bluuebunny/crossref_metadata_2025 huggingface dataset:
Mutations in the telomerase component NHP2 cause the premature ageing syndrome dyskeratosis congenita
Authors: Vulliamy Tom , Beswick Richard , Kirwan Michael , Marrone Anna , Digweed Martin , Walne Amanda , Dokal Inderjeet
Published: June 2008

Dyskeratosis congenita is a premature aging syndrome characterized by muco-cutaneous features and a range of other abnormalities, including early greying, dental loss, osteoporosis, and malignancy. Dyskeratosis congenita cells age prematurely and have very short telomeres. Patients have mutations in genes that encode components of the telomerase complex (dyskerin, TERC, TERT, and NOP10), important in the maintenance of telomeres. Many dyskeratosis congenita patients remain uncharacterized. Here, we describe the analysis of two other proteins, NHP2 and GAR1, that together with dyskerin and NOP10 are key components of telomerase and small nucleolar ribonucleoprotein (snoRNP) complexes. We have identified previously uncharacterized NHP2 mutations that can cause autosomal recessive dyskeratosis congenita but have not found any GAR1 mutations. Patients with NHP2 mutations, in common with patients bearing dyskerin and NOP10 mutations had short telomeres and low TERC levels. SiRNA-mediated knockdown of NHP2 in human cells led to low TERC levels, but this reduction was not observed after GAR1 knockdown. These findings suggest that, in human cells, GAR1 has a different impact on the accumulation of TERC compared with dyskerin, NOP10, and NHP2. Most of the mutations so far identified in patients with classical dyskeratosis congenita impact either directly or indirectly on the stability of RNAs. In keeping with this effect, patients with dyskerin, NOP10, and now NHP2 mutations have all been shown to have low levels of telomerase RNA in their peripheral blood, providing direct evidence of their role in telomere maintenance in humans.
Naturally acquired Duffy-binding protein-specific binding inhibitory antibodies confer protection from blood-stage
Plasmodium vivax
infection
Authors: King Christopher L. , Michon Pascal , Shakri Ahmad Rushdi , Marcotty Alexandra , Stanisic Danielle , Zimmerman Peter A. , Cole-Tobian Jennifer L. , Mueller Ivo , Chitnis Chetan E.
Published: June 2008

Individuals residing in malaria-endemic regions acquire protective immunity after repeated infection with malaria parasites; however, mechanisms of protective immunity and their immune correlates are poorly understood. Blood-stage infection with
Plasmodium vivax
depends completely on interaction of
P. vivax
Duffy-binding protein (PvDBP) with the Duffy antigen on host erythrocytes. Here, we performed a prospective cohort treatment/reinfection study of children (5–14 years) residing in a
P. vivax
-endemic region of Papua New Guinea (PNG) in which children were cleared of blood-stage infection and then examined biweekly for reinfection for 25 weeks. To test the hypothesis that naturally acquired binding inhibitory antibodies (BIAbs) targeting PvDBP region II (PvDBPII) provide protection against
P. vivax
infection, we used a quantitative receptor-binding assay to distinguish between antibodies that merely recognize PvDBP and those that inhibit binding to Duffy. The presence of high-level BIAbs (>90% inhibition of PvDBPII-Duffy binding,
n
= 18) before treatment was associated with delayed time to
P. vivax
reinfection diagnosed by light microscopy (
P
= 0.02), 55% reduced risk of
P. vivax
reinfection (Hazard's ratio = 0.45,
P
= 0.04), and 48% reduction in geometric mean
P. vivax
parasitemia (
P
< 0.001) when compared with children with low-level BIAbs (
n
= 148). Further, we found that stable, high-level BIAbs displayed strain-transcending inhibition by reducing reinfection with similar efficiency of PNG
P. vivax
strains characterized by six diverse PvDBPII haplotypes. These observations demonstrate a functional correlate of protective immunity
in vivo
and provide support for developing a vaccine against
P. vivax
malaria based on PvDBPII.
Yeast UCS proteins promote actomyosin interactions and limit myosin turnover in cells
Authors: Lord Matthew , Sladewski Thomas E. , Pollard Thomas D.
Published: June 2008

Two functions are proposed for the conserved family of UCS proteins: helping to fold myosin motor proteins and stimulating the motor function of folded myosins. We examined both functions in yeast. The fission yeast UCS protein (Rng3p) concentrates in nodes containing myosin-II (Myo2) and other proteins that condense into the cytokinetic contractile ring. Both the N-terminal (central) and C-terminal (UCS) domains of Rng3p can concentrate independently in contractile rings, but only full-length Rng3p supports contractile ring function
in vivo
. The presence of Rng3p in ATPase assays doubles the apparent affinity (
K
ATPase
) of both native Myo2 and recombinant heads of Myo2 for actin filaments. Rng3p promotes gliding of actin filaments by full-length Myo2 molecules, but not Myo2 heads alone. Myo2 isolated from mutant strains defective for Rng3p function is soluble and supports actin filament gliding. In budding yeast the single UCS protein (She4p) acts on both myosin-I isoforms (Myo3p and Myo5p) and one of two myosin-V isoforms (Myo4p). Myo5p turns over ≈10 times faster in
she4
Δ cells than wild-type cells, reducing the level of Myo5p in cells 10-fold and in cortical actin patches ≈4-fold. Nevertheless, Myo5p isolated from
she4
Δ cells has wild-type ATPase and motility activities. Thus, a fraction of this yeast myosin can fold
de novo
in the absence of UCS proteins, but UCS proteins promote myosin stability and interactions with actin.
Metaplastic control of the endocannabinoid system at inhibitory synapses in hippocampus
Authors: Edwards David A. , Zhang Longhua , Alger Bradley E.
Published: June 2008

