'Sometimes people let love conquer them': how love, intimacy, and trust in relationships between men who have sex with men influence perceptions of sexual risk and sexual decision-making.
Men who have sex with men account for a disproportionate burden of HIV incidence in the USA. Although much research has examined the drivers of sexual risk-taking, the emotional contexts in which men make sexual decisions has received little attention. In this three-phase, 10-week longitudinal qualitative study involving 25 gay and bisexual men, we used timeline-based interviews and quantitative web-based diaries about sexual and/or dating partners to examine how emotions influence HIV risk perceptions and sexual decision-making. Participants described love, intimacy, and trust as reducing HIV risk perceptions and facilitating engagement in condomless anal intercourse. Lust was not as linked with risk perceptions, but facilitated non condom-use through an increased willingness to engage in condomless anal intercourse, despite perceptions of risk. Results indicate that gay and bisexual men do not make sexual decisions in an emotional vacuum. Emotions influence perceptions of risk so that they do not necessarily align with biological risk factors. Emotional influences, especially the type and context of emotions, are important to consider to improve HIV prevention efforts among gay and bisexual men.


Peer influence: use of alcohol, tobacco, and prescription medications.
Risk-taking behavior (e.g., alcohol abuse, tobacco usage, misuse of prescription medications) among college students is a widespread problem. This study focused not only on the frequency of risky health behaviors in college students, but also the companions with whom they engaged in such behaviors. Three hundred and twelve college students completed a survey examining the frequency with which they engaged in alcohol, tobacco, and improper prescription medication use, as well as with whom they were most likely to engage in these behaviors. Results indicated that participants were most likely to take health risks when accompanied by someone they consider a friend. Results also indicated gender differences in risk-taking behaviors, as well as an interaction effect between companion and gender. This information would be useful when developing preventive interventions for college students. Implementing interventions that are specific to certain populations might generate greater success in reducing risk-taking behavior.


Gene activation in plastids by the CRE site-specific recombinase.
We developed a novel system for gene activation in plastids that uses the CRE/loxP site-specific recombination system to create a translatable reading frame by excision of a blocking sequence. To test the system, we introduced an inactive gfp* gene into the tobacco plastid genome downstream of the selectable spectinomcyin resistance (aadA) marker gene. The aadA gene is the blocking sequence, and is flanked by directly oriented loxP sites for excision by the CRE. In the non-activated state, gfp* is transcribed from the aadA promoter, but the mRNA is not translated due to the lack of an AUG translation initiation codon. Green Fluorescent Protein (GFP) expression is activated by excision of the aadA coding segment to link up the gfp* coding region with the translation initiation codon of aadA. Tobacco plants that carry the inactive gfp* gene do not contain detectable levels of GFP. However, activation of gfp* resulted in GFP accumulation, proving the utility of CRE-induced protein expression in tobacco chloroplasts. The gene activation system described here will be useful to probe plastid gene function and for the production of recombinant proteins in chloroplasts.


Risk of Guillain-Barré syndrome after 2010-2011 influenza vaccination.
Influenza vaccination has been implicated in Guillain Barré Syndrome (GBS) although the evidence for this link is controversial. A case-control study was conducted between October 2010 and May 2011 in seven Italian Regions to explore the relation between influenza vaccination and GBS. The study included 176 GBS incident cases aged ≥18 years from 86 neurological centers. Controls were selected among patients admitted for acute conditions to the Emergency Department of the same hospital as cases. Each control was matched to a case by sex, age, Region and admission date. Two different analyses were conducted: a matched case-control analysis and a self-controlled case series analysis (SCCS). Case-control analysis included 140 cases matched to 308 controls. The adjusted matched odds ratio (OR) for GBS occurrence within 6 weeks after influenza vaccination was 3.8 (95 % CI: 1.3, 10.5). A much stronger association with gastrointestinal infections (OR = 23.8; 95 % CI 7.3, 77.6) and influenza-like illness or upper respiratory tract infections (OR = 11.5; 95 % CI 5.6, 23.5) was highlighted. The SCCS analysis included all 176 GBS cases. Influenza vaccination was associated with GBS, with a relative risk of 2.1 (95 % CI 1.1, 3.9). According to these results the attributable risk in adults ranges from two to five GBS cases per 1,000,000 vaccinations.


Leveraging Home-Delivered Meal Programs to Address Unmet Needs for At-Risk Older Adults: Preliminary Data.
Home-delivered meal programs serve a predominantly homebound older adult population, characterized by multiple chronic conditions, functional limitations, and a variety of complex care needs, both medical and social. A pilot study was designed to test the feasibility of leveraging routine meal-delivery service in two home-delivered meal programs to proactively identify changes in older adult meal recipients' (clients') health, safety, and well-being and address unmet needs. Meal delivery personnel (drivers) were trained to use a mobile application to submit electronic alerts when they had a concern or observed a change in a client's condition. Alerts were received by care coordinators, who followed up with clients to offer support and help connect them to health and community services. Over a 12-month period, drivers submitted a total of 429 alerts for 189 clients across two pilot sites. The most frequent alerts were submitted for changes in health (56%), followed by self-care or personal safety (12%) and mobility (11%). On follow-up, a total of 132 referrals were issued, with most referrals for self-care (33%), health (17%), and care management services (17%). Focus groups conducted with drivers indicated that most found the mobile application easy to use and valued change of condition monitoring as an important contribution. Findings suggest that this is a feasible approach to address unmet needs for vulnerable older adults and may serve as an early-warning system to prevent further decline and improve quality of life. Efforts are underway to test the protocol across additional home-delivered meal programs. J Am Geriatr Soc 67:1946-1952, 2019.


Longitudinal measures of perfluoroalkyl substances (PFAS) in serum of Gullah African Americans in South Carolina: 2003-2013.
Charleston Harbor has elevated concentrations of PFAS in dolphins, but local human exposure data are limited. We sought to describe PFAS serum concentrations' temporal trends among Gullah African American residents of coastal South Carolina. Longitudinal measures of PFAS in blood serum from a Gullah clinical sample, without lupus, were examined using spaghetti plots and visit-to-visit change scores (e.g., differences in concentrations between visits) among the 68 participants with repeated measures available. We also modeled population-level trends among the 71 participants with any data using proportionate percentile models, accounting for clustering through robust standard errors. In a post-hoc analysis we examined heterogeneity of temporal trends by age through mixed-effects models for the log-transformed PFAS compounds. Population concentrations of PFOS dropped approximately 9 (95% CI: 8, 10) percent each year over 2003-2013. This was concordant with individual PFOS trajectories (median PFOS change score -21.7 ng/g wet weight, interquartile range of PFOS change scores: -32.8, -14.9) and reports for other populations over this time period. Several other compounds including PFOA, PFHxS, and PFuNDA also showed a population-level decrease. However, examination of individual trajectories suggested substantial heterogeneity. Post-hoc analyses indicated that PFAS trajectories were heterogeneous by age. Many PFAS compounds are decreasing in a sample of Gullah African Americans from coastal South Carolina. There may be age differences in the elimination kinetics of PFASs. The possible role of age as a modifier of PFAS serum trends merits further research.


Outcomes of idiopathic chronic large pericardial effusion.
Aim of this paper is to evaluate the outcomes of 'idiopathic' chronic large pericardial effusions without initial evidence of pericarditis. All consecutive cases of idiopathic chronic large pericardial effusions evaluated from 2000 to 2015 in three Italian tertiary referral centres for pericardial diseases were enrolled in a prospective cohort study. The term 'idiopathic' was applied to cases that performed a complete diagnostic evaluation to exclude a specific aetiology. A clinical and echocardiographic follow-up was performed every 3-6 months. 100 patients were included (mean age 61.3±14.6 years, 54 females, 44 patients were asymptomatic according to clinical evaluation) with a mean follow-up of 50 months. The baseline median size of the effusion (evaluated as the largest end-diastolic echo-free space) was 25 mm (IQR 8) and decreased to a mean value of 7 mm (IQR 19; p<0.0001) with complete regression in 39 patients at the end of follow-up. There were no new aetiological diagnoses. Adverse events were respectively: cardiac tamponade in 8 patients (8.0%), pericardiocentesis in 30 patients (30.0%), pericardial window in 12 cases (12.0%) and pericardiectomy in 3 patients (3.0%). Recurrence-free survival and complications-free survival was better in patients treated without interventions (log rank p=0.0038). The evolution of 'idiopathic' chronic large pericardial effusions is usually benign with reduction of the size of the effusion in the majority of cases, and regression in about 40% of cases. The risk of cardiac tamponade is 2.2%/year and recurrence/complications survival was better in patients treated conservatively without interventions.


Frequency analysis of the fibrillatory activity from surface ECG lead V1 and intracardiac recordings: implications for mapping of AF.
Fibrillatory waves observed in the surface electrograms may be a direct reflection of the electrophysiologic mechanism of the atrial fibrillation (AF). This study compared the fibrillatory waves in the surface ECG and the individual intracardiac mapping sites in different types of paroxysmal AF. Thirty patients with paroxysmal AF originating from the pulmonary veins (PVs) or superior vena cava (SVC) were enrolled. Frequency analysis was performed on the intracardiac electrograms recorded from various mapping sites in both atria sequentially with simultaneous surface electrogram recordings. The SVC-AF patients had a trend toward a higher DF in ECG lead V1 when compared with the PV-AF patients (7.35 +/- 2.09 vs. 5.89 +/- 0.79 Hz, P = 0.018). The mean dominant frequency (DF) of the LA mapping sites in the PV-AF patients was higher than that in the SVC-AF patients (7.06 +/- 0.66 vs. 6.13 +/- 0.96 Hz, P = 0.009), whereas the mean DF of the RA mapping sites was similar between the two groups (5.84 +/- 0.80 vs. 6.26 +/- 1.11 Hz, P = NS). The intra-class correlation coefficient (ICC) between the mean DF of the RA sites and V1 was higher (r = 0.21, P = 0.02) when compared with the mean DF of the LA sites (r = -0.007, P > 0.05). Furthermore, the maximal ICC was observed in the anterolateral RA free wall (r = 0.84, P < 0.001) and not the other anatomic sites of the RA and LA. The fibrillatory activity observed in ECG lead V1 correlated primarily with the activity of the anterolateral RA free wall and thus may be useful for detecting the AF source if it is close to that area.


