bluuebunny/crossref_metadata_2025 huggingface dataset:
Simultaneous assessment of loss of heterozygosity at multiple microsatellite loci using semi-automated fluorescence-based detection: subregional mapping of chromosome 4 in cervical carcinoma.
Authors: Hampton G M , Larson A A , Baergen R N , Sommers R L , Kern S , Cavenee W K
Published: July 2002

Detection of loss of heterozygosity (LOH) by comparison of normal and tumor genotypes using PCR-based microsatellite loci provides considerable advantages over traditional Southern blotting-based approaches. However, current methodologies are limited by several factors, including the numbers of loci that can be evaluated for LOH in a single experiment, the discrimination of true alleles versus "stutter bands," and the use of radionucleotides in detecting PCR products. Here we describe methods for high throughput simultaneous assessment of LOH at multiple loci in human tumors; these methods rely on the detection of amplified microsatellite loci by fluorescence-based DNA sequencing technology. Data generated by this approach are processed by several computer software programs that enable the automated linear quantitation and calculation of allelic ratios, allowing rapid ascertainment of LOH. As a test of this approach, genotypes at a series of loci on chromosome 4 were determined for 58 carcinomas of the uterine cervix. The results underscore the efficacy, sensitivity, and remarkable reproducibility of this approach to LOH detection and provide subchromosomal localization of two regions of chromosome 4 commonly altered in cervical tumors.
Fos and Jun bend the AP-1 site: effects of probe geometry on the detection of protein-induced DNA bending.
Authors: Kerppola T K
Published: July 2002

The effect of Fos and Jun binding on the structure of the AP-1 recognition site is controversial. Results from phasing analysis and phase-sensitive detection studies of DNA bending by Fos and Jun have led to opposite conclusions. The differences between these assays, the length of the spacer between two bends and the length of the sequences flanking the bends, are investigated here using intrinsic DNA bend standards. Both an increase in the spacer length as well as a decrease in the length of flanking sequences resulted in a reduction in the phase-dependent variation in electrophoretic mobilities. Probes with a wide separation between the bends and short flanking sequences, such as those used in the phase-sensitive detection studies, displayed no phase-dependent mobility variation. This shape-dependent variation in electrophoretic mobilities was reproduced by complexes formed by truncated Fos and Jun. Results from ligase-catalyzed cyclization experiments have been interpreted to indicate the absence of DNA bending in the Fos-Jun-AP-1 complex. However, truncated Fos and Jun can alter the relative rates of inter- and intramolecular ligation through mechanisms unrelated to DNA bending, confounding the interpretation of cyclization data. The analogous phase- and shape-dependence of the electrophoretic mobilities of the Fos-Jun-AP-1 complex and an intrinsic DNA bend confirm that Fos and Jun bend DNA, which may contribute to their functions in transcription regulation.
The highly conserved
skb1
gene encodes a protein that interacts with Shk1, a fission yeast Ste20/PAK homolog
Authors: Gilbreth Mary , Yang Peirong , Wang Dan , Frost Jeff , Polverino Anthony , Cobb Melanie H. , Marcus Stevan
Published: July 2002

The Shk1 protein kinase, a homolog of
Saccharomyces
cerevisiae
Ste20 and mammalian
p21
Cdc42/Rac
-activated kinases, is an essential component
of a Ras- and Cdc42-dependent signaling cascade required for cell
viability, normal morphology, and mitogen-activated protein
kinase-mediated sexual responses in the fission yeast,
Schizosaccharomyces pombe
. To identify
S.
pombe
proteins that modulate or mediate Shk1 functions, we
conducted a two-hybrid screen for Shk1-interacting proteins. One of the
genes identified as a result of this screen was
skb1
. We
show that Skb1 interacts with a region of the N-terminal regulatory
domain of Shk1 distinct from that to which Cdc42 binds, and that Shk1,
Cdc42, and Skb1 are able to form a ternary complex
in
vivo
.
S. pombe
cells carrying an
skb1
null mutation are less elongate in morphology than
wild-type cells and exhibit a moderate growth defect. The morphology
defect of the
skb1
deletion mutant is suppressed by
overexpression of Shk1. Overexpression of Skb1 causes wild-type
S. pombe
cells to become hyperelongated. Additional
genetic analyses described herein suggest that Skb1 is a component of
the morphology control branch of the Ras signaling cascade in
S.
pombe
and that it positively modulates Shk1 function. Homologs
of Skb1 are encoded by open reading frames in the genomes of
S.
cerevisiae
and
Caenorhabditis
elegans
and by an uncharacterized human cDNA sequence. Thus,
skb1
may be the first well-characterized member of a
highly conserved family of genes encoding potential
p21
Cdc42/Rac
-activated kinase regulators.
Complementary mutations in an antigenic peptide allow for crossreactivity of autoreactive T-cell clones
Authors: Ausubel Lara J. , Kwan Candice K. , Sette Alessandro , Kuchroo Vijay , Hafler David A.
Published: July 2002

