Potential impurities in drug substances: Compound-specific toxicology limits for 20 synthetic reagents and by-products, and a class-specific toxicology limit for alkyl bromides.
This paper provides compound-specific toxicology limits for 20 widely used synthetic reagents and common by-products that are potential impurities in drug substances. In addition, a 15 μg/day class-specific limit was developed for monofunctional alkyl bromides, aligning this with the class-specific limit previously defined for monofunctional alkyl chlorides. Both the compound- and class-specific toxicology limits assume a lifetime chronic exposure for the general population (including sensitive subpopulations) by all routes of exposure for pharmaceuticals. Inhalation-specific toxicology limits were also derived for acrolein, formaldehyde, and methyl bromide because of their localized toxicity via that route. Mode of action was an important consideration for a compound-specific toxicology limit. Acceptable intake (AI) calculations for certain mutagenic carcinogens assumed a linear dose-response for tumor induction, and permissible daily exposure (PDE) determination assumed a non-linear dose-response. Several compounds evaluated have been previously incorrectly assumed to be mutagenic, or to be mutagenic carcinogens, but the evidence reported here for such compounds indicates a lack of mutagenicity, and a non-mutagenic mode of action for tumor induction. For non-mutagens with insufficient data to develop a toxicology limit, the ICH Q3A qualification thresholds are recommended. The compound- and class-specific toxicology limits described here may be adjusted for an individual drug substance based on treatment duration, dosing schedule, severity of the disease and therapeutic indication.


IL-6 regulates adipose deposition and homeostasis in lymphedema.
Lymphedema (LE) is a morbid disease characterized by chronic limb swelling and adipose deposition. Although it is clear that lymphatic injury is necessary for this pathology, the mechanisms that underlie lymphedema remain unknown. IL-6 is a known regulator of adipose homeostasis in obesity and has been shown to be increased in primary and secondary models of lymphedema. Therefore, the purpose of this study was to determine the role of IL-6 in adipose deposition in lymphedema. The expression of IL-6 was analyzed in clinical tissue specimens and serum from patients with or without LE, as well as in two mouse models of lymphatic injury. In addition, we analyzed IL-6 expression/adipose deposition in mice deficient in CD4(+) cells (CD4KO) or IL-6 expression (IL-6KO) or mice treated with a small molecule inhibitor of IL-6 or CD4 depleting antibodies to determine how IL-6 expression is regulated and the effect of changes in IL-6 expression on adipose deposition after lymphatic injury. Patients with LE and mice treated with lymphatic excision of the tail had significantly elevated tissue and serum expression of IL-6 and its downstream mediator. The expression of IL-6 was associated with adipose deposition and CD4(+) inflammation and was markedly decreased in CD4KO mice. Loss of IL-6 function resulted in significantly increased adipose deposition after tail lymphatic injury. Our findings suggest that IL-6 is increased as a result of adipose deposition and CD4(+) cell inflammation in lymphedema. In addition, our study suggests that IL-6 expression in lymphedema acts to limit adipose accumulation.


Long-term efficacy and safety of etanercept in children with polyarticular-course juvenile rheumatoid arthritis: interim results from an ongoing multicenter, open-label, extended-treatment trial.
To evaluate the long-term efficacy and safety of etanercept in children with juvenile rheumatoid arthritis (JRA) participating in an ongoing multicenter, open-label, extended-treatment trial. All patients had been participants in an initial randomized efficacy and safety trial of etanercept. Etanercept was administered at a dosage of 0.4 mg/kg (maximum 25 mg) subcutaneously twice each week. Safety and efficacy evaluations were performed every 3-4 months. The JRA 30% definition of improvement (DOI) was defined as improvement of > or =30% in at least 3 of 6 response variables used to assess disease activity, with no more than 1 variable worsening by more than 30%. At the time of analysis, 48 of the 58 patients (83%) were still enrolled in the study; 43 of them (74%) had completed 2 years of treatment. Of these 43 patients, 81% met the JRA 30% DOI, 79% met the JRA 50% DOI, and 67% met the JRA 70% DOI. Ten children started low-dose methotrexate after year 1. Of the 32 children taking prednisone, the dosage was decreased to <5 mg/day in 26 (81%). Two children had serious infections (varicella with aseptic meningitis in one and complicated sepsis in the other). In general, adverse events were of the types seen in a general pediatric patient population. Children with severe, longstanding, methotrexate-resistant polyarticular JRA demonstrated sustained clinical improvement with >2 years of continuous etanercept treatment. Etanercept was generally well-tolerated. There were no increases in the rates of adverse events over time. However, children taking etanercept should be monitored closely for infections.


Trisomy 18 in a canine thyroid adenoma.
A canine thyroid adenoma showing trisomy 18 as the sole clonal cytogenetic abnormality (9 of 30 analyzed metaphases) is reported. Because trisomies are a recurrent cytogenetic finding in human benign thyroid tumors as well, it is suggested that the molecular relationship between these trisomies and the development of thyroid tumors can be determined by comparative gene mapping.


Effect of protein-modifying reagents on ecto-apyrase from rat brain.
We have tested several chemical modifiers to investigate which amino acid residues, present in the primary structure of the ecto-apyrase, could be involved in catalysis. Synaptosomes from cerebral cortex of rats were prepared and the ATP diphosphohydrolase activity was assayed in absence or the presence of the modifiers. Percentages of residual activity for ATPase and ADPase obtained when the following reagents were tested, are respectively: phenylglyoxal (an arginine group modifier) 17 and 30%; Woodward's reagent (a carboxylic group modifier) 33 and 23%; Koshland's reagent (a tryptophan group modifier) 10 and 12%; maleic anhidride (an amino group modifier) 11 and 25% and carbodiimide reagent (a carboxylic group modifier) 56 and 72%. Otherwise, PMSF, a seryl protein modifier and DTNB, a SH-group modifier did not affect either ATPase or ADPase activity. Inhibitions observed after treatment with phenylglyoxal and Woodward's reagent were significantly prevented when the synaptosomal fraction was preincubated with ATP and ADP, indicating that the arginine and the side chain of glutamate or aspartate (carboxyl groups) participate in the structure of the active site. This interpretation was confirmed by using GTP and GDP, two other apyrase substrates. Phenylglyoxal and Woodward's reagent also inhibited the GTPase and GDPase activities and this inhibition was prevented by preincubation with these substrates.


Central ghrelin signaling mediates the metabolic response of C57BL/6 male mice to chronic social defeat stress.
Chronic stressors promote metabolic disturbances, including obesity and metabolic syndrome. Ghrelin, a peptide that promotes appetite and the accumulation of adipose tissue, is also secreted in response to stressors to protect the brain and peripheral tissues from the effects of these stressors. Here we demonstrate that elevated ghrelin levels produced by chronic exposure to social stress are associated with increased caloric intake and body weight gain in male C57BL mice. In contrast, stressed mice lacking ghrelin receptors (GHSR KO mice) or C57BL mice receiving chronic intracerebroventricular delivery of the ghrelin receptor antagonist [d-Lys(3)]-GHRP-6 show attenuated weight gain and feeding responses under the same social stress paradigm. Interestingly, stressed GHSR KO mice showed depleted sc and intrascapular brown fat depots, whereas stressed young wild-type mice did not. In old wild-type mice, chronic social defeat increased visceral and intrascapular brown fat depots in association with increases in obesity markers like hyperleptinemia and hyperinsulinemia along with increased hypothalamic expression of neuropeptide Y and Agouti related peptide. Importantly, the elevated expression of these peptides persisted least for 2 weeks after cessation of the stressor regimen. In contrast, old GHSR KO mice did not show these alterations after chronic social defeat. These results suggest that ghrelin plays an important role in the metabolic adaptations necessary to meet the energetic demands posed by stressors, but chronic exposure to stress-induced ghrelin elevations ultimately could lead to long lasting metabolic dysfunctions.


Increased matrix metalloproteinase expression and activation following experimental acute pancreatitis.
The observation that matrix metalloproteinases (MMPs) are central to tissue remodeling and may contribute to organ failure prompted us to investigate the role of MMPs in acute pancreatitis. We hypothesize that increased expression and activation of MMP-2 and MMP-9 will correlate with organ injury following acute pancreatitis. Acute pancreatitis was induced in five male rats by retrograde infusion of 5% sodium taurocholate into the pancreatic duct. Sham laparotomy was performed on five rats serving as a control. Pancreatitis was confirmed by histology and serum amylase levels. MMP-2 and MMP-9 activity and expression were assayed by gelatin zymography in the lungs and ascitic fluid of each animal. Lung permeability was assayed by Evans blue dye extravasation. Lung activity of MMP-2 and MMP-9 was confirmed by a specific fluorogenic MMP substrate assay. Lung permeability increased twofold in the animals with severe pancreatitis compared with sham. Analysis of the zymograms from lung homogenate revealed a threefold increase in active MMP-2 in severe pancreatitis compared with sham and no change in MMP-9 activity. Gelatin zymograms of peritoneal fluid from severe pancreatitis animals demonstrated increased levels of active MMP-2 and MMP-9 compared with the sham group. Increases in MMP activity were confirmed by MMP activity assay using a fluorogenic substrate. This study demonstrates a correlation between severity of acute pancreatitis and active MMP-2 and MMP-9 levels in the peritoneal fluid and MMP-2 activity in lung homogenate. The MMP-mediated degradation of the basement membrane offers a potential pharmacologic and therapeutic target for halting the final biologic outcome of severe pancreatitis.


Ionizing radiation as a phytosanitary treatment against European corn borer (Lepidoptera: Crambidae) in ambient, low oxygen, and cold conditions.
The European corn borer, Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae), is a quarantine pest for several fresh commodities, including corn-on-the-cob, bell peppers, and green beans. Methyl bromide fumigation is the usual phytosanitary treatment, but the fumigant is under increasing regulation as a stratospheric ozone-depleting substance. Ionizing radiation is a relatively new commercial alternative that is currently used in several countries. The present research explored radiation doses that would provide quarantine security for commodities at risk of being infested by O. nubilalis. Radiotolerance of late pupae (the most tolerant stage infesting commodities) as determined by hatch of F1 eggs was not affected by host (meridic diet versus ear corn) or temperature (1 versus 13 degrees C) but was positively affected by low oxygen. Longevity was shorter for adults of irradiated than nonirradiated pupae. The minimum absorbed dose for phytosanitary irradiation against O. nubilalis could vary from 233 Gy for prevention of F1 pupation to 343 Gy for prevention of F1 egg hatch. Lower doses might be possible if greater risk of treatment failure was acceptable.


