bluuebunny/crossref_metadata_2025 huggingface dataset:
Recruitment of DNA methyltransferase I to DNA repair sites
Authors: Mortusewicz Oliver , Schermelleh Lothar , Walter Joachim , Cardoso M. Cristina , Leonhardt Heinrich
Published: June 2005

In mammalian cells, the replication of genetic and epigenetic information is directly coupled; however, little is known about the maintenance of epigenetic information in DNA repair. Using a laser microirradiation system to introduce DNA lesions at defined subnuclear sites, we tested whether the major DNA methyltransferase (Dnmt1) or one of the two
de novo
methyltransferases (Dnmt3a, Dnmt3b) are recruited to sites of DNA repair
in vivo
. Time lapse microscopy of microirradiated mammalian cells expressing GFP-tagged Dnmt1, Dnmt3a, or Dnmt3b1 together with red fluorescent protein-tagged proliferating cell nuclear antigen (PCNA) revealed that Dnmt1 and PCNA accumulate at DNA damage sites as early as 1 min after irradiation in S and non-S phase cells, whereas recruitment of Dnmt3a and Dnmt3b was not observed. Deletion analysis showed that Dnmt1 recruitment was mediated by the PCNA-binding domain. These data point to a direct role of Dnmt1 in the restoration of epigenetic information during DNA repair.
Probing charge transport at the single-molecule level on silicon by using cryogenic ultra-high vacuum scanning tunneling microscopy
Authors: Guisinger Nathan P. , Yoder Nathan L. , Hersam Mark C.
Published: June 2005

A cryogenic variable-temperature ultra-high vacuum scanning tunneling microscope is used for measuring the electrical properties of isolated cyclopentene molecules adsorbed to the degenerately p-type Si(100)-2×1 surface at a temperature of 80 K. Current–voltage curves taken under these conditions show negative differential resistance at positive sample bias, in agreement with previous observations at room temperature. Because of the enhanced stability of the scanning tunneling microscope at cryogenic temperatures, repeated measurements can be routinely taken over the same molecule. Taking advantage of this improved stability, we show that current–voltage curves on isolated cyclopentene molecules are reproducible and possess negligible hysteresis for a given tip–molecule distance. On the other hand, subsequent measurements with variable tip position show that the negative differential resistance voltage increases with increasing tip–molecule distance. By using a one-dimensional capacitive equivalent circuit and a resonant tunneling model, this behavior can be quantitatively explained, thus providing insight into the electrostatic potential distribution across a semiconductor-molecule-vacuum-metal tunnel junction. This model also provides a quantitative estimate for the alignment of the highest occupied molecular orbital of cyclopentene with respect to the Fermi level of the silicon substrate, thus suggesting that this experimental approach can be used for performing chemical spectroscopy at the single-molecule level on semiconductor surfaces. Overall, these results serve as the basis for a series of design rules that can be applied to silicon-based molecular electronic devices.
Premature ovarian aging in mice deficient for
Gpr3
Authors: Ledent Catherine , Demeestere Isabelle , Blum David , Petermans Julien , Hämäläinen Tuula , Smits Guillaume , Vassart Gilbert
Published: June 2005

After becoming competent for resuming meiosis, fully developed mammalian oocytes are maintained arrested in prophase I until ovulation is triggered by the luteotropin surge. Meiotic pause has been shown to depend critically on maintenance of cAMP level in the oocyte and was recently attributed to the constitutive Gs (the heterotrimeric GTP-binding protein that activates adenylyl cyclase) signaling activity of the G protein-coupled receptor GPR3. Here we show that mice deficient for
Gpr3
are unexpectedly fertile but display progressive reduction in litter size despite stable age-independent alteration of meiotic pause. Detailed analysis of the phenotype confirms premature resumption of meiosis,
in vivo
, in about one-third of antral follicles from
Gpr3

–
/
–

females, independently of their age. In contrast, in aging mice, absence of GPR3 leads to severe reduction of fertility, which manifests by production of an increasing number of nondeveloping early embryos upon spontaneous ovulation and massive amounts of fragmented oocytes after superovulation. Severe worsening of the phenotype in older animals points to an additional role of GPR3 related to protection (or rescue) of oocytes from aging.
Gpr3
-defective mice may constitute a relevant model of premature ovarian failure due to early oocyte aging.
Unzipped and loaded
Authors: Skibbens Robert V.
Published: June 2005

It is well known that the products of chromosome replication are paired to ensure that the sisters segregate away from each other during mitosis. A key issue is how cells pair sister chromatids but preclude the catastrophic pairing of nonsister chromatids. The identification of both replication factor C and DNA helicases as critical for sister chromatid pairing has brought new insights into this fundamental process.
Importance of integrin LFA-1 deactivation for the generation of immune responses
Authors: Semmrich Monika , Smith Andrew , Feterowski Carolin , Beer Sandra , Engelhardt Britta , Busch Dirk H. , Bartsch Bernadett , Laschinger Melanie , Hogg Nancy , Pfeffer Klaus , Holzmann Bernhard
Published: June 2005

