Early vitrectomy for progressive diabetic proliferations covering the macula.
The clinical course in 50 eyes was analysed after pars plana vitrectomy for progressive diabetic fibrovascular proliferations. Patients were assigned to pars plana vitrectomy if progression of proliferations occurred despite a photocoagulation treatment with a mean number of 3500 burns and additional peripheral cryoablation. All cases had visual impairment because of fibrovascular tissue covering the macula without detachment of the macula. Flat proliferations were present in all eyes without retinal elevation, vitreous detachment, or vitreous haemorrhage. The follow up intervals ranged from 13 months to 39 months (mean interval 24 months). Twelve months postoperatively, 36 eyes (72%) showed improved visual acuity, five eyes (10%) were worse, and nine eyes (18%) were unchanged. Thirty two eyes (64%) achieved a final visual acuity of 0.2 or better, and 45 eyes (90%) gained 0.05 or better. In only two eyes could reproliferation be observed. The postoperative course indicates that pars plana vitrectomy for diabetic fibrovascular proliferations covering the macula can preserve socially useful visual acuity of at least 0.05 in most cases.


Exposure to the gut microbiota drives distinct methylome and transcriptome changes in intestinal epithelial cells during postnatal development.
The interplay of epigenetic processes and the intestinal microbiota may play an important role in intestinal development and homeostasis. Previous studies have established that the microbiota regulates a large proportion of the intestinal epithelial transcriptome in the adult host, but microbial effects on DNA methylation and gene expression during early postnatal development are still poorly understood. Here, we sought to investigate the microbial effects on DNA methylation and the transcriptome of intestinal epithelial cells (IECs) during postnatal development. We collected IECs from the small intestine of each of five 1-, 4- and 12 to 16-week-old mice representing the infant, juvenile, and adult states, raised either in the presence or absence of a microbiota. The DNA methylation profile was determined using reduced representation bisulfite sequencing (RRBS) and the epithelial transcriptome by RNA sequencing using paired samples from each individual mouse to analyze the link between microbiota, gene expression, and DNA methylation. We found that microbiota-dependent and -independent processes act together to shape the postnatal development of the transcriptome and DNA methylation signatures of IECs. The bacterial effect on the transcriptome increased over time, whereas most microbiota-dependent DNA methylation differences were detected already early after birth. Microbiota-responsive transcripts could be attributed to stage-specific cellular programs during postnatal development and regulated gene sets involved primarily immune pathways and metabolic processes. Integrated analysis of the methylome and transcriptome data identified 126 genomic loci at which coupled differential DNA methylation and RNA transcription were associated with the presence of intestinal microbiota. We validated a subset of differentially expressed and methylated genes in an independent mouse cohort, indicating the existence of microbiota-dependent "functional" methylation sites which may impact on long-term gene expression signatures in IECs. Our study represents the first genome-wide analysis of microbiota-mediated effects on maturation of DNA methylation signatures and the transcriptional program of IECs after birth. It indicates that the gut microbiota dynamically modulates large portions of the epithelial transcriptome during postnatal development, but targets only a subset of microbially responsive genes through their DNA methylation status.


Precommitting to serve the underserved.
In many countries worldwide, especially in sub-Saharan Africa, a shortage of physicians limits the provision of lifesaving interventions. One existing strategy to increase the number of physicians in areas of critical shortage is conditioning medical school scholarships on a precommitment to work in medically underserved areas later. Current practice is usually to demand only one year of service for each year of funded studies. We show the effectiveness of scholarships conditional on such precommitment for increasing physician supplies in underserved areas. Then we defend these scholarships against ethical worries that they constitute slavery contracts; rely on involuntary, biased, or unauthorized early consent by a young signatory; put excessive strains on signed commitments; give rise to domination; and raise suspicion of slavery contracts. Importantly, we find that scholarships involving far longer commitment than current practice allows would also withstand these worries. Policymakers should consider introducing conditional scholarships, including long-term versions, as a means to increasing the supply of physicians to medically underserved areas.


Proteome analysis of a Lactococcus lactis strain overexpressing gapA suggests that the gene product is an auxiliary glyceraldehyde 3-phosphate dehydrogenase.
The sequence of the genome from the Lactococcus lactis subspecies lactis strain IL1403 shows the presence of two reading frames, gapA and gapB, putatively encoding glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Previous proteomic analysis of the L. lactis subspecies cremoris strain MG1363 has revealed two neighbouring protein spots, GapBI and GapBII, with amino terminal sequences identical to the product of gapA from the L. lactis subspecies cremoris strain LM0230 and that of the two IL1403 sequences. In order to assign the two protein spots to their respective genes we constructed an L. lactis strain that overexpessed the gapA gene derived from MG1363 upon nisin induction. Compared to the wild-type, the overexpressing strain had a 3.4-fold elevated level of specific GAPDH activity when grown in the presence of nisin. In both MG1363 and the gapA overexpressing strain the GAPDH activity was specific for NAD. No NADP dependent activity was detected. Proteome analysis of the gapA overexpressing strain revealed two new protein spots, GapAI and GapAII, not previously detected in proteome analysis of MG1363. Results from mass spectrometry analysis of GapA and GapB and comparison with the deduced protein sequences for the GAPDH isozymes from the genome sequence of strain IL1403 allowed us to assign GapA and GapB to their apparent IL1403 homologues encoded by gapA and gapB, respectively. Furthermore, we suggest that a homologue of a gapB product, represented by GapB, is the main source of GAPDH activity in L. lactis during normal growth.


Plasma proteomic profiles from disease-discordant monozygotic twins suggest that molecular pathways are shared in multiple systemic autoimmune diseases.
Although systemic autoimmune diseases (SAID) share many clinical and laboratory features, whether they also share some common features of pathogenesis remains unclear. We assessed plasma proteomic profiles among different SAID for evidence of common molecular pathways that could provide insights into pathogenic mechanisms shared by these diseases. Differential quantitative proteomic analyses (one-dimensional reverse-phase liquid chromatography-mass spectrometry) were performed to assess patterns of plasma protein expression. Monozygotic twins (four pairs discordant for systemic lupus erythematosus, four pairs discordant for juvenile idiopathic arthritis and two pairs discordant for juvenile dermatomyositis) were studied to minimize polymorphic gene effects. Comparisons were also made to 10 unrelated, matched controls. Multiple plasma proteins, including acute phase reactants, structural proteins, immune response proteins, coagulation and transcriptional factors, were differentially expressed similarly among the different SAID studied. Multivariate Random Forest modeling identified seven proteins whose combined altered expression levels effectively segregated affected vs. unaffected twins. Among these seven proteins, four were also identified in univariate analyses of proteomic data (syntaxin 17, α-glucosidase, paraoxonase 1, and the sixth component of complement). Molecular pathway modeling indicated that these factors may be integrated through interactions with a candidate plasma biomarker, PON1 and the pro-inflammatory cytokine IL-6. Together, these data suggest that different SAID may share common alterations of plasma protein expression and molecular pathways. An understanding of the mechanisms leading to the altered plasma proteomes common among these SAID may provide useful insights into their pathogeneses.


A review of epidemiological parameters from Ebola outbreaks to inform early public health decision-making.
The unprecedented scale of the Ebola outbreak in West Africa has, as of 29 April 2015, resulted in more than 10,884 deaths among 26,277 cases. Prior to the ongoing outbreak, Ebola virus disease (EVD) caused relatively small outbreaks (maximum outbreak size 425 in Gulu, Uganda) in isolated populations in central Africa. Here, we have compiled a comprehensive database of estimates of epidemiological parameters based on data from past outbreaks, including the incubation period distribution, case fatality rate, basic reproduction number (R 0 ), effective reproduction number (R t ) and delay distributions. We have compared these to parameter estimates from the ongoing outbreak in West Africa. The ongoing outbreak, because of its size, provides a unique opportunity to better understand transmission patterns of EVD. We have not performed a meta-analysis of the data, but rather summarize the estimates by virus from comprehensive investigations of EVD and Marburg outbreaks over the past 40 years. These estimates can be used to parameterize transmission models to improve understanding of initial spread of EVD outbreaks and to inform surveillance and control guidelines.