Correlation of KIT and PDGFRA mutational status with clinical benefit in patients with gastrointestinal stromal tumor treated with sunitinib in a worldwide treatment-use trial.
Several small studies indicated that the genotype of KIT or platelet-derived growth factor receptor-α (PDGFRA) contributes in part to the level of clinical effectiveness of sunitinib in gastrointestinal stromal tumor (GIST) patients. This study aimed to correlate KIT and PDGFRA mutational status with clinical outcome metrics (progression-free survival [PFS], overall survival [OS], objective response rate [ORR]) in a larger international patient population. This is a non-interventional, retrospective analysis in patients with imatinib-resistant or intolerant GIST who were treated in a worldwide, open-label treatment-use study (Study 1036; NCT00094029) in which sunitinib was administered at a starting dose of 50 mg/day on a 4-week-on, 2-week-off schedule. Molecular status was obtained in local laboratories with tumor samples obtained either pre-imatinib, post-imatinib/pre-sunitinib, or post-sunitinib treatment, and all available data were used in the analyses regardless of collection time. The primary analysis compared PFS in patients with primary KIT exon 11 versus exon 9 mutations (using a 2-sided log-rank test) and secondary analyses compared OS (using the same test) and ORR (using a 2-sided Pearson χ(2) test) in the same molecular subgroups. Of the 1124 sunitinib-treated patients in the treatment-use study, 230 (20%) were included in this analysis, and baseline characteristics were similar between the two study populations. Median PFS was 7.1 months. A significantly better PFS was observed in patients with a primary mutation in KIT exon 9 (n = 42) compared to those with a primary mutation in exon 11 (n = 143; hazard ratio = 0.59; 95 % confidence interval, 0.39-0.89; P = 0.011), with median PFS times of 12.3 and 7.0 months, respectively. Similarly, longer OS and higher ORR were observed in patients with a primary KIT mutation in exon 9 versus exon 11. The data available were limited to investigate the effects of additional KIT or PDGFRA mutations on the efficacy of sunitinib treatment. This large retrospective analysis confirms the prognostic significance of KIT mutation status in patients with GIST. This analysis also confirms the effectiveness of sunitinib as a post-imatinib therapy, regardless of mutational status. NCT01459757.


Quantitative assessment of the subepithelial collagen band does not increase the accuracy of diagnosis of collagenous colitis.
The thickness of eosinophilic band in collagenous colitis (CC) was assessed by 3 methods: histologic estimates (22 observers), conventional measurements using a calibrated micrometric scale (1 observer), and semiautomatic micrometric measurements (1 observer). By the histologic estimate technique, 7.4% of the results failed to diagnose CC; by calibrated micrometry, the failure was 6% and by semiautomatic micrometry, 6%. The main difficulty in measuring the thickness of the CC band is that the deeper border of the band appears fuzzy and hairy-irregular. CC should be defined not exclusively on the basis of the thickness of the collagen table, but as a microscopic constellation characterized by a distorted superficial cell arrangement, with areas of epithelial denudation and inflammatory cells in the superficial epithelium and the lamina propria. In agreement with Lazenby's statement: "Focusing solely on the collagen band can result in both over- and underdiagnosis"


Flavonoids induce apoptosis in human leukemia U937 cells through caspase- and caspase-calpain-dependent pathways.
Flavonoids are polyphenolic phytochemicals that are ubiquitous in plants and present in the common human diet. They may exert diverse beneficial effects, including antioxidant and anticarcinogenic activities. In this study we tested the apoptotic activity of 22 flavonoids and related compounds in leukemic U937 cells. Several flavones but none of the isoflavones or flavanones tested induced apoptotic cell death under these conditions, as determined by reduction in cell viability, flow cytometry, and oligonucleosomal DNA fragmentation. Structure-activity relationship showed that at least two hydroxylations in positions 3, 5, and 7 of the A ring were needed to induce apoptosis, whereas hydroxylation in 3' and/or 4' of the B ring enhanced proapoptotic activity. At lower concentrations, these compounds were also able to sensitize these cells to apoptosis induced by tumor necrosis factor-alpha. Regarding the mechanisms, galangin, luteolin, chrysin, and quercetin induced apoptosis in a way that required the activation of caspases 3 and 8, but not caspase 9. In contrast, an active role of calpains in addition to caspases was demonstrated in apoptosis induced by fisetin, apigenin, and 3,7-dihydroxyflavone. Our data show evidence of the proapoptotic properties of some flavonoids that could support their rational use as chemopreventive and therapeutic agents against carcinogenic disease.


Mitotic chromosome condensation in the sperm nucleus during postfertilization maturation division in Urechis eggs.
Changes in the morphology of the sperm nucleus in the egg cytoplasm are mong the immediate events in nucleocytoplasmic interactions during early embryogenesis. Soon after its entrance into the egg cytoplasm, the sperm nucleus of various organisms increases in size with the transformation of condensed chromatin to a diffuse state, resembling the chromatin of an interphase nucleus (2, 13, 15, 16). This is followed by a close association or fusion of male and female pronuclei (2, 13, 15, 16). Cytoplasmic influences on nuclear morphology have also been demonstrated clearly in nuclear transplantation and cell fusion studies (10, 11). Reactivation of the nucleus, such as the transplanted brain nucleus in Xenopus egg cytoplasm or the hen erythrocyte nucleus in interphase cytoplasm of HeLa cells, is accompanied by nuclear enlargement and chromatin dispersion (10, 11). However, premature mitotic-like chromosome condensation takes place in the nuclei of sperm or interphase cells fused with mitotic cells (9, 12). Thus, chromosome dispersion and condensation seem to depend on the state of the cytoplasm in which the nucleus is present. These observations imply that the initial morphological changes in the sperm nucleus after fertilization may very well be dependent on the state of maturation of eggs at the time of sperm entry. Unfertilized eggs of Urechis caupo, a marine echiuroid worm, are stored at the diakinesis stage. These eggs complete maturation division after insemination and this is followed by fusion of male and female pronuclei (5, 8). Therefore, Urechis caupo is a suitable organism in which to study the response of the sperm nucleus to the changing state of the egg cytoplasm during and after postfertilization maturation division.


Molecular genetic analysis of Down syndrome.
Down syndrome (DS) is caused by trisomy of all or part of human chromosome 21 (HSA21) and is the most common genetic cause of significant intellectual disability. In addition to intellectual disability, many other health problems, such as congenital heart disease, Alzheimer's disease, leukemia, hypotonia, motor disorders, and various physical anomalies occur at an elevated frequency in people with DS. On the other hand, people with DS seem to be at a decreased risk of certain cancers and perhaps of atherosclerosis. There is wide variability in the phenotypes associated with DS. Although ultimately the phenotypes of DS must be due to trisomy of HSA21, the genetic mechanisms by which the phenotypes arise are not understood. The recent recognition that there are many genetically active elements that do not encode proteins makes the situation more complex. Additional complexity may exist due to possible epigenetic changes that may act differently in DS. Numerous mouse models with features reminiscent of those seen in individuals with DS have been produced and studied in some depth, and these have added considerable insight into possible genetic mechanisms behind some of the phenotypes. These mouse models allow experimental approaches, including attempts at therapy, that are not possible in humans. Progress in understanding the genetic mechanisms by which trisomy of HSA21 leads to DS is the subject of this review.


Evaluating the Use of Genetics in Brugada Syndrome Risk Stratification.
The evolution of the current dogma surrounding Brugada syndrome (BrS) has led to a significant debate about the real usefulness of genetic testing in this syndrome. Since BrS is defined by a particular electrocardiogram (ECG) pattern, after ruling out certain possible causes, this disease has come to be defined more for what it is not than for what it is. Extensive research is required to understand the effects of specific individual variants, including modifiers, rather than necessarily grouping together, for example, "all SCN5A variants" when trying to determine genotype-phenotype relationships, because not all variants within a particular gene act similarly. Genetic testing, including whole exome or whole genome testing, and family segregation analysis should always be performed when possible, as this is necessary to advance our understanding of the genetics of this condition. All considered, BrS should no longer be considered a pure autosomal dominant disorder, but an oligogenic condition. Less common patterns of inheritance, such as recessive, X-linked, or mitochondrial may exist. Genetic testing, in our opinion, should not be used for diagnostic purposes. However, variants in SCN5A can have a prognostic value. Patients should be diagnosed and treated per the current guidelines, after an arrhythmologic examination, based on the presence of the specific BrS ECG pattern. The genotype characterization should come in a second stage, particularly in order to guide the familial diagnostic work-up. In families in which an SCN5A pathogenic variant is found, genetic testing could possibly contribute to the prognostic risk stratification.


Role of Microvascular Disruption in Brain Damage from Traumatic Brain Injury.
Traumatic brain injury (TBI) is acquired from an external force, which can inflict devastating effects to the brain vasculature and neighboring neuronal cells. Disruption of vasculature is a primary effect that can lead to a host of secondary injury cascades. The primary effects of TBI are rapidly occurring while secondary effects can be activated at later time points and may be more amenable to targeting. Primary effects of TBI include diffuse axonal shearing, changes in blood-brain barrier (BBB) permeability, and brain contusions. These mechanical events, especially changes to the BBB, can induce calcium perturbations within brain cells producing secondary effects, which include cellular stress, inflammation, and apoptosis. These secondary effects can be potentially targeted to preserve the tissue surviving the initial impact of TBI. In the past, TBI research had focused on neurons without any regard for glial cells and the cerebrovasculature. Now a greater emphasis is being placed on the vasculature and the neurovascular unit following TBI. A paradigm shift in the importance of the vascular response to injury has opened new avenues of drug-treatment strategies for TBI. However, a connection between the vascular response to TBI and the development of chronic disease has yet to be elucidated. Long-term cognitive deficits are common amongst those sustaining severe or multiple mild TBIs. Understanding the mechanisms of cellular responses following TBI is important to prevent the development of neuropsychiatric symptoms. With appropriate intervention following TBI, the vascular network can perhaps be maintained and the cellular repair process possibly improved to aid in the recovery of cellular homeostasis.


