Purification of Adult Hymenolepis diminuta (Cestoda) Mitochondrial NADPH→NAD+ Transhydrogenase.
The mitochondrial, inner-membrane-associated, reversible NADPH→NAD+ transhydrogenase of the energetically anaerobic adult cestode Hymenolepis diminuta connects NADPH generation, via a mitochondrial NADP+-specific "malic" enzyme, with NADH formation needed for electron transport. In reducing the pyridine nucleotide, the enzyme concomitantly catalyzes transmembrane proton translocation, thereby coupling NADH formation to ATP generation or NADPH formation to ATP hydrolysis. Detergent-solubilized transhydrogenase, from isolated mitochondrial membranes, was purified to apparent homogeneity using ion exchange and hydroxylapatite chromatographies. The enzyme displayed a monomeric Mr of ∼110 kDa and required phospholipid, without which activity was rapidly lost. Of the phospholipids examined, phosphatidylcholine was the most effective. Transhydrogenase-catalyzed NADH formation was inhibited by NAD(P)+ and adenylates, suggesting regulatory effects of the pyridine nucleotides and effects of pyridine nucleotide-simulating molecules. In keeping with its proton-translocating function, the enzyme was inhibited by dicyclohexylcarbodiimide. The isolated enzyme catalyzed neither NADH→NADP+ nor NADH→NAD+ transhydrogenations, thereby suggesting a need for a minimal coupling to electron transport for the NADH→NADP+ reaction as well as enzyme specificity. Anti-transhydrogenase monospecific antibodies proved inhibitory to NADPH→NAD+ transhydrogenation catalyzed by both isolated and membrane-associated enzymes. This purification study apparently represents a first for parasitic helminths or multicellular invertebrates generally and establishes a framework for evaluating the transhydrogenase as a potential site for specific chemotherapeutic attack.


Patient safety, satisfaction, and quality of hospital care: cross sectional surveys of nurses and patients in 12 countries in Europe and the United States.
To determine whether hospitals with a good organisation of care (such as improved nurse staffing and work environments) can affect patient care and nurse workforce stability in European countries. Cross sectional surveys of patients and nurses. Nurses were surveyed in general acute care hospitals (488 in 12 European countries; 617 in the United States); patients were surveyed in 210 European hospitals and 430 US hospitals. 33 659 nurses and 11 318 patients in Europe; 27 509 nurses and more than 120 000 patients in the US. Nurse outcomes (hospital staffing, work environments, burnout, dissatisfaction, intention to leave job in the next year, patient safety, quality of care), patient outcomes (satisfaction overall and with nursing care, willingness to recommend hospitals). The percentage of nurses reporting poor or fair quality of patient care varied substantially by country (from 11% (Ireland) to 47% (Greece)), as did rates for nurses who gave their hospital a poor or failing safety grade (4% (Switzerland) to 18% (Poland)). We found high rates of nurse burnout (10% (Netherlands) to 78% (Greece)), job dissatisfaction (11% (Netherlands) to 56% (Greece)), and intention to leave (14% (US) to 49% (Finland, Greece)). Patients' high ratings of their hospitals also varied considerably (35% (Spain) to 61% (Finland, Ireland)), as did rates of patients willing to recommend their hospital (53% (Greece) to 78% (Switzerland)). Improved work environments and reduced ratios of patients to nurses were associated with increased care quality and patient satisfaction. In European hospitals, after adjusting for hospital and nurse characteristics, nurses with better work environments were half as likely to report poor or fair care quality (adjusted odds ratio 0.56, 95% confidence interval 0.51 to 0.61) and give their hospitals poor or failing grades on patient safety (0.50, 0.44 to 0.56). Each additional patient per nurse increased the odds of nurses reporting poor or fair quality care (1.11, 1.07 to 1.15) and poor or failing safety grades (1.10, 1.05 to 1.16). Patients in hospitals with better work environments were more likely to rate their hospital highly (1.16, 1.03 to 1.32) and recommend their hospitals (1.20, 1.05 to 1.37), whereas those with higher ratios of patients to nurses were less likely to rate them highly (0.94, 0.91 to 0.97) or recommend them (0.95, 0.91 to 0.98). Results were similar in the US. Nurses and patients agreed on which hospitals provided good care and could be recommended. Deficits in hospital care quality were common in all countries. Improvement of hospital work environments might be a relatively low cost strategy to improve safety and quality in hospital care and to increase patient satisfaction.


Genomic approaches to studying CFTR transcriptional regulation.
The CFTR gene was identified over 20 years ago, and yet how the gene is transcriptionally regulated is not fully understood. Completion of the human genome sequence has encouraged a new generation of genomic techniques that can be used to identify and characterize the regulatory elements of the genome, which are often hidden in non-coding regions. In this chapter we describe two techniques that we have used to identify regulatory regions of the CFTR locus: DNase-chip, which utilizes DNase I-digested chromatin hybridized to tiled microarrays in order to locate regions of the CFTR locus that are "open" and thus likely regions of transcription factor binding; and quantitative chromosome conformation capture (q3C), which uses quantitative PCR analysis of digested and ligated, crosslinked chromosomes to measure physical interactions between distal genomic regions. When used together, these methods provide a powerful avenue to discover transcriptional regulatory elements within large genomic regions.


Cumulative fluid intake minus output is not associated with ventilator weaning duration or extubation outcomes in children.
The effect of fluid balance on respiratory outcomes for critically ill children has not been evaluated. The only indicator of fluid balance routinely recorded across our intensive care units was estimated fluid intake and output. We sought to determine whether cumulative intake minus output (I-O) at the start of weaning predicted weaning duration and whether cumulative I-O at extubation predicted extubation failure. Prospective observational study. Ten pediatric intensive care units. Cumulative I-O was recorded daily for 301 mechanically ventilated children (<18 yrs of age) from November 1999 through April 2001. Cumulative I-O was recorded during a study of weaning strategies and extubation failure in which mechanical ventilation of the majority of patients during weaning and extubation was managed according to a protocol that did not include fluid balance indicators. Outcomes were the time to successful removal of ventilatory support and the rate of initial extubation failure. Relationships between cumulative I-O and outcomes were assessed by means of proportional hazards and logistic regression. The mean cumulative I-O per kilogram of ideal body weight at the start of weaning was 101 mL (sd, 180). Cumulative I-O at the time weaning was initiated did not predict duration of mechanical ventilator weaning. The mean cumulative I-O per kilogram of ideal body weight at extubation was 136 mL (sd, 237). Cumulative I-O at extubation did not predict extubation outcome. There was an association between cumulative I-O at extubation and the duration of weaning in cases not managed by a protocol. Although routinely recorded, cumulative fluid I-O does not appear to have clinical utility in cases managed according to a mechanical ventilator protocol in which tidal volume and oxygenation on minimal levels of ventilator support are systematically tested.


Crystal structure of Mj1640/DUF358 protein reveals a putative SPOUT-class RNA methyltransferase.
The proteins in DUF358 family are all bacterial proteins, which are ∼200 amino acids long with unknown function. Bioinformatics analysis suggests that these proteins contain several conserved arginines and aspartates that might adopt SPOUT-class fold. Here we report crystal structure of Methanocaldococcus jannaschii DUF358/Mj1640 in complex with S-adenosyl-L-methionine (SAM) at 1.4 Å resolution. The structure reveals a single domain structure consisting of eight-stranded β-sheets sandwiched by six α-helices at both sides. Similar to other SPOUT-class members, Mj1640 contains a typical deep trefoil knot at its C-terminus to accommodate the SAM cofactor. However, Mj1640 has limited structural extension at its N-terminus, which is unique to this family member. Mj1640 forms a dimer, which is mediated by two parallel pairs of α-helices oriented almost perpendicular to each other. Although Mj1640 shares close structural similarity with Nep1, the significant differences in N-terminal extension domain and the overall surface charge distribution strongly suggest that Mj1640 might target a different RNA sequence. Detailed structural analysis of the SAM-binding pocket reveals that Asp157 or Glu183 from its own monomer or Ser43 from the associate monomer probably plays the catalytic role for RNA methylation.


Pseudomonas aeruginosa lectin LecB impairs keratinocyte fitness by abrogating growth factor signalling.
Lectins are glycan-binding proteins with no catalytic activity and ubiquitously expressed in nature. Numerous bacteria use lectins to efficiently bind to epithelia, thus facilitating tissue colonisation. Wounded skin is one of the preferred niches for Pseudomonas aeruginosa, which has developed diverse strategies to impair tissue repair processes and promote infection. Here, we analyse the effect of the P. aeruginosa fucose-binding lectin LecB on human keratinocytes and demonstrate that it triggers events in the host, upon binding to fucosylated residues on cell membrane receptors, which extend beyond its role as an adhesion molecule. We found that LecB associates with insulin-like growth factor-1 receptor and dampens its signalling, leading to the arrest of cell cycle. In addition, we describe a novel LecB-triggered mechanism to down-regulate host cell receptors by showing that LecB leads to insulin-like growth factor-1 receptor internalisation and subsequent missorting towards intracellular endosomal compartments, without receptor activation. Overall, these data highlight that LecB is a multitask virulence factor that, through subversion of several host pathways, has a profound impact on keratinocyte proliferation and survival.


