First Report of Shining Willow as a Host Plant for Septoria musiva.
During the summer of 2001, leaf spots resembling those caused by Septoria musiva Peck. were observed on shining willow (Salix lucida Mühl. subsp. lucida) at Leclerville, Québec, Canada (46°34'19″N,71°59'35″W). Affected leaves had brown, necrotic leaf spots (>5mm in diameter) surrounded by a darker brown halo. Conidia were cylindrical, straight to curved with 1 to 4 septa, 28 to 54 × 3.5 to 4 μm, and were produced in pycnidia located on the abaxial surface in the center of the leaf spots. The causal agent of this disease was successfully isolated by germinating the conidia on corn meal agar that was supplemented with streptomycin (50mg/ml) and chloramphenicol (300mg/ml) and followed with the transfer of the germinated conidia to potato dextrose agar. Leaf symptoms and morphology matched those of S. musiva, the cause of leaf spot and stem canker of hybrid poplars in North America (2,4). The internal transcribed spacers and the 5.8S portion of the rDNA were amplified using PCR with the ITS1 (5'-TCC GTA GGT GAA CCT GCG G-3') and ITS2 (5'-GCT GCG TTC TTC ATC GAT GC-3') primer pair on total genomic DNA extracted from a pure culture of the pathogen. The rDNA sequence obtained (GenBank Accession No. AY555277) had 100% identity at 506 base positions with the ITS1, 5.8S, and ITS2 of three S. musiva isolates from Québec and one from Wisconsin (GenBank Accession Nos. AY549464 to AY549467). To test for pathogenicity, excised leaf disks from plants propagated by softwood cuttings of the source plant and from one hybrid poplar clone (Populus maximowiczii × P. xjackii) were inoculated with 3 μl of a suspension of ground mycelium or sterile water (control). Disks were placed in a 24-well tissue culture plate with 1 ml of distilled water per well and incubated in a growth room maintained at 22°C with a 16-h photoperiod. After 1 month, symptoms were similar to those previously observed. Isolates collected from shining willow or hybrid poplar were able to induce S. musiva leaf spot symptoms on leaf disks excised from shining willow or the hybrid poplar clone. From symptomatic leaf disks, S. musiva was consistently reisolated. To our knowledge, this is the first report of S. musiva on a member of the genus Salix. S. didyma, S. salicicola, and S. salicina have been reported from leaves of species of Salix (1,3). Only a vague morphological description of S. didyma was found (3). Moreover, conidia of S. salicicola (20 to 50 × 2.5 to 3.5 μm) and S. salicina (40 to 60 μm long, unspecified width) overlap dimensions of S. musiva conidia (1). There is a need to reexamine the relationships between these species of Septoria. Evidently, the complete host range of S. musiva is not yet known. References: (1) L. Lanier et al. Mycologie et Pathologie Forestières. Masson. Paris, 1978. (2) M. E. Ostry. Eur. J. For. Pathol. 17:158, 1987. (3) P. A. Saccardo. Sylloge fungurum omnium hucusque cognitorum. Patavii: Sumptibus Auctoris, 1882. (4) L. J. Spielman et al. Plant Dis. 70:968, 1986.


Stress-induced glucocorticoids at the earliest stages of herpes simplex virus-1 infection suppress subsequent antiviral immunity, implicating impaired dendritic cell function.
The systemic elevation of psychological stress-induced glucocorticoids strongly suppresses CD8(+) T cell immune responses resulting in diminished antiviral immunity. However, the specific cellular targets of stress/glucocorticoids, the timing of exposure, the chronology of immunological events, and the underlying mechanisms of this impairment are incompletely understood. In this study, we address each of these questions in the context of a murine cutaneous HSV infection. We show that exposure to stress or corticosterone in only the earliest stages of an HSV-1 infection is sufficient to suppress, in a glucocorticoid receptor-dependent manner, the subsequent antiviral immune response after stress/corticosterone has been terminated. This suppression resulted in early onset and delayed resolution of herpetic lesions, reduced viral clearance at the site of infection and draining popliteal lymph nodes (PLNs), and impaired functions of HSV-specific CD8(+) T cells in PLNs, including granzyme B and IFN-gamma production and the ability to degranulate. In knockout mice lacking glucocorticoid receptors only in T cells, we show that these impaired CD8(+) T cell functions are not due to direct effects of stress/corticosterone on the T cells, but the ability of PLN-derived dendritic cells to prime HSV-1-specific CD8(+) T cells is functionally impaired. These findings highlight the susceptibility of critical early events in the generation of an antiviral immune response to neuroendocrine modulation and implicate dendritic cells as targets of stress/glucocorticoids in vivo. These findings also provide insight into the mechanisms by which the clinical use of glucocorticoids contributes to altered immune responses in patients with viral infections or tumors.


Infectious circular DNA of human adenovirus type 5: regeneration of viral DNA termini from molecules lacking terminal sequences.
A series of plasmids containing the entire human adenovirus genome with viral DNA termini joined 'head to tail' has been isolated. Several plasmids were able to generate infectious virus following transfection of human cells in spite of having small deletions and rearrangements at the junctions of termini. One plasmid has lost 2 bp of DNA from one end of the viral genome and 11 bp from the other end yet produced viruses with complete wild-type sequences at both ends of the genome. We propose a model for replication of viral DNA off circular templates in which regeneration of terminal information involves translocation of primer and polymerase during initiation of DNA replication. The model suggests a novel mechanism for extension of the 5' ends of linear DNA molecules which could be applicable to chromosomal telomeres.


Assessment of wool waste and hair waste as soil amendment and nutrient source.
A field and two container experiments were conducted to assess uncomposted wool and hair wastes as a nutrient source for crops and to evaluate their potential to improve soil biological and chemical properties. Overall, addition of wool or hair waste to soil increased yields of basil (Ocimum basilicum L. 'Trakia'), thorn apple (Datura innoxia Mill. 'Inka'), peppermint (Mentha x piperita L. 'Black Mitchum'), and garden sage (Salvia officinalis L. 'Desislava'), increased NH(4)-N and NO(3)-N in soil, increased total N (and protein) content in plant tissue, stimulated soil microbial biomass, and decreased mycorrhizae colonization of plant roots of thorn apple but not in basil. Wool and hair waste additions to soil altered slightly the content and composition of plant secondary metabolites (essential oils or alkaloids); however, overall the constituents remained within the "typical" range for the respective crops. Scanning electron microscopy (SEM) and energy dispersive X-ray (EDX) analysis demonstrated that wool and hair wastes decompose slowly under field or greenhouse conditions, and act as a slow release S, N, P, and K fertilizer. These results, along with the measured concentrations of NO(3)-N in soil at harvest, suggest that the addition of wool or hair waste of only 3.3 g kg(-1) of soil may support two to five harvests or crops under greenhouse conditions and two to four field seasons in field production systems, and would improve soil biological and chemical characteristics. Further research is needed to optimize the rate of application of these waste materials to the nutrient requirements of specific crops to avoid nitrate leaching into the ground water. In addition, the effect of wool and hair waste on other environmental end points should also be further investigated before specific recommendations for growers are provided.


Comparison of digitally reconstructed radiographs generated from axial and helical CT scanning modes: a phantom study.
Digitally reconstructed radiographs (DRRs) play a vital role for verifying patient position for many radiotherapy treatments. As DRRs are generated from CT scans, image quality may be affected by the scanning mode (axial or helical). The aim of this study was to investigate the high and low contrast resolution and the spatial linearity in DRRs as a function of CT scanning mode and to highlight the significance of this variation, if any. A commercial CT phantom (Fluke Biomedical Model 76-417) was scanned with a Siemens Somatom Sensation 4 CT scanner using six variations of field of view, scanning mode and helical pitch. The image quality of the DRR's produced from the scans was evaluated in terms of high contrast resolution using the modulation transfer function, low contrast resolution using Image Quality Score method and spatial linearity. The results indicated that the high contrast resolution for the axial mode was comparable to that for the helical modes. The low contrast resolution with axial scanning was comparable to scans performed with a helical pitch of 1 but deteriorated at pitches greater than 1. The field of view was not found to impact the low contrast resolution. When changing scanning parameters the impact on DRR quality should be considered. For DRRs, particularly where visualisation of low contrast structures is desired, a helical scan with a pitch of 1 is recommended.


Modelling aqueous solubility of sodium chloride in clays at thermodynamic conditions of hydraulic fracturing by molecular simulations.
To address the high salinity of flow-back water during hydraulic fracturing, we have studied the equilibrium partitioning of NaCl and water between the bulk phase and clay pores. In shale rocks, such a partitioning can occur between fractures with a bulk-like phase and clay pores. We use an advanced Grand Canonical Monte Carlo (GCMC) technique based on fractional exchanges of dissolved ions and water molecules. We consider a typical shale gas reservoir condition of a temperature of 365 K and pressure of 275 bar, and we represent clay pores by pyrophyllite and Na-montmorillonite slits of a width ranging from about 7 to 28 Å, covering clay pores from dry clay to clay pores with a bulk-like layer in the middle of the pore. We employ the Joung-Cheatham model for ions, SPC/E model for water and CLAYFF for the clay pores. We first determine the chemical potentials for NaCl and water in the bulk phase using Osmotic Ensemble Monte Carlo simulations. The chemical potentials are then used in GCMC to simulate the adsorption of ions and water molecules in the clay pores, and in turn to predict the salt solubility in confined solutions. Besides the thermodynamic properties, we evaluate the structure and in-plane diffusion of the adsorbed fluids, and ion conductivities.


