bluuebunny/crossref_metadata_2025 huggingface dataset:
RNA polymerase II 140wimp mutant and mutations in the TFIIH subunit XPB differentially affect homeotic gene expression in Drosophila
Authors: Gutiérrez Luis , Merino Carlos , Vázquez Martha , Reynaud Enrique , Zurita Mario
Published: August 2004

Mutations in the XPB and XPD helicases of the DNA repair/transcription factor TFIIH are involved in several human genetic disorders. An unanswered problem concerning the complexity of the phenotype–genotype relationship is why mutations in individual subunits of TFIIH produce specific phenotypes and not many others. In order to investigate this question we tested whether mutations in the Drosophila XPB homolog, haywire (hay), would modify homeotic derepression phenotypes. In this work, we report that mutations in hay and in the 140‐kDa subunit of the RNA polymerase II (RpII140wimp) act as dominant modifiers of the derepression phenotypes of the Sex combs reduced (Scr) and Ultrabithorax (Ubx) genes. The hay mutations only weakly suppress the Scr derepression phenotype caused by the AntpScx mutation but not by Polycomb. In contrast, the RpII140wimp mutation strongly suppresses both Scr derepression phenotypes. In addition, the RpII140wimp also generates phenotypes indicative of loss of Ubx function. On the other hand, all the derepression homeotic phenotypes are sensitive to the generalized reduction of transcription levels when the flies are grown with actinomycin D. We also show that different promoter control regions have differential sensitivity to different hay alleles. All these results support that although TFIIH is a basal transcription factor, mutations in the subunit encoded by hay have specific effects in the transcription of some genes. genesis 40:58–66, 2004. © 2004 Wiley‐Liss, Inc.
Pressure variations in peat as a result of gas bubble dynamics
Authors: Kellner Erik , Price Jonathan S. , Waddington J. M.
Published: August 2004

Transient high pore‐water pressures, up to 50 cm higher than ambient pressure, developed over the summer season at various depths in a shallow (1 m) fen peat. The excess pressures had a pattern of gradual increases and sharp drops, and their initiation and release typically corresponded to abrupt changes in atmospheric pressure. We conclude that these phenomena depend on gas bubbles (probably methane) generated by biological activity, both by clogging pores and by building up pressure as they grow. These transient and spatially discontinuous high‐pressure zones were found using pressure transducers in sealed (backfilled) pits, but not in piezometers open to the atmosphere. Piezometers may provide a conduit for the release of gas and pressure, thus rendering them unsuitable for measuring this phenomenon.

Although the development of localized zones of high pressure causes erratic and unpredictable hydraulic gradients, we suggest that their effect on the flow of water or solutes is offset by the reduced permeability caused by the bubbles, which allows them to be sustained. These zones, however, probably deflect flows driven by the dominant hydraulic gradients. Furthermore, they may cause the peat volume to adjust (swell). The use and interpretation of traditional methods for estimating hydraulic head and conductivity in peat soils thus require great caution. Copyright © 2004 John Wiley & Sons, Ltd.
Human hepatic and renal microsomes, cytochromes P450 1A1/2, NADPH:Cytochrome P450 reductase and prostaglandin H synthase mediate the formation of aristolochic acid‐DNA adducts found in patients with urothelial cancer
Authors: Stiborová Marie , Frei Eva , Hodek Petr , Wiessler Manfred , Schmeiser Heinz H.
Published: August 2004

Aristolochic acid (AA), a naturally occurring nephrotoxin and carcinogen, has been associated with the development of urothelial cancer in humans. Understanding which human enzymes are involved in AA activation and/or detoxication is important in the assessment of an individual's susceptibility to this plant carcinogen. Using the 32P postlabeling assay, we examined the ability of microsomal samples from 8 human livers and from 1 human kidney to activate AAI, the major component of the plant extract AA, to metabolites forming adducts in DNA. Microsomes of both organs generated DNA adduct patterns reproducing those found in renal tissues from humans exposed to AA. 7‐(deoxyadenosin‐N6‐yl)aristolactam I, 7‐(deoxyguanosin‐N2‐yl)aristolactam I and 7‐(deoxyadenosin‐N6‐yl)aristolactam II were identified as AA‐DNA adducts formed from AAI by all human hepatic and renal microsomes. To define the role of human microsomal enzymes in the activation of AAI, we investigated the modulation of AAI‐DNA adduct formation by cofactors and selective inhibitors of microsomal reductases, cytochrome P450 (CYP) enzymes, NADPH:CYP reductase and NADH:cytochrome b5 reductase. We also determined whether the activities of CYP and NADPH:CYP reductase in different human hepatic microsomal samples correlated with the levels of AAI‐DNA adducts formed by the same microsomal samples. On the basis of these studies, we attribute most of the activation of AAI in human hepatic microsomes to CYP1A2. In contrast to human hepatic microsomes, in human renal microsomes NADPH:CYP reductase is more effective in AAI activation. In addition, prostaglandin H synthase is another enzyme activating AAI in renal microsomes. The results demonstrate for the first time the potential of microsomal enzymes in human liver and kidney to activate AAI by nitroreduction.
1α,25(OH)2D3 Regulation of integrin expression is substrate dependent
Authors: Raz P. , Lohmann C. H. , Turner J. , Wang L. , Poythress N. , Blanchard C. , Boyan B. D. , Schwartz Z.
Published: August 2004