Probing the brain in autism using FMRI and diffusion tensor imaging.
Newly emerging theories suggest that the brain does not function as a cohesive unit in autism, and this discordance is reflected in the behavioral symptoms displayed by individuals with autism. While structural neuroimaging findings have provided some insights into brain abnormalities in autism, the consistency of such findings is questionable. Functional neuroimaging, on the other hand, has been more fruitful in this regard because autism is a disorder of dynamic processing and allows examination of communication between cortical networks, which appears to be where the underlying problem occurs in autism. Functional connectivity is defined as the temporal correlation of spatially separate neurological events. Findings from a number of recent fMRI studies have supported the idea that there is weaker coordination between different parts of the brain that should be working together to accomplish complex social or language problems. One of the mysteries of autism is the coexistence of deficits in several domains along with relatively intact, sometimes enhanced, abilities. Such complex manifestation of autism calls for a global and comprehensive examination of the disorder at the neural level. A compelling recent account of the brain functioning in autism, the cortical underconnectivity theory, provides an integrating framework for the neurobiological bases of autism. The cortical underconnectivity theory of autism suggests that any language, social, or psychological function that is dependent on the integration of multiple brain regions is susceptible to disruption as the processing demand increases. In autism, the underfunctioning of integrative circuitry in the brain may cause widespread underconnectivity. In other words, people with autism may interpret information in a piecemeal fashion at the expense of the whole. Since cortical underconnectivity among brain regions, especially the frontal cortex and more posterior areas, has now been relatively well established, we can begin to further understand brain connectivity as a critical component of autism symptomatology. A logical next step in this direction is to examine the anatomical connections that may mediate the functional connections mentioned above. Diffusion Tensor Imaging (DTI) is a relatively novel neuroimaging technique that helps probe the diffusion of water in the brain to infer the integrity of white matter fibers. In this technique, water diffusion in the brain is examined in several directions using diffusion gradients. While functional connectivity provides information about the synchronization of brain activation across different brain areas during a task or during rest, DTI helps in understanding the underlying axonal organization which may facilitate the cross-talk among brain areas. This paper will describe these techniques as valuable tools in understanding the brain in autism and the challenges involved in this line of research.


Assessing the environmental performance of English arable and livestock holdings using data from the Farm Accountancy Data Network (FADN).
Agri-environment schemes (AESs) have been implemented across EU member states in an attempt to reconcile agricultural production methods with protection of the environment and maintenance of the countryside. To determine the extent to which such policy objectives are being fulfilled, participating countries are obliged to monitor and evaluate the environmental, agricultural and socio-economic impacts of their AESs. However, few evaluations measure precise environmental outcomes and critically, there are no agreed methodologies to evaluate the benefits of particular agri-environmental measures, or to track the environmental consequences of changing agricultural practices. In response to these issues, the Agri-Environmental Footprint project developed a common methodology for assessing the environmental impact of European AES. The Agri-Environmental Footprint Index (AFI) is a farm-level, adaptable methodology that aggregates measurements of agri-environmental indicators based on Multi-Criteria Analysis (MCA) techniques. The method was developed specifically to allow assessment of differences in the environmental performance of farms according to participation in agri-environment schemes. The AFI methodology is constructed so that high values represent good environmental performance. This paper explores the use of the AFI methodology in combination with Farm Business Survey data collected in England for the Farm Accountancy Data Network (FADN), to test whether its use could be extended for the routine surveillance of environmental performance of farming systems using established data sources. Overall, the aim was to measure the environmental impact of three different types of agriculture (arable, lowland livestock and upland livestock) in England and to identify differences in AFI due to participation in agri-environment schemes. However, because farm size, farmer age, level of education and region are also likely to influence the environmental performance of a holding, these factors were also considered. Application of the methodology revealed that only arable holdings participating in agri-environment schemes had a greater environmental performance, although responses differed between regions. Of the other explanatory variables explored, the key factors determining the environmental performance for lowland livestock holdings were farm size, farmer age and level of education. In contrast, the AFI value of upland livestock holdings differed only between regions. The paper demonstrates that the AFI methodology can be used readily with English FADN data and therefore has the potential to be applied more widely to similar data sources routinely collected across the EU-27 in a standardised manner.


Suppression of mitogen-induced proliferation of normal spleen cells by macrophages from chickens inoculated with Marek's disease virus.
Spleen cells from chickens 7 days after inoculation with Marek's disease virus (MDV) responded poorly to stimulation by phytohemagglutinin (PHA). Addition of these cells to syngeneic normal spleen cells caused of marked suppression of the PHA response of the normal cells. The MDV spleen cells also inhibited the DNA synthesis of MSB-1 lymphoblastoid cells in vitro. The suppressive activity is attributed to the presence in MDV spleen cells of a population of suppressor cells with characteristics typical of macrophages. The suppressor cell activity was not removable by treatment with anti-T or anti-B serum with C, but it was reversible by treatment with carrageenan or carbonyl iron/magnet, by passage through glass wool column, and by adherence to plastic Petri dishes. The adherent MDV spleen cells also showed strong suppressor cell activity against syngeneic normal spleen cells.


Rise in testing and diagnosis associated with Scotland's Action Plan on Hepatitis C and introduction of dried blood spot testing.
A key aim of the Hepatitis C Action Plan for Scotland was to reduce the undiagnosed population through awareness-raising activities, for general practitioners and those at risk, and the introduction of dried blood spot (DBS) sampling in community drug services to overcome barriers to testing. This study evaluates the impact of these activities on testing and diagnosis. Data on hepatitis C virus (HCV) testing undertaken between January 1999 and December 2011 in Scotland's four largest health boards were analysed. Segmented regression analysis was used to examine changes in testing following the (1) launch of the Action Plan and (2) introduction of DBS testing. Between the pre-Action Plan and Action Plan periods, increases were observed in the average number of HCV tests (19 058-29 045), positive tests (1993-2405) and new diagnoses (1221-1367). Since July 2009, 26% of new diagnoses were made in drug services. The trend in the number of positive tests was raised during the Action Plan, compared to pre-Action Plan, particularly in drug services (rate ratio (RR)=1.4, p<0.001) and prisons (RR=1.2, p<0.001); no change was observed in general practice. Following introduction of DBS testing, there was a 3-fold increase in testing (RR=3.5, p<0.001) and 12-fold increase in positives (RR=12.1, p<0.001) in drug services. The introduction of DBS sampling in community drug services has made an appreciable contribution to efforts to diagnose the HCV-infected population in Scotland. These findings are important to other countries, with injecting-related HCV epidemics, needing to scale-up testing/case-finding initiatives.


Intercellular communication in malignant pleural mesothelioma: properties of tunneling nanotubes.
Malignant pleural mesothelioma is a particularly aggressive and locally invasive malignancy with a poor prognosis despite advances in understanding of cancer cell biology and development of new therapies. At the cellular level, cultured mesothelioma cells present a mesenchymal appearance and a strong capacity for local cellular invasion. One important but underexplored area of mesothelioma cell biology is intercellular communication. Our group has previously characterized in multiple histological subtypes of mesothelioma a unique cellular protrusion known as tunneling nanotubes (TnTs). TnTs are long, actin filament-based, narrow cytoplasmic extensions that are non-adherent when cultured in vitro and are capable of shuttling cellular cargo between connected cells. Our prior work confirmed the presence of nanotube structures in tumors resected from patients with human mesothelioma. In our current study, we quantified the number of TnTs/cell among various mesothelioma subtypes and normal mesothelial cells using confocal microscopic techniques. We also examined changes in TnT length over time in comparison to cell proliferation. We further examined potential approaches to the in vivo study of TnTs in animal models of cancer. We have developed novel approaches to study TnTs in aggressive solid tumor malignancies and define fundamental characteristics of TnTs in malignant mesothelioma. There is mounting evidence that TnTs play an important role in intercellular communication in mesothelioma and thus merit further investigation of their role in vivo.


The mechanism and site of action of lidocaine hydrochloride in guinea pig inner ear.
Lidocaine was applied to the round window (RW) in order to localize its site of action in the cochlea. Cochlear microphonic (CM), summating potential (SP), and compound action potential (CAP) input/output functions were measured to a 16 kHz tone burst to assess the functional changes of the cochlea. In separate experiments, the effect of lidocaine on the whole cell current of isolated outer hair cells (OHC) was studied. A dose of 2 microliters of 40 mM lidocaine in saline solution, when applied to the RW, caused a small change in all measured variables, indicating a passage of the drug through the RW membrane to sites of action. However, 160 mM of lidocaine further decreased CM, SP, and CAP by a total of 40% from the control. A partial recovery occurred for CM during the 30 min follow-up period. CAP and SP continued to decline. In isolated OHCs, lidocaine decreased the whole cell current in a dose-dependent fashion. The KD for lidocaine effect on OHCs was 7 mM. Our in vivo results indicate that lidocaine affects OHCs and reduces CM, causing a subsequent reduction in SP and CAP. The increased effect of lidocaine on CAP and SP, while CM is recovering, suggests an additional specific effect of lidocaine on the cochlear nerve and/or on inner hair cells. Considering that lidocaine alters OHC current (in isolated hair cells) and that lidocaine does not affect endocochlear potential [Laurikainen et al. Acta Otolaryngol (Stockh) 1991: 112: 800-9], the observed CM changes are most likely due to an in vivo effect on OHCs. Thus, the early effect of lidocaine on the cochlea appears to be due to a significant change in organ of Corti function, rather than to direct anesthesia of the cochlear nerve. Later, an independent effect of the drug may occur on neural tissues in the inner ear.


Multigene panels in prostate cancer risk assessment.
The aim of this review is to identify, synthesize, and appraise the literature on the analytic validity, clinical validity, and clinical utility of commercially available single nucleotide polymorphism (SNP) panel tests for assessing the risk of prostate cancer. MEDLINE®, Cochrane CENTRAL, Cochrane Database of Systematic Reviews, and Embase, from the beginning of each database to October 2011. Search strategies used combinations of controlled vocabulary (medical subject headings, keywords) and text words. Grey literature was identified. Three Key Questions (KQs) encompassing broad aspects of the analytic validity, clinical validity, and clinical utility of SNP-based panels were developed with the input of a Technical Expert Panel assembled by the Evidence-based Practice Center and approved by the Agency for Healthcare Research and Quality. Standard systematic review methodology was applied, with eligibility criteria developed separately for each KQ. From 1,998 unique citations, 14 were retained for data abstraction and quality assessment following title and abstract screening and full text screening. All focused on clinical validity (KQ2), and evaluated 15 individual panels with two to 35 SNPs. All had poor discriminative ability for predicting risk of prostate cancer and/or distinguishing between aggressive and asymptomatic/latent disease. The risk of bias of the studies was determined to be moderate. None of the panels had been evaluated in routine clinical settings. The evidence on currently available SNP panels does not permit meaningful assessment of analytic validity. The limited evidence on clinical validity is insufficient to conclude that the panels assessed would perform adequately as screening or risk stratification tests. No evidence is available on the clinical utility of current panels.


