bluuebunny/crossref_metadata_2025 huggingface dataset:
PYP‐1, inorganic pyrophosphatase, is required for larval development and intestinal function in C. elegans
Authors: Ko Kyung Min , Lee Wonhae , Yu Jae-Ran , Ahnn Joohong
Published: November 2007

Inorganic pyrophosphatase (PPase) catalyzes the hydrolysis of inorganic pyrophosphate (PPi) into phosphate (Pi), which provides a thermodynamic driving force for important biosynthetic reactions. The nematode Caenorhabditis elegans gene C47E12.4 encodes a PPase (PYP‐1) which shows 54% amino acid identity with human PPase. PYP‐1 exhibits specific enzyme activity and is mainly expressed in the intestinal and nervous system. A null mutant of pyp‐1 reveals a developmental arrest at early larval stages and exhibits gross defects in intestinal morphology and function. The larval arrest phenotype was successfully rescued by reintroduction of the pyp‐1 gene, suggesting that PYP‐1 is required for larval development and intestinal function in C. elegans.
A Stargardt disease‐3 mutation in the mouse Elovl4 gene causes retinal deficiency of C32–C36 acyl phosphatidylcholines
Authors: McMahon Anne , Jackson Shelley N. , Woods Amina S. , Kedzierski Wojciech
Published: November 2007

Stargardt disease‐3 (STGD3) is a juvenile dominant macular degeneration caused by mutations in elongase of very long chain fatty acid‐4. All identified mutations produce a truncated protein which lacks a motif for protein retention in endoplasmic reticulum, the site of fatty acid synthesis. In these studies of Stgd3‐knockin mice carrying a human pathogenic mutation, we examined two potential pathogenic mechanisms: truncated protein‐induced cellular stress and lipid product deficiency. Analysis of mutant retinas detected no cellular stress but demonstrated selective deficiency of C32–C36 acyl phosphatidylcholines. We conclude that this deficit leads to the human STGD3 pathology.
Anti‐obesity and anti‐tumor pro‐apoptotic peptides are sufficient to cause release of cytochrome c from vesicles
Authors: Sandoval Cristina M. , Salzameda Bridget , Reyes Kristine , Williams Taylor , Hohman Valerie S. , Plesniak Leigh A.
Published: November 2007

Peptides that target tissue for an apoptotic death have potential as therapeutics in a variety of disease conditions. The class of peptides described herein enters the cell through a specific receptor‐mediated interaction. Once inside the cell, the peptide migrates toward the mitochondria, where the membrane barrier is disrupted. These experiments demonstrate that upon treatment with these short peptides large unilamellar vesicles are not lysed, a graded mode of leakage is observed and the transient pores formed by these peptides are large enough to release entrapped cytochrome c from the vesicles.
Intracellular organelles and calcium homeostasis in rods and cones
Authors: SZIKRA TAMAS , KRIŽAJ DAVID
Published: November 2007

The role of intracellular organelles in Ca2+homeostasis was studied in salamander rod and cone photoreceptors under conditions that simulate photoreceptor activation by darkness and light. Sustained depolarization evoked a Ca2+gradient between the cell body and ellipsoid regions of the inner segment (IS). The standing pattern of calcium fluxes was created by interactions between the plasma membrane, endoplasmic reticulum (ER), and mitochondria. Pharmacological experiments suggested that mitochondria modulate both baseline [Ca2+]i in hyperpolarized cells as well as kinetics of Ca2+entry via L type Ca2+channels in cell bodies and ellipsoids of depolarized rods and cones. Inhibition of mitochondrial Ca2+sequestration by antimycin/oligomycin caused a three-fold reduction in the amount of Ca2+accumulated into intracellular organelles in both cell bodies and ellipsoids. A further 50% decrease in intracellular Ca2+content within cell bodies, but not ellipsoids, was observed after suppression of SERCA-mediated Ca2+uptake into the ER. Inhibition of Ca2+sequestration into the endoplasmic reticulum by thapsigargin or cyclopiazonic acid decreased the magnitude and kinetics of depolarization-evoked Ca2+signals in cell bodies of rods and cones and decreased the amount of Ca2+accumulated into internal stores. These results suggest that steady-state [Ca2+]i in photoreceptors is regulated in a region-specific manner, with the ER contribution predominant in the cell body and mitochondrial buffering [Ca2+] the ellipsoid. Local [Ca2+]i levels are set by interactions between the plasma membrane Ca2+channels and transporters, ER and mitochondria. Mitochondria are likely to play an essential role in temporal and spatial buffering of photoreceptor Ca2+.
Applying Circular Statistics to the Analysis of Monitoring Data
Authors: Kubiak Thomas , Jonas Cornelia
Published: November 2007

Abstract. Patterns of psychological variables in time have been of interest to research from the beginning. This is particularly true for ambulatory monitoring research, where large (cross-sectional) time-series datasets are often the matter of investigation. Common methods for identifying cyclic variations include spectral analyses of time-series data or time-domain based strategies, which also allow for modeling cyclic components. Though the prerequisites of these sophisticated procedures, such as interval-scaled time-series variables, are seldom met, their usage is common. In contrast to the time-series approach, methods from a different field of statistics, directional or circular statistics, offer another opportunity for the detection of patterns in time, where fewer prerequisites have to be met. These approaches are commonly used in biology or geostatistics. They offer a wide range of analytical strategies to examine “circular data,” i.e., data where period of measurement is rotationally invariant (e.g., directions on the compass or daily hours ranging from 0 to 24, 24 being the same as 0). In psychology, however, circular statistics are hardly known at all. In the present paper, we intend to give a succinct introduction into the rationale of circular statistics and describe how this approach can be used for the detection of patterns in time, contrasting it with time-series analysis. We report data from a monitoring study, where mood and social interactions were assessed for 4 weeks in order to illustrate the use of circular statistics. Both the results of periodogram analyses and circular statistics-based results are reported. Advantages and possible pitfalls of the circular statistics approach are highlighted concluding that ambulatory assessment research can benefit from strategies borrowed from circular statistics.
An Empirical Analysis of the Obtrusiveness of and Participants' Compliance with the Electronically Activated Recorder (EAR)
Authors: Mehl Matthias R. , Holleran Shannon E.
Published: November 2007