Astrobiological stoichiometry.
Chemical composition affects virtually all aspects of astrobiology, from stellar astrophysics to molecular biology. We present a synopsis of the research results presented at the "Stellar Stoichiometry" Workshop Without Walls hosted at Arizona State University April 11-12, 2013, under the auspices of the NASA Astrobiology Institute. The results focus on the measurement of chemical abundances and the effects of composition on processes from stellar to planetary scales. Of particular interest were the scientific connections between processes in these normally disparate fields. Measuring the abundances of elements in stars and giant and terrestrial planets poses substantial difficulties in technique and interpretation. One of the motivations for this conference was the fact that determinations of the abundance of a given element in a single star by different groups can differ by more than their quoted errors. The problems affecting the reliability of abundance estimations and their inherent limitations are discussed. When these problems are taken into consideration, self-consistent surveys of stellar abundances show that there is still substantial variation (factors of ∼ 2) in the ratios of common elements (e.g., C, O, Na, Al, Mg, Si, Ca) important in rock-forming minerals, atmospheres, and biology. We consider how abundance variations arise through injection of supernova nucleosynthesis products into star-forming material and through photoevaporation of protoplanetary disks. The effects of composition on stellar evolution are substantial, and coupled with planetary atmosphere models can result in predicted habitable zone extents that vary by many tens of percent. Variations in the bulk composition of planets can affect rates of radiogenic heating and substantially change the mineralogy of planetary interiors, affecting properties such as convection and energy transport.


Neuroinflammation in Alzheimer's disease.
Increasing evidence suggests that Alzheimer's disease pathogenesis is not restricted to the neuronal compartment, but includes strong interactions with immunological mechanisms in the brain. Misfolded and aggregated proteins bind to pattern recognition receptors on microglia and astroglia, and trigger an innate immune response characterised by release of inflammatory mediators, which contribute to disease progression and severity. Genome-wide analysis suggests that several genes that increase the risk for sporadic Alzheimer's disease encode factors that regulate glial clearance of misfolded proteins and the inflammatory reaction. External factors, including systemic inflammation and obesity, are likely to interfere with immunological processes of the brain and further promote disease progression. Modulation of risk factors and targeting of these immune mechanisms could lead to future therapeutic or preventive strategies for Alzheimer's disease.


Polygenic association between attention-deficit/hyperactivity disorder liability and cognitive impairments.
A recent genome-wide association study (GWAS) identified 12 independent loci significantly associated with attention-deficit/hyperactivity disorder (ADHD). Polygenic risk scores (PRS), derived from the GWAS, can be used to assess genetic overlap between ADHD and other traits. Using ADHD samples from several international sites, we derived PRS for ADHD from the recent GWAS to test whether genetic variants that contribute to ADHD also influence two cognitive functions that show strong association with ADHD: attention regulation and response inhibition, captured by reaction time variability (RTV) and commission errors (CE). The discovery GWAS included 19 099 ADHD cases and 34 194 control participants. The combined target sample included 845 people with ADHD (age: 8-40 years). RTV and CE were available from reaction time and response inhibition tasks. ADHD PRS were calculated from the GWAS using a leave-one-study-out approach. Regression analyses were run to investigate whether ADHD PRS were associated with CE and RTV. Results across sites were combined via random effect meta-analyses. When combining the studies in meta-analyses, results were significant for RTV (R2 = 0.011, β = 0.088, p = 0.02) but not for CE (R2 = 0.011, β = 0.013, p = 0.732). No significant association was found between ADHD PRS and RTV or CE in any sample individually (p > 0.10). We detected a significant association between PRS for ADHD and RTV (but not CE) in individuals with ADHD, suggesting that common genetic risk variants for ADHD influence attention regulation.


Which intervention design factors influence performance of community health workers in low- and middle-income countries? A systematic review.
Community health workers (CHWs) are increasingly recognized as an integral component of the health workforce needed to achieve public health goals in low- and middle-income countries (LMICs). Many factors influence CHW performance. A systematic review was conducted to identify intervention design related factors influencing performance of CHWs. We systematically searched six databases for quantitative and qualitative studies that included CHWs working in promotional, preventive or curative primary health services in LMICs. One hundred and forty studies met the inclusion criteria, were quality assessed and double read to extract data relevant to the design of CHW programmes. A preliminary framework containing factors influencing CHW performance and characteristics of CHW performance (such as motivation and competencies) guided the literature search and review.A mix of financial and non-financial incentives, predictable for the CHWs, was found to be an effective strategy to enhance performance, especially of those CHWs with multiple tasks. Performance-based financial incentives sometimes resulted in neglect of unpaid tasks. Intervention designs which involved frequent supervision and continuous training led to better CHW performance in certain settings. Supervision and training were often mentioned as facilitating factors, but few studies tested which approach worked best or how these were best implemented. Embedment of CHWs in community and health systems was found to diminish workload and increase CHW credibility. Clearly defined CHW roles and introduction of clear processes for communication among different levels of the health system could strengthen CHW performance.When designing community-based health programmes, factors that increased CHW performance in comparable settings should be taken into account. Additional intervention research to develop a better evidence base for the most effective training and supervision mechanisms and qualitative research to inform policymakers in development of CHW interventions are needed.


Forensic electrochemistry: the electroanalytical sensing of synthetic cathinone-derivatives and their accompanying adulterants in "legal high" products.
The production and abuse of new psychoactive substances, known as "legal highs" which mimic traditional drugs of abuse is becoming a global epidemic. Traditional analytical methodologies exist which can provide confirmatory analysis but there is a requirement for an on-the-spot analytical screening tool that could be used to determine whether a substance, or sample matrix contains such legal, or formally "legal highs". In this paper the electrochemical sensing of (±)-methcathinone and related compounds at a range of commercially available electrode substrates is explored. We demonstrate for the first time that this class of "legal highs" are electrochemically active providing a novel sensing protocol based upon their electrochemical oxidation. Screen-printed graphite sensing platforms are favoured due to their proven ability to be mass-produced providing large numbers of reliable and reproducible electrode sensing platforms that preclude the requirement of surface pre-treatment such as mechanical polishing as is the case in the use of solid/re-usable electrode substrates. Additionally they hold potential to be used on-site potentially being the basis of an on-site legal high screening device. Consequently the electroanalytical sensing of (±)-methcathinone (3a), (±)-4′-methylmethcathinone [3b, 4-MMC, (±)-mephedrone] and (±)-4′-methyl-N-ethylcathinone (3c, 4-MEC) is explored using screen-printed sensing platforms with the effect of pH explored upon the analytical response with their analytical efficiency evaluated towards the target legal highs. Interesting at pH values below 6 the voltammetric response quantitatively changes from that of an electrochemically irreversible response to that of a quasi-reversible signature which can be used analytically. It is demonstrated for the first time that the electroanalytical sensing of (±)-methcathinone (3a), (±)-mephedrone (3b) and 4-MEC (3c) are possible with accessible linear ranges found to correspond to 16–200 μg mL(−1) for 3a (at pH 12) and 16–350 μg mL(−1) for both 3b and 3c in pH 2, with limits of detection (3σ) found to correspond to 44.5, 39.8 and 84.2 μg mL(−1) respectively. Additionally adulterants that are commonly incorporated into cathinone legal highs are electrochemically explored at both pH 2 and 12.


Improved out-of-hospital cardiac arrest survival after the sequential implementation of 2005 AHA guidelines for compressions, ventilations, and induced hypothermia: the Wake County experience.
We assess survival from out-of-hospital cardiac arrest after community-wide implementation of 2005 American Heart Association guidelines. This was an observational multiphase before-after cohort in an urban/suburban community (population 840,000) with existing advanced life support. Included were all adults treated for cardiac arrest by emergency responders. Excluded were patients younger than 16 years and trauma patients. Intervention phases in months were baseline 16; phase 1, new cardiopulmonary resuscitation 12; phase 2, impedance threshold device 6; and phase 3, full implementation including out-of-hospital-induced hypothermia 12. Primary outcome was survival to discharge. Other survival and neurologic outcomes were compared between study phases, and adjusted odds ratios with 95% confidence intervals (CIs) for survival by phase were determined by multivariate regression. One thousand three hundred sixty-five cardiac arrest patients were eligible for inclusion: baseline n=425, phase 1 n=369, phase 2 n=161, phase 3 n=410. Across phases, patients had similar demographic, clinical, and emergency medical services characteristics. Overall and witnessed ventricular fibrillation and ventricular tachycardia survival improved throughout the study phases: respectively, baseline 4.2% and 13.8%, phase 1 7.3% and 23.9%, phase 2 8.1% and 34.6%, and phase 3 11.5% and 40.8%. The absolute increase for overall survival from baseline to full implementation was 7.3% (95% CI 3.7% to 10.9%); witnessed ventricular fibrillation/ventricular tachycardia survival was 27.0% (95% CI 13.6% to 40.4%), representing an additional 25 lives saved annually in this community. In the context of a community-wide focus on resuscitation, the sequential implementation of 2005 American Heart Association guidelines for compressions, ventilations, and induced hypothermia significantly improved survival after cardiac arrest. Further study is required to clarify the relative contribution of each intervention to improved survival outcomes.


A FAK scaffold inhibitor disrupts FAK and VEGFR-3 signaling and blocks melanoma growth by targeting both tumor and endothelial cells.
Melanoma has the highest mortality rate of all skin cancers and a major cause of treatment failure is drug resistance. Tumors heterogeneity requires novel therapeutic strategies and new drugs targeting multiple pathways. One of the new approaches is targeting the scaffolding function of tumor related proteins such as focal adhesion kinase (FAK). FAK is overexpressed in most solid tumors and is involved in multiple protein-protein interactions critical for tumor cell survival, tumor neovascularization, progression and metastasis. In this study, we investigated the anticancer activity of the FAK scaffold inhibitor C4, targeted to the FAK-VEGFR-3 complex, against melanomas. We compared C4 inhibitory effects in BRAF mutant vs BRAF wild type melanomas. C4 effectively caused melanoma tumor regression in vivo, when administered alone and sensitized tumors to chemotherapy. The most dramatic effect of C4 was related to reduction of vasculature of both BRAF wild type and V600E mutant xenograft tumors. The in vivo effects of C4 were assessed in xenograft models using non-invasive multimodality imaging in conjunction with histologic and molecular biology methods. C4 inhibited cell viability, adhesion and motility of melanoma and endothelial cells, specifically blocked phosphorylation of VEGFR-3 and FAK and disrupted their complexes. Specificity of in vivo effects for C4 were confirmed by a decrease in tumor FAK and VEGFR-3 phosphorylation, reduction of vasculogenesis and reduced blood flow. Our collective observations provide evidence that a small molecule inhibitor targeted to the FAK protein-protein interaction site successfully inhibits melanoma growth through dual targeting of tumor and endothelial cells and is effective against both BRAF wild type and mutant melanomas.


