From Pancreatic β-Cell Gene Networks to Novel Therapies for Type 1 Diabetes.
Completion of the Human Genome Project enabled a novel systems- and network-level understanding of biology, but this remains to be applied for understanding the pathogenesis of type 1 diabetes (T1D). We propose that defining the key gene regulatory networks that drive β-cell dysfunction and death in T1D might enable the design of therapies that target the core disease mechanism, namely, the progressive loss of pancreatic β-cells. Indeed, many successful drugs do not directly target individual disease genes but, rather, modulate the consequences of defective steps, targeting proteins located one or two steps downstream. If we transpose this to the T1D situation, it makes sense to target the pathways that modulate the β-cell responses to the immune assault-in relation to signals that may stimulate the immune response (e.g., HLA class I and chemokine overexpression and/or neoantigen expression) or inhibit the invading immune cells (e.g., PDL1 and HLA-E expression)-instead of targeting only the immune system, as it is usually proposed. Here we discuss the importance of a focus on β-cells in T1D, lessons learned from other autoimmune diseases, the "alternative splicing connection," data mining, and drug repurposing to protect β-cells in T1D and then some of the initial candidates under testing for β-cell protection.


Preparing soldiers for the stress of combat.
Protracted use of stressors during military training courses does not necessarily enhance a Soldier?s ability to regulate stress on the battlefield. Extensive stress during training can be a contributing factor to suboptimal neurologic and overall long-term health. Prolonged high-stress military training programs, as well as extended duration combat deployments, should be comprehensively scrutinized for opportunities to preserve health and increase combat effectiveness. Contemporary research in neuroscience and psychology can provide insight into training techniques that can be used to control stress and optimize performance in combat. Physical fitness training programs can elevate the stress threshold. Extensive situational training can also inoculate Soldiers to specific combat stressors. Training methods such as these will enable Soldiers to achieve higher levels of performance while under enemy fire and are encouraged for units deploying to combat. combat stress, military training, military deployment, physical training, post-traumatic stress disorder, sleep deprivation, stress inoculation training.


Exposure of Anopheles mosquitoes to trypanosomes reduces reproductive fitness and enhances susceptibility to Plasmodium.
During a blood meal, female Anopheles mosquitoes are potentially exposed to diverse microbes in addition to the malaria parasite, Plasmodium. Human and animal African trypanosomiases are frequently co-endemic with malaria in Africa. It is not known whether exposure of Anopheles to trypanosomes influences their fitness or ability to transmit Plasmodium. Using cell and molecular biology approaches, we found that Trypanosoma brucei brucei parasites survive for at least 48h after infectious blood meal in the midgut of the major malaria vector, Anopheles coluzzii before being cleared. This transient survival of trypanosomes in the midgut is correlated with a dysbiosis, an alteration in the abundance of the enteric bacterial flora in Anopheles coluzzii. Using a developmental biology approach, we found that the presence of live trypanosomes in mosquito midguts also reduces their reproductive fitness, as it impairs the viability of laid eggs by affecting their hatching. Furthermore, we found that Anopheles exposure to trypanosomes enhances their vector competence for Plasmodium, as it increases their infection prevalence. A transcriptomic analysis revealed that expression of only two Anopheles immune genes are modulated during trypanosome exposure and that the increased susceptibility to Plasmodium was microbiome-dependent, while the reproductive fitness cost was dependent only on the presence of live trypanosomes but was microbiome independent. Taken together, these results demonstrate multiple effects upon Anopheles vector competence for Plasmodium caused by eukaryotic microbes interacting with the host and its microbiome, which may in turn have implications for malaria control strategies in co-endemic areas.


Frequency and Predictors of Renal Transplantation Among Patients Rendered Surgically Anephric for Sporadic Renal Cancer.
To assess the frequency of renal transplantation in patients rendered surgically anephric during treatment of renal cancers as well as the clinicopathologic factors associated with receipt of transplantation. A retrospective review was conducted to identify patients rendered surgically anephric between 2001 and 2016 due to cancer in both renal units or cancer in an anatomically or functionally solitary kidney. Patient demographics, comorbidities, and cancer features were compared between patients who subsequently received a renal transplantation and those who did not. Time-to-event analysis was used to compare time to transplantation across varied identified parameters. Among 27 patients rendered anephric, 4 (15%) received a renal transplantation over a median follow-up of 21.6 months (interquartile range 7.2, 53.3). All transplanted patients were less than 70 years of age and had cT1a renal parenchymal mass at the time of nephrectomy. No patient undergoing completion nephrectomy for upper tract urothelial carcinoma received transplantation. Patients who were evaluated by the transplant service prior to nephrectomy were more likely to eventually undergo transplantation (60% vs 5%; P < .01). On time-to-event analyses, a cT1a renal parenchymal mass (P < .01) and a pre-nephrectomy transplant evaluation (P < .01) were associated with receipt of a transplant. Patients rendered anephric via nephrectomy for cancer are more likely to receive renal transplantation if they are less than 70 years old, have a cT1a renal parenchymal mass, and receive transplant consultation before nephrectomy. These data may inform future patient counseling.


Mannan-binding lectin activates C3 and the alternative complement pathway without involvement of C2.
Lectin pathway activation of C3 is known to involve target recognition by mannan-binding lectin (MBL) or ficolins and generation of classical pathway C3 convertase via cleavage of C4 and C2 by MBL-associated serine protease 2 (MASP-2). We investigated C3 activation in C2-deficient human sera and in sera with other defined defects of complement to assess other mechanisms through which MBL might recruit complement. The capacity of serum to support C3 deposition was examined by ELISA using microtiter plates coated with O antigen-specific oligosaccharides derived from Salmonella typhimurium, S. thompson, and S. enteritidis corresponding to serogroups B, C, and D (BO, CO, and DO). MBL bound to CO, but not to BO and DO, and efficiently supported C3 deposition in the absence of C2, C4, or MASP-2. The existence of an MBL-dependent C2 bypass mechanism for alternative pathway-mediated C3 activation was clearly demonstrated using CO, solid-phase mannan, and E. coli LPS. MASP-1 might contribute, but was not required for C3 deposition in the model used. Independent of MBL, specific antibodies to CO supported C3 deposition through classical and alternative pathways. MBL-dependent C2 bypass activation could be particularly important in various inherited and acquired complement deficiency states.


Oxaliplatin induces hyperexcitability at motor and autonomic neuromuscular junctions through effects on voltage-gated sodium channels.
Oxaliplatin, an effective cytotoxic treatment in combination with 5-fluorouracil for colorectal cancer, is associated with sensory, motor and autonomic neurotoxicity. Motor symptoms include hyperexcitability while autonomic effects include urinary retention, but the cause of these side-effects is unknown. We examined the effects on motor nerve function in the mouse hemidiaphragm and on the autonomic system in the vas deferens. In the mouse diaphragm, oxaliplatin (0.5 mM) induced multiple endplate potentials (EPPs) following a single stimulus, and was associated with an increase in spontaneous miniature EPP frequency. In the vas deferens, spontaneous excitatory junction potential frequency was increased after 30 min exposure to oxaliplatin; no changes in resting Ca(2+) concentration in nerve terminal varicosities were observed, and recovery after stimuli trains was unaffected. In both tissues, an oxaliplatin-induced increase in spontaneous activity was prevented by the voltage-gated Na(+) channel blocker tetrodotoxin (TTX). Carbamazepine (0.3 mM) also prevented multiple EPPs and the increase in spontaneous activity in both tissues. In diaphragm, beta-pompilidotoxin (100 microM), which slows Na(+) channel inactivation, induced multiple EPPs similar to oxaliplatin's effect. By contrast, blockers of K(+) channels (4-aminopyridine and apamin) did not replicate oxaliplatin-induced hyperexcitability in the diaphragm. The prevention of hyperexcitability by TTX blockade implies that oxaliplatin acts on nerve conduction rather than by effecting repolarisation. The similarity between beta-pompilidotoxin and oxaliplatin suggests that alteration of voltage-gated Na(+) channel kinetics is likely to underlie the acute neurotoxic actions of oxaliplatin.


Improving quality of Food Frequency Questionnaire response in low-income Mexican American children.
The authors evaluated the validity and reliability of the Block Kids Food Frequency Questionnaire (BKFFQ) and the Block Kid Screener (BKScreener) in Mexican American children living along the Texas-Mexico border who participated in the National Institutes of Health-funded Proyecto Bienestar Laredo. The Bienestar/NEEMA health program is a school-based diabetes and obesity control program, and the Proyecto Bienestar Laredo is the translation of the Bienestar/NEEMA health program to 38 elementary schools in Laredo, Texas. Par ticipants included 2,376 eight-year-old boys (48%) and girls (52%) from two school districts in Laredo. Two Food Frequency Questionnaire (BKFFQ and BKScreener) dietary intakes were collected, and an expert panel of nutritionist assigned a classification response quality of "Good," "Questionable," and "Poor," based on playfulness (systematic or nonrandom) patterns and completion rates. In addition, both instruments were assessed for reliability (test-retest) in 138 students from a San Antonio School District. Children's height, weight, percentage body fat, reported family history of diabetes, and Texas Assessments of Knowledge and Skills in reading and mathematics scores were collected. This study showed that for Mexican American children living along the Texas-Mexico border, within the time constraints of the classroom, BKScreener yielded better data than the BKFFQ.