Jeavons Syndrome: Clinical Features and Response to Treatment.
Jeavons syndrome is an underreported epileptic syndrome characterized by eyelid myoclonia, eyelid closure-induced seizures or electroencephalography paroxysms, and photosensitivity. Drug-resistant epilepsy is common, but the prognostic factors and clinical course leading to drug resistance have not been well characterized. We identified 30 patients who met the diagnostic criteria of Jeavons syndrome at a single institution between January 1, 2000 and December 15, 2016. Criteria for Jeavons syndrome included all of the following: (1) eyelid myoclonia with or without absences, (2) eye-closure-induced seizures or electroencephalography paroxysms, and (3) seizure onset after 12 months of age. We reviewed and described the epilepsy history, antiepileptic drug trials, and response to treatments. Mean age at seizure onset was 7.3 years, and 80% were female. Absence seizures (63%) and generalized tonic-clonic seizures (23%) were most common at onset. Diagnosis was delayed by an average of 9.6 years. After a median follow-up of two years, 80% of patients had drug resistant epilepsy and 70% experienced generalized tonic-clonic seizures. Generalized tonic-clonic seizures and seizure types other than absence seizures increased the risk of drug-resistant epilepsy (P values 0.049 and 0.03, respectively). Valproic acid, lamotrigine, ethosuximide, and levetiracetam were the most effective in reducing seizures by more than 50%. The diagnosis of Jeavons syndrome is often delayed. Generalized tonic-clonic seizures and seizure types other than absence seizures may be predictors of drug-resistant epilepsy among patients with Jeavons syndrome.


SIRT1/FoxO3 axis alteration leads to aberrant immune responses in bronchial epithelial cells.
Inflammation and ageing are intertwined in chronic obstructive pulmonary disease (COPD). The histone deacetylase SIRT1 and the related activation of FoxO3 protect from ageing and regulate inflammation. The role of SIRT1/FoxO3 in COPD is largely unknown. This study evaluated whether cigarette smoke, by modulating the SIRT1/FoxO3 axis, affects airway epithelial pro-inflammatory responses. Human bronchial epithelial cells (16HBE) and primary bronchial epithelial cells (PBECs) from COPD patients and controls were treated with/without cigarette smoke extract (CSE), Sirtinol or FoxO3 siRNA. SIRT1, FoxO3 and NF-κB nuclear accumulation, SIRT1 deacetylase activity, IL-8 and CCL20 expression/release and the release of 12 cytokines, neutrophil and lymphocyte chemotaxis were assessed. In PBECs, the constitutive FoxO3 expression was lower in patients with COPD than in controls. Furthermore, CSE reduced FoxO3 expression only in PBECs from controls. In 16HBE, CSE decreased SIRT1 activity and nuclear expression, enhanced NF-κB binding to the IL-8 gene promoter thus increasing IL-8 expression, decreased CCL20 expression, increased the neutrophil chemotaxis and decreased lymphocyte chemotaxis. Similarly, SIRT1 inhibition reduced FoxO3 expression and increased nuclear NF-κB. FoxO3 siRNA treatment increased IL-8 and decreased CCL20 expression in 16HBE. In conclusion, CSE impairs the function of SIRT1/FoxO3 axis in bronchial epithelium, dysregulating NF-κB activity and inducing pro-inflammatory responses.


Compliance to antipsychotic medication: a challenge for client, family and health care providers.
Compliance with anti-psychotic medications plays a significant role in managing clients with schizophrenia. It not only helps in controlling the symptoms, but also decreases the risk of relapses and ultimately improves quality of life (QoL) for the clients. However, compliance with anti-psychotic medication remains a challenge for the client, family and healthcare providers. Identification of these associated factors is vital to make appropriate plan to enhance medication compliance. In this paper, various factors are highlighted that are associated with medication compliance in clients with schizophrenia.


Influence of anterior cervical plate design on Dysphagia: a 2-year prospective longitudinal follow-up study.
To compare the incidence, prevalence, and rate of improvement of dysphagia in patients undergoing anterior cervical spine surgery with two different anterior instrumentation designs. The study subjects were 156 consecutive patients undergoing anterior cervical spine surgery with plate fixation. We compared the incidence of dysphagia among the two different plate groups both produced by the same manufacturer (Medtronic Danek); the Atlantis plate has thicker and wider plate dimensions than the Zephir plate. Dysphagia evaluations were performed prospectively by telephone interviews at 1, 2, 6, 12, and 24 months following the procedure. Risk factors such as gender, revision surgery, and number of surgical levels were compared between the groups and were not statistically different. Overall incidences of dysphagia were 49%, 37%, 20%, 15.4%, and 11% at 1, 2, 6, 12, and 24 months, respectively. Severe and disabling dysphagia is reported to be a relatively uncommon complication of anterior cervical surgery. However, a significant number of patients report mild to moderate discomfort including double-swallowing and catching sensation. Except at the 2-month follow-up point, the Atlantis plate group had higher incidences of dysphagia than the Zephir group at all time points (57% vs 50%, 36% vs 4%, 23% vs 14%, 17% vs 7%, 14% vs 0% at 1, 2, 6, 12, and 24 months, respectively). The Atlantis plate group had a 14% incidence of dysphagia at 2 years compared with the Zephir group, which had a 0% incidence at 2 years (P < 0.04). For primary surgeries, there was a higher incidence of dysphagia at all time points in the Atlantis group when compared with the Zephir group (58% vs 43%, 35% vs 30%, 22% vs 10%, 17% vs 0%, and 13% vs 0% at 1, 2, 6, 12, and 24 months, respectively) (P < 0.04 at 1 year). A regression analysis was performed. The resulting formulas predict the permanent rate of dysphagia for the Atlantis group is 13.6% and for the Zephir group is 3.58%. The use of a smaller and smoother profile plate such as the Zephir does reduce the incidence of dysphagia as compared with a slightly larger and less smooth plate such as the Atlantis.


Testing the psychometric properties of the Brisbane Practice Environment Measure using Exploratory Factor Analysis and Confirmatory Factor Analysis in an Australian registered nurse population.
A cross-sectional survey was conducted, and the construct validity and reliability of the Brisbane Practice Environment Measure in an Australian sample of registered nurses were examined. Nurses were randomly selected from the database of an Australian nursing organization. The original 33 items of the Brisbane Practice Environment Measure were utilized to inform the psychometric properties using confirmatory factor analysis. The Cronbach's alpha was 0.938 for the total scale and ranged 0.657-0.887 for the subscales. A five-factor structure of the measure was confirmed, χ(2) = 944.622, (P < 0.01), χ(2) /d.f. ratio = 2.845, Tucker Lewis Index 0.929, Root Mean Square Error = 0.061 and Comparative Fit Index = 0.906. The selected 28 items of the measure proved reliable and valid in measuring effects of the practice environment upon Australian nurses. The implications are that regular measurement of the practice environment using these 28 items might assist in the development of strategies which might improve job satisfaction and retention of registered nurses in Australia.


Factors affecting patient delay of diagnosis and completion of Direct Observation Therapy, Short-course (DOTS) among the migrant population in Shandong, China.
In China, the epidemiological and socioeconomic status of the migrant population suggests that the vulnerable population should be prioritized for tuberculosis (TB) control. A face-to-face interview using a structured questionnaire was performed on a total of 314 smear-positive pulmonary TB patients among the migrant population of 12 randomly selected counties in Shandong Province, China. From the results, the cases of patient delay of diagnosis accounted for 40.8%, and the completion rate of Direct Observation Therapy, Short-course (DOTS) was as low as 67.2%. There were 47.1% missed cases in the first diagnosis. Factors affecting detection and treatment were present in their socioeconomic status, working style, and the accessibility to related TB care. The findings indicated that migrant TB patients suffer delayed diagnosis, a low case detection rate and a low completion DOTS rate. Improvement of migrants' working conditions and accessibility to specialized TB care is essential and is expected to lead to better case detection and treatment completion.


Optical parametric chirped-pulse amplifier as an alternative to Ti:sapphire regenerative amplifiers.
We demonstrated a high-pulse energy, femtosecond-pulse source based on optical parametric chirped-pulse amplification. We successfully amplified 1-microm broadband oscillator pulses to 31 mJ and recompressed them to 310-fs pulse duration, at a 10-Hz repetition rate. The gain in our system is 6 x 10(7), achieved by the single passing of only 40 mm of gain material pumped by a commercial Q-switched Nd:YAG laser. This relatively simple system replaces a more complex Ti:sapphire regenerative-amplifier-based chirped-pulse amplification system. Numerous features in design and performance of optical parametric chirped-pulse amplifiers make them a preferred alternative to regenerative amplifiers based on Ti:sapphire in the front end of high-peak-power lasers.


Natural Uranium Tissue Content of Three Caucasian Males.
Uranium content and concentrations were measured in the tissues of three Caucasian male whole body donors to the U.S. Transuranium and Uranium Registries with no known intake other than from natural environmental sources. Average total body uranium content in the three cases was 81.3 ± 22.3 μg, of which 37.2 ± 2.1 μg (46%) was in the skeleton. The skeleton had a mean concentration of 3.79 ± 0.45 μg U kg(-1) wet weight and 11.72 ± 1.49 μg U kg(-1) ash. Distribution was in bone volume and not predominately on bone surfaces. Soft tissue concentrations ranged over about an order of magnitude, averaging about 0.5 μg kg wet weight for all tissues except the thoracic lymph nodes, which averaged 32.3 times the mean for soft tissue of the three cases. Observed thyroid tissue concentrations were about an order of magnitude greater than the average soft tissue concentration in two of the three background cases, suggestive of a possible long-term depot in this organ. Kidney content of uranium averaged 0.38 ± 0.21 μg for the three cases, an order of magnitude lower than the 7 μg recommended for Reference Man. The lower content and concentration in the kidney do not support a significant long-term depot for uranium in that organ. Assuming equilibrium between intake and excretion, the tissue data suggest a transfer coefficient from blood to skeleton of 0.14 with a residence half-life in the skeleton of 4,950 d (13.56 y), significantly greater than the 1,500 d (4.1 y) half-time proposed by ICRP.


Arginase overexpression in neurons and its effect on traumatic brain injury.
Arginine is a semi-essential amino acid which serves as a substrate for nitric oxide (NO) production by nitric oxide synthase (NOS) and a precursor for various metabolites including ornithine, creatine, polyamines, and agmatine. Arginase competes with nitric oxide synthase for substrate arginine to produce orthinine and urea. There is contradictory evidence in the literature on the role of nitric oxide in the pathophysiology of traumatic brain injury (TBI). These contradictory perspectives are likely due to different NOS isoforms - endothelial (eNOS), inducible (iNOS) and neuronal (nNOS) which are expressed in the central nervous system. Of these, the role of nNOS in acute injury remains less clear. This study aimed to employ a genetic approach by overexpressing arginase isoforms specifically in neurons using a Thy-1 promoter to manipulate cell autonomous NO production in the context of TBI. The hypothesis was that increased arginase would divert arginine from pathological NO production. We generated 2 mouse lines that overexpress arginase I (a cytoplasmic enzyme) or arginase II (a mitochondrial enzyme) in neurons of FVB mice. We found that two-weeks after induction of controlled cortical injury, overexpressing arginase I but not arginase II in neurons significantly reduced contusion size and contusion index compared to wild-type (WT) mice. This study establishes enhanced neuronal arginase levels as a strategy to affect the course of TBI and provides support for the potential role of neuronal NO production in this condition.


