bluuebunny/crossref_metadata_2025 huggingface dataset:
Use of Fatty Acid Methyl Ester Profiles to Compare Copper-Tolerant and Copper-Sensitive Strains of Pantoea ananatis
Authors: Nischwitz C. , Gitaitis R. , Sanders H. , Langston D. , Mullinix B. , Torrance R. , Boyhan G. , Zolobowska L.
Published: September 2007

A survey was conducted to evaluate differences in fatty acid methyl ester (FAME) profiles among strains of Pantoea ananatis, causal agent of center rot of onion (Allium cepa), isolated from 15 different onion cultivars in three different sites in Georgia. Differences in FAME composition were determined by plotting principal components (PCs) in two-dimensional plots. Euclidean distance squared (ED2) values indicated a high degree of similarity among strains. Plotting of PCs calculated from P. ananatis strains capable of growing on media amended with copper sulfate pentahydrate (200 μg/ml) indicated that copper-tolerant strains grouped into tight clusters separate from clusters formed by wild-type strains. However, unlike copper-sensitive strains, the copper-tolerant strains tended to cluster by location. A total of 80, 60, and 73% of the strains from Tift1, Tift2, and Tattnall, respectively, exhibited either confluent growth or partial growth on copper-amended medium. However, all strains were sensitive to a mixture of copper sulfate pentahydrate (200 μg/ml) and maneb (40 μg/ml). When copper-tolerant clones were analyzed and compared with their wild-type parents, in all cases the plotting of PCs developed from copper-tolerant clones formed tight clusters separate from clusters formed by the parents. Eigenvalues generated from these tests indicated that two components provided a good summary of the data, accounting for 98, 98, and 96% of the standardized variance for strains Pna 1-15B, Pna 1-12B, and Pna 2-5A, respectively. Furthermore, feature 4 (cis-9-hexadecenoic acid/2-hydroxy-13-methyltetradecanoic acid) and feature 7 (cis-9/trans-12/cis-7-octadecenoic acid) were the highest or second highest absolute values for PC1 in all three strains of the parents versus copper-tolerant clones, and hexadecanoic acid was the highest absolute value for PC2 in all three strains. Along with those fatty acids, dodecanoic acid and feature 3 (3-hydroxytetradecanoic acid/14-methylpentadecenoic acid) also had an impact on the differences observed between copper-sensitive parents and copper-resistant mutants. Finding these changes in bacterial fatty acid composition could lead to the development of a laboratory assay to identify copper-tolerant strains using gas chromatography as well as providing clues to further elucidate the mode of action of copper tolerance.
Comparison of Colletotrichum orbiculare and Several Allied Colletotrichum spp. for mtDNA RFLPs, Intron RFLP and Sequence Variation, Vegetative Compatibility, and Host Specificity
Authors: Liu B. , Wasilwa L. A. , Morelock T. E. , O'Neill N. R. , Correll J. C.
Published: September 2007

Based on spore morphology, appressorium development, sequence similarities of the rDNA, and similarities in amplified restriction fragment length polymorphism (AFLP), it has been proposed that Colletotrichum orbiculare, C. trifolii, C. lindemuthianum, and C. malvarum represent a single phylogenetic species, C. orbiculare. In the current study, the phylogenetic relationship among isolates in the C. orbiculare species complex was reassessed. In all, 72 isolates of C. orbiculare from cultivated cucurbit or weed hosts, C. trifolii from alfalfa, C. lindemuthianum from green bean, and C. malvarum from prickly sida (Sida spinosa) were examined for mitochondrial DNA (mtDNA) restriction fragment length polymorphisms (RFLPs), RFLPs and sequence variation of a 900-bp intron of the glutamine synthetase gene and a 200-bp intron of the glyceraldehyde-3-phosphate dehydrogenase gene, and vegetative compatibility. In addition, host specificity was examined in foliar inoculations on cucurbit, bean, and alfalfa hosts. Inoculations also were conducted on cucumber fruit. Genetically distinct isolates, based on vegetative compatibility, within the species complex (C. orbiculare, C. trifolii, and C. malvarum) had an identical mtDNA haplotype (haplotype A) when examined with each of three different restriction enzymes. Isolates of C. lindemuthianum had a very similar mtDNA haplotype to haplotype A, with a single polymorphism detected with the enzyme HaeIII. The four species represent a phylogenetically closely related group based on a statistical analysis of the 900- and 200-bp intron sequences. However, distinct RFLPs in the 900-bp intron were consistently associated with each species and could be used to qualitatively and quantitatively distinguish each species. Furthermore, each of the species showed distinct host specificity, with isolates of C. orbiculare (from cucurbits), C. lindemuthianum, and C. trifolii being pathogenic only on cucurbits, green bean, and alfalfa, respectively. Consequently, distinct and fixed nucleotide, or genotypic (intron sequences and RFLPs) and phenotypic (host specificity) characteristics can be used to distinguish C. orbiculare, C. lindemuthianum, and C. trifolii from one another; therefore, they should be recognized as distinct species. This species delineation is consistent with the most current species concepts in fungi. More isolates and further characterization is needed to determine whether C. orbiculare from cocklebur and C. malvarum represent distinct species. RFLPs of the 900-bp intron may represent a relatively inexpensive, reliable, and useful diagnostic tool for general species differentiation in the genus Colletotrichum.
Anisotropy, in Density and in Distance, of the Dispersal of Yellow Rust of Wheat: Experiments in Large Field Plots and Estimation
Authors: Soubeyrand S. , Enjalbert J. , Sanchez A. , Sache I.
Published: September 2007