The dynamic regulation of ligand binding is considered crucial for integrin function. However, the importance of activity regulation for integrin function in vivo is largely unknown. Here, we have applied gene targeting to delete the GFFKR sequence of the lymphocyte function-associated antigen–1 (LFA-1) αL subunit cytoplasmic domain in mouse germline. Lymphocytes from Lfa-1d/d mutant mice showed constitutive activation of LFA-1–mediated cell adhesion and impaired de-adhesion from intercellular adhesion molecule-1 that resulted in defective cell migration. In contrast, signaling through LFA-1 was not affected in Lfa-1d/d cells. T cell activation by superantigen-loaded and allogeneic APCs, cytotoxic T cell activity, T-dependent humoral immune responses, and neutrophil recruitment during aseptic peritonitis were impaired in Lfa-1d/d mice. Thus, deactivation of LFA-1 and disassembly of LFA-1–mediated cell contacts seem to be vital for the generation of normal immune responses.
An Electrostatic Engine Model for Autoinhibition and Activation of the Epidermal Growth Factor Receptor (EGFR/ErbB) Family
Authors: McLaughlin Stuart , Smith Steven O. , Hayman Michael J. , Murray Diana
Published: June 2005

We propose a new mechanism to explain autoinhibition of the epidermal growth factor receptor (EGFR/ErbB) family of receptor tyrosine kinases based on a structural model that postulates both their juxtamembrane and protein tyrosine kinase domains bind electrostatically to acidic lipids in the plasma membrane, restricting access of the kinase domain to substrate tyrosines. Ligand-induced dimerization promotes partial trans autophosphorylation of ErbB1, leading to a rapid rise in intracellular [Ca2+] that can activate calmodulin. We postulate the Ca2+/calmodulin complex binds rapidly to residues 645–660 of the juxtamembrane domain, reversing its net charge from +8 to −8 and repelling it from the negatively charged inner leaflet of the membrane. The repulsion has two consequences: it releases electrostatically sequestered phosphatidylinositol 4,5-bisphosphate (PIP2), and it disengages the kinase domain from the membrane, allowing it to become fully active and phosphorylate an adjacent ErbB molecule or other substrate. We tested various aspects of the model by measuring ErbB juxtamembrane peptide binding to phospholipid vesicles using both a centrifugation assay and fluorescence correlation spectroscopy; analyzing the kinetics of interactions between ErbB peptides, membranes, and Ca2+/calmodulin using fluorescence stop flow; assessing ErbB1 activation in Cos1 cells; measuring fluorescence resonance energy transfer between ErbB peptides and PIP2; and making theoretical electrostatic calculations on atomic models of membranes and ErbB juxtamembrane and kinase domains.
Functionally Active T1-T1 Interfaces Revealed by the Accessibility of Intracellular Thiolate Groups in Kv4 Channels
Authors: Wang Guangyu , Shahidullah Mohammad , Rocha Carmen A. , Strang Candace , Pfaffinger Paul J. , Covarrubias Manuel
Published: June 2005

Gating of voltage-dependent K+ channels involves movements of membrane-spanning regions that control the opening of the pore. Much less is known, however, about the contributions of large intracellular channel domains to the conformational changes that underlie gating. Here, we investigated the functional role of intracellular regions in Kv4 channels by probing relevant cysteines with thiol-specific reagents. We find that reagent application to the intracellular side of inside-out patches results in time-dependent irreversible inhibition of Kv4.1 and Kv4.3 currents. In the absence or presence of Kv4-specific auxiliary subunits, mutational and electrophysiological analyses showed that none of the 14 intracellular cysteines is essential for channel gating. C110, C131, and C132 in the intersubunit interface of the tetramerization domain (T1) are targets responsible for the irreversible inhibition by a methanethiosulfonate derivative (MTSET). This result is surprising because structural studies of Kv4-T1 crystals predicted protection of the targeted thiolate groups by constitutive high-affinity Zn2+ coordination. Also, added Zn2+ or a potent Zn2+ chelator (TPEN) does not significantly modulate the accessibility of MTSET to C110, C131, or C132; and furthermore, when the three critical cysteines remained as possible targets, the MTSET modification rate of the activated state is ∼200-fold faster than that of the resting state. Biochemical experiments confirmed the chemical modification of the intact α-subunit and the purified tetrameric T1 domain by MTS reagents. These results conclusively demonstrate that the T1–T1 interface of Kv4 channels is functionally active and dynamic, and that critical reactive thiolate groups in this interface may not be protected by Zn2+ binding.
INEFFICIENT STANDARD ADOPTION: INERTIA AND MOMENTUM REVISITED
Authors: CLEMENTS MATTHEW T.
Published: June 2005

This article examines the possibility that consumers will adopt an inefficient standard. When there are successive generations of consumers, the current generation will not consider the costs and benefits to past and future generations of adopting a new standard. If a standard is proprietary, the incentives of a firm to induce adoption of the standard generally do not match the social incentives. The divergence is caused by the firm's imperfect ability to appropriate the future surplus generated by the standard. (JEL D62, L1)
DO ECONOMICS DEPARTMENTS SEARCH OPTIMALLY IN FACULTY RECRUITING?
Authors: CHEN YONMIN , MCKINNISH TERRA
Published: June 2005

We find that it is optimal for higher‐quality departments to search broadly across many or even all fields in faculty recruiting, whereas it is optimal for lower‐quality departments to conduct narrower searches. We develop a simple search model in which optimal search scope is shown to increase in department quality. Using data from Job Openings for Economists, we find that higher‐ranked departments do conduct broader searches. We find that a 10‐place increase in department ranking is associated with 3.5–4.8 more Journal of Economic Literature subfields listed in a position announcement. (JEL J44, J64, D83, L8)
Livestock grazing affects the egg size of an insectivorous passerine
Authors: Evans Darren M , Redpath Stephen M , Evans Sharon A , Elston David A , Dennis Peter
Published: June 2005