Abstract. In this article, the authors provide an empirical analysis of the obtrusiveness of and participants' compliance with a relatively new psychological ambulatory assessment method, called the electronically activated recorder or EAR. The EAR is a modified portable audio-recorder that periodically records snippets of ambient sounds from participants' daily environments. In tracking moment-to-moment ambient sounds, the EAR yields an acoustic log of a person's day as it unfolds. As a naturalistic observation sampling method, it provides an observer's account of daily life and is optimized for the assessment of audible aspects of participants' naturally-occurring social behaviors and interactions. Measures of self-reported and behaviorally-assessed EAR obtrusiveness and compliance were analyzed in two samples. After an initial 2-h period of relative obtrusiveness, participants habituated to wearing the EAR and perceived it as fairly unobtrusive both in a short-term (2 days, N = 96) and a longer-term (10-11 days, N = 11) monitoring. Compliance with the method was high both during the short-term and longer-term monitoring. Somewhat reduced compliance was identified over the weekend; this effect appears to be specific to student populations. Important privacy and data confidentiality considerations around the EAR method are discussed.
The 16,000‐event 2003–2004 earthquake swarm in Ubaye (French Alps)
Authors: Jenatton Liliane , Guiguet Robert , Thouvenot François , Daix Nicolas
Published: November 2007

The Ubaye valley, one of the most active seismic zones in the French Alps, was visited in 2003–2004 by a prolific and protracted earthquake swarm with a maximum magnitude ML = 2.7. The seismic activity clustered along a 9‐km‐long, 3‐ to 8‐km‐deep rupture zone which trends NW‐SE across the valley and dips 80°SW. Focal mechanisms for the larger shocks show either normal faulting with a SW‐NE trending extension direction or NW‐SE strike slip with right lateral displacement. The activity initiated in the central part of the rupture zone, diffused to its periphery, and eventually concentrated in its southeastern deeper part. A permanent station situated above the swarm allowed us to monitor the entire phenomenon from its inception to its conclusion. The complete time series includes more than 16,000 events, with shocks down to magnitude ML = −1.3. It shows bursts of activity, separated by quiescent periods, with no well‐defined subswarms as observed in other similar studies. The Gutenberg‐Richter b value significantly varied between 1.0 and 1.5 in the course of the phenomenon.
Modeling study of ozone seasonal cycle in lower troposphere over east Asia
Authors: Li Jie , Wang Zifa , Akimoto Hajime , Gao Chao , Pochanart Pakpong , Wang Xiquan
Published: November 2007

On the basis of three mountain sites (Mount Tai, Hua and Huang) newly founded in east‐central China and several other sites from the Acid Deposition Monitoring Network in east Asia (EANET) and WMO World Data Centre for Greenhouse Gases (WDCGG), we investigate seasonal cycle of ozone over east Asia and its budgets in east‐central China by using a regional chemical transport model (NAQPMS). The observations show a striking ozone pattern of two sharp peaks in May‐June and September–October at three mountain sites in east‐central China which are higher than those observed at other mountain sites in Europe and North America. Ozone budgets analysis by the model confirms that maximum of net photochemical productions reaches 31.8, 15.1, and 11.4 ppbv/d at Mount Tai, Hua, and Huang, respectively. The net photochemical production dominates the formation of ozone maximums at Mount Tai and Hua in June, and the importing transport also plays a comparable importance at Mount Huang. In comparison with those in the western North Pacific, east‐central China shows stronger net photochemical productions, which are comparable to anthropogenic sources regions in Europe and North America.
Relative sensitivity of the Atlantic meridional overturning circulation to river discharge into Hudson Bay and the Arctic Ocean
Authors: Rennermalm Asa K. , Wood Eric F. , Weaver Andrew J. , Eby Michael , Déry Stephen J.
Published: November 2007

Increases in high‐latitude river discharge over the 20th century and projected continued increases during the 21st century may have an impact on the Atlantic meridional overturning circulation (AMOC), which could feed back to regional and global climate. Although the general trend in high‐latitude river discharge is positive, there is important geographical spread in the trends. While Eurasian rivers draining into the Arctic Ocean show positive trends over the 20th century, rivers draining into Hudson Bay show negative trends since 1964. Here the sensitivity of AMOC to changes in river discharge into Hudson Bay and the Arctic Ocean is studied with an intermediate‐complexity Earth system model. It is found that ocean freshening originating from Arctic rivers is more effective in slowing down the AMOC than freshening originating from Hudson Bay rivers, given the same magnitude of freshening in both regions. The lesser impact of Hudson Bay river discharge on AMOC is the result of a buildup of freshwater anomalies in the Labrador Sea affecting the northward flow of the Gulf Stream. This work highlights that not only the freshening magnitude but the region where this freshening takes place is crucial for the AMOC response to altered river discharge climatology.
Late Cenozoic metamorphic evolution and exhumation of Taiwan
Authors: Beyssac Olivier , Simoes Martine , Avouac Jean Philippe , Farley Kenneth A. , Chen Yue‐Gau , Chan Yu‐Chang , Goffé Bruno
Published: November 2007