The modifiability of neuronal response plasticity is called “metaplasticity.” In suppressing synaptic inhibition and facilitating induction of long-term excitatory synaptic plasticity, endocannabinoids (eCBs) act as agents of metaplasticity. We now report the discovery of a calcium-dependent mechanism that regulates eCB mobilization by metabotropic glutamate receptor (mGluR) activation. The switch-like mechanism primes cells to release eCBs and requires a transient rise in intracellular Ca
2+
concentration ([Ca
2+
]
i
) but not concurrent activation of mGluRs. Conversely, short-term, [Ca
2+
]
i
- dependent eCB release can be persistently enhanced by mGluR activation. Hence, eCBs are also objects of metaplasticity, subject to higher levels of physiological control.
Spatial shifts in spawning habitats of Arcto-Norwegian cod related to multidecadal climate oscillations and climate change
Authors: Sundby Svein , Nakken Odd
Published: June 2008

Sundby, S., and Nakken, O. 2008. Spatial shifts in spawning habitats of Arcto-Norwegian cod related to multidecadal climate oscillations and climate change. – ICES Journal of Marine Science, 65: 953–962. Arcto-Norwegian cod tend to produce strong year classes in warm years and poor year classes in cold years. This work shows that spawning intensity at various spawning sites along 1500 km of Norwegian coast is also influenced by climate variations. However, while the recruitment response to temperature is immediate and on an interannual time-scale, the response to changes in spawning site is slower, on a multidecadal time-scale. There have been two cold and two warm periods during the 20th century, cold from 1900 to 1920, warm from 1930 to 1950, cold from 1960 to 1970, and warm since the mid-1980s. A time-series for 1900–1976 on cod roe indices along the coast shows that the southernmost spawning areas are more important during cold periods, and the northernmost ones in warm periods, and coincide with high population fecundity. After 1976, qualitative observations show that there have been poor spawning fisheries in the southernmost spawning areas during the present warm period. From 2003, spawning has been observed along the coast of East Finnmark where it did not transpire during the previous 40 years.
Non-parametric estimation of residual moments and covariance
Authors: Evans Dafydd , Jones Antonia J
Published: June 2008

The aim of non-parametric regression is to model the behaviour of a response vector
Y
in terms of an explanatory vector
X
, based only on a finite set of empirical observations. This is usually performed under the additive hypothesis
Y
=
f
(
X
)+
R
, where
f
(
X
)=

(
Y
|
X
) is the true regression function and
R
is the true residual variable. Subject to a Lipschitz condition on
f
, we propose new estimators for the moments (scalar response) and covariance (vector response) of the residual distribution, derive their asymptotic properties and discuss their application in practical data analysis.
Mechanisms of substrate selectivity for Bacillus anthracis thymidylate kinase
Authors: Carnrot Cecilia , Wang Liya , Topalis Dimitri , Eriksson Staffan
Published: June 2008

Bacillus anthracis is well known in connection with biological warfare. The search for new drug targets and antibiotics is highly motivated because of upcoming multiresistant strains. Thymidylate kinase is an ideal target since this enzyme is at the junction of the de novo and salvage synthesis of dTTP, an essential precursor for DNA synthesis. Here the expression and characterization of thymidylate kinase from B. anthracis (Ba‐TMPK) is presented. The enzyme phosphorylated deoxythymidine‐5′‐monophosphate (dTMP) efficiently with K m and V max values of 33 μM and 48 μmol mg−1 min−1, respectively. The efficiency of deoxyuridine‐5′‐monophosphate phosphorylation was ∼10% of that of dTMP. Several dTMP analogs were tested, and D‐FMAUMP (2′‐fluoroarabinosyl‐5‐methyldeoxyuridine‐5′‐monophosphate) was selectively phosphorylated with an efficiency of 172% of that of D‐dTMP, but l‐FMAUMP was a poor substrate as were 5‐fluorodeoxyuridine‐5′‐monophosphate (5FdUMP) and 2′,3′‐dideoxy‐2′,3′‐didehydrothymidine‐5′‐monophosphate (d4TMP). No activity could be detected with 3′‐azidothymidine‐5′‐monophosphate (AZTMP). The corresponding nucleosides known as efficient anticancer and antiviral compounds were also tested, and d‐FMAU was a strong inhibitor with an IC50 value of 10 μM, while other nucleosides—l‐FMAU, dThd, 5‐FdUrd, d4T, and AZT, and 2′‐arabinosylthymidine—were poor inhibitors. A structure model was built for Ba‐TMPK based on the Staphylococcus aureus TMPK structure. Docking with various substrates suggested mechanisms explaining the differences in substrate selectivity of the human and the bacterial TMPKs. These results may serve as a start point for development of new antibacterial agents.
Roles of the interactions between Env and Gag proteins in the HIV‐1 replication cycle
Authors: Murakami Tsutomu
Published: June 2008

The Env and Gag proteins of HIV‐1 are the two major structural proteins of this retrovirus. The interactions between Env and Gag proteins and their regulation in HIV‐1 are required for several steps of the replication cycle, involving not only virus assembly, specifically Env incorporation, but also entry steps after virus maturation. A large number of host factors and certain membrane microdomains appear to engage both in transport/trafficking of Env and/or Gag proteins, and in the interactions of these two proteins. The present review briefly summarizes our current knowledge regarding the roles of the interactions between Env and Gag proteins in the virus replication cycle.
Cleavage of the NS2‐3 protein in the cells of cattle persistently infected with non‐cytopathogenic bovine viral diarrhea virus
Authors: Kameyama Ken‐ichiro , Sakoda Yoshihiro , Matsuno Keita , Ito Asako , Tajima Motoshi , Nakamura Shigeyuki , Kida Hiroshi
Published: June 2008

