bluuebunny/crossref_metadata_2025 huggingface dataset:
The efficacy of reboxetine in the treatment‐refractory patients with panic disorder: an open label study
Authors: Dannon P. N. , Iancu I. , Grunhaus L.
Published: November 2002

Background and Objective

Selective serotonin reuptake inhibitors (SSRIs) are currently the first‐line treatment for panic disorder, although up to 30% of patients either do not respond to SSRIs or withdraw due to adverse events. Reboxetine, a selective norepinephrine reuptake inhibitor (selective NRI), is effective in treating depression and may alleviate depression‐related anxiety. This study aimed to investigate the efficacy of reboxetine in the treatment of patients with panic disorder who did not respond to SSRIs.

Method

In this 6‐week, open‐label study, 29 adult outpatients with panic disorder who had previously failed to respond to SSRI treatment received reboxetine 2 mg/day, titrated to a maximum of 8 mg/day over the first 10 days. Efficacy was assessed using the Panic Self‐Questionnaire (PSQ), the Hamilton Rating Scale for Anxiety (HAM‐A), the 17‐item Hamilton Rating Scale for Depression (HRSD) and the Global Assessment of Functioning (GAF) Scale.

Results

The 24 patients who completed the study responded well to reboxetine treatment. Significant improvement (p < 0.001) was observed in the number of daily panic attacks, and on the scales measuring anxiety, depression and functioning. Reboxetine was generally well tolerated. Five patients withdrew due to adverse events.

Conclusions

Reboxetine appears to be effective in the treatment of SSRI‐refractory panic disorder patients and warrants further clinical investigation. Copyright © 2002 John Wiley & Sons, Ltd.
Acute effects of cigarettes in non‐deprived smokers on memory, calculation and executive functions
Authors: Sakurai Yasuhisa , Kanazawa Ichiro
Published: November 2002

To determine whether smoking one or two cigarettes daily has effects on cognitive functions and the blood nicotine level, we examined 20 smokers and 20 nonsmokers as part of a normal daily smoking regimen using psychological tests for memory, calculation and association. Subjects underwent Buschke's selective reminding test, a mental arithmetic task (multiplication between two digits and one digit) and an association task (letter fluency); the duration of each task being 2 min under the condition of pre‐smoking, one cigarette smoking and two cigarette smoking sessions, with an inter‐session interval of 50 min. The total recall or correct responses were evaluated. The results showed that performance deteriorated in the later sessions in the memory task, whereas it improved in the later sessions in the calculation and association tasks, with no group (smoker–nonsmoker) difference, nor interaction between tasks and groups in each task. Plasma nicotine increased by 10 ng/ml immediately after smoking a cigarette, and 14 ng/ml immediately after smoking two cigarettes, but decreased to the pre‐smoking level or slightly higher 10 min after smoking. These findings suggested that a daily dose of nicotine had little effect on the performance related to memory, calculation and mental association. Copyright © 2002 John Wiley & Sons, Ltd.
Evolution of mRNA polyadenylation between oocyte maturation and first embryonic cleavage in cattle and its relation with developmental competence
Authors: Brevini T.A.L. , Lonergan P. , Cillo F. , Francisci C. , Favetta L.A. , Fair T. , Gandolfi F.
Published: November 2002

In this study we analyzed the pattern of polyadenylation changes that takes place between the resumption of meiosis and the first cleavage of bovine oocytes. Moreover, we investigated whether the delayed occurrence of the first cleavage division, which characterizes embryos of low developmental competence, is accompanied by an altered polyadenylation pattern of individual transcripts. We determined the polyadenylation status of a group of genes that characterize physiological processes, involved in early differentiation (Oct‐4), compaction, and cavitation (β‐actin, plakophilin, connexin‐32, connexin‐43), energy metabolism (glucose transporter type 1, pyruvate dehydrogenase phosphatase), RNA processing (RNA poly(A) polymerase), and stress (heat shock protein 70). RNA was isolated from pools of 20 oocytes or embryos at the germinal vesicle (GV) stage, at the end of in vitro maturation, at the end of in vitro fertilization, and at the time of the first cleavage. Cleavage was assessed 27, 30, 36, 42 hr post insemination (hpi), and at the latter time the remaining uncleaved oocytes were retained as a group. Between oocyte isolation and first cleavage at 27 hpi (best quality embryos), the poly(A) tail of individual transcripts followed four patterns: no changes (β‐actin, PDP); gradual reduction (Cx‐43, Oct‐4, Plako); gradual elongation (Cx‐32, TPA); reduction followed by elongation (PAP, HSP‐70, Glut‐1). If the interval between insemination and first cleavage was longer than 27 hpi (progressively lower quality embryos) further changes of polyadenylation were observed, which differed for each gene considered. These data indicated that specific changes in polyadenylation contribute to the modulation of gene expression in bovine embryos at this stage of development. Defective developmental competence is accompanied by abnormal polyadenylation levels of specific maternal mRNAs with synchrony between polyadenylation and cleavage emerging as an apparently important factor. Mol. Reprod. Dev. 63: 510–517, 2002. © 2002 Wiley‐Liss, Inc.
Identification of a novel GPI‐anchored CRISP glycoprotein, MAK248, located on the posterior head and equatorial segment of cynomolgus macaque sperm
Authors: Yudin A.I. , Li M.‐W. , Robertson K.R. , Tollner T. , Cherr G.N. , Overstreet J.W.
Published: November 2002