Outpatient Quick Diagnosis Units for the evaluation of suspected severe diseases: an observational, descriptive study.
Hospitals in countries with public health systems have recently adopted organizational changes to improve efficiency and resource allocation, and reducing inappropriate hospitalizations has been established as an important goal. Our goal was to describe the functioning of a Quick Diagnosis Unit in a Spanish public university hospital after evaluating 1,000 consecutive patients. We also aimed to ascertain the degree of satisfaction among Quick Diagnosis Unit patients and the costs of the model compared to conventional hospitalization practices. Observational, descriptive study. Our sample comprised 1,000 patients evaluated between November 2008 and January 2010 in the Quick Diagnosis Unit of a tertiary university public hospital in Barcelona. Included patients were those who had potentially severe diseases and would normally require hospital admission for diagnosis but whose general condition allowed outpatient treatment. We analyzed several variables, including time to diagnosis, final diagnoses and hospitalizations avoided, and we also investigated the mean cost (as compared to conventional hospitalization) and the patients' satisfaction. In 88% of cases, the reasons for consultation were anemia, anorexia-cachexia syndrome, febrile syndrome, adenopathies, abdominal pain, chronic diarrhea and lung abnormalities. The most frequent diagnoses were cancer (18.8%; mainly colon cancer and lymphoma) and Iron-deficiency anemia (18%). The mean time to diagnosis was 9.2 days (range 1 to 19 days). An estimated 12.5 admissions/day in a one-year period (in the internal medicine department) were avoided. In a subgroup analysis, the mean cost per process (admission-discharge) for a conventional hospitalization was 3,416.13 Euros, while it was 735.65 Euros in the Quick Diagnosis Unit. Patients expressed a high degree of satisfaction with Quick Diagnosis Unit care. Quick Diagnosis Units represent a useful and cost-saving model for the diagnostic study of patients with potentially severe diseases. Future randomized study designs involving comparisons between controls and intervention groups would help elucidate the usefulness of Quick Diagnosis Units as an alternative to conventional hospitalization.


Breastfeeding and verbal ability of 3-year-olds in a multicity sample.
Our goal was to analyze the effect of maternal verbal ability and education on the association between breastfeeding and children's cognitive functioning. First, we hypothesized that maternal verbal abilities account for a large portion of the association between breastfeeding and child verbal abilities. Second, we hypothesized that after adjusting for maternal verbal abilities, a positive effect of breastfeeding will be most evident among highly educated mothers, because these mothers may have more opportunity to engage in cognitively stimulating parenting than do mothers with less education. With data on 1645 American-born mothers participating in a longitudinal birth cohort study, we used linear regression to determine the influence of breastfeeding for at least 1 month on the Peabody Picture Vocabulary Test-Third Edition scores of 3-year-old children. Models were adjusted for an extensive set of demographic characteristics, including mother's Peabody Picture Vocabulary Test and the Home Observation for Measurement of the Environment score. Mothers were categorized into 1 of 3 educational-status groups: no high school diploma, high school diploma, and some post-secondary education. In unadjusted mean comparisons, breastfed children had Peabody Picture Vocabulary Test scores that were 6.6 points higher than children who were not breastfed. After adjusting for demographic characteristics and maternal verbal ability, the coefficient dropped to 1.72. Among mothers with education beyond high school, the children's Peabody Picture Vocabulary Test scores in adjusted models were 2.2 points higher for breastfed children. Among mothers with a high school diploma or less, there were no significant differences in the children's Peabody Picture Vocabulary Test scores by breastfeeding status. These results were consistent in white, black, and Hispanic children. Maternal Peabody Picture Vocabulary Test scores mediate much of the association between breastfeeding and child verbal abilities. The beneficial effects of breastfeeding on children's cognition may emerge only when breastfeeding is done in conjunction with other positive parenting behaviors. The advantageous effects of breastfeeding do not seem to be solely attributable to the superior nutrient content of breast milk.


The collectivity of drinking cultures: is the theory applicable to African settings?
Skog's theory of collective drinking behaviour implies that countries with a strict informal social control of drinking alcohol would not exhibit 'collective displacement' of consumption (a linear association between population mean consumption and percentile values across the full range of the distribution), as do countries with less informal social control. This paper aimed to test this hypothesis by examining the alcohol consumption distributions in African countries with a strong informal control of alcohol. Data on alcohol consumption from the World Health Organization's general population surveys in 15 African countries were aggregated and analysed with respect to skewedness and collective displacement of the distribution. The distribution of consumption was strongly positively skewed, with 10-15% of the drinkers consuming more than twice the mean consumption. There was also clear evidence of a collective displacement of the consumption distribution, and the consumption mean was a strong predictor of the distribution percentile values across the full range of the distribution. Correspondingly, consumption mean predicted the prevalence of heavy drinkers. The distribution patterns of alcohol consumption in African countries are consistent with those observed previously in industrialized countries. These findings seem to counter Skog's theory of collective drinking behaviour and support the universality of the observation that the prevalence of problem drinking is linked closely to mean consumption.


Long-term safety and efficacy of ziv-aflibercept in retinal diseases.
To investigate the long-term safety of intravitreal ziv-aflibercept in eyes receiving six or more intravitreal injections of ziv-aflibercept, an off-label substitute to the approved aflibercept. Consecutive patients with retinal disease receiving six or more of intravitreal 0.05 mL ziv-aflibercept (1.25 mg) injections were followed monthly in three centres. Outcome measures were best-corrected visual acuity (BCVA) (logarithm of the minimum angle of resolution (logMar)) and central macular thickness (CMT) on spectral domain optical coherence tomography and monitoring for ocular inflammation, progression of lens opacities and intraocular pressure rise. Paired comparison was done using Wilcoxon signed-rank test calculator. Sixty-five eyes of 60 consecutive patients received a mean of 8.4 (6-17) intravitreal injections with a baseline mean logMAR BCVA of 0.98±0.56 and CMT 432.7±163.0 μm and followed for a mean of 9.2 months (range 6-18 months). After the sixth injection, mean BCVA improved to 0.57±0.36 (p=0.001) and CMT decreased to 274.8±117.8 μm (p=0.0001). At the 9-month follow-up, mean BCVA improved to 0.62±0.37 (p=0.0004) and mean CMT decreased to 292.0±160.9 μm (p<0.01) in 19 eyes. At 1 year, mean BCVA was 0.73±0.52 and CMT 311.6±232.5 μm in seven eyes. Intraocular pressures did not increase after injections. One subject developed transient mild iritis at the fourth injection but not on subsequent injections. No lens opacity progression or endophthalmitis was noted. Systemic adverse effects were not registered. Repeated intravitreal injections of ziv-aflibercept appear tolerable, safe and efficacious in the therapy of retinal disease.


Effects of an Acute Exercise Bout on Serum Hepcidin Levels.
Iron deficiency is a frequent and multifactorial disorder in the career of athletes, particularly in females. Exercise-induced disturbances in iron homeostasis produce deleterious effects on performance and adaptation to training; thus, the identification of strategies that restore or maintain iron homeostasis in athletes is required. Hepcidin is a liver-derived hormone that degrades the ferroportin transport channel, thus reducing the ability of macrophages to recycle damaged iron, and decreasing iron availability. Although it has been suggested that the circulating fraction of hepcidin increases during early post-exercise recovery (~3 h), it remains unknown how an acute exercise bout may modify the circulating expression of hepcidin. Therefore, the current review aims to determine the post-exercise expression of serum hepcidin in response to a single session of exercise. The review was carried out in the Dialnet, Elsevier, Medline, Pubmed, Scielo and SPORTDiscus databases, using hepcidin (and "exercise" or "sport" or "physical activity") as a strategy of search. A total of 19 articles were included in the review after the application of the inclusion/exclusion criteria. This search found that a single session of endurance exercise (intervallic or continuous) at moderate or vigorous intensity (60-90% VO2peak) stimulates an increase in the circulating levels of hepcidin between 0 h and 6 h after the end of the exercise bout, peaking at ~3 h post-exercise. The magnitude of the response of hepcidin to exercise seems to be dependent on the pre-exercise status of iron (ferritin) and inflammation (IL-6). Moreover, oxygen disturbances and the activation of a hypoxia-induced factor during or after exercise may stimulate a reduction of hepcidin expression. Meanwhile, cranberry flavonoids supplementation promotes an anti-oxidant effect that may facilitate the post-exercise expression of hepcidin. Further studies are required to explore the effect of resistance exercise on hepcidin expression.


MINT: software to identify motifs and short-range interactions in trajectories of nucleic acids.
Structural biology experiments and structure prediction tools have provided many high-resolution three-dimensional structures of nucleic acids. Also, molecular dynamics force field parameters have been adapted to simulating charged and flexible nucleic acid structures on microsecond time scales. Therefore, we can generate the dynamics of DNA or RNA molecules, but we still lack adequate tools for the analysis of the resulting huge amounts of data. We present MINT (Motif Identifier for Nucleic acids Trajectory) - an automatic tool for analyzing three-dimensional structures of RNA and DNA, and their full-atom molecular dynamics trajectories or other conformation sets (e.g. X-ray or nuclear magnetic resonance-derived structures). For each RNA or DNA conformation MINT determines the hydrogen bonding network resolving the base pairing patterns, identifies secondary structure motifs (helices, junctions, loops, etc.) and pseudoknots. MINT also estimates the energy of stacking and phosphate anion-base interactions. For many conformations, as in a molecular dynamics trajectory, MINT provides averages of the above structural and energetic features and their evolution. We show MINT functionality based on all-atom explicit solvent molecular dynamics trajectory of the 30S ribosomal subunit.


