Intrafraction displacement of prone versus supine prostate positioning monitored by real-time electromagnetic tracking.
Implanted radiofrequency transponders were used for real-time monitoring of the intrafraction prostate displacement between patients in the prone position and the same patients in the supine position. Thirteen patients had three transponders implanted transperineally and were treated prone with a custom-fitted thermoplastic immobilization device. After collecting data from the last fraction, patients were realigned in the supine position and the displacements of the transponders were monitored for 5-7 minutes. Fourier transforms were applied to the data from each patient to determine periodicity and its amplitude. To remove auto correlation from the stream of displacement data, the distribution of short-term and long-term velocity components were calculated from Poincaré plots of paired sequential vector displacements. The mean absolute displacement was significantly greater prone than supine in the superior-inferior (SI) plane (1.2 ± 0.6 mm vs. 0.6 ± 0.4 mm, p= 0.015), but not for the lateral or anterior-posterior (AP) planes. Displacements were least in the lateral direction. Fourier analyses showed the amplitude of respiratory oscillations was much greater for the SI and AP planes in the prone versus the supine position. Analysis of Poincaré plots confirmed greater short-term variance in the prone position, but no difference in the long-term variance. The centroid of the implanted transponders was offset from the treatment isocenter by > 5 mm for 1.9% of the time versus 0.8% of the time for supine. These results confirmed significantly greater net intrafraction prostate displacement of patients in the prone position than in the supine position, but most of the difference was due to respiration-induced motion that was most pronounced in the SI and AP directions. Because the respiratory motion remained within the action threshold and also within our 5 mm treatment planning margins, there is no compelling reason to choose one treatment position over the other.


Personal goals and factors related to QoL in Dutch homeless people: what is the role of goal-related self-efficacy?
Very little is known about the personal goals of homeless people and how these relate to their quality of life (QoL). By using survey data on 407 homeless adults upon entry to the social relief system in 2011, we examined the personal goals of homeless adults and the association between their perceived goal-related self-efficacy and their QoL. A hierarchical regression analysis was used to analyse the association between QoL and goal-related self-efficacy, relative to factors contributing to QoL, such as demographic characteristics, socioeconomic resources, health and service use. Results indicate that the majority of homeless adults had at least one personal goal for the coming 6 months and that most goals concerned housing and daily life (94.3%) and finances (83.6%). The QoL of homeless adults appeared to be lower in comparison with general population samples. General goal-related self-efficacy was positively related to QoL (β = 0.09, P = 0.042), independent of socioeconomic resources (i.e. income and housing), health and service use. The strongest predictors of QoL were psychological distress (β = -0.45, P < 0.001), income (β = 0.14, P = 0.002) and being institutionalised (β = 0.12, P = 0.004). In conclusion, the majority of homeless adults entering the social relief system have personal goals regarding socioeconomic resources and their goal-related self-efficacy is positively related to QoL. It is therefore important to take the personal goals of homeless people as the starting point of integrated service programmes and to promote their goal-related self-efficacy by strength-based interventions.


Long term complications following pelvic and para-aortic lymphadenectomy for endometrial cancer, incidence and potential risk factors: a single institution experience.
To determine the incidence of long term lymphadenectomy complications in primary surgery for endometrial cancer and to elucidate risk factors for these complications. A retrospective chart review was carried out for all patients with endometrial cancer managed at Parma University Hospital Unit of Gynecology and Obstetrics between 2010 and 2016. Inclusion criteria were surgical procedure including hysterectomy and lymphadenectomy (pelvic or pelvic and aortic). We identified patients with postoperative lymphocele and lower extremity lymphedema. Logistic regression analysis was used to identify predictive factors for postoperative complications. Of the 249 patients tested, 198 underwent pelvic lymphadenectomy (79.5%), and 51 (20.5%) of those underwent both pelvic and para-aortic lymphadenectomy. Among the 249 patients, 92 (36.9 %) developed lymphedema while 43 (17.3%) developed lymphocele. Multivariate analysis showed that addition of para-artic lymphadenectomy was an independent predictor for both lymphedema (odds ratio (OR) 2.764, 95% confidence interval (CI) 1.023 to 7.470) and lymphocele (OR 5.066, 95% CI 1.605 to 15.989). Moreover, postoperative adjuvant radiotherapy (OR 2.733, 95% CI 1.149 to 6.505) and identification of any positive lymph node (OR 19.391, 95% CI 1.486 to 253.0) were significantly correlated with lymphedema, while removal of circumflex iliac nodes (OR 8.596, 95% CI 1.144 to 65.591) was associated with lymphoceles occurrence. Although sentinel lymph node navigation is a promising option, lymphadenectomy represents the primary treatment in many patients with endometrial cancer. However, comprehensive nodal dissection remains associated with a high rate of long term complications, such as lymphedema and lymphocele. Avoiding risk factors that are related to the development of these postoperative complications is often difficult and, therefore, the strategy to assess lymph nodal status in these women must be tailored to obtain the maximum results in terms of oncological and functional outcome.


Combined experimental/theoretical refinement of indole ring geometry using deuterium magnetic resonance and ab initio calculations.
We have used experimental deuterium NMR spectra from labeled tryptophans in membrane-spanning gramicidin A (gA)(1) channels to refine the geometry of the indole ring and, specifically, the C2-(2)H bond direction. By using partial exchange in a cold organic acid, we were able to selectively deuterate ring positions C2 and C5 and, thereby, define unambiguous spectral assignments. In a backbone-independent analysis, the assigned spectra from four distinct labeled tryptophans were used to assess the geometry of the planar indole ring. We found that the C2-(2)H bond makes an angle of about 6 degrees with respect to the normal to the indole ring bridge, and the experimental geometry was confirmed by density functional calculations using a 6-311G** basis set. The precisely determined ring geometry and the experimental spectra in turn are the foundation for calculations of the orientation of each tryptophan indole ring, with respect to the bilayer membrane normal, and of a principal order parameter S(zz) for each ring. The results have general significance for revising the tryptophan ring geometry that is used in protein molecular modeling, as well as for the analysis of tryptophan ring orientations in membrane-spanning proteins. The experimental precision in the definition of the indole ring geometry demonstrates yet another practical application emanating from fundamental research on the robust gramicidin channel.


Occupational health services in selected International Commission on Occupational Health (ICOH) member countries.
International organizations have encouraged countries to organize occupational health services (OHS) for all working people. This study surveyed how that objective has been achieved in a sample of International Commission on Occupational Health (ICOH) member countries from all continents. A structured questionnaire was sent to ICOH national secretaries in 61 countries. The survey focused on (i) policies and strategies; (ii) systems, institutions, and infrastructures; (iii) contents and activities; (iv) human and other resources; (v) financing; and (v) future priorities in the development of national OHS systems. Of 47 respondents, 70% had drawn up a policy and strategy for OHS and 79% had a national institute of occupational health or safety. The calculated coverage of workers was 19% among the respondent countries. Sixty percent of respondent countries used multiple channels for service provision and 70% provided mixed contents of OHS with preventive and curative services. Almost all (94%) reported availability of multidisciplinary experts, but not in sufficient numbers. OHS is financed through combined employer plus insurance financing in 62% of respondents and through employer financing only in 38%. The countries identified well the needs for future development of OHS. In spite of documented policies for OHS, only slightly over one third of the surveyed countries had organized OHS for more than 50% of workers. The vast majority of workers of the world are underserved due to four gaps in OHS: implementation, coverage, content, and capacity.


Developing de novo human artificial chromosomes in embryonic stem cells using HSV-1 amplicon technology.
De novo artificial chromosomes expressing genes have been generated in human embryonic stem cells (hESc) and are maintained following differentiation into other cell types. Human artificial chromosomes (HAC) are small, functional, extrachromosomal elements, which behave as normal chromosomes in human cells. De novo HAC are generated following delivery of alpha satellite DNA into target cells. HAC are characterized by high levels of mitotic stability and are used as models to study centromere formation and chromosome organisation. They are successful and effective as gene expression vectors since they remain autonomous and can accommodate larger genes and regulatory regions for long-term expression studies in cells unlike other viral gene delivery vectors currently used. Transferring the essential DNA sequences for HAC formation intact across the cell membrane has been challenging for a number of years. A highly efficient delivery system based on HSV-1 amplicons has been used to target DNA directly to the ES cell nucleus and HAC stably generated in human embryonic stem cells (hESc) at high frequency. HAC were detected using an improved protocol for hESc chromosome harvesting, which consistently produced high-quality metaphase spreads that could routinely detect HAC in hESc. In tumour cells, the input DNA often integrated in the host chromosomes, but in the host ES genome, it remained intact. The hESc containing the HAC formed embryoid bodies, generated teratoma in mice, and differentiated into neuronal cells where the HAC were maintained. The HAC structure and chromatin composition was similar to the endogenous hESc chromosomes. This review will discuss the technological advances in HAC vector delivery using HSV-1 amplicons and the improvements in the identification of de novo HAC in hESc.


The mid- to long-term results of the anterior cruciate ligament reconstruction with hamstring tendons using Transfix technique.
In this study, mid to long-term results of anterior cruciate ligament reconstruction with hamstring tendons and Transfix technique were evaluated. Anterior cruciate ligament (ACL) reconstruction with four-strand hamstring tendon was performed with Transfix technique on 271 (198 males, 73 females; mean age 25.7; 17-52) patients with anterior cruciate ligament ruptures. The patients were followed up with clinical examination, Lysholm and Tegner activity scales, IKDC scoring system, KT-1000 test and radiological examination. The mean follow-up period was 82 (48-100) months; 204 (75%) patients had no subjective complaints. According to the KT-1000 test, only 14 (5%) patients had more than 5 mm laxity postoperatively, whereas, 161 (59%) patients had more than 5 mm laxity preoperatively. In addition to this, only 19 (7%) patients had Lysholm scores less than 80 postoperatively, whereas 154 (57%) patients scored less than 80 preoperatively. When compared with Tegner activity scale, 189 (70%) patients scored<6 preoperatively and only 24 (8%) postoperatively; 78 (29%) patients scored D preoperatively and only 5 (2%) patients scored D postoperatively on the basis of the IKDC scoring system. Our functional results were found to be satisfactory in more than 90% of patients. Commonly seen problems in ACL reconstruction such as inaccurate graft placement and tunnel widening were found to be consistent with the values in relevant literature. However, we demonstrated that the functional results and the stability of the knee were not related with tunnel widening. This study concludes that the reconstruction of ACL with hamstring tendons and the Transfix technique is reasonably successful, safe and causes low morbidity. Furthermore, we believe that proper graft preparation, accurate tunnel placement, notch-plasty, fixation and rehabilitation program are all as important as the choice of graft and fixation material.