To identify a sperm‐surface component that is highly antigenic, we immunized female cynomolgus macaques with glycosylphosphatidylinositol (GPI)‐anchored sperm surface proteins that were released following treatment with phosphatidylinositol‐specific phospholipase C (PI‐PLC). Five different adjuvants were used in combination with the PI‐PLC‐released proteins, and three of these proteins (24, 48, and 53 kDa) were shown to be potent antigens for immunization of female monkeys. The 53 kDa protein was found to be a surface coating protein and not a GPI‐anchored protein. Polyclonal antibodies to the 24 kDa protein and the 48 kDa protein were produced in rabbits. The two antibodies recognized both proteins on Western blots. The same rabbit antibodies recognized 28, 18, and 10 kDa bands on a Western blot of chemically reduced PI‐PLC‐released proteins, suggesting that the 48 kDa protein is a dimer of the 24 kDa protein, which we refer to as MAK248. Rabbit polyclonal antibodies developed to reduced fragments of the 24 kDa protein showed that the 18 and 10 kDa bands are proteolytic peptide fragments of the 24 kDa protein. Screening of tissues from male macaques showed that MAK248 is expressed only in the epididymis. Microsequencing of two proteolytic fragments of the 18 kDa component showed 100% amino acid homology to a 233 deduced amino acid sequence previously identified in human testes genome. Antibodies to MAK248 recognized a 24 kDa protein released from human sperm exposed to PI‐PLC. Antibodies to MAK248 recognized the equatorial segment and posterior head regions of capacitated cynomolgus macaque sperm. Structural analysis suggests that MAK248 is a novel CRISP protein and a member of the CAP (CRISP, Ag 5, PR‐1) family of proteins. Based on amino acid sequence homology, it is possible that MAK248 functions as a protease inhibitor. Mol. Reprod. Dev. 63: 488–499, 2002. © 2002 Wiley‐Liss, Inc.
Quantification and distribution of equine oocyte cortical granules during meiotic maturation and after activation
Authors: Carneiro Gustavo F. , Liu Irwin K.M. , Hyde Dallas , Anderson Gary B. , Lorenzo Pedro L. , Ball Barry A.
Published: November 2002

In vitro fertilization (IVF) is being routinely used in humans and several domestic species, however, limited success has been achieved in the horse. Although immature equine oocytes are capable of completing meiosis in vitro, subsequent fertilization, and embryonic development of those oocytes are questionable. The lack of development of these oocytes could be attributed to an impaired cytoplasmic maturation. In the horse, the study of oocyte cytoplasmic maturation and post‐fertilization development has been hindered by the lack of progress in IVF. In mammalian oocytes, migration of cortical granules (CG) has been used as an important criterion to evaluate cytoplasmic maturation. The aim of this study was to describe and quantify the CG distribution of equine oocytes during in vitro meiotic maturation and to assess activation of oocytes with calcium ionophore based upon fluorescein isothiocyanate (FITC)‐labeled Lens culinaris agglutinin (LCA) and laser confocal microscopy. The results of this study indicate that CG are distributed throughout the cytoplasm of oocytes at the germinal vesicle (GV) stage (immature). As maturation proceeds, a progressive centripetal migration of CG to the oocyte cortex occurs with the formation of a monolayer adjacent to the plasma membrane starting by the end of a 30 hr incubation period and increasing significantly after 36 hr. After activation, significant reduction in the number of CG was observed (P < 0.001) suggesting that oocytes cultured under the present conditions possess the ability to release CG in response to the ele‐vation of intracellular free calcium. Mol. Reprod. Dev. 63: 451–458, 2002. © 2002 Wiley‐Liss, Inc.
Inhibition of luteinizing hormone receptor expression during the development of caprine corpora lutea by administration of gonadotropin‐releasing hormone antagonist
Authors: Kawate Noritoshi , Tsuji Makoto , Tamada Hiromichi , Inaba Toshio , Sawada Tsutomu
Published: November 2002

The present study was conducted to examine effects of a potent GnRH antagonist (GA), which suppresses release of luteinizing hormone (LH), on LH receptor expression during the development of the caprine corpus luteum (CL). Goats were divided into control and GA‐treated groups. The goats were treated with saline or GA (50 μg/kg, s.c.) on days 0 (day of ovulation), 4 and 8 (control only), and CL collected on a subset of goats (n = 3 for each day) on days 0 (no saline), 4, 8, or 14 (control only). Ribonuclease protection assay and [125I]‐hCG binding assay were performed to quantitate mRNA and protein of the LH receptor in the CL, respectively. On day 4, CL weight, levels of LH receptor mRNA and protein in the GA‐treated group were similar to those of the control group. By day 8, CL weight and levels of LH receptor mRNA and protein in the GA‐treated group were reduced relative to those of the control group (P < 0.05). There was no difference of affinity of the LH receptor between both groups on day 8. These results suggest that the treatment with GA inhibits gene and protein expressions of the LH receptor during the development of CL in the goat, and thus, support an idea that endogenous LH participates in the increase of its own receptor. Mol. Reprod. Dev. 63: 444–450, 2002. © 2002 Wiley‐Liss, Inc.
Enhanced mass detection of oligonucleotides using reverse‐phase high‐performance liquid chromatography/electrospray ionization ion‐trap mass spectrometry
Authors: Deguchi Kisaburo , Ishikawa Masako , Yokokura Takefumi , Ogata Izumi , Ito Shinya , Mimura Tadao , Ostrander Chad
Published: November 2002

A new method providing enhanced sensitivity for the analysis of oligonucleotides using an on‐line coupled system of reversed‐phase high‐performance liquid chromatography (RP‐HPLC) and electrospray ionization ion‐trap mass spectrometry (ESI‐MS) has been developed. The presented method allows the use of the standard gradient elution of 0.1 M triethylammonium acetate (TEAA) buffer (adjusted to pH 7.0 with acetic acid) and acetonitrile that is typically used for the separation of oligonucleotides in RP‐HPLC. An added feature of this method is the ability to combine and mix additional 0.1 M imidazole in acetonitrile after the separation column for improved ESI‐MS performance. This is similar to the post‐column reaction method in liquid chromatography (LC) and the liquid sheath flow method in LC/ESI‐MS, both of which offer the advantage of not compromising the chromatographic separation conditions. The application of this new method is demonstrated to afford improved sensitivity for the analysis of oligonucleotides (20–50mer) via on‐line coupled HPLC/ESI‐MS analysis and purification systems. Copyright © 2002 John Wiley & Sons, Ltd.
Determination of melanotan‐II in rat plasma by liquid chromatography/tandem mass spectrometry: determination of pharmacokinetic parameters in rat following intravenous administration
Authors: Mock S. , Shen X. , Tamvakopoulos C.
Published: November 2002

