Simultaneous determination of anionic intermediates for Bacillus subtilis metabolic pathways by capillary electrophoresis electrospray ionization mass spectrometry.
A method for simultaneous determination of anionic metabolites based on capillary electrophoresis (CE) coupled to electrospray ionization mass spectrometry is described. To prevent current drop by the system, electroosmotic flow (EOF) reversal by using a cationic polymer-coated capillary was indispensable. A mixture containing 32 standards including carboxylic acids, phosphorylated carboxylic acids, phosphorylated saccharides, nucleotides, and nicotinamide and flavin adenine coenzymes of glycolysis and the tricarboxylic acid cycle pathways were separated by CE and selectively detected by a quadrupole mass spectrometer with a sheath-flow electrospray ionization interface. Key to the analysis was EOF reversal using a cationic polymer-coated capillary and an electrolyte system consisting of 50 mM ammonium acetate, pH 9.0. The relative standard deviations of the method were better than 0.4% for migration times and between 0.9% and 5.4% for peak areas. The concentration detection limits for these metabolites were between 0.3 and 6.7 micromol/L with pressure injection of 50 mbar for 30 s (30 nL); i.e., mass detection limits ranged from 9 to 200 fmol, at a signal-to-noise ratio of 3. This method was applied to the comprehensive analysis of metabolic intermediates extracted from Bacillus subtilis, and 27 anionic metabolites could be directly detected and quantified.


Optic disc and choroidal metastasis from sporadic medullary thyroid carcinoma: case report and review of the literature.
Choroidal metastasis is a very rare clinical manifestation of thyroid cancer. Herein, we report on a patient with non-familial medullary thyroid carcinoma (MTC), who presented with choroidal metastasis. A 63-year-old male patient with MTC presented with sudden loss of vision in his right eye for 1 month. The patient had a history of complete thyroidectomy and chemotherapy for disseminated MTC. Ophthalmological examination showed optic disc and choroidal metastases in the right eye and a small choroidal metastasis in the left eye. The patient was scheduled for external irradiation therapy for the ocular area but died 2 months after choroidal metastasis was diagnosed. Choroidal metastasis must be considered in MTC patients who develop loss of vision. This is a very rare clinical situation that generally occurs in the late advanced stages of the disease and carries a poor prognosis for these patients.


Differentiating between Alzheimer Disease Patients and Controls with Phase-difference-enhanced Imaging at 3T: A Feasibility Study.
To test the feasibility of the phase difference enhanced (PADRE) imaging for differentiation between Alzheimer disease (AD) patients and control subjects on 3T MR imaging. Fifteen patients with AD and 10 age-matched control subjects underwent two-dimensional fast field echo imaging to obtain PADRE images on a 3T MR scanner. A double Gaussian distribution model was used to determine the threshold phase value for differentiation between the physiologic and non-physiologic iron in the cerebral cortices, and PADRE images were processed with the threshold. Using a 4-point grading system, two readers independently assessed the signal of the four cerebral cortices on PADRE images: the cuneus, precuneus, superior frontal gyrus, and superior temporal gyrus. The difference in the signals in each cortex between the AD patients and age-matched control subjects was determined by using Mann-Whitney U test. Inter-rater reliability was determined by Kappa analysis. We also evaluated the correlation between Mini-Mental State Examination (MMSE) score and the hypointense grade, and between disease duration and the hypointense grade using the Spearman rank correlation test. The threshold phase value for differentiation between the physiologic and non-physiologic iron was -4.6% π (radian). The mean grades of the cuneus, precuneus, and superior temporal gyrus were significantly higher for the AD patients than for the control subjects (P = 0.002). Excellent inter-rater reliability was seen in the precuneus (kappa = 0.93), superior temporal gyrus (kappa = 0.94), and superior frontal gyrus (kappa = 0.93); good inter-rater reliability was observed in the cuneus (kappa = 0.75). We found a statistical correlation between MMSE score and the hypointense grade in superior temporal gyrus (STG) (P = 0.008), and no correlation between disease duration and the hypointense grade in any gyrus. Our results suggest the feasibility of PADRE imaging at 3T for differentiation between AD patients and control subjects.


Mapping Spatial Genetic Landscapes in Tissue Sections through Microscale Integration of Sampling Methodology into Genomic Workflows.
In cancer research, genomic profiles are often extracted from homogenized macrodissections of tissues, with the histological context lost and a large fraction of material underutilized. Pertinently, the spatial genomic landscape provides critical complementary information in deciphering disease heterogeneity and progression. Microscale sampling methods such as microdissection to obtain such information are often destructive to a sizeable fraction of the biopsy sample, thus showing limited multiplexability and adaptability to different assays. A modular microfluidic technology is here implemented to recover cells at the microscale from tumor tissue sections, with minimal disruption of unsampled areas and tailored to interface with genome profiling workflows, which is directed here toward evaluating intratumoral genomic heterogeneity. The integrated workflow-GeneScape-is used to evaluate heterogeneity in a metastatic mammary carcinoma, showing distinct single nucleotide variants and copy number variations in different tumor tissue regions, suggesting the polyclonal origin of the metastasis as well as development driven by multiple location-specific drivers.


Amplified lipid rafts of malignant cells constitute a target for inhibition of aberrantly active NFAT and melanoma tumor growth by the aminobisphosphonate zoledronic acid.
Nuclear factors of activated T cells (NFAT) are critical modulators of cancer cell growth and survival. However, the mechanisms of their oncogenic dysregulation and strategies for targeting in tumors remain elusive. Here, we report coupling of anti- apoptotic NFAT (NFAT2) activation to cholesterol-enriched lipid raft microdomains of malignant melanoma cells and interruption of this pathway by the aminobisphosphonate zoledronic acid (Zol). The pathway was indicated by capability of Zol to promote apoptosis and to retard in vivo outgrowth of tumorigenic melanoma cell variants through inhibition of permanently active NFAT2. NFAT2 inhibition resulted from disintegration of cholesterol-enriched rafts due to reduction of cellular cholesterol by Zol. Mechanistically, raft disruption abolished raft-localized robust store-operated Ca(2+) (SOC) entry, blocking constitutive activation of protein kinase B/Akt (PKB) and thereby reactivating the NFAT repressor glycogen synthase kinase 3β (GSK3β). Pro-apoptotic inactivation of NFAT2 also followed reactivation of GSK3β by direct inhibition of PKB or SOC, whereas GSK3β blockade prevented Zol-induced NFAT2 inhibition and cell death. The rescuing effect of GSK3β blockade was reproduced by recovery of entire SOC/PKB/GSK3β cascade after reconstitution of rafts by cholesterol replenishment of Zol-treated tumorigenic cells. Remarkably, these malignant cells displayed higher cholesterol and lipid raft content than non-tumorigenic cells, which expressed weak SOC, PKB and NFAT2 activities and resisted raft-ablating action of Zol. Together, the results underscore the functional relevance of amplified melanoma rafts for tumor-promoting NFAT2 signaling and reveal these distinctive microdomains as a target for in vitro and in vivo demise of tumorigenic cells through NFAT2 inhibition by the clinical agent Zol.


Likelihood ratios for modern screening mammography. Risk of breast cancer based on age and mammographic interpretation.
To determine the sensitivity, specificity, and likelihood ratios (LRs) for modern screening mammography by decade of age and mammographic interpretation. Cross-sectional. Nine counties in northern California. A total of 26,057 women aged 30 years and older who underwent a total of 41,747 first and subsequent screening mammographic examinations at the Mobile Mammography Screening Program of the University of California, San Francisco, from April 1985 to September 1991. Breast cancer risk profile, 2 standard mammographic views per breast, and follow-up of abnormal and normal mammograms of contacting women's physicians and by linkage to the regional Surveillance, Epidemiology, and End Results tumor registry. False-negative examinations were normal examinations that occurred within 13 months of a diagnosis of invasive breast cancer or ductal carcinoma in situ. The sensitivity of first screening mammography increased with age: 77.3% for ages 30 to 39 years, 86.7% for ages 40 to 49 years, 93.6% for ages 50 to 59 years, 94.1% for ages 60 to 69 years, and 91.2% for ages 70 years and older (P = .04). Specificity was similar for all ages, ranging from 92.6% to 95.2%. Of all abnormal first screening examinations, 92.9% were reported as "additional evaluation needed." The LRs for that category ranged from 5.2 to 8.8 and did not vary with age. Based on the risk of breast cancer before mammography, which increases with age, the risk of breast cancer after mammography associated with these LRs were 0.01 for ages 30 to 39 years, 0.02 for ages 40 to 49 years, 0.05 for ages 50 to 59 years, 0.07 for ages 60 to 69 years, and 0.07 for ages 70 years and older. The LRs for mammography reported as "suspicious for malignancy" ranged from 88 to 144 and did not vary across age groups. These LRs were associated with a risk of breast cancer about 10 times greater than when mammography was reported as "additional evaluation needed." Most abnormal first screening mammography are interpreted as "additional evaluation needed" and are associated with LRs of about 7. Given this low LR, the risk of breast cancer after mammography is primarily influenced by a woman's age-specific risk of breast cancer before mammography. The LRs for screening mammography interpreted as "suspicious for malignancy" are high (about 124) and are associated with a substantial increase in the risk of breast cancer irrespective of age, but these interpretations comprise only a small proportion of abnormal mammography.


A novel method for implementation of frameless StereoEEG in epilepsy surgery.
Stereoelectroencephalography (SEEG) is an invasive diagnostic procedure in epilepsy surgery that is usually implemented with frame-based methods. To describe a new technique of frameless SEEG and report a prospective case series at a single center. Image integration and planning of electrode trajectories were performed preoperatively on specialized software and exported to a Medtronic S7 StealthStation. Trajectories were implemented by frameless stereotaxy using percutaneous drilling and bolt insertion. Twenty-two patients went this technique, with the insertion of 187 intracerebral electrodes. Of 187 electrodes, 175 accurately reached their neurophysiological target, as measured by postoperative computed tomography reconstruction and multimodal image integration with preoperative magnetic resonance imaging. Four electrodes failed to hit their target due to extradural deflection, and 3 were subsequently resited satisfactorily. Eight electrodes were off target by a mean of 3.6 mm (range, 0.9-6.8 mm) due to a combination of errors in bolt trajectory implementation and bending of the electrode. There was 1 postoperative hemorrhage that was clinically asymptomatic and no postoperative infections. Sixteen patients were offered definitive cortical resections, and 6 patients were excluded from resective surgery. Frameless SEEG is a novel and safe method for implementing SEEG and is easily translated into clinical practice.


