What is the structure of kaolinite? Reconciling theory and experiment.
Density functional modeling of the crystalline layered aluminosilicate mineral kaolinite is conducted, first to reconcile discrepancies in the literature regarding the exact geometry of the inner and inner surface hydroxyl groups, and second to investigate the performance of selected exchange-correlation functionals in providing accurate structural information. A detailed evaluation of published experimental and computational structures is given, highlighting disagreements in space groups, hydroxyl bond lengths, and bond angles. A major aim of this paper is to resolve these discrepancies through computations. Computed structures are compared via total energy calculations and validated against experimental structures by comparing computed neutron diffractograms, and a final assessment is performed using vibrational spectra from inelastic neutron scattering. The density functional modeling is carried out at a sufficiently high level of theory to provide accurate structure predictions while keeping computational requirements low enough to enable the use of the structures in large-scale calculations. It is found that the best functional to use for efficient density functional modeling of kaolinite using the DMol3 software package is the BLYP functional. The computed structure for kaolinite at 0 K has C1 symmetry, with the inner hydroxyl group angled slightly above the a,b plane and the inner surface hydroxyls aligned close to perpendicular to that plane.


Fish cognition: a primate's eye view.
We provide selected examples from the fish literature of phenomena found in fish that are currently being examined in discussions of cognitive abilities and evolution of neocortex size in primates. In the context of social intelligence, we looked at living in individualized groups and corresponding social strategies, social learning and tradition, and co-operative hunting. Regarding environmental intelligence, we searched for examples concerning special foraging skills, tool use, cognitive maps, memory, anti-predator behaviour, and the manipulation of the environment. Most phenomena of interest for primatologists are found in fish as well. We therefore conclude that more detailed studies on decision rules and mechanisms are necessary to test for differences between the cognitive abilities of primates and other taxa. Cognitive research can benefit from future fish studies in three ways: first, as fish are highly variable in their ecology, they can be used to determine the specific ecological factors that select for the evolution of specific cognitive abilities. Second, for the same reason they can be used to investigate the link between cognitive abilities and the enlargement of specific brain areas. Third, decision rules used by fish could be used as 'null-hypotheses' for primatologists looking at how monkeys might make their decisions. Finally, we propose a variety of fish species that we think are most promising as study objects.


Response of Ancestral Soybean Lines and Commercial Cultivars to Rhizoctonia Root and Hypocotyl Rot.
Rhizoctonia root and hypocotyl rot is a common disease of soybean caused by Rhizoctonia solani. There are no commercial cultivars marketed as resistant to Rhizoctonia root and hypocotyl rot, and only a few sources of partial resistance to this disease have been reported. Ninety ancestral soybean lines, maturity groups (MGs) 000 to X, and 700 commercial cultivars, MGs II to IV, were evaluated for resistance to R. solani under greenhouse conditions. Most of the ancestral lines and cultivars evaluated were susceptible; however, 21 of the ancestral lines and 20 of the commercial cultivars were partially resistant. Of the 21 ancestral lines, CNS, Mandarin (Ottawa), and Jackson are in the pedigree of cultivars previously reported as being partially resistant to R. solani. In an additional study, dry root weights of 21 soybean cultivars were evaluated after inoculation with R. solani. Variation in dry root weight occurred among cultivars, but there was not a significant (P = 0.05) correlation between dry root weight and disease severity.


Suppression of ghost pulses in 40Gbps optical transmission systems with fixed-pattern phase modulation.
Novel fixed-pattern phase modulation scheme is proposed to suppress the IFWM-induced ghost pulses in 40Gbps RZ optical transmission systems through destructing the interference of the IFWM components in symmetric patterns. More than 3dBm launch power margin is achieved. It is a cost-effective method to improve the performance of 40Gbps RZ systems due to greater nonlinearity tolerance.


Long term epidemiological analysis of Citrobacter diversus in a neonatal intensive care unit.
A prolonged outbreak of Citrobacter diversus central nervous system infection among hospitalized term infants, peaking in 1979, ceased with establishment of nurse-patient cohorting. The outbreak was attributed to dissemination of an epidemic strain among infants in an antiquated neonatal intensive care unit. When C. diversus colonization recurred within the new neonatal intensive care unit in 1984, cohorting and bacteriologic surveillance were reinstituted. By utilizing biotypes, plasmid profiles and antibiograms, four different C. diversus strains were identified circulating during 1979. Strains recovered between 1984 and 1988 from neonatal intensive care unit infants were similar to those from community-acquired sources. A strain considered avirulent in 1979 was found causing bacteremia in two infants (one with central nervous system disease) in 1984 to 1988. During cohorting C. diversus acquisition was 0.019/patient-month; after cohorting ceased it was 0.017/patient-month. Multiple source introductions appeared to occur with different C. diversus strains, some causing infant disease. No efficacy of cohorting was evident.


Scientific report on the analysis of the 2-year compulsory intensified monitoring of atypical scrapie.
The European Commission asked EFSA whether the scientific data on the 2-year intensified monitoring in atypical scrapie (AS) outbreaks (2013-2020) provide any evidence on the contagiousness of AS, and whether they added any new knowledge on the epidemiology of AS. An ad hoc data set from intensified monitoring in 22 countries with index case/s of AS in sheep and/or goats (742 flocks from 20 countries, 76 herds from 11 countries) was analysed. No secondary cases were confirmed in goat herds, while 35 secondary cases were confirmed in 28 sheep flocks from eight countries. The results of the calculated design prevalence and of a model simulation indicated that the intensified monitoring had limited ability to detect AS, with no difference between countries with or without secondary cases. A regression model showed an increased, but not statistically significant, prevalence (adjusted by surveillance stream) of secondary cases in infected flocks compared with that of index cases in the non-infected flocks (general population). A simulation model of within-flock transmission, comparing a contagious (i.e. transmissible between animals under natural conditions) with a non-contagious scenario, produced a better fit of the observed data with the non-contagious scenario, in which each sheep in a flock had the same probability of developing AS in the first year of life. Based on the analyses performed, and considering uncertainties and data limitations, it was concluded that there is no new evidence that AS can be transmitted between animals under natural conditions, and it is considered more likely (subjective probability range 50-66%) that AS is a non-contagious, rather than a contagious disease. The analysis of the data of the EU intensified monitoring in atypical scrapie infected flocks/herds confirmed some of the known epidemiological features of AS but identified that major knowledge gaps still remain.


A comprehensive analysis of in vitro and in vivo genetic fitness of Pseudomonas aeruginosa using high-throughput sequencing of transposon libraries.
High-throughput sequencing of transposon (Tn) libraries created within entire genomes identifies and quantifies the contribution of individual genes and operons to the fitness of organisms in different environments. We used insertion-sequencing (INSeq) to analyze the contribution to fitness of all non-essential genes in the chromosome of Pseudomonas aeruginosa strain PA14 based on a library of ∼300,000 individual Tn insertions. In vitro growth in LB provided a baseline for comparison with the survival of the Tn insertion strains following 6 days of colonization of the murine gastrointestinal tract as well as a comparison with Tn-inserts subsequently able to systemically disseminate to the spleen following induction of neutropenia. Sequencing was performed following DNA extraction from the recovered bacteria, digestion with the MmeI restriction enzyme that hydrolyzes DNA 16 bp away from the end of the Tn insert, and fractionation into oligonucleotides of 1,200-1,500 bp that were prepared for high-throughput sequencing. Changes in frequency of Tn inserts into the P. aeruginosa genome were used to quantify in vivo fitness resulting from loss of a gene. 636 genes had <10 sequencing reads in LB, thus defined as unable to grow in this medium. During in vivo infection there were major losses of strains with Tn inserts in almost all known virulence factors, as well as respiration, energy utilization, ion pumps, nutritional genes and prophages. Many new candidates for virulence factors were also identified. There were consistent changes in the recovery of Tn inserts in genes within most operons and Tn insertions into some genes enhanced in vivo fitness. Strikingly, 90% of the non-essential genes were required for in vivo survival following systemic dissemination during neutropenia. These experiments resulted in the identification of the P. aeruginosa strain PA14 genes necessary for optimal survival in the mucosal and systemic environments of a mammalian host.


Apolipoprotein E polymorphism and atherosclerosis.
The apo E locus contributes to determining the variation in plasma cholesterol levels of healthy and diseased populations. It also influences the expression of hyperlipidemia and appears to modulate the susceptibility to atherosclerosis in a complex multifactorial interaction. There is evidence that the presence of apo E2 is protective, whereas that of apo E4 predisposes to coronary artery disease. The burden of proof, however, lies on future, well-designed clinical trials and prospective studies. The study of the biological significance of the apo E polymorphism in humans has emphasized the importance of gene-gene and gene-environment interactions in the pathogenesis of hyperlipidemia and atherosclerosis. The apo E polymorphism involves the coding region of the apo E gene and results in alterations of the gene product which, in turn, either directly or secondarily affect the metabolic fate of the lipoprotein particles. Rapid advances in knowledge over the last decade have provided a metabolic explanation for the observation of the opposite effects of the epsilon 4 and the epsilon 2 alleles on lipoprotein levels. Apo E2 has lower receptor binding affinity which results in delayed clearance of apo E2-bearing lipoprotein particles from plasma. Apo E4 is distributed differently from apo E3 between VLDL and HDL, is degraded more rapidly than apo E3, and may enhance the catabolism of E4-bearing particles, leading to other alterations in lipoprotein metabolism which result in elevated levels of LDL. In view of the significant opposite impacts of the epsilon 4 and the epsilon 2 alleles on plasma LDL cholesterol concentrations, it is evident that determination of the apo E phenotype will become a useful adjunct to the assessment of the cardiovascular risk profile of an individual. In addition, the relationship between the epsilon 2 allele and type III hyperlipoproteinemia provides a valuable model for the study of complex genetic interactions in the pathogenesis of hyperlipidemia. The further study of apo E and its interactions shows great promise for a deeper comprehension of the pathogenesis of atherosclerosis.