The NS2‐3 of BVDV is cleaved in cultured cells infected with cp BVDV but not in those infected with ncp BVDV when tested more than 10 hours post infection. However, it is not known whether cleavage of NS2‐3 occurs in vivo. In the present study, cleavage of NS2‐3 in cattle persistently infected with BVDV was investigated. All BVDV isolated from PI animals were of the ncp biotype, and NS2‐3 proteins were detected in bovine fetal muscular cells infected with these viruses. On the other hand, in the leukocytes of those PI animals, NS3 proteins, products of the cleavage of NS2‐3 proteins, were detected. In addition, the NS3 proteins were also detected in leukocytes artificially infected with ncp BVDV. These results reveal that the NS2‐3 protein of BVDV is cleaved in leukocytes. Furthermore, NS3 proteins were detected in many tissues of PI cattle, such as lymphoid tissue, brain, thyroid, lung, and kidney. These results suggest that the NS2‐3 protein of ncp BVDV cleaves in vivo.
Genetic analyses of the putative O and K antigen gene clusters of pandemic Vibrio parahaemolyticus
Authors: Okura Masatoshi , Osawa Ro , Tokunaga Akihiko , Morita Masatomo , Arakawa Eiji , Watanabe Haruo
Published: June 2008

Pandemic V. parahaemolyticus strains have rapidly changed their serotypes, but its determinants, especially K antigen, and the genes involved in serotype have been an open question. The purpose of this study was to gain insights into these points. Although V. parahaemolyticus is known to be lacking O‐side chain on its lipopolysaccharide, and O antigens are thought to be represented by core OS, the genome sequence of V. parahaemolyticus O3:K6 strain RIMD2210633 suggests that this bacterium potentially synthesizes O‐side chain. To explore possible relatedness between this O‐side chain biosynthesis gene cluster, which is similar in the serotypes of Vibrio cholerae, and of V. parahaemolyticus, we amplified both core OS and O‐side chain gene clusters of the strains belonging to various serotypes of V. parahaemolyticus by long PCR and performed PCR RFLP analyses. The results of our RFLP analyses suggest that the core OS biosynthesis gene cluster is related to the O antigens of pandemic V. parahaemolyticus and that the putative O‐side chain gene cluster is related to K antigens of pandemic V. parahaemolyticus. We then determined the sequence of these regions of a pandemic O4:K68 strain, and compared it with the corresponding sequence of RIMD2210633. In addition, PCR analysis showed the putative O4 and K68 antigen gene clusters are unique to the strains belonging to the O4 and K68 serotype respectively. The data implies that the pandemic O4:K68 V. parahaemolyticus strain emerged from the pandemic O3:K6 strain by replacement of the putative O and K antigen gene clusters.
Genetic diversity ofFrancisella tularensissubspeciesholarcticastrains isolated in Japan
Authors: Fujita Osamu , Uda Akihiko , Hotta Akitoyo , Okutani Akiko , Inoue Satoshi , Tanabayashi Kiyoshi , Yamada Akio
Published: June 2008

The recently developed MLVA has high discriminatory power for the typing of individual strains or isolates ofFrancisella tularensis. In the present study, MLVA was applied to 33 JapaneseF. tularensissubspeciesholarcticastrains to examine the genetic diversity ofF. tularensisisolated. Among the seven VNTR loci analyzed, Ft‐M2, Ft‐M10, and Ft‐M20 loci showed high genetic polymorphism in Japanese strains, whereas Ft‐M3 was most variable in non‐Japanese strains. These results provide novel extended information about the genomic diversity among the strains ofF. tularensisssp.holarcticadistributed in Japan and enable determination of whether a given isolate is indigenous to Japan by examining these loci using MLVA.
Electron microscopic examination of uncultured soil‐dwelling bacteria
Authors: Amako Kazunobu , Takade Akemi , Taniai Hiroaki , Yoshida Shin‐ichi
Published: June 2008

Bacteria living in soil collected from a rice paddy in Fukuoka, Japan, were examined by electron microscopy using a freeze‐substitution fixation method. Most of the observed bacteria could be categorized, based on the structure of the cell envelope and overall morphology, into one of five groups: (i) bacterial spore; (ii) Gram‐positive type; (iii) Gram‐negative type; (iv) Mycobacterium like; and (v) Archaea like. However, a few of the bacteria could not be readily categorized into one of these groups because they had unique cell wall structures, basically resembling those of Gram‐negative bacteria, but with the layer corresponding to the peptidoglycan layer in Gram‐negative bacteria being extremely thick, like that of the cortex of a bacterial spore. The characteristic morphological features found in many of these uncultured, soil‐dwelling cells were the nucleoid being in a condensed state and the cytoplasm being shrunken. We were able to produce similar morphologies in vitro using a Salmonella sp. by culturing under low‐temperature, low‐nutrient conditions, similar to those found in some natural environments. These unusual morphologies are therefore hypothesized to be characteristic of bacteria in resting or dormant stages.
‘LOSCS’ Lateral Offset Stacked Channel Simulations: Towards geometrical modelling of turbidite elementary channels
Authors: Labourdette Richard
Published: June 2008