The influence of chain elongation on Karplus-type relationships: a DFT study of scalar coupling constants in polyacetylene derivatives.
The coupling constants of a series of acetylenic derivatives have been calculated using the finite perturbation method. In the case of dimethylated derivatives a Karplus-type relationship has been obtained for coupling constants of hydrogen atoms separated up to 15 bonds. Additional relationships have been obtained between the interatomic distances and the coupling constants.


Haemophilus influenzae lobar pneumonia with underlying multiple myeloma: a case report.
Haemophilus influenzae is an uncommon but important cause of lobar pneumonia, specifically in patients whose host defence mechanisms are impaired by unrecognized underlying diseases. A case of H. influenzae lobar pneumonia in a patient with underlying multiple myeloma is presented. The clinical features, treatment and procedures which aid in making the diagnosis are briefly discussed.


Knowledge of attitude toward experience and satisfaction with electroconvulsive therapy in a sample of Iranian patients.
Despite the wide consensus over the safety and efficacy of electroconvulsive therapy (ECT), it still faces negative publicity and unfavorable attitudes of patients and families. Little is known about how the experience with ECT affects the patients' and their families' attitude toward it. The aim of this study was to examine a sample of Iranian patients and their families regarding their experience with ECT and to compare their knowledge and attitude toward ECT before and after this experience and their satisfaction with it. We surveyed 22 patients with major depressive disorder about to undergo ECT and 1 family member of each patient for their knowledge and attitude toward ECT and then surveyed them again after the trial of ECT to compare those variables while assessing their experience and satisfaction with ECT. Patients were rated using the Hamilton Depression Rating Scale and Mini-Mental Status Examination before and after the treatment. We found that, before ECT, family members had a more favorable attitude toward ECT than patients, but after ECT, the patients' attitude changed more positively compared with their families. Both patients and their families had a poor knowledge of ECT before the ECT trial, but their total knowledge increased afterward, although not in the areas of indications and therapeutic effects. The majority of patients and their families found ECT to be beneficial and were satisfied with it. Satisfaction with ECT was independent of treatment outcome. There was a high rate of perceived coercion to consent to ECT. Attention should be paid toward educating patients and their families about the ECT process, indications, risks, safety, and effects as well as informing them about their freedom of choice and right to refuse.


Abutment Material Effect on Peri-implant Soft Tissue Color and Perceived Esthetics.
The purpose of this study was to evaluate the effect of implant abutment material on peri-implant soft tissue color using intraoral spectrophotometric analysis and to compare the clinical outcomes with patient and clinician perception and satisfaction. Thirty patients and four prosthodontic faculty members participated. Abutments were zirconia, gold-hued titanium, and titanium. Peri-implant mucosa color of a single anterior implant restoration was compared to the patient's control tooth. Spectrophotometric analysis using SpectroShadeTM Micro data determined the color difference (ΔE, ΔL*, Δa*, Δb*) between the midfacial peri-implant soft tissue for each abutment material and the marginal gingiva of the control tooth. Color difference values of the abutment groups were compared using ANOVA (α = 0.05). Patient and clinician satisfaction surveys were also conducted using a color-correcting light source. The results of each patient and clinician survey question were compared using chi-square analysis (α = 0.05). Pearson correlation analyses identified the relationship between the total color difference (ΔE) and the patient/clinician perception and satisfaction, as well as between ΔE and tissue thickness. Zirconia abutments displayed significantly smaller spectrophotometric gingival color difference (ΔE) compared to titanium and gold-hued titanium abutments (respectively, 3.98 ± 0.99; 7.22 ± 3.31; 5.65 ± 2.11; p < 0.05). Among ΔL*, Δa*, and Δb*, only Δa* (red-green spectrum) showed significant difference between groups. There was no significant correlation between measured soft tissue thickness and ΔE, but thick gingival phenotype, determined by a probe test, demonstrated a smaller ΔE than thin phenotype (4.82 ± 1.49; 6.41 ± 3.27; p = 0.097). There was no statistical difference in patient or clinician satisfaction among abutment materials, and no correlation between ΔE and the patient and clinician satisfaction. Patient satisfaction was significantly higher than clinician, and patient-perceived differences were lower than clinicians' (p < 0.01). Clinicians' satisfaction was higher for gingival (pink) esthetics than crown (white) esthetics (p < 0.05). Peri-implant mucosa with zirconia abutments demonstrated significantly lower mean color difference compared to titanium or gold-hued titanium abutments as measured spectrophotometrically; however, no statistical difference in patient or clinician perception/satisfaction among abutment materials was demonstrated. Patients were significantly more satisfied than clinicians.


Photoreceptor cells in the vitiligo mouse die by apoptosis. TRPM-2/clusterin expression is increased in the neural retina and in the retinal pigment epithelium.
To determine the mechanism of photoreceptor cell death in the vitiligo mouse, a model of retinal degeneration in which the genetic defect is not retina specific but is instead caused by single point mutation in the microphthalmia (mi) gene that codes for a basic helix-loop-helix DNA transcription factor. Detection of apoptotic cells was performed in fixed retinal tissue using the TUNEL assay in animals 1, 2, 4, 6, 8, 16, 32, 40, and 52 weeks. Electron microscopic analysis was used to confirm the morphologic hallmarks of apoptosis, and Southern blot analysis was used to detect internucleosomal DNA fragmentation. Additionally, the expression of a gene associated with apoptosis, TRPM-2/clusterin, was examined. At ages beyond the time of normal retinal programmed cell death, vitiligo retinas had significantly more TUNEL-positive photoreceptor cells and more photoreceptor cells with condensed chromatin than controls. DNA internucleosomal fragmentation ladders were present in vitiligo retinas even as late as 15 weeks, a time well beyond developmental apoptosis in controls. TRPM-2/clusterin mRNA levels in vitiligo neural retinas were similar to controls initially but were two times greater than controls by 12 weeks. Surprisingly, TRPM-2/clusterin mRNA levels were elevated in the retinal pigment epithelium in the mutant; the expression at one week was two times greater than normals and remained elevated for many months, even though retinal pigment epithelial cells showed no morphologic evidence of apoptosis. The morphologic and biochemical data suggest that photoreceptor cells die by apoptosis in vitiligo mice. The increased retinal TRPM-2/clusterin mRNA levels may be a direct response to these events. The increased expression of this gene in the retinal pigment epithelium, however, may reflect its role in tissue regression and membrane remodeling. Mechanisms by which the mi gene defect might result in the vitiligo retinopathy are proposed.


The type 2 CCK/gastrin receptor antagonist YF476 acutely prevents NSAID-induced gastric ulceration while increasing iNOS expression.
YF476 differs from the proton pump inhibitor (PPI) esomeprazole in mode of action by antagonizing the type 2 receptor of cholecystokinin/gastrin (CCK-2R). YF476 protection against diclofenac-induced gastric ulcers was compared to esomeprazole and correlated with plasma levels of hormones related to gastric pH (gastrin, ghrelin, and somatostatin), gastric gene expression of these hormones, their receptors, and inducible nitric oxide synthase (iNOS). YF476 or esomeprazole pretreatments were followed by diclofenac. Four hours later, gastric tissue was excised and analyzed for ulcer index. An intragastrically implanted Bravo capsule measured pH for 5 days during YF476 plus pentagastrin treatment. Changes in gene expression were assayed for gastrin, ghrelin, and somatostatin; their receptors; and iNOS. YF476 acutely (within 4 h) protected against diclofenac-induced gastric ulcers equivalent to esomeprazole. Gastric pH recorded during 5 days in the presence of pentagastrin was 1.83 (±0.06). YF476 raised pH to 3.67 (±0.09) and plasma ghrelin, gastrin, and somatostatin increased. YF476 increased gene expression of somatostatin receptor and gastrin, while ghrelin receptor decreased; transcripts coding ghrelin, somatostatin, and CCK-2R remained unchanged. In the presence of diclofenac, esomeprazole increased expression of all these transcripts and that of iNOS, while YF476 yielded only decreased CCK-2R and increased iNOS transcripts. YF476 is a potential new preventative treatment for patients at risk of nonsteroidal antiinflammatory drug (NSAID)-induced ulceration. Gastric gene expressions of ghrelin, gastrin, and somatostatin and their receptors differ between esomeprazole and YF476. Despite these differences and different modes of action to raise gastric pH, both drugs acutely increase iNOS, suggesting iNOS expression parallels pH.


Risks and benefits of tumor necrosis factor-alpha inhibitors in the management of psoriatic arthritis: systematic review and metaanalysis of randomized controlled trials.
To evaluate the efficacy and safety of tumor necrosis factor-alpha (TNF-alpha) inhibitors in the management of psoriatic arthritis (PsA). We searched electronic databases to identify randomized controlled trials (RCT) of adalimumab, etanercept, and infliximab used in patients with PsA. Random effects metaanalysis was undertaken to produce pooled estimates of the relative risk, risk difference, or the weighted mean difference for efficacy and safety outcomes using Stata version 9.0. Six RCT met the inclusion criteria, including 982 patients. All 3 TNF-alpha inhibitors were significantly more effective than placebo on the basis of Psoriatic Arthritis Response Criteria (PsARC) and American College of Rheumatology response criteria ACR20, ACR50, and ACR70 ratings. There were no significant differences between TNF-alpha inhibitors and placebo in the proportions of patients experiencing withdrawal for any reason (RR 0.48, 95% CI 0.20-1.18), or withdrawal due to adverse events (RR 2.14, 95% CI 0.73-6.27), serious adverse events (RR 0.98, 95% CI 0.55-1.77), or upper respiratory tract infections (RR 0.91, 95% CI 0.65-1.28). Pooled rates for injection site reactions were significantly higher for adalimumab and etanercept than for placebo (RR 2.48, 95% CI 1.16-5.29), but there was no significant difference in the proportion of patients experiencing infusion reactions with infliximab (RR 1.03, 95% CI 0.48-2.20) compared against placebo. Indirect analysis did not demonstrate any significant differences between the TNF-alpha inhibitors. TNF-alpha inhibitors are effective treatments for PsA with no important added risks associated with their short-term use. There is still a need for longterm risk-benefit assessment of using these drugs for the management of PsA.