Livestock grazing is a major driver of ecosystem change, and has been associated with significant declines in various bird species worldwide. In Britain, there is particular concern that severe grazing pressure is deleteriously affecting vegetation and birds in upland regions. However, the mechanism by which grazing affects birds is unclear. Here, we report for the first time, to our knowledge, that sheep grazing pressure affects the egg size of a common upland passerine: the meadow pipit
Anthus pratensis
. We manipulated sheep stocking densities in a replicated field experiment, and found that plots with the highest stocking density contained nests with the smallest eggs, and that plots with low stocking density contained nests with the largest eggs. However, eggs laid in ungrazed plots were also small, suggesting that either too many sheep or their removal from upland areas might have a detrimental effect on pipit egg size. We found no significant effect on fledging success but the reduced post-fledging survival of young from smaller eggs, as seen in other studies, could partly explain declines in upland birds.
Environmental stochasticity in dispersal areas can explain the ‘mysterious’ disappearance of breeding populations
Authors: Penteriani Vincenzo , Otalora Fermín , Sergio Fabrizio , Ferrer Miguel
Published: June 2005

We present the results of an individual-based simulation model, showing that increasing the mortality of non-breeding dispersers within settlement areas can lead to the extinction of species and (meta)populations in a subtle way. This is because the areas where dispersers settle are generally unknown or difficult to detect. Consequently, fewer efforts are devoted to the conservation of these sites than to the conservation of breeding territories. Additionally, high mortality rates affecting the floater sector of a population become evident in the breeding sector only after several of years, when it is too difficult or too late to halt the decline. As a result, because most conservation projects on endangered species and populations mainly focus on breeding areas, many current efforts may be wasted in locations other than those in which conservation would be really necessary and effective.
Why are small males aggressive?
Authors: Morrell Lesley J , Lindström Jan , Ruxton Graeme D
Published: June 2005

Aggression is ubiquitous in the animal kingdom, whenever the interests of individuals conflict. In contests between animals, the larger opponent is often victorious. However, counter intuitively, an individual that has little chance of winning (generally smaller individuals) sometimes initiates contests. A number of hypotheses have been put forward to explain this behaviour, including the ‘desperado effect’ according to which, the likely losers initiate aggression due to lack of alternative options. An alternative explanation suggested recently is that likely losers attack due to an error in perception: they mistakenly perceive their chances of winning as being greater than they are. We show that explaining the apparently maladaptive aggression initiated by the likely loser can be explained on purely economic grounds, without requiring either the desperado effect or perception errors. Using a game-theoretical model, we show that if smaller individuals can accurately assess their chance of winning, if this chance is less than, but close to, a half, and if resources are scarce (or the contested resource is of relatively low value), they are predicted to be as aggressive as their larger opponents. In addition, when resources are abundant, and small individuals have some chance of winning, they may be more aggressive than their larger opponents, as it may benefit larger individuals to avoid the costs of fighting and seek alternative uncontested resources.
Supply-side invasion ecology: characterizing propagule pressure in coastal ecosystems
Authors: Verling Emma , Ruiz Gregory M , Smith L. David , Galil Bella , Miller A. Whitman , Murphy Kathleen R
Published: June 2005

The observed rates and deleterious impacts of biological invasions have caused significant alarm in recent years, driving efforts to reduce the risk (establishment) of new introductions. Characterizing the supply of propagules is key to understanding invasion risk and developing effective management strategies. In coastal ecosystems, ships' ballast water is an important transfer mechanism (vector) for marine and freshwater species. Commercial ships exhibit a high degree of variation in ballast water operations that affect both the quantity and quality of propagule supply, and thereby invasion risk. The per-ship inoculation size from ballast water depends upon both the volume discharged and the organism density. Moreover, propagule quality will vary among source regions (ports) and voyage routes, due to differences in species composition and transport conditions, respectively. We show that significant differences exist in (i) the frequency and volume of ballast water discharge among vessel types, (ii) the frequency of vessel types and routes (source regions) among recipient ports, and (iii) the transit success (survivorship) of zooplankton in ballast tanks among voyage routes. Thus, propagule supply is not a simple function of total ship arrivals. For ships, as well as other vectors, variation in propagule quantity and quality must be explicitly considered to estimate invasion risk and advance predictive ability.
An approach for genogrouping of Japanese isolates of aquabirnaviruses in a new genogroup, VII, based on the VP2/NS junction region
Authors: Nishizawa Toyohiko , Kinoshita Shinichi , Yoshimizu Mamoru
Published: June 2005

Aquabirnaviruses, represented by Infectious pancreatic necrosis virus (IPNV), have been isolated from epizootics in salmonids and a variety of aquatic animals in the world; six genogroups of aquabirnaviruses have been identified. In comparisons of nucleotide sequences of the VP2/NS junction region, maximum nucleotide diversities of 30·8 % were observed among 93 worldwide aquabirnavirus isolates. A phylogenetic tree revealed the existence of a new genogroup, VII, for Japanese aquabirnavirus isolates from marine fish and molluscan shellfish. Nucleotide diversities between genogroups VII and I–VI were 18·7 % or greater. At the nucleotide level, Japanese IPNV isolates from epizootics in salmonids were nearly identical to a genogroup I strain from the USA or Canada. It is suggested that Japanese IPNV isolates belonging to genogroup I were originally introduced from North American sources, whereas Japanese aquabirnavirus isolates of genogroup VII were from marine aquatic animals indigenous to Japan.
Isolation and cloning of the raccoon (Procyon lotor) papillomavirus type 1 by using degenerate papillomavirus-specific primers
Authors: Rector Annabel , Van Doorslaer Koenraad , Bertelsen Mads , Barker Ian K. , Olberg Rolf-Arne , Lemey Philippe , Sundberg John P. , Van Ranst Marc
Published: June 2005

