bluuebunny/crossref_metadata_2025 huggingface dataset:
Attention‐deficit/hyperactivity disorder (ADHD) as a risk factor for persistent nocturnal enuresis in children: A two‐year follow‐up study
Authors: BAEYENS DIETER , ROEYERS HERBERT , DEMEYERE ISOLDE , VERTÉ SYLVIE , HOEBEKE PIET , WALLE JOHAN VANDE
Published: November 2005

Aims: A previous prevalence study indicated that the prevalence of ADHD is highly increased in enuretic children. In the current 2‐y follow‐up study we investigate the relationship between both disorders further. Our goal is to determine whether the ADHD diagnoses can be reconfirmed and whether children with ADHD are more at risk for difficult‐to‐cure enuresis. Moreover, we explore the effect of medical enuresis parameters on the course of the voiding problem.

Methods: Eighty‐six children with enuresis were screened twice on the presence of ADHD with a 2‐y interval. A multi‐method, multi‐informant assessment of ADHD was used, the child's medical file was consulted, and a parent questionnaire on the child's current voiding problems was completed.

Results: Although 73% of all children with a 2‐y‐old diagnosis of ADHD still meet the disorder's criteria, only 66% of all subtype diagnoses can be reconfirmed. The odds that a child with ADHD still has voiding problems after 2 y are 3.17 times higher than for a child without ADHD. Although a slightly increased number of prescribed therapies in the ADHD group was noticed, no other significant differences in enuresis treatment methods were found between the groups. The medical parameters were not associated with treatment outcome.

Conclusion: Since 73% of ADHD diagnoses can be reconfirmed, the data suggest that the prevalence of the ADHD syndrome rather than reactive ADHD symptomatology is increased in enuretic children. Children with ADHD are at risk for persistent enuresis. Two‐year‐old medical enuresis parameters seem to have little effect on the current presence/absence of enuresis.
Emerging teacher strategies for mediating ‘Technology‐integrated Instructional Conversations’: a socio‐cultural perspective
Authors: Hennessy Sara , Deaney Rosemary , Ruthven Kenneth
Published: November 2005

This article draws on socio‐cultural learning theory as a conceptual framework for analysing how teachers structure classroom activities and interactions during ‘Technology‐integrated Instructional Conversations’ (TICs). It reports on a collaborative programme of small‐scale projects undertaken by 15 teacher‐researchers using various forms of computer‐based ICT to support subject teaching and learning. The participants developed, trialled and refined new pedagogic approaches and activities in six curriculum areas (English, classics, design technology, geography, history, science) at secondary level. A cross‐case analysis was conducted using lesson observations, follow‐up teacher interviews and teachers' research reports.

A typology of proactive and responsive pedagogic strategies for mediating pupil interactions with ICT was identified. These included exploiting the technology in new ways and circumventing its associated constraints. The strategies emerging illustrated how teachers structured activities judiciously; supported, guided and challenged; encouraged pupil collaboration, experimentation, reflection and analysis; avoided floundering and maintained a focus on subject learning; integrated the use of other resources; and developed information handling skills. These strategies and the gradual withdrawal of teacher support served to increase pupil participation and responsibility for their own learning. Pupils themselves played a role in structuring TICs through opportunistically soliciting teacher assistance and feedback.
Selective accumulation of differentiated CD8+ T cells specific for respiratory viruses in the human lung
Authors: de Bree Godelieve J. , van Leeuwen Ester M.M. , Out Theo A. , Jansen Henk M. , Jonkers René E. , van Lier René A.W.
Published: November 2005

The lungs are frequently challenged by viruses, and resident CD8+ T cells likely contribute to the surveillance of these pathogens. To obtain insight into local T cell immunity to respiratory viruses in humans, we determined the specificity, phenotype, and function of lung-residing CD8+ T cells and peripheral blood CD8+ T cells in a paired analysis. The lung contained markedly higher frequencies of influenza (FLU)-specific and respiratory syncytial virus (RSV)-specific CD8+ T cells when compared with the circulation. This contrasted with an equal distribution of cytomegalovirus- and Epstein-Bar virus–specific CD8+ T cells. Noticeably, a substantial fraction of the lung-residing FLU- and RSV-specific CD8+ T cells had progressed to a relatively late differentiation phenotype, reflected by low expression of CD28 and CD27. Lung-derived FLU-specific CD8+ T cells had low activation requirements, as expansion of these cells could be initiated by cognate peptide in the absence of helper cell–derived signals. Thus, the human lung contains high numbers of differentiated FLU- and RSV-specific memory CD8+ T cells that can readily expand upon reexposure to virus. Resident lung T cells may provide immediate immunological protection against pulmonary virus infections.
Where CD4+CD25+ T reg cells impinge on autoimmune diabetes
Authors: Chen Zhibin , Herman Ann E. , Matos Michael , Mathis Diane , Benoist Christophe
Published: November 2005

