bluuebunny/crossref_metadata_2025 huggingface dataset:
Post‐translational phosphorylation affects the IgE binding capacity of caseins
Authors: Bernard H. , Meisel H. , Creminon C. , Wal J.M.
Published: July 2002

IgE response specific to those molecular regions of casein that contain a major phosphorylation site was analyzed using native and modified caseins and derived peptides. This study included (i) the naturally occurring common variants A1 and A from β‐ and αs2‐caseins, respectively, which were purified in the native form and then dephosphorylated, (ii) a purified rare variant D of αs2‐casein which lacks one major phosphorylation site, and (iii) the native and dephosphorylated tryptic fragment f(1–25) from β‐casein. Direct and indirect ELISA using sera from patients allergic to milk showed that the IgE response to caseins is affected by modifying or eliminating the major phosphorylation site.
Inhibition of plant telomerase by telomere‐binding proteins from nuclei of telomerase‐negative tissues
Authors: Fulnečková Jana , Fajkus Jiřı́
Published: July 2002

The activity of telomerase in plant cells is precisely regulated in response to changes in cell division rate. To explore this regulatory mechanism, the effect on telomerase activity of protein extracts from nuclei of telomerase‐negative tissues was examined. An inhibition of telomerase activity was found which was species‐non‐specific. This inhibition was due to proteins which form salt‐stable, sequence‐specific complexes with the G‐rich telomeric strand and reduce its accessibility, as shown by gel retardation and by terminal transferase (TdT) extension of G‐rich telomeric and non‐telomeric (substrate) primers. A 40 kDa polypeptide was detected by SDS–PAGE after cross‐linking the complex formed by extracts from tobacco leaf nuclei. Such proteins may be involved in regulation of telomerase activity in plants.
Subcellular differences in post‐translational modification of barley 14‐3‐3 proteins
Authors: van Zeijl Mieke J. , Testerink Christa , Kijne Jan W. , Wang Mei
Published: July 2002

Expression and post‐translational modification of barley 14‐3‐3 isoforms, 14‐3‐3A, 14‐3‐3B and 14‐3‐3C, were investigated using isoform‐specific antibodies. Although all three isoforms were shown to be present in the cytosolic, the nuclear and the microsomal cell fractions, differences in post‐translational modification were identified for the different cell fractions. Germination‐related modifications of 14‐3‐3 proteins were observed in the cytosol and the microsomal fraction, but not in the nucleus. In vitro proteolytic cleavage of 14‐3‐3 proteins using trypsin suggests that for 14‐3‐3A this change was caused by proteolytic cleavage of the unconserved C‐terminal region.
The influence of glycerol and chloroplast lipids on the spectral shifts of pigments associated with NADPH:protochlorophyllide oxidoreductase from Avena sativa L
Authors: Klement Harald , Oster Ulrike , Rüdiger Wolfhart
Published: July 2002

Dark‐grown angiosperm seedlings lack chlorophylls, but accumulate protochlorophyllide a complexed with the light‐dependent enzyme NADPH:protochlorophyllide oxidoreductase. Previous investigators correlated spectral heterogeneity of in vivo protochlorophyllide forms and a shift of chlorophyllide forms from 680 to 672 nm (Shibata shift) occurring after irradiation, with intact membrane structures which are destroyed by solubilization. We demonstrate here that the various protochlorophyllide forms and the Shibata shift which disappear upon solubilization are restored if the reconstituted complex is treated with plastid lipids and 80% (w/v) glycerol. We hypothesize that the lipids can form a cubic phase and that this is the precondition in vitro and in vivo for the observed spectral properties before and after irradiation.
Involvement of apoptosis‐inducing factor during dolichyl monophosphate‐induced apoptosis in U937 cells
Authors: Yasugi Etsuko , Kumagai Tsukasa , Nishikawa Yoshihisa , Okuma Emiko , Saeki Kumiko , Oshima Mieko , Susin Santos A. , Kroemer Guido , Yuo Akira
Published: July 2002

Dolichyl monophosphate (Dol‐P) has been found to induce apoptosis in human leukemia U937 cells. During this apoptotic execution, the increase of plasma membrane fluidity (5–20 min), caspase‐3‐like protease activation (2–4 h), chromatin condensation and DNA ladder formation (3–4 h) were observed successively. Here, we report that reduction in mitochondrial transmembrane potential and translocation of apoptosis‐inducing factor (AIF) are early events (1–3 h) in the apoptotic process induced by Dol‐P in U937 cells. The AIF was concentrated around nuclei and partly translocated to the nuclei, which was confirmed by immunocytochemistry using specific anti‐AIF antibody. Both caspase‐8 and caspase‐3 inhibitors blocked only DNA fragmentation but not mitochondrial processes, AIF migration and chromatin condensation. These results indicate that mitochondrial changes are an early step in the apoptosis induced by Dol‐P and AIF is one of the important factors which induce chromatin condensation in nuclei.
Bi‐substrate analogue ligands for affinity chromatography of protein kinases
Authors: Loog Mart , Uri Asko , Järv Jaak , Ek Pia
Published: July 2002