Nerve fibers in culture and their interactions with non-neural cells visualized by immunofluorescence.
Cultures of embryonic mouse spinal cord explants, alone or in combination with rat myotubes, were stained by indirect immunofluorescence using antibodies against three structural proteins to: (a) reveal the distribution of these proteins among different cell types, and (b) test the usefulness of antibody staining to reveal the gross morphology of the neurite network in complex cultures. Affinity column purified antibodies were used against chicken gizzard actin, porcine brain tubulin, and skeletal muscle alpha-actinin. Neurites were stained intensely by anti-actin as was the stress fiber pattern of underlying fibroblasts. With anti-tubulin, the staining of neurites was an order of magnitude more intense than the staining of the microtubule pattern of background fibroblasts. Neurite cell bodies and astrocyte-like glia cells were stained with anti-tubulin and their nuclei remained unstained. Anti-tubulin could thus be used to trace even the finest extensions of nerve processes in spinal cord and spinal cord-muscle cultures. Furthermore, it could be combined with the histochemical reaction for acetylcholinesterase (AChE, EC 22.214.171.124) to demonstrate AChE-positive neurons and specialized nerve-muscle contact sites. The staining of neural elements with anti-alpha-actinin was generally much weaker than with anti-actin and anti-tubulin. Neurites were stained only moderately in comparison to myotube Z lines in the same culture. However, a distinct staining of the periphery of dorsal root ganglion cells was observed. Thus, a protein immunologically related to muscle alpha-actinin is present in the nervous system. In myotubes, Z lines were stained intensely with anti-alpha-actinin while I bands were only faintly stained with anti-actin. In isolated myofibrils, both structures were stained intensely with the same antibody preparations.


The influence of footwear on walking biomechanics in individuals with chronic ankle instability.
The effects of footwear on the walking kinematics, kinetics and electromyography (EMG) of individuals with chronic ankle instability (CAI) at different speeds are still unknown. The objective of this cross-sectional study was to evaluate the kinematic, kinetic and electromyography differences between shod and barefoot walking at comfortable (CW) and fast (FW) speeds in individuals with CAI. Twenty-one individuals with CAI walked on a 5-meter walkway shod and barefoot at CW and FW speeds. A force plate was used to record the ground reaction forces, a 3-D motion analysis system to record the lower limb kinematics and a surface EMG system to collect the gluteus medius, vastus lateralis, gastrocnemius lateralis, gastrocnemius medialis, peroneus longus and tibialis anterior muscles activity. The dependent variables were ankle and knee angles and moments and normalized muscle activity. The shod and barefoot data during CW and FW were compared using a one-dimensional non-parametric mapping analysis. The main results of this study were that individuals with CAI exhibited more ankle dorsiflexion angle, knee extension and tibialis anterior muscle activation during the beginning of the stance phase during shod compared to barefoot walking. Also, the biomechanical effects of shoes are similar during walking at FW and CW. The biomechanical deficits associated with CAI were partly attenuated during the shod compared to the barefoot condition and these effects were similar at CW and FW. These findings are compatible with the concept that locomotor interventions using suitable shoes may enhance gait abilities in individuals with CAI.


Hydrolysis of Formyl Fluoride Catalyzed by Sulfuric Acid and Formic Acid in the Atmosphere.
Formyl fluoride (HFCO) is an important atmospheric molecule, and its reaction with the OH radical is an important pathway when degradation of HFCO is considered in earth's troposphere. Here, we study the hydrolysis of formyl fluoride (HFCO + H2O) with sulfuric acid (H2SO4) and formic acid (HCOOH) acting as catalysts by utilizing M06-2X, CCSD(T)-F12a, and conventional transitional state theory with Eckart tunneling to explore the atmospheric impact of the above-said hydrolysis reactions. Our calculated results show that H2SO4 has a remarkably catalytic role in the gas-phase hydrolysis of HFCO, as the energy barriers of the HFCO + H2O reaction are reduced from 39.22 and 41.19 to 0.26 and -0.63 kcal/mol with respect to the separate reactants, respectively. In addition, we also find that H2SO4 can significantly accelerate the decomposition of FCH(OH)2 into hydrogen fluoride (HF) and HCOOH. This is because while the barrier height for the unimolecular decomposition of FCH(OH)2 into HF and HCOOH is 31.63 kcal/mol, the barrier height for the FCH(OH)2 + H2SO4 reaction is predicted to be -5.99 kcal/mol with respect to separate reactants. Nevertheless, the comparative relative rate analysis shows that the reaction between HFCO and the OH radical is still the most dominant pathway when the tropospheric degradation of HFCO is taken into account and that the gas-phase hydrolysis of HFCO may only occur with the help of H2SO4 when the atmospheric concentration of OH is about 101 molecules cm-3 or less. Having an understanding from the present study that the gas-phase hydrolysis of HFCO in the presence of H2SO4 has very limited role possibly in the absence of sunlight, we also prefer here to emphasize that the HFCO + H2O + H2SO4 reaction may occur on the surface of secondary organic aerosols for the formation of HCOOH.


Between- and within-herd variation in blood and milk biomarkers in Holstein cows in early lactation.
Both blood- and milk-based biomarkers have been analysed for decades in research settings, although often only in one herd, and without focus on the variation in the biomarkers that are specifically related to herd or diet. Biomarkers can be used to detect physiological imbalance and disease risk and may have a role in precision livestock farming (PLF). For use in PLF, it is important to quantify normal variation in specific biomarkers and the source of this variation. The objective of this study was to estimate the between- and within-herd variation in a number of blood metabolites (β-hydroxybutyrate (BHB), non-esterified fatty acids, glucose and serum IGF-1), milk metabolites (free glucose, glucose-6-phosphate, urea, isocitrate, BHB and uric acid), milk enzymes (lactate dehydrogenase and N-acetyl-β-D-glucosaminidase (NAGase)) and composite indicators for metabolic imbalances (Physiological Imbalance-index and energy balance), to help facilitate their adoption within PLF. Blood and milk were sampled from 234 Holstein dairy cows from 6 experimental herds, each in a different European country, and offered a total of 10 different diets. Blood was sampled on 2 occasions at approximately 14 days-in-milk (DIM) and 35 DIM. Milk samples were collected twice weekly (in total 2750 samples) from DIM 1 to 50. Multilevel random regression models were used to estimate the variance components and to calculate the intraclass correlations (ICCs). The ICCs for the milk metabolites, when adjusted for parity and DIM at sampling, demonstrated that between 12% (glucose-6-phosphate) and 46% (urea) of the variation in the metabolites' levels could be associated with the herd-diet combination. Intraclass Correlations related to the herd-diet combination were generally higher for blood metabolites, from 17% (cholesterol) to approximately 46% (BHB and urea). The high ICCs for urea suggest that this biomarker can be used for monitoring on herd level. The low variance within cow for NAGase indicates that few samples would be needed to describe the status and potentially a general reference value could be used. The low ICC for most of the biomarkers and larger within cow variation emphasises that multiple samples would be needed - most likely on the individual cows - for making the biomarkers useful for monitoring. The majority of biomarkers were influenced by parity and DIM which indicate that these should be accounted for if the biomarker should be used for monitoring.


Effect of antihypertensive treatment on kidney function in diabetic nephropathy.
The effect of long term, aggressive antihypertensive treatment on kidney function in diabetic nephropathy was studied prospectively in 11 insulin dependent diabetics (mean age 30). During the mean pretreatment period of 32 (range 23-66) months the glomerular filtration rate decreased significantly and albuminuria and the arterial blood pressure increased significantly. During the 72 (range 32-91) month period of antihypertensive treatment the average arterial blood pressure fell from 143/96 mm Hg to 129/84 mm Hg and albuminuria decreased from 1038 micrograms/min to 504 micrograms/min. The rate of decline in the glomerular filtration rate decreased from 0.89 (range 0.44-1.46) ml/min/month before treatment to 0.22 (range 0.01-0.40) ml/min/month during treatment. The rate of decline in the glomerular filtration rate was significantly smaller during the second three years compared with the first three years in patients who received long term antihypertensive treatment (greater than or equal to 6 years). One patient died from acute myocardial infarction (glomerular filtration rate 46 ml/min/1.74 m2). Effective antihypertensive treatment postpones renal insufficiency in diabetic nephropathy.


Ancient genomes suggest the eastern Pontic-Caspian steppe as the source of western Iron Age nomads.
For millennia, the Pontic-Caspian steppe was a connector between the Eurasian steppe and Europe. In this scene, multidirectional and sequential movements of different populations may have occurred, including those of the Eurasian steppe nomads. We sequenced 35 genomes (low to medium coverage) of Bronze Age individuals (Srubnaya-Alakulskaya) and Iron Age nomads (Cimmerians, Scythians, and Sarmatians) that represent four distinct cultural entities corresponding to the chronological sequence of cultural complexes in the region. Our results suggest that, despite genetic links among these peoples, no group can be considered a direct ancestor of the subsequent group. The nomadic populations were heterogeneous and carried genetic affinities with populations from several other regions including the Far East and the southern Urals. We found evidence of a stable shared genetic signature, making the eastern Pontic-Caspian steppe a likely source of western nomadic groups.


Rapid and slow nitric oxide responses during conducted vasodilation in the in vivo intestine and brain cortex microvasculatures.
Conduction of arteriolar vasodilation is initiated by activation of nitric oxide (NO) mechanisms, but dependent on conduction of hyperpolarization. Most studies have used brief (<1 second) activation of the initial vasodilation to evaluate the fast conduction processes. However, most arteriolar mechanisms involving NO production persist for minutes. In this study, fast and slower components of arteriolar conduction in the in vivo rat brain and small intestine were compared using three-minute stimulation of NO-dependent vasodilation and measurement of [NO] at the distal sites. Within 10-15 seconds, both vasculatures had a rapidly conducted vasodilation and dilation at distance had a fast but small [NO] component. A slower but larger distal vasodilation occurred after 60-90 seconds in the intestine, but not the brain, and was associated with a substantial increase in [NO]. This slowly developed dilation appeared to be caused by flow mediated responses of larger arterioles as smaller arterioles dilated to lower downstream resistance. These results indicate while the intestinal and cerebral arterioles have a fast conducted vasodilation system, the intestinal arterioles also have a slower but larger dilation of major arterioles that is NO related and dependent on the conduction of vasodilation between small arterioles.