Foxp3 is required for the generation and activity of CD4+CD25+ regulatory T (T reg) cells, which are important controllers of autoimmunity, including type-1 diabetes. To determine where T reg cells affect the diabetogenic cascade, we crossed the Foxp3 scurfy mutation, which eliminates T reg cells, with the BDC2.5 T cell receptor (TCR) transgenic mouse line. In this model, the absence of T reg cells did not augment the initial activation or phenotypic characteristics of effector T cells in the draining lymph nodes, nor accelerate the onset of T cell infiltration of the pancreatic islets. However, this insulitis was immediately destructive, causing a dramatic progression to overt diabetes. Microarray analysis revealed that T reg cells in the insulitic lesion adopted a gene expression program different from that in lymph nodes, whereas T reg cells in draining or irrelevant lymph nodes appeared very similar. Thus, T reg cells primarily impinge on autoimmune diabetes by reining in destructive T cells inside the islets, more than during the initial activation in the draining lymph nodes.
PKCβ regulates BCR-mediated IKK activation by facilitating the interaction between TAK1 and CARMA1
Authors: Shinohara Hisaaki , Yasuda Tomoharu , Aiba Yuichi , Sanjo Hideki , Hamadate Megumi , Watarai Hiroshi , Sakurai Hiroaki , Kurosaki Tomohiro
Published: November 2005

The B cell antigen receptor (BCR)–mediated activation of IκB kinase (IKK) and nuclear factor–κB require protein kinase C (PKC)β; however, the mechanism by which PKCβ regulates IKK is unclear. Here, we demonstrate that another protein kinase, TGFβ-activated kinase (TAK)1, is essential for IKK activation in response to BCR stimulation. TAK1 interacts with the phosphorylated CARMA1 (also known as caspase recruitment domain [CARD]11, Bimp3) and this interaction is mediated by PKCβ. IKK is also recruited to the CARMA1–Bcl10–mucosal-associated lymphoid tissue 1 adaptor complex in a PKCβ-dependent manner. Hence, our data suggest that phosphorylation of CARMA1, mediated by PKCβ, brings two key protein kinases, TAK1 and IKK, into close proximity, thereby allowing TAK1 to phosphorylate IKK.
The Molecular Immunology of Mucositis: Implications for Evidence‐Based Research in Alternative and Complementary Palliative Treatments
Authors: Chiappelli Francesco
Published: November 2005

The terms ‘mucositis’ and ‘stomatitis’ are often used interchangeably. Mucositis, however, pertains to pharyngeal‐esophago‐gastrointestinal inflammation that manifests as red, burn‐like sores or ulcerations throughout the mouth. Stomatitis is an inflammation of the oral tissues proper, which can present with or without sores, and is made worse by poor dental hygiene. Mucositis is observed in a variety of immunosuppressed patients, but is most often consequential to cancer therapy. It appears as early as the third day of intervention, and is usually established by Day 7 of treatment. Mucositis increases mortality and morbidity and contributes to rising health care costs. The precise immune components involved in the etiology of mucositis are unclear, but evidence‐based research (EBR) data has shown that applications of granulocyte–macrophage‐colony stimulating factor prevent the onset or the exacerbation of oropharyngeal mucositis. The molecular implications of this observation are discussed from the perspective of future developments of complementary and alternative treatments for this condition. It must be emphasized that this article is meant to be neither a review on mucositis and the various treatments for it, nor a discussion paper on its underlying molecular immunology. It is a statement of the implications of EBR for CAM‐based interventions for mucositis. It explores and discusses the specific domain of molecular immunology in the context of mucositis and its direct implications for EBR research in CAM‐based treatments for mucositis.
Immunology and Homeopathy. 1. Historical Background
Authors: Bellavite Paolo , Conforti Anita , Piasere Valeria , Ortolani Riccardo
Published: November 2005

Homeopathy was born as an experimental discipline, as can be seen from the enormous amount of homeopathic data collected over more than two centuries. However, the medical tradition of homeopathy has been separated from that of conventional science for a long time. Conventional scientific wisdom dictates that homeopathy should have no effect above placebo but experiments on ultra‐high dilutions of solutes together with some clinical data suggest the intriguing possibility that it might do in some circumstances. Today, an osmotic process between disciplines, previously seen as in conflict, is facilitated because over the last few decades homeopathy has initiated the methods of current medical science and a substantial number of experimental studies—at molecular, cellular and clinical levels—are available. One area of dialogue and of common progress is that of inflammation and immunity, probably because these are closely related to the traditional ‘vital force’ of the body′s self‐healing power. In a series of papers we review the historical origins of homeopathy, the laboratory and animal models related to the field of immunopharmacology, the clinical evidence in favor and against the use of homeopathy in the inflammatory diseases and the hypotheses regarding its action mechanism(s). Finally, we will enlighten the specific characteristics of the homeopathic approach, which places great emphasis on identifying a cure for the whole organism.
The pharmacokinetic and pharmacodynamic consequences of the co‐administration of lamotrigine and a combined oral contraceptive in healthy female subjects
Authors: Sidhu Jagdev , Job Sarah , Singh Sunita , Philipson Richard
Published: November 2005

Aims

To assess the pharmacokinetic and pharmacodynamic effects of co‐administration of a combined oral contraceptive (ethinyloestradiol plus levonorgestrel) and lamotrigine.

Methods

Over a period of 130 days, healthy female subjects took lamotrigine (titrated up to 300 mg day−1) and the combined oral contraceptive, either individually or as co‐therapy. Plasma ethinyloestradiol and levonorgestrel concentrations were measured in the presence and absence of lamotrigine, and serum lamotrigine concentrations were measured in the presence and absence of the combined oral contraceptive. Serum concentrations of follicle‐stimulating hormone (FSH), luteinizing hormone (LH), progesterone, oestradiol and sex hormone binding globulin were also determined.

