bluuebunny/crossref_metadata_2025 huggingface dataset:
Acute radiation toxicity assessment of a 3‐D conformal head and neck radiation treatment technique
Authors: Kearvell R , Kuan R , Preston R , Tanner P , Buck H , Hedges R
Published: September 2004

Patients undergoing radiation treatment for head and neck cancer have many critical structures within or adjacent to the treatment area. Avoiding these critical structures is more efficient and easier using 3‐D conformal planning. At Sir Charles Gairdner Hospital an investigation into the acute radiation toxicity was undertaken when the head and neck treatment technique was changed from the conventional 2‐D plan to the conformal 3‐D plan. Although the primary target volume (PTV), fraction size and overall dose remain the same for the two techniques, differing beam configurations mean that treatment fields enter and exit through different parts of the head and neck. Ten patients were initially assessed to gain a baseline appearance of their head and neck region, and then graded weekly according to the toxicity criteria during treatment and at 4, 8 and 12 weeks after treatment. The results of the assessment indicate that there is no increase in toxicity as a result of treating using the conformal head and neck technique.
Assessing outcomes of cancer care: Lessons to be learned from a retrospective review of the management of small cell lung cancer at the Cancer Therapy Centre, Liverpool Hospital, January 1996−July 2000
Authors: Ellis PM , Delaney G , Della‐Fiorentina S , Moylan E
Published: September 2004

The purpose of the present paper was to review the outcomes of care of small cell lung cancer (SCLC) at one Sydney teaching hospital. A retrospective cohort study was carried out of patients with SCLC seen between January 1996 and July 2000. The main outcomes were relapse‐free and overall survival. Secondary outcomes of interest were the uniformity of staging investigations, initial treatment, use of prophylactic cranial irradiation (PCI), patterns of relapse and treatment received following relapse. One hundred and three patients with SCLC were treated at the Liverpool Hospital Cancer Therapy Centre during this period. There were 58 men (56%) and 45 women (44%). Forty‐two patients (41%) had limited stage disease (LD) and 61 (59%) had extensive stage disease (ED). There was considerable variation in staging investigations. There was little variation in systemic treatment of SCLC. Only 32 of 42 patients with limited stage SCLC were candidates for thoracic radiotherapy and only seven patients received PCI. Median relapse‐free survival was 11.2 months (95% confidence interval (CI): 7.7−14.8) for patients with LD and 6 months (95%CI: 4.4−7.5) for ED. Median overall survival was 15.1 months (95%CI: 11−19.1) for patients with LD and 8.9 months (95%CI: 7.5−10.2) for ED. Some health outcomes similar to that reported in clinical trials can be achieved in clinical practice. Measuring health outcomes is an important process of maintaining quality of care.
Atypical diffusion‐weighted magnetic resonance findings in glioblastoma multiforme
Authors: Batra A , Tripathi RP
Published: September 2004

The present report describes two patients with glioblastoma multiforme with the tumour demonstrating low central apparent diffusion coefficient values similar to those found in cerebral abscesses. Although conventional MR images were fairly specific for tumour, the cases illustrate the need for exercising caution when using diffusion‐weighted (DW) MR images for the differentiation of necrotic brain tumours from abscesses. The DW MR imaging information should always be integrated with the conventional spin‐echo MR images.
Role of embolization in the management of splenic trauma
Authors: Lui B , Schlicht S , Vrazas J
Published: September 2004

The management of splenic trauma is determined by factors related to a patient's clinical state. Currently, non‐surgical management is favoured because of morbidity related to laparotomy and splenectomy. However, the presence of a pseudoaneurysm might necessitate selective splenic arteriography and embolization. Even though this also has associated morbidity, conservation of viable splenic tissue is a desired result.
Sarcoidosis of the thoracic and lumbar spine: Imaging findings with an emphasis on magnetic resonance imaging
Authors: Lisle D , Mitchell K , Crouch M , Windsor M
Published: September 2004

Vertebral sarcoidosis is exceedingly rare with only a few cases reported in the published literature. The case of a 40‐year‐old man with vertebral sarcoidosis is presented, emphasizing the role of MRI and biopsy in confirming the diagnosis. A brief review of the published literature is also presented.
Concurrent emphysematous pyelonephritis and emphysematous cholecystitis in type 2 diabetes
Authors: Bhansali A , Bhadada S , Shridhar C , Choudhary S , Khandelwal N
Published: September 2004

Concurrence of emphysematous pyelonephritis and emphysematous cholecystitis in a patient with type 2 diabetes has not been reported in the published literature, to the authors’ knowledge. A patient who had both at the time of presentation is reported. The mechanism of gas formation and the management strategies are discussed.
Direct thrombin injection into aneurysmal sac in a patient with a type II endoleak
Authors: El‐Saeity N , Benfayed W , Ramesh N , Feeley M , Torreggiani WC
Published: September 2004

Endovascular grafting of abdominal aortic aneurysms provides a good alternative to open surgery, especially in high‐risk patients. Endoleaks are a well‐recognized complication and are typically diagnosed on CT. We descibe a case in which a patient's endoleak was evaluated by MRI and successfully treated by direct thrombin injection into the site of the leak.
Cerebellar metastases from prostatic carcinoma
Authors: Schoenwaelder M , Waugh J , Russell P
Published: September 2004