Structural basis of lipid-driven conformational transitions in the KvAP voltage-sensing domain.
Voltage-gated ion channels respond to transmembrane electric fields through reorientations of the positively charged S4 helix within the voltage-sensing domain (VSD). Despite a wealth of structural and functional data, the details of this conformational change remain controversial. Recent electrophysiological evidence showed that equilibrium between the resting ('down') and activated ('up') conformations of the KvAP VSD from Aeropyrum pernix can be biased through reconstitution in lipids with or without phosphate groups. We investigated the structural transition between these functional states, using site-directed spin-labeling and EPR spectroscopic methods. Solvent accessibility and interhelical distance determinations suggest that KvAP gates through S4 movements involving an ∼3-Å upward tilt and simultaneous ∼2-Å axial shift. This motion leads to large accessibly changes in the intracellular water-filled crevice and supports a new model of gating that combines structural rearrangements and electric-field remodeling.


Intraslab earthquakes: dehydration of the Cascadia slab.
We simultaneously invert travel times of refracted and wide-angle reflected waves for three-dimensional compressional-wave velocity structure, earthquake locations, and reflector geometry in northwest Washington state. The reflector, interpreted to be the crust-mantle boundary (Moho) of the subducting Juan de Fuca plate, separates intraslab earthquakes into two groups, permitting a new understanding of the origins of intraslab earthquakes in Cascadia. Earthquakes up-dip of the Moho's 45-kilometer depth contour occur below the reflector, in the subducted oceanic mantle, consistent with serpentinite dehydration; earthquakes located down-dip occur primarily within the subducted crust, consistent with the basalt-to-eclogite transformation.


Thermal optimality of net ecosystem exchange of carbon dioxide and underlying mechanisms.
• It is well established that individual organisms can acclimate and adapt to temperature to optimize their functioning. However, thermal optimization of ecosystems, as an assemblage of organisms, has not been examined at broad spatial and temporal scales. • Here, we compiled data from 169 globally distributed sites of eddy covariance and quantified the temperature response functions of net ecosystem exchange (NEE), an ecosystem-level property, to determine whether NEE shows thermal optimality and to explore the underlying mechanisms. • We found that the temperature response of NEE followed a peak curve, with the optimum temperature (corresponding to the maximum magnitude of NEE) being positively correlated with annual mean temperature over years and across sites. Shifts of the optimum temperature of NEE were mostly a result of temperature acclimation of gross primary productivity (upward shift of optimum temperature) rather than changes in the temperature sensitivity of ecosystem respiration. • Ecosystem-level thermal optimality is a newly revealed ecosystem property, presumably reflecting associated evolutionary adaptation of organisms within ecosystems, and has the potential to significantly regulate ecosystem-climate change feedbacks. The thermal optimality of NEE has implications for understanding fundamental properties of ecosystems in changing environments and benchmarking global models.


IKBKE Is Required during KRAS-Induced Pancreatic Tumorigenesis.
Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest malignancies lacking effective therapeutic strategies. Here, we show that the noncanonical IκB-related kinase, IKBKE, is a critical oncogenic effector during KRAS-induced pancreatic transformation. Loss of IKBKE inhibits the initiation and progression of pancreatic tumors in mice carrying pancreatic-specific KRAS activation. Mechanistically, we demonstrate that this protumoral effect of IKBKE involves the activation of GLI1 and AKT signaling and is independent of the levels of activity of the NF-κB pathway. Further analysis reveals that IKBKE regulates GLI1 nuclear translocation and promotes the reactivation of AKT post-inhibition of mTOR in PDAC cells. Interestingly, combined inhibition of IKBKE and mTOR synergistically blocks pancreatic tumor growth. Together, our findings highlight the functional importance of IKBKE in pancreatic cancer, support the evaluation of IKBKE as a therapeutic target in PDAC, and suggest IKBKE inhibition as a strategy to improve efficacy of mTOR inhibitors in the clinic. Cancer Res; 77(2); 320-9. ©2017 AACR.


Quality of life among perinatally HIV-affected and HIV-unaffected school-aged and adolescent Ugandan children: a multi-dimensional assessment of wellbeing in the post-HAART era.
To examine quality of life (QOL) in perinatally HIV-infected (PHIV) or HIV-exposed uninfected (PHEU) vs. healthy HIV-unexposed uninfected (HUU) children during school-age/adolescence. PHIV infection was diagnosed via DNA PCR. Current HIV status was confirmed by HIV rapid diagnostic test. Three HIV groups were defined: PHIV, PHEU, and HUU. QOL was assessed with proxy and self-report versions of the PedsQL™ 4.0 instrument at 6-18 years of age. QOL scores ranged from zero (least QOL) to 100 (highest QOL) in the following dimensions: combined QOL inventory (CQOLI), multi-dimensional vigor (MDV), general wellbeing (GWB), present functioning, and general cognitive functioning (CF). Multivariable linear regression models estimated HIV-related percent differences (β) in QOL scores and 95% confidence intervals (CI). Compared to HUU CQOLI deficits ranged from 6.5 to 9.2% (95% CI -15.4, -1.6), GWB deficit ranged from 6.5 to 10.5% (95% CI -16.0, -1.3), MDV deficit ranged from 6.8 to 11.6% (95% CI -14.5, 0.9), and CF deficit ranged from 9.7 to 13.1% for PHIV children. QOL deficits of similar magnitude and direction in most domains were observed for PHIV compared to PHEU. However, self-reported indicators of GWB (β = -3.5; 95% CI -9.0, 2.0) and present functioning (β = 4.0; 95% CI -4.6, 12.5) were similar for PHIV compared to PHEU. QOL scores were generally similar for PHEU compared to HUU. PHEU and HUU had similar QOL profile but PHIV predicted sustained deficits in multiple QOL domains. PHIV and PHEU children were similar with respect to general wellbeing and present functioning. Psychosocial and scholastic interventions in combination with HIV care are likely to improve QOL in PHIV.


The dystrotelin, dystrophin and dystrobrevin superfamily: new paralogues and old isoforms.
Dystrophins and dystrobrevins are distantly related proteins with important but poorly understood roles in the function of metazoan muscular and neuronal tissues. Defects in them and their associated proteins cause a range of neuromuscular disorders. Members of this superfamily have been discovered in a relatively serendipitous way; we set out to compile a comprehensive description of dystrophin- and dystrobrevin-related sequences from available metazoan genome sequences, validated in representative organisms by RT-PCR, or acquired de novo from key species. Features of the superfamily revealed by our survey include: a) Dystrotelin, an entirely novel branch of the superfamily, present in most vertebrates examined. Dystrotelin is expressed in the central nervous system, and is a possible orthologue of Drosophila DAH. We describe the preliminary characterisation of its function, evolution and expression. b) A novel vertebrate member of the dystrobrevin family, gamma-dystrobrevin, an ancient branch now extant only in fish, but probably present in our own ancestors. Like dystrophin, zebrafish gamma-dystrobrevin mRNA is localised to myosepta. c) The extent of conservation of alternative splicing and alternative promoter use in the dystrophin and dystrobrevin genes; alternative splicing of dystrophin exons 73 and 78 and alpha-dystrobrevin exon 13 are conserved across vertebrates, as are the use of the Dp116, Dp71 and G-utrophin promoters; the Dp260 and Dp140 promoters are tetrapod innovations. d) The evolution of the unique N-terminus of DRP2 and its relationship to Dp116 and G-utrophin. e) A C-terminally truncated common ancestor of dystrophin and utrophin in cyclostomes. f) A severely restricted repertoire of dystrophin complex components in ascidians. We have refined our understanding of the evolutionary history and isoform diversity of the five previously reported vertebrate superfamily members and describe two novel members, dystrotelin and gamma-dystrobrevin. Dystrotelins, dystrophins and dystrobrevins are roughly equally related to each other. Vertebrates therefore have a repertoire of seven superfamily members (three dystrophins, three dystrobevins, and one dystrotelin), with one lost in tetrapods. Most invertebrates studied have one member from each branch. Although the basic shared function which is implied by the common architecture of these distantly related proteins remains unclear, it clearly permeates metazoan biology.


Molecular characterization of a phloem-specific gene encoding the filament protein, phloem protein 1 (PP1), from Cucurbita maxima.
Sieve elements in the phloem of most angiosperms contain proteinaceous filaments and aggregates called P-protein. In the genus Cucurbita, these filaments are composed of two major proteins: PP1, the phloem filament protein, and PP2, the phloem lactin. The gene encoding the phloem filament protein in pumpkin (Cucurbita maxima Duch.) has been isolated and characterized. Nucleotide sequence analysis of the reconstructed gene gPP1 revealed a continuous 2430 bp protein coding sequence, with no introns, encoding an 809 amino acid polypeptide. The deduced polypeptide had characteristics of PP1 and contained a 15 amino acid sequence determined by N-terminal peptide sequence analysis of PP1. The sequence of PP1 was highly repetitive with four 200 amino acid sequence domains containing structural motifs in common with cysteine proteinase inhibitors. Expression of the PP1 gene was detected in roots, hypocotyls, cotyledons, stems, and leaves of pumpkin plants. PP1 and its mRNA accumulated in pumpkin hypocotyls during the period of rapid hypocotyl elongation after which mRNA levels declined, while protein levels remained elevated. PP1 was immunolocalized in slime plugs and P-protein bodies in sieve elements of the phloem. Occasionally, PP1 was detected in companion cells. PP1 mRNA was localized by in situ hybridization in companion cells at early stages of vascular differentiation. The developmental accumulation and localization of PP1 and its mRNA paralleled the phloem lactin, further suggesting an interaction between these phloem-specific proteins.


Impact of climate change on three-dimensional dynamic critical load functions.
Changes in climate and atmospheric deposition of base cations can alter the ionic composition of soil and surface waters, and therefore affect the structure and function of sensitive ecosystems. However, these drivers are not generally explicitly considered in the calculation of critical loads or dynamic critical loads to evaluate the recovery of ecosystems from elevated acidic deposition. Here we explore the importance of accounting for these changes in calculating dynamic critical loads for ecosystems. We developed three-dimensional dynamic critical load surfaces as a function of nitrate, sulfur, and base cation deposition under current and future climate change scenarios for the Hubbard Brook Experimental Forest, New Hampshire. This case study indicates that dynamic critical loads for nitrate and sulfur will be lower under conditions of potential climate change or decreases in base cation deposition. This analysis suggests that greater emission controls may be needed to protect sensitive forest ecosystems from acidic deposition under a future climate change or conditions of lower atmospheric deposition of base cations, particularly for watersheds experiencing elevated leaching losses of nitrate. This study should facilitate more informed policy decisions on emission control strategies and assessments of ecosystem recovery.