The Taiwan mountain belt is composed of a Cenozoic slate belt (Hsuehshan Range units, HR, and Backbone Slates, BS) and of accreted polymetamorphic basement rocks (Tananao Complex, TC). Ongoing crustal shortening has resulted from the collision between the Chinese continental margin and the Luzon volcanic arc, which initiated ∼6.5 Ma ago. The grade and age of metamorphism and exhumation are a key record of the development of the orogenic wedge. Because the Taiwan mountain belt is mostly composed by accreted sediments lacking metamorphic index minerals, quantitative constraints on metamorphism are sparse. By contrast, these rocks are rich in carbonaceaous material (CM) and are therefore particularly appropriate for RSCM (Raman Spectroscopy of CM) thermometry. We apply this technique in addition to (U‐Th)/He thermochronology on detrital zircons to assess peak metamorphic temperatures (T) and the late exhumational history respectively, along different transects in central and southern Taiwan. In the case of the HR units, we find evidence for high metamorphic T of at least 340°–350°C and locally up to 475°C, and for relative rapid exhumation with zircon (U‐Th)/He ages in the range of 1.5–2 Ma. Farther east, the BS were only slightly metamorphosed (T < 330 °C), and zircons are not reset for (U‐Th)/He. From the eastern BS to the inner TC schists, T gradually increases from ∼350°C up to ∼500°C following an inverted metamorphic gradient. Available geochronological constraints and the continuous thermal gradient from the BS to the basement rocks of the TC suggest that the high RSCM T of the TC were most probably acquired during the last orogeny, and were not inherited from a previous thermal event. Zircons yield (U‐Th)/He ages of ∼0.5–1.2 Ma. Peak metamorphic T and the timing of exhumation do not show along‐strike variations over the TC in the studied area. In contrast, exhumation is laterally diachronous and decreases southward in the case of the HR units. In particular, our data imply that the HR units have been exhumed by a minimum of 15 km over the last few Ma. In the case of the BS, they show far less cumulated exhumation and much slower cooling rates. We propose that most of the deformation and exhumation of the Taiwan mountain belt is sustained through two underplating windows located beneath the Hsuehshan Range and the TC. Our data show significant departures from the predictions of the prevailing model in Taiwan, which assumes a homogeneous critical wedge with dominant frontal accretion. Our study sheds new light on how the mountain belt has grown as a possible result of underplating mostly.
Estimating the performance of international regulatory regimes: Methodology and empirical application to international water management in the Naryn/Syr Darya basin
Authors: Siegfried Tobias , Bernauer Thomas
Published: November 2007

We develop a methodology for estimating the performance of international regulatory regimes in the context of transboundary surface waters. Our performance metric relies on assessments, over time, of actual performance, counterfactual performance, and optimal performance. The metric is of practical relevance as a diagnostic tool for policy evaluation. Thus it provides a starting point for policy improvement. To demonstrate the empirical relevance of this methodology we examine international water management in the Naryn/Syr Darya basin, a major international river system in Central Asia. The emphasis is on the Toktogul reservoir, the main reservoir in the Naryn/Syr Darya basin, and its downstream effects. The biggest policy challenge in this case has been to design and implement international trade‐offs among water releases for upstream hydropower production in winter and water releases for downstream irrigation in summer. We find that the international regime in place since 1998 is characterized by low average performance and high variability.
Isotopic fractionation of methane and ethane hydrates between gas and hydrate phases
Authors: Hachikubo Akihiro , Kosaka Tomoko , Kida Masato , Krylov Alexey , Sakagami Hirotoshi , Minami Hirotsugu , Takahashi Nobuo , Shoji Hitoshi
Published: November 2007

Isotopic fractionation of carbon and hydrogen in methane and ethane during the formation of gas hydrates was investigated. The gas hydrate samples were experimentally prepared in a pressure cell and isotopic compositions of both residual and hydrate‐bound gases were measured. δD of hydrate‐bound molecules of methane and ethane hydrates was several per mil lower than that of residual gas molecules in the formation processes, while there was no difference in the case of δ13C. These isotopic differences in δD are enough small for discussing the source types of hydrate‐bound gases using the δ13C‐δD diagram of Whiticar et al. [1986]. These results may provide useful insight into the formation process of gas hydrates.
Anti‐Atlas Moroccan Chain as the source of lithogenic‐derived micronutrient fluxes to the deep Northeast Atlantic Ocean
Authors: Chavagnac V. , Waniek J. J. , Atkin D. , Milton J. A. , Leipe T. , Green D. R. H. , Bahlo R. , Hayes T. E. F. , Schulz‐Bull D. E.
Published: November 2007

Identifying the source of atmospheric dust is crucial to better understand the global marine biogeochemical cycle as phytoplankton growth relies on dissolved micronutrient elements available in the open ocean. Mineralogical, geochemical and Sr isotope analyses of a one year‐time series (April 2003–April 2004) of deep ocean particle flux at the Madeira Abyssal Plain in the subtropical northeast Atlantic are presented. The lithogenic fraction has a high occurrence of palygorskite and smectite and an absence of kaolinite together with Sr isotopic compositions similar to the Northeast Atlantic aerosols. This indicates the Anti‐Atlas Moroccan chain of Paleozoïc age as the source region. The lithogenic fraction supplies 4 times more Fe during a dust event than during the spring‐summer aeolian input. A continuous input of only 1% of the lithogenic iron made available over a year period, could lead to an increase in primary productivity of 40% relative to today's value.
Climatic control on the peak discharge of glacier outburst floods
Authors: Ng Felix , Liu Shiyin , Mavlyudov Bulat , Wang Yanguo
Published: November 2007