Access to care for women veterans: delayed healthcare and unmet need.
Timely access to healthcare is essential to ensuring optimal health outcomes, and not surprisingly, is at the heart of healthcare reform efforts. While the Veterans Health Administration (VA) has made improved access a priority, women veterans still underutilize VA healthcare relative to men. Eliminating access disparities requires a better understanding of the barriers to care that women veterans' experience. We examined the association of general and veteran-specific barriers on access to healthcare among women veterans. Cross-sectional, population-based national telephone survey of 3,611 women veterans. Delayed healthcare or unmet healthcare need in the prior 12 months. Of women veterans, 19% had delayed healthcare or unmet need, with higher rates in younger age groups (36%, 29%, 16%, 7%, respectively, in 18-34, 35-49, 50-64, and 65-plus age groups; p < 0.001). Among those delaying or going without care, barriers that varied by age group were: unaffordable healthcare (63% of 18-34 versus 12% of 65-plus age groups); inability to take off from work (39% of those <50); and transportation difficulties (36% of 65-plus). Controlling for age, race/ethnicity, regular source of care, and health status, being uninsured (OR = 6.5; confidence interval [CI] 3.0-14.0), knowledge gaps about VA care (OR = 2.1; 95% CI 1.1-4.0), perception that VA providers are not gender-sensitive (OR = 2.4; CI 1.2-4.7), and military sexual assault history (OR = 2.1; CI 1.1-4.0) predicted delaying or foregoing care, whereas VA use and enrollment priority did not. Both general and veteran-specific factors impact women veterans' access to needed services. Many of the identified access barriers are potentially modifiable through expanded VA healthcare and social services. Health reform efforts should address these barriers for VA nonusers. Efforts are also warranted to improve women veterans' knowledge of availability and affordability of VA healthcare, and to enhance the gender-sensitivity of this care.


Evolution with stochastic fitness and stochastic migration.
Migration between local populations plays an important role in evolution - influencing local adaptation, speciation, extinction, and the maintenance of genetic variation. Like other evolutionary mechanisms, migration is a stochastic process, involving both random and deterministic elements. Many models of evolution have incorporated migration, but these have all been based on simplifying assumptions, such as low migration rate, weak selection, or large population size. We thus have no truly general and exact mathematical description of evolution that incorporates migration. We derive an exact equation for directional evolution, essentially a stochastic Price equation with migration, that encompasses all processes, both deterministic and stochastic, contributing to directional change in an open population. Using this result, we show that increasing the variance in migration rates reduces the impact of migration relative to selection. This means that models that treat migration as a single parameter tend to be biassed - overestimating the relative impact of immigration. We further show that selection and migration interact in complex ways, one result being that a strategy for which fitness is negatively correlated with migration rates (high fitness when migration is low) will tend to increase in frequency, even if it has lower mean fitness than do other strategies. Finally, we derive an equation for the effective migration rate, which allows some of the complex stochastic processes that we identify to be incorporated into models with a single migration parameter. As has previously been shown with selection, the role of migration in evolution is determined by the entire distributions of immigration and emigration rates, not just by the mean values. The interactions of stochastic migration with stochastic selection produce evolutionary processes that are invisible to deterministic evolutionary theory.


Viral dark matter and virus-host interactions resolved from publicly available microbial genomes.
The ecological importance of viruses is now widely recognized, yet our limited knowledge of viral sequence space and virus-host interactions precludes accurate prediction of their roles and impacts. In this study, we mined publicly available bacterial and archaeal genomic data sets to identify 12,498 high-confidence viral genomes linked to their microbial hosts. These data augment public data sets 10-fold, provide first viral sequences for 13 new bacterial phyla including ecologically abundant phyla, and help taxonomically identify 7-38% of 'unknown' sequence space in viromes. Genome- and network-based classification was largely consistent with accepted viral taxonomy and suggested that (i) 264 new viral genera were identified (doubling known genera) and (ii) cross-taxon genomic recombination is limited. Further analyses provided empirical data on extrachromosomal prophages and coinfection prevalences, as well as evaluation of in silico virus-host linkage predictions. Together these findings illustrate the value of mining viral signal from microbial genomes.


Chondroitin sulfate proteoglycans prevent immune cell phenotypic conversion and inflammation resolution via TLR4 in rodent models of spinal cord injury.
Chondroitin sulfate proteoglycans (CSPGs) act as potent inhibitors of axonal growth and neuroplasticity after spinal cord injury (SCI). Here we reveal that CSPGs also play a critical role in preventing inflammation resolution by blocking the conversion of pro-inflammatory immune cells to a pro-repair phenotype in rodent models of SCI. We demonstrate that enzymatic digestion of CSPG glycosaminoglycans enhances immune cell clearance and reduces pro-inflammatory protein and gene expression profiles at key resolution time points. Analysis of phenotypically distinct immune cell clusters revealed CSPG-mediated modulation of macrophage and microglial subtypes which, together with T lymphocyte infiltration and composition changes, suggests a role for CSPGs in modulating both innate and adaptive immune responses after SCI. Mechanistically, CSPG activation of a pro-inflammatory phenotype in pro-repair immune cells was found to be TLR4-dependent, identifying TLR4 signalling as a key driver of CSPG-mediated immune modulation. These findings establish CSPGs as critical mediators of inflammation resolution failure after SCI in rodents, which leads to prolonged inflammatory pathology and irreversible tissue destruction.


Post-transcriptional repression of the Drosophila midkine and pleiotrophin homolog miple by HOW is essential for correct mesoderm spreading.
The even spreading of mesoderm cells in the Drosophila embryo is essential for its proper patterning by ectodermally derived signals. In how germline clone embryos, defects in mesoderm spreading lead to a partial loss of dorsal mesoderm derivatives. HOW is an RNA-binding protein that is thought to regulate diverse mRNA targets. To identify direct HOW targets, we implemented a series of selection methods on mRNAs whose levels were elevated in how germline clone embryos during the stage of mesoderm spreading. Four mRNAs were found to be specifically elevated in the mesoderm of how germline clone embryos, and to exhibit specific binding to HOW via their 3' UTRs. Importantly, overexpression of three of these genes phenocopied the mesoderm-spreading phenotype of how germline clone embryos. Further analysis showed that overexpressing one of these genes, miple (a Drosophila midkine and pleiotrophin heparin-binding growth factor), in the mesoderm led to abnormal scattered MAPK activation, a phenotype that might explain the abnormal mesoderm spreading. In addition, the number of EVE-positive cells, which are responsive to receptor tyrosine kinase (RTK) signaling, was increased following Miple overexpression in the mesoderm and appeared to be dependent on Heartless function. In summary, our analysis suggests that HOW downregulates the levels of a number of mRNA species in the mesoderm in order to enable proper mesoderm spreading during early embryogenesis.


Genes involved in yellow pigmentation of Cronobacter sakazakii ES5 and influence of pigmentation on persistence and growth under environmental stress.
Cronobacter spp. are opportunistic food-borne pathogens that are responsible for rare but highly fatal cases of meningitis and necrotizing enterocolitis in neonates. While the operon responsible for yellow pigmentation in Cronobacter sakazakii strain ES5 was described recently, the involvement of additional genes in pigment expression and the influence of pigmentation on the fitness of Cronobacter spp. have not been investigated. Thus, the aim of this study was to identify further genes involved in pigment expression in Cronobacter sakazakii ES5 and to assess the influence of pigmentation on growth and persistence under conditions of environmental stress. A knockout library was created using random transposon mutagenesis. The screening of 9,500 mutants for decreased pigment production identified 30 colorless mutants. The mapping of transposon insertion sites revealed insertions in not only the carotenoid operon but also in various other genes involved in signal transduction, inorganic ions, and energy metabolism. To determine the effect of pigmentation on fitness, colorless mutants (DeltacrtE, DeltacrtX, and DeltacrtY) were compared to the yellow wild type using growth and inactivation experiments, a macrophage assay, and a phenotype array. Among other findings, the colorless mutants grew at significantly increased rates under osmotic stress compared to that of the yellow wild type while showing increased susceptibility to desiccation. Moreover, DeltacrtE and DeltacrtY exhibited increased sensitivity to UVB irradiation.


Ethnography/ethics.
This paper situates discussion of the ethics of ethnographic research against the background of a theoretical and methodological debate about the relationship between ethics and method, and about the relationships between research methods and their objects. In particular, the paper investigates the implications of folding together the ethical and the empirical in research and argues that this requires the development of new ethico-ethnographic methods for the investigation of ethico-moral objects. The paper falls into three main parts. The first considers calls for what has come to be known as empirical ethics, that is, for a more empirically informed bioethics, by way of an exploration of the integration of ethnographic methods in bioethics, and concludes that approaches which see the ethical and the empirical as 'complementary' do not do justice to the methodological implications of enfolding the ethical and the ethnographic. The second part juxtaposes this with calls for the integration of ethics in ethnography and, similarly, argues that the enfolding of the ethical and the empirical in ethnography calls for the development of new methods. The paper goes on to problematise the 'negotiational' approaches to informed consent preferred by many ethnographers, arguing that the concept of negotiation, rather than offering a solution to the problem of consent, is itself ethically complex and in need of analysis. The paper argues that, in the context of ethnographic research, the possibility of negotiational forms of consent depends upon engagement between researchers and researched, with unavoidably 'ethical' concepts such as 'respect', 'recognition', 'dignity', 'justice' and so on, and that this poses methodological challenges to ethnography. The paper's third section explores the implications of these arguments for research practice, using The Genethics Club as an example.


Clostridium spores for tumor-specific drug delivery.
Insufficient blood supply of rapidly growing tumors leads to the presence of hypoxia, a well-known feature in solid tumors. Hypoxia is known to decrease the efficiency of currently used anti-cancer modalities like surgery, chemotherapy and radiotherapy. Therefore, hypoxia seems to be a major limitation in current anti-cancer therapy. The use of non-pathogenic clostridia to deliver toxic agents to the tumor cells takes advantage of this unique physiology. These strictly anaerobic, Gram-positive, spore-forming bacteria give, after systemic administration, a selective colonization of hypoxic/necrotic areas within the tumor. Moreover, they can be genetically modified to secrete therapeutic proteins like cytosine deaminase or tumor necrosis factor-alpha. The specificity of this protein delivery system can be further increased when expression is controlled by the use of a radio-inducible promoter, leading to increased spatial and temporal regulation of protein expression. This approach of bacterial vector systems to target protein expression to the tumor can be considered very safe since bacteria can be eliminated at any moment by the addition of proper antibiotics. The Clostridium-based delivery system thus presents an alternative therapeutic modality to deliver anti-tumor agents specifically to the tumor site. This high selectivity offers a major advantage in comparison with the classical gene therapy systems.