Turbidite hydrocarbon reservoirs are complex features, that need to be described in detail and represented as clearly as possible. The morphology and internal distribution of elementary distributary channels are dependent on depositional settings, leading to diverse arrangements at different scales. Reservoir modelling usually requires a description of sedimentary heterogeneity on a scale smaller than that given by seismic resolution. This is because seismic data only display the outside geometry of their lateral stack, i.e. a turbidite fairway. Complexes of Laterally Offset Stacked Turbidite Channels (LOSCs) also require a description based on the scales of individual channel bodies. The most common representation of channels in a fairway is by stochastic object modelling; i.e. populating the observed fairway with realistic forms representing individual channels, but with no established consistency between the individual channels. On the other hand, one essential characteristic of LOSCs is that it evolves by progressive migration laterally and/or downdip. Stochastic object modelling provides an inadequate representation of this progressive evolution, and consequently, a poor rendering of the heterogeneity distribution in the reservoir. The method we propose consists of defining a realistic succession of individual channels that can accurately build the fairway observed on seismic. ‘Realism’ is defined using criteria from the shape of individual channels, and based on the amount of displacement necessary between successive episodes. Depending on seismic resolution, the system can be constrained by one or several positions of individual channels (the most recent position of the channel is often filled with shale, therefore visible on seismic and usable as a control point). The final result is a deterministic succession of channels laterally stacked to build the seismically observed envelope. Even with no calibration, the resulting architecture respects the general ‘texture’ of the complex and provides a better simulation of flow pathways than that achieved by random object modelling.
Vincristine pharmacokinetics is related to clinical outcome in children with standard risk acute lymphoblastic leukemia
Authors: Lönnerholm Gudmar , Frost Britt‐Marie , Abrahamsson Jonas , Behrendtz Mikael , Castor Anders , Forestier Erik , Heyman Mats , Uges Donald R. A. , de Graaf Siebold S. N.
Published: June 2008

Vincristine is a key drug in the treatment of childhood and adult acute lymphoblastic leukemia (ALL), and many other childhood malignancies. Despite decades of wide clinical use, no data on the correlation between vincristine pharmacokinetics and long‐term clinical outcome have been published. We here report clinical data (median follow‐up time 10·5 years, range 7·3–12 years) for 86 children with B‐cell precursor ALL, in whom vincristine kinetics were studied on treatment day 1. The median total plasma clearance was 429 and 331 ml/min per m2 and the area under the plasma concentration‐time curve (AUC) was 4·49 and 5·40 mg/l × min in relapse and non‐relapse patients, respectively (not significant). In standard risk patients, where treatment depends more heavily on vincristine than in other subgroups, the relative risk (RR) of relapse was significantly increased for patients with clearance values above median (RR 5·2; P = 0·036), or AUC values below median (RR 5·8; P = 0·025). Our data suggest a relationship between the antileukemic effect and the systemic exposure of the drug, which warrants further studies.
Dose–response relationship for risk of non‐vertebral fracture with inhaled corticosteroids
Authors: Weatherall M. , James K. , Clay J. , Perrin K. , Masoli M. , Wijesinghe M. , Beasley R.
Published: June 2008

Objective
To determine the strength of association between the dose of inhaled corticosteroids (ICS) and risk of non‐vertebral fracture in adults.

Methods
A systematic review and meta‐analysis of case–control studies of non‐vertebral fractures in adults, in which at least two doses of ICS were reported as the dose of beclomethasone dipropionate (BDP) or equivalent.

Results
Five case–control studies were identified, with a total of 43 783 cases and 259 936 controls. There was an association between the risk of non‐vertebral fracture and increasing dose of BDP. The random‐effects odds ratio of relative risk for a non‐vertebral fracture was 1.12 (95% confidence interval 1.00–1.26) per 1000 μg increase in the daily dose of BDP or equivalent.

Conclusion
In older adults, the relative risk of non‐vertebral fractures increases by about 12% for each 1000 μg/day increase in the dose of BDP or equivalent. The magnitude of this risk was considerably less than other common risk factors for fracture in the older adult.
A comparison of phosphorus and DOC leachates from different types of leaf litter in an urban environment
Authors: WALLACE TODD A. , GANF GEORGE G. , BROOKES JUSTIN D.
Published: June 2008

1. The leaching rates of filterable reactive phosphorus (FRP) and dissolved organic carbon (DOC) from five leaf litter types commonly occurring in urban environments in Mediterranean regions of Southern Australia were compared. The relative composition, bioavailability and oxygen demand of this DOC were also assessed. Four tree species were assessed, including the native river red gum (Eucalyptus camaldulensis) and three introduced deciduous species, the English elm (Ulmus procera), London plane (Platanus acerifolia) and white poplar (Populus alba). Grass cuttings (mixed species) were selected as a common garden waste.

2. Except for English elm, the majority of FRP and DOC was released within the first 48 h. Grass cuttings released the highest amount of FRP with white poplar releasing the most DOC. Species that released relatively high amounts of DOC (white poplar, English elm, river red gum) released relatively low amounts of FRP. Conversely, species that released relatively low amounts of DOC (grass cuttings and London plane) tended to release relatively high amounts of FRP.

3. Analysis of DOC composition, combined with the differing oxygen demand and DOC depletion curves, demonstrated that there were substantial differences in the DOC leached from the leaf litter of the different species. Biochemical oxygen demand and the biodegradability of the DOC was positively correlated with the proportion of hydrophilic and hydrophobic acids present in the leachate.

4. These results demonstrate that simple measurements of nutrient release per gram of leaf litter would be insufficient to predict the ecological impact on receiving waters resulting from changes in dominant vegetation. Furthermore, the use of traps to prevent particulate leaf material from entering streams may have limited potential for reducing the load of dissolved nutrients. We conclude that any changes to vegetation type which substantially alter the timing of leaf fall or the composition of leaf leachates should be avoided.
Involvement of endogenous leukotriene B4 and platelet‐activating factor in polymorphonuclear leucocyte recruitment to dermal inflammatory sites in rats
Authors: Bélanger Caroline , Elimam Hanan , Lefebvre Julie , Borgeat Pierre , Marleau Sylvie
Published: June 2008

