Inhibition of airway smooth muscle tone by Chinese herbal medicines.
The aim of the present study was to elucidate whether Chinese traditional herbal drugs, Gorei-San (TJ-17) and Toki-Shakuyaku-San (TJ-23), affect airway smooth muscle tone and, if so, to determine what the mechanism of action is. Rabbit tracheal segments were isolated and the contractile responses to electrical field stimulation and acetylcholine were measured before and after the application of TJ-17 or TJ-23 under isometric conditions in vitro. Ouabain-sensitive rubidium-86 (86Rb) uptake by tissues in response to each drug was also measured. Each herbal medicine attenuated the contractile responses to electrical field stimulation and acetylcholine in a concentration-dependent manner, the maximal inhibition of acetylcholine-induced contraction being 37.5+/-4.9% for TJ-17 and 42.4+/-5.3% for TJ-23 (p<0.05 for each). These effects were not altered by mechanical removal of the epithelium, indomethacin, the nitric oxide synthase inhibitor NG -nitro-L-arginine methyl ester, the cyclic adenosine monophosphate (cAMP)-dependent protein kinase inhibitor adenosine 3'5'-cyclic monophosphorothioate (Rp-cAMPS), the cyclic guanosine monophosphate (cGMP)-dependent protein kinase inhibitor KT5823, or the calcium (Ca2+)-activated potassium (K+) channel inhibitor charybdotoxin, but were greatly inhibited in the presence of the sodium (Na+)-K+ adenosine triphosphatase (ATPase) inhibitor ouabain. Incubation of tissues with TJ-17 and TJ-23 dose dependently increased ouabain-sensitive 86Rb uptake. The results of the study suggest that both Gorei-San and Toki-Shakuyaku-San reduce airway smooth muscle tone via a postjunctional mechanism probably through stimulation of the sodium pump and the subsequent hyperpolarization/repolarization of the cell membrane. These effects may contribute to the antiasthmatic properties of these herbal medicines.


Contextual behavior and neural circuits.
Animals including humans engage in goal-directed behavior flexibly in response to items and their background, which is called contextual behavior in this review. Although the concept of context has long been studied, there are differences among researchers in defining and experimenting with the concept. The current review aims to provide a categorical framework within which not only the neural mechanisms of contextual information processing but also the contextual behavior can be studied in more concrete ways. For this purpose, we categorize contextual behavior into three subcategories as follows by considering the types of interactions among context, item, and response: contextual response selection, contextual item selection, and contextual item-response selection. Contextual response selection refers to the animal emitting different types of responses to the same item depending on the context in the background. Contextual item selection occurs when there are multiple items that need to be chosen in a contextual manner. Finally, when multiple items and multiple contexts are involved, contextual item-response selection takes place whereby the animal either chooses an item or inhibits such a response depending on item-context paired association. The literature suggests that the rhinal cortical regions and the hippocampal formation play key roles in mnemonically categorizing and recognizing contextual representations and the associated items. In addition, it appears that the fronto-striatal cortical loops in connection with the contextual information-processing areas critically control the flexible deployment of adaptive action sets and motor responses for maximizing goals. We suggest that contextual information processing should be investigated in experimental settings where contextual stimuli and resulting behaviors are clearly defined and measurable, considering the dynamic top-down and bottom-up interactions among the neural systems for contextual behavior.


3D Cell Culture in Alginate Hydrogels.
This review compiles information regarding the use of alginate, and in particular alginate hydrogels, in culturing cells in 3D. Knowledge of alginate chemical structure and functionality are shown to be important parameters in design of alginate-based matrices for cell culture. Gel elasticity as well as hydrogel stability can be impacted by the type of alginate used, its concentration, the choice of gelation technique (ionic or covalent), and divalent cation chosen as the gel inducing ion. The use of peptide-coupled alginate can control cell-matrix interactions. Gelation of alginate with concomitant immobilization of cells can take various forms. Droplets or beads have been utilized since the 1980s for immobilizing cells. Newer matrices such as macroporous scaffolds are now entering the 3D cell culture product market. Finally, delayed gelling, injectable, alginate systems show utility in the translation of in vitro cell culture to in vivo tissue engineering applications. Alginate has a history and a future in 3D cell culture. Historically, cells were encapsulated in alginate droplets cross-linked with calcium for the development of artificial organs. Now, several commercial products based on alginate are being used as 3D cell culture systems that also demonstrate the possibility of replacing or regenerating tissue.


Protective Effect of Propolis in Proteinuria, Crystaluria, Nephrotoxicity and Hepatotoxicity Induced by Ethylene Glycol Ingestion.
Propolis is a natural honeybee product with wide biological activities and potential therapeutic properties. The aim of the study is to evaluate the protective effect of propolis extract on nephrotoxicity and hepatotoxicity induced by ethylene glycol in rats. Five groups of rats were used. Group 1 received drinking water, group 2 received 0.75% ethylene-glycol in drinking water, group 3 received 0.75% ethylene-glycol in drinking water along with cystone 500 mg/kg/body weight (bw) daily, group 4 received 0.75% ethylene-glycol in drinking water along with propolis extract at a dose of 100 mg/kg/bw daily, and group 5 received 0.75% ethylene-glycol in drinking water along with propolis extract at a dose of 250 mg/kg/bw daily. The treatment continued for a total of 30 d. Urinalyses for pH, crystals, protein, creatinine, uric acid and electrolytes, and renal and liver function tests were performed. Ethylene-glycol increased urinary pH, urinary volume, and urinary calcium, phosphorus, uric acid and protein excretion. It decreased creatinine clearance and magnesium and caused crystaluria. Treatment with propolis extract or cystone normalized the level of magnesium, creatinine, sodium, potassium and chloride. Propolis is more potent than cystone. Propolis extract alleviates urinary protein excretion and ameliorates the deterioration of liver and kidney function caused by ethylene glycol. Propolis extract has a potential protective effect against ethylene glycol induced hepatotoxicity and nephrotoxicity and has a potential to treat and prevent urinary calculus, crystaluria and proteinuria.


Coping with genetic diversity: the contribution of pathogen and human genomics to modern vaccinology.
Vaccine development faces major difficulties partly because of genetic variation in both infectious organisms and humans. This causes antigenic variation in infectious agents and a high interindividual variability in the human response to the vaccine. The exponential growth of genome sequence information has induced a shift from conventional culture-based to genome-based vaccinology, and allows the tackling of challenges in vaccine development due to pathogen genetic variability. Additionally, recent advances in immunogenetics and genomics should help in the understanding of the influence of genetic factors on the interindividual and interpopulation variations in immune responses to vaccines, and could be useful for developing new vaccine strategies. Accumulating results provide evidence for the existence of a number of genes involved in protective immune responses that are induced either by natural infections or vaccines. Variation in immune responses could be viewed as the result of a perturbation of gene networks; this should help in understanding how a particular polymorphism or a combination thereof could affect protective immune responses. Here we will present: i) the first genome-based vaccines that served as proof of concept, and that provided new critical insights into vaccine development strategies; ii) an overview of genetic predisposition in infectious diseases and genetic control in responses to vaccines; iii) population genetic differences that are a rationale behind group-targeted vaccines; iv) an outlook for genetic control in infectious diseases, with special emphasis on the concept of molecular networks that will provide a structure to the huge amount of genomic data.


Constructing stable spatial maps of the world.
To interact rapidly and effectively with our environment, our brain needs access to a neural representation--or map--of the spatial layout of the external world. However, the construction of such a map poses major challenges to the visual system, given that the images on our retinae depend on where the eyes are looking, and shift each time we move our eyes, head, and body to explore the world. Much research has been devoted to how the stability is achieved, with the debate often polarized between the utility of spatiotopic maps (that remain solid in external coordinates), as opposed to transiently updated retinotopic maps. Our research suggests that the visual system uses both strategies to maintain stability. fMRI, motion-adaptation, and saccade-adaptation studies demonstrate and characterize spatiotopic neural maps within the dorsal visual stream that remain solid in external rather than retinal coordinates. However, the construction of these maps takes time (up to 500 ms) and attentional resources. To solve the immediate problems created by individual saccades, we postulate the existence of a separate system to bridge each saccade with neural units that are 'transiently craniotopic'. These units prepare for the effects of saccades with a shift of their receptive fields before the saccade starts, then relaxing back into their standard position during the saccade, compensating for its action. Psychophysical studies investigating localization of stimuli flashed briefly around the time of saccades provide strong support for these neural mechanisms, and show quantitatively how they integrate information across saccades. This transient system cooperates with the spatiotopic mechanism to provide a useful map to guide interactions with our environment: one rapid and transitory, bringing into play the high-resolution visual areas; the other slow, long-lasting, and low-resolution, useful for interacting with the world.


Role of alcohol in hospitalized road trauma in Viet nam.
To assess the blood alcohol concentration (BAC; dependent variable) of patients with road traffic injuries (RTIs) presenting at 3 provincial and central hospitals in the Socialist Republic of Viet Nam by age, sex, and road user type (independent variables). This survey formed part of the Viet Nam Road Traffic Injury Prevention Project, funded by Bloomberg Philanthropies. RTIs are a leading cause of death and disability in Viet Nam, with 14,690 deaths and 143,940 injuries reported by the Ministry of Health (MOH) in 2010. Research estimates suggest that motorcycle riders and passengers account for 60 percent of fatalities. Alcohol has long been suspected of being a leading cause of road traffic collisions and injuries. However, until now data on this relationship have been limited. A descriptive cross-sectional study measuring BAC in all consenting patients with RTIs presenting at 3 provincial or central hospitals between July 2009 and September 2010. All results were anonymous and summary information on key variables was sent to MOH and the World Health Organization (WHO) on a monthly basis. Of the 36,418 patients with RTIs presenting to these 3 hospitals between July 2009 and September 2010, BAC analysis was completed on 14,990 patients (41.2%), representing all patients with RTIs 15 years of age and above who consented to anonymous testing. BAC results ranged from 0 to 0.589 g/dL blood, with a mean of 0.0441 g/dL being the average concentration among all tested patients. Of all patients tested, 56.8 percent had no detectable alcohol in their system. Motorcycle riders were most commonly represented in the tested sample (70.7%), with 27.8 percent having a BAC above the legal limit (0.05 g/dL). Car or other vehicle drivers represented 2.7 percent of the sample, with 63.4 percent tested having a BAC above 0 g/dL, the legal limit for these road users. The results of this preliminary study indicate that 29.1 percent of all car drivers and motorcycle riders presenting at hospitals with RTIs exceeded the legal BAC limit for operating a motor vehicle. Though further study is required, this is suggestive that strengthening the enforcement of drink-driving laws is an urgent national road safety priority.