Handgrip force steadiness in young and older adults: a reproducibility study.
Force steadiness is a quantitative measure of the ability to control muscle tonus. It is an independent predictor of functional performance and has shown to correlate well with different degrees of motor impairment following stroke. Despite being clinically relevant, few studies have assessed the validity of measuring force steadiness. The aim of this study was to explore the reproducibility of handgrip force steadiness, and to assess age difference in steadiness. Intrarater reproducibility (the degree to which a rating gives consistent result on separate occasions) was investigated in a test-retest design with seven days between sessions. Ten young and thirty older adults were recruited and handgrip steadiness was tested at 5%, 10% and 25% of maximum voluntary contraction (MVC) using Nintendo Wii Balance Board (WBB). Coefficients of variation were calculated from the mean force produced (CVM) and the target force (CVT). Area between the force curve and the target force line (Area) was also calculated. For the older adults we explored reliability using intraclass correlation coefficient (ICC) and agreement using standard error of measurement (SEM), limits of agreement (LOA) and smallest real difference (SRD). A systematic improvement in handgrip steadiness was found between sessions for all measures (CVM, CVT, Area). CVM and CVT at 5% of MVC showed good to high reliability, while Area had poor reliability for all percentages of MVC. Averaged ICC for CVM, CVT and Area was 0.815, 0.806 and 0.464, respectively. Averaged ICC on 5%, 10%, and 25% of MVC was 0.751, 0.667 and 0.668, respectively. Measures of agreement showed similar trends with better results for CVM and CVT than for Area. Young adults had better handgrip steadiness than older adults across all measures. The CVM and CVT measures demonstrated good reproducibility at lower percentages of MVC using the WBB, and could become relevant measures in the clinical setting. The Area measure had poor reproducibility. Young adults have better handgrip steadiness than old adults.


Genomic analysis of Klebsiella pneumoniae isolates from Malawi reveals acquisition of multiple ESBL determinants across diverse lineages.
ESBL-producing Klebsiella pneumoniae (KPN) pose a major threat to human health globally. We carried out a WGS study to understand the genetic background of ESBL-producing KPN in Malawi and place them in the context of other global isolates. We sequenced genomes of 72 invasive and carriage KPN isolates collected from patients admitted to Queen Elizabeth Central Hospital, Blantyre, Malawi. We performed phylogenetic and population structure analyses on these and previously published genomes from Kenya (n = 66) and from outside sub-Saharan Africa (n = 67). We screened for presence of antimicrobial resistance (AMR) genetic determinants and carried out association analyses by genomic sequence cluster, AMR phenotype and time. Malawian isolates fit within the global population structure of KPN, clustering into the major lineages of KpI, KpII and KpIII. KpI isolates from Malawi were more related to those from Kenya, with both collections exhibiting more clonality than isolates from the rest of the world. We identified multiple ESBL genes, including blaCTX-M-15, several blaSHV, blaTEM-63 and blaOXA-10, and other AMR genes, across diverse lineages of the KPN isolates from Malawi. No carbapenem resistance genes were detected; however, we detected IncFII and IncFIB plasmids that were similar to the carbapenem resistance-associated plasmid pNDM-mar. There are multiple ESBL genes across diverse KPN lineages in Malawi and plasmids in circulation that are capable of carrying carbapenem resistance. Unless appropriate interventions are rapidly put in place, these may lead to a high burden of locally untreatable infection in vulnerable populations.


Validity of Physical Activity Monitors for Estimating Energy Expenditure During Wheelchair Propulsion.
It is unknown if activity monitors can detect the increased energy expenditure (EE) of wheelchair propulsion at different speeds or on different surfaces. Individuals who used manual wheelchairs (n = 14) performed 5 wheeling activities: on a level surface at 3 speeds, on a rubberized track at 1 fixed speed and on a sidewalk course at a self-selected speed. EE was measured using a portable indirect calorimetry system and estimated by an Actical (AC) worn on the wrist and a SenseWear (SW) activity monitor worn on the upper arm. Repeated- measures ANOVA was used to compare measured EE to the estimates from the standard AC prediction equation and SW using 2 different equations. Repeated-measures ANOVA demonstrated a significant main effect between measured EE and estimated EE. There were no differences between the criterion method and the AC across the 5 activities. The SW overestimated EE when wheeling at 3 speeds on a level surface, and during sidewalk wheeling. The wheelchair-specific SW equation improved the EE prediction during low intensity activities, but error progressively increased during higher intensity activities. During manual wheelchair propulsion, the wrist-mounted AC provided valid estimates of EE, whereas the SW tended to overestimate EE.


GABAA receptor autoimmunity: A multicenter experience.
We sought to validate methods for detection and confirmation of GABAA receptor (R)-IgG and clinically characterize seropositive cases. Archived serum and CSF specimens (185 total) suspected to harbor GABAAR-IgG were evaluated by indirect immunofluorescence assay (IFA). Twenty-six specimens from 19 patients appeared suspicious for GABAAR-IgG positivity by IFA, based on prior reports and comparison with commercial GABAAR antibody staining. Aliquots of those specimens were tested at the University of Oxford, United Kingdom, and Euroimmun, Lubeck, Germany, for GABAAR-IgG by cell-based assays (CBAs) using HEK293-indicator cells transfected with plasmids encoding different GABAAR subunits. Eight specimens (of 26 tested; 4 serums, 4 CSFs) from 5 patients were confirmed by CBA to be GABAAR-IgG positive. Patient IgGs were always reactive with α1β3 GABAAR subunits. One more patient was identified clinically after this validation study. Median age for the 6 patients at serologic diagnosis was 44 years (range, 1-71 years), and 4 of them were male. Among the 4 for whom clinical information was available (2 treated by the authors), all had encephalitis and antiepileptic drug refractory seizures. Three out of 4 patients treated with a combination of immunotherapies had good outcomes. The fourth, recognized to have an autoimmune cause late in the clinical course, had severe permanent neurologic sequelae and brain atrophy. Though not as common as NMDA-R encephalitis, GABAAR encephalitis generally has a characteristic clinical-radiologic presentation and is treatable, making accurate laboratory diagnosis critical.


Clinical and Genome-Wide Analysis of Serum Platinum Levels after Cisplatin-Based Chemotherapy.
Serum platinum is measurable for years after completion of cisplatin-based chemotherapy (CBC). We report the largest investigation of serum platinum levels to date of 1,010 testicular cancer survivors (TCS) assessed 1-35 years after CBC and evaluate genetic contributions to these levels. Eligible TCS given 300 or 400 (±15) mg/m2 cisplatin underwent extensive audiometric testing, clinical examination, completed questionnaires, and had crude serum platinum levels measured. Associations between serum platinum and various risk factors and toxicities were assessed after fitting a biexponential model adjusted for follow-up time and cumulative cisplatin dose. A genome-wide association study (GWAS) was performed using the serum platinum residuals of the dose and time-adjusted model. Serum platinum levels exceeded the reference range for approximately 31 years, with a strong inverse relationship with creatinine clearance at follow-up (age-adjusted P = 2.13 × 10-3). We observed a significant, positive association between residual platinum values and luteinizing hormone (age-adjusted P = 6.58 × 10-3). Patients with high residual platinum levels experienced greater Raynaud phenomenon than those with medium or low levels (age-adjusted ORhigh/low = 1.46; P = 0.04), as well as a higher likelihood of developing tinnitus (age-adjusted ORhigh/low = 1.68, P = 0.07). GWAS identified one single-nucleotide polymorphism (SNP) meeting genome-wide significance, rs1377817 (P = 4.6 × 10-8, a SNP intronic to MYH14). This study indicates that residual platinum values are correlated with several cisplatin-related toxicities. One genetic variant is associated with these levels.


A multicenter phase II trial (SAKK 36/06) of single-agent everolimus (RAD001) in patients with relapsed or refractory mantle cell lymphoma.
Mantle cell lymphoma accounts for 6% of all B-cell lymphomas and is generally incurable. It is characterized by the translocation t(11;14) leading to cyclin D1 over-expression. Cyclin D1 is downstream of the mammalian target of rapamycin threonine kinase and can be effectively blocked by mammalian target of rapamycin inhibitors. We set out to examine the single agent activity of the orally available mammalian target of rapamycin inhibitor everolimus in a prospective, multicenter trial in patients with relapsed or refractory mantle cell lymphoma (NCT00516412). Eligible patients who had received a maximum of three prior lines of chemotherapy were given everolimus 10 mg for 28 days (one cycle) for a total of six cycles or until disease progression. The primary endpoint was the best objective response. Adverse reactions, progression-free survival and molecular response were secondary endpoints. Thirty-six patients (35 evaluable) were enrolled and treatment was generally well tolerated with Common Terminology Criteria grade ≥ 3 adverse events (>5%) including anemia (11%), thrombocytopenia (11%) and neutropenia (8%). The overall response rate was 20% (95% CI: 8-37%) with two complete remissions and five partial responses; 49% of the patients had stable disease. At a median follow-up of 6 months, the median progression-free survival was 5.5 months (95% CI: 2.8-8.2) overall and 17.0 (6.4-23.3) months for 18 patients who received six or more cycles of treatment. Three patients achieved a lasting complete molecular response, as assessed by polymerase chain reaction analysis of peripheral blood. Everolimus as a single agent is well tolerated and has anti-lymphoma activity in relapsed or refractory mantle cell lymphoma. Further studies of everolimus in combination with chemotherapy or as a single agent for maintenance treatment are warranted.