A 67‐year‐old man presented with worsening headaches and gait disturbance, and with minimal clinical signs. A brain MRI showed multiple solid cerebellar metastases. The pathological diagnosis of metastatic carcinoma of the prostate was further suggested by an elevation in prostate specific antigen, and was pathologically confirmed following the neurosurgical removal of the tumours. The rarity of this presentation, as documented in the clinical literature, is reviewed.
Deciding to provide a ‘reader’ in examinations for the General Certificate of Secondary Education (GCSE): questions about validity and ‘inclusion’
Authors: Woods Kevin
Published: September 2004

In England, Wales and Northern Ireland, the General Certificate of Secondary Education (GCSE) was designed as an ‘inclusive’ qualification, access to which has been supported by providing a ‘reader’ to some candidates during examinations. A candidate reading age criterion of ten years has been employed by the GCSE awarding bodies to determine eligibility for this provision. In this paper, Kevin Woods, a member of the Educational Support and Inclusion Research and Teaching Group in the Faculty of Education at the University of Manchester, examines the rationale and evidence for adopting this criterion and describes an investigation of the reading needs of a cross‐sectional sample of 38 GCSE examination candidates in trial examinations. The investigation found a low level of candidate need for a reader, with candidate reading age and self‐prediction being unreliable indicators of this need. Kevin Woods highlights the implications for the assessment process used to determine eligibility for a reader in GCSE examinations and pays particular attention to the feasibility and validity of including all students as eligible.
Stepping out of the classroom: involving teachers in the evaluation of national special education policy
Authors: Bourke Roseanna , Kearney Alison , Bevan‐Brown Jill
Published: September 2004

Since Richard Rose relaunched BJSE's Research Section as a forum for the discussion of practitioner research, BJSE has published a series of very successful accounts of individual teacher enquiry and action research. In this article from New Zealand, Roseanna Bourke, now working at the Ministry of Education in Wellington, and her two colleagues, Alison Kearney and Jill Bevan‐Brown, both from the Department of Learning and Teaching at Massey University, present an alternative view of the involvement of teachers in research.

In 1999, the Ministry of Education asked a team of researchers from Massey University to evaluate the introduction of New Zealand's first national policy for special education, known as Special Education 2000. The 16‐member research team was contracted to provide a three‐year, multi‐method, longitudinal evaluation incorporating both quantitative and qualitative approaches. This article looks specifically at one aspect of the approach used in the evaluation – the use of teachers as fieldworkers. Teachers across the country were trained as fieldworkers to assist in the collection of data in schools and early childhood centres. Training was given to all teachers on the research aims, methods, ethics, including cultural sensitivity, and rationale for the evaluation. By participating in large‐scale research and acting as ‘fieldworkers’, 68 teachers had the opportunity to develop their skills as researchers, while learning about other schools' perceptions and experiences of the special education policy. The experience provided a means for teachers to become apprenticed into a research culture, and to talk with principals and other teachers about common areas of interest. It also ensured that the research had knowledgeable and credible fieldworkers to collect data in schools.
Feature: UNESCO World Heritage and the Joggins cliffs of Nova Scotia
Authors: Falcon‐Lang Howard J. , Calder John H.
Published: September 2004

UNESCO World Heritage status is the highest honour that may be bestowed on a palaeontological site. In addition to heightening conservation status, it confers international recognition of a locality's ‘outstanding universal value’ and often triggers the release of substantial regional development funds. Despite these incentives it is, perhaps, not surprising that only a handful of fossil sites have successfully navigated the World Heritage selection process. In this feature, we draw on our recent experience of developing a World Heritage bid for the Pennsylvanian ‘Coal Age’ locality of Joggins, Nova Scotia. As well as demonstrating the huge significance of Joggins, we hope that our findings will provide valuable guidelines for the assessment of World Heritage fossil sites in general.
In vitro total and methane gas production as influenced by rate of nitrogen application, season of harvest and perennial ryegrass cultivar
Authors: Lovett D. K. , Bortolozzo A. , Conaghan P. , O'Kiely P. , O'Mara F. P.
Published: September 2004

The effects of rate of inorganic nitrogen (N) fertilization (0, 80 or 160 kg N ha−1 per regrowth), season of harvest (regrowths 1, 2 and 3) and perennial ryegrass (Lolium perenne L.) cultivar [classified as having either a normal or elevated water soluble carbohydrate (WSC) concentration genotype] on in vitro gas production and digestibility were assessed. Increased N fertilizer application significantly decreased total gas production (TGP), methane (CH4) production and organic matter digestibility (OMD). The results suggest that the decreases in TGP and CH4 production were associated with a restriction in organic matter (OM) fermentation and an altered crude protein (CP) to structural carbohydrate ratio rather than a modification in the stoichiometry of fermentation. Season of harvest only significantly (P < 0·05) altered in vitro OMD and CH4 production at 8 h, despite altering the chemical composition of the herbage. Cultivar effects on all measured in vitro parameters were not significant presumably because the elevated WSC concentration trait was not expressed strongly in the study.
Turf quality and reliability in varieties of four turfgrass species in contrasting Italian environments
Authors: Russi L. , Annicchiarico P. , Martiniello P. , Tomasoni C. , Piano E. , Veronesi F.
Published: September 2004