Lakes impounded by natural ice dams occur in many glacier regions. Their sudden emptying along subglacial paths can unleash ∼1 km3 of floodwater, but predicting the peak discharge of these subglacial outburst floods (‘jökulhlaups’) is notoriously difficult. To study how environmental factors control jökulhlaup magnitude, we use thermo‐mechanical modelling to interpret a 40‐year flood record from Merzbacher Lake in the Tian Shan. We show that the mean air temperature during each flood modulates its peak discharge, by influencing both the rate of meltwater input to the lake as it drains, and the lake‐water temperature. The flood devastation potential thus depends sensitively on weather, and this dependence explains how regional climatic warming drives the rising trend of peak discharges in our dataset. For other subaerial ice‐dammed lakes worldwide, regional warming will also promote higher‐impact jökulhlaups by raising the likelihood of warm weather during their occurrence, unless other factors reduce lake volumes at flood initiation to outweigh this effect.
Impacts of large‐scale oscillations on pan‐Arctic terrestrial net primary production
Authors: Zhang Ke , Kimball John S. , McDonald Kyle C. , Cassano John J. , Running Steven W.
Published: November 2007

Analyses of regional climate oscillations and satellite remote sensing derived net primary production (NPP) and growing season dynamics for the pan‐Arctic region indicate that the oscillations influence NPP by regulating seasonal patterns of low temperature and moisture constraints to photosynthesis. Early‐spring (Feb–Apr) patterns of the Arctic Oscillation (AO) are proportional to growing season onset (r = −0.653; P = 0.001), while growing season patterns of the Pacific Decadal Oscillation (PDO) are proportional to plant‐available moisture constraints to NPP (Im) (r = −0.471; P = 0.023). Relatively strong, negative PDO phases from 1988–1991 and 1998–2002 coincided with prolonged regional droughts indicated by a standardized moisture stress index. These severe droughts resulted in widespread reductions in NPP, especially for relatively drought prone boreal forest and grassland/cropland ecosystems. The influence of AO and PDO patterns on northern vegetation productivity appears to be decreasing and increasing, respectively, as low temperature constraints to plant growth relax and NPP becomes increasingly limited by available water supply under a warming climate.
Rotational stability of tidally deformed planetary bodies
Authors: Matsuyama I. , Nimmo F.
Published: November 2007

We consider the true polar wander (rotational variations driven by mass redistribution) of tidally deformed planetary bodies. The rotation pole of bodies without tidal deformation is stabilized by the component of the rotational bulge which retains a memory for prior rotational states, that is, a remnant rotational bulge. For planetary bodies with tidal deformation, the additional stabilizing effect of a remnant tidal bulge results in less permissive excursions of the rotation pole. The magnitude of the load driving reorientation is parameterized by Q, the ratio between the degree‐2 gravitational potential of the load and the remnant rotational bulge. Reorientation is favored if the initial load longitude is close to 90°, that is, close to the center of the leading or trailing hemisphere. As an illustration of the new theory, we consider reorientation driven by internal loading on Saturn's moon Enceladus. Small loads (∣Q∣ ≈ 1) are inconsistent with significant reorientation because of the small present‐day angular separation between the load and the rotation axis. Larger loads (∣Q∣ ≈ 2) permit reorientations approaching 90°. Large reorientation scenarios are consistent with the present‐day equatorial location of a geologically inferred ancient polar terrain.
Murine thymic stromal lymphopoietin promotes the differentiation of regulatory T cells from thymic CD4+CD8−CD25− naïve cells in a dendritic cell‐independent manner
Authors: Lee June‐Yong , Lim Yu‐Mi , Park Min‐Jung , Min So‐Youn , Cho Mi‐La , Sung Young‐Chul , Park Se‐Ho , Kim Ho‐Youn , Cho Young‐Gyu
Published: November 2007

Human thymic stromal lymphopoietin (TSLP) activates dendritic cells (DCs), which promote the proliferation and differentiation of CD4+ T cells. However, murine TSLP (mTSLP) can act directly on CD4+ T cells and bring about their differentiation. We studied the role of mTSLP in the generation of CD4+CD25+FoxP3+ T cells from thymocytes. mTSLP promoted the differentiation of CD4+ single‐positive thymocytes into CD4+CD25+FoxP3+ T cells. Although we cannot exclude an effect of TSLP mediated through DCs due to co‐stimulatory effects, mTSLP appears to act directly on thymocytes. T‐cell receptor and TSLP receptor signaling act synergistically on thymocytes to generate CD4+CD25+FoxP3+ T cells. mTSLP may play an important role in maintaining immune tolerance by promoting the conversion of thymocytes into natural regulatory T cells via escape from negative selection.
Recombinant antimicrobial peptides efficiently produced using novel cloning and purification processes
Authors: Metlitskaia Luba , Cabralda Jennifer E. , Suleman Dinar , Kerry Cynthia , Brinkman Jacquelyn , Bartfeld Daniel , Guarna M. Marta
Published: November 2007

Endogenous antimicrobial peptides are ubiquitous components of animal and plant host defences. These peptides, usually cationic and amphipathic, kill target cells rapidly and are efficacious against antibiotic‐resistant and clinically relevant pathogens. A practical challenge in the development of cationic peptides as therapeutics is to meet the production requirements for large quantities of highly purified drug substance at competitive costs. While chemical peptide synthesis can be used to manufacture cationic peptides, we have developed cost‐effective methods for recombinant production by expressing fusion proteins comprised of multiple copies of the peptides. The fusion proteins accumulate in Escherichia coli inclusion bodies and constitute over 50% of the total cellular proteins. Active antimicrobial peptides are released by chemical reagents and purified by chromatography, combining both standard and novel approaches. Challenges of industrial‐scale manufacturing of therapeutics were considered in the development of this process.
Biotransformation of geraniol by Rhodococcus sp. strain GR3
Authors: Chatterjee Tanaya
Published: November 2007