Osteoblasts are attachment‐dependent cells that interact with their surface through integrin‐mediated mechanisms. Their differentiation is regulated by 1,25‐dihydroxyvitamin D3 [1α,25(OH)2D3] and is affected by substrate chemistry and microtopography, suggesting that 1α,25(OH)2D3 may regulate integrin expression in a surface‐specific manner. To test this hypothesis, osteoblast‐like human MG63 cells were grown on tissue culture plastic and on grit‐blasted and acid‐etched titanium disks with a complex microtopography to induce osteoblast differentiation. Expression of α2, α5, αv, β1, and β3 integrins were quantified by real‐time polymerase chain reaction (PCR) as a function of time in culture and treatment with 1α,25(OH)2D3. Results were correlated with expression of osteocalcin, a marker of a differentiated osteoblast. Osteocalcin mRNA increased with time and 1α,25(OH)2D3 treatment and these changes were greater in cultures on the titanium disks. Integrin expression varied with time in culture and this was also surface dependent. At each time point, β1 and α2 mRNAs were greater on titanium than on plastic, whereas α5 expression was reduced and αv,β3 expression was unaffected. 1α,25(OH)2D3 increased β1 mRNA on both surfaces at all time points, but it increased α2 expression only in 8‐d cultures. 1α,25(OH)2D3 caused reduced α5 expression only in cultures grown on plastic for 8 d, and had no effect on either αv or β3 expression regardless of surface. These results show that integrin expression in human osteoblast‐like cells is differentially modulated by 1α,25(OH)2D3 in a time‐dependent manner that is sensitive to the surface on which the cells are grown. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 71A: 217–225, 2004
Exploring the predictability of the ‘Short Rains’ at the coast of East Africa
Authors: Hastenrath Stefan , Polzin Dierk , Camberlin Pierre
Published: August 2004

The boreal autumn ‘Short Rains’ at the coast of East Africa are deficient when there is weak development of a zonal circulation cell along the Indian Ocean equator, an anomalously low sea‐surface temperature in the western portion of the basin, and in the high phase of the southern oscillation. Such large‐scale circulation departures and their precursors are described by compact indices. September values of these indices for the period 1958–96 are used to explore the predictability of an index (RON) of October–November rainfall at the coast of East Africa. Regressions with cross‐validation over the entire 1958–96 period are evaluated for the early (1958–77) and late (1978–96) halves of the record. In complementary experiments, the entire record is separated into 1958–77 as a training period and 1978–96 as a verification period. In all experiments, correlation of calculated versus observed rainfall is high for the early record and low for the late half of the record, a behaviour not noted in cross‐validation over the entire 39 year time span. The 11‐year sliding correlations of the indicated circulation indices with RON all reveal a drastic deterioration of relationships from the early to the late half of the record, although the equatorial zonal circulation cell appears to remain strong throughout. Copyright © 2004 Royal Meteorological Society
Measured wind speed trends on the west coast of Canada
Authors: Tuller Stanton E.
Published: August 2004

Trends in measured wind speed are discussed for four stations on the west coast of Canada. Periods of record vary with the station. They begin in the late 1940s or the 1950s and run through to the early to mid 1990s. The most prominent feature of the time series was a decline in mean annual and winter wind speeds at Cape St James, Victoria International Airport, and Vancouver International Airport during the middle portion of the record. Declines in mean annual wind speed are matched by increases in the percentage of calms and decreases in high wind speed observations. The pressure gradient between Victoria, Vancouver and Comox, the Pacific North American index, the Pacific decadal oscillation index, and other climate elements in British Columbia and the northwestern USA show trends at roughly the same time, indicating a natural cause of the wind speed decrease. Comox Airport mean wind speeds increased, however, perhaps the result of reduced friction in the vicinity of the anemometer outweighing the decrease in the regional pressure gradient. Copyright © 2004 Royal Meteorological Society
Energy and radiation balance of a central European city
Authors: Christen Andreas , Vogt Roland
Published: August 2004

Results from an experimental network of seven energy balance stations in and around a European city are presented. The network of micrometeorological stations was part of the Basel Urban Boundary Layer Experiment (BUBBLE) carried out in the city of Basel, Switzerland. Three urban sites provided turbulent flux densities and radiation data over dense urban surfaces. Together with a suburban site and three rural reference sites, this network allowed the simultaneous comparison of urban, suburban, and rural energy balance partitioning during one month of summertime measurements. The partitioning is analysed together with long‐term data to evaluate the magnitude of the urban flux density modification, and to document characteristic values in their diurnal and yearly course. Simple empirical relations between flux densities and surface characteristics are presented. The energy balance partitioning is addressed separately for daytime and nocturnal situations. All four components of the surface radiation budget are analysed. Moreover, the vertical flux density divergences within the urban canopy layer are discussed. Copyright © 2004 Royal Meteorological Society
Improving precipitation measurement
Authors: Strangeways Ian
Published: August 2004