Matrix metalloproteinase-7-dependent release of tumor necrosis factor-alpha in a model of herniated disc resorption.
Herniated disc (HD), one of the major causes of low back pain, is often resolved spontaneously without surgical intervention. Resorption is associated with a marked increase in infiltrating macrophages, and the matrix metalloproteinases (MMP) MMP-3 and MMP-7 have been implicated in this phenomenon. We developed a murine organ culture model in which intact intervertebral discs were cocultured with peritoneal macrophages to investigate the role of MMPs in HD resorption. Using macrophages isolated from MMP-null mice, we report that macrophage-produced MMP-7 was required for proteoglycan degradation, loss of wet weight, and macrophage infiltration of cocultured discs. The inability of MMP-7-deficient macrophages to infiltrate discs could not be attributed to a defect in macrophage migration. MMP-7 was required for the release of the cytokine TNF-alpha from peritoneal macrophages. The generation of soluble TNF-alpha was essential for the induction of MMP-3 in disc cocultures, which in turn is required for the generation of a macrophage chemoattractant and subsequent macrophage infiltration. TNF-alpha release from macrophages was necessary but insufficient for disc resorption, which required macrophage infiltration. We conclude that there is extensive communication between macrophages and chondrocytes in HD resorption and that an essential component of this communication is the requirement for MMPs to release soluble bioactive factors.


The 15-Country Collaborative Study of Cancer Risk Among Radiation Workers in the Nuclear Industry: design, epidemiological methods and descriptive results.
Radiation protection standards are based mainly on risk estimates from studies of atomic bomb survivors in Japan. The validity of extrapolations from the relatively high-dose acute exposures in this population to the low-dose, protracted or fractionated environmental and occupational exposures of primary public health concern has long been the subject of controversy. A collaborative retrospective cohort study was conducted to provide direct estimates of cancer risk after low-dose protracted exposures. The study included nearly 600,000 workers employed in 154 facilities in 15 countries. This paper describes the design, methods and results of descriptive analyses of the study. The main analyses included 407,391 nuclear industry workers employed for at least 1 year in a participating facility who were monitored individually for external radiation exposure and whose doses resulted predominantly from exposure to higher-energy photon radiation. The total duration of follow-up was 5,192,710 person-years. There were 24,158 deaths from all causes, including 6,734 deaths from cancer. The total collective dose was 7,892 Sv. The overall average cumulative recorded dose was 19.4 mSv. A strong healthy worker effect was observed in most countries. This study provides the largest body of direct evidence to date on the effects of low-dose protracted exposures to external photon radiation.


Lipid-based Transfection Reagents Exhibit Cryo-induced Increase in Transfection Efficiency.
The advantages of lipid-based transfection reagents have permitted their widespread use in molecular biology and gene therapy. This study outlines the effect of cryo-manipulation of a cationic lipid-based formulation, Lipofectamine 2000, which, after being frozen and thawed, showed orders of magnitude higher plasmid delivery efficiency throughout eight different cell lines, without compromising cell viability. Increased transfection efficiency with the freeze-thawed reagent was also seen with 2'-O-methyl phosphorothioate oligonucleotide delivery and in a splice-correction assay. Most importantly, a log-scale improvement in gene delivery using the freeze-thawed reagent was seen in vivo. Using three different methods, we detected considerable differences in the polydispersity of the different nucleic acid complexes as well as observed a clear difference in their surface spreading and sedimentation, with the freeze-thawed ones displaying substantially higher rate of dispersion and deposition on the glass surface. This hitherto overlooked elevated potency of the freeze-thawed reagent facilitates the targeting of hard-to-transfect cells, accomplishes higher transfection rates, and decreases the overall amount of reagent needed for delivery. Additionally, as we also saw a slight increase in plasmid delivery using other freeze-thawed transfection reagents, we postulate that freeze-thawing might prove to be useful for an even wider variety of transfection reagents.


Effect of Phenylephrine on the Accommodative System.
Accommodation is controlled by the action of the ciliary muscle and mediated primarily by parasympathetic input through postganglionic fibers that originate from neurons in the ciliary and pterygopalatine ganglia. During accommodation the pupil constricts to increase the depth of focus of the eye and improve retinal image quality. Researchers have traditionally faced the challenge of measuring the accommodative properties of the eye through a small pupil and thus have relied on pharmacological agents to dilate the pupil. Achieving pupil dilation (mydriasis) without affecting the accommodative ability of the eye (cycloplegia) could be useful in many clinical and research contexts. Phenylephrine hydrochloride (PHCl) is a sympathomimetic agent that is used clinically to dilate the pupil. Nevertheless, first investigations suggested some loss of functional accommodation in the human eye after PHCl instillation. Subsequent studies, based on different measurement procedures, obtained contradictory conclusions, causing therefore an unexpected controversy that has been spread almost to the present days. This manuscript reviews and summarizes the main research studies that have been performed to analyze the effect of PHCl on the accommodative system and provides clear conclusions that could help clinicians know the real effects of PHCl on the accommodative system of the human eye.


Identification and developmental expression of a novel low molecular weight neuronal intermediate filament protein expressed in Xenopus laevis.
Xenopus laevis is a valuable model system for the study of vertebrate neuroembryogenesis. However, very few well-characterized nervous system-specific molecular markers are available for studies in this organism. We screened a X. laevis adult brain cDNA library using a cDNA probe for mouse low molecular weight neurofilament protein (NF-L) in order to identify neuron-specific intermediate filament proteins. Clones for two distinct neuron-specific intermediate filament proteins were isolated and sequenced. One of these encoded for a Xenopus NF-L (XNF-L) and the other for a novel neuron-specific Xenopus intermediate filament protein (XNIF) that was present earlier and more abundantly than XNF-L during development. XNIF contained a central rod domain with multiple sequence features characteristic of IF proteins. The XNF-L was very similar to mouse NF-L, with a 77% sequence identity in the rod domain and the presence of a polyglutamic acid region in the tail domain, characteristic of type IV neurofilament proteins. In contrast, XNIF showed only 60% identity to mouse NF-L in the rod domain and lacked the glutamic acid-rich sequence in the tail domain. XNIF also had a very low (approximately 38%) sequence identity in the head and tail domains as compared to NF-L and other neurofilament proteins (45% identity to the head domain of alpha-internexin). In the adult frog, XNIF mRNA is detected by Northern blots only within the nervous system and by in situ hybridization histochemistry exclusively in neurons, particularly in the medullary reticular system and spinal cord. Antisera raised against the unique tail region of XNIF detected a single distinct 60 kDa band in Western blots of nervous system cytoskeletal preparations, and this XNIF immunoreactivity was concentrated in axons in the PNS and in small perikarya in the dorsal root ganglion. In contrast, NF-L immunoreactivity was principally in the large perikarya in the dorsal root ganglion. In development, XNIF mRNA appears more abundant than XNF-L mRNA in all premetamorphic stages examined. XNIF mRNA is first detectable at stage 24 (26 hr), whereas stable expression of XNF-L is at stage 35/36 (50 hr). XNIF immunoreactivity is detectable within the cement gland, within many neuronal cell bodies and axon tracts within the developing nervous system, and within all cellular layers of the developing retina. The availability of these two distinct neuron-specific intermediate filament proteins, with different temporal and spatial expression patterns, should provide new markers as well as targets for functional perturbation in the developing X. laevis nervous system.


MBT/Pas mouse: a relevant model for the evaluation of Rift Valley fever vaccines.
Currently, there are no worldwide licensed vaccines for Rift Valley fever (RVF) that are both safe and effective. Development and evaluation of vaccines, diagnostics and treatments depend on the availability of appropriate animal models. Animal models are also necessary to understand the basic pathobiology of infection. Here, we report the use of an inbred MBT/Pas mouse model that consistently reproduces RVF disease and serves our purpose for testing the efficacy of vaccine candidates; an attenuated Rift Valley fever virus (RVFV) and a recombinant RVFV-capripoxvirus. We show that this model is relevant for vaccine testing.


Mindfulness-based Cognitive Therapy (MBCT) in bipolar disorder: preliminary evaluation of immediate effects on between-episode functioning.
Bipolar disorder is highly recurrent and rates of comorbidity are high. Studies have pointed to anxiety comorbidity as one factor associated with risk of suicide attempts and poor overall outcome. This study aimed to explore the feasibility and potential benefits of a new psychological treatment (Mindfulness-based Cognitive Therapy: MBCT) for people with bipolar disorder focusing on between-episode anxiety and depressive symptoms. The study used data from a pilot randomized trial of MBCT for people with bipolar disorder in remission, focusing on between-episode anxiety and depressive symptoms. Immediate effects of MBCT versus waitlist on levels of anxiety and depression were compared between unipolar and bipolar participants. The results suggest that MBCT led to improved immediate outcomes in terms of anxiety which were specific to the bipolar group. Both bipolar and unipolar participants allocated to MBCT showed reductions in residual depressive symptoms relative to those allocated to the waitlist condition. Analyses were based on a small sample, limiting power. Additionally the study recruited participants with suicidal ideation or behaviour so the findings cannot immediately be generalized to individuals without these symptoms. The study, although preliminary, suggests an immediate effect of MBCT on anxiety and depressive symptoms among bipolar participants with suicidal ideation or behaviour, and indicates that further research into the use of MBCT with bipolar patients may be warranted.


Lidocaine accelerates neuroelectrical recovery after incomplete global ischemia in rabbits.
The use of high-dose lidocaine for cerebral protection during ischemia has produced varied results. Our study uses a new, single carotid artery preparation in the rabbit to produce incomplete global ischemia by graded carotid occlusion; specific electroencephalographic changes are used as the end point for the extent of blood flow reduction sustained during 20 minutes. We monitored arterial pressure, intracranial pressure, and internal carotid blood flow that were recorded with an electromagnetic flowmeter after surgical ligation of the opposite internal and the two vertebral arteries, and we studied the electroencephalogram and somatosensory-evoked potentials elicited by stimulation of the sciatic nerve. Low-dose lidocaine (0.2 mg/kg/min) infused throughout the experiment significantly accelerated the time course of the return of electroencephalographic and evoked-potential amplitudes toward control. Deep halothane anesthesia alone elicited the slowest recovery, suggesting that the action of lidocaine was independent of its general anesthetic effect. There were very small differences among the groups in the measured arterial pressure, intracranial pressure, and cerebral blood flow, suggesting that lidocaine changed recovery rate without markedly modifying any characteristic of the postischemic cerebral perfusion. The protective effect of lidocaine may be the result of a specific blockade of Na+ channels or a decrease in excitatory neurotransmitter release, either of which would cause a delay in the onset of the events that lead to neuronal damage during ischemia.