Microbial degradation of geraniol, a natural monoterpene alcohol, was studied using a Rhodococcus sp. strain GR3 isolated from soil. The bioconversion product was identified as geranic acid [(2E)‐3,7‐dimethylocta‐2,6‐dienoic acid] and its structure was established by 1H‐NMR, Fourier‐transform IR spectrometry and GC‐MS. The optimum temperature for this bioconversion was found to be 30 °C, and the reaction proceeds to a saturation with a time constant of 12.5 h. No appreciable degradation of product was observed using this bacterium.
Inactivation of the bovine‐spongiform‐encephalopathy (BSE) agent by the acid and alkaline processes used in the manufacture of bone gelatine
Authors: Grobben Adrianus H. , Steele Phillip J. , Somerville Robert A. , Taylor David M.
Published: November 2007

A validation study was carried out to determine the capacity of the traditional acid and alkaline processes used in the manufacture of bovine bone gelatine to remove and/or inactivate the transmissible agent that causes BSE (bovine spongiform encephalopathy). Using an accurately scaled down laboratory process that precisely mimicked the minimum conditions of the industrial processes, gelatine (gelatin) was manufactured from industrial starting material that had been spiked with mouse brain infected with the 301V strain of mouse‐passaged BSE agent. Clearance factors were determined by titrating the infectivity levels of the infected mouse brain tissue, the gelatine extracts, and the final sterilized gelatine solution. The infectivity level of the spiked starting material was 108.4 mouse intracerebral ID50/kg (ID50 is the dose at which half of the challenged animals were infected). Clearance factors of 102.6 and 103.7 ID50 were demonstrated for the first stages of the acid and alkaline processes respectively during which the bones are converted to crude gelatine. It was further demonstrated that the complete acid and alkaline processes both reduced infectivity to undetectable levels, giving clearance factors of 104.8 ID50 for the acid process, and 104.9 ID50 for the alkaline process.
Modified alginate and chitosan for lactic acid bacteria immobilization
Authors: Le‐Tien Canh , Millette Mathieu , Mateescu Mircea‐Alexandru , Lacroix Monique
Published: November 2007

Beads with enhanced‐stability acid media, which were based on alginate and chitosan functionalized by succinylation (increasing the anionic charges able to retain protons) or by acylation (improving matrix hydrophobicity), were developed for immobilization of bacterial cells. Beads (3 mm diameter) formed by ionotropic gelation with CaCl2 presented good mechanical characteristics. After 30 min incubation of viable free Lactobacillus rhamnosus cells in simulated gastric fluid (pH 1.5), we noticed that the level of viable bacteria was undetectable. Bacterial immobilization in native‐alginate‐based beads generated a viable‐cell count of 22–26%, whereas, when entrapped in succinylated alginate and chitosan beads, the percentage of viable cells was of 60 and 66%, respectively. Best viability (87%) was found for bacteria immobilized in N‐palmitoylaminoethyl alginate, which affords a high protective effect, probably due to long alkyl pendants that improve the beads' hydrophobicity, limiting hydration in the acidic environment.
Development of a highly efficient system for assessing recombinant gene expression in plant cell suspensions via Agrobacterium tumefaciens transformation
Authors: Fuentes Alejandro , Ramos Pedro Luis , Ayra Camilo , Rodríguez Meilyn , Ramírez Nadia , Pujol Merardo
Published: November 2007

A transient gene‐expression system was developed and used to characterize promoter strength, to verify suitability of bacterial gene modifications for expression in plant cells, and to express active antibody molecules. The system is based on suspension tobacco cells transformed by Agrobacterium in a transient way. Conditions such as pre‐culture of tobacco cells and the co‐cultivation period were identified as determinants to achieve high expression levels. Under established conditions the activity strength of CaMV (cauliflower mosaic virus) 35 S and ToMoTV (tomato mottle taino virus) AL1 promoters were compared. A modified cry gene sequence from Bacillus thuringiensis was expressed and detected by Western‐blot analysis. A monoclonal antibody against anti‐(hepatitis B virus surface antigen) was produced in such quantities as to allow testing of biological activity and preliminary characterization.
Optimization of penicillin G acylase multipoint immobilization on to glutaraldehyde–chitosan beads
Authors: Adriano Wellington S. , Filho Edilson H. C. , Silva James A. , Gonçalves Luciana R. B.
Published: November 2007

The objective of this work was to study the immobilization of penicillin G acylase from Escherichia coli on to chitosan–glutaraldehyde beads by multipoint covalent binding. This process was optimized using a 23 experimental design. The parameters selected for the present study were the concentrations of glutaraldehyde, phenylacetic acid and sodium borohydride. Three responses were chosen, namely immobilization yield and stabilization factors of enzyme derivatives at high temperature and at alkaline pH. All the runs at the maximum (+1) and minimum (−1) levels were performed at random. Three experiments were performed at the centre point, coded as zero, for experimental‐error estimation. With respect to immobilization yield, the main effectors were the concentrations of glutaraldehyde and phenylacetic acid. For stabilization factors at 50°C and at alkaline pH, the main effectors were the concentrations of glutaraldehyde and sodium borohydride and the interaction between them.
A detailed understanding of the enhanced hypothermic productivity of interferon‐γ by Chinese‐hamster ovary cells
Authors: Fox Stephen R. , Tan Hong Kiat , Tan Mei Chee , Wong S. C. Niki C. , Yap Miranda G. S. , Wang Daniel I. C.
Published: November 2007