Cancer risks in BRCA2 mutation carriers.
Carriers of germline mutations in the BRCA2 gene are known to be at high risk of breast and ovarian cancers, but the risks of other cancers in mutation carriers are uncertain. We investigated these risks in 173 breast-ovarian cancer families with BRCA2 mutations identified at 20 centers in Europe and North America. Other cancer occurrence was determined in a final cohort of 3728 individuals, among whom 681 persons had breast or ovarian cancer and 3047 persons either were known mutation carriers, were first-degree relatives of known mutation carriers, or were first-degree relatives of breast or ovarian cancer patients. Incidence rates were compared with population-specific incidence rates, and relative risks (RRs) to carriers, together with 95% confidence intervals (CIs), were estimated by use of a maximum likelihood approach. Three hundred thirty-three other cancers occurred in this cohort. Statistically significant increases in risks were observed for prostate cancer (estimated RR = 4.65; 95% CI = 3.48-6.22), pancreatic cancer (RR = 3.51; 95% CI = 1. 87-6.58), gallbladder and bile duct cancer (RR = 4.97; 95% CI = 1. 50-16.52), stomach cancer (RR = 2.59; 95%CI = 1.46-4.61), and malignant melanoma (RR = 2.58; 95% CI = 1.28-5.17). The RR for prostate cancer for men below the age of 65 years was 7.33 (95% CI = 4.66-11.52). Among women who had already developed breast cancer, the cumulative risks of a second, contralateral breast cancer and of ovarian cancer by the age of 70 years were estimated to be 52.3% (95% CI = 41.7%-61.0%) and 15.9% (95% CI = 8.8%-22.5%), respectively. In addition to the large risks of breast and ovarian cancers, BRCA2 mutations may be associated with increased risks of several other cancers.


Do Response Styles Affect Estimates of Growth on Social-Emotional Constructs? Evidence from Four Years of Longitudinal Survey Scores.
Survey respondents employ different response styles when they use the categories of the Likert scale differently despite having the same true score on the construct of interest. For example, respondents may be more likely to use the extremes of the response scale independent of their true score. Research already shows that differing response styles can create a construct-irrelevant source of bias that distorts fundamental inferences made based on survey data. While some initial studies examine the effect of response styles on survey scores in longitudinal analyses, the issue of how response styles affect estimates of growth is underexamined. In this study, we conducted empirical and simulation analyses in which we scored surveys using item response theory (IRT) models that do and do not account for response styles, and then used those different scores in growth models and compared results. Generally, we found that response styles can affect estimates of growth parameters including the slope, but that the effects vary by psychological construct, response style, and IRT model used.


A combinatorial analysis using observational data identifies species that govern ecosystem functioning.
Understanding the relationship between biodiversity and ecosystem functioning has so far resulted from two main approaches: the analysis of species' functional traits, and the analysis of species interaction networks. Here we propose a third approach, based on the association between combinations of species or of functional groups, which we term assembly motifs, and observed ecosystem functioning. Each assembly motif describes a biotic environment in which species interactions have particular effects on a given ecosystem function. Clustering species in functional groups generates a classification of ecosystems based on their assembly motif. We evaluate the quality of each species clustering, that is its ability to predict an ecosystem function, by the coefficient of determination of the ecosystem classification. An iterative process then enables identifying the species clustering in functional groups that best accounts for the functioning of the observed ecosystems. We test this approach using experimental and simulated datasets. We show that our combinatorial analysis makes it possible to identify the combinations of functional groups of species whose interactions govern ecosystem functioning without any a priori knowledge of the species themselves or their interactions. Our combinatorial approach reproduces the associative learning of empirical ecologists, and proves to be powerful and parsimonious.


[Quadratus lumborum block for both cholecystectomy and right-sided nephrectomy].
The quadratus lumborum block (QLB) is a unilateral facial plane block, which extends from T4 to L1 at the paravertebral space. Injecting local anesthetic between the facial plane of the quadratus lumborum muscle and the psoas major muscle provides the block of the referred dermatomes. However, the number of published studies for QLB used in various surgical procedures is limited. In this case report, we share the results of a 46-years-old ASA I female patient, who underwent open surgery for cholecystectomy and right-sided nephrectomy in the same session. After general anesthesia was induced, QLB was performed in the left lateral decubitus position. A convex probe was placed in transversely between the iliac crest and the costal margin at the midclavicular line. 20cc of 0.25% bupivacaine was injected to the facial plane between the quadratus lumborum and psoas major muscles. The surgery lasted 4 hours and completed uneventfully. In the postoperative period, the patient was provided with morphine PCA. After 24 hours, the VAS score was 0, and the total demanded morphine dose was 13 mg. This case report recommends that QLB may be an adequate choice in the postoperative pain management for patients undergoing cholecystectomy and nephrectomy.


Brief Report: Coherent Motion Processing in Autism: Is Dot Lifetime an Important Parameter?
Contrasting reports of reduced and intact sensitivity to coherent motion in autistic individuals may be attributable to stimulus parameters. Here, we investigated whether dot lifetime contributes to elevated thresholds in children with autism. We presented a standard motion coherence task to 31 children with autism and 31 typical children, with both limited and unlimited lifetime conditions. Overall, children had higher thresholds in the limited lifetime condition than in the unlimited lifetime condition. However, children with autism were affected by this manipulation to the same extent as typical children and were equally sensitive to coherent motion. Our results suggest that dot lifetime is not a critical stimulus parameter and speak against pervasive difficulties in coherent motion perception in children with autism.


Only the Lonely: A Study of Loneliness Among University Students in Norway.
Loneliness is a major public health concern among college and university students, the evidence is inconsistent regarding whether there is an increasing trend or not. Furthermore, knowledge of the demographic determinants for loneliness are limited. The present study assesses recent trends of loneliness from 2014 to 2018, and explores demographic risk indicators of loneliness among students. Data was drawn from two waves of a national student health survey from 2014 and 2018 for higher education in Norway (the SHoT-study). In 2018, all 162,512 fulltime students in Norway were invited to participate and 50,054 students (69.1% women) aged 18-35 years were included (response rate = 30.8%). Loneliness was measured by "The Three-Item Loneliness Scale" (T-ILS) and one item from the Hopkins Symptom Checklist-25 (HSCL-25). Age showed a curvilinear association with loneliness, with the youngest and oldest students reporting the highest level of loneliness across all measures. Other significant demographic determinants of loneliness were being female, single and living alone. There was a considerable increase in loneliness from 2014 (16.5%) to 2018 (23.6%, p < .001), and the increase was particularly strong for males, for whom the proportion of feeling "extremely" lonely had more than doubled. The high rate of loneliness and the increasing trends indicate the need for preventive interventions in the student population.


Communal child-rearing: The role of nurses in school health.
Child-rearing remains a concern within our communities, especially because families of today lack primary parents due to multifaceted challenges such as working mothers, diseases and violence. Health-promoting school initiatives are necessary because they allow a multifaceted approach to child-rearing. They further provide a conducive environment for continued schoolchild-rearing moving from home to school. This study promotes an integrated approach to school care using the African concept of Ubuntu - solidarity and sense of community - as a point of departure. The socio-ecological model was used, which includes the work of the school healthcare nurse in contributing to holistic health services. An integrative review was conducted in January 2013, which included methodology studies, a theory review and a variety of studies related to school health. The studies were categorised according to school health, Ubuntu and the socio-ecological model. The role of school healthcare nurses entails acting as a liaison officer between a variety of stakeholders who work together to shape the future of children. Ubuntu, together with the socio-ecological model, can assist us to involve an entire community to raise children. This knowledge serves as a background to the planning of a school health programme. The role of the nurse in school health can also assist in collaborative efforts to formulate the programme and develop the competencies that will inform school health nurse training curricula.


Genome size variation in the pine fusiform rust pathogen Cronartium quercuum f.sp. fusiforme as determined by flow cytometry.
The genome size of the pine fusiform rust pathogen Cronartium quercuum f.sp. fusiforme (Cqf) was determined by flow cytometric analysis of propidium iodide-stained, intact haploid pycniospores with haploid spores of two genetically well characterized fungal species, Sclerotinia sclerotiorum and Puccinia graminis f.sp. tritici, as size standards. The Cqf haploid genome was estimated at ~90 Mb, similar to other Pucciniales species for which reference genome sequences are available. Twenty-three Cqf pycniospore samples were compared that comprised three samples obtained from naturally occurring pine galls and 20 samples obtained after artificial inoculation with parental isolates and their progeny. Significant variation in genome size (>10% of mean) was detected among unrelated as well as sibling Cqf samples. The unexpected plasticity in Cqf genome size observed among sibling samples is likely to be driven by meiosis between parental genomes that differ in size.


Individual cell migration serves as the driving force for optic vesicle evagination.
The cellular mechanisms underlying organ formation are largely unknown. We visualized early vertebrate eye morphogenesis at single-cell resolution by in vivo imaging in medaka (Oryzias latipes). Before optic vesicle evagination, retinal progenitor cells (RPCs) modulate their convergence in a fate-specific manner. Presumptive forebrain cells converge toward the midline, whereas medial RPCs remain stationary, predetermining the site of evagination. Subsequent optic vesicle evagination is driven by the active migration of individual RPCs. The analysis of mutants demonstrated that the retina-specific transcription factor rx3 determines the convergence and migration behaviors of RPCs. Hence, the migration of individual cells mediates essential steps of organ morphogenesis.


Effects of primary care team social networks on quality of care and costs for patients with cardiovascular disease.
Cardiovascular disease is the leading cause of mortality and morbidity in the United States. Primary care teams can be best suited to improve quality of care and lower costs for patients with cardiovascular disease. This study evaluates the associations between primary care team communication, interaction, and coordination (ie, social networks); quality of care; and costs for patients with cardiovascular disease. Using a sociometric survey, 155 health professionals from 31 teams at 6 primary care clinics identified with whom they interact daily about patient care. Social network analysis calculated variables of density and centralization representing team interaction structures. Three-level hierarchical modeling evaluated the link between team network density, centralization, and number of patients with a diagnosis of cardiovascular disease for controlled blood pressure and cholesterol, counts of urgent care visits, emergency department visits, hospital days, and medical care costs in the previous 12 months. Teams with dense interactions among all team members were associated with fewer hospital days (rate ratio [RR] = 0.62; 95% CI, 0.50-0.77) and lower medical care costs (-$556; 95% CI, -$781 to -$331) for patients with cardiovascular disease. Conversely, teams with interactions revolving around a few central individuals were associated with increased hospital days (RR = 1.45; 95% CI, 1.09-1.94) and greater costs ($506; 95% CI, $202-$810). Team-shared vision about goals and expectations mediated the relationship between social network structures and patient quality of care outcomes. Primary care teams that are more interconnected and less centralized and that have a shared team vision are better positioned to deliver high-quality cardiovascular disease care at a lower cost.