Long-distance dispersal of spores generally presents anisotropy. This anisotropy can appear in the mean number of spores deposited along a given direction (anisotropy in density) and in the mean distance that a spore travels in a given direction (anisotropy in distance). Specific experiments together with a statistical methodology are proposed to study this effect. The experiments are based on the use of a point source of a traceable inoculum and susceptible trap plots in large resistant field plots. The anisotropy is characterized by two functions: a directional density function and a mean distance function which are related with the anisotropies in density and distance, respectively. A nonparametric approach is developed to estimate these functions and to help in choosing a parametric model. Then, the parametric model is estimated. In two field experiments, migrations up to 175 and 225 m from the source were detected, with ≈25% of the trap plots infected. Whatever the experiment, the two estimated anisotropies presented different shapes (i.e., the number of spores dispersed in a given direction was not proportional to the mean distance travelled by these spores).
Effects of Diffuse Colonization of Grape Berries byUncinula necatoron Bunch Rots, Berry Microflora, and Juice and Wine Quality
Authors: Gadoury David M. , Seem Robert C. , Wilcox Wayne F. , Henick-Kling Thomas , Conterno Lorenza , Day Andrea , Ficke Andrea
Published: September 2007

Production of grape (principally cultivars of Vitis vinifera) for high-quality wines requires a high level of suppression of powdery mildew (Uncinula necator syn. Erysiphe necator). Severe infection of either fruit or foliage has well-documented and deleterious effects upon crop and wine quality. We found that berries nearly immune to infection by U. necator due to the development of ontogenic resistance may still support diffuse and inconspicuous mildew colonies when inoculated ≈3 weeks post-bloom. Fruit with diffuse mildew colonies appear to be healthy and free of powdery mildew in late-season vineyard assessments with the naked eye. Nonetheless, presence of these colonies on berries was associated with (i) elevated populations of spoilage microorganisms; (ii) increased evolution of volatile ethyl acetate, acetic acid, and ethanol; (iii) increased infestation by insects known to be attracted to the aforementioned volatiles; (iv) increased rotting by Botrytis cinerea; and (v) increased frequency of perceived defects in wines prepared from fruit supporting diffuse powdery mildew colonies. Prevention of diffuse infection requires extending fungicidal protection until fruit are fully resistant to infection. Despite a perceived lack of improvement in disease control due to the insidious nature of diffuse powdery mildew, potential deleterious effects upon crop and wine quality thereby would be avoided.
Detection, Distribution, Sporulation, and Survival of Phytophthora ramorum in a California Redwood-Tanoak Forest Soil
Authors: Fichtner E. J. , Lynch S. C. , Rizzo D. M.
Published: September 2007

Recovery of Phytophthora ramorum from soils throughout sudden oak death-affected regions of California illustrates that soil may serve as an inoculum reservoir, but the role of soil inoculum in the disease cycle is unknown. This study addresses the efficacy of soil baiting, seasonal pathogen distribution under several epidemiologically important host species, summer survival and chlamydospore production in soil, and the impact of soil drying on pathogen survival. The efficacy of rhododendron leaves and pears as baits for detection of soilborne propagules were compared. Natural inoculum associated with bay laurel (Umbellularia californica), tanoak (Lithocarpus densiflorus), and redwood (Sequoia sempervirens) were determined by monthly baiting. Summer survival and chlamydospore production were assessed in infected rhododendron leaf disks incubated under bay laurel, tanoak, and redwood at either the surface, the litter/soil interface, or in soil. Rhododendron leaf baits were superior to pear baits for sporangia detection, but neither bait detected chlamydospores. Most inoculum was associated with bay laurel and recovery was higher in soil than litter. Soil-incubated inoculum exhibited over 60% survival at the end of summer and also supported elevated chlamydospore production. P. ramorum survives and produces chlamydospores in forest soils over summer, providing a possible inoculum reservoir at the onset of the fall disease cycle.
Chaotic stirring in quasi-turbulent flows
Authors: Lekien Francois , Coulliette Chad
Published: September 2007

Transport in laminar flows is governed by chaotic stirring and striation in long thin filaments. In turbulent flows, isotropic mixing dominates and tracers behave like stochastic variables. In this paper, we investigate the quasi-turbulent, intermediate regime where both chaotic stirring and turbulent mixing coexist. In these flows, the most common in nature, aperiodic Lagrangian coherent structures (LCSs) delineate particle transport and chaotic stirring. We review the recent developments in LCS theory and apply these techniques to measured surface currents in Monterey Bay, California. In the bay, LCSs can be used to optimize the release of drifting buoys or to minimize the impact of a coastal pollution source.
Ntr1 activates the Prp43 helicase to trigger release of lariat-intron from the spliceosome
Authors: Tanaka Naoko , Aronova Anna , Schwer Beate
Published: September 2007

DEAD/H-box NTPases remodel the spliceosome at multiple steps during the pre-mRNA splicing cycle. The RNA-dependent NTPase Prp43 catalyzes dissociation of excised lariat-intron from the spliceosome, but it is unclear how Prp43 couples the energy of ATP hydrolysis to intron release. Here, we report that activation of Prp43’s inherently feeble helicase activity by the splicing factor Ntr1 is required for lariat-intron release. Lethal Prp43 mutants T384A and T384V, which are active for ATP hydrolysis and fail to dissociate lariat-intron from spliceosomes, are refractory to stimulation of RNA unwinding by Ntr1. An N-terminal 120-amino-acid segment of Ntr1 suffices for binding to Prp43 and for stimulating its helicase activity. We identify missense mutations in Prp43 and Ntr1 that disrupt protein–protein interaction and impair Ntr1 enhancement of Prp43 RNA unwinding. Our results demonstrate for the first time that regulating the motor activity of a DEAH-box protein by an accessory factor is critical for mRNA splicing.
Rational design of memory in eukaryotic cells
Authors: Ajo-Franklin Caroline M. , Drubin David A. , Eskin Julian A. , Gee Elaine P.S. , Landgraf Dirk , Phillips Ira , Silver Pamela A.
Published: September 2007