A method is described for the determination of melanotan‐II (MT‐II), a synthetic cyclic heptapeptide analog of α‐melanocyte‐stimulating hormone (α‐MSH), in rat plasma by liquid chromatography/tandem mass spectrometry (LC/MS/MS). The analyte is recovered from plasma by solid‐phase extraction (SPE) and subsequently analyzed by LC/MS/MS. A SPE procedure using OASIS 96‐well plates is used for extraction of MT‐II from rat plasma. Using the described approach a limit of quantitation of 5 ng/mL was achieved in rat plasma. This level of sensitivity allowed the determination of pharmacokinetic parameters, following intravenous administration of MT‐II in rat. Copyright © 2002 John Wiley & Sons, Ltd.
PROPERTIES OF THE VERTEBRATE SKELETAL MUSCLE TUBULAR SYSTEM AS A SEALED COMPARTMENT
Authors: Launikonis Bradley S. , Stephenson D. George
Published: November 2002

Confocal imaging of impermeant fluorescent dyes trapped in the tubular (t‐) system of skeletal muscle fibres of rat and cane toad was used to examine changes in the morphology of the t‐system upon mechanical skinning, the time course of dye loss from the sealed t‐system in mechanically skinned fibres and the influence of rapid application and removal of glycerol on the morphology of the sealed t‐system. In contrast to intact fibres, which have a t‐system open to the outside, the sealed t‐system of toad mechanically skinned fibres consistently displayed local swellings (vesicles). The occurrence of vesicles in the sealed t‐system of rat‐skinned fibres was infrequent. Application and removal of 200–400mM glycerol to the sealed t‐system did not produce any obvious changes in its morphology. The dyes fluo‐3, fura‐2 and Oregon green 488 were lost from the sealed t‐system of toad fibres at different rates suggesting that the mechanism of organic anion transport across the tubular wall was not by indiscriminate bulk transport. The rate of fluo‐3 and fura‐2 loss from the sealed t‐system of rat fibres was greater in rat than in toad fibres and could be explained by differences in surface area: volume ratio of the t‐system in the two fibre types. Based on the results presented here and on other results from this laboratory, an explanation is given for the formation of numerous vesicles in toad‐skinned fibres and lack of vesicle formation in rat‐skinned fibres. This explanation can also help with better understanding the mechanism responsible for vacuole formation in intact fibres.
PARASITOPHOROUS VACUOLE: MORPHOFUNCTIONAL DIVERSITY IN DIFFERENT COCCIDIAN GENERA (A SHORT INSIGHT INTO THE PROBLEM)
Authors: Beyer Tamara V. , Svezhova Nadezhda V. , Radchenko Antonina I. , Sidorenko Nina V.
Published: November 2002

We present a short insight into the problem of parasitophorous vacuole (PV) formation as a most peculiar kind of cell vacuolization occurring in the course of intracellular development of coccidian pathogens of the genera Eimeria, Isospora, Toxoplasma, Sarcocystis, Cryptosporidium, Epieimeria, and Karyolysus. The review focuses on the morpho‐functional diversity of PVs in these parasites. By the present time, the PVs containing different parasite genera and species have been examined to different extent. The membrane of the PV (PVM) obviously derives from the host cell plasmalemma. But soon after parasite penetration, the morphofunctional organization and biochemical composition of the PVM drastically changes: its proteins are selectively excluded and those of the parasite are incorporated. As the result, the PV becomes not fusigenic for lysosomes or any other vacuoles or vesicles, because host cell surface markers necessary for membrane fusion are eliminated from the PVM during parasite invasion.

The pattern of the PVs is parasite specific and demonstrates a broad diversity within the same genera and species and even at different stages of the endogenous development. The PV is far from being an indifferent membrane vesicle containing the parasite. Instead, it represents a dynamic system that reflects the innermost events of host—parasite relationships, thus promoting the accomplishing of the parasite life cycle, which, in its turn, is a necessary prerequisite of the parasite eventual survival as a species.
VACUOLATION IN T‐TUBULES AS A MODEL FOR TUBULAR‐VESICULAR TRANSFORMATIONS IN BIOMEMBRANE SYSTEMS
Authors: Krolenko Sergei A. , Lucy Jack A.
Published: November 2002

This review outlines the basic properties of T‐tubules in skeletal muscle cells, and the factors that govern reversible vacuolation in T‐tubules under experimental conditions. Comparable membranous transformations, involving the plasma membrane or occurring intracellularly, in non‐muscle cells are then considered. Finally, the mechanisms of similar transformations in various model membrane systems are discussed. In view of the similarities between reversible vacuolation in the T‐system and membrane transformations occurring in a variety of non‐muscle cells, it is suggested that reversible vacuolation in T‐tubules may be regarded as a general model for tubular‐vesicular transformations in biomembranes.
Systemic tissue inhibitor of metalloproteinase‐1 gene delivery reduces neointimal hyperplasia in balloon‐injured rat carotid artery
Authors: Furman C , Luo Z , Walsh K , Duverger N , Copin C , Fruchart J.C , Rouis M
Published: November 2002

