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At least 271 records · Page 15Linked to original sources
PubMed2002

The amino acid residues asparagine 354 and isoleucine 372 of human farnesoid X receptor confer the receptor with high sensitivity to chenodeoxycholate.

The critical steps in bile acid metabolism have remarkable differences between humans and mice. It is known that human cholesterol 7 alpha-hydroxylase, the enzyme catalyzing the rate-limiting step of bile acid synthesis, is more sensitive to bile acid suppression. In addition, hepatic bile acid export in humans is more dependent on the bile salt export pump (BSEP). To explore the molecular basis for these species differences, we analyzed the function of the ligand-binding domain (LBD) of human and murine farnesoid X receptor (FXR), a nuclear receptor for bile acids. We observed a strong interspecies difference in bile acid-mediated FXR function; in the coactivator association assay, chenodeoxycholate (CDCA) activated human FXR-LBD with 10-fold higher affinity and 3-fold higher maximum response than murine FXR-LBD. Consistently, in HepG2 cells human FXR-LBD increased reporter expression more robustly in the presence of CDCA. The basis for these differences was investigated by preparing chimeric receptors and by site-directed mutagenesis. Remarkably, the double replacements of Lys(366) and Val(384) in murine FXR (corresponding to Asn(354) and Ile(372) in human FXR) with Asn(366) and Ile(384) explained the difference in both potency and maximum activation; compared with the wild-type murine FXR-LBD, the double mutant gained 8-fold affinity and more than 250% maximum response to CDCA in vitro. This mutant also increased reporter expression to an extent comparable with that of human FXR-LBD in HepG2 cells. These results demonstrate that Asn(354) and Ile(372) are critically important for FXR function and that murine FXR can be "humanized" by substituting with the two corresponding residues of human FXR. Consistent with the difference in FXR-LBD transactivation, CDCA induced endogenous expression of human BSEP by 10-12-fold and murine BSEP by 2-3-fold in primary hepatocytes. This study not only provides the identification of critical residues for FXR function but may also explain the species difference in bile acids/cholesterol metabolism.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2006

Personality disorder and the outcome of depression: meta-analysis of published studies.

BACKGROUND: There is conflicting evidence about the influence of personality disorder on outcome in depressive disorders. AIMS: Meta-analysis of studies in which a categorical assessment of personality disorder or no personality disorder was made in people with depressive disorders, and categorical outcome (recovered/not recovered) also determined. METHOD: Systematic electronic search of the literature for relevant publications. Hand searches of Journal of Affective Disorders and recent reviews, with subsequent meta-analysis of selected studies. RESULTS: Comorbid personality disorder with depression was associated with a doubling of the risk of a poor outcome for depression compared with no personality disorder (random effects model OR=2.18, 95% CI 1.70-2.80), a robust finding maintained with only Hamilton-type depression criteria at outcome (OR=2.20, 95% CI 1.61-3.01). All treatments apart from electroconvulsive therapy (ECT) showed this poor outcome, and the ECT group was small. CONCLUSIONS: Combined depression and personality disorder is associated with a poorer outcome than depression alone.

Antidepressive Agents↗
PubMed1992

Model misspecification and multipoint linkage analysis.

Pairwise linkage analysis is robust to genetic model misspecification provided dominance is correctly specified, the primary effect being inflation of the recombination fraction. By contrast, we show that multipoint analysis under misspecified models is not robust when a putative disease locus is placed between close flanking markers, with potentially spuriously negative multipoint lod scores being produced. The problem is due to incorrect attribution of segregation of a disease allele and the consequent conclusion of (unlikely) double crossovers between flanking markers. As a possible solution, we propose the use of high disease allele frequencies, as this allows probabilistically for nonsegregation (through parental homozygosity or dual matings). We show analytically and through analysis of pedigree data simulated under a two-locus heterogeneity model that using a disease allele frequency of 0.05 in the dominant case and 0.25 in the recessive case is quite robust in producing positive multipoint lod scores with close flanking markers across a broad range of conditions including varying allele frequencies, epistasis, genetic heterogeneity and phenocopies.