Intramuscular botulinum toxin prior to comprehensive rehabilitation has no added value for improving motor impairments, gait kinematics and goal attainment in walking children with spastic cerebral palsy.
Botulinum toxin (BoNT-A) is widely used in combined treatment for spastic cerebral palsy, but its added value preceding comprehensive rehabilitation for motor impairments, gait, and goal attainment has not been studied. A comparative multi-centre trial, in which two groups underwent comprehensive rehabilitation (i.e. high-intensive functional physiotherapy, and indicated casting/orthoses). One group received intramuscular BoNT-A prior to rehabilitation, and the other group did not receive BoNT-A. Children with spastic cerebral palsy, Gross Motor Function Classification System (GMFCS) levels I-III, age range 4-12 years, indicated for BoNT-A treatment regarding mobility problems. Sixty-five children participated (37 boys), mean age 7.3 years (standard deviation (SD) 2.3, range 4-12 years), equally distributed across GMFCS levels. Forty-one children received BoNT-A+ comprehensive rehabilitation and 24 received comprehensive rehabilitation only. Functional leg muscle strength, passive range of motion, angle of catch, cerebral palsy-related pain, walking speed, kinematic gait parameters, goal attainment, and proxy-reported general functioning were assessed at baseline, primary end-point (12 weeks) and 24-week follow-up. Statistical analyses were performed with linear mixed models. At the primary end-point there were no statistically significant differences in treatment effects between the groups, except for the angle-of-catch of the rectus femoris, which was in favour of comprehensive rehabilitation without BoNT-A (12° difference, 95% confidence interval (95% CI) 2:23, p = 0.025). Results at follow-up were similar. At the group level, treating with BoNT-A prior to comprehensive rehabilitation did not add to the clinical effectiveness of rehabilitation. Thus, BoNT-A prescription and use should be critically reconsidered in this cerebral palsy age- and GMFCS-subgroup.


A comparative study of three cytotoxicity test methods for nanomaterials using sodium lauryl sulfate.
The biocompatibility evaluation of nanomaterials is essential for their medical diagnostic and therapeutic usage, where a cytotoxicity test is the simplest form of biocompatibility evaluation. Three methods have been commonly used in previous studies for the cytotoxicity testing of nanomaterials: trypan blue exclusion, colorimetric assay using water soluble tetrazolium (WST), and imaging under a microscope following calcein AM/ethidium homodimer-1 staining. However, there has yet to be a study to compare each method. Therefore, in this study three methods were compared using the standard reference material of sodium lauryl sulfate (SLS). Each method of the cytotoxicity test was carried out using mouse fibroblasts of L-929 exposed to different concentrations of SLS. Compared to the gold standard trypan blue exclusion test, both colorimetric assay using water soluble tetrazolium (WST) and imaging under microscope with calcein AM/ethidium homodimer-1 staining showed results that were not statistically different. Also, each method exhibited various advantages and disadvantages, which included the need of equipment, time taken for the experiment, and provision of additional information such as cell morphology. Therefore, this study concludes that all three methods of cytotoxicity testing may be valid, though careful consideration will be needed when selecting tests with regard to time, finances, and the amount of information required by the researcher(s).


Overnight Polysomnographic Characteristics and Oxygen Saturation of Healthy Infants, 1 to 18 Months of Age, Born and Residing At High Altitude (2,640 Meters).
Approximately 8% of the world population resides above 1,600 m, with about 10 million people living above 2,500 m in Colombia. However, reference values for polysomnography (PSG) and oxygen saturation (Spo2) of children < 2 years old residing at high altitude are currently unavailable. Healthy infants aged 1 to 18 months born and residing at high altitude (Bogotá: 2,640 m) underwent overnight PSG. Four age groups were defined: group 1, < 45 days; group 2, 3 to 4 months; group 3, 6 to 7 months; and group 4, 10 to 18 months. Of 122 children enrolled, 50 had three consecutive PSG tests and were analyzed as a longitudinal subcohort. A total of 281 PSG tests were performed in 122 infants (56% girls): group 1, 106 PSG tests; group 2, 89 PSG tests; group 3, 61 PSG tests; and group 4, 25 PSG tests. Active sleep diminished and quiet sleep increased with maturation. Apnea-hypopnea indexes (total, central, and obstructive) were highest in group 1 (21.4, 12.4, and 6.8/h total sleep time, respectively) and diminished with age (P < .001). Mean Spo2 during waking and sleep increased with age (P < .001). Nadir Spo2 values during respiratory events were lower in younger infants. Longitudinal assessments of 50 infants confirmed the temporal trends described for the cross-sectional dataset. Healthy infants (≤ 18 months old) born and residing at high altitude show preserved sleep architecture but higher apnea-hypopnea indexes and more prominent desaturation with respiratory events than do those living at low altitude. The current study findings can be used as reference values for infants at high altitude.


Novel inhibitors complexed with glutamate dehydrogenase: allosteric regulation by control of protein dynamics.
Mammalian glutamate dehydrogenase (GDH) is a homohexameric enzyme that catalyzes the reversible oxidative deamination of l-glutamate to 2-oxoglutarate using NAD(P)(+) as coenzyme. Unlike its counterparts from other animal kingdoms, mammalian GDH is regulated by a host of ligands. The recently discovered hyperinsulinism/hyperammonemia disorder showed that the loss of allosteric inhibition of GDH by GTP causes excessive secretion of insulin. Subsequent studies demonstrated that wild-type and hyperinsulinemia/hyperammonemia forms of GDH are inhibited by the green tea polyphenols, epigallocatechin gallate and epicatechin gallate. This was followed by high throughput studies that identified more stable inhibitors, including hexachlorophene, GW5074, and bithionol. Shown here are the structures of GDH complexed with these three compounds. Hexachlorophene forms a ring around the internal cavity in GDH through aromatic stacking interactions between the drug and GDH as well as between the drug molecules themselves. In contrast, GW5074 and bithionol both bind as pairs of stacked compounds at hexameric 2-fold axes between the dimers of subunits. The internal core of GDH contracts when the catalytic cleft closes during enzymatic turnover. None of the drugs cause conformational changes in the contact residues, but all bind to key interfaces involved in this contraction process. Therefore, it seems likely that the drugs inhibit enzymatic turnover by inhibiting this transition. Indeed, this expansion/contraction process may play a major role in the inter-subunit communication and allosteric regulation observed in GDH.


Understanding human genetic variation in the era of high-throughput sequencing.
The EMBO/EMBL symposium 'Human Variation: Cause and Consequence' highlighted advances in understanding the molecular basis of human genetic variation and its myriad implications for biology, human origins and disease. As high-throughput sequencing allows us to define genetic variation and its functional consequences at genome-wide resolution for a large number of people, important questions need to be asked about how to use new technologies to maximize the translational relevance of genetic research for society and the individual patient.


Autobiographical and text recall in the elderly: an investigation of a processing resource deficit.
This study examined different explanations of age-related impairments in recall of details from text and autobiographical events. An interpretation of Central Executive Capacity Deficit was supported and explored further. This suggests that details are more demanding of capacity than main points, and that ability to appropriately integrate details with context is likely to be impaired. An implication was that irrelevant and false information may occur, and this was supported in both autobiographical and text recall. The effects were then examined in relation to various measures of ability. The aim was to determine whether declining capacity (as indicated by "Fluid Intelligence" measures) predicted ability to recall in a detailed manner. The difficulty with details was predicted independently by chronological age and by measures of fluid (e.g. AH4 intelligence test) and the more crystallized verbal ability (Mill Hill vocabulary test). Only a measure of the specificity of autobiographical recall was predicted solely by measures of fluid intelligence. Decreased specificity was not a result of faster decay of memory for details, as there was little difference across the lifespan. The resource deficit appears to affect retrieval and appropriate implementation of detail. It was concluded that lower-ability elderly subjects have decreased Central Executive resources, which leads to poor (often inappropriate) integration of details with central thematic points, but that subjects' verbal ability, which does not decline with age, still has an important part to play.


Medical examiners, coroners, and biologic terrorism: a guidebook for surveillance and case management.
Medical examiners and coroners (ME/Cs) are essential public health partners for terrorism preparedness and response. These medicolegal investigators support both public health and public safety functions and investigate deaths that are sudden, suspicious, violent, unattended, and unexplained. Medicolegal autopsies are essential for making organism-specific diagnoses in deaths caused by biologic terrorism. This report has been created to 1) help public health officials understand the role of ME/Cs in biologic terrorism surveillance and response efforts and 2) provide ME/Cs with the detailed information required to build capacity for biologic terrorism preparedness in a public health context. This report provides background information regarding biologic terrorism, possible biologic agents, and the consequent clinicopathologic diseases, autopsy procedures, and diagnostic tests as well as a description of biosafety risks and standards for autopsy precautions. ME/Cs' vital role in terrorism surveillance requires consistent standards for collecting, analyzing, and disseminating data. Familiarity with the operational, jurisdictional, and evidentiary concerns involving biologic terrorism-related death investigation is critical to both ME/Cs and public health authorities. Managing terrorism-associated fatalities can be expensive and can overwhelm the existing capacity of ME/Cs. This report describes federal resources for funding and reimbursement for ME/C preparedness and response activities and the limited support capacity of the federal Disaster Mortuary Operational Response Team. Standards for communication are critical in responding to any emergency situation. This report, which is a joint collaboration between CDC and the National Association of Medical Examiners (NAME), describes the relationship between ME/Cs and public health departments, emergency management agencies, emergency operations centers, and the Incident Command System.