Results

Of the 22 enrolled subjects, 16 had evaluable pharmacokinetic data. The mean (90% CI) ratios of lamotrigine area under the concentration‐time curve from 0 to 24 h (AUC(0,24 h)) and maximum observed concentration (Cmax) of lamotrigine when it was given with the combined oral contraceptive and during monotherapy were 0.48 (0.44, 0.53) and 0.61 (0.57, 0.66), respectively. Ethinyloestradiol pharmacokinetics were unchanged by lamotrigine, the mean combined oral contraceptive + lamotrigine : combined oral contraceptive alone ratios (90% CI) of the AUC(0,24 h) and Cmax of levonorgestrel were 0.81 (0.76, 0.86) and 0.88 (0.82, 0.93), respectively. FSH and LH concentrations were increased (by 4.7‐fold and 3.4‐fold, respectively) in the presence of lamotrigine, but the low serum progesterone concentrations suggested that suppression of ovulation was maintained. Intermenstrual bleeding was reported by 7/22 (32%) of subjects during co‐administration of lamotrigine and combined oral contraceptive.

Conclusions

A clinically relevant pharmacokinetic interaction was observed during co‐administration of a combined oral contraceptive and lamotrigine. A dosage adjustment for lamotrigine may need to be considered when these agents are co‐administered. A modest decrease in the plasma concentration of levonorgestrel was also observed but there was no corresponding hormonal evidence of ovulation.
Refractory anaemia with excess of blasts (RAEB): analysis of reclassification according to the WHO proposals
Authors: Germing Ulrich , Strupp Corinna , Kuendgen Andrea , Aivado Manuel , Giagounidis Aristoteles , Hildebrandt Barbara , Aul Carlo , Haas Rainer , Gattermann Norbert
Published: November 2005

The French–American–British (FAB) classification assigns patients with myelodysplastic syndromes to the category of refractory anaemia with excess blasts (RAEB) if they have a medullary blast count of 5–20%, and/or a peripheral blast count of 2–5%. The new World Health Organization (WHO) classification subdivides RAEB into RAEB I with a medullary blast count ≤10% and a peripheral blast count ≤5% and RAEB II with >10% medullary and/or >5% peripheral blasts. RAEB II is also diagnosed if Auer rods are present. In 558 patients, we analysed these subtypes of RAEB in terms of haematological characteristics, karyotype anomalies and prognosis. RAEB I was diagnosed in 256 and RAEB II in 302 patients. In the RAEB II group, 22% of patients had >5% peripheral blasts or the presence of Auer rods. The median survival was 16 months for RAEB I as compared with 9 months for RAEB II. Patients with Auer rods, regardless of their medullary and peripheral blast count, had no worse prognosis. No significant differences were identified between the RAEB subtypes with respect to clinical, morphological, haematological and cytogenetic parameters. The survival data support the WHO reclassification of RAEB based on peripheral and medullary blast counts and Auer rods. The WHO classification is useful for diagnosis and provides risk stratification, supported by cytogenetic data for clinical decision making, identifying those RAEB patients with an unfavourable prognosis who should be offered chemotherapy or stem cell transplantation.
Soluble thrombomodulin is antithrombotic in the presence of neutralising antibodies to protein C and reduces circulating activated protein C levels in primates
Authors: Tanaka Kenichi A. , Fernández José A. , Marzec Ulla M. , Kelly Andrew B. , Mohri Mitsunobu , Griffin John H. , Hanson Stephen R. , Gruber Andras
Published: November 2005

We studied whether there was a relationship between the anticoagulant effects of recombinant human soluble thrombomodulin (rhsTM) and activation of protein C in a primate model of acute vascular graft thrombosis in 11 baboons (Papio species). Baboons were pretreated with 0·1, 1 and 5 mg/kg of rhsTM, with or without co‐injection of a neutralising monoclonal antibody to protein C (HPC4) in the 1 mg/kg rhsTM group. Subsequently, thrombogenic polyester grafts were deployed for 3 h into chronic exteriorised arteriovenous shunts. Thrombus growth in the graft, plasma‐activated protein C (APC) levels, coagulation and thrombosis markers were determined. In untreated baboons, baseline circulating APC levels more than doubled and graft thrombi propagated until reaching equilibrium in about 1 h. Treatment with rhsTM reduced thrombus propagation rates, prolonged the clotting and bleeding times, decreased thrombin–antithrombin complex, β‐thromboglobulin and fibrinopeptide A levels, and, surprisingly, also decreased systemic APC levels, in a dose‐dependent manner. In the presence of HPC4 antibody to inhibit APC generation, the acute antithrombotic activity of rhsTM on graft thromboses was not attenuated for up to 80 min, but sustained thrombus accumulation was observed over a 180‐min period. These findings suggest that, in contrast to the prevailing hypotheses, the primary antithrombotic activity of rhsTM is independent of protein C, at least in this primate model. Direct inhibition of thrombin's prothrombotic activity upon complex formation with rhsTM might explain the molecular mechanism of the observed antithrombotic effect.
Prevalence of high‐grade CIN following mild dyskaryotic smears in different age groups
Authors: Giannopoulos T. , Butler‐Manuel S. , Tailor A. , Demetriou E. , Daborn L.
Published: November 2005

Objective:  The new guidelines of the British Society of Colposcopy and Cervical Pathology suggest that women should be offered colposcopy after only one mildly dyskaryotic smear. This is expected to generate increased workload for the colposcopy clinics, at least in the short term. The main objective of this study was to estimate the incidence of high‐grade cervical intraepithelial neoplasia (CIN) in women with mildly dyskaryotic smears and investigate whether there is any variation in different age groups. The rationale was to determine whether we could reduce the burden on colposcopy services by prioritizing the mild dyskaryotic referrals by age, as we hypothesized that high‐grade CIN is less frequent in younger women.