Association of Liver Injury From Specific Drugs, or Groups of Drugs, With Polymorphisms in HLA and Other Genes in a Genome-Wide Association Study.
We performed a genome-wide association study (GWAS) to identify genetic risk factors for drug-induced liver injury (DILI) from licensed drugs without previously reported genetic risk factors. We performed a GWAS of 862 persons with DILI and 10,588 population-matched controls. The first set of cases was recruited before May 2009 in Europe (n = 137) and the United States (n = 274). The second set of cases were identified from May 2009 through May 2013 from international collaborative studies performed in Europe, the United States, and South America. For the GWAS, we included only cases with patients of European ancestry associated with a particular drug (but not flucloxacillin or amoxicillin-clavulanate). We used DNA samples from all subjects to analyze HLA genes and single nucleotide polymorphisms. After the discovery analysis was concluded, we validated our findings using data from 283 European patients with diagnosis of DILI associated with various drugs. We associated DILI with rs114577328 (a proxy for A*33:01 a HLA class I allele; odds ratio [OR], 2.7; 95% confidence interval [CI], 1.9-3.8; P = 2.4 × 10-8) and with rs72631567 on chromosome 2 (OR, 2.0; 95% CI, 1.6-2.5; P = 9.7 × 10-9). The association with A*33:01 was mediated by large effects for terbinafine-, fenofibrate-, and ticlopidine-related DILI. The variant on chromosome 2 was associated with DILI from a variety of drugs. Further phenotypic analysis indicated that the association between DILI and A*33:01 was significant genome wide for cholestatic and mixed DILI, but not for hepatocellular DILI; the polymorphism on chromosome 2 was associated with cholestatic and mixed DILI as well as hepatocellular DILI. We identified an association between rs28521457 (within the lipopolysaccharide-responsive vesicle trafficking, beach and anchor containing gene) and only hepatocellular DILI (OR, 2.1; 95% CI, 1.6-2.7; P = 4.8 × 10-9). We did not associate any specific drug classes with genetic polymorphisms, except for statin-associated DILI, which was associated with rs116561224 on chromosome 18 (OR, 5.4; 95% CI, 3.0-9.5; P = 7.1 × 10-9). We validated the association between A*33:01 terbinafine- and sertraline-induced DILI. We could not validate the association between DILI and rs72631567, rs28521457, or rs116561224. In a GWAS of persons of European descent with DILI, we associated HLA-A*33:01 with DILI due to terbinafine and possibly fenofibrate and ticlopidine. We identified polymorphisms that appear to be associated with DILI from statins, as well as 2 non-drug-specific risk factors.


Prostate-specific antigen for prostate cancer staging in a population-based register.
Previous studies have shown a relationship between serum prostate-specific antigen (PSA) level and prostate tumour volume. Reports based on selected case series have also indicated that serum PSA may be used for staging, although a varying prevalence of metastasizing tumours complicates the interpretation of these studies. In order to determine the accuracy of the serum level of PSA in predicting the presence of metastases we performed a prospective cohort study of a geographically defined population of men with prostate cancer. Serum level of PSA and the results of investigations for regional lymph node and distant metastases were recorded for all 8328 men with prostate cancer registered in the Swedish National Prostate Cancer Register 1996-1997. The prevalence of lymph node metastases among men who had undergone lymph node exploration was 4%, 16% and 33% for well, moderately and poorly differentiated tumours. The corresponding prevalence of distant metastases was 12%, 30% and 48%. With serum PSA <20 ng/ml as a cut-off point the negative likelihood ratios for well and moderately differentiated tumours were found to be 0.47 and 0.45 for lymph node metastases and 0.24 and 0.18 for distant metastases, resulting in post-test probabilities >92% for the exclusion of metastases. In men with poorly differentiated tumours, the negative likelihood ratio would need to be even lower to safely exclude disseminated disease. For well to moderately differentiated tumours, further investigations to assess the presence of metastases may be omitted with no great risk for understaging if serum PSA <20 ng/ml.


Tocopherols and tocotrienols as anticancer treatment for lung cancer: future nutrition.
Nutrition has been known for ages to shield the immune system against several formulations that deregulate normal DNA repair mechanisms, and induce tumorigenesis. Vitamins and in specific Vit E and its members tocopherols (α-, β-, γ-, δ-) and tocotrienols (α-, β-, γ-, δ-) have demonstrated strong association with the prevention of cancer and inhibition of tumor, both in vitro and in vivo. Vitamin E has also demonstrated effective role against chemotherapy resistant cancer cell evolution and a protective role in acute interstitial disease. Several formulations of Vitamin E have been investigated conjugated with different carriers as nano-formulations and administered in different forms. Additionally, several tumorigenic pathways have been investigated separately in an effort to identify which member of Vitamin E inhibits efficiently every pathway. Vitamin E presented efficiency against specific subhistology types of lung cancer. Finally, in the current work up to date information regarding novel formulations with Vitamin E and inhibition pathways are going to be presented and commented.


Do vaccination interventions have effects? A study on how poultry vaccination interventions change smallholder farmer knowledge, attitudes, and practice in villages in Kenya and Tanzania.
Poultry are important for many poor households in developing countries, but there are many constraints to poultry production, including disease. One of the most important diseases of chickens is Newcastle disease (ND). Even though there are effective vaccines against this disease available in most countries, uptake by small-scale poultry keepers is often low. In this study, two areas in Kenya and Tanzania were studied, where some villages had received additional support to get vaccination and other villages had not. In Kenya, 320 households from 10 villages were interviewed, of which half of the villages had active promotion of vaccination through village-based advisors. In Tanzania, 457 households were interviewed, of which 241 came from villages that have had active support through either a project or government extension services. Knowledge about vaccines and the attitudes towards vaccinating against ND was evaluated using mixed multivariable logistic models. Results indicate that in Kenya, the most important determinants for understanding the function of a vaccine were having had support in the village and to have knowledge about ND signs, while in Tanzania gender and previous vaccine use were important in addition to having had support. Attitudes towards vaccination were mainly determined by knowledge, where more knowledge about how vaccines work in general or about ND contributed to more positive attitudes. Among Kenyan farmers that had never used the vaccine before, the amount of birds they lost to disease and predators also influenced attitudes. In conclusion, this study supports the notion that knowledge is a very important component of extension support and that simply making vaccines available may not be sufficient for high levels of uptake.


Understanding charge transfer and recombination by interface engineering for improving the efficiency of PbS quantum dot solar cells.
In quantum dot heterojunction solar cells (QDHSCs), the QD active layer absorbs sunlight and then transfers the photogenerated electrons to an electron-transport layer (ETL). It is generally believed that the conduction band minimum (CBM) of the ETL should be lower than that of the QDs to enable efficient charge transfer from the QDs to the collection electrode (here, FTO) through the ETL. However, by employing Mg-doped ZnO (Zn1-xMgxO) as a model ETL in PbS QDHSCs, we found that an ETL with a lower CBM is not necessary to realize efficient charge transfer in QDHSCs. The existence of shallow defect states in the Zn1-xMgxO ETL can serve as additional charge-transfer pathways. In addition, the conduction band offset (CBO) between the ETL and the QD absorber has been, for the first time, revealed to significantly affect interfacial recombination in QDHSCs. We demonstrate that a spike in the band structure at the ETL/QD interface is useful for suppressing interfacial recombination and improving the open-circuit voltage. By varying the Mg doping level in ZnO, we were able to tune the CBM, defect distribution and carrier concentration in the ETL, which play key roles in charge transfer and recombination and therefore the device performance. PbS QDHSCs based on the optimized Zn1-xMgxO ETL exhibited a high power conversion efficiency of 10.6%. Our findings provide important guidance for enhancing the photovoltaic performance of QD-based solar cells.


The burden of disease related to indoor air in the Netherlands: do different methods lead to different results?
Dutch policy makers needed a knowledge base for prioritising control of different indoor air pollutants. Several burden of disease (BoD) estimates were available, but it was not known if they could be applied to The Netherlands. To quantify the BoD related to indoor air in The Netherlands, and to compare the outcomes with a previous study (EnVIE), which used a different BoD methodology. The largest BoD was attributable to environmental tobacco smoke. The next most important indoor air pollutants were radon and thoron from soils and building materials, followed by dampness and carbon monoxide (CO). Formaldehyde exposure did not contribute to the total BoD, according to our estimates. The EnVIE estimate was three times higher, and the most important indoor air pollutants by BoD were combustion products from outdoor sources, bioaerosols due to dampness and by outdoor sources, volatile organic compounds, radon from soils, pathogens and CO. The differences in estimates were primarily caused by the different selection or definition of substances in indoor air, rather than the differing BoD methodology. Indoor air exposure is associated with a considerable BoD in The Netherlands; approximately 1500 healthy life years per 1 million inhabitants are lost due to 1 year of exposure. The results from the different BoD studies examined here are difficult to compare--even the relative order of most important pollutants depends on choices and assumptions in the assessment. A careful evaluation of BoD estimates is required before they can be used in policy making.


Magnetic resonance anatomy of the proximal metacarpal region of the horse described from images acquired from low- and high-field magnets.
While low-field magnetic resonance (MR) images can provide useful information in the investigation of proximal metacarpal region pain, an in-depth knowledge of anatomy and comparison with more detailed high-field images are essential to understand the meaning of different signal intensities within tissues. This anatomic description is based on low-field and high-field MR examination of 30 cadaver metacarpal regions of mature horses with no history of carpal or proximal metacarpal pain. Normal MR anatomy is described and is illustrated by high-field and low-field MR images in transverse, sagittal and dorsal planes. Normal anatomic variations of soft tissue and osseous structures are discussed. Differences between the signal intensity and definition of tissues on high-field and low-field MR images and in different pulse sequences are highlighted. Several structures could be evaluated in both high-field and low-field images that cannot easily be imaged using radiography and ultrasonography, including the abaxial margins of the suspensory ligament, the interosseous ligaments between the metacarpal bones and the carpometacarpal ligaments. Structures that have previously not been described in detail were also identified.