Metalloproteinases (MMP)‐2 and MMP‐9 play a role in smooth muscle cell (SMC) migration from the media to the intima following arterial injury. Intravenous administration of adenovirus encoding tissue inhibitor of metalloproteinase‐1 (TIMP‐1) into balloon‐injured rat arteries (3×1011 viral particles/rat; n=7) resulted in a transient expression of TIMP‐1 and a significant inhibition of neointima thickening within 16 days (∼40% vs. control; P=0.012). Three days after injury, the number of intimal SMCs was decreased by ∼98% in TIMP‐1‐treated rats. However, no alteration was seen in intimal SMC proliferation after 13 days of injury. Therefore, our results show that systemic gene transfer of TIMP‐1 is a promising approach in early restenosis treatment.
Characterization of a novel human UDP‐GalNAc transferase, pp‐GalNAc‐T101
Authors: Cheng Lamei , Tachibana Kouichi , Zhang Yan , Guo Jian-ming , Kahori Tachibana Kahori , Kameyama Akihiko , Wang Han , Hiruma Toru , Iwasaki Hiroko , Togayachi Akira , Kudo Takashi , Narimatsu Hisashi
Published: November 2002

A novel member of the human UDP‐N‐acetyl‐D‐galactosamine:polypeptide N‐acetylgalactosaminyltransferase (pp‐GalNAc‐T) gene family was cloned as a homolog of human pp‐GalNAc‐T7, and designated pp‐GalNAc‐T10. pp‐GalNAc‐T10 transcript was found in the small intestine, stomach, pancreas, ovary, thyroid gland and spleen. In a polypeptide GalNAc‐transfer assay, recombinant pp‐GalNAc‐T10 transferred GalNAc onto a panel of mucin‐derived peptide substrates. Furthermore, pp‐GalNAc‐T10 demonstrated strong transferase activity with glycopeptide substrates.
Amyloid formation of native folded protein induced by peptide‐based graft copolymer
Authors: Koga Tomoyuki , Taguchi Kazuhiro , Kogiso Masaki , Kobuke Yoshiaki , Kinoshita Takatoshi , Higuchi Masahiro
Published: November 2002

We report here that a native folded holo‐myoglobin, when incubated with a synthetic amyloidogenic peptide in aqueous solutions, forms fibrils. These fibrils took a cross‐β form (inter‐strand spacing: 4.65 Å and inter‐sheet spacing: 10.65 Å) and bound the amyloidophilic dye Congo red as did the authentic amyloid fibrils. In contrast such fibril formation of myoglobin did not occur in the absence of the peptide. These results suggest the possibility that inter‐molecular interaction of native protein with the amyloidogenic peptide trigger the amyloid formation even for the non‐pathogenic native protein like myoglobin, which itself exists as a globular form, under certain conditions.
Mechanisms for 2‐methoxyestradiol‐induced apoptosis of prostate cancer cells
Authors: Bu Shizhong , Blaukat Andree , Fu Xin , Heldin Nils-Erik , Landström Maréne
Published: November 2002

Prostate and breast carcinomas are sex hormone‐related carcinomas, which are known to be associated with an over‐expression of the proto‐oncogene Bcl‐2. Here, we report that 2‐methoxyestradiol (2‐ME), an endogenous metabolite of estrogen that does not bind to nuclear estrogen receptors, effectively induces apoptosis in Bcl‐2‐expressing human prostate and breast carcinoma cells in vitro and in a rat prostate tumor model in vivo. In several cell lines derived from prostate, breast, liver and colorectal carcinomas, 2‐ME treatment led to an activation of c‐Jun N‐terminal kinase (JNK) and phosphorylation of Bcl‐2, which preceded the induction of apoptosis. In summary, our data suggest that 2‐ME induces apoptosis in epithelial carcinomas by causing phosphorylation of JNK, which appears to be correlated with phosphorylation of Bcl‐2.
Apoptosis‐specific protein (ASP 45 kDa) is distinct from human Apg5, the homologue of the yeast autophagic gene apg5
Authors: Yung Hong Wa , Xue Luzheng , Tolkovsky Aviva M
Published: November 2002

We have examined whether the apoptosis‐specific protein p45ASP and human Apg5 are identical proteins. Like p45ASP, myc‐hApg5 cross‐reacted with a c‐Jun antibody and ∼50% of myc‐hApg5 was bound to a Triton X‐100‐insoluble fraction in HeLa cells. However, soluble myc‐hApg5 was degraded during apoptosis induced by staurosporine or TNFα/cycloheximide whilst expression of soluble p45ASP was stabilised. Furthermore, myc‐hApg5 degradation was blocked by the caspase inhibitor Boc‐Asp(OMe)FMK whilst p45ASP expression was eliminated. Moreover, myc‐hApg5 (∼32 kDa) never assumed the size of p45ASP (45 kDa). It is therefore likely that p45ASP and human Apg5 are distinct proteins although they do share some common characteristics.
Increase in cellular glutamate levels stimulates exocytosis in pancreatic β‐cells
Authors: Høy Marianne , Maechler Pierre , Efanov Alexander M , Wollheim Claes B , Berggren Per-Olof , Gromada Jesper
Published: November 2002

Glutamate has been implicated as an intracellular messenger in the regulation of insulin secretion in response to glucose. Here we demonstrate by measurements of cell capacitance in rat pancreatic β‐cells that glutamate (1 mM) enhanced Ca2+‐dependent exocytosis. Glutamate (1 mM) also stimulated insulin secretion from permeabilized rat β‐cells. The effect was dose‐dependent (half‐maximum at 5.1 mM) and maximal at 10 mM glutamate. Glutamate‐induced exocytosis was stronger in rat β‐cells and clonal INS‐1E cells compared to β‐cells isolated from mice and in parental INS‐1 cells, which correlated with the expressed levels of glutamate dehydrogenase. Glutamate‐induced exocytosis was inhibited by the protonophores FCCP and SF6847, by the vacuolar‐type H+‐ATPase inhibitor bafilomycin A1 and by the glutamate transport inhibitor Evans Blue. Our data provide evidence that exocytosis in β‐cells can be modulated by physiological increases in cellular glutamate levels. The results suggest that stimulation of exocytosis is associated with accumulation of glutamate in the secretory granules, a process that is dependent on the transgranular proton gradient.
Protein kinase inhibitors block the stimulation of the AMP‐activated protein kinase by 5‐amino‐4‐imidazolecarboxamide riboside
Authors: Fryer Lee G.D , Parbu-Patel Asha , Carling David
Published: November 2002