Toxoplasma gondii protease TgSUB1 is required for cell surface processing of micronemal adhesive complexes and efficient adhesion of tachyzoites.
Host cell invasion by Toxoplasma gondii is critically dependent upon adhesive proteins secreted from the micronemes. Proteolytic trimming of microneme contents occurs rapidly after their secretion onto the parasite surface and is proposed to regulate adhesive complex activation to enhance binding to host cell receptors. However, the proteases responsible and their exact function are still unknown. In this report, we show that T. gondii tachyzoites lacking the microneme subtilisin protease TgSUB1 have a profound defect in surface processing of secreted microneme proteins. Notably parasites lack protease activity responsible for proteolytic trimming of MIC2, MIC4 and M2AP after release onto the parasite surface. Although complementation with full-length TgSUB1 restores processing, complementation of Δsub1 parasites with TgSUB1 lacking the GPI anchor (Δsub1::ΔGPISUB1) only partially restores microneme protein processing. Loss of TgSUB1 decreases cell attachment and in vitro gliding efficiency leading to lower initial rates of invasion. Δsub1 and Δsub1::ΔGPISUB1 parasites are also less virulent in mice. Thus TgSUB1 is involved in micronemal protein processing and regulation of adhesive properties of macromolecular adhesive complexes involved in host cell invasion.


RegTransBase--a database of regulatory sequences and interactions based on literature: a resource for investigating transcriptional regulation in prokaryotes.
Due to the constantly growing number of sequenced microbial genomes, comparative genomics has been playing a major role in the investigation of regulatory interactions in bacteria. Regulon inference mostly remains a field of semi-manual examination since absence of a knowledgebase and informatics platform for automated and systematic investigation restricts opportunities for computational prediction. Additionally, confirming computationally inferred regulons by experimental data is critically important. RegTransBase is an open-access platform with a user-friendly web interface publicly available at http://regtransbase.lbl.gov. It consists of two databases - a manually collected hierarchical regulatory interactions database based on more than 7000 scientific papers which can serve as a knowledgebase for verification of predictions, and a large set of curated by experts transcription factor binding sites used in regulon inference by a variety of tools. RegTransBase captures the knowledge from published scientific literature using controlled vocabularies and contains various types of experimental data, such as: the activation or repression of transcription by an identified direct regulator; determination of the transcriptional regulatory function of a protein (or RNA) directly binding to DNA or RNA; mapping of binding sites for a regulatory protein; characterization of regulatory mutations. Analysis of the data collected from literature resulted in the creation of Putative Regulons from Experimental Data that are also available in RegTransBase. RegTransBase is a powerful user-friendly platform for the investigation of regulation in prokaryotes. It uses a collection of validated regulatory sequences that can be easily extracted and used to infer regulatory interactions by comparative genomics techniques thus assisting researchers in the interpretation of transcriptional regulation data.


Changes in the retreatment radiation tolerance of the spinal cord with time after the initial treatment.
To estimate, from experimental data, the retreatment radiation 'tolerances' of the spinal cord at different times after initial treatment. A model was developed to show the relationship between the biological effective doses (BEDs) for two separate courses of treatment with the BED of each course being expressed as a percentage of the designated 'retreatment tolerance' BED value, denoted [Formula: see text] and [Formula: see text]. The primate data of Ang et al. ( 2001 ) were used to determine the fitted parameters. However, based on rodent data, recovery was assumed to commence 70 days after the first course was complete, and with a non-linear relationship to the magnitude of the initial BED (BEDinit). The model, taking into account the above processes, provides estimates of the retreatment tolerance dose after different times. Extrapolations from the experimental data can provide conservative estimates for the clinic, with a lower acceptable myelopathy incidence. Care must be taken to convert the predicted [Formula: see text] value into a formal BED value and then a practical dose fractionation schedule. Used with caution, the proposed model allows estimations of retreatment doses with elapsed times ranging from 70 days up to three years after the initial course of treatment.


Musculoskeletal impairment survey in Rwanda: design of survey tool, survey methodology, and results of the pilot study (a cross sectional survey).
Musculoskeletal impairment (MSI) is an important cause of morbidity and mortality worldwide, especially in developing countries. Prevalence studies for MSI in the developing world have used varying methodologies and are seldom directly comparable. This study aimed to develop a new tool to screen for and diagnose MSI and to pilot test the methodology for a national survey in Rwanda. A 7 question screening tool to identify cases of MSI was developed through literature review and discussions with healthcare professionals. To validate the tool, trained rehabilitation technicians screened 93 previously identified gold standard 'cases' and 86 'non cases'. Sensitivity, specificity and positive predictive value were calculated. A standardised examination protocol was developed to determine the aetiology and diagnosis of MSI for those who fail the screening test. For the national survey in Rwanda, multistage cluster random sampling, with probability proportional to size procedures will be used for selection of a cross-sectional, nationally representative sample of the population. Households to be surveyed will be chosen through compact segment sampling and all individuals within chosen households will be screened. A pilot survey of 680 individuals was conducted using the protocol. : The screening tool demonstrated 99% sensitivity and 97% specificity for MSI, and a positive predictive value of 98%. During the pilot study 468 out of 680 eligible subjects (69%) were screened. 45 diagnoses were identified in 38 persons who were cases of MSI. The subjects were grouped into categories based on diagnostic subgroups of congenital (1), traumatic (17), infective (2) neurological (6) and other acquired(19). They were also separated into mild (42.1%), moderate (42.1%) and severe (15.8%) cases, using an operational definition derived from the World Health Organisation's International Classification of Functioning, Disability and Health. : The screening tool had good sensitivity and specificity and was appropriate for use in a national survey. The pilot study showed that the survey protocol was appropriate for measuring the prevalence of MSI in Rwanda. This survey is an important step to building a sound epidemiological understanding of MSI, to enable appropriate health service planning.


Does partial cystectomy compromise oncologic outcomes for patients with bladder cancer compared to radical cystectomy? A matched case-control analysis.
To our knowledge long-term oncologic outcomes following partial cystectomy for urothelial carcinoma remain to be defined. We evaluated patterns of recurrence and survival among matched patients treated with partial vs radical cystectomy for bladder cancer. We identified 86 patients who underwent partial cystectomy for pT1-4N0-1Mx urothelial carcinoma between 1980 and 2006 at our institution. They were matched 1:2 to patients undergoing radical cystectomy based on age, gender, pathological T stage and receipt of neoadjuvant chemotherapy. Survival was estimated using Kaplan-Meier analysis and compared with the log rank test. Median postoperative followup was 6.2 years (range 0 to 27). No difference was noted for 10-year distant recurrence-free survival (61% vs 66%, p = 0.63) or cancer specific survival (58% vs 63%, p = 0.67) between patients treated with partial and radical cystectomy, respectively. Interestingly, 4 of 86 patients (5%) who underwent partial cystectomy showed extravesical pelvic tumor recurrence postoperatively vs 29 of 167 (17%) who underwent radical cystectomy (p = 0.004). In addition, 33 of 86 patients (38%) were diagnosed with intravesical recurrence of tumor after partial cystectomy and 16 of 86 (19%) initially treated with partial cystectomy ultimately underwent radical cystectomy. Our matched analysis demonstrated no difference in metastasis-free or cancer specific survival between select patients undergoing partial cystectomy and those undergoing radical cystectomy. Nevertheless, patients treated with partial cystectomy remain at risk for intravesical recurrence and, thus, they should be counseled and surveilled accordingly.


Infection and vertical transmission of Kamiti river virus in laboratory bred Aedes aegypti mosquitoes.
Kamiti river virus (KRV) is an insect-only Flavivirus that was isolated from field-collected Ae. macintoshi mosquitoes in 1999, and is closely related to cell fusing agent virus. Both of these viruses belong to the family Flaviviridae, which also contains other viruses of medical importance, such as yellow fever virus, West Nile virus and dengue. Because Ae. macintoshi is the only known natural host to KRV, the main objective of this study was to establish the possibility that other mosquito hosts of the virus exist, by determining its ability to infect Ae. aegypti mosquitoes under laboratory conditions. The study also sought to determine the rates of infection and, subsequently, vertical transmission as a possible means of its maintenance and propagation in nature, given that it neither grows in vertebrate cells or mice. The mosquitoes were infected by the virus either as larvae or adults. Virus assay was done by re-isolation in tissue culture and indirect immunofluoresce assay methods. KRV infected Ae. aegypti mosquitoes, with the observed rates as high as 74 to 96 %. The virus was also transmitted vertically in these mosquitoes. Vertical transmission rates of 3.90 % were observed for the 2nd and 3rd ovarian cycles combined. These results suggest that Ae. aegypti mosquitoes are likely to be infected with KRV in nature, and that vertical transmission is the natural means by which it is maintained and propagated in this host, and possibly others.


Global MicroRNA Expression Profiling Identifies Unique MicroRNA Pattern of Radioresistant Glioblastoma Cells.
Glioblastoma multiforme (GBM) is the most aggressive intracranial tumor characterized with infaust prognosis. Despite advances in neurosurgical and radiotherapeutic techniques and chemotherapy, the median overall survival ranges between 12-15 months from diagnosis. The main cause of treatment failure is considered the presence of tumor cells resistant to conventional therapy, mainly radiotherapy. MicroRNAs (miRNAs) are small, non-coding RNAs that function as post-transcriptional regulators of gene expression and have been repeatedly proven to play important roles in pathogenesis and biological features of many cancers, including GBM and its radioresistant phenotype. In our study, we established radioresistant cells from the commonly used human GBM cell lines T98G, U87MG and U251. Consequently, we performed global miRNA expression profiling in both radioresistant and parental cell lines and identified 113 miRNAs with significantly different expression (p<0.05) between these two groups (73 miRNAs were up-regulated, 40 miRNAs were down-regulated). Some of these miRNAs have been previously described in relation to ionizing radiation, and others were herein identified for the first time. We believe that after deeper functional investigation of identified miRNAs in relation to radioresistance, these miRNAs present potential predictive biomarkers or therapeutic targets in GBM.


Nitrate reductase-mediated early nitric oxide burst alleviates oxidative damage induced by aluminum through enhancement of antioxidant defenses in roots of wheat (Triticum aestivum).
• Nitric oxide (NO) is an important signaling molecule involved in the physiological processes of plants. The role of NO release in the tolerance strategies of roots of wheat (Triticum aestivum) under aluminum (Al) stress was investigated using two genotypes with different Al resistances. • An early NO burst at 3 h was observed in the root tips of the Al-tolerant genotype Jian-864, whereas the Al-sensitive genotype Yang-5 showed no NO accumulation at 3 h but an extremely high NO concentration after 12 h. Stimulating NO production at 3 h in the root tips of Yang-5 with the NO donor relieved Al-induced root inhibition and callose production, as well as oxidative damage and ROS accumulation, while elimination of the early NO burst by NO scavenger aggravated root inhibition in Jian-864. • Synthesis of early NO in roots of Jian-864 was mediated through nitrate reductase (NR) but not through NO synthase. Elevated antioxidant enzyme activities were induced by Al stress in both wheat genotypes and significantly enhanced by NO donor, but suppressed by NO scavenger or NR inhibitor. • These results suggest that an NR-mediated early NO burst plays an important role in Al resistance of wheat through modulating enhanced antioxidant defense to adapt to Al stress.