A critical role for leukotriene B4 (LTB4) and/or platelet‐activating factor (PAF) in regulating polymorphonuclear cell (PMN) trafficking to inflammatory sites has been reported in a number of experimental inflammatory models. In vitro, newly synthesized LTB4 and PAF were shown to act in an autocrine/paracrine or intracrine fashion to enhance intracellular arachidonic acid availability and leukotriene biosynthesis. This suggested potentially cooperative effects of these lipid mediators in regulating PMN extravasation. The present study aimed to elucidate whether endogenous LTB4 and PAF may both act to regulate plasma extravasation and PMN trafficking to inflammatory sites in experimental inflammation. With this aim, we have used selective and potent PAF and LTB4 receptor antagonist pretreatments in dermal and pulmonary inflammation models in rats. Our results show additive inhibitory effects of dual LTB4 and PAF receptor blockade in either PAF‐ or LTB4‐elicited cutaneous PMN accumulation compared to single‐drug administration. Furthermore, the combined administration of the drugs inhibited the PMN accumulation induced by the chemically unrelated soluble agonists tumour necrosis factor‐α and C5a. Finally, in a model of pulmonary inflammation induced by the intravenous injection of Sephadex beads, lung neutrophilia was reduced by 63% following the administration of LTB4 and PAF antagonists, in contrast with the lack of effect of single drug administration. Our results strongly support a role of both endogenous LTB4 and PAF in regulating PMN trafficking to inflammatory sites in various experimental conditions.
Human Langerhans’ cells and dermal‐type dendritic cells generated from CD34 stem cells express different toll‐like receptors and secrete different cytokines in response to toll‐like receptor ligands
Authors: Rozis George , Benlahrech Adel , Duraisingham Sai , Gotch Frances , Patterson Steven
Published: June 2008

Langerhans’ cells (LC) and dermal dendritic cells (dDC) are located in the superficial and deeper layers of the skin respectively and represent the main dendritic cell (DC) populations of the skin. LC‐like and dDC‐like DC can be generated from CD34 stem cells and this system is widely used as a model for investigating these cells and in therapeutic vaccination. Here we report toll‐like receptor (TLR) expression in human LC and dDC derived from CD34 stem cells. In vitro‐generated DC expressed TLR‐1, 2, 4, 6, 8 and 10. LC, but not dDC, expressed TLR‐5, whereas only dDC expressed TLR‐3. Maturation of LC was mediated by TLR‐2, 4 and 5 ligands, but not by a TLR‐3 ligand. dDC maturation was induced by TLR‐3 and ‐4, but not with TLR‐5 ligand and only weakly by a TLR‐2 ligand. Stimulated LC secreted interleukin (IL)‐1β, low levels of tumour necrosis factor‐α (TNF‐α) and IL‐8, but not IL‐6 or IL‐10. dDC secreted TNF‐α, IL‐6, IL‐8 and IL‐10, but little IL‐1β. IL‐12p70 was not produced by ligand‐stimulated dDC or LC, but was secreted by monocyte‐derived DC (mdDC) stimulated with lipopolysaccharide (LPS). Thus, in vitro‐generated LC and dDC detect different pathogen‐associated molecules and show different cytokine‐secretion profiles in response to TLR ligands.
β2‐Adrenergic receptor regulates Toll‐like receptor‐4‐induced nuclear factor‐κB activation through β‐arrestin 2
Authors: Kizaki Takako , Izawa Tetsuya , Sakurai Takuya , Haga Shukoh , Taniguchi Naoyuki , Tajiri Hisao , Watanabe Kenji , Day Noorbibi K. , Toba Kenji , Ohno Hideki
Published: June 2008

Toll‐like receptors (TLRs) play an important role in innate immunity while, β2‐adrenergic receptors (β2AR) provide the key linkages for the sympathetic nervous system to regulate the immune system. However, their role in macrophages remains uncertain. Here, we demonstrate the cross‐talk between β2AR and TLR signalling pathways. Expression of β2AR was down‐regulated by TLR4 ligand lipopolysaccharide (LPS) stimulation. To investigate the physiological consequence of this down‐regulation RAW264 cells, a macrophage cell line, were transfected with a β2AR expression vector (RAWar). Both LPS‐stimulated inducible nitric oxide synthase (NOS II) expression and NO production were markedly suppressed in the RAWar cells. The activation of nuclear factor‐κB (NF‐κB) and degradation of the inhibitor of NF‐κB (IκBα) in response to LPS were markedly decreased in these cells. The level of β‐arrestin 2, which regulates β2AR signalling, was also reduced in RAW264 cells after stimulation with LPS, but not in RAWar cells. Overexpression of β‐arrestin 2 (RAWarr2) also inhibited NO production and NOS II expression. Furthermore, we demonstrated that β‐arrestin 2 interacted with cytosolic IκBα and that the level of IκBα coimmunoprecipitated by anti‐β‐arrestin 2 antibodies was decreased in the RAW264 cells but not in RAWar or RAWarr2 cells. These findings suggest that LPS‐stimulated signals suppress β2AR expression, leading to down‐regulation of β‐arrestin 2 expression, which stabilizes cytosolic IκBα and inhibits the NF‐κB activation essential for NOS II expression, probably to ensure rapid and sufficient production of NO in response to microbial attack.
Functional analysis of a tripartite stability element within the CD40 ligand 3′ untranslated region
Authors: Laughlin Jennifer , Oghlidos Sanaz , Porter Joseph F. , Matus‐Nicodemos Rodrigo , Sinquett Frank L. , Marcelli Valentina , Covey Lori R.
Published: June 2008