Partial sequences of a novel papillomavirus were amplified from a cutaneous lesion biopsy of a raccoon (Procyon lotor), by using PCR with degenerate papillomavirus-specific primers. The Procyon lotor papillomavirus type 1 (PlPV-1) DNA was amplified with long template PCR in two overlapping fragments, together encompassing the entire genome, and the complete PlPV-1 genomic sequence was determined. The PlPV-1 genome consists of 8170 bp, and contains the typical papillomaviral open reading frames, encoding five early proteins and two late capsid proteins. Besides the classical non-coding region (NCR1) between the end of L1 and the start of E6, PlPV-1 contains an additional non-coding region (NCR2) of 1065 bp between the early and late protein region, which has previously also been described for the canine oral papillomavirus (COPV) and the Felis domesticus papillomavirus (FdPV-1). Phylogenetic analysis places PlPV-1 together with COPV and FdPV-1 in a monophyletic branch which encompasses the Lambda papillomavirus genus.
Deletion of open reading frames 9, 10 and 11 from the avian adenovirus CELO genome: effect on biodistribution and humoral responses
Authors: Le Goff Frédérick , Méderlé-Mangeot Isabelle , Jestin André , Langlois Patrick
Published: June 2005

In this study, the in vivo effect of the 3·6 kbp deletion of the three open reading frames (ORF) 9, 10 and 11 found at the right end of the CELO genome was examined. Groups of chickens were inoculated oronasally with 105–107 p.f.u. per animal of wild-type virus and two recombinant CELO strains (rCELO) expressing luciferase and secreted alkaline phosphatase (SEAP). The tissue biodistribution, assessed by PCR, was similar for both wild-type and recombinant viruses. The infectious viral particle titre was determined by a p.f.u. counting method and the antibody responses to the CELO vector and the SEAP antigen were evaluated by ELISA. Infectious particle titres in tissues from chickens inoculated with the wild-type CELO virus increased up to 6 days post-inoculation, and declined until 11 days while titres in organs from chickens inoculated with the rCELO strain were low and only detectable at 4 days post-inoculation. Moreover, although anti-CELO antibody levels were three times lower in sera from chickens inoculated with rCELO, antibodies directed to the heterologous SEAP antigen were detected. Based on these results, no differences in tropism were observed, but the level of production of viral particles and the humoral responses appeared to decrease. Viruses replicate less efficiently with a deletion performed at the right end of the CELO genome. Nevertheless, the presence of antibodies directed to heterologous antigens makes the CELO virus an advantageous candidate for avian vaccination.
Analysis of larval antigens of Oestrus ovis for the diagnosis of oestrosis by enzyme‐linked immunosorbent assay
Authors: Alcaide M. , Reina D. , Frontera E. , Navarrete I.
Published: June 2005

Abstract.  A serodiagnostic test for the diagnosis of infestation by the sheep nasal bot fly, Oestrus ovis (Linné) was examined. The enzyme‐linked immunosorbent assay (ELISA) technique was used to analyze and compare the production of immunoglobulin G (IgG) antibodies against excretory‐secretory products (ESP) and crude extract (CE) antigens from all the different larval stages of O. ovis in the sera of 276 adult sheep sampled in summer (n = 135) and winter (n = 141). ESP from first stage larvae was the most sensitive, coating antigen in winter and ESP from second stage larvae during summer. The most specific values were obtained by ESP against L1 in winter and by CE against L3 in summer. These results show that the stage of larval development has a significant impact on the humoral immune response over the course of a season. A significant correlation (P < 0.001) was found between the number of O. ovis larvae and the serum antibody levels using all differents antigens, except L3 CE. In Spain, where a long favourable period exists for the evolution and development of the different stage larvae between March and November, the ELISA test using L1 ESP antigen during winter and L2 ESP antigen in summer may be used for ovine oestrosis immunodiagnosis.
Duration of development of two species of carrion‐breeding scuttle flies and forensic implications
Authors: Disney R. H. L.
Published: June 2005

Abstract.  A procedure for rearing carrion‐breeding scuttle flies (Diptera: Phoridae) under naturally fluctuating temperatures is described. Data on the duration of development of the egg stage, egg plus feeding larval stages, postfeeding larvae and total development from egg to adult, at different temperature regimes (means of daily means) are tabulated for Megaselia giraudii (Egger) and M. rufipes (Meigen). The variation is greater than previously supposed and involves larvae derived from the same egg batch leaving the bait in successive waves. Despite this variation the tables allow the provision of percentage probabilities of estimations of the earliest oviposition dates for specimens collected from a forensic case.
Ectopic bone formation facilitated by human mesenchymal stem cells and osteogenic cytokines via nutrient vessel injection in a nude rat model
Authors: Fukui Masashi , Akita Sadanori , Akino Kozo
Published: June 2005