Turf quality is a complex character of fundamental importance in turfgrass evaluation. It takes into account aesthetic and functional aspects, and depends on several individual components which may vary with time. The use of a synthetic and simple quality score is necessary when evaluating large numbers of varieties. The quality of 110 turfgrass varieties belonging to four species (Lolium perenne, Poa pratensis, Festuca arundinacea and F. rubra) was assessed in the second and third years of growth at three Italian locations, i.e. Lodi (Po Valley, continental climate), Perugia (central Italy, sub‐Mediterranean climate) and Foggia (southern Italy, Mediterranean climate), using a visual score ranging from 9 (outstanding turf) to 1 (very poor turf). A randomized block design with three replicates was used, with varieties of the same species being blocked to facilitate the comparison within species. On average, F. rubra had the lowest turf quality score in summer, whereas the other species had the lowest quality score in winter and the highest quality score in summer and autumn. Components of variance were large for genotype (i.e. variety) main effects in all species. A variety × location interaction was found in F. arundinacea and higher‐order interactions were also found for P. pratensis and F. rubra. Genotype × environment effects were small relative to genotypic effects in L. perenne. Variety × year interactions were small in all cases, whereas variety × location and variety × season interactions were greater. Stability of turf quality across sites, seasons and years was measured for each variety in terms of environmental variance, i.e. the variance of score values across sites, years and seasons. Mean scores and stability values of varieties were integrated into an index of reliability that estimated the lowest score value expected in 0·80 of cases. For each species, a small subset of highly reliable varieties could be identified. Implications of the results for the testing and breeding of varieties in Italy are discussed.
Effect of heading date of perennial ryegrass cultivars on tillering and tiller development in spring and summer
Authors: Laidlaw A. S.
Published: September 2004

Three cultivars (two diploid and one tetraploid) in each of three maturity groups (early, intermediate and late) of perennial ryegrass were sown in 10 m2 plots, replicated four times, in Northern Ireland in June 1997 in a study of the effect of heading date on tiller development (including initiation to flower) and turnover of tillers produced at specific times in spring in 1998 and 1999. The plots were harvested seven times in each year. Annual dry‐matter production was similar for all groups in each year. In spring and early summer of both years, tiller density of the diploid cultivars was 1·5 times greater than that of the tetraploid cultivars and the mean tiller density over all swards in June was about 0·40 times greater than that in April. Maximum proportions of reproductive tillers in the early, intermediate and late maturity groups, determined from apical dissections, were found in early April, mid‐May and early June, respectively. Although a high proportion of tillers, which were present when annual observations commenced in spring, was decapitated at the first harvest in the early group, the previous population density was maintained by rapid production of new tillers during May, including those from suppressed tiller buds during reproduction.

It is concluded that the relationship between heading date and rate of tiller turnover (including flowering) at specified times in spring is important in sward management throughout the early part of the growing season and should be taken into account in tiller‐based grass growth models.
Stress and recovery of hill pastures in the North Island of New Zealand
Authors: Nicholas P. K. , Kemp P. D. , Barker D. J.
Published: September 2004

Moisture and treading treatments were imposed on intact turves that were relocated to a glasshouse after being removed from three hill pastures of different soil fertility in the North Island of New Zealand. The experiment consisted of a 2‐month stress phase, where the treatments were wetting (W), wetting and treading (WT), drying (D) and control (C). In this phase, herbage accumulation rate, tiller density and leaf extension rate were lower on the D turves, and herbage accumulation rate and tiller density were lower on the WT turves than for the C turves. Herbage accumulation rate was higher on the W treatment than on the C treatment.

In the 2‐month recovery phase, herbage accumulation rate and leaf extension rate on the D turves were higher than those of the C treatment. Herbage accumulation rate and tiller density took longer to recover on the WT turves but by the end of the recovery period tiller density on these turves exceeded that of the C turves and the original tiller densities on the WT turves. Changes (increase or decrease) in leaf extension rate were associated with the W treatment and tiller density with the WT treatment. Moisture was limiting on the D and C turves, but on the W and WT turves, where moisture was adequate for plant growth, nutrients were limiting, notably phosphorus on the W and WT turves and sulphur on the W turves.

The D treatment turves recovered very quickly once the stress was removed but the WT turves were slower to recover. Under the experimental conditions applied, the hill pasture turves were more resilient to the drying treatment than the wetting and treading treatment.
Modelling the comparative productivity and profitability of grass and legume systems of silage production in northern Europe
Authors: Topp C. F. E. , Doyle C. J.
Published: September 2004

A pre‐existing mechanistic model of grass growth, developed to simulate grass production in the UK, was extended to include a variety of grass–legume mixtures and used to predict comparative yields across a range of sites in northern Europe, using experimental data from a series of experiments at fifteen sites. Specifically, predictions were made of the comparative yields of grass (Lolium perenne L.), white clover (Trifolium repens L.), red clover (T. pratense L.), lucerne (Medicago sativa L.), lotus (Lotus corniculatus L.) and galega (Galega orientalis Lam.). The results indicated that the model could generate simulated differences in yields between forage crops, which accorded with observed rankings. However, the predictions were more accurate for total yields than individual cuts, for a country than for an individual site, and for monocultures than for grass–legume mixtures. Nevertheless, the results appeared sufficiently robust to be used to prepare forecasts of yield productivity in different agro‐climatic zones in northern Europe. Using estimates of the production costs and economic values for the forages, it was also possible to assess their comparative profitability.