Delineating the angiogenic gene expression profile before pulmonary vascular remodeling in a lamb model of congenital heart disease.
Disordered angiogenesis is implicated in pulmonary vascular remodeling secondary to congenital heart diseases (CHD). However, the underlying genes are not well delineated. We showed previously that an ovine model of CHD with increased pulmonary blood flow (PBF, Shunt) has an "angiogenesis burst" between 1 and 4 wk of age. Thus we hypothesized that the increased PBF elicited a proangiogenic gene expression profile before onset of vessel growth. To test this we utilized microarray analysis to identify genes that could be responsible for the angiogenic response. Total RNA was isolated from lungs of Shunt and control lambs at 3 days of age and hybridized to Affymetrix gene chips for microarray analyses (n = 8/group). Eighty-nine angiogenesis-related genes were found to be upregulated and 26 angiogenesis-related genes downregulated in Shunt compared with control lungs (cutting at 1.2-fold difference, P < 0.05). We then confirmed upregulation of proangiogenic genes FGF2, Angiopoietin2 (Angpt2), and Birc5 at mRNA and protein levels and upregulation of ccl2 at mRNA level in 3-day Shunt lungs. Furthermore, we found that pulmonary arterial endothelial cells (PAEC) isolated from fetal lambs exhibited increased expression of FGF2, Angpt2, Birc5, and ccl2 and enhanced angiogenesis when exposed to elevated shear stress (35 dyn/cm²) compared with cells exposed to more physiological shear stress (20 dyn/cm²). Finally, we demonstrated that blocking FGF2, Angpt2, Birc5, or ccl2 signaling with neutralizing antibodies or small interfering RNA (siRNA) significantly decreased the angiogenic response induced by shear stress. In conclusion, we have identified a "proangiogenic" gene expression profile in a lamb model of CHD with increased PBF that precedes onset of pulmonary vascular remodeling. Our data indicate that FGF2, Angpt2, Birc5, and ccl2 may play important roles in the angiogenic response.


Parasite infection rather than tactile stimulation is the proximate cause of cleaning behaviour in reef fish.
Cleaning behaviour is a popular example of non-kin cooperation. However, quantitative support for this is generally sparse and the alternative, that cleaners are parasitic, has also been proposed. Although the behaviour involves some of the most complex and highly developed interspecific communication signals known, the proximate causal factors for why clients seek cleaners are controversial. However, this information is essential to understanding the evolution of cleaning. I tested whether clients seek cleaners in response to parasite infection or whether clients seek cleaners for tactile stimulation regardless of parasite load. Parasite loads on client fish were manipulated and clients exposed to cleaner fish and control fish behind glass. I found that parasitized client fish spent more time than unparasitized fish next to a cleaner fish. In addition, parasitized clients spent more time next to cleaners than next to control fish, whereas unparasitized fish were not attracted to cleaners. This study shows, I believe for the first time, which is somewhat surprising, that parasite infection alone causes clients to seek cleaning by cleaners and provides insight into how this behaviour evolved.


Unsedated Office-Based Thulium Laser Therapy in Patients With Vocal Fold Cysts.
The objective is to report 3 cases of vocal fold cysts treated with thulium laser in an office setting under local anesthesia. The surgical technique, which consists of surgical marsupialization, is described in detail with emphasis on the laser setting. All 3 patients had complete regression of the lesion and marked improvement in voice quality and good closure of the vocal folds during phonation. The authors review the literature on the application of laser therapy in an office setting and advocate this as an alternative treatment option for patients with vocal fold cysts who are at high risk for general anesthesia or unwilling to undergo the conventional microsurgery.


The Interdisciplinary Rehabilitation Care Team and the Role of Physical Therapy in Survivor Exercise.
Rehabilitation professionals offer expertise in functional assessment, treatment of impairments and functional limitations, and disability prevention. To optimize recovery, and often prior to participating in community-based exercise programming, survivors may need rehabilitation services from a range of healthcare professionals, including physiatrists, nurses, nutritionists, psychologists, and speech, occupational, and physical therapists. Survivors with physical impairments and functional limitations may benefit from interdisciplinary rehabilitation and physical therapy, including tailored therapeutic exercise interventions. A literature review was conducted using the key words cancer survivor, cancer rehabilitation, impairment, fatigue, lymphedema, chemotherapy-induced peripheral neuropathy, and exercise. MEDLINE®, EMBASE, Cochrane Database of Systematic Reviews, and CINAHL® databases were searched. Nurses play a critical role in identifying survivors whose function or fitness is compromised to the point where participation in community-based exercise programming would be inappropriate or unsafe. The interdisciplinary rehabilitation care team can help facilitate the survivor's transition to community-based exercise programming.


A comprehensive comparison of general RNA-RNA interaction prediction methods.
RNA-RNA interactions are fast emerging as a major functional component in many newly discovered non-coding RNAs. Basepairing is believed to be a major contributor to the stability of these intermolecular interactions, much like intramolecular basepairs formed in RNA secondary structure. As such, using algorithms similar to those for predicting RNA secondary structure, computational methods have been recently developed for the prediction of RNA-RNA interactions. We provide the first comprehensive comparison comprising 14 methods that predict general intermolecular basepairs. To evaluate these, we compile an extensive data set of 54 experimentally confirmed fungal snoRNA-rRNA interactions and 102 bacterial sRNA-mRNA interactions. We test the performance accuracy of all methods, evaluating the effects of tool settings, sequence length, and multiple sequence alignment usage and quality. Our results show that-unlike for RNA secondary structure prediction--the overall best performing tools are non-comparative energy-based tools utilizing accessibility information that predict short interactions on this data set. Furthermore, we find that maintaining high accuracy across biologically different data sets and increasing input lengths remains a huge challenge, causing implications for de novo transcriptome-wide searches. Finally, we make our interaction data set publicly available for future development and benchmarking efforts.


Nicotine vaccines for smoking cessation.
By reducing the amount of nicotine that reaches the brain when a person smokes a cigarette, nicotine vaccines may help people to stop smoking or to prevent recent quitters from relapsing. The aims of this review are to assess the efficacy of nicotine vaccines for smoking cessation and for relapse prevention, and to assess the frequency and type of adverse events associated with the use of nicotine vaccines. We searched the Cochrane Tobacco Addiction Review Group specialised register for trials, using the term 'vaccine' in the title or abstract, or in a keyword (date of most recent search April 2012). To identify any other material including reviews and papers potentially relevant to the background or discussion sections, we also searched MEDLINE, EMBASE, and PsycINFO, combining terms for nicotine vaccines with terms for smoking and tobacco use, without design limits or limits for human subjects. We searched the Annual Meeting abstracts of the Society for Research on Nicotine and Tobacco up to 2012, using the search string 'vaccin'. We searched Google Scholar for 'nicotine vaccine'. We also searched company websites and Google for information related to specific vaccines. We searched clinicaltrials.gov in March 2012 for 'nicotine vaccine' and for the trade names of known vaccine candidates. We included randomized controlled trials of nicotine vaccines, at Phase II and Phase III trial stage and beyond, in adult smokers or recent ex-smokers. We included studies of nicotine vaccines used as part of smoking cessation or relapse prevention interventions. We extracted data on the type of participants, the dose and duration of treatment, the outcome measures, the randomization procedure, concealment of allocation, blinding of participants and personnel, reporting of outcomes, and completeness of follow-up.Our primary outcome measure was a minimum of six months abstinence from smoking. We used the most rigorous definition of abstinence, and preferred cessation rates at 12 months and biochemically validated rates where available. We have used the risk ratio (RR) to summarize individual trial outcomes. We have not pooled the current group of included studies as they cover different vaccines and variable regimens. There are no nicotine vaccines currently licensed for public use, but there are a number in development. We found four trials which met our inclusion criteria, three comparing NicVAX to placebo and one comparing NIC002 (formerly NicQbeta) to placebo. All were smoking cessation trials conducted by pharmaceutical companies as part of the drug development process, and all trials were judged to be at high or unclear risk of bias in at least one domain. Overall, 2642 smokers participated in the included studies in this review. None of the four included studies detected a statistically significant difference in long-term cessation between participants receiving vaccine and those receiving placebo. The RR for 12 month cessation in active and placebo groups was 1.35 (95% Confidence Interval (CI) 0.82 to 2.22) in the trial of NIC002 and 1.74 (95% CI 0.73 to 4.18) in one NicVAX trial. Two Phase III NicVAX trials, for which full results were not available, reported similar quit rates of approximately 11% in both groups. In the two studies with full results available, post hoc analyses detected higher cessation rates in participants with higher levels of nicotine antibodies, but these findings are not readily generalisable. The two studies with full results showed nicotine vaccines to be well tolerated, with the majority of adverse events classified as mild or moderate. In the study of NIC002, participants receiving the vaccine were more likely to report mild to moderate adverse events, most commonly flu-like symptoms, whereas in the study of NicVAX there was no significant difference between the two arms. Information on adverse events was not available for the large Phase III trials of NicVAX.Vaccine candidates are likely to undergo significant changes before becoming available to the general public, and those included in this review may not be the first to reach market; this limits the external validity of the results reported in this review in terms of both effectiveness and tolerability. There is currently no evidence that nicotine vaccines enhance long-term smoking cessation. Rates of serious adverse events recorded in the two trials with full data available were low, and the majority of adverse events reported were at mild to moderate levels. The evidence available suggests nicotine vaccines do not induce compensatory smoking or affect withdrawal symptoms. No nicotine vaccines are currently licensed for use in any country but a number are under development.Further trials of nicotine vaccines are needed, comparing vaccines with placebo for smoking cessation. Further trials are also needed to explore the potential of nicotine vaccines to prevent relapse. Results from past, current and future research should be reported in full. Adverse events and serious adverse events should continue to be carefully monitored and thoroughly reported.


Is there a difference between capsule endoscopy and computed tomography as a first-line study in obscure gastrointestinal bleeding?
Capsule endoscopy (CE) is currently recommended as the first-line study in the evaluation of obscure gastrointestinal bleeding (OGIB), while computed tomography (CT) is often thought of as complementary to CE. This study evaluated CT as a first-line study in OGIB and compared it with CE. Ninety-nine patients with OGIB who received both CE and CT were included. CT included conventional CT and CT enterography (CTE). Patients were divided into two groups: the CT before CE group (CT first group n=75) and the CE before CT group (CE first group n=24). The two groups were compared retrospectively. Overt OGIB was present in 92% of patients. Mucosal lesions (46%) were the most common diagnoses, while tumors accounted for 7%. The diagnostic yield of CE was significantly higher than that of CT for both groups (CT first group, p<0.001; CE first group, p=0.013). In the CT first group, the diagnostic yield using both CT and CE (48/75; 64%) was significantly higher than that for CT alone (12/75; 16%, p=0.005). In the CE first group, the diagnostic yield with both CT and CE versus CE alone was 70.9% versus 62.5%, respectively, with a significant difference (p=0.045). There was no significant clinical difference associated with the order in which the tests were performed. However, CE and CT, when used together, had a significantly greater diagnostic yield than did CE or CT when used alone.