Tobacco psychopharmacology and public health policy: it takes a community.
This commentary is based upon the author's lecture given as the 2010 recipient of the award named in honor of Drs. Joseph V. Brady and Charles R. Schuster, given by the Psychopharmacology and Substance Abuse Division (Division 28) of the American Psychological Association (APA). The focus is on the contributions of many behavioral pharmacology researchers who collaborated very much in the spirit of an interactive community dedicated to the common cause of advancing science in service of public health. Division 28 and its members hold a prominent place in this account because, throughout the 1980s and 1990s, the Division was the lead scientific forum for bringing together researchers addressing the behavioral pharmacology of tobacco and nicotine. The commentary provides an overview of how advances utilizing animal and human models of dependence and withdrawal came to inform public health policy and more recently, tobacco product regulation. The commentary also recounts how efforts by the tobacco industry collided with those of nonindustry researchers, including Division 28 members, and how this was taken up in congressional hearings that addressed behavioral pharmacology research on tobacco. The review concludes with an overview of current challenges to behavioral pharmacology researchers to assist in guiding the regulation of tobacco products by the United States Food and Drug Administration and other national regulatory authorities, as well as guiding the implementation of the international tobacco treaty-the World Health Organization Framework Convention on Tobacco Control.


Specificity and timescales of cortical adaptation as inferences about natural movie statistics.
Adaptation is a phenomenological umbrella term under which a variety of temporal contextual effects are grouped. Previous models have shown that some aspects of visual adaptation reflect optimal processing of dynamic visual inputs, suggesting that adaptation should be tuned to the properties of natural visual inputs. However, the link between natural dynamic inputs and adaptation is poorly understood. Here, we extend a previously developed Bayesian modeling framework for spatial contextual effects to the temporal domain. The model learns temporal statistical regularities of natural movies and links these statistics to adaptation in primary visual cortex via divisive normalization, a ubiquitous neural computation. In particular, the model divisively normalizes the present visual input by the past visual inputs only to the degree that these are inferred to be statistically dependent. We show that this flexible form of normalization reproduces classical findings on how brief adaptation affects neuronal selectivity. Furthermore, prior knowledge acquired by the Bayesian model from natural movies can be modified by prolonged exposure to novel visual stimuli. We show that this updating can explain classical results on contrast adaptation. We also simulate the recent finding that adaptation maintains population homeostasis, namely, a balanced level of activity across a population of neurons with different orientation preferences. Consistent with previous disparate observations, our work further clarifies the influence of stimulus-specific and neuronal-specific normalization signals in adaptation.


[Cataract surgery: or why are there some patients excluded].
Cataracts are the main cause of blindness in the world, although they can be treated with relatively simple and inexpensive surgery. This study was carried out in 1997 and 1998 in five cities in the state of São Paulo, Brazil, to identify the reasons for persons not having cataract surgery. The population studied were patients seen at a community project for the rehabilitation of cataract-caused blindness, Projeto Zona Livre de Catarata (the Cataract-free-Zone Project). A questionnaire was used to interview 776 individuals with cataracts who sought assistance at the project and who had a visual acuity of 20/100 or less in the better eye. Six hundred and eighty-three patients had previously sought ophthalmic care, most frequently (27%) at public health services. The main reasons for subjects not having had cataract surgery were financial (69% of respondents) and the feeling of "still having good eyesight" (69% of respondents). Among patients who said they were afraid of surgery, the main reason was concern about being left blind. All the subjects whom the project deemed suitable for surgery agreed to undergo the procedure. Apparently, there is a gap between searching for ophthalmic services and the surgical resolution of cataracts. The predominant reasons for not having surgery were financial and logistical. There is a need to facilitate access to cataract surgery by decentralizing social services and by developing community projects to prevent cataract-caused blindness.


Charcot-Marie-Tooth disease: extensive cranial nerve involvement on CT and MR imaging.
We report a case of genetically verified Charcot-Marie-Tooth disease in which the patient had cranial nerve symptoms. CT and MR imaging demonstrated enlargement of several cranial nerves, as well as their skull-base foramina, with faint contrast material enhancement identified.


Association of p120, a tyrosine kinase substrate, with E-cadherin/catenin complexes.
p120 was originally identified as a substrate of pp60src and several receptor tyrosine kinases, but its function is not known. Recent studies revealed that this protein shows homology to a group of proteins, beta-catenin/Armadillo and plakoglobin (gamma-catenin), which are associated with the cell adhesion molecules cadherins. In this study, we examined whether p120 is associated with E-cadherin using the human carcinoma cell line HT29, as well as other cell lines, which express both of these proteins. When proteins that copurified with E-cadherin were analyzed, not only alpha-catenin, beta-catenin, and plakoglobin but also p120 were detected. Conversely, immunoprecipitates of p120 contained E-cadherin and all the catenins, although a large subpopulation of p120 was not associated with E-cadherin. Analysis of these immunoprecipitates suggests that 20% or less of the extractable E-cadherin is associated with p120. When p120 immunoprecipitation was performed with cell lysates depleted of E-cadherin, beta-catenin was no longer coprecipitated, and the amount of plakoglobin copurified was greatly reduced. This finding suggests that there are various forms of p120 complexes, including p120/E-cadherin/beta-catenin and p120/E-cadherin/plakoglobin complexes; this association profile contrasts with the mutually exclusive association of beta-catenin and plakoglobin with cadherins. When the COOH-terminal catenin binding site was truncated from E-cadherin, not only beta-catenin but also p120 did not coprecipitate with this mutated E-cadherin. Immunocytological studies showed that p120 colocalized with E-cadherin at cell-cell contact sites, even after non-ionic detergent extraction. Treatment of cells with hepatocyte growth factor/scatter factor altered the level of tyrosine phosphorylation of p120 as well as of beta-catenin and plakoglobin. These results suggest that p120 associates with E-cadherin at its COOH-terminal region, but the mechanism for this association differs from that for the association of beta-catenin and plakoglobin with E-cadherin, and thus, that p120, whose function could be modulated by growth factors, may play a unique role in regulation of the cadherin-catenin adhesion system.


Identification of quantitative trait loci associated with acylsugar accumulation using intraspecific populations of the wild tomato, Lycopersicon pennellii.
Lycopersicon pennellii LA716, a wild relative of tomato, is resistant to a number of insect pests due to the accumulation of acylsugars exuded from type IV trichomes. These acylsugars are a class of compounds including both acylglucoses and acylsucroses. Intraspecific populations between L. pennellii LA716 and L. pennellii LA1912, the latter an accession that assorts for low-level acylsugar accumulation, were created to study the inheritance of type IV trichome density, acylsugar accumulation levels, percentage of acylsugars that are acylglucoses, and leaf area. The F2 population was subsequently used to determine genomic regions associated with these traits. The relative proportion of acylglucoses and acylsucroses was found to be largely controlled by a single locus near TG549 on chromosome 3. One locus on chromosome 10 showed significant associations with acylsugar levels. In addition, 1 locus on chromosome 4 showed significant associations with leaf area. Ten additional loci showed modest associations with one or more of the traits examined, 5 of which have been previously reported.


Noninvasive quantification of T2 and concentrations of ascorbate and glutathione in the human brain from the same double-edited spectra.
The transverse relaxation times (T(2)) and concentrations of Ascorbate (Asc) and glutathione (GSH) were measured from a single dataset of double-edited spectra that were acquired at several TEs at 4 T in the human brain. Six TEs between 102 and 152 ms were utilized to calculate T(2) for the group of 12 subjects scanned five times each. Spectra measured at all six TEs were summed to quantify the concentration in each individual scan. LCModel fitting was optimized for the quantification of the Asc and GSH double-edited spectra. When the fitted baseline was constrained to be flat, T(2) was found to be 67 ms (95% confidence interval, 50-83 ms) for GSH and ≤115 ms for Asc using the sum of spectra measured over 60 scans. The Asc and GSH concentrations quantified in each of the 60 scans were 0.62 ± 0.08 and 0.81 ± 0.11 µmol/g [mean ± standard deviation (SD), n = 60], respectively, using 10 µmol/g N-acetylaspartate as an internal reference and assuming a constant influence of N-acetylaspartate and antioxidant T(2) relaxation in the reference solution and in vivo. The T(2) value of GSH was measured for the first time in the human brain. The data are consistent with short T(2) for both antioxidants. These T(2) values are essential for the absolute quantification of Asc and GSH concentrations measured at long TE, and provide a critical step towards addressing assumptions about T(2), and therefore towards the quantification of concentrations without the possibility of systematic bias.


A worksite prevention program for construction workers: design of a randomized controlled trial.
A worksite prevention program was developed to promote the work ability of construction workers and thereby prolong a healthy working life. The objective of this paper is to present the design of a randomized controlled trial evaluating the effectiveness of that intervention program compared with usual care for construction workers. The study is designed as a randomized controlled trial with a follow-up of one year. Employees eligible for this study are construction workers performing actual construction work. The worksite intervention will be compared with usual care. This intervention was developed by using the Intervention Mapping approach and consists of the following components: (1) two individual training sessions of a physical therapist to lower the physical workload, (2) a Rest-Break tool to improve the balance between work and recovery, and (3) two empowerment training sessions to increase the influence of the construction workers at the worksite. Outcome measures are assessed at baseline, 3, 6, and 12 months. The primary outcome measures of this study are work ability and health-related quality of life. Secondary outcome measures include need for recovery, musculoskeletal complaints, work engagement and self efficacy. Cost-effectiveness will be evaluated from the company perspective. Moreover, a process evaluation will be conducted. The feasibility of the intervention and the study has been enhanced by creating an intervention program that explicitly appeals to construction workers and will not interfere too much with the ongoing construction. The feasibility and effectiveness of this worksite prevention program will be investigated by means of an effect- and a process evaluation. If proven effective, this worksite prevention program can be implemented on a larger scale within the construction industry. NTR1278.