The ability to logically engineer novel cellular functions promises a deeper understanding of biological systems. Here we demonstrate the rational design of cellular memory in yeast that employs autoregulatory transcriptional positive feedback. We built a set of transcriptional activators and quantitatively characterized their effects on gene expression in living cells. Modeling in conjunction with the quantitative characterization of the activator-promoter pairs accurately predicts the behavior of the memory network. This study demonstrates the power of taking advantage of components with measured quantitative parameters to specify eukaryotic regulatory networks with desired properties.
N-cofilin is associated with neuronal migration disorders and cell cycle control in the cerebral cortex
Authors: Bellenchi Gian Carlo , Gurniak Christine B. , Perlas Emerald , Middei Silvia , Ammassari-Teule Martine , Witke Walter
Published: September 2007

Many neuronal disorders such as lissencephaly, epilepsy, and schizophrenia are caused by the abnormal migration of neurons in the developing brain. The role of the actin cytoskeleton in neuronal migration disorders has in large part remained elusive. Here we show that the F-actin depolymerizing factor n-cofilin controls cell migration and cell cycle progression in the cerebral cortex. Loss of n-cofilin impairs radial migration, resulting in the lack of intermediate cortical layers. Neuronal progenitors in the ventricular zone show increased cell cycle exit and exaggerated neuronal differentiation, leading to the depletion of the neuronal progenitor pool. These results demonstrate that mutations affecting regulators of the actin cytoskeleton contribute to the pathology of cortex development.
The prepattern transcription factor Irx3 directs nephron segment identity
Authors: Reggiani Luca , Raciti Daniela , Airik Rannar , Kispert Andreas , Brändli André W.
Published: September 2007

The nephron, the basic structural and functional unit of the vertebrate kidney, is organized into discrete segments, which are composed of distinct renal epithelial cell types. Each cell type carries out highly specific physiological functions to regulate fluid balance, osmolarity, and metabolic waste excretion. To date, the genetic basis of regionalization of the nephron has remained largely unknown. Here we show that Irx3, a member of the Iroquois (Irx) gene family, acts as a master regulator of intermediate tubule fate. Comparative studies in Xenopus and mouse have identified Irx1, Irx2, and Irx3 as an evolutionary conserved subset of Irx genes, whose expression represents the earliest manifestation of intermediate compartment patterning in the developing vertebrate nephron discovered to date. Intermediate tubule progenitors will give rise to epithelia of Henle’s loop in mammals. Loss-of-function studies indicate that irx1 and irx2 are dispensable, whereas irx3 is necessary for intermediate tubule formation in Xenopus. Furthermore, we demonstrate that misexpression of irx3 is sufficient to direct ectopic development of intermediate tubules in the Xenopus mesoderm. Taken together, irx3 is the first gene known to be necessary and sufficient to specify nephron segment fate in vivo.
X-ray powder diffraction structural characterization of Pb1 − x
Ba
x
Zr0.65Ti0.35O3 ceramic
Authors: Mir M. , Mastelaro V. R. , Neves P. P. , Doriguetto A. C. , Garcia D. , Lente M. H. , Eiras J. A. , Mascarenhas Y. P.
Published: September 2007

The structure of Pb1 − x
Ba
x
Zr0.65Ti0.35O3 (PBZT) ceramic materials with 0.00 ≤ x ≤ 0.40 was studied using synchrotron X-ray powder diffraction data. According to the Rietveld refinements, the structure of PBZT ceramics with x = 0.00, 0.10 and 0.20 at room temperature was rhombohedral R3c. A phase transition from rhombohedral to cubic was observed at 543 and 463 K for x = 0.10 and 0.20, respectively. The refinement for the compositions x = 0.30 and x = 0.40 showed a cubic structure from 10 to 450 K, in good agreement with the dielectric properties of these samples.
Knowledge-based model of hydrogen-bonding propensity in organic crystals
Authors: Galek Peter T. A. , Fábián László , Motherwell W. D. Samuel , Allen Frank H. , Feeder Neil
Published: September 2007

A new method is presented to predict which donors and acceptors form hydrogen bonds in a crystal structure, based on the statistical analysis of hydrogen bonds in the Cambridge Structural Database (CSD). The method is named the logit hydrogen-bonding propensity (LHP) model. The approach has a potential application in identifying both likely and unusual hydrogen bonding, which can help to rationalize stable and metastable crystalline forms, of relevance to drug development in the pharmaceutical industry. Whilst polymorph prediction techniques are widely used, the LHP model is knowledge-based and is not restricted by the computational issues of polymorph prediction, and as such may form a valuable precursor to polymorph screening. Model construction applies logistic regression, using training data obtained with a new survey method based on the CSD system. The survey categorizes the hydrogen bonds and extracts model parameter values using descriptive structural and chemical properties from three-dimensional organic crystal structures. LHP predictions from a fitted model are made using two-dimensional observables alone. In the initial cases analysed, the model is highly accurate, achieving ∼ 90% correct classification of both observed hydrogen bonds and non-interacting donor–acceptor pairs. Extensive statistical validation shows the LHP model to be robust across a range of small-molecule organic crystal structures.
Synthesis, characterization and phase transitions in the inorganic–organic layered perovskite-type hybrids [(C
n
H2n + 1NH3)2PbI4], n = 4, 5 and 6
Authors: Billing David G. , Lemmerer Andreas
Published: September 2007