The significance of needle placement site in acupuncture.
Traditional acupuncturists claim that correct needling of classical acupuncture loci is associated with a characteristic set of sensations usually referred to as 'Teh Chi'. The studies reported here examine this claim. In the first a multiple adjective sensation rating scale was developed and administered to 125 patients receiving acupuncture treatment. The results were subjected to principal components analysis and the first factor to emerge provided some support for the constellation of sensations corresponding to Teh Chi. In the second study the scale was used in a randomised controlled mixed single/double blind experiment in which 65 volunteers were stimulated at three classical and three non-classical (sham) needling sites by either a trained acupuncturist (single blind) or an anaesthetist (double blind). The results of the second study did not support the contention that the sensations of Teh Chi occur more frequently at classical acupuncture needling sites. The implications of the results for the practice of acupuncture are discussed.


Effects of nutrient variability on the genetic-based resistance of Eucalyptus globulus to a mammalian herbivore and on plant defensive chemistry.
Plant resistance to herbivores can be influenced not only by the independent effects of plant genotype and environmental variation, but by interactions between the two. The main aim of this study was to assess the effects of environmental variability (nutrient treatment) on the known genetic-based expression of resistance and defensive chemistry of Eucalyptus globulus to browsing by the generalist mammalian herbivore Trichosurus vulpecula. In a captive feeding trial, we measured intake of seedlings from one relatively resistant (Blue Gum Hill) and one relatively susceptible (St Helens) population of E. globulus grown under two nutrient treatments (no fertiliser, plus fertiliser). There was a significant genotypexfertiliser interaction effect on intake of E. globulus foliage by T. vulpecula, and the predicted genetic-based resistance of the two populations was expressed only for the non-fertilised treatment. Expression of resistance largely reflected the combined and inverse effects of nitrogen and condensed tannin concentrations. The expression of plant secondary metabolite concentration differed between compounds, but in all cases the effects of plant genotype and fertiliser treatment were independent. The formylated phloroglucinol compounds differed significantly between genotypes but not between fertiliser treatments. In contrast, the effect of plant genotype on the expression of condensed tannins was weak but they were significantly reduced by fertiliser. Essential oils were influenced by both plant genotype and fertiliser treatment and were significantly higher in the fertilised seedlings than in the non-fertilised seedlings. This study highlights interactive effects of plant genotype and environment in influencing the phenotypic expression of resistance in a eucalypt species to a mammalian browser. It also demonstrates that this interactive effect is the net result of independent effects of genotype and environment on plant chemistry and finally, that different groups of compounds within a plant can respond very differently to variation in environmental conditions.


Genetic Risk Score for Coronary Disease Identifies Predispositions to Cardiovascular and Noncardiovascular Diseases.
The taxonomy of cardiovascular (CV) diseases is divided into a broad spectrum of clinical entities. Many such diseases coincide in specific patient groups and suggest shared predisposition. This study focused on coronary artery disease (CAD) and investigated the genetic relationship to CV and non-CV diseases with reported CAD comorbidity. This study examined 425,196 UK Biobank participants to determine a genetic risk score (GRS) based on 300 CAD associated variants (CAD-GRS). This score was associated with 22 traits, including risk factors, diseases secondary to CAD, as well as comorbid and non-CV conditions. Sensitivity analyses were performed in individuals free from CAD or stable angina diagnosis. Hypercholesterolemia (odds ratio [OR]: 1.27; 95% CI: 1.26 to 1.29) and hypertension (OR: 1.11; 95% CI: 1.10 to 1.12) were strongly associated with the CAD-GRS, which indicated that the score contained variants predisposing to these conditions. However, the CAD-GRS was also significant in patients with CAD who were free of CAD risk factors (OR: 1.37; 95% CI: 1.30 to 1.44). The study observed significant associations between the CAD-GRS and peripheral arterial disease (OR: 1.28; 95% CI: 1.23 to 1.32), abdominal aortic aneurysms (OR: 1.28; 95% CI: 1.20 to 1.37), and stroke (OR: 1.08; 95% CI: 1.05 to 1.10), which remained significant in sensitivity analyses that suggested shared genetic predisposition. The score was also associated with heart failure (OR: 1.25; 95% CI: 1.22 to 1.29), atrial fibrillation (OR: 1.08; 95% CI: 1.05 to 1.10), and premature death (OR: 1.04; 95% CI: 1.02 to 1.06). These associations were abolished in sensitivity analyses that indicated that they were secondary to prevalent CAD. Finally, an inverse association was observed between the score and migraine headaches (OR: 0.94; 95% CI: 0.93 to 0.96). A wide spectrum of CV conditions, including premature death, might develop consecutively or in parallel with CAD for the same genetic roots. In conditions like heart failure, the study found evidence that the CAD-GRS could be used to stratify patients with no or limited genetic overlap with CAD risk. Increased genetic predisposition to CAD was inversely associated with migraine headaches.


Molecular analysis of the GSTT1 gene polymorphism in patients with clinical manifestation of atherosclerosis.
Atherosclerosis is a chronic inflammatory disease formed by the accumulation of lipids in the innermost layer and large-caliber artery (tunica intima). This accumulation, along with platelet factors, stimulates the proliferation of muscle cells in this region. Over than 400 genes may be related to the pathology since they regulate endothelial function, coagulation, inflammation, metabolism of amino acids, lipids, and carbohydrates. Glutathione S-transferases (GST) are enzymes that catalyze the polymorphic detoxification of metabolites produced by oxidative stress within the cells, which is induced by reactive oxygen species. GSTs are one of the defense mechanisms against oxidative stress damage. Due to genetic, cultural, and environmental factors, the rate of atherosclerosis is higher; however, an early diagnosis is crucial for the prevention and treatment of several complications related to the disease. The present study aimed to analyze the frequency of GSTT1 genotypes regarding the presence or absence of the polymorphism in patients with clinical manifestation of atherosclerosis. We collected 200 samples of peripheral blood of patients with the previous diagnosis of atherosclerosis based on clinical examination and imaging, and 100 samples of peripheral blood to compose the control group of patients without clinical manifestation of atherosclerosis. The polymorphism was assessed by PCR and analyzed on the agarose gel stained with 2.0% ethidium bromide. The frequency of the GSTT1 gene polymorphism was compared using the chi-square test (P < 0.05) and the G-test. In the case group, we detected 85.5% of patients with the GSTT1 genotype present and 14.5% of patients with the null genotype. A significant difference was observed between groups (case vs control) for the presence of the GSTT1 polymorphism. According to the analysis of the variable alcohol consumption, we found that in the case group the presence of the GSTT1 gene was higher in individuals who reported not drinking alcohol. In this study, the presence of the GSTT1 gene polymorphism in male patients with atherosclerosis was 1.5 times higher when compared to female patients. Regarding the variable time of smoking, we found that this genotype was more frequent in smokers for both case and control groups.


Interannual variability in reproductive traits of the Patagonian toothfish Dissostichus eleginoides around the sub-Antarctic island of South Georgia.
Commercial fisheries data, collected as part of an observer programme and covering the period 1997-2014, were utilized in order to define key reproductive traits and spawning dynamics of the Patagonian toothfish Dissostichus eleginoides at South Georgia. Multi-year spawning site fidelity of D. eleginoides was revealed through the identification of previously unknown spawning hotspots. Timing of female spawning was shown to have shifted later, leading to a shorter spawning duration. A decrease in length and mass of female and male spawning fish and a reduced number of large spawning fish was found, evidence of a change in size structure of spawning D. eleginoides. During the study period fewer later maturity stage females (including spawning stage) were observed in conjunction with increased proportions of early stage female D. eleginoides. The findings are discussed in the context of reproductive success, with consideration of the possible effects such spawning characteristics and behaviours may have on egg and larval survival. This work presents the first long-term assessment of D. eleginoides spawning dynamics at South Georgia and provides valuable knowledge for both the ecology of the species and for future fisheries management of this commercially important species.


Defining Advance Care Planning for Adults: A Consensus Definition From a Multidisciplinary Delphi Panel.
Despite increasing interest in advance care planning (ACP) and previous ACP descriptions, a consensus definition does not yet exist to guide clinical, research, and policy initiatives. The aim of this study was to develop a consensus definition of ACP for adults. We convened a Delphi panel of multidisciplinary, international ACP experts consisting of 52 clinicians, researchers, and policy leaders from four countries and a patient/surrogate advisory committee. We conducted 10 rounds using a modified Delphi method and qualitatively analyzed panelists' input. Panelists identified several themes lacking consensus and iteratively discussed and developed a final consensus definition. Panelists identified several tensions concerning ACP concepts such as whether the definition should focus on conversations vs. written advance directives; patients' values vs. treatment preferences; current shared decision making vs. future medical decisions; and who should be included in the process. The panel achieved a final consensus one-sentence definition and accompanying goals statement: "Advance care planning is a process that supports adults at any age or stage of health in understanding and sharing their personal values, life goals, and preferences regarding future medical care. The goal of advance care planning is to help ensure that people receive medical care that is consistent with their values, goals and preferences during serious and chronic illness." The panel also described strategies to best support adults in ACP. A multidisciplinary Delphi panel developed a consensus definition for ACP for adults that can be used to inform implementation and measurement of ACP clinical, research, and policy initiatives.