We previously identified a cis‐acting element within the 3′ untranslated region of CD40 ligand messenger RNA (mRNA) that is composed of three complex binding sites and acts to increase mRNA stability in both in vitro and in vivo systems. We now demonstrate the functional consequences of the three binding sites with respect to increasing both luciferase activity and mRNA stability in a heterologous transcript expressed in a T‐cell line. The internal region B was shown to be a bona fide stability element because its presence increased luciferase activity fourfold over the unmodified transcript and its removal from the XbaI–HaeIII region resulted in rapid degradation of the transcript. Region A contained both a binding site for a polypyrimidine‐tract‐binding protein (PTB)‐mediated complex (Complex I) and an upstream, adjacent sequence that was a negative regulator of mRNA stability. Region C bound Complex II, which contained both PTB and heterogeneous nuclear ribonucleoproteinL (hnRNPL), and was less effective as a stability element on its own compared to region B. Our findings demonstrate differential levels of activity for the three binding sites relative to the turnover of CD40 ligand mRNA, suggesting that the lack of binding of Complex I/II during the early stages of T‐cell activation contributes to the rapid degradation of the CD40 ligand mRNA transcript.
Vδ2 T‐lymphocyte responses in cord blood samples from Italy and Côte d’Ivoire
Authors: Cairo Cristiana , Mancino Giorgio , Cappelli Giulia , Pauza C. David , Galli Elena , Brunetti Ercole , Colizzi Vittorio
Published: June 2008

Cord blood T lymphocytes are immature and their functional defect partially reflects a suboptimal level of costimulatory signals provided by neonatal antigen‐presenting cells. Neonatal Vδ2 T lymphocytes, a small component of cellular immunity involved in the response against bacteria, protozoa, virus‐infected cells and tumours, are also considered to be immature. Cord blood Vδ2 T lymphocytes are mostly naïve, proliferate poorly and do not produce cytokines in response to the model phosphoantigen isopentenyl pyrophosphate. We cultured cord blood mononuclear cells with the aminobisphosphonate Pamidronate or with live bacille Calmette–Guérin, and showed that both elicit a strong cord blood Vδ2 T‐cell proliferative response, inducing the expression of activation markers and promoting the differentiation from naïve to memory cells. Our results suggest that cord blood Vδ2 T cells are not inherently unresponsive and can mount strong responses to aminobisphosphonates and mycobacteria. Neonatal Vδ2 T lymphocytes may be important participants in responses to microbial infections early in life.
Generation and characterization of transgenic mice expressing a T‐cell receptor specific for the tumour‐associated antigen MDM2
Authors: Xue Shao‐An , Bendle Gavin M. , Holler Angelika , Stauss Hans J.
Published: June 2008

T‐cell‐based antigen‐specific immunotherapy targeting tumour‐associated antigens offers the potential for cancer immunotherapy. However, the majority of identified tumour‐associated antigens are also expressed at low levels in normal tissues and mechanisms of tolerance induction are likely to affect the quality of T‐cell responses to such antigens. In this study a T‐cell receptor transgenic model was developed to determine the magnitude of T‐cell tolerance to the tumour‐associated antigen murine double minute‐2 (MDM2), a widely expressed protein that is found at elevated levels in many tumours. The analysis of transgenic mice showed that thymic deletion was responsible for purging large numbers of MDM2‐specific T cells from the repertoire. However, some T cells with specificity for MDM2 were able to escape thymic deletion and persisted in the peripheral T‐cell pool. Functional analysis revealed that these T cells displayed defects in antigen‐driven expansion. This functional impairment of the MDM2‐specific T cells was maintained following adoptive transfer of the T cells into hosts that were unable to present the T‐cell‐receptor‐recognized antigen. This study demonstrates that thymic deletion and the functional impairment of T cells present in the periphery both operate to establish T‐cell tolerance to the tumour‐associated antigen MDM2. Furthermore, the tolerant phenotype was stable and did not require continuous MDM2 peptide presentation in normal tissues.
Physicochemical and functional properties of barley β‐glucan as affected by different extraction procedures
Authors: Ahmad Asif , Anjum Faqir Muhammad , Zahoor Tahir , Nawaz Haq , Din Ahmad
Published: June 2008

Four methods were evaluated for extraction of β‐glucan from barley. Gum pellets obtained by respective methods were examined for yield, purity and various physicochemical and functional characteristics. Highest yield and recovery was found in samples that were extracted by hot water treatment. This method also removed the maximum impurities from gum pellets and contained highest amount of soluble fibre, while alkali‐extracted sample contained highest amount of insoluble fibre. All samples contained an appreciable amount of calcium and phosphorus. Extracted pellets had good water retention and foaming capacity. A significant variation in extraction methods was observed with respect to water binding capacity and foaming capacity. However, the same level of foaming stability was observed in all extraction methods. Alkaline and acidic extraction methods adversely affect the viscosity properties of the samples. A significant affect of extraction methods was also observed on colour parameters of β‐glucan. A negative correlation exists between foaming capacity and viscosity, whereas a positive correlation observed between soluble fibre and viscosity.
Generalist foraging of pollinators: diet expansion at high density
Authors: Fontaine Colin , Collin Carine L. , Dajoz Isabelle
Published: June 2008

Plant–pollinator interactions are crucial for the reproduction of most angiosperms. When faced with perturbations, plant–pollinator networks might be robust mainly due to the presence of highly generalist species. Perturbations can, in turn, affect how pollinators exploit their food sources and therefore their degree of generalization.

In particular, generalization of pollinators might vary with forager density. At high densities floral resources available in plant communities would be more rapidly depleted than at low densities. According to optimal foraging theory (OFT), this decrease in resources should lead to an increase in diet breadth.

We investigated the impact of pollinator density on diet breadth of Bombus terrestris. We recorded the individual foraging behaviour of bumblebees from eight colonies. They foraged in presence of either one or several conspecifics on experimental plant communities composed of five plant species. Diet breadth was calculated as the number of plant species visited, as well as by the Levins index. We analyzed the effect of forager density on diet breadth at both the individual and the colony levels.

We found that forager density affected degree of generalization at individual and colony levels. A more competitive environment increased individual generalization, as predicted by OFT, and decreased the variability in generalization across colonies. Moreover, we found that bumblebee size was positively related to the amplitude of diet breadth change.