Culturing CHO (Chinese‐hamster ovary) cells at low temperature leads to growth arrest in the G0/G1 phase of the cell cycle and, in many cases, causes an increase in the specific productivity of recombinant protein. Controlled proliferation is often used to increase CHO specific productivity, and thus there is speculation that the enhanced productivity at low temperature is due to G0/G1‐phase growth arrest. However, we show that the positive effect of low temperature on recombinant protein production is due to elevated mRNA levels and not due to growth arrest and that a cell line can still exhibit growth‐associated productivity at low temperatures. Using a CHO cell producing recombinant human IFN‐γ (interferon‐γ), we show that productivity increases as the percentage of cells in the S phase of the cell cycle increases, at both 32 and 37°C. The increased productivity is due to higher recombinant IFN‐γ mRNA levels. We also show that, for a given cell‐cycle distribution, specific productivity increases as the temperature is lowered from 37 to 32°C. Thus specific productivity is maximized when cells are actively growing (high percentage of S‐phase cells) and also exposed to low temperature. These findings have important implications for cell‐culture optimization and cell‐line engineering, providing evidence that a CHO cell line capable of actively growing at low temperature would provide improved total protein production relative to the current growth strategies, namely 37°C active growth or low‐temperature growth arrest.
Active hypothermic growth: a novel means for increasing total interferon‐γ production by Chinese‐hamster ovary cells
Authors: Fox Stephen R. , Yap Mei Xia , Yap Miranda G. S. , Wang Daniel I. C.
Published: November 2007

When grown under hypothermic conditions, CHO (Chinese‐hamster ovary) cells become growth‐arrested in the G0/G1 phase of the cell cycle and also often exhibit increased recombinant‐protein production. We have shown in the accompanying paper [Fox, Tan, Tan, Wong, Yap and Wang (2005) Biotechnol. Appl. Biochem. 41, 255–264] that the positive effect of low temperature on recombinant‐protein production is due to elevated mRNA levels and not due to G0/G1‐phase growth arrest and that a cell line can still show growth‐associated productivity at low temperature. This finding led to the hypothesis that improved total production of recombinant protein would be achieved by stimulating cells to actively grow at low temperature, a culture condition previously unreported in the literature. In the present study we have validated this hypothesis by stimulating hypothermic (32°C) growth through the use of different growth factors. Hypothermic growth was stimulated in fetal‐bovine‐serum‐supplemented adherent cultures using basic fibroblast growth factor or insulin. Hypothermic growth was also stimulated in suspension cultures normally grown in protein‐free medium by using supplementation with fetal bovine serum. These methods resulted in up to 7.7‐ and 4.9‐fold increases in total recombinant‐protein production compared with the 37 and 32°C control cultures respectively. This proof‐of‐concept study will motivate the creation of cell lines capable of growing at low temperatures for use in industrial processes.
Characterization of different strains of poliovirus and influenza virus by differential scanning calorimetry
Authors: Krell Tino , Manin Catherine , Nicolaï Marie‐Claire , Pierre‐Justin Catherine , Bérard Yves , Brass Olivier , Gérentes Lionel , Leung‐Tack Patricia , Chevalier Michel
Published: November 2007

Vaccines against poliomyelitis and influenza contain inactivated forms of poliovirus and influenza virus. These antigens are generated on an industrial scale from the purified active viruses that have been analysed in this study by DSC (differential scanning calorimetry). Multiple unfolding transitions are seen for influenza virus A/New Caledonia/20/99 (H1N1), A/Panama/2007/99 (H3N2) and B/Shangdong/7/97. These data, combined with previously reported data on other influenza viruses, indicates that each influenza virus strain has a characteristic unfolding behaviour. Only minor changes were seen in the thermogram of βPL (β‐propiolactone)‐inactivated influenza virus, which is consistent with the proposition that βPL reacts mainly with the nucleotide fraction of the virus. We demonstrate that a peak annotation of the thermogram of the native virus is possible using bromelain‐treated virus and virosomes. At pH 1.5–2.5, poliovirus of type I unfolds in a single unfolding event with respective Tm (midpoint of protein unfolding transition) values between 34 and 45 °C. At pH 2, polioviruses of type II unfold equally in a single event, but, compared with the type I virus, with a Tm value increased by 3.7 °C. At neutral pH, the DSC thermogram of type I poliovirus was very ‘noisy’. Data obtained offer the possibility of precisely characterizing and identifying different viral strains.
Chemical glycosidation of trypsin with O‐carboxymethyl‐poly‐β‐cyclodextrin: catalytic and stability properties
Authors: Villalonga Maria L. , Reyes Gretel , Fragoso Alex , Cao Roberto , Fernández Leyden , Villalonga Reynaldo
Published: November 2007

The polysaccharide O‐carboxymethyl‐poly‐β‐cyclodextrin was synthesized (molecular mass 13000 Da, 40% carboxy groups) and attached to the surface of bovine pancreatic trypsin. The resulting neoglycoenzyme retained high proteolytic and esterolytic activity and contained approx. 1.0 mol of polymer/mol of enzyme. The optimum temperature for trypsin activity was increased by 10°C after this transformation. Thermostability of the polymer–enzyme complex was increased by about 14°C over 10 min incubation. The conjugate was also more resistant to thermal inactivation at different temperatures, ranging from 45 to 60°C, demonstrating the influence of supramolecular and polymer–protein electrostatic interactions on trypsin thermostabilization. Additionally, the conjugate was 36‐fold more resistant to the action of the anionic surfactant SDS. This modification also protected the enzyme from autolysis at alkaline pH.
Artificial nerve conduits in peripheral‐nerve repair
Authors: Kannan Ruben Y. , Salacinski Henryk J. , Butler Peter E. M. , Seifalian Alexander M.
Published: November 2007