Evaluation of the effect of calcium gluconate and bovine thrombin on the temporal release of transforming growth factor beta 1 and platelet-derived growth factor isoform BB from feline platelet concentrates.
There are not reported regarding the protocols for obtaining platelet concentrates (PC) in cats for medical purposes. The objectives of this study were: 1) to describe a manual method for producing two kinds of PC in cats (PC-A and PC-B), 2) to describe the cellular population of the PC, 3) to measure and compare the effect of calcium gluconate (CG) and bovine thrombin (BT) on the temporal release of transforming growth factor beta 1 (TGF-β1) and platelet-derived growth factor type BB (PDGF-BB) at 3 and 12 hours post-activation and 4) to establish correlations between the cellular population of both PCs and the concentration of growth factors (GF). Blood samples were taken from 16 cats for complete blood count, plasma collection and PC preparation. The PC were arbitrarily divided into two fractions, specifically, PC-A (lower fraction) and PC-B (upper fraction). The platelet counts were significantly different (P<0.05) between the PC and whole blood but not between the PC fractions. The TGF-β1 concentration efficiencies for PC-A and PC-B activated with CG were 42.86% and 46.54%, and activated with BT were 42.88% and 54.64%, respectively. The PDGF-BB concentration efficiencies for PC-A and PC-B activated with CG were 61.36% and 60.61%, and activated with BT were 65.64% and 72.12%, respectively. The temporal release of GFs showed no statistically significant difference (P>0.05) between the activating substances at the time or for any PC fraction. Whatever the activation means, these preparations of cat PC provide significant concentrations of platelets and GFs for possible clinical or experimental use.


Genetic diversity, inter-gene pool introgression and nutritional quality of common beans (Phaseolus vulgaris L.) from Central Africa.
The Great Lakes region of Central Africa is a major producer of common beans in Africa. The region is known for high population density and small average farm size. The common bean represents the most important legume crop of the region, grown on over a third of the cultivated land area, and the per capita consumption is among the highest in the world for the food crop. The objective of this study was to evaluate the genetic diversity in a collection of 365 genotypes from the Great Lakes region of Central Africa, including a large group of landraces from Rwanda as well as varieties from primary centers of diversity and from neighboring countries of Central Africa, such as the Democratic Republic of Congo and Uganda, using 30 fluorescently labeled microsatellite markers and automated allele detection. In addition, the landraces were evaluated for their seed iron and zinc concentration to determine if genetic diversity influenced nutritional quality. Principal coordinate and neighbor-joining analyses allowed the separation of the landraces into 132 Andean and 195 Mesoamerican (or Middle American) genotypes with 32 landraces and 6 varieties intermediate between the gene pools and representing inter-gene pool introgression in terms of seed characteristics and alleles. Genetic diversity and the number of alleles were high for the collection, reflecting the preference for a wide range of seed types in the region and no strong commercial class preference, although red, red mottled and brown seeded beans were common. Observed heterozygosity was also high and may be explained by the common practice of maintaining seed and plant mixtures, a coping strategy practiced by Central African farmers to reduce the effects of abiotic and biotic stresses. Finally, nutritional quality differed between the gene pools with respect to seed iron and zinc concentration, while genotypes from the intermediate group were notably high in both minerals. In conclusion, this study has shown that Central African varieties of common bean are a source of wide genetic diversity with variable nutritional quality that can be used in crop improvement programs for the region.


Endothelial alpha3beta1-integrin represses pathological angiogenesis and sustains endothelial-VEGF.
Integrin alpha3beta1 is a major receptor for laminin. The expression levels of laminins-8 and -10 in the basement membrane surrounding blood vessels are known to change during tumor angiogenesis. Although some studies have suggested that certain ligands of alpha3beta1 can affect angiogenesis either positively or negatively, either a direct in vivo role for alpha3beta1 in this process or its mechanism of action in endothelial cells during angiogenesis is still unknown. Because the global genetic ablation of alpha3-integrin results in an early lethal phenotype, we have generated conditional-knockout mice where alpha3 is deleted specifically in endothelial cells (ec-alpha3-/-). Here we show that ec-alpha3-/- mice are viable, fertile, and display enhanced tumor growth, elevated tumor angiogenesis, augmented hypoxia-induced retinal angiogenesis, and increased vascular endothelial growth factor (VEGF)-mediated neovascularization ex vivo and in vivo. Furthermore, our data provide a novel method by which an integrin may regulate angiogenesis. We show that alpha3beta1 is a positive regulator of endothelial-VEGF and that, surprisingly, the VEGF produced by endothelial cells can actually repress VEGF-receptor 2 (Flk-1) expression. These data, therefore, identify directly that endothelial alpha3beta1 negatively regulates pathological angiogenesis and implicate an unexpected role for low levels of endothelial-VEGF as an activator of neovascularization.


Visual Cues of Object Properties Differentially Affect Anticipatory Planning of Digit Forces and Placement.
Studies on anticipatory planning of object manipulation showed initial task failure (i.e., object roll) when visual object shape cues are incongruent with other visual cues, such as weight distribution/density (e.g., symmetrically shaped object with an asymmetrical density). This suggests that shape cues override density cues. However, these studies typically only measured forces, with digit placement constrained. Recent evidence suggests that when digit placement is unconstrained, subjects modulate digit forces and placement. Thus, unconstrained digit placement might be modulated on initial trials (since it is an explicit process), but not forces (since it is an implicit process). We tested whether shape and density cues would differentially influence anticipatory planning of digit placement and forces during initial trials of a two-digit object manipulation task. Furthermore, we tested whether shape cues would override density cues when cues are incongruent. Subjects grasped and lifted an object with the aim of preventing roll. In Experiment 1, the object was symmetrically shaped, but with asymmetrical density (incongruent cues). In Experiment 2, the object was asymmetrical in shape and density (congruent cues). In Experiment 3, the object was asymmetrically shaped, but with symmetrical density (incongruent cues). Results showed differential modulation of digit placement and forces (modulation of load force but not placement), but only when shape and density cues were congruent. When shape and density cues were incongruent, we found collinear digit placement and symmetrical force sharing. This suggests that congruent and incongruent shape and density cues differentially influence anticipatory planning of digit forces and placement. Furthermore, shape cues do not always override density cues. A continuum of visual cues, such as those alluding to shape and density, need to be integrated.


Liver-specific mRNA for Insig-2 down-regulated by insulin: implications for fatty acid synthesis.
Insig-1 and -2 are closely related proteins of the endoplasmic reticulum (ER) that block proteolytic activation of sterol regulatory element-binding proteins (SREBPs), transcription factors that activate the synthesis of cholesterol and fatty acids in liver and other organs. When cellular cholesterol levels are high, Insig proteins bind and trap SREBP cleavage-activating protein (SCAP), retaining it in the ER and preventing it from escorting SREBPs from ER to the site of proteolytic activation in the Golgi complex. Here, we report the discovery of a liver-specific transcript of Insig-2, designated Insig-2a. This transcript and the ubiquitous transcript, designated Insig-2b, differ through the use of different promoters that produce different noncoding first exons that splice into a common second exon. Although the Insig-2a and -2b mRNAs encode identical proteins, they differ in patterns of regulation. Insig-2a is the predominant transcript in livers of fed animals, and it is selectively down-regulated by insulin. Insig-2a mRNA increases when mice are fasted, and it declines when they are refed. The transcript also increases in livers of rats whose insulin-secreting pancreatic beta cells have been destroyed by streptozotocin, and it is reduced when insulin is injected. The insulin-mediated fall in Insig-2a may allow SREBP-1c to be processed, thereby allowing insulin to stimulate fatty acid synthesis, even under conditions in which hepatic cholesterol levels are elevated.


Development and assessment of a novel ex vivo corneal culture technique involving an agarose-based dome scaffold for use as a model of in vivo corneal wound healing in dogs and rabbits.
To develop and assess a novel ex vivo corneal culture technique involving an agarose-based dome scaffold (ABDS) for use as a model of in vivo corneal wound healing in dogs and rabbits. Corneas from clinically normal dogs (paired corneas from 8 dogs and 8 single corneas) and rabbits (21 single corneas). 8 single dog corneas (DCs), 1 DC from each pair, and 10 rabbit corneas (RCs) were wounded with an excimer laser; 1 DC from each pair and 11 RCs remained unwounded. Corneas were cultured for 21 days on ABDSs (8 pairs of DCs and all RCs) or on flat-topped scaffolds (8 single DCs). The surface area of corneal fluorescein retention was measured every 6 (DCs) or 12 (RCs) hours until full corneal epithelialization was detected. Changes in corneal clarity were evaluated at 0, 7, 14, and 21 days. Median time to full epithelialization for wounded dog and rabbit corneas was 48 and 60 hours, respectively; among wounded DCs, time to full epithelization did not differ by scaffold type. After 21 days of culture on ABDSs, all DCs and RCs that epithelialized developed a circular, diffuse, cloud-like pattern of optical haze, whereas DCs cultured on flat-topped scaffolds developed a focal, crater-like region of optical haze. All corneas on the ABDSs maintained convex curvature throughout the study. Wounded ex vivo DCs and RCs cultured on ABDSs reliably epithelialized, formed optical haze (consistent with in vivo wound healing), and maintained convex curvature. This culture technique may be adaptable to other species.


Effects of pulse-delivered inhaled nitric oxide administration on pulmonary perfusion and arterial oxygenation in dorsally recumbent isoflurane-anesthetized horses.
To image the spatial distribution of pulmonary blood flow by means of scintigraphy, evaluate ventilation-perfusion (VA/Q) matching and pulmonary blood shunting (Qs/Qt) by means of the multiple inert gas elimination technique (MIGET), and measure arterial oxygenation and plasma endothelin-1 concentrations before, during, and after pulse-delivered inhaled nitric oxide (PiNO) administration to isoflurane-anesthetized horses in dorsal recumbency. 3 healthy adult Standardbreds. Nitric oxide was pulsed into the inspired gases in dorsally recumbent isoflurane-anesthetized horses. Assessment of VA/Q matching, Qs/Qt, and Pao2 content was performed by use of the MIGET, and spatial distribution of pulmonary blood flow was measured by perfusion scintigraphy following IV injection of technetium Tc 99m-labeled macroaggregated human albumin before, during, and 30 minutes after cessation of PiNO administration. During PiNO administration, significant redistribution of blood flow from the dependent regions to the nondependent regions of the lungs was found and was reflected by improvements in VA/Q matching, decreases in Qs/Qt, and increases in Pao2 content, all of which reverted to baseline values at 30 minutes after PiNO administration. Administration of PiNO in anesthetized dorsally recumbent horses resulted in redistribution of pulmonary blood flow from dependent atelectatic lung regions to nondependent aerated lung regions. Because hypoxemia is commonly the result of atelectasis in anesthetized dorsally recumbent horses, the addition of nitric oxide to inhaled gases could be used clinically to alleviate hypoxemia in horses during anesthesia.