Posture, flexibility and grip strength in horse riders.
Since the ability to train the horse to be ambidextrous is considered highly desirable, rider asymmetry is recognized as a negative trait. Acquired postural and functional asymmetry can originate from numerous anatomical regions, so it is difficult to suggest if any is developed due to riding. The aim of this study was therefore to assess symmetry of posture, strength and flexibility in a large population of riders and to determine whether typical traits exist due to riding. 127 right handed riders from the UK and USA were categorized according to years riding (in 20 year increments) and their competition level (using affiliated test levels). Leg length, grip strength and spinal posture were measured and recorded by a physiotherapist. Standing and sitting posture and trunk flexibility were measured with 3-D motion capture technology. Right-left differences were explored in relation to years riding and rider competitive experience. Significant anatomical asymmetry was found for the difference in standing acromion process height for a competition level (-0.07±1.50 cm Intro/Prelim; 0.02±1.31 cm Novice; 0.43±1.27 cm Elementary+; p=0.048) and for sitting iliac crest height for years riding (-0.23±1.36 cm Intro/Prelim; 0.01±1.50 cm Novice; 0.86±0.41 cm Elementary+; p=0.021). For functional asymmetry, a significant interaction was found for lateral bending ROM for years riding x competition level (p=0.047). The demands on dressage riders competing at higher levels may predispose these riders to a higher risk of developing asymmetry and potentially chronic back pain rather than improving their symmetry.


Impaired endothelium-dependent relaxations in rabbits subjected to aortic coarctation hypertension.
Rabbits were rendered hypertensive by suprarenal coarctation of the abdominal aorta. Seven days later, endothelium-dependent and endothelium-independent vascular relaxations were examined in vascular rings taken from hypertensive (thoracic aorta, carotid artery) and normotensive (abdominal aorta) regions. Relaxation of phenylephrine-contracted rings in response to endothelium-dependent agonists (acetylcholine, A23187) was impaired, compared with that in sham-operated and intact controls, in regions exposed to the elevated blood pressure (i.e., above the coarctation). Responses to acetylcholine and A23187 in the abdominal aorta, below the coarctation, were not altered. The diminished endothelium-dependent responses in the thoracic aorta were not affected by pretreatment with the cyclooxygenase inhibitor indomethacin. In contrast to acetylcholine and A23187, responses to the endothelium-independent agonist nitroprusside were not attenuated in vessels from hypertensive regions, indicating that the defect occurred in the endothelium. The EC50 for acetylcholine-induced relaxations of thoracic aorta correlated significantly with mean arterial pressure above the coarctation, indicating that the extent to which endothelium-dependent relaxation is impaired is in proportion to the degree of blood pressure elevation. This study suggests that the diminished relaxations by endothelium-dependent agonists is a local response to the elevation of blood pressure and is not due to a circulating factor.


A₃ adenosine receptor allosteric modulator induces an anti-inflammatory effect: in vivo studies and molecular mechanism of action.
The A3 adenosine receptor (A3AR) is overexpressed in inflammatory cells and in the peripheral blood mononuclear cells of individuals with inflammatory conditions. Agonists to the A3AR are known to induce specific anti-inflammatory effects upon chronic treatment. LUF6000 is an allosteric compound known to modulate the A3AR and render the endogenous ligand adenosine to bind to the receptor with higher affinity. The advantage of allosteric modulators is their capability to target specifically areas where adenosine levels are increased such as inflammatory and tumor sites, whereas normal body cells and tissues are refractory to the allosteric modulators due to low adenosine levels. LUF6000 administration induced anti-inflammatory effect in 3 experimental animal models of rat adjuvant induced arthritis, monoiodoacetate induced osteoarthritis, and concanavalin A induced liver inflammation in mice. The molecular mechanism of action points to deregulation of signaling proteins including PI3K, IKK, IκB, Jak-2, and STAT-1, resulting in decreased levels of NF-κB, known to mediate inflammatory effects. Moreover, LUF6000 induced a slight stimulatory effect on the number of normal white blood cells and neutrophils. The anti-inflammatory effect of LUF6000, mechanism of action, and the differential effects on inflammatory and normal cells position this allosteric modulator as an attractive and unique drug candidate.


Relationship between somatic mosaicism of Pax6 mutation and variable developmental eye abnormalities-an analysis of CRISPR genome-edited mouse embryos.
The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein (Cas) system is a rapid gene-targeting technology that does not require embryonic stem cells. To demonstrate dosage effects of the Pax6 gene on eye formation, we generated Pax6-deficient mice with the CRISPR/Cas system. Eyes of founder embryos at embryonic day (E) 16.5 were examined and categorized according to macroscopic phenotype as class 1 (small eye with distinct pigmentation), class 2 (pigmentation without eye globes), or class 3 (no pigmentation and no eyes). Histologically, class 1 eyes were abnormally small in size with lens still attached to the cornea at E16.5. Class 2 eyes had no lens and distorted convoluted retinas. Class 3 eyes had only rudimentary optic vesicle-like tissues or histological anophthalmia. Genotyping of neck tissue cells from the founder embryos revealed somatic mosaicism and allelic complexity for Pax6. Relationships between eye phenotype and genotype were developed. The present results demonstrated that development of the lens from the surface ectoderm requires a higher gene dose of Pax6 than development of the retina from the optic vesicle. We further anticipate that mice with somatic mosaicism in a targeted gene generated by CRISPR/Cas-mediated genome editing will give some insights for understanding the complexity in human congenital diseases that occur in mosaic form.


Real-Time Quaking-Induced Conversion Detection of Bovine Spongiform Encephalopathy Prions in a Subclinical Steer.
Bovine spongiform encephalopathy (BSE) belongs to a group of fatal prion diseases that result from the misfolding of the cellular prion protein (PrPC) into a pathogenic form (PrPSc) that accumulates in the brain. In vitro assays such as serial protein misfolding amplification and real-time quaking-induced conversion (RT-QuIC) allow assessment of the conversion of PrPC to PrPSc. RT-QuIC can be used for the detection of prions in a variety of biological tissues from humans and animals. However, there is no such comparison of RT-QuIC data between BSE positive and presymptomatic cattle. Further, the current study assesses prion distribution in multiple brain regions of clinically ill or subclinical animals. Here, we compare RT-QuIC reactions seeded with brain samples collected from experimentally inoculated cattle that were clinically ill or subclinically affected with BSE. The results demonstrate RT-QuIC seeding in various brain regions of an animal with subclinical BSE despite being determined negative by immunohistochemistry. Bioassay of the subclinical animal and RT-QuIC of brainstem from inoculated knockout (PRNP-/-) cattle were used to confirm infectivity in the subclinical animal and determine that RT-QuIC reactions were not the result of residual inoculum, respectively. These results confirm that RT-QuIC is a highly sensitive prion detection assay that can detect prions in a steer prior to the onset of clinical signs of BSE.


FGF4, a direct target of LEF1 and Wnt signaling, can rescue the arrest of tooth organogenesis in Lef1(-/-) mice.
Lymphoid enhancer factor (LEF1), a nuclear mediator of Wnt signaling, is required for the formation of organs that depend on inductive interactions between epithelial and mesenchymal tissues. In previous tissue recombination experiments with normal and Lef1(-/-) tooth germs, we found that the effect of LEF1 expression in the epithelium is tissue nonautonomous and transferred to the subjacent mesenchyme. Here we examine the molecular basis for LEF1 function and find that the epithelium of the developmentally arrested Lef1(-/-) tooth rudiments fails to express Fgf4, Shh, and Bmp4, but not Wnt10a. We identify the Fgf4 gene as a direct transcriptional target for LEF1 and show that beads soaked with recombinant FGF4 protein can fully overcome the developmental arrest of Lef1(-/-) tooth germs. In addition, we find that FGF4 beads induce rapidly the expression of Fgf3 in dental mesenchyme and that both epithelial and mesenchymal FGF proteins induce the delayed expression of Shh in the epithelium. Taken together, these data indicate that a single target of LEF1 can account for the function of LEF1 in tooth development and for a relay of a Wnt signal reception to a cascade of FGF signaling activities, allowing for a sequential and reciprocal communication between epithelium and mesenchyme.


Getting serious about the early-life epilepsies: Lessons from the world of pediatric oncology.
Early-life epilepsies represent a group of many individually rare and often complex developmental brain disorders associated with lifelong devastating consequences and high risk for early mortality. The quantity and quality of evidence needed to guide the evaluation and treatment to optimize outcomes of affected children is minimal; most children are treated within an evidence-free practice zone based solely on anecdote and lore. The remarkable advances in diagnostics and therapeutics are implemented haphazardly with no systematic effort to understand their effects and value. This stands in stark contrast to the evidence-rich practice of the Children's Oncology Group, where standard of care treatments are identified through rigorous, multicenter research studies, and the vast majority of patients are treated on protocols developed from that research. As a consequence, overall mortality for childhood cancers has declined from ∼90% in the 1950s to ∼20% today. The situations of these 2 rare disease specialties are contrasted, and some suggestions for moving early-life epilepsy onto a fast track for success are offered. Chief amongst these is that early-life epilepsy should be treated with the same urgency as pediatric cancer. The best diagnostics and evidence-based treatments should be used in a systematic fashion right from the start, not after the child and family have been subjected to the ravages of the disorder for months or years. This will require unity and cooperation among physicians, researchers, and institutions across state and national borders.