Chelating effect of citric acid is negligible for development of enamel erosions.
Citric acid (CA) is a component in beverages responsible for dental erosion. The aim of this study was to examine the influence of CA with different pH, titratable acid and buffer capacity (ß), and the impact of the chelating effect of CA on development of enamel erosions. In a superfusion model, hydroxy apatite (HAp) dissolution of bovine enamel was measured in four experiments (EXP 1-4) with 27 experimental groups (n = 8 per group). The samples were superfused with different CA variations and respective controls. EXP-1: Dilution series of HCl (pH 2.15-3.02). EXP-2: Dilution series of natural CA (56-1.75 mmol l(-1); pH 2.15-3.02). EXP-3: CA solutions (56 and 14 mmol l(-1), ß: 39.7 and 10.2 mmol l(-1) pH(-1), respectively) with different titratable acidity at equal pH values. EXP-4: CA concentrations (56-1.75 mmol l(-1)) neutralized to pH 7. CA led to higher HAp-dissolution than HCl. With higher pH, the difference in HAp-dissolution rate between the two acids became increasingly smaller. At equal pH, HAp-dissolution was higher for the CA with the higher amount of titratable acid. However, no clear correlation between erosion and titratable acid or ß could be found. Only minimal amounts of HAp were dissolved by neutralized CA compared to CA with natural pH. Under the chosen conditions chelating effects of CA do not have a relevant influence of HAp-dissolution of enamel. Moreover, amount of HAp-dissolution by CA is not attributed to a single factor alone. The interplay between the different parameters of CA seems to be responsible for its erosive potential. The erosive potential of solutions containing citric acid with unknown concentrations could not be predicted using a single parameter alone, and should at best determined in experimental set-ups.


Neural substrates of contextual interference during motor learning support a model of active preparation.
When individuals acquire new skills, initial performance is typically better and tasks are judged to be easier when the tasks are segregated and practiced by block, compared to when different tasks are randomly intermixed in practice. However, subsequent skill retention is better for a randomly practiced group, an effect known as contextual interference (CI). The present study examined the neural substrates of CI using functional magnetic resonance imaging (fMRI). Individuals learned a set of three 4-element sequences with the left hand according to a block or random practice schedule. Behavioral retest for skill retention confirmed the presence of a typical CI effect with the random group outperforming the block group. Using a go/no-go fMRI paradigm, sequence preparation during the premovement study period was separated from movement execution. Imaging data for the two groups were compared for the first 1/3 and final 1/3 of training trials. Toward the end of training, behavioral performance between the two groups was similar, although the random group would later display a performance advantage on retention testing. During study time, the random group showed greater activity in sensorimotor and premotor regions compared to the block group. These areas are associated with motor preparation, sequencing, and response selection. This pattern of recruitment is consistent with the hypothesis that CI benefits in a sequencing task are due to improved capacity to actively prepare motor responses.


A Community-Based Culture Collection for Targeting Novel Plant Growth-Promoting Bacteria from the Sugarcane Microbiome.
The soil-plant ecosystem harbors an immense microbial diversity that challenges investigative approaches to study traits underlying plant-microbe association. Studies solely based on culture-dependent techniques have overlooked most microbial diversity. Here we describe the concomitant use of culture-dependent and -independent techniques to target plant-beneficial microbial groups from the sugarcane microbiome. The community-based culture collection (CBC) approach was used to access microbes from roots and stalks. The CBC recovered 399 unique bacteria representing 15.9% of the rhizosphere core microbiome and 61.6-65.3% of the endophytic core microbiomes of stalks. By cross-referencing the CBC (culture-dependent) with the sugarcane microbiome profile (culture-independent), we designed a synthetic community comprised of naturally occurring highly abundant bacterial groups from roots and stalks, most of which has been poorly explored so far. We then used maize as a model to probe the abundance-based synthetic inoculant. We show that when inoculated in maize plants, members of the synthetic community efficiently colonize plant organs, displace the natural microbiota and dominate at 53.9% of the rhizosphere microbial abundance. As a result, inoculated plants increased biomass by 3.4-fold as compared to uninoculated plants. The results demonstrate that abundance-based synthetic inoculants can be successfully applied to recover beneficial plant microbes from plant microbiota.


'Understanding and being understood' as a creative caring phenomenon--in care of patients with stroke and aphasia.
Five care providers particularly successful at communicating with patients with communication difficulties were video-recorded together with three patients with aphasia after stroke, during morning care activities. The care providers were then interviewed immediately after the video-recordings, about their experiences of communicating with such patients. The interviews with the care providers were interpreted by means of a phenomenological hermeneutic method. Co-creating was the main theme found. Care providers invite the patient to participate in the creative act of communication. They have a communicative attitude and show interest in the patients' personal desires. The care providers encounter the patient as a presence in a caring communion. In part, care providers communicate by continuously conveying their presence to the patient and even creating availability in a close and open intersubjective relationship. A relaxed and supportive atmosphere facilitates reciprocity between care provider and patient. The communication is not technical or strategic; instead care providers share the patients' experiences in a silent dialogue. This silent dialogue involves sharing the patients' feelings and thus receiving messages from the patient.


Induction of severe autoimmune disease in normal mice by simultaneous action of multiple immunostimulators.
Either of two immunostimulating factors (lpr, lipopolysaccharide) enhanced the pathogenic autoimmune responses of MRL/n mice, but the serologic and immunopathologic characteristics differed. In contrast, either factor acting alone, caused minimal immunopathology in normal mice, despite autoantibody induction. Combined immunostimulation, however, caused fatal glomerulonephritis in normal-background C57BL/6 mice. These results show the profound influence of the background genome on the effects of immunostimulating agents, and show that resistance to autoimmune disease in immunologically normal mice is not absolute.


Family Caregivers' Reflections on Experiences of Assisted Suicide in Switzerland: A Qualitative Interview Study.
Thousands of family members worldwide are annually involved in assisted dying. Family participation in assisted dying has rarely been investigated and families' needs typically are not considered in assisted dying legislation and clinical guidelines. To explore family caregivers' reflections on experiences of assisted suicide in Switzerland. A cross-sectional qualitative interview study conducted in the Italian- and French-speaking regions of Switzerland. Interpretation and analysis were performed using qualitative content analysis. Twenty-eight close relatives and family carers of 18 patients who died by assisted suicide in Switzerland were interviewed. Family members perceived their involvement in assisted suicide as characterized by five phases; 1) contemplation, 2) gaining acceptance, 3) gaining permission, 4) organization, and 5) aftermath. Families can participate in these phases at diverse levels and with varying degrees of involvement. Important triggers for families and patients for transition between phases include patients' experiences of their life-threatening illnesses and related treatments, their increasing awareness of approaching death, and family member recognition of their loved one's unbearable suffering. Participating in assisted suicide created further demanding tasks for families in addition to their role of caregivers. Families appeared to be involved in the preparation of assisted suicide along with patients, irrespective of their personal values regarding assisted dying. Support for family members is essential if they are involved in tasks preparatory to assisted suicide. Clinical guidelines and policies concerning assisted dying should acknowledge and address family needs.


Isolation and characterization of Schistosoma haematobium egg antigens.
Schistosoma haematobium soluble egg antigen (ShSEA) was prepared from eggs isolated from the livers of hamsters or mice infected for at least 3 months. Immunoaffinity purified S. haematobium egg antigens (ShSh) were isolated by first passing ShSEA through a column containing anti-S. mansoni hamster IgG coupled to CNBr-activated Sepharose 4B, and recycling the unbound fraction until no more bound material could be eluted with an acid wash. The unbound fraction was then filtered through a second antibody affinity column containing anti-S. haematobium hamster IgG, and in the acid eluate the ShSh antigens were obtained. This antigenic preparation was shown by PAGE to contain at least 6 distinct bands ranging in molecular weight (Mr) from 116 to less than 31 Kd. A 40 Kd polypeptide was identified by both silver staining and EITB as specific for S. haematobium eggs. In addition, a 55 Kd worm-egg shared antigen was identified as a prominent band in EITB expressed during a primary S. haematobium hamster infection. The sera from hamsters harboring patent S. haematobium or S. mansoni infections were reacted by ELISA with ShSh antigens. The anti-Sh sera showed significantly higher absorbance values than the anti-Sm sera, demonstrating that only a minor population of S. mansoni cross-reactive egg antigens is still present in the ShSh antigens. Sera collected weekly for 13 weeks from hamsters with a primary infection of S. haematobium were then tested by ELISA against ShSh, ShSEA and SmSEA antigens. Antibody levels against both ShSEA and SmSEA were shown to increase early in infection (2 weeks). Moreover, antibody levels to ShSh did not increase until week 5 post-infection. These findings suggest that the purification procedure utilized results in the elimination of most of the S. mansoni worm antigens cross-reactive with S. haematobium eggs. The ShSh antigens had shown a high degree of sensitivity and stage-species specificity also suggesting their potential as antigens for the immunodiagnosis of schistosomiasis haematobia.


Volume of blood required to detect common neonatal pathogens.
To determine the minimum volume of blood and the absolute number of organisms required for detection of bacteremia and fungemia by an automated colorimetric blood culture system (BacT/Alert, Organon Teknika). Common neonatal pathogens, Escherichia coli, Streptococcus agalactiae (group B streptococcus (GBS): one American Type Culture Collection (ATCC) strain and one clinical isolate), Staphylococcus epidermidis, and Candida albicans, were seeded into blood to produce bacteremia or fungemia with low colony counts (1 to 3 colony-forming units (CFU) per milliliter) and ultra-low colony counts (<1 CFU/ml). For each organism, 96 culture bottles were inoculated with either 0.25, 0.5, 1.0, or 4.0 ml of the two seeded blood concentrations. Blood culture bottles were incubated in the BacT/Alert device for 5 days, and time to positivity was noted when applicable. All bottles were subcultured on plated media. The Poisson statistic was used to calculate the probability of finding at least one viable CFU per inoculated culture bottle. The fraction of culture bottles with positive findings per group was divided by the probability of one or more organisms present to give the positivity index. Plated subculture identified no growth of organisms not detected by the colorimetric detection system. The false-positive rate for the automated device was less than 1%. The positivity index for the GBS clinical isolate was 1.13, for the GBS ATCC isolate 0.96, for S. epidermidis 0.94, for C. albicans 0.97, and for E. coli 0.95. There was a statistically significant difference with time to positivity and inocula volume (p <0.01), but the difference was not clinically important. If one or two viable colony-forming units are in the blood inoculated into culture media, the BacT/Alert system will detect growth rapidly. Because there appears to be a sizable subset of neonates who are at risk of sepsis with a colony count less than 4 CFU/ml, then a 0.5 ml inoculum of blood into the culture media is inadequate for sensitive and timely detection of bacteremia. One to two milliliters of blood should increase microorganism recovery in the face of low-colony-count sepsis.