The study indicated that red clover, whether grown singly or in a mixture with grass economically outperformed all silage systems based on pure grass swards, even those receiving 400 kg N ha−1. White clover and lucerne were also consistently more profitable than grass‐based systems, receiving 200 kg N ha−1. In relative terms, lucerne appeared to be more profitable in the south and east of the region, while white clover outperformed lucerne in the north and west. In the north and east of the region, there was some evidence that galega might also hold promise as a silage crop, but lotus was not found to be viable under north European environmental conditions.
Performance of cultivated perennial grass mixtures under different grazing intensities in the alpine region of the Qinghai‐Tibetan Plateau
Authors: Dong S. K. , Kang M. Y. , Hu Z. Z. , Long R. , Pu X. P.
Published: September 2004

In 1998, seeds of four cultivated native perennial grasses, Bromus inermis (B), Clinelymus nutans (C), Elymus nutans (E) and Agropyron cristatum (A), were sown as mixtures with different seed rates in three combinations (B + C, B + E + A and B + E + C + A) in a field study in the north‐eastern part of the Qinghai‐Tibetan Plateau, China. A grazing experiment was conducted in 2000 to investigate the performance of these grass mixtures at leaf, plant and sward scales under different grazing intensities (GI). Four GIs, expressed as the proportion of herbage consumed by Tibetan lambs in relation to the available biomass (IP), were applied in the summer: no grazing, and 0·30, 0·50 and 0·70 of IP. Tiller numbers of the grasses increased and leaf photosynthetic rate, sward leaf area index (LAI) and herbage mass declined with increase in GI. No effect of GI on specific leaf area was observed. Interactions between GI and grass mixtures on the dynamics of species composition, swards’ LAI and herbage mass were found. Large fluctuations in species composition with different GIs showed the poor species compatibility and sward persistence of these grass mixtures under high GI by sheep.
Analysis of postarousal EEG activity during somnambulistic episodes
Authors: Zadra Antonio , Pilon Mathieu , Joncas Steve , Rompré Sylvie , Montplaisir Jacques
Published: September 2004

Early studies found that electroencephalographic (EEG) recordings during somnambulistic episodes were characterized by a combination of alpha, theta, and delta frequencies, without evidence of clear wakefulness. Three postarousal EEG patterns associated with slow‐wave sleep (SWS) arousals were recently identified in adults with sleepwalking and sleep terrors. The goal of the present study was to evaluate the distribution of these postarousal EEG patterns in 10 somnambulistic patients (three males, seven females, mean age: 25.1, SD: 4.1) evaluated at baseline and following 38 h of sleep deprivation. A total of 44 behavioral arousals were recorded in the laboratory; seven episodes at baseline (five from SWS, two from stage 2 sleep) and 37 episodes during recovery sleep (30 from SWS, seven from stage 2 sleep). There was no significant difference in the distribution of postarousal EEG patterns identified during baseline and recovery sleep. One pattern, comprised of diffuse rhythmic and synchronous delta activity, was preferentially associated with relatively simple behavioral episodes but did not occur during episodes from stage 2 sleep. Overall, delta activity was detected in 48% of the behavioral episodes from SWS and in 22% of those from stage 2. There was no evidence of complete awakening during any of the episodes. The results support the view of somnambulism as a disorder of arousal and suggest that sleepwalkers’ atypical arousal reactions can manifest themselves in stage 2 sleep in addition to SWS.
Comparative utility of instruments for monitoring sleepiness‐related performance decrements in the operational environment
Authors: Balkin Thomas J. , Bliese Paul D. , Belenky Gregory , Sing Helen , Thorne David R. , Thomas Maria , Redmond Daniel P. , Russo Michael , Wesensten Nancy J.
Published: September 2004

As both military and commercial operations increasingly become continuous, 24‐h‐per‐day enterprises, the likelihood of operator errors or inefficiencies caused by sleep loss and/or circadian desynchrony also increases. Avoidance of such incidents requires the timely application of appropriate interventions – which, in turn, depend on the ability to measure and monitor the performance capacity of individuals in the operational environment. Several factors determine the potential suitability of candidate measures, including their relative sensitivity, reliability, content validity, intrusiveness and cumbersomeness/fieldability. In the present study, the relative sensitivity (defined as the ratio of effect size to 95% confidence interval) of several measures to the effects of sleep loss was compared in a sleep restriction experiment, in which groups were allowed 3, 5, 7, or 9 h time in bed (TIB) across seven consecutive nights. Of the measures compared, the Psychomotor Vigilance Test was among the most sensitive to sleep restriction, was among the most reliable with no evidence of learning over repeated administrations, and possesses characteristics that make it among the most practical for use in the operational environment.
Slow horizontal eye movement at human sleep onset
Authors: Porte Helene Sophrin
Published: September 2004