Global conservation significance of Ecuador's Yasuní National Park.
The threats facing Ecuador's Yasuní National Park are emblematic of those confronting the greater western Amazon, one of the world's last high-biodiversity wilderness areas. Notably, the country's second largest untapped oil reserves--called "ITT"--lie beneath an intact, remote section of the park. The conservation significance of Yasuní may weigh heavily in upcoming state-level and international decisions, including whether to develop the oil or invest in alternatives. We conducted the first comprehensive synthesis of biodiversity data for Yasuní. Mapping amphibian, bird, mammal, and plant distributions, we found eastern Ecuador and northern Peru to be the only regions in South America where species richness centers for all four taxonomic groups overlap. This quadruple richness center has only one viable strict protected area (IUCN levels I-IV): Yasuní. The park covers just 14% of the quadruple richness center's area, whereas active or proposed oil concessions cover 79%. Using field inventory data, we compared Yasuní's local (alpha) and landscape (gamma) diversity to other sites, in the western Amazon and globally. These analyses further suggest that Yasuní is among the most biodiverse places on Earth, with apparent world richness records for amphibians, reptiles, bats, and trees. Yasuní also protects a considerable number of threatened species and regional endemics. Yasuní has outstanding global conservation significance due to its extraordinary biodiversity and potential to sustain this biodiversity in the long term because of its 1) large size and wilderness character, 2) intact large-vertebrate assemblage, 3) IUCN level-II protection status in a region lacking other strict protected areas, and 4) likelihood of maintaining wet, rainforest conditions while anticipated climate change-induced drought intensifies in the eastern Amazon. However, further oil development in Yasuní jeopardizes its conservation values. These findings form the scientific basis for policy recommendations, including stopping any new oil activities and road construction in Yasuní and creating areas off-limits to large-scale development in adjacent northern Peru.


Daughter preference and contraceptive-use in matrilineal tribal societies in Meghalaya, India.
Although son preference in patrilineal society is an established fact, daughter preference in matrilineal society is not thoroughly examined. Very few studies have been carried out on the issue. This paper attempts to explore the daughter preference and contraceptive-use in matrilineal tribal societies in Meghalaya, India. Data from the National Family Health Survey 1998-1999 have been used in this study because, among the large-scale surveys, only this dataset allows identification of matrilineal sample. Mean, percentage, and standard deviation are computed in the present study. Further, the data have been cross-tabulated, and logistic regression has been run through SPSS (version 15). Among the ever-married matrilineal women, 17% desired more sons than daughters but 18.2% desired more daughters than sons. About 11% of ever-married women could achieve their desired sex composition of children. However, a very striking finding suggests that, even after achieving desired sex composition of children, as high as 61.8% of women were still not using contraception mainly because of programme factors while one-fourth were still depending on temporary methods. The rest 13.2% adopted terminal method of contraception, which calls for immediate attention of planners. With the increase in the number of sons but without daughter, contraceptive-use drastically decreased. The most desired sex composition of children seems to be two daughters and a son. Absence of daughter with increase in the total number of sons increased the desire for additional children. Every woman with two or more sons but without daughter wanted the next child to be a daughter. Thus, there are ample evidences to draw the conclusion that there is, in fact, a daughter preference in the matrilineal tribal societies in Meghalaya, India. Policy-makers may, thus, target the women who have achieved fertility and should ensure that daughter preference does not lead to the negligence to sons.


The influence of enrichment devices on development of osteoarthritis in a surgically induced murine model.
This study measured the influence of three different environmental enrichment devices (EEDs) on the severity of osteoarthritis (OA) in a surgically induced murine model. The development of OA requires joint movement after surgical instability induced by destabilization of the medial meniscus at 10 weeks of age. We evaluated the hypothesis that animals behavioral activity levels may influence the severity of the disease by investigating the effect of different EEDs on mouse activity and correlating this to OA severity. Thirty male 129S6/SvEvTac mice were housed in groups of five and provided with nesting material and one of three different EEDs: a heavy plastic tube (CPVC), Shepherd Shack (SS), or Tecniplast Mouse House (TMH). We videorecorded the cages throughout the study and constructed an ethogram. Eight weeks after surgery we euthanized the mice and performed a histologic examination of the knees to score the severity of OA based on the different housing systems, correlating the scores with behavioral activity levels for each cage. OA was higher in the mice with CPVC and TMH devices in their cages, whereas the mice with SS devices exhibited less cartilage damage; however, although we observed increased behavioral activity in mice with the CPVC tube and TMH and less in mice with the SS, the statistical results were not significant. The histological results of OA and the ethogram correlated to support our hypothesis that the type of EED plays an indirect role in the severity of the disease by modifying the activity levels of mice. In activity-dependent studies, the impact of an EED needs to be evaluated before change the environment.


Amyloid formation by the pro-inflammatory S100A8/A9 proteins in the ageing prostate.
The conversion of soluble peptides and proteins into polymeric amyloid structures is a hallmark of many age-related degenerative disorders, including Alzheimer's disease, type II diabetes and a variety of systemic amyloidoses. We report here that amyloid formation is linked to another major age-related phenomenon--prostate tissue remodelling in middle-aged and elderly men. By using multidisciplinary analysis of corpora amylacea inclusions in prostate glands of patients diagnosed with prostate cancer we have revealed that their major components are the amyloid forms of S100A8 and S100A9 proteins associated with numerous inflammatory conditions and types of cancer. In prostate protease rich environment the amyloids are stabilized by dystrophic calcification and lateral thickening. We have demonstrated that material closely resembling CA can be produced from S100A8/A9 in vitro under native and acidic conditions and shows the characters of amyloids. This process is facilitated by calcium or zinc, both of which are abundant in ex vivo inclusions. These observations were supported by computational analysis of the S100A8/A9 calcium-dependent aggregation propensity profiles. We found DNA and proteins from Escherichia coli in CA bodies, suggesting that their formation is likely to be associated with bacterial infection. CA inclusions were also accompanied by the activation of macrophages and by an increase in the concentration of S100A8/A9 in the surrounding tissues, indicating inflammatory reactions. These findings, taken together, suggest a link between bacterial infection, inflammation and amyloid deposition of pro-inflammatory proteins S100A8/A9 in the prostate gland, such that a self-perpetuating cycle can be triggered and may increase the risk of malignancy in the ageing prostate. The results provide strong support for the prediction that the generic ability of polypeptide chains to convert into amyloids could lead to their involvement in an increasing number of otherwise apparently unrelated diseases, particularly those associated with ageing.


Strategies to support substance use disorder care transitions from acute-care to community-based settings: A Scoping review and typology.
Acute-care interventions that identify patients with substance use disorders (SUDs), initiate treatment, and link patients to community-based services, have proliferated in recent years. Yet, much is unknown about the specific strategies being used to support continuity of care from emergency department (ED) or inpatient hospital settings to community-based SUD treatment. In this scoping review, we synthesize the existing literature on patient transition interventions, and form an initial typology of reported strategies. We searched Pubmed, Embase, CINAHL and PsychINFO for peer-reviewed articles published between 2000-2021 that studied interventions linking SUD patients from ED or inpatient hospital settings to community-based SUD services. Eligible articles measured at least one post-discharge treatment outcome and included a description of the strategy used to promote linkage to community care. Detailed information was extracted on the components of the transition strategies and a thematic coding process was used to categorize strategies into a typology based on shared characteristics. Facilitators and barriers to transitions of care were synthesized using the Consolidated Framework for Implementation Research. Forty-five articles met inclusion criteria. 62% included ED interventions and 44% inpatient interventions. The majority focused on patients with opioid (71%) followed by alcohol (31%) use disorder. The transition strategies reported across studies were heterogeneous and often not well described. An initial typology of ten transition strategies, including five pre- and five post-discharge transition strategies is proposed. The most common strategy was scheduling an appointment with a community-based treatment provider prior to discharge. A range of facilitators and barriers were described, which can inform efforts to improve hospital-to-community transitions of care. Strategies to support transitions from acute-care to community-based SUD services, although critical for ensuring continuity of care, vary greatly across interventions and are inconsistently measured and described. More research is needed to classify SUD care transition strategies, understand their components, and explore which lead to the best patient outcomes.


Road safety of passing maneuvers: A bivariate extreme value theory approach under non-stationary conditions.
Observed accidents have been the main resource for road safety analysis over the past decades. Although such reliance seems quite straightforward, the rare nature of these events has made safety difficult to assess, especially for new and innovative traffic treatments. Surrogate measures of safety have allowed to step away from traditional safety performance functions and analyze safety performance without relying on accident records. In recent years, the use of extreme value theory (EV) models in combination with surrogate safety measures to estimate accident probabilities has gained popularity within the safety community. In this paper we extend existing efforts on EV for accident probability estimation for two dependent surrogate measures. Using detailed trajectory data from a driving simulator, we model the joint probability of head-on and rear-end collisions in passing maneuvers. We apply the Block Maxima method and estimate several extremal univariate and bivariate models, including the logistic copula. In our estimation we account for driver specific characteristics and road infrastructure variables. We show that accounting for these factors improve the head-on and rear-end collision probabilities estimation. This work highlights the importance of considering driver and road heterogeneity in evaluating related safety events, of relevance to interventions both for in-vehicle and infrastructure-based solutions. Such features are essential to keep up with the expectations from surrogate safety measures for the integrated analysis of accident phenomena, which show to significantly improve from the best known stationary extreme value models.


The discoidin I gene family of Dictyostelium discoideum is linked to genes regulating its expression.
The discoidin I protein has been studied extensively as a marker of early development in the cellular slime mold Dictyostelium discoideum. However, like most other developmentally regulated proteins in this system, no reliable information was available on the linkage of the discoidin genes to other known genes. Analysis of the linkage of the discoidin I genes by use of restriction fragment length polymorphisms revealed that all three discoidin I genes as well as a pseudogene are located on linkage group II. This evidence is consistent with the discoidin I genes forming a gene cluster that may be under the control of a single regulatory element. The discoidin I genes are linked to three genetic loci (disA, motA, daxA) that affect the expression of the discoidin I protein. Linkage of the gene family members to regulatory loci may be important in the coordinate maintenance of the gene family and regulatory loci. A duplication affecting the entire discoidin gene family is also linked to group II; this appears to be a small tandem duplication. This duplication was mapped using a DNA polymorphism generated by insertion of the Tdd-3 mobile genetic element into a Tdd-2 element flanking the gamma gene. A probe for Tdd-2 identified a restriction fragment length polymorphism in strain AX3K that was consistent with generation by a previously proposed Tdd-3 insertion event. A putative duplication or rearrangement of a second Tdd-2 element on linkage group IV of strain AX3K was also identified. This is the first linkage information available for mobile genetic elements in D. discoideum.