The AMP‐activated protein kinase (AMPK) is the central component of a protein kinase cascade that plays a major role in energy sensing. AMPK is activated pharmacologically by 5‐amino‐4‐imidazolecarboxamide (AICA) riboside monophosphate (ZMP), which mimics the effects of AMP on the AMPK cascade. Here we show that uptake of AICA riboside into cells, mediated by the adenosine transport system, is blocked by a number of protein kinase inhibitors. Under these conditions, ZMP does not accumulate to sufficient levels to stimulate AMPK. Our results demonstrate that careful interpretation is required when using AICA riboside in conjunction with protein kinase inhibitors to investigate the physiological role of AMPK.
NRSF/REST confers transcriptional repression of the GPR10 gene via a putative NRSE/RE‐1 located in the 5′ promoter region
Authors: Kemp Daniel M. , Lin Julia C. , Ubeda Mariano , Habener Joel F.
Published: November 2002

The G protein‐coupled receptor GPR10 is highly localized to areas of the brain. In an effort to reveal transcriptional determinants of this tissue specificity, we recognized a putative NRSE (neuron‐restrictive silencer element) located in the 5′ promoter region of the gene. The cognate NRSE binding protein NRSF (neuron‐restrictive silencer factor) restricts gene expression to mature neurons and endocrine cells by repressing their transcription in non‐neuronal/‐endocrine cells. In cell lines where NRSF‐mediated gene repression has been functionally established, the activity of the GPR10 promoter was repressed in a manner consistent with NRSE‐dependent regulation. A specific point mutation to confer non‐functionality of the NRSE revealed a 10‐fold de‐repression of reporter gene expression. In contrast, in the GPR10‐expressing cell line GH3, mRNA transcripts of NRSF were undetectable and suppression of promoter activity was not observed. However, transfection of a rat NRSF expression vector resulted in significant repression of transcription, which was reversed by mutation of the NRSE. In conclusion, we demonstrate that the GPR10 gene is specifically regulated by NRSF, and suggest this to be a contributory factor in the tissue‐specific distribution of GPR10 in vivo.
Calcium‐binding analysis and molecular modeling reveal echis coagulation factor IX/factor X‐binding protein has the Ca‐binding properties and Ca ion‐independent folding of other C‐type lectin‐like proteins
Authors: Atoda Hideko , Kaneko Hiroki , Mizuno Hiroshi , Morita Takashi
Published: November 2002

Many biologically active heterodimeric proteins of snake venom consist of two C‐type lectin‐like subunits. One of these proteins, habu IX/X‐bp, is a Gla domain‐binding protein whose subunits both bind to a Ca2+ ion, with a total of two Ca2+‐binding sites. The molecular modeling and Ca2+‐binding analysis of echis IX/X‐bp revealed that it lacks one of two Ca2+‐binding sites, though the folding of this subunit is conserved. It is concluded that heterodimeric C‐type lectin‐like proteins function independent of Ca2+ and have essentially a similar folding to habu IX/X‐bp.
Spectral imaging and linear un‐mixing enables improved FRET efficiency with a novel GFP2–YFP FRET pair
Authors: Zimmermann Timo , Rietdorf Jens , Girod Andreas , Georget Virginie , Pepperkok Rainer
Published: November 2002

Spectral variants of the green fluorescent protein (GFP) have been extensively used as reporters to image molecular interactions in living cells by fluorescence resonance energy transfer (FRET). However, those GFP variants which are the most efficient donor acceptor pairs for FRET measurements show a high degree of spectral overlap which has hampered in the past their use in FRET applications. Here we use spectral imaging and subsequent un‐mixing to quantitatively separate highly overlapping donor and acceptor emissions in FRET measurements. We demonstrate the method in fixed and living cells using a novel GFP based FRET pair (GFP2–YFP (yellow)), which has an increased FRET efficiency compared to the most commonly used FRET pair consisting of cyan fluorescent protein and YFP. Moreover, GFP2 has its excitation maximum at 396 nm at which the YFP acceptor is excited only below the detection level and thus this FRET pair is ideal for applications involving sensitized emission.
Gating of the expressed Cav3.1 calcium channel
Authors: Lacinová L’ , Klugbauer N , Hofmann F
Published: November 2002

Intramembrane charge movement originating from Cav3.1 (T‐type) channel expressed in HEK 293 cells was investigated. Ion current was blocked by 1 mM La3+. Charge movement was detectable for depolarizations above ∼−70 mV and saturated above +60 mV. The voltage dependence of charge movement followed a single Boltzmann function with half‐maximal activation voltage +12.9 mV and +12.3 mV and with slopes of 22.4 mV and 18.1 mV for the ON‐ and OFF‐charge movement, respectively. Inactivation of ICa by prolonged depolarization pulse did not immobilize intramembrane charge movement in the Cav3.1 channel.
Expression and regulation of peroxiredoxin 5 in human osteoarthritis
Authors: Wang Min-Xia , Wei Aiqun , Yuan Jun , Trickett Annette , Knoops Bernard , Murrell George A.C
Published: November 2002