T cells recognize antigen by formation of a trimolecular complex in which the T-cell receptor (TCR) recognizes a specific peptide antigen within the groove of a major histocompatibility complex (MHC) molecule. It has generally been assumed that T-cell recognition of two distinct MHC–antigen complexes is due to similarities in the three-dimensional structure of the complexes. Here we report results of experiments examining the crossreactivity of TCRs recognizing the myelin basic protein peptide MBPp85–99 and several of its analogs in the context of MHC. We demonstrate that single conservative amino acid substitutions of the antigenic peptide at the predominant TCR contact residues at positions 91 and 93 totally abrogate reactivity of specific T-cell clones. Yet, when a conservative substitution is made at position 91 concomitant with a substitution at position 93, the T-cell clones regain reactivity equivalent with that of the original stimulating peptide. Thus, the exact nature of the amino acid side chains engaging one TCR functional pocket may change the apparent selectivity of the other predominant TCR functional pocket, thus suggesting a remarkable degree of receptor plasticity. This ability of the TCR–MHC–peptide complex to undergo conformational changes provides a conceptual framework for reconciling the apparent paradox of the extreme selectivity of the TCR and its remarkable crossreactivity with different MHC–peptide complexes.
Dictyostelium
myosin II null mutant can still cap Con A receptors
Authors: Aguado-Velasco Carmen , Bretscher Mark S.
Published: July 2002

Cross-linked antigens on the surface of a motile cell cap at the trailing end of the cell. In
Dictyostelium discoideum,
myosin II null mutants have previously been reported to be unable to cap Con A receptors, although they are able to locomote. This finding implicated myosin II as an essential component of the capping mechanism, although not of the machinery for locomotion. Here we show that myosin II null mutants do cap Con A receptors, albeit less efficiently than does wild type. This shows that cap formation is not absolutely dependent on myosin II and that a close mechanistic relationship between capping, particle movement, and cell migration may still exist.
Workshop on Schizophrenia
Authors: Barondes Samuel H. , Alberts Bruce M. , Andreasen Nancy C. , Bargmann Cornelia , Benes Francine , Goldman-Rakic Patricia , Gottesman Irving , Heinemann Stephen F. , Jones Edward G. , Kirschner Marc , Lewis David , Raff Martin , Roses Allen , Rubenstein John , Snyder Solomon , Watson Stanley J. , Weinberger Daniel R. , Yolken Robert H.
Published: July 2002

On November 29–30, 1995, the National Academy of Sciences and the Institute of Medicine brought together experts in schizophrenia and specialists in other areas of the biological sciences in a workshop aimed at promoting the application of the latest biological information to this clinical problem. The workshop paid particular attention to evidence of pathology in the brains of people with schizophrenia, and to the possibility that this reflects an abnormality in brain development that eventually leads to the appearance of symptoms. The participants were impressed with the complexity of the problem, and felt that multiple approaches would be required to understand this disease. They recommended that a major focus should be on the search for predisposing genes, but that there should be parallel research in many other areas.
Modular synthesis of
de novo-
designed metalloproteins for light-induced electron transfer
Authors: Rau Harald K. , DeJonge Niels , Haehnel Wolfgang
Published: July 2002

The design and chemical synthesis of two
de novo
four-helix bundle proteins is described; each protein has two bound cofactors. Their construction from purified peptides is based on the modular assembly of different amphiphilic helices by chemoselective coupling to a cyclic peptide template. In the hydrophobic interior of the antiparallel four-helix bundle these proteins contain a heme in a binding pocket with two ligating histidine residues. A ruthenium-tris(bipyridine) complex is covalently bound to different positions at the hydrophilic side of one of the heme-binding helices. Laser-induced electron transfer across the varied distance through this helix has been studied and compared with a pathway analysis. The UV-visible, CD, and mass spectra are consistent with the structure and orientation predetermined by the template.
Ab initio
calculation of electronic coupling in the photosynthetic reaction center
Authors: Zhang Linda Yu , Friesner Richard A.
Published: July 2002

We have carried out an
ab initio
electronic structure calculations of electron transfer couplings between chromophores in the bacterial photosynthetic reaction center. The couplings agree remarkably well with parameters obtained from recent quantum dynamical modeling of experimental data assuming an explicit intermediate mechanism. We also have computed couplings on the M-side of the reaction center and have found that the interaction of the primary donor to the M-side intermediate bacteriochlorophyll is quite small because of destructive interference of the two localized coupling matrix elements. This may explain the slow rate of electron transfer down the M-side of the reaction center.
Multiple domains of repressor activator protein 1 contribute to facilitated binding of glycolysis regulatory protein 1
Authors: López M. Cecilia , Smerage Jeffery B. , Baker Henry V.
Published: July 2002

The function of repressor activator protein 1 (Rap1p) at glycolytic enzyme gene upstream activating sequence (UAS) elements in
Saccharomyces cerevisiae
is to facilitate binding of glycolysis regulatory protein 1 (Gcr1p) at adjacent sites. Rap1p has a modular domain structure. In its amino terminus there is an asymmetric DNA-bending domain, which is distinct from its DNA-binding domain, which resides in the middle of the protein. In the carboxyl terminus of Rap1p lie its silencing and putative activation domains. We carried out a molecular dissection of Rap1p to identify domains contributing to its ability to facilitate binding of Gcr1p. We prepared full-length and three truncated versions of Rap1p and tested their ability to facilitate binding of Gcr1p by gel shift assay. The ability to detect ternary complexes containing Rap1p⋅DNA⋅Gcr1p depended on the presence of binding sites for both proteins in the probe DNA. The DNA-binding domain of Rap1p, although competent to bind DNA, was unable to facilitate binding of Gcr1p. Full-length Rap1p and the amino- and carboxyl-truncated versions of Rap1p were each able to facilitate binding of Gcr1p at an appropriately spaced binding site. Under these conditions, Gcr1p displayed an approximately 4-fold greater affinity for Rap1p-bound DNA than for otherwise identical free DNA. When spacing between Rap1p- and Gcr1p-binding sites was altered by insertion of five nucleotides, the ability to form ternary Rap1p⋅DNA⋅Gcr1p complexes was inhibited by all but the DNA-binding domain of Rap1p itself; however, the ability of each individual protein to bind the DNA probe was unaffected.
Evidence for the load-dependent mechanical efficiency of individual myosin heads in skeletal muscle fibers activated by laser flash photolysis of caged calcium in the presence of a limited amount of ATP
Authors: Sugi Haruo , Iwamoto Hiroyuki , Akimoto Tsuyoshi , Ushitani Hiroko
Published: July 2002