Slow eye movement (SEM) at human sleep onset has been studied as a function of EEG and behavioral state but has not been subjected to systematic physical measurement. This study counted the frequency of horizontal SEM and quantified its physical properties during hypnagogic EEG stages 3–8 (stage 1 sleep). SEM amplitudes, peak velocities, and durations were digitally measured. As in previous studies SEM occurred throughout stage 1 sleep, coinciding with all tonic states and with all phasic events that classically define stage 1. Within stage 1 the physical properties of SEM varied significantly. In hypnagogic stage 3, where EEG alpha dominates up to half of any 30‐s epoch, the typical SEM was moderate in amplitude, slow, and short‐lasting, and SEM peak velocity described a weak function of SEM amplitude. In stages 5 and 6 combined – characterized by a low‐voltage delta–theta EEG, sustained alpha suppression, and no more than one vertex sharp wave (VSW) per 30‐s epoch – SEM velocity was maximal, and described a strong function of amplitude. Despite low absolute velocities, stage 5/6 SEM could approximate saccade‐like morphologies. In stages 7 and 8 combined, characterized by VSW bursts and incipient spindles or K complexes, SEM was maximal in amplitude and duration but not faster than SEM in stage 5/6. These properties of human SEM are discussed in relation to the anatomy and physiology of SEM in macaque.
The 24‐h growth hormone rhythm in men: sleep and circadian influences questioned
Authors: Brandenberger Gabrielle , Weibel Laurence
Published: September 2004

The 24‐h rhythm of growth hormone (GH) is thought to be controlled primarily by sleep processes with a weak circadian component. This concept has been recently questioned in sleep‐deprived persons. To test the notion of a high sleep‐dependency of GH release, we established simultaneous 24‐h rhythms of GH and melatonin, a circadian marker, in night workers who form a model for challenging sleep and circadian processes. Ten day‐active subjects and 11 night workers were studied during their usual sleep–wake schedule, with sleep from 23:00 to 07:00 hours and 07:00 to 15:00 hours, respectively. Experiments were conducted in sleep rooms under continuous nutrition, bed rest, and dim light. Melatonin and GH were measured every 10 min over 24 h. In day‐active subjects, melatonin and GH showed the well‐known 24‐h profiles, with a major sleep‐related GH pulse accounting for 52.8 ± 3.5% of the 24‐h GH production and the onset of the melatonin surge occurring at 21:53 hours ± 18 min. In night workers, melatonin showed variable circadian adaptation, with the onset of secretion varying between 21:45 and 05:05 hours. The sleep‐related GH pulse was lowered, but the reduction was compensated for by the emergence of large individual pulses occurring unpredictably during waking periods, so that the total amount of GH secreted during the 24 h was constant. One cannot predict the degree of GH adaptation from the highly variable melatonin shift. These results argue against the concept that sleep processes exert a predominant influence on GH release whatever the conditions. When sleep and circadian processes are misaligned, the blunting of the sleep‐related GH pulse is counteracted, as in sleep‐deprived persons, by a compensatory mechanism promoting GH pulses during wakefulness.
DOMINO1, a member of a small plant‐specific gene family, encodes a protein essential for nuclear and nucleolar functions
Authors: Lahmy Sylvie , Guilleminot Jocelyne , Cheng Ching‐Ming , Bechtold Nicole , Albert Sylvie , Pelletier Georges , Delseny Michel , Devic Martine
Published: September 2004

Arabidopsis embryos carrying the domino1 mutation grow slowly in comparison with wild type embryos and as a consequence reach only the globular stage at desiccation. The primary defect of the mutation at the cellular level is the large size of the nucleolus that can be observed soon after fertilization in the nuclei of both the embryo and the endosperm. The ultrastructure of mutant nucleoli is drastically different from wild type and points to a fault in ribosome biogenesis. DOMINO1 encodes a protein, which belongs to a plant‐specific gene family sharing a common motif of unknown function, present in the tomato DEFECTIVE CHLOROPLASTS AND LEAVES (LeDCL) protein. Using a GFP protein fusion, we show that DOMINO1 is targeted to the nucleus. We propose that inactivation of DOMINO1 has a negative effect on ribosome biogenesis and on the rate of cell division.
Multiple gene detection by in situ RT‐PCR in isolated plant cells and tissues
Authors: Pesquet Edouard , Barbier Odile , Ranocha Philippe , Jauneau Alain , Goffner Deborah
Published: September 2004