Decreased free 1,25-dihydroxycholecalciferol index in patients with the nephrotic syndrome.
The serum concentrations of calcium, phosphorus, parathyroid hormone, vitamin D3 metabolites and their transport protein (DBP) were measured in 18 patients with the nephrotic syndrome (mean daily proteinuria 8.8 g). The glomerular filtration rate was normal in 13 patients while the remaining 5 had a mild degree of renal failure. The serum concentrations of total protein, albumin and DBP were significantly decreased in patients with the nephrotic syndrome. The serum calcium concentration was decreased but the calculated ionized calcium concentration remained normal. The serum concentrations of 25-hydroxycholecalciferol (5.3 +/- 3.1 micrograms/l) and 1,25-dihydroxycholecalciferol [1,25-(OH)2D3 (20 +/- 12 ng/l)] were significantly lower in patients with the nephrotic syndrome and normal glomerular filtration rates than in normal controls (14.4 +/- 4 micrograms/l and 42 +/- 13 ng/l, respectively). The free 1,25-(OH)2D3 index was also significantly below normal (0.9 +/- 0.4 vs. 1.8 +/- 0.4). Total and free 1,25-(OH)2D3 were still further reduced in patients with mild renal failure. The nephrotic syndrome thus results in mild vitamin D depletion with decreased free 1,25-(OH)2D3 concentrations but generally without secondary hyperparathyroidism.


The RENAL (Randomised Evaluation of Normal vs. Augmented Level of Replacement Therapy) study: statistical analysis plan.
The Randomised Evaluation of Normal vs. Augmented Level of Replacement Therapy (RENAL) study is the largest interventional trial ever conducted in patients with acute renal failure. To develop and report a pre-determined statistical analysis plan which the investigators will adhere to in analysing the data from the trial. The data collected by the researchers as part of the trial protocol was reviewed and formally assessed. Information relevant to baseline characteristics was selected and, for each item, statistically relevant descriptive elements were described. Information relevant to the process of care and delivery of prescribed trial therapy was similarly classified and, for each item, appropriate descriptive statistical analysis was planned with appropriate comparison between groups. Finally, trial outcomes were selected, and an appropriate statistical comparison between groups was planned and described. A standard analysis plan for the RENAL trial results was developed, which allows a comprehensive description of baseline characteristics, features of the process of care and trial treatment delivery, and pre-determined statistical assessment of relevant outcome measures in a way that is transparent, available to the public, verifiable and predetermined before the actual analysis of data. We have developed a pre-determined statistical analysis plan for the RENAL trial. This plan will be adhered to in order to avoid introducing any analysis bias associated with prior knowledge of study findings.


Plasma fluctuations as Markovian noise.
Noise theory is used to study the correlations of stationary Markovian fluctuations that are homogeneous and isotropic in space. The relaxation of the fluctuations is modeled by the diffusion equation. The spatial correlations of random fluctuations are modeled by the exponential decay. Based on these models, the temporal correlations of random fluctuations, such as the correlation function and the power spectrum, are calculated. We find that the diffusion process can give rise to the decay of the correlation function and a broad frequency spectrum of random fluctuations. We also find that the transport coefficients may be estimated by the correlation length and the correlation time. The theoretical results are compared with the observed plasma density fluctuations from the tokamak and helimak experiments.


Insulin-like growth factor-I has different effects on myogenin induction and cell cycle progression in human alveolar and embryonal rhabdomyosarcoma cells.
Alveolar rhabdomyosarcoma (RMS) has a much poorer outcome than embryonal RMS. In this study, we found that IGF-I affected the induction of myogenin and cell cycle progression in alveolar RMS cells, but not in embryonal RMS cells. IGF-I enhanced the induction of myogenin protein in alveolar RMS SJ-Rh30 and KP-RMS-MS cells as it did in myoblast C2C12 cells, but not in embryonal RMS RD or KP-RMS-KH cells. IGF-I induction of myogenin protein was blocked by anti-IGF-IR monoclonal antibody alphaIR-3 and the mTOR-specific inhibitor rapamycin. In Rh30mTOR-rr cells, which stably express a rapamycin-resistant mutant mTOR, rapamycin did not inhibit IGF-I induction of myogenin protein. These data suggest that IGF-I induces myogenin in alveolar RMS cells through the IGF-IR/mTOR pathway. In C2C12 cells, IGF-I induces myogenin protein followed by cell cycle arrest leading to myogenic differentiation. IGF-I promoted G1-S cell cycle progression without any signs of terminal differentiation in alveolar RMS cells. On the other hand, IGF-I promoted neither cell cycle arrest nor G1-S cell cycle progression in embryonal RMS cells. In alveolar RMS SJ-Rh30 cells, 4E-BP1, one of two effectors downstream of mTOR, was continuously hyperphosphorylated by IGF-I, whereas in embryonal RMS RD cells, 4E-BP1 was only transiently hyperphosphorylated. These findings suggest that the different effects of IGF-I on myogenin induction and cell cycle progression in alveolar and embryonal RMS cells are due to a difference of phosphorylation status of 4E-BP1. These different responses to IGF-I help to explain immunohistochemical and clinical behavioral differences between alveolar and embryonal RMS.


Vaccine acceptance: Science, policy, and practice in a 'post-fact' world.
Suboptimal vaccination uptake may be associated with outbreaks of vaccine-preventable diseases in many parts of the world. Researchers and practitioners working on improving vaccine acceptance and uptake gathered together for the fifth annual meeting on vaccine acceptance, organized by the Fondation Mérieux at its conference centre in Veyrier-du-Lac, France, to share their experiences in building, improving and sustaining vaccine confidence and uptake. The importance and value of truly listening to people and seeking to understand the perspectives of vaccine hesitant people was emphasized throughout the meeting. The benefits of social marketing, which can be used to influence behavior that benefit individuals and communities for the greater social good, and its integration into strategies aimed at improving vaccine acceptance and uptake, were discussed. Healthcare professionals (HCPs) need tools and training to help them engage effectively in vaccination acceptance conversations with parents and other patients. Two potential tools, motivational interviewing (MI) and AIMS (Announce, Inquire, Mirror, Secure), were presented. Examples of MI approaches that have successfully improved vaccination acceptance and uptake included a project in Canada aimed at parents just after the birth of their baby. The role of mandates to increase vaccination uptake in the short-term was discussed, but to achieve sustainable vaccination uptake this must be complemented with other strategies. These annual meetings have led to the creation of an informal community of practice that facilitates cross-pollination between the various disciplines and different settings of those involved in this area of research and implementation. It was agreed that we must continue our efforts to promote vaccine acceptance and thus increase vaccination uptake, by fostering more effective vaccination communication, monitoring of the media conversation on vaccination, designing and rigorously evaluating targeted interventions, and surveillance of vaccine acceptance and uptake with pertinent, reliable measures.


Why is tick-borne encephalitis increasing? A review of the key factors causing the increasing incidence of human TBE in Sweden.
The highest annual incidence of human tick-borne encephalitis (TBE) in Sweden ever recorded by the Swedish Institute for Communicable Disease Control (SMI) occurred last year, 2011. The number of TBE cases recorded during 2012 up to 6th August 2012 indicates that the incidence for 2012 could exceed that of 2011. In this review of the ecology and epidemiology of TBE in Sweden our main aim is to analyse the possible reasons behind the gradually increasing incidence of human TBE during the last 20 years. The main TBE virus (TBEV) vector to humans in Sweden is the nymphal stage of the common tick Ixodes ricinus. The main mode of transmission and maintenance of TBEV in the tick population is considered to be when infective nymphs co-feed with uninfected but infectible larvae on rodents. In most locations the roe deer, Capreolus capreolus is the main host for the reproducing adult I. ricinus ticks. The high number of roe deer for more than three decades has resulted in a very large tick population. Deer numbers have, however, gradually declined from the early 1990s to the present. This decline in roe deer numbers most likely made the populations of small rodents, which are reservoir-competent for TBEV, gradually more important as hosts for the immature ticks. Consequently, the abundance of TBEV-infected ticks has increased. Two harsh winters in 2009-2011 caused a more abrupt decline in roe deer numbers. This likely forced a substantial proportion of the "host-seeking" ticks to feed on bank voles (Myodes glareolus), which at that time suddenly had become very numerous, rather than on roe deer. Thus, the bank vole population peak in 2010 most likely caused many tick larvae to feed on reservoir-competent rodents. This presumably resulted in increased transmission of TBEV among ticks and therefore increased the density of infected ticks the following year. The unusually warm, humid weather and the prolonged vegetation period in 2011 permitted nymphs and adult ticks to quest for hosts nearly all days of that year. These weather conditions stimulated many people to spend time outdoors in areas where they were at risk of being attacked by infective nymphs. This resulted in at least 284 human cases of overt TBE. The tick season of 2012 also started early with an exceptionally warm March. The abundance of TBEV-infective "hungry" ticks was presumably still relatively high. Precipitation during June and July was rich and will lead to a "good mushroom season". These factors together are likely to result in a TBE incidence of 2012 similar to or higher than that of 2011.


Co-transmission of conduct problems with attention-deficit/hyperactivity disorder: familial evidence for a distinct disorder.
Common disorders of childhood and adolescence are attention-deficit/hyperactivity disorder (ADHD), oppositional defiant disorder (ODD) and conduct disorder (CD). For one to two cases in three diagnosed with ADHD the disorders may be comorbid. However, whether comorbid conduct problems (CP) represents a separate disorder or a severe form of ADHD remains controversial. We investigated familial recurrence patterns of the pure or comorbid condition in families with at least two children and one definite case of DSM-IV ADHDct (combined-type) as part of the International Multicentre ADHD Genetics Study (IMAGE). Using case diagnoses (PACS, parental account) and symptom ratings (Parent/Teacher Strengths and Difficulties [SDQ], and Conners Questionnaires [CPTRS]) we studied 1009 cases (241 with ADHDonly and 768 with ADHD + CP), and their 1591 siblings. CP was defined as > or =4 on the SDQ conduct-subscale, and T > or = 65, on Conners' oppositional-score. Multinomial logistic regression was used to ascertain recurrence risks of the pure and comorbid conditions in the siblings as predicted by the status of the cases. There was a higher relative risk to develop ADHD + CP for siblings of cases with ADHD + CP (RRR = 4.9; 95%CI: 2.59-9.41); p < 0.001) than with ADHDonly. Rates of ADHDonly in siblings of cases with ADHD + CP were lower but significant (RRR = 2.9; 95%CI: 1.6-5.3, p < 0.001). Children with ADHD + CP scored higher on the Conners ADHDct symptom-scales than those with ADHDonly. Our finding that ADHD + CP can represent a familial distinct subtype possibly with a distinct genetic etiology is consistent with a high risk for cosegregation. Further, ADHD + CP can be a more severe disorder than ADHDonly with symptoms stable from childhood through adolescence. The findings provide partial support for the ICD-10 distinction between hyperkinetic disorder (F90.0) and hyperkinetic conduct disorder (F90.1).


