bluuebunny/crossref_metadata_2025 huggingface dataset:
Localization of FMRP‐associated mRNA granules and requirement of microtubules for activity‐dependent trafficking in hippocampal neurons
Authors: Antar L. N. , Dictenberg J. B. , Plociniak M. , Afroz R. , Bassell G. J.
Published: August 2005

Fragile X syndrome is caused by the absence of the fragile X mental‐retardation protein (FMRP), an mRNA‐binding protein, which may play important roles in the regulation of dendritic mRNA localization and/or synaptic protein synthesis. We have recently applied high‐resolution fluorescence imaging methods to document the presence, motility and activity‐dependent regulation of FMRP granule trafficking in dendrites and spines of cultured hippocampal neurons. In this study, we show that FMRP granules distribute to F‐actin‐rich compartments, including filopodia, spines and growth cones during the staged development of hippocampal neurons in culture. Fragile X mental‐retardation protein granules were shown to colocalize with ribosomes, ribosomal RNA and MAP1B mRNA, a known FMRP target, which encodes a protein important for microtubule and actin stabilization. The levels of FMRP within dendrites were reduced by disruption of microtubule dynamics, but not by disruption of F‐actin. Direct measurements of FMRP transport kinetics using fluorescence recovery after photobleaching in living neurons showed that microtubules were required to induce the mGluR‐dependent translocation into dendrites. This study provides further characterization of the composition and regulated trafficking of FMRP granules in dendrites of hippocampal neurons.
Differential translation and fragile X syndrome
Authors: Vanderklish P. W. , Edelman G. M.
Published: August 2005

Fragile X syndrome (FXS) is caused by the transcriptional silencing of the Fmr1 gene, which encodes a protein (FMRP) that can act as a translational suppressor in dendrites, and is characterized by a preponderance of abnormally long, thin and tortuous dendritic spines. According to a current theory of FXS, the loss of FMRP expression leads to an exaggeration of translation responses linked to group I metabotropic glutamate receptors. Such responses are involved in the consolidation of a form of long‐term depression that is enhanced in Fmr1 knockout mice and in the elongation of dendritic spines, resembling synaptic phenotypes over‐represented in fragile X brain. These observations place fragile X research at the heart of a long‐standing issue in neuroscience. The consolidation of memory, and several distinct forms of synaptic plasticity considered to be substrates of memory, requires mRNA translation and is associated with changes in spine morphology. A recent convergence of research on FXS and on the involvement of translation in various forms of synaptic plasticity has been very informative on this issue and on mechanisms underlying FXS. Evidence suggests a general relationship in which the receptors that induce distinct forms of efficacy change differentially regulate translation to produce unique spine shapes involved in their consolidation. We discuss several potential mechanisms for differential translation and the notion that FXS represents an exaggeration of one ‘channel’ in a set of translation‐dependent consolidation responses.
Therapeutic implications of the mGluR theory of fragile X mental retardation
Authors: Bear M. F.
Published: August 2005

Evidence is reviewed that the consequences of group 1 metabotropic glutamate receptor (Gp1 mGluR) activation are exaggerated in the absence of the fragile X mental retardation protein, likely reflecting altered dendritic protein synthesis. Abnormal mGluR signaling could be responsible for remarkably diverse psychiatric and neurological symptoms in fragile X syndrome, including delayed cognitive development, seizures, anxiety, movement disorders and obesity.
FMRP RNA targets: identification and validation
Authors: Darnell J. C. , Mostovetsky O. , Darnell R. B.
Published: August 2005

The Fragile X Syndrome is caused by the loss of function of the FMR1 gene (Pieretti et al. 1991. Cell 66, 817–822; O'Donnell & Warren 2002. Annu Rev Neurosci 25, 315–338]. Identification of the RNA targets to which FMRP binds is a key step in understanding the function of the protein and the cellular defects caused by its absence (Darnell et al. 2004 Ment Retard Dev Disabil Res Rev 10, 49–52). Here we discuss the current understanding of FMRP as an RNA‐binding protein, the different approaches that have been taken to identify FMRP RNA targets and the relevance of some of these approaches to FMRP biology. In addition, we present evidence that point mutations in the K‐homology (KH)1 or KH2 domains of FMRP abrogate its polyribosome association in transfected neuroblastoma cells but that the deletion of the RGG box does not. This suggests that RNA binding by the RGG box of FMRP may mediate other aspects of cellular mRNA metabolism such as mRNA localization or that it may have a role downstream of polyribosome association.
Biochemical network models simplified by balanced truncation
Authors: Liebermeister Wolfram , Baur Ulrike , Klipp Edda
Published: August 2005

Modelling of biochemical systems usually focuses on certain pathways, while the concentrations of so‐called external metabolites are considered fixed. This approximation ignores feedback loops mediated by the environment, that is, via external metabolites and reactions. To achieve a more realistic, dynamic description that is still numerically efficient, we propose a new methodology: the basic idea is to describe the environment by a linear effective model of adjustable dimensionality. In particular, we (a) split the entire model into a subsystem and its environment, (b) linearize the environment model around a steady state, and (c) reduce its dimensionality by balanced truncation, an established method for large‐scale model reduction. The reduced variables describe the dynamic modes in the environment that dominate its interaction with the subsystem. We compute metabolic response coefficients that account for complexity‐reduced dynamics of the environment. Our simulations show that a dynamic environment model can improve the simulation results considerably, even if the environment model has been drastically reduced and if its kinetic parameters are only approximately known. The speed‐up in computation gained by model reduction may become vital for parameter estimation in large cell models.
Molecular dynamics structures of peptide nucleic acid·DNA hybrid in the wild‐type and mutated alleles of Ki‐ras proto‐oncogene
Authors: Rathinavelan Thenmalarchelvi , Yathindra Narayanarao
Published: August 2005