Selective cholinesterase inhibitors from Buxus sempervirens L. and their molecular docking studies.
In this work, two alkaloids namely (+)-buxabenzamidienine (1) and (+)-buxamidine (2) were isolated from Buxus sempervirens, using bioassay-guided fractionation and isolation method. Their acetyl- (AChE) and butyrylcholinesterase (BChE) inhibitory activities were studied and the compounds were found to be quite selective inhibitors of AChE. IC50 values of compound 1 for electric eel AChE and horse BChE were 0.787 and 7.68 mM, respectively; while the corresponding IC50 of compound 2 were 1.70 and 549.98 mM, respectively. Theoretical (quantum mechanical, homology modelling and docking) calculations were performed in order to explain their interactions with different AChE (electric eel and human) and BChE (horse and human). The x-ray crystal structures of electric eel AChE, human AChE, human BChE and a model of horse BChE constructed by homology with human BChE were used for docking of compounds 1 and 2. Density functional theory (DFT) calculations of the compounds were performed at the B3LYP/6- 31G** level using the program Spartan™, and their HOMO and LUMO energy levels were calculated. Docking studies exhibited that compound 1 interacts with the acyl-binding pocket of the active site gorge of huAChE, and including several other hydrophobic interactions.


Tracking deforming objects using particle filtering for geometric active contours.
Tracking deforming objects involves estimating the global motion of the object and its local deformations as a function of time. Tracking algorithms using Kalman filters or particle filters have been proposed for finite dimensional representations of shape, but these are dependent on the chosen parametrization and cannot handle changes in curve topology. Geometric active contours provide a framework which is parametrization independent and allow for changes in topology. In the present work, we formulate a particle filtering algorithm in the geometric active contour framework that can be used for tracking moving and deforming objects. To the best of our knowledge, this is the first attempt to implement an approximate particle filtering algorithm for tracking on a (theoretically) infinite dimensional state space.


Effect of two randomised exercise programmes on bone mass of healthy postmenopausal women.
The effect of two structured exercise programmes on the bone mass of 48 healthy postmenopausal white women aged 50-62 was studied after one year. Volunteers were randomised to group 1 (control), group 2 (aerobic exercise), or group 3 (aerobic and strengthening exercises). Before and after the training programme each subject had evaluations of bone mass (determined by neutron activation analysis and expressed as calcium bone index) and maximum oxygen uptake attained on a multistage exercise treadmill test. After one year both exercise groups had higher levels of fitness and greater bone mass than controls. Mean values (2 SEM) for changes in the calcium bone index were -0.011 (0.037), 0.039 (0.035), and 0.066 (0.036) for groups 1, 2, and 3, respectively. Analysis of variance on the observed data and analysis of covariance adjusting changes to the initial mean value for the whole group showed significant differences between each exercise group and the controls but no difference between the exercise groups themselves. Both exercise groups showed a significant improvement in maximum oxygen uptake. This study suggests that exercise may modify bone loss in healthy postmenopausal women.


Influence of Delayed Conductance on Neuronal Synchronization.
In the brain, the excitation-inhibition balance prevents abnormal synchronous behavior. However, known synaptic conductance intensity can be insufficient to account for the undesired synchronization. Due to this fact, we consider time delay in excitatory and inhibitory conductances and study its effect on the neuronal synchronization. In this work, we build a neuronal network composed of adaptive integrate-and-fire neurons coupled by means of delayed conductances. We observe that the time delay in the excitatory and inhibitory conductivities can alter both the state of the collective behavior (synchronous or desynchronous) and its type (spike or burst). For the weak coupling regime, we find that synchronization appears associated with neurons behaving with extremes highest and lowest mean firing frequency, in contrast to when desynchronization is present when neurons do not exhibit extreme values for the firing frequency. Synchronization can also be characterized by neurons presenting either the highest or the lowest levels in the mean synaptic current. For the strong coupling, synchronous burst activities can occur for delays in the inhibitory conductivity. For approximately equal-length delays in the excitatory and inhibitory conductances, desynchronous spikes activities are identified for both weak and strong coupling regimes. Therefore, our results show that not only the conductance intensity, but also short delays in the inhibitory conductance are relevant to avoid abnormal neuronal synchronization.


Penalized likelihood estimation of a trivariate additive probit model.
This article proposes a penalized likelihood method to estimate a trivariate probit model, which accounts for several types of covariate effects (such as linear, nonlinear, random, and spatial effects), as well as error correlations. The proposed approach also addresses the difficulty in estimating accurately the correlation coefficients, which characterize the dependence of binary responses conditional on covariates. The parameters of the model are estimated within a penalized likelihood framework based on a carefully structured trust region algorithm with integrated automatic multiple smoothing parameter selection. The relevant numerical computation can be easily carried out using the SemiParTRIV() function in a freely available R package. The proposed method is illustrated through a case study whose aim is to model jointly adverse birth binary outcomes in North Carolina.


Dendritic nanotubes self-assembled from stiff polysaccharides as drug and probe carriers.
Dendritic nanotubes (DNTs) with hydrophobic cavities were constructed directly from rigid branched β-1,3-d-glucan (AF1) in aqueous solution, and the AF1 sample was isolated from the fruiting bodies of Auricularia auricula-judae, a household nutritional food. The structure of AF1 dendritic nanotubes was demonstrated with a transmission electron microscope (TEM) and a scanning electron microscope (SEM), and a schematic diagram was proposed to describe the formation process, which was supported by the results of static/dynamic light scattering (SLS/DLS) and atomic force microscopy (AFM). In solution, a sequential self-assembly of the AF1 chains in a parallel manner occurred to form lamellas followed by self-curling into nanotubes with the mean diameters from 20 to 80 nm, depending on the concentration and molecular weight of AF1, through hydrogen bonding and hydrophilic/hydrophobic interaction. As a result of the dendritic structure, the AF1 aggregates exhibited highly condensed hydrophobic regions, which could be used as carriers to achieve a high concentration of the target molecules. In our findings, the anticancer drug DOX and the fluorescent probe TPA-BMO could be loaded into the hydrophobic region of DNTs. Interestingly, DOX-loaded DNTs of AF1 exhibited high drug loading capacity and pH-triggered sustained release behaviors (>23 days) with reduced cytotoxicity in vitro. Moreover, the bioimaging experiment demonstrated that TPA-BMO-loaded DNTs of AF1 induced stronger fluorescence intensity than TPA-BMO alone, and maintained a longer duration time (18 days) in vivo. Therefore, the DNTs of AF1 have promising applications as bioactive carriers, especially in the fields of drug delivery and bioimaging.


Long persistent bcr-abl positive transcript detected by polymerase chain reaction after marrow transplant for chronic myelogenous leukemia without clinical relapse: a study of 64 patients.
We report here the results of polymerase chain reaction (PCR) for bcr-abl transcript and clinical details derived from 64 chronic myelogenous leukemia (CML) patients after allogeneic bone marrow transplantation (BMT). A total of 139 samples (2 to 220 weeks after BMT) were analyzed and bcr-abl transcript was detected in 99 samples from 52 patients. Patients were defined as bcr-abl early negative (EN) if they had > or = 1 negative PCR result < or = 1 year post-BMT (n = 13), and bcr-abl late positive (LP) if they had > or = 1 positive PCR result > or = 1 year post-BMT (n = 21). Among LP patients, only two patients had hematologic/cytogenetic (clinical) relapse. Another 19 LP patients remained in clinical remission 7 to 130 weeks after positive analysis for bcr-abl transcript, including 5 patients who had persistent bcr-abl transcript detectable even 2 years after BMT. To estimate the relationship between clinical data and residual bcr-abl transcript, EN patients are compared with LP patients. However, no clinical data studied were significantly associated with the persistent PCR positivity. If only patients in chronic phase are compared, the t-test showed significant correlation between leukocyte count just before BMT and sustained bcr-abl transcript (P < .05). These results suggest that PCR positivity is frequently observed in CML patients who sustain clinical remission after BMT, without being predictive of imminent clinical relapse. Tumor burden at the time of BMT may play an important role in the latency of bcr-abl positivity after BMT.


Development and Psychometric Validation of the FACE-Q Skin, Lips, and Facial Rhytids Appearance Scales and Adverse Effects Checklists for Cosmetic Procedures.
Patient-reported outcomes data are needed to determine the efficacy of cosmetic procedures. To describe the development and psychometric evaluation of 8 appearance scales and 2 adverse effect checklists for use in minimally invasive cosmetic procedures. We performed a psychometric study to select the most clinically sensitive items for inclusion in item-reduced scales and to examine reliability and validity with patients. Recruitment of the sample for this study took place from June 6, 2010, through July 28, 2014. Data analysis was performed from December 11, 2014, to December 22, 2015. Pretreatment and posttreatment patients 18 years and older who were consulting for any type of facial aesthetic treatment were studied. Patients were from plastic surgery and dermatology outpatient clinics in the United States and Canada (field-test sample) and a clinical trial of a minimally invasive lip treatment in the United Kingdom and France (clinical trial sample). The FACE-Q scales that measure appearance of the skin, lips, and facial rhytids (ie, overall, forehead, glabella, lateral periorbital area, lips, and marionette lines), with scores ranging from 0 (lowest) to 100 (highest), and the FACE-Q adverse effects checklists for problems after skin and lip treatment. Of 783 patients recruited, 503 field-test patients (response rate, 90%) and 280 clinical trial participants were studied. The mean (SD) age of the patients was 47.4 (14.0) years in the field-test sample and 47.7 (12.3) years in the clinical trial sample. Most of the patients were female (429 [85.3%] in the field-test sample and 274 [97.9%] in the clinical trial sample). Rasch Measurement Theory analyses led to the refinement of 8 appearance scales with 66 total items. All FACE-Q scale items had ordered thresholds and acceptable item fit. Reliability, measured with the Personal Separation Index (range, 0.88-0.95) and Cronbach α (range, 0.93-0.98), was high. Lower scores for appearance scales that measured the skin (r = -0.48, P < .001), lips (r = -0.21, P = .001), and lip rhytids (r = -0.32, P < .001) correlated with the reporting of more skin- and lip-related adverse effects. Higher scores for the 8 appearance scales correlated (range, 0.70-0.28; P < .001) with higher scores on the core 10-item FACE-Q satisfaction with facial appearance scale. In the pretreatment group, older age was significantly correlated with lower scores on 5 of the 6 rhytids scales (exception was forehead rhytids) (range, -0.28 to -0.65; P = .03 to <.001). Pretreatment patients reported significantly lower scores on 7 of the 8 appearance scales compared with posttreatment patients (exception was skin) (P < .001 to .005 on independent sample t tests). The FACE-Q appearance scales and adverse effects checklists can be used in clinical practice, research, and quality improvement to incorporate cosmetic patients' perspective in outcome assessments.