Acupuncture for chronic nonpulsatile tinnitus: A randomized clinical trial.
There is challenge to find an effective treatment for tinnitus. Few studies were done on the effects of acupuncture on tinnitus. This study evaluated the effect of acupuncture on chronic non-pulsatile tinnitus. This randomized double-blind clinical trial was conducted from December 2014 to September 2015. Patients suffering from chronic non-pulsatile tinnitus were randomly allocated into two groups: acupuncture vs. placebo. They were treated in 15 sessions and at the end of the fifteenth sessions and 3 weeks after completion of the treatment, visual analog scale (VAS) for tinnitus loudness and tinnitus severity index (TSI) questionnaires were completed. The case group included 26 males and 18 females, and in the control group there were 27 males and 17 females: with mean age of 49.11±1.07 and 55.20±8.33 years, respectively (p=0.005). TSI and VAS before treatment were 43.84±2.81 and 9.56±0.43 in cases and 43.52±2.94 and 9.54±0.45 in controls, respectively. Both measures improved after 15 sessions in cases to 24.82±1.04 and 2.88±0.33, and to 33.16±1.24 and 7.86±0.23 in controls. The changes of TSI and VAS were significant in all groups (p<0.001). TSI and VAS in acupuncture group were lower than placebo group in each session (p<0.001), except TSI in the tenth session (p=0.392). Acupuncture is effective in reducing the loudness and severity of tinnitus and can be a useful treatment for nonpulsatile chronic tinnitus.


Aqueous humor-borne factor upregulates Bcl-2 expression in corneal endothelial cells.
To determine if factors present in the aqueous humor (AH) protect the corneal endothelium from apoptosis. Mouse and human corneal endothelial cells were cultured in vitro, and apoptosis was induced by nutrient deprivation. AH and supernatant from iris/ciliary body (I/CB) cell cultures were tested for their effect on corneal endothelial cell apoptosis. The effect of I/CB supernatant on Fas, Bax, and Bcl-2 gene transcription was evaluated by Northern blotting. I/CB supernatant was subjected to proteinase analysis to identify the apoptosis inhibitory factor(s). Rabbit AH and supernatant from mouse I/CB cell cultures inhibited the apoptosis of mouse and human immortalized corneal endothelial cell lines. The inhibition of apoptosis was associated with an upregulation of Bcl-2 gene transcription and Bcl-2 protein expression. Bax gene expression was not significantly affected by I/CB cell supernatant. Induction of apoptosis by stimulation of the Fas receptor was unaffected by I/CB cell supernatant. Protease analyses indicated that the apoptosis inhibitory factor was a protein. The inability of corneal endothelial cells to undergo mitosis renders the corneal endothelium vulnerable to loss of physiological function through cellular attrition. However, a protein(s) produced by I/CB cells and present in the AH, upregulates Bcl-2 gene transcription and protects the corneal endothelial cells from apoptosis.


Furosin, an ellagitannin, suppresses RANKL-induced osteoclast differentiation and function through inhibition of MAP kinase activation and actin ring formation.
Phenolic compounds including tannins and flavonoids have been implicated in suppression of osteoclast differentiation/function and prevention of bone diseases. However, the effects of hydrolysable tannins on bone metabolism remain to be elucidated. In this study, we found that furosin, a hydrolysable tannin, markedly decreased the differentiation of both murine bone marrow mononuclear cells and Raw264.7 cells into osteoclasts, as revealed by the reduced number of tartrate resistant acid phosphatase (TRAP)-positive multinucleated cells and decreased TRAP activity. Furosin appears to target at the early stage of osteoclastic differentiation while having no cytotoxic effect on osteoclast precursors. Analysis of the inhibitory mechanisms of furosin revealed that it inhibited the receptor activator of nuclear factor-kappaB ligand (RANKL)-induced activation of p38 mitogen-activated protein kinase (p38MAPK) and c-Jun N-terminal kinase (JNK)/activating protein-1 (AP-1). Furthermore, furosin reduced resorption pit formation in osteoclasts, which was accompanied by disruption of the actin rings. Taken together, these results demonstrate that naturally occurring furosin has an inhibitory activity on both osteoclast differentiation and function through mechanisms involving inhibition of the RANKL-induced p38MAPK and JNK/AP-1 activation as well as actin ring formation.


The pragmatic language abilities of children with ADHD following a play-based intervention involving peer-to-peer interactions.
Children with Attention Deficit Hyperactivity Disorder (ADHD) commonly experience significant pragmatic language deficits which put them at risk of developing emotional and social difficulties. This study aimed to examine the pragmatic language exhibited in a peer-to-peer interaction between the children with ADHD and their playmates following a pilot play-based intervention. Participants were children (aged 5-11 years) diagnosed as having ADHD (n = 14) and their self-selected typically-developing playmate. Pragmatic language was measured using the Pragmatic Protocol (PP) and the Structured Multidimensional Assessment Profiles (S-MAPs). Children's structural language was also screened and compared against their pragmatic language skills pre-post play-based intervention. The pragmatic language of children with ADHD improved significantly from pre-post intervention as measured by both the PP and S-MAPs. Both children with and without structural language difficulties improved significantly from pre- to post-intervention using S-MAPs; only children with structural language difficulties improved significantly using PP. The findings support the notion that pragmatic skills may improve following a play-based intervention that is characterized by didactic social interaction. As pragmatic language is a complex construct, it is proposed that clinicians and researchers reconsider the working definition of pragmatic language and the operationalization thereof in assessments.


Hypoxia-inducible factor 1 (HIF-1) pathway.
Hypoxia-inducible factor 1 (HIF-1) is a basic helix-loop-helix-PAS domain transcription factor that is expressed in all metazoan organisms and is composed of HIF-1alpha and HIF-1beta subunits. Under hypoxic conditions, HIF-1 regulates the transcription of hundreds of genes in a cell type-specific manner. The HIF-1alpha subunit is regulated by O2-dependent hydroxylation of proline residue 402, 564, or both, by prolyl hydroxylase domain protein 2 (PHD2), which promotes binding of the von Hippel-Lindau protein (VHL), leading to ubiquitination and proteasomal degradation; and O2-dependent hydroxylation of asparagine residue 803 by factor inhibiting HIF-1 (FIH-1), which blocks the binding of the 300-kilodalton coactivator protein (p300) and CREB binding protein (CBP). The hydroxylation reactions, which utilize O2 and alpha-ketoglutarate as substrates and generate CO2 and succinate as by-products, provide a mechanism by which changes in cellular oxygenation are transduced to the nucleus as changes in HIF-1 activity. Hydroxylase activity is inhibited in the presence of low concentrations of O2, high concentrations of tricarboxylic acid cycle intermediates (isocitrate, oxaloacetate, succinate, or fumarate), or chelators of Fe(II). Receptor for activated C kinase 1 (RACK1) competes with heat shock protein 90 (HSP90) for binding to HIF-1alpha and mediates O2-independent ubiquitination and proteasomal degradation. A growing number of proteins and small molecules have been identified that regulate HIF-1 activity by modulating the physical or functional interaction of PHD2, VHL, FIH-1, RACK1, or HSP90 with HIF-1alpha.


Training safer orthopedic surgeons. Construct validation of a virtual-reality simulator for hip fracture surgery.
Virtual-reality (VR) simulation in orthopedic training is still in its infancy, and much of the work has been focused on arthroscopy. We evaluated the construct validity of a new VR trauma simulator for performing dynamic hip screw (DHS) fixation of a trochanteric femoral fracture. 30 volunteers were divided into 3 groups according to the number of postgraduate (PG) years and the amount of clinical experience: novice (1-4 PG years; less than 10 DHS procedures); intermediate (5-12 PG years; 10-100 procedures); expert (> 12 PG years; > 100 procedures). Each participant performed a DHS procedure and objective performance metrics were recorded. These data were analyzed with each performance metric taken as the dependent variable in 3 regression models. There were statistically significant differences in performance between groups for (1) number of attempts at guide-wire insertion, (2) total fluoroscopy time, (3) tip-apex distance, (4) probability of screw cutout, and (5) overall simulator score. The intermediate group performed the procedure most quickly, with the lowest fluoroscopy time, the lowest tip-apex distance, the lowest probability of cutout, and the highest simulator score, which correlated with their frequency of exposure to running the trauma lists for hip fracture surgery. This study demonstrates the construct validity of a haptic VR trauma simulator with surgeons undertaking the procedure most frequently performing best on the simulator. VR simulation may be a means of addressing restrictions on working hours and allows trainees to practice technical tasks without putting patients at risk. The VR DHS simulator evaluated in this study may provide valid assessment of technical skill.


Impact of headache on sickness absence and utilisation of medical services: a Danish population study.
The aim was to study the extent and type of health service utilisation, medication habits, and sickness absence due to the primary headaches. This was a cross sectional epidemiological survey of headache disorders in a general population. Headache was diagnosed according to a structured interview and a neurological examination using the criteria of the International Headache Society. A random sample of 25-64 year-old individuals was drawn from the Danish National Central Person Registry. All subjects were living in the Copenhagen County. 740 subjects participated (76% of the sample); 119 had migraine and 578 had tension type headache. Among subjects with migraine 56% had, at some time, consulted their general practitioner because of the migraine. The corresponding percentage among subjects with tension type headache was 16. One or more specialists had been consulted by 16% of migraine sufferers and by 4% of subjects with tension type headache. The consultation rates of chiropractors and physiotherapists were 5-8%. Hospital admissions and supplementary laboratory investigations due to headache were rare (< 3%). Half of the migraine sufferers and 83% of subjects with tension type headache in the previous year had managed with at least one type of drug in the current year. Acetylsalicylic acid preparations and paracetamol were the most commonly used analgesics. Prophylaxis of migraine was used by 7%. In the preceding year 43% of employed migraine sufferers and 12% of employed subjects with tension type headache had missed one or more days of work because of headache. Most common was 1-7 days off work. The total loss of workdays per year due to migraine in the general population was estimated at 270 days per 1000 persons. For tension type headache the corresponding figure was 820. Women were more likely to consult a practitioner than men, whereas no significant sex difference emerged as regards absenteeism from work. The impact of the headache disorders on work performance in the general population is substantial, and the disorders merit increased attention.