Although a contracting muscle regulates its energy output depending on the load imposed on it (“Fenn effect”), the mechanism underlying the load-dependent energy output remains obscure. To explore the possibility that the mechanical efficiency, with which chemical energy derived from ATP hydrolysis is converted into mechanical work, of individual myosin heads changes in a load-dependent manner, we examined the auxotonic shortening of glycerinated rabbit psoas muscle fibers, containing ATP molecules almost equal in number to the myosin heads, after laser-flash photolysis of caged calcium. Immediately before laser-flash activation, almost all of the myosin heads in the fiber are in the state M⋅ADP⋅Pi, and can undergo only one ATP hydrolysis cycle after activation. When the fibers were activated to shorten under various auxotonic loads, the length, force, and power output changes were found to be scaled according to the auxotonic load. Both the power and energy outputs were maximal under a moderate auxotonic load. The amount of M⋅ADP⋅Piutilized at a time after activation was estimated from the amount of isometric force developed after interruption of fiber shortening. This amount was minimal in the isometric condition and increased nearly in proportion to the distance of fiber shortening. These results are taken as evidence that the efficiency of chemomechanical energy conversion in individual myosin heads changes in a load-dependent manner.
Cross-modal reorganization of callosal connectivity without altering thalamocortical projections
Authors: Pallas Sarah L. , Littman Thomas , Moore David R.
Published: July 2002

Mammalian cerebral cortex is composed of a multitude of different areas that are each specialized for a unique purpose. It is unclear whether the activity pattern and modality of sensory inputs to cortex play an important role in the development of cortical regionalization. The modality of sensory inputs to cerebral cortex can be altered experimentally. Neonatal diversion of retinal axons to the auditory thalamus (cross-modal rewiring) results in a primary auditory cortex (AI) that resembles the primary visual cortex in its visual response properties and topography. Functional reorganization could occur because the visual inputs use existing circuitry in AI, or because the early visual inputs promote changes in AI’s circuitry that make it capable of constructing visual receptive field properties. The present study begins to distinguish between these possibilities by exploring whether the callosal connectivity of AI is altered by early visual experience. Here we show that early visual inputs to auditory thalamus can reorganize callosal connections in auditory cortex, causing both a reduction in their extent and a reorganization of the pattern. This result is distinctly different from that in deafened animals, which have widespread callosal connections, as in early postnatal development. Thus, profound changes in cortical circuitry can result simply from a change in the modality of afferent input. Similar changes may underlie cortical compensatory processes in deaf and blind humans.
Transcriptional regulation of the rat Müllerian inhibiting substance type II receptor in rodent Leydig cells
Authors: Teixeira Jose , Kehas David J. , Antun Reem , Donahoe Patricia K.
Published: July 2002

Müllerian inhibiting substance (MIS) causes regression of the fetal Müllerian duct on binding a heteromeric complex of types I and II cell-surface receptors in the fetal urogenital ridge. The MIS type II receptor (MISRII), which provides specificity for MIS, is also expressed in the adult testis, ovary, and uterus. The rat MISRII promoter was cloned to study the molecular mechanisms underlying its temporal and cell-specific expression. The 1.6-kilobase (kb) promoter contained no recognizable TATA or CAAT box, but there was a consensus Sp1 site upstream of the transcription initiation site. Two binding sites for the orphan nuclear receptor steroidogenic factor-1 (SF-1) are occupiedin vitroby using nuclear extracts from R2C cells, an MIS-responsive rat Leydig cell line that expresses endogenous MISRII, with differing affinities, indicating that the distal SF-1 site is bound more avidly than is the proximal SF-1 site. R2C cells transfected with MISRII promoter/luciferase reporter constructs show a 12-fold induction with the 1.6-kb fragment and deletion of sequences upstream of −282-bp lowered luciferase expression to one-third. Mutation of both SF-1 sites greatly inhibited luciferase expression, whereas mutation of either site alone resulted in continuing activation by endogenous SF-1, indicating redundancy.In vitrobinding and transcriptional analyses suggest that a proximal potential Smad-responsive element and an uncharacterized element also contribute to activation of the MISRII gene. R2C cells and MISRII promoter regulation can now be used to uncover endogenous transcription factors responsible for receptor expression or repression.
Le traitement des voyelles nasalisées et des groupes consonantiques par l'apprenti‐lecteur francophone
Authors: Magnan Annie , Bianchéri Patricia
Published: July 2002