Patient-reported Outcomes and Quality of Life in Anemic and Symptomatic Patients With Myelofibrosis: Results From the MOMENTUM Study.
Myelofibrosis (MF) is a chronic myeloproliferative neoplasm that typically manifests with debilitating symptoms that progressively worsen, negatively impacting patients' quality of life. Fatigue is a multifactorial and burdensome MF-related symptom due to its severity, persistence, and prevalence, with anemia a contributing factor and major unmet need. Clinical trials of the Janus kinase (JAK)1/JAK2/activin A receptor type 1 inhibitor momelotinib have shown consistent anemia benefits, in addition to improvements in MF-related symptoms. The phase 3 MOMENTUM trial in symptomatic and anemic patients met its primary end point, with a greater proportion having a Myelofibrosis Symptom Assessment Form (MFSAF) Total Symptom Score (TSS) reduction ≥50% at week 24 with momelotinib versus danazol. To support the positive primary end point result, we conducted longitudinal, responder, and time-to-event analyses of patient-reported outcomes from MOMENTUM, as measured by the MFSAF, European Organization for Research and Treatment of Cancer Quality of Life Questionnaire (EORTC QLQ-C30), and Patient-Reported Outcomes Measurement Information System (PROMIS) assessments. These analyses demonstrated rapid and durable response benefits with momelotinib, with achievement of first TSS response by day 29 and continued improvement over time. Improvements favored momelotinib versus danazol for each MFSAF individual item, and greater improvements were observed for disease- and cancer-related fatigue and physical functioning at week 24, with significant results for multiple items/domains across the 3 assessments. These findings are consistent in demonstrating that momelotinib provides substantial symptom benefit.


Childhood lichen planus with nail involvement: a case.
Childhood lichen planus is a very rare entity which is characterized by violaceous, scaly, flat-topped polygonal papules commonly involving the flexor aspects of the wrists and legs, and oral and genital mucous embranes. The incidence of lichen planus peaks between the ages of 30 and 60 years, where as children comprise only 2% to 3% of reported cases. We report a nine-year-old girl with widespread lichen planus, involving the nails, with no mucosal involvement and regressing after treatment with a starting dosage of 20 mg/day systemic flucortolon (Ultralan) therapy.


Source space estimation of oscillatory power and brain connectivity in tinnitus.
Tinnitus is the perception of an internally generated sound that is postulated to emerge as a result of structural and functional changes in the brain. However, the precise pathophysiology of tinnitus remains unknown. Llinas' thalamocortical dysrhythmia model suggests that neural deafferentation due to hearing loss causes a dysregulation of coherent activity between thalamus and auditory cortex. This leads to a pathological coupling of theta and gamma oscillatory activity in the resting state, localised to the auditory cortex where normally alpha oscillations should occur. Numerous studies also suggest that tinnitus perception relies on the interplay between auditory and non-auditory brain areas. According to the Global Brain Model, a network of global fronto-parietal-cingulate areas is important in the generation and maintenance of the conscious perception of tinnitus. Thus, the distress experienced by many individuals with tinnitus is related to the top-down influence of this global network on auditory areas. In this magnetoencephalographic study, we compare resting-state oscillatory activity of tinnitus participants and normal-hearing controls to examine effects on spectral power as well as functional and effective connectivity. The analysis is based on beamformer source projection and an atlas-based region-of-interest approach. We find increased functional connectivity within the auditory cortices in the alpha band. A significant increase is also found for the effective connectivity from a global brain network to the auditory cortices in the alpha and beta bands. We do not find evidence of effects on spectral power. Overall, our results provide only limited support for the thalamocortical dysrhythmia and Global Brain models of tinnitus.


Amyloid-like Self-Assembly of a Cellular Compartment.
Most vertebrate oocytes contain a Balbiani body, a large, non-membrane-bound compartment packed with RNA, mitochondria, and other organelles. Little is known about this compartment, though it specifies germline identity in many non-mammalian vertebrates. We show Xvelo, a disordered protein with an N-terminal prion-like domain, is an abundant constituent of Xenopus Balbiani bodies. Disruption of the prion-like domain of Xvelo, or substitution with a prion-like domain from an unrelated protein, interferes with its incorporation into Balbiani bodies in vivo. Recombinant Xvelo forms amyloid-like networks in vitro. Amyloid-like assemblies of Xvelo recruit both RNA and mitochondria in binding assays. We propose that Xenopus Balbiani bodies form by amyloid-like assembly of Xvelo, accompanied by co-recruitment of mitochondria and RNA. Prion-like domains are found in germ plasm organizing proteins in other species, suggesting that Balbiani body formation by amyloid-like assembly could be a conserved mechanism that helps oocytes function as long-lived germ cells.


Use of alcohol swabs to clean injecting sites among people who regularly inject drugs in Australia.
Cleaning drug injection sites with alcohol swabs prior to injecting reduces risk of abscesses and other skin and soft tissue infections (SSTI). Better understanding of swabbing behaviours can inform interventions to improve injecting hygiene. We aimed to determine the socio-demographic, drug use and injecting risk exposure correlates of swabbing prior to injecting and reasons for not swabbing. The Illicit Drug Reporting System recruited participants who had injected drugs at least monthly in the past six months in June-July 2017 from all Australian capital cities via needle and syringe programs and word-of-mouth. A structured interview was used to collect information on drug use and related behaviour, as well as swabbing practices. Logistic regression was used to identify factors associated with not swabbing at last injection. Of 853 respondents, one-quarter (26%) reported that they did not swab prior to their last injection. In adjusted analyses, crystal methamphetamine as the last drug injected, past month receptive or distributive syringe sharing, and past month re-use of one's own needle were significantly associated with not swabbing at last injection. Among participants who did not swab at last injection, swabbing was frequently considered unnecessary and a small number disliked using alcohol swabs. Efforts are needed to increase awareness of the importance of injecting hygiene in preventing SSTI. Interventions to increase swabbing should be included as part of a wider package of injecting hygiene education, particularly in light of associations with receptive and/or distributive syringe sharing.


Self-Assembly of Supramolecular Light-Harvesting Arrays from Symmetric Perylene-3,4-dicarboximide Trefoils.
Unlike the widely studied perylene-3,4:9,10-bis(dicarboximide) (PDI) dyes, self-assembly of the corresponding perylene-3,4-dicarboximide (PMI) dyes into large arrays and studies of their excited state properties have received far less attention. Two symmetric PMI trefoils were synthesized by connecting the 9-position of the perylene core either directly (1) or through a phenylene linker (2) to the 1,3,5-positions of a central benzene ring. Synchrotron-based small- and wide-angle X-ray scattering measurements in methylcyclohexane show that trefoil 1 self-assembles into cofacial trimers (13) on average, while trefoil 2 forms much larger assemblies that are tridecamers (213) on average. Their photophysics were characterized using steady-state as well as transient absorption and emission spectroscopy. Time-resolved spectroscopy reveals that both 13 and 213 initially form excitonically coupled excited states that subsequently relax to excimer states having 20 and 8.4 ns lifetimes, respectively, which decay to ground-state primarily nonradiatively. The data are consistent with stronger electronic coupling between the PMI molecules in 213 relative to 13.


Liquid crystal clad near-infrared metamaterials with tunable negative-zero-positive refractive indices.
Near-infrared metamaterials that possess a reconfigurable index of refraction from negative through zero to positive values are presented. Reconfigurability is achieved by cladding thin layers of liquid crystal both as a superstrate and a substrate on an established negative-index metamaterial, and adjusting the permittivity of the liquid crystal. Numerical results show that the index of refraction for the proposed structure can be changed over the range from ?1 to +1.8 by tuning the liquid crystal permittivity from 2 to 6 at a wavelength of 1.4 ?m.


The reliability of the N400 in single subjects: implications for patients with disorders of consciousness.
Functional neuroimaging assessments of residual cognitive capacities, including those that support language, can improve diagnostic and prognostic accuracy in patients with disorders of consciousness. Due to the portability and relative inexpensiveness of electroencephalography, the N400 event-related potential component has been proposed as a clinically valid means to identify preserved linguistic function in non-communicative patients. Across three experiments, we show that changes in both stimuli and task demands significantly influence the probability of detecting statistically significant N400 effects - that is, the difference in N400 amplitudes caused by the experimental manipulation. In terms of task demands, passively heard linguistic stimuli were significantly less likely to elicit N400 effects than task-relevant stimuli. Due to the inability of the majority of patients with disorders of consciousness to follow task commands, the insensitivity of passive listening would impede the identification of residual language abilities even when such abilities exist. In terms of stimuli, passively heard normatively associated word pairs produced the highest detection rate of N400 effects (50% of the participants), compared with semantically-similar word pairs (0%) and high-cloze sentences (17%). This result is consistent with a prediction error account of N400 magnitude, with highly predictable targets leading to smaller N400 waves, and therefore larger N400 effects. Overall, our data indicate that non-repeating normatively associated word pairs provide the highest probability of detecting single-subject N400s during passive listening, and may thereby provide a clinically viable means of assessing residual linguistic function. We also show that more liberal analyses may further increase the detection-rate, but at the potential cost of increased false alarms.


Estimating the residency expansion required to avoid projected primary care physician shortages by 2035.
The purpose of this study was to calculate the projected primary care physician shortage, determine the amount and composition of residency growth needed, and estimate the impact of retirement age and panel size changes. We used the 2010 National Ambulatory Medical Care Survey to calculate utilization of ambulatory primary care services and the US Census Bureau to project demographic changes. To determine the baseline number of primary care physicians and the number retiring at 66 years, we used the 2014 American Medical Association Masterfile. Using specialty board and American Osteopathic Association figures, we estimated the annual production of primary care residents. To calculate shortages, we subtracted the accumulated primary care physician production from the accumulated number of primary care physicians needed for each year from 2015 to 2035. More than 44,000 primary care physicians will be needed by 2035. Current primary care production rates will be unable to meet demand, resulting in a shortage in excess of 33,000 primary care physicians. Given current production, an additional 1,700 primary care residency slots will be necessary by 2035. A 10% reduction in the ratio of population per primary care physician would require more than 3,000 additional slots by 2035, whereas changing the expected retirement age from 66 years to 64 years would require more than 2,400 additional slots. To eliminate projected shortages in 2035, primary care residency production must increase by 21% compared with current production. Delivery models that shift toward smaller ratios of population to primary care physicians may substantially increase the shortage.