The low affinity of peptide nucleic acid (PNA) to hybridize with DNA in the presence of a mismatch endows PNA with a high degree of discriminatory capacity that has been exploited in therapeutics for the selective inhibition of the expression of point‐mutated genes. To obtain a structural basis for this intriguing property, molecular dynamics simulations are carried out on PNA·DNA duplexes formed at the Ki‐ras proto‐oncogene, comprising the point‐mutated (GAT), and the corresponding wild‐type (GGT) codon 12. The designed PNA forms an A…C mismatch with the wild‐type sequence and a perfect A…T pair with the point mutated sequence. Results show that large movements in the pyrimidine base of the A…C mismatch cause loss of stacking, especially with its penultimate base, concomitant with a variable mismatch hydrogen bond, including its occasional absence. These, in turn, bring about dynamic water interactions in the vicinity of the mismatch. Enthalpy loss and the disproportionate entropy gain associated with these are implicated as the factors contributing to the increase in free energy and diminished stability of PNA·DNA duplex with the A…C mismatch. Absence of these in the isosequential DNA duplex, notwithstanding the A…C mismatch, is attributed to the differences in topology of PNA·DNA vis‐à‐vis DNA duplexes. It is speculated that similar effects might be responsible for the reduced stability observed in PNA·DNA duplexes containing other base pair mismatches, and also in mismatch containing PNA·RNA duplexes.
The cytochrome P450scc system opens an alternate pathway of vitamin D3 metabolism
Authors: Slominski Andrzej , Semak Igor , Zjawiony Jordan , Wortsman Jacobo , Li Wei , Szczesniewski Andre , Tuckey Robert C.
Published: August 2005

We show that cytochrome P450scc (CYP11A1) in either a reconstituted system or in isolated adrenal mitochondria can metabolize vitamin D3. The major products of the reaction with reconstituted enzyme were 20‐hydroxycholecalciferol and 20,22‐dihydroxycholecalciferol, with yields of 16 and 4%, respectively, of the original vitamin D3 substrate. Trihydroxycholecalciferol was a minor product, likely arising from further metabolism of dihydroxycholecalciferol. Based on NMR analysis and known properties of P450scc we propose that hydroxylation of vitamin D3 by P450scc occurs sequentially and stereospecifically with initial formation of 20(S)‐hydroxyvitamin D3. P450scc did not metabolize 25‐hydroxyvitamin D3, indicating that modification of C25 protected it against P450scc action. Adrenal mitochondria also metabolized vitamin D3 yielding 10 hydroxyderivatives, with UV spectra typical of vitamin D triene chromophores. Aminogluthimide inhibition showed that the three major metabolites, but not the others, resulted from P450scc action. It therefore appears that non‐P450scc enzymes present in the adrenal cortex to some extent contribute to metabolism of vitamin D3. We conclude that purified P450scc in a reconstituted system or P450scc in adrenal mitochondria can add one hydroxyl group to vitamin D3 with subsequent hydroxylation being observed for reconstituted enzyme but not for adrenal mitochondria. Additional vitamin D3 metabolites arise from the action of other enzymes in adrenal mitochondria. These findings appear to define novel metabolic pathways involving vitamin D3 that remain to be characterized.
The human cytomegalovirus‐encoded chemokine receptor US28 induces caspase‐dependent apoptosis
Authors: Pleskoff Olivier , Casarosa Paola , Verneuil Laurence , Ainoun Fadela , Beisser Patrick , Smit Martine , Leurs Rob , Schneider Pascal , Michelson Susan , Ameisen Jean Claude
Published: August 2005

Viral subversion of apoptosis regulation plays an important role in the outcome of host/virus interactions. Although human cytomegalovirus (HCMV) encodes several immediate early (IE) antiapoptotic proteins (IE1, IE2, vMIA and vICA), no proapoptotic HCMV protein has yet been identified. Here we show that US28, a functional IE HCMV‐encoded chemokine receptor, which may be involved in both viral dissemination and immune evasion, constitutively induces apoptosis in several cell types. In contrast, none of nine human cellular chemokine receptors, belonging to three different subfamilies, induced any significant level of apoptosis. US28‐induced cell death involves caspase 10 and caspase 8 activation, but does not depend on the engagement of cell‐surface death receptors of the tumour necrosis factor receptor/CD95 family. US28 cell‐death induction is prevented by coexpression of C‐FLIP, a protein that inhibits Fas‐associated death domain protein (FADD)‐mediated activation of caspase 10 and caspase 8, and by coexpression of the HCMV antiapoptotic protein IE1. The use of US28 mutants indicated that the DRY sequence of its third transmenbrane domain, required for constitutive G‐protein signalling, and the US28 intracellular terminal domain required for constitutive US28 endocytosis, are each partially required for cell‐death induction. Thus, in HCMV‐infected cells, US28 may function either as a chemokine receptor, a phospholipase C activator, or a proapoptotic factor, depending on expression levels of HCMV and/or cellular antiapoptotic proteins.
Novel brain 14‐3‐3 interacting proteins involved in neurodegenerative disease
Authors: Mackie Shaun , Aitken Alastair
Published: August 2005