A powerful association test of multiple genetic variants using a random-effects model.
There is an emerging interest in sequencing-based association studies of multiple rare variants. Most association tests suggested in the literature involve collapsing rare variants with or without weighting. Recently, a variance-component score test [sequence kernel association test (SKAT)] was proposed to address the limitations of collapsing method. Although SKAT was shown to outperform most of the alternative tests, its applications and power might be restricted and influenced by missing genotypes. In this paper, we suggest a new method based on testing whether the fraction of causal variants in a region is zero. The new association test, T REM , is derived from a random-effects model and allows for missing genotypes, and the choice of weighting function is not required when common and rare variants are analyzed simultaneously. We performed simulations to study the type I error rates and power of four competing tests under various conditions on the sample size, genotype missing rate, variant frequency, effect directionality, and the number of non-causal rare variant and/or causal common variant. The simulation results showed that T REM was a valid test and less sensitive to the inclusion of non-causal rare variants and/or low effect common variants or to the presence of missing genotypes. When the effects were more consistent in the same direction, T REM also had better power performance. Finally, an application to the Shanghai Breast Cancer Study showed that rare causal variants at the FGFR2 gene were detected by T REM and SKAT, but T REM produced more consistent results for different sets of rare and common variants.


Efficacy of Ultrasound-Guided Serratus Plane Block on Postoperative Quality of Recovery and Analgesia After Video-Assisted Thoracic Surgery: A Randomized, Triple-Blind, Placebo-Controlled Study.
The optimal regional technique for analgesia and improved quality of recovery after video-assisted thoracic surgery (a procedure associated with considerable postoperative pain) has not been established. The main objective in this study was to compare quality of recovery in patients undergoing serratus plane block (SPB) with either ropivacaine or normal saline on the first postoperative day. Secondary outcomes were analgesic outcomes, including postoperative pain intensity and opioid consumption. Ninety patients undergoing video-assisted thoracic surgery were randomized to receive ultrasound-guided SPB with 0.4 mL/kg of either 0.375% ropivacaine (SPB group) or normal saline (control group) after anesthetic induction. The primary outcome was the 40-item Quality of Recovery (QoR-40) score at 24 hours after surgery. The QoR-40 questionnaire was completed by patients the day before surgery and on postoperative days 1 and 2. Pain scores, opioid consumption, and adverse events were assessed for 2 days postoperatively. Eighty-five patients completed the study: 42 in the SPB group and 43 in the control group. The global QoR-40 scores on both postoperative days 1 and 2 were significantly higher in the SPB group than in the control group (estimated mean difference 8.5, 97.5% confidence interval [CI], 2.1-15.0, and P = .003; 8.5, 97.5% CI, 2.0-15.1, and P = .004, respectively). The overall mean difference between the SPB and control groups was 8.5 (95% CI, 3.3-13.8; P = .002). Pain scores at rest and opioid consumption were significantly lower up to 6 hours after surgery in the SPB group than in the control group. Cumulative opioid consumption was significantly lower up to 24 hours postoperatively in the SPB group. Single-injection SPB with ropivacaine enhanced the quality of recovery for 2 days postoperatively and improved postoperative analgesia during the early postoperative period in patients undergoing video-assisted thoracic surgery.


Genetic control of mammalian meiotic recombination. I. Variation in exchange frequencies among males from inbred mouse strains.
Genetic background effects on the frequency of meiotic recombination have long been suspected in mice but never demonstrated in a systematic manner, especially in inbred strains. We used a recently described immunostaining technique to assess meiotic exchange patterns in male mice. We found that among four different inbred strains--CAST/Ei, A/J, C57BL/6, and SPRET/Ei--the mean number of meiotic exchanges per cell and, thus, the recombination rates in these genetic backgrounds were significantly different. These frequencies ranged from a low of 21.5 exchanges in CAST/Ei to a high of 24.9 in SPRET/Ei. We also found that, as expected, these crossover events were nonrandomly distributed and displayed positive interference. However, we found no evidence for significant differences in the patterns of crossover positioning between strains with different exchange frequencies. From our observations of >10,000 autosomal synaptonemal complexes, we conclude that achiasmate bivalents arise in the male mouse at a frequency of 0.1%. Thus, special mechanisms that segregate achiasmate chromosomes are unlikely to be an important component of mammalian male meiosis.


Quality of life of patients with chronic lymphocytic leukaemia in the Netherlands: results of a longitudinal multicentre study.
To describe the health-related quality of life (HRQoL) of an unselected population of patients with chronic lymphocytic leukaemia (CLL) including untreated patients. HRQoL was measured by the EORTC QLQ-C30 including the CLL16 module, EQ-5D, and VAS in an observational study over multiple years. All HRQoL measurements per patient were connected and analysed using area under the curve analysis over the entire study duration. The total patient group was compared with the general population, and three groups of CLL patients were described separately, i.e. patients without any active treatment ("watch and wait"), chlorambucil treatment only, and patients with other treatment(s). HRQoL in the total group of CLL patients was compromised when compared with age- and gender-matched norm scores of the general population. CLL patients scored statistically worse on the VAS and utility score of the EQ-5D, all functioning scales of the EORTC QLQ-C30, and the symptoms of fatigue, dyspnoea, sleeping disturbance, appetite loss, and financial difficulties. In untreated patients, the HRQoL was slightly reduced. In all treatment stages, HRQoL was compromised considerably. Patients treated with chlorambucil only scored worse on the EORTC QLQ-C30 than patients who were treated with other treatments with regard to emotional functioning, cognitive functioning, bruises, uncomfortable stomach, and apathy. CLL patients differ most from the general population on role functioning, fatigue, concerns about future health, and having not enough energy. Once treatment is indicated, HRQoL becomes considerably compromised. This applies to all treatments, including chlorambucil, which is considered to be a mild treatment.


Isometric contraction of arm flexor muscles as a method of evaluating cardiac vagal tone in man.
1. We have previously shown that brief voluntary isometric contractions of upper arm flexor muscles performed for one respiratory cycle elicit a significant decrease in the R-R interval. The present study was designed to determine if similar changes are produced by non-voluntary electrically evoked contractions and, if so, to establish the consistency and repeatability of the associated changes in the R-R interval. 2. The heart rate (R-R interval) response to voluntary or non-voluntary brief isometric contraction equivalent to 40% of the maximum voluntary contraction was studied in 10 healthy young male subjects during controlled ventilation at supine rest. 3. The absolute values of R-R intervals occurring in any one of 10 arbitrary phases of a respiratory cycle were measured and plotted by a computer. 4. Both voluntary and non-voluntary contractions elicited similar changes in heart rate and R-R interval, which were greater during expiration than during inspiration. 5. This confirms our previous finding that the magnitude of the R-R interval changes, with brief isometric contraction, is positively related to the degree of cardiac vagal tone. 6. Analysis of the variability between repeated tests initiated in either inspiration or expiration revealed that there was significantly less variability with the electrically induced contraction. 7. It was concluded that electrically induced contractions of 40% maximal voluntary contraction are a viable alternative to voluntary contractions and provide a more controllable means of measuring cardiac vagal withdrawal.


Regulator soluble factors of human IgE synthesis. I. Production and isolation.
The system of IgE response undoubtedly has a special interest, given the role of this immunoglobulin on the pathogenesis of allergic diseases. In the last few years the existence of a regulator mechanism that maintains the synthesis of IgE in low but effective levels has been confirmed. This mechanism consists in the equilibrium between the actions of suppressors and enhancers. These two soluble factors present in the serum and capable of suppressing and enhancing IgE response in control rats have been identified; both have a highly selective activity on IgE, exerting their effect in an antigen-independent form and require specie-specificity. Motivated by these findings, we have applied this experimental system used in rats to humans. Healthy controls were selected as low responders of IgE, atopics as high responders, and asthmatic pollinosis considered as the prototype of allergic disease mediated by IgE. After obtaining the factors mentioned by means of a bi-directional mixed culture in a pool of lymphocytes, we proceeded to their isolation by affinity chromatography in Con-A Sepharose columns. The glycoproteins obtained were afterwards submitted to different techniques of characterization: immunoelectrophoresis in polycrilamide gel, chromatography technique, etc.


Barriers to the use of emergency medical services for ST-elevation myocardial infarction: Determining why many patients opt for self-transport.
Access to timely ST-elevation myocardial infarction (STEMI) care is facilitated by paramedics and emergency medical services (EMS). However, a large proportion of STEMI patients do not access care through EMS. This study sought to identify patient-reported factors for their decision to use (or not use) EMS. Semi-structured interviews were conducted with a sample of STEMI patients admitted to a large tertiary care centre between November 2011 and January 2012. Participants were grouped according to mode of transportation to hospital at time of index event (EMS vs self-transport). Participant responses were classified using a published framework (modified for a STEMI population) as barriers or facilitators to EMS use, and compared between groups. Data were collected on 61 patients (32 EMS, 29 self-transport). Mean age was 60.3 (SD 11.5), and 23% were female. EMS users were more likely to have a Killip Class >1 (25% vs 4%; P = 0.03). Self-transport patients were more likely to perceive EMS as slower (48% vs 0%) and express concerns over resources misuse (34% vs 3%; P = 0.002), when compared to EMS patients. Patients who accessed EMS were more likely to acknowledge the benefits of EMS (44% vs 7%; P = 0.001) and were more likely to have been encouraged by a family member to call EMS (34% vs 4%; P = 0.003). STEMI patient perceptions are a key factor in determining EMS use. Health care stakeholders should target the identified barriers to improve utilization of EMS, and develop strategies to optimize care for patients who do not access EMS.


Genome sequencing in microfabricated high-density picolitre reactors.
The proliferation of large-scale DNA-sequencing projects in recent years has driven a search for alternative methods to reduce time and cost. Here we describe a scalable, highly parallel sequencing system with raw throughput significantly greater than that of state-of-the-art capillary electrophoresis instruments. The apparatus uses a novel fibre-optic slide of individual wells and is able to sequence 25 million bases, at 99% or better accuracy, in one four-hour run. To achieve an approximately 100-fold increase in throughput over current Sanger sequencing technology, we have developed an emulsion method for DNA amplification and an instrument for sequencing by synthesis using a pyrosequencing protocol optimized for solid support and picolitre-scale volumes. Here we show the utility, throughput, accuracy and robustness of this system by shotgun sequencing and de novo assembly of the Mycoplasma genitalium genome with 96% coverage at 99.96% accuracy in one run of the machine.