Sacral neuromodulation therapy: a promising treatment for adolescents with refractory functional constipation.
Sacral neuromodulation therapy has been successfully applied in adult patients with urinary and fecal incontinence and in adults with constipation not responding to intensive conservative treatment. No data, however, are available on sacral neuromodulation therapy as a treatment option in adolescents with refractory functional constipation. This study aimed to describe the short-term results of sacral neuromodulation in adolescents with chronic functional constipation refractory to intensive conservative treatment. This is a retrospective review. This study took place at the Department of Surgery, Maastricht University Medical Centre, The Netherlands. Thirteen patients (all girls, age 10-18 years) with functional constipation according to the ROME III criteria not responding to intensive oral and rectal laxative treatment were assigned for sacral neuromodulation. When improvement of symptoms was observed during the testing phase, a permanent stimulator was implanted. Patients were prospectively followed up to at least 6 months after implantation of the permanent stimulator by interviews, bowel diaries, and Cleveland Clinic constipation score. Improvement was defined as spontaneous defecation ≥ 2 times a week. At presentation, none of the patients had spontaneous defecation or felt the urge to defecate. All patients had severe abdominal pain. Regular school absenteeism was present in 10 patients. After the testing phase, all but 2 patients had spontaneous defecation ≥ 2 times a week with a reduction in abdominal pain. After implantation, 11 (of 12) had a normal spontaneous defecation pattern of ≥ 2 times a week without medication, felt the urge to defecate, and perceived less abdominal pain without relapse of symptoms until 6 months after implantation. The average Cleveland Clinic constipation score decreased from 20.9 to 8.4. One lead revision and 2 pacemaker relocations were necessary. This study is limited by its small sample size, single-institution bias, and retrospective nature. Sacral neuromodulation appears to be a promising new treatment option in adolescents with refractory functional constipation not responding to intensive conservative therapy. Larger randomized studies with long-term follow-up are required.


Arthroscopic versus open acromioplasty: a prospective, randomized, blinded study.
The purpose of this study is to determine whether arthroscopic acromioplasty is equivalent or superior to open acromioplasty, in a prospective, randomized, controlled, blinded clinical trial. Seventy-one patients with a clinical diagnosis of impingement syndrome were randomized to arthroscopic or open acromioplasty. Nine were excluded because of full-thickness rotator cuff tears diagnosed after randomization. Sixty-two patients (49 men and 13 women) with a minimum follow-up of 12 months (mean, 25 months) were included. The patient groups were virtually identical with regard to duration of symptoms, shoulder functional demands, age, sex, hand dominance, mechanism of onset, range of motion, strength, joint laxity, and the presence of a compensation claim. Patients were prospectively randomized to arthroscopic or open acromioplasty after stratification for age (>50 years),associated ligamentous laxity, and the presence of an ongoing compensation claim. The main outcome measure was visual analog scales for pain and function. Also recorded were UCLA shoulder scores and visual analog scales for postoperative improvement, patient satisfaction, and a variety of clinical measures. An independent blinded examiner assessed all patients. There was no significant difference between open and arthroscopic acromioplasty in visual analog scales for postoperative improvement (P =.30), patient satisfaction (P =.94), UCLA shoulder score (P =.69), or strength (P =.62); however, open was superior to arthroscopic acromioplasty for pain and function (P =.01). Overall, 67% of patients had a good or excellent result. This increased to 87% when unsettled compensation claims were excluded. Repeat (open) acromioplasty was performed in 5 patients in the unsuccessful arthroscopic group without improvement. Open acromioplasty was equivalent to arthroscopic acromioplasty for UCLA scores and patient satisfaction. For pain and function, both gave significant improvement but the open technique may be superior. Unsettled compensation is a predictor of poor outcome.


A global approach to tumor immunology.
Biological and clinical advances in the understanding of tumor immunology suggest that immune responsiveness of human tumors is a complex biological phenomenon that could be best studied by a real-time comparison of tumor/host interactions in the tumor microenvironment through a high-throughput discovery-driven approach. This conclusion is derived from our recognition that too many hypotheses or, in other words, no solid single hypothesis exist, based on experimental results, to further drive experimentation in human subjects. Functional genomic studies entertained during the last few years consolidated the belief that in humans the interactions between tumor and immune cells are too complex to be approached exclusively with a hypothesis driven method. We believe that immune cells suit cancer cells in a Yin and Yang balance by opposing and yet mutually depending on each other. Indeed, immune infiltration in tumors may play a dual role modulating in different circumstances cancer cell growth or destruction through a physiological modulation of inflammation. It is reasonable to question what induces inflammation at the tumor site. We hypothesize that inflammation is primarily driven by the phenotype of tumor cells that can modulate their microenvironment through cell-to-cell interactions or the secretion of soluble factors. Thus, in analogy the observation of immune cells within tumors parallels the presence of paramedics, police and firemen at the scene of an accident, which is reactive to and not causative of the occurrence. In this review we will explore this hypothesis by reporting and summarizing most of our recent work in the frame of available literature on the subject.


Cold Ischemia Score: An mRNA Assay for the Detection of Extended Cold Ischemia in Formalin-Fixed, Paraffin-Embedded Tissue.
Although there are thousands of formalin-fixed paraffin-embedded (FFPE) tissue blocks potentially available for scientific research, many are of questionable quality, partly due to unknown preanalytical variables. We analyzed FFPE tissue biospecimens as part of the National Cancer Institute (NCI) Biospecimen Preanalytical Variables program to identify mRNA markers denoting cold ischemic time. The mRNA was extracted from colon, kidney, and ovary cancer FFPE blocks (40 patients, 10-12 hr fixation time) with 1, 2, 3, and 12 hr cold ischemic times, then analyzed using qRT-PCR for 23 genes selected following a literature search. No genes tested could determine short ischemic times (1-3 hr). However, a combination of three unstable genes normalized to a more stable gene could generate a "Cold Ischemia Score" that could distinguish 1 to 3 hr cold ischemia from 12 hr cold ischemia with 62% sensitivity and 84% specificity.


Discontinuation of medication overuse in headache patients: recovery of therapeutic responsiveness.
It is generally accepted that ongoing medication overuse nullifies the effect of prophylactic treatment, although few data support this contention. We set out to describe the treatment outcome in patients withdrawn from medication overuse and relate any improvement to a renewed effect of prophylaxis. For patients with probable medication-overuse headache (pMOH), treated and dismissed from the Danish Headache Centre in 2002 and 2003, we assed, from prospective headache diaries, the headache frequency before and after withdrawal of offending drugs and compared these frequencies with the headache frequency at dismissal. Among 1326 patients, 337 had pMOH. Eligible were 175, mean age 49 years, male/female ratio 1 : 2.7. Overall, there was a 46% decrease in headache frequency from the first visit to dismissal (P < 0.0001). Patients with no improvement 2 months after complete drug withdrawal (N = 88) subsequently responded to pharmacological and/or non-pharmacological prophylaxis with a 26% decrease in headache frequency as measured from the end of withdrawal to dismissal (P < 0.0001). At dismissal, 47% were on prophylaxis. Former non-responders to medical prophylaxis had a 49% decrease in headache frequency from first visit to dismissal (P < 0.0001), whereas those who had never received prophylaxis had a 56% reduction (P < 0.0001). This difference was not statistically significant (P = 0.22). Almost all MOH patients benefit from drug withdrawal, either just from the withdrawal or by transformation from therapeutic non-responsiveness to responsiveness. According to the International Classification of Headache Disorders, 2nd edn, the MOH diagnosis requires improvement after drug withdrawal. Our data suggest that these diagnostic criteria are too strict.


Metabolic acidosis does not contribute to chronic renal injury in the rat.
1. Metabolic acidosis invariably accompanies chronic renal failure, and short periods of metabolic acidosis cause renal growth and proteinuria in normal rats. Rates of ammoniagenesis are increased in chronic renal failure, and it has been suggested that this contributes to disease progression. This study assessed (i) whether prolonged acidosis causes chronic renal injury in the normal kidney and (ii) whether abrogation of acidosis slows disease progression in the remnant kidney. 2. Metabolic acidosis was induced in normal rats by dietary hydrochloric acid. Urinary excretion of total protein, lysozyme and albumin increased, peaking at week 8 but returning to baseline by week 14. At killing after 14 weeks, kidney weights, glomerular filtration rates and serum creatinine were the same in both groups, but kidney/body weight and kidney/heart weight ratios were greater in the acidotic group. All kidneys were normal by light microscopy. 3. Rats subjected to five-sixths nephrectomy were given sufficient dietary bicarbonate to abolish uraemic acidosis, and their outcome was compared with that of non-alkalinized remnants (controls). Proteinuria, glomerular filtration rates, blood pressure, histological injury and time to the development of terminal uraemia were no better in bicarbonate-supplemented animals than in controls. 4. These data demonstrate that metabolic acidosis neither causes nor exacerbates chronic renal injury. We conclude that the treatment of uraemic acidosis is unlikely to influence disease progression in patients with chronic renal failure.


Communism and the meaning of social memory: towards a critical-interpretive approach.
Using a case study of representations of communism in Romania, the paper offers a sketch of a critical-interpretive approach for exploring and engaging with the social memory of communism. When one considers the various contemporary appraisals, responses to and positions towards the communist period one identifies and one is obliged to deal with a series of personal and collective moral/political quandaries. In their attempt to bring about historical justice, political elites create a world that conforms more to their needs and desires than to the diversity of meanings of communism, experiences and dilemmas of lay people. This paper argues that one needs to study formal aspects of social memory as well as "lived", often conflicting, attitudinal and mnemonic stances and interpretive frameworks. One needs to strive to find the meaning of the social memory of communism in the sometimes contradictory, paradoxical attitudes and meanings that members of society communicate, endorse and debate. Many of the ethical quandaries and dilemmas of collective memory and recent history can be better understood by describing the discursive and sociocultural processes of meaning-making and meaning-interpretation carried out by members of a polity.