Antimicrobial agent resistance in mycobacteria: molecular genetic insights.
The primary theme emerging from molecular genetic work conducted with Mycobacterium tuberculosis and several other mycobacterial species is that resistance is commonly associated with simple nucleotide alterations in target chromosomal genes rather than with acquisition of new genetic elements encoding antibiotic-altering enzymes. Mutations in an 81-bp region of the gene (rpoB) encoding the beta subunit of RNA polymerase account for rifampin resistance in 96% of M. tuberculosis and many Mycobacterium leprae isolates. Streptomycin resistance in about one-half of M. tuberculosis isolates is associated with missense mutations in the rpsL gene coding for ribosomal protein S12 or nucleotide substitutions in the 16S rRNA gene (rrs). Mutations in the katG gene resulting in catalase-peroxidase amino acid alterations nad nucleotide substitutions in the presumed regulatory region of the inhA locus are repeatedly associated with isoniazid-resistant M. tuberculosis isolates. A majority of fluoroquinolone-resistant M. tuberculosis isolates have amino acid substitutions in a region of the DNA gyrase A subunit homologous to a conserved fluoroquinolone resistance-determining region. Multidrug-resistant isolates of M. tuberculosis arise as a consequence of sequential accumulation of mutations conferring resistance to single therapeutic agents. Molecular strategies show considerable promise for rapid detection of mutations associated with antimicrobial resistance. These approaches are now amenable to utilization in an appropriately equipped clinical microbiology laboratory.


Search asymmetries? What search asymmetries?
In order to establish a search asymmetry, one must run an experiment with a symmetric design and get asymmetric results. Given an asymmetric design, one expects asymmetric results, and such results do not imply an asymmetry in the search mechanisms. In this paper, I argue that a number of experiments purporting to show search asymmetries contain built-in design asymmetries. A saliency model of visual search predicts the results of these experiments, using only a simple measure of target-distractor similarity, without reliance on asymmetric search mechanisms. These results have implications for search mechanisms and for other experiments purporting to show search asymmetries.


Allograft inflammatory factor-1 in myeloid cells drives autoimmunity in type 1 diabetes.
Allograft inflammatory factor-1 (AIF1) is a calcium-responsive cytoplasmic scaffold protein that directs hematopoiesis and immune responses within dendritic cells (DC) and macrophages. Although the role of AIF1 in transplant rejection and rheumatoid arthritis has been explored, little is known about its role in type 1 diabetes. Here, we show that in vivo silencing of AIF1 in NOD mice restrained infiltration of immune cells into the pancreas and inhibited diabetes incidence. Analyses of FACS-sorted CD45neg nonleukocyte populations from resected pancreatic islets showed markedly higher expression of insulin in the AIF1-silenced groups. Evaluation of CD45+ leukocytes revealed diminished infiltration of effector T cells and DC in the absence of AIF1. Transcriptional profiling further revealed a marked decrease in cDC1 DC-associated genes CD103, BATF3, and IRF8, which are required for orchestrating polarized type 1 immunity. Reduced T cell numbers within the islets were observed, with concomitant lower levels of IFN-γ and T-bet in AIF1-silenced cohorts. In turn, there was a reciprocal increase in functionally suppressive pancreas-resident CD25+Foxp3+CD4+ Tregs. Taken together, results show that AIF1 expression in myeloid cells plays a pivotal role in promoting type 1 diabetes and that its suppression restrains insulitis by shifting the immune microenvironment toward tolerance.


The importance of considering community-level effects when selecting insecticidal malaria vector products.
Insecticide treatment of nets, curtains or walls and ceilings of houses represent the primary means for malaria prevention worldwide. Direct personal protection of individuals and households arises from deterrent and insecticidal activities which divert or kill mosquitoes before they can feed. However, at high coverage, community-level reductions of mosquito density and survival prevent more transmission exposure than the personal protection acquired by using a net or living in a sprayed house. A process-explicit simulation of malaria transmission was applied to results of 4 recent Phase II experimental hut trials comparing a new mosaic long-lasting insecticidal net (LLIN) which combines deltamethrin and piperonyl butoxide with another LLIN product by the same manufacturer relying on deltamethrin alone. Direct estimates of mean personal protection against insecticide-resistant vectors in Vietnam, Cameroon, Burkina Faso and Benin revealed no clear advantage for combination LLINs over deltamethrin-only LLINs (P = 0.973) unless both types of nets were extensively washed (Relative mean entomologic inoculation rate (EIR) ± standard error of the mean (SEM) for users of combination nets compared to users of deltamethrin only nets = 0.853 ± 0.056, P = 0.008). However, simulations of impact at high coverage (80% use) predicted consistently better impact for the combination net across all four sites (Relative mean EIR ± SEM in communities with combination nets, compared with those using deltamethrin only nets = 0.613 ± 0.076, P < 0.001), regardless of whether the nets were washed or not (P = 0.467). Nevertheless, the degree of advantage obtained with the combination varied substantially between sites and their associated resistant vector populations. Process-explicit simulations of community-level protection, parameterized using locally-relevant experimental hut studies, should be explicitly considered when choosing vector control products for large-scale epidemiological trials or public health programme procurement, particularly as growing insecticide resistance necessitates the use of multiple active ingredients.


MicroRNA-mediated integration of haemodynamics and Vegf signalling during angiogenesis.
Within the circulatory system, blood flow regulates vascular remodelling, stimulates blood stem cell formation, and has a role in the pathology of vascular disease. During vertebrate embryogenesis, vascular patterning is initially guided by conserved genetic pathways that act before circulation. Subsequently, endothelial cells must incorporate the mechanosensory stimulus of blood flow with these early signals to shape the embryonic vascular system. However, few details are known about how these signals are integrated during development. To investigate this process, we focused on the aortic arch (AA) blood vessels, which are known to remodel in response to blood flow. By using two-photon imaging of live zebrafish embryos, we observe that flow is essential for angiogenesis during AA development. We further find that angiogenic sprouting of AA vessels requires a flow-induced genetic pathway in which the mechano-sensitive zinc finger transcription factor klf2a induces expression of an endothelial-specific microRNA, mir-126, to activate Vegf signalling. Taken together, our work describes a novel genetic mechanism in which a microRNA facilitates integration of a physiological stimulus with growth factor signalling in endothelial cells to guide angiogenesis.


A Chemical Probe for Tudor Domain Protein Spindlin1 to Investigate Chromatin Function.
Modifications of histone tails, including lysine/arginine methylation, provide the basis of a "chromatin or histone code". Proteins that contain "reader" domains can bind to these modifications and form specific effector complexes, which ultimately mediate chromatin function. The spindlin1 (SPIN1) protein contains three Tudor methyllysine/arginine reader domains and was identified as a putative oncogene and transcriptional coactivator. Here we report a SPIN1 chemical probe inhibitor with low nanomolar in vitro activity, exquisite selectivity on a panel of methyl reader and writer proteins, and with submicromolar cellular activity. X-ray crystallography showed that this Tudor domain chemical probe simultaneously engages Tudor domains 1 and 2 via a bidentate binding mode. Small molecule inhibition and siRNA knockdown of SPIN1, as well as chemoproteomic studies, identified genes which are transcriptionally regulated by SPIN1 in squamous cell carcinoma and suggest that SPIN1 may have a role in cancer related inflammation and/or cancer metastasis.


β-arrestin-2 is an essential regulator of pancreatic β-cell function under physiological and pathophysiological conditions.
β-arrestins are critical signalling molecules that regulate many fundamental physiological functions including the maintenance of euglycemia and peripheral insulin sensitivity. Here we show that inactivation of the β-arrestin-2 gene, barr2, in β-cells of adult mice greatly impairs insulin release and glucose tolerance in mice fed with a calorie-rich diet. Both glucose and KCl-induced insulin secretion and calcium responses were profoundly reduced in β-arrestin-2 (barr2) deficient β-cells. In human β-cells, barr2 knockdown abolished glucose-induced insulin secretion. We also show that the presence of barr2 is essential for proper CAMKII function in β-cells. Importantly, overexpression of barr2 in β-cells greatly ameliorates the metabolic deficits displayed by mice consuming a high-fat diet. Thus, our data identify barr2 as an important regulator of β-cell function, which may serve as a new target to improve β-cell function.


Testing the use of practice facilitation in a cluster randomized stepped-wedge design trial to improve adherence to cardiovascular disease prevention guidelines: HealthyHearts NYC.
HealthyHearts NYC (HHNYC) will evaluate the effectiveness of practice facilitation as a quality improvement strategy for implementing the Million Hearts' ABCS treatment guidelines for reducing cardiovascular disease (CVD) among high-risk patients who receive care in primary care practices in New York City. ABCS refers to (A) aspirin in high-risk individuals; (B) blood pressure control; (C) cholesterol management; and (S) smoking cessation. The long-term goal is to create a robust infrastructure for implementing and disseminating evidence-based practice guidelines (EBPG) in primary care practices. We are using a stepped-wedge cluster randomized controlled trial design to evaluate the implementation process and the impact of practice facilitation (PF) versus usual care on ABCS outcomes in 250 small primary care practices. Randomization is at the practice site level, all of which begin as part of the control condition. The intervention consists of one year of PF that includes a combination of one-on-one onsite visits and shared learning across practice sites. PFs will focus on helping sites implement evidence-based components of patient-centered medical home (PCMH) and the chronic care model (CCM), which include decision support, provider feedback, self-management tools and resources, and linkages to community-based services. We hypothesize that practice facilitation will result in superior clinical outcomes compared to usual care; that the effects of practice facilitation will be mediated by greater adoption of system changes in accord with PCMH and CCM; and that there will be increased adaptive reserve and change capacity. NCT02646488.