Spatial imaging of carbon reactivity centers in Pd/C catalytic systems.
Gaining insight into Pd/C catalytic systems aimed at locating reactive centers on carbon surfaces, revealing their properties and estimating the number of reactive centers presents a challenging problem. In the present study state-of-the-art experimental techniques involving ultra high resolution SEM/STEM microscopy (1 Å resolution), high brilliance X-ray absorption spectroscopy and theoretical calculations on truly nanoscale systems were utilized to reveal the role of carbon centers in the formation and nature of Pd/C catalytic materials. Generation of Pd clusters in solution from the easily available Pd2dba3 precursor and the unique reactivity of the Pd clusters opened an excellent opportunity to develop an efficient procedure for the imaging of a carbon surface. Defect sites and reactivity centers of a carbon surface were mapped in three-dimensional space with high resolution and excellent contrast using a user-friendly nanoscale imaging procedure. The proposed imaging approach takes advantage of the specific interactions of reactive carbon centers with Pd clusters, which allows spatial information about chemical reactivity across the Pd/C system to be obtained using a microscopy technique. Mapping the reactivity centers with Pd markers provided unique information about the reactivity of the graphene layers and showed that >2000 reactive centers can be located per 1 μm2 of the surface area of the carbon material. A computational study at a PBE-D3-GPW level differentiated the relative affinity of the Pd2 species to the reactive centers of graphene. These findings emphasized the spatial complexity of the carbon material at the nanoscale and indicated the importance of the surface defect nature, which exhibited substantial gradients and variations across the surface area. The findings show the crucial role of the structure of the carbon support, which governs the formation of Pd/C systems and their catalytic activity.


Interventions Targeting Glucocorticoid-Krüppel-like Factor 15-Branched-Chain Amino Acid Signaling Improve Disease Phenotypes in Spinal Muscular Atrophy Mice.
The circadian glucocorticoid-Krüppel-like factor 15-branched-chain amino acid (GC-KLF15-BCAA) signaling pathway is a key regulatory axis in muscle, whose imbalance has wide-reaching effects on metabolic homeostasis. Spinal muscular atrophy (SMA) is a neuromuscular disorder also characterized by intrinsic muscle pathologies, metabolic abnormalities and disrupted sleep patterns, which can influence or be influenced by circadian regulatory networks that control behavioral and metabolic rhythms. We therefore set out to investigate the contribution of the GC-KLF15-BCAA pathway in SMA pathophysiology of Taiwanese Smn-/-;SMN2 and Smn2B/- mouse models. We thus uncover substantial dysregulation of GC-KLF15-BCAA diurnal rhythmicity in serum, skeletal muscle and metabolic tissues of SMA mice. Importantly, modulating the components of the GC-KLF15-BCAA pathway via pharmacological (prednisolone), genetic (muscle-specific Klf15 overexpression) and dietary (BCAA supplementation) interventions significantly improves disease phenotypes in SMA mice. Our study highlights the GC-KLF15-BCAA pathway as a contributor to SMA pathogenesis and provides several treatment avenues to alleviate peripheral manifestations of the disease. The therapeutic potential of targeting metabolic perturbations by diet and commercially available drugs could have a broader implementation across other neuromuscular and metabolic disorders characterized by altered GC-KLF15-BCAA signaling.


Acute coronary syndrome (Takotsubo cardiomyopathy) following electroconvulsive therapy in the absence of significant coronary artery disease: case report and review of the literature.
We report a case of myocardial infarction occurring in a 45-year-old woman in the absence of coronary artery disease during a course of electroconvulsive therapy (ECT) for major depression. After the third ECT session, the patient reported substernal chest pain, and although the electrocardiogram was normal, cardiac enzymes were found to be elevated. Cardiac workup to determine etiology during hospital stay showed no evidence of coronary artery disease on catheterization. Cardiac echocardiograph and computed tomography findings were consistent with a diagnosis of Takotsubo cardiomyopathy. This cardiac syndrome is a stress-induced syndrome occurring in the absence of coronary artery stenosis and associated with surging of catecholamines. It is diagnosed on the basis of the shape of the apical ballooning observed on cardiac imaging, which resembles a "takotsubo" or octopus fishing pot in Japan, where this syndrome was first recognized. Coincidentally, after termination of her ECT course, this patient was also diagnosed with Loeys-Dietz syndrome, an inherited connective tissue disorder that can predispose sufferers to aortic dissection. This case illustrates the importance of careful workup of chest pain in the setting of ECT, even if electrocardiogram findings seem unremarkable initially.


Differences in perception of dysentery and enteric fever and willingness to receive vaccines among rural residents in China.
Enteric diseases including dysentery and enteric fever remain significant public health problems in China. While vaccines offer great potential in controlling these diseases, greater understanding of factors influencing acceptance of vaccines is needed to create effective enteric disease control programs in rural China. Cross-sectional quantitative study with randomly sampled households from two sites in China, one experiencing high rates of shigellosis (Zengding) and the other of typhoid/paratyphoid (Lingchuan). Sociobehavioral survey data were collected through face-to-face interviews from 501 respondents (56% female) in Zhengding regarding dysentery and 624 in Lingchuan (51% female) regarding enteric fever. Vaccine acceptability was measured by expressed need for vaccination and willingness to pay. Comparative and associative analyses were conducted to assess disease perception, vaccination service satisfaction, likelihood of improvements in water and sanitation, and vaccine acceptability. Nearly all respondents in Lingchuan considered enteric fever to be prevalent in the community, while only one half of the respondents in Zhengding considered dysentery to be problematic (p < 0.01). Nevertheless, more respondents in Zhengding were fearful that a household member would acquire dysentery than were Lingchuan respondents worried that a household member would acquire enteric fever (p < 0.01). Perceived vulnerability of specific subgroups (odds ratios ranging from 1.6 to 8.1), knowing someone who died of the disease (odds ratio reached infinity) and satisfaction with past vaccination services (odds ratios reached infinity) were consistently associated with perceived need for vaccines of target populations of all age groups while the association between perception of sanitary improvement and vaccine need was limited. Perceived need for a vaccine was associated with willingness to pay for the vaccine. Perceptions of enhanced vulnerability of specific subgroups to a disease and satisfactory experiences with vaccination services may increase the perceived need for a vaccine, leading to increased willingness to pay for vaccine. Vaccines are not perceived as important for the elderly.


Neural basis for motor learning in the vestibuloocular reflex of primates. II. Changes in the responses of horizontal gaze velocity Purkinje cells in the cerebellar flocculus and ventral paraflocculus.
1. We made extracellular recordings from Purkinje cells in the flocculus and ventral paraflocculus of awake monkeys before and after motor learning in the vestibuloocular reflex (VOR). Three samples were recorded 1) after miniaturizing spectacles had reduced the gain of the VOR (eye speed divided by head speed) to 0.4; 2) when the gain of the VOR was near 1.0; and 3) after magnifying spectacles had increased the gain of the VOR to 1.6. 2. We studied Purkinje cells that showed stronger modulation of simple-spike firing rate during horizontal than during vertical pursuit. These cells corresponded to the previously identified "horizontal gaze velocity Purkinje cells" or HGVP-cells. During pursuit of smooth target motion with the head stationary, HGVP-cells showed strong modulation of firing rate with increases for ipsiversive eye motion (toward the side of recording). When the monkey canceled his VOR by tracking a target that moved exactly with him during sinusoidal head rotation in the horizontal plane, HGVP-cells again showed strong modulation of firing rate with increases for ipsiversive head motion. 3. The responses of HGVP-cells during pursuit with the head stationary and during cancellation of the VOR reveal separate components of firing rate related to eye and head velocity. We used these two behavioral conditions to test for effects of motor learning on the head and eye velocity components of the simple-spike firing of HGVP-cells. Our data confirm the previous observation that motor learning causes the sensitivity to head velocity to be larger when the gain of the VOR is high and smaller when the gain of the VOR is low. Thus we agree with the previous conclusion that changes in the vestibular sensitivity of HGVP-cells, measured during sinusoidal head motion at low frequencies, are in the wrong direction to cause changes in the gain of the VOR. 4. To determine whether the simple-spike output from the HGVP-cells plays a role in the VOR after motor learning, we recorded simple-spike firing during the VOR evoked by transient, rapid changes in head velocity in darkness. When the gain of the VOR was low, firing rate increased during the VOR evoked by ipsiversive head motion and decreased during the VOR evoked by contraversive head motion. When the gain of the VOR was high, the direction selectivity of the responses was reversed.(ABSTRACT TRUNCATED AT 400 WORDS)


Effects of pelargonidin-3-O-glucoside and its metabolites on lipopolysaccharide-stimulated cytokine production by THP-1 monocytes and macrophages.
Epidemiological evidence suggests cardioprotective effects of anthocyanin consumption. This study examined the predominant strawberry anthocyanin, pelargonidin-3-O-glucoside (Pg-3-glc), and three of its plasma metabolites (protocatechuic acid [PCA], 4-hydroxybenzoic acid, and phloroglucinaldehyde [PGA]) for effects on the production of selected cytokines by lipopolysaccharide-stimulated THP-1 monocytes and macrophages. Concentrations of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, IL-8 and IL-10 were determined using a cytometric bead array kit. PCA at 0.31, 1.25 and 20 μM and PGA at 5 and 20 μM decreased the concentration of IL-6 in the monocyte cultures, but there were no effects on TNF-α, IL-1β, IL-8 and IL-10 and there were no effects of the other compounds. In the macrophage cultures, PGA at 20 μM decreased the concentrations of IL-6 and IL-10, but there was no effect on TNF-α, IL-1β and IL-8 and there were no effects of the other compounds. In conclusion, while the effects of PGA were only observed at the higher, supraphysiological concentration and are thus considered of limited physiological relevance overall, the anti-inflammatory properties of PCA were observed at both the lower, physiologically relevant, and the higher concentrations; however, effects were modest and limited to IL-6 and monocytes. These preliminary data suggest potential for physiologically attainable PCA concentrations to modulate IL-6 production by monocytes.