Transparent Films Made of Highly Scattering Particles.
Today, colloids are widely employed in various products from creams and coatings to electronics. The ability to control their chemical, optical, or electronic features by controlling their size and shape explains why these materials are so widely preferred. Nevertheless, altering some of these properties may also lead to some undesired side effects, one of which is an increase in optical scattering upon concentration. Here, we address this strong scattering issue in films made of binary colloidal suspensions. In particular, we focus on raspberry-type polymeric particles made of a spherical polystyrene core decorated by small hemispherical domains of acrylate with an overall positive charge, which display an unusual stability against aggregation in aqueous solutions. Their solid films display a brilliant red color due to Bragg scattering but appear completely white on account of strong scattering otherwise. To suppress the scattering and induce transparency, we prepared films by hybridizing them with oppositely charged PS particles with a size similar to that of the bumps on the raspberries. We report that the smaller PS particles prevent raspberry particle aggregation in solid films and suppress scattering by decreasing the spatial variation of the refractive index inside the film. We believe that the results presented here provide a simple strategy to suppress strong scattering of larger particles to be used in optical coatings.


Diabetes insipidus is an unfavorable prognostic factor for response to glucocorticoids in patients with autoimmune hypophysitis.
Autoimmune hypophysitis (AH) has a variable clinical presentation and natural history; likewise, its response to glucocorticoid therapy is often unpredictable. To identify clinical and radiological findings associated with response to glucocorticoids. 12 consecutive patients with AH, evaluated from 2008 to 2016. AH was the exclusion diagnosis after ruling out other pituitary masses and secondary causes of hypophysitis. Mean follow-up time was 30 ± 27 months (range 12-96 months). MRI identified two main patterns of presentation: global enlargement of the pituitary gland or panhypophysitis (n = 4, PH), and pituitary stalk abnormality only, or infundibulo-neuro-hypophysitis (n = 8, INH). Multiple tropin defects were more common in PH (100%) than those in INH (28% P = 0.014), whereas diabetes insipidus was more common in INH (100%) than that in PH (50%; P = 0.028). All 4 PH and 4 out of 8 INH were treated with glucocorticoids. Pituitary volume significantly reduced in all PH patients (P = 0.012), defective anterior pituitary function recovered only in the two patients without diabetes insipidus (50%) and panhypopituitarism persisted, along with diabetes insipidus, in the remaining 2 (50%). In all INH patients, either treated or untreated, pituitary stalk diameter reduced (P = 0.008) but diabetes insipidus persisted in all. Glucocorticoid therapy may improve anterior pituitary function in a subset of patients but has no effect on restoring posterior pituitary function. Diabetes insipidus appears as a negative prognostic factor for response to glucocorticoids.


Prevalence of antibodies to bovine viral diarrhoea virus in bulk tank milk and associated risk factors in Scottish dairy herds.
Bulk tank milk samples were collected from 374 dairy farms in Scotland in 2007/2008 along with questionnaire data relating to the management of the farm. Milk samples were tested for antibodies to bovine viral diarrhoea virus (BVDV) using a commercially available (Svanova) kit and percentage positivity scores calculated according to the manufacturer's guidelines. There were 220 farms that did not routinely vaccinate for bovine viral diarrhoea (BVD), and these were distributed according to the Swedish BVD eradication classes as 12.7 per cent, 22.3 per cent, 44.5 per cent and 20.5 per cent for Classes 0, 1, 2 and 3, respectively. A more sophisticated statistical method (finite mixture modelling) which does not depend on arbitrary thresholds and categories suggested a 73 per cent prevalence of herds with high mean levels of antibodies. Risk factor analysis suggested that routine vaccination for BVD, suspicion of BVD, housing of pregnant cows with calves, total number of cows and the proportion of cows that were dry were all associated with increased BVDV antibodies in bulk milk. The inclusion of BVD within the farm's health plan was associated with decreased BVDV antibodies in the bulk milk.


DNA crosslinking damage and cancer - a tale of friend and foe.
Interstrand crosslinks (ICLs) represent a major challenge for DNA replication and transcription by preventing DNA strand separation. Cells deficient in ICL repair are hypersensitive to a variety of bifunctional alkylating agents and exhibit excessive genomic instability. Patients with deficient ICL repair, such as those with Fanconi anemia, are predisposed to a broad spectrum of cancers. The profound cellular toxicity of ICLs is exploited clinically in cancer chemotherapy. Therefore, understanding the mechanism of ICL repair and its impact on cancer development and treatment is very important. Studies of diseases with defective ICL repair, especially Fanconi anemia, have revealed unique ICL repair mechanisms in humans. In this review, we describe pathways and factors involved in ICL damage response and their implications in cancer development and treatment.


Screening marine natural products for selective inhibitors of key kynurenine pathway enzymes.
Kynurenine, a metabolite of tryptophan along the 'kynurenine pathway', is at a branch point of the pathway which can lead to the synthesis of both quinolinic acid (QUIN) and kynurenic acid (KYNA). KYNA is an antagonist of glutamate receptors; however, QUIN is a selective agonist of NMDA receptors, and has been shown to act as an excitotoxic agent. A high QUIN/KYNA ratio has been implicated in a variety of neurological diseases in which excitotoxic neuronal cell death is found, e.g. AIDS-related dementia, stroke, etc. Inhibiting the key enzymes of this pathway (i.e. kynureninase and kynurenine 3-hydroxylase) would lower the QUIN/KYNA ratio, which may potentially have neuroprotective effects. We have developed high through-put assays for kynurenine pathway enzymes which allow us to screen extracts from marine organisms for selective enzyme inhibitors. Active metabolites are purified, isolated and identified by HPLC, high-field NMR and mass spectral techniques. Extracts from a sponge of the Aka species were found to contain a selective inhibitor of kynureninase. We have recently purified and identified the active principal as being serotonin sulfate. Related indoleamines, serotonin and 5-hydroxyindoleacetic acids are inactive. This finding may be suggestive of a novel interaction between the serotoninergic and excitatory amino acid pathways.


Biodegradation of phenanthrene by indigenous microorganisms in soils from Livingstone Island, Antarctica.
Biodegradation of polycyclic aromatic hydrocarbons (PAHs) in soils has been linked to history of exposure to PAHs and prevailing environmental conditions. This work assessed the capacity of indigenous microorganisms in soils collected in Livingstone Island (South Shetlands Islands, Antarctica) with no history of pollution (∑PAHs: 0.14-1.47 ng g(-1) dw) to degrade (14) C-phenanhthrene at 4, 12 and 22 °C. The study provides evidence of the presence of phenanthrene-degrading microorganisms in all studied soils. Generally, the percentage of (14) C-phenanhthrene mineralized increased with increasing temperature. The highest extent of (14) C-phenanhthrene mineralization (47.93%) was observed in the slurried system at 22 °C. This work supports findings of the presence of PAH-degrading microorganisms in uncontaminated soils and suggests the case is the same for uncontaminated Antarctic remote soils.


Multidisciplinary management improves anxiety, depression, medication adherence, and quality of life among patients with epilepsy in eastern China: A prospective study.
The objective of this study was to investigate the effect of a multidisciplinary program on anxiety, depression, medication adherence, and quality of life in patients with epilepsy in eastern China. A cohort of 184 patients with epilepsy from the epilepsy clinic of a tertiary hospital in eastern China completed this program, out of which 92 were randomized into the intervention group and 92 the control group. Patients in both groups received standard antiepileptic drugs (AEDs), while those of the intervention group received an additional 12-month multidisciplinary program developed by a group of the epileptologist, pharmacist, psychiatrist, and epilepsy specialist nurse. Patients were assessed both before and after the 12-month period. The Beck Depression Inventory (BDI), the Beck Anxiety Inventory (BAI), and the eight-item Morisky Medication Adherence Scale (MMAS-8) were used to assess the severity of depression, anxiety, and medication adherence, respectively, along with Quality of life in Epilepsy-31 (QOLIE-31) and self-reported seizure frequency for life quality and seizure severity. The 12-month multidisciplinary program significantly reduced the number of patients with severe depression (p = 0.013) and anxiety (p = 0.002), increased the number of patients with moderate-to-high AED adherence (p = 0.006) and the overall QOLIE-31 score (p < 0.001) in the intervention group. Both groups demonstrated a significant increase in the number of patients with a low seizure frequency after the 12-month period (p < 0.001). The 12-month multidisciplinary program offers an effective management strategy in improving psychiatric comorbidities, medication adherence, and quality of life in patients with epilepsy in eastern China.


Silencing of HPV 18 oncoproteins With RNA interference causes growth inhibition of cervical cancer cells.
Silencing the expression of human papillomavirus (HPV) oncoproteins should have therapeutic benefits for cervical cancer. The authors' objective was to study RNA interference of the HPV 18 E6/E7 bicistronic mRNA with E6 small interfering RNA (siRNA) and E7 siRNA and determine the effect of each siRNA on oncoprotein expression, resultant cell growth, and downstream molecular effects. RNA interference was used to knockdown HPV 18 E6 and E7 oncoproteins on the HPV 18 positive cervical cancer cell lines HeLa and C4I. Western blotting was used to assay for each oncoprotein expression and select downstream molecular targets. Cell cycle analyses, cell viability assays, and colony formation assays were performed to determine the effect of treatment by both HPV 18 E6 siRNA and E7 siRNA. The transfection reagent oligofectamine and Tax siRNA were used as negative controls. Transfection with E6 siRNA caused complete loss of E6 but not E7 oncoprotein. However, E7 siRNA induced complete loss of both E6 and E7 oncoproteins. E6 siRNA mediated the reexpression of p53 protein and a moderate decrease in phosphorylated retinoblastoma protein expression (pRb), resulting in decreased colony formation. Transfection with E7 siRNA mediated a robust increase in p53 expression and complete loss of pRb, resulting in a marked decrease in colony formation compared to the E6 siRNA (P =.001). Flow cytometry revealed significantly increased apoptotic cells with E7 siRNA compared to E6 siRNA and control. RNA interference targeting the E7 portion of the bicistronic HPV 18 mRNA can silence both E6 and E7 oncoproteins and is most effective in cervical cancer growth inhibition.