Three inorganic–organic layered perovskite-type hybrids of the general formula [(C
n
H2n + 1NH3)2PbI4], n = 4, 5 and 6, display a number of reversible first-order phase transitions in the temperature range from 256 to 393 K. [(C4H9NH3)2PbI4] has a single phase transition, [(C5H11NH3)2PbI4] has two phase transitions and [(C6H13NH3)2PbI4] has three phase transitions. In all three cases, the lowest-temperature phase transition is thermochromic and the crystals change colour from yellow in their lowest-temperature phase to orange in their higher-temperature phase for [(C4H9NH3)2PbI4] and [(C6H13NH3)2PbI4], and from orange to red for [(C5H11NH3)2PbI4]. The structural details associated with this phase transition have been investigated via single-crystal X-ray diffraction, SC-XRD, for all three compounds.
Three-dimensional hydrogen-bonded frameworks in organic crystals: a topological study
Authors: Baburin Igor A. , Blatov Vladislav A.
Published: September 2007

1551 homomolecular single hydrogen-bonded frameworks in organic crystals have been classified into 148 topological types of three-periodic nets. Different representations of hydrogen-bonded frameworks as nets of molecular centroids, edge or ring nets are discussed. To study the influence of hydrogen bonds on the topology of molecular packings, 42 270 molecular crystals without hydrogen bonds have been considered. The topologies of molecular packings are found to be independent of hydrogen bonding. Analysis of 231 homomolecular frameworks composed of crystallographically different molecules shows that molecules not related by symmetry tend to form the same hydrogen-bond pattern. The relations between net topological types, space-group symmetry of crystals, site symmetry and point-group symmetry of molecules are discussed. As a result, a set of rules for the crystal design of molecular frameworks is proposed.
Inversion of 4f-states in CeB6 thermally excited at 430 K
Authors: Makita Ryoko , Tanaka Kiyoaki , Ōnuki Yoshichika , Tatewaki Hiroshi
Published: September 2007

The 4f states of Ce in a typical Kondo crystal, CeB6, are split into an excited state Γ7 and the ground state Γ8, with an excitation energy at 560 K. The electron-density distribution of the thermally excited state was measured at 430 K using a four-circle diffractometer equipped with a small furnace. In contrast to the previous results at lower temperature, electrons are transferred from B6 to Ce at 430 K. X-ray atomic-orbital analysis revealed that the 5d-Γ8 orbitals (the energy level of which is similar to that of the B-2p orbitals) are fully occupied and the 4f-Γ7 orbitals are more populated than the 4f-Γ8 orbitals. Fully occupied 5d-Γ8 makes the 4f-Γ8 states unstable and the energy levels of 4f-Γ7 and 4f-Γ8 are inverted.
Structures of tetrabromothiophene and tetrabromoselenophene: the influence of the heteroatom on the heterophene packing
Authors: Helmholdt Robert B. , Sonneveld Ed J. , Vande Velde Christophe M. L. , Blockhuys Frank , Lenstra Albert T. H. , Geise Herman J. , Peschar René
Published: September 2007

The crystal structures of C4Br4S and C4Br4Se have been determined from X-ray powder diffraction data, using direct-space search techniques. In the case of C4Br4S two crystalline phases occur, a stable orthorhombic and a metastable monoclinic phase. For the orthorhombic phase two different structural models were found that fit the experimental data equally well. The diversity in crystal structure models and packings of C4Br4S is explained.
Revision of pyrrhotite structures within a common superspace model
Authors: Izaola Zunbeltz , González Santiago , Elcoro Luis , Perez-Mato J. M. , Madariaga Gotzon , García Alberto
Published: September 2007

The structure of pyrrhotite (Fe1 − x
S with 0.05 ≤ x ≤ 0.125) has been reinvestigated in the framework of the superspace formalism. A common model with a centrosymmetric superspace group is proposed for the whole family. The atomic domains in the internal space representing the Fe atoms are parametrized as crenel functions that fulfil the closeness condition. The proposed model explains the x-dependent space groups observed and the basic features of the structures reported up to now. Our model yields for any x value a well defined ordered distribution of Fe vacancies in contrast to some of the structural models proposed in the literature. A new (3 + 1)-dimensional refinement of Fe0.91S using the deposited dataset [Yamamoto & Nakazawa (1982). Acta Cryst. A38, 79–86] has been performed as a benchmark of the model. The consistency of the proposed superspace symmetry and its validity for other compositions has been further checked by means of ab initio calculations of both atomic forces and equilibrium atomic positions in non-relaxed and relaxed structures, respectively.
Hexagonal ferrites: a unified model of the (TS)
n
T series in superspace
Authors: Orlov Ivan , Palatinus Lukáš , Arakcheeva Alla , Chapuis Gervais
Published: September 2007

Hexagonal ferrites represent an extensive family of mixed-layer magnetic materials with periods up to 1500 Å along the stacking direction, probably constituting the largest unit cells in the inorganic realm. The (TS)
n
T subfamily includes P\bar{3}m1 and R\bar{3}m structures that can be derived from Y ferrite Ba2
M
2Fe12O22 (M = Zn, Fe, Co, Mg, Mn) by introducing stacking faults. A unified (3 + 1)-dimensional superspace model is proposed for all members of the (TS)
n
T family. The model belongs to the superspace group X\bar{3}m1(00\gamma) with \textstyle X = \{\left({{1}\over{3}},{{2}\over{3}},0,{{1}\over{3}}\right), \textstyle\left({{2}\over{3}},{{1}\over{3}},0,{{2}\over{3}}\right)\}, has a unit cell of the basic structure with a = 5.88, c = 4.84 Å and modulation vector {\bf q} = {{4n+3}\over{9n+6}}{\bf c}^*, where n is rational for periodic structures and irrational for the aperiodic ones. The model was tested on calculated data of one of the principal members of the (TS)
n
T family, the Y ferrite (n = \infty). The fit obtained with the superspace model was excellent. The model allowed a reduction of refinable parameters by 19% with respect to the ordinary refinement without a significant increase of the refinement R values.
Structure determination of structurally complex Ag36Li64 gamma-brass
Authors: Noritake Tatsuo , Aoki Masakazu , Towata Shin-ichi , Takeuchi Tsunehiro , Mizutani Uichiro
Published: September 2007