Novel affinity ligands, consisting of ATP‐resembling part coupled with specificity determining peptide fragment, were proposed for purification of protein kinases. Following this approach affinity sorbents based on two closely similar ligands AdoC–Aoc–Arg4–Lys and AdoC–Aoc–Arg4–NH(CH2)6NH2, where AdoC stands for adenosine‐5′‐carboxylic acid and Aoc for amino‐octanoic acid, were synthesized and tested for purification of recombinant protein kinase A catalytic subunit directly from crude cell extract. Elution of the enzyme with MgATP as well as L‐arginine yielded homogeneous protein kinase A preparation in a single purification step. Also protein kinase A from pig heart homogenate was selectively isolated using MgATP as eluting agent. Protein kinase with acidic specificity determinant (CK2) as well as other proteins possessing nucleotide binding site (L‐type pyruvate kinase) or sites for wide variety of different ligands (bovine serum albumin) did not bind to the column, pointing to high selectivity of the bi‐functional binding mode of the affinity ligand.
Nuclear localization of a hypoxia‐inducible novel non‐symbiotic hemoglobin in cultured alfalfa cells1
Authors: Seregélyes Csaba , Mustárdy László , Ayaydin Ferhan , Sass László , Kovács László , Endre Gabriella , Lukács Noémi , Kovács Izabella , Vass Imre , Kiss György B. , Horváth Gábor V. , Dudits Dénes
Published: July 2002

We have isolated a 483‐bp‐long full‐length cDNA clone encoding a non‐symbiotic hemoglobin called Mhb1, the first one found in alfalfa. This non‐symbiotic hemoglobin is a single copy gene localized in linkage group 4 in diploid Medicago genome. The Mhb1 mRNA was found only in the roots of alfalfa plants. The Mhb1 gene was inducible by hypoxia and showed no induction by cold stress treatment. The Mhb1 transcript level increased at the G2/M boundary in a synchronized alfalfa cell suspension culture. The majority of Mhb1 protein was shown to be localized in the nucleus and smaller amounts were detected in the cytoplasm. A potential link to the nitric oxide signalling pathway is also discussed.
Antioxidative galloyl esters as enzyme inhibitors of p‐hydroxybenzoate hydroxylase
Authors: Abe Ikuro , Kashiwagi Kenji , Noguchi Hiroshi
Published: July 2002

Gallic acid and its esters were evaluated as enzyme inhibitors of recombinant p‐hydroxybenzoate hydroxylase (PHBH), a NADPH‐dependent flavin monooxygenase from Pseudomonas aeruginosa. n‐Dodecyl gallate (DG) (IC50=16 μM) and (−)‐epigallocatechin‐3‐O‐gallate (EGCG) (IC50=16 μM), a major component of green tea polyphenols, showed the most potent inhibition, while product‐like gallic acid did not inhibit the enzyme significantly (IC50>250 μM). Inhibition kinetics revealed that both DG and EGCG inhibited PHBH in a non‐competitive manner (K
I=18.1 and 14.0 μM, respectively). The enzyme inhibition was caused by specific binding of the antioxidative gallate to the enzyme, and by scavenging reactive oxygen species required for the monooxygenase reaction. Molecular modeling predicted that EGCG binds to the enzyme in the proximity of the FAD binding site via formation of three hydrogen bonds.
Malignant transformation of B lymphoma cell line BJAB by Epstein–Barr virus‐encoded small RNAs
Authors: Yamamoto Norio , Takizawa Takenori , Iwanaga Yoichi , Shimizu Norio , Yamamoto Naoki
Published: July 2002

EBV‐encoded small RNAs (EBERs) are non‐polyadenylated and abundantly transcribed RNAs, whose functions have not yet been fully elucidated. Here, we report that the EBV‐negative B lymphoma cell line, BJAB, was rendered more malignant and resistant to apoptosis by EBERs. EBER‐transfected cells exhibited enhanced growth potential in SCID mice as well as in soft agar, and showed resistance to apoptotic stimuli in comparison with the vector control. EBERs inhibited the activity of the double‐stranded RNA‐dependent protein kinase, PKR, which is reputed to act as a tumor‐suppressor. These results suggest that EBERs play an important role in the pathogenesis of EBV‐associated malignancies through the inhibition of PKR.
Antibodies to the CFTR modulate the turgor pressure of guard cell protoplasts via slow anion channels
Authors: Leonhardt Nathalie , Bazin Ingrid , Richaud Pierre , Marin Elena , Vavasseur Alain , Forestier Cyrille
Published: July 2002

The plasma membrane guard cell slow anion channel is a key element at the basis of water loss control in plants allowing prolonged osmolite efflux necessary for stomatal closure. This channel has been extensively studied by electrophysiological approaches but its molecular identification is still lacking. Recently, we described that this channel was sharing some similarities with the mammalian ATP‐binding cassette protein, cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel [Leonhardt, N. et al. (1999) Plant Cell 11, 1141–1151]. Here, using the patch‐clamp technique and a bioassay, consisting in the observation of the change in guard cell protoplasts volume, we demonstrated that a functional antibody raised against the mammalian CFTR prevented ABA‐induced guard cell protoplasts shrinking and partially inhibited the slow anion current. Moreover, this antibody immunoprecipitated a polypeptide from guard cell protein extracts and immunolabeled stomata in Vicia faba leaf sections. These results indicate that the guard cell slow anion channel is, or is closely controlled by a polypeptide, exhibiting one epitope shared with the mammalian CFTR.
Binding of a peptide from a Streptococcus dysgalactiae MSCRAMM to the N‐terminal F1 module pair of human fibronectin involves both modules
Authors: Schwarz-Linek Ulrich , Plevin Michael J. , Pickford Andrew R. , Höök Magnus , Campbell Iain D. , Potts Jennifer R.
Published: July 2002