Adenophostin A can stimulate Ca2+ influx without depleting the inositol 1,4,5-trisphosphate-sensitive Ca2+ stores in the Xenopus oocyte.
Adenophostin A possesses the highest known affinity for the inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) receptor (InsP3R). The compound shares with Ins(1,4,5)P3 those structural elements essential for binding to the InsP3R. However, its adenosine 2'-phosphate moiety has no counterpart in the Ins(1,4,5)P3 molecule. To determine whether its unique structure conferred a distinctive biological activity, we characterized the adenophostin-induced Ca2+ signal in Xenopus oocytes using the Ca2+-gated Cl- current assay. In high concentrations, adenophostin A released Ca2+ from Ins(1,4, 5)P3-sensitive stores and stimulated a Cl- current that depended upon the presence of extracellular Ca2+. We used this Cl- current as a marker of Ca2+ influx. In low concentrations, however, adenophostin A stimulated Ca2+ influx exclusively. In contrast, Ins(1,4,5)P3 and (2-hydroxyethyl)-alpha-D-glucopyranoside 2',3, 4-trisphosphate, an adenophostin A mimic lacking most of the adenosine moiety, always released intracellular Ca2+ before causing Ca2+ influx. Ins(1,4,5)P3 could still release Ca2+ during adenophostin A-induced Ca2+ influx, confirming that the Ins(1,4, 5)P3-sensitive intracellular Ca2+ stores had not been emptied. Adenophostin- and Ins(1,4,5)P3-induced Ca2+ influx were not additive, suggesting that both agonists stimulated a common Ca2+ entry pathway. Heparin, which blocks binding to the InsP3R, prevented adenophostin-induced Ca2+ influx. These data indicate that adenophostin A can stimulate the influx of Ca2+ across the plasma membrane without inevitably emptying the Ins(1,4,5)P3-sensitive intracellular Ca2+ stores.


Does clinical error contribute to unnecessary antibiotic use?
Patient expectations and physician attitudes are often cited as factors in the overuse of antibiotics. This study examined whether clinical error might also be important. In treating 517 patients with sore throat, family physicians estimated the probability that group A streptococcus infection was present. Two thirds of antibiotics prescribed were to culture-negative patients and therefore considered unnecessary. Physicians overestimated the probability that a group A streptococcal infection was present by an average 33.2% in these cases, compared with 6.9% otherwise (p < 0.001). The rate of unnecessary prescribing was 5.1% when the physician estimate differed from the true probability of a group A streptococcal infection by <10%, 16.0% for an error of 10-29%, 35.6% for an error of 30-49%, and 78.3% when the chance of the infection was overestimated by 50% or more. Clinical error in estimating the likelihood of group A streptococcal infection probably contributes to unnecessary antibiotic use in patients with sore throat.


A human alcohol dehydrogenase gene (ADH6) encoding an additional class of isozyme.
The human alcohol dehydrogenase (ADH; alcohol:NAD+ oxidoreductase, EC 126.96.36.199) gene family consists of five known loci (ADH1-ADH5), which have been mapped close together on chromosome 4 (4q21-25). ADH isozymes encoded by these genes are grouped in three distinct classes in terms of their enzymological properties. A moderate structural similarity is observed between the members of different classes. We isolated an additional member of the ADH gene family by means of cross-hybridization with the ADH2 (class I) cDNA probe. cDNA clones corresponding to this gene were derived from PCR-amplified libraries as well. The coding sequence of a 368-amino-acid-long open reading frame was interrupted by introns into eight exons and spanned approximately 17 kilobases on the genome. The gene contains a glucocorticoid response element at the 5' region. The transcript was detected in the stomach and liver. The deduced amino acid sequence of the open reading frame showed about 60% positional identity with known human ADHs. This extent of homology is comparable to interclass similarity in the human ADH family. Thus, the newly identified gene, which is designated ADH6, governs the synthesis of an enzyme that belongs to another class of ADHs presumably with a distinct physiological role.


Autologous hematopoietic stem-cell transplantation for relapsed or refractory Hodgkin's disease in children and adolescents.
To determine the treatment outcome and clinical factors that are of prognostic significance for children and adolescents with relapsed or refractory Hodgkin's disease (HD) who received treatment with high-dose chemotherapy and autologous hematopoietic stem-cell transplantation (HSCT). Fifty-three consecutive children and adolescents 21 years of age or younger with relapsed or refractory HD underwent HSCT. At day 100 after transplantation, 29 patients (55%) were in a complete remission or maintained a continuous complete response, six (11%) had a partial response, and 11 (21%) failed to respond or had progressive disease. The failure-free survival (FFS) at 5 years was 31%, and overall survival was 43%. Twenty-one patients died of progressive HD, and nine died secondary to transplantation-related complications, including two secondary leukemias. Prognostic factors important for FFS were normal pretransplantation lactate dehydrogenase levels (5-year FFS = 42%), compared with patients with elevated LDH levels (5-year FFS = 0%) (P < .001), and disease sensitivity at the time of HSCT with FFS in untreated relapse, sensitive disease, and resistant disease 44%, 35%, and 9%, respectively (P = .06). There was no statistically significant difference in FFS or overall survival between age subgroups that were analyzed (< 13, 13 to 18, 19 to 21) or in comparison with an adult cohort. HSCT is an effective treatment modality that can result in long-term cures and should be considered for children and adolescents with relapsed HD.


Effects of environmental heat on conception rates in lactating dairy cows: critical periods of exposure.
Environmental heat can reduce conception rates (the proportion of services that result in pregnancy) in lactating dairy cows. The study objectives were to identify periods of exposure relative to the service date in which environmental heat is most closely associated with conception rates, and to assess whether the total time cows are exposed to high environmental heat within each 24-h period is more closely associated with conception rates than is the maximum environmental heat for each 24-h period. A retrospective observational study was conducted in 25 predominantly Holstein-Friesian commercial dairy herds located in Australia. Associations between weather and conception rates were assessed using 16,878 services performed over a 21-mo period. Services were classified as successful based on rectal palpation. Two measures of heat load were defined for each 24-h period: the maximum temperature-humidity index (THI) for the period, and the number of hours in the 24-h period when the THI was >72. Conception rates were reduced when cows were exposed to a high heat load from the day of service to 6 d after service, and in wk -1. Heat loads in wk -3 to -5 were also associated with reduced conception rates. Thus, management interventions to ameliorate the effects of heat load on conception rates should be implemented at least 5 wk before anticipated service and should continue until at least 1 wk after service. High autocorrelations existed between successive daily values in both measures, and associations between day of heat load relative to service day and conception rates differed substantially when ridge regression was used to account for this autocorrelation. This indicates that when assessing the effects of heat load on conception rates, the autocorrelation in heat load between days should be accounted for in analyses. The results suggest that either weekly averages or totals summarizing the daily heat load are adequate to describe heat load when assessing effects on conception rates in lactating dairy cows.


Prader-willi syndrome: A case report and a Chinese literature review.
Prader-Willi syndrome (PWS) is a genetic disorder, resulting from lack of gene expression on the paternally inherited chromosome 15. It is important to determine diagnostic methods for PWS for early treatment. In this study, we report a newborn with Prader-willi syndrome. We further summarized the genetic testing results in the Chinese literature and the relevance of high resolution chromosome and genome-wide copy number variation analysis. There is a heterozygosis deletion of a 5 Mb region in the paternal chromosome 15q11.3-q13.3 by genome-wide copy number variation analysis. However, there is no abnormality in high resolution chromosome karyotype analysis. In conclusion, genome-wide copy number variation analysis is an effective and specific diagnosis method, which will provide scientific evidence for the clinical diagnosis and early treatment of PWS.


Using peptide loop insertion mutagenesis for the evolution of proteins.
The insertion of peptide loops into the polypeptide chain of proteins at surface-exposed regions is an attractive avenue to modify the protein's properties or to evolve new functionalities. The strategy of peptide loop insertion has, for example, been used to create new binding sites in protein scaffolds and has led to the isolation of proteins with excellent binding affinities for various biological structures. Peptide loops have also been inserted into enzymes to modulate their catalytic properties. We recently used loop insertion mutagenesis to evolve a mutant of O(6)-alkylguanine-DNA-alkyltransferase (AGT) that reacts with the nonnatural substrate O(6)-propargylguanine for applications in molecular imaging. In this chapter, we describe in detail the protocols that we have applied (1) to identify sites in AGT that are permissive to loop insertion, (2) to manipulate DNA to create large loop insertion libraries, and (3) to identify mutants with desired properties. The experimental procedures are general and can easily be adapted for the insertion of peptide loops into other classes of enzymes or into any protein of interest.


Characterization of the antibody response elicited by immunization with pneumococcal surface protein A (PspA) as recombinant protein or DNA vaccine and analysis of protection against an intranasal lethal challenge with Streptococcus pneumoniae.
Pneumococcal surface protein A (PspA) is an important candidate for a vaccine against pneumococcal infections. DNA vaccines expressing PspA were shown to protect mice against intraperitoneal and colonization challenge models in mice. We now show that a DNA vaccine expressing PspA from clade 4 (pSec-pspA4Pro) is also able to elicit protection against an intranasal lethal challenge model at levels similar to the recombinant protein PspA4Pro adjuvanted with alum. PspA4Pro + alum induced an IgG response characterized by a high IgG1/IgG2a ratio, leading to a lack of binding of anti-PspA IgG2a antibodies to intact pneumococci in vitro, which is in contrast to the response elicited by pSec-pspA4Pro. Epitopes recognized by the sera were mapped and antibodies induced by immunization with PspA4Pro + alum showed positive reaction with several synthetic peptides, mostly located in the first half of the protein. On the other hand, antibodies induced by the DNA vaccine showed reactivity with only two peptides. Though both strategies were protective against the intranasal lethal challenge model, the elicited humoral responses differ significantly, with the detection of important differences in the Fc (IgG1/IgG2a ratios) and Fab (recognized epitopes) regions of the induced antibodies.