Approaches for discovering anti-prion compounds: lessons learned and challenges ahead.
Recent years have witnessed major advances in our understanding of the molecular bases of prion diseases. These studies not only highlight the protein misfolding as a potential initiator of a neurodegenerative process, they also provide a foundation for considering whether such a process can be common to many neurodegenerative diseases, including Alzheimer's disease. This makes prion diseases a sort of prototype of neurodegenerative disease, endowed with some intrinsic positive features in terms of drug development. Thanks to the fact that disappearance of the scrapie protein can serve as a clear readout of drug efficiency, phenotypic approaches have high potential for prion disease drug discovery. In this review, the authors discuss phenotypic screening and how it lends itself to drug repositioning. Furthermore, they discuss the advantages of working with a molecule with proven safety, tolerability and drug-like properties in combination with a reliable phenotypic screening and how it could improve the success rate for prion drug development. They also provide examples of several interesting candidates that have been identified using this approach, including quinacrine, astemizole, guanabenz and doxycycline. The availability of persistently scrapie-infected murine neuroblastoma cells has greatly helped to identify compounds that inhibit prion formation. However, a human neuronal model infected with the human isoform would ultimately serve as the ideal disease model toward the discovery of effective drugs.


A Crystallin Fold in the Interleukin-4-inducing Principle of Schistosoma mansoni Eggs (IPSE/α-1) Mediates IgE Binding for Antigen-independent Basophil Activation.
The IL-4-inducing principle from Schistosoma mansoni eggs (IPSE/α-1), the major secretory product of eggs from the parasitic worm S. mansoni, efficiently triggers basophils to release the immunomodulatory key cytokine interleukin-4. Activation by IPSE/α-1 requires the presence of IgE on the basophils, but the detailed molecular mechanism underlying activation is unknown. NMR and crystallographic analysis of IPSEΔNLS, a monomeric IPSE/α-1 mutant, revealed that IPSE/α-1 is a new member of the βγ-crystallin superfamily. We demonstrate that this molecule is a general immunoglobulin-binding factor with highest affinity for IgE. NMR binding studies of IPSEΔNLS with the 180-kDa molecule IgE identified a large positively charged binding surface that includes a flexible loop, which is unique to the IPSE/α-1 crystallin fold. Mutational analysis of amino acids in the binding interface showed that residues contributing to IgE binding are important for IgE-dependent activation of basophils. As IPSE/α-1 is unable to cross-link IgE, we propose that this molecule, by taking advantage of its unique IgE-binding crystallin fold, activates basophils by a novel, cross-linking-independent mechanism.


Effects of Pesticide Mixtures on Host-Pathogen Dynamics of the Amphibian Chytrid Fungus.
Anthropogenic and natural stressors often interact to affect organisms. Amphibian populations are undergoing unprecedented declines and extinctions with pesticides and emerging infectious diseases implicated as causal factors. Although these factors often co-occur, their effects on amphibians are usually examined in isolation. We hypothesized that exposure of larval and metamorphic amphibians to ecologically relevant concentrations of pesticide mixtures would increase their post-metamorphic susceptibility to the fungus Batrachochytrium dendrobatidis (Bd), a pathogen that has contributed to amphibian population declines worldwide. We exposed five anuran species (Pacific treefrog, Pseudacris regilla; spring peeper, Pseudacris crucifer; Cascades frog, Rana cascadae; northern leopard frog, Lithobates pipiens; and western toad, Anaxyrus boreas) from three families to mixtures of four common insecticides (chlorpyrifos, carbaryl, permethrin, and endosulfan) or herbicides (glyphosate, acetochlor, atrazine, and 2,4-D) or a control treatment, either as tadpoles or as newly metamorphic individuals (metamorphs). Subsequently, we exposed animals to Bd or a control inoculate after metamorphosis and compared survival and Bd load. Bd exposure significantly increased mortality in Pacific treefrogs, spring peepers, and western toads, but not in Cascades frogs or northern leopard frogs. However, the effects of pesticide exposure on mortality were negligible, regardless of the timing of exposure. Bd load varied considerably across species; Pacific treefrogs, spring peepers, and western toads had the highest loads, whereas Cascades frogs and northern leopard frogs had the lowest loads. The influence of pesticide exposure on Bd load depended on the amphibian species, timing of pesticide exposure, and the particular pesticide treatment. Our results suggest that exposure to realistic pesticide concentrations has minimal effects on Bd-induced mortality, but can alter Bd load. This result could have broad implications for risk assessment of amphibians; the outcome of exposure to multiple stressors may be unpredictable and can differ between species and life stages.


Examining the effects of HIV self-testing compared to standard HIV testing services: a systematic review and meta-analysis.
HIV self-testing (HIVST) is a discreet and convenient way to reach people with HIV who do not know their status, including many who may not otherwise test. To inform World Health Organization (WHO) guidance, we assessed the effect of HIVST on uptake and frequency of testing, as well as identification of HIV-positive persons, linkage to care, social harm, and risk behaviour. We systematically searched for studies comparing HIVST to standard HIV testing until 1 June 2016. Meta-analyses of studies reporting comparable outcomes were conducted using a random-effects model for relative risks (RR) and 95% confidence intervals. The quality of evidence was evaluated using GRADE. After screening 638 citations, we identified five randomized controlled trials (RCTs) comparing HIVST to standard HIV testing services among 4,145 total participants from four countries. All offered free oral-fluid rapid tests for HIVST and were among men. Meta-analysis of three RCTs showed HIVST doubled uptake of testing among men (RR = 2.12; 95% CI: 1.51, 2.98). Meta-analysis of two RCTs among men who have sex with men showed frequency of testing nearly doubled (Rate ratio = 1.88; 95% CI: 1.17; 3.01), resulting in two more tests in a 12-15-month period (Mean difference = 2.13; 95% CI: 1.59, 2.66). Meta-analysis of two RCTs showed HIVST also doubled the likelihood of an HIV-positive diagnosis (RR = 2.02; 95% CI: 0.37, 10.76, 5.32). Across all RCTs, there was no indication of harm attributable to HIVST and potential increases in risk-taking behaviour appeared to be minimal. HIVST is associated with increased uptake and frequency of testing in RCTs. Such increases, particularly among those at risk who may not otherwise test, will likely identify more HIV-positive individuals as compared to standard testing services alone. However, further research on how to support linkage to confirmatory testing, prevention, treatment and care services is needed. WHO now recommends HIVST as an additional HIV testing approach.


Quantitative phase retrieval of a complex-valued object using variable function orders in the fractional Fourier domain.
We propose a novel and effective method to quantitatively recover a complex-valued object from diffraction intensity maps recorded in the fractional Fourier domain. A wavefront modulation is introduced in the wave path, and several diffraction intensity maps are recorded through variable function orders in the fractional Fourier transform. A new phase retrieval algorithm is then proposed, and advantages of the proposed algorithm are also discussed. A proof-of-principle study is presented to show the feasibility and effectiveness of the proposed method.


Effects of free, cued and modelled reflection on medical students' diagnostic competence.
Structured reflection while practising the diagnosing of cases has been shown to improve medical students' learning of clinical diagnosis. The present study investigated whether additional instructional guidance increases the benefits of reflection by comparing the effects of free, cued and modelled reflection on learning. Fifty-eight Year 5 and 57 Year 6 medical students participated in a three-phase experiment. During the learning phase, participants diagnosed eight clinical cases under different experimental conditions: free reflection; cued reflection, and modelled reflection. In an immediate test and a delayed test administered 1 week later, they diagnosed new sets of eight different cases, four of which presented diseases they had studied during the learning phase. Learning was measured according to diagnostic accuracy on the cases that involved the four diseases that appeared in all phases. Repeated-measures analysis of variance (anova) of mean scores for diagnostic accuracy (range: 0-1) showed a significant main effect of experimental condition (p < 0.001), year of training (p = 0.013), and performance moment (p = 0.003), without significant interaction effects. Overall, the modelled reflection group and the cued reflection group did not differ in performance (p = 1.00), but both outperformed the free reflection group (p < 0.001 for both comparisons). Overall performance increased in the delayed test relative to the immediate test (p = 0.004) and to the learning phase (p = 0.03), but did not differ in the latter two phases. Both Year 6 and Year 5 students rated studying examples of reflection as less effortful than either cued or free reflection in the learning phase (p < 0.001 for all comparisons). Students apparently learn more with less effort by studying correct structured reflection while practising the diagnosing of cases than by reflecting without any instructional guidance. Examples of reflection and cued reflection were more beneficial for learning than free reflection and may represent a useful instructional strategy for clinical teaching.


'Candidatus Planktophila limnetica', an actinobacterium representing one of the most numerically important taxa in freshwater bacterioplankton.
Actinobacteria often constitute a large fraction of the bacterioplankton in freshwater systems. Cultivation-independent methods have revealed that the so-called acI lineage frequently represents the most numerous taxon among assemblages of freshwater Actinobacteria and even among total freshwater bacterioplankton. Bacteria affiliated with this uncultivated lineage have been detected in freshwater habitats located in various continents and climatic zones but have never been found among terrestrial or offshore marine systems. So far, this ecologically important lineage of freshwater Actinobacteria is not represented by a recognized taxon. In this study, we established a stable mixed culture containing a strain affiliated with the acI lineage from a freshwater lake in Austria. The proportion of the strain in the culture could be increased by manipulation of the medium composition by more than one order of magnitude, however all subsequent attempts to isolate this strain into pure culture were unsuccessful. Some of the phenotypic traits of this acI strain were determined and its taxonomic position within the Actinobacteria was analysed. Phylogenetic analysis of this organism's 16S rRNA gene revealed a distant relationship with cultivated organisms and recognized species (89 % gene sequence similarity with the latter). Furthermore, this analysis did not support a clear assignment of the strain to any of the recognized families within the phylum Actinobacteria. It is suggested that a candidate taxon, 'Candidatus Planktophila limnetica' is established to represent this strain.