Although rainfall has been measured for centuries scientifically and in isolated brief episodes over millennia for agriculture, it is still not measured adequately even today for climatology, water resources, and other precise applications. This paper outlines the history of raingauges, their errors, and describes the field testing over 3 years of a ‘first guess’ design for an aerodynamic rain collector proposed by Folland in 1988. Although shown to have aerodynamic advantage over a standard 5″ gauge, the new rain collector was found to suffer from outsplash in heavy rain. To study this problem, and to derive general basic design rules for aerodynamic gauges, its performance was investigated in turbulent, real‐world conditions rather than in the controlled and simplified environment of a wind tunnel or mathematical model as in the past. To do this, video records were made using thread tracers to indicate the path of the wind, giving new insight into the complex flow of natural wind around and within raingauges. A new design resulted, and 2 years of field testing have shown that the new gauge has good aerodynamic and evaporative characteristics and minimal outsplash, offering the potential for improved precipitation measurement. Copyright © 2004 Royal Meteorological Society
Energy balance at a cold Alpine firn saddle, Seserjoch, Monte Rosa
Authors: Suter Stephan , Hoelzle Martin , Ohmura Atsumu
Published: August 2004

Little knowledge exists to date on the complex interactions between the atmosphere and glaciers leading to the formation of cold firn in high‐elevation accumulation areas in the Alps. Although the occurrence of cold firn and ice together is not a widespread phenomenon in the Alps, it is of particular significance as it relates to many glaciological issues, and has a major impact on glaciers in their role as an environmental and climatic archive. The atmospheric impact on cold firn was investigated using an energy balance study at the cold, 4300 m high firn saddle of Seserjoch, Monte Rosa area (Italy and Switzerland). Measurements of short‐ and longwave radiation, wind speed and wind direction, air temperature, humidity, snow height and snow and firn temperatures were conducted between September 1998 and August 2000 under difficult meteorological conditions. A 1 year time series of energy balance measurements spanning the period of May 1999 to April 2000 is presented. The results show that net radiation and turbulent heat fluxes are major elements of the energy balance. The heat fluxes due to surface melt in summer, and re‐freezing events (re‐freezing of meltwater at the surface, or rime accretion) cannot be neglected. The magnitude of these fluxes is difficult to interpret, as these are derived indirectly from the energy balance and, hence, also include instrumental and methodological errors in the energy balance calculation. Precise high‐resolution measurements of surface temperatures in summer made it possible to identify individual surface melt events and the prevailing meteorological conditions favouring or hindering surface melt. Copyright © 2004 Royal Meteorological Society
Porous‐Conductive Chitosan Scaffolds for Tissue Engineering, 1
Authors: Wan Ying , Wu Hua , Wen Dijiang
Published: August 2004

Summary: Novel porous‐conductive chitosan scaffolds were fabricated by incorporating conductive polypyrrole (PPy) particles into a chitosan matrix and employing a phase separation technique to build pores inside the scaffolds. Conductive polypyrrole particles were prepared with a microemulsion method using FeCl3 as a dopant. The preparation conditions were optimized to obtain scaffolds with controlled pore size and porosity. The conductivity of the scaffolds was investigated using a standard four‐point probe technique. It was found that several kinds of scaffolds showed a conductivity close to 10−3 S · cm−1 with a low polypyrrole loading of around 2 wt.‐%. The main mechanical properties, such as tensile strength, breaking elongation and Young's modulus of the scaffolds, were examined both in the dry and in the hydrated states. The results indicated that a few different kinds of scaffolds exhibited the desired mechanical strength for some tissue engineering applications. The miscibility of polypyrrole and chitosan was also evaluated using a dynamic mechanical method. The presence of significant phase separation was detected in non‐porous PPy/chitosan scaffolds but enhanced miscibility in porous PPy/chitosan scaffolds was observed.

Variation in conductivity of the scaffolds with polypyrrole content.

image

Variation in conductivity of the scaffolds with polypyrrole content.
Photopolymerization of acrylamide initiated by the three‐component system safranine/triethanolamine/diphenyliodonium chloride: The effect of the aggregation of the salt
Authors: Gómez María L. , Montejano Hernán A. , del Valle Bohorquez María , Previtali Carlos M.
Published: August 2004

The effects of diphenyliodonium chloride (DPI) on the polymerization of acrylamide photoinitiated by the dye safranine with triethanolamine as a coinitiator were investigated in aqueous solutions. The salt notably increased the polymerization rate. This accelerating effects increased appreciably at concentrations of DPI at which the light scattering of the solutions became important. The effects of DPI on the photophysical properties of the dye were also investigated. Although the absorption and fluorescence were scarcely affected, the triplet yield increased by 60%. However, the polymerization rate increased by a factor much higher than that of the triplet yield. The results obtained at high concentrations of the salt could be ascribed to the presence of aggregates of the hydrophobic cations. The lower limit established for the formation of the aggregates was a DPI concentration of approximately 1 × 10−3 M. Possible mechanisms for the action of the salt were examined. © 2004 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 42: 4916–4920, 2004
Thermal self‐initiation of styrene in the presence of TEMPO radicals: Bulk and miniemulsion
Authors: Pan Gaofeng , Sudol E. David , Dimonie Victoria L. , El‐Aasser Mohamed S.
Published: August 2004