Somatic estrogen receptor α mutations that induce dimerization promote receptor activity and breast cancer proliferation.
Physiologic activation of the estrogen receptor alpha (ERα) is mediated by estradiol (E2) binding in the ligand binding pocket of the receptor, repositioning helix 12 (H12) to facilitate binding of coactivator proteins in the unoccupied coactivator binding groove. In breast cancer, activation of ERα is often observed through point mutations that lead to the same H12 repositioning in the absence of E2. Through expanded genetic sequencing of breast cancer patients, we identified a collection of mutations located far from H12 but nonetheless capable of promoting E2-independent transcription and breast cancer cell growth. Using machine learning and computational structure analyses, this set of mutants was inferred to act distinctly from the H12-repositioning mutants and instead was associated with conformational changes across the ERα dimer interface. Through both in vitro and in cell assays of full length ERα protein and isolated ligand binding domain (LBD), we found that these mutants promoted ERα dimerization, stability, and nuclear localization. Point mutations that selectively disrupted dimerization abrogated E2-independent transcriptional activity of these dimer-promoting mutants. The results revealed a distinct mechanism for activation of ERα function through enforced receptor dimerization and suggested dimer disruption as a potential therapeutic strategy to treat ER-dependent cancers.


A review of differences between basal ganglia and cerebellar control of movements as revealed by functional imaging studies.
The role of the basal ganglia and cerebellum in the control of movements is unclear. We summarize results from three groups of PET studies of regional CBF. The results show a double dissociation between (i) selection of movements, which induces differential effects in the basal ganglia but not the cerebellum, and (ii) sensory information processing, which involves the cerebellum but not the basal ganglia. The first set of studies concerned motor learning of a sequence of finger movements; there was a shift of activation in the anterior-posterior direction of the basal ganglia which paralleled changes in the motor areas of the frontal cortex. During new learning, the dorsolateral prefrontal cortex and striatum (caudate nucleus and anterior putamen) were activated. When subjects had to select movements, the premotor cortex and mid-putamen were activated. With automatic (overlearned) movements, the sensorimotor cortex and posterior putamen were activated. When subjects paid attention to overlearned actions, activation shifted back to the dorsolateral prefrontal cortex and striatum. The cerebellum was not activated when subjects made new decisions, attended to their actions or selected movements. These results demonstrate components of basal ganglia-(thalamo)-cortical loops in humans. According to earlier studies in animals we propose that the basal ganglia may be concerned with selecting movements or the selection of appropriate muscles to perform a movement selected by cortical areas (e.g. premotor cortex). Secondly, a visuomotor co-ordination task was examined. In the absence of visual control over arm movements, subjects were required to use a computer mouse to either generate new lines or to re-trace lines on a computer screen. The neocerebellum (hemispheres of the posterior lobe, cerebellar nuclei and cerebellar vermis), not the basal ganglia, was more engaged when lines were re-traced (compared with new line generation). Animal experiments have shown that error detection (deviation from given lines) and correction occurs during line re-tracing but not line generation. Our data suggest that the neocerebellum (not the basal ganglia) is involved in monitoring and optimizing movements using sensory (proprioceptive) feedback. Thirdly, the relative contribution of sensory information processing to the signal during active/passive execution of a motor task (flexion and extension of the elbow) was examined; it was found that 80-90% of the neocerebellar signal could be attributed to sensory information processing. The basal ganglia were not involved in sensory information processing. They may be concerned with movement/ muscle selection (efferent motor component); the neocerebellum may be concerned with monitoring the outcome (afferent sensory component) and optimizing movements using sensory (feedback) information.


Extracellular Vesicle MicroRNA in Malignant Pleural Effusion.
Lung and breast cancer are the two most common causes of malignant pleural effusion (MPE). MPE diagnosis plays a crucial role in determining staging and therapeutic interventions in these cancers. However, our understanding of the pathogenesis and progression of MPE at the molecular level is limited. Extracellular Vesicles (EVs) and their contents, including microRNAs (miRNAs), can be isolated from all bodily fluids, including pleural fluid. This study aims to compare EV-miRNA patterns of expression in MPE caused by breast (BA-MPE) and lung (LA-MPE) adenocarcinomas compared to the control group of heart-failure-induced effusions (HF-PE). We conducted an analysis of 24 pleural fluid samples (8 LA-MPE, 8 BA-MPE, and 8 HF-PE). Using NanoString technology, we profiled miRNAs within EVs isolated from 12 cases. Bioinformatic analysis demonstrated differential expression of miR-1246 in the MPE group vs. HF-PE group and miR-150-5p and miR-1246 in the BA-MPE vs. LA-MPE group, respectively. This difference was demonstrated and validated in an independent cohort using real-time PCR (RT-PCR). miRNA-1246 demonstrated 4-fold increased expression (OR: 3.87, 95% CI: 0.43, 35) in the MPE vs. HF-PE group, resulting in an area under the curve of 0.80 (95% CI: 0.60, 0.99). The highest accuracy for differentiating MPE vs. HF-PE was seen with a combination of miRNAs compared to each miRNA alone. Consistent with prior studies, this study demonstrates dysregulation of specific EV-based miRNAs in breast and lung cancer; pleural fluid provides direct access for the analysis of these EV-miRNAs as biomarkers and potential targets and may provide insight into the underlying pathogenesis of tumor progression. These findings should be explored in large prospective studies.


Effect of Exogenously Supplied Foliar Potassium on Phloem Loading in Beta vulgaris L.
The effect of foliar application of K(+) on processes associated with phloem loading was investigated in source leaves of sugar beet (Beta vulgaris L.). KCI was supplied exogenously at concentrations of up to 100 millimolar in the solution bathing the abraded upper epidermis of source leaves. K(+) added at concentrations below 30 millimolar generally promoted the rate of export of material derived from (14)CO(2) but not from exogenously applied [(14)C]sucrose. Paralleling promotion of export, the level of material derived from photosynthesis, which was released into the bathing solution, also increased in response to addition of K(+) to the free space. Net photosynthetic rate was not affected. K(+) at 5 and 15 millimolar concentrations did not stimulate uptake of [(14)C]sucrose into source leaf discs.The data suggest that the promotion of export rate by K(+) results primarily from an effect on the site regulating efflux of sucrose into the apoplast prior to loading into the minor veins, rather than on the loading site itself. A change in the level of sucrose in the free space appears to precede a change in export rate.


Effect of excess iodine intake on thyroid diseases in different populations: A systematic review and meta-analyses including observational studies.
Although several reports concerning the association of iodine excess and thyroid disease have appeared, no systematic review of the association between iodine excess intake and thyroid diseases, especially hyperthyroidism and hypothyroidism, has yet been reported. We conducted a systematic search of Ovid MEDLINE, PubMed, Cochrane Central Register of Controlled Trials databases, Ichushi-Web and CiNii database for intervention trials and observational studies. Search terms were constructed from related words for excess AND iodine intake or excretion AND thyroid hormones or diseases AND study designs. After considering the qualitative heterogeneity among studies, a meta-analysis was conducted and odds ratios and 95% confidence intervals (CI) were estimated in random-effects models. A protocol was registered with PROSPERO (No. CRD42015028081). 50 articles were included, including three intervention trials, six case-control studies, six follow-up studies and 35 cross-sectional studies. Three cross-sectional studies in adults included in meta-analysis. Odds ratio of overt and subclinical hypothyroidism between excess and adequate populations were 2.78 (CI:1.47 to 5.27) and 2.03 (CI:1.58 to 2.62) in adults, respectively. Source of excess iodine status was mainly iodized salt or water in included studies. Although universal salt iodization has improved goiter rates, chronic exposure to excess iodine from water or poorly monitored salt are risk factors for hypothyroidism in free-living populations. Monitoring of both iodine concentration in salt as well as the iodine concentration in local drinking water are essential to preventing thyroid diseases. Hypothyroidism should be also carefully monitored in areas with excess iodine. Because of the low quality and limited number of included studies, further evidence and review are required.


Bacterial diversity in a finished compost and vermicompost: differences revealed by cultivation-independent analyses of PCR-amplified 16S rRNA genes.
Bacterial communities are important catalysts in the production of composts. Here, it was analysed whether the diversity of bacteria in finished composts is stable and specific for the production process. Single-strand conformation polymorphism (SSCP) based on polymerase chain reaction amplified partial 16S rRNA genes was used to profile and analyse bacterial communities found in total DNA extracted from finished composts. Different batches of compost samples stored over a period of 12 years and a 1-year-old vermicompost were compared to each other. According to digital image analysis, clear differences could be detected between the profiles from compost and vermicompost. Differences between three different periods of compost storage and between replicate vermicompost windrows were only minor. A total of 41 different 16S rRNA genes were identified from the SSCP profiles by DNA sequencing, with the vast majority related to yet-uncultivated bacteria. Sequences retrieved from compost mainly belonged to the phyla Actinobacteria and Firmicutes. In contrast, vermicompost was dominated by bacteria related to uncultured Chloroflexi, Acidobacteria, Bacteroidetes and Gemmatimonadetes. The differences were underscored with specific gene probes and Southern blot hybridizations. The results confirmed that different substrates and composting processes selected for specific bacterial communities in the finished products. The specificity and consistency of the bacterial communities inhabiting the compost materials suggest that cultivation-independent bacterial community analysis is a potentially useful indicator to characterize the quality of finished composts in regard to production processes and effects of storage conditions.