Inferring Gene Regulatory Networks from a Population of Yeast Segregants.
Constructing gene regulatory networks is crucial to unraveling the genetic architecture of complex traits and to understanding the mechanisms of diseases. On the basis of gene expression and single nucleotide polymorphism data in the yeast, Saccharomyces cerevisiae, we constructed gene regulatory networks using a two-stage penalized least squares method. A large system of structural equations via optimal prediction of a set of surrogate variables was established at the first stage, followed by consistent selection of regulatory effects at the second stage. Using this approach, we identified subnetworks that were enriched in gene ontology categories, revealing directional regulatory mechanisms controlling these biological pathways. Our mapping and analysis of expression-based quantitative trait loci uncovered a known alteration of gene expression within a biological pathway that results in regulatory effects on companion pathway genes in the phosphocholine network. In addition, we identify nodes in these gene ontology-enriched subnetworks that are coordinately controlled by transcription factors driven by trans-acting expression quantitative trait loci. Altogether, the integration of documented transcription factor regulatory associations with subnetworks defined by a system of structural equations using quantitative trait loci data is an effective means to delineate the transcriptional control of biological pathways.


Simplified method for detecting tritium contamination in plants and soil.
Cost-effective methods are needed to identify the presence and distribution of tritium near radioactive waste disposal and other contaminated sites. The objectives of this study were to (i) develop a simplified sample preparation method for determining tritium contamination in plants and (ii) determine if plant data could be used as an indicator of soil contamination. The method entailed collection and solar distillation of plant water from foliage, followed by filtration and adsorption of scintillation-interfering constituents on a graphite-based solid phase extraction (SPE) column. The method was evaluated using samples of creosote bush [Larrea tridentata (Sessé & Moc. ex DC.) Coville], an evergreen shrub, near a radioactive disposal area in the Mojave Desert. Laboratory tests showed that a 2-g SPE column was necessary and sufficient for accurate determination of known tritium concentrations in plant water. Comparisons of tritium concentrations in plant water determined with the solar distillation-SPE method and the standard (and more laborious) toluene-extraction method showed no significant difference between methods. Tritium concentrations in plant water and in water vapor of root-zone soil also showed no significant difference between methods. Thus, the solar distillation-SPE method provides a simple and cost-effective way to identify plant and soil contamination. The method is of sufficient accuracy to facilitate collection of plume-scale data and optimize placement of more sophisticated (and costly) monitoring equipment at contaminated sites. Although work to date has focused on one desert plant, the approach may be transferable to other species and environments after site-specific experiments.


Assessment of right ventricular regional contraction and comparison with the left ventricle in normal humans: a cine magnetic resonance study with presaturation myocardial tagging.
Right ventricular regional contractility has been thought to be difficult to assess precisely. Cine magnetic resonance imaging with presaturation myocardial tagging was employed to quantitate the contraction of the right ventricular free wall and to identify normal performance compared with the left ventricle. Nine normal volunteers, aged 27-39 years, were examined in a 1.5 Tesla superconductive magnet, and short axis and four-chamber sections at the mid-ventricular level were imaged with cine magnetic resonance sequences. Tags, applied at end diastole as two parallel black lines, intersected the mid-portion of the free wall, dividing it into upper, centre, and lower segments in the short axis section, and anterior, middle, and posterior segments in the four-chamber section. From a series of cine magnetic resonance images at 50 ms intervals over a cardiac cycle, end diastolic, and early, mid-, and end systolic images were chosen for calculation of the endocardial, epicardial, and mean percent fractional shortening (%FS) in the six segments. There was (1) a gradual increase in %FS in systole in both sections (P < 0.001, < 0.005); (2) a poor transmural gradient of contractility; (3) a predominance of meridional shortening (whole length, mean end systolic %FS (SD): short axis, 17.4 (3.1)%; four-chamber, 30.1 (4.1)%; P < 0.001) in contrast to dominant circumferential shortening in the left ventricular lateral wall; (4) lower predominance of contractility in the short axis section (P < 0.001), and a middle dip of contractility in the four-chamber section (P < 0.005). Heterogeneity of contractility was closely correlated with the myocardial fibre architecture, and with wall stress determined by its thickness and curvature. It was proved that right ventricular regional function could be analysed non-invasively using cine magnetic resonance imaging with myocardial tagging.


Natural Sensations Evoked in Distal Extremities Using Surface Electrical Stimulation.
Electrical stimulation is increasingly relevant in a variety of medical treatments. In this study, surface electrical stimulation was evaluated as a method to non-invasively target a neural function, specifically natural sensation in the distal limbs. Electrodes were placed over the median and ulnar nerves at the elbow and the common peroneal and lateral sural cutaneous nerves at the knee. Strength-duration curves for sensation were compared between nerves. The location, modality, and intensity of each sensation were also analyzed. In an effort to evoke natural sensations, several patterned waveforms were evaluated. Distal sensation was obtained in all but one of the 48 nerves tested in able-bodied subjects and in the two nerves from subjects with an amputation. Increasing the pulse amplitude of the stimulus caused an increase in the area and magnitude of the sensation in a majority of subjects. A low frequency waveform evoked a tapping or tapping-like sensation in 29 out of the 31 able-bodied subjects and a sensation that could be considered natural in two subjects with an amputation. This waveform performed better than other patterned waveforms that had proven effective during implanted extra-neural stimulation. Surface electrical stimulation has the potential to be a powerful, non-invasive tool for activation of the nervous system. These results suggest that a tapping sensation in the distal extremity can be evoked in most able-bodied individuals and that targeting the nerve trunk from the surface is a valid method to evoke sensation in the phantom limb of individuals with an amputation for short term applications.


Division of labour and terminal differentiation in a novel Bacillus thuringiensis strain.
A major challenge in bacterial developmental biology has been to understand the mechanisms underlying cell fate decisions. Some differentiated cell types display cooperative behaviour. Cooperation is one of the greatest mysteries of evolutionary biology and microbes have been considered as an excellent system for experimentally testing evolution theories. Bacillus thuringiensis (Bt) is a spore-forming bacterium, which is genetically closely related to B. anthracis, the agent of anthrax, and to B. cereus, an opportunistic human pathogen. The defining feature that distinguishes Bt from its relatives is its ability to produce crystal inclusions in the sporulating cells. These toxins are solubilized after ingestion and are cooperative public goods in insect hosts. In this study, we describe a Bt strain LM1212 that presents the unique ability to terminally differentiate into crystal producers and spore formers. Transcriptional analysis based on lacZ and gfp reporter genes suggested that this phenotype is the consequence of a new type of cell differentiation associated with a novel regulation mode of cry gene expression. The differentiating crystal-producer phenotype has higher spore productivity than a typical Bt strain and is better able to compete with Cry toxin null 'cheaters'. Potentially, this division of labour provides additional fitness benefits in terms of spore viability or durability of Cry toxin.


Variability of maximal aortic aneurysm diameter measurements on CT scan: significance and methods to minimize.
We noted substantial differences when measuring repeatedly the same abdominal aortic aneurysm (AAA) on the same computed tomography (CT) scan. This study quantitated this variability, and methods to minimize it were developed. The CT maximal diameter of 25 AAAs was measured by eight experienced observers, including six vascular surgeons and two radiologists, using two methods: an unstandardized protocol, and a standardized protocol using fine calipers to carefully measure the largest diameter perpendicular to the estimated aneurysm centerline, from outer aneurysm wall to outer wall. The average measurement difference between observers was calculated for each method. The average difference between each observer's measurement and the official radiology report value was also calculated. Agreement between the two measurement methods was assessed with Bland-Altman plots. The difference in maximal diameter measurements between each observer averaged 4.0 +/- 5.1 mm (range, 0.0-35.0 mm) with the unstandardized method. The mean measurement difference with the standardized protocol was significantly lower, and averaged 2.8 +/- 4.4 mm (range, 0.0-26.0 mm; P<.05). Measurements taken from the official radiology report differed from each of the observer's standardized measurement by an average of 5.0 +/- 6.3 mm (range, 0.0-28.0 mm). This difference was similar for both the unstandardized and standardized methods. Bland-Altman plots confirmed the wide variation of the maximal diameter measurements when the unstandardized method was compared with the standardized method (95% confidence interval, -9-9 mm). Routine CT maximal diameter measurement of AAAs can have substantial interobserver variability. Standardized measurement protocols can decrease, but not eliminate, this measurement variability. Thus apparent size changes based on CT measurements may represent measurement artifact rather than actual aneurysm growth or shrinkage, particularly when a standardized system is not used.


Use of the PROMIS® Depression scale and the Beck Depression Inventory in patients with heart failure.
This study evaluated agreement between the Patient-Reported Outcomes Measurement Information System® (PROMIS®) Depression scale and the Beck Depression Inventory (BDI-II) in patients with heart failure and comorbid major depression. The BDI-II and the computerized adaptive test version of the PROMIS® Depression scale were administered at baseline to 158 participants in a randomized controlled trial of cognitive behavior therapy for major depression in patients with heart failure. A crosswalk table (Choi, Schalet, Cook, & Cella, 2014) was used to transform the PROMIS® scores into "linked" BDI-II equivalent scores. Bland-Altman plots, histograms, and scatterplots were used to visualize the agreement between these scores at baseline and 6 months, and intraclass correlation coefficients (ICCs) were calculated for each occasion to quantify the agreement. Treatment effects and change scores were also examined. The measures agreed moderately at baseline (ICC = 0.52, p < .0001) and strongly at 6 months (ICC = 0.77, p < .0001), but on average, the linked and observed BDI-II scores differed by 3.1 points at baseline (p < .0001) and -0.17 points at 6 months (p = .78). The discrepancies were considerably larger in many individual cases on both occasions. The PROMIS® Depression scale is likely to play an important role in research on depression in patients with heart failure, but for now, it should be used in addition to rather than instead of the BDI-II in studies in which the BDI-II would ordinarily be used. Additional research is needed to evaluate the validity and utility of the PROMIS® Depression scale in patients with heart failure. (PsycINFO Database Record (c) 2019 APA, all rights reserved).