The crystal structure of the Ag36Li64 gamma-brass was determined by analyzing the powder diffraction pattern taken using a synchrotron radiation beam with wavelength 0.50226 Å. It turned out that the compound contained 52 atoms in its unit cell with the space group I\bar 43m and that the Li atom enters exclusively into inner tetrahedral (IT) and cubo-octahedral (CO) sites, whereas the Ag atom enters into those on outer tetrahedral (OT) and octahedral (OH) sites in the 26-atom cluster. Small amounts of Li also exist in OT and OH sites, resulting in chemical disorder. We discovered that the volumes of the IT and CO polyhedra shrink, while those of the OT and OH polyhedra expand relative to those of the corresponding polyhedra in the original b.c.c. (body-centered cubic) structure. This feature is universal and is found in other gamma-brasses such as Cu5Zn8 and Al8V5, for which the structure data are available. Among these gamma-brasses, we revealed the unique bond-length distribution for pairs connecting the atom on OH sites and that on CO sites, depending on the degree of d–p orbital hybridization between the transition metal elements such as Ag, Cu and V on OH sites, and the non-transition metal elements such as Li, Zn and Al on CO sites. It is suggested that this may hold a clue to resolving why some gamma-brasses such as the present Ag–Li and Cu–Zn possess a finite solid solution, but others such as Al8V5 and Mn3In exist as line compounds.
Order–disorder transition in monoclinic sulfur: a precise structural study by high-resolution neutron powder diffraction. Corrigendum
Authors: David W. I. F. , Ibberson R. M. , Cox S. F. J. , Wood P. T.
Published: September 2007

Revised lattice parameters for Table 1 of the paper by David et al. (2006), Acta Cryst. B62, 953–959, are given.
Cell cycle‐dependent expression of potassium channels and cell proliferation in rat mesenchymal stem cells from bone marrow
Authors: Deng X. L. , Lau C. P. , Lai K. , Cheung K. F. , Lau G. K. , Li G. R.
Published: September 2007

Abstract.  Objective: Recently, our team has demonstrated that voltage‐gated delayed rectifier K+ current (IKDR) and Ca2+‐activated K+ current (IKCa) are present in rat bone marrow‐derived mesenchymal stem cells; however, little is known of their physiological roles. The present study was designed to investigate whether functional expression of IKDR and IKCa would change with cell cycle progression, and whether they could regulate proliferation in undifferentiated rat mesenchymal stem cells (MSCs). Materials and Methods: Membrane potentials and ionic currents were recorded using whole‐cell patch clamp technique, cell cycling was analysed by flow cytometry, cell proliferation was assayed with DNA incorporation method and the related genes were down‐regulated by RNA interference (RNAi) and examined using RT‐PCR. Results: It was found that membrane potential hyperpolarized, and cell size increased during the cell cycle. In addition, IKDR decreased, while IKCa increased during progress from G1 to S phase. RT‐PCR revealed that the mRNA levels of Kv1.2 and Kv2.1 (likely responsible for IKDR) reduced, whereas the mRNA level of KCa3.1 (responsible for intermediate‐conductance IKCa) increased with the cell cycle progression. Down‐regulation of Kv1.2, Kv2.1 or KCa3.1 with the specific RNAi, targeted to corresponding gene inhibited proliferation of rat MSCs. Conclusion: These results demonstrate that membrane potential, IKDR and IKCa channels change with cell cycle progression and corresponding alteration of gene expression. IKDR and intermediate‐conductance IKCa play an important role in maintaining membrane potential and they participate in modulation of proliferation in rat MSCs.
Inhibition of COX‐2 with NS‐398 decreases colon cancer cell motility through blocking epidermal growth factor receptor transactivation: possibilities for combination therapy
Authors: Banu N. , Buda A. , Chell S. , Elder D. , Moorghen M. , Paraskeva C. , Qualtrough D. , Pignatelli M.
Published: September 2007

Abstract.  The use of non‐steroidal anti‐inflammatory drugs has proved of great interest in the prevention and treatment of colorectal cancer, although their precise mechanisms of action remain unclear. Overexpression of cyclooxygenase‐2 (COX‐2) and subsequent prostaglandin production promote metastasis and have been shown to increase cell motility in vitro. Objective: We have aimed to elucidate whether specific inhibition of COX‐2 with NS‐398 (NS‐398 is a selective inhibitor of COX‐2) would be able to inhibit motility of colorectal cancer cells and whether this was modulated through epidermal growth factor receptor (EGFR) transactivation. Materials and Methods: A transwell filter assay was used to study cell motility. Expression of COX‐2, EGFR phosphorylation and prostaglandin E2 (PGE2) receptors were assessed by Western blot analysis and reverse transcriptase‐polymerase chain reaction. PGE2 concentrations after NS‐398 treatment were estimated by enzyme immunoassay. Results: Treatment with NS‐398 significantly reduced PGE2 levels and reduced cell migration in the HT29 and HCA7 colorectal carcinoma cell lines and this effect was rescued by addition of PGE2. Furthermore, specific inhibition of COX‐2 with NS‐398 reduced EGFR phosphorylation in colorectal cancer cells. Direct inhibition of EGFR activity with AG1478 reduced PGE2‐stimulated motility, clearly demonstrating that PGE2 acts via the EGFR‐signalling pathway. The novel combination of NS‐398 and AG1478 dramatically reduced migration of colorectal cancer cells. Conclusion: The data presented indicate that the use of NS‐398 in chemoprevention and adjuvant therapy for colorectal cancer may work in part, through the inhibition of cell motility. Furthermore, our data suggest that the combined use of non‐steroidal anti‐inflammatory drugs with EGFR antagonists could be explored further for future use in the clinic.
The effects of iron deficiency on neutrophil/monocyte apoptosis in children
Authors: Berrak S. G. , Angaji M. , Turkkan E. , Canpolat C. , Timur C. , Eksioglu‐Demiralp E.
Published: September 2007