Correlation of loss of heterozygosity with cytogenetic analysis using G-banding and fluorescence in situ hybridization in aneuploid cultures from two human testicular germ-cell tumors.
A detailed karyotype analysis using fluorescence in situ hybridization (FISH), with 24 chromosome-specific paint probes has been carried out on newly established cell lines from two testicular tumors, an i(12p)-positive teratoma, and an i(12p)-negative combined seminoma/teratoma. This has been correlated with loss of heterozygosity (LOH) and allelic imbalance, using DNA RFLP analysis to clarify the genetic changes and to identify any common regions of deletion or rearrangement. With G-banding alone, a total of 11 breakpoints were recognized. After FISH, the position of seven required revision, and 21 new ones were identified. The chromosomes involved most frequently in both tumors were numbers 1, 12, and 18. Breakpoints in 11q and 16q were also seen in both, and seven or more copies of 12p per cell were found in all clones. LOH was found for 18q in both tumors, and overall was much more frequent in underrepresented regions (one or two copies). On the whole, there was good agreement between the cytogenetic and DNA RFLP data; loci showing allelic imbalance generally had an odd number of copies of the chromosome region in which they were known to be located. Combined data on the chromosome 1 translocations in both tumors suggested that rearrangements were more complicated than cytogenetics alone had predicted.


Effects of cell number on teratoma formation by human embryonic stem cells.
Teratoma formation is a critical obstacle to safe clinical translation of human embryonic stem (ES) cell-based therapies in the future. As current methods of isolation are unable to yield 100% pure population of differentiated cells from a pluripotent donor source, potential development of these tumors is a significant concern. Here we used non-invasive reporter gene imaging to investigate the relationship between human ES cell number and teratoma formation in a xenogenic model of ES cell transplantation. Human ES cells (H9 line) were stably transduced with a double fusion (DF) reporter construct containing firefly luciferase and enhanced green fluorescent protein (Fluc-eGFP) driven by a human ubiquitin promoter. Immunodeficient mice received intramyocardial (n = 35) or skeletal muscle (n = 35) injection of 1 x 10(2), 1 x 10(3), 1 x 10(4), 1 x 10(5) or 1 x 10(6) DF positive ES cells suspended in saline for myocardium and Matrigel for skeletal muscle. Cell survival and proliferation were monitored via bioluminescence imaging (BLI) for an 8 week period following transplantation. Mice negative for Fluc signal after 8 weeks were followed out to day 365 to confirm tumor absence. Significantly, in this study, a minimum of 1 x 10(5) ES cells in the myocardium and 1 x 10(4) cells in the skeletal muscle was observed to be requisite for teratoma development, suggesting that human ES cell number may be a critical factor in teratoma formation. Engraftment and tumor occurrence were also observed to be highly dependent on ES cell number. We anticipate these results should yield useful insights to the safe and reliable application of human ES cell derivatives in the clinic.


Bacterial colonization and extinction on marine aggregates: stochastic model of species presence and abundance.
Organic aggregates provide a favorable habitat for aquatic microbes, are efficiently filtered by shellfish, and may play a major role in the dynamics of aquatic pathogens. Quantifying this role requires understanding how pathogen abundance in the water and aggregate size interact to determine the presence and abundance of pathogen cells on individual aggregates. We build upon current understanding of the dynamics of bacteria and bacterial grazers on aggregates to develop a model for the dynamics of a bacterial pathogen species. The model accounts for the importance of stochasticity and the balance between colonization and extinction. Simulation results suggest that while colonization increases linearly with background density and aggregate size, extinction rates are expected to be nonlinear on small aggregates in a low background density of the pathogen. Under these conditions, we predict lower probabilities of pathogen presence and reduced abundance on aggregates compared with predictions based solely on colonization. These results suggest that the importance of aggregates to the dynamics of aquatic bacterial pathogens may be dependent on the interaction between aggregate size and background pathogen density, and that these interactions are strongly influenced by ecological interactions and pathogen traits. The model provides testable predictions and can be a useful tool for exploring how species-specific differences in pathogen traits may alter the effect of aggregates on disease transmission.


A new system of computer-assisted navigation leading to reduction in operating time in uncemented total hip replacement in a matched population.
Computer-assisted navigation techniques are used to optimise component placement and alignment in total hip replacement. It has developed in the last 10 years but despite its advantages only 0.3% of all total hip replacements in England and Wales are done using computer navigation. One of the reasons for this is that computer-assisted technology increases operative time. A new method of pelvic registration has been developed without the need to register the anterior pelvic plane (BrainLab hip 6.0) which has shown to improve the accuracy of THR. The purpose of this study was to find out if the new method reduces the operating time. This was a retrospective analysis of comparing operating time in computer navigated primary uncemented total hip replacement using two methods of registration. Group 1 included 128 cases that were performed using BrainLab versions 2.1-5.1. This version relied on the acquisition of the anterior pelvic plane for registration. Group 2 included 128 cases that were performed using the newest navigation software, BrainLab hip 6.0 (registration possible with the patient in the lateral decubitus position). The operating time was 65.79 (40-98) minutes using the old method of registration and was 50.87 (33-74) minutes using the new method of registration. This difference was statistically significant. The body mass index (BMI) was comparable in both groups. The study supports the use of new method of registration in improving the operating time in computer navigated primary uncemented total hip replacements.


[A study of MTT-hybrid assay using human bone and soft tissue tumor cells].
We investigated a new chemosensitivity test, MTT-hybrid assay, which was a hybrid of MTT colorimetric assay and double-layered soft agar colony assay, using human bone and soft tissue tumor cells. MTT formazan crystals produced by viable cells in the soft agar medium were solubilized by SDS at 60 degrees C. The absorbance (560 nm) is directly proportional to the cell number over a wide range. The absorbance increased in proportion to colonial growth of osteosarcoma cells, while it decreased in a human diploid cell strain in a few days. Drug sensitivity of tumor cells is supposed to be assessed without contaminating normal cells by MTT-hybrid assay in primary tumor samples. Good correlation of IC50 was observed between MTT-hybrid assay and colony assay. The MTT-hybrid assay shows potential value as a rapid predictive test for chemotherapeutic agents in an individual patient.


Welcoming talent? A comparative study of immigrant entrepreneurs' entry policies in France, Germany and the Netherlands.
This article explores the admission policies for self-employed non-EU immigrants wanting to start or move their business to the European Union (EU). Selecting immigrant entrepreneurs is a specific and understudied policy strand in the battle for talent. No common EU policy is available (yet) and although national policies do show some similarity, they differ in respect of how and who decides if an entrepreneur serves a national economic interest. By presenting a first-time model for defining the level of welcoming, this study adds an instrument to the toolbox of both scholars and policy makers for evaluating immigration policies. Whether a policy is welcoming depends on material criteria, such as entry conditions giving the entrepreneur a fair chance and on the formal criteria of the applicable procedures and the actors involved in the decision-making process. The body of the article constitutes of a legal comparison between French, German and Dutch entry policies for non-EU entrepreneurs. The article concludes that a future EU policy on welcoming immigrant entrepreneurs must set standards for a large variety of entrepreneurs, allow for the economic interest to be broadly defined and have, at the least, transparent and practical procedures.


The structure of the Helicobacter pylori ferric uptake regulator Fur reveals three functional metal binding sites.
Fur, the ferric uptake regulator, is a transcription factor that controls iron metabolism in bacteria. Binding of ferrous iron to Fur triggers a conformational change that activates the protein for binding to specific DNA sequences named Fur boxes. In Helicobacter pylori, HpFur is involved in acid response and is important for gastric colonization in model animals. Here we present the crystal structure of a functionally active HpFur mutant (HpFur2M; C78S-C150S) bound to zinc. Although its fold is similar to that of other Fur and Fur-like proteins, the crystal structure of HpFur reveals a unique structured N-terminal extension and an unusual C-terminal helix. The structure also shows three metal binding sites: S1 the structural ZnS₄ site previously characterized biochemically in HpFur and the two zinc sites identified in other Fur proteins. Site-directed mutagenesis and spectroscopy analyses of purified wild-type HpFur and various mutants show that the two metal binding sites common to other Fur proteins can be also metallated by cobalt. DNA protection and circular dichroism experiments demonstrate that, while these two sites influence the affinity of HpFur for DNA, only one is absolutely required for DNA binding and could be responsible for the conformational changes of Fur upon metal binding while the other is a secondary site.