Aberrant response inhibition and task switching in psychopathic individuals.
Deficits in cognitive control have been considered a core dysfunction of psychopathy, responsible for disrupted self-control. We investigated cognitive control impairments, including difficulties with task switching, failure of response inhibition, and inability to adjust speed of responding. Participants included 16 subjects with psychopathic traits (Ps), and 22 healthy controls (HCs). We recorded behavioral responses during a Task Switching paradigm, a probe of flexible behavioral adaptation to changing contexts; and a Go/NoGo Task, which assesses response inhibition and indexes behavioral impulsivity. During task switching, Ps evidenced impairments shifting set when conflicting (incongruent) information was presented, but performed as well as HCs in the absence of such conflict. In addition, when they encountered these difficulties, they failed to adjust their speed of responding. Ps presented also with deficits in response inhibition, with many commission errors on the Go/NoGo Task. This study identified impairments in response inhibition and in set shifting in psychopathic individuals. When shifting set, they evidenced difficulties refocusing on a new task when it was incongruent with the previous task. These deficits interfere with regulation of ongoing behavior and disrupt self-regulation. Our findings suggest abnormal neural processing during suppression of inappropriate responses in psychopathic individuals.


Low pressure giant pulmonary artery aneurysms in the adult: natural history and management strategies.
To describe aspects of the natural history and pathophysiology of giant low pressure pulmonary artery aneurysms and to propose potential surgical strategies. Cross sectional retrospective review. Supraregional tertiary referral centre. All adult patients referred for assessment of giant pulmonary artery aneurysm retrospectively identified from the Mayo Adult Congenital Heart Disease Clinic database. Patient data were reviewed from hospital records, including echocardiograms, magnetic resonance images, radiographs, and histology slides. Four patients were identified with a median age of 52 years (range 37-64 years). Presenting symptoms were effort related dyspnoea, chest discomfort, and hoarseness in one patient. All patients had pulmonary regurgitation and clinical evidence of right ventricular enlargement in association with a pulsatile mass at the upper left sternal edge. Transthoracic echocardiography showed the giant pulmonary artery aneurysm involving the main pulmonary artery and proximal branches, and confirmed severe pulmonary regurgitation in all patients. None of the patients had intimal tearing, medial dissection, or pulmonary arterial rupture. The pulmonary valve was replaced to relieve symptoms and preserve right ventricular function. Pulmonary arterial histology showed medial degeneration of elastic fibres with accumulation of basophilic ground substance. Rupture or dissection of these low pressure aneurysms is rare. The timing of surgical intervention should be determined by changes in right ventricular size and function resulting from pulmonary regurgitation or pulmonary stenosis, and not the size of the aneurysm.


Autoimmune Encephalitis in Children: A Case Series at a Tertiary Care Center.
Autoimmune encephalitis is the third most common cause of encephalitis in children. We provide a detailed account of presenting symptoms, diagnosis, and response to treatment in pediatric autoimmune encephalitis patients evaluated at University of California San Francisco within a 2.5-year period. Eleven were identified: anti-N-methyl-d-aspartate receptor (NMDAR) encephalitis (n = 4), antibody-negative autoimmune encephalitis (n = 4), steroid-responsive encephalopathy associated with thyroiditis (SREAT) (n = 2), and glial fibrillary acidic protein (GFAP)-associated encephalitis (n = 1). Most common presenting symptoms included seizures and behavior changes (54%). More than 90% of patients showed improvement following first-line immunotherapy (high-dose corticosteroids, intravenous immunoglobulin, and/or plasma exchange). A total of 64% received second-line treatment with rituximab, cyclophosphamide, or mycophenolate mofetil. One patient with NMDAR encephalitis died despite escalating immunotherapy. None of the patients showed complete recovery after median follow-up of 9 months (range 0.5-66). Children with autoimmune encephalitis have a diverse clinical presentation and may lack an identifiable autoantibody. Majority of patients show a good response to immunotherapy; however, recovery can be delayed.


Caudal ropivacaine-clonidine: A better post-operative analgesic approach.
The aim was to determine qualitative and quantitative aspects of caudal block, haemodynamic effects, and post-operative pain relief of ropivacaine 0.25% versus ropivacaine 0.25% with clonidine for lower abdominal surgeries in paediatric patients. A double-blind study was conducted among 44 paediatric patients in the Department of Anaesthesiology and Intensive Care of our institute. A total of 44 ASA-I paediatric patients between the ages of 1 and 9 years, scheduled for elective hernia surgery, were enrolled in this randomised double-blind study. The caudal block was administered with ropivacaine 0.25% (Group I) and ropivacaine 0.25% and clonidine 2 µg/kg (Group II) after induction with general anaesthesia. Haemodynamic parameters were observed before, during and after the surgical procedure. Post-operative analgesic duration, total dose of rescue analgesia, pain scores and any side effects were looked for and recorded. All the results were tabulated and analysed statistically. The variables in the two groups were compared using the non-parametric tests. For all statistical analyses, the level of significance was P < 0.05. Forty-four patients were enrolled in this study and their data were subjected to statistical analysis: 22 patients in both the groups were comparable with regard to demographic data, haemodynamic parameters and other vitals and were statistically non-significant (P>0.05). The duration of analgesia was significantly prolonged in Group II (P<0.05). The dose requirement for post-operative pain relief was also significantly lesser in Group II. The incidences of side effects were almost comparable and non-significant. A caudal block with 0.25% of isobaric ropivacaine combined with 2 µg/kg of clonidine provides efficient analgesia intra-operatively and prolonged duration of analgesia post-operatively.


Phosphorylation of drebrin by cyclin-dependent kinase 5 and its role in neuronal migration.
Cyclin-dependent kinase 5 (Cdk5)-p35 is a proline-directed Ser/Thr kinase which plays a key role in neuronal migration, neurite outgrowth, and spine formation during brain development. Dynamic remodeling of cytoskeletons is required for all of these processes. Cdk5-p35 phosphorylates many cytoskeletal proteins, but it is not fully understood how Cdk5-p35 regulates cytoskeletal reorganization associated with neuronal migration. Since actin filaments are critical for the neuronal movement and process formation, we aimed to find Cdk5 substrates among actin-binding proteins. In this study, we isolated actin gels from mouse brain extracts, which contain many actin-binding proteins, and phosphorylated them by Cdk5-p35 in vitro. Drebrin, a side binding protein of actin filaments and well known for spine formation, was identified as a phosphorylated protein. Drebrin has two isoforms, an embryonic form drebrin E and an adult type long isoform drebrin A. Ser142 was identified as a common phosphorylation site to drebrin E and A and Ser342 as a drebrin A-specific site. Phosphorylated drebrin is localized at the distal area of total drebrin in the growth cone of cultured primary neurons. By expressing nonphosphorylatable or phosphorylation mimicking mutants in developing neurons in utero, the reversible phosphorylation/dephosphorylation reaction of drebrin was shown to be involved in radial migration of cortical neurons. These results suggest that Cdk5-p35 regulates neuronal migration through phosphorylation of drebrin in growth cone processes.


Insulin secretion and insulin sensitivity in relation to glucose tolerance: lessons from the Botnia Study.
Recently, a new stage in glucose tolerance, impaired fasting glucose (IFG) (fasting plasma glucose level of 6.1-6.9 mmol/l), was introduced in addition to impaired glucose tolerance (IGT) (2-h glucose level of 7.8-11.0 mmol/l). It is not clear whether IFG and IGT differ with respect to insulin secretion or sensitivity. To address this question, we estimated insulin secretion (by measuring both insulin levels and the ratio of insulin-to-glucose levels in 30-min intervals) and insulin sensitivity (by using the homeostasis model assessment [HOMA] index) from an oral glucose tolerance test (OGTT) in 5,396 individuals from the Botnia Study who had varying degrees of glucose tolerance. There was poor concordance between IFG and IGT: only 36% (303 of 840) of the subjects with IFG had IGT, whereas 62% (493 of 796) of the subjects with IGT did not have IFG. Compared with subjects with normal glucose tolerance (NGT), subjects with IFG were more insulin resistant (HOMA-insulin resistance [IR] values 2.64 +/- 0.08 vs. 1.73 +/- 0.03, P < 0.0005), had greater insulin responses during an OGTT (P = 0.0001), had higher waist-to-hip ratios (P < 0.005), had higher triglyceride and total cholesterol concentrations (P < 0.0005), and had lower HDL cholesterol concentrations (P = 0.0001). Compared with subjects with IFG, subjects with IGT had a lower incremental 30-min insulin-to-glucose area during an OGTT (13.8 +/- 1.7 vs. 21.7 +/- 1.7, P = 0.0008). Compared with subjects with IGT, subjects with mild diabetes (fasting plasma glucose levels <7.8 mmol/l) showed markedly impaired insulin secretion that could no longer compensate for IR and elevated glucose levels. A progressive decline in insulin sensitivity was observed when moving from NGT to IGT and to subjects with diabetes (P < 0.05 for trend), whereas insulin secretion followed an inverted U-shaped form. We conclude that IFG is characterized by basal IR and other features of the metabolic syndrome, whereas subjects with IGT have impaired insulin secretion in relation to glucose concentrations. An absolute decompensation of beta-cell function characterizes the transition from IGT to mild diabetes.


Effect of nitrous acid on lung function in asthmatics: a chamber study.
Nitrous acid, a component of photochemical smog and a common indoor air pollutant, may reach levels of 100 ppb where gas stoves and unvented portable kerosene heaters are used. Nitrous acid is a primary product of combustion and may also be a secondary product by reaction of nitrogen dioxide with water. Because the usual assays for nitrogen dioxide measure several oxides of nitrogen (including nitrous acid) together, previous studies of indoor nitrogen dioxide may have included exposure to and health effects of nitrous acid. To assess the respiratory effects of nitrous acid exposure alone, we carried out a double-blinded crossover chamber exposure study with 11 mildly asthmatic adult subjects. Each underwent 3-hr exposures to 650 ppb nitrous acid and to filtered room air with three 20-min periods of moderate cycle exercise. Symptoms, respiratory parameters during exercise, and spirometry after exercise were measured. A statistically significant decrease in forced vital capacity was seen on days when subjects were exposed to nitrous acid. This effect was most marked at 25 min and 85 min after exposure began. Aggregate respiratory and mucous membrane symptoms were also significantly higher with nitrous acid. We conclude that this concentration and duration of exposure to nitrous acid alters lung mechanics slightly, does not induce significant airflow obstruction, and produces mild irritant symptoms in asthmatics.