In TEMPO (2,2,6,6,‐tetramethyl‐1‐piperidinyloxy) controlled styrene radical polymerizations, the thermal self‐initiation reaction of styrene monomer is one of the main sources for the deviations from ideal living polymerization. However, it is also important because it continuously generates radicals to compensate for the loss of radicals caused by the termination reactions and thereby maintains a reasonable reaction rate. In this report, different initial TEMPO concentrations were used in styrene miniemulsion polymerizations without any added initiator. The consumption rate of TEMPO or radical generation rate was calculated from the length of the induction period and the increasing total number of polymer chains. It was found that there is little difference between the miniemulsions and the corresponding bulk systems in terms of the length of the induction period, which increases linearly with initial TEMPO concentration. After the induction period, the consumption rate of TEMPO or radical generation rate was reduced to a lower level, and a faster initial polymerization rate was found in the bulk system compared to the corresponding miniemulsion system. © 2004 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 42: 4921–4932, 2004
Determination of tributyrin and its metabolite butyrate in Wistar rat plasma samples by gas chromatography/mass spectrometry
Authors: Su Jie , Zhang Ningning , Ho Paul C.
Published: August 2004

A gas chromatographic (GC) method with mass spectrometric (MS) detection was developed for the determination of tributyrin and its metabolite butyrate in rat plasma. Following precipitation of plasma protein with acetonitrile, the analytes in the samples were separated on a DB‐5ms capillary column with helium as carrier gas. Phenylmethylsulfonyl fluoride (PMSF), an inhibitor for serine proteases, papain and acetylcholinesterase, was found to be essential to inhibit the activity of enzyme(s) responsible for the hydrolysis of tributyrin in both rat and human blood samples. The enzyme inhibitor in 5 mM (final concentration) was added immediately into the blood samples after collection to prevent the hydrolysis. The linear concentration ranges for tributyrin and butyrate were 0.1–2.0 and 1–20 μM, respectively. The coefficients of variation for intra‐day and inter‐day assays for tributyrin were all <10%, and those for butyrate were also <10%, except for the lowest concentration (1 μM), which was less than 20%. The accuracy of all concentration determinations ranged from 96.0–110.0%. The limit of quantification (LOQ) was 0.1 μM for tributyrin and 1.0 μM for butyrate. This method could detect tributyrin and butyrate simultaneously, and represents an improvement in sensitivity for the detection of tributyrin compared with the previous gas chromatography‐flame ionization detection (GC‐FID) method. Copyright © 2004 John Wiley & Sons, Ltd.
A new and sensitive on‐line liquid chromatography/mass spectrometric approach for top‐down protein analysis: the comprehensive analysis of human growth hormone in an E. coli lysate using a hybrid linear ion trap/Fourier transform ion cyclotron resonance mass spectrometer
Authors: Wu Shiaw‐Lin , Jardine Ian , Hancock William S. , Karger Barry L.
Published: August 2004

A sensitive, integrated top‐down liquid chromatography/mass spectrometry (LC/MS) approach, suitable for the near complete characterization of specific proteins in complex protein mixtures, such as inclusion bodies of an E. coli lysate, has been successfully developed using a hybrid linear ion trap/Fourier transform ion cyclotron resonance (FTICR) mass spectrometer. In particular, human growth hormone (hGH) (200 fmol) was analyzed with high sequence coverage (>95%), including the sites of disulfide linkages. The high mass accuracy and resolution of the FTICR mass spectrometer was used to reveal high charge state ions of hGH (22 kDa). The highly charged intact protein ions (such as the 17+ species) were captured and fragmented in the linear ion trap cell. The fragment ions from MS/MS spectra were then successfully analyzed in the FTICR cell in an on‐line LC/MS run. Peptide fragments from the N‐terminal and C‐terminal regions, as well as large interior fragments, were captured and identified. The results allowed the unambiguous assignment of disulfide bonds Cys53–Cys165 and Cys182–Cys189, indicative of proper folding of hGH. The disulfide bond assignments were also confirmed by analysis of the tryptic digest of a sample of hGH purified from inclusion bodies. On‐line LC/MS with the linear ion trap/FTICR yields high mass accuracy in both the MS and MS/MS modes (within 2 ppm with external calibration). The approach should prove useful in biotechnology applications to characterize correctly folded proteins, both in the early protein expression and the later processed stages, using only a single automated on‐line LC/MS top‐down method. Copyright © 2004 John Wiley & Sons, Ltd.
Product variety and firm survival in the microcomputer software industry
Authors: Cottrell Tom , Nault Barrie R.
Published: August 2004

This article provides an analysis of product variety and scope economies in the microcomputer software industry by using detailed firm‐level and product‐level information on firms' bundling of functionalities over application categories and computing platforms. We find that the management of product variety through the way different application categories are integrated in products and the platforms on which these products are offered can be as important as the significance of scope economies at the more aggregated firm level. Specifically, we find that there is little evidence of firm benefits from economies of scope in production, but there is substantial evidence that products benefit from economies of scope in consumption. In addition, we find that firms with products that encapsulate more application categories perform better, and those with products that cover more computing platforms perform worse. Finally, changes in product variety through new product introductions improve firm performance, but extensions to existing products hinder the performance of the firm and the product. We conclude that research in scope economies can benefit from a more detailed model of the evolution of product variety that includes data and analysis at the firm level and at the product level. Copyright © 2004 John Wiley & Sons, Ltd.
Improving patients' knowledge of the fetal anomaly scan
Authors: Smith R. P. , Titmarsh S. , Overton T. G.
Published: August 2004

Objectives

To determine the level of patient knowledge regarding the 20‐week screening ultrasound examination, which is performed on over 95% of women in our unit, and to ascertain whether the use of targeted education could improve patient knowledge and expectations, to determine from where patients obtained information on the scan and to ascertain whether patients felt they had chosen to undergo the scan.