Profiling microRNAs in lung tissue from pigs infected with Actinobacillus pleuropneumoniae.
MicroRNAs (miRNAs) are a class of non-protein-coding genes that play a crucial regulatory role in mammalian development and disease. Whereas a large number of miRNAs have been annotated at the structural level during the latest years, functional annotation is sparse. Actinobacillus pleuropneumoniae (APP) causes serious lung infections in pigs. Severe damage to the lungs, in many cases deadly, is caused by toxins released by the bacterium and to some degree by host mediated tissue damage. However, understanding of the role of microRNAs in the course of this infectious disease in porcine is still very limited. In this study, the RNA extracted from visually unaffected and necrotic tissue from pigs infected with Actinobacillus pleuropneumoniae was subjected to small RNA deep sequencing. We identified 169 conserved and 11 candidate novel microRNAs in the pig. Of these, 17 were significantly up-regulated in the necrotic sample and 12 were down-regulated. The expression analysis of a number of candidates revealed microRNAs of potential importance in the innate immune response. MiR-155, a known key player in inflammation, was found expressed in both samples. Moreover, miR-664-5p, miR-451 and miR-15a appear as very promising candidates for microRNAs involved in response to pathogen infection. This is the first study revealing significant differences in composition and expression profiles of miRNAs in lungs infected with a bacterial pathogen. Our results extend annotation of microRNA in pig and provide insight into the role of a number of microRNAs in regulation of bacteria induced immune and inflammatory response in porcine lung.


A phase I trial of stereotactic external beam radiation for subfoveal choroidal neovascular membranes in age-related macular degeneration.
Effective treatment for neovascular age-related macular degeneration (AMD) is currently limited. Radiation therapy, a therapeutic approach with known antiangiogenic properties, has been investigated as a modality to prevent severe visual loss in AMD. Most of the studies using external beam radiation employed <25 Gy to the whole eye, which is below the dose of radiation that is toxic to the retina and optic nerve ( approximately 50 Gy and approximately 59 Gy, respectively). Stereotactic fractionated external beam radiation (St-EBR) is a method that allows radiation to be delivered to a small, defined area. We investigated the effects of St-EBR in incremental doses up to 40 Gy on neovascular AMD. Patients with clinical signs and fluorescein angiography demonstrating neovascular AMD, visual acuity (VA) better than 20/400 and ineligible for laser treatment (MPS criteria) or who refused to have laser photocoagulation were enrolled in the study. Each patient was treated with radiation at incremental dosages from 20 Gy to 40 Gy. After completion of the radiation course, all patients were followed-up at 3 and 7 weeks and 3, 6, and 12 months. Best-corrected VA (ETDRS), slit-lamp and fluorescein angiographic evaluations were performed at each visit. 94 eyes of 89 patients were treated from October 1997 to April 2000. The VA was 0.82+/-0.35 before treatment, 0.83+/-0.36 at 6 months, and 0.89+/-0.33 at 12 months. No patients suffered any significant acute side effects. No significant benefits in either VA or in membrane size were derived from increasing the doses of radiation. Our results are consistent with trends of a palliative benefit of radiotherapy in neovascular AMD and support further investigation of radiotherapy. Since there is no evidence that therapeutic effectiveness is dose dependent, our data provide no justification for potentially dangerous escalations in radiation dosage for treating neovascular AMD.


Infratentorial traumatic brain hemorrhage: may outcome be predicted by initial GCS?
Overall prognosis of nonpenetrating traumatic brain hemorrhage may be predicted by neurologic function days after insult. The relationship between immediate function and outcome has not been examined for infratentorial traumatic brain hemmorhage (iTBH) identified on computed tomographic (CT) scan. Given the severity of brain stem injury, it is conceivable that immediate function may be predictive. A retrospective review of 1,500 brain injuries occurring at our institution identified 18 patients (1.2%) with iTBH on CT scan (eight brain stem, five cerebellum, five both). Demographics, supratentorial injuries, and outcome at 6 months (Glasgow Outcome Scale) were recorded. Initial Glasgow Coma Scale (GCS) in 11 patients was less than 5 (group I). Seven patients had GCS scores greater than or equal to 6 (group II). Nine patients in group I either died or were vegetative. In group II, one died; none were vegetative (p < 0.02). Regression analysis demonstrated a strong correlation between initial GCS and Glasgow Outcome Scale scores at 6 months for all patients (p < 0.001). We conclude that initial GCS score may be predictive of long-term outcome in patients with CT scan evidence of iTBH-a relationship to be explored further for prognostic information.


The First Heartbeat-Origin of Cardiac Contractile Activity.
The amniote embryonic heart starts as a crescent of mesoderm that transitions through a midline linear heart tube in the course of developing into the four chambered heart. It is unusual in having to contract rhythmically while still undergoing extensive morphogenetic remodeling. Advances in imaging have allowed us to determine when during development this contractile activity starts. In the mouse, focal regions of contractions can be detected as early as the cardiac crescent stage. Calcium transients, required to trigger contraction, can be detected even earlier, prior to contraction. In this review, we outline what is currently known about how this early contractile function is initiated and the impact early contractile function has on cardiac development.


The effects of land transport on animal welfare.
Animal welfare during and as a result of transport can be assessed by using a range of behavioural, physiological, pathological and carcass-quality indicators that are described in this paper. Measures of the extent of any disease, injury or mortality resulting from, or exacerbated by, transport are important because health is an important part of welfare. Many of the indicators are measures of stress as they involve long-term adverse effects on the individual. Factors affecting the welfare of animals before, during and after transport which are discussed are: definition of the responsibilities and competence, attitudes to animals and need for training of staff; planning of journeys and methods of payment of staff; laws and retailers' codes; genetics, especially selection for high productivity; rearing conditions and experience; the mixing of animals from different social groups; handling and loading procedures; driving methods; space allowance; increased susceptibility to disease and efforts to minimise the spread of disease.


[Restoring medicine as a scientific and humanist profession].
There is an urgent need, in our society, to recover the real meaning of medicine as a complex of science and humanistic values committed to person's health care. Following the initiative of the American Board of Internal Medicine, the American College of Physicians and the American Society of Internal Medicine (ACP-ASIM) and the European Federation of Internal Medicine, the Medical Professionalism Project has launched a comprehensive declaration to rephrase the social contract between medicine and the society, emphasising the principles and responsibilities that must orient the thoughts and actions of the good physician. The importance, soundness and opportunity of this chart, stimulates an ample dissemination of these concepts and to incorporate them as moral assets, integrating the quality, as experts on a scientific discipline, with the humanistic values provided in these era of increasing bioethical demands.


Temporal changes in bone composition, architecture, and strength following estrogen deficiency in osteoporosis.
Using an ovariectomized (OVX) ovine model, we provide an analysis of the timing of changes in bone following estrogen deficiency. The expression of genes known to regulate osteoclastogenesis, matrix production, and mineralization, as measured by real-time RT-PCR, was significantly increased by 12 months; and increased expression was maintained through to 31 months post-OVX compared to controls. FTIR spectroscopy confirmed that mineralized crystals were less mature than in controls 12 months post-OVX and were even less so by 31 months. The mineral-to-matrix ratio was significantly reduced by 31 months, while the ratio of mature to immature collagen cross-linking was initially increased at 12 months and subsequently reduced at 31 months post-OVX. In contrast, trabecular number, thickness, and separation were unchanged at 12 months. Significant reductions in trabecular number and thickness and a significant increase in trabecular separation were observed 31 months after OVX. Most notably perhaps these combined changes led to a significant reduction in the compressive strength of trabecular bone after 31 months. The results indicate that there is an initial increase in bone turnover, which is accompanied by a change in bone composition. This is followed by a continued increase in bone resorption and relative reduction in bone formation, leading to deterioration in bone microarchitecture. Ultimately, these cumulative changes led to a significant reduction in the compressive strength of bones following 31 months of estrogen deficiency. These findings provide important insight into the time sequence of changes during osteoporosis.


Emergency department crowding and thrombolysis delays in acute myocardial infarction.
We estimate the effect of emergency department (ED) crowding on door-to-needle time for patients given intravenous thrombolysis for suspected acute myocardial infarction. This was a retrospective observational study of patients thrombolyzed in the ED for suspected acute myocardial infarction in 1998 to 2000 in 25 community and teaching hospital EDs in Ontario. EDs located close together and sharing a common ambulance diversion system were grouped into networks consisting of 2 to 5 hospitals each. At patient registration in an ED, the ambulance diversion status of all EDs in the network was determined. Network crowding was calculated as the percentage of EDs that were diverting ambulances on patient registration, categorized as none (0%), moderate (<60%), and high (> or =60%). Door-to-needle time was defined as time from ED registration to drug administration. Multivariable quantile regression and logistic regression were carried out; covariates included age, sex, ECG characteristics, previous acute myocardial infarction, vital signs, time of presentation, and hospital type. A total of 3,452 thrombolysis patients were included: mean age was 62.9 years, and 73% were male patients. Overall median door-to-needle time was 43 minutes (interquartile ratio 27 to 80). Median door-to-needle time was 40, 45, and 47 minutes in conditions of none, moderate, and high network crowding, respectively ( P <.001). The adjusted odds ratios for door-to-needle time delay (>30 minutes) and major delay (>60 minutes) were 1.32 (95% confidence interval [CI] 0.98 to 1.79) and 1.40 (95% CI 1.12 to 1.75), respectively, for high network crowding compared with none, and 1.21 (95% CI 0.89 to 1.63) and 1.06 (95% CI 0.86 to 1.29), respectively, for moderate crowding compared with none. In multivariate analyses, moderate and high crowding conditions were associated with increased median door-to-needle time (3.0 minutes [95% CI 0.1 to 6.0] and 5.8 minutes [95% CI 2.7 to 9.0], respectively). ED crowding is associated with increased door-to-needle times for patients with suspected acute myocardial infarction and may represent a barrier to improving cardiac care in EDs.