Host invasion by a number of pathogenic bacteria such as staphylococci and streptococci involves binding to fibronectin, a ubiquitous extracellular matrix protein. On the bacterial side, host extracellular matrix adherence is mediated by MSCRAMMs (microbial surface components recognizing adhesive matrix molecules) which, in some cases, have been identified to be important virulence factors. In this study we used nuclear magnetic resonance spectroscopy to characterize the interaction of B3, a synthetic peptide derived from an adhesin of Streptococcus dysgalactiae, with the N‐terminal module pair 1F12F1 of human fibronectin. 1F12F1 chemical shift changes occurring on formation of the 1F12F1/B3 complex indicate that both modules bind to the peptide and that a similar region of each module is involved. A similar surface of the 4F15F1 module pair had previously been identified as the binding site for a fibronectin‐binding peptide from Staphylococcus aureus.
Oligosaccharyltransferase is highly specific for the hydroxy amino acid in Asn‐Xaa‐Thr/Ser
Authors: Breuer Wilhelm , Klein Roger A , Hardt Birgit , Bartoschek Achim , Bause Ernst
Published: July 2002

Pig liver oligosaccharyltransferase (OST), which is involved in the en bloc transfer of the Dol‐PP‐linked GlcNAc2‐Man9‐Glc3 precursor on to asparagine residues in the Asn‐Xaa‐Thr/Ser sequence, is highly stereospecific for the conformation of the 3‐carbon atom in the hydroxy amino acid. Moreover, substitution of the hydroxy group by either SH as in cysteine, or NH2 as in β,γ‐diamino‐butanoic acid as reported previously [Bause, E. et al., Biochem. J. 312 (1995) 979–985], followed by the determination of the pH optimum for enzymatic activity, indicates that neither a negative nor a positive charge in the hydroxy amino acid position is tolerated by the enzyme. Binding of the threonine β‐methyl group by OST is also specific, with serine, L‐threo‐β‐hydroxynorvaline and L‐β‐hydroxynorleucine containing tripeptides all bound much less efficiently than the threonine peptide itself. The data are interpreted in terms of a highly stereospecific hydrophobic binding pocket for the threonine CH3‐CH(OH) group.
Direct evidence that lactogenic hormones induce homodimerization of membrane‐anchored prolactin receptor in intact Nb2‐11C rat lymphoma cells
Authors: Sakal Edna , Elberg Gerard , Gertler Arieh
Published: July 2002

The ability of full‐size prolactin receptor (PRLR) from Nb2 rat lymphoma cell line to undergo lactogenic hormone‐induced dimerization in intact cells or in a partially purified microsomal fraction was tested. The stoichiometry of ovine placental lactogen (oPL) binding to PRLR was documented by SDS‐PAGE of the covalently cross‐linked complexes between [125I]oPL and intact Nb2‐11C cells. The molecular masses of the specific bands were 82 and 141 kDa, corresponding to PRLR:oPL and (PRLR)2:oPL complexes. These results provide direct evidence for the occurrence of hormone‐induced receptor dimerization in intact cells. Gel‐filtration studies revealed that under non‐denaturing conditions, the purified receptor forms high‐molecular‐mass aggregates (190 and 540 kDa) composed of receptor dimers and oligomers. Since this aggregation was not dependent on the presence of lactogenic hormone, it is possible that the receptor in the intact cells may already exist as a non‐covalent dimer or oligomer and that hormone‐induced dimerization stabilizes the complex or changes its conformation.
Stimulation of respiration by methylene blue in rat liver mitochondria
Authors: Visarius Theresa M , Stucki Jörg W , Lauterburg Bernhard H
Published: July 2002

The effect of methylene blue on isolated rat liver mitochondria in the presence and absence of chloroacetaldehyde was investigated. Fatty acid oxidation was inhibited by chloroacetaldehyde and subsequently stimulated by methylene blue. Assessment of tightly coupled mitochondria revealed decreasing respiratory control ratios induced by increasing concentrations of methylene blue and methylene blue provoked mitochondrial swelling. In uncoupled mitochondria, methylene blue promoted a concentration‐dependent stimulation of respiration. These findings provide evidence that methylene blue, the redox dye currently used as an antidote for encephalopathy associated with alkylating chemotherapy, uncouples oxidative phosphorylation and acts as an electron transfer mediator to stimulate mitochondrial respiration.
Characterization of membrane‐associated prothrombin activator in normal and injured murine tissues
Authors: Shikamoto Yasuo , Shibusawa Satoshi , Okuyama Izumi , Morita Takashi
Published: July 2002

Thrombin is a multifunctional enzyme involved in coagulation, cell modulation and inflammation. We recently reported a novel membrane‐associated prothrombin activator, abbreviated as MAPA, found in cultured fibroblasts and glial cell lines. In this study, we examined the physiological role of this enzyme. MAPA‐like activity was detected in the liver, kidney, lung and heart but not in the spleen or brain in normal mice. To examine whether MAPA participates in biological reactions, hepatic and renal injury were induced by administration of CCl4 and HgCl2, respectively. MAPA‐like activity was specifically increased in the injured tissues: the activity was elevated by about 100‐fold in 48 h in the liver and increased by about 5‐fold in 12 h in the kidney. Their enzymatic properties were the same as those of MAPA in 8C feline kidney fibroblast cells. Phospholipids are required for activation of prothrombin by MAPA obtained from both 8C cells and tissues. These results suggest that MAPA activates prothrombin on the cell surface in injured tissue and participates in inflammation and regeneration associated with tissue injury.
Mutants affected in the putative diacylglycerol binding site of yeast protein kinase C
Authors: Jacoby Jörg J , Schmitz Hans-Peter , Heinisch Jürgen J
Published: July 2002