Methods

An anonymous questionnaire was issued to all patients attending for their 20‐week scan over a 3‐week period in July 2002 (n = 220). There were nine questions, focusing on maternal choice, information and knowledge. As a result of the information obtained, we undertook a program of education for hospital and community medical and midwifery staff. A printed information sheet given to the patients at the time of booking their 20‐week scan was introduced. We then issued the questionnaire for another 3‐week period in July 2003 (n = 171).

Results

As a result of our education program, there was a significant increase in the number of women answering various knowledge‐based questions correctly about the 20‐week scan, in those using the written information provided as a source of information (P < 0.0001), and in those who felt they had been involved in the decision to have the scan (P = 0.003).

Conclusions

Patients need to be better informed about the 20‐week scan, and choose whether or not to have it. Health professionals are pivotal in this process. While written information helps to reinforce the information given, alone it has only a small effect. Copyright © 2004 ISUOG. Published by John Wiley & Sons, Ltd.
Intranuclear localization of the Ki‐67 reactive antigen in HeLa cells. Flow cytometric analysis
Authors: Sasaki Kohsuke , Matsumura Kouji , Murakami Tomoyuki , Shinozaki Fumihiko , Takahashi Manabu
Published: August 2004

Summary— The intranuclear localization of the Ki‐67 reactive antigen was immunocytochemically investigated using flow cytometry. HeLa S3 cells were immunocytochemically stained with the monoclonal antibody, Ki‐67, after in situ treatments with various kinds of compounds, namely: HCl; NaCl; RNase; Sl nuclease and DNase I. The only treatment that markedly diminished the immunofluorescence intensity of the cells was exposure to DNase I. Nuclear fluorescence was no longer observed in the cells digested with relatively high concentrations of DNase I. These results suggest that the antigen recognized by Ki‐67 is closely associated with DNA, but is not directly associated with either the nuclear matrix or histones.
Study of the heterogeneity of a human non‐small cell bronchopulmonary epidermoid primary carcinoma: determination of the DNA content of NSCLCN6L2 cells and of cloned sub‐populations
Authors: Gratas Catherine , Robillard Nelly , Roussakis Christos , Audouin Anne‐Françoise , Verbis Jean‐François
Published: August 2004

Summary— A continuous cell line, NSCLCN6L2, was established in vitro from a human bronchopulmonary epidermoid carcinoma and then cloned on agar gel and by selective media. The DNA content of each sub‐population was compared with that of the parent line by flow cytometry. This study showed the heterogeneity of the NSCLCN6L2 line and the possibility of selection by cloning distinct ploidy sub‐populations (group 1: diploid lines; group 2: hypotetraploid lines; group 3: a diploid and tetraploid line); it also allowed the time‐course of the evolving lines to be followed. Correlation of these results with other properties of the different sub‐populations will provide a better understanding of their biological behavior, particularly of their chemosensitivity.
Response of meristem cells to hyperoxygenation
Authors: Giménez‐Abián M. Inmaculada , López‐Sáez Jorge F. , De La Torre Consuelo
Published: August 2004

Summary— Cycle parameters were estimated in Allium cepa L meristems proliferating under steady state conditions in tap water aerated with either 21 or 100% O2 and containing 7.9 and 39.0 mg/1 of dissolved oxygen, respectively. Cycle time was slightly but consistently reduced under hyperoxygenation, as shown in synchronous cells labelled as binucleate following a short treatment with 5 mM caffeine. By autoradiography after 3H‐thymidine treatment followed by microdensitometry of unlabelled interphases it was determined that cellular frequencies at the last third of the S period (S3) were statistically different (P<0.05) in control and hyperoxygenated conditions. Estimation of absolute timing confirmed that S3 duration was nearly halved under the increased O2 tension. Finally, G1 time was not modified by hyperoxygenation, though this cycle stage lengthened under hypoxia.
Immunochemical localization and association with spermatozoa of androgen‐regulated proteins of MR 24000 secreted by the mouse epididymis
Authors: Jimenez Clément , Ghyselinck Norbert B. , Depeiges Annie , Dufaure Jean‐Pierre
Published: August 2004

Summary— A polyclonal monospecific immune serum was raised against androgen‐regulated proteins with Mr 24000 secreted by the mouse caput epididymidis (6). Sections of frozen tissues from the different regions of the epididymis have been studied by indirect immuno‐ fluorescence. Results indicate that the antigens are secretory proteins produced by the epithelial cells of the caput epididymidis, essentially in the medial and distal segments. Accumulation of the antigens was observed in the lumen of the caput and the corpus epididymal duct. Subsequently, their association with the sperm surface occurred and persisted down to the cauda epididymidis.
Redistribution and recycling of internalized membrane in seminal vesicle secretory cells
Authors: Mata Lucinda R. , Christensen Erik I.
Published: August 2004