Comparison of left ventricular and biventricular pacing: Rationale and clinical implications.
Cardiac resynchronization therapy constitutes a cornerstone in advanced heart failure treatment, when there is evidence of dyssynchrony, especially by electrocardiography. However, it is plagued both by persistently high (~30%) rates of nonresponse and by deterioration of right ventricular function, owing to iatrogenic dyssynchrony in the context of persistent apical pacing to ensure delivery of biventricular pacing. Left ventricular pacing has long been considered an alternative to standard biventricular pacing and can be achieved as easily as inserting a single pacing electrode in the coronary sinus. Although monoventricular left ventricular pacing has been proven to yield comparable results with the standard biventricular modality, it is the advent of preferential left ventricular pacing, combining both the powerful resynchronization potential of multipolar coronary sinus and right-sided electrodes acting in concert and the ability to preserve intrinsic, physiological right ventricular activation. In this review, we aim to present the underlying principles and modes for delivering left ventricular pacing, as well as to highlight advantages of preferential over monoventricular configuration. Finally, current clinical evidence, following implementation of automated algorithms, regarding performance of left ventricular as compared with biventricular pacing will be discussed. It is expected that the field of preferential left ventricular pacing will grow significantly over the following years, and its combination with other advanced pacing modalities may promote clinical status and prognosis of patients with advanced dyssynchronous heart failure.


Calibration of respiratory inductive plethysmograph during natural breathing.
We describe a single-posture method for deriving the proportionality constant (K) between rib cage (RC) and abdominal (AB) amplifiers of the respiratory inductive plethysmograph (RIP). Qualitative diagnostic calibration (QDC) is based on equations of the isovolume maneuver calibration (ISOCAL) and is carried out during a 5-min period of natural breathing without using mouthpiece or mask. In this situation, K approximates the ratio of standard deviations (SD) of the uncalibrated changes of AB-to-RC volume deflections. Validity of calibration was evaluated by 1) analyzing RIP waveforms during an isovolume maneuver and 2) comparing changes of tidal volume (VT) amplitude and functional residual capacity (FRC) level measured by spirometry (SP) with RIP values. Comparisons of VT(RIP) to VT(SP) were also obtained in a variety of postures during natural (uninstructed) preferential RC and AB breathing and with voluntary changes of VT amplitude and FRC level. VT(RIP)-to-VT(SP) comparisons were equal to or closer than published reports for single posture, ISOCAL, multiple- and linear-regression procedures. QDC of RIP in supine posture with comparisons to SP in that posture and others showed better accuracy in horizontal than upright postures.


Adverse effects of danazol prophylaxis on the lipid profiles of patients with hereditary angioedema.
Hereditary angioedema (HAE) is a rare disorder caused by the deficiency of the C1-inhibitor gene (C1INH) . Patients experience recurrent bouts of edema, which can occur in almost any region of the body. As regards the treatment of the disease, danazol (an attenuated androgen) is used, among other agents, for long-term prophylaxis. The aim of this study was to investigate the possible adverse effects of danazol on serum lipid profile, as well as to ascertain whether danazol treatment is associated with an increased risk of atherosclerosis. Serum concentrations of total cholesterol, high-density lipoprotein (HDL), low-density lipoprotein (LDL), triglycerides, apolipoprotein A-I, apolipoprotein B-100, and lipoprotein(a) were compared between danazol-treated patients with HAE and 2 control groups (ie, patients who did not receive long-term danazol prophylaxis and untreated healthy subjects). Serum concentrations of HDL ( P = .0002 and P < .0001) and apolipoprotein A-I ( P = .0015 and P < .0001) were significantly lower, whereas LDL ( P = .0129 and P = .0127) and apolipoprotein B-100 ( P = .0456 and P = .0013) were higher in the danazol-treated patients compared with the 2 control groups, respectively. No significant difference was found in total cholesterol, triglyceride, or lipoprotein(a) levels. Patients who received danazol had an 11.6 (95% CI, 2.7-49.7) times higher risk for abnormally low HDL levels and a 4.4 (95% CI, 1.2-16.0) times lower risk for high LDL concentrations. Our findings indicate that the long-term use of danazol is associated with an increased risk for early atherosclerosis in patients with HAE. Consequently, monitoring of HDL and LDL levels at regular intervals is recommended during follow-up.


A modular dose delivery system for treating moving targets with scanned ion beams: Performance and safety characteristics, and preliminary tests.
The purpose of this study was to develop a modular dose-delivery system (DDS) for scanned-ion radiotherapy that mitigates against organ motion artifacts by synchronizing the motion of the beam with that of the moving anatomy. We integrated a new motion synchronization system and an existing DDS into two centers. The modular approach to integration utilized an adaptive layer of software and hardware interfaces. The method of synchronization comprised three major tasks, namely, the creation of 3D treatment plans (each representing one phase of respiratory motion and together comprising a 4D plan), monitoring anatomic motion during treatment, and synchronization of the beam to anatomic motion. The synchronization was accomplished in real time by repeatedly selecting and delivering a 3D plan, i.e., the one that most closely corresponded to the current anatomic state, until all plans were delivered. The performance characteristics of the motion mitigation system were tested by delivering 4D treatment plans to a moving phantom and comparing planned and measured dose distributions. Dosimetric performance was considered acceptable when the gamma-index pass rate was >90%, homogeneity-index value was >95%, and conformity-index value was >60%. Selected safety characteristics were tested by introducing errors during treatment and testing DDS response. Acceptable dosimetric performance and safety characteristics were observed for all treatment plans. We demonstrated, for the first time, that a modular prototype system, synchronizing scanned ion beams with moving targets can deliver conformal, motion-compensated dose distributions. The prototype system was implemented and characterized at GSI and CNAO.


A hot pepper cDNA encoding ascorbate peroxidase is induced during the incompatible interaction with virus and bacteria.
Capsicum annuum L. is infected by a number of viruses, including the tobacco mosaic virus (TMV). To study the defense-related genes that are induced by TMV in hot peppers, the pepper plant, which is susceptible to P1.2 but resistant to the P0 pathotype of TMV, was inoculated with TMV-P0. Differential screening isolated the genes that were specifically up- or down-regulated during the hypersensitive response (HR). The CaAPX1 cDNA clone that putatively encodes a polypeptide of cytosolic ascorbate peroxidase was selected as an up-regulated gene. It was isolated for further study. The full-length cDNA for CaAPX1, which is 972 bp long, contained the open-reading frame of 250-amino acid residues. A genomic Southern blot analysis showed that there were only limited copies of the CaAPX1 gene in the hot pepper genome. In hot pepper cv. Bugang, which is resistant to TMV-P0 and susceptible to TMV-P1.2, the CaAPX1 gene transcript was accumulated by TMV-P0, but not by TMV-P1.2 inoculation. CaAPX1 transcripts began to accumulate 24 h post-inoculation of TMV-P0, and increased gradually until 96 h. To investigate whether each transcript is induced by other stimuli, the plants were treated with various chemicals and wounding. A striking induction of the CaAPX1 transcript was observed at 2 h. It subsided 12 h after salicylic acid (SA), ethephon, and methyl jasmonate (MeJA) treatments. The response of the gene upon other pathogen infection was also examined by a bacterial pathogen (Xanthomonas campestris pv. vesicatoria race 3) inoculation. The CaAPX1 gene was induced in a hot pepper (C. annuum cv. ECW 20R) that was resistant to this bacterial pathogen, but not in a susceptible hot pepper (C. annuum cv. ECW). These results suggest the possible role(s) for the CaAPX1 gene in plant defense against viral and bacterial pathogen.


Roles of UGT, P450, and Gut Microbiota in the Metabolism of Epacadostat in Humans.
Epacadostat (EPA, INCB024360) is a first-in-class, orally active, investigational drug targeting the enzyme indoleamine 2,3-dioxygenase 1 (IDO1). In Phase I studies, EPA has demonstrated promising clinical activity when used in combination with checkpoint modulators. When the metabolism of EPA was investigated in humans, three major, IDO1-inactive, circulating plasma metabolites were detected and characterized: M9, a direct O-glucuronide of EPA; M11, an amidine; and M12, N-dealkylated M11. Glucuronidation of EPA to form M9 is the dominant metabolic pathway, and in vitro, this metabolite is formed by UGT1A9. However, negligible quantities of M11 and M12 were detected when EPA was incubated with a panel of human microsomes from multiple tissues, hepatocytes, recombinant human cytochrome P450s (P450s), and non-P450 enzymatic systems. Given the reductive nature of M11 formation and the inability to define its source, the role of gut microbiota was investigated. Analysis of plasma from mice dosed with EPA following pretreatment with either antibiotic (ciprofloxacin) to inhibit gut bacteria or 1-aminobenzotriazole (ABT) to systemically inhibit P450s demonstrated that gut microbiota is responsible for the formation of M11. Incubations of EPA in human feces confirmed the role of gut bacteria in the formation of M11. Further, incubations of M11 with recombinant P450s showed that M12 is formed via N-dealkylation of M11 by CYP3A4, CYP2C19, and CYP1A2. Thus, in humans three major plasma metabolites of EPA were characterized: two primary metabolites, M9 and M11, formed directly from EPA via UGT1A9 and gut microbiota, respectively, and M12 formed as a secondary metabolite via P450s from M11.


Expectations and outcomes of gastric feeding tubes.
To compare expected outcomes with actual outcomes from tube feeding in adult patients. This prospective cohort study was conducted in two North Carolina hospitals. Surrogates were interviewed shortly after feeding tube insertion and at 3- and 6-month follow-up; chart abstraction and death certificate review also were carried out. Participants were surrogate decision-makers for consecutive adult patients who received new feeding tubes. There were 288 patients with surrogate decision-makers enrolled. Mean age was 65 years; 30% had a primary diagnosis of stroke, 16% neurodegenerative disorder, 20% head and neck cancer, and 30% other diagnoses. At 3 months, 21% of patients had died, and 6-month mortality was 30%. At 3 months, 38% of survivors were residing in a nursing home, and 27% had the feeding tube removed. Patients were impaired in most activities of daily living (ADLs) with little change over time. Medical complications were common: 25% of patients had decubitus ulcers at 3 months, and 24% had at least one episode of pneumonia. Perceived global quality of life was poor at 4.6 (on a 0-10 scale) at baseline, and surrogates anticipated this would improve to 8.0 with tube feeding. Family surrogates' expectations for improvement from the feeding tube were very high at baseline and remained so at 3 and 6 months. Families' high expectations of benefit from tube feeding are in contrast to clinical outcomes. Providers and families need better information about the outcomes of this common procedure.