Role of Borrelia burgdorferi linear plasmid 25 in infection of Ixodes scapularis ticks.
The tick-borne bacterium Borrelia burgdorferi has over 20 different circular and linear plasmids. Some B. burgdorferi plasmids are readily lost during in vitro culture or genetic manipulation. Linear plasmid 25, which is often lost in laboratory strains, is required for the infection of mice. Strains missing linear plasmid 25 (lp25(-)) are able to infect mice if the BBE22 gene on lp25 is provided on a shuttle vector. In this study, we examined the role of lp25 and BBE22 in tick infections. We tested the hypothesis that complementation with BBE22 in spirochetes lacking lp25 would restore the ability of spirochetes to infect ticks. A natural tick infection cycle was performed by feeding larvae on mice injected with the parental, lp25(-), or lp25(-) BBE22-complemented spirochete strains. In addition, larvae and nymphs were artificially infected with different strains to study tick infections independent of mouse infections. B. burgdorferi missing lp25 was significantly impaired in its ability to infect larval and nymphal ticks. When an lp25(-) strain was complemented with BBE22, the ability to infect ticks was partially restored. Complementation with BBE22 allowed spirochetes lacking lp25 to establish short-term infections in ticks, but in most cases the infection prevalence was lower than that of the wild-type strain. In addition, the number of infected ticks decreased over time, suggesting that another gene(s) on lp25 is required for long-term persistence in ticks and completion of a natural infection cycle.


Discoloration of Resin Composites Induced by Coffee and Tomato Sauce and Subjected to Surface Polishing: An In Vitro Study.
BACKGROUND The purpose of this study is to evaluate and compare color stability of 3 resin based restorative materials when immersed into staining media at 0, 3, 30, 45, 60, and 75 days, and to assess the efficacy of surface polishing in reducing possible discoloration. MATERIAL AND METHODS Ninety composite discs were prepared in a custom-made mold (5 mm diameter and 2 mm thick) from 3 different light cured composites (Filtek™ Z250, Harmonize™, and G-aenial). Color differences of all specimens were measured by a spectrophotometer according to the CIE L*a*b* system. After baseline color measurements, 30 discs from each composite group were randomly divided into 3 subgroups of 10 specimens each. Subgroups (n=10) were immersed for 75 days into different staining solutions: coffee, tomato sauce, and distilled water (control). Solutions were changed every week and color measurements were repeated every 15 days. At day 75, color measurements of the specimens were performed before and after surface polishing with aluminum oxide discs. The experimental data were statistically evaluated using repeated measures one-way ANOVA test followed by Tukey's multiple pairwise comparison with a significance level of 5%. RESULTS Of the 3 composites, G-aenial reported the highest color change deviation when immersed into coffee (ΔE=8.674), and tomato sauce (ΔE=7.737) at day 75, followed by Harmonize that also exhibited a significant difference for coffee (ΔE=4.7) and tomato sauce (ΔE=3.8) when compared to distilled water. While Filtek™ Z250 did not show any significant difference between the 3 storage solutions (P>0.05). Only G-aenial had significant color change (P<0.05) after polishing with aluminum oxide discs for all tested samples, whereas Filtek™ Z250 and Harmonize presented no significant difference after surface polishing (P>0.05). CONCLUSIONS Under the tested experimental conditions, Filtek™ Z250, among the 3 resin-based composites, exhibited the highest color stability when subjected to coffee and tomato sauce as well. Whereas, G-aenial presented the highest color deviation when immersed in both staining media. Surface polishing effectively reduced coffee and tomato sauce discolorations for G-aenial's specimens and had no significant effect regarding Filtek™ Z250 and Harmonize.


Bone forming capacity of cell- and growth factor-based constructs at different ectopic implantation sites.
The aim of this study was to compare the effect of implantation site (i.e., subcutaneous, SQ vs. intramuscular, IM) on bone forming capacity of cell-based and growth factor-based scaffolds in athymic nude rats after an implantation period of 8 weeks. Cell-based scaffolds consisted of porous hydroxyapatite/tricalcium phosphate (HA/TCP) scaffolds seeded with either human adipose tissue-derived mesenchymal stem cells (AT-MSCs) only or both AT-MSCs and human umbilical vein endothelial cells (HUVECs), which were precultured in osteogenic medium for 7 days. Growth factor-based scaffolds consisted of porous HA/TCP scaffolds with 20 µg preadsorbed bone morphogenetic protein-2 (BMP-2). Histological and histomorphometrical analysis were used to assess bone formation. A differentiation experiment was performed in parallel to compare the in vitro osteogenic capacity of cell-based scaffolds. The results showed that cell-based scaffolds showed evident osteogenic differentiation in vitro, with only marginal differences between AT-MSCs only and AT-MSCs/HUVECs. In vivo, none of the cell-based scaffolds showed bone formation, irrespective of the site of implantation. In contrast, all growth factor-based scaffolds showed bone formation at both implantation sites without differences in the amount of formed bone. In conclusion, the results of this study demonstrated that the bone forming capacity of HA/TCP scaffolds with pre-adsorbed BMP-2 was equal at different ectopic implantation sites. Further, despite obvious in vitro osteogenic differentiation of AT-MSCs and AT-MSCS/HUVECs on HA/TCP scaffolds, no bone formation of these cell-based scaffolds was observed in vivo. This indicates further investigation on bone formation mechanisms of AT-MSCs is needed before AT-MSCs can be used as a cytotherapeutic treatment in clinics.


The Macrophage: A Disputed Fortress in the Battle against Mycobacterium tuberculosis.
Mycobacterium tuberculosis (Mtb), the etiological agent of human tuberculosis (TB), has plagued humans for thousands of years. TB still remains a major public health problem in our era, causing more than 4,400 deaths worldwide every day and killing more people than HIV. After inhaling Mtb-contaminated aerosols, TB primo-infection starts in the terminal lung airways, where Mtb is taken up by alveolar macrophages. Although macrophages are known as professional killers for pathogens, Mtb has adopted remarkable strategies to circumvent host defenses, building suitable conditions to survive and proliferate. Within macrophages, Mtb initially resides inside phagosomes, where its survival mostly depends on its ability to take control of phagosomal processing, through inhibition of phagolysosome biogenesis and acidification processes, and by progressively getting access to the cytosol. Bacterial access to the cytosolic space is determinant for specific immune responses and cell death programs, both required for the replication and the dissemination of Mtb. Comprehension of the molecular events governing Mtb survival within macrophages is fundamental for the improvement of vaccine-based and therapeutic strategies in order to help the host to better defend itself in the battle against the fierce invader Mtb. In this mini-review, we discuss recent research exploring how Mtb conquers and transforms the macrophage into a strategic base for its survival and dissemination as well as the associated defense strategies mounted by host.


Validation of a spectral light scattering method to differentiate large from small particles in intraocular lenses.
A psychophysical approach has been designed to measure straylight from intraocular lenses (IOLs) in vitro. This approach uses a clinical straylight meter (C-Quant) and an observer's eye as optical detector. Based on this, we introduced a method for study of straylight-wavelength dependency for IOLs. This dependency can be used to distinguish between 2 types of scattering particles (small and large) as defined by Mie theory. Validation was performed using a turbidity standard and scattering filters. Several IOLs were analyzed to identify potential scattering sources. Large particles were found to predominate in scattering from the studied lenses. This was confirmed by straylight-angular dependency found in these IOLs.


RhoA protects the mouse heart against ischemia/reperfusion injury.
The small GTPase RhoA serves as a nodal point for signaling through hormones and mechanical stretch. However, the role of RhoA signaling in cardiac pathophysiology is poorly understood. To address this issue, we generated mice with cardiomyocyte-specific conditional expression of low levels of activated RhoA (CA-RhoA mice) and demonstrated that they exhibited no overt cardiomyopathy. When challenged by in vivo or ex vivo ischemia/reperfusion (I/R), however, the CA-RhoA mice exhibited strikingly increased tolerance to injury, which was manifest as reduced myocardial lactate dehydrogenase (LDH) release and infarct size and improved contractile function. PKD was robustly activated in CA-RhoA hearts. The cardioprotection afforded by RhoA was reversed by PKD inhibition. The hypothesis that activated RhoA and PKD serve protective physiological functions during I/R was supported by several lines of evidence. In WT mice, both RhoA and PKD were rapidly activated during I/R, and blocking PKD augmented I/R injury. In addition, cardiac-specific RhoA-knockout mice showed reduced PKD activation after I/R and strikingly decreased tolerance to I/R injury, as shown by increased infarct size and LDH release. Collectively, our findings provide strong support for the concept that RhoA signaling in adult cardiomyocytes promotes survival. They also reveal unexpected roles for PKD as a downstream mediator of RhoA and in cardioprotection against I/R.


A question of balance: achieving appropriate nutrient levels in biofortified staple crops.
The biofortification of staple crops with vitamins is an attractive strategy to increase the nutritional quality of human food, particularly in areas where the population subsists on a cereal-based diet. Unlike other approaches, biofortification is sustainable and does not require anything more than a standard food-distribution infrastructure. The health-promoting effects of vitamins depend on overall intake and bioavailability, the latter influenced by food processing, absorption efficiency and the utilisation or retention of the vitamin in the body. The bioavailability of vitamins in nutritionally enriched foods should ideally be adjusted to achieve the dietary reference intake in a reasonable portion. Current vitamin biofortification programmes focus on the fat-soluble vitamins A and E, and the water-soluble vitamins C and B9 (folate), but the control of dosage and bioavailability has been largely overlooked. In the present review, we discuss the vitamin content of nutritionally enhanced foods developed by conventional breeding and genetic engineering, focusing on dosage and bioavailability. Although the biofortification of staple crops could potentially address micronutrient deficiency on a global scale, further research is required to develop effective strategies that match the bioavailability of vitamins to the requirements of the human diet.


Insulin self-administration technique with disposable syringe among patients with diabetes mellitus followed by the family health strategy.
This cross-sectional study aimed to describe the most common correct and incorrect self-administration techniques for insulin using disposable syringes by patients cared for by the Family Health Strategy (FHS), relate the findings to sociodemographic variables and also identify the professional responsible for teaching this technique. A total of 169 patients were selected by simple random sampling in 37 FHS units in a city in the state of Minas Gerais, Brazil from August to October 2006. The results identified errors in all the steps recommended by the American Diabetes Association and Brazilian Diabetes Association for the safe administration of insulin, from hand washing to compression on the injection site. The FHS favors the development of interventions focused on the needs of the clientele registered at the unit, stimulating self-care. Results from this study can contribute to the planning of these interventions.