Summary— This study was performed to clarify the fate of membrane constituents internalized from the apical domain in secretory cells, in particular their possible recycling and the compartments involved in it. Glycoproteins of the apical membrane of seminal vesicle secretory cells from guinea‐pig were covalently labeled in vitro (0°C, 20 min) with 3H‐galactose and the epithelium incubated for 15 min (37°C, first incubation) to allow endocytosis. The label which was not internalized was then exposed to enzymatic hydrolysis (0°C, 30 min) and the epithelium re‐incubated to allow membrane movement for 15 and 30 min (37°C, 2nd incubation). After each step of the protocol, tissue pieces were fixed and processed for electron microscope autoradiography and the results studied by morphometric analysis. Following labeling, 99% of the silver grains were associated with the apical domain of the cell membrane (AD). After the 1st incubation at 37°C, 30° of the grains were inside the cells in association with the cytoplasmic vesicles (Cyt ves), secretory vacuoles (SV), Golgi vesicles (GV), Golgi cisternae (GC), multivesicular bodies (MVB), lysosomes (LYS), and the cell membrane basolateral domain (BLD). About 58% of non‐internalized radioactivity was removed by hydrolysis. During the 2nd incubation at 37°C the concentration of label increased in BLD and LYS, decreased in SV and MVB, and fluctuated in GC, GV and AD. The distribution of grains observed at 15 min, as compared using the χ‐square test, was highly significantly different from that expected without recycling. The results show that cell membrane glycoproteins internalized at the cell apex recycle back to the membrane apical domain and are consistent with the involvement of GC and SV in the recycling pathway. Membrane shuttle between the apical and basolateral domains of the cell membrane is also suggested by these observations.
Cellular localization of an embryonic interferon, ovine trophoblastin and its mRNA in sheep embryos during early pregnancy
Authors: Guillomot Michel , Michel Catherine , Gaye Pierre , Charlier Nadia , Trojan Jean , Martal Jacques
Published: August 2004

Summary— The ovine embryo produces an interferon named ovine Trophoblastin (oTP) which is involved in the maternal recognition of pregnancy and ensures the maintenance of progesterone secretion by the corpus luteum. We have used indirect immunohistofluorescence and in situ hybridization on histological sections to investigate the fate of this protein and its mRNA in ovine embryos from days 3 to 25 of pregnancy. The level of expression was measured by image analysis of the autoradiographs after in situ hybridization. Both techniques clearly demonstrated that oTP and its mRNA were specifically localized in the extra‐embryonic trophoblast. Neither the embryonic cells, nor the yolk sac or the amniotic tissues produced the protein or its mRNA. The protein could be detected by d 11 of pregnancy in the elongated blastocyst. Maximum of expression is observed at d 14 and the level decreased by d 16 of pregnancy. The arrest of expression occurred in the regions of trophoblast which have established cellular contacts with the uterine epithelium during the implantation process.
Behavior of confluent endothelial cells after irradiation. Modulation of wound repair by heparin and acidic fibroblast growth factor
Authors: Klein‐Soyer Claude , Beretz Alain , Cazenave Jean‐Pierre , Driot Francis , Maffrand Jean‐Pierre
Published: August 2004

Summary— Image analysis was used to study the repair process of a circular mechanical lesion of confluent human endothelial cells in culture after irradiation (10 Gy) prior to wounding. Coverage of denuded areas 48 and 96 h after injury of endothelial cells was identical in control and irradiated cultures, although the labeling index was lowered by 80 to 95% in irradiated cultures. The cell density of non damaged irradiated areas was decreased by 50%. When cultures were submitted to increasing doses of radiation (5.0–30 Gy), the labeling index of the cells diminished rapidly between 0 and 5.0 Gy and reached a plateau at 10 Gy. The decrease in cell density (50% of control at 96 h) was identical at each dose of radiation. Thus cell migration alone could be sufficient for the repair of the lesion, while cell proliferation would mainly maintain the original cell density. The addition of heparin to the culture medium slowed down cell migration and proliferation, but the speed of repair was identical in irradiated and non‐irradiated cultures. Acidic fibroblast growth factor plus heparin accelerated equally the repair process whether the cultures were irradiated or not. In irradiated cultures the presence of acidic fibroblast growth factor and heparin maintained cell density in confluent areas at a level similar to that in non‐irradiated damaged control cultures without addition of mitogens. Thus heparin and acidic fibroblast growth factor play a role in cell proliferation, in the maintenance of the cell monolayer integrity and in restoring a continuous layer by rapid cell migration and elongation after irradiation.
Immunocytochemical demonstration of quantitative differences in the distribution of lysozyme in human airway secretory granule phenotypes
Authors: Hinnrasky Jocelyne , Chevillard Martine , Puchelle Édith
Published: August 2004

Summary— The distribution of lysozyme in the different secretory granules (SG) of human tracheal and bronchial submucosal gland serous cells was studied by light and electron microscopy, using a post‐embedding immunogold technique. SG were differentiated into 5 phenotypes according to their structure and staining electron density. All the SG‐phenotypes were reactive to lysozyme. In the heterogeneous SG‐phenotypes, quantitative immunocytochemistry showed that the density of lysozyme labeling was significantly higher in the electron‐dense central core compared to the electron‐lucent peripheral rim. At the tracheal level, the density of lysozyme did not vary significantly within the different SG‐phenotypes, whereas at the bronchial level, the differences were significant. Moreover, the lysozyme labeling density was much higher in the bronchial than in the tracheal SG.
Electron microscopic study of the optic tentacle collar cells of the snail Helix aspersa and immunolocalization of methionine enkephalin‐like material in their granular content
Authors: Magdelaine Sylvie , Marchand Claude‐Roland , Griffond Bernadette
Published: August 2004