In an attempt to study the functional similarities between protein kinase C from the yeast Saccharomyces cerevisiae and its human homologues we have started in vitro mutagenesis to alter specific domains. Here we report on the exchange of four cysteine residues by serines in yeast Pkc1p that have been shown to be essential for diacylglycerol (DAG) binding and activation by this compound in humans. The mutant yeast protein leads to sensitivity to caffeine and low concentrations of SDS when expressed in a pkc1 deletion strain. However, sensitivity to staurosporine was not affected. Our data indicate that the conserved DAG binding domain serves an important function in yeast Pkc1p.
Bicarbonate protects the donor side of photosystem II against photoinhibition and thermoinactivation
Authors: Klimov V.V , Baranov S.V , Allakhverdiev S.I
Published: July 2002

The rate of photoinhibition of photosystem II (PSII) activities (photoinduced change of chlorophyll fluorescence yield, ΔF, and photoreduction of 2,6‐dichlorophenol‐indophenol) in O2‐evolving pea subchloroplast membrane fragments in medium depleted of CO2 was considerably decreased upon addition of 5 mM NaHCO3 before the light treatment. A similar effect was observed when 100 μM MnCl2 was added before the photoinhibition. In PSII membrane fragments depleted of Mn, the pre‐illumination led to irreversible loss of the capability of PSII to be reactivated by Mn2+, and the rate of the photoinhibition was decreased by a factor of 2 or 5 if the pre‐illumination was done in the presence of 0.2 μM MnCl2 (≈4 Mn per PSII reaction center) added alone or in combination with 5 mM NaHCO3, respectively. A similar protective effect of bicarbonate was also revealed in the dark, during thermoinactivation of O2‐evolving PSII at 40°C: the rate of thermoinactivation of ΔF was decreased by a factor of 3 if 5 mM NaHCO3 was added to the medium. The results are consistent with the idea that bicarbonate is an essential component of the Mn‐containing water‐oxidizing complex of PSII, which decreases its susceptibility to photoinhibition and thermoinactivation.
Telomerase from Saccharomyces cerevisiae contains several protein subunits and may have different activities depending on the protein content
Authors: Petrov Andrey V , Dokudovskaya Svetlana S , Sokolov Kirill A , Lavrik Olga I , Favre Alain , Dontsova Olga A , Bogdanov Alexey A
Published: July 2002

Telomerase is a ribonucleoprotein responsible for maintaining telomeres during the cell cycle [1, 2]. Here we describe a two‐step purification procedure for the Saccharomyces cerevisiae telomerase complex. We have found that the properties (processivity, nuclease activity) of telomerase depend on the isolation procedure. Using a cross‐linking approach, we have revealed several proteins that could be components of the telomerase complex. Furthermore, spectra of cross‐linked proteins differ in processive and non‐processive telomerase complexes.
Factors limiting display of foreign peptides on the major coat protein of filamentous bacteriophage capsids and a potential role for leader peptidase
Authors: Malik Pratap , Terry Tamsin D. , Bellintani Francesca , Perham Richard N.
Published: July 2002

Many small peptides can be displayed on every copy of the major coat protein in recombinant filamentous bacteriophages but larger peptides can only be accommodated in hybrid virions mixed with wild‐type protein subunits. A peptide insert of 12 residues capable of display at high copy number in a hybrid virion was found to be incapable of supporting recombinant virion assembly, a defect that could not be overcome by over‐expressing leader peptidase in the same Escherichia coli cell. In contrast, over‐expressing leader peptidase did increase the copy number of two 9‐residue peptides that were poorly incorporated into hybrid virions. The factors that limit peptide display are varied and not restricted to the early stages of viral assembly.
Chlorina and viridis mutants of barley (Hordeum vulgare L.) allow assignment of long‐wavelength chlorophyll forms to individual Lhca proteins of photosystem I in vivo
Authors: Knoetzel Jürgen , Bossmann Björn , Grimme L.Horst
Published: July 2002

The isolated subcomplex LHCI‐730 of plant photosystem I (PSI) chlorophyll (Chl) a/b binding antenna is a heterodimer of Lhca1 and Lhca4 and has a 77 K fluorescence emission peak at 730 nm (F730). Recently, three Chl spectral forms with 77 K fluorescence emission peaks at 720 nm, 730 nm and 742 nm were identified in native PSI. In an attempt to assign the two longest wavelength emission maxima to peripheral PSI antenna proteins, we performed 77 K fluorescence emission spectroscopy on intact leaves of chlorina and viridis mutants from barley which lack individual LHCI‐730 proteins. This approach indicates that F732 is found only in Lhca1 and F742 only in Lhca4, when these proteins are associated with the PSI reaction centre.
Carcinoembryonic antigen‐related cell‐cell adhesion molecule C‐CAM is greatly increased in serum and urine of rats with liver diseases
Authors: Lucka Lothar , Sel Serdar , Danker Kerstin , Horstkorte Rüdiger , Reutter Werner
Published: July 2002

C‐CAM (rat cell CAM/human CD66a) is ubiquitous and multifunctional. It is involved in intercellular adhesion, signal transduction and cell growth inhibition. Structurally, it is related to the carcinoembryonic antigen. In the present study serum, bile and urine of rats with liver diseases were analyzed for the presence of cell CAM. After bile duct ligation and during galactosamine (GalN) hepatitis we found that large amounts of liver membrane‐bound C‐CAM are secreted or shed into blood. The serum level of another liver membrane‐bound protein, LI‐cadherin, is not increased. It was shown that C‐CAM is also present in bile fluid, and for the first time that C‐CAM is present in the urine of rats with liver diseases. A particularly high concentration was measured in the urine of rats suffering from GalN hepatitis.
Application of fluorescence polarization to the steady‐state enzyme kinetic analysis of calpain II
Authors: Sem Daniel S. , McNeeley Patricia A.
Published: July 2002