Methods:  The study sample included all women who were referred for colposcopy with a cervical smear suggesting mild dyskaryosis (with or without koilocytosis) from April 2000 to March 2003.

Results:  We studied 510 women. They were divided into three age groups (<20, 20–25 and >25 years). The overall prevalence of high‐grade CIN (CIN II and III) was 28.7%.The positive predictive value of a mildly dyskaryotic smear for high‐grade CIN was similar in all groups.

Conclusions:  Our results show that we are not in a position to prioritize our referrals by age group and reduce the initial pressure for colposcopies. We are also concerned that with the implementation of the new guidelines, a significant number of women <25 years will be carrying high‐grade CIN for more than 5 years before they are first screened.
Outcome of women with index smear showing mild dyskaryosis: effects of age and evidence of HPV infection
Authors: Winn C. M. , Jones H.
Published: November 2005

Objective:  Recent national guidelines (NHSCSP Document 20) recommend colposcopy referral after one mildly dyskaryotic smear, compared with the current practice of cytological surveillance and referral if the abnormality persists. The aim of this study was to identify the percentage of women whose first abnormal smear, showing mild dyskaryosis, returned to normal with cytological surveillance. Colposcopy could therefore be avoided in this group. This study also assessed whether age or human papillomavirus (HPV) status affected this outcome and the impact of non‐attenders on the reliability of surveillance.

Methods:  This was a retrospective study examining the follow‐up of 1484 women whose first abnormal smear showed mild dyskaryosis between 1996 and 1998. The possible outcomes were: persisting abnormality referred to colposcopy, follow‐up by cytology alone (negative follow‐up), lost to follow‐up or moved out of the area. Results were further assessed in terms of age (over or under 35 years) and cytological evidence of HPV effect.

Results:  In this study 50.9% of women, presenting with a mildly dyskaryotic smear, returned to normal without colposcopy within the follow‐up period of 6–8 years. Age (over/under 35) or cytological evidence of HPV did not significantly affect this figure.

Conclusions:  Immediate colposcopy would overtreat 50% of the study group resulting in 159 extra colposcopies in this unit per year. High‐grade abnormalities were twice as prevalent (22% versus 11%) in the younger age group, suggesting that younger rather than older women would benefit from immediate referral.
Distinct modes of DNA damage response in S. pombe G0 and vegetative cells
Authors: Mochida Satoru , Yanagida Mitsuhiro
Published: November 2005

Upon nitrogen‐starvation, mostly G2 vegetative (VE) fission yeast cells promote two rounds of division and enter the G0 state with 1C DNA via an uncommitted G1. Whilst G0 cells are permanently arrested, they keep viability through recycling the intracellular nitrogen. We here show that, whilst the DNA damages are efficiently repaired in G0 cells, neither Chk1 activation nor Cdc2 implication for Crb2 (53BP1 like) do not occur. ATR‐like Rad3 and non‐hyperphosphorylated Crb2 participate the repair processes in G0 cells that are more sensitive to UV and γ‐ray than in VE cells. The sensitivity like in VE cells is restored after replication in the nitrogen‐replenished medium, suggesting that the damage hyper‐sensitive nature of G0 cells is due to the error‐prone repair for single DNA duplex chromosome. The double‐strand break (DSB) repair in G0 cells required Pku80, one of non‐homologous end joining (NHEJ) proteins. S. pombe G0 cells upon DNA damages thus respond distinctively from VE cells in regard with regulation of checkpoint proteins and the mode of repair that is dependent upon the use of NHEJ.
Ex vivo study of the adhesion of an epoxy‐based sealer to human dentine submitted to irradiation with Er : YAG and Nd : YAG lasers
Authors: Sousa‐Neto M. D. , Silva Coelho F. I. , Marchesan M. A. , Alfredo E. , Silva‐Sousa Y. T. C.
Published: November 2005

Aim  To evaluate the adhesion of an epoxy‐based sealer to human dentine submitted to irradiation with Er : YAG or Nd : YAG laser at various parameters.

Methodology  Ninety maxillary canine teeth were sectioned transversely at the cemento‐enamel junction and at the root tip to leave an 8‐mm‐long cylinder. The tooth specimen was centred in a metallic ring (16 mm diameter and 8 mm height) and embedded in acrylic resin. The root canals were prepared using a low‐speed handpiece and a conical diamond bur, which was attached to a paralleling device. This bur was lowered to a depth previously determined by a silicone stop. Specimens were divided into nine groups: group I, dentine was treated with 2 mL of 17% EDTAC for 5 min. Groups II–V were irradiated with Er : YAG laser at the following parameters: group II – 8 Hz and 200 mJ input (120 mJ output); group III – 8 Hz and 400 mJ input (240 mJ output); group IV – 16 Hz and 200 mJ input (120 mJ output); group V – 16 Hz and 400 mJ input (240 mJ output). Groups VI–IX were irradiated with Nd : YAG laser at the following parameters: group VI – 10 Hz and 1 W input (0.4 W output); group VII – 10 Hz and 2 W input (0.8 W output); group VIII – 15 Hz and 1 W input (0.4 W output); group IX – 15 Hz and 2 W input (0.8 W output). The root canals were filled with an epoxy‐based root canal sealer and submitted to a push‐out test.

Results  Statistical analysis showed significant differences (P < 0.01) between Er : YAG and Nd : YAG laser treatments at the higher frequencies compared with 17% EDTAC. Greater adhesion values were obtained for groups IV and V (Er : YAG laser) and groups VIII and IX (Nd : YAG laser), which were statistically different from groups II and III (Er : YAG laser) and groups VI and VII (Nd : YAG laser). Treatment with only 17% EDTAC had the lowest adhesion values.