Templated synthesis of nylon nucleic acids and characterization by nuclease digestion.
Nylon nucleic acids containing oligouridine nucleotides with pendent polyamide linkers and flanked by unmodified heteronucleotide sequences were prepared by DNA templated synthesis. Templation was more efficient than the single-stranded synthesis: Coupling step yields were as high as 99.2%, with up to 7 amide linkages formed in the synthesis of a molecule containing 8 modified nucleotides. Controlled digestion by calf spleen phosphodiesterase enabled the mapping of modified nucleotides in the sequences. A combination of complete degradation of nylon nucleic acids by snake venom phosphodiesterase and dephosphorylation of the resulting nucleotide fragments by bacterial alkaline phosphatase, followed by LCMS analysis, clarified the linear structure of the oligo-amide linkages. The templated synthesis strategy afforded nylon nucleic acids in the target structure and was compatible with the presence heteronucleotides. The complete digestion procedure produced a new species of DNA analogues, nylon ribonucleosides, which display nucleosides attached via a 2'-alkylthio linkage to each diamine and dicarboxylate repeat unit of the original nylon nucleic acids. The binding affinity of a nylon ribonucleoside octamer to the complementary DNA was evaluated by thermal denaturing experiments. The octamer was found to form stable duplexes with an inverse dependence on salt concentration, in contrast to the salt-dependent DNA control.


The cytoprotective enzyme heme oxygenase-1 suppresses Ebola virus replication.
Ebola virus (EBOV) is the causative agent of a severe hemorrhagic fever in humans with reported case fatality rates as high as 90%. There are currently no licensed vaccines or antiviral therapeutics to combat EBOV infections. Heme oxygenase-1 (HO-1), an enzyme that catalyzes the rate-limiting step in heme degradation, has antioxidative properties and protects cells from various stresses. Activated HO-1 was recently shown to have antiviral activity, potently inhibiting the replication of viruses such as hepatitis C virus and human immunodeficiency virus. However, the effect of HO-1 activation on EBOV replication remains unknown. To determine whether the upregulation of HO-1 attenuates EBOV replication, we treated cells with cobalt protoporphyrin (CoPP), a selective HO-1 inducer, and assessed its effects on EBOV replication. We found that CoPP treatment, pre- and postinfection, significantly suppressed EBOV replication in a manner dependent upon HO-1 upregulation and activity. In addition, stable overexpression of HO-1 significantly attenuated EBOV growth. Although the exact mechanism behind the antiviral properties of HO-1 remains to be elucidated, our data show that HO-1 upregulation does not attenuate EBOV entry or budding but specifically targets EBOV transcription/replication. Therefore, modulation of the cellular enzyme HO-1 may represent a novel therapeutic strategy against EBOV infection.


Treatment of discogenic back pain with autologous bone marrow concentrate injection with minimum two year follow-up.
The purpose of this study is to assess safety and feasibility of intradiscal bone marrow concentrate (BMC) injections to treat discogenic pain as an alternative to surgery. A total of 26 patients (11 male, 15 female, aged 18-61 years, 13 single level, 13 two level) that met inclusion criteria of chronic (> 6 months) discogenic low back pain, degenerative disc pathology assessed by magnetic resonance imaging (MRI) with modified Pfirrmann grade of IV-VII at one or two levels, candidate for surgical intervention (failed conservative treatment and radiologic findings) and a visual analogue scale (VAS) pain score of 40 mm or more at initial visit. Initial Oswestry Disability Index (ODI) and VAS pain score average was 56.5 % and 80.1 mm (0-100), respectively. Adverse event reporting, ODI score, VAS pain score, MRI radiographic changes, progression to surgery and cellular analysis of BMC were noted. Retrospective cell analysis by flow cytometry and colony forming unit-fibroblast (CFU-F) assays were performed to characterise each patient's BMC and compare with clinical outcomes. The BMC was injected into the nucleus pulposus of the symptomatic disc(s) under fluoroscopic guidance. Patients were evaluated clinically prior to treatment and at three, six, 12 and 24 months and radiographically prior to treatment and at 12 months. There were no complications from the percutaneous bone marrow aspiration or disc injection. Of 26 patients, 24 (92 %) avoided surgery through 12 months, while 21 (81 %) avoided surgery through two years. Of the 21 surviving patients, the average ODI and VAS scores were reduced to 19.9 and 27.0 at three months and sustained to 18.3 and 22.9 at 24 months, respectively (p ≤ 0.001). Twenty patients had follow-up MRI at 12 months, of whom eight had improved by at least one Pfirrmann grade, while none of the discs worsened. Total and rate of pain reduction were linked to mesenchymal stem cell concentration through 12 months. Only five of the 26 patients elected to undergo surgical intervention (fusion or artificial disc replacement) by the two year milestone. This study provides evidence of safety and feasibility in the non-surgical treatment of discogenic pain with autologous BMC, with durable pain relief (71 % VAS reduction) and ODI improvements (> 64 %) through two years.


The zebrafish mutant bumper shows a hyperproliferation of lens epithelial cells and fibre cell degeneration leading to functional blindness.
The development of the eye lens is one of the classical paradigms of induction during embryonic development in vertebrates. But while there have been numerous studies aimed at discovering the genetic networks controlling early lens development, comparatively little is known about later stages, including the differentiation of secondary lens fibre cells. The analysis of mutant zebrafish isolated in forward genetic screens is an important way to investigate the roles of genes in embryogenesis. In this study we describe the zebrafish mutant bumper (bum), which shows a transient, tumour-like hyperproliferation of the lens epithelium as well as a progressively stronger defect in secondary fibre cell differentiation, which results in a significantly reduced lens size and ectopic location of the lens within the neural retina. Interestingly, the initial hyperproliferation of the lens epithelium in bum spontaneously regresses, suggesting this mutant as a valuable model to study the molecular control of tumour progression/suppression. Behavioural analyses demonstrate that, despite a morphologically normal retina, larval and adult bum(-/-) zebrafish are functionally blind. We further show that these fish have defects in their craniofacial skeleton with normal but delayed formation of the scleral ossicles within the eye, several reduced craniofacial bones resulting in an abnormal skull shape, and asymmetric ectopic bone formation within the mandible. Genetic mapping located the mutation in bum to a 4cM interval on chromosome 7 with the closest markers located at 0.2 and 0cM, respectively.


Utility of syndecan-1 (CD138) expression in the diagnosis of undifferentiated malignant neoplasms: a tissue microarray study of 1,754 cases.
Syndecan-1, a heparan sulfate-rich membrane glycoprotein, is expressed in plasma cells and is considered a reliable marker of plasmacytic differentiation. However, it has not been widely tested in non-hematolymphoid tissues, and thus its utility in the setting of an undifferentiated malignant neoplasm has not been evaluated. The authors conducted an extensive study of CD138 staining in over 1,700 normal, benign, and malignant non-hematolymphoid tissues, using five tissue microarrays. Immunohistochemical staining was performed with two commercially available CD138 monoclonal antibodies directed against syndecan-1 (Serotec, Oxford, UK, and DAKO, Carpenteria, CA). In addition to the specific membrane staining, many normal tissues and epithelial tumors showed strong cytoplasmic immunoreactivity. A small subset of mesenchymal neoplasms also showed membrane and cytoplasmic immunoreactivity. In squamous cell carcinoma of the head and neck, renal cell carcinoma, and prostate adenocarcinoma, the intensity of CD138 staining inversely correlated with the histologic grade of the carcinoma. However, statistically significant staining differences and their correlation with histologic grades differed depending on whether the Serotec or the DAKO antibody was used. These results indicate that CD138 immunoreactivity is widespread in normal and neoplastic epithelial tissues, as well as a variety of undifferentiated epithelial and mesenchymal processes. The authors conclude that the expression of syndecan-1, although relatively specific to plasma cells within the hematolymphoid system, should be interpreted with extreme caution in the setting of an undifferentiated neoplasm. Furthermore, the two commercially available monoclonal CD138 antibodies tested in this study showed significant differences in their immunoreactivity in different tumor types.


Functional assessment of isolated mitochondria in vitro.
Mitochondria play a pivotal role in cellular function, not only as a major site of ATP production, but also by regulating energy expenditure, apoptosis signaling, and production of reactive oxygen species. Altered mitochondrial function is reported to be a key underlying mechanism of many pathological states and in the aging process. Functional measurements of intact mitochondria isolated from fresh tissue provides distinct information regarding the function of these organelles that complements conventional mitochondrial assays using previously frozen tissue as well as in vivo assessment using techniques such as magnetic resonance and near-infrared spectroscopy. This chapter describes the process by which mitochondria are isolated from small amounts of human skeletal muscle obtained by needle biopsy and two approaches used to assess mitochondrial oxidative capacity and other key components of mitochondrial physiology. We first describe a bioluminescent approach for measuring the rates of mitochondrial ATP production. Firefly luciferase catalyzes a light-emitting reaction whereby the substrate luciferin is oxidized in an ATP-dependent manner. A luminometer is used to quantify the light signal, which is proportional to ATP concentration. We also review a method involving polarographic measurement of oxygen consumption. Measurements of oxygen consumption, which previously required large amounts of tissue, are now feasible with very small amounts of sample obtained by needle biopsy due to recent advances in the field of high-resolution respirometry. We illustrate how careful attention to substrate combinations and inhibitors allows an abundance of unique functional information to be obtained from isolated mitochondria, including function at various energetic states, oxidative capacity with electron flow through distinct complexes, coupling of oxygen consumption to ATP production, and membrane integrity. These measurements, together with studies of mitochondrial DNA abundance, mRNA levels, protein expression, and synthesis rates of mitochondrial proteins provide insightful mechanistic information about mitochondria in a variety of tissue types.


Use of resonance ionization spectroscopy to detect DNA bands on ultrathin spin-coated gels.
Development of alternative electrophoresis procedures are necessary for large volume sequencing and mapping studies. The use of stable isotopes as DNA labels and ultrathin gels promises to greatly increase the rate of sequencing. Spin coating is presented as an alternative method for producing ultrathin polyacrylamide gels. The technique has the potential of producing gels of micron to submicron thicknesses by varying the viscosity of the acrylamide solution and the spinning speed. Thirty micron thick 6% (weight %) gels were produced in this manner. Tin-labeled DNA oligomers were electrophoresed and detected using sputter-initiated resonance ionization spectroscopy (SIRIS). The usefulness of SIRIS and laser atomization RIS (LARIS) to sample the surface and deeper layers of 240 microns thick gels was investigated. With LARIS, whole cross-sections of the gel can be atomized, possibly allowing complete sampling of labels.