L-glutamine with D-glucose stimulates oxidative metabolism and NaCl absorption in piglet jejunum.
To explore the relationship between intestinal fluid absorption and oxidative metabolism, we measured the effects of amino acids and glucose on piglet jejunal ion transport and oxygen consumption (QO2) in vitro. Jejunal QO2 was stimulated by L-glutamine and D-glucose but not by the nonmetabolizable organic solutes methyl beta-D-glucoside or L-phenylalanine. QO2 was maximally enhanced by the combination of D-glucose and L-glutamine (5 mM). Even though 5 mM L-glutamine was previously found to be insufficient to stimulate NaCl absorption, 5 mM L-glutamine enhanced jejunal NaCl flux when combined with equimolar mucosal D-glucose. Either D-glucose or methyl beta-D-glucoside caused an increase in short-circuit current (Isc), an increase in Na+ absorption in excess of Isc, and a decrease in Cl- secretion, when L-glutamine was substituted for D-glucose (10 mM) on the serosal side. This relationship suggests that mucosal sugars, if combined with L-glutamine, enhance neutral NaCl absorption as well as electrogenic Na+ flow. (Aminooxy)acetate, an inhibitor of alanine aminotransferase, abolished the stimulation of QO2 and the NaCl-absorptive response to L-glutamine. We conclude that the oxidative metabolism fueled by L-glutamine is linked to a NaCl-absorptive mechanism in the intestine. We propose that the CO2 produced by glutamine metabolism yields carbonic acid, which dissociates to H+ and HCO3-, which may stimulate parallel antiports in the apical membrane.


Systematic review with network meta-analysis: the efficacy of anti-TNF agents for the treatment of Crohn's disease.
Anti-tumour necrosis factor-alpha agents (anti-TNF) are effective therapies for the treatment of Crohn's disease (CD), but their comparative efficacy is unknown. To perform a network meta-analysis comparing the efficacy of anti-TNF therapies in CD. After screening 506 studies, reviewers extracted information on 10 studies. Traditional meta-analysis (TMA) was used to compare each anti-TNF agent to placebo. Bayesian network meta-analysis (NMA) was performed to compare the effects of anti-TNF agents to placebo. In addition, sample sizes for comparative efficacy trials were calculated. Compared to placebo, TMA revealed that anti-TNF agents result in a higher likelihood of induction of remission and response (RR: 1.66, 95% CI: 1.17-2.36 and RR: 1.43, 95% CI: 1.17-1.73, respectively) as well as maintenance of remission and response (RR: 1.78, 95% CI: 1.51-2.09 and RR: 1.68, 95% CI: 1.46-1.93, respectively). NMA found nonsignificant trends between infliximab and adalimumab or certolizumab pegol. Among subcutaneous therapies, NMA demonstrated superiority of adalimumab to certolizumab pegol for induction of remission (RR: 2.93, 95% CrI: 1.21-7.75). Sample size calculations suggest that adequately powered head-to-head comparative efficacy trials would require greater than 3000 patients. All anti-TNF agents are effective for induction and maintenance of response and remission in the treatment of CD. Although adalimumab is superior to certolizumab pegol for induction of remission, there is no evidence of clinical superiority among anti-TNF agents. Head-to-head trials among the anti-TNF agents are impractical in terms of size and cost.


Creation of highly aligned electrospun poly-L-lactic acid fibers for nerve regeneration applications.
Aligned, electrospun polymer fibers have shown considerable promise in directing regenerating axons in vitro and in vivo. However, in several studies, final electrospinning parameters are presented for producing aligned fiber scaffolds, and alignment where minimal fiber crossing occurs is not achieved. Highly aligned species are necessary for neural tissue engineering applications to ensure that axonal extension occurs through a regenerating environment efficiently. Axonal outgrowth on fibers that deviate from the natural axis of growth may delay axonal extension from one end of a scaffold to the other. Therefore, producing aligned fiber scaffolds with little fiber crossing is essential. In this study, the contributions of four electrospinning parameters (collection disk rotation speed, needle size, needle tip shape and syringe pump flow rate) were investigated thoroughly with the goal of finding parameters to obtain highly aligned electrospun fibers made from poly-L-lactic acid (PLLA). Using an 8 wt% PLLA solution in chloroform, a collection disk rotation speed of 1000 revolutions per minute (rpm), a 22 gauge, sharp-tip needle and a syringe pump rate of 2 ml h(-1) produced highly aligned fiber (1.2-1.6 microm in diameter) scaffolds verified using a fast Fourier transform and a fiber alignment quantification technique. Additionally, the application of an insulating sheath around the needle tip improved the rate of fiber deposition (electrospinning efficiency). Optimized scaffolds were then evaluated in vitro using embryonic stage nine (E9) chick dorsal root ganglia (DRGs) and rat Schwann cells (SCs). To demonstrate the importance of creating highly aligned scaffolds to direct neurite outgrowth, scaffolds were created that contained crossing fibers. Neurites on these scaffolds were directed down the axis of the aligned fibers, but neurites also grew along the crossed fibers. At times, these crossed fibers even stopped further axonal extension. Highly aligned PLLA fibers generated under optimized electrospinning conditions guided neurite and SC growth along the aligned fibers. Schwann cells demonstrated the bipolar phenotype seen along the fibers. Using a novel technique to determine fiber density, an increase in fiber density correlated to an increase in the number of neurites, but average neurite length was not statistically different between the two different fiber densities. Together, this work presents methods by which to produce highly aligned fiber scaffolds efficiently and techniques for assessing neurite outgrowth on different fiber scaffolds, while suggesting that crossing fibers may be detrimental in fostering efficient, directed axonal outgrowth.


Reactivation of Chagas' disease in a human immunodeficiency virus-infected patient leading to severe heart disease with a late positive direct microscopic examination of the blood.
We report a human immunodeficiency virus (HIV)-infected man with chronic Chagas' disease who developed a congestive heart failure that could not be clinically controlled. Endomyocardial biopsy revealed severe myocarditis and the xenodiagnosis result was positive, but Trypanosoma cruzi by direct microscopic examination of the blood was found only four months after the symptoms had started. Treatment with benznidazole was effective in reducing parasitemia, stabilizing the clinical status, and controlling tissue damage related to the parasite. Although the finding of T. cruzi trypomastigotes by direct microscopic examination of the blood has been considered the mark of Chagas' reactivation in immunocompromised patients with chronic disease, in this case it was a late finding.


Hypothalamic stem cells control ageing speed partly through exosomal miRNAs.
It has been proposed that the hypothalamus helps to control ageing, but the mechanisms responsible remain unclear. Here we develop several mouse models in which hypothalamic stem/progenitor cells that co-express Sox2 and Bmi1 are ablated, as we observed that ageing in mice started with a substantial loss of these hypothalamic cells. Each mouse model consistently displayed acceleration of ageing-like physiological changes or a shortened lifespan. Conversely, ageing retardation and lifespan extension were achieved in mid-aged mice that were locally implanted with healthy hypothalamic stem/progenitor cells that had been genetically engineered to survive in the ageing-related hypothalamic inflammatory microenvironment. Mechanistically, hypothalamic stem/progenitor cells contributed greatly to exosomal microRNAs (miRNAs) in the cerebrospinal fluid, and these exosomal miRNAs declined during ageing, whereas central treatment with healthy hypothalamic stem/progenitor cell-secreted exosomes led to the slowing of ageing. In conclusion, ageing speed is substantially controlled by hypothalamic stem cells, partially through the release of exosomal miRNAs.


GRASP65, a protein involved in the stacking of Golgi cisternae.
NEM prevents mitotic reassembly of Golgi cisternae into stacked structures. The major target of NEM is a 65 kDa protein conserved from yeast to mammals. Antibodies to this protein and a recombinant form of it block cisternal stacking in a cell-free system, justifying its designation as a Golgi ReAssembly Stacking Protein (GRASP65). One of the two minor targets of NEM is GM130, previously implicated in the docking of transport vesicles and mitotic fragmentation of the Golgi stack. GRASP65 is complexed with GM130 and is tightly bound to Golgi membranes, even under mitotic conditions when both are heavily phosphorylated. These results link vesicle docking, stacking of Golgi cisternae, and the disruption of both of these interactions during mitosis.


Combinations of susceptible genotypes and individual responses to toxicants.
The variation in individual responses to exogenous agents has been shown to be exceptionally wide. It is because of this large diversity of responsiveness that risk factors to environmentally induced diseases have been difficult to pinpoint, particularly at low exposure levels. Opportunities now exist for studies of host factors in environmentally induced cancer or other diseases in which an environmental component can be presumed. Many of the studies have shown an elevated disease proneness for individuals carrying the potential at-risk alleles of metabolic genes, but a number of controversial results have also been reported. One possible explanation for the divergent findings is lack of knowledge of the other potentially relevant genotypes for a given exposure. This paper gives an overview of the published data on combinations of metabolic genotypes in relation to individual susceptibility to environmental toxicants.


Improved restoration from multiple images of a single object: application to fluorescence microscopy.
We present an approach for the combined restoration of multiple different images of a single object. A linear Tikhonov filter adapted for this purpose is derived in detail. Nonlinear constrained algorithms can also be adapted, and we illustrate this possibility for an iterative constrained Tikhonov algorithm. Both the linear and the iterative constrained Tikhonov algorithms were used to analyze performance in fluorescence confocal imaging by use of simulated and experimental data. One can improve the quality of restored confocal images significantly if the signal that normally is rejected by the detection pinhole of a confocal laser scanning microscope is also recorded on a separate detector such that the two recorded signals are used together for image restoration according to the proposed algorithms.


Recognition of viral glycoproteins by influenza A-specific cross-reactive cytolytic T lymphocytes.
Two populations of cytolytic T lymphocytes (CTL) generated after influenza A virus infection can be distinguished into one with specificity for the sensitizing hemagglutinin type and a second with cross-reactivity for antigens induced by other type-A influenza viruses. The molecules carrying the antigenic determinants recognized by the cross-reactive CTL were studied. In L-929 cells abortively infected with fowl plague virus, matrix (M) protein synthesis is specifically inhibited, whereas the envelope glycoproteins, hemagglutinin and neuraminidase, are synthesized and incorporated into the plasma membrane. These target cells were lysed by cross-reactive CTL. The envelope proteins of type A/Victoria virus were separated from the other virion components and reconstituted into lipid vesicles that lacked M protein that subsequently were used to prepare artificial target cells. Target-cell formation with vesicles was achieved by addition of fusion-active Sendai virus. These artificial target cells were also susceptible to lysis by cross-reactive CTL. In contrast to previous observations that suggested that the M protein of influenza viruses is recognized by these effector cells, we present evidence that the antigencic determinants induced by the viral glycoproteins are recognized.