Predictors of Recurrence, and Progression-Free and Overall Survival following Open versus Robotic Radical Cystectomy: Analysis from the RAZOR Trial with a 3-Year Followup.
The RAZOR (Randomized Open versus Robotic Cystectomy) trial revealed noninferior 2-year progression-free survival for robotic radical cystectomy. This update was performed with extended followup for 3 years to determine potential differences between the approaches. We also report 3-year overall survival and sought to identify factors predicting recurrence, and progression-free and overall survival. We analyzed the per protocol population of 302 patients from the RAZOR study. Cumulative recurrence was estimated using nonbladder cancer death as the competing risk event and the Gray test was applied to assess significance in differences. Progression-free survival and overall survival were estimated by the Kaplan-Meier method and compared with the log rank test. Predictors of outcomes were determined by Cox proportional hazard analysis. Estimated progression-free survival at 36 months was 68.4% (95% CI 60.1-75.3) and 65.4% (95% CI 56.8-72.7) in the robotic and open groups, respectively (p=0.600). At 36 months overall survival was 73.9% (95% CI 65.5-80.5) and 68.5% (95% CI 59.8-75.7) in the robotic and open groups, respectively (p=0.334). There was no significant difference in the cumulative incidence rates of recurrence (p=0.802). Patient age greater than 70 years, poor performance status and major complications were significant predictors of 36-month progression-free survival. Stage and positive margins were significant predictors of recurrence, and progression-free and overall survival. Surgical approach was not a significant predictor of any outcome. This analysis showed no difference in recurrence, 3-year progression-free survival or 3-year overall survival for robotic vs open radical cystectomy. It provides important prospective data on the oncologic efficacy of robotic radical cystectomy and high level data for patient counseling.


Effect of consumption of fatty acids, calcium, vitamin D and boron with regular physical activity on bone mechanical properties and corresponding metabolic hormones in rats.
The consumption of fatty acids, nutrients, and regular physical activity, individually influence bone mechanical properties in rats. To investigate their effects in combination, male rats were divided into the seven groups: G1: regular food and drinking water; G2: same as Gr.1 + physical activity (Whole body vibration; WBV); G3: same as Gr.2 + Calcium, Vit. D, Boron; G4: same as Gr.3 + canola oil; G5: same as Gr.3 + sunflower oil; G6: same as Gr.3 + mix of sunflower oil and canola oil; and G7: same as Gr.3 + coconut oil; and treated for 8 weeks. Analysis between the control with the groups 2 and 3 revealed that vibration in the G2 increased the body weight (P = 0.04), with no other major difference in plasma and bone indices. Comparison between the control with the G4-G7 (the oil groups) revealed that the rats in the G5 had a lower body weight (15 % less) and a significant increase in plasma levels of Estradiol in the G7 was noted. In addition, levels of Testosterone in the G4 and G7, and Free Testosterone in the G7 had a remarkable increase. Similar trend was observed for plasma levels of Vit. D in the G4 and G5. The stiffness and the breaking strength of the femur in the G7, and the breaking strength of the lumbar in the G7 compared to the control and the G4 and G5 was significantly higher and tended to increase in comparison to the G6. Better and stronger measurements observed for coconut oil is warranted to further study its effect on biomechanical properties of bones.


Cough in the Athlete: CHEST Guideline and Expert Panel Report.
Cough is a common symptom experienced by athletes, particularly after exercise. We performed a systematic review to assess the following in this population: (1) the main causes of acute and recurrent cough, either exercise-induced or not, (2) how cough is assessed, and (3) how cough is treated in this population. From the systematic review, suggestions for management were developed. This review was performed according to the CHEST methodological guidelines and Grading of Recommendations Assessment, Development and Evaluation framework until April 2015. To be included, studies had to meet the following criteria: participants had to be athletes and adults and adolescents aged ≥ 12 years and had to complain of cough, regardless of its duration or relationship to exercise. The Expert Cough Panel based their suggestions on the data extracted from the review and final grading by consensus according to a Delphi process. Only 60 reports fulfilled the inclusion criteria, and the results of our analysis revealed only low-quality evidence on the causes of cough and how to assess and treat cough specifically in athletes. Although there was no formal evaluation of causes of cough in the athletic population, the most common causes reported were asthma, exercise-induced bronchoconstriction, respiratory tract infection (RTI), upper airway cough syndrome (UACS) (mostly from rhinitis), and environmental exposures. Cough was also reported to be related to exercise-induced vocal cord dysfunction among a variety of less common causes. Although gastroesophageal reflux disease (GERD) is frequent in athletes, we found no publication on cough and GERD in this population. Assessment of the causes of cough was performed mainly with bronchoprovocation tests and suspected disease-specific investigations. The evidence to guide treatment of cough in the athlete was weak or nonexistent, depending on the cause. As data on cough in athletes were hidden in a set of other data (respiratory symptoms), evidence tables were difficult to produce and were done only for cough treatment in athletes. The causes of cough in the athlete appear to differ slightly from those in the general population. It is often associated with environmental exposures related to the sport training environment and occurs predominantly following intense exercise. Clinical history and specific investigations should allow identification of the cause of cough as well as targeting of the treatment. Until management studies have been performed in the athlete, current guidelines that exist for the general population should be applied for the evaluation and treatment of cough in the athlete, taking into account specific training context and anti-doping regulations.


Supervised lifestyle intervention for people with metabolic syndrome improves outcomes and reduces individual risk factors of metabolic syndrome: A systematic review and meta-analysis.
Metabolic syndrome is characterised by a clustering of metabolic risk factors including abdominal obesity, raised triglycerides, lowered HDL cholesterol, hypertension and impaired glucose tolerance. Multifaceted lifestyle interventions including diet and exercise are recommended as the first-line treatment for the metabolic syndrome. To investigate the effects of lifestyle interventions that include both diet interventions and supervised exercise on outcomes for people with metabolic syndrome. A systematic review and meta-regression was conducted. PubMed, EMBASE, MEDLINE and CINAHL were searched from the earliest date possible until November 2018 to identify randomised controlled trials examining the effects of lifestyle interventions compared to usual care on patient health outcomes and components of metabolic syndrome. Post-intervention means and standard deviations were pooled using inverse variance methods and random-effects models to calculate mean differences (MD), standardised mean differences (SMD) and 95% confidence intervals (CI). Searching identified 2598 articles, of which 15 articles reporting data from 10 trials, with 1160 participants were included in this review. Compared to usual care, supervised lifestyle intervention demonstrated significant improvements in all but one of the components of metabolic syndrome. Reductions were seen in waist circumference (-4.9 cm, 95%CI -8.0 to -1.7), systolic blood pressure (-6.5 mmHg, 95%CI -10.7 to -2.3), diastolic blood pressure (-1.9 mmHg, 95%CI -3.6 to -0.2), triglycerides (SMD -0.46, 95%CI -0.88 to -0.04) and fasting glucose (SMD -0.68, 95%CI -1.20 to -0.15). Prevalence of metabolic syndrome was reduced by 39% in intervention group participants compared to control group participants (Risk Ratio 0.61, 95%CI 0.38 to 0.96). Improvements in quality of life were not statistically significant. There is low to moderate quality evidence that supervised multifaceted lifestyle intervention improves multiple risk factors of metabolic syndrome, as well as reducing prevalence of the disease. Health services should consider implementing lifestyle intervention programs for people with metabolic syndrome to improve health outcomes and prevent progression to chronic disease.


The metabolic syndrome and its components in 178 patients treated for craniopharyngioma after 16 years of follow-up.
Patients with craniopharyngioma are at an increased risk for cardio- and cerebrovascular mortality. The metabolic syndrome (MetS) is an important cardiometabolic risk factor, but barely studied in patients with craniopharyngioma. We aimed to investigate the prevalence of and risk factors for the MetS and its components in patients with craniopharyngioma. Cross-sectional study with retrospective data. We studied the prevalence of and risk factors for the MetS and its components in 110 Dutch (median age 47 years, range 18-92) and 68 Swedish (median age 50 years, range 20-81) patients with craniopharyngioma with ≥3 years of follow-up (90 females (51%); 83 patients with childhood-onset craniopharyngioma (47%); median follow-up after craniopharyngioma diagnosis 16 years (range 3-62)). In Dutch patients aged 30-70 years and Swedish patients aged 45-69 years, we examined the prevalence of the MetS and its components relative to the general population. Sixty-nine (46%) of 149 patients with complete data demonstrated the MetS. Prevalence of the MetS was significantly higher in patients with craniopharyngioma compared with the general population (40% vs 26% (P < 0.05) for Dutch patients; 52% vs 15% (P < 0.05) for Swedish patients). Multivariable logistic regression analysis identified visual impairment as a borderline significant predictor of the MetS (OR 2.54, 95% CI 0.95-6.81; P = 0.06) after adjustment for glucocorticoid replacement therapy and follow-up duration. Age, female sex, tumor location, radiological hypothalamic damage, 90Yttrium brachytherapy, glucocorticoid replacement therapy and follow-up duration significantly predicted components of the MetS. Patients with craniopharyngioma are at an increased risk for the MetS, especially patients with visual impairment.


Definition and diagnosis of vertebral fracture.
The majority of vertebral deformities are asymptomatic and thus definition is normally based on radiographic appearance. There are no internationally agreed criteria for vertebral fracture based on radiographic appearance. A variety of morphometric methods have been developed to ascertain changes in vertebral shape based on a reduction in vertebral height. Criteria have to be established to determine what cutoff constitutes abnormality. An alternative is to adopt a semiquantitative grading using clinical observations of a experienced radiologist. This approach enhances specificity by excluding nonosteoporotic causes of changing vertebral shape. Such methods, however, may be subject to difficulties in standardization.


Estimation of oxygen saturation from erythrocytes by high-resolution spectroscopic optical coherence tomography.
We demonstrate the feasibility of estimating hemoglobin oxygen saturation (SaO(2)) from erythrocytes over a visible wavelength band using high-resolution spectroscopic optical coherence tomography (SOCT). We take SOCT spectra from individual cells on in vitro human blood phantoms and calculate SaO(2) by fitting the spectra to an analytical model. We also use diffusive spectroscopy (DS) measurements, which is validated by direct measurement of oxygen partial pressure, as a comparison. The result shows a reliable linear association between the SaO(2) derived from SOCT and the DS measurement, indicating this methodology has potential on in vivo applications to evaluate blood oxygenation.


