whether analyzed by condition or all groups together. CONCLUSIONS: Elevated levels of IL-6 and IL-8 in cervical mucus of infertile groups may play a role in etiology of idiopathic and immunologic infertility. These findings suggest that the measurements of cytokines (especially IL-6 and IL-8) in cervical mucus may provide a tool for specific diagnosis and possibly treatment of infertility, especially idiopathic infertility.

Antibodies↗
PubMed1996

Frequency of the apolipoprotein E epsilon 4 allele in a case-control study of early onset Parkinson's disease.

OBJECTIVES: It has been suggested that Parkinson's disease and Alzheimer's disease may share a common or at least overlapping aetiology. The prevalence of dementia among cases of Parkinson's disease is known to be greater than expected in the general population. The frequency of the apolipoprotein epsilon 4 allele in a large case-control study of early onset Parkinson's disease has been examined. METHODS: 215 patients and 212 population based controls were recruited from the Republic of Ireland between 1992 and 1994. Cases had to have disease onset at 55 years or younger and be born after 1925. RESULTS: The frequency of the epsilon 4 allele was almost identical between cases of Parkinson's disease (14.6%) and healthy controls (13.3%). There was no relation between epsilon 4 status and disease onset, disease duration, Hoehn and Yahr score, and disease progression. The frequency of the epsilon 4 allele was not increased among 10 patients with Parkinson's disease with dementia (10.0%) compared with the other patients without dementia (14.8%). There was no association between epsilon 4 allele status and either a history of smoking, family history of dementia, or Parkinson's disease, or being born in a rural area. The odds ratio for the ApoE epsilon 4 allele associated with Parkinson's disease was 1.10 (95% confidence interval (95% CI) 0.68-1.79), adjusting for age group, sex, and residential status. The pooled odds ratio from a meta-analysis of six studies of ApoE epsilon 4 status and Parkinson's disease was 0.94 (95% CI 0.69-1.27). CONCLUSIONS: The results from our study as well as the pooled meta-analysis exclude any important role for ApoE epsilon 4 status in the development of Parkinson's disease. Our results similarly do not support its role either in dementia associated with Parkinson's disease or disease prognosis.

Age of Onset↗
PubMed1996

Micronucleus induction in human lymphocytes: comparative effects of X rays, alpha particles, beta particles and neutrons and implications for biological dosimetry.

The cytokinesis-block micronucleus assay in peripheral blood lymphocytes has the potential for being a simple and rapid method of biological dosimetry. This technique has been used to study the induction of micronuclei in the blood from 12 donors after exposure to a range of radiations with track-averaged LET values ranging from 0.26 to 44 keV microns -1. Data based on the average response of the 12 individuals for 250 kVp X rays were found to agree well with results published previously from other laboratories using similar techniques. Low dose-limiting RBE values relative to 250 kVp X rays for the radiations studied were found to be 0.50 for strontium/yttrium-90 beta particles, 6.9 for 20-23 keV microns -1 alpha particles and 17 for 24 keV neutrons. The pattern of the variation of individual radiosensitivity was found to be complex and dependent on dose, and the evaluation of individual radiosensitivity based on the response at one dose only can be misleading. It is concluded that, although the cytokinesis-block micronucleus assay in blood lymphocytes is a radiobiologically appropriate technique to use for biological dosimetry, its practical implementation may be limited by a need to perform individual pre-exposure calibrations.

Adult↗
PubMed1995

Inhibition and inactivation of vanadium bromoperoxidase by the substrate hydrogen peroxide and further mechanistic studies.

Hydrogen peroxide, which is a substrate of vanadium bromoperoxidase (V-BrPO), has been shown to be a noncompetitive inhibitor of V-BrPO. Hydrogen peroxide inhibition increases with increasing pH. The inhibition is reversible under the conditions of the initial steady-state kinetic experiments. Analysis of the inhibition constants (KiiH2O2, KisH2O2) versus H+ concentration indicates that an ionizable group with a pKa between 6.5 and 7 is involved in the inhibition. The origin of the oxygen atoms in the dioxygen produced by the V-BrPO-catalyzed bromide-assisted disproportionation of hydrogen peroxide has been shown through H2(18)O2 labeling experiments to originate from the same molecule of hydrogen peroxide. V-BrPO-catalyzed bromination is shown to be an electrophilic (Br+) as opposed to a radical (Br.) process. The stoichiometry of H2O2 consumed to MCD reacted or to O2 produced is reported. The concentration of hydrogen peroxide also affects the competition of dioxygen formation during MCD bromination; competitive dioxygen formation is strongly enhanced at high pH. Turnover of V-BrPO under conditions of very high hydrogen peroxide concentration leads to irreversible inactivation at pH 4 and pH 5. Much less inactivation occurs during turnover at long reaction times at higher pH (> pH 6), and the inactivation can be fully reversed by subsequent addition of vanadate.