Patterns of longitudinal brain atrophy in the logopenic variant of primary progressive aphasia.
The logopenic variant of primary progressive aphasia (PPA) is characterised by impaired sentence repetition and word retrieval difficulties. Post mortem studies, amyloid imaging and CSF tau/Aβ measurements suggest Alzheimer's disease (AD) pathology as the underlying cause. Relatively little is known about patterns of progression in patients with the logopenic variant of PPA. 21 patients (3 with post mortem confirmation of AD and 5 with positive amyloid PIB-PET scans) were studied with longitudinal T1-weighted MR imaging (mean interscan interval 1.2years) using volumetric analysis and voxel-based morphometry (VBM). Baseline imaging showed asymmetrical (left greater than right) involvement of the posterior superior temporal and inferior parietal lobes as well as posterior cingulate and medial temporal lobes. The whole brain rate of volume loss was 2.0% per year with a greater rate of left hemisphere atrophy (2.3%/year) than right hemisphere (1.6%/year). Longitudinal VBM analysis showed increasing involvement of other areas in the left hemisphere (temporal, parietal, frontal and caudate) and atrophy of areas in the right hemisphere that had been involved earlier in the disease in the left hemisphere, particularly posterior cingulate/precuneus. With disease progression there was worsening of anomia, sentence repetition and sentence comprehension but consistent with the spread of imaging changes also deficits in single word comprehension, single word repetition and verbal memory. This study shows that the logopenic variant of PPA remains an asymmetrical disease, with spread through the left hemisphere language network but also involvement to a lesser degree of regions in the right hemisphere that mirror the earlier left hemisphere changes.


Biomechanics of shear-sensitive adhesion in climbing animals: peeling, pre-tension and sliding-induced changes in interface strength.
Many arthropods and small vertebrates use adhesive pads for climbing. These biological adhesives have to meet conflicting demands: attachment must be strong and reliable, yet detachment should be fast and effortless. Climbing animals can rapidly and reversibly control their pads' adhesive strength by shear forces, but the mechanisms underlying this coupling have remained unclear. Here, we show that adhesive forces of stick insect pads closely followed the predictions from tape peeling models when shear forces were small, but strongly exceeded them when shear forces were large, resulting in an approximately linear increase of adhesion with friction. Adhesion sharply increased at peel angles less than ca 30°, allowing a rapid switch between attachment and detachment. The departure from classic peeling theory coincided with the appearance of pad sliding, which dramatically increased the peel force via a combination of two mechanisms. First, partial sliding pre-stretched the pads, so that they were effectively stiffer upon detachment and peeled increasingly like inextensible tape. Second, pad sliding reduces the thickness of the fluid layer in the contact zone, thereby increasing the stress levels required for peeling. In combination, these effects can explain the coupling between adhesion and friction that is fundamental to adhesion control across all climbing animals. Our results highlight that control of adhesion is not solely achieved by direction-dependence and morphological anisotropy, suggesting promising new routes for the development of controllable bio-inspired adhesives.


Lysosomal sequestration (trapping) of lipophilic amine (cationic amphiphilic) drugs in immortalized human hepatocytes (Fa2N-4 cells).
Lipophilic (logP > 1) and amphiphilic drugs (also known as cationic amphiphilic drugs) with ionizable amines (pKa > 6) can accumulate in lysosomes, a process known as lysosomal trapping. This process contributes to presystemic extraction by lysosome-rich organs (such as liver and lung), which, together with the binding of lipophilic amines to phospholipids, contributes to the large volume of distribution characteristic of numerous cardiovascular and central nervous system drugs. Accumulation of lipophilic amines in lysosomes has been implicated as a cause of phospholipidosis. Furthermore, elevated levels of lipophilic amines in lysosomes can lead to high organ-to-blood ratios of drugs that can be mistaken for active drug transport. In the present study, we describe an in vitro fluorescence-based method (using the lysosome-specific probe LysoTracker Red) to identify lysosomotropic agents in immortalized hepatocytes (Fa2N-4 cells). A diverse set of compounds with various physicochemical properties were tested, such as acids, bases, and zwitterions. In addition, the partitioning of the nonlysosomotropic atorvastatin (an anion) and the lysosomotropics propranolol and imipramine (cations) were quantified in Fa2N-4 cells in the presence or absence of various lysosomotropic or nonlysosomotropic agents and inhibitors of lysosomal sequestration (NH4Cl, nigericin, and monensin). Cellular partitioning of propranolol and imipramine was markedly reduced (by at least 40%) by NH4Cl, nigericin, or monensin. Lysosomotropic drugs also inhibited the partitioning of propranolol by at least 50%, with imipramine partitioning affected to a lesser degree. This study demonstrates the usefulness of immortalized hepatocytes (Fa2N-4 cells) for determining the lysosomal sequestration of lipophilic amines.


A Comparison of Risk and Protective Factors Related to Depressive Symptoms among American Indian and Caucasian Older Adults.
Despite efforts to reduce health disparities, many American Indian and Alaska Native (AI/AN) populations, including older adults, experience elevated levels of depression and associated suicide. Although adverse childhood experiences (ACEs) and social support are well-documented risk and protective factors for depression in the general population, little is known about AI/AN populations, especially older adults. The purpose of this study was to examine factors related to depression among a sample of AI older adults in the Midwest. Data were collected using a self-administered survey completed by 479 AI and Caucasian respondents over the age of 50. The survey included standardized measures such as the Geriatric Depression Scale-Short Form, ACE Questionnaire, and Multidimensional Scale of Perceived Social Support. Hierarchical multivariate regression analyses were conducted to evaluate the main hypotheses of the study. Results indicate that two dimensions of ACEs (childhood neglect and household dysfunction) were positively associated with depressive symptoms; social support was negatively associated with depressive symptoms. Perceived health and living alone were also significant predictors of depressive symptoms. ACEs may play a significant role in depression among AI/AN populations across the life course and into old age. Social support offers a promising mechanism to bolster resilience among AI/AN older adults.


Participation in sports in relation to adolescent growth and development.
Puberty is defined by physical growth, development of secondary sexual characteristics, and maturation of psychosocial skills. The initiation and rate of progression of pubertal events varies among adolescents, but pubertal changes occur in a predictable stepwise manner. Factors including individual differences in physical and psychosocial development, stage of development based on age (early, middle, and late), and the rate of pubertal development, may all contribute to the way in which adolescents experience sports activities. During adolescence, gender differences also become more apparent and may significantly impact sports participation. As practitioners evaluate overall development and adolescent readiness for sports participation, they should consider the different areas of development including: somatic, neurologic, cognitive, psychosocial-function in an integrated and interdependent approach.


Evaluation of Thick-Slab Overlapping MIP Images of Contrast-Enhanced 3D T1-Weighted CUBE for Detection of Intracranial Metastases: A Pilot Study for Comparison of Lesion Detection, Interpretation Time, and Sensitivity with Nonoverlapping CUBE MIP, CUBE, and Inversion-Recovery-Prepared Fast-Spoiled Gradient Recalled Brain Volume.
Early and accurate identification of cerebral metastases is important for prognostication and treatment planning although this process is often time consuming and labor intensive, especially with the hundreds of images associated with 3D volumetric imaging. This study aimed to evaluate the benefits of thick-slab overlapping MIPs constructed from contrast-enhanced T1-weighted CUBE (overlapping CUBE MIP) for the detection of brain metastases in comparison with traditional CUBE and inversion-recovery prepared fast-spoiled gradient recalled brain volume (IR-FSPGR-BRAVO) and nonoverlapping CUBE MIP. A retrospective review of 48 patients with cerebral metastases was performed at our institution from June 2016 to October 2017. Brain MRIs, which were acquired on multiple 3T scanners, included gadolinium-enhanced T1-weighted IR-FSPGR-BRAVO and CUBE, with subsequent generation of nonoverlapping CUBE MIP and overlapping CUBE MIP. Two blinded radiologists identified the total number and location of metastases on each image type. The Cohen κ was used to determine interrater agreement. Sensitivity, interpretation time, and lesion contrast-to-noise ratio were assessed. Interrater agreement for identification of metastases was fair-to-moderate for all image types (κ = 0.222-0.598). The total number of metastases identified was not significantly different across the image types. Interpretation time for CUBE MIPs was significantly shorter than for CUBE and IR-FSPGR-BRAVO, saving at least 50 seconds per case on average (P < .001). The mean lesion contrast-to-noise ratio for both CUBE MIPs was higher than for IR-FSPGR-BRAVO. The mean contrast-to-noise ratio for small lesions (<4 mm) was lower for nonoverlapping CUBE MIP (1.55) than for overlapping CUBE MIP (2.35). For both readers, the sensitivity for lesion detection was high for all image types but highest for overlapping CUBE MIP and CUBE (0.93-0.97). This study suggests that the use of overlapping CUBE MIP or nonoverlapping CUBE MIP for the detection of brain metastases can reduce interpretation time without sacrificing sensitivity, though the contrast-to-noise ratio of lesions is highest for overlapping CUBE MIP.


Synthesis, anticonvulsant and toxicity evaluation of 2-(1H-indol-3-yl)acetyl-N-(substituted phenyl)hydrazine carbothioamides and their related heterocyclic derivatives.
A series of new 5-(1H-indol-3-yl)methyl-4-(substituted aryl)-2,4-dihydro-3H-1,2,4-triazole-3-thiones (4a-g), 5-(1H-indol-3-yl)methyl-N-(substituted aryl)-1,3,4-oxadiazol-2-amines (5a-g) and 5-(1H-indol-3-yl)methyl-N-(substituted aryl)-1,3,4-thiadiazol-2-amines (6a-g) were prepared by treating 2-(1H-indol-3-yl)acetyl-N-(substituted phenyl) hydrazine carbothioamides (3a-g) with suitable reagents. All the newly synthesized compounds were screened for their anticonvulsant activity in the MES model and were compared with the standard drugs phenytoin sodium and carbamazepine. Out of the twenty-one compounds studied, 4b, 4e, 4f, 5b, 5d, 5g, 6b, 6d and 6e showed comparable MES activity to phenytoin and carbamazepine after 0.5 h. Compound 5b showed to be more potent than carbamazepine after 4 h. Compounds 4a, 4c, 4d, 5a, 5c, 5e, 5f, 6f and 6g showed lower neurotoxicity than phenytoin.