The effects of betalain-rich cacti (dragon fruit and cactus pear) on endothelial and vascular function: a systematic review of animal and human studies.
Dragon fruit (Hylocereus) and cactus pear (Opuntia) are cacti species that have been widely used globally as a reliable source of food as well as traditional folk remedies. They have become of scientific interest recently due to their high levels of bioactive phytochemical compounds, in particular betalains. Earlier systematic reviews have explored the impact of supplementation of these cactus species on obesity, type-2 diabetes mellitus and cardiovascular risk factors: body weight and composition, serum triglycerides, cholesterol, blood glucose and blood pressure. However, effects on vascular health and endothelial function have yet to be reviewed. In order to address this gap in the literature, a systematic review has been conducted to evaluate the physiological effects of Hylocereus and Opuntia cacti on endothelial and vascular function in in vivo animal models and human studies. An electronic search was performed in the following databases: PubMed (MEDLINE), EMBASE (via Ovid), CINAHL, Scopus, Web of Science®, and The Cochrane Library (CENTRAL). All journals were searched since inceptions up to January 2020 without language restriction. Outcomes of interest were blood pressure, arterial stiffness, vascular reactivity and biochemical markers of endothelial dysfunction. Two investigators independently performed the study selection and data extraction. From 394 references, only 16 studies (9 animal and 7 human) fulfilled the eligibility criteria. Animal studies suggested a potential increase in vasodilation and serum nitric oxide and a reduction in vascular stiffness and blood pressure. The small number of human studies showed a reduction in heart rate as well as an increase in heart rate variability. Although these findings appear to indicate improvement in vascular health, there is a severe lack of robust, randomised human intervention studies to identify underlying mechanisms, optimal dose and long-term effects of cacti consumption.


Chronic obstructive pulmonary disease in stage I non-small cell lung cancer that underwent anatomic resection: the role of a recurrence promoter.
Despite the use of anatomic resection, the post-surgical recurrence rate remains high in early-stage non-small cell lung cancer (NSCLC). Chronic inflammation plays a role in the mechanism that promotes tumor initiation. This study aimed to investigate the association between recurrence outcome and chronic inflammation-related co-morbidities in early-stage resected NSCLC. A review of medical records for recurrence outcome and co-morbidities, in terms of chronic obstructive pulmonary disease (COPD), DM, asthma and cardiovascular diseases, was performed with 181 patients with stage I NSCLC that underwent anatomic resection. Subjects with T descriptors as T2a disease (49.5 vs. 28.0%, p < 0.05) and the presence of COPD (42.4 vs. 20.7%, p < 0.01) had a higher risk of tumor recurrence. Univariate analysis for recurrence-free survival showed T descriptor as T2a (21.5 months vs. NR, p < 0.05) and the presence of COPD (20.5 months vs. NR, p < 0.01) as significant factors predicting reduced survival. The presence of COPD (HR: 1.98; 95% CI, 1.29-.02, p < 0.01) and T descriptor as T2a (HR: 2.01; 95% CI, 1.04-3.91, p < 0.05) remain independent predictors of reduced recurrence-free survival in the Cox regression model. Patients with COPD were at higher risk of brain recurrence (OR: 7.88; 95% CI, 1.50-41.3, p < 0.01). In contrast, patients without COPD showed a tendency toward recurrence in bone and liver (OR: 4.13; 95% CI, 1.08-15.8, p = 0.05). Subjects with COPD and T2a disease had a higher risk of recurrence. The role of COPD as a recurrence promoter merits further prospective investigation.


Single-photon propagation through dielectric bandgaps.
Theoretical models of photon traversal through quarter-wave dielectric stack barriers that arise due to Bragg reflection predict the saturation of the propagation time with the barrier length, known as the Hartman effect. This saturation is sensitive to the addition of single dielectric layers, varying significantly from sub-luminal to apparently super-luminal and vice versa. Our research tests the suitability of photonic bandgaps as an optical model for the tunneling process. Of particular importance is our observation of subtle structural changes in dielectric stacks drastically affecting photon traversal times, allowing for apparent sub- and super-luminal effects. We also introduce a simple model to link HOM visibility to wavepacket distortion that allows us to exclude this as a possible cause of the loss of contrast in the barrier penetration process.


Barley Yellow Dwarf Luteoviruses and Their Predominant Aphid Vectors in Winter Wheat Grown in South Carolina.
Barley yellow dwarf is recognized as an important disease problem in winter wheat production in the southeastern United States, but there is relatively little known about the ecology and epidemiology of barley yellow dwarf virus (BYDV) in this region. From 1991 to 1993, and in 1996 and 1997, winter wheat was sampled for BYDV throughout the principal wheat production areas in South Carolina. In addition, in 1997, a small number of samples were collected from fields in North Carolina and Kentucky. Plant samples were assayed to determine the BYDV serotype and, subsequently, coat protein sequences of isolates within the same serotype were compared using restriction fragment length polymorphisms. Representative BYDV isolates from South Carolina and type isolates from New York were compared in aphid transmission experiments using aphid species collected from South Carolina and laboratory colonies maintained in New York. The predominant BYDV serotype in South Carolina (in all years) was PAV, accounting for 94% of the total BYDV-infected samples analyzed. The RPV serotypes were more abundant in samples collected from western North Carolina and Kentucky. PAV isolates from all regions were identical to the New York BYDV-PAV in terms of serology and restriction fragment patterns. Furthermore, the aphid transmission phenotypes were similar for South Carolina and New York BYDV isolates. The predominant aphids colonizing winter wheat in South Carolina included Schizaphis graminum, Rhopalosiphum rufiabdominalis, R. padi, and Sitobion avenea. The South Carolina clones of R. padi and S. avenae were similar to the New York laboratory clones in their abilities to transmit various BYDV isolates from New York and South Carolina. In contrast to the New York clone of Schizaphis graminum that can vector SGV, PAV, and RPV, the S. graminum clone from South Carolina was not a vector of any BYDV serotype tested. R. rufiabdominalis was found to be an efficient vector of PAV, RPV, and RMV isolates, but did not transmit MAV or SGV.


A Comprehensive Assessment of the Effects of Transgenic Cry1Ac/Cry1Ab Rice Huahui 1 on Adult Micraspis discolor (Fabricius) (Coleoptera: Coccinellidae).
Micraspis discolor (Fabricius) (Coleoptera: Coccinellidae) is a widely distributed coleoptera predator in southern Asia in rice ecosystem, and adult M. discolor feed on both rice pollen and soft-bodied arthropods. Bitrophic bioassay and tritrophic bioassay were conducted to evaluate the potential impact of Cry1Ac/Cry1Ab-expressing rice Huahui 1 and its non-transgenic counterpart Minghui 63 on fitness parameters of adult M. discolor. The results showed that the survival, and fecundity of this beetle' adults were not different when they fed on Bt rice or non-Bt rice pollen or Nilaparvata lugens (Stål) reared on Bt rice or non-Bt rice. Toxicity assessment to ensure M. discolor adults were not sensitive to Cry1Ab or Cry1Ac protein independent from the pollen background, M. discolor adults were fed with an artificial diet containing Cry1Ac, Cry1Ab or both protein approximately 10 times higher concentration than in Huahui 1 rice pollen. No difference was detected for any of the life-table parameters tested between Cry protein-containing and pure diet. Artificial diet containing E-64 (N-(trans-Epoxysuccinyl)-L-leucine 4-guanidinobutylamide) was included as a positive control. In contrast, the pre-oviposition and fecundity of M. discolor were significantly adversely affected by feeding on E-64-containing diet. In both bioassays, the uptakes of Cry protein by adult M. discolor were tested by ELISA measurements. These results indicated that adults of M. discolor are not affected by Cry1Ab- or Cry1Ac-expressing rice pollen and are not sensitive to Cry protein at concentrations exceeding the levels in rice pollen in Huahui1. This suggests that M. discolor adults would not be harmed by Cry1Ac/Cry1Ab rice if Bt rice Huahui 1 were commercialized.