Consistency in recognizing microinvasion in breast carcinomas is improved by immunohistochemistry for myoepithelial markers.
Microinvasion is the smallest morphologically identifiable stage of invasion. Its presence and distinction from in situ carcinoma may have therapeutic implications, and clinical staging also requires the recognition of this phenomenon. Microinvasion is established on the basis of several morphological criteria, which may be difficult and not perfectly reproducible among pathologists. The aim of this study was to assess the consistency of diagnosing microinvasion in the breast on traditional haematoxylin and eosin (HE) stained slides and to evaluate whether immunohistochemistry (IHC) for myoepithelial markers could improve this. Digital images were generated from representative areas of 50 cases stained with HE and IHC for myoepithelial markers. Cases were specifically selected from the spectrum of in situ to microinvasive cancers. Twenty-eight dedicated breast pathologists assessed these cases at different magnifications through a web-based platform in two rounds: first HE only and after a washout period by both HE and IHC. Consistency in the recognition of microinvasion significantly improved with the use of IHC. Concordance rates increased from 0.85 to 0.96, kappa from 0.5 to 0.85, the number of cases with 100% agreement rose from 9/50 to 25/50 with IHC and the certainty of diagnosis also increased. The use of IHC markedly improves the consistency of identifying microinvasion. This corroborates previous recommendations to use IHC for myoepithelial markers to clarify cases where uncertainty exists about the presence of microinvasion. Microinvasive carcinoma is a rare entity, and seeking a second opinion may avoid overdiagnosis.


Cell-specific optoporation with near-infrared ultrafast laser and functionalized gold nanoparticles.
Selective targeting of diseased cells can increase therapeutic efficacy and limit off-target adverse effects. We developed a new tool to selectively perforate living cells with functionalized gold nanoparticles (AuNPs) and near-infrared (NIR) femtosecond (fs) laser. The receptor CD44 strongly expressed by cancer stem cells was used as a model for selective targeting. Citrate-capped AuNPs (100 nm in diameter) functionalized with 0.01 orthopyridyl-disulfide-poly(ethylene glycol) (5 kDa)-N-hydroxysuccinimide (OPSS-PEG-NHS) conjugated to monoclonal antibodies per nm(2) and 5 μM HS-PEG (5 kDa) were colloidally stable in cell culture medium containing serum proteins. These AuNPs attached mostly as single particles 115 times more to targeted CD44(+) MDA-MB-231 and CD44(+) ARPE-19 cells than to non-targeted CD44(-) 661W cells. Optimally functionalized AuNPs enhanced the fs laser (800 nm, 80-100 mJ cm(-2) at 250 Hz or 60-80 mJ cm(-2) at 500 Hz) to selectively perforate targeted cells without affecting surrounding non-targeted cells in co-culture. This novel highly versatile treatment paradigm can be adapted to target and perforate other cell populations by adapting to desired biomarkers. Since living biological tissues absorb energy very weakly in the NIR range, the developed non-invasive tool may provide a safe, cost-effective clinically relevant approach to ablate pathologically deregulated cells and limit complications associated with surgical interventions.


An antigenic structure of the trans-activator protein encoded by human T-cell leukemia virus type-I (HTLV-I), as defined by a panel of monoclonal antibodies.
In order to study an antigenic structure of the trans-activator protein encoded by human T-cell leukemia virus type-I (HTLV-I), tax1 antigen, we generated and characterized a panel of rat anti-tax1 monoclonal antibodies (MAbs) designated WATM-1, WATM-2, WATM-3, and WATM-4. These MAbs were derived from WKA rats immunized with HTLV-I-transformed (HTLV-I+) syngeneic T cells. Immunoblot assays showed that: (1) All the MAbs reacted with the tax1 antigen in HTLV-I+ cell lines and a recombinant tax1 antigen, PX141 (containing entire tax1 polypeptide); (2) WATM-3 and WATM-4, but not WATM-1 or WATM-2, reacted with a truncated tax1 antigen, XD59 (tax1 amino acids 180-338); (3) None of them reacted with another truncated tax1 antigen, XD128 (tax1 amino acids 1-47 and 286-353); and (4) each of the four MAbs had different reactivity with tax1-related antigens in the range 38-41 kDa expressed in simian cell lines infected with various HTLV-I-related simian retroviruses (STLV-I). None of the MAbs reacted with HTLV-II tax antigen. Human sera containing anti-tax1 antibodies interfered specifically with the antigen-specific binding of all the MAbs. These results suggest that the present rat MAbs are directed against various epitopes on the tax1 antigen. An antigenic structure of the tax1 antigen deduced from reactivity of a panel of anti-tax1 MAbs including the present rat MAbs is discussed.


A cyclophilin homology domain-independent role for Nup358 in HIV-1 infection.
The large nucleoporin Nup358/RanBP2 forms eight filaments that project from the nuclear pore into the cytoplasm where they function as docking platforms for nucleocytoplasmic transport receptors. RNAi screens have implicated Nup358 in the HIV-1 life cycle. The 164 C-terminal amino acids of this 3,224 amino acid protein are a cyclophilin homology domain (Nup358Cyp), which has potential to bind the HIV-1 capsid and regulate viral progress to integration. Here we examined the virological role of Nup358 in conditional knockout mouse cells and in RNAi-depleted human CD4⁺ T cells. Cre-mediated gene knockout was toxic and diminished HIV-1 infectivity. However, cellular health and HIV-1 susceptibility were coordinately preserved if, prior to gene inactivation, a transposon was used to express all of Nup358 or only the N-terminal 1340 amino acids that contain three FG repeats and a Ran-binding domain. HIV-1, but not N74D capsid-mutant HIV-1, was markedly sensitive to TNPO3 depletion, but they infected 1-1340 segment-complemented Nup358 knockout cells equivalently. Human and mouse CypA both rescued HIV-1 in CypA gene⁻/⁻ Jurkat cells and TRIM-Nup358Cyp fusions derived from each species were equally antiviral; each also inhibited both WT and N74D virus. In the human CD4⁺T cell line SupT1, abrupt Nup358 depletion reduced viral replication but stable Nup358-depleted cells replicated HIV-1 normally. Thus, human CD4⁺ T cells can accommodate to loss of Nup358 and preserve HIV-1 susceptibility. Experiments with cylosporine, viruses with capsids that do not bind cyclophilins, and growth arrest did not uncover viral dependency on the C-terminal domains of Nup358. Our data reinforce the virological importance of TNPO3 and show that Nup358 supports nuclear transport functions important for cellular homeostasis and for HIV-1 nuclear import. However, the results do not suggest direct roles for the Nup358 cyclophilin or SUMO E3 ligase domains in engaging the HIV-1 capsid prior to nuclear translocation.


Protocol for the 'e-Nudge trial': a randomised controlled trial of electronic feedback to reduce the cardiovascular risk of individuals in general practice [ISRCTN64828380].
Cardiovascular disease (including coronary heart disease and stroke) is a major cause of death and disability in the United Kingdom, and is to a large extent preventable, by lifestyle modification and drug therapy. The recent standardisation of electronic codes for cardiovascular risk variables through the United Kingdom's new General Practice contract provides an opportunity for the application of risk algorithms to identify high risk individuals. This randomised controlled trial will test the benefits of an automated system of alert messages and practice searches to identify those at highest risk of cardiovascular disease in primary care databases. Patients over 50 years old in practice databases will be randomised to the intervention group that will receive the alert messages and searches, and a control group who will continue to receive usual care. In addition to those at high estimated risk, potentially high risk patients will be identified who have insufficient data to allow a risk estimate to be made. Further groups identified will be those with possible undiagnosed diabetes, based either on elevated past recorded blood glucose measurements, or an absence of recent blood glucose measurement in those with established cardiovascular disease. The intervention will be applied for two years, and outcome data will be collected for a further year. The primary outcome measure will be the annual rate of cardiovascular events in the intervention and control arms of the study. Secondary measures include the proportion of patients at high estimated cardiovascular risk, the proportion of patients with missing data for a risk estimate, and the proportion with undefined diabetes status at the end of the trial.


The presence of respiratory disorders in individuals with low back pain: A systematic review.
Inspiratory muscles, such as the diaphragm, play a key role in both respiration and spinal control. Therefore, diaphragm dysfunctions are often related to low back pain (LBP). However, few is known on the association between the presence of LBP and the presence of respiratory disorders (RD). To perform a systematic review on the relation between RD and LBP. Systematic review. Two reviewers searched on PubMed/MEDLINE for studies concerning LBP and RD, from 1950 up to January 2016. The search string consisted of the following key words: low back pain, dyspnea, respiratory problems, lung diseases, comorbidity, pulmonary disease, chronic obstructive, smoking, asthma, allergy, sinusitis, respiratory tract infection and hyperventilation. The aim was to evaluate a potential correlation, co-occurrence or causality between RD and LBP. A total of 16 articles were included. A significant correlation between the presence of LBP and the presence of RD such as dyspnea, asthma, different forms of allergy, and respiratory infections was found. No correlation was found between Chronic Obstructive Pulmonary Disease (COPD) and LBP, and no articles were found on the correlation between hyperventilation and LBP. This is the first study providing an overview of the literature on the relation between LBP and RD. Immunological, biomechanical, psychosocial and socio-economic factors might explain this correlation. Smoking is likely to contribute. Future studies must reveal the causative relationship. Therapy, level 2a.


The pros and cons of using photographs in nursing research.
Traditional approaches to research can sometimes face difficulties in engaging participants, being flexible and ultimately eliciting data about people's experiences. Visual methods are alternative approaches that use image - often photographs - as stimuli or structure in interviews. However, little has been published in the literature to guide nurse researchers in applying and evaluating these methods. To increase nurse researchers' awareness of visual methods and their potential. The paper explores visual methods, with a focus on those that use photographs. It discusses the benefits of using photographs in research, such as reducing the gap between the researcher and participant and helping participants to express meaningful data. It also examines associated ethical, analytical and practical difficulties. The paper illustrates discussion points with reflections from research and compares interviews that do and do not use photographs. Using photographs is a good alternative to more traditional approaches but the exact benefits are difficult to evidence because of the complexities of the interactions involved. for practice This paper should increase nurse researchers' awareness of visual methods, help them to make informed choices about research methods and encourage the use of visual methods in health research.