Shaping smoking cessation in hard-to-treat smokers.
Contingency management (CM) effectively treats addictions by providing abstinence incentives. However, CM fails for many who do not readily become abstinent and earn incentives. Shaping may improve outcomes in these hard-to-treat (HTT) individuals. Shaping sets intermediate criteria for incentive delivery between the present behavior and total abstinence. This should result in HTT individuals having improving, rather than poor, outcomes. We examined whether shaping improved outcomes in HTT smokers (never abstinent during a 10-visit baseline). Smokers were stratified into HTT (n = 96) and easier-to-treat (ETT [abstinent at least once during baseline]; n = 50) and randomly assigned to either CM or CM with shaping (CMS). CM provided incentives for breath carbon monoxide (CO) levels <4 ppm (approximately 1 day of abstinence). CMS shaped abstinence by providing incentives for COs lower than the 7th lowest of the participant's last 9 samples or <4 ppm. Interventions lasted for 60 successive weekday visits. Cluster analysis identified 4 groups of participants: stable successes, improving, deteriorating, and poor outcomes. In comparison with ETT, HTT participants were more likely to belong to 1 of the 2 unsuccessful clusters (odds ratio [OR] = 8.1, 95% CI [3.1, 21]). This difference was greater with CM (OR = 42, 95% CI [5.9, 307]) than with CMS, in which the difference between HTT and ETT participants was not significant. Assignment to CMS predicted membership in the improving (p = .002) as compared with the poor outcomes cluster. Shaping can increase CM's effectiveness for HTT smokers.


MDMA, methamphetamine, and CYP2D6 pharmacogenetics: what is clinically relevant?
In vitro human studies show that the metabolism of most amphetamine-like psychostimulants is regulated by the polymorphic cytochrome P450 isozyme CYP2D6. Two compounds, methamphetamine and 3,4-methylenedioxymethamphetamine (MDMA), were selected as archetypes to discuss the translation and clinical significance of in vitro to in vivo findings. Both compounds were chosen based on their differential interaction with CYP2D6 and their high abuse prevalence in society. Methamphetamine behaves as both a weak substrate and competitive inhibitor of CYP2D6, while MDMA acts as a high affinity substrate and potent mechanism-based inhibitor (MBI) of the enzyme. The MBI behavior of MDMA on CYP2D6 implies that subjects, irrespective of their genotype/phenotype, are phenocopied to the poor metabolizer (PM) phenotype. The fraction of metabolic clearance regulated by CYP2D6 for both drugs is substantially lower than expected from in vitro studies. Other isoenzymes of cytochrome P450 and a relevant contribution of renal excretion play a part in their clearance. These facts tune down the potential contribution of CYP2D6 polymorphism in the clinical outcomes of both substances. Globally, the clinical relevance of CYP2D6 polymorphism is lower than that predicted by in vitro studies.


Single nucleotide polymorphism in sugar pathway and disease resistance genes in sugarcane.
Single nucleotide polymorphism in sugar pathway and disease resistance genes showing genetic association with sugar content and red rot resistance would be useful in marker-assisted genetic improvement of sugarcane. Validation and genotyping of potential sequence variants in candidate genes are necessary to understand their functional significance and trait association potential. We discovered, characterized, validated and genotyped SNPs and InDels in sugar pathway and disease resistance genes of Saccharum complex and sugarcane varieties using amplicon sequencing and CAPS assays. The SNPs were abundant in the non-coding 3'UTRs than 5'UTRs and coding sequences depicting a strong bias toward C to T transition substitutions than transversions. Sequencing of cloned amplicons validated 61.6 and 45.2 % SNPs detected in silico in 21 sugar pathway and 16 disease resistance genes, respectively. Sixteen SNPs in four sugar pathway genes and 10 SNPs in nine disease resistance genes were validated through cost-effective CAPS assay. Functional and adaptive significance of SNP and protein haplotypes identified in sugar pathway and disease resistance genes was assessed by correlating their allelic variation with missense amino acid substitutions in the functional domains, alteration in protein structure models and possible modulation of catalytic enzyme activity in contrasting high and low sugar and moderately red rot resistant and highly susceptible sugarcane genotypes. A strong genetic association of five SNPs in the sugar pathway and disease resistance genes, and an InDel marker in the promoter sequence of sucrose synthase-2 gene, with sugar content and red rot resistance, was evident. The functionally relevant SNPs and InDels, detected and validated in sugar pathway and disease resistance genes, and genic CAPS markers designed, would be of immense use in marker-assisted genetic improvement of sugarcane for sugar content and disease resistance.


Barriers to Alzheimer disease drug discovery and development in academia.
The drug discovery and the drug development processes represent a continuum of recursive activities that range from initial drug target identification to final Food and Drug Administration approval and marketing of a new therapeutic. Drug discovery, as its name implies, is more exploratory and less focused in many cases, whereas drug development has a clinically defined endpoint and a specific disease goal. Academia has historically made major contributions to this process at the early discovery phases. However, current trends in the organization of the pharmaceutical industry suggest an expanded role for academia in the near future. Megamergers among major pharmaceutical corporations indicate their movement toward a focus on end-stage clinical trials, manufacturing, and marketing. There has been a parallel increase in outsourcing of intermediate steps to specialty small pharmaceutical, biotechnology, and contract service companies. The new paradigm suggests that academia will play an increasingly important role at the proof-of-principle stage of basic and clinical drug discovery research, in training the future skilled work force, and in close partnerships with small pharmaceutical and biotechnology companies. However, academic drug discovery research faces a set of barriers to progress, the relative importance of which varies with the home institution and the details of the research area. These barriers fall into four general categories: (1) the historical administrative structure and environment of academia; (2) the structure and emphasis of peer review panels that control research funding by government and private agencies; (3) the organization and operation of the academic infrastructure; and (4) the structure and availability of specialized resources and information management. Selected examples of barriers to drug discovery and drug development research and training in academia are presented, as are some specific recommendations designed to minimize or circumvent these barriers. In some cases, precedents established by other disease-focused areas may be relevant to Alzheimer disease and related disorders, but the overall impact of any changes requires adaptation at the top of the administrative structures in academia and funding agencies to support and encourage cooperative efforts among faculty investigators.


Direct pathway cloning of the sodorifen biosynthetic gene cluster and recombinant generation of its product in E. coli.
Serratia plymuthica WS3236 was selected for whole genome sequencing based on preliminary genetic and chemical screening indicating the presence of multiple natural product pathways. This led to the identification of a putative sodorifen biosynthetic gene cluster (BGC). The natural product sodorifen is a volatile organic compound (VOC) with an unusual polymethylated hydrocarbon bicyclic structure (C16H26) produced by selected strains of S. plymuthica. The BGC encoding sodorifen consists of four genes, two of which (sodA, sodB) are homologs of genes encoding enzymes of the non-mevalonate pathway and are thought to enhance the amounts of available farnesyl pyrophosphate (FPP), the precursor of sodorifen. Proceeding from FPP, only two enzymes are necessary to produce sodorifen: an S-adenosyl methionine dependent methyltransferase (SodC) with additional cyclisation activity and a terpene-cyclase (SodD). Previous analysis of S. plymuthica found sodorifen production titers are generally low and vary significantly among different producer strains. This precludes studies on the still elusive biological function of this structurally and biosynthetically fascinating bacterial terpene. Sequencing and mining of the S. plymuthica WS3236 genome revealed the presence of 38 BGCs according to antiSMASH analysis, including a putative sodorifen BGC. Further genome mining for sodorifen and sodorifen-like BGCs throughout bacteria was performed using SodC and SodD as queries and identified a total of 28 sod-like gene clusters. Using direct pathway cloning (DiPaC) we intercepted the 4.6 kb candidate sodorifen BGC from S. plymuthica WS3236 (sodA-D) and transformed it into Escherichia coli BL21. Heterologous expression under the control of the tetracycline inducible PtetO promoter firmly linked this BGC to sodorifen production. By utilizing this newly established expression system, we increased the production yields by approximately 26-fold when compared to the native producer. In addition, sodorifen was easily isolated in high purity by simple head-space sampling. Genome mining of all available genomes within the NCBI and JGI IMG databases led to the identification of a wealth of sod-like pathways which may be responsible for producing a range of structurally unknown sodorifen analogs. Introduction of the S. plymuthica WS3236 sodorifen BGC into the fast-growing heterologous expression host E. coli with a very low VOC background led to a significant increase in both sodorifen product yield and purity compared to the native producer. By providing a reliable, high-level production system, this study sets the stage for future investigations of the biological role and function of sodorifen and for functionally unlocking the bioinformatically identified putative sod-like pathways.


Direct photoaffinity labeling of Kir6.2 by [gamma-(32)P]ATP-[gamma]4-azidoanilide.
ATP-sensitive potassium (K(ATP)) channels are under complex regulation by intracellular ATP and ADP. The potentiatory effect of MgADP is conferred by the sulfonylurea receptor subunit of the channel, SUR, whereas the inhibitory effect of ATP appears to be mediated via the pore-forming subunit, Kir6.2. We have previously reported that Kir6.2 can be directly labeled by 8-azido-[gamma-(32)P]ATP. However, the binding affinity of 8-azido-ATP to Kir6.2 was low probably due to modification at 8' position of adenine. Here we demonstrate that Kir6.2 can be directly photoaffinity labeled with higher affinity by [gamma-(32)P]ATP-[gamma]4-azidoanilide ([gamma-(32)P]ATP-AA), containing an unmodified adenine ring. Photoaffinity labeling of Kir6.2 by [gamma-(32)P]ATP-AA is not affected by the presence of Mg(2+), consistent with Mg(2+)-independent ATP inhibition of K(ATP) channels. Interestingly, SUR1, which can be strongly and specifically photoaffinity labeled by 8-azido-ATP, was not photoaffinity labeled by ATP-AA. These results identify key differences in the structure of the nucleotide binding sites on SUR1 and Kir6.2.


