Polymeric microspheres as protein transduction reagents.
Discovering the function of an unknown protein, particularly one with neither structural nor functional correlates, is a daunting task. Interaction analyses determine binding partners, whereas DNA transfection, either transient or stable, leads to intracellular expression, though not necessarily at physiologically relevant levels. In theory, direct intracellular protein delivery (protein transduction) provides a conceptually simpler alternative, but in practice the approach is problematic. Domains such as HIV TAT protein are valuable, but their effectiveness is protein specific. Similarly, the delivery of intact proteins via endocytic pathways (e.g. using liposomes) is problematic for functional analysis because of the potential for protein degradation in the endosomes/lysosomes. Consequently, recent reports that microspheres can deliver bio-cargoes into cells via a non-endocytic, energy-independent pathway offer an exciting and promising alternative for in vitro delivery of functional protein. In order for such promise to be fully exploited, microspheres are required that (i) are stably linked to proteins, (ii) can deliver those proteins with good efficiency, (iii) release functional protein once inside the cells, and (iv) permit concomitant tracking. Herein, we report the application of microspheres to successfully address all of these criteria simultaneously, for the first time. After cellular uptake, protein release was autocatalyzed by the reducing cytoplasmic environment. Outside of cells, the covalent microsphere-protein linkage was stable for ≥90 h at 37 °C. Using conservative methods of estimation, 74.3% ± 5.6% of cells were shown to take up these microspheres after 24 h of incubation, with the whole process of delivery and intracellular protein release occurring within 36 h. Intended for in vitro functional protein research, this approach will enable study of the consequences of protein delivery at physiologically relevant levels, without recourse to nucleic acids, and offers a useful alternative to commercial protein transfection reagents such as Chariot™. We also provide clear immunostaining evidence to resolve residual controversy surrounding FACS-based assessment of microsphere uptake.


Detection of Type VII Collagen Autoantibodies Before the Onset of Bullous Systemic Lupus Erythematosus.
Anti-type VII collagen autoantibodies are often detectable in patients with bullous systemic lupus erythematosus (BSLE). However, the timing of their appearance preceding the onset of disease is unknown to date. We report the case of a 50-year-old woman with a history of SLE who was seen with vesicles and bullae around her lips, trunk, axillae, arms, and thighs. Histologic analysis and immunofluorescence and immunoblot studies confirmed the diagnosis of BSLE. Immunoblotting and enzyme-linked immunosorbent assay studies of the patient's serum obtained 3 months before the onset of BSLE showed the presence of anti-type VII collagen autoantibodies. Levels of anti-type VII collagen IgG increased after bullous lesions appeared. Within 1 month after initiating dapsone therapy and increasing the dosage of prednisone, skin lesions promptly resolved. One year after the onset of BSLE, the anti-type VII collagen IgG decreased below levels observed before the inception of the bullous lesions. Anti-type VII collagen autoantibodies can precede the clinical appearance of BSLE. The subsequent increase and decrease in levels of circulating anti-type VII collagen autoantibodies, which mirrored skin disease activity, support a potential role in their initiation of disease.


Human immunocompetent Organ-on-Chip platforms allow safety profiling of tumor-targeted T-cell bispecific antibodies.
Traditional drug safety assessment often fails to predict complications in humans, especially when the drug targets the immune system. Here, we show the unprecedented capability of two human Organs-on-Chips to evaluate the safety profile of T-cell bispecific antibodies (TCBs) targeting tumor antigens. Although promising for cancer immunotherapy, TCBs are associated with an on-target, off-tumor risk due to low levels of expression of tumor antigens in healthy tissues. We leveraged in vivo target expression and toxicity data of TCBs targeting folate receptor 1 (FOLR1) or carcinoembryonic antigen (CEA) to design and validate human immunocompetent Organs-on-Chips safety platforms. We discovered that the Lung-Chip and Intestine-Chip could reproduce and predict target-dependent TCB safety liabilities, based on sensitivity to key determinants thereof, such as target expression and antibody affinity. These novel tools broaden the research options available for mechanistic understandings of engineered therapeutic antibodies and assessing safety in tissues susceptible to adverse events.


Add-on Cannabidiol Treatment for Drug-Resistant Seizures in Tuberous Sclerosis Complex: A Placebo-Controlled Randomized Clinical Trial.
Efficacy of cannabidiol has been demonstrated in seizures associated with Lennox-Gastaut and Dravet syndromes but appears not yet to have been established in conditions with primarily focal seizures, such as tuberous sclerosis complex (TSC). To evaluate efficacy and safety of 25-mg/kg/day and 50-mg/kg/day cannabidiol dosages vs placebo against seizures associated with TSC. This double-blind, placebo-controlled randomized clinical trial (GWPCARE6) enrolled patients between April 6, 2016, and October 4, 2018; follow-up was completed on February 15, 2019. The trial was conducted at 46 sites in Australia, Poland, Spain, the Netherlands, United Kingdom, and United States. Eligible patients (aged 1-65 years) were those with a clinical diagnosis of TSC and medication-resistant epilepsy who had had at least 8 TSC-associated seizures during the 4-week baseline period, with at least 1 seizure occurring in at least 3 of the 4 weeks, and were currently taking at least 1 antiepileptic medication. Patients received oral cannabidiol at 25 mg/kg/day (CBD25) or 50 mg/kg/day (CBD50) or a matched placebo for 16 weeks. The prespecified primary outcome was the change from baseline in number of TSC-associated seizures for cannabidiol vs placebo during the treatment period. Of 255 patients screened for eligibility, 31 were excluded and 224 were randomized. Of the 224 included patients (median [range] age, 11.4 [1.1-56.8] years; 93 female patients [41.5%]), 75 were randomized to CBD25, 73 to CBD50, and 76 to placebo, with 201 completing treatment. The percentage reduction from baseline in the type of seizures considered the primary end point was 48.6% (95% CI, 40.4%-55.8%) for the CBD25 group, 47.5% (95% CI, 39.0%-54.8%) for the CBD50 group, and 26.5% (95% CI, 14.9%-36.5%) for the placebo group; the percentage reduction from placebo was 30.1% (95% CI, 13.9%-43.3%; P < .001) for the CBD25 group and 28.5% (95% CI, 11.9%-42.0%; nominal P = .002) for the CBD50 group. The most common adverse events were diarrhea (placebo group, 19 [25%]; CBD25 group, 23 [31%]; CBD50 group, 41 [56%]) and somnolence (placebo group, 7 [9%]; CBD25 group, 10 [13%]; CBD50 group, 19 [26%]), which occurred more frequently with cannabidiol than placebo. Eight patients in CBD25 group, 10 in CBD50 group, and 2 in the placebo group discontinued treatment because of adverse events. Twenty-eight patients taking cannabidiol (18.9%) had elevated liver transaminase levels vs none taking placebo. Cannabidiol significantly reduced TSC-associated seizures compared with placebo. The 25-mg/kg/day dosage had a better safety profile than the 50-mg/kg/day dosage. ClinicalTrials.gov Identifier: NCT02544763.


Teaching communications skills to medical students: Introducing the fine art of medical practice.
Like many other people based professions, communications skills are essential to medical practice also. Traditional medical teaching in India does not address communication skills which are most essential in dealing with patients. Communication skills can be taught to medical students to increase clinical competence. To teach basic communication and counseling skills to fourth-year undergraduate students to increase their clinical competence. A total of 48, fourth-year MBBS students participated in the study. They were given training in basic communication and counseling skills and taught the patient interview technique according to Calgary-Cambridge guide format. Improvement in communication was assessed by change in pre- and post-training multiple choice questions, clinical patient examination, and Standardized Patient Satisfaction Questionnaire (SPSQ) scores. About 88% of the students in the sample were convinced of the importance of learning communication skills for effective practice. Almost 90% students were communicating better after training, as tested by improved SPSQ. As judged by Communication Skill Attitude Scale, student's positive attitude toward learning communication skill indicated that there is a necessity of communication skill training during undergraduate years. The ability to communicate effectively is a core competency for medical practitioners. Inculcating habits of good communications skill during formative years will help the medical students and future practitioners. Regular courses on effective communication should be included in the medical school curriculum.


Influence of organic matter decomposition on soluble carbon and its copper-binding capacity.
Bulk and low molecular weight (LMW) (<1 kDa) water-extractable carbon were collected from fresh and microbially degraded wheat straw (Triticum aestivum L.) and crimson clover (Trifolium incarnatum L.) residues to monitor early-stage humification over an 8-wk incubation. Copper complexation parameters were determined for both bulk and LMW water-extractable C for both plant materials in a separate 1-wk incubation. Humification progressed through increasing molar absorptivity (A285) and phenolic and total acidity (TA), and through an increase in average molecular size and degree of polymerization as determined by ultrafiltration and changes in fluorescence peak locations. Such dynamic transformations demonstrate that while humification is a bulk property, with C breakdown and stabilization occurring simultaneously and continuously in soil, its early stages can be effectively monitored for fresh plant residues. Significant changes consistently occurred during the first 7 d of the incubation and were more pronounced for LMW fractions than bulk extracts. For both residues, water-extractable C extracted initially and following a 7-d incubation desorbed and complexed 0.11 to 0.55 mmol resin-bound Cu g(-1) C. Low molecular weight water-extractable C generated the higher values within this range, and values increased consistently following incubation. Potential concerns regarding LMW soluble Cu complexes include percolation through soils or runoff into adjacent water bodies as well as effects on plant root development.


High-resolution X-ray structure of the trimeric Scar/WAVE-complex precursor Brk1.
The Scar/WAVE-complex links upstream Rho-GTPase signaling to the activation of the conserved Arp2/3-complex. Scar/WAVE-induced and Arp2/3-complex-mediated actin nucleation is crucial for actin assembly in protruding lamellipodia to drive cell migration. The heteropentameric Scar/WAVE-complex is composed of Scar/WAVE, Abi, Nap, Pir and a small polypeptide Brk1/HSPC300, and recent work suggested that free Brk1 serves as a homooligomeric precursor in the assembly of this complex. Here we characterized the Brk1 trimer from Dictyostelium by analytical ultracentrifugation and gelfiltration. We show for the first time its dissociation at concentrations in the nanomolar range as well as an exchange of subunits within different DdBrk1 containing complexes. Moreover, we determined the three-dimensional structure of DdBrk1 at 1.5 Å resolution by X-ray crystallography. Three chains of DdBrk1 are associated with each other forming a parallel triple coiled-coil bundle. Notably, this structure is highly similar to the heterotrimeric α-helical bundle of HSPC300/WAVE1/Abi2 within the human Scar/WAVE-complex. This finding, together with the fact that Brk1 is collectively sandwiched by the remaining subunits and also constitutes the main subunit connecting the triple-coil domain of the HSPC300/WAVE1/Abi2/ heterotrimer to Sra1(Pir1), implies a critical function of this subunit in the assembly process of the entire Scar/WAVE-complex.