Bromides↗
PubMed1995

Epigenetic self-regulation of developmental excision of an internal eliminated sequence on Paramecium tetraurelia.

Differentiation of the somatic macronucleus of ciliates after sexual events involves the programmed excision of thousands of single-copy internal eliminated sequences (IESs) from the germ-line genome. We have studied two cell lines of Paramecium tetraurelia that have identical germ-line genomes but differ in their macronuclear genomes. In the IES- cell line, a 222-bp IES interrupting a coding sequence is reproducibly excised during macronuclear differentiation, whereas it is not in the IES+ cell line. In a cross between the two lines, the developmental alternative in maternally inherited, suggesting that it is epigenetically controlled by the old (prezygotic) macronucleus in each cell. Transformation of the macronucleus of both lines with plasmids carrying fragments of either version of the gene shows that the presence of the IES sequence in the old macronucleus results in retention of the IES in the new macronuclear genome of sexual progeny. This could be attributable to (1) inhibition of excision, or (2) repair of a double-strand gap left in the genomic sequence after constitutive excision of the IES, by a polymerization mechanism using a homologous IES+ template from the old macronucleus. The latter possibility is ruled out by experiments showing that modified IESs can inhibit excision without being copied in the new macronuclear genome. Possible mechanisms are discussed in the light of a quantitative analysis of excision inhibition by the maternal IES sequence.

Animals↗
PubMed1995

Levels of interferon-gamma and tumor necrosis factor-alpha in sera and cervical mucus of fertile and infertile women: implication in infertility.

Concentrations of two immune cytokines, namely interferon-gamma (INF-gamma) and tumor necrosis factor-alpha (TNF-alpha), were determined in the sera and cervical mucus samples of fertile (n = 16), idiopathic infertile (n = 44), and immunoinfertile women (n = 45) to investigate their role, if any, in female infertility. Sera of idiopathic infertile women demonstrated significantly (P < 0.0001) higher levels of INF-gamma compared to those in fertile controls, whether expressed as pg/ml or pg/mg serum protein. Similarly, sera of immunoinfertile women demonstrated significantly (P = 0.0008) higher levels of INF-gamma compared to fertile controls and idiopathic infertile women. Cervical mucus of idiopathic infertile women also demonstrated significantly (P < 0.0001) higher concentrations of INF-gamma compared to those in fertile controls. Cervical mucus of immunoinfertile women demonstrated significantly (P < 0.0001) higher concentrations of INF-gamma compared to those in fertile controls and idiopathic infertile women. INF-gamma levels in serum did not significantly (P > 0.05) correlate (r = 0.12-0.43) with the concentrations in cervical mucus, when all the three groups were compared together. However, when the serum levels were compared with the cervical mucus concentrations by condition, only the idiopathic infertile group showed a significant (P = 0.005) correlation (r = 0.70). Serum levels of TNF-alpha did not differ significantly (P > 0.05) among three groups of women. Cervical mucus concentrations of TNF-alpha, however, varied among groups with levels being significantly (P = 0.04) higher-in idiopathic infertile women compared with fertile controls and in immunoinfertile women significantly (P = 0.0007) higher than in fertile controls as well as idiopathic infertile women. TNF-alpha levels in serum correlated (r = 0.65) significantly (P < 0.001) with the concentrations in cervical mucus when all the three groups were compared together or individually by infertility condition. These findings suggest the involvement of cytokines in infertility, and thus may have potential applications in diagnosis and treatment of female infertility.

Adult↗
PubMed1994

Resistance to secondary amyloidosis in A/J mice is not significantly associated with allelic variants linked to the serum amyloid A gene cluster.

We have tested the hypothesis that structural allelic variants of serum amyloid A confer relative resistance to secondary amyloidosis in the A/J mouse. F2 mice, previously generated from amyloid-resistant (A/J) and amyloid-susceptible (C57BL/6J) strains and categorized with respect to amyloid susceptibility, were genotyped by polymerase chain reaction (PCR) amplification of the polymorphic D7Nds5 microsatellite. This microsatellite is closely linked to the SAA gene cluster and can discriminate between D7Nds5 alleles of A/J and C57BL/6J origin. The distribution of D7Nds5 genotypes in relation to splenic amyloid load did not deviate significantly from that expected of a random distribution, indicating that A/J amyloid resistance is not determined by variants at, or close to, D7Nds5. Therefore, structural alleles in the tightly-linked SAA gene cluster do not confer amyloid resistance in this mouse model.

Alleles↗