Animals↗
PubMed1995

Dose-response relationship of levodopa with mood and anxiety in fluctuating Parkinson's disease: a double-blind, placebo-controlled study.

We investigated the effect of levodopa on mood and anxiety in eight Parkinson's disease patients with motor fluctuations. Each patient received 0.0-, 0.5-, and 1.0-mg/kg/hr levodopa infusions in randomly assigned order under double-blind conditions on consecutive days. Mood elevation and anxiety reduction based on half-hourly patient rating and a corresponding increase in tapping speed occurred with active drug infusion but not placebo infusion. The effects were dose related. The higher-dose infusion rate produced more rapid onset, greater magnitude, and longer duration of response. We conclude that mood and anxiety fluctuations related to levodopa dosing are robust pharmacologic, and not placebo, effects.

Affect↗
PubMed1997

New strategy for the construction of single-stranded plasmids with single mutagenic lesions.

Single-stranded DNA vectors containing single adducts offer a unique opportunity to study the biochemistry and genetics of trans lesion synthesis, a process during which a DNA polymerase synthesizes across a lesion. We describe a new and general strategy to produce high-quality single-stranded plasmids containing a single adduct within a predetermined sequence context starting with a short oligonucleotide containing the lesion of interest. These vectors are isolated from the corresponding double-stranded constructs by selective enzymatic degradation in vitro of the nonadducted uracil-containing strand. Efficient and complete removal of this strand was achieved using uracil DNA glycosilase to generate AP sites followed by the action of the AP endonuclease associated with exonuclease III and the robust 3'-->5' exonuclease activity associated with T7 DNA polymerase. We show the utility of these constructs for the study of trans lesion synthesis in vitro and in vivo in the case of the highly carcinogenic N-2-acetylaminofluorene adducts located within frameshift mutation hot spots. The possibility to construct both single-stranded and double-stranded plasmids, with the same origin of replication (i.e., ColE1), will allow a direct comparison between single-stranded and double-stranded DNA replication in site-specific mutagenesis studies.

DNA Adducts↗
PubMed2004

4-1BB and OX40 act independently to facilitate robust CD8 and CD4 recall responses.

Mice deficient in OX40 or 4-1BB costimulatory pathways show defects in T cell recall responses, with predominant effects on CD4 vs CD8 T cells, respectively. However, OX40L can also stimulate CD8 T cells and 4-1BBL can influence CD4 T cells, raising the possibility of redundancy between the two TNFR family costimulators. To test this possibility, we generated mice deficient in both 4-1BBL and OX40L. In an adoptive transfer model, CD4 T cells expressed 4-1BB and OX40 sequentially in response to immunization, with little or no overlap in the timing of their expression. Under the same conditions, CD8 T cells expressed 4-1BB, but no detectable OX40. Thus, in vivo expression of 4-1BB and OX40 can be temporally and spatially segregated. In the absence of OX40L, there were decreased CD4 T cells late in the primary response and no detectable secondary expansion of adoptively transferred CD4 T cells under conditions in which primary expansion was unaffected. The 4-1BBL had a minor effect on the primary response of CD4 T cells in this model, but showed larger effects on the secondary response, although 4-1BBL(-/-) mice show less impairment in CD4 secondary responses than OX40L(-/-) mice. The 4-1BBL(-/-) and double knockout mice were similarly impaired in the CD8 T cell response, whereas OX40L(-/-) and double knockout mice were similarly impaired in the CD4 T cell response to both protein Ag and influenza virus. Thus, 4-1BB and OX40 act independently and nonredundantly to facilitate robust CD4 and CD8 recall responses.

4-1BB Ligand↗
PubMed2005

On-chip PCR amplification of very long templates using immobilized primers on glassy surfaces.