Antimicrobial residues, non-typhoidal Salmonella, Vibrio spp. and associated microbiological hazards in retail shrimps purchased in Ho Chi Minh city (Vietnam).
We investigated antimicrobial residues, non-typhoidal Salmonella (NTS), Vibrio spp. and their associated antimicrobial resistance (AMR), in shrimps locally purchased in Ho Chi Minh City (Vietnam). In addition, we investigated the relationship between AMR in NTS, Vibrio spp. and antimicrobial residue in the same sample. A total of 40 samples of shrimp heads/shells from different retail sources was cultured using ISO 6579-1:2017 (NTS) and ISO/TS 21872-1:2007 (Vibrio spp.). Phenotypic antimicrobial susceptibility was investigated using Vitek (NTS, 34 antimicrobials) and disk diffusion (Vibrio spp., 12 antimicrobials). A total of 9 (22.5%) samples contained antimicrobial residue, including tetracyclines, fluoroquinolones, sulfonamides and macrolides (in 7.5%, 7.5%, 2.5% and 2.5% of samples, respectively). Shrimp samples from supermarkets had a higher prevalence of antimicrobial residue than those purchased in street markets (50% vs. 13.3%) (p = 0.049). A total of 30 (75%) samples were contaminated with NTS. All samples contained Vibrio spp., with V. parahaemolyticus being most common (87.5% samples). A total of 58.9% NTS isolates were multidrug resistant. With regards to the highest priority, critically important antimicrobials, the highest resistance corresponded to quinolones (14.4-47.8%), followed by 3rd and 4th generation cephalosporins (3.3-7.8%). Vibrio spp. isolates were characterised by their high resistance against ampicillin (82.7%) and 3rd generation cephalosporins (8.3-16.5%). Extended Spectrum Beta-Lactamase (ESBL) activity was detected in 28.1% V. parahaemolyticus isolates. Half of ESBL-positive V. parahaemolyticus strains harboured bla CTX-M1. We found an association between the presence of residues and the number of resistances for NTS (p = 0.075) and Vibrio spp. isolates (p = 0.093) from the same sample. These findings suggest that the presence of residues may contribute to the selection of AMR in foodborne pathogens in shrimps. Authorities should strengthen policies aiming at restricting inappropriate antimicrobial usage in shrimp farming, and step up monitoring of antimicrobial residues and food-borne pathogens at retail in Vietnam.


Capillary electrophoresis assay for ubiquitin carboxyl-terminal hydrolases with chemically synthesized ubiquitin-valine as substrate.
Ubiquitin is expressed in eukaryotic cells as precursors, fused via its carboxyl terminus either to other ubiquitin sequences in linear polyubiquitin arrays or to specific ribosomal proteins. In some of the polyubiquitin fusions a single amino acid (e.g., valine in humans) is attached to the carboxyl terminus. These gene products are rapidly (probably cotranslationally) cleaved by ubiquitin carboxyl-terminal hydrolase (UCH) enzymes; therefore, although ubiquitin precursors are suitable substrates for assays of UCH activity, they are difficult to isolate from nucleated cells. While the recombinant approach allows the production of ubiquitin precursors in prokaryotic cells (which do not contain the ubiquitin system), proteins produced in this manner require purification and may also be susceptible to modification by bacterial enzymes, e.g., adventitious proteolysis. As an alternative we have chemically synthesized human ubiquitin-valine. In the assay described here the cleavage of ubiquitin-valine to ubiquitin (77 and 76 residue proteins, respectively) by a purified recombinant Drosophila UCH was monitored by capillary electrophoresis. Mass spectrometry verified the precise cleavage of ubiquitin-valine, confirming that this synthetic protein is a UCH substrate. Synthetic ubiquitin-valine may serve as a generic substrate for UCHs allowing the purification and identification of new members of this enzyme family.


Encoding of olfactory information with oscillating neural assemblies.
In the brain, fast oscillations of local field potentials, which are thought to arise from the coherent and rhythmic activity of large numbers of neurons, were observed first in the olfactory system and have since been described in many neocortical areas. The importance of these oscillations in information coding, however, is controversial. Here, local field potential and intracellular recordings were obtained from the antennal lobe and mushroom body of the locust Schistocerca americana. Different odors evoked coherent oscillations in different, but usually overlapping, ensembles of neurons. The phase of firing of individual neurons relative to the population was not dependent on the odor. The components of a coherently oscillating ensemble of neurons changed over the duration of a single exposure to an odor. It is thus proposed that odors are encoded by specific but dynamic assemblies of coherently oscillating neurons. Such distributed and temporal representation of complex sensory signals may facilitate combinatorial coding and associative learning in these, and possibly other, sensory networks.


UPLC-MS-Based Metabolomics Profiling for α-Glucosidase Inhibiting Property of Parkia speciosa Pods.
Parkia speciosa is a food plant that grows indigenously in Southeast Asia. A great deal of interest has been paid to this plant due to its traditional uses in the treatment of several diseases. The pods contain many beneficial secondary metabolites with potential applications in medicine and cosmetics. However, studies on their phytochemical properties are still lacking. Therefore, the present study was undertaken to profile the bioactive compounds of P. speciosa pods collected from six different regions of Malaysia through ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) and α-glucosidase inhibitory potential. This study applied metabolomics to elucidate the differences between P. speciosa populations found naturally in the different locations and to characterize potential α-glucosidase inhibitors from P. speciosa pods. P. speciosa collected from different regions of Malaysia showed good α-glucosidase inhibitory activity, with a median inhibitory concentration (IC50) of 0.45-0.76 μg/mL. The samples from the northern and northeastern parts of Peninsular Malaysia showed the highest activity. Using UHPLC-QTOF-MS/MS analysis, 25 metabolites were identified in the pods of P. speciosa. The findings unveiled that the pods of P. speciosa collected from different locations exhibit different levels of α-glucosidase inhibitory activity. The pods are a natural source of potent antidiabetic bioactive compounds.


Pain Symptomatology and Management in Pediatric Ehlers-Danlos Syndrome: A Review.
Ehlers-Danlos syndromes (EDS) are a group of connective tissue disorders that manifest with hyperextensibility of joints and skin, and general tissue fragility. While not a major criterion for clinical diagnosis, pain is a frequently endorsed symptom across subtypes of EDS. As such, the present review aims to summarize research to date on pain characteristics and management, and the relationship between such pain symptomatology and quality of life in pediatric EDS. Characteristics of pain, including theorized etiology, relative intensity and extent of pain are described, as well as descriptions of frequently endorsed pain sites (musculoskeletal, and non-musculoskeletal). Interventions related to the management of musculoskeletal (e.g., pharmaceutical intervention, physical therapy) and non-musculoskeletal pain (e.g., pharmaceutical and psychological interventions) are discussed, highlighting the need for additional research related to pediatric pain management in the context of hypermobility syndromes. In addition, the relationship between pain in pediatric EDS and quality of life is described. Finally, limitations of literature to date are described and recommendations for future lines of research are outlined.


Evolutionary Changes in Sensitivity to Hormonally Induced Gonadal Sex Reversal in a Frog Species.
The Japanese frog Glandirana rugosa is unique in that it shows geographic variation in sex chromosome differentiation and heterogametic sex determination. To elucidate the cause of interpopulation differences in gonadal sex differentiation, we investigated hormonally induced sex reversal and the expression patterns of genes associated with sex determination during early tadpole development. We found that sex reversal was easily induced in XX females and XY males of 2 forms (West-Japan and East-Japan) of G. rugosa with the ancestral homomorphic sex chromosomes under male heterogametic sex determination. During sex reversal, expression of CYP19 and/or FOXL2 was dependent on the phenotypic sex of the gonad. In contrast, sex reversal was not induced in ZW females of a population with a heteromorphic ZW sex chromosome system or in XX females or XY males in a population with a heteromorphic XY sex chromosome system. The latter 2 populations are evolutionarily derived forms. These results indicate an evolutionary direction for the gonadal sex differentiation mechanism. The original system was highly sensitive to sex hormones and allowed almost complete sex reversal. From this ancestral form, a new system evolved that was resistant to hormones and showed a change in the heterogametic sex and the sex chromosome differentiation mechanism.


Imprecision in medical communication: study of a doctor talking to patients with serious illness.
Uncertainty is believed to be a central feature in illness experiences. Conversations between a consultant hematologist and 61 seriously ill patients were transcribed, entered on a database and scrutinized for patterns of language uncertainty by linguistic concordancing analysis. Transcripts were then discussed in detail with the hematologist, and techniques of protocol analysis were used to gain insight into his thought processes during consultations. The main findings were that the doctor used many more expressions of uncertainty than did patients: that evaluative terms were widely used to reassure rather than to worry patients; and that patients and doctor together used certain key terms ambiguously, in a manner which allowed the doctor to feel that facts were not misrepresented while perhaps permitting the patient to feel reassured.


Axl is a prognostic marker in oral squamous cell carcinoma.
Overexpression of the receptor tyrosine kinase Axl is implicated in several diseases. The present study was conducted to determine the biologic and clinical significance of Axl in oral squamous cell carcinoma (OSCC). The expression of Axl was examined in a panel of OSCC cell lines. Activation of Axl by Gas6 treatment and silencing of Axl via Axl shRNA were used to examine the effect of Axl on OSCC cell line. Expression of Axl in cancer tissues were examined by immunohistochemical staining. The associations between Axl expression and clinicopathologic features and prognosis were analyzed. Varied Axl expression was noted in OSCC cell lines. Compared with control cells, modulated Axl signal affected epithelial-mesenchymal gene expression and cell invasion and migration. The immunoreactivity of Axl was low in normal epithelium, and a progressively increased positive percentage was noted, from normal/hyperplastic epithelium (10.9%) to dysplasia (30.8%) to cancer tissue (54.5%). Axl expression correlated with lymph node status (P = .001) and clinical stage (P = .014) of OSCC. Patients with high expression of Axl showed poor prognosis compared with those with low Axl expression patients (P < .001). In multivariate prognostic analysis according to the Cox proportional hazard regression model, Axl expression remained as an independent prognostic factor (P = .037; CI, 1.042-3.839). Our data indicated that Axl signal promotes OSCC carcinogenesis and progression. The expression of Axl is a valuable marker for OSCC aggressiveness and clinical outcome.


Assay of steroids by liquid chromatography-tandem mass spectrometry in monitoring 21-hydroxylase deficiency.
Immunoassays of steroid hormones are still used in the diagnosis and monitoring of patients with congenital adrenal hyperplasia. However, cross-reactivity between steroids can give rise to falsely elevated steroid levels. Here, we compare the use of immunoassays and liquid chromatography-tandem mass spectrometry (LC-MS/MS) in the monitoring of patients with classic 21-hydroxylase deficiency (21OHD). Steroid profiles in different mutation groups (genotypes) were also compared. Fifty-five patients with classic 21OHD (38 women) were studied. Blood samples were collected in the morning after an overnight medication fast. LC-MS/MS and immunoassays were employed to assay 17-hydroxyprogesterone (17OHP), testosterone and androstenedione. In addition, 21-deoxycortisol (21DF), 11-deoxycortisol (11DF), corticosterone, deoxycorticosterone, cortisone and cortisol were analyzed by LC-MS/MS. Testosterone, androstenedione and 17OHP levels were consistently lower (by about 30-50%) when measured by LC-MS/MS compared with immunoassays, with exception of testosterone in men. There was a significant correlation between 21DF and 17OHP (r = 0.87, P < 0.001), but three patients had undetectable 21DF. Subjects with no enzyme activity had significantly lower mean 11DF concentrations than subjects with residual activity. The use of LC-MS/MS gives a more specific view of adrenal steroid levels in 21OHD compared with immunoassays, which seem to considerably overestimate the levels of 17OHP and androstenedione. Falsely elevated levels of 17OHP and androstenedione could lead to overtreatment with glucocorticoids.