Conclusions  An increase in frequency, independent of power settings, of the lasers used in this study increased adhesion of an epoxy‐based root canal sealer.
Effect of mineral trioxide aggregate on cytokine production by peritoneal macrophages
Authors: Rezende T. M. B. , Vargas D. L. , Cardoso F. P. , Sobrinho A. P. R. , Vieira L. Q.
Published: November 2005

Aim  To test the effect of two commercial brands of grey mineral trioxide aggregate (ProRoot® and MTA‐Ângelus®) on cytokine production by M1 and M2 inflammatory macrophages.

Methodology  M1 (from C57BL/6 mice) and M2 peritoneal inflammatory macrophages (from C57BL/6 IL12p40−/− mice) were obtained and cultured in vitro in the presence of MTA. The cellular viability and the production of tumour necrosis factor‐α, interleukin (IL)‐12 and IL‐10 in response to stimulation with interferon‐γ and Fusobacterium nucleatum or Peptostreptococcus anaerobius were evaluated. Data were analysed by Mann–Whitney, Kruskal–Wallis and anova tests.

Results  The cements did not interfere with cellular viability or with cytokine production by either type of macrophage. However, M2 macrophages produced higher levels of IL‐10 when stimulated with F. nucleatum than M1 macrophages (P < 0.05).

Conclusions  The brands of MTA evaluated did not interfere in the cytokine response by M1 or M2 macrophages to the two bacteria tested. However, a difference in cytokine production between the two types of macrophages was found.
The effect of environmental factors on the distribution of Neoparamoeba pemaquidensis in Tasmania
Authors: Douglas‐Helders M , Nowak B , Butler R
Published: November 2005

Aquaculture in Tasmania is mostly carried out in estuaries. These estuarine habitats show a great variety and form unique environments in which Neoparamoeba pemaquidensis, the amoebic gill disease (AGD)‐causing protozoan, may or may not survive. Tasmania is divided into two zones, one where AGD is present and one where AGD is absent, but any ecological data to rationalize this distribution is lacking. In in vitro trials N. pemaquidensis strains were exposed to different concentrations of ammonium sulphate, copper sulphate, copper sulphate and tannin, and different Neoparamoeba densities, salinities and temperatures. A trial using field water samples investigated the survival of N. pemaquidensis in waters sourced from AGD‐free and AGD‐positive zones, and water analysis was performed to determine any differences. Significantly decreased protozoan survival was found with exposure to increasing copper sulphate concentrations from 10 to 100 000μm (P < 0.001), salinity of 15‰ (P < 0.001), low Neoparamoeba densities of 625 and 1250 cells mL−1 (P = 0.0005), and water sourced from Macquarie Harbour (P < 0.001). The water chemistry of this AGD‐free zone showed significantly lower dissolved calcium and magnesium concentrations which may contribute to this area being AGD‐free. Understanding of the ecology of N. pemaquidensis will enable better control and prevention strategies for Tasmanian salmon growers.
Modulation of β‐actin, insulin‐like growth factor 1 and glucose‐6‐phosphate dehydrogenase gene expression during vibriosis of sea bream, Sparus (=Rhabdosargus) sarba Forsskål
Authors: Deane E E , Woo N Y S
Published: November 2005

Gene expression changes, related to growth regulation, were investigated in silver sea bream, Sparus (=Rhabdosargus) sarba, subjected to vibriosis. Using reverse transcriptase polymerase chain reaction assays, transcript levels of β‐actin, insulin‐like growth factor 1 (IGF‐1) and glucose‐6‐phosphate dehydrogenase (G6PDH) were analysed. A tissue‐specific response for β‐actin was found, as transcript levels in the kidney increased during early‐ and mid‐stage vibriosis, but remained unchanged in liver tissue. The gene transcript levels of IGF‐1 were found to decrease in both liver and kidney from an early stage of vibriosis. The gene transcript levels of G6PDH increased at early and mid stages of vibriosis in liver and kidney before decreasing during the late stage of vibriosis. Further studies on G6PDH demonstrated that chemical inhibition of G6PDH in silver sea bream fibroblasts resulted in increased cell death upon exposure to hydrogen peroxide. The results from the present study demonstrate the complexity of growth‐related gene expression events that occur in fish tissues during disease.
Detection of Renibacterium salmoninarum in chinook salmon, Oncorhynchus tshawytscha (Walbaum), using quantitative PCR
Authors: Powell M , Overturf K , Hogge C , Johnson K
Published: November 2005

A quantitative polymerase chain reaction (QPCR) assay has been developed to detect varying levels of Renibacterium salmoninarum, the causative agent of bacterial kidney disease. This assay allows for the direct enumeration of bacterial DNA or RNA copy number within tissues and body fluids. The assay can be applied non‐lethally and can be used to determine whether R. salmoninarum is transcriptionally active. The presence of R. salmoninarum in kidney tissues from 430 chinook salmon collected from five Idaho Fish and Game operated hatcheries was initially evaluated using the widely employed enzyme‐linked immunosorbent assay (ELISA) with two sets of Kirkegaard and Perry Laboratories polyclonal antibodies, ‘mother batches’ 1 and 2. The same tissue samples were then analysed using the novel QPCR assay and the results compared. At moderate to high levels of infection [optical density (OD > 0.5)], ELISA values and estimated DNA copy number were highly correlated (r2 > 0.80), although correlation to specific antibody batches varied. However, lower ELISA values (OD < 0.5) observed with either antibody batch did not correlate well with the QPCR assay (R2 ≤ 0.43). Negative controls, run concurrently with the PCR assay, did not indicate extraneous DNA contamination in the PCR samples.
Selective acquisition of novel mating type and vegetative incompatibility genes via interspecies gene transfer in the globally invading eukaryote Ophiostoma novo‐ulmi
Authors: PAOLETTI MATHIEU , BUCK KENNETH W. , BRASIER CLIVE M.
Published: November 2005