Nonenzymatic glycosylation of human serum albumin and its effect on antibodies profile in patients with diabetes mellitus.
Albumin glycation and subsequent formation of advanced glycation end products (AGEs) correlate with diabetes and associated complications. Human Serum Albumin (HSA) was modified with D-glucose for a 40 day period under sterile conditions at 37°C. Modified samples along with native HSA (unmodified) were analyzed for structural modifications by UV and fluorescence, FTIR, Liquid chromatography mass spectrometry (LCMS) and X-ray crystallography. New-Zealand white female rabbits immunized with AGEs, represent auto-antibodies formation as assessed by competitive and direct binding enzyme-linked immunosorbent assay (ELISA). Neo-epitopesagainst In-vitro formed AGEs were characterized in patients with diabetes mellitus type 2 (n = 50), type 1 (n = 50), gestational diabetes (n = 50) and type 2 with chronic kidney disease (CKD) with eGFR level 60-89 mL/min (n = 50) from serum direct binding ELISA. Glycated-HSA showed amarked increase in hyperchromicity of 65.82%,71.98%, 73.62% and 76.63% at λ280 nm along with anincreasein fluorescence intensity of 65.82%, 71.98%, 73.62% and 76.63% in glycated-HSA compared to native. FTIR results showed theshifting of Amide I peak from 1656 cm_1 to 1659 cm_1 and Amide II peak from 1554 cm_1 to 1564 cm_1 in glycated-HSA, with anew peak appearance of carbonyl group at 1737 cm-1. LCMS chromatogram of glycated-HSA showed thepresence of carboxymethyl lysine (CML) at 279.1 m/z. Immunological analysis showed high antibody titre>1:12,800 in theserum of rabbits immunized with glycated-HSA (modified with 400 mg/dL glucose) and inhibition of 84.65% at anantigen concentration of 20μg/mL. Maximum serum auto-antibody titre was found in T2DM (0.517±0.086), T1DM (0.108±0.092), GDM (0.611±0.041) and T2DM+CKD (0.096±0.25) patients immunized with glycated-HSA (modified with 400 mg/dL glucose). Non-enzymatic glycosylation of HSA manifests immunological complications in diabetes mellitus due to change in its structure that enhances neo-epitopes generation.


Jinx, an MCMV susceptibility phenotype caused by disruption of Unc13d: a mouse model of type 3 familial hemophagocytic lymphohistiocytosis.
Mouse cytomegalovirus (MCMV) susceptibility often results from defects of natural killer (NK) cell function. Here we describe Jinx, an N-ethyl-N-nitrosourea-induced MCMV susceptibility mutation that permits unchecked proliferation of the virus, causing death. In Jinx homozygotes, activated NK cells and cytotoxic T lymphocytes (CTLs) fail to degranulate, although they retain the ability to produce cytokines, and cytokine levels are markedly elevated in the blood of infected mutant mice. Jinx was mapped to mouse chromosome 11 on a total of 246 meioses and confined to a 4.60-million basepair critical region encompassing 122 annotated genes. The phenotype was ascribed to the creation of a novel donor splice site in Unc13d, the mouse orthologue of human MUNC13-4, in which mutations cause type 3 familial hemophagocytic lymphohistiocytosis (FHL3), a fatal disease marked by massive hepatosplenomegaly, anemia, and thrombocytopenia. Jinx mice do not spontaneously develop clinical features of hemophagocytic lymphohistiocytosis (HLH), but do so when infected with lymphocytic choriomeningitis virus, exhibiting hyperactivation of CTLs and antigen-presenting cells, and inadequate restriction of viral proliferation. In contrast, neither Listeria monocytogenes nor MCMV induces the syndrome. In mice, the HLH phenotype is conditional, which suggests the existence of a specific infectious trigger of FHL3 in humans.


Inhibition of protein interactions with the beta 2 sliding clamp of Escherichia coli DNA polymerase III by peptides from beta 2-binding proteins.
The sliding clamp of the Escherichia coli replisome is now understood to interact with many proteins involved in DNA synthesis and repair. A universal interaction motif is proposed to be one mechanism by which those proteins bind the E. coli sliding clamp, a homodimer of the beta subunit, at a single site on the dimer. The numerous beta(2)-binding proteins have various versions of the consensus interaction motif, including a related hexameric sequence. To determine if the variants of the motif could contribute to the competition of the beta-binding proteins for the beta(2) site, synthetic peptides derived from the putative beta(2)-binding motifs were assessed for their abilities to inhibit protein-beta(2) interactions, to bind directly to beta(2), and to inhibit DNA synthesis in vitro. A hierarchy emerged, which was consistent with sequence similarity to the pentameric consensus motif, QL(S/D)LF, and peptides containing proposed hexameric motifs were shown to have activities comparable to those containing the consensus sequence. The hierarchy of peptide binding may be indicative of a competitive hierarchy for the binding of proteins to beta(2) in various stages or circumstances of DNA replication and repair.


Measuring Teacher Job Satisfaction: Assessing Invariance in the Teacher Job Satisfaction Scale (TJSS) Across Six Countries.
Work and organizational psychology has long been concerned with measuring job satisfaction in organizational contexts, and this has carried across to the field of education, leading to a research focus on the work-related satisfaction of teachers. Today, a myriad of organizations continue to assess employees' job satisfaction on a routine basis (Liu, Borg, & Spector, 2004). Unfortunately, a sort of balkanization of the field has resulted in the production of dozens of specific measurement tools, making it difficult to cross-compare samples and contexts. The present paper tested the measurement invariance of the Teacher Job Satisfaction Scale (TJSS) in six international cohorts (Netherlands, United States, Russia China, Italy and Palestine) of in-service teachers (N = 2,819). Confirmatory factor analysis and multi-group invariance tests were applied. The TJSS-9 displayed robust psychometric proprieties and no substantial departures from measurement invariance (configural and metric). Future research is required to further test equivalence across additional countries, with view to developing a truly international tool for measuring job satisfaction in teaching.


The feasibility of robotic-assisted concomitant procedures during mitral valve operations.
In this study, we present our clinical experience and midterm results with the robotic-assisted concomitant procedures during mitral valve operations. Between March 2010 and February 2018, a total of 34 patients (8 males, 26 females; mean age 58.3 years; range, 34 to 78 years) who underwent robotic-assisted concomitant procedures during mitral valve surgery were retrospectively analyzed. Demographic characteristics of the patients, comorbidities, medical, and surgical histories, operative and laboratory results, electrocardiographic findings, postoperative intensive care unit and ward outcomes, and cardiac follow-up data were recorded. Atrial fibrillation-related medication use, stroke, or other thromboembolic events, and electrocardiographic reports in patients who underwent cryoablation were reviewed at three and 12 months after the operation. A total of 76 robotic-assisted concomitant procedures were performed during mitral valve repair (n=11) or replacement (n=23) in 34 patients. These procedures were cryoablation (n=29), tricuspid valve repair (n=6), tricuspid valve replacement (n=2), left atrial appendage ligation (n=32), atrial septal defect and patent foramen ovale closure (n=5), and left atrial thrombectomy (n=2). The mean preoperative EuroSCORE values were 5.1±2.5. The mean duration of cardiopulmonary bypass and cross-clamp was 156±69.4 min and 101±42 min, respectively. Normal sinus rhythm was restored in 85% of the patients (24/28) after cryoablation and two patients (5.8%) had permanent pacemaker within a year during follow-up. There was one (2.9%) mortality in the early postoperative period due to hemorrhage related to the posterior left ventricular wall rupture. No blood product was used in 82.4% of the patients. One patient had a transient cerebral event and symptoms regressed completely within two months. Technological improvements and growing experience can decrease the suspects related to prolonged operational duration during robotic-assisted cardiac surgery. Concomitant procedures in addition to mitral valve operations can be performed with low complication rates in centers with experience of robotic surgery.


Ethics of alternative trial designs and methods in low-resource settings.
This editorial introduces articles in this Special Issue, which are based on presentations given at the 2017 meeting of the Global Forum of Bioethics in Research meeting. The main themes presented at the meeting were the use of cluster randomized trials, stepped-wedge cluster randomized trials, and controlled human infection models in research conducted in low-resource settings. The editorial sets out which ethical issues may arise in the context of alternative trial designs and describes the articles in this issue that addresses some or more of the ethical issues, such as justification of the research design, risk-benefit evaluations and consent.


The impact of operative time on complications after plastic surgery: a multivariate regression analysis of 1753 cases.
Little evidence within plastic surgery literature supports the precept that longer operative times lead to greater morbidity. The authors investigate surgery duration as a determinant of morbidity, with the goal of defining a clinically relevant time for increased risk. A retrospective chart review was conducted of patients who underwent a broad range of complex plastic surgical procedures (n = 1801 procedures) at UT Southwestern Medical Center in Dallas, Texas, from January 1, 2008 to January 31, 2012. Adjusting for possible confounders, multivariate logistic regression assessed surgery duration as an independent predictor of morbidity. To define a cutoff for increased risk, incidence of complications was compared among quintiles of surgery duration. Stratification by type of surgery controlled for procedural complexity. A total of 1753 cases were included in multivariate analyses with an overall complication rate of 27.8%. Most operations were combined (75.8%), averaging 4.9 concurrent procedures. Each hour increase in surgery duration was associated with a 21% rise in odds of morbidity (P < .0001). Compared with the first quintile of operative time (<2.0 hours), there was no change in complications until after 3.1 hours of surgery (odds ratio, 1.6; P = .017), with progressively greater odds increases of 3.1 times after 4.5 hours (P < .0001) and 4.7 times after 6.8 hours (P < .0001). When stratified by type of surgery, longer operations continued to be associated with greater morbidity. Surgery duration is an independent predictor of complications, with a significantly increased risk above 3 hours. Although procedural complexity undoubtedly affects morbidity, operative time should factor into surgical decision making.


Isolation of antirhinoviral sesquiterpenes from ginger (Zingiber officinale).
The dried rhizomes of Indonesian ginger, Zingiber officinale, were investigated for antirhinoviral activity in the plaque reduction test. Fractionation by solvent extraction, solvent partition, and repeated chromatography guided by bioassay, allowed the isolation of several sesquiterpenes with antirhinoviral activity. The most active of these was beta-sesquiphellandrene [2] with an IC50 of 0.44 microM vs. rhinovirus IB in vitro.