This paper presents the application of fluorescence polarization to the determination of dissociation constants for competitive inhibitors that bind to enzymes. This steady‐state enzyme kinetic study measures the inhibition of the conversion of a fluorescently tagged substrate to a lower molecular weight fluorescent product by calpain II. It relies on the measurement of a parameter proportional to velocity, which is sufficient for this type of analysis. The strengths and limitations of the method are discussed. Inhibition constants for filamin and spectrin determined by this method are 125 nM and 13 nM respectively.
Binding properties of the human homeodomain protein OTX2 to a DNA target sequence
Authors: Briata Paola , Ilengo Cristina , Bobola Nicoletta , Corte Giorgio
Published: July 2002

OTX2, a homeodomain protein essential in mouse for the development of structures anterior to rhombomere 3, binds with high affinity to a DNA element (called OTS) present in the human tenascin‐C promoter. Here we investigate the binding properties of the full length recombinant human OTX2 and of several deletion mutants to the OTS element. We demonstrate that, upon binding of the protein to its DNA target site, a second molecule of OTX2 is recruited to the complex and that a nearby second binding site is not necessary for this interaction. OTX2 sequences located within a region carboxyl‐terminal to the homeodomain are necessary in addition to the homeodomain for binding to DNA. Furthermore, OTX2 dimerization requires the same protein domains necessary for DNA binding.
Nitric oxide mediates brain mitochondrial maturation immediately after birth
Authors: Almeida Angeles , Bolaños Juan P , Medina José M
Published: July 2002

The possible role of nitric oxide (NO) in brain mitochondrial maturation was studied. Within the first 5 min after birth, a sharp increase in ATP concentrations was observed, coinciding with an increase in mitochondrial complex II‐III (succinate‐cytochrome c reductase) activity, while complex I (NADH‐CoQ1 reductase) and complex IV (cytochrome c oxidase) activities remained unchanged. Under the same circumstances, cGMP concentrations were increased by 5 min after birth, correlating significantly with ATP concentrations. Since ATP concentrations also correlated significantly with mitochondrial complex II‐III activity, these three parameters may be associated. Inhibition of NO synthase activity brought about by the administration of N
ω‐nitro‐L‐arginine monomethyl ester to mothers prevented the postnatal increase in cGMP and ATP levels and complex II‐III activity. These results suggest that early postnatal mitochondrial maturation in the brain is a NO‐mediated process.
Serotonin 5‐HT1B and 5‐HT1D receptors form homodimers when expressed alone and heterodimers when co‐expressed
Authors: Xie Zhidong , Lee Samuel P. , O'Dowd Brian F. , George Susan R.
Published: July 2002

The serotonin (5‐hydroxytryptamine (5‐HT)) 1B and 1D receptor subtypes share a high amino acid sequence identity and have similar ligand binding properties. In this study, we demonstrate that both receptor subtypes exist as monomers and homodimers when expressed alone and as monomers and heterodimers when co‐expressed. Gene expression studies have shown that there are brain regions where the 5‐HT1B and 5‐HT1D receptors are co‐localized and where heterodimerization may occur physiologically. This is the first direct visualization of the physical association between G protein‐coupled receptors of different subtypes.
Urinary thymine dimers and 8‐oxo‐2′‐deoxyguanosine in psoriasis
Authors: Ahmad Jabeen , Cooke Marcus S. , Hussieni Amina , Evans Mark D. , Patel Kayuri , Burd Robert M. , Bleiker Tanya O. , Hutchinson Peter E. , Lunec Joseph
Published: July 2002

Psoralen in conjunction with UVA (PUVA) is perhaps the most effective treatment for psoriasis. It is, however, a risk factor for skin cancer in these patients and there is a need to develop non‐invasive assays reflective of treatment‐induced DNA damage. We report here the assessment of two important lesions, thymine dimer (T〈〉T) and 8‐oxo‐2′‐deoxyguanosine (8‐OHdG), in the urine of psoriasis patients. It was found that, once corrected for urine concentration, the psoriatic group had significantly higher (P<0.0001) urinary levels of thymine dimers compared to the control group. No significant differences in urinary 8‐OHdG levels were noted between the psoriatic, atopic dermatitis and control groups. Therefore biomonitoring of therapy from the very start with this simple and non‐invasive assay could perhaps be an effective measure of the risk involved with the treatment allowing optimization for minimal‐risk therapy.
Synonymous codon substitutions affect ribosome traffic and protein folding during in vitro translation
Authors: Komar Anton A , Lesnik Thierry , Reiss Claude
Published: July 2002

To investigate the possible influence of the local rates of translation on protein folding, 16 consecutive rare (in Escherichia coli) codons in the chloramphenicol acetyltransferase (CAT) gene have been replaced by frequent ones. Site‐directed silent mutagenesis reduced the pauses in translation of CAT in E. coli S30 extract cell‐free system and led to the acceleration of the overall rate of CAT protein synthesis. At the same time, the silently mutated protein (with unaltered protein sequence) synthesized in the E. coli S30 extract system was shown to possess 20% lower specific activity. The data suggest that kinetics of protein translation can affect the in vivo protein‐folding pathway, leading to increased levels of protein misfolding.
Dietary and hypothyroid hypercholesterolemia induces hepatic apolipoprotein E expression in the rat: direct role of cholesterol
Authors: Santillo Mariarosaria , Migliaro Angelo , Mondola Paolo , Laezza Chiara , Damiano Simona , Stingo Stefania , Fiorentino Laura , Andreozzi Adriana , Vitale Mario , Bifulco Maurizio
Published: July 2002