The Dutch elm disease fungus Ophiostoma novo‐ulmi, which has destroyed billions of elm trees worldwide, originally invaded Europe as a series of clonal populations with a single mating type (MAT‐2) and a single vegetative incompatibility (vic) type. The populations then rapidly became diverse with the appearance of the MAT‐1 type and many vegetative incompatibility types. Here, we have investigated the mechanism using isolates from sites in Portugal at which the rapid evolution of O. novo‐ulmi populations from clonality to heterogeneity was well established. We show by genetic mapping of vic and MAT loci with AFLP markers and by sequence analysis of MAT loci that this diversification was due to selective acquisition by O. novo‐ulmi of the MAT‐1 and vic loci from another species, Ophiostoma ulmi. A global survey showed that interspecies transfer of the MAT‐1 locus occurred on many occasions as O. novo‐ulmi spread across the world. We discuss the possibility that fixation of the MAT‐1 and vic loci occurred in response to spread of deleterious viruses in the originally clonal populations. The process demonstrates the potential of interspecies gene transfer for facilitating rapid adaptation of invasive organisms to a new environment.
Carbon isotope evidence for CAM photosynthesis in the Mesozoic
Authors: Decker John E. , de Wit Maarten J.
Published: November 2005

Crassulacean acid metabolism (CAM) photosynthesis, a flexible adaptation to water‐stress, has not been shown to occur before the Pleistocene, despite suggestions that CAM first evolved in the Mesozoic or earlier. Here we report on multiple (≥7) positive organic carbon isotope excursions (δ13Corg > −20‰) in Late Triassic and Early Jurassic (c.200 Ma) sedimentary rocks from an arid terrestrial ecosystem in South Africa. These excursions are interpreted as evidence of the episodic dominance of CAM plants in response to heightened aridity in the heartland of Gondwana, and hint at climatic instability in a greenhouse world.
Sex ratio selection and multi-factorial sex determination in the housefly: a dynamic model
Authors: Kozielska M. , Pen I. , Beukeboom L. W. , Weissing F. J.
Published: November 2005

Sex determining (SD) mechanisms are highly variable between different taxonomic groups and appear to change relatively quickly during evolution. Sex ratio selection could be a dominant force causing such changes. We investigate theoretically the effect of sex ratio selection on the dynamics of a multi-factorial SD system. The system considered resembles the naturally occurring three-locus system of the housefly, which allows for male heterogamety, female heterogamety and a variety of other mechanisms. Sex ratio selection is modelled by assuming cost differences in the production of sons and daughters, a scenario leading to a strong sex ratio bias in the absence of constraints imposed by the mechanism of sex determination. We show that, despite of the presumed flexibility of the SD system considered, equilibrium sex ratios never deviate strongly from 1 : 1. Even if daughters are very costly, a male-biased sex ratio can never evolve. If sons are more costly, sex ratio can be slightly female biased but even in case of large cost differences the bias is very small (<10% from 1 : 1). Sex ratio selection can lead to a shift in the SD mechanism, but cannot be the sole cause of complete switches from one SD system to another. In fact, more than one locus remains polymorphic at equilibrium. We discuss our results in the context of evolution of the variable SD mechanism found in natural housefly populations.
Bone Marrow Mesenchymal Stem Cells: From Characterization of Neural Differentiation to a Possible Therapeutic Use in Neurodegeneration
Authors: Arnhold S. , Klein H. , BenMime L. , Addicks K.
Published: November 2005

Bone marrow derived stromal cells are of mesenchymal origin and precursor cells for skeletal tissue components such as chondroblasts and osteoblasts. Furthermore, under experimental conditions, a differentiation potency into myogenic and neuronal cells could be demonstrated. Because of their multipotency these cells represent a population of non‐haematogen stem cells that can be regarded as an alternative to human embryonic stem cells for future autologous cell replacement therapies. For a closer look at the differentiation capacity of these cells, rat and human bone marrow stromal cells were isolated from the femur bone and kept in the cell culture applying different cultivation protocols. In a cultivation medium with a serum content of 20%, the majority of these cells express a variety of neuronal markers such as ß‐III Tubulin and NeuN as well as the astrocyte marker GFAP, while a minority of about 20% express the marker for neural precursor cells nestin. Cultivation in a chemically defined serum free medium results in the differentiation of a markedly higher percentage of nestin positive neural precursor‐like cells. Using bFGF in combination with B27 these cells can be forced to form three dimensionally organized spheres. In order to elucidate a possible therapeutical potency of the bone marrow derived cells the synthesis of neurotrophic factors such as BDNF and NGF were analysed using the ELISA technique. Furthermore, they can be infected using a third generation adenoviral vector with high efficiency and show migratory activity in vitro. After injection of bone marrow derived mesenchymal stem cells into the lateral ventricle of adult rats they adhere to the ependymocytes and pass the ependymal barrier in order to settle in the subventricular space.
Preliminary Results Concerning the Expression of Carbohydrates in the Olfactory System of the Sturgeon
Authors: Quinteiro P. S. , Yáñez J. , Salazar I.
Published: November 2005