Summary— An ultrastructural and immunocytological study was carried out on the collar cells of the optic tentacle of Helix aspersa. These cells are supposed to be the source of a reproduction controlling hormone. The immunocytological study was performed using an anti‐methionine enkephalin antibody obtained from rabbits in our laboratory. The collar cells are characterized by an enlarged rough endoplasmic reticulum, numerous mitochondria and Golgi bodies surrounded by secretory vesicles, suggesting an intense synthesizing activity. Their principal feature consists of numerous various‐sized granules where methionine enkephalin immunoreactivity is localized. No classical neurosecretory granules are observed while synapse‐like structures are often encountered. The cells should not be regarded as neurosecretory cells but rather as glandular cells which could ensure different functions, one in relation to reproduction, and another in relation to perception processes, particularly as they contain methionine enkephalin‐like material.
Construction and characterization of a DNA library from mouse chromosomes 19 purified by flow cytometry
Authors: Baron Bruno , Métézeau Philippe , Kiefer‐Gachelin Hélène , Goldberg Michel E.
Published: August 2004

Summary—

In spite of the constant development concerning physical mapping of eukaryotic genomes, the mouse chromosome 19 remains poorly characterized. In order to improve the possibilities for studying this chromosome, we have constructed a chromosome‐specific EcoRI DNA fragment library from mouse chromosomes 19 sorted by flow cytometry. The resulting library contains about 3 × 104 recombinant clones. The identified inserts range in size from about 0.2–10 kb, with a 4 kb average size and with no observable redundancy. The purity of the library has been analyzed by flow‐blot. For that purpose, chromosomes from 2 cell lines, 1 with a normal karyotype and 1 with translocated chromosome 19, were sorted on nylon filters and hybridized with 9 clones of the library. Results show that 5 clones out of the 9 clearly originate from sorted chromosomes 19 and 3 and are likely to be derived from its DNA, thus indicating that the library of chromosome 19 is of high purity.
Tendon collagen fibrillogenesis is a multistep assembly process as revealed by quick‐freezing and freeze‐substitution
Authors: Mallein‐Gérin Frédéric , Garrone Robert
Published: August 2004

Summary—

The ultrastructure of chick embryo tendons has been examined after quick‐freezing by liquid helium and freeze‐substitution. Several stages of collagen assemblies were observed: intracellular packing of SLS‐like aggregates surrounded by membrane containing areas with a clathrin coat; fine non cross‐striated filaments connecting the cell membrane at 1 pole of the cells and collagen fibrils; tufts of filaments directly linked to collagen fibrils. This study reveals that some stages are more constant and abundant than supposed (the intracellular SLS‐like aggregates) and that other extracellular assemblies that were hypothesized but usually badly preserved by conventional electron microscopy are clearly captured by the method.
Effects of cerulenin, an inhibitor of fatty acid synthesis on reconstitution of the dental basement membrane
Authors: Goldberg Michel , Lecolle Sylvie , Lesot Hervé , Ruch Jean‐Victor
Published: August 2004

Summary—

When trypsin‐dissociated enamel organs and dental papillae were recombined in the presence of cerulenin — an antibiotic which is a potent inhibitor of fatty acid synthesis — the newly synthesized basement membrane seemed defective in a dose‐dependent manner. The three‐dimensional relationship between the basement membrane components and the plasma membrane appears to be regulated in part by lipids.
Location of Drosophila C virus target organs in Drosophila host population by an immunofluorescence technique
Authors: Lautié‐Harivel Nicole , Thomas‐Orillard Michèle
Published: August 2004

Summary—

Using the immunofluorescence technique we attempted to locate, in the Drosophila host, Drosophila C virus (DCV) target organs after injection of adult flies. Two kinds of organs were infected: those which play a role in reproductive function, including the fat body and follicular cells, and others, including thoracic muscle fibers, tracheal cells, and the digestive tract. These organs correspond to those found in previous tests. Fat body proteins of a DCV‐free host population seemed to cross‐react with antivirus C antibody. This immune response depended on the origin of the host population. It is known that, when DCV is ingested from the first larval instar, it may have beneficial effects upon host development and reproduction. As DCV has a narrow host spectrum, it is suggested that it is well adapted to its natural host. Hypotheses are proposed to explain how the host resists viral infection and may in fact benefit from such an infection.
Calcium phosphate deposits in domes of human pancreatic adenocarcinoma (Capan‐1) cell cultures
Authors: Hollande Etienne , Di Donato Jean‐He´le`ne Levrat , Fanjul Marjorie , Palevody Claude , Daumas Marguerite , Puech Jacques , Ratovo Ginette
Published: August 2004