The pre- and postoperative illness trajectory in patients with pituitary tumours.
Experiences and need of support during surgery and start of replacement therapy in patients with pituitary tumours are highly unknown. This study aimed at exploring patient experiences during pre- and postoperative care and recovery after pituitary surgery in patients with a pituitary tumour. Within a qualitative study design, 16 consecutive patients who underwent surgery for pituitary tumours were repeatedly interviewed. In total, 42 interviews were performed before and after surgery. Analysis was performed using qualitative interpretation. Suffering a pituitary tumour was overwhelming for many patients and struggling with existential issues was common. Patients expressed loneliness and vulnerability before and after surgery. How professionals handled information in connection with diagnosis greatly affected the patients. Other patients with the same diagnosis were experienced as the greatest support. Normalisation of bodily symptoms and relationships with others were reported during postoperative recovery. However, a fear that the tumour would return was present. Patients with pituitary tumours need structured support, including peer support, which acknowledges physical, cognitive as well as emotional and existential concerns. Information related to diagnosis and surgery should be adapted in relation to the loneliness and the existential seriousness of the situation. Care and support for patients with pituitary tumours should preferably be organised based on continuity and an unbroken care pathway from the first pre-operative evaluation through to postoperative care and the start of a life-long endocrine treatment and tumour surveillance.


Raman Spectroscopy of Organic-Inorganic Halide Perovskites.
Micro-Raman spectroscopy provides laterally resolved microstructural information for a broad range of materials. In this Letter, we apply this technique to tri-iodide (CH3NH3PbI3), tribromide (CH3NH3PbBr3), and mixed iodide-bromide (CH3NH3PbI3-xBrx) organic-inorganic halide perovskite thin films and discuss necessary conditions to obtain reliable data. We explain how to measure Raman spectra of pristine CH3NH3PbI3 layers and discuss the distinct Raman bands that develop during moisture-induced degradation. We also prove unambiguously that the final degradation products contain pure PbI2. Moreover, we describe CH3NH3PbI3-xBrx Raman spectra and discuss how the perovskite crystallographic symmetries affect the Raman band intensities and spectral shapes. On the basis of the dependence of the Raman shift on the iodide-to-bromide ratio, we show that Raman spectroscopy is a fast and nondestructive method for the evaluation of the relative iodide-to-bromide ratio.


Antepartum dalteparin versus no antepartum dalteparin for the prevention of pregnancy complications in pregnant women with thrombophilia (TIPPS): a multinational open-label randomised trial.
Thrombophilias are common disorders that increase the risk of pregnancy-associated venous thromboembolism and pregnancy loss and can also increase the risk of placenta-mediated pregnancy complications (severe pre-eclampsia, small-for-gestational-age infants, and placental abruption). We postulated that antepartum dalteparin would reduce these complications in pregnant women with thrombophilia. In this open-label randomised trial undertaken in 36 tertiary care centres in five countries, we enrolled consenting pregnant women with thrombophilia at increased risk of venous thromboembolism or with previous placenta-mediated pregnancy complications. Eligible participants were randomly allocated in a 1:1 ratio to either antepartum prophylactic dose dalteparin (5000 international units once daily up to 20 weeks' gestation, and twice daily thereafter until at least 37 weeks' gestation) or to no antepartum dalteparin (control group). Randomisation was done by a web-based randomisation system, and was stratified by country and gestational age at randomisation day with a permuted block design (block sizes 4 and 8). At randomisation, site pharmacists (or delegates) received a randomisation number and treatment allocation (by fax and/or e-mail) from the central web randomisation system and then dispensed study drug to the local coordinator. Patients and study personnel were not masked to treatment assignment, but the outcome adjudicators were masked. The primary composite outcome was independently adjudicated severe or early-onset pre-eclampsia, small-for-gestational-age infant (birthweight <10th percentile), pregnancy loss, or venous thromboembolism. We did intention-to-treat and on-treatment analyses. This trial is registered with ClinicalTrials.gov, number NCT00967382, and with Current Controlled Trials, number ISRCTN87441504. Between Feb 28, 2000, and Sept 14, 2012, 292 women consented to participate and were randomly assigned to the two groups. Three women were excluded after randomisation because of ineligibility (two in the antepartum dalteparin group and one in the control group), leaving 146 women assigned to antepartum dalteparin and 143 assigned to no antepartum dalteparin. Some patients crossed over to the other group during treatment, and therefore for on-treatment and safety analysis there were 143 patients in the dalteparin group and 141 in the no dalteparin group. Dalteparin did not reduce the incidence of the primary composite outcome in both intention-to-treat analysis (dalteparin 25/146 [17·1%; 95% CI 11·4-24·2%] vs no dalteparin 27/143 [18·9%; 95% CI 12·8-26·3%]; risk difference -1·8% [95% CI -10·6% to 7·1%)) and on-treatment analysis (dalteparin 28/143 [19·6%] vs no dalteparin 24/141 [17·0%]; risk difference +2·6% [95% CI -6·4 to 11·6%]). In safety analysis, the occurrence of major bleeding did not differ between the two groups. However, minor bleeding was more common in the dalteparin group (28/143 [19·6%]) than in the no dalteparin group (13/141 [9·2%]; risk difference 10·4%, 95% CI 2·3-18·4; p=0·01). Antepartum prophylactic dalteparin does not reduce the occurrence of venous thromboembolism, pregnancy loss, or placenta-mediated pregnancy complications in pregnant women with thrombophilia at high risk of these complications and is associated with an increased risk of minor bleeding. Canadian Institutes of Health Research, Heart and Stroke Foundation of Canada, and Pharmacia and UpJohn.


Scoping review of the impacts of urban agriculture on the determinants of health.
There has been an increasing interest in urban agriculture (UA) practice and research in recent years. Scholars have already reported numerous beneficial and potential adverse impacts of UA on health-related outcomes. This scoping review aims to explore these impacts and identify knowledge gaps for future UA studies. A systematic search was conducted in seven electronic bibliographic databases to identify relevant peer-reviewed studies. Articles were screened and assessed for eligibility. From eligible studies, data were extracted to summarize, collate, appraise the quality and make a narrative account of the findings. A total of 101 articles (51 quantitative, 29 qualitative, and 21 mixed methods studies) were included in our final analysis. Among these articles, 38 and 37% reported findings from North America and Sub-Saharan Africa respectively. Quantitative studies revealed evidence of positive impacts of UA on food security, nutrition outcomes, physical and mental health outcomes, and social capital. The qualitative studies reported a wide range of perceived benefits and motivations of UA. The most frequently reported benefits include contributions to social capital, food security, health and/or wellbeing. However, the evidence must be interpreted with caution since the quality of most of the studies was assessed as weak to moderate. While no definitive conclusions can be drawn about the adverse impacts of UA on health, paying particular attention to contamination of UA soil is recommended. More peer-reviewed studies are needed in areas where UA is practiced such as Latin America and Caribbean. The inconsistency and the lack of strong quality in the methodology of the included studies are proof that more rigorous studies are also needed in future research. Nevertheless, the substantial existing evidence from this review corroborate that UA can influence different determinants of health such as food security, social capital, health and well-being in a variety of contexts.


[Fluorine speciation in soil and the remediation of fluorine contaminated soil].
In China, fluoride pollution in soil is severe and poses a serious threat to human health and ecological security. However, how to control fluorine-contaminated soil has not received widespread attention. Here, we summarized fluorine speciation in soil and its main chemical reactions in water-soil system and reviewed the research progress on the remediation of fluorine-contaminated soils. Then, we proposed the focus of future research on fluorine-contaminated soil remediation. The aim of this review was providing the reference for the remediation of fluoride-contaminated soil. There are five forms of fluorine in soils, with the proportion of residual fluorine being over 90%. The reactions of fluorine in the soil solution mainly include precipitation-dissolution, complexation-dissociation, and adsorption-desorption. At present, the remediation technology of fluorine contaminated soil mainly focused on chemo-immobilization, chemical leaching, electrokinetic remediation, and phytoremediation. Clarifying the combined forms of fluorine in soil, screening functional microorga-nisms and plants, developing the combined remediation technology will be the focus of future research. Ultimately, on site fluorine-contaminated soil remediation could be implemented.


Waveguide propagation-loss measurement technique.
A novel nondestructive propagation-loss measurement technique for single-mode waveguides is demonstrated. The method calculates the total propagation-loss coefficient from the measured ratio of the retroreflected and transmitted light from the waveguide and is independent of the coupling efficiency of the input light. A surface-wave transducer is utilized as an acousto-optic modulator in a lock-in scheme to enhance the measurement ease and precision.


Offending behaviour in adults with Asperger syndrome.
Considerable speculation is evident both within the scientific literature and popular media regarding possible links between Asperger syndrome and offending. A survey methodology that utilised quantitative data collection was employed to investigate the prevalence of offending behaviour amongst adults with Asperger Syndrome in a large geographical area of South Wales, UK; qualitative interviews were then conducted with a sub-sample of those identified. A small number of participants meeting the study criteria were identified. For those who had offended, their experience of the criminal justice system was essentially negative. Possible implications of the results were discussed.


Differential modulation of cell adhesion by interaction between adhesive and counter-adhesive proteins: characterization of the binding of vitronectin to osteonectin (BM40, SPARC).
Heparin-binding forms of vitronectin, a multifunctional adhesive glycoprotein, are associated with the extracellular matrix (ECM) at different locations in the body and serve to promote cell adhesion and the regulation of pericellular proteolysis at sites of angiogenesis. In the present study we characterized the interactions of vitronectin with the counter-adhesive protein osteonectin (also termed SPARC or BM40). Osteonectin and vitronectin were both found associated with the ECM of cultured endothelial cells and were localized in vessel wall sections of kidney tissue. In vitro, the heparin-binding multimeric isoform of vitronectin bound to immobilized osteonectin in a saturable manner with half-maximal binding at 30-40 nM. Preincubation of plasma vitronectin with plasminogen activator inhibitor 1 (PAI-1), which provoked multimer formation, induced the binding of vitronectin to osteonectin. Binding was optimal at physiological ionic strength, and binary complexes were stabilized by tissue transglutaminase-mediated cross-linking. In a concentration-dependent fashion, PAI-1, CaCl2, heparin and heparan sulphate, but not other glycosaminoglycans, interfered with the binding of vitronectin to osteonectin. Using vitronectin-derived synthetic peptides as well as mutant forms of recombinant osteonectin, we found that the heparin-binding region of vitronectin interacted with the C-terminal region of osteonectin that contains a high-affinity Ca2+-binding site with counter-adhesive properties. Adhesion of cultured endothelial cells was partly abrogated by osteonectin and was correspondingly reversed by vitronectin in a concentration-dependent manner. These results indicate that specific interactions between vitronectin and osteonectin modulate cell adhesion and might thereby regulate endothelial cell function during angiogenesis.