A novel Pyrin-Associated Autoinflammation with Neutrophilic Dermatosis mutation further defines 14-3-3 binding of pyrin and distinction to Familial Mediterranean Fever.
Pyrin-Associated Autoinflammation with Neutrophilic Dermatosis (PAAND) is a recently described monogenic autoinflammatory disease. The causal p.S242R MEFV mutation disrupts a binding motif of the regulatory 14-3-3 proteins within pyrin. Here, we investigate a family with clinical features consistent with PAAND in whom the novel p.E244K MEFV mutation, located in the +2 site of the 14-3-3 binding motif in pyrin, has been found. Multiplex cytokine analyses were performed on p.E244K patient and control serum. Peripheral blood mononuclear cells were stimulated ex vivo with lipopolysaccharide (LPS). In vitro, inflammasome complex formation was evaluated by flow cytometry of Apoptosis-associated Speck-like protein containing a Caspase recruitment domain (ASC) specks. Interleukin-1β (IL-1β) and IL-18 production was quantified by ELISA. The ability of the p.E244K pyrin mutation to interact with 14-3-3 was assessed by immunoprecipitation. PAAND p.E244K patient serum displayed a different cytokine profile compared with patients with Familial Mediterranean Fever (FMF). In overexpression models, p.E244K pyrin was associated with decreased 14-3-3 binding and increased ASC speck formation. THP-1 monocytes expressing PAAND pyrin mutations demonstrated spontaneous caspase-1-dependent IL-1β and IL-18 secretion, as well as cell death, which were significantly greater than those of wild-type and the FMF-associated mutation p.M694V. In PAAND, disruption of the +2 position of a 14-3-3 binding motif in pyrin results in its constitutive activation, with spontaneous production of IL-1β and IL-18, associated with inflammatory cell death. The altered serum cytokine profile may explain the different clinical features exhibited by PAAND patients compared with those with FMF.


Autologous chondrocyte implantation versus ACI using 3D-bioresorbable graft for the treatment of large full-thickness cartilage lesions of the knee.
In autologous chondrocyte implantation (ACI), the periosteum patch which is sutured over the cartilage defect has been identified as a major source of complications such as periosteal hypertrophy. In the present retrospective study, we compared midterm results of first-generation ACI with a periosteal patch to second generation ACI using a biodegradable collagen fleece (BioSeed-C) in 82 patients suffering from chronic posttraumatic and degenerative cartilage lesions of the knee. Clinical outcome was assessed in 42 patients of group 1 and in 40 patients of group 2 before implantation of the autologous chondrocytes and at a minimum follow-up of 2 years using the ICRS score, the modified Cincinnati score and the Lysholm score. Although patients treated with BioSeed-C had more previous surgical procedures on their respective knees, highly significant improvements (P < 0.001) were assessed in both groups at comparable outcome levels: the ICRS score improved from grade D (poor) preoperatively to grade C (fair); the modified Cincinnati knee score from 3.26 to 6.4 (group 1) and 3.3 and 6.88 (group 2). Lysholm score improved from 33 to 70 points (group 1) and from 47 to 78 points (group 2), respectively. Revision surgery was due to symptomatic periosteal hypertrophy (n = 4), graft failure (n = 3), plica syndrome (n = 2) synovectomy (n = 1) (group 1); and graft failure (n = 2), debridement (n = 1), synovectomy (n = 2) (group 2). These results suggest that BioSeed-C is an equally effective treatment option for focal degenerative chondral lesions of the knee in this challenging and complex patient profile.


Efficacy and safety of anti-tuberculosis drugs in HIV-positive patients: a prospective study.
To assess the efficacy and safety of anti-tuberculosis drugs in HIV-positive patients at a tertiary care teaching hospital. As a part of an ongoing study of opportunistic infections (OIs) in HIV-positive patients, drug treatment in patients suffering from tuberculosis was assessed to determine its efficacy and safety. Based on prevalence data for last three years, a purposive sampling of study population was carried out in this observational, prospective, single centre study. Tuberculosis (TB) was the most common OI observed. The selected patients were followed up for a period of one year to evaluate the clinical course and outcome of OIs, and the efficacy and safety of drugs used was checked. Tuberculosis was observed in 89 out of 134 enrolled patients. These included 79 adults and 10 children. Males (66.2%) were commonly affected. Extra pulmonary TB (73%) was the most common manifestation with abdominal TB observed in 55 (61.7%) patients. All patients were treated in accordance with the Revised National Tuberculosis Control Programme (RNTCP) guidelines as recommended by National AIDS Control Organization (NACO), India. Outcome of TB was assessable in 70 patients. Majority (82.8%) of the patients were cured, while 12 patients (17.1%) died during the course of treatment. A total of 149 ADRs were observed in 67 (75.2%) patients. Majority of ADRs (n = 147) were non-serious and did not warrant a change in therapy. Discoloration of urine was the most common ADR observed. TB is the most common opportunistic infection in HIV-positive patients with abdominal TB being the most common manifestation. RNTCP and NACO guidelines are adhered to in these patients. Anti-tuberculosis drugs are well tolerated and effective in majority of the patients.


Complementation of a deletion in the rubella virus p150 nonstructural protein by the viral capsid protein.
Rubella virus (RUB) replicons with an in-frame deletion of 507 nucleotides between two NotI sites in the P150 nonstructural protein (DeltaNotI) do not replicate (as detected by expression of a reporter gene encoded by the replicon) but can be amplified by wild-type helper virus (Tzeng et al., Virology 289:63-73, 2001). Surprisingly, virus with DeltaNotI was viable, and it was hypothesized that this was due to complementation of the NotI deletion by one of the virion structural protein genes. Introduction of the capsid (C) protein gene into DeltaNotI-containing replicons as an in-frame fusion with a reporter gene or cotransfection with both DeltaNotI replicons and RUB replicon or plasmid constructs containing the C gene resulted in replication of the DeltaNotI replicon, confirming the hypothesis that the C gene was the structural protein gene responsible for complementation and demonstrating that complementation could occur either in cis or in trans. Approximately the 5' one-third of the C gene was necessary for complementation. Mutations that prevented translation of the C protein while minimally disturbing the C gene sequence abrogated complementation, while synonymous codon mutations that changed the C gene sequence without affecting the amino acid sequence at the 5' end of the C gene had no effect on complementation, indicating that the C protein, not the C gene RNA, was the moiety responsible for complementation. Complementation occurred at a basic step in the virus replication cycle, because DeltaNotI replicons failed to accumulate detectable virus-specific RNA.


An electromyographic evaluation of subdividing active-assistive shoulder elevation exercises.
Active-assistive range of motion exercises to gain shoulder elevation have been subdivided into gravity-minimized and upright-assisted exercises, yet no study has evaluated differences in muscular demands. Compared with gravity-minimized exercises, upright-assisted exercises will generate larger electromyographic (EMG) activity. Compared with all active-assistive exercises, upright active forward elevation will generate more EMG activity. Controlled laboratory study. Fifteen healthy individuals participated in this study. The supraspinatus, infraspinatus, and anterior deltoid were evaluated. The independent variables were 11 exercises performed in random order. The dependent variable was the maximum EMG amplitude of each muscle that was normalized to a maximal voluntary isometric contraction (MVIC). Each muscle demonstrated significant differences between exercises (P < .001), with upright active forward elevation producing the greatest EMG for all muscles (95% confidence interval [CI], 12% to 50% MVIC). The orders of exercise varied by muscle, but the 5 gravity-minimized exercises always generated the lowest EMG activity. The upright-assisted exercises (95% CI, 23% to 42% MVIC) for the anterior deltoid generated more EMG activity than did the gravity-minimized exercises (95% CI, 9% to 21% MVIC) (P < .05). The infraspinatus and supraspinatus demonstrated increasing trends in EMG activity from gravity minimized to upright assisted (P > .05). The results suggest a clear distinction between gravity-minimized exercises and upright-assisted exercises for the anterior deltoid but not for the supraspinatus and infraspinatus. Between the 2 types of assisted exercises, the results also suggest a clear distinction in terms of active elevation of the arm for the supraspinatus and anterior deltoid but not for the infraspinatus. Muscle activation levels increase as support is removed, but subdivision of active-assistive range of motion to protect the supraspinatus and infraspinatus may not be necessary.


Folic acid and vitamin B(12) supplementation lowers plasma homocysteine but has no effect on serum bone turnover markers in elderly women: a randomized, double-blind, placebo-controlled trial.
An elevated homocysteine level is a newly recognized risk factor for osteoporosis. Older individuals may have elevated homocysteine levels due to inadequate folate intake and/or lower absorption of vitamin B(12). The aim of this study was to determine whether there is an impact of folic acid and vitamin B(12) supplementation on homocysteine levels and, subsequently, on bone turnover markers in older women with mildly to moderately elevated homocysteine levels. It is hypothesized that supplementation with folic acid and vitamin B(12) will improve homocysteine levels and, in turn, positively modify bone turnover markers in this population. This randomized, double-blind, placebo-controlled trial included 31 women (65 to 93 years) with homocysteine levels greater than 10 μmol/L. Participants were randomly assigned to receive either a daily folic acid (800 μg) and vitamin B(12) (1000 μg) (n = 17) or a matching placebo (n = 14) for 4 months. The results showed significantly lower homocysteine concentrations in the vitamin group compared to the placebo group (10.6 vs 18.5 μmol/L, P = .007). No significant difference in serum alkaline phosphatase or C-terminal cross-linking telopeptide of type I collagen was found between the vitamin and placebo groups before or after supplementation. The use of folic acid and vitamin B(12) as a dietary supplement to improve homocysteine levels could be beneficial for older women, but additional research must be conducted in a larger population and for a longer period to determine if there is an impact of supplementation on bone turnover markers or other indicators of bone health.


Radiation-induced radicals in glucose-1-phosphate. I. Electron paramagnetic resonance and electron nuclear double resonance analysis of in situ X-irradiated single crystals at 77 K.
Electron magnetic resonance analysis of radiation-induced defects in dipotassium glucose-1-phosphate dihydrate single crystals in situ X-irradiated and measured at 77 K shows that at least seven different carbon-centered radical species are trapped. Four of these (R1-R4) can be fully or partly characterized in terms of proton hyperfine coupling tensors. The dominant radical (R2) is identified as a C1-centered species, assumedly formed by a scission of the sugar-phosphate junction and the concerted formation of a carbonyl group at the neighboring C2 carbon. This structure is chemically identical to a radical recently identified in irradiated sucrose single crystals. Radical species R1 and R4 most likely are C3- and C6-centered species, respectively, both formed by a net hydrogen abstraction. R3 is suggested to be chemically similar to but geometrically different from R4. Knowledge of the identity of the sugar radicals present at 77 K provides a first step in elucidating the formation mechanism of the phosphoryl radicals previously detected after X-irradiation at 280 K. In paper II, the chemical identity, precise conformation, and possible formation mechanisms of these radical species are investigated by means of DFT calculations and elementary insight into the radiation chemistry of sugar and sugar derivatives is obtained.