Apolipoprotein E (apo E) exerts a protective effect against atherosclerosis, related to its role in intracellular cholesterol removal and remnants clearance. In this study we investigated the effect of dietary and hypothyroid hypercholesterolemia, induced respectively by a high cholesterol diet and by propylthiouracil, on hepatic apo E expression in Wistar male rats. The Northern and Western blot analysis of hepatic mRNA and protein levels showed a 2–3‐fold increase of apo E in hypercholesterolemic rats compared to controls. The incubation of FAO rat hepatoma cells with 25‐OH cholesterol and mevalonate led to a three‐fold increase of apo E mRNA, demonstrating a direct role of cholesterol on apo E expression. This effect was completely abolished by elevating intracellular cAMP levels with forskolin. Immunoblot and immunofluorescence analysis revealed that 25‐OH cholesterol/mevalonate strongly increased also apo E protein synthesis and secretion in FAO cells. Our data demonstrate that hypercholesterolemia, apart of the cause (diet or hypothyroidism) induces liver apo E expression in the rat and that this effect can be directly related, via cAMP, to cholesterol.
Doppler blood flow velocity waveforms of the umbilical artery during variable decelerations in labor
Authors: Sakai M. , Kozuma S. , Okai T. , Kagawa H. , Ryo E. , Taketani Y.
Published: July 2002

Objective: To investigate the changes in resistance indices of blood flow velocity waveforms of the umbilical artery (UmA‐RIs) during variable decelerations (VDs) vs. early decelerations (EDs). Methods: UmA‐RIs were measured before and during uterine contractions in 30 cases with VDs, nine cases with EDs and 20 cases without decelerations. In 12 out of 30 cases with VDs, UmA‐RI was determined at the following five stages of a VD; before the deceleration, downward, bottom, upward and resumption stages. Results: UmA‐RIs during VDs were significantly higher compared with those at preceding periods. UmA‐RIs at downward and bottom stages of VDs were significantly higher than those at the upward stage, although no differences in heart rate were found between downward and upward stages. UmA‐RIs during EDs or uterine contractions without decelerations did not differ from those preceding them. Conclusion: UmA‐RIs increased during VDs, particularly at downward and bottom stages during which the cord seemed to be actually compressed.
Evaluation of interobserver agreement of cardiotocograms
Authors: Bernardes J. , Costa‐Pereira A. , Ayres‐de‐Campos D. , Geijn H.P.van , Pereira‐Leite L.
Published: July 2002

Objective: To evaluate interobserver agreement in visual analysis of each cardiotocographic event. Methods: Three experts independently divided 16 antepartum and 17 intrapartum cardiotocograms into baseline segments, accelerations and decelerations, according to the FIGO guidelines. Baseline segments were further classified as having normal, reduced or increased variability and decelerations as early, late and variable. Uterine activity was divided into tonus and contractions. Agreement was assessed by the proportions of agreement (pa) with 95% confidence intervals. Results: Reproducibility in assessment of baseline segments with normal variability, accelerations and uterine activity was acceptable (pa = 0.56–0.71) whereas that of other segments was not (pa = 0.14–0.45). Conclusions: Analysis of most cardiotocographic events is poorly reproducible, even when experts use the FIGO guidelines. This may be explained by some still ambiguous guidelines, by eyeball limitations in evaluation of subtle events, and by the incapacity of busy clinicians to assess complex and multiple cardiotocographic events in a systematic and disciplined fashion.
Objective computerized fetal heart rate analysis
Authors: Bernardes J , Ayres‐de‐Campos D , Costa‐Pereira A , Pereira‐Leite L , Garrido A
Published: July 2002

Objective: To assess the validity of a computerized methodology for cardiotocogram analysis based on a recently described reproducible visual estimation of the baseline. Methods: Forty‐two antepartum and 43 intrapartum cardiotocograms (CTGs) acquired by a personal computer were selected. Antepartum tracings were performed in the 48 h that preceded an elective cesarean section, and intrapartum tracings were performed until delivery. FHR baselines were estimated by an expert, according to an objective and reproducible methodology. Using these baselines, automated detection of accelerations and decelerations and estimation of variability was performed by the personal computer. A quantitative adaptation of the FIGO guidelines for fetal monitoring was used to classify tracings. Perinatal outcome was classified according to the Apgar score and umbilical arterial pH. Validity was then assessed by the proportions of agreement (PA), kappa statistic (κ), sensitivity and specificity, with 95% confidence intervals (95% CI). Cases showing a disagreement between CTG and perinatal classification were reviewed and an adjustment in baseline definition was tested. Results: The initial overall PA and κ between CTG and perinatal classification were, respectively, 0.79 (95% CI: 0.69–0.87) and 0.62 (95% CI: 0.41–0.83). The overall PA and κ, after baseline adjustment were, respectively, 0.89 (95% CI: 0.81–0.95) and 0.78 (95% CI: 0.58–0.98). Sensitivities and specificities ranged between 79% (95% CI: 60–92%) and 100% (95% CI: 95–100%). Conclusions: Good clinical prediction may be possible with an objective methodology for cardiotocogram analysis based on a recently described reproducible baseline estimation.
Sonographic diagnosis of intrauterine growth restriction (IUGR) by fetal transverse cerebellar diameter (TCD)/abdominal circumference (AC) ratio
Authors: Tongsong T. , Wanapirak C. , Thongpadungroj T.
Published: July 2002