Vaccine message framing and parents' intent to immunize their infants for MMR.
Emphasizing societal benefits of vaccines has been linked to increased vaccination intentions in adults. It is unclear if this pattern holds for parents deciding whether to vaccinate their children. The objective was to determine whether emphasizing the benefits of measles-mumps-rubella (MMR) vaccination directly to the vaccine recipient or to society differentially impacts parents' vaccine intentions for their infants. In a national online survey, parents (N = 802) of infants <12 months old were randomly assigned to receive 1 of 4 MMR vaccine messages: (1) the Centers for Disease Control and Prevention Vaccine Information Statement (VIS), (2) VIS and information emphasizing the MMR vaccine's benefits to the child, (3) VIS and information emphasizing societal benefits, or (4) VIS and information emphasizing benefits both to the child and society. Parents reported their likelihood of vaccinating their infants for MMR on a response scale of 0 (extremely unlikely) to 100 (extremely likely). Compared with the VIS-only group (mean intention = 86.3), parents reported increased vaccine intentions for their infants when receiving additional information emphasizing the MMR vaccine's benefits either directly to the child (mean intention = 91.6, P = .01) or to both the child and society (mean intention = 90.8, P = .03). Emphasizing the MMR vaccine's benefits only to society did not increase intentions (mean intention = 86.4, P = .97). We did not see increases in parents' MMR vaccine intentions for their infants when societal benefits were emphasized without mention of benefits directly to the child. This finding suggests that providers should emphasize benefits directly to the child. Mentioning societal benefits seems to neither add value to, nor interfere with, information highlighting benefits directly to the child.


H19 lncRNA Promotes Skeletal Muscle Insulin Sensitivity in Part by Targeting AMPK.
Skeletal muscle plays a pivotal role in regulating systemic glucose homeostasis in part through the conserved cellular energy sensor AMPK. AMPK activation increases glucose uptake, lipid oxidation, and mitochondrial biogenesis, leading to enhanced muscle insulin sensitivity and whole-body energy metabolism. Here we show that the muscle-enriched H19 long noncoding RNA (lncRNA) acts to enhance muscle insulin sensitivity, at least in part, by activating AMPK. We identify the atypical dual-specificity phosphatase DUSP27/DUPD1 as a potentially important downstream effector of H19. We show that DUSP27, which is highly expressed in muscle with previously unknown physiological function, interacts with and activates AMPK in muscle cells. Consistent with decreased H19 expression in the muscle of insulin-resistant human subjects and rodents, mice with genetic H19 ablation exhibit muscle insulin resistance. Furthermore, a high-fat diet downregulates muscle H19 via both posttranscriptional and epigenetic mechanisms. Our results uncover an evolutionarily conserved, highly expressed lncRNA as an important regulator of muscle insulin sensitivity.


Feeding low or pharmacological concentrations of zinc oxide changes the hepatic proteome profiles in weaned piglets.
Pharmacological levels of zinc oxide can promote growth and health of weaning piglets, but the underlying molecular mechanisms are yet not fully understood. The aim of this study was to determine changes in the global hepatic protein expression in response to dietary zinc oxide in weaned piglets. Nine half-sib piglets were allocated to three dietary zinc treatment groups (50, 150, 2500 mg/kg dry matter). After 14 d, pigs were euthanized and liver samples taken. The increase in hepatic zinc concentration following dietary supplementation of zinc was accompanied by up-regulation of metallothionein mRNA and protein expression. Global hepatic protein profiles were obtained by two-dimensional difference gel electrophoresis following matrix-assisted laser desorption ionization/time-of-flight mass spectrometry. A total of 15 proteins were differentially (P<0.05) expressed between groups receiving control (150 mg/kg) or pharmacological levels of zinc (2500 mg/kg) with 7 down- (e.g. arginase1, thiosulfate sulfurtransferase, HSP70) and 8 up-regulated (e.g. apolipoprotein AI, transferrin, C1-tetrahydrofolate synthase) proteins. Additionally, three proteins were differentially expressed with low zinc supply (50 mg/kg Zn) in comparison to the control diet. The identified proteins were mainly associated with functions related to cellular stress, transport, metabolism, and signal transduction. The differential regulation was evaluated at the mRNA level and a subset of three proteins of different functional groups was selected for confirmation by western blotting. The results of this proteomic study suggest that zinc affects important liver functions such as blood protein secretion, protein metabolism, detoxification and redox homeostasis, thus supporting the hypothesis of intermediary effects of pharmacological levels of zinc oxide fed to pigs.


ICA-based artefact removal and accelerated fMRI acquisition for improved resting state network imaging.
The identification of resting state networks (RSNs) and the quantification of their functional connectivity in resting-state fMRI (rfMRI) are seriously hindered by the presence of artefacts, many of which overlap spatially or spectrally with RSNs. Moreover, recent developments in fMRI acquisition yield data with higher spatial and temporal resolutions, but may increase artefacts both spatially and/or temporally. Hence the correct identification and removal of non-neural fluctuations is crucial, especially in accelerated acquisitions. In this paper we investigate the effectiveness of three data-driven cleaning procedures, compare standard against higher (spatial and temporal) resolution accelerated fMRI acquisitions, and investigate the combined effect of different acquisitions and different cleanup approaches. We applied single-subject independent component analysis (ICA), followed by automatic component classification with FMRIB's ICA-based X-noiseifier (FIX) to identify artefactual components. We then compared two first-level (within-subject) cleaning approaches for removing those artefacts and motion-related fluctuations from the data. The effectiveness of the cleaning procedures was assessed using time series (amplitude and spectra), network matrix and spatial map analyses. For time series and network analyses we also tested the effect of a second-level cleaning (informed by group-level analysis). Comparing these approaches, the preferable balance between noise removal and signal loss was achieved by regressing out of the data the full space of motion-related fluctuations and only the unique variance of the artefactual ICA components. Using similar analyses, we also investigated the effects of different cleaning approaches on data from different acquisition sequences. With the optimal cleaning procedures, functional connectivity results from accelerated data were statistically comparable or significantly better than the standard (unaccelerated) acquisition, and, crucially, with higher spatial and temporal resolution. Moreover, we were able to perform higher dimensionality ICA decompositions with the accelerated data, which is very valuable for detailed network analyses.


Potential chemical markers for the identification of irradiated sausages.
Hydrocarbons, gas compounds, and off-odor volatiles were determined for irradiated (0 or 5 kGy) commercial sausages with different fat contents (16% and 29%) during a 60-d storage period at 4 °C. Total of 4 hydrocarbons (C14:1, C15:0, C16:2, and C17:1) were detected only in irradiated sausages: the amount of C16:2 was the highest, followed by C17:1, C14:1, and C15:0. The concentrations of hydrocarbons decreased significantly (P < 0.05) with storage, but were still detectable at the end of 60-d storage. Irradiated sausages produced significantly higher amounts of CO than the nonirradiated ones. CH(4) was detected only in irradiated sausages. Dimethyl disulfide was detected only in irradiated sausages and its concentration decreased significantly (P < 0.05) with storage. Fat content of sausages showed a significant effect on the production and retention of hydrocarbons, gas compounds, and sulfur volatiles in irradiated sausages during storage. Some hydrocarbons (C16:2, C17:1, C14:1, and C15:0), CH(4) , and dimethyl disulfide were only found in irradiated sausages indicating that these compounds can be used as potential markers for irradiated sausages.


Validation of cross-cultural child mental health and psychosocial research instruments: adapting the Depression Self-Rating Scale and Child PTSD Symptom Scale in Nepal.
The lack of culturally adapted and validated instruments for child mental health and psychosocial support in low and middle-income countries is a barrier to assessing prevalence of mental health problems, evaluating interventions, and determining program cost-effectiveness. Alternative procedures are needed to validate instruments in these settings. Six criteria are proposed to evaluate cross-cultural validity of child mental health instruments: (i) purpose of instrument, (ii) construct measured, (iii) contents of construct, (iv) local idioms employed, (v) structure of response sets, and (vi) comparison with other measurable phenomena. These criteria are applied to transcultural translation and alternative validation for the Depression Self-Rating Scale (DSRS) and Child PTSD Symptom Scale (CPSS) in Nepal, which recently suffered a decade of war including conscription of child soldiers and widespread displacement of youth. Transcultural translation was conducted with Nepali mental health professionals and six focus groups with children (n=64) aged 11-15 years old. Because of the lack of child mental health professionals in Nepal, a psychosocial counselor performed an alternative validation procedure using psychosocial functioning as a criterion for intervention. The validation sample was 162 children (11-14 years old). The Kiddie-Schedule for Affective Disorders and Schizophrenia (K-SADS) and Global Assessment of Psychosocial Disability (GAPD) were used to derive indication for treatment as the external criterion. The instruments displayed moderate to good psychometric properties: DSRS (area under the curve (AUC)=0.82, sensitivity=0.71, specificity=0.81, cutoff score ≥ 14); CPSS (AUC=0.77, sensitivity=0.68, specificity=0.73, cutoff score ≥ 20). The DSRS items with significant discriminant validity were "having energy to complete daily activities" (DSRS.7), "feeling that life is not worth living" (DSRS.10), and "feeling lonely" (DSRS.15). The CPSS items with significant discriminant validity were nightmares (CPSS.2), flashbacks (CPSS.3), traumatic amnesia (CPSS.8), feelings of a foreshortened future (CPSS.12), and easily irritated at small matters (CPSS.14). Transcultural translation and alternative validation feasibly can be performed in low clinical resource settings through task-shifting the validation process to trained mental health paraprofessionals using structured interviews. This process is helpful to evaluate cost-effectiveness of psychosocial interventions.