Cette étude porte sur le traitement des syllabes complexes par l'apprenti‐lecteur francophone. On s'intéresse au traitement de structures phonologiques CV avec voyelle nasale (CAN) (expérience 1) et au traitement des groupes consonantiques (CRA) (expérience 2) placées à l'initiale d'un item. On propose à des enfants bons et faibles lecteurs de première et deuxième année d'apprentissage une tâche de jugement d'identité entre un item‐cible présenté en modalité auditive et un item‐test présenté en modalité visuelle. Les résultats montrent la tendance des jeunes lecteurs à simplifier les mots polysyllabiques qu'ils ont à lire en les transformant en suite de syllabes ouvertes élémentaires de type CV. Ces résultats confirment l'hypothèse selon laquelle les caractéristiques linguistiques spécifiques à une langue influencent le mode de traitement des mots écrits et donc leur acquisition.
EEG Aspects of Cognitive Processes: A Contribution to the Proteus‐like Nature of Consciousness
Authors: Petsche Hellmuth , Etlinger Susan C.
Published: July 2002

Probability mapping of amplitude and coherence based on spectral analysis yields information on the cooperation of brain areas in cognitive processes. The method is based on comparing these two parameters during the performance of a mental task with the respective values of the EEG at rest. For each of 6 frequency bands spanning the entire EEG spectrum the data are represented on schematic brain maps with significant changes of amplitude (entered at the sites of the 19 electrodes) and coherence (between all respective electrodes). These maps have proven to be largely task‐specific in both group and individual studies. They are considered to objectify electrical manifestations of differential attention.
The Effects of Age‐based and Grade‐based Sampling on the Relative Standing of Countries in International Comparative Studies of Student Achievement
Authors: O'Leary Michael
Published: July 2002

Abstract The investigation reported in this article was prompted by discrepancies between the published outcomes from two international tests of science achievement: the Second International Assessment of Educational Progress (IAEP2) administered in 1991 and the Third International Mathematics and Science Study (TIMSS) administered in 1995. One finding was that while average science achievement for Irish 13 year‐olds was reported to be at the low end of the distribution for the 20 participating countries in IAEP2, it was around the middle of the distribution for the 40 or so countries that participated in TIMSS in the early grades of secondary schooling. Initial comparisons suggested that there were also inconsistencies in outcomes for some of the 11 other countries that participated in both surveys, e.g. France, Portugal, and Switzerland. Analyses described in the article reveal that when sampling/population definition differences between the two surveys are accounted for, science achievement in Ireland was not at the low level suggested by initial interpretations of the IAEP2 data but was closer to the levels reported in TIMSS. While the sampling issue did not fully account for discrepancies with respect to the IAEP2/TIMSS outcomes for some countries, it is argued that the findings outlined in this article have a number of implications for policy‐makers using data from future international comparative studies of student achievement.
Linking and Optimality in the Norwegian Presentational Focus Construction
Authors: Lødrup Helge
Published: July 2002

The presentational focus construction in Norwegian can have an agent in object position, as inDet arbeidet en mann i skogen‘It worked a man in the-woods”. This creates problems for linking theory, in which a robust generalization says that agents are not realized as objects. Optimality Theory makes possible a new approach to this classical problem in Scandinavian generative grammar. The constraint that agents are not realized as objects must compete with other constraints. In Norwegian, it is ranked below a requirement that a presentational focus must be realized in object position. The Norwegian situation is compared to languages with different constraint rankings.
Virtual and Actual Existentials in English, Swedish and Icelandic
Authors: Twardzisz Piotr
Published: July 2002

The focus of my analysis is the so-called existential construction. The languages examined are English, Swedish and Icelandic. The present article assumes the perspective of Ronald W. Langacker's cognitive grammar as the theoretical background. First of all, the assumption is that the unstressed, initial pronounthere, or its Scandinavian equivalents, are semantically definable as abstract-setting subjects of their respective sentences, with, possibly, the exception of Icelandic það. Secondly, the conceptualization of the existential scenes in the three languages is a dynamic process in each case. The dynamicity of the semantics of existential scenes is the result of assuming two planes, the actual and a virtual one, and establishing correspondences between them. The actual plane reflects our direct apprehension of reality. A virtual plane consists in the dynamic re-assignment of roles to the actual elements introduced by means of the virtual abstract-setting subject.
Reassessing naltrexone maintenance as a treatment for illicit heroin users
Authors: Hulse G. K. , Basso M. R.
Published: July 2002

Most studies investigating the efficacy of naltrexone maintenance as a treatment for illicit heroin users have reported poor outcomes. Many of these studies, however, classify patients who periodically return to heroin use while receiving naltrexone maintenance as treatment failures. This study investigated 6‐month outcome status in 100 illicit heroin users who commenced naltrexone maintenance in a community‐based out‐patient treatment programme. The study aimed to assess patient status at 6 months using contrasting outcome criteria. In the first analysis periodic heroin use was not considered to constitute treatment failure (hence naltrexone was continued and they remained under observation). In the second analysis, return to heroin use was considered to represent treatment failure. Using the first approach, we found that 60% of patients were still on naltrexone maintenance at 6 months, with only a small number (28%) having returned to heroin use. In contrast, when return to heroin use for 7 or more consecutive days was regarded as a treatment failure, only 31% of patients were “retained in treatment” with the majority (62%) having returned to heroin use. It is argued that since periods of heroin use are commonly associated with attempting to manage heroin dependence, the analysis that allowed periodic heroin use during treatment, and produced positive outcome data, represented a more valid assessment of naltrexone maintenance as a treatment for illicit heroin users. It is suggested that this outcome criterion should therefore be employed in future research. Poor outcomes reported in many previous studies may be an artefact of using inappropriate outcome criteria to assess illicit heroin use.
The impact of naloxone/lofexidine combination treatment on the opiate withdrawal syndrome
Authors: Bearn J. , Bennett J. , Martin T. , Gossop M. , Strang J.
Published: July 2002