Abstract.  Objectives: Iron is essential for DNA synthesis; its deficiency may lead to impaired DNA synthesis and subsequent alterations in levels of apoptosis. Here, we have aimed to investigate effects of iron deficiency anaemia (IDA) on apoptotic response of phagocytic cells and to understand whether the effect is reversible after iron supplementation. Materials and methods: Forty‐nine IDA patients and 26 healthy controls, aged between 6 months and 12 years with similar demographic status, were considered. Neutrophil‐ and monocyte‐apoptotic responses of IDA patients and the control group were compared by flow cytometry. Then, IDA patients were provided with oral iron supplementation. On day 15 of iron therapy, neutrophil‐ and monocyte‐apoptotic responses of IDA patients were rechecked and were compared to those of control group. Results: Neutrophil‐ and monocyte‐apoptotic responses in terms of early and late percentages of apoptosis, and percentages of necrotic cells, were significantly less in IDA patients compared to the control group. The significantly low apoptotic responses of IDA patients rose to levels of the control group by day 15 of iron therapy. Besides, the effect of IDA on apoptotic responses was found to be more enhanced in severe IDA patients that those of mild IDA patients. Conclusion: Correction of differences after iron supplementation therapy implies that IDA might be a cause for changes in neutophil‐ and monocyte‐apoptotic responses. The impact of this diminution of apoptotic cellular function in IDA should be further investigated, with longitudinal studies, in order to document the impact of any severe and/or long‐lasting IDA on autoimmunity and malignancy.
Simulated microgravity inhibits the proliferation and osteogenesis of rat bone marrow mesenchymal stem cells
Authors: Dai Z. Q. , Wang R. , Ling S. K. , Wan Y. M. , Li Y. H.
Published: September 2007

Abstract. Objectives: Microgravity is known to affect the differentiation of bone marrow mesenchymal stem cells (BMSCs). However, a few controversial findings have recently been reported with respect to the effects of microgravity on BMSC proliferation. Thus, we investigated the effects of simulated microgravity on rat BMSC (rBMSC) proliferation and their osteogeneic potential.Materials and methods: rBMSCs isolated from marrow using our established effective method, based on erythrocyte lysis, were identified by their surface markers and their proliferation characteristics under normal conditions. Then, they were cultured in a clinostat to simulate microgravity, with or without growth factors, and in osteogenic medium. Subsequently, proliferation and cell cycle parameters were assessed using methylene blue staining and flow cytometry, respectively; gene expression was determined using Western blotting and microarray analysis.Results: Simulated microgravity inhibited population growth of the rBMSCs, cells being arrested in the G0/G1phase of cell cycle. Growth factors, such as insulin‐like growth factor‐I, epidermal growth factor and basic fibroblastic growth factor, markedly stimulated rBMSC proliferation in normal gravity, but had only a slight effect in simulated microgravity. Akt and extracellular signal‐related kinase 1/2 phosphorylation levels and the expression of core‐binding factor α1 decreased after 3 days of clinorotation culture. Microarray and gene ontology analyses further confirmed that rBMSC proliferation and osteogenesis decreased under simulated microgravity.Conclusions: The above data suggest that simulated microgravity inhibits population growth of rBMSCs and their differentiation towards osteoblasts. These changes may be responsible for some of the physiological changes noted during spaceflight.
Human ovarian surface epithelial cells immortalized with hTERT maintain functional pRb and p53 expression
Authors: Li N. F. , Broad S. , Lu Y. J. , Yang J. S. , Watson R. , Hagemann T. , Wilbanks G. , Jacobs I. , Balkwill F. , Dafou D. , Gayther S. A.
Published: September 2007

Abstract. Objective: Cell immortalization is considered to be a prerequisite status for carcinogenesis. Normal human ovarian surface epithelial (OSE) cells, which are thought to be the origin of most of human ovarian carcinomas, have a very limited lifespan in culture. Establishment of immortalized OSE cell lines has, in the past, required inactivation of pRb and p53 functions. However, this often leads to increased chromosome instability during prolonged culture.Materials and Methods: In this study, we have used a retroviral infection method to overexpress human telomerase reverse transcriptase (hTERT) gene, in primary normal OSE cells, under optimized culture conditions.Results: In vitroandin vivoanalysis of hTERT‐immortalized cell lines confirmed their normal epithelial characteristics. Gene expression profiles and functional analysis of p16INK4A, p15INK4B, pRb and p53 confirmed the presence of their intact functions. Our study suggests that inactivation of pRb and p53 is not necessary for OSE immortalization. Furthermore, down‐regulation of p15INK4Bin the immortalized cells may indicate a functional role for this protein in them.Conclusion: These immortal OSE cell lines are likely to be an important tool for studying human OSE biology and carcinogenesis.
Skin keratinocytes pre‐treated with embryonic stem cell‐conditioned medium or BMP4 can be directed to an alternative cell lineage
Authors: Grinnell K. L. , Bickenbach J. R.
Published: September 2007