With the number of functional genomic approaches in plant biology increasing daily, the demand for rapid and reliable RNA localization techniques for gene characterization is being felt. We present herein a novel, liquid phase in situ RT‐PCR (IS‐RT‐PCR) protocol using a combination of gene‐specific fluorescent primers and spectral confocal microscopy to localize target RNA in epicotyl sections and xylogenic suspension cultures of Zinnia elegans. Potential sources of artefacts from fixation to gene detection were systematically eliminated using both fluorescent primers and nucleotides for 18S rRNA gene detection, resulting in a set of optimal parameters for IS‐RT‐PCR that may be readily adapted to any target gene. By judiciously choosing fluorescent primers with non‐overlapping fluorochromes, we have shown that our technique is readily adapted to multiplex IS‐RT‐PCR, enabling the simultaneous localization of more than one gene within a complex tissue or heterogeneous cell population. A 6‐carboxy‐2′,4,4′,5′,7,7′‐hexachlorofluorescein (6‐HEX)‐labelled primer and a tetrachloro‐6‐carboxy‐fluorescein (TET)‐labelled primer were designed for two marker genes associated with programmed cell death in tracheary elements (TEs): an endonuclease (Zen1) and a cysteine protease (ZcP4), respectively. An additional Cyan5 (Cy5)‐labelled primer was used to monitor 18SrRNA expression. As expected, the 18S signal was constitutively expressed throughout epicotyls sections and living cells in xylogenic in vitro cultures, whereas Zen1 and ZcP4 were co‐localized in forming TEs both in planta and in vitro. Analogous to clustering analysis of gene expression using microarrays to elucidate common metabolic pathways and developmental processes, this novel technique is perfectly adapted to gaining a better understanding of gene function via the coordinated expression of genes in specific cell types of complex tissues and cell populations.
A dehydration‐induced NAC protein, RD26, is involved in a novel ABA‐dependent stress‐signaling pathway
Authors: Fujita Miki , Fujita Yasunari , Maruyama Kyonoshin , Seki Motoaki , Hiratsu Keiichiro , Ohme‐Takagi Masaru , Tran Lam‐Son Phan , Yamaguchi‐Shinozaki Kazuko , Shinozaki Kazuo
Published: September 2004

Arabidopsis thaliana RD26 cDNA, isolated from dehydrated plants, encodes a NAC protein. Expression of the RD26 gene was induced not only by drought but also by abscisic acid (ABA) and high salinity. The RD26 protein is localized in the nucleus and its C terminal has transcriptional activity. Transgenic plants overexpressing RD26 were highly sensitive to ABA, while RD26‐repressed plants were insensitive. The results of microarray analysis showed that ABA‐ and stress‐inducible genes are upregulated in the RD26‐overexpressed plants and repressed in the RD26‐repressed plants. Furthermore, RD26 activated a promoter of its target gene in Arabidopsis protoplasts. These results indicate that RD26 functions as a transcriptional activator in ABA‐inducible gene expression under abiotic stress in plants.
Adjustment of diurnal starch turnover to short days: depletion of sugar during the night leads to a temporary inhibition of carbohydrate utilization, accumulation of sugars and post‐translational activation of ADP‐glucose pyrophosphorylase in the following light period
Authors: Gibon Yves , Bläsing Oliver E. , Palacios‐Rojas Natalia , Pankovic Dejana , Hendriks Janneke H.M. , Fisahn Joachim , Höhne Melanie , Günther Manuela , Stitt Mark
Published: September 2004

A larger proportion of the fixed carbon is retained as starch in the leaf in short days, providing a larger store to support metabolism and carbon export during the long night. The mechanisms that facilitate this adjustment of the sink–source balance are unknown. Starchless pgm mutants were analysed to discover responses that are triggered when diurnal starch turnover is disturbed. Sugars accumulated to high levels during the day, and fell to very low levels by the middle of the night. Sugars rose rapidly in the roots and rosette after illumination, and decreased later in the light period. Global transcript profiling revealed only small differences between pgm and Col0 at the end of the day but large differences at the end of the night, when pgm resembled Col0 after a 4–6 h prolongation of the night and many genes required for biosynthesis and growth were repressed [Plant J. 37 (2004) 914]. It is concluded that transient sugar depletion at the end of the night inhibits carbon utilization at the start of the ensuing light period. A second set of experiments investigated the stimulation of starch synthesis in response to short days in wild‐type Col0. In short days, sugars were very low in the roots and rosette at the end of the dark period, and after illumination accumulated rapidly in both organs to levels that were higher than in long days. The response resembles pgm, except that carbohydrate accumulated in the leaf as starch instead of sugars. A similar response was found after transfer from long to short days. Inclusion of sugar in the rooting medium attenuated the stimulation of starch synthesis. Post‐translational activation of ADP‐glucose pyrophosphorylase (AGPase) was increased in pgm, and in Col0 in short days. It is concluded that starch synthesis is stimulated in short day conditions because sugar depletion at the end of the night triggers a temporary inhibition of growth and carbohydrate utilization in the first part of the light period, leading to transient accumulation of sugar and activation of AGPase.
Purification and cloning of an esterase from the weed black‐grass (Alopecurus myosuroides), which bioactivates aryloxyphenoxypropionate herbicides
Authors: Cummins Ian , Edwards Robert
Published: September 2004

Carboxyesterases which activate aryloxyphenoxypropionate (AOPP) graminicides to their bioactive herbicidal acids by hydrolysing the respective ester precursors have been identified in black‐grass (Alopecurus myosuroides), a problem weed of cereal crops in Northern Europe. The dominant 40 kDa carboxyesterase was purified 1700‐fold and identified as a serine hydrolase by affinity labelling with a biotinylated fluorophosphonate suicide substrate. MS–MS sequencing of a peptide digest identified it to be a member of the GDSL family of serine hydrolases. The full‐length A. myosuroides hydrolase (Amgdsh1) was cloned by RACE‐PCR and expressed in the yeast Pichia pastoris as a secreted enzyme. Expression was associated with activity towards AOPP esters. AmGDSH1 was predicted to be glycosylated and exported to the apoplast in planta. Based on the analysis of related sequences in monocotyledonous plants an alternative classification of the GDSL plant hydrolase superfamily is suggested and their importance in endogenous metabolism and herbicide bioactivation in crops and weeds discussed.
Biological functions of ent‐ and syn‐copalyl diphosphate synthases in rice: key enzymes for the branch point of gibberellin and phytoalexin biosynthesis
Authors: Otomo Kazuko , Kenmoku Hiromichi , Oikawa Hideaki , König Wilfried A. , Toshima Hiroaki , Mitsuhashi Wataru , Yamane Hisakazu , Sassa Takeshi , Toyomasu Tomonobu
Published: September 2004