In vivo studies using bone marrow‐derived mesenchymal stem cells are still uncommon. Applications for bone defect replacement in undesirable clinical circumstances such as large defects, bacterial or other pathogen‐contaminated fields, and irradiated surgical wound bed necessitate vascularized bone regeneration. Use of a fascial flap including regenerated bone would be a very powerful tool for treatment. It would be especially beneficial in cases where normal bone regeneration is not expected due to a lack of sufficient blood supply, extensive surgical scaring, or bacterial contamination. In this study, we used nude rats in which the superficial epigastric flap of the experimental group was used to wrap around a mixture of human mesenchymal stem cells, bone morphogenetic protein‐2, and basic fibroblast growth factor cytokines in a gelatin carrier. These rats showed significantly higher bone mineral density at 4 weeks compared to the other experimental groups containing phosphate buffered saline, human mesenchymal stem cells alone, or the two cytokines alone (p < 0.01). There were no remarkable histologic differences up to 7 days. At 2 weeks, more progressive vascularity and perivascular tissue deposits were seen in the experimental group. Basophilic mineral structure surrounded the fibroblast‐like mesenchymal stem cells at 4 weeks, presumably osteoblastic or osteoclastic cell lining. Bone marker immunohistochemistry against alkaline phosphatase and osteocalcin revealed diffuse and distinct immunoreactivity in osteoblastic cells in the experimental group at 4 weeks. Further transcriptional expression of polyomavirus enhancer binding protein 2αA suggested that the human transplanted cells proceeded to osteogenic lineage in 4 weeks. These results may be useful as a new approach for bone regeneration.
Intraepidermal nerve fiber density in rat foot pad: neuropathologic–neurophysiologic correlation
Authors: Lauria Giuseppe , Lombardi Raffaella , Borgna Monica , Penza Paola , Bianchi Roberto , Savino Costanza , Canta Annalisa , Nicolini Gabriella , Marmiroli Paola , Cavaletti Guido
Published: June 2005

Abstract  Quantification of cutaneous innervation in rat footpad is a useful tool to investigate sensory small‐diameter nerve fibers, which are affected early in peripheral neuropathies. The aim of this work was to provide normative reference data on the density of intraepidermal nerve fibers (IENFs) and Langerhans cells in the hindpaw footpad of Sprague–Dawley and Wistar rats. We also evaluated the sensibility of IENF density by comparing neuropathologic findings with neurophysiologic examination and the presence of peripheral neuropathy in two well‐characterized animal models of neuropathy. IENF density was quantified in 22 Sprague–Dawley rats and 13 Wistar rats and compared with 19 age‐matched Sprague–Dawley rats with streptozotocin‐induced diabetic neuropathy and 30 age‐matched Wistar rats with cisplatin‐ or paclitaxel‐induced neuropathy. Antidromic tail sensory nerve conduction velocity (SNCV) was assessed in all animals. IENF and Langerhans cell densities were constant in healthy Sprague–Dawley rats at any age, and they were similar to those observed in healthy Wistar rats. In neuropathic rats, both SNCV and IENF density were significantly reduced with respect to controls. Quantification of IENF density was significantly correlated with changes in conduction velocity. Diabetic neuropathy rats alone showed a significantly higher density of Langerhans cells compared with controls. Our study demonstrated that IENF density quantification correlates with SNCV changes and suggests that this might represent a useful outcome measurement in experimental neuropathies.
Cepharanthine potently enhances the sensitivity of anticancer agents in K562 cells
Authors: Ikeda Ryuji , Che Xiao‐Fang , Yamaguchi Tatsuya , Ushiyama Mina , Zheng Chun‐Lei , Okumura Hiroshi , Takeda Yasuo , Shibayama Yoshihiko , Nakamura Kazuo , Jeung Hei‐Cheul , Furukawa Tatsuhiko , Sumizawa Tomoyuki , Haraguchi Misako , Akiyama Shin‐ichi , Yamada Katsushi
Published: June 2005

A major impediment to cancer treatment is the development of resistance by the tumor. P‐glycoprotein (P‐gp) and multidrug resistance protein 1 (MRP1) are involved in multidrug resistance. In addition to the extrusion of chemotherapeutic agents through these transporters, it has been reported that there are differences in the intracellular distribution of chemotherapeutic agents between drug resistant cells and sensitive cells. Cepharanthine is a plant alkaloid that effectively reverses resistance to anticancer agents. It has been previously shown that cepharanthine is an effective agent for the reversal of resistance in P‐gp‐overexpressing cells. Cepharanthine has also been reported to have numerous pharmacological effects besides the inhibition of P‐gp. It has also been found that cepharanthine enhanced sensitivity to doxorubicin (ADM) and vincristine (VCR), and enhanced apoptosis induced by ADM and VCR of P‐gp negative K562 cells. Cepharanthine changed the distribution of ADM from cytoplasmic vesicles to nucleoplasm in K562 cells by inhibiting the acidification of cytoplasmic organelles. Cepharanthine in combination with ADM should be useful for treating patients with tumors. (Cancer Sci 2005; 96: 372–376)
Chemokines in tumor progression and metastasis
Authors: Tanaka Toshiyuki , Bai Zhongbin , Srinoulprasert Yuttana , Yang BoGi , Hayasaka Haruko , Miyasaka Masayuki
Published: June 2005

Although chemokines have been thought of primarily as leukocyte attractants, a growing body of evidence indicates that they also contribute to a number of tumor‐related processes, such as tumor cell growth, angiogenesis/angiostasis, local invasion, and metastasis. The current knowledge of the possible involvement of chemokines and their receptors in these cellular events are reviewed here. The operating mechanism of chemokines in relation to metastatic processes in vivo are also discussed. (Cancer Sci 2005; 96: 317–322)
Effects of lengthening contraction on calcium kinetics and skeletal muscle contractility in humans
Authors: Nielsen J. S. , Madsen K. , Jørgensen L. V. , Sahlin K.
Published: June 2005

Aim: We have tested the hypothesis that the altered muscle contractility after lengthening contractions (LC) is caused by altered calcium (Ca2+) kinetics.

Methods: Subjects (n = 8) performed 100 drop jumps and muscle contractility was measured pre‐ and post‐exercise by maximal voluntary contraction (MVC) and transcutaneous electrical stimulation (1, 20 and 50 Hz). Muscle biopsies were analysed for muscle metabolites, rates of SR Ca2+uptake (CaU) and release (CaR) and myosin heavy chain (MHC) composition.