Expansion of the SOS regulon of Vibrio cholerae through extensive transcriptome analysis and experimental validation.
The SOS response is an almost ubiquitous response of cells to genotoxic stresses. The full complement of genes in the SOS regulon for Vibrio species has only been addressed through bioinformatic analyses predicting LexA binding box consensus and in vitro validation. Here, we perform whole transcriptome sequencing from Vibrio cholerae treated with mitomycin C as an SOS inducer to characterize the SOS regulon and other pathways affected by this treatment. Comprehensive transcriptional profiling allowed us to define the full landscape of promoters and transcripts active in V. cholerae. We performed extensive transcription start site (TSS) mapping as well as detection/quantification of the coding and non-coding RNA (ncRNA) repertoire in strain N16961. To improve TSS detection, we developed a new technique to treat RNA extracted from cells grown in various conditions. This allowed for identification of 3078 TSSs with an average 5'UTR of 116 nucleotides, and peak distribution between 16 and 64 nucleotides; as well as 629 ncRNAs. Mitomycin C treatment induced transcription of 737 genes and 28 ncRNAs at least 2 fold, while it repressed 231 genes and 17 ncRNAs. Data analysis revealed that in addition to the core genes known to integrate the SOS regulon, several metabolic pathways were induced. This study allowed for expansion of the Vibrio SOS regulon, as twelve genes (ubiEJB, tatABC, smpA, cep, VC0091, VC1190, VC1369-1370) were found to be co-induced with their adjacent canonical SOS regulon gene(s), through transcriptional read-through. Characterization of UV and mitomycin C susceptibility for mutants of these newly identified SOS regulon genes and other highly induced genes and ncRNAs confirmed their role in DNA damage rescue and protection. We show that genotoxic stress induces a pervasive transcriptional response, affecting almost 20% of the V. cholerae genes. We also demonstrate that the SOS regulon is larger than previously known, and its syntenic organization is conserved among Vibrio species. Furthermore, this specific co-localization is found in other γ-proteobacteria for genes recN-smpA and rmuC-tatABC, suggesting SOS regulon conservation in this phylum. Finally, we comment on the limitations of widespread NGS approaches for identification of all RNA species in bacteria.


A novel strategy for repairing preganglionic cervical root avulsion in brachial plexus injury by sural nerve grafting.
In this study, the authors evaluated the efficacy of a new surgical strategy for reconnecting the injured brachial plexus with the spinal cord using fibrin glue containing acidic fibroblast growth factor as an adhesive and neurotrophic agent. Eighteen patients with preganglionic brachial plexus injuries, each with varying degrees of upper limb dysfunction, underwent cervical laminectomy with or without sural nerve grafting. The treatment of each avulsed root varied according to the severity of the injury. Some patients also underwent a second-stage operation involving supraclavicular brachial plexus exploration for reconnection with the corresponding segment of cervical spinal cord at the trunk level. Muscle strength was graded both pre- and postoperatively with the British Medical Research Council scale, and the results were analyzed with the Friedman and Wilcoxon signed-rank tests. Muscle strength improvements were observed in 16 of the 18 patients after 24 months of follow-up. Significant improvements in mean muscle strength were observed in patients from all repair method groups at 12 and 24 months postoperatively (p < 0.05). Statistical significance was not reached in the groups with insufficient numbers of cases. The authors' new surgical strategy yielded clinical improvement in muscle strength after preganglionic brachial plexus injury, such that nerve regeneration may have taken place. Reconnection of the brachial plexus to the cervical spinal cord is possible. Functional motor recovery, observed through increases in Medical Research Council-rated muscle strength in the affected arm, is likewise possible.


Ant distribution in relation to ground water in north Florida pine flatwoods.
Longleaf pine savannas are one of the most threatened ecosystems in the world, yet are understudied. Ants are a functionally important and diverse group of insects in these ecosystems. It is largely unknown how local patterns of species diversity and composition are determined through the interaction of this dominant animal group with abiotic features of longleaf pine ecosystems. Here we describe how an important abiotic variable, depth to water table, relates to ant species distributions at local scales. Pitfall trapping studies across habitat gradients in the Florida coastal plains longleaf pine flatwoods showed that the ant community changed with mild differences in habitat. In this undulating landscape, elevation differences were less than 2 m, and the depth to the water table ranged from < 20 cm to 1.2 m. The plant species composing the ground cover were zoned in response to depth to water, and shading by canopy trees increased over deeper water tables. Of the 27 ant species that were analyzed, depending on the statistical test, seven or eight were significantly more abundant over a deep water table, eight to ten over a shallow one, and nine to eleven were not significantly patterned with respect to depth to water. Ant species preferring sites with shallow groundwater also preferred the shadier parts of the sites, while those preferring sites with deeper groundwater preferred the sunnier parts of the sites. This suggests that one group of species prefers hot-dry conditions, and the other cooler-moist. Factor analysis and abundance-weighted mean site characteristics generally confirmed these results. These results show that ant communities in this region respond to subtle differences in habitat, but whether these differences arise from founding preferences, survival, competition, or some combination of these is not known.


Multi-component HPLC fingerprinting of Radix Salviae Miltiorrhizae and its LC-MS-MS identification.
The multi-component fingerprinting method of Radix Salviae Miltiorrhizae (Root of Salvia miltiorrhiza BGE.), an important and popular medicinal herb in traditional Chinese medicine, was studied using reverse-phase HPLC and LC-MS-MS. Extract containing both the water-soluble phenolic compounds and nonpolar diterpenoid compounds known to be the herb's main bioactive components was prepared by a two-step extractive procedure. An HPLC fingerprinting method which can simultaneously separate these two types of compounds was established with gradient elution mode and photodiode array detection at 280 nm. Eighteen peaks in this HPLC fingerprint were structurally identified by employing LC-MS-MS techniques. The electrospray ionization (ESI) MS-MS spectra of most salvianolic acids displayed a characteristic behavior of loss of danshensu and caffeic acid moieties, while those of tanshinones showed a particular behavior of loss of H2O which is quite different from the fragmentation pattern in electron ionization mass spectrometry (EI-MS). The HPLC fingerprints of 7 batches of crude drugs showed similar separation pattern and provided much chemical information of the pharmacologically-active compounds in the crude drugs, which is useful for the authentication and quality evaluation of this medicinal herb.


Leveraging Methylome-Environment Interaction to Detect Genetic Determinants of Disease.
The association between DNA methylation and a trait of interest may depend on an environmental exposure, and incorrectly accounting for this dependence can lead to a reduction in power of the standard tests used in epigenome-wide association studies. We present the M-ME test to jointly test for the main effect of DNA methylation and methylation-environment interaction. Through simulation, we compare the power and type 1 error of the M-ME test to a standard marginal test (M test) and a standard interaction test (ME test) under 1,800 different underlying models. These models allow for methylation-environment correlation and measurement error in the exposure. In many true underlying models, either the M test or the ME test has very low power, but the M-ME test has optimal or nearly optimal power to detect a DNA methylation effect in all models considered, including those with methylation- environment dependence and measurement error in the exposure. Type 1 error inflation occurs in the tests when the exposure is measured with error and correlated with DNA methylation. The M-ME test is an attractive choice for studies aiming to detect any DNA methylation association when little is known about the epigenetic associations a priori.


Patient blood management interventions do not lead to important clinical benefits or cost-effectiveness for major surgery: a network meta-analysis.
Patient blood management (PBM) interventions aim to improve clinical outcomes by reducing bleeding and transfusion. We assessed whether existing evidence supports the routine use of combinations of these interventions during and after major surgery. Five systematic reviews and a National Institute of Health and Care Excellence health economic review of trials of common PBM interventions enrolling participants of any age undergoing surgery were updated. The last search was on June 1, 2019. Studies in trauma, burns, gastrointestinal haemorrhage, gynaecology, dentistry, or critical care were excluded. The co-primary outcomes were: risk of receiving red cell transfusion and 30-day or hospital all-cause mortality. Treatment effects were estimated using random-effects models and risk ratios (RR) with 95% confidence intervals (CIs). Heterogeneity assessments used I2. Network meta-analyses used a frequentist approach. The protocol was registered prospectively (PROSPERO CRD42018085730). Searches identified 393 eligible randomised controlled trials enrolling 54 917 participants. PBM interventions resulted in a reduction in exposure to red cell transfusion (RR=0.60; 95% CI 0.57, 0.63; I2=77%), but had no statistically significant treatment effect on 30-day or hospital mortality (RR=0.93; 95% CI 0.81, 1.07; I2=0%). Treatment effects were consistent across multiple secondary outcomes, sub-groups and sensitivity analyses that considered clinical setting, type of intervention, and trial quality. Network meta-analysis did not demonstrate additive benefits from the use of multiple interventions. No trial demonstrated that PBM was cost-effective. In randomised trials, PBM interventions do not have important clinical benefits beyond reducing bleeding and transfusion in people undergoing major surgery.