We isolated two novel 14‐3‐3 binding proteins using 14‐3‐3 ζ as bait in a yeast two‐hybrid screen of a human brain cDNA library. One of these encoded the C‐terminus of a neural specific armadillo‐repeat protein, δ‐catenin (neural plakophilin‐related arm‐repeat protein or neurojungin). δ‐Catenin from brain lysates was retained on a 14‐3‐3 affinity column. Mutation of serine 1072 in the human protein and serine 1094 in the equivalent site in the mouse homologue (in a consensus binding motif for 14‐3‐3) abolished 14‐3‐3 binding to δ‐catenin in vitro and in transfected cells. δ‐catenin binds to presenilin‐1, encoded by the gene most commonly mutated in familial Alzheimer's disease. The other clone was identified as the insulin receptor tyrosine kinase substrate protein of 53 kDa (IRSp53). Human IRSp53 interacts with the gene product implicated in dentatorubral‐pallidoluysian atrophy, an autosomal recessive disorder associated with glutamine repeat expansion of atrophin‐1.
The A‐subunit of surface‐bound Shiga toxin stimulates clathrin‐dependent uptake of the toxin
Authors: Torgersen Maria L. , Lauvrak Silje U. , Sandvig Kirsten
Published: August 2005

Shiga toxin can be internalized by clathrin‐dependent endocytosis in different cell lines, although it binds specifically to the glycosphingolipid Gb3. It has been demonstrated previously that the toxin can induce recruitment of the toxin–receptor complex to clathrin‐coated pits, but whether this process is concentration‐dependent or which part of the toxin molecule is involved in this process, have so far been unresolved issues. In this article, we show that the rate of Shiga toxin uptake is dependent on the toxin concentration in several cell lines [HEp‐2, HeLa, Vero and baby hamster kidney (BHK)], and that the increased rate observed at higher concentrations is strictly dependent on the presence of the A‐subunit of cell surface‐bound toxin. Surface‐bound B‐subunit has no stimulatory effect. Furthermore, this increase in toxin endocytosis is dependent on functional clathrin, as it did not occur in BHK cells after induction of antisense to clathrin heavy chain, thereby blocking clathrin‐dependent endocytosis. By immunofluorescence, we show that there is an increased colocalization between Alexa‐labeled Shiga toxin and Cy5‐labeled transferrin in HeLa cells upon addition of unlabeled toxin. In conclusion, the data indicate that the Shiga toxin A‐subunit of cell surface‐bound toxin stimulates clathrin‐dependent uptake of the toxin. Possible explanations for this phenomenon are discussed.
Properties and significance of apoFNR as a second form of air‐inactivated [4Fe‐4S]·FNR of Escherichia coli
Authors: Achebach Stephanie , Selmer Thorsten , Unden Gottfried
Published: August 2005

The active form of the oxygen sensor fumarate nitrate reductase regulator (FNR) of Escherichia coli contains a [4Fe‐4S] cluster which is converted to a [2Fe‐2S] cluster after reaction with air, resulting in inactivation of FNR. Reaction of reconstituted [4Fe‐4S]·FNR with air resulted within 5 min in conversion to apoFNR. The rate was comparable to the rate known for [4Fe‐4S]·FNR/[2Fe‐2S]·FNR cluster conversion, suggesting that apoFNR is a product of [2Fe‐2S]·FNR decomposition and a final form of air‐inactivated FNR in vitro. Formation of apoFNR and the redox state of the cysteinyl residues were determined in vitro by alkylation. FNR contains five cysteinyl residues, four of which (Cys20, Cys23, Cys29 and Cys122) ligate the FeS clusters. Alkylated FNR and proteolytic fragments thereof were analyzed by MALDI‐TOF. ApoFNR formed by air inactivation of [4Fe‐4S]·FNR in vitro contained one or two disulfides. Only disulfide pairs Cys16/20 and Cys23/29 were formed; Cys122 was never part of a disulfide. The same type of disulfide was found in apoFNR obtained during isolation of FNR, suggesting that cysteine disulfide formation follows a fixed pattern. ApoFNR, including the form with two disulfides, can be reconstituted to [4Fe‐4S]·FNR after disulfide reduction. The experiments suggest that apoFNR is a major form of FNR under oxic conditions.
Solution structure of human proinsulin C‐peptide
Authors: Munte Claudia Elisabeth , Vilela Luciano , Kalbitzer Hans Robert , Garratt Richard Charles
Published: August 2005

The C‐peptide of proinsulin is important for the biosynthesis of insulin, but has been considered for a long time to be biologically inert. Recent studies in diabetic patients have stimulated a new debate about its possible regulatory role, suggesting that it is a hormonally active peptide. We describe structural studies of the C‐peptide using 2D NMR spectroscopy. In aqueous solution, the NOE patterns and chemical shifts indicate that the ensemble is a nonrandom structure and contains substructures with defined local conformations. These are more clearly visible in 50% H2O/50% 2,2,2‐trifluoroethanol. The N‐terminal region (residues 2–5) forms a type I β‐turn, whereas the C‐terminal region (residues 27–31) presents the most well‐defined structure of the whole molecule including a type III′β‐turn. The C‐terminal pentapeptide (EGSLQ) has been suggested to be responsible for chiral interactions with an as yet uncharacterized, probably a G‐protein‐coupled, receptor. The three central regions of the molecule (residues 9–12, 15–18 and 22–25) show tendencies to form β‐bends. We propose that the structure described here for the C‐terminal pentapeptide is consistent with the previously postulated CA knuckle, believed to represent the active site of the C‐peptide of human proinsulin.
Interacciones Planta‐Nodriza en Lophophora diffusa (Cactaceae) en un Desierto Subtropical de México1
Authors: Zúñiga Bertha , Malda Guadalupe , Suzán Humberto
Published: August 2005