Injuries to the nervous system are the result of mechanical, thermal, chemical or congenital pathologies and, if function is not restored, they lead to loss of muscle function, pain and impaired sensation. Current treatment modalities essentially coapt the two nerves ends together or place a nerve graft between the cut ends. However, clinical results have never been optimal, and therefore a quest for better options has taken place. In this review article we look at the synthetic and biomimetic options currently being tested as potential nerve grafts.
Electrochemical studies on polysorbate‐20 (Tween 20)‐entrapped haemoglobin and its application in a hydrogen peroxide biosensor
Authors: Ma Xiang , Chen Ting , Liu Lifang , Li Genxi
Published: November 2007

Haemoglobin (Hb) was entrapped in polysorbate 20 and then modified on a pyrolytic graphite electrode. Electrochemical studies revealed that a pair of stable and well‐defined redox peaks attributed to the direct redox reaction of Hb could be observed in a phosphate buffer solution (pH 6.0). The anodic and cathodic peaks were located at −236 and −316 mV (versus a saturated calomel reference electrode) separately. The formal potential, E0′, was linearly varied with pH in the range from 3.0 to 10.0 with a slope of −48.0 mV·pH−1. Moreover, the protein was capable of catalysing the reduction of H2O2. Accordingly, an unmediated biosensor for H2O2 was prepared with a linear range from 8.0×10−7 to 1.0×10−3 M. This biosensor exhibited good stability, sensitivity and reproducibility.
Antitumour activity of cationic‐liposome‐conjugated adenovirus containing the CCL19 [chemokine (C‐C motif) ligand 19] gene
Authors: Cao Mei , Deng Hong‐Xin , Zhao Jian , Fan Lin‐yu , Jiang Yu , Wen Yan‐Jun , Li Jiong , Lei Song , Mao Yong‐qiu , Ding Zhen‐Yu , Wei Yu‐quan
Published: November 2007

CCL19 [chemokine (C‐C motif) ligand 19; also known as MIP‐3β (macrophage inflammatory protein‐3β) or ELC (Epstein–Barr‐virus‐induced molecule 1 ligand chemokine)], one of the immunostimulatory cytokines, chemoattracts both DCs (dendritic cells) and T‐lymphocytes. Adenoviral vector is one of the most used gene delivery vectors for cancer therapy because of its high gene‐transfection efficiency. However, its wider application is limited, owing to immune responses that reduce transgene expression and decrease the efficacy of repeated administration. We constructed the recombinant replication deficient adenoviral vectors containing the CCL19 gene (Ad‐CCL19) and combined them with PEG‐PE [poly(ethylene glycol)–phosphatidylethanolamine]‐modified cationic liposomes (Ad‐CCL19/PEG‐PE) for immunotherapy against murine fibrosarcoma. Although there were hardly any therapeutic differences between Ad‐CCL19‐ and Ad‐CCL19/PEG‐PE‐treated mice that were observed at the second administration, the final results demonstrated that Ad‐CCL19/PEG‐PE‐treated mice survived much longer. The antitumour efficacy may be related to the high level of CCL19 after the final administration and lasting expression of IFN‐γ (interferon‐γ) and IL‐12 (interleukin‐12) in the Ad‐CCL19/PEG‐PE‐treated group, which were measured by reverse‐transcription PCR and ELISA. The results demonstrated that PEG‐PE‐cationic‐liposome‐conjugated adenovirus could prolong the expression of the therapeutic gene in vivo and may enhance the antitumour efficacy.
Effect of ascorbic acid on microcirculation in patients with Type II diabetes: a randomized placebo-controlled cross-over study
Authors: LU Qing , BJÖRKHEM Ingemar , WRETLIND Bengt , DICZFALUSY Ulf , HENRIKSSON Peter , FREYSCHUSS Anna
Published: November 2007

Manifestations of vascular disease, including microvascular changes, constitute the major part of the morbidity and mortality in diabetic patients. Oxidative stress has been suggested to play an important role in the vascular dysfunction of diabetic patients. Furthermore, epidemiological observations indicate a beneficial effect of an increased dietary intake of antioxidants. The present study tested the hypothesis that the antioxidant ascorbic acid influences microcirculatory function in patients with Type II diabetes. Patients with Type II diabetes were treated with 1 g of ascorbic acid three times a day for 2 weeks in a randomized placebo-controlled double-blind cross-over design. Microvascular reactivity was assessed by vital capillaroscopy and PRH (post-occlusive reactive hyperaemia). hs-CRP (high-sensitivity C-reactive protein), IL-6 (interleukin-6), IL-1ra (interleukin-1 receptor antagonist) and ox-LDL (oxidized low-density lipoprotein) were analysed. The results showed no significant change in microvascular reactivity assessed after 2 weeks of ascorbic acid treatment. TtP (time to peak) was 12.0±3.3 s before and 11.2±3.5 s after ascorbic acid (n=17). In comparison, TtP was 11.5±2.9 s before and 10.6±2.8 s after placebo (not significant). IL-1ra, IL-6, hs-CRP and ox-LDL did not change significantly after ascorbic acid, neither as absolute or relative values. In conclusion, in contrast with some studies reported previously, we could not demonstrate an effect of continuous oral treatment with ascorbic acid on microvascular reactivity assessed at the level of individual capillaries. Furthermore, we found no indication of an effect on inflammatory cytokines or ox-LDL.
Inhibitory effect of pitavastatin (NK-104) on the C-reactive-protein-induced interleukin-8 production in human aortic endothelial cells
Authors: KIBAYASHI Etsuko , URAKAZE Masaharu , KOBASHI Chikaaki , KISHIDA Mika , TAKATA Michiyo , SATO Akira , YAMAZAKI Katsuya , KOBAYASHI Masashi
Published: November 2007