Lectins have been used in several areas of biomedicine and are particularly useful for histochemistry. Their ability to map the distributions of various kinds of cell‐ borne glycan, and thereby to identify particular cell populations, has allowed clarification of a number of issues in neuroscience. In the case of the olfactory system, for example, lectins may be involved in development, in the continual regeneration of olfactory neurons and even in the processing of olfactory information. Whereas in mammals and amphibians lectins have been widely employed for the study of the olfactory and accessory olfactory systems, little information regarding lectin labelling is available in fish. We have used a panel of 11 lectins (UEA, BSI‐B4, DBA, LEA, VVA, SBA, PSA, WGA, WGA‐s, ECA, LTA) in paraformaldehyde fixed and paraffin embedded specimens of a Chondrostei fish, the sturgeon (Acipenser baeri). Our preliminary findings show that the olfactory receptor cells, the olfactory nerve fibres and their terminals in the olfactory bulbs were labelled with BSI‐B4, DBA, VVA, SBA, PSA, WGA and WGA‐s. The presence of glycoproteins, whose terminal sugars are detected by lectin binding, might be related to the reception of an odour stimulus and its transduction into a nervous signal or to the histogenesis of the olfactory system.
The Expression of Connexins during Growth and Development of Skeletal Elements in the Dog
Authors: Schnapper A. , Brehm R. , Bergmann M. , Meyer W.
Published: November 2005

Efficient and well‐regulated intercellular communication is essential to organize the activities of the different cell populations involved during growth and concomitant structural maturation of the skeleton. For direct cell‐cell communication gap junctions composed of connexins (Cx) are present. So far 21 Cx types have been identified in mammals, which differ in molecular permeability as well as tissue expression pattern. To characterize the Cx distribution in the developing canine skeleton, samples from humerus, scapula and lumbar vertebrae of Beagle dogs aged 1.5, 3, 4.5 months and 1.5 years were fixed in Bouin's solution, decalcified with 25% EDTA and embedded in paraffin wax. Using immunohistochemistry and in situ hybridization, the expression of Cx 26, 30, 31, 32, 43, and 45 was studied in bone tissue plus associated epiphyseal and growth plate cartilage. Of all bone cells, osteoblasts showed the largest Cx expression repertoire: they were positive for Cx 26, 30, 32 and 43. After transformation into bone lining cells Cx43 expression was lost while in osteocytes only Cx 26 and 32 was detected. A wide variety of Cx reactions was also found in osteoclasts: Cx 26, 30, 32, and 43. In contrast, chondrocytes showed a much more restricted Cx expression pattern. Positive reactions were only seen in hypertrophic chondrocytes of epiphyseal cartilage (Cx 26, 31, 32) and in the proliferative (Cx32) and hypertrophic zone (Cx 26 and 32) of growth plate cartilage. Additionally, Cx31 was strongly expressed in the cellular layer of the periosteum covering areas of intensive bone modelling in the puppies. Age‐ and microlocation‐dependent variations in expression and/or staining intensity were also observed in the other Cx types. These results demonstrated that canine bone cells are capable of synthesizing a wide variety of Cx types during skeletal development, thus propagating different cellular signals via gap junctions in a differentially regulated and requirement orientated manner.
Investigation of Senescence in Canine Fibroblasts
Authors: Streit S. , Salomon F. ‐V. , Stahl T.
Published: November 2005

Introduction and Aims:  Primary and secondary cell cultures of canine dermal fibroblasts were used to test the suitability of several senescence‐linked cellular markers to characterize the chronological age of dogs. In further studies these markers will be used to test the hypothesis that senescence‐linked changes at the cellular level are representative for the observed inverse relationship of body mass and life expectancy in different dog breeds.

Methods:  In this study standardized skin samples of Beagles were used to analyse the cell morphology and several senescence‐linked parameters as Cumulative Population Doubling Level, Highest Population Doubling Level, Culture Doubling Time and Life Span Completed to determine the replicative potential of fibroblasts. The samples were divided by the age of the donors into three groups (‘young’, ‘middle’ and ‘old’).

Results:  There are notable differences between the replicative potential of young and old Beagles at the cellular level. Preliminary results show that in fibroblast cultures obtained from the young group the highest population doubling level is higher and the culture doubling time is shorter then in cultures of the old group. Cultures from the young and middle‐aged group cannot be distinguished by the measured parameters. For final results the complete statistical analysis of sampled data has to be carried out.

Conclusions:  Our results demonstrate a relationship between the chronological age of the skin donor and the replicative potential of the dermal fibroblasts. Therefore cellular senescence‐linked markers could serve as suitable tools to compare the age of individual dogs of different breeds.
Histamine and VIP‐positive Mast Cells in Renal Blood Vessels of Domestic Swine
Authors: Vodenicharov A. , Gulubova M. , Ilieski V.
Published: November 2005

Introduction:  Histamine and vasoactive intestinal polypeptide (VIP) are well known biogenic amines (substances), which rapidly elute from the mast cell granules under physiological conditions. They play an important role in the smooth muscle cells motility.

Aim:  The aim of the study was to investigate histamine‐ and VIP‐positive mast cells in the wall of pig kidney blood vessels in concern to their role of motility of smooth muscle cells and renal haemodynamics, respectively.