One of the two species from the genus Lophophora is the false peyote Lophophora diffusa, an endemic cactus species of the xerophytic shrubland at Querétaro, México, considered threatened from illegal extraction due to its hallucinogenic and medicinal properties. We analyzed the spatial distribution of L. diffusa and its association with the locally dominant shrub species with the system SADIE (Spatial Analysis by Distance IndicEs). We also studied the principal microclimatic factors (light, temperature, and humidity) beneath the canopy of the possible nurse plant species. The use of SADIE is a new approach to study the mechanisms of spatial distribution. Lophophora diffusa and the shrub species presented an aggregated distribution with patches and gaps, as indicated by the values of Ia= 4.179 for L. diffusa and Ia= 1.660 for the vegetation. Lophophora diffusa was positively associated with the arboreal vegetation, particularly with Larrea tridentata and Acacia sororia, but was negatively associated with Celtis pallida and Myrtillocactus geometrizans. Microclimate evaluation indicated that C. pallida canopy significantly reduced radiation and temperature compared to the other species (L. tridentata, A.sororia y P. laevigata). We had expected L. diffusa to exhibit a positive spatial association with C. pallida; however, the reduction in light availability apparently limited seedling establishment of L. diffusa.
Microbial Biomass, Respiration, and Decomposition of Hura crepitans L. (Euphorbiaceae) Leaves in a Tropical Stream1
Authors: Abelho Manuela , Cressa Claudia , Graça Manuel A. S.
Published: August 2005

The processing of leaves in temperate streams has been the subject of numerous studies but equivalent tropical ecosystems have received little attention. We investigated leaf breakdown of a tropical tree species (Hura crepitans, Euphorbiaceae), in a tropical stream using leaf bags (0.5 mm mesh) over a period of 24 days. We followed the loss of mass and the changes in adenosine triphosphate (ATP) concentrations and respiration rates associated with the decomposing leaves. The breakdown rate was fast (k=−0.0672/d, kd=−0.0031/degree‐day), with 81 percent loss of the initial mass within 24 days. This high rate was probably related to the stable and high water temperature (22°C) favoring strong biological activity. Respiration rates increased until day 16 (1.1 mg O2/h/g AFDM), but maximum ATP concentrations were attained at day 9 (725 nmol ATP/g AFDM) when leaf mass remaining was 52 percent. To determine the relative importance of fungi and bacteria during leaf decomposition, ATP concentrations, and respiration rates were determined in samples treated with antibiotics, after incubation in the stream. The results of the samples treated with the antifungal or the bacterial antibiotic suggest a higher contribution of the fungal community for total microbial biomass and a higher contribution of the bacterial community for microbial respiration rates, especially during the later stages of leaf decomposition. However, these results should be analyzed with caution since both antibacterial and antifungal agents did not totally eliminate microbial activity and biomass.
Foliage Density of Branches and Distribution of Plant‐Dwelling Spiders1
Authors: De Souza Andréa Lúcia Teixeira , Martins Rogério Parentoni
Published: August 2005

We compared the abundance of foliage‐living spiders among seven widespread plant species comprising a gradient of levels of structural complexity in a tropical savannah‐like region in southeastern Brazil. Spider abundance among plant species was positively related to the foliage density of their branches. A field experiment using artificial branches was carried out to isolate foliage density effects on spider abundance, thus controlling both biomass and texture effects. Artificial branches were attached to branches of three plant species with similar foliage density, Baccharis dracunculifolia, Diplusodon virgatus, and Microlicia fasciculata. Two treatments were set up: artificial branches with higher foliage density attracted more spiders than those with lower foliage density. The guild structure of hunting spiders was compared among vegetative branches of three plant species with different levels of foliage density: B. dracunculifolia, D. virgatus, and Bidens gardneri. Stalker, and ambusher spiders were more abundant on branches of B. dracunculifolia, which had the highest foliage density. Foliage‐runners constituted the dominant guild on D. virgatus and B. gardneri, which have lower foliage density branches. Our results suggest that branch architecture is the most important factor determining the abundance of plant‐dwelling spiders in the study area independently of branch biomass, leaf surface area or texture.
The Influence of Ants on the Foraging Behavior of Birds in an Agroforest1
Authors: Philpott Stacy M. , Greenberg Russell , Bichier Peter
Published: August 2005

Ants limit bird foraging success via interference or exploitative competition. We compared bird foraging (number and duration of visits, bird species visiting) on ant (Azteca instabilis)‐infested and ant‐free tropical trees (Inga michelianaandAlchornea latifolia). Ants did not affect the number of bird visits or the number of species visiting. Ant presence shortened visit duration (overall and for insectivores) only onA. latifoliawhere ant activity was higher. Ants may thus hinder bird foraging on some tropical trees potentially shaping how predators affect arthropod communities; yet ant effects depend on bird foraging guild and ant activity.
COSMIC HOLOGRAPHIC BOUNDS WITH UV AND IR CUTOFFS
Authors: MYUNG YUN SOO
Published: August 2005