The role of transmural ventricular heterogeneities in cardiac vulnerability to electric shocks.
Transmural electrophysiological heterogeneities have been shown to contribute to arrhythmia induction in the heart; however, their role in defibrillation failure has never been examined. The goal of this study is to investigate how transmural heterogeneities in ionic currents and gap-junctional coupling contribute to arrhythmia generation following defibrillation strength shocks. This study used a 3D anatomically realistic bidomain model of the rabbit ventricles. Transmural heterogeneity in ionic currents and reduced sub-epicardial intercellular coupling were incorporated based on experimental data. The ventricles were paced apically, and truncated-exponential monophasic shocks of varying strength and timing were applied via large external electrodes. Simulations demonstrate that inclusion of transmural heterogeneity in ionic currents results in an increase in vulnerability to shocks, reflected in the increased upper limit of vulnerability, ULV, and the enlarged vulnerable window, VW. These changes in vulnerability stem from increased post-shock dispersion in repolarisation as it increases the likelihood of establishment of re-entrant circuits. In contrast, reduced sub-epicardial coupling results in decrease in both ULV and VW. This decrease is caused by altered virtual electrode polarisation around the region of sub-epicardal uncoupling, and specifically, by the increase in (1) the amount of positively polarised myocardium at shock-end and (2) the spatial extent of post-shock wavefronts.


Seoul hantavirus in brown rats in the Netherlands: implications for physicians--Epidemiology, clinical aspects, treatment and diagnostics.
The recent discovery of Seoul hantavirus (SEOV) presence in wild rat populations in the Netherlands has direct implications for Dutch clinicians and hantavirus diagnostics. SEOV is amongst the Old World hantaviruses which cause haemorrhagic fever and renal syndrome (HFRS) in humans. HFRS is characterised by a classical triad of fever, acute kidney injury and haemorrhage, but can show different signs and symptoms in specific cases. SEOV is transmitted from infected rats to humans by inhalation of aerosolised excreta. When compared with the known circulating hantaviruses in the Netherlands, Puumala (PUUV) and Tula (TULV), SEOV causes a more severe form of HFRS. Data from cohort studies undertaken in China and Northern Europe show differences in signs and symptoms at onset of disease, (haemorrhagic) complications and mortality. Furthermore, routine diagnostics currently available for hantavirus diagnosis in the Netherlands are not optimised for SEOV detection. The clinical outcome of an SEOV and PUUV infection will greatly benefit from an early diagnosis which will reduce the costs of unnecessary tests and treatments as well. The discovery of SEOV circulation in the Netherlands follows recent findings of SEOV infections in both rodents and humans in England, Wales, France, Belgium and Sweden, indicating the emerging character of SEOV and a high importance of this hantavirus for Public Health in large areas of Europe. Here, we review the current knowledge on the clinical manifestation of SEOV versus PUUV infections in humans, the treatment of clinical cases and diagnostics.


Predictors of long-term shunt-dependent hydrocephalus after aneurysmal subarachnoid hemorrhage. Clinical article.
The purpose of this study was to identify predictors of shunt-dependent hydrocephalus after aneurysmal subarachnoid hemorrhage (SAH). The authors evaluated the incidence of shunt-dependent hydrocephalus in a consecutive cohort of 580 patients with SAH who were admitted to the Neurological Intensive Care Unit of Columbia University Medical Center between July 1996 and September 2002. Patient demographics, 24-hour admission variables, initial CT scan characteristics, daily transcranial Doppler variables, and development of in-hospital complications were analyzed. Odds ratios and 95% CIs for candidate predictors were calculated using multivariate nominal logistic regression. Admission glucose of at least 126 mg/dl (adjusted OR 1.6; 95% CI 1.0-2.6), admission brain CT scan with a bicaudate index of at least 0.20 (adjusted OR 1.43; 95% CI 1.0-2.0), Fisher Grade 4 (adjusted OR 2.71; 95% CI 1.2-5.7), fourth ventricle hemorrhage (adjusted OR 1.78; 95% CI 1.1-2.7), and development of nosocomial meningitis (adjusted OR 2.2; 95% CI 1.4-3.7) were independently associated with shunt dependency. These data suggest that permanent CSF diversion after aneurysmal SAH may be independently predicted by hyperglycemia at admission, findings on the admission CT scan (Fisher Grade 4, fourth ventricle intraventricular hemorrhage, and bicaudate index ≥ 0.20), and development of nosocomial meningitis. Future research is needed to assess if tight glycemic control, reduction of fourth ventricle clot burden, and prevention of nosocomial meningitis may reduce the need for permanent CSF diversion after aneurysmal SAH.


Keratocystic odontogenic tumours of the jaws and associated pathologies: a 10-year clinicopathologic audit in a referral teaching hospital in Kenya.
To establish the pattern of occurrence and the clinicopathological features of keratocystic odontogenic tumour (KCOT) over a 10-year period. Patients from the University of Nairobi Dental Hospital treated for KCOT were included in the study over a 10-year period. The study highlights the demographic, clinico-radiological and histological features of these tumours. A total of 22 confirmed cases of KCOTs were recorded with equal gender prevalence; (M:F = 1.44:1). The age range of the patients was from 10 to 69 years with a peak in the second decade of life (mean = 27.5 yrs). Of the 22 cases, 15 (68.2%) occurred in the mandible of which eight (53.3%) involved the body, five (33.4%) the angle and ramus. Six (27.3%) occurred in the maxilla, and one (4.5%) was in both jaws and was associated with Gorlin-Goltz Syndrome. The most common presenting complaint in most patients was swelling 54.6%, and in 18.2% was incidental finding. Eight (36.4%) cases showed satellite cysts upon pathologic evaluation. Thirteen (59.1%) cases were managed by surgical excision, while nine (40.9%) were managed by enucleation. Based on the outcome of this study, KCOT present mostly in body, angle and ramus of the mandible and its peak is in the second decade of life.


Distant nodal metastasis: is it always an unresectable disease?
This article aims at analyzing the published literature concerning the treatment of patients with gastric cancer and distant nodal metastases, actually considered metastatic disease. A systematic search was undertaken using Medline, Embase, Cochrane and Web-of-Science libraries. No specific restriction on year of publication was used; preference was given to English papers. Both clinical series and literature reviews were selected. Only 11 papers address the issue of surgery for nodal basins outside the D2 dissection area. From these papers, in selected cases extended surgery may prove useful in prolonging survival, when a comprehensive therapeutic pathway including chemotherapy is scheduled. In conclusion, in presence of nodal metastases outside the loco-regional nodes, surgery may be considered for metastatic nodes in stations 13 and 16, in selected cases.


A prospective postmarket study to evaluate the safety and efficacy of a new peripherally inserted central catheter stabilization system.
An initial postmarket study of the SecurAcath used with 5 Fr peripherally inserted central catheters (PICCs) was conducted with 68 adult patients at 3 different institutions in the United States. PICCs were placed in both outpatients and inpatients, with patients in critical care and medical/surgical units, home care, and extended care facilities. Sixty-two (91.2%) of the patients completed therapy without a securement-related device malfunction or device-related adverse event associated with the securement system. The device was readily accepted by both patients and nursing staff. The SecurAcath represents a novel, safe, and effective method for catheter securement.


18 F-fluorothymidine uptake in follicular lymphoma and error-prone DNA repair.
We observed a disproportional 18 F-fluorothymidine (F-FLT) uptake in follicular lymphoma (FL) relative to its low cell proliferation. We tested the hypothesis that the 'excess' uptake of 18 F-FLT in FL is related to error-prone DNA repair and investigated whether this also contributes to 18 F-FLT uptake in diffuse large B cell lymphoma (DLBCL). We performed immunohistochemical stainings to assess the pure DNA replication marker MIB-1 as well as markers of both DNA replication and repair like PCNA, TK-1 and RPA1 on lymph node biopsies of 27 FLs and 35 DLBCLs. In 7 FL and 15 DLBCL patients, 18 F-FLT-PET had been performed. 18 F-FLT uptake was lower in FL than in DLBCL (median SUVmax 5.7 vs. 8.9, p = 0,004), but the ratio of 18 F-FLT-SUVmax to percentage of MIB-1 positive cells was significantly higher in FL compared with DLBCL (p = 0.001). The median percentage of MIB-1 positive cells was 10% (range, 10% to 20%) in FL and 70% (40% to 80%) in DLBCL. In contrast, the median percentages of PCNA, TK-1 and RPA1 positive cells were 90% (range, 80 to 100), 90% (80 to 100) and 100% (80 to 100) in FL versus 90% (60 to 100), 90% (60 to 100) and 100% (80 to 100) in DLBCL, respectively. This is the first demonstration of a striking discordance between 18 F-FLT uptake in FL and tumour cell proliferation. High expression of DNA replication and repair markers compared with the pure proliferation marker MIB-1 in FL suggests that this discordance might be due to error-prone DNA repair. While DNA repair-related 18 F-FLT uptake considerably contributes to 18 F-FLT uptake in FL, its contribution to 18 F-FLT uptake in highly proliferative DLBCL is small. This apparently high contribution of DNA repair to the 18 F-FLT signal in FL may hamper studies where 18 F-FLT is used to assess response to cytostatic therapy or to distinguish between FL and transformed lymphoma.


Long-term valproate and lamotrigine treatment may be a marker for reduced growth and bone mass in children with epilepsy.
To determine whether long-term treatment with valproate (VPA) and/or lamotrigine (LTG) in children with epilepsy is associated with altered growth and/or bone metabolism. Twenty-seven boys and 26 girls, aged 3 to 17 years (9.2 +/- 3.9, mean +/- SD), with epilepsy treated with VPA and/or LTG for > or =2 years were evaluated for growth, nutrient intakes, physical activity, bone mineral density (BMD), and blood biochemical indices of mineral and bone metabolism. Twenty-three (43.4%) of the children had a body height below the 10th percentile. Z-scores for BMD below -1.5 occurred in 24.4% of the children. When patients were divided into two groups according to daily activity score, a significantly lower Z-score for total body BMD (p = 0.007), percentile for body height (p = 0.05), and plasma parathyroid hormone (PTH; p = 0.04), osteocalcin (p = 0.04) and 25-hydroxyvitamin D (25OHD) (p = 0.01) were found in the inactive compared with the active group. Z-score for total body BMD was correlated with daily activity score (r = 0.43, p = 0.008). Plasma intact osteocalcin and intact PTH values correlated significantly (r = 0.36, p = 0.02). Plasma 1,25-dihydroxyvitamin D was within normal range for all subjects. When patients were divided into LTG-alone, VPA-alone, and LTG-plus-VPA treatment groups, significantly lower (p < 0.05) plasma osteocalcin and percentile for body height were found in the VPA-plus-LTG treatment group. Long-term VPA and LTG therapy, particularly when combined, is associated with short stature, low BMD, and reduced bone formation. These alterations may be mediated primarily through reduced physical activity rather than through a direct link to the VPA and/or LTG therapy.