The elusive silica/water interface: isolated silanols under water as revealed by vibrational sum frequency spectroscopy.
It has been long recognized that the surface chemistry of silica, and in particular the type and relative amount of surface bound silanol groups, plays a critical role in many of the properties associated with the material, where a typical example is the discrepant adsorption behavior observed depending on the pretreatment history of the surface. However, in spite of its importance, the direct probing of specific surface silanol groups under water has been hampered by instrumental limitations. Here we make use of vibrational sum frequency spectroscopy (VSFS) to first, identify under water the OH stretch of isolated surface silanols, and second, explore its acid/base behavior and dependence on the surface pretreatment method. The properties of other types of silanol groups (i.e. hydrogen bonded/geminal) are also inferred from the data. The ability to directly probe these functional groups under water represents a crucial step to further improving our understanding of this widely used mineral oxide.


Identification of homophenylalanine biosynthetic genes from the cyanobacterium Nostoc punctiforme PCC73102 and application to its microbial production by Escherichia coli.
L-Homophenylalanine (L-Hph) is a useful chiral building block for synthesis of several drugs, including angiotensin-converting enzyme inhibitors and the novel proteasome inhibitor carfilzomib. While the chemoenzymatic route of synthesis is fully developed, we investigated microbial production of L-Hph to explore the possibility of a more efficient and sustainable approach to L-Hph production. We hypothesized that L-Hph is synthesized from L-Phe via a mechanism homologous to 3-methyl-2-oxobutanoic acid conversion to 4-methyl-2-oxopentanoic acid during leucine biosynthesis. Based on bioinformatics analysis, we found three putative homophenylalanine biosynthesis genes, hphA (Npun_F2464), hphB (Npun_F2457), and hphCD (Npun_F2458), in the cyanobacterium Nostoc punctiforme PCC73102, located around the gene cluster responsible for anabaenopeptin biosynthesis. We constructed Escherichia coli strains harboring hphABCD-expressing plasmids and achieved the fermentative production of L-Hph from L-Phe. To our knowledge, this is the first identification of the genes responsible for homophenylalanine synthesis in any organism. Furthermore, to improve the low conversion efficiency of the initial strain, we optimized the expression of hphA, hphB, and hphCD, which increased the yield to ∼630 mg/liter. The L-Hph biosynthesis and L-Leu biosynthesis genes from E. coli were also compared. This analysis revealed that HphB has comparatively relaxed substrate specificity and can perform the function of LeuB, but HphA and HphCD show tight substrate specificity and cannot complement the LeuA and LeuC/LeuD functions, and vice versa. Finally, the range of substrate tolerance of the L-Hph-producing strain was examined, which showed that m-fluorophenylalanine, o-fluorophenylalanine, and L-tyrosine were accepted as substrates and that the corresponding homoamino acids were generated.


Diets based on virgin olive oil or fish oil but not on sunflower oil prevent age-related alveolar bone resorption by mitochondrial-related mechanisms.
Aging enhances frequency of chronic diseases like cardiovascular diseases or periodontitis. Here we reproduced an age-dependent model of the periodontium, a fully physiological approach to periodontal conditions, to evaluate the impact of dietary fat type on gingival tissue of young (6 months old) and old (24 months old) rats. Animals were fed life-long on diets based on monounsaturated fatty acids (MUFA) as virgin olive oil, n-6 polyunsaturated fatty acids (n-6PUFA), as sunflower oil, or n-3PUFA, as fish oil. Age-related alveolar bone loss was higher in n-6PUFA fed rats, probably as a consequence of the ablation of the cell capacity to adapt to aging. Gene expression analysis suggests that MUFA or n-3PUFA allowed mitochondria to maintain an adequate turnover through induction of biogenesis, autophagy and the antioxidant systems, and avoiding mitochondrial electron transport system alterations. The main finding is that the enhanced alveolar bone loss associated to age may be targeted by an appropriate dietary treatment. The mechanisms involved in this phenomenon are related with an ablation of the cell capacity to adapt to aging. Thus, MUFA or n-3PUFA might allow mitochondrial maintaining turnover through biogenesis or autophagy. They might also be able to induce the corresponding antioxidant systems to counteract age-related oxidative stress, and do not inhibit mitochondrial electron transport chain. From the nutritional and clinical point of view, it is noteworthy that the potential treatments to attenuate alveolar bone loss (a feature of periodontal disease) associated to age could be similar to some of the proposed for the prevention and treatment of cardiovascular diseases, a group of pathologies recently associated with age-related periodontitis.


Fasting hyperglycaemia following trans-catheter arterial chemo-embolization for hepatocellular carcinoma in cirrhosis.
Following a hyperosmolar diabetic coma in a cirrhotic patient with hepatocellular carcinoma undergoing transcatheter arterial chemo-embolization, we assessed the prevalence, severity, causes and prognostic impact of impaired glucose metabolism following transcatheter arterial chemo-embolization. Plasma glucose, pancreatic and thyroid hormones, cortisol, growth hormone, ACTH and TSH concentrations were determined before and after transcatheter arterial chemo-embolization in 98 patients (70 with a normal fasting glucose, 7 with mild fasting hyperglycaemia and 21 diabetics) undergoing 226 transcatheter arterial chemo-embolization procedures. Child status, body temperature, serum ALT and amylase levels, tumour size, gelfoam embolization and disease aetiology were recorded. Liver function was assessed before and after transcatheter arterial chemo-embolization by measuring monoethylglycinexylidide formation after i.v. lidocaine. A significant rise in glucose levels (p < 0.0001) was observed in 30/98 patients. Hyperglycaemia was more frequent in diabetics (67%) and patients with mild fasting hyperglycaemia (71%). Glucose concentrations doubled in 12 patients; 4 required long-term insulin. Fever, a previously altered carbohydrate metabolism and raised ALT levels were prognostic factors for hyperglycaemia (p < 0.01). Plasma C-peptide, glucose/insulin and glucose/C-peptide ratios, were increased after transcatheter arterial chemo-embolization (p < 0.05). Transcatheter arterial chemo-embolization was followed by a reduction in the monoethylglycinexylidide formation capacity (p < 0.05), particularly in hyperglycaemia patients (p < 0.02). Transcatheter arterial chemo-embolization is frequently followed by a derangement in glucose metabolism which is potentially severe, associated with preceding glucose imbalance, fever and a transient deterioration in liver function.


Refining analyses of copy number variation identifies specific genes associated with developmental delay.
Copy number variants (CNVs) are associated with many neurocognitive disorders; however, these events are typically large, and the underlying causative genes are unclear. We created an expanded CNV morbidity map from 29,085 children with developmental delay in comparison to 19,584 healthy controls, identifying 70 significant CNVs. We resequenced 26 candidate genes in 4,716 additional cases with developmental delay or autism and 2,193 controls. An integrated analysis of CNV and single-nucleotide variant (SNV) data pinpointed 10 genes enriched for putative loss of function. Follow-up of a subset of affected individuals identified new clinical subtypes of pediatric disease and the genes responsible for disease-associated CNVs. These genetic changes include haploinsufficiency of SETBP1 associated with intellectual disability and loss of expressive language and truncations of ZMYND11 in individuals with autism, aggression and complex neuropsychiatric features. This combined CNV and SNV approach facilitates the rapid discovery of new syndromes and genes involved in neuropsychiatric disease despite extensive genetic heterogeneity.


Enrichment of two isoflavone aglycones in black soymilk by Rhizopus oligosporus NTU 5 in a plastic composite support bioreactor.
A plastic composite support (PCS) bioreactor was implemented to evaluate the effects on isoflavone deglycosylation in black soymilk fermented by Rhizopus oligosporus NTU 5. Evaluation for the optimal PCS for mycelia immobilisation was conducted, which led to the significant results that the most mycelium weight (0.237 g per PCS, P < 0.05) is held by an S-type PCS; therefore, it was selected for black soymilk fermentation. It was found that the PCS fermentation system without pH control exhibits better efficiency of isoflavone bioconversion (daidzin to daidzein, and genistin to genistein) than the one with pH control at pH 6.5. As for the long-run fermentation, those without pH control indeed accelerate the isoflavone bioconversion by continuously releasing β-glucosidase into soymilk. Deglycosylation can be completed in 8 to 24 h and sustained for at least 34 days as 26 batches. The non-pH-control fermentation system also exhibits the highest total phenolic content (ranged from 0.147 to 0.340 mg GAE mL(-1) sample) when compared to the pH-controlled and suspended ones. Meanwhile, the black soymilk from the 22nd batch with 8 h fermentation demonstrated the highest DPPH radical scavenging effect (54.7%). A repeated-batch PCS fermentation system was established to accelerate the deglycosylation rate of isoflavone in black soymilk. © 2015 Society of Chemical Industry.


Tribute to P. L. Lutz: putting life on 'pause'--molecular regulation of hypometabolism.
Entry into a hypometabolic state is an important survival strategy for many organisms when challenged by environmental stress, including low oxygen, cold temperatures and lack of food or water. The molecular mechanisms that regulate transitions to and from hypometabolic states, and stabilize long-term viability during dormancy, are proving to be highly conserved across phylogenic lines. A number of these mechanisms were identified and explored using anoxia-tolerant turtles as the model system, particularly from the research contributions made by Dr Peter L. Lutz in his explorations of the mechanisms of neuronal suppression in anoxic brain. Here we review some recent advances in understanding the biochemical mechanisms of metabolic arrest with a focus on ideas such as the strategies used to reorganize metabolic priorities for ATP expenditure, molecular controls that suppress cell functions (e.g. ion pumping, transcription, translation, cell cycle arrest), changes in gene expression that support hypometabolism, and enhancement of defense mechanisms (e.g. antioxidants, chaperone proteins, protease inhibitors) that stabilize macromolecules and promote long-term viability in the hypometabolic state.