Clinical studies in opiate‐dependent patients suggest that detoxification treatment with opiate antagonists may accelerate the resolution of the opiate withdrawal syndrome, permitting early induction onto naltrexone maintenance treatment. The present open study compared the clinical efficacy of daily naloxone injections in conjunction with lofexidine, with conventional lofexidine monotherapy, in 49 polysubstance‐misusing opiate‐dependent patients. Overall, the addition of naloxone did not confer substantial benefit over lofexidine monotherapy, although area‐under‐the‐curve analysis showed that withdrawal severity in the naloxone/lofexidine combination group was significantly less than in the lofexidine monotherapy group, who experienced more severe withdrawal symptoms on days 4, 7, 9 and 13 of treatment. There were no significant differences in rates of completion of detoxification. Blood pressure remained within normal limits in both groups. Naltrexone maintenance treatment acceptability was low; only four patients continued with treatment for 5 or more days. The modest benefit of adding naloxone to lofexidine compared to the findings of previous opiate antagonist detoxification treatment studies is discussed in the context of the hypothesis that a critical level of opiate receptor occupancy is required to accelerate resolution of opiate withdrawal; the neurochemical mechanisms which may promote this are discussed.
Phenylalkylamine abuse among opiate addicts attending a methadone treatment programme in the Republic of Ireland
Authors: Kavanagh P. , Dunne J. , Feely J. , Maguire R. , Corrigan D. , Keating J. J. , Meegan M. J. , Clancy J. M. , Burdett J.
Published: July 2002

Since the early 1990s ring‐substituted derivatives of amphetamine have been abused widely in the Republic of Ireland. The main ring‐substituted amphetamines being abused include methylenedioxyamphetamine (MDA), methylenedioxymethamphetamine (MDMA) and methylenedioxyethamphetamine (MDEA). A newer illicit synthetic analogue, which has been seized to a lesser extent by Irish police, is N‐methyl‐1‐(3,4‐methylenedioxyphenyl)‐2‐butanamine (MBDB). The work presented here involved the determination of the type of ring‐substituted amphetamines being abused by a group of recovering opiate abusers participating in a methadone maintenance programme in a Dublin Drug Rehabilitation Centre. Urine samples which tested positive for amphetamines and ring‐substituted amphetamines via EMIT immunoassay were subjected to further analysis using GC‐MS with MBTFA flash derivatization. It was found that the methylenedioxypropanamines were being abused, as was amphetamine itself. However, no abuse of methylenedioxybutanamines or thioamphetamines was observed.
Book and Media Reviews
Authors: BALL DAVID
Published: July 2002

Books reviewed in this article:

Book reviews in this column will primarily be of titles focusing completely, or in part, on biological aspects of addiction. However, significant titles of general relevance to the addictions field will also be included, even if they are not “biological”, as will titles of general methodological and clinical relevance, even if they are not on “addictions”. Similar considerations will apply to other media (software, audio tapes and CDs, videos, etc.). However, specific “addictions” software applications seem to be relatively uncommon and, as these items are rarely reviewed elsewhere, we will endeavour to include reviews of some of the older programmes that are still useful, as well as new titles that appear. I would appreciate suggesti ons of any items suitable for reviews, but especially software and other media of specifi c relevance to the addictions. Please contact: Dr David Ball, National Addiction Centre, 4 Windsor Walk, London SE5 8AF, UK. Email: media@addictionbiology.com

Peer Review in Health Sciences F. GODLEE & T. JEFFERSON

Controlling our Destinies: historical, philosophical, ethical and theological perspectives on the Human Genome Project P.R. SLOAN (Ed.)

The Human Brain: essentials of behavioural neuroscience J. BEATTY

Modern Chromatographic Analysis of Vitamins, 3rd edition A. P. DE LEENHEER, W.E LAMBERT & J.F. VAN BOCXLAER

Multivariate Applications in Substance Use Research: new methods for new questions J.S. ROSE, L. CHASSIN, C.C. PRESSON & S.J. SHERMAN (Eds)
Background genotype modulates the effects of γ‐PKC on the development of rapid tolerance to ethanol‐induced hypothermia
Authors: Bowers Barbara J. , Collins Allan C. , Wehner Jeanne M.
Published: July 2002