Giving Voice to Patients: Developing a Discussion Method to Involve Patients in Translational Research.
Biomedical research policy in recent years has often tried to make such research more 'translational', aiming to facilitate the transfer of insights from research and development (R&D) to health care for the benefit of future users. Involving patients in deliberations about and design of biomedical research may increase the quality of R&D and of resulting innovations and thus contribute to translation. However, patient involvement in biomedical research is not an easy feat. This paper discusses the development of a method for involving patients in (translational) biomedical research aiming to address its main challenges. After reviewing the potential challenges of patient involvement, we formulate three requirements for any method to meaningfully involve patients in (translational) biomedical research. It should enable patients (1) to put forward their experiential knowledge, (2) to develop a rich view of what an envisioned innovation might look like and do, and (3) to connect their experiential knowledge with the envisioned innovation. We then describe how we developed the card-based discussion method 'Voice of patients', and discuss to what extent the method, when used in four focus groups, satisfied these requirements. We conclude that the method is quite successful in mobilising patients' experiential knowledge, in stimulating their imaginaries of the innovation under discussion and to some extent also in connecting these two. More work is needed to translate patients' considerations into recommendations relevant to researchers' activities. It also seems wise to broaden the audience for patients' considerations to other actors working on a specific innovation.


Healthy co-twins of patients with affective disorders show reduced risk-related activation of the insula during a monetary gambling task.
Healthy first-degree relatives of patients with affective disorders are at increased risk for affective disorders and express discrete structural and functional abnormalities in the brain reward system. However, value-based decision making is not well understood in these at-risk individuals. We investigated healthy monozygotic and dizygotic twins with or without a co-twin history of affective disorders (high-risk and low-risk groups, respectively) using functional MRI during a gambling task. We assessed group differences in activity related to gambling risk over the entire brain. We included 30 monozygotic and 37 dizygotic twins in our analysis. Neural activity in the anterior insula and ventral striatum increased linearly with the amount of gambling risk in the entire cohort. Individual neuroticism scores were positively correlated with the neural response in the ventral striatum to increasing gambling risk and negatively correlated with individual risk-taking behaviour. Compared with low-risk twins, the high-risk twins showed a bilateral reduction of risk-related activity in the middle insula extending into the temporal cortex with increasing gambling risk. Post hoc analyses revealed that this effect was strongest in dizygotic twins. The relatively old average age of the mono- and dizygotic twin cohort (49.2 yr) may indicate an increased resilience to affective disorders. The size of the monozygotic high-risk group was relatively small (n = 13). The reduced processing of risk magnitude in the middle insula may indicate a deficient integration of exteroceptive information related to risk-related cues with interoceptive states in individuals at familial risk for affective disorders. Impaired risk processing might contribute to increased vulnerability to affective disorders.


Protocol: Chromatin immunoprecipitation (ChIP) methodology to investigate histone modifications in two model diatom species.
In this report we describe a chromatin immunoprecipitation (ChIP) protocol for two fully sequenced model diatom species Phaeodactylum tricornutum and Thalassiosira pseudonana. This protocol allows the extraction of satisfactory amounts of chromatin and gives reproducible results. We coupled the ChIP assay with real time quantitative PCR. Our results reveal that the two major histone marks H3K4me2 and H3K9me2 exist in P. tricornutum and T. pseudonana. As in other eukaryotes, H3K4me2 marks active genes whereas H3K9me2 marks transcriptionally inactive transposable elements. Unexpectedly however, T. pseudonana housekeeping genes also show a relative enrichment of H3K9me2. We also discuss optimization of the procedure, including growth conditions, cross linking and sonication. Validation of the protocol provides a set of genes and transposable elements that can be used as controls for studies using ChIP in each diatom species. This protocol can be easily adapted to other diatoms and eukaryotic phytoplankton species for genetic and biochemical studies.


Clinical validation of an autoantibody test for lung cancer.
Autoantibodies may be present in a variety of underlying cancers several years before tumours can be detected and testing for their presence may allow earlier diagnosis. We report the clinical validation of an autoantibody panel in newly diagnosed patients with lung cancer (LC). Three cohorts of patients with newly diagnosed LC were identified: group 1 (n = 145), group 2 (n = 241) and group 3 (n = 269). Patients were individually matched by gender, age and smoking history to a control individual with no history of malignant disease. Serum samples were obtained after diagnosis but before any anticancer treatment. Autoantibody levels were measured against a panel of six tumour-related antigens (p53, NY-ESO-1, CAGE, GBU4-5, Annexin 1 and SOX2). Assay sensitivity was tested in relation to demographic variables and cancer type/stage. The autoantibody panel demonstrated a sensitivity/specificity of 36%/91%, 39%/89% and 37%/90% in groups 1, 2 and 3, respectively, with good reproducibility. There was no significant difference between different LC stages, indicating that the antigens included covered the different types of LC well. This assay confirms the value of an autoantibody panel as a diagnostic tool and offers a potential system for monitoring patients at high risk of LC.


Antibody-like Biorecognition Sites for Proteins from Surface Imprinting on Nanoparticles.
Natural antibodies are used widely for important applications such as biomedical analysis, cancer therapy, and directed drug delivery, but they are expensive and may have limited stability. This study describes synthesis of antibody-like binding sites by molecular imprinting on silica nanoparticles (SiNP) using a combination of four organosilane monomers with amino acid-like side chains providing hydrophobic, hydrophilic, and H-bonding interactions with target proteins. This approach provided artificial antibody (AA) nanoparticles with good selectivity and specificity to binding domains on target proteins in a relatively low-cost synthesis. The AAs were made by polymer grafting onto SiNPs for human serum albumin (HSA) and glucose oxidase (GOx). Binding affinity, selectivity, and specificity was compared to several other proteins using adsorption isotherms and surface plasmon resonance (SPR). The Langmuir-Freundlich adsorption model was used to obtain apparent binding constants (KLF) from binding isotherms of HSA (6.7 × 10(4)) and GOx (4.7 × 10(4)) to their respective AAs. These values were 4-300 fold larger compared to a series of nontemplate proteins. SPR binding studies of AAs with proteins attached to a gold surface confirmed good specificity and revealed faster binding for the target proteins compared to nontarget proteins. Target proteins retained their secondary structures upon binding. Binding capacity of AAHSA for HSA was 5.9 mg HSA/g compared to 1.4 mg/g for previously report imprinted silica beads imprinted with poly(aminophenyl)boronic acid. Also, 90% recovery for HSA spiked into 2% calf serum was found for AAHSA.


Racial-ethnic disparities in acute blood pressure after intracerebral hemorrhage.
To assess race-ethnic differences in acute blood pressure (BP) following intracerebral hemorrhage (ICH) and the contribution to disparities in ICH outcome. BPs in the field (emergency medical services [EMS]), emergency department (ED), and at 24 hours were compared and adjusted for group differences between non-Hispanic black (black), non-Hispanic white (white), and Hispanic participants in the Ethnic Racial Variations of Intracerebral Hemorrhage case-control study. Outcome was obtained by modified Rankin Scale (mRS) score at 3 months. We analyzed race-ethnic differences in good outcome (mRS ≤ 2) and mortality after adjusting for baseline differences and included BP recordings in this model. Of 2,069 ICH cases enrolled, 30% were white, 37% black, and 33% Hispanic. Black and Hispanic patients had higher EMS and ED systolic and diastolic BPs compared with white patients (p = 0.0001). Although attenuated, at 24 hours after admission, black patients had higher systolic and diastolic BPs. After adjusting for baseline differences, significant race/ethnic differences persisted for EMS systolic, ED systolic and diastolic, and 24-hours diastolic BP. Only ED systolic and diastolic BP was associated with poor functional outcome, and no BP predicted mortality. We found no race-ethnic differences in 3-month functional outcome or mortality after adjusting for group differences, including acute BPs. Although black and Hispanic patients had higher BPs than white patients at presentation, we did not find race-ethnic disparities in 3-month functional outcome or mortality. ED systolic and diastolic BP was associated with poor functional outcome, but not mortality, in this race-ethnically diverse population.


Molecular mechanisms involved in tumor repopulation after radiotherapy.
Radiotherapy remains one of most important treatment modalities for solid tumors. Current radiotherapy is mostly based on a set of concepts called the 4"R"s, which were established when there was lack of understanding of the underlying molecular mechanisms. However, progress made in the past two decades are beginning to allow us to see some of the molecular details involved in tumor response to radiation therapy. In this review, we will attempt to summarize some of the key discoveries in molecular radiation biology that have direct relevance to radiotherapy. We will focus our discussion on areas such as radiation induced tumor vasculogenesis, stem cell mobilization, and cellular repopulation. We hope our discussion will stimulate further studies in this important area of cancer research.


Multicenter prospective cohort study of the incidence of adverse events associated with cosmetic dermatologic procedures: lasers, energy devices, and injectable neurotoxins and fillers.
Common noninvasive to minimally invasive cosmetic dermatologic procedures are widely believed to be safe given the low incidence of reported adverse events, but reliable incidence data regarding adverse event rates are unavailable to date. To assess the incidence of adverse events associated with noninvasive to minimally invasive cosmetic dermatologic procedures, including those involving laser and energy devices, as well as injectable neurotoxins and fillers. A multicenter prospective cohort study (March 28, 2011, to December 30, 2011) of procedures performed using laser and energy devices, as well as injectable neurotoxins and soft-tissue augmentation materials, among 8 geographically dispersed US private and institutional dermatology outpatient clinical practices focused on cosmetic dermatology, with a total of 23 dermatologists. Participants represented a consecutive sample of 20 399 cosmetic procedures. Data acquisition was for 3 months (13 weeks) per center, with staggered start dates to account for seasonal variation. Web-based data collection daily at each center to record relevant procedures, by category type and subtype. Adverse events were detected by (1) initial observation by participating physicians or staff; (2) active ascertainment from patients, who were encouraged to self-report after their procedure; and (3) follow-up postprocedural phone calls to patients by staff, if appropriate. When adverse events were not observed by physicians but were suspected, follow-up visits were scheduled within 24 hours to characterize these events. Detailed information regarding each adverse event was entered into an online form. The main outcome was the total incidence of procedure-related adverse events (total adverse events divided by total procedures performed), as verified by clinical examination. Forty-eight adverse events were reported, for a rate of 0.24% (95% CI, 0.18%-0.31%). Overall, 36 procedures resulted in at least 1 adverse event, for a rate of 0.18% (95% CI, 0.13%-0.25%). No serious adverse events were reported. Adverse events were infrequently associated with known risk factors. Noninvasive to minimally invasive cosmetic dermatologic procedures, including energy, neurotoxin, and filler procedures, are safe when performed by experienced board-certified dermatologists. Adverse events occur in less than 1% of patients, and most of these are minor and transient.