We introduce the cosmic holographic bounds with two uv and ir cutoff scales, to deal with both the inflationary universe in the past and dark energy in the future. To describe quantum fluctuations of inflation on sub-horizon scales, we use the Bekenstein–Hawking energy bound. The Hubble bounds are introduced for classical fluctuations of inflation on super-horizon scales. It turns out that the Hubble entropy bound is satisfied with the entanglement entropy and the Hubble temperature bound leads to a condition for the slow-roll inflation. In order to describe the dark energy, we introduce the holographic energy density which is the one saturating the Bekenstein–Hawking energy bound for a weakly gravitating system. Here the uv (ir) cutoff is given by the Planck scale (future event horizon), respectively. As a result, we find the close connection between inflation and dark energy.
ON THE DIFFEOMORPHISMS FOR AKBULUT'S KNOT CONCORDANCE SURGERY
Authors: TANGE MOTOO
Published: August 2005

Akbulut generalized Fintushel–Stern knot surgery in a certain way. He showed that an operation C(K#(-K)) → { diffeo type of X} is not injective, where C(K#(-K)) is self-concordance set of K#(-K) and K is trefoil knot. In the present paper we prove that the same phenomenon occurs for any knot K and for some other knots L instead of K#(-K).
Constitutive eNOS-derived nitric oxide is a determinant of endothelial junctional integrity
Authors: Predescu Dan , Predescu Sanda , Shimizu Jun , Miyawaki-Shimizu Kayo , Malik Asrar B.
Published: August 2005

Basal vascular endothelial permeability is normally kept low in part by the restrictiveness of interendothelial junctions (IEJs). We investigated the possible role of nitric oxide (NO) in controlling IEJ integrity and thereby regulating basal vascular permeability. We determined the permeability of continuous endothelia in multiple murine vascular beds, including lung vasculature, of wild-type mice, endothelial nitric oxide synthase (eNOS) null mice, and mice treated with NOS inhibitor N-nitro-l-arginine methyl ester (l-NAME). Light and electron microscopic studies revealed that l-NAME treatment resulted in IEJs opening within a few minutes with a widespread response within 30 min. We observed a 35% increase in transendothelial transport of albumin, using as tracer dinitrophenylated albumin in mouse lungs and other organs studied. To rule out the involvement of blood cells in the mechanism of increased endothelial permeability, vascular beds were flushed free of blood, treated with l-NAME, and perfused with the tracer. The open IEJs observed in these studies indicated a direct role for NO in preserving the normal structure of endothelial junctions. We also used the electron-opaque tracer lanthanum chloride to assess vascular permeability. Lanthanum chloride was presented by perfusion to various vascular beds of mice lacking NO. Open IEJs were seen only in capillary and venular endothelial segments of mice lacking NO, and there was a concomitant increase in vascular permeability to the tracer. Together, these data demonstrate that constitutive eNOS-derived NO is a crucial determinant of IEJ integrity and thus serves to maintain the low basal permeability of continuous endothelia.
More than just a barrier: urothelium as a drug target for urinary bladder pain
Authors: Birder Lori A.
Published: August 2005

Although the urinary bladder urothelium has classically been thought of as a passive barrier to ions/solutes, a number of novel properties have been recently attributed to these cells. Studies have revealed that the urothelium is involved in sensory mechanisms (i.e., ability to express a number of sensor molecules or respond to thermal, mechanical, and chemical stimuli) and can release chemical mediators. Localization of afferent nerves next to the urothelium suggests these cells may be targets for transmitters released from bladder nerves or that chemicals released by urothelial cells may alter afferent excitability. Taken together, these and other findings highlighted in this review suggest a sensory function for the urothelium. Elucidation of mechanisms impacting on urothelial function may provide insights into the pathology of bladder dysfunction.
Adenosine Down-Regulates Giant Depolarizing Potentials in the Developing Rat Hippocampus by Exerting a Negative Control on Glutamatergic Inputs
Authors: Safiulina Victoria F. , Kasyanov Alexander M. , Giniatullin Rashid , Cherubini Enrico
Published: August 2005

Adenosine is a widespread neuromodulator that can be directly released in the extracellular space during sustained network activity or can be generated as the breakdown product of adenosine triphosphate (ATP). Whole cell patch-clamp recordings were performed from CA3 principal cells and interneurons in hippocampal slices obtained from P2–P7 neonatal rats to study the modulatory effects of adenosine on giant depolarizing potentials (GDPs) that constitute the hallmark of developmental networks. We found that GDPs were extremely sensitive to the inhibitory action of adenosine (IC50 = 0.52 μM). Adenosine also contributed to the depressant effect of ATP as indicated by DPCPX-sensitive changes of ATP-induced reduction of GDP frequency. Similarly, adenosine exerted a strong inhibitory action on spontaneous glutamatergic synaptic events recorded from GABAergic interneurons and on interictal bursts that developed in CA3 principal cells after blockade of γ-aminobutyric acid type A (GABAA) receptors with bicuculline. All these effects were prevented by DPCPX, indicating the involvement of inhibitory A1 receptors. In contrast, GABAergic synaptic events were not changed by adenosine. Consistent with the endogenous role of adenosine on network activity, DPCPX per se increased the frequency of GDPs, interictal bursts, and spontaneous glutamatergic synaptic events recorded from GABAergic interneurons. Moreover, the adenosine transport inhibitor NBTI and the adenosine deaminase blocker EHNA decreased the frequency of GDPs, thus providing further evidence that endogenous adenosine exerts a powerful control on GDP generation. We conclude that, in the neonatal rat hippocampus, the inhibitory action of adenosine on GDPs arises from the negative control of glutamatergic, but not GABAergic, inputs.
Molecular cytogenetic insights into the ageing syndrome Hutchinson-Gilford Progeria (HGPS)
Authors: Corso C. , Parry E.M. , Faragher R.G.A. , Seager A. , Green M.H.L. , Parry J.M.
Published: August 2005