Synaptic connectivity and neuronal morphology: two sides of the same coin.
Neurons often possess elaborate axonal and dendritic arbors. Why do these arbors exist and what determines their form and dimensions? To answer these questions, I consider the wiring up of a large highly interconnected neuronal network, such as the cortical column. Implementation of such a network in the allotted volume requires all the salient features of neuronal morphology: the existence of branching dendrites and axons and the presence of dendritic spines. Therefore, the requirement of high interconnectivity is, in itself, sufficient to account for the existence of these features. Moreover, the actual lengths of axons and dendrites are close to the smallest possible length for a given interconnectivity, arguing that high interconnectivity is essential for cortical function.


Extension of diffuse low-grade gliomas beyond radiological borders as shown by the coregistration of histopathological and magnetic resonance imaging data.
OBJECTIVE Magnetic resonance imaging tends to underestimate the extent of diffuse low-grade gliomas (DLGGs). With the aim of studying the presence of tumor cells outside the radiological border, the authors developed a method of correlating MRI findings with histological data in patients with suspected DLGGs in whom en bloc resections were performed. METHODS Five patients with suspected DLGG suitable for en bloc resection were recruited from an ongoing prospective study. Sections of the entire tumor were immunostained with antibodies against mutated IDH1 protein (IDH1-R132H). Magnetic resonance images were coregistered with corresponding IDH1 images. The growth pattern of tumor cells in white and gray matter was assessed in comparison with signal changes on corresponding MRI slices. RESULTS Neuropathological assessment revealed DLGG in 4 patients and progression to WHO Grade III glioma in 1 patient. The tumor core consisted of a high density of IDH1-R132H-positive tumor cells and was located in both gray and white matter. Tumor cells infiltrated along the peripheral fibers of the white matter tracts. In all cases, tumor cells were found outside the radiological tumor border delineated on T2-FLAIR MRI sequences. CONCLUSIONS The authors present a new method for the coregistration of histological and radiological characteristics of en bloc-removed infiltrative brain tumors that discloses tumor invasion at the radiological tumor borders. This technique can be applied to evaluate the sensitivity of alternative imaging methods to detect scattered tumor cells at tumor borders. Accurate methods for detection of infiltrative tumor cells will improve the possibility of performing radical tumor resection. In future studies, the method could also be used for in vivo studies of tumor invasion.


Male genital mutilation in the high-mountain goblin spider, Unicorn catleyi.
Male genital mutilation is a common mechanism by which males reduce sperm competition by plugging female insemination ducts with different parts of its own genital system. This behavior is frequent in many spider families but is uncommon in Haplogynae. The reproductive biology of Dysderoidea is not well studied and the data is fragmentary; male genital mutilation has been reported only for one species of Oonopidae. This study provides evidence of male genital mutilation in Unicorn catleyi Platnick and Brescovit (Araneae: Oonopidae). Pieces of the embolus were found in the female posterior receptaculum. This behavior is a strategy used by the males in order to guarantee their paternity and not for escape from female attacks as has been reported for other species of Araneae, since cannibalism is unlikely in this species. The presence of embolus in the posterior receptaculum suggests this is the first place where sperm is received. The similarity of the female genitalia of U. catleyi to those of Orsolobidae, along with sclerotization of the seminal duct in the male copulatory bulb that is also present in Orchestina, Xiombarg, and Orsolobidae, provide strong evidence of the basal position of this genus in the family Oonopidae.


Representative taxa in field trials for environmental risk assessment of genetically modified maize.
When assessing the benefits and risks of transgenic crops, one consideration is their relative effects on non-target arthropod (NTA) abundance and functions within agroecosystems. Several laboratory and field trials have been conducted in Spain since the late 1990s to assess this issue. A consideration in the design of field trials is whether it is necessary to sample most NTAs living in the crop or only representative taxa that perform main ecological functions and have a good capacity to detect small changes in their abundance. Small changes in the field abundance of an effective representative taxon should be detectable using standard experimental protocols. The ability of a species to reveal differences across treatments may be analysed by examining the detectable treatment effects for surveyed non-target organisms. Analysis of data from several NTAs recorded in 14 field trials conducted over 10 years using complete block designs allowed us to select a number of representative taxa capable of detecting changes in the density or activity of arthropod herbivores, predators, parasitoids and decomposers in transgenic and non-transgenic maize varieties. The most suitable NTA as representative taxa (with detectable treatment effects below 50%) included leafhoppers among arthropod herbivores, Orius spp., Araneae, and Carabidae among predators, chalcidids, particularly the family Mymaridae, among parasitoids and Chloropidae as decomposer. Details of sampling techniques for each sampled taxa and their advantages and disadvantages are discussed. It is concluded that abundance of taxa is the most influential factor determining their capacity to detect changes caused by genetically modified varieties.


Workshop on partial breast irradiation: state of the art and the science, Bethesda, MD, December 8-10, 2002.
Breast conserving surgery followed by radiation therapy has been accepted as an alternative to mastectomy in the management of patients with early-stage breast cancer. Over the past decade there has been increasing interest in a variety of radiation techniques designed to treat only the portion of the breast deemed to be at high risk for local recurrence (partial-breast irradiation [PBI]) and to shorten the duration of treatment (accelerated partial-breast irradiation [APBI]). To consider issues regarding the equivalency of the various radiation therapy approaches and to address future needs for research, quality assurance, and training, the National Cancer Institute, Division of Cancer Treatment and Diagnosis, Radiation Research Program, hosted a Workshop on PBI in December 2002. Although 5- to 7-year outcome data on patients treated with PBI and APBI are now becoming available, many issues remain unresolved, including clinical and pathologic selection criteria, radiation dose and fractionation and how they relate to the standard fractionation for whole breast irradiation, appropriate target volume, local control within the untreated ipsilateral breast tissue, and overall survival. This Workshop report defines the issues in relation to PBI and APBI, recommends parameters for consideration in clinical trials and for reporting of results, serves to enhance dialogue among the advocates of the various radiation techniques, and emphasizes the importance of education and training in regard to results of PBI and APBI as they become emerging clinical treatments.


Verification of a Method for Measuring Parkinson's Disease Related Temporal Irregularity in Spiral Drawings.
Parkinson's disease (PD) is a progressive movement disorder caused by the death of dopamine-producing cells in the midbrain. There is a need for frequent symptom assessment, since the treatment needs to be individualized as the disease progresses. The aim of this paper was to verify and further investigate the clinimetric properties of an entropy-based method for measuring PD-related upper limb temporal irregularities during spiral drawing tasks. More specifically, properties of a temporal irregularity score (TIS) for patients at different stages of PD, and medication time points were investigated. Nineteen PD patients and 22 healthy controls performed repeated spiral drawing tasks on a smartphone. Patients performed the tests before a single levodopa dose and at specific time intervals after the dose was given. Three movement disorder specialists rated videos of the patients based on the unified PD rating scale (UPDRS) and the Dyskinesia scale. Differences in mean TIS between the groups of patients and healthy subjects were assessed. Test-retest reliability of the TIS was measured. The ability of TIS to detect changes from baseline (before medication) to later time points was investigated. Correlations between TIS and clinical rating scores were assessed. The mean TIS was significantly different between healthy subjects and patients in advanced groups (p-value = 0.02). Test-retest reliability of TIS was good with Intra-class Correlation Coefficient of 0.81. When assessing changes in relation to treatment, TIS contained some information to capture changes from Off to On and wearing off effects. However, the correlations between TIS and clinical scores (UPDRS and Dyskinesia) were weak. TIS was able to differentiate spiral drawings drawn by patients in an advanced stage from those drawn by healthy subjects, and TIS had good test-retest reliability. TIS was somewhat responsive to single-dose levodopa treatment. Since TIS is an upper limb high-frequency-based measure, it cannot be detected during clinical assessment.


Cost-effective evaluation and treatment for carotid disease.
To compare carotid endarterectomy (CEA) based solely on Duplex ultrasonography (DU) with CEA based on DU and arteriography. Retrospective case series analysis in a regional tertiary care center. Consecutive sample of 194 patients undergoing 218 CEAs from January 1, 1993, through June 30, 1995, with either preoperative DU plus arteriography (165 CEAs) or DU only (53 CEAs). Concordance of the 2 diagnostic imaging techniques and influence of these on the conduct of surgery, surgical outcome, and resource cost. There was agreement (kappa = 0.85) between DU and arteriography in the detection of a carotid occlusion or a stenosis greater than 45%. Arteriography demonstrated 26 aortic arch branch lesions (15.8%), 22 intracranial abnormalities (13.3%), and 6 type C ulcers (3.6%), in addition to 1 nonoccluded internal carotid artery (ICA) (0.61%) and 1 contralateral severe ICA stenosis (0.61%) inaccurately estimated by Duplex. These findings prompted 3 changes (1.8%) in surgical therapy, including 2 decisions in favor of CEA and 1 subclavian-carotid bypass added to CEA. There was no difference in the stroke and death rate for CEA based solely on DU compared with CEA based on DU and arteriography (P = .43). The mean total hospital cost was $5,534 for DU only CEA vs $7,608 for DU plus arteriogram CEA (mean difference = $2,074, P < .01). The addition of carotid arteriography to a diagnostic Duplex ultrasound study that already suggested the need for CEA did not change the operative plan in 98% (162/165) of the cases. Carotid endarterectomy based solely on DU is appropriate and cost-effective.


Healthy recovery: A stepped wedge cluster randomised controlled trial of a healthy lifestyle intervention for people attending residential alcohol and other drug treatment.
Unhealthy lifestyle behaviours contribute to the poor health of people attending alcohol and other drug (AOD) treatment. Healthy Recovery is an 8-session group-based intervention that targets smoking, diet and physical inactivity as part of an integrated healthy lifestyle approach. The current study aimed to examine the effectiveness of Healthy Recovery when delivered within residential AOD treatment settings. The study design was a stepped-wedge cluster randomised trial. Participants were 151 current smokers attending residential AOD programs provided by the Australian Salvation Army (n = 71, Control condition; n = 80, Intervention condition). The primary outcome was number of cigarettes smoked per day. Secondary outcomes examined other smoking behaviours (7-day point prevalence, use of nicotine replacement therapy [NRT]), diet (servings and variety of fruit and vegetables), and physical activity. The mean number of cigarettes smoked per day was significantly lower in the Intervention condition at 2-, 5-, and 8-month follow-up. There were also significant differences in favour of the Intervention condition for number of quit attempts, use of NRT and variety of fruit. There were no other significant differences for other dietary or physical activity variables. Healthy Recovery had a positive impact on smoking behaviours. Future research should consider strategies to further promote smoking cessation (e.g. promoting longer-term use of NRT), as well as addressing physical activity and dietary behaviours. The introduction of broader organisational approaches (e.g. smoke free policies, organised group exercise and cooking activities) might help to enhance healthy lifestyle approaches within AOD treatment settings.