Comparison of characteristics between patients with H7N9 living in rural and urban areas of Zhejiang Province, China: a preliminary report.
A total of 134 cases of H7N9 influenza infection were identified in 12 provinces of China between March 25 and September 31, 2013. Of these, 46 cases occurred in Zhejiang Province. We carried out a preliminary comparison of characteristics between rural and urban H7N9 cases from Zhejiang Province, China. Field investigations were conducted for each confirmed H7N9 case. A standardized questionnaire was used to collect information about demographics, exposure history, clinical signs and symptoms, timelines of medical visits and care after onset of illness. Of the 46 H7N9 cases in Zhejiang Province identified between March 25 and September 31, 2013, there were 16 rural cases and 30 urban cases. Compared to urban cases, there was a higher proportion of females among the rural cases [11/16 (69%) vs. 6/30 (20%), P = 0.001]. Among the rural cases, 14/15 (93%) with available data had a history of recent poultry exposure, which was significantly higher than that among urban cases (64%, P = 0.038). More patients from the rural group had a history of breeding poultry compared with those from the urban group [38% (6/16) vs. 10% (3/30), respectively; P = 0.025]. Interestingly, the median number of medical visits of patients from rural areas was higher than that of patients from urban areas (P = 0.046). There was no difference between the two groups in terms of age distribution, fatality rate, incubation period, symptoms, and underlying medical conditions. In conclusion, compared to patients from urban areas, more patients from rural areas were female, had an exposure history, had a history of breeding poultry, and had a higher number of medical visits. These findings indicate that there are different exposure patterns between patients living in rural and urban areas and that more rural cases were infected through backyard poultry breeding.


Calcification, storm damage and population resilience of tabular corals under climate change.
Two facets of climate change--increased tropical storm intensity and ocean acidification--are expected to detrimentally affect reef-building organisms by increasing their mortality rates and decreasing their calcification rates. Our current understanding of these effects is largely based on individual organisms' short-term responses to experimental manipulations. However, predicting the ecologically-relevant effects of climate change requires understanding the long-term demographic implications of these organism-level responses. In this study, we investigate how storm intensity and calcification rate interact to affect population dynamics of the table coral Acropora hyacinthus, a dominant and geographically widespread ecosystem engineer on wave-exposed Indo-Pacific reefs. We develop a mechanistic framework based on the responses of individual-level demographic rates to changes in the physical and chemical environment, using a size-structured population model that enables us to rigorously incorporate uncertainty. We find that table coral populations are vulnerable to future collapse, placing in jeopardy many other reef organisms that are dependent upon them for shelter and food. Resistance to collapse is largely insensitive to predicted changes in storm intensity, but is highly dependent on the extent to which calcification influences both the mechanical properties of reef substrate and the colony-level trade-off between growth rate and skeletal strength. This study provides the first rigorous quantitative accounting of the demographic implications of the effects of ocean acidification and changes in storm intensity, and provides a template for further studies of climate-induced shifts in ecosystems, including coral reefs.


Association of Cognitive Impairment in Patients on 3-Hydroxy-3-Methyl-Glutaryl-CoA Reductase Inhibitors.
Atherosclerotic cardiovascular diseases are the leading cause of death in the United States. A reduction in cholesterol with 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors (statin) significantly reduces mortality and morbidity. Statins may be associated with cognitive impairment or dementia. Our aim was to study the association of cognitive impairment or dementia in patients who were on a statin. Electronic medical records of 3,500 adult patients in our suburban internal medicine office were reviewed. There were 720 (20.6%) patients in the statin treatment group. Dementia or cognitive impairment was an associated comorbid condition in 7.9% patients in the statin treatment group compared to 3.1% patients in the non-statin group (P < 0.001). Analysis of all of the patients with cognitive impairment or dementia showed that among the age ranges of 51 years through 100 years, the patients in the statin treatment group had a higher prevalence of cognitive impairment or dementia compared to the non-statin group. In the statin treatment group, we found significantly higher prevalence of hyperlipidemia (86.3%), hypertension (69.6%), diabetes mellitus (36.0%), osteoarthritis (31.5%), coronary artery disease (26.1%), hypothyroidism (21.5%) and depression (19.3%) compared to the non-statin group (P < 0.001). About 39.9% of the patients with dementia or cognitive impairment were on statin therapy compared to 18.9% patients who had no dementia or cognitive impairment and were on statin therapy (P < 0.001). Among the patients with cognitive deficit or dementia in the statin treatment group, the majority of the patients were either on atorvastatin (43.9%) or simvastatin (35.1%), followed by rosuvastatin (12.2%) and pravastatin (8.8%). We found greater odds of dementia or cognitive impairment with each year increase in age (1.3 times), in women (2.2 times), African American race (2.7 times), non-consumption of moderate amount of alcohol (two times), diabetes mellitus (1.6 times), hypothyroidism (1.7 times), cerebrovascular accident (3.2 times), and other rheumatological diseases (1.8 times). The association of dementia or cognitive impairment was significantly higher in the patients who were on statin therapy compared to the patients who were not on a statin.


The Role of Pre-existing Cross-Reactive Central Memory CD4 T-Cells in Vaccination With Previously Unseen Influenza Strains.
Influenza vaccination is a common approach to prevent seasonal and pandemic influenza. Pre-existing antibodies against close viral strains might impair antibody formation against previously unseen strains-a process called original antigenic sin. The role of this pre-existing cellular immunity in this process is, despite some hints from animal models, not clear. Here, we analyzed cellular and humoral immunity in healthy individuals before and after vaccination with seasonal influenza vaccine. Based on influenza-specific hemagglutination inhibiting (HI) titers, vaccinees were grouped into HI-negative and -positive cohorts followed by in-depth cytometric and TCR repertoire analysis. Both serological groups revealed cross-reactive T-cell memory to the vaccine strains at baseline that gave rise to the majority of vaccine-specific T-cells post vaccination. On the contrary, very limited number of vaccine-specific T-cell clones was recruited from the naive pool. Furthermore, baseline quantity of vaccine-specific central memory helper T-cells and clonotype richness of this population directly correlated with the vaccination efficacy. Our findings suggest that the deliberate recruitment of pre-existing cross-reactive cellular memory might help to improve vaccination outcome.


Acute myeloid leukaemia blast cells with a tyrosine kinase domain mutation of FLT3 are less sensitive to lestaurtinib than those with a FLT3 internal tandem duplication.
FLT3 tyrosine kinase domain mutations (FLT3/TKDs) are associated with a favourable prognosis in acute myeloid leukaemia (AML), unlike FLT3 internal tandem duplications (FLT3/ITDs) that have a poor prognosis. Whilst FLT3/ITD+ cells are more susceptible to the cytotoxic effects of FLT3 inhibitors than wild type (WT) cells, the sensitivity of FLT3/TKD+ cells to therapeutic agents is unclear, as is the importance of the mutant level. We therefore studied the effect of cytarabine and the FLT3 inhibitor lestaurtinib, either alone or in combination, on in vitro survival of blast cells from 36 cases of AML (14 FLT3/WT, 11 FLT3/ITD+ and 11 FLT3/TKD+). All three groups showed similar sensitivity to the cytotoxic effects of cytarabine but FLT3/ITD mutant level was inversely correlated with cytarabine cytotoxicity (P = 0.04) whereas FLT3/TKD mutant level had no impact. FLT3/TKD+ cells showed a similar response to lestaurtinib as FLT3/WT cells, whereas FLT3/ITD+ cells were more sensitive (P = 0.004). There was no correlation between mutant level and lestaurtinib sensitivity for either FLT3/ITD+ or FLT3/TKD+ cells. Synergistic cytotoxicity of lestaurtinib plus cytarabine was demonstrated in all three groups. These results suggest that FLT3/TKD+ and FLT3/WT cases should not be differentiated when considering patients for treatment with FLT3 inhibitors.


Genome sequence of Halomonas hydrothermalis Y2, an efficient ectoine-producer isolated from pulp mill wastewater.
Halophilic microorganisms have great potentials towards biotechnological applications. Halomonas hydrothermalis Y2 is a halotolerant and alkaliphilic strain that isolated from the Na+-rich pulp mill wastewater. The strain is dominant in the bacterial community of pulp mill wastewater and exhibits metabolic diversity in utilizing various substrates. Here we present the genome sequence of this strain, which comprises a circular chromosome 3,933,432 bp in size and a GC content of 60.2%. Diverse genes that encoding proteins for compatible solutes synthesis and transport were identified from the genome. With a complete pathway for ectoine synthesis, the strain could produce ectoine from monosodium glutamate and further partially secreted into the medium. In addition, around 20% ectoine was increased by deleting the ectoine hydroxylase (EctD). The genome sequence we report here will provide genetic information regarding adaptive mechanisms of strain Y2 to its harsh habitat, as well as facilitate exploration of metabolic strategies for diverse compatible solutes, e.g., ectoine production.


Gallstones: genetics versus environment.
The aim of this study was to determine if a significant genetic component contributes to the pathogenesis of symptomatic gallstones. Gallstones represent a polygenic disorder that affects more than 30,000,000 Americans and results in more than 750,000 cholecystectomies in the United States annually. Risk factors include age, gender, race, parity, obesity, and diabetes. A family history of gallstones also has been identified as a risk factor suggesting that genetics play a role in gallstone formation. However, the role of genetics in the pathogenesis of gallstone formation has not been determined. A gallbladder disease-specific questionnaire was administered to 904 healthy unrelated adult volunteers (association study). The questionnaire ascertained a history of cholecystectomy and gallstone disease in first-degree relatives, as well as medical history, demographic, and anthropometric data. A logistic regression model was used to identify risk factors for symptomatic gallstone disease in a multivariate analysis. A maximum likelihood based variance decomposition approach was then used in 1,038 individuals from 358 families (family study) to estimate the additive genetic heritability of symptomatic gallstone disease. In the association study significant risk factors for symptomatic gallstone disease were female gender (relative risk 8.8, P <.003), obesity (BMI > 30, relative risk 3.7, P <.001), age > 50 (relative risk 2.5, P <.001), and a positive family history of previous cholecystectomy in a first-degree family member (relative risk 2.2, P <.01). In the family study the additive genetic heritability of symptomatic gallstones was 29% (P <.02), age and gender were significant covariates and explained 9.3% of the phenotypic variation in gallbladder disease. These data suggest that genetic factors are responsible for at least 30% of symptomatic gallstone disease. However, the true role of heredity in gallstone pathogenesis is probably higher because data based on symptomatic gallbladder disease underestimates the true prevalence in the population.