Recent data have indicated that CRP (C-reactive protein) plays a role in atherosclerosis, in addition to being a marker for inflammatory diseases. IL-8 (interleukin-8), a CXC chemokine, is present in human coronary atheroma and promotes monocyte–endothelial cell adhesion. In the present study, we examined the effect of pitavastatin (NK-104), a synthetic statin (3-hydroxy-3-methylglutaryl CoA reductase inhibitor), on IL-8 production induced by CRP in human AoEC (aortic endothelial cells). We also investigated whether CRP can induce IL-8 production and if the activation of signalling pathways are functionally related. The concentrations of IL-8 in the media after stimulation with CRP were measured by ELISA, and the expression of IL-8 mRNA was assessed by Northern blot. The phosphorylation of MAPKs (mitogen-activated protein kinases) was determined by Western blot. The production of IL-8 induced by CRP (10 μg/ml) was enhanced significantly and was inhibited by pitavastatin. The expression of IL-8 mRNA was increased in a dose-dependent manner after stimulation with CRP (1–100 μg/ml), whereas expression of IL-8 mRNA induced by CRP (50 μg/ml) was significantly diminished by 5 μM pitavastatin. Furthermore, specific MAPK inhibitors (PD98059, SB203580 and SP600125) inhibited the expression of IL-8 mRNA induced by CRP (50 μg/ml). The phosphorylation of all three MAPKs [ERK (extracellular-signal-regulated kinase), p38 MAPK and JNK (c-Jun N-terminal kinase)] induced by CRP (10 μg/ml) was also significantly inhibited by pitavastatin. Our results suggest that CRP may play a role in atherosclerosis via IL-8 production and pitavastatin may prevent the progression of atherosclerosis not only by lowering plasma low-density lipoprotein cholesterol levels, but also by suppressing IL-8 production in endothelial cells through the inhibition of MAPK (ERK, p38 MAPK and JNK) pathways.
Haemodynamically significant plaque formation and regional endothelial dysfunction in cholesterol-fed ApoE−/− mice
Authors: JOHANSSON Maria E. , HÄGG Ulrika , WIKSTRÖM Johannes , WICKMAN Anna , BERGSTRÖM Göran , GAN Li-ming
Published: November 2007

Flow-mediated vasodilation is suggested as one of the mechanisms involved in arterial expansive remodelling, which is thought to be a defence mechanism in atherogenesis. In the present study, we tested the hypothesis that lumen obstructive plaque formation is associated with failure of NO (nitric oxide)-dependent vasodilation in conduit vessels. Cardiac function and aortic root flow velocities were assessed using high-resolution echocardiography and two-dimensional-guided pulsed Doppler in ApoE−/− (apolipoprotein E-deficient) mice fed a standard or high-cholesterol diet. Endothelial function in the proximal and mid-descending aortic regions was studied using a myograph technique. Flow velocity at the aortic root of cholesterol-fed ApoE−/− mice was significantly increased as a result of lumen narrowing, detected via histological analysis. NO-dependent vasodilatory responses were selectively impaired in the atherosclerosis-prone vascular regions in cholesterol-fed ApoE−/− mice. In conclusion, consumption of a high-cholesterol diet results in lumen obstructive plaque formation in ApoE−/− mice, which significantly alters aortic root haemodynamics. This phenomenon is associated with impaired NO-dependent vasodilation in vessel segments known to be prone to atherosclerosis.
Relative risks of the metabolic syndrome according to the degree of insulin resistance in apparently healthy Korean adults
Authors: PARK Seung-Ha , LEE Won-Young , RHEE Eun-Jung , JEON Woo-Kyu , KIM Byung-Ik , RYU Seung-Ho , KIM Sun-Woo
Published: November 2007

A new simple criterion for diagnosing metabolic syndrome was proposed in the third report of the NCEP (National Cholesterol Education Program). In the present study, we analysed the association between metabolic syndrome and insulin resistance to investigate the effects of the latter on the prevalence of metabolic syndrome based on the new criteria recommended in the ATP (Adult Treatment Panel) III report. A total of 7057 participants (4472 men and 2585 women), who underwent medical screening at the Sungkyunkwan University Kangbuk Samsung Hospital, were investigated. Fasting insulin levels were measured and components of the metabolic syndrome as defined by the ATP III report were determined. We also applied the criteria for abdominal obesity as defined by APC-WC (Asia–Pacific criteria for waist circumference). The prevalence of metabolic syndrome as defined by ATP III was 5.3% (5.0% in men and 5.8% in women) and 8.9% (8.1% in men and 10.3% in women) by APC-WC. The odds ratio for the metabolic syndrome was significantly higher in subjects with higher insulin resistance than in those with lower insulin resistance. The mean levels of HOMA (homoeostatic model assessment) and fasting insulin were significantly higher in those with more of the components of the metabolic syndrome. A high HOMA (≥2.56) and fasting insulin concentration (≥9.98 μIU/ml; where IU is international unit) were found to be independent risk factors of the metabolic syndrome by multiple regression analysis after adjusting for age, sex and body mass index (P<0.001). These results show that the metabolic syndrome is significantly correlated with the insulin resistance index, and that appropriate values of HOMA and fasting insulin concentration may serve as a helpful guide for the management of metabolic syndrome.