Hutchinson-Gilford Progeria Syndrome (HGPS) is an extremely rare genetic disorder characterized by premature ageing in childhood and serves as a valuable model for the human ageing process in general. Most recently, point mutations in the lamin A (LMNA) gene on chromosome 1q have been associated with the disease, however how these mutations relate to the complex phenotype of HGPS remains to be established. It has been shown that fibroblasts from HGPS patients are frequently resistant to immortalization with telomerase (hTERT), consistent with the idea that the loss of a dominant acting HGPS gene is a pre-requisite for immortalization. In this study we report the first detailed cytogenetic analysis of hTERT-immortalised HGPS cell lines from three patients and one corresponding primary fibroblast culture. Our results provide evidence for a cytogenetic mosaicism in HGPS with a distinctive pattern of chromosome aberrations in all the HGP clones. Chromosome 11 alterations were observed at a high frequency in each immortalised HGPS cell line but were also present at a lower frequency in the corresponding primary cells. Moreover, we were able to identify the 11q13→q23 region as a potential site of breakage. Our results are therefore consistent with a role of chromosome 11 alterations in the escape from senescence observed in HGPS cells. In addition to this defined rearrangement, we consistently observed complex chromosomal rearrangements, suggesting that HGPS displays features of chromosomal instability.
Radiosensitivity detected by the micronucleus test is not generally increased in sporadic prostate cancer patients
Authors: Varga D. , Michel I. , Patino-Garcia B. , Paiss T. , Vogel W. , Maier C.
Published: August 2005

The micronucleus test (MNT) has shown increased micronuclei (MN) frequencies in BRCA associated and sporadic breast cancer patients, Ataxia telangiectasia and Nijmegen Breakage Syndrome patients, demonstrating a common cellular phenotype of increased radiosensitivity. Some genes, causative of these diseases, have also recently been associated with prostate cancer. In order to investigate if prostate cancer exhibits the cellular phenotype of increased radiosensitivity, we performed MNT analysis on 22 sporadic prostate cancer patients and 43 male controls. We determined the baseline MN frequency, in order to see in vivo chromosomal damage without radiation, and induced (after irradiation with 2 Gy) frequency of MN, both in binucleated cells (BNC) obtained from cultured peripheral blood lymphocytes. An automated image analysis system was used to score the MN employing two different classifiers (Classifier A and B) for detection of BNC. The mean baseline frequencies were 48/43 MN/1000 BNC (A/B) for the controls and 42/50 (A/B) for prostate cancer patients. The induced MN frequencies amounted to 107/111 MN/1000 BNC (A/B) for controls and 111/114 MN/1000 BNC (A/B) for prostate cancer patients. The obtained MN frequencies did not result in a statistically significant difference between unselected cases and controls. However, restricting the analysis to young patients (50–60 years, N = 7) and age-matched controls (N = 7) revealed marginally significant higher MN frequencies in patients. We conclude that increased radiosensitivity is not a property of prostate cancer patients in general.
Structure of equine 2′-5′oligoadenylate synthetase (OAS) gene family and FISH mapping of OAS genes to ECA8p15→p14 and BTA17q24→q25
Authors: Perelygin A.A. , Lear T.L. , Zharkikh A.A. , Brinton M.A.
Published: August 2005

Mammalian 2′-5′ oligoadenylate (2-5A) synthetases are important mediators of the antiviral activity of interferons. Both human and mouse 2-5A synthetase gene families encode four forms of enzymes: small, medium, large and ubiquitin-like. In this study, the structures of four equine OAS genes were determined using DNA sequences derived from fifteen cDNA and four BAC clones. Composition of the equine OAS gene family is more similar to that of the human OAS family than the mouse Oas family. Two OAS-containing bovine BAC clones were identified in GenBank. Both equine and bovine BAC clones were physically assigned by FISH to horse and cattle chromosomes, ECA8p15→p14 and BTA17q24→ q25, respectively. The comparative mapping data confirm conservation of synteny between ungulates, humans and rodents.
Differences in the pattern of X-linked gene expression between fetal bovine muscle and fibroblast cultures derived from the same muscle biopsies
Authors: Nino-Soto M.I. , Nuber U.A. , Basrur P.K. , Ropers H.-H. , King W.A.
Published: August 2005