Molecular characterization and expression of urotensin II and its receptor in the flounder (Platichthys flesus): a hormone system supporting body fluid homeostasis in euryhaline fish.
Urotensin II (UII) is a potent vasoconstrictor in mammals, but the source of circulating UII remains unclear. Investigations of the caudal neurosecretory system (CNSS), considered the major source of UII in fish, alongside target tissue expression of UII receptor (UT), can provide valuable insights into this highly conserved regulatory system. We report UII gene characterization, expression of the first fish UT, and responses to salinity challenge in flounder. The 12-aa UII peptide shares 73% sequence identity with pig and human UII. Flounder UT receptor shares 56.7% identity with rat. Although the CNSS is the major site of UII expression, RT-PCR revealed expression of UII and UT in all tissues tested. Around 30-40% of large CNSS Dahlgren cells expressed UII, alone or in combination with urotensin I and/or corticotrophin releasing hormone. Immunolocalization of UT in osmoregulatory tissues (gill, kidney) was associated with vascular elements. There were no consistent differences in CNSS UII expression or plasma UII between seawater (SW)- and freshwater (FW)-adapted fish, although gill and kidney UT expression was lower in FW animals. After acute transfer from SW to FW, plasma UII and kidney and gill UT expression were reduced, whereas UT expression in kidney was increased after reverse transfer. UII appears to be more important to combat dehydration and salt-loading in SW than the hemodilution faced in FW. Potentially, altered target tissue sensitivity through changes in UT expression, is an important physiological controlling mechanism, not only relevant for migratory fish but also likely conserved in mammals.


A comparison of physical and psychological features of responders and non-responders to cervical facet blocks in chronic whiplash.
Cervical facet block (FB) procedures are often used as a diagnostic precursor to radiofrequency neurotomies (RFN) in the management of chronic whiplash associated disorders (WAD). Some individuals will respond to the FB procedures and others will not respond. Such responders and non-responders provided a sample of convenience to question whether there were differences in their physical and psychological features. This information may inform future predictive studies and ultimately the clinical selection of patients for FB procedures. This cross-sectional study involved 58 individuals with chronic WAD who responded to cervical FB procedures (WAD_R); 32 who did not respond (WAD_NR) and 30 Healthy Controls (HC)s. Measures included: quantitative sensory tests (pressure; thermal pain thresholds; brachial plexus provocation test); nociceptive flexion reflex (NFR); motor function (cervical range of movement (ROM); activity of the superficial neck flexors during the cranio-cervical flexion test (CCFT). Self-reported measures were gained from the following questionnaires: neuropathic pain (s-LANSS); psychological distress (General Health Questionnaire-28), post-traumatic stress (PDS) and pain catastrophization (PCS). Individuals with chronic whiplash attended the laboratory once the effects of the blocks had abated and symptoms had returned. Following FB procedures, both WAD groups demonstrated generalized hypersensitivity to all sensory tests, decreased neck ROM and increased superficial muscle activity with the CCFT compared to controls (p < 0.05). There were no significant differences between WAD groups (all p > 0.05). Both WAD groups demonstrated psychological distress (GHQ-28; p < 0.05), moderate post-traumatic stress symptoms and pain catastrophization. The WAD_NR group also demonstrated increased medication intake and elevated PCS scores compared to the WAD_R group (p < 0.05). Chronic WAD responders and non-responders to FB procedures demonstrate a similar presentation of sensory disturbance, motor dysfunction and psychological distress. Higher levels of pain catastrophization and greater medication intake were the only factors found to differentiate these groups.


Cooking methods and esophageal squamous cell carcinoma in high-risk areas of Iran.
Cooking methods have been implicated in the etiology of gastrointestinal cancers, reflecting exposure to potential carcinogens as results of cooking. We used a validated food frequency questionnaire and a pretested cooking method questionnaire in 3 groups: 40 esophageal squamous cell carcinoma (ESCC) cases from a high-risk area in northeast of Iran, 40 healthy subjects from the same high-risk area, and 40 healthy subjects from a low-risk area in Southern Iran. We compared the frequency of boiling, grilling, and frying, and the frying score among these 3 groups. We also calculated "frying index" by multiplying the frequency of each fried food item by its frying score. Mean frying to boiling ratios were 18.2:1, 12.8:1, and 2.6:1 for cases, high-risk controls, and low-risk controls, respectively (P < 0.01). Reuse of cooking oil for frying was reported in 37.5% of the ESCC cases, 25% of high-risk controls, and 7.5% of low-risk controls (P < 0.001). Frying index was higher in the high-risk than in the low-risk controls (P < 0.001) and in cases than in the high-risk controls (P < 0.05) after adjusting for smoking, opium use, rural residence, education, and ethnicity. High-temperature cooking and frying may be associated with increased risk of ESCC in high-risk areas.


The type 1 insulin-like growth factor receptor is over-expressed in bladder cancer.
To analyse bladder cancer biopsies and investigate the pattern of expression of the type 1 insulin-like growth factor receptor (IGF1R), a receptor tyrosine kinase that mediates tumour cell proliferation, motility and protection from apoptosis. Formalin-fixed specimens of bladder cancer (40 whole-mount, 80 cores on a tumour microarray) and normal bladder (15 samples) were stained immunohistochemically for the IGF1R. The IGF1R expression was also measured by quantitative reverse transcription-polymerase chain reaction (Q-RT-PCR) on RNA extracted from fresh frozen bladder cancers (61) and benign bladder (12). Of the 15 samples of normal bladder, 14 showed negligible (1+) or light (2+) IGF1R immunostaining. By contrast moderate (3+) or heavy (4+) staining for IGF1R was detected in 89 (74%) of the 120 samples of malignant urothelium. Q-RT-PCR showed significantly higher levels of steady-state IGF1R mRNA in tumours (all cases, Ta-T4) than in normal bladder (P < 0.05), indicating up-regulation at the transcriptional level. This difference was particularly evident when comparing normal urothelium with superficial (Ta-T1) or invasive (T2-4) tumours; only the latter showed significant IGF1R over-expression at the RNA level (P < 0.05 vs normal bladder). The IGF1R is up-regulated in bladder cancer compared with non-malignant bladder, and might contribute to a propensity for invasion.


A clinical threat in patients with granulomatosis polyangiitis in remission: Subglottic stenosis.
Granulomatosis with polyangiitis (GPA) is a systemic necrotizing granulomatous disease that involves small- and medium-sized arteries and affects the main respiratory tracts and kidneys. Upper respiratory tract involvement usually occurs in 90% of patients, who most frequently present with symptoms of chronic sinusitis. Subglottic stenosis (SS) is a rare and severe complication that is usually observed in approximately 15% of patients. Here we present a case of SS in a patient with limited form of GPA during remission.


Risks of Stroke Recurrence and Mortality After First and Recurrent Strokes in Denmark: A Nationwide Registry Study.
To examine risks of stroke recurrence and mortality after first and recurrent stroke. Using Danish nationwide health registries, we included patients (age ≥18 years) with first-time ischemic stroke (N = 105,397) or intracerebral hemorrhage (N = 13,350) during 2004-2018. Accounting for the competing risk of death, absolute risks of stroke recurrence were computed separately for each stroke subtype and within strata of age groups, sex, stroke severity, body mass index, smoking, alcohol, the Essen stroke risk score, and atrial fibrillation. Mortality risks were computed after first and recurrent stroke. After adjusting for competing risks, the overall 1-year and 10-year risks of recurrence were 4% and 13% following first-time ischemic stroke and 3% and 12% following first-time intracerebral hemorrhage. For ischemic stroke, the risk of recurrence increased with age, was higher for men and following mild than more severe stroke. The most marked differences were across Essen risk scores, for which recurrence risks increased with increasing scores. For intracerebral hemorrhage, risks were similar for both sexes and did not increase with Essen risk score. For ischemic stroke, the 1-year and 10-year risks of all-cause mortality were 17% and 56% after a first-time stroke and 25% and 70% after a recurrent stroke; corresponding estimates for intracerebral hemorrhage were 37% and 70% after a first-time event and 31% and 75% after a recurrent event. The risk of stroke recurrence was substantial following both subtypes, but risks differed markedly among patient subgroups. The risk of mortality was higher after a recurrent than first-time stroke.


Chemical characteristics of an Ilex Kuding tea polysaccharide and its protective effects against high fructose-induced liver injury and vascular endothelial dysfunction in mice.
The present study was designed to investigate the protective effects of Ilex Kuding tea polysaccharides (IKTP) on high fructose (HF)-induced liver injury and vascular endothelial dysfunction in mice. IKTP were identified as acidic heteropolysaccharides by FT-IR and HPLC. Healthy male Kunming mice fed 20% fructose in drinking water for 8 consecutive weeks significantly displayed dyslipidemia, hepatic steatosis, oxidative stress and vascular endothelial dysfunction. However, continuous administration of IKTP at 200, 400 and 800 mg per kg bw in HF-fed mice could prevent the damage caused by HF-diets, especially at dosages of 400 and 800 mg per kg bw (p < 0.01). IKTP significantly reduced the HF-induced elevation of the serum TC, TG, LDL-C, TXA2 and ET-1 levels, as well as AST and ALT activities, while markedly increased the HF-induced decline of HDL-C, PGI2 and eNOS levels in the serum compared to HF-fed mice. Meanwhile, the hepatic MDA level was lowered while SOD and GSH-Px activities were increased in IKTP-treated mice, compared to HF-fed mice. Histopathology of the liver and cardiovascular aortic by H&E or oil red O staining confirmed the liver steatosis and the vascular injury induced by HF-diets and the protective effects of IKTP. These findings suggest that HF causes oxidative damage, and IKTP alleviates liver injury and vascular endothelial dysfunction.