Abnormal response to granulocyte colony-stimulating factor (G-CSF) in canine cyclic hematopoiesis is not caused by altered G-CSF receptor expression.
A decrease in responsiveness to granulocyte colony-stimulating factor (G-CSF) has been implicated in the pathophysiology of cyclic hematopoiesis. Using the canine model of cyclic neutropenia, we examined the response of neutrophil precursors to G-CSF in vitro and G-CSF receptor expression in neutrophils from grey collie dogs to determine whether the abnormal response observed to G-CSF in vivo in this disorder is present at the level of the progenitor cell and is caused by defective G-CSF receptor expression. Bone marrow mononuclear cells from grey collie dogs required sevenfold higher G-CSF concentrations than normal dog cells to achieve half-maximal colony growth [56 pmol/L v 8 pmol/L). Receptor binding assays with 125I-labeled G-CSF and Scatchard analyses of the equilibrium binding data were consistent with expression of a single class of high-affinity receptors for G-CSF on neutrophils from both normal dogs and grey collies with similar receptor numbers (56 to 446 sites/cell v 78 to 199 sites/cell) and binding affinities (28 to 206 pmol/L v 84 to 195 pmol/L). Chemical cross-linking studies identified a G-CSF binding protein of approximately 120 kD on neutrophils from grey collies, similar in size to that on normal dog neutrophils. No abnormal G-CSF receptor mRNA transcripts were detected in neutrophils from grey collie dogs by Northern blot analysis. Treatment of both normal and grey collie neutrophils with G-CSF rapidly induced tyrosine phosphorylation of an 80-kD protein that behaved like canine c-rel. These results demonstrate that the abnormal responsiveness to G-CSF in canine cyclic hematopoiesis is present in neutrophil precursors and is not associated with demonstrable alterations in the number, binding affinity, or overall size of the G-CSF receptor in neutrophils, or with defective tyrosine phosphorylation of p80. These data suggest that cyclic hematopoiesis is caused by a defect in the G-CSF signal transduction pathway at a point distal to G-CSF receptor binding that does not involve the early biochemical events leading to p80 tyrosine phosphorylation.


Changes in use of time, activity patterns, and health and wellbeing across retirement: design and methods of the life after work study.
Retirement is a major life transition during which people restructure everyday activities; however little is known about this. The primary aim of the Life After Work study is to comprehensively measure changes in time use and patterns of physical activity and sedentary behaviour, and its associations with health and wellbeing, across the retirement transition. A target sample of 120 participants aged 50 years and over will be recruited in two Australian state capital cities, Adelaide and Brisbane. Participants will undertake a battery of assessments approximately 3 months prior to retirement, and 3, 6 and 12 months post-retirement. Measures will include self-reported use of time (using the Multimedia Activity Recall for Children and Adults), objectively assessed physical activity and sedentary behaviour (using Actigraph GT3X+ accelerometers), self-reported health and well-being (using a battery of questionnaires including the Short-Form Health Survey (SF-36), Australian Unity Personal Well-being Index (AUPWI), Depression Anxiety Stress Scales 21 (DASS21), Short Warwick-Edinburgh Mental Well-being Scale, UCLA Loneliness Scale, Rosenberg Self-Esteem Scale), retirement circumstances and socio-demographic characteristics, objectively assessed anthropometric measures (height, weight and waist circumference), and resting blood pressure. Multivariate mixed models will be used to examine changes in use of time, health and well-being across retirement. The results will provide important new information that will inform the development of lifestyle and policy interventions to address and improve health and well-being in retirement.


Polarization measurements in high-energy deuteron photodisintegration.
We present measurements of the recoil proton polarization for the d(gamma-->,p-->)n reaction at straight theta(c.m.) = 90 degrees for photon energies up to 2.4 GeV. These are the first data in this reaction for polarization transfer with circularly polarized photons. The induced polarization p(y) vanishes above 1 GeV, contrary to meson-baryon model expectations, in which resonances lead to large polarizations. However, the polarization transfer Cx does not vanish above 1 GeV, inconsistent with hadron helicity conservation. Thus, we show that the scaling behavior observed in the d(gamma,p)n cross sections is not a result of perturbative QCD. These data should provide important tests of new nonperturbative calculations in the intermediate energy regime.


The atypical chemokine receptor ACKR3/CXCR7 is a broad-spectrum scavenger for opioid peptides.
Endogenous opioid peptides and prescription opioid drugs modulate pain, anxiety and stress by activating opioid receptors, currently classified into four subtypes. Here we demonstrate that ACKR3/CXCR7, hitherto known as an atypical scavenger receptor for chemokines, is a broad-spectrum scavenger of opioid peptides. Phylogenetically, ACKR3 is intermediate between chemokine and opioid receptors and is present in various brain regions together with classical opioid receptors. Functionally, ACKR3 is a scavenger receptor for a wide variety of opioid peptides, especially enkephalins and dynorphins, reducing their availability for the classical opioid receptors. ACKR3 is not modulated by prescription opioids, but we show that an ACKR3-selective subnanomolar competitor peptide, LIH383, can restrain ACKR3's negative regulatory function on opioid peptides in rat brain and potentiate their activity towards classical receptors, which may open alternative therapeutic avenues for opioid-related disorders. Altogether, our results reveal that ACKR3 is an atypical opioid receptor with cross-family ligand selectivity.


ARX, a novel Prd-class-homeobox gene highly expressed in the telencephalon, is mutated in X-linked mental retardation.
Investigation of a critical region for an X-linked mental retardation (XLMR) locus led us to identify a novel Aristaless related homeobox gene (ARX ). Inherited and de novo ARX mutations, including missense mutations and in frame duplications/insertions leading to expansions of polyalanine tracts in ARX, were found in nine familial and one sporadic case of MR. In contrast to other genes involved in XLMR, ARX expression is specific to the telencephalon and ventral thalamus. Notably there is an absence of expression in the cerebellum throughout development and also in adult. The absence of detectable brain malformations in patients suggests that ARX may have an essential role, in mature neurons, required for the development of cognitive abilities.


Differential distribution of salivary agglutinin and amylase in the Golgi apparatus and secretory granules of human salivary gland acinar cells.
The secretory granules of salivary glands often display complex internal substructures, yet little is known of the molecular organization of their contents or the mechanisms involved in packaging of the secretory proteins. We used post-embedding immunogold labeling with antibodies to two secretory proteins, agglutinin and alpha-amylase, to determine their distribution in the Golgi apparatus and secretory granules of the human submandibular gland acinar cells. With monoclonal antibodies specific for carbohydrate epitopes of the agglutinin, reactivity was found in the trans Golgi saccules, trans Golgi network, and immature and mature secretory granules. In the granules, labeling was seen in regions of low and medium electron density, but not in the dense cores. Reactivity seen on the apical and basolateral membranes of acinar and duct cells was attributed to a shared epitope on a membrane glycoprotein. Labeling with a polyclonal antibody to amylase was found in the Golgi saccules, immature and mature secretory granules, but not in the trans Golgi network. In the granules, amylase was present in the dense cores and in areas of medium density, but not in the regions of low density. These results indicate that these two proteins are distributed differently within the secretory granules, and suggest that they follow separate pathways between the Golgi apparatus and forming secretory granules. Small vesicles and tubular structures that labeled only with the antibodies to the agglutinin were observed on both faces of the Golgi apparatus and in the vicinity of the cell membrane. These structures may represent constitutive secretion vesicles involved in transport of the putative membrane glycoprotein to the cell membrane.


The Effect of a Backpack Hip Strap on Energy Expenditure While Walking.
Objective To examine the effect of backpack hip strap use on walking energy expenditure while carrying a loaded backpack. Background Previous studies have demonstrated that energy cost increases as the mass of the load carried increases. However, few investigations have focused on backpack carriage design. Methods Fifteen young, healthy, male subjects walked at a self-selected pace for 10 minutes in two backpack loading conditions: with a hip strap (strapped) and without a hip strap (nonstrapped). Oxygen consumption (VO2), rating of perceived exertion (RPE), respiratory exchange ratio (RER), and heart rate (HR) were monitored throughout each 10-minute trial. Change scores from the 4th to 10th minute were calculated for each variable. A t test was used to evaluate the difference between conditions for each variable. Results The changes in VO2 (-0.62 ± 0.40 vs. 0.33 ± 0.23, p = .04) and RPE (1 ± 0.25 vs. 2 ± 0.21, p < .01) from the 4th to the 10th minute were different for the strapped versus nonstrapped condition. There was no difference in the change in RER (0.04 ± 0.01 vs. 0.03 ± 0.01, p > .05) or HR (3.53 ± 0.93 vs. 4.07 ± 1.39, p > .05) for the strapped versus unstrapped condition. Conclusions Wearing a hip strap reduced the energy expenditure and perceived exertion in as little as 10 minutes of walking compared to the nonstrapped condition. Future work should consider the effect of a hip strap on these variables while hiking for extended periods. Application Wearing a hip strap may increase the comfort and reduce the energy required of wearing a backpack. This is useful information for backpack designers, military personnel, and recreational hikers.