The sex determination system in mammals creates an imbalance between males and females in the number of X chromosomes. This imbalance is compensated through transcriptional silencing of one of the two X chromosomes in female diploid cells by epigenetic modifications. Although common for mammals, X inactivation shows marked species-specific differences in mechanisms and end results, and provides a unique opportunity to study epigenetic regulation of gene expression. The aim of the present study was to establish the expression pattern of selected X-linked genes in bovine fetal muscle tissue and muscle fibroblast cultures in order to follow possible modifications at the transcriptional level attributable to in vitro culture. We used heterologous cDNA microarray hybridization and quantitative real-time PCR to study the pattern of expression of X-linked genes including SLC25A6, GAB3, MECP2, RPS4X, JARID1C, UBE1, BIRC4 and SLC16A2. Quantitative real-time PCR analysis in fetal bovine muscle showed higher transcript levels in females for all X-linked genes tested with the exception of SLC25A6, with differences being significant for RPS4X, JARID1C and UBE1. The expression in fibroblast cultures derived from the same samples differed, with significantly higher levels for UBE1, GAB3 and BIRC4, while the rest of the panel of X-linked genes remained unchanged. The changed expression pattern in vitro, probably reflecting modifications in the epigenetic mechanisms that regulate transcriptional activity and gene silencing in X inactivation, has important implications for the advancement of new biotechnologies such as somatic cell nuclear transfer and stem cell therapy.
ESTs from brain and testis of White Leghorn and red junglefowl: annotation, bioinformatic classification of unknown transcripts and analysis of expression levels
Authors: Savolainen P. , Fitzsimmons C. , Arvestad L. , Andersson L. , Lundeberg J.
Published: August 2005

We report the generation, assembly and annotation of expressed sequence tags (ESTs) from four chicken cDNA libraries, constructed from brain and testis tissue dissected from red junglefowl and White Leghorn. 21,285 5′-end ESTs were generated and assembled into 2,813 contigs and 9,737 singletons, giving 12,549 tentative unique transcripts. The transcripts were annotated using BLAST by matching to known chicken genes or to putative homologues in other species using the major gene/protein databases. The results for these similarity searches are available on www.sbc.su.se/∼arve/chicken. 4,129 (32.9%) of the transcripts remained without a significant match to gene/protein databases, a proportion of unmatched transcripts similar to earlier non-mammalian EST studies. To estimate how many of these transcripts may represent novel genes, they were studied for the presence of coding sequence. It was shown that most of the unique chicken transcripts do not contain coding parts of genes, but it was estimated that at least 400 of the transcripts contain coding sequence, indicating that 3.2% of avian genes belong to previously unknown gene families. Further BLAST search against dbEST left 1,649 (13.1%) of the transcripts unmatched to any library. The number of completely unmatched transcripts containing coding sequence was estimated at 180, giving a measure of the number of putative novel chicken genes identified in this study. 84.3% of the identified transcripts were found only in testis tissue, which has been poorly studied in earlier chicken EST studies. Large differences in expression levels were found between the brain and testis libraries for a large number of transcripts, and among the 525 most frequently represented transcripts, there were at least 20 transcripts with significant difference in expression levels between red junglefowl and White Leghorn.
Antigen-Induced Reduction in Mast Cell and Basophil Functional Responses due to Reduced Syk Protein Levels
Authors: Kepley Christopher L.
Published: August 2005

Background: The high-affinity IgE receptor, FcεRI, is unresponsive on mast cells and basophils from people in several populations through an unknown mechanism. Similarly, FcεRI-positive basophils from ‘nonreleasers’ are IgE-unresponsive and are deficient in the tyrosine kinase Syk. Objective: To test the hypothesis that cross-linking FcεRI on mast cells and basophils leads to FcεRI nonresponsiveness through reduction in Syk protein levels. Methods: Human mast cells and basophils were used to determine if FcεRI hyporesponsiveness correlated with reduced Syk levels. Results: It is shown that suboptimal antigen challenge, that did not lead to significant mediator release, induced nonresponsiveness and correlated with reduced Syk. Other IgE-associated signaling molecules were unaffected by the same treatment. The ability of IgE-unresponsive mast cells to regain FcεRI responsiveness is paralleled by increased cellular Syk levels in vitro. The reduction of Syk levels with suboptimal antigen concentrations was calcium independent and mediated through a proteasome-dependent mechanism. Conclusion: These findings confirm and extend our knowledge about a novel regulatory mechanism for maintaining FcεRI in a quiescent state. This mechanism may also explain why low concentrations of allergen given to patients during allergen immunotherapy induce FcεRI nonresponsiveness and therapeutic benefit without inducing systemic anaphylaxis.
NOS 1 Is Required for Allergen-Induced Expression of NOS 2 in Mice
Authors: Iijima Hiroaki , Tulic Meri K. , Duguet Alexandre , Shan Jichuan , Carbonara Paolo , Hamid Qutayba , Eidelman David H.
Published: August 2005