A decentralized clerkship: strategies for standardizing content and instruction.
Course decentralization in a required family medicine clerkship occurred because additional teaching and clinical resources were needed to meet the educational goals and objectives of an entire class of medical students. The decentralized teaching effort, which consisted of a variety of practice models, had the potential for an inconsistent exposure to the required educational content of the clerkship. Course monitoring and evaluation also was difficult because of the logistics of the various teaching sites. In order to minimize inconsistent content exposure, efforts to standardize course requirements occurred through criterion-based faculty and teaching site selection, initial faculty development, and provision of appropriate library resources. Continuous standardization consisted of required periodic faculty development, frequent evaluation of faculty and teaching sites, and comparative studies of student performance. Standardization of some of the students' patient care experience resulted from the use of predetermined list of required clinical encounters. A computer assisted evaluation of students' patient experience log defined the actual student patient care population at various teaching sites and compared this to course requirements. Deficits in patient care experiences were addressed by providing either the appropriate patient encounters or alternative experiences through print, audiovisual, or teaching sessions.


The respiratory syncytial virus matrix protein possesses a Crm1-mediated nuclear export mechanism.
The respiratory syncytial virus (RSV) matrix (M) protein is localized in the nucleus of infected cells early in infection but is mostly cytoplasmic late in infection. We have previously shown that M localizes in the nucleus through the action of the importin beta1 nuclear import receptor. Here, we establish for the first time that M's ability to shuttle to the cytoplasm is due to the action of the nuclear export receptor Crm1, as shown in infected cells, and in cells transfected to express green fluorescent protein (GFP)-M fusion proteins. Specific inhibition of Crm1-mediated nuclear export by leptomycin B increased M nuclear accumulation. Analysis of truncated and point-mutated M derivatives indicated that Crm1-dependent nuclear export of M is attributable to a nuclear export signal (NES) within residues 194 to 206. Importantly, inhibition of M nuclear export resulted in reduced virus production, and a recombinant RSV carrying a mutated NES could not be rescued by reverse genetics. That this is likely to be due to the inability of a nuclear export deficient M to localize to regions of virus assembly is indicated by the fact that a nuclear-export-deficient GFP-M fails to localize to regions of virus assembly when expressed in cells infected with wild-type RSV. Together, our data suggest that Crm1-dependent nuclear export of M is central to RSV infection, representing the first report of such a mechanism for a paramyxovirus M protein and with important implications for related paramyxoviruses.


Cloning and characterization of Pros45, the Drosophila SUG1 proteasome subunit homolog.
The proteasome plays essential roles in a variety of cellular processes, including degradation of the bulk of cellular proteins, degradation of short-lived proteins such as cell cycle regulators, generation of antigenic peptides, and mediating programmed cell death. One of the best characterized subunits of the 26S proteasome is encoded by the yeast gene SUG1. We report here the cloning and characterization of the Drosophila homolog of this gene, Pros45. At the protein level, Pros45 is highly conserved with respect to its homologs in a variety of taxa: it shows 74% identity to yeast Sug1; 86% to mouse m56/mSug1/FZA-B; 87% to human Trip1; and 97% to moth 18-56. Using a genomic clone as a probe for in situ hyridization to polytene chromesomes, we demonstrated that Pros45 maps to 19F, near the base of the X chromosome. Use of a pros45 cDNA clone as a probe revealed a second site of hybridization at 99CD. Pros45 mRNA is found in the unfertilized egg and in all cells of the early embryo. By the end of embryogenesis, Pros45 is expressed predominantly in the central nervous system. Targeted expression of Pros45 in a variety of different cells using the Gal4 UAS P-element system failed to generate an overt phenotype. This study provides the foundation for further examination of the role of the 26S proteasome in homeostasis and development in Drosophila.


Emerging Comorbidities in Adult Asthma: Risks, Clinical Associations, and Mechanisms.
Asthma is a heterogeneous disease with many phenotypes, and age at disease onset is an important factor in separating the phenotypes. Most studies with asthma have been performed in patients being otherwise healthy. However, in real life, comorbid diseases are very common in adult patients. We review here the emerging comorbid conditions to asthma such as obesity, metabolic syndrome, diabetes mellitus type 2 (DM2), and cardiac and psychiatric diseases. Their role as risk factors for incident asthma and whether they affect clinical asthma are evaluated. Obesity, independently or as a part of metabolic syndrome, DM2, and depression are risk factors for incident asthma. In contrast, the effects of comorbidities on clinical asthma are less well-known and mostly studies are lacking. Cross-sectional studies in obese asthmatics suggest that they may have less well controlled asthma and worse lung function. However, no long-term clinical follow-up studies with these comorbidities and asthma were identified. These emerging comorbidities often occur in the same multimorbid adult patient and may have in common metabolic pathways and inflammatory or other alterations such as early life exposures, systemic inflammation, inflammasome, adipokines, hyperglycemia, hyperinsulinemia, lung mechanics, mitochondrial dysfunction, disturbed nitric oxide metabolism, and leukotrienes.


Extracellular Matrix Disparities in an Nkx2-5 Mutant Mouse Model of Congenital Heart Disease.
Congenital heart disease (CHD) affects almost one percent of all live births. Despite diagnostic and surgical reparative advances, the causes and mechanisms of CHD are still primarily unknown. The extracellular matrix plays a large role in cell communication, function, and differentiation, and therefore likely plays a role in disease development and pathophysiology. Cell adhesion and gap junction proteins, such as integrins and connexins, are also essential to cellular communication and behavior, and could interact directly (integrins) or indirectly (connexins) with the extracellular matrix. In this work, we explore disparities in the expression and spatial patterning of extracellular matrix, adhesion, and gap junction proteins between wild type and Nkx2-5 +/R52G mutant mice. Decellularization and proteomic analysis, Western blotting, histology, immunostaining, and mechanical assessment of embryonic and neonatal wild type and Nkx2-5 mutant mouse hearts were performed. An increased abundance of collagen IV, fibronectin, and integrin β-1 was found in Nkx2-5 mutant neonatal mouse hearts, as well as increased expression of connexin 43 in embryonic mutant hearts. Furthermore, a ventricular noncompaction phenotype was observed in both embryonic and neonatal mutant hearts, as well as spatial disorganization of ECM proteins collagen IV and laminin in mutant hearts. Characterizing such properties in a mutant mouse model provides valuable information that can be applied to better understanding the mechanisms of congenital heart disease.


The use of the UCLan Mountain Medicine and other Masters programmes as a recruitment tool for healthcare organisations with difficulties in recruitment and retention.
A variety of incentives are used to stimulate recruitment to remote and rural locations. In this presentation, we share experiences at the University of Central Lancashire of forming partnerships with National Health Service (NHS) organisations to offer investment in careers as a recruitment and retention tool. Qualitative structured interviews. NHS organisations priorities were to find cost-effective and successful strategies to recruit and retain workers. Many had tried financial incentives, including 'golden handshakes' and 'golden handcuffs' but found them ineffective or unaffordable. Prospective employee priorities were multifactorial, including a desire for flexibility, manageable workload and the ability to develop their personal and career interests. Whilst rates of pay were important, one-off lump sum payments were viewed as having lower value. This partnership approach has helped us develop MSc programmes that fit with the needs of their services and innovatively support their recruitment aims. We have also given voice to the needs of our learners, for example by encouraging job planning approaches that facilitate the prolonged blocks of leave required for acclimatisation of practitioners of Mountain Medicine to travel to high altitude. When explored, advertised one-off lump sum payments amounts were perceived as being misleading due to tax deductions, diminishing their utility as a 'feel good' factor in retention. Conversely, gradual investment over time, using academic study as an enabler for flexible job planning combined with a feeling that their employer supported some of their drivers and values contributed to a greater sense of commitment by employees.


Echocardiographic epicardial fat thickness is related to altered blood pressure responses to exercise stress testing.
Hypertensive response at peak exercise and blunted blood pressure (BP) recovery, altered BP responses obtained from exercise stress testing, have been suggested as risk factors for future onset of hypertension in previous studies. Epicardial fat, a new cardiometabolic risk factor, has been linked to hypertension in some recent studies. In this study, we tested the primary hypothesis suggesting that the epicardial fat thickness (EFT) is related to altered BP responses to treadmill exercise testing. We also evaluated the sensitivity and specificity of the EFT as a predictor of hypertensive response to peak exercise. Normotensive subjects underwent to treadmill stress testing and transthoracic echocardiography. Hypertensive response to peak treadmill exercise testing was defined as ≥ 210/105 mmHg and ≥ 190/105 mmHg at peak exercise in males and females, respectively. BP recovery index (BPRI) was defined as the ratio of the BP at the 3rd minute of the recovery phase to BP at peak exercise. EFT was measured by echocardiography. Thirty-two subjects with hypertensive response to peak exercise constituted Group 1 and 48 subjects with normal response constituted Group 2. The mean EFT of subjects in Group 1 was significantly higher (8.2 ± 1.1 mm vs 5.1 ± 1.5 mm; p = 0.0001) than subjects in Group 2. In correlation analysis performed in Group 1, EFT was found to be significantly correlated with BPRI (r = 0.51, p < 0.003). An EFT of ≥ 6.5 mm predicted the hypertensive response to peak exercise test with 68.8% sensitivity and 87.5% specificity (receiving operator characteristic area under curve: 0.879, 95% CI 0.793-0.965, p < 0.001). Patients with EFT ≥ 6.5 mm showed a significantly increased BPRI (0.89 ± 0.07 vs 0.74 ± 0.09, p < 0.0001) and peak systolic BP (198.4 ± 15.3 mmHg vs 169.4 ± 19.8 mmHg, p < 0.0001). There were significant differences in metabolic equivalents, maximum heart rate, homeostatic model assessment of insulin resistance, high-density lipoprotein-cholesterol, waist circumference and age values between two patients groups dichotomized according to the cut-off value of EFT. BPRI was the only independent variable related to EFT in the multivariate analysis (odds ratio = 1.4, 95% CI 2.75-7.16, p = 0.001). EFT was found to be related to altered BP responses to exercise stress testing. The echocardiographic measurement of EFT may serve as a useful non-invasive indicator of heightened risk of future hypertension.