Abstract.  Objectives: In this study, we have investigated whether secreted factors from embryonic stem cells (ESCs) could reprogramme keratinocytes and increase their potential to be directed into alternative cell lineages. Materials and methods: Contact and non‐contact co‐cultures of skin keratinocytes and murine ESCs were used initially to confirm any reprogramming ability of ESC‐conditioned medium (CM). Immunofluoresence was used to assess nuclear expression of octamer‐4 (Oct‐4), as well as to confirm neuronal protein expression in neuroectodermally directed keratinocytes. Transcript expression changes were evaluated using semiquantitative reverse transcription‐polymerase chain reaction. Western blotting, accompanied by densitometry analysis, was used to evaluate protein expression following morphology changes. Results: We found that keratinocytes treated with ESC‐CM changed their morphology and were stimulated to express the pluripotency regulator, Oct‐4, and its target transcripts, Sox‐2, Nanog, Utf1 and Rex‐1. We demonstrate that at least one of the reprogramming factors is bone morphogenetic factor‐4 (BMP4). Pre‐treated keratinocytes could be specifically directed to differentiate into cells of the neuronal lineage. The majority of responsive keratinocytes were the epidermal stem cell population, with a small percentage of transit‐amplifying cells also being affected. Conclusions: Our results suggest that ESC‐CM contains a number of factors, including BMP4, which are capable of reprogramming mouse skin keratinocytes to make them more developmentally potent, as evidenced by their ability to be re‐differentiated into cells of the neuronal lineage. Our findings also imply a continuum of differentiation within the basal keratinocyte population. An increase in developmental potential combined with directed differentiation could increase the therapeutic relevancy of somatic cells.
Inhibition of NF‐κB activation by the histone deacetylase inhibitor 4‐Me2N‐BAVAH induces an early G1 cell cycle arrest in primary hepatocytes
Authors: Papeleu P. , Wullaert A. , Elaut G. , Henkens T. , Vinken M. , Laus G. , Tourwé D. , Beyaert R. , Rogiers V. , Vanhaecke T.
Published: September 2007

Abstract.  Objective: Benzoylaminoalkanohydroxamic acids, including 5‐(4‐dimethylaminobenzoyl)aminovaleric acid hydroxamide (4‐Me2N‐BAVAH), are structural analogues of Trichostatin A, a naturally occurring histone deacetylase inhibitor (HDACi). 4‐Me2N‐BAVAH has been shown to induce histone hyperacetylation and to inhibit proliferation in Friend erythroleukaemia cells in vitro. However, the molecular mechanisms have remained unidentified. Materials and Methods: In this study, we evaluated the effects of 4‐Me2N‐BAVAH on proliferation in non‐malignant cells, namely epidermal growth factor‐stimulated primary rat hepatocytes. Results and Conclusion: We have found that 4‐Me2N‐BAVAH inhibits HDAC activity at non‐cytotoxic concentrations and prevents cells from responding to the mitogenic stimuli of epidermal growth factor. This results in an early G1 cell cycle arrest that is independent of p21 activity, but instead can be attributed to inhibition of cyclin D1 transcription through a mechanism involving inhibition of nuclear factor‐kappaB activation. In addition, 4‐Me2N‐BAVAH delays the onset of spontaneous apoptosis in primary rat hepatocyte cultures as evidenced by down‐regulation of the pro‐apoptotic proteins Bid and Bax, and inhibition of caspase‐3 activation.
Plasma aldosterone response to the low‐dose adrenocorticotrophin (ACTH 1–24) stimulation test
Authors: Honour J. W. , Bridges N. A. , Conway‐Phillips E. , Hindmarsh P. C.
Published: September 2007

Background  Endocrine tests for adrenal insufficiency use pharmacological doses of stimulant such as ACTH. More physiological tests have often used high‐dose protocols for sampling frequency.

Aims  To evaluate the response of plasma aldosterone concentration to low doses (125, 250 and 500 ng/m2 body surface area) of synthetic ACTH.

Design  A randomised trial in six normal adult males aged 18–27 years.

Materials and methods  Aldosterone concentration was measured by radioimmunoassay in serum from blood samples taken at 10 min intervals for 90 min.

Results  All three doses produced a significant rise in plasma aldosterone concentration (125 ng/m2, P = 0·003; 250 ng/m2, P < 0·001; 500 ng/m2, P < 0·001) but there was no effect of dose on either the peak or incremental plasma aldosterone concentration. Mean time to peak was similar between the doses and the two higher doses were associated with a longer secretory profile (125 ng/m2 56 (26 SD) mins, 250 ng/m2 74 (19) mins, 500 ng/m2 77 (21) mins; F = 3·39; P = 0·04). Peaks of 100% were detected within 30 min of drug administration and peak response was associated with the prestimulation plasma aldosterone concentration (r = 0·45; P = 0·003). The between‐ and within‐individual coefficients of variation for prestimulation concentrations were 37·0% and 32·8%, and for the peak response were 27·2% and 27·2%, respectively.