The role of γ‐PKC in initial sensitivity and in the development of rapid tolerance to the hypothermic effects of ethanol were investigated in γ‐PKC null mutant mice. Effects of the single gene mutation were evaluated on three different genetic backgrounds. Null mutants from a C57BL/6J X 129/SvJ mixed genetic background failed to develop rapid tolerance after 4 days of i.p. ethanol injections. However, when the null mutation was introgressed onto a C57BL/6J background for six generations to create a congenic line, the expression of rapid tolerance unexpectedly reoccurred in the null mutant mice. Subsequent outcrossing of the γ‐PKC null mutation to a C57BL/6J X 129/SvEvTac mixed background did not restore the no tolerance phenotype. These observations, taken together with similar results reported previously concerning the development of chronic tolerance to ethanol in these same genotypes, 1 indicate that the gene coding for γ‐PKC has pleiotropic effects in the expression of both rapid and chronic tolerance to ethanol‐induced hypothermia. However, the impact of γ‐PKC is modulated by the background genotype. These results stress the necessity of understanding interactions with genetic background when interpreting the effects of single gene mutations on complex behavioral traits.
Assembly and Intracellular Targeting of the βγ Subunits of Heterotrimeric G Proteins
Authors: Rehm Armin , Ploegh Hidde L.
Published: July 2002

The assembly in living cells of heterotrimeric guanine nucleotide binding proteins from their constituent α, β, and γ subunits is a complex process, compounded by the multiplicity of the genes that encode them, and the diversity of receptors and effectors with which they interact. Monoclonal anti-β antibodies (ARC5 and ARC9), raised against immunoaffinity purified βγ complexes, recognize β subunits when not associated with γ and can thus be used to monitor assembly of βγ complexes. Complex formation starts immediately after synthesis and is complete within 30 min. Assembly occurs predominantly in the cytosol, and association of βγ complexes with the plasma membrane fraction starts between 15–30 min of chase. Three pools of β subunits can be distinguished based on their association with γ subunits, their localization, and their detergent solubility. Association of β and α subunits with detergent-insoluble domains occurs within 1 min of chase, and increases to reach a plateau of near complete detergent resistance within 30 min of chase. Brefeldin A treatment does not interfere with delivery of βγ subunits to detergent-insoluble domains, suggesting that assembly of G protein subunits with their receptors occurs distally from the BFA-imposed block of intracellular membrane trafficking and may occur directly at the plasma membrane.
KLP38B: A Mitotic Kinesin-related Protein That Binds PP1
Authors: Alphey Luke , Parker Louise , Hawcroft Gillian , Guo Yiquan , Kaiser Kim , Morgan Gareth
Published: July 2002

We have identified a new member of the kinesin superfamily in Drosophila, KLP38B (kinesin-like protein at 38B). KLP38B was isolated through its two-hybrid interaction with the catalytic subunit of type 1 serine/threonine phosphoprotein phosphatase (PP1). We demonstrate that recombinant KLP38B and PP1 associate in vitro. This is the first demonstration of direct binding of a kinesin-related protein to a regulatory enzyme.

Though most closely related to the Unc-104 subfamily of kinesin-related proteins, KLP38B is expressed only in proliferating cells. KLP38B mutants show cell proliferation defects in many tissues. KLP38B is required for normal chromatin condensation as embryos from KLP38B mutant mothers have undercondensed chromatin at metaphase and anaphase. This is the first time that a kinesin-related protein has been shown to have such a role. Incomplete lethality of a strong KLP38B allele suggests partial redundancy with one or more additional kinesin-related proteins.
Interleukin 4 Gene–defective Mice Reconstituted with Wild-type Bone Marrow Fail to Produce Normal Immunoglobulin E Levels
Authors: Lange Claudia , Schüler Thomas , Blankenstein Thomas
Published: July 2002

The ability to reconstitute interleukin (IL)-4−/− mice with bone marrow of IL-4+/+ mice was investigated. The absence of the IL-4−/− gene in donor or recipient cells did not impair the reconstitution. All immunoglobulin (Ig) subsets occurred at normal serum levels except for IgE and to some extent IgG1. IgE production did not recover in the reconstituted mice over prolonged time. However, these mice were competent for IgE production, because a single intrasplenic injection of IL-4 restored IgE levels, which then remained constant. Wild-type mice reconstituted with wild-type bone marrow constantly had IgE serum levels comparable to untreated animals. In wild-type mice reconstituted with IL-4−/− bone marrow, IgE levels dropped gradually and disappeared by week 12. We make three unrelated but nonetheless important conclusions: (a) (immunoregulation) the tightly regulated IL-4 gene should be expressed constantly in low amounts (and with apparent absence of antigen stimulation) to keep the normal threshold of IgE; (b) (ontogeny of the immune system) an early unidentified source of IL-4 must be postulated which is lost in adult mice; and (c) (bone marrow transfer/gene therapy) under certain circumstances, the genotype of the recipient influences the reconstitution.
Transplanted Long-Term Cultured Pre-Bi Cells Expressing Calpastatin Are Resistant to B Cell Receptor–Induced Apoptosis
Authors: Ruiz-Vela Antonio , Serrano Fernando , González Manuel A. , Abad José Luis , Bernad Antonio , Maki Masatoshi , Martínez-A Carlos
Published: July 2002

Long-term cultured pre-B cells are able to differentiate into immunoglobulin (Ig)M-positive B cells (IgM+ cells) when transplanted into severe combined immunodeficient (SCID) mice. Based on previous studies, here we report the development of a reconstitution assay in nonobese diabetic/SCID (NOD/SCID) mice using pre-B cells, which allows us to study the role of calpains (calcium-activated endopeptidases) during B cell development as well as in B cell clonal deletion. Using this model, we show that calpastatin (the natural inhibitor of calpains) inhibits B cell receptor–induced apoptosis in IgM+ cells derived from transplanted mice. We thus hypothesize an important function for calpain in sculpting the B cell repertoire.
Eliciting the Low-Activity Aldehyde Dehydrogenase Asian Phenotype by an Antisense Mechanism Results in an Aversion to Ethanol
Authors: Garver Eric , Tu Guang-chou , Cao Qing-Na , Aini Maria , Zhou Feng , Israel Yedy
Published: July 2002