Comparative Efficacy of Insecticides on Bactrocera tryoni and Zeugodacus cucumis (Diptera: Tephritidae) in Laboratory and Semifield Trials in Fruiting Vegetables.
In-field management of Bactrocera tryoni (Froggatt) and Zeugodacus cucumis (French) (Diptera: Tephritidae) in fruiting vegetable crops has relied almost exclusively on organophosphate cover sprays. Laboratory and semifield trials were performed to compare a number of alternative insecticides for efficacy against these species. A novel semifield method was used whereby the insecticides were applied to crops as cover sprays under field conditions, and treated plants bearing fruit were transferred to large cages and exposed to fruit flies. Efficacy was assessed in terms of numbers of pupae developing from treated fruit. A laboratory cage method was also used to assess effects on adult mortality and comparative effects of 1- and 3-d-aged residues. The neonicotinoids clothianidin and thiacloprid were very effective against B. tryoni and Z. cucumis. Clothianidin was the only insecticide other than dimethoate to affect adult mortality. The synthetic pyrethroid alpha-cypermethrin was also very effective, particularly in semifield trials, although higher incidence of aphid and whitefly infestation was observed in this treatment compared to others. Cyantraniliprole was effective against B. tryoni, but less effective against Z. cucumis. Imidacloprid, bifenthrin, spinetoram, and abamectin were all relatively less effective, although all demonstrated a suppressive effect.


A Net Mold-Based Method of Biomaterial-Free Three-Dimensional Cardiac Tissue Creation.
Ischemic cardiomyopathy poses a significant public health burden due to the irreversible loss of functional cardiac tissue. Alternative treatment strategies include creation of three-dimensional (3D) cardiac tissues to both replace and augment injured native tissue. In this study, we utilize a net mold-based method to create a biomaterial-free 3D cardiac tissue and compare it to current methods using biomaterials. Cardiomyocytes, fibroblasts, and endothelial cells were combined using a hanging drop method to create spheroids. For the net mold patch method, spheroids were seeded into a net mold-based system to create biomaterial-free 3D cardiac patches. For the gel patch, spheroids were embedded in a collagen gel. Immunohistochemistry revealed increased alignment, vascularization, collagen I expression, cell viability, and higher density of cells in the net mold patch compared with the gel patch. Furthermore, in vivo testing in a left anterior descending artery ligation rat model found increased ejection fraction and smaller scar area following implantation of the net mold patch. We present a novel and simple reproducible method to create biomaterial-free 3D net mold patches that may potentially improve the treatment of heart failure in the future.


Localisation and functional significance of ryanodine receptors during beta-adrenoceptor stimulation in the guinea-pig sino-atrial node.
Recent evidence shows that calcium released from the sarcoplasmic reticulum (SR) plays an important role in the regulation of heart rate. The aim of this study was to investigate the subcellular distribution of ryanodine receptors in the guinea-pig sino-atrial (SA) node and to determine their functional role in the regulation of pacemaker frequency in response to beta-adrenoceptor stimulation. Monoclonal antibodies raised against the cardiac ryanodine receptor were used with confocal microscopy to investigate ryanodine receptor distribution in single guinea-pig SA node cells. The functional role of ryanodine receptors was investigated in both multicellular SA node/atrial preparations and in single SA node cells. Ryanodine receptor labelling was observed in all SA node cells studied and showed both subsarcolemmal and intracellular staining. In the latter, labelling appeared as transverse bands with a regular periodicity of approximately 2 microm. This interval resembled that of the expected sarcomere spacing but did not, however, depend on the presence of transverse tubules. The bands of ryanodine receptors appeared to be located in the region of the Z lines, based on co-distribution studies with antibodies to alpha-actinin, myomesin and binding sites for phalloidin. Functional studies on single SA node cells showed that application of ryanodine (2 micromol/l) reduced the rate of firing of spontaneous action potentials (measured using the perforated patch clamp technique) and this was associated with changes in action potential characteristics. Ryanodine also significantly decreased the positive chronotropic actions of isoprenaline in both multicellular and single cell preparations. In single cells exposed to 100 nmol/l isoprenaline, ryanodine caused a decrease in the rate of firing and this was associated with a decrease in the amplitude of the measured calcium transients. These findings are the first to show immunocytochemical evidence for the presence and organisation of ryanodine receptor calcium release channels in mammalian SA node cells. This study also provides evidence of a role for ryanodine sensitive sites in the beta-adrenergic modulation of heart rate in this species.


Weight and age at menarche.
In the National Child Development Study (1958 cohort) information on their age at menarche and their weights and heights measured at 7, 11, and 16 years was available for 4427 girls. The distribution of age at menarche was not influenced by social class. Weight adjusted for height did not play an important part in the timing of sexual maturation of the girls in the study. Relative weight (weight expressed as a percentage of standard weight) at the ages of 7 and 11 years explained only 3.2%, and 4.9%, respectively of the variation in age at menarche, and changes in relative weight between these two ages accounted for 2%. Girls with early menarche were more likely to be overweight at ages 7, 11, and 16 years than those with late menarche, although early menarche was also reported by girls who were underweight or of average weight. These findings support the hypothesis that in well nourished populations the relation between menarche and body size is largely regulated by genetic factors and that nutrition is less important.


Time-Reversal-Breaking Weyl Fermions in Magnetic Heusler Alloys.
Weyl fermions have recently been observed in several time-reversal-invariant semimetals and photonics materials with broken inversion symmetry. These systems are expected to have exotic transport properties such as the chiral anomaly. However, most discovered Weyl materials possess a substantial number of Weyl nodes close to the Fermi level that give rise to complicated transport properties. Here we predict, for the first time, a new family of Weyl systems defined by broken time-reversal symmetry, namely, Co-based magnetic Heusler materials XCo_{2}Z (X=IVB or VB; Z=IVA or IIIA). To search for Weyl fermions in the centrosymmetric magnetic systems, we recall an easy and practical inversion invariant, which has been calculated to be -1, guaranteeing the existence of an odd number of pairs of Weyl fermions. These materials exhibit, when alloyed, only two Weyl nodes at the Fermi level-the minimum number possible in a condensed matter system. The Weyl nodes are protected by the rotational symmetry along the magnetic axis and separated by a large distance (of order 2π) in the Brillouin zone. The corresponding Fermi arcs have been calculated as well. This discovery provides a realistic and promising platform for manipulating and studying the magnetic Weyl physics in experiments.


Avoidant personality disorder symptoms in first-degree relatives of schizophrenia patients predict performance on neurocognitive measures: the UCLA family study.
Whether avoidant personality disorder symptoms are related to neurocognitive impairments that aggregate in relatives of schizophrenics is unknown. We report the relationship between avoidant personality disorder symptoms and neurocognitive performance in the first-degree relatives of probands with schizophrenia. 367 first-degree relatives of probands with schizophrenia and 245 relatives of community controls were interviewed for the presence of avoidant personality symptoms and symptoms of paranoid and schizotypal personality disorders and administered neurocognitive measures. Relationships between neurocognitive measures and avoidant symptoms were analyzed using linear mixed models. Avoidant dimensional scores predicted performance on the span of apprehension (SPAN), 3-7 Continuous Performance Test (3-7 CPT), and Trail Making Test (TMT-B) in schizophrenia relatives. These relationships remained significant on the SPAN even after adjustment for paranoid or schizotypal dimensional scores and on the TMT-B after adjustment for paranoid dimensional scores. Moreover, in a second set of analyses comparing schizophrenia relatives to controls there were significant or trending differences in the degree of the relationship between avoidant symptoms and each of these neurocognitive measures even after adjustments for paranoid and schizotypal dimensional scores. The substantial correlation between avoidant and schizotypal symptoms suggests that these personality disorders are not independent. Avoidant and in some cases schizotypal dimensional scores are significant predictors of variability in these neurocognitive measures. In all analyses, higher levels of avoidant symptoms were associated with worse performance on the neurocognitive measures in relatives of schizophrenia probands. These results support the hypothesis that avoidant personality disorder may be a schizophrenia spectrum phenotype.


Studies on the pathogenesis of fever. VI. The interaction of leucocytes and endotoxin in vitro.
Study in vitro of the interaction of bacterial endotoxin with rabbit polymorphonuclear leucocytes has resulted in the following findings: 1. Incubation of endotoxin and leucocytes in saline results in: (a) the release of leucocytic pyrogen, and (b) the inactivation of endotoxin. 2. Cell-free extracts of leucocytes also inactivate endotoxin. 3. Incubation of leucocytes in "physiological" saline causes rapid discharge of leucocytic pyrogen. In contrast, relatively little pyrogen is released by leucocytes incubated in fresh serum. 4. The release of leucocytic pyrogen in serum is markedly stimulated by the presence of endotoxin. 5. Leucocytes obtained from tolerant rabbits interact with endotoxin in essentially the same manner as leucocytes from normal rabbits. The pertinence of these findings to the pathogenesis of fever and to related information concerning human leucocytes has been discussed.


Significance of Comorbid Psychological Stress and Depression on Outcomes After Cardiac Rehabilitation.
Depression is associated with increased mortality in stable coronary heart disease. Cardiac rehabilitation and exercise training has been shown to decrease depression, psychological stress, and mortality after a coronary heart disease event. The presence of depression at completion of cardiac rehabilitation and exercise training is associated with increased mortality. However, it is unknown if depression with comorbid psychological risk factors such as anxiety or hostility confers an additional mortality disadvantage. We evaluated the mortality effect of anxiety and hostility on depression after cardiac rehabilitation and exercise training. We studied 1150 patients with coronary heart disease following major coronary heart disease events who had completed formal cardiac rehabilitation and exercise training. Using Kellner questionnaires, stress levels were measured in 1 of 3 domains: anxiety, hostility, and depression (with an aggregated overall psychological stress score) and divided into 3 groups: nondepressed (n = 1072), depression alone (n = 18), and depression with anxiety or hostility (n = 60). Subjects were analyzed for all-cause mortality over 161 months of follow-up (mean 6.4 years) by National Death Index. Depression after cardiac rehabilitation was not common (6.8%; mortality 20.8%) but when present, frequently associated with either anxiety or hostility (77% of depressed patients; mortality 22.0%). After adjustment for age, sex, ejection fraction, and baseline peak oxygen consumption, depression alone (hazard ratio [HR] 1.73, P = .04), as well as depression with comorbid psychological stress, was associated with higher mortality (HR 1.98, P = .03). Furthermore, our data showed an increased mortality when both anxiety and hostility were present in addition to depression after cardiac rehabilitation (HR 2.41, P = .04). After cardiac rehabilitation, depression, when present, is usually associated with other forms of psychological stress, which confers additional mortality. More measures are needed to address psychological stress after cardiac rehabilitation.