Identification of a novel archaebacterial thioredoxin: determination of function through structure.
As part of a high-throughput, structural proteomic project we have used NMR spectroscopy to determine the solution structure and ascertain the function of a previously unknown, conserved protein (MtH895) from the thermophilic archeon Methanobacterium thermoautotrophicum. Our findings indicate that MtH895 contains a central four-stranded beta-sheet core surrounded by two helices on one side and a third on the other. It has an overall fold superficially similar to that of a glutaredoxin. However, detailed analysis of its three-dimensional structure along with molecular docking simulations of its interaction with T7 DNA polymerase (a thioredoxin-specific substrate) and comparisons with other known members of the thioredoxin/glutaredoxin family of proteins strongly suggest that MtH895 is more akin to a thioredoxin. Furthermore, measurement of the pK(a) values of its active site thiols along with direct measurements of the thioredoxin/glutaredoxin activity has confirmed that MtH895 is, indeed, a thioredoxin and exhibits no glutaredoxin activity. We have also identified a group of previously unknown proteins from several other archaebacteria that have significant (34-44%) sequence identity with MtH895. These proteins have unusual active site -CXXC- motifs not found in any known thioredoxin or glutaredoxin. On the basis of the results presented here, we predict that these small proteins are all members of a new class of truncated thioredoxins.


A switch in the control of growth of the wing imaginal disks of Manduca sexta.
Insulin and ecdysone are the key extrinsic regulators of growth for the wing imaginal disks of insects. In vitro tissue culture studies have shown that these two growth regulators act synergistically: either factor alone stimulates only limited growth, but together they stimulate disks to grow at a rate identical to that observed in situ. It is generally thought that insulin signaling links growth to nutrition, and that starvation stops growth because it inhibits insulin secretion. At the end of larval life feeding stops but the disks continue to grow, so at that time disk growth has become uncoupled from nutrition. We sought to determine at exactly what point in development this uncoupling occurs. Growth and cell proliferation in the wing imaginal disks and hemolymph carbohydrate concentrations were measured at various stages in the last larval instar under experimental conditions of starvation, ligation, rescue, and hormone treatment. Here we show that in the last larval instar of M. sexta, the uncoupling of nutrition and growth occurs as the larva passes the critical weight. Before this time, starvation causes a decline in hemolymph glucose and trehalose and a cessation of wing imaginal disks growth, which can be rescued by injections of trehalose. After the critical weight the trehalose response to starvation disappears, and the expression of insulin becomes decoupled from nutrition. After the critical weight the wing disks loose their sensitivity to repression by juvenile hormone, and factors from the abdomen, but not the brain, are required to drive continued growth. During the last larval instar imaginal disk growth becomes decoupled from somatic growth at the time that the endocrine events of metamorphosis are initiated. These regulatory changes ensure that disk growth continues uninterrupted when the nutritive and endocrine signals undergo the drastic changes associated with metamorphosis.


Single-nucleotide polymorphism identification and genotyping in Camelina sativa.
Camelina sativa, a largely relict crop, has recently returned to interest due to its potential as an industrial oilseed. Molecular markers are key tools that will allow C. sativa to benefit from modern breeding approaches. Two complementary methodologies, capture of 3' cDNA tags and genomic reduced-representation libraries, both of which exploited second generation sequencing platforms, were used to develop a low density (768) Illumina GoldenGate single nucleotide polymorphism (SNP) array. The array allowed 533 SNP loci to be genetically mapped in a recombinant inbred population of C. sativa. Alignment of the SNP loci to the C. sativa genome identified the underlying sequenced regions that would delimit potential candidate genes in any mapping project. In addition, the SNP array was used to assess genetic variation among a collection of 175 accessions of C. sativa, identifying two sub-populations, yet low overall gene diversity. The SNP loci will provide useful tools for future crop improvement of C. sativa.


Group B Streptococcus Degrades Cyclic-di-AMP to Modulate STING-Dependent Type I Interferon Production.
Induction of type I interferon (IFN) in response to microbial pathogens depends on a conserved cGAS-STING signaling pathway. The presence of DNA in the cytoplasm activates cGAS, while STING is activated by cyclic dinucleotides (cdNs) produced by cGAS or from bacterial origins. Here, we show that Group B Streptococcus (GBS) induces IFN-β production almost exclusively through cGAS-STING-dependent recognition of bacterial DNA. However, we find that GBS expresses an ectonucleotidase, CdnP, which hydrolyzes extracellular bacterial cyclic-di-AMP. Inactivation of CdnP leads to c-di-AMP accumulation outside the bacteria and increased IFN-β production. Higher IFN-β levels in vivo increase GBS killing by the host. The IFN-β overproduction observed in the absence of CdnP is due to the cumulative effect of DNA sensing by cGAS and STING-dependent sensing of c-di-AMP. These findings describe the importance of a bacterial c-di-AMP ectonucleotidase and suggest a direct bacterial mechanism that dampens activation of the cGAS-STING axis.


Comparing survival curves using an easy to interpret statistic.
Here, I describe a statistic for comparing two survival curves that has a clear and obvious meaning and has a long history in biostatistics. Suppose we are comparing survival times associated with two treatments A and B. The statistic operates in such a way that if it takes on the value 0.95, then the interpretation is that a randomly chosen patient treated with A has a 95% chance of surviving longer than a randomly chosen patient treated with B. This statistic was first described in the 1950s, and was generalized in the 1960s to work with right-censored survival times. It is a useful and convenient measure for assessing differences between survival curves. Software for computing the statistic is readily available on the Internet.


A novel protein modification pathway related to the ubiquitin system.
Ubiquitin conjugation is known to target protein substrates primarily to degradation by the proteasome or via the endocytic route. Here we describe a novel protein modification pathway in yeast which mediates the conjugation of RUB1, a ubiquitin-like protein displaying 53% amino acid identity to ubiquitin. We show that RUB1 conjugation requires at least three proteins in vivo. ULA1 and UBA3 are related to the N- and C-terminal domains of the E1 ubiquitin-activating enzyme, respectively, and together fulfil E1-like functions for RUB1 activation. RUB1 conjugation also requires UBC12, a protein related to E2 ubiquitin-conjugating enzymes, which functions analogously to E2 enzymes in RUB1-protein conjugate formation. Conjugation of RUB1 is not essential for normal cell growth and appears to be selective for a small set of substrates. Remarkably, CDC53/cullin, a common subunit of the multifunctional SCF ubiquitin ligase, was found to be a major substrate for RUB1 conjugation. This suggests that the RUB1 conjugation pathway is functionally affiliated to the ubiquitin-proteasome system and may play a regulatory role.


Differential regulation of endogenous pro-inflammatory cytokine genes by medroxyprogesterone acetate and norethisterone acetate in cell lines of the female genital tract.
Medroxyprogesterone acetate (MPA) and norethisterone (NET) and its derivatives are widely used in female reproductive therapy, but little is known about their mechanisms of action via steroid receptors in the female genital tract. MPA used as a contraceptive has been implicated in effects on local immune function. However, the relative effects of progesterone (Prog), MPA and norethisterone acetate (NET-A) on cytokine gene expression in the female genital tract are unknown. Using two epithelial cell lines generated from normal human vaginal (Vk2/E6E7) and ectocervical (Ect1/E6E7) cells as in vitro cell culture model systems for mucosal immunity of the female cervicovaginal environment, we investigated steroid receptor expression and activity as well as regulation of cytokine/chemokine genes by MPA and NET-A, as compared to the endogenous hormone Prog. We show that the Prog, androgen, glucocorticoid and estrogen receptors (PR, AR, GR and ER, respectively) are expressed in both the Vk2/E6E7 and Ect1/E6E7 cell lines, and that the GR and AR are transcriptionally active. This study is the first to show ligand-, promoter- and cell-specific regulation of IL-6, IL-8 and RANTES (regulated-upon-activation, normal T cell expressed and secreted) gene expression by Prog, MPA and NET-A in these cell lines. Moreover, we show that the repression of the TNF-α-induced RANTES gene by MPA in the Ect1/E6E7 cell line is mediated by the AR. Collectively, these data demonstrate that cell lines from different anatomical sites of the female genital tract respond differently to Prog and the synthetic progestins, most likely due to differential actions via different steroid receptors. The results highlight the importance of choice of progestins for immune function in the cervicovaginal environment. They further suggest that choice of progestins in endocrine therapy may have implications for women's risk of susceptibility to infections due to differential actions on genes involved in inflammation and immune function.


Costs of planning and implementing the CDC's Colorectal Cancer Screening Demonstration Program.
The Centers for Disease Control and Prevention (CDC) initiated the Colorectal Cancer Screening Demonstration Program (CRCSDP) to explore the feasibility of establishing a large-scale colorectal cancer screening program for underserved populations in the United States. The authors of the current report provide a detailed description of the total program costs (clinical and nonclinical) incurred during both the start-up and service delivery (screening) phases of the 4-year program. Tailored cost questionnaires were completed by staff at the 5 CRCSDP sites. Cost data were collected for clinical services and nonclinical programmatic activities (program management, data collection, and tracking, etc). In-kind contributions also were measured and were assigned monetary values. Nearly $11.3 million was expended by the 5 sites over 4 years, and 71% was provided by the CDC. The proportion of funding spent on clinical service delivery and service delivery/patient support comprised the largest proportion of cost during the implementation phase (years 2-4). The per-person nonclinical cost comprised a substantial portion of total costs for all sites. The cost per person screened varied across the 5 sites and by screening method. Overall, economies of scale were observed, with lower costs resulting from larger numbers of individuals screened. Programs incur substantial variable costs related to clinical services and semivariable costs related to nonclinical services. Therefore, programs that serve large populations are likely to achieve a lower cost per person.