Results: The rates of torque relaxation and CaU were positively related to muscle fibre type composition (% MHC II). Muscle creatine (Cr) decreased and the ratio between phosphocreatine (PCr) and Cr increased 3 and 24 h post‐exercise (P < 0.05 vs. pre‐exercise). LC resulted in reduced MVC (−19%), twitch torque (−41%) and 20/50 Hz torque ratio (−30%) and a faster relaxation rate (P < 0.05). The contractile parameters recovered partially but remained altered 24 h post‐exercise (P < 0.05). The average CaR was unchanged after LC (P > 0.05). However, the response varied between subjects and the relative post‐exercise CaR was significantly related to the degree of LFF (post/pre 20/50 Hz force ratio) and to the decline in twitch force (post/pre twitch ratio). CaU was lower in seven of eight subjects after LC (P > 0.05).

Conclusion: The decline in torque after LC could not be explained by metabolic factors since PCr/Cr ratio increased. The relation between CaR and fatigue suggests that the mechanism of fatigue in part may be attributed to intrinsic changes in the SR Ca2+release channel. The faster torque relaxation after LC could not be explained by an increased rate of CaU.
Anti-inflammatory properties of heat shock protein 70 and butyrate on Salmonella-induced interleukin-8 secretion in enterocyte-like Caco-2 cells
Authors: Malago J J , Koninkx J F J G , Tooten P C J , Van Liere E A , Van Dijk J E
Published: June 2005

Intestinal epithelial cells secrete the chemokine interleukin (IL)-8 in the course of inflammation. Because heat shock proteins (Hsps) and butyrate confer protection to enterocytes, we investigated whether they modulate Salmonella enterica serovar Enteritidis (S. serovar Enteritidis)-induced secretion of IL-8 in enterocyte-like Caco-2 cells. Caco-2 cells incubated with or without butyrate (0–20 m M, 48 h) were infected with S. serovar Enteritidis after (1 h at 42°C, 6 h at 37°C) or without prior heat shock (37°C). Levels of Hsp70 production and IL-8 secretion were analysed using immunostaining of Western blots and enzyme-linked immunosorbent assay (ELISA), respectively. The cells secreted IL-8 in response to S. serovar Enteritidis and produced Hsp70 after heat shock or incubation with butyrate. The IL-8 secretion was inhibited by heat shock and butyrate concentrations as low as 0·2 m M for crypt-like and 1 m M for villous-like cells. In a dose-dependent manner, higher butyrate concentrations enhanced IL-8 secretion to maximal levels followed by a gradual but stable decline. This decline was associated with increasing production of Hsp70 and was more vivid in crypt-like cells. In addition, the higher concentrations abolished the heat shock inhibitory effect. Instead, they promoted the IL-8 production in heat-shocked cells even in the absence of S. serovar Enteritidis. We conclude that heat shock and low concentrations of butyrate inhibit IL-8 production by Caco-2 cells exposed to S. serovar Enteritidis. Higher butyrate concentrations stimulate the chemokine production and override the inhibitory effect of the heat shock. The IL-8 down-regulation could in part be mediated via production of Hsp70.
Major immunoreactive domains of human ribosomal P proteins lie N-terminal to a homologous C-22 sequence: application to a novel ELISA for systemic lupus erythematosus
Authors: Lin J L J , Dubljevic V , Fritzler M J , Toh Ban-Hock
Published: June 2005

The aim of this study was to identify immunoreactive domains on human ribosomal P0, P1 and P2 proteins, other than the C-22 peptide, to develop a novel ELISA using a combination of these proteins and to compare this ELISA with one using the C-22 peptide. Human recombinant P0, P1, P2 and mutant P0 lacking the homologous C-22 peptide (N-P0) were produced in bacteria and tested by ELISA and immunoblotting using sera from 48 patients with systemic lupus erythematosus (SLE), 48 with an unrelated inflammatory disorder (Crohn's disease) and 47 healthy controls. ELISA with P0, P1 and P2, premixed at equimolar concentrations, gave higher OD readings than each protein tested individually. Eighteen SLE sera tested positive by ELISA with premixed P0, P1, P2 but only 3 tested positive with the C-22 peptide. Twenty-two SLE sera reacted positively, as determined by immunoblotting, with 5 different P protein combinations: P1P2, P0P1P2, P1, P0P1, P0 and P1. Only sera reactive with all three P proteins reacted with the C-22 peptide, with absent or minimal reactivity with N-P0. Native antigens yielded sensitivity (6/48, 13%) similar to the C-22 peptide assay. An ELISA with premixed P1 and P2 gave higher OD values than the arithmetic means with P1 or P2. Fifteen SLE patients had antibodies to double stranded (ds)-DNA, of which 6 also had antibodies to P0P1P2 by ELISA but 12 reactive with P0P1P2 did not have discernable ds-DNA antibodies. Ribosomal P autoantibodies react mainly with epitopes N-terminal to a homologous C-22 peptide. An ELISA with premixed P0, P1 and P2 has 5-fold greater sensitivity (38%) for SLE than an assay with the conventional C-22 peptide (7%). The combined sensitivity for SLE for antibodies to P0P1P2 and ds-DNA is 56%, higher than C-22 and ds-DNA, 38%. Only one of the SLE patients had neuropsychiatric lupus.
Innate immune responses in Lyme borreliosis: enhanced tumour necrosis factor-α and interleukin-12 in asymptomatic individuals in response to live spirochetes
Authors: Sjöwall J , Carlsson A , Vaarala O , Bergström S , Ernerudh J , Forsberg P , Ekerfelt C
Published: June 2005