Reactive oxygen species (ROS) are implicated in the pathogenesis of osteoarthritis (OA). However, little is known about the antioxidant defence system in articular cartilage. We investigated the expression and regulation of peroxiredoxin 5 (PRDX5), a newly discovered thioredoxin peroxidase, in human normal and osteoarthritic cartilage. Our results show that human cartilage constitutively expresses PRDX5. Moreover, the expression is up‐regulated in OA. Inflammatory cytokines tumour necrosis factor α and interleukin 1 β contribute to this up‐regulation by increasing intracellular ROS production. The present study suggests that PRDX5 may play a protective role against oxidative stress in human cartilage.
Maturation of the activities of recombinant mite allergens Der p 1 and Der f 1, and its implication in the blockade of proteolytic activity
Authors: Takai Toshiro , Mineki Reiko , Nakazawa Takuya , Takaoka Masatoshi , Yasueda Hiroshi , Murayama Kimie , Okumura Ko , Ogawa Hideoki
Published: November 2002

Recombinant pro‐Der p 1 expressed in yeast Pichia pastoris was convertible into the prosequence‐removed mature Der p 1 with full activities of cysteine protease and IgE‐binding with or without N‐glycosylation of the mature sequence as well as pro‐Der f 1. The active recombinant variants will be the basis for various future studies. The major N‐terminus of pro‐Der p 1 with low proteolytic activity was the putative signal‐cleavage site, while that of pro‐Der f 1 contained not only the equivalent site but also 21 residues downstream, and pro‐Der f 1 retained significant activity. Contribution of the N‐terminal region of the Der p 1 prosequence including an N‐glycosylation motif on effective inhibition of proteolytic activity of pro‐Der p 1 was suggested.
Transcription initiation sites and promoter structure of the human TRAIL‐R3 gene1
Authors: Ruiz de Almodóvar Carmen , López-Rivas Abelardo , Redondo Juan Miguel , Rodrı́guez Antonio
Published: November 2002

TRAIL‐R3 is a decoy receptor for TRAIL (tumor necrosis factor‐related apoptosis‐inducing ligand), a member of the tumor necrosis factor ligand family. In several cell types decoy receptors inhibit TRAIL‐induced apoptosis by binding TRAIL and preventing its binding to TRAIL pro‐apoptotic receptors. Here we report the cloning of the promoter region of human TRAIL‐R3 and the mapping of the transcriptional start sites. This gene contains a consensus TATA box and the minimal promoter lies within the first 33 nucleotides upstream of the transcription start site. Transient transfection assays of luciferase reporter plasmids demonstrate that human TRAIL‐R3 promoter can be induced in doxorubicin‐treated MCF‐7 cells in a p53‐independent manner.
Generation of a novel proteolysis resistant vascular endothelial growth factor165 variant by a site‐directed mutation at the plasmin sensitive cleavage site
Authors: Lauer Gereon , Sollberg Stephan , Cole Melanie , Krieg Thomas , Eming Sabine A
Published: November 2002

Vascular endothelial growth factor (VEGF) is a potent angiogenic mediator in tissue repair. In non‐healing human wounds plasmin cleaves and inactivates VEGF165. In the present study, we generated recombinant VEGF165 mutants resistant to plasmin proteolysis. Substitution of Arg110 with Ala110 or Gln110, and Ala111 with Pro111 yielded plasmin‐resistant and biologically active VEGF165 mutants. In addition, substitution of Ala111 with Pro111 resulted in a substantial degree of stabilization when incubated in wound fluid obtained from non‐healing wounds. These results suggest that the plasmin cleavage site Arg110/Ala111 and the carboxyl‐terminal domain play an important role in the mitogenic activity of VEGF165.
NMR structure of the unliganded Bombyx mori pheromone‐binding protein at physiological pH
Authors: Lee Donghan , Damberger Fred F , Peng Guihong , Horst Reto , Güntert Peter , Nikonova Larisa , Leal Walter S , Wüthrich Kurt
Published: November 2002

The nuclear magnetic resonance structure of the unliganded pheromone‐binding protein (PBP) from Bombyx mori at pH above 6.5, BmPBPB, consists of seven helices with residues 3–8, 16–22, 29–32, 46–59, 70–79, 84–100, and 107–124, and contains the three disulfide bridges 19–54, 50–108, and 97–117. This polypeptide fold encloses a large hydrophobic cavity, with a sufficient volume to accommodate the natural ligand bombykol. The polypeptide folds in free BmPBPB and in crystals of a BmPBP–bombykol complex are nearly identical, indicating that the B‐form of BmPBP in solution represents the active conformation for ligand binding.
Identification of the nuclear localization signal of p21cip1 and consequences of its mutation on cell proliferation
Authors: Rodrı́guez-Vilarrupla Aina , Dı́az Carmen , Canela Núria , Rahn Hans-Peter , Bachs Oriol , Agell Neus
Published: November 2002

Overexpression of p21cip1 induces cell cycle arrest. Although this ability has been correlated with its nuclear localization, the evidence is not conclusive. The mutants that were used to inhibit its nuclear translocation could no longer bind to several proteins known to interact with the last 25 amino acids of p21cip1. Here we used point mutation analysis and fusion of the proteins to DsRed to identify which amino acids are essential for the nuclear localization of p21cip1. We conclude that amino acids RKR140–142 are essential for nuclear translocation of p21cip1. While wild‐type DsRed‐p21 induces cell cycle arrest in 95% of transfected cells, overexpression of cytoplasmatic p21AAA140–142 arrested only 20% of transfected cells. We conclude that cytoplasmatic p21, with no deletion in the C‐terminal region, had a much lower capacity to arrest the cell cycle.
N‐Myristoylethanolamine–cholesterol (1:1) complex: first evidence from differential scanning calorimetry, fast‐atom‐bombardment mass spectrometry and computational modelling
Authors: Ramakrishnan M , Kenoth Roopa , Kamlekar Ravi Kanth , Chandra M.Sharath , Radhakrishnan T.P , Swamy Musti J
Published: November 2002