Synthesis. Our study sheds light on an additional level of complexity of plant–pollinator systems, showing that pollinator diet breadth is a flexible trait which results from behavioural adaptation to resource availability. More generally, changes in foraging insects’ diet via competition for resources are likely to ensue from anthropogenic ecosystem disturbance and to impact upon the functioning of plant–pollinator networks.
Thermal acclimation of photosynthesis in black spruce [Picea mariana (Mill.) B.S.P.]
Authors: WAY DANIELLE A. , SAGE ROWAN F.
Published: June 2008

We investigated the thermal acclimation of photosynthesis and respiration in black spruce seedlings [Picea mariana (Mill.) B.S.P.] grown at 22/14 °C [low temperature (LT)] or 30/22 °C [high temperature (HT)] day/night temperatures. Net CO2 assimilation rates (Anet) were greater in LT than in HT seedlings below 30 °C, but were greater in HT seedlings above 30 °C. Dark and day respiration rates were similar between treatments at the respective growth temperatures. When respiration was factored out of the photosynthesis response to temperature, the resulting gross CO2 assimilation rates (Agross) was lower in HT than in LT seedlings below 30 °C, but was similar above 30 °C. The reduced Agross of HT seedlings was associated with lower needle nitrogen content, lower ribulose 1·5‐bisphosphate carboxylase/oxygenase (Rubisco) maximum carboxylation rates (Vcmax) and lower maximum electron transport rates (Jmax). Growth treatment did not affect Vcmax : Jmax. Modelling of the CO2 response of photosynthesis indicated that LT seedlings at 40 °C might have been limited by heat lability of Rubisco activase, but that in HT seedlings, Rubisco capacity was limiting. In sum, thermal acclimation of Anet was largely caused by reduced respiration and lower nitrogen investments in needles from HT seedlings. At 40 °C, photosynthesis in LT seedlings might be limited by Rubisco activase capacity, while in HT seedlings, acclimation removed this limitation.
MYBL2 is a new regulator of flavonoid biosynthesis in Arabidopsis thaliana
Authors: Dubos Christian , Le Gourrierec José , Baudry Antoine , Huep Gunnar , Lanet Elodie , Debeaujon Isabelle , Routaboul Jean‐Marc , Alboresi Alessandro , Weisshaar Bernd , Lepiniec Loïc
Published: June 2008

In Arabidopsis thaliana, several MYB and basic helix‐loop‐helix (BHLH) proteins form ternary complexes with TTG1 (WD‐Repeats) and regulate the transcription of genes involved in anthocyanin and proanthocyanidin (PA) biosynthesis. Similar MYB‐BHLH‐WDR (MBW) complexes control epidermal patterning and cell fates. A family of small MYB proteins (R3‐MYB) has been shown to play an important role in the regulation of epidermal cell fates, acting as inhibitors of the MBW complexes. However, so far none of these small MYB proteins have been demonstrated to regulate flavonoid biosynthesis. The genetic and molecular analyses presented here demonstrated that Arabidopsis MYBL2, which encodes a R3‐MYB‐related protein, is involved in the regulation of flavonoid biosynthesis. The loss of MYBL2 activity in the seedlings of two independent T‐DNA insertion mutants led to a dramatic increase in the accumulation of anthocyanin. In addition, overexpression of MYBL2 in seeds inhibited the biosynthesis of PAs. These changes in flavonoid content correlate well with the increased level of mRNA of several structural and regulatory anthocyanin biosynthesis genes. Interestingly, transient expression analyses in A. thaliana cells suggested that MYBL2 interacts with MBW complexes in planta and directly modulates the expression of flavonoid target genes. These results are fully consistent with the molecular interaction of MYBL2 with BHLH proteins observed in yeast. Finally, MYBL2 expression studies, including its inhibition by light‐induced stress, allowed us to hypothesise a physiological role for MYBL2. Taken together, these results bring new insights into the transcriptional regulation of flavonoid biosynthesis and provide new clues and tools for further investigation of its developmental and environmental regulation.
Comparison of bacterial pellets and microbial markers for the estimation of the microbial nitrogen and amino acids flows from single flow continuous culture fermenters fed diets containing two‐stage olive cake
Authors: Molina‐Alcaide E. , Moumen A. , Martín‐García I. , Carro M. D.
Published: June 2008

The effects of using effluent bacteria (EB) and solid‐ (SAB) and liquid‐ (LAB) associated bacteria and diaminopimelic acid (DAPA) or purine bases (PB) and partially substituting alfalfa hay (AH) by a concentrate including olive cake on the microbial N flow (MNF) and amino acids (AA) flow were investigated with continuous culture fermenters fed AH and a mixture of AH and a concentrate containing barley grains and two‐stage olive cake (2:1 ratio) without (AHCO) or with polyethylene glycol (PEG) (AHCOP). The MNF was not different among diets with SAB or LAB (p = 0.302 and 0.203, respectively) and DAPA, but differed with PB (p = 0.021 and 0.014, respectively). With EB both markers detected similar differences, AHCOP showing a higher value (p < 0.05) than AH and AHCO. The MNF was higher (p < 0.001) with PB than DAPA. Daily flow of non‐essential AA was not different (p = 0.356) among diets but essential AA flow was higher (p < 0.05) for AH and AHCOP than for AHCO. The SAB presented lower (p < 0.05) total AA than LAB and higher total AA (p < 0.05) for diet AH than AHCO. The AA profile of EB was similar to that of LAB, but alanine and leucine were higher (p < 0.05) in EB than in LAB. Microbial contribution to AA flow was 45.4%, 55.6% and 58.1% for diets AH, AHCO and AHCOP respectively. With both markers, microbial AA flow was higher (p < 0.05) for diet AHCOP compared with AH (451 and 355 mg/day, respectively), but not different (p > 0.05) for AHCOP and AHCO (389 mg/day). The results would indicate that olive cake could be used in the practical feeding of small ruminants without negatively affecting microbial AA N supply.
Carriage ofCandidaspecies in oral cavities of HIV infected patients in South India
Authors: Girish Kumar C. P. , Menon Thangam , Rajasekaran S. , Sekar B. , Prabu D.
Published: June 2008