Summary— Human pancreatic cells of the Capan‐1 line form domes in culture during the stationary growth stage. The domes are thought to be a result of the transport of water and electrolytes by the Capan‐1 cells. In older Capan‐1 cultures, the epithelial sheets formed thickenings from several layers of cells of which the outermost ones were joined by tight type junctions. In the intracellular space, deposits of insoluble calcium salts were observed. Culture of Capan‐1 cells in the presence of fibroblasts prolonged survival of the cultures with intact domes for more than 80 days. The Capan‐1 cells proliferated forming multilayers and closed cavities which we called super‐domes. X‐ray spectrometry and electron diffraction analysis showed that the abundant deposits inside these cavities consisted of calcium phosphate in an apatite structure. The number of these deposits increased with time in culture, and they appeared to be formed at the sites of contact with an extracellular matrix consisting of cell debris. Deposits were not observed within the culture medium. Cells from domes were stained cytochemically for ATPases and alkaline phosphatases and examined by light and electron microscopy. The Capan‐1 cells surrounding the domes were differentiated, polarized cells containing placental type alkaline phosphatases on their apical membranes and Ca2+‐ATPases on their basolateral membranes. These enzymes were thought to play a role in the accumulation of phosphate and Ca2+ ions in the dome cavities, which then formed crystals in the presence of organic compounds produced by lysis of cells of the deepest layers of the super‐domes. The crystals of hydroxyapatite observed in standard Capan‐1 cell cultures and those cocultured with fibroblasts were assumed to be a result of transepithelial transport of Ca2+ and phosphate ions by these cells.
In vitro induction of early mouse embryo intracisternal particles (ε particles) in cultured cell lines
Authors: Lasneret Jacqueline , Dianoux Laurent , Lesser Joe¨l , Peries Jorge , Canivet Martine
Published: August 2004

Summary— The experimental induction of ε particles, retrovirus‐like structures corresponding to the small IA particles of the mouse, was studied by electron microscopy in rodent‐cultured cell lines. Among the chemicals tested, only IdUr was shown to be an effective inducer, but not cycloheximide, puromycine, deoxy‐fluorouracile or 5‐azacytine. However, only two mouse‐derived cell lines: KiBALB and FG 10, among 27 cell lines of mouse, rat and mink origins tested, expressed ε particles upon IdUr treatment. Epsilon particles thus respond to chemical inducers very differently in comparison with large IAP. Moreover, the addition of interferon previously shown to attenuate IAP production, had no effect on that of ε particles.
Acid secretion and membrane reorganization in single gastric parietal cell in primary culture
Authors: Mangeat Paul , Gusdinar Tutus , Sahuquet Alain , Hanzel David K , Forte John G , Magous Richard
Published: August 2004

Summary— A digitally‐enhanced videomicroscopy study of rabbit gastric parietal cells in primary culture was performed using alternate observations with differential interference contrast and fluorescence optics of cells mounted and perfused on a temperature‐controlled microscope stage. The effect of histamine, a physiological effector of acid secretion, was followed. Isolated parietal cells possess an internal apical vacuole, which kept the cell in a pseudopolarized state. This apical vacuole is a site of acid secretion. This was demonstrated by the direct visualization of the uptake of the fluorescent weak base 9‐amino acridine and of the concomitant enormous swelling of the acid vacuole which reached an estimated size of 3–7 times the normal cell volume. This morphological change of shape and acidification of apical vacuoles was fully reversible and cells could respond to successive stimulations. A quantitative study of these events provided a value of the acid accumulation index for each single cell in response to histamine. Individual cell response varied within a factor of 7. The cellular localization of the proton pump complex responsible for acid secretion and of the major components of the secretory microvilli, actin and ezrin, a histamine‐dependent phosphorylation target of protein kinase A, were detected by indirect immunofluorescence microscopy in resting and stimulated cells. Both actin and ezrin colocalized at the apical vacuole membrane in resting and stimulated cells, whereas the proton pump shifted from an intracytoplasmic pool to the apical vacuole membrane upon stimulation.
Protein phosphatase activities in vivo in Xenopus laevis oocyte: Inhibition by okadaic acid
Authors: Cormier Patrick , Mulner‐Lorillon Odile , Ozon Rene´ , Belle´ Robert
Published: August 2004

Summary— Protein phosphatase activities were analyzed in vivo in Xenopus oocytes. The dephosphorylation of microinjected β casein was inhibited when the tumor promoter okadaic acid was microinjected into oocytes. Inhibition was dose dependent and reversible; 50 % of activity was recovered 15–30 minutes after microinjection.
Sequence analysis of cell cycle control (cdc2) protein kinases among protein serine/threonine kinases
Authors: Guerrucci Marie‐Anne , Soldano Henri , Bellé Robert
Published: August 2004

Summary—

Among protein serine/threonine kinases, the CDC2 proteins are both well characterized as protein serine/threonine kinases and are functionally involved in the control of cell division. Protein serine/threonine kinase sequences were analysed using Fourier transform of the coded sequences. Characteristic code/frequency pairs were extracted from a set of well defined protein serine/threonine kinases. The characteristic frequencies 0.179, 0.250 and 0.408 distinguished protein serine/threonine kinases from proteins which did not have the biological activity. Pertinent patterns in the sequence, responsible for the code/frequency pairs detection were searched and found to be correlated with the putative catalytic domain of the proteins.

Protein serine/threonine kinases involved in cell division control, CDC2 protein kinases, were compared to the other protein serine/threonine kinases. Specific code/frequency pairs were extracted from the sequences and could be related to the function or regulation of the kinases in cell division. Two CDC2 related proteins CDC2(Mm) from mice and CDC2(Gg) from chicken were shown to fit well with the CDC2 proteins, whereas KIN28, PHO85 and PSKJ3, which skare sequence homology but not functional activity with the CDC2 proteins, were clearly excluded from the CDC2 proteins by the characteristic code/frequency pairs. Pertinent patterns in the CDC2 proteins were analysed and mapped on the CDC2 related protein sequences. Four patterns were correlated with the code/frequency detection and therefore, could be associated to the regulation of the CDC2‐related proteins.