In this paper we describe a novel method for visualizing very long DNA fragments (for example >6 kb) which are difficult to spot with commonly used arrayers or capillary samplers with very small nanoliter volumes, using directly bound primers on "on-chip" polymerase chain reaction (PCR). We have used the genomes of the M13 bacteriophage (7.2 kb) the human mitochondrion (16.5 kb) as examples of long DNA templates to test the PCR and were able to elicit robust reactivity. Over 75% of the immobilized primers could be elongated to their fullest extent. In addition we were able to elicit the PCR reaction with double stranded templates in which one primer was immobilized and the other suspended in the reaction solution. These synthesized PCR products were visualized by either confocal microarray scanning or fluorescence microscopy using Cy5-dye fluorescence of the modified free primer, or the fluorescence of intercalating dyes.

Adsorption↗
PubMed2004

Uniform-sign cross-peak double-quantum-filtered correlation spectroscopy.

We detail the uniform-sign cross-peak double-quantum-filtered correlation spectroscopy (UC2QF COSY) experiment, a new through-bond correlation method for disordered solids. This experiment is a refocused version of the popular double-quantum-filtered correlation spectroscopy experiment in liquids. Its key feature is that it provides in-phase and doubly absorptive line shapes, which renders it robust for chemical shift correlation in solids. Both theory and experiment point to distinct advantages of this protocol, which are illustrated by several experiments under challenging conditions, including fast magic-angle spinning (30kHz), anisotropic molecular motion, and (13)C correlation spectroscopy at the natural abundance isotope level.

Journal Article↗
PubMed2001

A new modified Smith predictor: the concept, design and tuning.

The paper presents a new modified Smith predictor (MSP) for processes with a long time delay. The MSP appears as an extension of the double controller-scheme (DCS) proposed by Tian and Gao. The important feature of the MSP is that the trade-off between disturbance rejection and robustness to variations in process parameters can be adjusted by means of a single free parameter. The main contribution of the paper concerns tuning of the MSP, which relies on a combination of magnitude optimum criterion with process parameterisation based on multiple integrals of the open-loop step response. In a simulation study the performance of the MSP is compared with that of two known controllers for time delay systems, i.e. DCS of Tian and Gao and Hägglund's predictive PI controller. The results show the advantage of the MSP compared to the two other controllers.

Journal Article↗
PubMed2007

Trans protein splicing of cyanobacterial split inteins in endogenous and exogenous combinations.

Inteins are autocatalytic protein domains that post-translationally excise from protein precursors and ligate their flanking regions with a peptide bond, in a process called protein splicing. Intein-containing DNA polymerases of cyanobacteria and nanoarchaea are naturally split into two separate genes at their intein domain. Such naturally occurring split inteins rapidly self-associate and reconstitute protein-splicing activity in trans. Here, we analyze the in vitro protein-splicing activity of three naturally split inteins from diverse cyanobacteria: Oscillatoria limnetica, Thermosynechococcus vulcanus, and Nostoc sp. PCC7120. N- and C-terminal halves of these split inteins were mixed in nine combinations, resulting in three endogenous (wild-type) and six exogenous combinations. Protein splicing was detected in all split-intein combinations, despite a 30-50% sequence variation between the homologous proteins. Splicing activity proceeded under a variety of conditions, including the presence of denaturants and reductants and high temperature, ionic strength, and viscosity. Still, in a high concentration of salt (2 M) or urea (6 M), specific combinations spliced significantly better than others. Additionally, copper ions were found to inhibit trans splicing in a reversible double-lock reaction. Our comparative analysis of naturally split inteins in endogenous and exogenous combinations demonstrates the modularity of trans protein-splicing elements and their robust activity. It suggests tight interactions between split-intein halves and conditions for modifying the specificity of intein parts. These results promote the biotechnological use of split inteins for controlled assembly of protein fragments either in vivo or in vitro and under moderate or extreme conditions.

Amino Acid Sequence↗
PubMed1994

Parathyroid hormone-related protein in the cardiovascular system.