Differences in Thallium-201 uptake in reperfused and nonreperfused myocardial infarction.
The respective importance of flow and cellular viability in determining initial myocardial thallium uptake was studied in reperfused and nonreperfused experimental myocardial infarction. Open-chest dogs were subjected to permanent coronary artery occlusion of 70-minute (n = 3) or 5-hour duration (n = 5), or to a 3-hour temporary occlusion followed by reflow (n = 14). Thallium uptake 10 minutes after intravenous injection was compared directly with radioactive microspheres in myocardial samples from excised hearts. Triphenyl tetrazolium chloride staining was used to differentiate necrotic and viable samples with confirmation by electron microscopy. In nonreperfused infarcts, thallium uptake occurred despite necrosis, and a close correlation was found between thallium uptake and regional myocardial blood flow. In reperfused infarcts, thallium uptake again occurred, but was reduced relative to flow in necrotic myocardium and, to a lesser extent, in reperfused viable areas. However, because of the high levels of reflow, actual thallium uptake was often more than 50% of normal in reperfused necrotic regions. This study demonstrates that the presence of thallium uptake is an unreliable indicator of myocardial injury and that reperfused necrotic tissue may have remarkably high levels of thallium uptake.


Autoimmune disorders of the neuromuscular junction.
The neuromuscular junction lies beyond the protection of the blood-brain barrier and is particularly vulnerable to antibody-mediated attack. In myasthenia gravis, the expression of acetylcholine receptors (AChRs) in the thymus is under the control of the autoimmune regulator protein (AIRE), and polymorphisms in the AChR correlate with early onset of disease. In some 'AChR seronegative' patients, thymic abnormalities associated with complement-activating antibodies binding only clustered AChRs have been demonstrated, and in others anti-muscle-specific kinase (MuSK) antibodies that show pathogenic effects in vivo. In Guillain-Barré syndrome, newly described antibodies bind to complex gangliosides. General immunosuppression is still the main treatment, but novel treatments that reduce complement-mediated damage or inhibit the binding of pathogenic antibodies are beginning to look promising.


Manganese: recent advances in understanding its transport and neurotoxicity.
The present review is based on presentations from the meeting of the Society of Toxicology in San Diego, CA (March 2006). It addresses recent developments in the understanding of the transport of manganese (Mn) into the central nervous system (CNS), as well as brain imaging and neurocognitive studies in non-human primates aimed at improving our understanding of the mechanisms of Mn neurotoxicity. Finally, we discuss potential therapeutic modalities for treating Mn intoxication in humans.


Unpacking Transient Event Dynamics in Electrophysiological Power Spectra.
Electrophysiological recordings of neuronal activity show spontaneous and task-dependent changes in their frequency-domain power spectra. These changes are conventionally interpreted as modulations in the amplitude of underlying oscillations. However, this overlooks the possibility of underlying transient spectral 'bursts' or events whose dynamics can map to changes in trial-average spectral power in numerous ways. Under this emerging perspective, a key challenge is to perform burst detection, i.e. to characterise single-trial transient spectral events, in a principled manner. Here, we describe how transient spectral events can be operationalised and estimated using Hidden Markov Models (HMMs). The HMM overcomes a number of the limitations of the standard amplitude-thresholding approach to burst detection; in that it is able to concurrently detect different types of bursts, each with distinct spectral content, without the need to predefine frequency bands of interest, and does so with less dependence on a priori threshold specification. We describe how the HMM can be used for burst detection and illustrate its benefits on simulated data. Finally, we apply this method to empirical data to detect multiple burst types in a task-MEG dataset, and illustrate how we can compute burst metrics, such as the task-evoked timecourse of burst duration.


Impact of a spreading epidemic on medical students.
The emergence of severe acute respiratory syndrome (SARS) had caused fear and anxiety of unprecedented proportion. To examine the impact of SARS on the medical students in a private medical university, a self-reporting questionnaire study was carried out to assess the factual knowledge, anxiety level and perception of the crisis, among the students. The two-week study (between 12 and 23 May, 2003) was carried out three weeks after the first reported SARS-related death in Malaysia. Ninety-one Phase I (junior) and 113 Phase II (senior) students completed the questionnaires. A large majority of students of Phase I and II were correct in their factual knowledge and were sensible in their perception of the future and the handling of the crisis by government(s). However, phase 1 students expressed significantly greater degree of anxiety compared to Phase II in relation to attendance and personal protection in hospital, and in meeting people coughing in public places. The lesser degree of anxiety expressed by phase II senior students may be due in part, to a more realistic assessment of SARS risk brought about by maturity, time spent in hospital and interaction with clinical lecturers and medical staff.


Evidence from vergence eye movements that disparities defined by luminance and contrast are sensed by independent mechanisms.
We recorded the initial disparity vergence responses (DVRs) elicited by 1-D sinusoidal gratings differing in phase at the two eyes by 1/4 wavelength and defined by luminance modulation (LM) or contrast modulation (CM) of dynamic binary noise. Both LM and CM stimuli elicited DVRs, but those to CM had longer latency (on average by ∼20 ms). DVRs showed sigmoidal dependence on depth of modulation, with higher thresholds for CM than for LM. With both LM and CM stimuli, fixing the modulation at one eye well above threshold rendered the DVR hypersensitive to low-level modulation at the other eye (dichoptic facilitation). Disparities defined by LM at one eye and CM at the other generated weak DVRs in the "wrong" direction, consistent with mediation entirely by distortion products associated with the CM stimulus. These (reversed) DVRs could be nulled by adding LM to the CM stimulus (in phase), and the greater the depth of the CM, the greater the added LM required for nulling, exactly as predicted by a simple compressive non-linearity. We conclude that disparities defined by LM and by CM are sensed by independent cortical mechanisms, at least for the purposes of generating short-latency vergence eye movements to disparity steps.


Rosetting Responses of Plasmodium-infected Erythrocytes to Antimalarials.
In malaria, rosetting is a phenomenon involving the cytoadherence of uninfected erythrocytes to infected erythrocytes (IRBC) harboring the late erythrocytic stage of Plasmodium spp. Recently, artesunate-stimulated rosetting has been demonstrated to confer a survival advantage to P. falciparum late-stage IRBC. This study investigated the rosetting response of P. falciparum and P. vivax clinical isolates to ex vivo antimalarial treatments. Brief exposure of IRBC to chloroquine, mefloquine, amodiaquine, quinine, and lumefantrine increased the rosetting rates of P. falciparum and P. vivax. Furthermore, the ex vivo combination of artesunate with mefloquine and piperaquine also resulted in increased the rosetting rates. Drug-mediated rosette-stimulation has important implications for the therapeutic failure of rapidly cleared drugs such as artesunate. However, further work is needed to establish the ramifications of increased rosetting rates by drugs with longer half-lifves, such as chloroquine, mefloquine, and piperaquine.


Differential pupillary constriction and awareness in the absence of striate cortex.
The fact that the pupil constricts differentially to visual stimuli in the absence of changes in light energy makes it a valuable tool for studying normal function as well as residual capacity in hemianopic subjects. When pupillometrically effective stimuli such as equiluminant gratings or coloured patches with an abrupt onset and offset are presented to the 'blind' hemifield, a hemianopic subject with damage largely restricted to striate cortex (V1) sometimes reports being 'aware' of the transient onset/offset, although without 'seeing' as such. The question addressed here is whether the pupil still responds in the condition of blindsight in its strict sense--i.e. discriminative capacity in the absence of acknowledged awareness--when stimuli are deliberately designed to eliminate awareness. This was accomplished by making stimulus onset and offset slow and gradual. The results with a well-studied hemianope, G.Y., demonstrate that there is still a pupillary constriction to isoluminant achromatic gratings and red-coloured stimuli, although reduced in size, in the absence of acknowledged awareness.


BALL-SNPgp-from genetic variants toward computational diagnostics.
In medical research, it is crucial to understand the functional consequences of genetic alterations, for example, non-synonymous single nucleotide variants (nsSNVs). NsSNVs are known to be causative for several human diseases. However, the genetic basis of complex disorders such as diabetes or cancer comprises multiple factors. Methods to analyze putative synergetic effects of multiple such factors, however, are limited. Here, we concentrate on nsSNVs and present BALL-SNPgp, a tool for structural and functional characterization of nsSNVs, which is aimed to improve pathogenicity assessment in computational diagnostics. Based on annotated SNV data, BALL-SNPgp creates a three-dimensional visualization of the encoded protein, collects available information from different resources concerning disease relevance and other functional annotations, performs cluster analysis, predicts putative binding pockets and provides data on known interaction sites. BALL-SNPgp is based on the comprehensive C ++ framework Biochemical Algorithms Library (BALL) and its visualization front-end BALLView. Our tool is available at www.ccb.uni-saarland.de/BALL-SNPgp email@example.com.


Science, institutional archives and open access: an overview and a pilot survey on the Italian cancer research institutions.
The Open Archive Initiative (OAI) refers to a movement started around the '90 s to guarantee free access to scientific information by removing the barriers to research results, especially those related to the ever increasing journal subscription prices. This new paradigm has reshaped the scholarly communication system and is closely connected to the build up of institutional repositories (IRs) conceived to the benefit of scientists and research bodies as a means to keep possession of their own literary production. The IRs are high-value tools which permit authors to gain visibility by enabling rapid access to scientific material (not only publications) thus increasing impact (citation rate) and permitting a multidimensional assessment of research findings. A survey was conducted in March 2010 to mainly explore the managing system in use for archiving the research finding adopted by the Italian Scientific Institutes for Research, Hospitalization and Health Care (IRCCS) of the oncology area within the Italian National Health Service (Servizio Sanitario Nazionale, SSN). They were asked to respond to a questionnaire intended to collect data about institutional archives, metadata formats and posting of full-text documents. The enquiry concerned also the perceived role of the institutional repository DSpace ISS, built up by the Istituto Superiore di Sanità (ISS) and based on a XML scheme for encoding metadata. Such a repository aims at acting as a unique reference point for the biomedical information produced by the Italian research institutions. An in-depth analysis has also been performed on the collection of information material addressed to patients produced by the institutions surveyed. The survey respondents were 6 out of 9. The results reveal the use of different practices and standard among the institutions concerning: the type of documentation collected, the software adopted, the use and format of metadata and the conditions of accessibility to the IRs. The Italian research institutions in the field of oncology are moving the first steps towards the philosophy of OA. The main effort should be the implementation of common procedures also in order to connect scientific publications to researchers curricula. In this framework, an important effort is represented by the project of ISS aimed to set a common interface able to allow migration of data from partner institutions to the OA compliant repository DSpace ISS.