Scoping Review of School-to-Work Transition for Youth With Intellectual Disabilities: A Practice Gap.
Youth with intellectual disabilities (ID) have persistently poor work outcomes. Occupational therapy can support school-to-work transition but is underrepresented in transition practice. To identify and describe interventions within the scope of occupational therapy for youth with ID who are transitioning from school to work. MEDLINE, ERIC, PsycINFO, and CINAHL were searched, and hand searching was performed in relevant peer-reviewed journals. Included were peer-reviewed, English-language articles published from 2004 to 2017 describing studies focused on youth with ID with no significant co-occurring physical diagnoses who were transitioning from U.S.-based school settings to paid employment. Data extraction was managed using Google Drive. Data were organized on extraction sheets by trained reviewers. The quality of each study was assessed using questions adapted from the Critical Appraisal Skills Program checklist. A total of 35 articles were included, 7 of which used randomized controlled designs. All articles described interventions aligned with the Occupational Therapy Practice Framework: Domain and Process (3rd ed.), but specific mention of occupational therapy was notably absent from the literature. Interventions had little and generally low-level evidence supporting their use. Significant and concerning gaps exist in the literature on school-to-work transition for youth with ID, likely impeding evidence-based practice. No included article mentioned occupational therapy or had a contributor who was an occupational therapy practitioner. Practitioners should advocate for occupational therapy's role in transition and contribute reports of occupational therapy transition services for youth with ID to the literature. This study demonstrates that occupational therapy is poorly represented in literature describing transition services for youth with ID. Although the articles described interventions within the occupational therapy domain, these interventions were not provided by occupational therapy practitioners and did not have a strong evidence base.


Preparation and activity of synthetic unmodified mammalian tRNAi(Met) in initiation of translation in vitro.
Translation of eukaryotic mRNA is initiated by a unique amino-acyl tRNA, Met-tRNAi(Met), which passes through a complex series of highly specific interactions with components of the translation apparatus during the initiation process. To facilitate in vitro biochemical and molecular biological analysis of these interactions in fully reconstituted translation initiation reactions, we generated mammalian tRNAi(Met) by in vitro transcription that lacked all eight base modifications present in native tRNAi(Met). Here we report a method for in vitro transcription and aminoacylation of synthetic unmodified initiator tRNAi(Met) that is active in every stage of the initiation process, including aminoacylation by methionyl-tRNA synthetase, binding of Met-tRNAi(Met) to eIF2-GTP to form a ternary complex, binding of the ternary complexes to 40S ribosomal subunits to form 43S complexes, binding of the 43S complex to a native capped eukaryotic mRNA, and scanning on its 5' untranslated region to the correct initiation codon to form a 48S complex, and finally joining with a 60S subunit to assemble an 80S ribosome that is competent to catalyze formation of the first peptide bond using the [35S]methionine residue attached to the acceptor terminus of the tRNAi(Met) transcript.


Evaluation of antibacterial activity of Australian basidiomycetous macrofungi using a high-throughput 96-well plate assay.
The production of antimicrobial compounds by macrofungi is not unexpected because they have to compete with other organisms for survival in their natural hostile environment. Previous studies have indicated that macrofungi contain secondary metabolites with a range of pharmacological activities including antimicrobial agents. To investigate macrofungi for antimicrobial activity due to the increasing need for new antimicrobials as a result of resistance in the bacterial community to existing treatments. Forty-seven different specimens of macrofungi were collected across Queensland, Australia. Freeze-dried fruiting bodies were sequentially extracted with three solvents: water, ethanol, and hexane. These extracts were tested against representative Gram+ve, Staphylococcus aureus and Gram-ve, Escherichia coli bacteria. Overall water and ethanol extracts were more effective against S. aureus than E. coli, whereas a small number of hexane extracts showed better results for their antimicrobial potential against E. coli at higher concentrations only. Encouraging results were found for a number of macrofungi in the genera Agaricus (Agaricaceae), Amanita (Amanitaceae), Boletus (Boletaceae), Cantharellus (Cantharellaceae), Fomitopsis (Fomitopsidaceae), Hohenbuehelia (Pleurotaceae), Lentinus (Polyporaceae), Ramaria (Gomphaceae), and Strobilomyces (Boletaceae) showing good growth inhibition of the pathogens tested. The present study establishes the antimicrobial potential of a sample of Australian macrofungi that can serve as potential candidates for the development of new antibiotics.


Sigma-1 Receptor Agonists Acting on Aquaporin-Mediated H2O2 Permeability: New Tools for Counteracting Oxidative Stress.
Sigma1 Receptor (S1R) is involved in oxidative stress, since its activation is triggered by oxidative or endoplasmic reticulum stress. Since specific aquaporins (AQP), called peroxiporins, play a relevant role in controlling H2O2 permeability and ensure reactive oxygen species wasted during oxidative stress, we studied the effect of S1R modulators on AQP-dependent water and hydrogen peroxide permeability in the presence and in the absence of oxidative stress. Applying stopped-flow light scattering and fluorescent probe methods, water and hydrogen peroxide permeability in HeLa cells have been studied. Results evidenced that S1R agonists can restore water permeability in heat-stressed cells and the co-administration with a S1R antagonist totally counteracted the ability to restore the water permeability. Moreover, compounds were able to counteract the oxidative stress of HeLa cells specifically knocked down for S1R. Taken together these results support the hypothesis that the antioxidant mechanism is mediated by both S1R and AQP-mediated H2O2 permeability. The finding that small molecules can act on both S1R and AQP-mediated H2O2 permeability opens a new direction toward the identification of innovative drugs able to regulate cell survival during oxidative stress in pathologic conditions, such as cancer and degenerative diseases.


Exposure to and experiences with a computerized decision support intervention in primary care: results from a process evaluation.
Trials evaluating the effects of interventions usually provide little insight into the factors responsible for (lack of) changes in desired outcomes. A process evaluation alongside a trial can shed light on the mechanisms responsible for the outcomes of a trial. The aim of this study was to investigate exposure to and experiences with a computerized decision support system (CDSS) intervention, in order to gain insight into the intervention's impact and to provide suggestions for improvement. A process evaluation was conducted as part of a large-scale cluster-randomized controlled trial investigating the effects of the CDSS NHGDoc on quality of care. Data on exposure to and experiences with the intervention were collected during the trial period among participants in both the intervention and control group - whenever applicable - by means of the NHGDoc server and an electronic questionnaire. Multiple data were analyzed using descriptive statistics. Ninety-nine percent (n = 229) of the included practices generated data for the NHGDoc server and 50 % (n = 116) responded to the questionnaire: both general practitioners (GPs; n = 112; 49 %) and practice nurses (PNs; n = 52; 37 %) participated. The actual exposure to the NHGDoc system and specific heart failure module was limited with 52 % of the GPs and 42 % of the PNs reporting to either never or rarely use the system. Overall, users had a positive attitude towards CDSSs. The most perceived barriers to using NHGDoc were a lack of learning capacity of the system, the additional time and work it requires to use the CDSS, irrelevant alerts, too high intensity of alerts and insufficient knowledge regarding the system. Several types of barriers may have negatively affected the impact of the intervention. Although users are generally positive about CDSSs, a large share of them is insufficiently aware of the functions of NHGDoc and, finds the decision support not always useful or relevant and difficult to integrate into daily practice. In designing CDSS interventions we suggest to more intensely involve the end-users and increase the system's flexibility and learning capacity. To improve implementation a proper introduction of a CDSS among its target group including adequate training is advocated. Clinical trials NCT01773057 .


Glomerella Leaf Spot Caused by a Nonhomothallic Strain of Glomerella cingulata on Highbush Blueberry Nursery Plants in Buenos Aires, Argentina.
In February 2009, irregular-shaped leaf spots affected blueberry (Vaccinium corymbosum L. 'Blue Crisp', 'Misty', and 'Sharp Blue') nursery plants in Buenos Aires. Single-spore cultures on potato dextrose agar and oat agar showed aerial white mycelium that turned light and dark gray, dark brown acervuli with setae, and a salmon-to-orange conidial mass. Septate, dark brown, 62 to 78 μm long setae were abundant in the acervulus. Conidia were unicellular, hyaline, straight, cylindrical, round at the ends, and averaged 15.2 (12.1 to 16.9) × 5.4 (4.9 to 6.2) μm. Dark brown, ovate to clavate, 10.25 × 6.25 μm (9 to 12 × 5 to 8) appressoria with a noticeable pore formed on slides near the edge of the cover glass. Dark subglobose structures were recorded immersed in the culture medium. No asci or ascospores were observed, indicating a nonhomothallic condition. The fungus was identified as Colletotrichum gloeosporioides (Penz.) Penz & Sacc. (teleomorph Glomerella cingulata (Stoneman) Spauld. & H. Schrenk) with traits similar to those already described (1). DNA was obtained from mycelium with a standard DNA extraction kit and the ribosomal, internal transcribed spacer (ITS) 1 and ITS2 regions were PCR amplified and sequenced with primers ITS1 and ITS4 (2). A BLASTN algorithm search revealed 100% identity of the sequence (535 bp long) with G. cingulata/C. gloeosporioides from citrus and mango and one from coffee identified as C. kahawae (GenBank Accession No. JF908919). The nucleotide sequence was deposited in GenBank (Accession No. JQ340087). Pathogenicity was verified on young plants and detached leaves of highbush blueberry 'Emerald', 'Misty', 'O'Neal', and 'Santa Fe', olive (Olea europaea 'Arbequina'), and marketed fruits of apple, mango, orange, and tomato. Disinfected healthy leaves were inoculated with a 9-mm2 mycelial block and incubated at 24°C with 12 h of light. Young plants were infected by placing the disinfected end of the branches within a micropipette tip filled with mycelium and kept under greenhouse conditions. Asymptomatic fruits of apple, mango, orange, and tomato were inoculated by placing a mycelial block on a small wound made on their surface. Detached leaves of highbush blueberry 'Emerald', 'O'Neal', 'Misty', and 'Santa Fe' showed 0.1 to 1.5 × 0.8 to 2 cm necrotic lesions after 3 days, covering 43 to 100% of the 'Emerald' leaf area after 8 days. Young plants of blueberry 'Emerald' and 'Misty' showed 1.5 to 3 cm necrotic lesions, acervuli, a salmon-orange conidial mass, and death of leaves at 25 days. On olive 'Arbequina', leaf necrotic lesions reached 0.1 to 3.5 cm after 5 days. Symptoms developed slowly on infected tomato fruits while inoculated fruits of apple, mango, and orange showed dark brown lesions that measured 2 to 7 × 1 to 3.5 cm at 5 days. No symptoms were observed on controls. The fungus was reisolated from inoculated plant parts. The disease was previously cited in Argentina (3), but to our knowledge, this is the first report of a nonhomothallic strain of G. cingulata from highbush blueberry colonizing and deteriorating fruits of apple, mango, orange, and tomato. References: (1) J. M. E. Mourde. No 315. CMI Descriptions of Pathogenic Fungi and Bacteria. Kew, Surrey, UK, 1971. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990. (3) E.R. Wright et al. OEPP/EPPO Bull. 28:219, 1998.