A mutation in the gene encoding for the liver mitochondrial aldehyde dehydrogenase (ALDH2–2), present in some Asian populations, lowers or abolishes the activity of this enzyme and results in elevations in blood acetaldehyde upon ethanol consumption, a phenotype that greatly protects against alcohol abuse and alcoholism. We have determined whether the administration of antisense phosphorothioate oligonucleotides (ASOs) can mimic the low-activity ALDH2–2 Asian phenotype. Rat hepatoma cells incubated for 24 h with an antisense oligonucleotide (ASO-9) showed reductions in ALDH2 mRNA levels of 85% and ALDH2 (half-life of 22 h) activity of 55% equivalent to a >90% inhibition in ALDH2 synthesis. Glutamate dehydrogenase mRNA and activity remained unchanged. Base mismatches in the oligonucleotide rendered ASO-9 virtually inactive, confirming an antisense effect. Administration of ASO-9 (20 mg/kg/day for 4 d) to rats resulted in a 50% reduction in liver ALDH2 mRNA, a 40% inhibition in ALDH2 activity, and a fourfold (P < 0.001) increase in circulating plasma acetaldehyde levels after ethanol (1 g/kg) administration. Administration of ASO-9 to rats by osmotic pumps led to an aversion (−61%, P < 0.02) to ethanol. These studies provide a proof of principle that specific inhibition of gene expression can be used to mimic the protective effects afforded by the ALDH2–2 phenotype.
Cyclic AMP Levels, Adenylyl Cyclase Activity, and Their Stimulation by Serotonin Quantif ied in Intact Neurons
Authors: Sudlow Leland C. , Gillette Rhanor
Published: July 2002

In molluscan central neurons that express cAMP-gated Na+ current (INa,cAMP), estimates of the cAMP binding affinity of the channels have suggested that effective native intracellular cAMP concentrations should be much higher than characteristic of most cells. Using neurons of the marine opisthobranch snail Pleurobranchaea californica, we applied theory and conventional voltage clamp techniques to use INa,cAMP to report basal levels of endogenous cAMP and adenylyl cyclase, and their stimulation by serotonin. Measurements were calibrated to iontophoretic cAMP injection currents to enable expression of the data in molar terms. In 30 neurons, serotonin stimulated on average a 23-fold increase in submembrane [cAMP], effected largely by an 18-fold increase in adenylyl cyclase activity. Serotonin stimulation of adenylyl cyclase and [cAMP] was inversely proportional to cells' resting adenylyl cyclase activity. Average cAMP concentration at the membrane rose from 3.6 to 27.6 μM, levels consistent with the expected cAMP dissociation constants of the INa,cAMP channels. These measures confirm the functional character of INa,cAMP in the context of high levels of native cAMP. Methods similar to those employed here might be used to establish critical characters of cyclic nucleotide metabolism in the many cells of invertebrates and vertebrates that are being found to express ion currents gated by direct binding of cyclic nucleotides.
Waveform relaxation as a dynamical system
Authors: Bjørhus Morten , Stuart Andrew
Published: July 2002

In this paper the properties of waveform relaxation are studied when applied to the dynamical system generated by an autonomous ordinary differential equation. In particular, the effect of the waveform relaxation on the invariant sets of the flow is analysed. Windowed waveform relaxation is studied, whereby the iterative technique is applied on successive time intervals of lengthTTand a fixed, finite, number of iterations taken on each window. This process does not generate a dynamical system onR+\mathbb {R}^+since two different applications of the waveform algorithm over different time intervals do not, in general, commute. In order to generate a dynamical system it is necessary to consider the timeTTmap generated by the relaxation process. This is done, andC1C^1-closeness of the resulting map to the timeTTmap of the underlying ordinary differential equation is established. Using this, various results from the theory of dynamical systems are applied, and the results discussed.
On Cherednik-Macdonald-Mehta identities
Authors: Etingof Pavel , Kirillov Alexander
Published: July 2002

In this note we give a proof of Cherednik’s generalization of Macdonald–Mehta identities for the root systemAn−1A_{n-1}, using representation theory of quantum groups. These identities give an explicit formula for the integral of a product of Macdonald polynomials with respect to a “difference analogue of the Gaussian measure”. They were suggested by Cherednik, who also gave a proof based on representation theory of affine Hecke algberas; our proof gives a nice interpretation for these identities in terms of representations of quantum groups and seems to be simpler than that of Cherednik.
Organ/patient geometric variation in external beam radiotherapy and its effects
Authors: Yan Di , Lockman David
Published: July 2002