Material and Methods:  Extra‐ and intraorgan renal blood vessels taken immediately after slaughtering of eight 6–8 months pigs were studied. Tissue samples were immersion fixed in 0.05 Mcacodylate buffer, pH 7.2, containing 2% paraformaldehyde and 0.2% glutaraldehyde for 5 days at 4°C. After fixation, tissue blocks were rinsed in 30% sucrose in distilled water at 4°C for 1–3 days. Cryostat sections (20–30 μm thick) were prepared and thawed in 0.05 M cacodylate buffer, pH 2, overnight. Immunocytochemical procedure started for free‐floating sections which were incubated in 1.2% hydrogen peroxide in methanol for 30 min, and rinsed in 0.1 M phosphate buffered saline (PBS), pH 7.4, for 15 min. Afterwards, sections were incubated with the primary antibodies in a dilution of 1:100 (histamine) and in vasoactive intestinal polypeptide (VIP) 1:1000 for 24 h at room temperature (RT), then rinsed in 0.1 M PBS, pH 7.4. Incubation succeeded: first, with biotinylated anti‐rabbit (histamine) and anti‐mouse VIP antibodies in a dilution of 1:20 for 4 h, and rinses in 0.1 M PBS, pH 7.4, and in 0.05 M tris‐HCl buffer, pH 7.5, for 10 min. Peroxidase activity was localized by using a mixture of 3 mg 3,3’ diaminobenzidine (DAB), in 15 ml 0.005 M Tris‐HCl buffer, pH 7.5, and 36 ml 1 % hydrogen peroxide for 10–20 min. Finally, a rinse in 0.01 M PBS, pH 7.4, was undertaken.

Immunochemicals:  The antibodies used were rabbit anti‐histamine (H7403) and rabbit anti‐vasoactive intestinal peptide (VIP), or (PEPA41) (Sigma, Saint Louis, Missouri, USA).

Results and Discussion:  It was established that VIP‐positive mast cells were located in the media and in the adventitia of large arterial vessels and renal vein, as well. Histamine positive mast cells were found mostly in the intima of renal artery and in the other layers of the wall. Both VIP and histamine mast cells were observed next to small arteries and veins in the fat tissue, around extraorgan blood vessels. Our finding suggest to mast cells participation in the smooth muscle cells motility and the maintenance of local homeostasis, as well.
The Development of the Metanephric Kidney in the Pig
Authors: Bragulla H.
Published: November 2005

Aims:  The metanephric kidneys of the pig are used as xenotransplants in human medicine. In order for transplants to fit within the host organisms, the subcapsular blastema and blood vessels are crucial for the development of new nephrons to sustain the organ functions. The aim of this study is to obtain data concerning the post‐natal development of metanephric nephrons in the porcine kidney.

Materials and Methods:  The metanephric kidneys of six porcine fetuses with a crown‐rump length ranging from 40 mm to 220 mm of eight piglets aged between 6 to 10 weeks and of three adult pigs were studied. Eight lectins as well as anti‐actin and anti‐myosin antibodies were used for lectin‐ and immunohistochemistry to study the subcapsular metanephric blastema, to visualize the blood‐urine barrier in the nephrons and collecting tubules, and to study the blood vessels in both the renal cortex and marrow.

Results and Conclusions:  A subcapsular metanephric blastema was still present in the kidney of 10‐week‐old piglets. Dense condensation of mesenchymal cells surrounded the terminal branches of the collecting ducts and showed first signs of mesenchymal‐epithelial transformation. Characteristic comma‐shaped and s‐shaped bodies were found in and underneath the subcapsular blastema. In the fibrous renal capsule of six‐week‐old piglets, a first faint binding reaction of anti‐actin was visible and intensified in the fibrous renal capsule in ten‐week‐old piglets and in adult pigs. In addition, the smooth‐muscle layers of the blood vessels were stained by the anti‐actin and anti‐myosin antibodies. The lectins showed various affinities to the endothelium of blood vessels and to the epithelial cells lining of the capsules of the metanephric renal corpuscles, the various parts of the renal tubules, as well as the collecting tubules and the renal pelvis. The affinity of the epithelial cells to a specific lectin varies in neighbouring cells, indicating different cell activities or cell cycles.
Computed Tomography as a New Tool for Investigating Leg Weakness in Turkeys
Authors: Buda S. , Grönemeyer D. , Hafez H. M. , Matzke M. , Walter A. , Budras K. D.
Published: November 2005

Introduction:  Leg weakness causes high economic losses in commercial poultry. The unspecific term includes, amongst others, diseases of the skeletal system caused by genetic, nutritional or microbiological factors. The analysis of computed tomography images may be a new tool to discriminate and evaluate skeletal abnormalities in turkeys and can therefore provide valuable hints in the treatment of turkey stocks suffering from leg weakness.

Material and Methods:  Legs of healthy turkeys and those showing clinical signs of leg weakness were examined using a Tomoscan M/EG/Compact (Philips Medical Systems) CT system. The slice thickness and distance were set to 2 mm. In addition to the evaluation of the obtained images, the software system 3D‐Doctor was used to create three‐dimensional objects of the investigated samples.

Results:  Differences between physiological and abnormal parts of the leg bones are distinguishable in the CT‐images and three‐dimensional reconstructed objects. Bone density can be calculated and thus provides a base to estimate possible nutritional demands or deficiencies. The three‐dimensional reconstruction compensates for the difficulties in understanding the detailed anatomical CT cross‐sections by which the examiner is confronted while studying the CT images.

Conclusion:  Although computed tomography of poultry is too expensive for routine herd diagnosis, and far less for single‐animal diagnosis, it is a valuable tool for investigating skeletal abnormalities and is very useful in examining samples related to leg weakness.