Mitochondrial DNA variation among Anopheles albimanus populations.
Barriers to gene flow between Pacific and Atlantic coast populations of Anopheles albimanus were reported in an earlier study of variation in the intergenic spacer of the nuclear ribosomal DNA. We examined the distribution of mitochondrial DNA haplotypes among A. albimanus populations to test for gene flow barriers with an independent genetic marker. A region of the NADH dehydrogenase subunit 5 gene was amplified by the polymerase chain reaction (PCR) in 1,105 mosquitoes collected from 16 locations in Guatemala and in single collections from Mexico, Honduras, Nicaragua, Costa Rica, Panama, Colombia, and Venezuela. The PCR products were tested for variation using single strand conformation polymorphism analysis and 45 haplotypes were detected. Haplotype frequencies did not vary between coasts in Guatemala. Populations within approximately 200 km of one another were panmictic. However, at distances > 200 km, FST and geographic distances were correlated suggesting that populations are isolated by distance.


Derisking the Cu-Mediated 18F-Fluorination of Heterocyclic Positron Emission Tomography Radioligands.
Molecules labeled with fluorine-18 (18F) are used in positron emission tomography to visualize, characterize and measure biological processes in the body. Despite recent advances in the incorporation of 18F onto arenes, the development of general and efficient approaches to label radioligands necessary for drug discovery programs remains a significant task. This full account describes a derisking approach toward the radiosynthesis of heterocyclic positron emission tomography (PET) radioligands using the copper-mediated 18F-fluorination of aryl boron reagents with 18F-fluoride as a model reaction. This approach is based on a study examining how the presence of heterocycles commonly used in drug development affects the efficiency of 18F-fluorination for a representative aryl boron reagent, and on the labeling of more than 50 (hetero)aryl boronic esters. This set of data allows for the application of this derisking strategy to the successful radiosynthesis of seven structurally complex pharmaceutically relevant heterocycle-containing molecules.


Endometrial preparation methods for frozen-thawed embryo transfer are associated with altered risks of hypertensive disorders of pregnancy, placenta accreta, and gestational diabetes mellitus.
What were the risks with regard to the pregnancy outcomes of patients who conceived by frozen-thawed embryo transfer (FET) during a hormone replacement cycle (HRC-FET)? The patients who conceived by HRC-FET had increased risks of hypertensive disorders of pregnancy (HDP) and placenta accreta and a reduced risk of gestational diabetes mellitus (GDM) in comparison to those who conceived by FET during a natural ovulatory cycle (NC-FET). Previous studies have shown that pregnancy and live-birth rates after HRC-FET and NC-FET are comparable. Little has been clarified regarding the association between endometrium preparation and other pregnancy outcomes. A retrospective cohort study of patients who conceived after HRC-FET and those who conceived after NC-FET was performed based on the Japanese assisted reproductive technology registry in 2014. The pregnancy outcomes were compared between NC-FET (n = 29 760) and HRC-FET (n = 75 474) cycles. Multiple logistic regression analyses were performed to investigate the potential confounding factors. The pregnancy rate (32.1% vs 36.1%) and the live birth rate among pregnancies (67.1% vs 71.9%) in HRC-FET cycles were significantly lower than those in NC-FET cycles. A multiple logistic regression analysis showed that pregnancies after HRC-FET had increased odds of HDPs [adjusted odds ratio, 1.43; 95% confidence interval (CI), 1.14-1.80] and placenta accreta (adjusted odds ratio, 6.91; 95% CI, 2.87-16.66) and decreased odds for GDM (adjusted odds ratio, 0.52; 95% CI, 0.40-0.68) in comparison to pregnancies after NC-FET. Our study was retrospective in nature, and some cases were excluded due to missing data. The implication of bias and residual confounding factors such as body mass index, alcohol consumption, and smoking habits should be considered in other observational studies. Pregnancies following HRC-FET are associated with higher risks of HDPs and placenta accreta and a lower risk of GDM. The association between the endometrium preparation method and obstetrical complication merits further attention. No funding was obtained for this work. The authors declare no conflicts of interest in association with the present study. Not applicable.


Effect of counselling on the psychiatric morbidity associated with mastectomy.
A controlled trial was conducted to determine whether counselling by a specialist nurse prevented the psychiatric morbidity associated with mastectomy and breast cancer. Seventy-five patients were counselled by the nurse and monitored during follow-up, while 77 patients received only the care normally given by the surgical unit. Counselling failed to prevent morbidity, but the nurse's regular monitoring of the women's progress led her to recognise and refer 76% of those who needed psychiatric help. Only 15% of the control group whose condition warranted help were recognised and referred. Consequently, 12 to 18 months after mastectomy there was much less psychiatric morbidity in the counselled group (12%) than in the control group (39%). These findings highlight the high degree of psychiatric morbidity in patients who have undergone mastectomy and indicate the need to find ways of reducing this morbidity.


Intervertebral cages for degenerative spinal diseases.
Interbody fusion techniques have been used for many years for the treatment of a variety of lumbar spine diagnoses. Part of the interest in increasing methods of interbody fusion has stemmed from concern that posterior fusion alone may allow micro-motion, which may generate pain in a ruptured or degenerated disc. Stabilization of the anterior segment led to the development of interbody fusion cages. These devices were designed to stabilize the spine while bony ingrowth from the vertebrae to the bone graft occurred. There are a variety of techniques for cage insertion, including open and laparoscopic techniques anteriorly, and open posterior approach. A lateral approach for cage placement has also been reported. The purpose of this paper is to present a review of the literature on lumbar intervertebral fusion performed using interbody cages. The reported results for these procedures vary, but in general the majority of patients have had favorable results. The complications are similar to those encountered with traditional interbody fusion procedures using bone grafts. There is a learning curve associated with the procedures, particularly with the laparoscopic techniques. Appropriate training for the spine surgeon as well as the access surgeon is important. There is a great deal of disparity in reports on using the cages as stand-alone devices as well as on laparoscopic approaches. Overall, the use of interbody cages for fusion appears to be a viable treatment, yielding good results. Fusion cages appear to have a role in spine care; however, as with any procedure, patient selection and proper training of the surgeon are critical.


The Behavior of Metropolis-Coupled Markov Chains When Sampling Rugged Phylogenetic Distributions.
Bayesian phylogenetic inference relies on the use of Markov chain Monte Carlo (MCMC) to provide numerical approximations of high-dimensional integrals and estimate posterior probabilities. However, MCMC performs poorly when posteriors are very rugged (i.e., regions of high posterior density are separated by regions of low posterior density). One technique that has become popular for improving numerical estimates from MCMC when distributions are rugged is Metropolis coupling (MC$^3$). In MC$^3$, additional chains are employed to sample flattened transformations of the posterior and improve mixing. Here, we highlight several underappreciated behaviors of MC3. Notably, estimated posterior probabilities may be incorrect but appear to converge, when individual chains do not mix well, despite different chains sampling trees from all relevant areas in tree space. Counterintuitively, such behavior can be more difficult to diagnose with increased numbers of chains. We illustrate these surprising behaviors of MC$^3$ using a simple, non-phylogenetic example and phylogenetic examples involving both constrained and unconstrained analyses. To detect and mitigate the effects of these behaviors, we recommend increasing the number of independent analyses and varying the temperature of the hottest chain in current versions of Bayesian phylogenetic software. Convergence diagnostics based on the behavior of the hottest chain may also help detect these behaviors and could form a useful addition to future software releases.


Test, Treat, Track, Test, and Treat Active Surveillance toward Elimination of Schistosomiasis: A Feasibility Study.
We assessed the feasibility of using a test, treat, track, test, and treat (5T) active surveillance strategy to identify and treat individuals with schistosomiasis in three very low-prevalence villages in Kafr El Sheikh Governorate, Egypt. Primary index cases (PICs) were identified using the point-of-care circulating cathodic antigen (POC-CCA) assay in schools, in rural health units (retesting individuals with positive Kato-Katz examinations over the previous 6 months), and at potential water transmission sites identified by PICs and field observations. Primary cases identified potential second-generation cases-people with whom they shared water activities-who were then tracked, tested, and treated if infected. Those sharing water activities with second-generation cases were also tested. The yield of PICs from the three venues were 128 of 3,576 schoolchildren (3.6%), 42 of 696 in rural health units (6.0%), and 83 of 1,156 at water contact sites (7.2%). There were 118 second- and 19 third-generation cases identified. Persons testing positive were treated with praziquantel. Of 388 persons treated, 368 (94.8%) had posttreatment POC-CCA tests 3-4 weeks after treatment, and 81.8% (301) became negative. The 67 persons remaining positive had negative results after a second treatment. Therefore, all those found positive, treated, and followed up were negative following one or two treatments. Analysis of efforts as expressed in person-hours indicates that 4,459 person-hours were required for these 5T activities, with nearly 65% of that time spent carrying out interviews, treatments, and evaluations following treatment. The 5T strategy appears feasible and acceptable as programs move toward elimination.