Conclusions  The response of plasma aldosterone concentrations to low‐dose ACTH administration requires a blood sampling protocol of 0, 10, 20 and 30 min to capture concentrations near the peak response. The high‐dose protocol would have missed the response. Over the dose range studied no dose–response was observed so the selection of dose should be based on the dose effective to release steroids in the glucocorticoid pathway if this study is to be used in conjunction with such evaluation.
Testing the differential predation hypothesis for the invasion of rusty crayfish in a stream community: laboratory and field experiments
Authors: KUHLMANN MARK L. , BADYLAK STEPHANIE M. , CARVIN ERIN L.
Published: September 2007

1. We tested the hypothesis that the non‐native rusty crayfish (Orconectes rusticus) is less vulnerable to predators than two native species (O. propinquus and O. obscurus) it is replacing in streams of the upper Susquehanna River catchment (New York, U.S.A.).

2. We used laboratory experiments to compare species‐specific predation rates by smallmouth bass (Micropterus dolomieu) on crayfish of equal size and field tethering experiments to compare relative predation rates between native O. propinquus and non‐native O. rusticus by the suite of crayfish predators in our system. We predicted that crayfish size would affect predation rate but that predation rates would be equal among species when size was controlled.

3. We also tested for two potential artefacts of tethering. We tethered crayfish in cages to test whether the ability to escape from tethers is size specific, and we tested whether tethering alters differential predation among crayfish species by the smallmouth bass.

4. In the laboratory, smallmouth bass predation on rusty crayfish was lower than on either of the native species. In the field, predation risk for tethered crayfish was inversely related to size but did not differ among species when size was taken into account. Because rusty crayfish in the field experiment were slightly larger than the native species, as in nature, mortality was overall lower for the rusty crayfish.

5. In cages, smaller crayfish were more probably to escape from tethers than larger ones, an artefact that may partially confound results from our tethering experiments. Unexpectedly, tethering nearly eliminated predation by smallmouth bass. This artefact prevented us from testing for an interaction of tethering with differential predation and means that the results of field tethering experiments do not include any contribution from smallmouth bass predation.

6. Our experiments highlight the importance of explicitly considering potential artefacts that could confound results.

7. Our results indicate that differential predation contributes to the rusty crayfish's invasion of a stream community. In our study system, predation rates on rusty crayfish are lower than for native species mostly because of selection by predators for smaller crayfish; species‐specific characteristics such as behaviour that further reduce predation may also contribute, especially where smallmouth bass predation is important.
Effects of an induced drought on soil carbon dioxide (CO2) efflux and soil CO2 production in an Eastern Amazonian rainforest, Brazil
Authors: SOTTA ELENEIDE DOFF , VELDKAMP EDZO , SCHWENDENMANN LUITGARD , GUIMARÃES BRENDA ROCHA , PAIXÃO ROSIENE KEILA , RUIVO MARIA de LOURDES P. , LOLA da COSTA ANTONIO CARLOS , MEIR PATRICK
Published: September 2007

In the next few decades, climate of the Amazon basin is expected to change, as a result of deforestation and rising temperatures, which may lead to feedback mechanisms in carbon (C) cycling that are presently unknown. Here, we report how a throughfall exclusion (TFE) experiment affected soil carbon dioxide (CO2) production in a deeply weathered sandy Oxisol of Caxiuanã (Eastern Amazon). Over the course of 2 years, we measured soil CO2 efflux and soil CO2 concentrations, soil temperature and moisture in pits down to 3 m depth. Over a period of 2 years, TFE reduced on average soil CO2 efflux from 4.3±0.1 μmol CO2 m−2 s−1 (control) to 3.2±0.1 μmol CO2 m−2 s−1 (TFE). The contribution of the subsoil (below 0.5 m depth) to the total soil CO2 production was higher in the TFE plot (28%) compared with the control plot (17%), and it did not differ between years. We distinguished three phases of drying after the TFE was started. The first phase was characterized by a translocation of water uptake (and accompanying root activity) to deeper layers and not enough water stress to affect microbial activity and/or total root respiration. During the second phase a reduction in total soil CO2 efflux in the TFE plot was related to a reduction of soil and litter decomposers activity. The third phase of drying, characterized by a continuing decrease in soil CO2 production was dominated by a water stress‐induced decrease in total root respiration. Our results contrast to results of a drought experiment on clay Oxisols, which may be related to differences in soil water retention characteristics and depth of rooting zone. These results show that large differences exist in drought sensitivity among Amazonian forest ecosystems, which primarily seem to be affected by the combined effects of texture (affecting water holding capacity) and depth of rooting zone.
Modelling past and present geographical distribution of the marine gastropodPatella rusticaas a tool for exploring responses to environmental change
Authors: LIMA FERNANDO P. , RIBEIRO PEDRO A. , QUEIROZ NUNO , XAVIER RAQUEL , TARROSO PEDRO , HAWKINS STEPHEN J. , SANTOS ANTÓNIO M.
Published: September 2007

A climate envelope approach was used to model the distributions of the intertidal gastropodPatella rustica, to test the robustness of forecast responses to climate change. The model incorporated variables that were likely to determine the abundance and the northern range limit of this species in the NE Atlantic. The model was built using classification and regression tree analysis (CART) trained with historical distribution data from the mid 1950s and a set of corresponding climatic and oceanographic variables. Results indicated air and sea temperature, in particular during the reproductive and settlement periods, as the main determinants of the Atlantic distribution ofP. rustica. The model was subsequently fed with contemporary climatic data and its output was compared with the current distribution and abundance ofP. rustica, assessed during a 2002–2003 survey. The model correctly hindcasted the recent collapse of a distributional gap in northern Portugal, as well as an increase in abundance at locations within its range. The predicted northward expansion of the northern range limit did not occur because the absence of the species was confirmed in a survey encompassing the whole Atlantic French coast up to Brest. Stretches of unsuitable habitat too long to be overcome by dispersal are the likely mechanism controlling the northern limit of the distribution of this intertidal species.