Simian immunodeficiency virus-infected macaques treated with highly active antiretroviral therapy have reduced central nervous system viral replication and inflammation but persistence of viral DNA.
During the era of highly active antiretroviral therapy (HAART), the prevalence of HIV-associated central nervous system (CNS) disease has increased despite suppression of plasma viremia. In a simian immunodeficiency virus (SIV) model system in which all animals develop AIDS and 90% develop CNS disease by 3 months after inoculation, pigtailed macaques were treated with a regimen of tenofovir disoproxil fumarate, saquinavir, atazanavir, and an integrase inhibitor starting at 12 days after inoculation and were euthanized at approximately 175 days after inoculation. Plasma and cerebrospinal fluid (CSF) viral loads declined rapidly after the initiation of HAART. Brain viral RNA was undetectable at necropsy, but viral DNA levels were not different from those in untreated SIV-infected macaques. CNS inflammation was significantly reduced, with decreased brain expression of major histocompatibility complex class II and glial fibrillary acidic protein and reduced levels of CSF CCL2 and interleukin 6. Brain from treated macaques had significantly lower levels of interferon beta, type 1 interferon-inducible gene myxovirus (influenza) resistance A, and indolamine 2,3-dioxygenase messenger RNA, suggesting that immune hyperactivation was suppressed, and fewer CD4(+) and CD8(+) T cells, suggesting that trafficking of T cells from peripheral blood was reduced. Brain levels of CD68 protein and tumor necrosis factor alpha and interferon gamma RNA were reduced but were not significantly lower, indicating continued CNS inflammation. These data, generated in a rigorous, high-viral-load SIV-infected macaque model, showed that HAART provided benefits with respect to CNS viral replication and inflammation but that no change in the level of viral DNA and continued CNS inflammation occurred in some macaques.


Negative regulation of yeast WASp by two SH3 domain-containing proteins.
WASp family proteins promote actin filament assembly by activating Arp2/3 complex and are regulated spatially and temporally to assemble specialized actin structures used in diverse cellular processes. Some WASp family members are autoinhibited until bound by activating ligands; however, regulation of the budding yeast WASp homolog (Las17/Bee1) has not yet been explored. We isolated full-length Las17 and characterized its biochemical activities on yeast Arp2/3 complex. Purified Las17 was not autoinhibited; in this respect, it is more similar to SCAR/WAVE than to WASp proteins. Las17 was a much stronger activator of Arp2/3 complex than its carboxyl-terminal (WA) fragment. In addition, actin polymerization stimulated by Las17-Arp2/3 was much less sensitive to the inhibitory effects of profilin compared to polymerization stimulated by WA-Arp2/3. Two SH3 domain-containing binding partners of Las17, Sla1 and Bbc1, were purified and were shown to cooperate in inhibiting Las17 activity. The two SLA1 SH3 domains required for this inhibitory activity in vitro were also required in vivo, in combination with BBC1, for cell viability and normal actin organization. Full-length Las17 is not autoinhibited and activates Arp2/3 complex more strongly than its WA domain alone, revealing an important role for the Las17 amino terminus in Arp2/3 complex activation. Two of the SH3 domain-containing ligands of Las17, Sla1 and Bbc1, cooperate to inhibit Las17 activity in vitro and are required for a shared function in actin organization in vivo. Our results show that, like SCAR/WAVE, WASp proteins can be controlled by negative regulation through the combined actions of multiple ligands.


Cyanine dye mediated mitochondrial targeting enhances the anti-cancer activity of small-molecule cargoes.
Organelle-specific delivery systems are of significant clinical interest. We demonstrate the use of common cyanine dyes Cy3 and Cy5 as vectors for targeting and delivering cargoes to mitochondria in cancer cells. Specifically, conjugation to the dyes can increase cytotoxicity by up to 1000-fold.


Slx1-Slx4 are subunits of a structure-specific endonuclease that maintains ribosomal DNA in fission yeast.
In most eukaryotes, genes encoding ribosomal RNAs (rDNA) are clustered in long tandem head-to-tail repeats. Studies of Saccharomyces cerevisiae have indicated that rDNA copy number is maintained through recombination events associated with site-specific blockage of replication forks (RFs). Here, we describe two Schizosaccharomyces pombe proteins, homologs of S. cerevisiae Slx1 and Slx4, as subunits of a novel type of endonuclease that maintains rDNA copy number. The Slx1-Slx4-dependent endonuclease introduces single-strand cuts in duplex DNA on the 3' side of junctions with single-strand DNA. Deletion of Slx1 or Rqh1 RecQ-like DNA helicase provokes rDNA contraction, whereas simultaneous elimination of Slx1-Slx4 endonuclease and Rqh1 is lethal. Slx1 associates with chromatin at two foci characteristic of the two rDNA repeat loci in S. pombe. We propose a model in which the Slx1-Slx4 complex is involved in the control of the expansion and contraction of the rDNA loci by initiating recombination events at stalled RFs.


Clinical significance of occult hepatitis B infection in progression of liver disease and carcinogenesis.
Occult hepatitis B infection (OBI) is defined as long-lasting persistence of hepatitis B virus (HBV) DNA in the liver of patients with hepatitis B surface antigen (HBsAg)-negative status, with or without serological markers of previous exposure (antibodies to HBsAg and/or to hepatitis B core antigen). Over the past two decades, significant progress has been made in understanding OBI and its clinical implications. OBI as a cause of chronic liver disease in patients with HBsAg-negative status is becoming an important disease entity. In conditions of immunocompetence, OBI is inoffensive in itself and detection of HBV DNA in the liver does not always indicate active hepatitis. However, when other factors that cause liver damage, such as hepatitis C virus infection, obesity and alcohol abuse are present, the minimal lesions produced by the immunological response to OBI might worsen the clinical course of the underlying liver disease. Several lines of evidence suggest that OBI is associated with progression of liver fibrosis and the development of hepatocellular carcinoma in patients with chronic liver disease. The major interest in OBI is primarily associated with the growing, widely discussed evidence of its clinical impact. The aim of this review is to highlight recent data for OBI, with a major focus on disease progression or carcinogenesis in patients with chronic liver disease.


Disrupted spermine homeostasis: a novel mechanism in polyglutamine-mediated aggregation and cell death.
Our data suggest a novel mechanism whereby pathological-length polyglutamine (polyQ) proteins promote the spermine synthetic pathway, increasing polyQ-aggregation and cell death. As detected in a cell-free turbidity assay, spermine promotes aggregation of thio-polyQ62 in a dose-dependent manner. Using a stable neuronal cell line expressing pathological-length [polyQ57-yellow fluorescent protein (YFP) (Q57)] or non-pathological-length [polyQ19-YFP (Q19)] polyglutamine protein, we show that multiple steps in the production of polyamines are affected in Q57 cells, suggesting dysfunctional spermine homeostasis. As the building block for spermine synthesis, arginine transport is significantly increased in neuronal cell lines stably expressing Q57. Q57 lines displayed upregulated basal and inducible arginase I activities that were not seen in polyQ19-YFP lines. Normal induction of spermidine/spermine N-acetyltransferase in Q19 lines regulating back-conversion of spermine, thereby reducing spermine levels, however, was not observed in Q57 lines. Pharmacological activation of ornithine decarboxylase (ODC), a key enzyme of the polyamine synthetic pathway, increased cellular aggregates and increased cell death in Q57 cells not observed in Q19 cells. Inhibition of ODC by difluoromethylornithine prevented basal and induced cell death in Q57 cells, demonstrating a central role for polyamines in this process.


High-resolution image reconstruction in fluorescence microscopy with patterned excitation.
We discuss data treatment strategies in structured illumination microscopy, using simulated and experimental data. In the setup, the illumination pattern is generated by projecting a movable pinhole mask into the sample, and a wide-field fluorescence microscope image is acquired for each pattern position. The structured illumination data obtained from a two-dimensional illumination pattern can be treated by projection strategies such as in video confocal microscopy (sum, maximum, maximum minus minimum, and superconfocal), by a scaled subtraction of the out-of-focus estimate, or by a modified version of the Fourier-space treatment, as is known for data from one-dimensional structured illumination. We investigate the influence of some data processing strategies on unwanted effects such as residual patterning and local deviations from linearity in the reconstructed intensity.


An ex-ovo chicken embryo culture system suitable for imaging and microsurgery applications.
Understanding the relationships between genetic and microenvironmental factors that drive normal and malformed embryonic development is fundamental for discovering new therapeutic strategies. Advancements in imaging technology have enabled quantitative investigation of the organization and maturing of the body plan, but later stage embryonic morphogenesis is less clear. Chicken embryos are an attractive vertebrate animal model system for this application because of its ease of culture and surgical manipulation. Early embryos can be cultured for a short time on filter paper rings, which enables complete optical access for cell patterning and fate studies. Studying advanced developmental processes such as cardiac morphogenesis are traditionally performed through a window of the eggshell, but this technique limits optical access due to window size. We previously developed a simple method to culture whole embryos ex-ovo on hexagonal weigh boats for up to 10 days, which enabled high resolution imaging via ultrasonography. These cultures were difficult to transport, limiting the types of imaging tools available for live experiments. We here present an improved shell-less culture system with a cost-effective, portable environmental chamber. Eggs were cracked onto a hammock created by a polyurethane membrane (cling wrap) affixed circumferentially to a plastic cup partially filled with sterile water. The dimensions of the circumference and depth of the hammock were both critical to maintain surface tension, while the mechanics of the hammock and water beneath helped dampen vibrations induced by transportation. A small footprint circulating water bath was also developed to enable continuous temperature control during experimentation. We demonstrate the ability to culture embryos in this way for at least 14 days without morphogenic defect or delay and employ this system in several microsurgical and imaging applications.


Juvenile offences among hospitalized adolescent inhalant users in Istanbul: a comparison regarding place of residence.
In this research, juvenile offenses and associated behaviors among adolescent inhalant users in Istanbul were investigated and inhalant users living in the street were compared to inhalant users living with their families. An interview questionnaire developed by the researchers was administered to 200 male adolescent inhalant users who were hospitalized during 2002-2003. More than half of the sample had committed juvenile offenses at least once in their lifetime, 16.3% had entered a house of corrections, 91.5% had friends who committed juvenile offenses, and the majority had been taken to a police station at least once in their lifetime. The rates for juvenile offenses, being taken to the police station, committing crimes to obtain money to buy drugs, and obtaining income through illegal activities were higher among adolescents living in the street than adolescents living with their families. Although the juvenile offense rate was higher among adolescents living in the street, it can be suggested that both groups live in subcultures that have a tendency towards crime, and inhalant use is part of these subcultures. Juvenile offense interventions can be useful for all inhalant users.