Manganese provides antioxidant protection for sperm cryopreservation that may offer new consideration for clinical fertility.
Reactive oxygen species (ROS) are generated by sperm metabolism. While, ROS are required for maturation, capacitation and acrosome reaction, they also modify many peroxidable cellular compounds. There is production of ROS during cryopreservation and frozen spermatozoa are highly sensitive to lipid peroxidation (LPO). Antioxidants exert a protective effect on the plasma membrane of frozen bovine sperm preserving both metabolic activity and cellular viability. Manganese (Mn(++)) is proved to be a chain breaking antioxidant in biological system. Therefore, we examined the role of (Mn(++)) during cryopreservation of cattle bull semen. Semen was divided into four parts and cryopreserved in egg-yolk-citrate extender + glycerol (EYC-G), EYC-G + 100 microM of Mn(++), EYC-G + 150 microM of Mn(++) and EYC-G + 200 microM of Mn(++). After four hours of cooling and 24 hrs of freezing, the spermatozoa were examined for percentage motility, Hypo-osmotic swelling (HOS), LPO and protein leakage. Addition of manganese to the semen during cryopreservation showed a protective effect and accounted for an increase in semen quality parameters [percentage motility, HOS percent and decrease in malondialdehyde (MDA) production and protein leakage]. The effect of manganese on motility and HOS was non-significant (p < 0.05) in cooled spermatozoa but significant with 150 microM of Mn(++) in frozen-thawed spermatozoa. MDA production and protein leakage decreased to a significant and maximum level (p < 0.05) on addition of 200 microM of manganese. The addition of manganese to EYC-G dilutor will improve the quality/fertility of semen, which will result in improvement of in vitro fertilization and artificial insemination success rate.


Lifestyle factors and the risk of a second breast cancer after ductal carcinoma in situ.
Little information exists on lifestyle factors that affect prognosis after treatment for ductal carcinoma in situ (DCIS) breast cancer. Improved understanding of the role of lifestyle factors is important to survivors wishing to reduce their risk of a second breast cancer diagnosis. We examined the association between body mass index (BMI), physical activity, and alcohol intake, and risk of a second breast cancer diagnosis among 1,925 DCIS survivors in the Wisconsin In Situ Cohort. Exposures were self-reported during biennial patient interviews. Second breast cancer diagnoses were validated via pathology report. Cox proportional hazards regression was used to estimate the association between prediagnosis, postdiagnosis, and change in exposure levels and the risk of a second diagnosis, with adjustment for patient, tumor, and treatment factors. Over a mean of 6.7 years of follow-up, 162 second breast cancer diagnoses were reported, including 57 invasive events, 60 in situ events, and 45 diagnoses of unknown stage. A significant trend of increasing risk of a second diagnosis was found over increasing categories of postdiagnosis alcohol intake (Ptrend = 0.02). Among premenopausal women, increased prediagnosis BMI was associated with a reduced risk of a second diagnosis (HR = 0.93; 95% confidence interval, 0.88-0.99). DCIS survivors may reduce their risk of a second diagnosis by reducing postdiagnosis alcohol consumption. The population of DCIS survivors is projected to surpass one million by the year 2016. Our results suggest that these women may be able to reduce their risk of a second diagnosis through moderation of alcohol consumption.


Avoiding Treatment Interruptions: What Role Do Australian Community Pharmacists Play?
To explore the reported practice of Australian community pharmacists when dealing with medication supply requests in absence of a valid prescription. Self-administered questionnaire was posted to 1490 randomly selected community pharmacies across all Australian states and territories. This sample was estimated to be a 20% of all Australian community pharmacies. Three hundred eighty five pharmacists participated in the study (response rate achieved was 27.9% (there were 111 undelivered questionnaires). Respondents indicated that they were more likely to provide medications to regular customers without a valid prescription compared to non-regular customers (p<0.0001). However, supply was also influenced by the type of prescription and the medication requested. In the case of type of prescription (Standard, Authority or Private) this relates to the complexity/probability of obtaining a valid prescription from the prescriber at a later date (i.e. supply with an anticipated prescription). Decisions to supply and/or not supply related to medication type were more complex. For some cases, including medication with potential for abuse, the practice and/or the method of supply varied significantly according to age and gender of the pharmacist, and pharmacy location (p<0.05). Although being a regular customer does not guarantee a supply, results of this study reinforce the importance for patients having a regular pharmacy, where pharmacists were more likely to continue medication supply in cases of patients presenting without a valid prescription. We would suggest, more flexible legislation should be implemented to allow pharmacists to continue supplying of medication when obtaining a prescription is not practical.


Building a new life: a chaplain's theory based case study of chronic illness.
This article presents the case study of spiritual care for a patient suffering from Parkinson's disease who was referred to the chaplain in an out-patient depression research program. The chaplain's interventions were informed by an application of narrative theory, and the article demonstrates how this theory enabled the chaplain to help a patient develop new coping strategies for dealing with chronic disease. Using narrative theory, the chaplain assisted the patient to develop a new sense of identity as a spiritual, contingent self as the disease eroded his physical self and former life. The article includes a description of a patient's spiritual needs, chaplain interventions, and an outcomes measure of those interventions. The author argues that narrative theory provides chaplains with a language to identify and craft the unique intervention that spiritual care has in the life trajectory of this Parkinson's patient and other patients dealing with chronic illnesses.


Characteristics of the adult learner.
This article provides an overview of adult learner characteristics, with an emphasis on those characteristics studied in diabetes patient education research. A selected review of the conceptual and research literature on general adult education and adult learning was conducted, with particular attention to diabetes patient education studies. Characteristics reviewed included learning styles, literacy level, age/aging, ethnicity or culture, gender, and knowledge. Studies of the learning style of group vs individual education indicated some positive benefits for group learning; questions remain about optimal size or periodicity. Studies evaluating the benefits of culturally specific interventions for diabetes management have yielded some information. Characteristics related to gender and age have been studied, but often in pilot or feasibility studies without the power to answer the study questions. There continue to be many gaps in knowledge related to adult learner characteristics in diabetes education. Lessons from both general adult learning literature and patient education literature from other chronic diseases should be evaluated and incorporated. The complexities of these learner characteristics create challenges in designing studies. However, evidence to support the need for effective educational interventions is of great importance for implementing change in health care.


Reciprocal rewards stabilize cooperation in the mycorrhizal symbiosis.
Plants and their arbuscular mycorrhizal fungal symbionts interact in complex underground networks involving multiple partners. This increases the potential for exploitation and defection by individuals, raising the question of how partners maintain a fair, two-way transfer of resources. We manipulated cooperation in plants and fungal partners to show that plants can detect, discriminate, and reward the best fungal partners with more carbohydrates. In turn, their fungal partners enforce cooperation by increasing nutrient transfer only to those roots providing more carbohydrates. On the basis of these observations we conclude that, unlike many other mutualisms, the symbiont cannot be "enslaved." Rather, the mutualism is evolutionarily stable because control is bidirectional, and partners offering the best rate of exchange are rewarded.


Regulation of tissue factor pathway inhibitor-2 (TFPI-2) expression by lysine-specific demethylase 1 and 2 (LSD1 and LSD2).
Tissue factor pathway inhibitor-2 (TFPI-2) is a major inhibitor of extracellular matrix degradation. Decreases in TFPI-2 contribute to malignant tumor cell production, and TFPI-2 is a presumed tumor suppressor. TFPI-2 gene transcription is regulated by two epigenetic mechanisms: DNA methylation of the promoter and K4 methylation of histone 3 (H3). Lysine-specific demethylase 1 (LSD1) and LSD2 demethylate H3K4me2/1. LSD1 has been implicated in TFPI-2 regulation through both epigenetic mechanisms, but the involvement of LSD2 remains unknown. We prepared a monoclonal anti-LSD2 antibody that clearly distinguishes LSD2 from LSD1. Knockdown of LSD1 or LSD2 by siRNAs increased TFPI-2 protein and mRNA. Simultaneous knockdown of both LSD1 and LSD2 showed additive effects. Bisulfite sequencing revealed that CpG sites in the TFPI-2 promoter region were unmethylated. These results indicate that LSD2 also contributes to TFPI-2 regulation through histone modification, and that further studies of the involvement of LSD2 in tumor malignancy are warranted.


Virtual Genome Walking across the 32 Gb Ambystoma mexicanum genome; assembling gene models and intronic sequence.
Large repeat rich genomes present challenges for assembly using short read technologies. The 32 Gb axolotl genome is estimated to contain ~19 Gb of repetitive DNA making an assembly from short reads alone effectively impossible. Indeed, this model species has been sequenced to 20× coverage but the reads could not be conventionally assembled. Using an alternative strategy, we have assembled subsets of these reads into scaffolds describing over 19,000 gene models. We call this method Virtual Genome Walking as it locally assembles whole genome reads based on a reference transcriptome, identifying exons and iteratively extending them into surrounding genomic sequence. These assemblies are then linked and refined to generate gene models including upstream and downstream genomic, and intronic, sequence. Our assemblies are validated by comparison with previously published axolotl bacterial artificial chromosome (BAC) sequences. Our analyses of axolotl intron length, intron-exon structure, repeat content and synteny provide novel insights into the genic structure of this model species. This resource will enable new experimental approaches in axolotl, such as ChIP-Seq and CRISPR and aid in future whole genome sequencing efforts. The assembled sequences and annotations presented here are freely available for download from https://tinyurl.com/y8gydc6n . The software pipeline is available from https://github.com/LooseLab/iterassemble .


Effects of harvesting flowers from shrubs on the persistence and abundance of wild shrub populations at multiple spatial extents.
Wildflower harvesting is an economically important activity of which the ecological effects are poorly understood. We assessed how harvesting of flowers affects shrub persistence and abundance at multiple spatial extents. To this end, we built a process-based model to examine the mean persistence and abundance of wild shrubs whose flowers are subject to harvest (serotinous Proteaceae in the South African Cape Floristic Region). First, we conducted a general sensitivity analysis of how harvesting affects persistence and abundance at nested spatial extents. For most spatial extents and combinations of demographic parameters, persistence and abundance of flowering shrubs decreased abruptly once harvesting rate exceeded a certain threshold. At larger extents, metapopulations supported higher harvesting rates before their persistence and abundance decreased, but persistence and abundance also decreased more abruptly due to harvesting than at smaller extents. This threshold rate of harvest varied with species' dispersal ability, maximum reproductive rate, adult mortality, probability of extirpation or local extinction, strength of Allee effects, and carrying capacity. Moreover, spatial extent interacted with Allee effects and probability of extirpation because both these demographic properties affected the response of local populations to harvesting more strongly than they affected the response of metapopulations. Subsequently, we simulated the effects of harvesting on three Cape Floristic Region Proteaceae species and found that these species reacted differently to harvesting, but their persistence and abundance decreased at low rates of harvest. Our estimates of harvesting rates at maximum sustainable yield differed from those of previous investigations, perhaps because researchers used different estimates of demographic parameters, models of population dynamics, and spatial extent than we did. Good demographic knowledge and careful identification of the spatial extent of interest increases confidence in assessments and monitoring of the effects of harvesting. Our general sensitivity analysis improved understanding of harvesting effects on metapopulation dynamics and allowed qualitative assessment of the probability of extirpation of poorly studied species.