Health beliefs associated with readiness for genetic counseling among high risk breast cancer survivors.
We used the Health Belief Model (HBM) to explore factors associated with readiness for genetic counseling among breast cancer survivors. Breast cancer survivors meeting NCCN genetic counseling referral criteria completed questionnaires capturing demographic and clinical information and factors guided by the HBM, including health beliefs, psychosocial variables, and cues to action. Using logistic regression, we examined whether the above variables differed based on readiness group (pre-contemplators, who did not plan to make a genetic counseling appointment, and contemplators, who planned to make a genetic counseling appointment in the next 1-6 months). Of 111 participants, 57% were pre-contemplators and 43% were contemplators. Higher cancer worry was associated with increased odds of being a contemplator (OR = 2.99; 95% CI = 1.37-6.54) and higher perceived barriers to genetic counseling were associated with decreased odds of being a contemplator (OR = 0.31; 95% CI = 0.11-0.85). Those who reported a family member encouraged them to get tested were more likely to be contemplators (OR = 3.57; 95% CI = 1.19-10.70). Our results suggest key factors for predicting genetic counseling readiness include cancer worry, perceived barriers, and family influence. There is need for increased genetic counseling awareness. Better understanding of factors related to survivors' decisions about counseling can inform tailored interventions to improve uptake and ultimately reduce cancer recurrence risk.


Prepuncture ultrasound-measured distance: an accurate reflection of epidural depth in infants and small children.
Epidural cannulation is technically difficult in children who have small anatomic structures. Ultrasound information regarding the distance of skin-to-ligament flavum may be useful, leading to an increase in success rate without dural puncture. This study was performed to assess whether ultrasound-measured, skin-to-ligament flavum distance would reflect the needle depth during epidural puncture in infants and children. The study compromised 180 children, aged 2 to 84 months, undergoing urologic surgery. After induction of anesthesia, ultrasound images of the longitudinal median and transverse views were acquired from L4-L5 in lateral decubitus position. Measured distance of skin-to-ligament flavum in each view was compared with the perpendicular skin-to-epidural depth, which was obtained from needle depth and angle by use of a trigonometric ratio equation. Additionally, we evaluated the ultrasound visibility of the ligament flavum and dura mater, number of puncture attempts, and complications. The correlation coefficient between measured distance and perpendicular epidural depth was slightly higher in longitudinal median view (R2 = 0.848) than in transverse view (R2 = 0.788). The visibility of ligament flavum and dura mater was "good" in 91 and 170 of 180 patients, respectively, and "sufficient" in the remaining subjects. The epidural space was located on first puncture attempt in 179 of 180 cases (99.4%). No incidents of dural puncture or bloody tap occurred. Ultrasound, particularly in the longitudinal median view, provides accurate information on the distance of skin-to-ligament flavum in infants and children. With reference to the measured distance, epidural puncture can be performed with minimal risk of dural puncture (upper limit of 95% CI = 1.67%).


Facile synthesis of nano-Li4 Ti5O12 for high-rate Li-ion battery anodes.
One of the most promising anode materials for Li-ion batteries, Li4Ti5O12, has attracted attention because it is a zero-strain Li insertion host having a stable insertion potential. In this study, we suggest two different synthetic processes to prepare Li4Ti5O12 using anatase TiO2 nanoprecursors. TiO2 powders, which have extraordinarily large surface areas of more than 250 m2 g-1, were initially prepared through the urea-forced hydrolysis/precipitation route below 100°C. For the synthesis of Li4Ti5O12, LiOH and Li2CO3 were added to TiO2 solutions prepared in water and ethanol media, respectively. The powders were subsequently dried and calcined at various temperatures. The phase and morphological transitions from TiO2 to Li4Ti5O12 were characterized using X-ray powder diffraction and transmission electron microscopy. The electrochemical performance of nanosized Li4Ti5O12 was evaluated in detail by cyclic voltammetry and galvanostatic cycling. Furthermore, the high-rate performance and long-term cycle stability of Li4Ti5O12 anodes for use in Li-ion batteries were discussed.


High-efficiency x-ray gratings with asymmetric-cut multilayers.
We present the fabrication and analysis of efficient and highly dispersive gratings for the x-ray and extreme ultraviolet (EUV) regime. We show that an asymmetric-cut multilayer structure can act as a near-perfect blazed grating. The precision and high line density are achieved by layer deposition of materials, which can be controlled to the angstrom level. We demonstrate this in the EUV regime with two structures made by cutting and polishing magnetron-sputtered multilayer mirrors of over 2000 bilayers thick, each with a period of 6.88 nm. These were cut at angles of 2.9° and 7.8° to the surface. Within the 3% bandwidth rocking curve of the multilayer, the angular dispersion of the diffracted wave was in agreement with the grating equation for elements with 7250 and 19,700 line pairs/mm, respectively. The dependence of the measured efficiency was in excellent agreement with a formulation of dynamical diffraction theory for multilayered structures. At a wavelength of 13.2 nm, the efficiency of the first-order diffraction was over 95% of the reflectivity of the uncut multilayer. We predict that such structures should also be effective at shorter x-ray wavelengths. Both the Laue (transmitting) and Bragg (reflecting) geometries are incorporated in our formalism, which is applied to the analysis of multilayer Laue lenses and focusing and dispersing Bragg optics.


Near full-length genome characterization of HIV type 1 unique BC recombinant forms from Southern Brazil.
Abstract In southern Brazil, CRF31_BC is the major HIV-1 recombinant form and URFs_BC occurs very infrequently. The near full-length genomes of four samples displaying a similar recombination profile in the integrase region and different profiles in the protease/reverse transcriptase regions were analyzed to elucidate their mosaic structure and to describe for the first time the near full-length genomes of URFs_BC circulating in Brazil. For this purpose, overlapping PCR amplifications followed by direct sequencing were carried out. Despite the observation that these samples share a similar recombination profile in the integrase region, phylogenetic, bootscan, and informative site analyses revealed that the four samples displayed distinct mosaic structures. Additional recombination events occurred mainly in the gag, reverse transcriptase, and env regions. Our data provide the first description of the near full-length genomes of URFs_BC in Brazil and offer important insights about the complexity of HIV-1 recombinant strains currently in circulation in the south of Brazil.


Host processing of branched DNA intermediates is involved in targeted transposition of IS911.
A simplified system using bacterial insertion sequence IS911 has been developed to investigate targeted insertion next to DNA sequences resembling IS ends. We show here that these IR-targeted events occur by an unusual mechanism. In the circular IS911 transposition intermediate the two IRs are abutted to form an IR/IR junction. IR-targeted insertion involves transfer of a single end of the junction to the target IR to generate a branched DNA structure. The single-end transfer (SET) intermediate, but not the final insertion product, can be detected in an in vitro reaction. SET intermediates must be processed by the bacterial host to obtain the final insertion products. Sequence analysis of these IR-targeted insertion products and of those obtained in vivo revealed high levels of DNA sequence conversion in which mutations from one IR were transferred to another. These sequence changes cannot be explained by the classic transposition pathway. A model is presented in which the four-way Holliday-like junction created by SET is processed by host-mediated branch migration, resolution, repair and replication. This pathway resembles those described for processing other branched DNA structures such as stalled replication forks.


HRCT of fibrosing lung disease.
The use of high-resolution computed tomography (HRCT) has brought increased diagnostic discrimination to the evaluation of lung disease, particularly fibrosing lung diseases. Once the presence of a predominantly fibrosing lung disease has been established on evaluation of a HRCT, a stepwise approach is proposed that can refine the potential HRCT diagnoses from a list of over 100 different interstitial lung diseases to one of only five fibrosing lung diseases. Within the category of the fibrosing lung diseases, the recognition of idiopathic pulmonary fibrosis (IPF) is key. IPF is the most prevalent idiopathic interstitial pneumonia and has a mortality greater than any of the other diffuse lung diseases. Several diagnostic dilemmas are explored including challenges with the recent IPF diagnosis and management guidelines (2011), as well as with the 'difficult to characterize' fibrosing diseases such as smoking-related lung fibrosis, unclassifiable disease and acute exacerbations of fibrosing lung disease.


Waking and sleeping following water deprivation in the rat.
Wake-sleep (W-S) states are affected by thermoregulation. In particular, REM sleep (REMS) is reduced in homeotherms under a thermal load, due to an impairment of hypothalamic regulation of body temperature. The aim of this work was to assess whether osmoregulation, which is regulated at a hypothalamic level, but, unlike thermoregulation, is maintained across the different W-S states, could influence W-S occurrence. Sprague-Dawley rats, kept at an ambient temperature of 24°C and under a 12 h∶12 h light-dark cycle, were exposed to a prolonged osmotic challenge of three days of water deprivation (WD) and two days of recovery in which free access to water was restored. Two sets of parameters were determined in order to assess: i) the maintenance of osmotic homeostasis (water and food consumption; changes in body weight and fluid composition); ii) the effects of the osmotic challenge on behavioral states (hypothalamic temperature (Thy), motor activity, and W-S states). The first set of parameters changed in WD as expected and control levels were restored on the second day of recovery, with the exception of urinary Ca(++) that almost disappeared in WD, and increased to a high level in recovery. As far as the second set is concerned, WD was characterized by the maintenance of the daily oscillation of Thy and by a decrease in activity during the dark periods. Changes in W-S states were small and mainly confined to the dark period: i) REMS slightly decreased at the end of WD and increased in recovery; ii) non-REM sleep (NREMS) increased in both WD and recovery, but EEG delta power, a sign of NREMS intensity, decreased in WD and increased in recovery. Our data suggest that osmoregulation interferes with the regulation of W-S states to a much lesser extent than thermoregulation.