Innate immunity is important for early defence against borrelia spirochetes and should play a role in the clinical outcome of the infection. In order to study early cytokine responses, in vitro differentiated dendritic cells (DCs) and whole blood cells from 21 patients with different clinical outcomes of Lyme neuroborreliosis were stimulated with live borrelia spirochetes. The borrelia-induced secretion of interleukin (IL)-4, IL-10, IL-12p70, interferon (IFN)-γ and tumour necrosis factor (TNF)-α in DCs and IL-1β, IL-6, IL-8, IL-10, IL-12p70, TNF-α, regulated upon activation normal T cell expressed and secreted (RANTES), monocyte chemoattractant protein (MCP)-1, macrophage inflammatory protein (MIP)-1α, MIP-1β and eotaxin in whole blood cells was measured by enzyme-linked immunospot (ELISPOT) and multiplex arrays, respectively. We found increased numbers of TNF-α-secreting DCs (P = 0·018) in asymptomatic seropositive individuals compared to patients with subacute neuroborreliosis and seronegative controls. Asymptomatic individuals were also found to have elevated levels of IL-12p70 (P = 0·031) in whole blood cell supernatants compared to seronegative controls. These results are in line with previous experiments using cells of the adaptive immune response, indicating that strong T helper type 1 (Th1) proinflammatory responses might be associated with a successful resolution of Lyme disease.
Gene polymorphisms and serological markers of patients with active Crohn's disease in a clinical trial of antisense to ICAM-1
Authors: Yacyshyn B R , Schievella A , Sewell K L , Tami J A
Published: June 2005

Serological profiles for anti-Saccharomyces cerevisiae antibodies (ASCA)/ perinuclear antineutrophil cytoplasmic antibodies (pANCA) and gene polymorphisms in tumour necrosis factor (TNF)-α and intercellular adhesion molecule-1 (ICAM-1) are associated with occurrence and/or outcome in Crohn's disease. The aim of the study was to characterize the ASCA/pANCA profile, soluble ICAM-1 expression and single nucleotide gene polymorphisms (SNPs) in TNF-α and ICAM-1 genes. Crohn's patients with moderate disease activity were enrolled in a clinical trial of Alicaforsen (ISIS 2302). Peripheral blood samples were collected prospectively for serum studies and for potential analysis of gene polymorphisms. A multivariate analysis was performed to compare treatment effect with the biomarkers studied. Serological testing for ASCA/pANCA was obtained for 257 patients at baseline: 37% were ASCA+/pANCA– (Crohn's pattern), 9% had both markers, 15% were ASCA–/pANCA+ and 39% had neither marker. When the data were analysed by multiple regression analysis, a trend was found within the Alicaforsen-treated groups for greater rates of remission in the ASCA+/pANCA– subgroup versus all other serological profiles (25 versus 14%, P = 0·068), but not versus the placebo remission rate (18·8%). Gene polymorphisms were assessed in 64 patients, 21 from the placebo group. ICAM-1 assessment revealed no over-representation. However, three unique TNF-α SNPs were identified that correlated significantly with remission; sites 290 (P = 0·0253), −2735 (P = 0·0317) and −3090 (P = 0·0067). Although the overall clinical trial was negative, we have identified a trend towards clinical remission with Alicaforsen therapy in a subgroup of patients with Crohn's disease expressing ASCA+/pANCA–. Furthermore, we have identified three TNF-α SNPs that may also predict a positive therapeutic outcome.
Mainland colonization by island lizards
Authors: Nicholson Kirsten E. , Glor Richard E. , Kolbe Jason J. , Larson Allan , Blair Hedges S. , Losos Jonathan B.
Published: June 2005

Aim We investigate biogeographic relationships within the lizard genusAnolisDaudin, 1802 to test the hypothesis that the mainland (Central and South American) Norops‐clade species descended from a West IndianAnolisancestor. Previous hypotheses have suggested that close island relatives of mainland Norops species (the CubanAnolis sagreiand JamaicanA. grahamiseries) represent over‐water dispersal from a mainland ancestor. These previous hypotheses predict that theA. sagreiandA. grahamiseries should be phylogenetically nested within a Norops clade whose ancestral geography traces to the mainland. If Norops is West Indian in origin, then West Indian species should span the deepest phylogenetic divergences within the Norops clade.

Location Central and South America and West Indian islands.

Methods The phylogenetic relationships ofAnolislizards are reconstructed from aligned DNA sequences using both parsimony and Bayesian approaches. Hypotheses are tested in two ways: (1) by reconstructing the ancestral geographic location for the Norops clade usingPagel & Lutzoni's (2002)Bayesian approach, and (2) by testing alternative topological arrangements via Wilcoxon Signed‐Ranks tests (Templeton, 1983) and Shimodaira–Hasegawa tests (Shimodaira & Hasegawa, 1999).

Results Our evidence supports an origin of mainland Norops anoles from a West Indian ancestor. A West Indian ancestor to the Norops clade is statistically supported, and alternatives to the biogeographic pattern [Cuban (Jamaican, Mainland)] are statistically rejected by Shimodaira–Hasegawa tests, although not by Wilcoxon Signed‐Ranks tests.