The interaction of N‐myristoylethanolamine (NMEA) with cholesterol is investigated by differential scanning calorimetry (DSC), fast‐atom‐bombardment mass spectrometry (FAB‐MS) and computational modelling. Addition of cholesterol to NMEA leads to a new phase transition at 55°C besides the chain‐melting transition of NMEA at 72.5°C. The enthalpy of the new transition increases with cholesterol content up to 50 mol%, but decreases thereafter, vanishing at 80 mol%. The enthalpy of the chain‐melting transition of NMEA decreases with an increase in cholesterol; the transition disappears at 50 mol%. FAB‐MS spectra of mixtures of NMEA and cholesterol provide clear signatures of the formation of {[NMEA+cholesterol]+} {[NMEA+cholesterol+Na]+}. These results are consistent with the formation of a 1:1 complex between NMEA and cholesterol. Molecular modelling studies support this experimental finding and provide a plausible structural model for the complex, which highlights multiple H‐bond interactions between the hydroxy group of cholesterol and the hydroxy and carbonyl groups of NMEA besides appreciable dispersion interaction between the hydrocarbon domains of the two molecules.
HspBP1, a homologue of the yeast Fes1 and Sls1 proteins, is an Hsc70 nucleotide exchange factor
Authors: Kabani Mehdi , McLellan Catherine , Raynes Deborah A , Guerriero Vince , Brodsky Jeffrey L
Published: November 2002

The yeast FES1 and SLS1 genes encode conserved nucleotide exchange factors that act on the cytoplasmic and endoplasmic reticulum luminal Hsp70s, Ssa1p and BiP, respectively. We report here that mammalian HspBP1 is homologous to Fes1p and that HspBP1 promotes nucleotide dissociation from both Ssa1p and mammalian Hsc70. In contrast, Fes1p inefficiently strips nucleotide from mammalian Hsc70, and unlike HspBP1 does not inhibit chaperone‐mediated protein refolding in vitro. Together, our data indicate that HspBP1 is a member of this new class of nucleotide exchange factors that exhibit varying degrees of compartment and species specificity.
Angiotensin‐I‐converting enzyme inhibitory peptides from tryptic hydrolysate of bovine αS2‐casein
Authors: Tauzin Jérôme , Miclo Laurent , Gaillard Jean-Luc
Published: November 2002

Angiotensin‐I‐converting enzyme (ACE) inhibitory activity of a tryptic digest of bovine αS2‐casein (αS2‐CN) was extensively investigated. Forty‐three peptide peaks were isolated and tested. Seven casokinins (i.e. CN‐derived ACE inhibitory peptides) were identified and their IC50 values were determined. Four peptides exhibited an IC50 value lower than 20 μM. Peptides αS2‐CN (f174–181) and αS2‐CN (f174–179) had IC50 values of 4 μM. Surprisingly, deletion of the C‐terminal dipeptide of two of these casokinins did not significantly alter their inhibitory activity.
Maximum principle for elliptic operators and applications
Authors: Tahraoui Rabah
Published: November 2002

Letfbe a signed function defined on some bounded domainΩ. We give sufficient conditions ensuring the positivity ofu, solution of the following equation:

- \Delta u = f \quad \text{in }\Omega , \qquad u\mid_{\partial \Omega } = 0.

Soit une fonctionfde signe non constant sur un domaine bornéΩ. On donne des conditions suffisantes assurant la stricte positivité de la solution de l’équation :

- \Delta u = f \quad \text{in }\Omega , \qquad u\mid_{\partial \Omega } = 0.
Nehari's problem and competing species systems
Authors: Conti M. , Terracini S. , Verzini G.
Published: November 2002

We consider an optimal partition problem in
N
-dimensional domains related to a method introduced by Nehari [22]. We prove existence of the minimal partition and some extremality conditions. Moreover we show some connections between the variational problem, the behaviour of competing species systems with large interaction and changing sign solutions to elliptic superlinear equations.

Nous étudions un problème de partition optimale dans

\mathbb{R}^{N}

qui se relie à une métode introduite par Nehari [22]. Nous établissons l’existence de la partition optimale et quelques conditions d’extremalité. En plus nous démontrons un lien entre le problème variationel, le comportement asymptotique de certains systèmes quand l’interaction tends à l’infini et les solutions qui changent de signe pour les equations elliptiques surlinéaires.
Local behavior and global existence of positive solutions of auλ⩽−Δu⩽uλ
Authors: Taliaferro Steven D.
Published: November 2002

We study the behavior near the origin of

C^2

positive solutions

u(x)

of

\tag{$\ast$}
au^{\lambda } \leq - \Delta u \leq u^{\lambda }

in a punctured neighborhood of the origin in

\mathbb{R}^{n}

(

n>2

) where the constants

λ

and

a

satisfy

\frac{n}{n - 2} < \lambda < \frac{n + 2}{n - 2}

and

0<a<1

. We also study the existence of

C^2

positive solutions of (

\ast

) in

\mathbb{R}^{n}

. In both cases we show that changing

a

from one value in the open interval

(0,1)

to another value in

(0,1)

can have a dramatic effect.

On étudie le comportement prés de l’ origine des solutions positives de classe

C^2

u(x)

de

\tag{$\ast$}
au^{\lambda } \leq - \Delta u \leq u^{\lambda }

dans un voisinage épointé de l’ origine dans

\mathbb{R}^{n}

(

n>2

) où les constantes

λ

et

a*

satisfont

\frac{n}{n - 2} < \lambda < \frac{n + 2}{n - 2}

et

0<a<1

. On étudie aussi l’ existence de solutions positives de classe

C^2

de (

\ast

) dans

\mathbb{R}^{n}

. Dans les deux cas nous montrons que changer la valeur de

a

dans l’intervalle ouvert

(0,1)

peut avoir un effet dramatique.
Efficacy of zolpidem in insomnia
Authors: Priest RG , Terzano MG , Parrino L , Boyer P
Published: November 2002