Factors affecting use of permethrin-treated bed nets during a randomized controlled trial in western Kenya.
Adherence with permethrin-treated bed net (ITN) use and their proper deployment was directly observed in 2,178 individuals (784 households) participating in a large-scale trial of ITNs on child mortality in western Kenya. The ITNs were distributed free of charge to ensure high coverage, resulting in a ratio of 1.46 persons per ITN. Approximately 30% of ITNs present were unused. The overall percentage adherence was 72.3%. The probability of adherence by individuals depended strongly on age (relative risk [RR] = 0.86, 95% confidence limit [CL] = 0.78-0.94), in which children less than five years of age were less likely to use ITNs than older individuals, and temperature, in which ITNs were more likely to be used in periods of cooler weather. A marginally significant diminution in adherence during the second year of the project was also observed (RR = 0.83, 95% CL = 0.68-1.01). Mosquito numbers, relative wealth, number of house occupants, and the educational level of the head of the household had no effect on adherence. In unstructured questioning of house residents, excessive heat was often cited as a reason for not deploying the child's ITN. The most important reason for non-adherence was disruption of sleeping arrangements, indicating that ITNs were not readily redeployed in the face of shifting sleeping patterns due to visitors, funerals, house construction, and other events. Challenges faced by health education programs to maximize adherence with ITN use are discussed.


Prevalence of Single and Multiple Natural NS3, NS5A and NS5B Resistance-Associated Substitutions in Hepatitis C Virus Genotypes 1-4 in Italy.
Natural resistance-associated substitutions (RASs) are reported with highly variable prevalence across different HCV genotypes (GTs). Frequency of natural RASs in a large Italian real-life cohort of patients infected with the 4 main HCV-GTs was investigated. NS3, NS5A and NS5B sequences were analysed in 1445 HCV-infected DAA-naïve patients. Sanger-sequencing was performed by home-made protocols on 464 GT1a, 585 GT1b, 92 GT2c, 199 GT3a, 16 GT4a and 99 GT4d samples. Overall, 20.7% (301/1455) of patients showed natural RASs, and the prevalence of multiclass-resistance was 7.3% (29/372 patients analysed). NS3-RASs were particularly common in GT1a and GT1b (45.2-10.8%, respectively), mainly due to 80K presence in GT1a (17%). Almost all GTs showed high prevalence of NS5A-RASs (range: 10.2-45.4%), and especially of 93H (5.1%). NS5A-RASs with fold-change >100x were detected in 6.8% GT1a (30H/R-31M-93C/H), 10.3% GT1b (31V-93H), 28.4% GT2c (28C-31M-93H), 8.5% GT3a (30K-93H), 45.5% GT4a (28M-30R-93H) and 3.8% GT4d (28V-30S-93H). Sofosbuvir RAS 282T was never detected, while the 159F and 316N RASs were found in GT1b (13.4-19.1%, respectively). Natural RASs are common in Italian patients infected with HCV-GTs 1-4. High prevalence of clinically-relevant RASs (such as Y93H) supports the appropriateness of HCV resistance-test to properly guide DAA-based therapy.


Connectome hubs at resting state in children and adolescents: Reproducibility and psychopathological correlation.
Functional brain hubs are key integrative regions in brain networks. Recently, brain hubs identified through resting-state fMRI have emerged as interesting targets to increase understanding of the relationships between large-scale functional networks and psychopathology. However, few studies have directly addressed the replicability and consistency of the hub regions identified and their association with symptoms. Here, we used the eigenvector centrality (EVC) measure obtained from graph analysis of two large, independent population-based samples of children and adolescents (7-15 years old; total N=652; 341 subjects for site 1 and 311 for site 2) to evaluate the replicability of hub identification. Subsequently, we tested the association between replicable hub regions and psychiatric symptoms. We identified a set of hubs consisting of the anterior medial prefrontal cortex and inferior parietal lobule/intraparietal sulcus (IPL/IPS). Moreover, lower EVC values in the right IPS were associated with psychiatric symptoms in both samples. Thus, low centrality of the IPS was a replicable sign of potential vulnerability to mental disorders in children. The identification of critical and replicable hubs in functional cortical networks in children and adolescents can foster understanding of the mechanisms underlying mental disorders.


Ptbp2 represses adult-specific splicing to regulate the generation of neuronal precursors in the embryonic brain.
Two polypyrimidine tract RNA-binding proteins (PTBs), one near-ubiquitously expressed (Ptbp1) and another highly tissue-restricted (Ptbp2), regulate RNA in interrelated but incompletely understood ways. Ptbp1, a splicing regulator, is replaced in the brain and differentiated neuronal cell lines by Ptbp2. To define the roles of Ptbp2 in the nervous system, we generated two independent Ptbp2-null strains, unexpectedly revealing that Ptbp2 is expressed in neuronal progenitors and is essential for postnatal survival. A HITS-CLIP (high-throughput sequencing cross-linking immunoprecipitation)-generated map of reproducible Ptbp2-RNA interactions in the developing mouse neocortex, combined with results from splicing-sensitive microarrays, demonstrated that the major action of Ptbp2 is to inhibit adult-specific alternative exons by binding pyrimidine-rich sequences upstream of and/or within them. These regulated exons are present in mRNAs encoding proteins associated with control of cell fate, proliferation, and the actin cytoskeleton, suggesting a role for Ptbp2 in neurogenesis. Indeed, neuronal progenitors in the Ptbp2-null brain exhibited an aberrant polarity and were associated with regions of premature neurogenesis and reduced progenitor pools. Thus, Ptbp2 inhibition of a discrete set of adult neuronal exons underlies early brain development prior to neuronal differentiation and is essential for postnatal survival.


Evaluation and implementation of chemotherapy regimen validation in an electronic health record.
Computerized provider order entry of chemotherapy regimens is quickly becoming the standard for prescribing chemotherapy in both inpatient and ambulatory settings. One of the difficulties with implementation of chemotherapy regimen computerized provider order entry lies in verifying the accuracy and completeness of all regimens built in the system library. Our goal was to develop, implement, and evaluate a process for validating chemotherapy regimens in an electronic health record. We describe our experience developing and implementing a process for validating chemotherapy regimens in the setting of a standard, commercially available computerized provider order entry system. The pilot project focused on validating chemotherapy regimens in the adult inpatient oncology setting and adult ambulatory hematologic malignancy setting. A chemotherapy regimen validation process was defined as a result of the pilot project. Over a 27-week pilot period, 32 chemotherapy regimens were validated using the process we developed. Results of the study suggest that by validating chemotherapy regimens, the amount of time spent by pharmacists in daily chemotherapy review was decreased. In addition, the number of pharmacist modifications required to make regimens complete and accurate were decreased. Both physician and pharmacy disciplines showed improved satisfaction and confidence levels with chemotherapy regimens after implementation of the validation system. Chemotherapy regimen validation required a considerable amount of planning and time but resulted in increased pharmacist efficiency and improved provider confidence and satisfaction.


Repeat Emergency Department Visits for Individuals With Intellectual and Developmental Disabilities and Psychiatric Disorders.
Although individuals with intellectual and developmental disabilities (IDD) and psychiatric concerns are more likely than others to visit hospital emergency departments (EDs), the frequency of their returns to the ED within a short time is unknown. In this population-based study we examined the likelihood of this group returning to the ED within 30 days of discharge and described these visits for individuals with IDD + psychiatric disorders (n = 3,275), and persons with IDD only (n = 1,944) compared to persons with psychiatric disorders only (n = 41,532). Individuals with IDD + psychiatric disorders, and individuals with IDD alone were more likely to make 30-day repeat ED visits. Improving hospital care and postdischarge community linkages may reduce 30-day returns to the ED among adults with IDD.


Global and Regional Respiratory Mechanics During Robotic-Assisted Laparoscopic Surgery: A Randomized Study.
Pneumoperitoneum and nonphysiological positioning required for robotic surgery increase cardiopulmonary risk because of the use of larger airway pressures (Paws) to maintain tidal volume (VT). However, the quantitative partitioning of respiratory mechanics and transpulmonary pressure (PL) during robotic surgery is not well described. We tested the following hypothesis: (1) the components of driving pressure (transpulmonary and chest wall components) increase in a parallel fashion at robotic surgical stages (Trendelenburg and robot docking); and (2) deep, when compared to routine (moderate), neuromuscular blockade modifies those changes in PLs as well as in regional respiratory mechanics. We studied 35 American Society of Anesthesiologists (ASA) I-II patients undergoing elective robotic surgery. Airway and esophageal balloon pressures and respiratory flows were measured to calculate respiratory mechanics. Regional lung aeration and ventilation was assessed with electrical impedance tomography and level of neuromuscular blockade with acceleromyography. During robotic surgical stages, 2 crossover randomized groups (conditions) of neuromuscular relaxation were studied: Moderate (1 twitch in the train-of-four stimulation) and Deep (1-2 twitches in the posttetanic count). Pneumoperitoneum was associated with increases in driving pressure, tidal changes in PL, and esophageal pressure (Pes). Steep Trendelenburg position during robot docking was associated with further worsening of the respiratory mechanics. The fraction of driving pressures that partitioned to the lungs decreased from baseline (63% ± 15%) to Trendelenburg position (49% ± 14%, P < .001), due to a larger increase in chest wall elastance (Ecw; 12.7 ± 7.6 cm H2O·L) than in lung elastance (EL; 4.3 ± 5.0 cm H2O·L, P < .001). Consequently, from baseline to Trendelenburg, the component of Paw affecting the chest wall increased by 6.6 ± 3.1 cm H2O, while PLs increased by only 3.4 ± 3.1 cm H2O (P < .001). PL and driving pressures were larger at surgery end than at baseline and were accompanied by dorsal aeration loss. Deep neuromuscular blockade did not change respiratory mechanics, regional aeration and ventilation, and hemodynamics. In robotic surgery with pneumoperitoneum, changes in ventilatory driving pressures during Trendelenburg and robot docking are distributed less to the lungs than to the chest wall as compared to routine mechanical ventilation for supine patients. This effect of robotic surgery derives from substantially larger increases in Ecw than ELs and reduces the risk of excessive PLs. Deep neuromuscular blockade does not meaningfully change global or regional lung mechanics.