Objective: To evaluate the validity of TCD/AC ratio in predicting IUGR. Study design: Prospective descriptive analysis. Setting: Maharaj Nakorn Chiang Mai Hospital, Chiang Mai University. Subjects: Women meeting the inclusion criteria consisting of: (1) singleton pregnancies after 28 weeks of gestation; (2) known accurate gestational age; and (3) clinically suspected IUGR. Methods: The patients were sonographically examined for TCD/AC ratio. The best cut‐off value of TCD/AC ratio in predicting IUGR was determined by a receiver operating characteristic (ROC) curve. The fetus with a TCD/AC ratio greater than the cut‐off value would be antenatally diagnosed as IUGR for every gestational week. Standard definition of IUGR was a low birthweight, less than the l0th percentile. Results: One hundred and sixty‐seven pregnancies with suspected IUGR were analyzed. The prevalence of IUGR among the study group was 51.5%. The best cut‐off value of the TCD/AC ratio for predicting IUGR was 15.4%, giving the sensitivity, specificity, positive predictive value and negative predictive value of 73.26%, 80.25%, 79.75%, and 73.86%, respectively. Conclusion: The sonographic fetal TCD/AC ratio as a gestational age‐independent method can be helpful in antenatal diagnosis of IUGR, especially in pregnancy with uncertain gestational age.
Daily variations of immunoreactive melatonin in the visual system of crayfish
Authors: Balzer Ivonne , Espínola Iván R. , Fuentes‐Pardo Beatriz
Published: July 2002

In crustaceans, melatonin has been detected in the central nervous system and some other organs. The aim of this study was to analyze the melatonin content in the visual system of Procambarus clarkii, by means of radioimmunoassay, at different day‐night phases. We have also studied the action of exogenous melatonin on the main properties of the electroretinogram (ERG) circadian rhythm. Experiments were conducted with 25 specimens maintained under controlled conditions of 16°C and 12 h of light alternating with 12 h of darkness. Eyes where cut in dim red light and shock frozen with liquid nitrogen and pulverized in a mortar until a homogenous powder was obtained. Melatonin was extracted with acetone, followed by centrifugation, diluted with an equal volume of equa bidest to ensure freezing at −80°C for at least 90 min and lyophilization at the same temperature. Lyophilizates, after having been dissolved in RIA buffer, were used for determinations of melatonin. Long‐term recordings of electrical responses to light (ERG) were obtained for 10 or more consecutive days. At the 5th day, a single dose of melatonin was injected and its effects on amplitude and period of the ERG circadian rhythm were measured. Melatonin concentrations differed considerably depending on the circadian time and attained a maximum during dark phase. Among the crustaceans, Procambarus clarkii represents the first case in which melatonin peaks during the night following the typical pattern known in the majority of organisms. After melatonin injection, period and amplitude of the ERG circadian rhythm were increased. This effect suggests the involvement of melatonin in the oscillators underlying the generation and expression of circadian rhythms in crayfish.
Innovation implementation in high technology manufacturing: A chaos‐theoretic empirical analysis
Authors: Jayanthi Shekhar , Sinha Kingshuk K.
Published: July 2002

This paper conceptualizes the process of innovation implementation in high technology manufacturing, a natural setting of multiple and ongoing innovation implementation. Building on the developments in organizational learning theory, we frame the process of innovation implementation in high technology manufacturing as a problem of balancing between exploitation and exploration. Through the application of a logistic difference equation, we provide insights into the dynamics of balancing between exploitation and exploration, and show that innovation implementation in high technology manufacturing can be conceptualized as a chaotic process, in a scientific sense. Using time series data from a wafer fabrication plant, the high technology manufacturing plant that served as our research site over a period of 125 weeks, we test this conceptualization. We find empirical support for the conceptualization of innovation implementation in high technology manufacturing as a chaotic process. We discuss the managerial implications of our study's findings, and the directions for the future research.
Optimization tools for design and marketing of new/improved products using the house of quality
Authors: Vairaktarakis George L.
Published: July 2002

Quality function deployment (QFD) has helped many firms realize significant reduction in product design costs and development time. The QFD process includes ranking customer preferences, rating the competitors, and parts deployment for the new/improved product. Prior to this research, such activities have been performed based on expert opinion, or the “best‐in‐class” approach. We develop and solve optimization models for the identification of consensus rankings and ratings, that take into account the priorities and perceptions of all the customers in a target market. Then, based on the consensus rankings, we identify a parts mix for the new/improved product that satisfies a budget constraint and matches or exceeds the performance expectations of all customers surveyed in the target market. Finally, we show how the QFD charts can be used to identify competitors that are falsely perceived as superior, as well as areas where the firm's marketing strategies have had the desired effects. Such insights are useful in developing the future marketing strategy of the firm.
Neonatal 6‐hydroxydopamine treatment affects GABAA receptor subunit expression during postnatal development of the rat cerebellum
Authors: Podkletnova I. , Alho H. , Mäkelä R. , Lüddens H. , Helén P. , Korpi E.R.
Published: July 2002