PTH-related protein (PTHrP) is expressed in a stretch-responsive manner in several types of smooth muscle. We previously demonstrated the production of PTHrP in adult rat heart muscle. In this study, we demonstrate the production of PTHrP in the cardiovascular systems of several mammalian species, including human. We demonstrate PTHrP by immunohistology, using a panel of murine monoclonal antibodies to PTHrP epitopes that span the entire length of the human PTHrP amino acid sequence, quantitate the concentration of PTHrP in the rat cardiovascular system by region-specific RIAs, and measure the relative levels of PTHrP messenger RNA (mRNA) by competitive polymerase chain reaction. Immunohistology studies demonstrated the presence of PTHrP in the cardiovascular systems of humans, rats, pigs, and rabbits. The most robust expression was found in atria, followed by the large vessels, then ventricles. No difference was seen between the left and right sides of the heart. Double staining procedures revealed that PTHrP and atrial naturietic peptide were coexpressed in some cells. Using RIAs and polymerase chain reaction, we demonstrated that atria contained a higher concentration of PTHrP than ventricles and that the relative PTHrP concentrations correlated to its mRNA concentrations in these two tissues. The concentrations of PTHrP in the smooth muscle surrounding the aorta and vena cava were comparable to those in atria. However, in these large vessels, the higher PTHrP levels did not correspond to its mRNA levels. Whereas the immunoreactive concentrations of PTHrP were similar in the atria, aorta, and vena cava, the mRNA levels in the aorta and vena cava were 3-fold lower than those in the atria. Certain PTHrP epitopes appeared to be differentially expressed in specific cardiovascular tissues. A comparison of region-specific assays showed that immunoreactivity measured by immunoassays to PTHrP-(38-64) and PTHrP-(109-141) were 3- to 5-fold greater than that determined by an immunoassay to PTHrP-(1-34). Our observations demonstrate that the atria, aorta, and vena cava contain the greatest amounts of PTHrP in the cardiovascular system. The discrepancy between the concentrations of PTHrP and its mRNA present in the aorta and vena cava suggest that the two may be regulated differently in these tissues. The widespread distribution of PTHrP suggests an important function for the protein in the cardiovascular system, possibly functioning as the calcium counterpart for the atrial natriuretic-sodium regulatory axis.

Animals↗
PubMed1989

Testes are asymmetric in the testicular hemicastration response of the male rat.

An enquiry was made into variability in the testicular hemicastration response of the mature rat (doubling of testicular vein testosterone concentration from the remaining testis 24 h after hemicastration). The response was shown to be asymmetric, being more reliable and more robust when the left testis was removed first. Apparently the testicular hemicastration response, which has been shown to be neurally controlled, shares the asymmetry which has been reported recently in the neural regulation of unilateral ovariectomy.

Animals↗
PubMedDate not supplied

Systematic review of systematic reviews of acupuncture published 1996-2005.

Systematic reviews of acupuncture have tended to support its use, but few applied rigorous inclusion criteria. We tested the credibility of conclusions of systematic reviews of acupuncture published since 1996 by applying rigorous inclusion criteria. Reinterpretation used randomised and double blind trials with valid outcomes or design, and with information available from at least four trials or from 200 patients. Qualified support for acupuncture was originally reported in 12 out of 35 systematic reviews, and strong support was found in another six. Applying stricter inclusion criteria, however, showed that none of the 35 reviews supported acupuncture, predominantly because there were too few patients in the randomised, double blind studies. Six reviews with more than 200 patients in randomised, double blind studies had good evidence of no benefit. Systematic reviews of acupuncture have overstated effectiveness by including studies likely to be biased. They provide no robust evidence that acupuncture works for any indication.

Acupuncture Therapy↗
PubMed2000

Wall Street comes to Washington: market watchers evaluate the health care system.

Purchasers are tolerating double-digit premium increases for health plans and agreeing to employee demands for open-access products and wide choice of doctors largely because of the robust economy and tight labor markets, according to a panel of analysts at the fifth annual Wall Street roundtable convened by the Center for Studying Health System Change (HSC). The analysts, who specialize in health care companies, also discussed the challenges facing managed care and how health plans are evolving. This Issue Brief reports on these and other trends, including employers considering defined contribution approaches, techniques to control pharmaceutical spending and the impact of the Internet on the health care system.