Where the action is: monitoring local trends in sexual behaviour.
To assess the role in an overall monitoring strategy of the PLACE method of estimating local trends in sexual behaviour among individuals at social venues in areas at increased risk of HIV transmission. Public venues identified by community informants as places where people meet new sexual partners were visited and characterised in Karaganda, Kazakhstan, in 2002 and 2003, and in a township in South Africa in 2000 and 2003. At a subset of venues, a representative sample of individuals socialising at the venue were interviewed about their sexual behaviour. The age distribution and partnerships rates of those socialising at these venues were compared by year and with national data from Kazakhstan and South Africa obtained via household surveys. Women and men socialising at venues were younger and had higher rates of new and concurrent partnerships than men and women interviewed in national household surveys. There was little apparent change in sexual behaviour between 2002 and 2003 in Karaganda. In the South African township, there was a reduction in sexual partnerships and an increase in condom use, possibly due to a local AIDS prevention programme. Findings from the PLACE method supplement national data on sexual behaviours with data from key populations in high transmission areas, inform local targeting of interventions, and, when subsequent rounds of PLACE are implemented, can evaluate change in target populations.


Novel Techniques for Observing Structural Dynamics of Photoresponsive Liquid Crystals.
We discuss in this article the experimental measurements of the molecules in liquid crystal (LC) phase using the time-resolved infrared (IR) vibrational spectroscopy and time-resolved electron diffraction. Liquid crystal phase is an important state of matter that exists between the solid and liquid phases and it is common in natural systems as well as in organic electronics. Liquid crystals are orientationally ordered but loosely packed, and therefore, the internal conformations and alignments of the molecular components of LCs can be modified by external stimuli. Although advanced time-resolved diffraction techniques have revealed picosecond-scale molecular dynamics of single crystals and polycrystals, direct observations of packing structures and ultrafast dynamics of soft materials have been hampered by blurry diffraction patterns. Here, we report time-resolved IR vibrational spectroscopy and electron diffractometry to acquire ultrafast snapshots of a columnar LC material bearing a photoactive core moiety. Differential-detection analyses of the combination of time-resolved IR vibrational spectroscopy and electron diffraction are powerful tools for characterizing structures and photoinduced dynamics of soft materials.


A new variant of the beta subunit of the high-affinity receptor for immunoglobulin E (Fc epsilon RI-beta E237G): associations with measures of atopy and bronchial hyper-responsiveness.
The high affinity receptor for IgE (Fc epsilon RI) has a central role in mast cell degranulation and IgE mediated allergy. A systematic search through the coding regions of the beta subunit of Fc epsilon RI (Fc epsilon RI-beta) has identified a novel coding polymorphism in exon seven. An adenine to guanine substitution changes amino acid residue 237 from glutamic acid to glycine (E237G), in the cytoplasmic tail of the protein. E237G is predicted to introduce a hydrophobicity change within the C-terminus of Fc epsilon RI-beta. It is adjacent to the immunoreceptor tyrosine activation motif (ITAM), and may affect the intracellular signalling capacity of Fc epsilon RI. E237G was detected in 53 subjects from an Australian general population sample of 1004 individuals (5.3%). E237G positive subjects had a significantly elevated skin test response to grass (p = 0.0004) and house dust mite (p = 0.04), RAST to grass (p = 0.002) and bronchial reactivity to methacholine (p = 0.0009). The relative risk of individuals with E237G having asthma compared to subjects without the variant was 2.3 (95% CI 1.26-4.19; p = 0.005).


Tracking metal ions through a Cu/Ag efflux pump assigns the functional roles of the periplasmic proteins.
Copper is an essential nutrient for all aerobic organisms but is toxic in excess. At the host-pathogen interface, macrophages respond to bacterial infection by copper-dependent killing mechanisms, whereas the invading bacteria are thought to counter with an up-regulation of copper transporters and efflux pumps. The tripartite efflux pump CusCBA and its metallochaperone CusF are vital to the detoxification of copper and silver ions in the periplasm of Escherichia coli. However, the mechanism of efflux by this complex, which requires the activation of the inner membrane pump CusA, is poorly understood. Here, we use selenomethionine (SeM) active site labels in a series of biological X-ray absorption studies at the selenium, copper, and silver edges to establish a "switch" role for the membrane fusion protein CusB. We determine that metal-bound CusB is required for activation of cuprous ion transfer from CusF directly to a site in the CusA antiporter, showing for the first time (to our knowledge) the in vitro activation of the Cus efflux pump. This metal-binding site of CusA is unlike that observed in the crystal structures of the CusA protein and is composed of one oxygen and two sulfur ligands. Our results suggest that metal transfer occurs between CusF and apo-CusB, and that, when metal-loaded, CusB plays a role in the regulation of metal ion transfer from CusF to CusA in the periplasm.


Association of interleukin-1 gene polymorphisms with sudden sensorineural hearing loss and Ménière's disease.
Sudden sensorineural hearing loss (SSNHL) and Ménière's disease are the most common inner ear diseases in which the causes are unknown. As recent magnetic resonance imaging has demonstrated disruption of the blood-labyrinth barrier in these inner ear diseases, inflammatory reaction associated with increased permeability of the blood vessels may be involved. The genotypes of interleukin 1A (IL1A) (-889C/T; rs1800587) and interleukin 1B (IL1B) (-511C/T; rs16944) were determined using an allele-specific primer-polymerase chain reaction method in 72 patients with SSNHL, 68 patients with Ménière's disease, and 2202 control subjects living almost in the same area as the patients. A significantly higher prevalence of the IL1A-889T allele was observed in SSNHL and Ménière's disease compared with controls, although no significant difference in distribution of IL1B-511C/T genotypes was observed between the patients and controls. Adjusted odd ratios for SSNHL and Ménière's disease risks in the -889TT genotypes were 25.89 (95% confidence interval (CI) 12.19-54.98) and 18.20 (95% CI 7.80-42.46), respectively, after age and gender were taken as moderator variables. Our results suggested that IL1A is closely associated with susceptibility of SSNHL and Ménière's disease.


Association between obesity and atopy in Chinese schoolchildren.
Despite parallel increases in asthma and obesity prevalence, there is little data on obesity as a risk factor for atopy. The latter is an important phenotype in asthmatic patients. This study investigates the association between asthma traits, atopy and obesity-related markers in Chinese adolescents. 486 schoolchildren were recruited among participants of our population-based study on the epidemiology of obesity, and their allergy status was ascertained using a standardized questionnaire. Subjects' anthropometry was recorded on-site, and fasting blood was collected for allergen-specific immunoglobulin E (IgE), lipids and systemic inflammatory biomarkers. 98 (20.2%) subjects were classified as overweight or obese. Obesity status was not associated with asthma, allergic rhinitis or eczema (p > 0.25). Atopy was not associated with age-adjusted body mass index (BMI) or waist circumference. Atopy and presence of allergen-specific IgE did not differ between overweight or obese children and those with normal BMI (p > 0.25), although subgroup analysis suggested that cockroach sensitization was more common among males who were obese or overweight (p = 0.045). White cell count (WCC) was higher among atopic than nonatopic children (mean values 6.5 x 10(9)/l vs. 6.2 x 10(9)/l, p = 0.006). Logistic regression revealed WCC to be the only risk factor for atopy (OR 18.97, p = 0.004). Obesity is not associated with asthma or atopy in Chinese children. High WCC is an important risk factor for atopy in both males and females. Gender does not exert any consistent effect on the association between obesity and allergen sensitization in children.


Secular trends in menarcheal age in India-evidence from the Indian human development survey.
Evidence from a number of countries in Europe and North America point towards the secular declining trend in menarcheal age with considerable spatial variations over the past two centuries. Similar trends were reported in several developing countries from Asia, Africa and Latin America. However, data corroborating any secular trend in the menarcheal age of the Indian population remained sparse and inadequately verified. We examined secular trends, regional heterogeneity and association of socioeconomic, anthropometric and contextual factors with menarcheal age among ever-married women (15-49 years) in India. Using the pseudo cohort data approach, we fit multiple linear regression models to estimate secular trends in menarcheal age of 91394 ever-married women using the Indian Human Development Survey. The mean age at menarche among Indian women was 13.76 years (95 % CI: 13.75, 13.77) in 2005. It declined by three months from 13.83 years (95% CI: 13.81, 13.85) among women born prior to 1955-1964, to nearly 13.62 years (95% CI: 13.58, 13.67) among women born during late 1985-1989. However, these aggregate national figures mask extensive spatial heterogeneity as mean age at menarche varied from 15.0 years in Himachal Pradesh during 1955-1964 (95% CI: 14.89-15.11) to about 12.1 years in Assam (95% CI: 11.63-12.56) during 1985-1989. The regression analysis established a reduction of nearly one month per decade, suggesting a secular decline in age at menarche among Indian women. Notably, the menarcheal age was significantly associated with the area of residence, geographic region, linguistic groups, educational attainment, wealth status, caste and religious affiliations among Indian women.


Population genetics of seaside Sparrow (Ammodramus maritimus) subspecies along the gulf of Mexico.
Seaside Sparrows (Ammodramus maritimus) along the Gulf of Mexico are currently recognized as four subspecies, including taxa in Florida (A. m. juncicola and A. m. peninsulae) and southern Texas (Ammodramus m. sennetti), plus a widespread taxon between them (A. m. fisheri). We examined population genetic structure of this "Gulf Coast" clade using microsatellite and mtDNA data. Results of Bayesian analyses (Structure, GeneLand) of microsatellite data from nine locations do not entirely align with current subspecific taxonomy. Ammodramus m. sennetti from southern Texas is significantly differentiated from all other populations, but we found evidence of an admixture zone with A. m. fisheri near Corpus Christi. The two subspecies along the northern Gulf Coast of Florida are significantly differentiated from both A. m. sennetti and A. m. fisheri, but are not distinct from each other. We found a weak signal of isolation by distance within A. m. fisheri, indicating this population is not entirely panmictic throughout its range. Although continued conservation concern is warranted for all populations along the Gulf Coast, A. m. fisheri appears to be more secure than the far smaller populations in south Texas and the northern Florida Gulf Coast. In particular, the most genetically distinct populations, those in Texas south of Corpus Christi, occupy unique habitats within a very small geographic range.