Rice (Oryza sativa L.) produces ent‐copalyl diphosphate (ent‐CDP) and syn‐CDP as precursors for several classes of phytoalexins and the phytohormones, gibberellins (GAs). It has recently been shown that a loss‐of‐function mutation of OsCPS1, a gene encoding a putative ent‐CDP synthase, results in a severely GA‐deficient dwarf phenotype in rice. To clarify the biological functions of the ent‐ and syn‐CDP synthases involved in the biosynthesis of phytoalexins and/or GAs, we isolated two cDNAs, OsCyc1 and OsCyc2, encoding putative diterpene cyclases from ultraviolet (UV)‐irradiated rice leaves (cv. Nipponbare). The production of phytoalexins in rice leaves is known to be highly induced by UV treatment. Using a bacterial expression system, we demonstrated that OsCyc1 encodes syn‐CDP synthase and that OsCyc2 and OsCPS1 encode ent‐CDP synthase. The level of expression of the OsCyc1 and OsCyc2 transcripts in rice leaves increased drastically in response to UV treatment, whereas expression of the OsCPS1 transcript was not induced by UV light. These results suggest that OsCyc1, OsCyc2 and OsCPS1 are responsible for the biosynthesis of momilactones A and B and oryzalexin S, oryzalexins A–F and phytocassanes A–E, and GAs, respectively. Our results strongly suggest the presence of two ent‐CDP synthase isoforms in rice, one that participates in the biosynthesis of GAs and a second that is involved in the biosynthesis of phytoalexins.
Freezing‐sensitive tomato has a functional CBF cold response pathway, but a CBF regulon that differs from that of freezing‐tolerant Arabidopsis
Authors: Zhang Xin , Fowler Sarah G. , Cheng Hongmei , Lou Yigong , Rhee Seung Y. , Stockinger Eric J. , Thomashow Michael F.
Published: September 2004

Many plants increase in freezing tolerance in response to low temperature, a process known as cold acclimation. In Arabidopsis, cold acclimation involves action of the CBF cold response pathway. Key components of the pathway include rapid cold‐induced expression of three homologous genes encoding transcriptional activators, CBF1, 2 and 3 (also known as DREB1b, c and a, respectively), followed by expression of CBF‐targeted genes, the CBF regulon, that increase freezing tolerance. Unlike Arabidopsis, tomato cannot cold acclimate raising the question of whether it has a functional CBF cold response pathway. Here we show that tomato, like Arabidopsis, encodes three CBF homologs, LeCBF1–3 (Lycopersicon esculentum CBF1–3), that are present in tandem array in the genome. Only the tomato LeCBF1 gene, however, was found to be cold‐inducible. As is the case for Arabidopsis CBF1–3, transcripts for LeCBF1–3 did accumulate in response to mechanical agitation, but not in response to drought, ABA or high salinity. Constitutive overexpression of LeCBF1 in transgenic Arabidopsis plants induced expression of CBF‐targeted genes and increased freezing tolerance indicating that LeCBF1 encodes a functional homolog of the Arabidopsis CBF1–3 proteins. However, constitutive overexpression of either LeCBF1 or AtCBF3 in transgenic tomato plants did not increase freezing tolerance. Gene expression studies, including the use of a cDNA microarray representing approximately 8000 tomato genes, identified only four genes that were induced 2.5‐fold or more in the LeCBF1 or AtCBF3 overexpressing plants, three of which were putative members of the tomato CBF regulon as they were also upregulated in response to low temperature. Additional experiments indicated that of eight tomato genes that were likely orthologs of Arabidopsis CBF regulon genes, none were responsive to CBF overexpression in tomato. From these results, we conclude that tomato has a complete CBF cold response pathway, but that the tomato CBF regulon differs from that of Arabidopsis and appears to be considerably smaller and less diverse in function.
Characterization of AtCHX17, a member of the cation/H+ exchangers, CHX family, from Arabidopsis thaliana suggests a role in K+ homeostasis
Authors: Cellier Françoise , Conéjéro Geneviève , Ricaud Lilian , Luu Doan Trung , Lepetit Marc , Gosti Françoise , Casse Francine
Published: September 2004