Delivery of Health Care↗
PubMed1998

Dose-dependent cortisol-induced increases in plasma leptin concentration in healthy humans.

BACKGROUND: Leptin is a hormone that regulates fat metabolism and appetite. The secretion of leptin is regulated by adiposity and, in the rodent, by factors such as insulin, beta-adrenergic agonists, and glucocorticoids (GCs). Increased secretion of the endogenous human GC, cortisol, occurs during stress and in disorders such as major depression. Pharmacological GCs can robustly increase plasma leptin concentrations in humans, leading us to hypothesize that cortisol may serve as a physiological regulator of human leptin secretion. METHODS: A randomized double-blind placebo-controlled comparison of 2 fixed oral dosages of cortisol (40 mg/d and 160 mg/d), given for 4 days to matched groups of healthy subjects (n=47). Low-dose treatment approximated GC output during mild stress, while high-dose treatment approximated GC output during maximal stress, spanning a range of GC secretion relevant to physiological stress. RESULTS: Cortisol produced dose-dependent and time-dependent increases in plasma leptin concentrations (time x treatment condition x body mass index; F6,123=10.73; P<.001). Initial treatment-induced increases in plasma leptin concentration returned toward baseline values during 4 treatment days, suggesting tolerance to this GC effect in these healthy subjects. CONCLUSIONS: The results indicate an important role for GCs in the short-term regulation of human leptin secretion. Glucocorticoid-induced increases in leptin secretion suggest a mechanism that may contribute to anorexia and weight loss during stress and disease states such as major depression, if these conditions are associated with sustained increases in plasma leptin concentrations.

Adipose Tissue↗
PubMed1995

Femtosecond molecular dynamics of tautomerization in model base pairs.

Hydrogen bonds commonly lend robustness and directionality to molecular recognition processes and supramolecular structures. In particular, the two or three hydrogen bonds in Watson-Crick base pairs bind the double-stranded DNA helix and determine the complementarity of the pairing. Watson and Crick pointed out, however, that the possible tautomers of base pairs, in which hydrogen atoms become attached to the donor atom of the hydrogen bond, might disturb the genetic code, as the tautomer is capable of pairing with different partners. But the dynamics of hydrogen bonds in general, and of this tautomerization process in particular, are not well understood. Here we report observations of the femtosecond dynamics of tautomerization in model base pairs (7-azaindole dimers) containing two hydrogen bonds. Because of the femtosecond resolution of proton motions, we are able to examine the cooperativity of formation of the tautomer (in which the protons on each base are shifted sequentially to the other base), and to determine the characteristic timescales of the motions in a solvent-free environment. We find that the first step occurs on a timescale of a few hundred femtoseconds, whereas the second step, to form the full tautomer, is much slower, taking place within several picoseconds; the timescales are changed significantly by replacing hydrogen with deuterium. These results establish the molecular basis of the dynamics and the role of quantum tunnelling.

Base Composition↗
PubMed2003

Rad54p is a chromatin remodeling enzyme required for heteroduplex DNA joint formation with chromatin.

In eukaryotic cells, the repair of DNA double-strand breaks by homologous recombination requires a RecA-like recombinase, Rad51p, and a Swi2p/Snf2p-like ATPase, Rad54p. Here we find that yeast Rad51p and Rad54p support robust homologous pairing between single-stranded DNA and a chromatin donor. In contrast, bacterial RecA is incapable of catalyzing homologous pairing with a chromatin donor. We also show that Rad54p possesses many of the biochemical properties of bona fide ATP-dependent chromatin-remodeling enzymes, such as ySWI/SNF. Rad54p can enhance the accessibility of DNA within nucleosomal arrays, but it does not seem to disrupt nucleosome positioning. Taken together, our results indicate that Rad54p is a chromatin-remodeling enzyme that promotes homologous DNA pairing events within the context of chromatin.

Adenosine Triphosphatases↗