Identification and characterization of glycanated and non-glycanated forms of biglycan and decorin in the human intervertebral disc.
Immunological studies revealed the presence of several different forms of biglycan and decorin in human intervertebral-disc tissues (annulus fibrosus, nucleus pulposus and cartilage end-plate). In the young intervertebral disc, glycosaminoglycan-containing (glycanated) forms of both biglycan and decorin represented a greater proportion of the total proteoglycan population present in extracts of annulus fibrosus and cartilage end-plate compared with extracts of nucleus pulposus, in which they were barely detectable. In older discs the glycanated forms of biglycan and decorin represented only a small proportion of the total proteoglycan present. Immunochemical analyses with an antibody to chondroitin/dermatan sulphate isomers indicated differences in the glycosaminoglycans substituted on glycanated forms of small proteoglycans found in different disc tissues. Dermatan sulphate was the predominant glycosaminoglycan present on biglycan and decorin in annulus fibrosus extracts, whereas chondroitin 4-sulphate was present in both small proteoglycans isolated from cartilage end-plate. In addition, immunochemical analyses with antibodies against core protein epitopes identified two non-glycanated forms of both biglycan and decorin. These non-glycanated forms of the small proteoglycans were found in all three regions of the disc. The two nonglycanated forms of biglycan had estimated molecular masses of 37 and 41 kDa and those of decorin were 43 and 45 kDa, respectively. These non-glycanated forms of biglycan and decorin increased in proportion with aging. N-terminal sequence analysis indicated that the larger non-glycanated form of decorin was a degradation product of its glycanated precursor. However, no N-terminal sequence information was obtainable from the other non-glycanated form of decorin or the two non-glycanated forms of biglycan. These data are consistent with the hypothesis that some of the non-glycanated forms of decorin and biglycan are degradation products of native precursors. However, the possibility remains that several different post-translationally modified forms of decorin and biglycan are synthesized by intervertebral-disc tissues.


Brain drain and health workforce distortions in Mozambique.
Trained human resources are fundamental for well-functioning health systems, and the lack of health workers undermines public sector capacity to meet population health needs. While external brain drain from low and middle-income countries is well described, there is little understanding of the degree of internal brain drain, and how increases in health sector funding through global health initiatives may contribute to the outflow of health workers from the public sector to donor agencies, non-governmental organisations (NGOs), and the private sector. An observational study was conducted to estimate the degree of internal and external brain drain among Mozambican nationals qualifying from domestic and foreign medical schools between 1980-2006. Data were collected 26-months apart in 2008 and 2010, and included current employment status, employer, geographic location of employment, and main work duties. Of 723 qualifying physicians between 1980-2006, 95.9% (693) were working full-time, including 71.1% (493) as clinicians, 20.5% (142) as health system managers, and 6.9% (48) as researchers/professors. 25.5% (181) of the sample had left the public sector, of which 62.4% (113) continued working in-country and 37.6% (68) emigrated from Mozambique. Of those cases of internal migration, 66.4% (75) worked for NGOs, 21.2% (24) for donor agencies, and 12.4% (14) in the private sector. Annual incidence of physician migration was estimated to be 3.7%, predominately to work in the growing NGO sector. An estimated 36.3% (41/113) of internal migration cases had previously held senior-level management positions in the public sector. Internal migration is an important contributor to capital flight from the public sector, accounting for more cases of physician loss than external migration in Mozambique. Given the urgent need to strengthen public sector health systems, frank reflection by donors and NGOs is needed to assess how hiring practices may undermine the very systems they seek to strengthen.


Droplet digital PCR for detection and quantification of circulating tumor DNA in plasma of head and neck cancer patients.
During posttreatment surveillance of head and neck cancer patients, imaging is insufficiently accurate for the early detection of relapsing disease. Free circulating tumor DNA (ctDNA) may serve as a novel biomarker for monitoring tumor burden during posttreatment surveillance of these patients. In this exploratory study, we investigated whether low level ctDNA in plasma of head and neck cancer patients can be detected using Droplet Digital PCR (ddPCR). TP53 mutations were determined in surgically resected primary tumor samples from six patients with high stage (II-IV), moderate to poorly differentiated head and neck squamous cell carcinoma (HNSCC). Subsequently, mutation specific ddPCR assays were designed. Pretreatment plasma samples from these patients were examined on the presence of ctDNA by ddPCR using the mutation-specific assays. The ddPCR results were evaluated alongside clinicopathological data. In all cases, plasma samples were found positive for targeted TP53 mutations in varying degrees (absolute quantification of 2.2-422 mutational copies/ml plasma). Mutations were detected in wild-type TP53 background templates of 7667-156,667 copies/ml plasma, yielding fractional abundances of down to 0.01%. Our results show that detection of tumor specific TP53 mutations in low level ctDNA from HNSCC patients using ddPCR is technically feasible and provide ground for future research on ctDNA quantification for the use of diagnostic biomarkers in the posttreatment surveillance of HNSCC patients.


Embryonic development of the Drosophila brain: formation of commissural and descending pathways.
The establishment of initial axonal pathways in the embryonic brain of Drosophila melanogaster was investigated at the cellular and molecular level using antibody probes, enhancer detector strains and axonal pathfinding mutants. During embryogenesis, two bilaterally symmetrical cephalic neurogenic regions form, which are initially separated from each other and from the ventral nerve cord. The brain commissure that interconnects the two brain hemispheres is pioneered by axons that project towards the midline in close association with an interhemispheric cellular bridge. The descending longitudinal pathways that interconnect the brain to the ventral nerve cord are prefigured by a chain of longitudinal glial cells and a cellular bridge between brain and subesophageal ganglion; pioneering descending and ascending neurons grow in close association with these structures. The formation of the embryonic commissural and longitudinal pathways is dependent on cells of the CNS midline. Mutations in the commissureless gene, which affects growth cone guidance towards the midline, result in a marked reduction of the brain commissure. Mutations in the single-minded gene and in other spitz group genes, which affect the differentiation of CNS midline cells, result in the absence or aberrant projection of longitudinal pathways. The analysis of axon pathway formation presented here reveals remarkable similarities as well as distinct differences in the embryonic development of the brain and the segmental ganglia, and forms the basis for a comprehensive genetic and molecular genetic dissection of axonal pathfinding processes in the developing brain.


Phase I trial and pharmacokinetic study of bevacizumab in pediatric patients with refractory solid tumors: a Children's Oncology Group Study.
We conducted a pediatric phase I trial of the vascular endothelial growth factor (VEGF)-neutralizing antibody bevacizumab (BV). Primary aims included estimating the maximum-tolerated dose (MTD) and determining the dose-limiting toxicities (DLTs), pharmacokinetics, and biologic effects of BV in children with cancer. BV (5, 10, 15 mg/kg) was administered intravenously every 2 weeks in 28-day courses to children with refractory solid tumors. Twenty-one patients enrolled, 20 (median age, 13 years) were eligible, and 18 completed one course and were fully assessable for toxicity. A total of 67 courses were administered (median, three courses per patient; range, one to 16 courses). Treatment was well tolerated with no DLTs observed. Non-DLTs included infusional reaction, rash, mucositis, proteinuria, and lymphopenia. Increases in systolic and diastolic blood pressure not meeting Common Terminology Criteria for Adverse Events (CTCAEv3) pediatric-specific criteria for hypertension were observed. There was no hemorrhage or thrombosis. Growth perturbation was not detected in a limited sample over the first course. The serum exposure to BV as measured by area under the concentration-time curve (AUC) seemed to increase in proportion to dose. The median clearance of BV was 4.1 mL/d/kg (range, 3.1 to 15.5 mL/d/kg), and the median half-life was 11.8 days (range, 4.4 to 14.6 days). No objective responses were observed. Exploratory analyses on circulating endothelial mobilization and viability are consistent with the available adult data. BV is well tolerated in children. Phase II pediatric studies of BV in combination with chemotherapy in dosing schedules similar to adults are planned.


Adenine-DNA adducts derived from the highly tumorigenic Dibenzo[a,l]pyrene are resistant to nucleotide excision repair while guanine adducts are not.
The structural origins of differences in susceptibilities of various DNA lesions to nucleotide excision repair (NER) are poorly understood. Here we compared, in the same sequence context, the relative NER dual incision efficiencies elicited by two stereochemically distinct pairs of guanine (N(2)-dG) and adenine (N(6)-dA) DNA lesions, derived from enantiomeric genotoxic diol epoxides of the highly tumorigenic fjord region polycyclic aromatic hydrocarbon dibenzo[a,l]pyrene (DB[a,l]P). Remarkably, in cell-free HeLa cell extracts, the guanine adduct with R absolute chemistry at the N(2)-dG linkage site is ∼35 times more susceptible to NER dual incisions than the stereochemically identical N(6)-dA adduct. For the guanine and adenine adducts with S stereochemistry, a similar but somewhat smaller effect (factor of ∼15) is observed. The striking resistance of the bulky N(6)-dA in contrast to the modest to good susceptibilities of the N(2)-dG adducts to NER is interpreted in terms of the balance between lesion-induced DNA distorting and DNA stabilizing van der Waals interactions in their structures, that are partly reflected in the overall thermal stabilities of the modified duplexes. Our results are consistent with the hypothesis that the high genotoxic activity of DB[a,l]P is related to the formation of NER-resistant and persistent DB[a,l]P-derived adenine adducts in cellular DNA.


Comparison of dopamine and norepinephrine in the treatment of shock.
Both dopamine and norepinephrine are recommended as first-line vasopressor agents in the treatment of shock. There is a continuing controversy about whether one agent is superior to the other. In this multicenter, randomized trial, we assigned patients with shock to receive either dopamine or norepinephrine as first-line vasopressor therapy to restore and maintain blood pressure. When blood pressure could not be maintained with a dose of 20 microg per kilogram of body weight per minute for dopamine or a dose of 0.19 microg per kilogram per minute for norepinephrine, open-label norepinephrine, epinephrine, or vasopressin could be added. The primary outcome was the rate of death at 28 days after randomization; secondary end points included the number of days without need for organ support and the occurrence of adverse events. The trial included 1679 patients, of whom 858 were assigned to dopamine and 821 to norepinephrine. The baseline characteristics of the groups were similar. There was no significant between-group difference in the rate of death at 28 days (52.5% in the dopamine group and 48.5% in the norepinephrine group; odds ratio with dopamine, 1.17; 95% confidence interval, 0.97 to 1.42; P=0.10). However, there were more arrhythmic events among the patients treated with dopamine than among those treated with norepinephrine (207 events [24.1%] vs. 102 events [12.4%], P<0.001). A subgroup analysis showed that dopamine, as compared with norepinephrine, was associated with an increased rate of death at 28 days among the 280 patients with cardiogenic shock but not among the 1044 patients with septic shock or the 263 with hypovolemic shock (P=0.03 for cardiogenic shock, P=0.19 for septic shock, and P=0.84 for hypovolemic shock, in Kaplan-Meier analyses). Although there was no significant difference in the rate of death between patients with shock who were treated with dopamine as the first-line vasopressor agent and those who were treated with norepinephrine, the use of dopamine was associated with a greater number of adverse events. (ClinicalTrials.gov number, NCT00314704.)