The Arabidopsis genome contains many sequences annotated as encoding H+‐coupled cotransporters. Among those are the members of the cation:proton antiporter‐2 (CPA2) family (or CHX family), predicted to encode Na+,K+/H+ antiporters. AtCHX17, a member of the CPA2 family, was selected for expression studies, and phenotypic analysis of knockout mutants was performed. AtCHX17 expression was only detected in roots. The gene was strongly induced by salt stress, potassium starvation, abscisic acid (ABA) and external acidic pH. Using the β‐glucuronidase reporter gene strategy and in situ RT‐PCR experiments, we have found that AtCHX17 was expressed preferentially in epidermal and cortical cells of the mature root zones. Knockout mutants accumulated less K+ in roots in response to salt stress and potassium starvation compared with the wild type. These data support the hypothesis that AtCHX17 is involved in K+ acquisition and homeostasis.
Dissecting large and complex genomes: flow sorting and BAC cloning of individual chromosomes from bread wheat
Authors: Šafář Jan , Bartoš Jan , Janda Jaroslav , Bellec Arnaud , Kubaláková Marie , Valárik Miroslav , Pateyron Stéphanie , Weiserová Jitka , Tušková Radka , Číhalíková Jarmila , Vrána Jan , Šimková Hana , Faivre‐Rampant Patricia , Sourdille Pierre , Caboche Michel , Bernard Michel , Doležel Jaroslav , Chalhoub Boulos
Published: September 2004

The analysis of the complex genome of common wheat (Triticum aestivum, 2n = 6x = 42, genome formula AABBDD) is hampered by its large size (∼17 000 Mbp) and allohexaploid nature. In order to simplify its analysis, we developed a generic strategy for dissecting such large and complex genomes into individual chromosomes. Chromosome 3B was successfully sorted by flow cytometry and cloned into a bacterial artificial chromosome (BAC), using only 1.8 million chromosomes and an adapted protocol developed for this purpose. The BAC library (designated as TA‐3B) consists of 67 968 clones with an average insert size of 103 kb. It represents 6.2 equivalents of chromosome 3B with 100% coverage and 90% specificity as confirmed by genetic markers. This method was validated using other chromosomes and its broad application and usefulness in facilitating wheat genome analysis were demonstrated by target characterization of the chromosome 3B structure through cytogenetic mapping. This report on the successful cloning of flow‐sorted chromosomes into BACs marks the integration of flow cytogenetics and genomics and represents a great leap forward in genetics and genomic analysis.
High humidity suppresses ssi4‐mediated cell death and disease resistance upstream of MAP kinase activation, H2O2 production and defense gene expression
Authors: Zhou Fasong , Menke Frank L.H. , Yoshioka Keiko , Moder Wolfgang , Shirano Yumiko , Klessig Daniel F.
Published: September 2004

The Arabidopsis ssi4 mutant, which exhibits spontaneous lesion formation, constitutive expression of pathogenesis‐related (PR) genes and enhanced resistance to virulent bacterial and oomycete pathogens, contains a gain‐of‐function mutation in a TIR‐NBS‐LRR type R gene. Epistatic analyses revealed that both PR gene expression and disease resistance are activated via a salicylic acid (SA)‐ and EDS1‐dependent, but NPR1‐ and NDR1‐independent signaling pathway. In this study, we demonstrate that in moderate relative humidity (RH; 60%), the ssi4 mutant accumulates H2O2 and SA prior to lesion formation and displays constitutive activation of the MAP kinases AtMPK6 and AtMPK3. It also constitutively expresses a variety of defense‐associated genes, including those encoding the WRKY transcription factors AtWRKY29 and AtWRKY6, the MAP kinases AtMPK6 and AtMPK3, the powdery mildew R proteins RPW8.1 and RPW8.2, EDS1 and PR proteins. All of these ssi4‐induced responses, as well as the chlorotic, stunted morphology and enhanced disease resistance phenotype, are suppressed by high RH (95%) growth conditions. Thus, a humidity sensitive factor (HSF) appears to function at an early point in the ssi4 signaling pathway. All ssi4 phenotypes, except for MAP kinase activation, also were suppressed by the eds1‐1 mutation. Thus, ssi4‐induced MAP kinase activation occurs downstream of the HSF but either upstream of EDS1 or on a separate branch of the ssi4 signaling pathway. SA is a critical signaling component in ssi4‐mediated defense responses. However, exogenously supplied SA failed to restore lesion formation in high RH‐grown ssi4 plants, although it induced defense gene expression. Thus, additional signals also are involved.
Optimal timing for living‐related liver transplantation in children
Authors: Kimura Takuya , Hasegawa Toshimichi , Sasaki Takashi , Shimizu Yoshiyuki , Mushiake Sohtarou , Fukuzawa Masahiro , Okada Akira
Published: September 2004

Abstract:  Background:  The pediatric end stage liver disease (PELD) score has been used widely to prioritize children awaiting cadaveric liver transplantation (LTx). To establish the objective parameter for optimal timing of living‐related LTx (LRLTx), we have assessed our cases using the PELD score.

Methods:  From 1997 to 2002, 24 children were evaluated 28 times for the indication of LRLTx. Among them, 15 were for jaundice and nine for growth failure, hepatopulmonary syndrome, and variceal bleeding. Nine of 24 children underwent LRLTx. They were divided into several groups according to their clinical course. The PELD score consisted of age, albumin, total bilirubin, prothrombine time‐international ratio (INR) and growth failure. A cut‐off value was obtained by the highest positive and negative predictive value.

Results:  The PELD score in cases whose indication for LRLTx was approved was significantly higher compared with the cases who were not, and a cut‐off value of 4 was obtained. The PELD score in cases who were alive after LRLTx was significantly lower compared with the cases who died after LRLTx or evaluation of the indication, and a cut‐off value of 22 was established.

Conclusion:  LRLTx may be considered when the PELD score exceeds 4, and LRLTx may be required immediately when the PELD score exceeds 22.