Neurotoxic elimination of noradrenergic terminals by 6‐hydroxydopamine (6‐OHDA) leads to alteration of the granule cell layer formation. We have studied the developmental expression of GABAA receptor subunits in rat cerebellum after neonatal administration of 6‐OHDA during the first postnatal month of life. 6‐OHDA was injected subcutaneously. The expression of GABAA receptor subunits was studied by in situ hybridization and immunohistochemistry. The alterations were observed in the neocerebellum — the part of the cerebellum which starts development postnatally. The migration of granule cells was delayed, and the total area of the granule cell layer in the neocerebellum from 6‐OHDA‐treated rats was reduced to 22.6±5% of the corresponding area from control rats. In situ hybridization with subunit‐specific antisense oligonucleotide probes was performed for α1, α2, α3, α5, α6, β1, β2, γ1 and γ2 subunits of the GABAA receptor. In neocerebellum, 6‐OHDA treatment caused a significant reduction in the α1, α6 and γ2 subunit mRNA levels. The expression of the other subunits was not changed. It has been shown that in the postnatal cerebellum α1 and α6 subunits can be detected in granule cells only when the cells had migrated to their final destination. Our findings indicate that a noradrenergic influence may be necessary for the normal maturation and migration of cerebellar granule cells.
Altered electrophysiological characteristics of developing rat cortical neurones after chronic methylmercury chloride treatment
Authors: Világi Ildikó , Dóczi Judit , Banczerowski‐Pelyhe Ilona
Published: July 2002

The effect of methylmercury chloride on the excitability of developing cortical neurons was tested. Methylmercury was administered in the drinking water to pregnant rats during gestation and suckling period and the offspring were investigated. The electrical characteristics of the neuronal membranes as well as the synaptic responses evoked by electrical stimulation of the corpus callosum were measured in brain slices. Slices prepared from the somatosensory cortex of 4‐week‐old rats were analyzed using sharp electrode intracellular microelectrophysiological recording technique. Long‐lasting treatment with low doses of methylmercury chloride caused a slight decrease in the membrane potential and in the amplitude of spikes together with an enhanced excitability. Some of the treated animals were grown up without any further methylmercury application, and their offspring (second generation) were also studied electrophysiologically. These untreated offspring had normal neuronal characteristics. The altered membrane characteristics detected in the 4‐week‐old MeHg treated animals might be the consequence of the abnormal developmental processes taking place in the presence of MeHg which may alter the normal neuronal excitability. Besides this, the acute toxic effect of Hg2+, which was present in the brain at the time of investigations, has to be taken into consideration.
The Zoology Department at Washington University (1944–1954): from undergraduate to graduate studies with Viktor Hamburger
Authors: Dunnebacke Thelma H.
Published: July 2002

Beginning from an undergraduate's perspective and continuing through graduate school, this student's experiences in the Department of Zoology at Washington University in St. Louis, Missouri was a time of many rewarding experiences. Now, on this occasion of his 100th birthday, I wish to express my appreciation to the Chairman, Dr. Viktor Hamburger, for his teachings, his encouragement, and his friendship that has lasted over the past 56 years.
N‐tert‐butyl‐alpha‐phenylnitrone, a free radical scavenger with anticholinesterase activity does not improve the cognitive performance of scopolamine‐challenged rats
Authors: Milivojevic Nataša , Babic Ksenja , Milatovic Dejan , Dettbarn Wolf‐D. , Sket Dusan , Zivin Marko
Published: July 2002

Reversible inhibitors of acetylcholinesterase improve spatial learning and memory in animal models of cognitive impairment. Here we investigate if the beneficial effects of free radical scavenger N‐tert‐butyl‐alpha‐phenylnitrone (PBN) on cognitive performance could be explained by its recently discovered anticholinesterase activity. Morris water maze experiment was performed to examine the effect of PBN on the impairment of spatial learning and memory induced by the antagonist of cholinergic muscarinic transmission scopolamine. In situ hybridization histochemistry experiment was performed to study its effects on the induction of immediate early gene expression (c‐fos, c‐jun) by dopamine D1 receptor agonist SKF‐82958 and on the augmentation of the SKF‐82958‐induced expression of these genes by scopolamine. In both experiments, the effects of PBN were compared to the effects of reversible anticholinesterase physostigmine. We found that physostigmine but not PBN significantly reversed the cognitive impairment in scopolamine‐challenged rats, prevented the induction of c‐fos and c‐jun mRNAs by SKF‐82958 and attenuated the augmentation of the SKF‐82958‐induced expression of these genes by scopolamine. The present experiments did not reveal a significant in vivo anticholinesterase activity of PBN.
Deliberate self-poisoning presenting at a rural hospital in Northern Ireland 1976–1996: relationship to prescribing
Authors: Kelly C.B. , Weir J. , Rafferty T. , Galloway R.
Published: July 2002

summary

Purpose – This study reports on a project to monitor deliberate self-poisoning in a rural area of Northern Ireland over a 20-year period. Comparison is made with reports from large urban centres. In addition, a local prescribing database allows assessment of any association between psychotropic drug prescription and use for deliberate self-poisoning. Materials and methods – Frequency of self-poisoning, demographic details and drugs used were recorded for all episodes of deliberate self-poisoning occurring at Craigavon Area Hospital for the years 1976, 1986, 1991 and 1996. It was possible to compare prescriptions of psychotropic drugs with their use for deliberate self-poisoning between the years 1991 and 1996 in the region served by the hospital, using the Defined Daily Dose (DDD) system. Results – In this rural area the pattern of deliberate self-poisoning has changed, as in urban centres, with a rise in frequency and the male/female ratio approaching unity. The pattern of drug use has altered, with paracetamol overtaking benzodiazepines as the most commonly used agent. More recently, antidepressants have become the second most frequently used drug class for this purpose. Psychotropic medications used for self-poisoning altered in proportion to their prescription between the years 1991 and 1996. Conclusions – In the face of a continuing rise in deliberate self-poisoning, which is effecting both urban and rural areas, care should be taken to prescribe the least toxic agent available as this is associated with likely frequency of self-poisoning for most classes of psychotropic drug.