Fifty‐four patients with human immunodeficiency virus (HIV) infection were studied to assess the load of oral carriage ofCandidaspp. The mean oralCandidacarriage density (30 305.93 ± 56 643.93 CFU ml−1) in HIV patients was significantly higher than that seen in the control population (93.48 ± 358.48 CFU ml−1;P = 0.000). The meanCandidaload in HIV patients with oral thrush (46 591.43 ± 65 002.57 CFU ml−1) was significantly higher than in the HIV subjects without oral thrush (306.32 ± 699.50 CFU ml−1;P = 0.000). Non‐C. albicans Candidaspecies (56%) were more predominant than theC. albicans(44%) isolates. 25S rDNA PCR analysis ofC. albicansrevealed preponderance of genotype A strains. Interestingly, 42.6% of rinse specimens grew multipleCandidaspecies, with the combination ofC. albicansandC. krusei(39.1%) being the most frequent.
Preoperative evaluation of hepatic functional reserve by converted ICGR15 calculated from 99mTc‐GSA scintigraphy
Authors: Kawamura Hideki , Kamiyama Toshiya , Nakagawa Takahito , Nakanishi Kazuaki , Yokoo Hideki , Tahara Munenori , Kamachi Hirofumi , Toi Hirofumi , Matsushita Michiaki , Todo Satoru
Published: June 2008

Background and Aim:  Conversion of data from technetium 99 m diethylenetriaminepentaacetic acid galactosyl human serum albumin (99mTc‐GSA) scintigraphy to ICGR15 (indocyanin green retention at 15 min) is an easy and convenient method for obtaining parameters to determine the appropriate and safe extent of liver resection. We investigated a conversion method which also accounts for LHL15 (receptor index: uptake ratio of the liver to the liver plus heart at 15 min) and HH15 (blood clearance index: uptake ratio of the heart at 15 min to that at 3 min) characteristics.

Methods:  Cases included 282 patients undergoing hepatic resection following 99mTc‐GSA scintigraphy and an ICG tolerance test. Degree of liver dysfunction was classified as A, B, or C according to criteria of the Liver Cancer Study Group of Japan.

Results:  HH15 demonstrated a larger distribution in patients with liver damage A, while LHL15 demonstrated a larger distribution in patients with liver damage B. In liver damage A, the conversion formula ICGR15 = 87.0–79.6 × LHL15 was obtained, and in liver damage B, the conversion formula ICGR15 = −23.3 + 72.4 × HH15 was obtained, and correlation with ICGR15 was higher (r = 0.61, P < 0.0001) than when the data were not segregated by liver damage severity. Furthermore, postoperative hyperbilirubinemia significantly occurred in cases where both ICGR15 and converted ICGR15 were high.

Conclusions:  Conversion models based on data segregated by severity of liver damage were more closely correlated with ICGR15 than conversion models not based on segregated data. By using this converted ICGR15, preoperative estimation of hepatic functional reserve can become more reliable.
Huang‐lian‐jie‐du‐tang, a traditional Chinese medicine prescription, induces cell‐cycle arrest and apoptosis in human liver cancer cells in vitro and in vivo
Authors: Hsu Ya‐Ling , Kuo Po‐Lin , Tzeng Tz‐Fei , Sung Shu‐Chiao , Yen Ming‐Hong , Lin Liang‐Tzung , Lin Chun‐Ching
Published: June 2008

Background and Aim:  Huang‐lian‐jie‐du‐tang (HLJDT; Japanese name, oren‐gedoku‐to) is a traditional Chinese medicine prescription known to possess anti‐inflammatory activity. Our study reports here for the first time the anticancer effect of HLJDT in two human liver cancer cell lines, Hep G2 and PLC/PRF/5.

Methods:  Inhibition of cell proliferation by HLJDT was measured by sodium 3′‐(1‐(phenylamino‐carbonyl)‐3,4‐tetrazolium)‐bis(4‐methoxy‐6‐nitro) benzene‐sulfonic acid hydrate (XTT) assay. Clonogenic assay was used to elucidate the possible differences in long‐term effects of HLJDT on human liver cancer cells. Cell cycle distribution was determined by flow cytometry. Apoptosis was detected using electrophoresis and terminal deoxynucleotidyl transferase‐mediated deoxyuridine triphosphate nick endlabeling (TUNEL) assay. Protein expressions were determined by immunoblot assay. The activity of nuclear factor–kappa B (NF‐κB) was determined by Trans‐AM ELISA kit. In vivo tumor activity was assessed by xenograft study.

Results:  HLJDT significantly increased the expression of inactivated phospho‐Cdc2 and phospho‐Cdc25C, and decreased the levels of cyclin A, cyclin B1, Cdc2, and Cdc25C, thereby contributing to cell‐cycle arrest. HLJDT increased the expression of Bax and Bak, but decreased the level of Bcl‐2 and Bcl‐XL, and subsequently triggered the mitochondrial apoptotic pathway. In addition, HLJDT also inhibited cell‐survival signaling by enhancing the amount of IκBα in the cytoplasm, reducing the level and activity of NF‐κB in the nucleus, and subsequently attenuating the expression of Bcl‐XL in Hep G2 and PLC/PRF/5 cells. The inhibitory effect mediated by HLJDT on cell growth was also demonstrated in a nude mouse model, in which the liver cancer cells induced tumor xenograft shrank considerably following treatment with HLJDT.

Conclusions:  Taken together, these results suggest a potential anticancer effect of HLJDT against human liver cancer cells.