De-repression of myelin-regulating gene expression after status epilepticus in mice lacking the C/EBP homologous protein CHOP.
The C/EBP homologous protein CHOP is normally present at low levels in cells but increases rapidly after insults such as DNA damage or endoplasmatic reticulum stress where it contributes to cellular homeostasis and apoptosis. By forming heterodimers with other transcription factors, CHOP can either act as a dominant-negative regulator of gene expression or to induce the expression of target genes. Recent work demonstrated that seizure-induced hippocampal damage is significantly worse in mice lacking CHOP and these animals go on to develop an aggravated epileptic phenotype. To identify novel CHOP-controlled target genes which potentially influence the epileptic phenotype, we performed a bioinformatics analysis of tissue microarrays from chop-deficient mice after prolonged seizures. GO analysis revealed genes associated with biological membranes were prominent among those in the chop-deficient array dataset and we identified myelin-associated genes to be particularly de-repressed. These data suggest CHOP might act as an inhibitor of myelin-associated processes in the brain and could be targeted to influence axonal regeneration or reorganisation.


Induction chemotherapy followed by allogeneic HCT versus upfront allogeneic HCT for advanced myelodysplastic syndrome: A propensity score matched analysis.
To reduce post-transplant relapse, acute myeloid leukemia (AML) type remission induction chemotherapy has been attempted to reduce disease burden before allogeneic hematopoietic cell transplantation (HCT) in patients with advanced myelodysplastic syndrome (MDS). However, the efficacy of induction chemotherapy before HCT is unclear. We retrospectively analyzed the Japanese registration data of 605 adult patients, who had received allogeneic HCT for advanced MDS between 2001 and 2016, to compare the post-transplant relapse between patients who received induction chemotherapy followed by allogeneic HCT and those who received upfront HCT. Propensity score matching identified 230 patients from each cohort. There were no significant differences in overall survival and non-relapse mortality between the two groups. The cumulative incidence of relapse was significantly higher in patients who received induction chemotherapy than those who received upfront HCT. In the subgroup analyses, upfront HCT had a significantly reduced relapse incidence among patients with poor cytogenetics, those with higher international prognostic scoring system at diagnosis, and those who received reduced-intensity conditioning. Our results suggested that AML type remission induction chemotherapy before HCT did not improve post-transplant relapse and survival for adult patients with advanced MDS. Upfront HCT is preferable for patients with a poor karyotype.


Attaining realistic and substantial reductions in HIV incidence: model projections of combining microbicide and male circumcision interventions in rural Uganda.
This study simulates the potential impact of male circumcision and female microbicide interventions, singularly and in combination, in rural Uganda. A dynamic model was parameterised and fitted to setting-specific data, and used to estimate the impact on HIV transmission of a microbicide and/or male circumcision intervention over 15 years. The proportion of circumcised men or women using microbicides was assumed to increase linearly from 0.18 (male circumcision) or 0.00 (microbicide use) to the final proportion 10 years later, then remain constant for 5 years. Women using microbicides were assumed to use the product in 80% of penile-vaginal sex acts. Male circumcision or microbicide use was assumed to reduce the per-act probability of HIV acquisition in men or women, respectively, by 60%. Independently, to obtain a 30% relative reduction in HIV incidence at 15 years, the model suggests that the final proportion of women using microbicides would need to be 0.91 (95% CI 0.75 to 1.00) or the proportion of circumcised men would need to be 0.96 (0.83 to impact not possible). The same impact could be achieved by combining the interventions, eg, if the proportion of women using microbicides was 0.49 (0.39 to 0.56) and the proportion of circumcised men was 0.67 (0.57 to 0.74). Under these assumptions, as separate interventions it is unlikely that increases in either the proportion of men circumcised or of women using microbicides could reduce HIV incidence by 30% or more at 15 years. A combination-prevention strategy using complementary interventions may be a more feasible approach to achieve substantial reductions in incidence.


Responsible use of organoids in precision medicine: the need for active participant involvement.
Organoids are three-dimensional multicellular structures grown in vitro from stem cells and which recapitulate some organ function. They are derivatives of living tissue that can be stored in biobanks for a multitude of research purposes. Biobank research on organoids derived from patients is highly promising for precision medicine, which aims to target treatment to individual patients. The dominant approach for protecting the interests of biobank participants emphasizes broad consent in combination with privacy protection and ex ante (predictive) ethics review. In this paradigm, participants are positioned as passive donors; however, organoid biobanking for precision medicine purposes raises challenges that we believe cannot be adequately addressed without more ongoing involvement of patient-participants. In this Spotlight, we argue why a shift from passive donation towards more active involvement is particularly crucial for biobank research on organoids aimed at precision medicine, and suggest some approaches appropriate to this context.


Expression of DOF genes identifies early stages of vascular development in Arabidopsis leaves.
The sequence of events underlying the formation of vascular networks in the leaf has long fascinated developmental biologists. In Arabidopsis leaves, vascular-precursor procambial cells derive from the elongation of morphologically inconspicuous ground cells that selectively activate expression of the HD-ZIP III gene ATHB8. Inception of ATHB8 expression operationally defines acquisition of a typically irreversible preprocambial cell state that preludes to vein formation. A view of the constellation of genes whose expression is activated at preprocambial stages would therefore be particularly desirable; however, very few preprocambial gene expression profiles have been identified. Here, we show that expression of three genes encoding members of the DOF family of plant-specific transcription factors is activated at stages overlapping onset of ATHB8 expression. Expression of DOF genes is initiated in wide domains that become confined to sites of vein development. Congruence between DOF expression fields and zones of vein formation persists upon experimental manipulation of leaf vascular patterning, suggesting that DOF expression identifies consistently recurring steps in vein ontogeny. Our results contribute to defining preprocambial cell identity at the molecular level.


Sex difference in the association of the angiotensin converting enzyme I/D polymorphism and body mass index.
Angiotensin converting enzyme (ACE) catayzes the formation of angiotensin I to angiotensin II. A polymorphism has been identified in intron 16 in which a 287 base-pair alu sequence was found to be present (insertion or I) or absent (deletion or D) in the population. ACE and the components of the renin-angiotensin system are expressed in adipose tissue and therefore the I/D polymorphism within ACE may be associated with obesity. This study involved genotyping two groups with differing body mass indexes (BMI <or=25 and BMI >or=30) that were composed primarily of middle-age Caucasian subjects (n=421). The male groups differed significantly in allele frequency at the ACE locus with the I allele more frequent in the BMI >or=30 group (p<0.05). While the female BMI >or=30 group also had a higher I allele frequency than the BMI <or=25 group, the difference is not significant. Our results demonstrate a sex difference in the association of the I/D polymorphism and BMI. Our findings differ from some other previous studies using various population groups which indicate that the ACE I/D polymorphism may be differentially associated with obesity depending on multiple factors.


Is preclinical data sharing the new norm?
Is preclinical data sharing the new norm? In my experience, it is certainly becoming more commonplace. However, it is not yet standard practice and remains the preserve of special projects. Here, I expound the benefits of sharing proprietary preclinical data using examples of successful initiatives. The main barriers to data sharing are then described, with suggestions for how these might be overcome. To maximise the benefits and minimise the risks involved, I suggest that organisations look to develop standard operating procedures for data sharing.


High-throughput single-cell sequencing identifies photoheterotrophs and chemoautotrophs in freshwater bacterioplankton.
Recent discoveries suggest that photoheterotrophs (rhodopsin-containing bacteria (RBs) and aerobic anoxygenic phototrophs (AAPs)) and chemoautotrophs may be significant for marine and freshwater ecosystem productivity. However, their abundance and taxonomic identities remain largely unknown. We used a combination of single-cell and metagenomic DNA sequencing to study the predominant photoheterotrophs and chemoautotrophs inhabiting the euphotic zone of temperate, physicochemically diverse freshwater lakes. Multi-locus sequencing of 712 single amplified genomes, generated by fluorescence-activated cell sorting and whole genome multiple displacement amplification, showed that most of the cosmopolitan freshwater clusters contain photoheterotrophs. These comprised at least 10-23% of bacterioplankton, and RBs were the dominant fraction. Our data demonstrate that Actinobacteria, including clusters acI, Luna and acSTL, are the predominant freshwater RBs. We significantly broaden the known taxonomic range of freshwater RBs, to include Alpha-, Beta-, Gamma- and Deltaproteobacteria, Verrucomicrobia and Sphingobacteria. By sequencing single cells, we found evidence for inter-phyla horizontal gene transfer and recombination of rhodopsin genes and identified specific taxonomic groups involved in these evolutionary processes. Our data suggest that members of the ubiquitous betaproteobacteria Polynucleobacter spp. are the dominant AAPs in temperate freshwater lakes. Furthermore, the RuBisCO (ribulose 1,5-bisphosphate carboxylase/oxygenase) gene was found in several single cells of Betaproteobacteria, Bacteroidetes and Gammaproteobacteria, suggesting that chemoautotrophs may be more prevalent among aerobic bacterioplankton than previously thought. This study demonstrates the power of single-cell DNA sequencing addressing previously unresolved questions about the metabolic potential and evolutionary histories of uncultured microorganisms, which dominate most natural environments.


White matter ischemic changes in hyperacute ischemic stroke: voxel-based analysis using diffusion tensor imaging and MR perfusion.
The purpose of this study was to evaluate changes in fractional anisotropy (FA), as measured by diffusion tensor imaging, of white matter (WM) infarction and hypoperfusion in patients with acute ischemic stroke using a quantitative voxel-based analysis. In this prospective study, diffusion tensor imaging and dynamic susceptibility contrast perfusion sequences were acquired in 21 patients with acute ischemic stroke who presented within 6 hours of symptom onset. The coregistered FA, apparent diffusion coefficient, and dynamic susceptibility contrast time to maximum (Tmax) maps were used for voxel-based quantification using a region of interest approach in the ipsilateral affected side and in the homologous contralateral WM. The regions of WM infarction versus hypoperfusion were segmented using a threshold method. Data were analyzed by regression and ANOVA. There was an overall significant mean difference (P<0.001) for the apparent diffusion coefficient, Tmax, and FA values between the normal, hypoperfused, and infarcted WM. The mean±SD of FA was significantly higher (P<0.001) in hypoperfused WM (0.397±0.019) and lower (P<0.001) in infarcted WM (0.313±0.037) when compared with normal WM (0.360±0.020). Regression tree analysis of hypoperfused WM showed the largest mean FA difference at Tmax above versus below 5.4 s with a mean difference of 0.033 (P=0.0096). Diffusion tensor imaging-FA was decreased in regions of WM infarction and increased in hypoperfused WM in patients with hyperacute acute ischemic stroke. The significantly increased FA values in the hypoperfused WM with Tmax≥5.4 s are suggestive of early ischemic microstructural changes.