Simulation of synaptic depression, posttetanic potentiation, and presynaptic facilitation of synaptic potentials from sensory neurons mediating gill-withdrawal reflex in Aplysia.
The defensive gill-withdrawal reflex in Aplysia has proven to be an attractive system for analyzing the neural mechanisms underlying simple forms of learning such as habituation, sensitization, and classic conditioning. Previous studies have shown that habituation is associated with synaptic depression and sensitization with presynaptic facilitation of transmitter release from sensory neurons mediating the reflex. The synaptic depression, in turn, is associated with a decrease in Ca2+ currents in the sensory neurons, whereas presynaptic facilitation with increased Ca2+ currents produced indirectly by a decrease in a novel serotonergic sensitive K+ current. The present work represents an initial quantitative examination of the extent to which these mechanisms account for each of these types of synaptic plasticity. To address these issues a lumped parameter mathematical model of the sensory neuron release process was constructed. Major components of this model include Ca2+-channel inactivation, Ca2+-mediated neurotransmitter release and mobilization, and readily releasable and upstream feeding pools of neurotransmitter. In the model, release of neurotransmitter has a linear function of Ca2+ concentration and is not affected directly by residual Ca2+. The model not only simulates the data of synaptic depression and recovery from depression, but also qualitatively predicts other features of neurotransmitter release that it was not designed to fit. These include features of synaptic depression with high and low levels of transmitter release, posttetanic potentiation, a steep relationship between action potential duration and transmitter release, enhanced release produced by broadening the sensory neuron action potential (presynaptic facilitation), and dramatic synaptic depression with two closely spaced tetraethylammonium (TEA) spikes. The model cannot account fully for synaptic depression with empirically observed somatic Ca2+-current kinetics. Rather a large component of synaptic depression is due to reduction to the pools of releasable neurotransmitter (depletion). In the model when spike durations are greater than 15-20 ms, spike broadening produces little facilitation. However, when spike durations are more physiological, spike broadening leads to enhanced transmitter release.


Diet and environment shape fecal bacterial microbiota composition and enteric pathogen load of grizzly bears.
Diet and environment impact the composition of mammalian intestinal microbiota; dietary or health disturbances trigger alterations in intestinal microbiota composition and render the host susceptible to enteric pathogens. To date no long term monitoring data exist on the fecal microbiota and pathogen load of carnivores either in natural environments or in captivity. This study investigates fecal microbiota composition and the presence of pathogenic Escherichia coli and toxigenic clostridia in wild and captive grizzly bears (Ursus arctos) and relates these to food resources consumed by bears. Feces were obtained from animals of two wild populations and from two captive animals during an active bear season. Wild animals consumed a diverse diet composed of plant material, animal prey and insects. Captive animals were fed a regular granulated diet with a supplement of fruits and vegetables. Bacterial populations were analyzed using quantitative PCR. Fecal microbiota composition fluctuated in wild and in captive animals. The abundance of Clostridium clusters I and XI, and of C. perfringens correlated to regular diet protein intake. Enteroaggregative E. coli were consistently present in all populations. The C. sordellii phospholipase C was identified in three samples of wild animals and for the first time in Ursids. This is the first longitudinal study monitoring the fecal microbiota of wild carnivores and comparing it to that of captive individuals of the same species. Location and diet affected fecal bacterial populations as well as the presence of enteric pathogens.


Microarray profiling of diaphyseal bone of rats suffering from hypervitaminosis A.
Vitamin A is the only known compound that produces spontaneous fractures in rats. In an effort to resolve the molecular mechanism behind this effect, we fed young male rats high doses of vitamin A and performed microarray analysis of diaphyseal bone with and without marrow after 1 week, i.e., just before the first fractures appeared. Of the differentially expressed genes in cortical bone, including marrow, 98% were upregulated. In contrast, hypervitaminotic cortical bone without marrow showed reduced expression of 37% of differentially expressed genes. Gene ontology (GO) analysis revealed that only samples containing bone marrow were associated with a GO term, which principally represented extracellular matrix. This is consistent with the histological findings of increased endosteal/marrow osteoblast number. Fourteen genes, including Cyp26b1, which is known to be upregulated by vitamin A, were selected and verified by real-time PCR. In addition, immunohistochemical staining of bone sections confirmed that the bone-specific molecule osteoadherin was upregulated. Further analysis of the major gene-expression changes revealed apparent augmented Wnt signaling in the sample containing bone marrow but reduced Wnt signaling in cortical bone. Moreover, induced expression of hypoxia-associated genes was found only in samples containing bone marrow. Together, these results highlight the importance of compartment-specific analysis of bone and corroborate previous observations of compartment-specific effects of vitamin A, with reduced activity in cortical bone but increased activity in the endosteal/marrow compartment. We specifically identify potential key osteoblast-, Wnt signaling-, and hypoxia-associated genes in the processes leading to spontaneous fractures.


Isolation and characterisation of a cDNA clone for human apolipoprotein CI and assignment of the gene to chromosome 19.
We have synthesised a mixed oligonucleotide 17 bases long and used it to isolate cDNA clones for apolipoprotein CI (apo CI) from an adult liver cDNA library. The partial sequence of one of these clones confirms its identity. We have used this probe and Southern blotting techniques to identify the human apo CI gene in DNA from a series of rodent X human somatic cell hybrids. Our results provide evidence for the assignment of this gene to human chromosome 19.


An investigation of the expression and adhesin function of H7 flagella in the interaction of Escherichia coli O157 : H7 with bovine intestinal epithelium.
Enterohaemorrhagic Escherichia coli O157 : H7 is a bacterial pathogen that can cause haemorrhagic colitis and haemolytic uremic syndrome. In the primary reservoir host, cattle, the terminal rectum is the principal site of E. coli O157 colonization. In this study, bovine terminal rectal primary epithelial cells were used to examine the role of H7 flagella in epithelial adherence. Binding of a fliC(H7) mutant O157 strain to rectal epithelium was significantly reduced as was binding of the flagellated wild-type strain following incubation with H7-specific antibodies. Complementation of fliC(H7) mutant O157 strain with fliC(H7) restored the adherence to wild-type levels; however, complementation with fliC(H6) did not restore it. High-resolution ultrastructural and imunofluorescence studies demonstrated the presence of abundant flagella forming physical contact points with the rectal epithelium. Binding to terminal rectal epithelium was specific to H7 by comparison with other flagellin types tested. In-cell Western assays confirmed temporal expression of flagella during O157 interaction with epithelium, early expression was suppressed during the later stages of microcolony and attaching and effacing lesion formation. H7 flagella are expressed in vivo by individual bacteria in contact with rectal mucosa. Our data demonstrate that the H7 flagellum acts as an adhesin to bovine intestinal epithelium and its involvement in this crucial initiating step for colonization indicates that H7 flagella could be an important target in intervention strategies.


Distinct reaction pathways followed upon reduction of oxy-heme oxygenase and oxy-myoglobin as characterized by Mössbauer spectroscopy.
Activation of O(2) by heme-containing monooxygenases generally commences with the common initial steps of reduction to the ferrous heme and binding of O(2) followed by a one-electron reduction of the O(2)-bound heme. Subsequent steps that generate reactive oxygen intermediates diverge and reflect the effects of protein control on the reaction pathway. In this study, Mössbauer and EPR spectroscopies were used to characterize the electronic states and reaction pathways of reactive oxygen intermediates generated by 77 K radiolytic cryoreduction and subsequent annealing of oxy-heme oxygenase (HO) and oxy-myoglobin (Mb). The results confirm that one-electron reduction of (Fe(II)-O(2))HO is accompanied by protonation of the bound O(2) to generate a low-spin (Fe(III)-O(2)H(-))HO that undergoes self-hydroxylation to form the alpha-meso-hydroxyhemin-HO product. In contrast, one-electron reduction of (Fe(II)-O(2))Mb yields a low-spin (Fe(III)-O(2)(2-))Mb. Protonation of this intermediate generates (Fe(III)-O(2)H(-))Mb, which then decays to a ferryl complex, (Fe(IV)=O(2-))Mb, that exhibits magnetic properties characteristic of the compound II species generated in the reactions of peroxide with heme peroxidases and with Mb. Generation of reactive high-valent states with ferryl species via hydroperoxo intermediates is believed to be the key oxygen-activation steps involved in the catalytic cycles of P450-type monooxygenases. The Mössbauer data presented here provide direct spectroscopic evidence supporting the idea that ferric-hydroperoxo hemes are indeed the precursors of the reactive ferryl intermediates. The fact that a ferryl intermediate does not accumulate in HO underscores the determining role played by protein structure in controlling the reactivity of reaction intermediates.


Nesfatin-1--role as possible new potent regulator of food intake.
Nesfatin-1 is an 82 amino acid peptide recently discovered in the brain which is derived from nucleobindin2 (NUCB2), a protein that is highly conserved across mammalian species. Nesfatin-1 has received much attention over the past two years due to its reproducible food intake-reducing effect that is linked with recruitment of other hypothalamic peptides regulating feeding behavior. A growing amount of evidence also supports that various stressors activate fore- and hindbrain NUCB2/nesfatin-1 circuitries. In this review, we outline the central nervous system distribution of NUCB2/nesfatin-1, and recent developments on the peripheral expression of NUCB2/nesfatin-1, in particular its co-localization with ghrelin in gastric X/A-like cells and insulin in ss-cells of the endocrine pancreas. Functional studies related to the characteristics of nesfatin-1's inhibitory effects on dark phase food intake are detailed as well as the central activation of NUCB2/nesfatin-1 immunopositive neurons in the response to psychological, immune and visceral stressors. Lastly, potential clinical implications of targeting NUCB2/nesfatin-1 signaling and existing gaps in knowledge to ascertain the role and mechanisms of action of nesfatin-1 are presented.


Population History and Mitochondrial Genetic Substructure of the Rama Amerindians from Nicaragua.
The Rama are a coastal population from southern Nicaragua who in large part were able to resist, at least for a time, the cultural changes and social reorganization brought on by colonial and modern influences. Historical information leaves the Rama origins and biological relationships with nearby extinct and extant groups ambiguous. The objective of this study was to examine the internal genetic microdifferentiation based on the first hypervariable region of the mitochondrial DNA (mtDNA) from a sample of approximately 20% of the population, and to expand the few available historical and anthropological data on the Rama by exploring the effects of cultural practices and historical events on genetic structure, providing an integrative perspective on the Rama genetic history. When considering differences in the spatial distribution and genetic diversity of the mtDNA haplotypes together with historical information on the Rama, a noteworthy pattern emerges. (a) Haplotypes are differentially distributed among a central Rama community (Punta Águila) compared with the other five peripheral communities (analysis of molecular variance: FCT = 0.10, p < 0.001), and their distribution is consistent with the historical relocation of this population after their split from Punta Gorda in the 18th century. (b) Differential genetic signatures found among central and peripheral Rama communities resemble two population histories: one of stability (haplogroup A2) and other of expansion (haplogroup B2), supporting the possibility that these patterns of genetic microdifferentiation between central and peripheral populations resulted from the 18th-century unification in southern Nicaragua of the Rama and a group of Voto migrants from Costa Rica that later split off and moved to the Bay of Bluefields.