The efficacy of zolpidem, a non benzodiazepine hypnotic agent with a short elimination half life, was reviewed, analysing more than 50 international clinical trials published since 1986. The hypnotic activity of zolpidem has been explored in different patient populations including normal volunteers, general practice outpatients and psychiatric out- or in-patients with varying sleep disorders; both transient and chronic. Assessment methods used have included objective and subjective measures of hypnotic efficacy for different treatment durations, with results confirming that 10 mg is superior to placebo. Zolpidem was shown to be superior in most trials on sleep parameters such as total sleep time, sleep onset latency and nocturnal awakenings, but total REM sleep and REM latency were usually unmodified. Zolpidem maintained normal sleep physiology as demonstrated by the preservation of slow wave stages and no, or minimal, effects on sleep architecture after abrupt discontinuation. Consequently, 10 mg is the recommended dose for the short-term treatment of insomnia in the non-elderly; in elderly patients 5 mg has been shown to be effective at inducing sleep whilst giving an optimum safety profile.
General safety profile of zolpidem: safety in elderly, overdose and rebound effects
Authors: Allain H , Monti J
Published: November 2002

The different aspects of general safety of zolpidem are reviewed after analysing the safety data obtained in adults as well as in elderly patients. The low incidence of reported adverse events during the first three years of utilisation of zolpidem have corroborated the results of the clinical studies. The most frequent adverse effects are central nervous system (CNS) related but they can be largely prevented by respecting the recommended doses and prescription rules. In cases of acute overdoses, no severe complications have been attributed to zolpidem taken alone. In most of the studies carried out within the recommended treatment duration, abrupt treatment discontinuation was not associated with clinically significant rebound insomnia, and the available pre-clinical and clinical data indicate that the risk of abuse or dependence is minimal when prescribed according to the recommended doses of 10 mg in adults and 5 mg in elderly. Thus, with now well-established arguments on a large patient basis going in the same direction, the safety of zolpidem can authorise and justify its wide prescription in the short-term treatment of insomnia.
Detecting anxiety and depression in hospitalised elderly patients using a brief inventory
Authors: Huber P. , Mulligan R. , Mackinnon A. , Nebuloni-French T. , Michel J.P.
Published: November 2002

This study validated a French language version of an inventory designed to detect symptoms of depression and anxiety [Goldberg et al, 1987] in a sample of elderly French-speaking inpatients at risk for one of these disorders. Latent trait analysis was used to replicate the structure of the symptoms in the inventory, and receiver operating characteristic analysis was used to assess the performance of the inventory as a screening measure for Major Depressive Episode and Generalised Anxiety Disorder according to DSM-IV criteria. Reflecting the ascertainment of individuals in the sample as being at risk for a disorder, prevalence of individual symptoms was high although the general structure of the inventory was found to be comparable to that found in samples of both community elderly and younger medical patients. ROC analyses showed that the subscales of inventory performed satisfactorily as screening measures for anxiety or depression but lacked specificity for each disorder. In addition to providing further evidence for the utility of this inventory to detect general psychiatric distress in elderly persons, this study provides a valid means of detecting symptoms of depression and anxiety in French speaking groups.
Biphasic vasomotor reflex responses of the hand skin following intradermal injection of melittin into the forearm skin
Authors: Koyama Natsu , Hirata Kazuhiko , Hori Koichiro , Dan Kenjiro , Yokota Toshikatsu
Published: November 2002

Melittin is the main toxin of honeybee venom. Previously, we have reported that intradermal injection of melittin into the volar aspect of forearm in humans produces a temporary pain and a subsequent sustained increase in the skin temperature due to axon reflex. To clarify the interaction between nociceptive inputs and vascular changes, we studied the influence of noxious stimulation by intradermal melittin on the vasomotor control of the distal extremities in human volunteers. Temperature changes of the bilateral palmar surface were recorded by means of a computer‐assisted infrared thermography. Unexpectedly, we found a biphasic response of skin temperature. The skin temperature of both fingers and hands decreased immediately after the melittin injection and then increased well above the control level, prior to the injection. There was a considerable individual variation in the baseline skin temperature, prior to melittin. The skin temperature in a finger/hand with lower preinjection value increased more markedly in the second phase. Consequently, the individual variation in the peak temperature of the second phase was less pronounced. The initial decrease was interpreted as sympathetic vasoconstrictor reflex induced by noxious stimulation and the later increase as release of sympathetic vasomotor tone.
Poststroke shoulder pain: a prospective study of the association and risk factors in 152 patients from a consecutive cohort of 205 patients presenting with stroke
Authors: Gamble Giles E , Barberan Elisa , Laasch Hans‐Ulrich , Bowsher David , Tyrrell Pippa J , Jones Anthony K.P
Published: November 2002

BACKGROUND AND PURPOSE Shoulder pain is known to retard rehabilitation after stroke. Its causes and prognosis are uncertain. This study describes the incidence of poststroke shoulder pain prospectively, in an unselected stroke population in the first 6 months after stroke and identifies risk factors for developing pain.

METHODS 297 patients with possible stroke were screened and stroke diagnosed in 205 cases. The 152 patients entered the study of which 123 patients were assessed up to 6 months. This cohort, with a mean age of 70.6 years, was examined at 2 weeks, 2, 4, and 6 months. A history of shoulder pain, Barthel score, anxiety and depression score were recorded. Full neurological and rheumatological examination was undertaken, using the contralateral side as a control. Pain outcome and stroke outcome was recorded at subsequent visits.

RESULTS 52 (40%) patients developed shoulder pain on the same side of their stroke. There was a strong association between pain and abnormal shoulder joint examination, ipsilateral sensory abnormalities and arm weakness. Shoulder pain had resolved or improved at 6 months in 41 (80%) of the patients with standard current treatment.

CONCLUSIONS Shoulder pain after stroke occurred in 40% of 123 patients surviving, consenting and not too unwell to participate. This included 52 patients of an original cohort of 205 patients presenting with stroke. Eighty percent of patients made a good recovery with standard treatment Patients with sensory and or motor deficits represent at risk sub‐groups.