Treatment variation in positioning of the organ/patient with respect to the radiation beams causes a temporal dose variation in critical normal tissues adjacent to the treatment target. This temporal variation induces uncertainties in understanding the normal tissue dose response, thereby limiting reliable treatment evaluation and optimization. The aim of this study is to model and analyze the temporal variation of organ dose distribution, and its effect on the biological effective dose. The study mainly focuses on the temporal dose variation caused by intertreatment organ motion/deformation and daily setup error. Sensitivity of the biological effective dose to organ/patient geometric variation, dose distribution, and treatment fractionation will be investigated. Significant deviation of the biological effective dose could be expected in a critical normal structure, even if the cumulative dose deviation in this structure is negligible. Patients with similar geometric variation characteristics can experience significantly different biological effective dose, and the differences are sensitive to the dose distribution and the total number of treatment fractions.
The thermal characteristics of different diodes on in vivo patient dosimetry
Authors: Welsh Keith T. , Reinstein L. E.
Published: July 2002

Diode sensitivity variations with temperature (SVWT) have been reported to vary from small negative values up to 0.6% per °C. Thus it is possible for diode calibration factors established at room temperature (∼20 °C) to yield errors in the range of −1% to +9% when diodes are placed on a patient's skin (∼30 °C) for in vivo entrance dose measurements. In this study we simulated several skin temperatures using a temperature‐controlled aluminum surface in contact with a section of Bolus. The internal temperatures of several diodes with different buildup thickness were monitored as a function of time when placed in contact with the heated bolus. Our results indicate that for different combinations of room temperature (18 °C–23 °C) and patient skin temperature (28 °C–34 °C) diodes reached 90% of their equilibrium temperature within 3–5 min. In addition, the range of typical skin temperatures was determined by measurements performed on a number of actual patients under clinical conditions. Based on the results of our experiments a protocol was developed to minimize the temperature based errors for in vivo dosimetry.
Breast image registration using a textural transformation
Authors: Sivaramakrishna Radhika
Published: July 2002

Small breast tumors could be detected early by registering two longitudinal breast images and subtracting them. The noise and low contrast in breast images permit few control points for image registration. We propose a new textural transformation, called the “starbyte” transformation, to solve this image registration problem. The starbyte transform is computed using local neighborhood properties around every pixel in an image and converts breast images to texture maps with clearly demarcable regions from which accurate control points can be easily identified. A complete registration algorithm is presented which consists of first starbyte transforming the original images, extracting control points (both manually and automatically), and performing unwarping using thin plate splines. The performance of the algorithm was rigorously tested on portions of mammograms warped by six different types of distortions. Limited studies have also been performed with added noise and simulated tumors. Preliminary results are also presented on roughly corresponding 2D slices taken from 3D MRIs of volunteers imaged twice with breast repositioning between the two images.
A numerical method to predict the effects of frequency-dependent attenuation and dispersion on speed of sound estimates in cancellous bone
Authors: Wear Keith A.
Published: July 2002

Many studies have demonstrated that time-domain speed-of-sound (SOS) measurements in calcaneus are predictive of osteoporotic fracture risk. However, there is a lack of standardization for this measurement. Consequently, different investigators using different measurement systems and analysis algorithms obtain disparate quantitative values for calcaneal SOS, impairing and often precluding meaningful comparison and/or pooling of measurements. A numerical method has been developed to model the effects of frequency-dependent attenuation and dispersion on transit-time-based SOS estimates. The numerical technique is based on a previously developed linear system analytic model for Gaussian pulses propagating through linearly attenuating, weakly dispersive media. The numerical approach is somewhat more general in that it can be used to predict the effects of arbitrary pulse shapes and dispersion relationships. The numerical technique, however, utilizes several additional assumptions (compared with the analytic model) which would be required for the practical task of correcting existing clinical databases. These include a single dispersion relationship for all calcaneus samples, a simple linear model relating phase velocity to broadband ultrasonic attenuation, and a constant calcaneal thickness. Measurements on a polycarbonate plate and 30 human calcaneus samples were in good quantitative agreement with numerical predictions. In addition, the numerical approach predicts that in cancellous bone, frequency-dependent attenuation tends to be a greater contributor to variations in transit-time-based SOS estimates than dispersion. This approach may be used to adjust previously acquired individual measurements so that SOS data recorded with different devices using different algorithms may be compared in a meaningful fashion.
Sources of distortion product otoacoustic emissions revealed by suppression experiments and inverse fast Fourier transforms in normal ears
Authors: Konrad-Martin Dawn , Neely Stephen T. , Keefe Douglas H. , Dorn Patricia A. , Gorga Michael P.
Published: July 2002

Primary and secondary sources combine to produce the 2f1-f2 distortion product otoacoustic emission (DPOAE) measured in the ear canals of humans. DPOAEs were obtained in nine normal-hearing subjects using a fixed-f2 paradigm in which f1 was varied. The f2 was 2 or 4 kHz, and absolute and relative primary levels were varied. Data were obtained with and without a third tone (f3) placed 15.6 Hz below 2f1-f2. The level of f3 was varied in order to suppress the stimulus frequency otoacoustic emission (SFOAE) coming from the 2f1-f2 place. These data were converted from the complex frequency domain into an equivalent time representation using an inverse fast Fourier transform (IFFT). IFFTs of unsuppressed DPOAE data were characterized by two or more peaks. Relative amplitudes of these peaks depended on overall primary level and on primary-level differences. The suppressor eliminated later peaks, but early peaks remained relatively unaltered. Results are interpreted to mean that the DPOAE measured in humans includes components from the f2 place (intermodulation distortion) and DP place (in the form of a SFOAE). These findings build on previous work by providing evidence that multiple peaks in the IFFT are due to a secondary source at the DP place.