Main conclusions Our data support the hypothesis of a West Indian origin for mainland Norops. This result contradicts previous hypotheses and suggests that island forms may be an important source for mainland biodiversity.
Probability samples of area births versus clinic populations for reproductive epidemiology studies
Authors: Savitz David A. , Dole Nancy , Kaczor Diane , Herring Amy H. , Siega‐Riz Anna Maria , Kaufman Jay , Thorp John M.
Published: June 2005

Studies of pregnancy outcome are generally based either on geographically defined populations, often from birth records, or on clinic or hospital populations. We compared women recruited into a clinic‐based study, the Pregnancy, Infection, and Nutrition (PIN) Study, with women who resided in the geographical area of the study (Alamance, Orange, and Wake Counties in North Carolina) and gave birth over the corresponding time period (1996–2000). Clinic participants were more likely to be Black, younger, have lower education, be unmarried, have a more frequent history of adverse pregnancy outcome, obtain prenatal care later, and smoke more cigarettes. Despite that profile, the proportion of clinic participants delivering preterm was somewhat lower than among area women overall (10.8% vs. 11.3%). Black/White risk ratios for preterm birth were markedly different for area (1.6) vs. clinic women (1.1), whereas other predictors were similar. Patterns may differ across groups for many reasons, including self‐selection of clinics and varying clinical practices.
Predictors of vaginal bleeding during the first two trimesters of pregnancy
Authors: Yang Juan , Savitz David A. , Dole Nancy , Hartmann Katherine E. , Herring Amy H. , Olshan Andrew F. , Thorp John M.
Published: June 2005

This study evaluates maternal age, race, cigarette smoking, prior spontaneous abortion, prior induced abortion, and prior preterm birth in relation to vaginal bleeding during the first two trimesters of pregnancy. Information on vaginal bleeding and predictors came from the Pregnancy, Infection, and Nutrition Study, which enrolled 2806 pregnant women at 24–29 weeks’ gestation during 1995–2000 in central North Carolina, USA. Generalised estimating equations were applied to take into account repeated episodes of vaginal bleeding during pregnancy. Women with advanced maternal age and passive smoking exposure were more likely to experience more intense vaginal bleeding during pregnancy, as were women with prior preterm birth. More intense bleeding was also more likely to be reported among women with multiple prior spontaneous abortions or multiple prior induced abortions, but not among women with a single prior spontaneous or induced abortion. The combination of prior spontaneous and induced abortion showed a dose–response association with the occurrence of vaginal bleeding during pregnancy.
Spatial and temporal variation of reactive oxygen species and antioxidant enzymes in o‐hydroxyethylorutin‐treated tomato leaves inoculated with Botrytis cinerea
Authors: Małolepsza U.
Published: June 2005

Pretreatment of detached tomato leaves with o‐hydroxyethylorutin reduced the percentage leaf area affected, and delayed the appearance of necrosis, following inoculation with conidial suspensions in droplets of the grey mould fungus Botrytis cinerea. o‐Hydroxyethylorutin delayed, but did not inhibit, in vitro germination of conidia, although overall percentage germination was reduced compared with water controls. Both the reactive oxygen species (ROS) – superoxide anions and hydrogen peroxide – increased twice as much in o‐hydroxyethylorutin‐treated leaf tissue 2 and 6 h postinoculation with B. cinerea conidia in tissues under inoculation drops, as well as in surrounding tissues, whereas in plants not pretreated with the compound ROS generation was noticed later, and only in tissues under inoculation drops. Compared with these compounds, changes in the levels of hydroxyl radicals, lipid peroxidation and the activity of the enzymes superoxide dismutase, guaiacol peroxidase, ascorbate peroxidase and catalase were largely unchanged. In stimulating ROS in inoculated tomato tissue, B. cinerea appeared to be affected directly pre‐ and postinfection, but indirect effects increasing host resistance cannot be excluded.
Comparative analysis of early H2O2 accumulation in compatible and incompatible wheat–powdery mildew interactions
Authors: Li A. L. , Wang M. L. , Zhou R. H. , Kong X. Y. , Huo N. X. , Wang W. S. , Jia J. Z.
Published: June 2005

Comparative analysis was carried out for hydrogen peroxide (H2O2) accumulation in response to powdery mildew (Blumeria graminis[Erysiphe graminis] f.sp. tritici) primary germ tube (PGT) and appressorial germ tube (AGT) contact, and in papilla (cell wall apposition) and hypersensitive responses (HR) in wheat (Triticum aestivum). Using primary leaves of three susceptible wheat lines (Bainong 3217; Beijing 837; Jingshuang 16) and five resistant lines [Mardler (Pm2 + Pm6); Ulka/8 × Cc (Pm2); Mardler/7 × Bainong 3217 (Pm2); Pm16; Pm16/7 × Beijing 837 (Pm16)], early H2O2 accumulation between 10 and 48 h after inoculation was studied. Strong H2O2 accumulation was found in effective papillae and associated cytosolic vesicles in both susceptible and resistant wheat lines, suggesting the important role of H2O2 in effective papillae formed as a general plant defence against powdery mildew. High frequency of effective papilla formation was observed in all five resistant lines. Among resistant lines, hypersensitive cell death was detected earlier in Mardler and Ulka/8 × Cc than in Pm16 and Pm16/7 × Beijing 837. In all cases this was associated with H2O2 accumulation in attacked epidermal cells. Interestingly, penetration resistance but not HR appeared to be mediated by the Pm2 gene in the Mardler/7 × Bainong 3217 line, suggesting that Pm2 may govern an HR‐independent defence pathway in this genetic background. As effective papillae and HR did not occur in the same cells, papilla deposition may be independent of the HR response, despite the fact that both defence mechanisms were associated with high H2O2 accumulation. In wheat powdery mildew‐incompatible interactions, HR acts as a second line of defence to contain infection when the papilla defence fails.