Interactions of alpha-helices with lipid bilayers: a review of simulation studies.
Membrane proteins, of which the majority seem to contain one or more alpha-helix, constitute approx. 30% of most genomes. A complete understanding of the nature of helix/bilayer interactions is necessary for an understanding of the structural principles underlying membrane proteins. This review describes computer simulation studies of helix/bilayer interactions. Key experimental studies of the interactions of alpha-helices and lipid bilayers are briefly reviewed. Surface associated helices are found in some membrane-bound enzymes (e.g. prostaglandin synthase), and as stages in the mechanisms of antimicrobial peptides and of pore-forming bacterial toxins. Transmembrane alpha-helices are found in most integral membrane proteins, and also in channels formed by amphipathic peptides or by bacterial toxins. Mean field simulations, in which the lipid bilayer is approximated as a hydrophobic continuum, have been used in studies of membrane-active peptides (e.g. alamethicin, melittin, magainin and dermaseptin) and of simple membrane proteins (e.g. phage Pf1 coat protein). All atom molecular dynamics simulations of fully solvated bilayers with transmembrane helices have been applied to: the constituent helices of bacteriorhodopsin; peptide-16 (a simple model TM helix); and a number of pore-lining helices from ion channels. Surface associated helices (e.g. melittin and dermaseptin) have been simulated, as have alpha-helical bundles such as bacteriorhodopsin and alamethicin. From comparison of the results from the two classes of simulation, it emerges that a major theoretical challenge is to exploit the results of all atom simulations in order to improve the mean field approach.


Activators of PPARs and LXR decrease the adverse effects of exogenous glucocorticoids on the epidermis.
While glucocorticoids (GC) exert beneficial effects (anti-inflammatory), they also have adverse effects on the epidermis including decreased epidermal differentiation, decreased keratinocyte proliferation, and decreased cutaneous permeability barrier homeostasis. Thus, the purpose of this study was to develop strategies to prevent these GC toxicities using simultaneous topical treatments in clobetasol-treated mice. While a triple-lipid mixture of stratum corneum lipids (ceramide, free fatty acid and cholesterol) was previously shown to reverse the GC-induced abnormality in cutaneous barrier function [J Invest Dermatol, 120 (2003) 456], this lipid mixture did not prevent the GC-induced abnormalities in either keratinocyte proliferation or differentiation. As activators of PPARalpha, beta/delta, gamma and LXR, regulate keratinocyte proliferation and differentiation and improve permeability barrier homeostasis, we next assessed the effects of these activators during concurrent GC treatment. Co-application of either ciglitazone (PPARgamma activator), clofibrate (PPARalpha activator) or 22R (OH) cholesterol (LXR activator) with clobetasol prevented the decrease in involucrin, filaggrin and loricrin expression. By contrast, a PPARbeta/delta activator (GW501516) normalized only the expression of involucrin and filaggrin but not loricrin. Moreover, topical application of PPARalpha, beta/delta or LXR activators partially prevented the decrease in keratinocyte proliferation in GC-treated murine skin, as measured using PCNA, while no effect was seen after co-treatment with PPARgamma activators. Finally, PPARgamma and PPARbeta/delta activators but not PPARalpha and LXR activators improved permeability barrier homeostasis in GC-treated mice. Together, these studies demonstrate that PPAR and LXR activators can prevent several of the adverse effects of topical GC on the epidermis.


A1899, PK-THPP, ML365, and Doxapram inhibit endogenous TASK channels and excite calcium signaling in carotid body type-1 cells.
Sensing of hypoxia and acidosis in arterial chemoreceptors is thought to be mediated through the inhibition of TASK and possibly other (e.g., BKCa ) potassium channels which leads to membrane depolarization, voltage-gated Ca-entry, and neurosecretion. Here, we investigate the effects of pharmacological inhibitors on TASK channel activity and [Ca2+ ]i -signaling in isolated neonatal rat type-1 cells. PK-THPP inhibited TASK channel activity in cell attached patches by up to 90% (at 400 nmol/L). A1899 inhibited TASK channel activity by 35% at 400 nmol/L. PK-THPP, A1899 and Ml 365 all evoked a rapid increase in type-1 cell [Ca2+ ]i . These [Ca2+ ]i responses were abolished in Ca2+ -free solution and greatly attenuated by Ni2+ (2 mM) suggesting that depolarization and voltage-gated Ca2+ -entry mediated the rise in [Ca2+ ]i. Doxapram (50 μmol/L), a respiratory stimulant, also inhibited type-1 cell TASK channel activity and increased [Ca2+ ]i. . We also tested the effects of combined inhibition of BKCa and TASK channels. TEA (5 mmol/L) slightly increased [Ca2+ ]i in the presence of PK-THPP and A1899. Paxilline (300 nM) and iberiotoxin (50 nmol/L) also slightly increased [Ca2+ ]i in the presence of A1899 but not in the presence of PK-THPP. In general [Ca2+ ]i responses to TASK inhibitors, alone or in combination with BKCa inhibitors, were smaller than the [Ca2+ ]i responses evoked by hypoxia. These data confirm that TASK channel inhibition is capable of evoking membrane depolarization and robust voltage-gated Ca2+ -entry but suggest that this, even with concomitant inhibition of BKCa channels, may be insufficient to account fully for the [Ca2+ ]i -response to hypoxia.


Tolerogenic function of dimeric forms of HLA-G recombinant proteins: a comparative study in vivo.
HLA-G is a natural tolerogenic molecule involved in the best example of tolerance to foreign tissues there is: the maternal-fetal tolerance. The further involvement of HLA-G in the tolerance of allogeneic transplants has also been demonstrated and some of its mechanisms of action have been elucidated. For these reasons, therapeutic HLA-G molecules for tolerance induction in transplantation are actively investigated. In the present study, we studied the tolerogenic functions of three different HLA-G recombinant proteins: HLA-G heavy chain fused to β2-microglobulin (B2M), HLA-G heavy chain fused to B2M and to the Fc portion of an immunoglobulin, and HLA-G alpha-1 domain either fused to the Fc part of an immunoglobulin or as a synthetic peptide. Our results demonstrate the tolerogenic function of B2M-HLA-G fusion proteins, and especially of B2M-HLA-G5, which were capable of significantly delaying allogeneic skin graft rejection in a murine in vivo transplantation model. The results from our studies suggest that HLA-G recombinant proteins are relevant candidates for tolerance induction in human transplantation.


Maternal occupational exposure to polycyclic aromatic hydrocarbons and the risk of isolated congenital heart defects among offspring.
Although there is evidence in experimental model systems that exposure to polycyclic aromatic hydrocarbons (PAHs) is linked with congenital heart defects (CHDs), few studies have examined the association in humans. We conducted a case-control study to examine the association between maternal exposure to PAHs and CHDs in offspring using data from the National Birth Defects Prevention Study (NBDPS) (1997-2011). We obtained detailed information on maternal occupation during the month before to three months after conception. Expert raters, masked to case-control status, assessed job descriptions to assign categorical levels of exposure. Categories were quantitatively mapped to estimate cumulative exposure to PAHs, incorporating exposure intensity, frequency, work duration, and work hours. Quartiles were generated for cumulative maternal exposure to PAHs. Crude and adjusted odds ratios (ORs) with 95% confidence intervals (CIs) were estimated using unconditional logistic regression for quartiles of PAH exposure and six CHD groupings (e.g. conotruncal) and specific subtypes (e.g. tetralogy of Fallot [ToF]). Final models were adjusted for maternal age, race/ethnicity, education, smoking, anticonvulsant use, folic acid supplementation, and study center. There were 4,775 case and 7,734 control infants eligible for the study. The prevalence of occupational exposure to PAHs was 10.2% among both case and control mothers. In adjusted analysis, compared to mothers with no occupational PAH exposure, those in the highest quartile of exposure were more likely to have offspring in the conotruncal heart defects group (OR 1.41; 95% CI 1.00-2.00), and with ToF (OR 1.83; 95% CI 1.21-2.78). Women in the highest quartile of estimated cumulative occupational PAH exposure during early pregnancy were more likely to have offspring with conotruncal heart defects, specifically ToF, compared to women with no occupational PAH exposure. Other comparisons between PAHs and other CHDs subgroups did not show any statistically precise associations.


Single-shot diffraction data from the Mimivirus particle using an X-ray free-electron laser.
Free-electron lasers (FEL) hold the potential to revolutionize structural biology by producing X-ray pules short enough to outrun radiation damage, thus allowing imaging of biological samples without the limitation from radiation damage. Thus, a major part of the scientific case for the first FELs was three-dimensional (3D) reconstruction of non-crystalline biological objects. In a recent publication we demonstrated the first 3D reconstruction of a biological object from an X-ray FEL using this technique. The sample was the giant Mimivirus, which is one of the largest known viruses with a diameter of 450 nm. Here we present the dataset used for this successful reconstruction. Data-analysis methods for single-particle imaging at FELs are undergoing heavy development but data collection relies on very limited time available through a highly competitive proposal process. This dataset provides experimental data to the entire community and could boost algorithm development and provide a benchmark dataset for new algorithms.


The neurology of empty categories aphasics' failure to detect ungrammaticality.
A direct investigation into the grammatical abilities of Broca's and Wernicke's aphasics sought to obtain critical evidence for a revised model of the functional neuroanatomy of language. We examined aphasics' ability to make grammaticality judgments on a set of theoretically selected, highly complex syntactic structures that involve, most prominently, fine violations of constraints on syntactic movement. Although both groups have been thought to possess intact abilities in this domain, we discovered severe deficits; Broca's and Wernicke's aphasics (whose performances differed) exhibited clear, delineated, and grammatically characterizable deficits - they follow from the Trace-Deletion Hypothesis, which is motivated by independent comprehension results. These conclusions have both linguistic and neurological implications; Linguistically, they show that the aphasic deficit interacts with more than one module of the grammar. Namely, it manifests not only when the thematic module is called for in interpretive tasks but also when constraints on syntactic movement are tapped in a study of judgment. Neurologically, the results support a view of receptive grammatical mechanisms in the left cortex, which is functionally more restrictive than currently assumed; neuroanatomically, however, it is more distributed.