Background: Although increased nitric oxide (NO) production in asthma is mediated largely by upregulation of the inducible form of nitric oxide synthase (iNOS, or NOS 2), some studies have suggested an important role for the usually constitutive neural NOS isoform (nNOS, or NOS 1). Aim: To investigate how NOS 1 may influence allergic inflammation, we used NOS 1 knockout mice and their wild-type (WT) controls. Methods: Mice were sensitized and challenged with ovalbumin (OVA) using a protocol known to upregulate NOS 2 in the airways. Results: In addition to expected increases in NOS 2 activity, OVA challenge led to increases in calcium-dependent NOS activity, which was accounted for by increased expression of NOS 1 at both mRNA (n = 5, p < 0.001) and protein levels (n = 5, p < 0.01). In NOS-1-deficient mice, OVA challenge induced less eosinophilia (n = 7, p < 0.05) and much less NO production (n = 10, p < 0.01) than in WT controls, reflecting not only the expected absence of NOS 1, but also lack of upregulation of NOS 2. This interaction appeared to be stimulus specific as NOS-1-deficient mice did upregulate NOS 2 following exposure to lipopolysaccharide (n = 5, p < 0.001). Conclusions: These findings underscore the importance of NOS 1 in allergic airway inflammation and suggest a mechanism by which NOS 1 may influence overall NO production in the airways.
Transvaginal Ultrasonography of the Endometrium in Postmenopausal Japanese Women
Authors: Tsuda Hiroshi , Nakamura Hiroaki , Inoue Takeshi , Kawamura Naoki , Adachi Ken-ichi , Bandera Christina A.
Published: August 2005

Aims: To determine the cut-off level of endometrial thickness for detecting endometrial disease on a large scale of screening and to examine the usefulness of transvaginal ultrasonography (TVS) for endometrial disease screening in asymptomatic postmenopausal Japanese women. Methods: The study involved 1,400 postmenopausal women in whom endometrial thickness was measured with TVS, and then compared with the histopathological diagnosis of endometrial specimens. Results: The prevalence of endometrial disease in asymptomatic and symptomatic cases was 2.3 and 21%. In symptomatic cases, at a cut-off limit of 3 mm for endometrial thickness, the sensitivity for detecting endometrial disease was 94%, specificity 70% and positive predictive value 46%. With an endometrial thickness ≤3 mm, the probability that endometrial disease would be overlooked was 0.025.In asymptomatic cases, the corresponding figures at a cut-off limit of 3 mm were 90, 84, 12% and 0.003. Conclusions: TVS does not appear to be an effective screening method in asymptomatic postmenopausal Japanese women because of the low prevalence of endometrial disease in this population. We recommend a normal cut-off level for endometrial thickness of 4 mm in symptomatic postmenopausal Japanese women.
Revised Uvulopalatopharyngoplasty with Uvula Preservation and Its Clinical Study
Authors: Han Demin , Ye Jingying , Lin Zhonghui , Wang Jun , Wang Jiangyong , Zhang Yuhuan
Published: August 2005

Objectives: A revised uvulopalatopharyngoplasty (UPPP) is reported for reducing the surgical complications without compromising the response. Methods and Materials: A new method of performing UPPP was designed, in which the uvula is preserved, but a larger portion of the soft palate is removed. This new procedure is called the ‘revised UPPP with uvula preservation’ (Han’s UPPP or H-UPPP), the characteristics of which are as follows: complete preservation of the uvula; larger portion of the soft palate resected (with the levator palati and tensor palati remaining intact); the adipose tissue in the space of the velum palati is removed; the preserved uvula will become a ‘normal’ one by the contraction of scar tissue on both sides of the musculus palato-uvularis and the lower margin of the tensor palati. 68 cases of obstructive sleep apnea/hypopnea syndrome had been treated with H-UPPP between September 1998 and May 2001. Results: Polysomnography was performed for all 68 cases 6 months postoperatively with a surgical response of 69.12%, defined as a >50% reduction of the apnea-hypopnea index and <20 apneic pauses/h; no palatopharyngeal incompetency or palatopharyngeal stenosis. Anatomic measurements showed that the preserved uvula in H-UPPP begins to retract 2 weeks postoperatively, and the preserved uvula becomes ‘normal’ in 3–6 months. Conclusions: These results suggest that H-UPPP is an effective surgery and produces fewer complications compared with the classic UPPP.
The Impact of an Out-Reach Clinic on Referral of Patients with Renal Impairment
Authors: Craig Kathrine J. , Riley Stephen G. , Thomas Beverley , Penney Michael , Donovan Kieron L. , Phillips Aled O.
Published: August 2005

Background: Early diagnosis and prompt treatment of a number of renal diseases may delay renal failure, obviate the need for renal replacement therapy and reduce co-morbidity. The aim of this study was to examine the impact of out-reach renal clinics on patterns of referral of patients with renal impairment to a nephrologist. Methods: The number of patients with renal impairment was determined as defined by serum creatinine levels >150 µmol/l in three centres within a single NHS trust over two separate 1-week periods. None of the centres studied has a local nephrologist, however one centre (hospital A) has renal out-reach clinics, another is geographically close to a renal unit (hospital B), while the third unit (hospital C) has no nephrology presence and is geographically furthest from the renal unit. In addition, retrospective as well as follow-up data on the renal function of all patients with renal impairment was collected. Results: In hospital A, there was a lower proportion of patients with unreferred renal impairment than in the other two hospitals. Within the unreferred patient group there were significantly more patients whose renal function improved during the follow-up period. A considerable proportion of patients with documented deterioration in renal function remained unknown to nephrology services 6 months after initial presentation. Other than the presence of an onsite nephrology service, the only other factor found to be significantly different in those patients not referred to nephrologists was age: as in all three centres, those not referred were significantly older. Conclusion: Inequity of access to renal services is an important obstacle to early referral of patients with impaired renal function. Out-reach renal services provide a model which significantly improves referral patterns.