The Role of Platelet Homeostasis in a Novel Topical PRP Formulation.
Topical platelet-rich plasma (PRP) must demonstrate stability to insure biologic activity in aesthetic medicine. The objective of this research was to evaluate the role of platelet homeostasis in a novel PRP topical cosmetic formulation to provide facial appearance improvement. The stability of the topical PRP formulation was evaluated in vitro followed by clinical in vivo testing. The in vitro evaluation examined platelet stability and morphology over a 90-day period within the preservative cosmetic base utilizing ELISA and light microscopy (LM)/scanning electron microscopy (SEM). The in vivo clinical study enrolled 20 subjects in a 120-day double blind split face study to evaluate the effect of 5–7x concentrated PRP compared to 2–3x concentrated PRP on facial photoaging. Cosmetic effect was evaluated by the subject and the dermatologist investigator on a 5-point ordinal scale at baseline, week 8, and week 16. 90-day stability for the topical PRP formulation was verified via ELISA and LM/SEM. ELISA showed the PRP was more inactive than control conditions via analyte concentration curves (PDGF-AB, EGF, and P-Selectin). LM/SEM demonstrated the PRP had less aggregation/activation over time within the cosmetic base and that refrigeration is superior to room-temperature storage thus delaying full platelet degranulation. The in vivo clinical study demonstrated parity between 20ml and 60ml PRP in terms of clinical efficacy. Platelets remain viable for up to 90 days in a refrigerated cosmetic vehicle with demonstrated topical clinical PRP facial benefits. PRP kits of 20ml and 60ml volumes for topical PRP are equally efficacious. J Drugs Dermatol. 2020;19(12): doi:10.36849/JDD.2020.5495.


Therapeutic protective effects of IL-12 combined with soluble IL-4 receptor against established infections of herpes simplex virus type 1 in thermally injured mice.
The effect of combination therapy between IL-12 and soluble IL-4R (sIL-4R) on the established infection of HSV-1 in thermally injured mice (TI mice) was investigated. All of the TI mice infected with lethal amounts of HSV-1 died when IL-12 was given therapeutically at a dose of 500 U/mouse. However, 80% of these mice treated prophylactically with IL-12 survived compared with 0% survival of the same mice treated with saline. The therapeutic administration of IL-12 to TI mice currently infected with HSV-1 caused an 80% survival of these mice when the treatment was combined with sIL-4R. Although IL-12 did not stimulate IFN-gamma production in cultures of splenic T cells from TI mice, IFN-gamma was produced by stimulation with IL-12 when the producer cells were prepared from TI mice that had been treated previously with sIL-4R. After stimulation with anti-CD3 mAb, splenic T cells from TI mice with the established infection of HSV-1 produced IL-4 into their culture fluids. However, IL-4 was not produced by splenic T cells that were prepared from the same infected mice treated with IL-12 and sIL-4R in combination. The results obtained herein indicate that the efficacies of the combination therapy against the established infection of HSV-1 may result from the IFN-gamma production stimulated by IL-12 in TI mice that are treated with sIL-4R for reducing burn-associated type 2 T cell responses.


Patient-Reported Outcomes in Cancer Drug Development and US Regulatory Review: Perspectives From Industry, the Food and Drug Administration, and the Patient.
Data reported directly by patients about how they feel and function are rarely included in oncology drug labeling in the United States, in contrast to Europe and to nononcology labeling in the United States, where this practice is more common. Multiple barriers exist, including challenges unique to oncology trials, and industry's concerns regarding cost, logistical complexities, and the Food and Drug Administration's (FDA's) rigorous application of its 2009 guidance on the use of patient-reported outcome (PRO) measures. A panel consisting of representatives of industry, FDA, the PRO Consortium, clinicians, and patients was assembled at a 2014 workshop cosponsored by FDA to identify practical recommendations for overcoming these barriers. Key recommendations included increasing proactive encouragement by FDA to clinical trial sponsors for including PROs in drug development programs; provision of comprehensive PRO plans by sponsors to FDA early in drug development; promotion of an oncology-specific PRO research agenda; development of an approach to existing ("legacy") PRO measures, when appropriate (focused initially on symptoms and functional status); and increased FDA and industry training in PRO methodology. FDA has begun implementing several of these recommendations.


Deletion of ELOVL5 leads to fatty liver through activation of SREBP-1c in mice.
Elongation of very long chain fatty acids (ELOVL)5 is one of seven mammalian fatty acid condensing enzymes involved in microsomal fatty acid elongation. To determine the in vivo substrates and function of ELOVL5, we generated Elovl5(-/-) mice. Studies using liver microsomal protein from wild-type and knockout mice demonstrated that the elongation of gamma-linolenic (C18:3, n-6) to dihomo-gamma-linolenic (C20:3, n-6) and stearidonic (C18:4, n-3) to omega3-arachidonic acid (C20:4, n-3) required ELOVL5 activity. Tissues of Elovl5(-/-) mice accumulated the C18 substrates of ELOVL5 and the levels of the downstream products, arachidonic acid (C20:4, n-6) and docosahexaenoic acid (DHA, C22:6, n-3), were decreased. A consequence of decreased cellular arachidonic acid and DHA concentrations was the activation of sterol regulatory element-binding protein (SREBP)-1c and its target genes involved in fatty acid and triglyceride synthesis, which culminated in the development of hepatic steatosis in Elovl5(-/-) mice. The molecular and metabolic changes in fatty acid metabolism in Elovl5(-/-) mice were reversed by dietary supplementation with arachidonic acid and DHA. These studies demonstrate that reduced ELOVL5 activity leads to hepatic steatosis, and endogenously synthesized PUFAs are key regulators of SREBP-1c activation and fatty acid synthesis in livers of mice.


Screening and early psychological intervention for depression in schools : systematic review and meta-analysis.
Depression in children and adolescents is considerably undertreated, and the school may be a good setting for identifying and treating depression. We conducted a meta-analysis of studies in which students were screened for depression, and those with depressive symptoms were treated with a psychological intervention. Only randomised controlled trials were included. Eight studies met the inclusion criteria. Five studies focused on younger children (7-14 years) and three studies were aimed at adolescents (12-19 years). In total 5803 students were screened, of whom 7.2% were included in the intervention studies (95% CI: 7.1-7.3). The 'numbers-needed-to-screen' was 31 (95% CI: 27-32), which means that 31 students had to be screened in order to generate one successfully treated case of depression. The effects of the psychological treatments at post-test were compared to control conditions in the 8 studies comprising 12 contrast groups, with a total of 413 students. The mean effect size was 0.55 (95% CI: 0.35-0.76). There were not enough studies to examine whether specific psychotherapies were superior to other psychotherapies. Although the number of studies is small and their quality is limited, screening and early intervention at schools may be an effective strategy to reduce the burden of disease from depression in children and adolescents. More research on the (negative) effects of these interventions is needed.


L-asparaginase-induced abnormality in plasma glucose level in patients of acute lymphoblastic leukemia admitted to a tertiary care hospital of Odisha.
The objective of this study was to evaluate any abnormal change in plasma glucose levels in patients treated with L-asparaginase (L-Asp)-based chemotherapy regimen in patients of acute lymphoblastic leukemia (ALL). This retrospective, hospital-based study was conducted in patients of ALL, admitted to the Clinical Haematology Department of a tertiary care hospital of Odisha from August 2014 to July 2015. Indoor records of 146 patients on multi-centered protocol-841 were evaluated for any alteration in plasma glucose level, time of onset of hypo/hyperglycemia, and persistence of plasma glucose alteration. Twenty-one percent of patients showed abnormal plasma glucose level. Most of these patients developed hypoglycemia and were of lower age group. Most of these patients developed hypoglycemia and were of lower age group, whereas a majority of higher age group patients developed hyperglycemia. In majority of the cases, abnormal glucose developed after three doses of L-Asp. Hypoglycemia subsided whereas hyperglycemia persisted till the end of our observation period. L-Asp produces more incidences of hypoglycemia than hyperglycemia in a good number of ALL patients towards which clinicians should be more vigilant. However, hyperglycemia persists for a longer duration than hypoglycemia.


Conditional transgenic models define how MYC initiates and maintains tumorigenesis.
MYC is one of the most commonly overexpressed oncogenes in human cancer. The targeted inactivation of MYC is a possible therapy for neoplasia. Conditional transgenic mouse model systems are tractable methods to precisely dissect how and when the inactivation of MYC might be effective in the treatment for human cancer. From these model systems, several general principles emerge. MYC inactivation stereotypically results in the proliferative arrest, differentiation and/or apoptosis of tumor cells. The specific consequences of MYC inactivation appear to depend both on the type of cancer as well as the constellation of genetic events unique to a given tumor. Tumors can escape from dependence upon MYC by acquiring compensatory genetic events. MYC inactivation can uncover the stem cell properties of tumor cells that differentiate into normal appearing cells. In some cases, these differentiated cells are actually dormant tumor cells that recover their neoplastic properties upon MYC reactivation. In other cases, even brief MYC inactivation is sufficient to induce sustained tumor regression. Insights from conditional transgenic mouse models will be useful in the development of therapies that target MYC for the treatment of cancer.


6-Thioguanine inhibits rotavirus replication through suppression of Rac1 GDP/GTP cycling.
Rotavirus infection has emerged as an important cause of complications in organ transplantation recipients and might play a role in the pathogenesis of inflammatory bowel disease (IBD). 6-Thioguanine (6-TG) has been widely used as an immunosuppressive drug for organ recipients and treatment of IBD in the clinic. This study aims to investigate the effects and mode-of-action of 6-TG on rotavirus replication. Human intestinal Caco2 cell line, 3D model of human primary intestinal organoids, laboratory rotavirus strain (SA11) and patient-derived rotavirus isolates were used. We have demonstrated that 6-TG significantly inhibits rotavirus replication in these intestinal epithelium models. Importantly, gene knockdown or knockout of Rac1, the cellular target of 6-TG, significantly inhibited rotavirus replication, indicating the supportive role of Rac1 for rotavirus infection. We have further demonstrated that 6-TG can effectively inhibit the active form of Rac1 (GTP-Rac1), which essentially mediates the anti-rotavirus effect of 6-TG. Consistently, ectopic over-expression of GTP-Rac1 facilitates but an inactive Rac1 (N17) or a specific Rac1 inhibitor (NSC23766) inhibits rotavirus replication. In conclusion, we have identified 6-TG as an effective inhibitor of rotavirus replication via the inhibition of Rac1 activation. Thus, for transplantation patients or IBD patients infected with rotavirus or at risk of rotavirus infection, the choice of 6-TG as a treatment appears rational.