Dual roles of the C2B domain of synaptotagmin I in synchronizing Ca2+-dependent neurotransmitter release.
Although the vesicular protein synaptotagmin I contains two Ca2+-binding domains (C2A and C2B), Ca2+ binding to the C2B domain is more important for triggering synchronous neurotransmitter release. We have used point mutagenesis to determine the functional contributions of the five negatively charged aspartate (Asp) residues that constitute the Ca2+-binding sites in the C2B domain of synaptotagmin I. Transfecting wild-type synaptotagmin I DNA into cultured hippocampal neurons from synaptotagmin I knock-out mice rescued Ca2+-dependent synchronous transmitter release and reduced a slower, asynchronous component of release, indicating that synaptotagmin I suppresses asynchronous release. Mutating either the second or third Asp residues of the C2B domain potently inhibited the ability of synaptotagmin I to rescue synchronous release but did not change its ability to suppress asynchronous release. Synaptotagmin I with mutations in the first or fourth Asp residues of the C2B domain partially rescued synchronous release and partially suppressed asynchronous release, whereas neutralizing the fifth Asp residue had no effect on the ability of synaptotagmin I to rescue transmitter release. Thus, we conclude that the C2B domain of synaptotagmin I regulates neurotransmitter release in at least two ways. Synchronous release absolutely requires binding of Ca2+ to the second and third Asp residues in this domain. For the suppression of asynchronous release, Ca2+ binding to the C2B domain of synaptotagmin I apparently is not necessary because mutation of the second Asp residue inhibits Ca2+ binding, yet still allows this protein to suppress asynchronous release.


Toward a better follow-up of ovarian recovery in young women after chemotherapy with a hypersensitive antimüllerian hormone assay.
To evaluate the utility of a hypersensitive assay for measuring low antimüllerian hormone (AMH) levels in young cancer patients during the ovarian recovery phase of their chemotherapy. Retrospective study. Academic medical center. Fifty-eight samples drawn at least 3 months after the end of chemotherapy in 30 women having either breast cancer (n=13) or hematologic malignancies (n=17) were selected to constitute two equally size groups: amenorrhea (n=30 samples) or spontaneous cycle (n=28 samples). None. Serum AMH levels were measured by a conventional AMH ELISA (EIA AMH/MIS) and a hypersensitive ELISA (PicoAMH, AnshLabs) on the same sample. Using a conventional assay, serum AMH was detectable (≥3 pmol/L) in 6.7% and in 10.7% of the samples corresponding to amenorrheic or cycling patients, respectively (nonsignificant). By contrast, with PicoAMH, serum AMH was detectable (≥0.07 pmol/L) in 71.4% of the samples from cycling women vs. 16.7% of the samples from amenorrheic patients. Multivariate regression analysis showed that among putative contributors, only the menstrual status (r=0.307) and serum FSH level (r=-0.546) were independently correlated to a detectable serum AMH with the picoAMH assay exclusively. The picoAMH assay, allowing measurement of very low AMH concentrations in human serum, should refine postchemotherapy ovarian follow-up in young women.


Continuous subtherapeutic insulin counteracts hypothalamopituitary-gonadal alterations in diabetic rats.
In experimental animal models, gonadal axis lesions are probably responsible for reproductive disorders associated with diabetes mellitus. The pathogenesis of these disorders is not yet known, but it is assumed that insulin deficiency plays an important role. To check this hypothesis, we have investigated the hypothalamopituitary-gonadal axis of insulin-treated streptozocin-induced diabetic (STZ-D) rats and compared it with that of untreated diabetic and control animals. Insulin was delivered by subcutaneously implanted osmotic minipumps. Furthermore, to determine whether possible beneficial insulin effects are selectively limited to the gonadal axis or act generally, we also studied retinal microangiopathy. The hypothalamopituitary-gonadal axis of insulin-treated diabetic animals was almost unchanged. On the contrary, retinal microangiopathy was only slightly influenced by subtherapeutic insulin doses. In conclusion, continuous administration of insulin at subtherapeutic doses can successfully counteract most of the effects of diabetes on the gonadal axis. Thus, the gonadal-axis impairment in STZ-D animals appears to be related to the fall of plasma insulin below a critical level. Furthermore, the various organ systems may respond to different plasma insulin threshold levels.


Optical coherence tomography is a useful tool in the differentiation between true edema and pseudoedema of the optic disc.
To assess the usefulness of spectral-domain optical coherence tomography (SD-OCT) peripapillary retinal nerve fiber layer (RNFL) thickness measurement in discriminating early phase optic disc edema (ODE) from pseudoedema (PODE). Hospital-based, multicenter, cross-sectional study involving external patients referred for recent identification of "presumed ODE". Patients underwent SD-OCT optic nerve head (ONH) RNFL thickness measurement at their first evaluation. In 155 of these, the causative etiology was subsequently ascertained and the respective eyes (one per patient) were assigned to the ODE (95 eyes) or PODE (60 eyes) group. Admission SD-OCT data were retrieved and used for the analysis. ROC curve analysis was used to calculate specificity, sensitivity and predictive value (PV) of the RNFL values. The PODE group was significantly younger than the ODE group (p = 0.007). The average and any single-quadrant RNFL thickness was significantly higher in the ODE group compared with the PODE and control groups. The average and the inferior quadrant thicknesses tested the most powerful parameters to differentiate ODE from PODE. A cutoff value of ≥ 110 μm for the average area, or of ≥ 150 μm for the inferior quadrant was associated with maximal sensitivity and specificity with positive PV greater than 80%. The SD-OCT evaluation of the peripapillary RNFL achieved good specificity, sensitivity and positive PV in discriminating between ODE and PODE. Despite the correct differential diagnosis between these categories still relies on a careful medical history taking and other ancillary testing, we proved the usefulness of SD-OCT RNFL measurement in supporting the diagnostic process.


Escape of TLR5 Recognition by Leptospira spp.: A Rationale for Atypical Endoflagella.
Leptospira (L.) interrogans are invasive bacteria responsible for leptospirosis, a worldwide zoonosis. They possess two periplasmic endoflagellae that allow their motility. L. interrogans are stealth pathogens that escape the innate immune recognition of the NOD-like receptors NOD1/2, and the human Toll-like receptor (TLR)4, which senses peptidoglycan and lipopolysaccharide (LPS), respectively. TLR5 is another receptor of bacterial cell wall components, recognizing flagellin subunits. To study the contribution of TLR5 in the host defense against leptospires, we infected WT and TLR5 deficient mice with pathogenic L. interrogans and tracked the infection by in vivo live imaging of bioluminescent bacteria or by qPCR. We did not identify any protective or inflammatory role of murine TLR5 for controlling pathogenic Leptospira. Likewise, subsequent in vitro experiments showed that infections with different live strains of L. interrogans and L. biflexa did not trigger TLR5 signaling. However, unexpectedly, heat-killed bacteria stimulated human and bovine TLR5, but did not, or barely induced stimulation via murine TLR5. Abolition of TLR5 recognition required extensive boiling time of the bacteria or proteinase K treatment, showing an unusual high stability of the leptospiral flagellins. Interestingly, after using antimicrobial peptides to destabilize live leptospires, we detected TLR5 activity, suggesting that TLR5 could participate in the fight against leptospires in humans or cattle. Using different Leptospira strains with mutations in the flagellin proteins, we further showed that neither FlaA nor Fcp participated in the recognition by TLR5, suggesting a role for the FlaB. FlaB have structural homology to Salmonella FliC, and possess conserved residues important for TLR5 activation, as shown by in silico analyses. Accordingly, we found that leptospires regulate the expression of FlaB mRNA according to the growth phase in vitro, and that infection with L. interrogans in hamsters and in mice downregulated the expression of the FlaB, but not the FlaA subunits. Altogether, in contrast to different bacteria that modify their flagellin sequences to escape TLR5 recognition, our study suggests that the peculiar central localization and stability of the FlaB monomers in the periplasmic endoflagellae, associated with the downregulation of FlaB subunits in hosts, constitute an efficient strategy of leptospires to escape the TLR5 recognition and the induced immune response.


Associations between sleep disturbance and suicidal ideation in adolescents admitted to an inpatient psychiatric unit.
The goals of our study were to: 1) describe the incidence of disturbances in sleep quality, sleep hygiene, sleep-related cognitions and nightmares; and 2) investigate the association between these sleep-related disturbances and suicidal ideation (SI), in adolescents admitted to a psychiatric inpatient unit. Our sample consisted of 50 adolescents between the ages of 12 and 17 years (32 females and 18 males; 41 Caucasian and nine African American). Our cross-sectional design involved the administration of the Adolescent Sleep Wake Scale (ASWS), the Adolescent Sleep Hygiene Scale (ASHS), the Dysfunctional Beliefs and Attitudes about Sleep-Short version for use with children (DBAS-C10), the Disturbing Dreams and Nightmare Scale (DDNSI), and the Suicidal Ideation Questionnaire Jr (SIQ-JR). Analyses were conducted using Pearson correlations, as well as univariate and multivariate regression. Results indicated that our sample experienced sleep disturbances and SI to a greater degree than non-clinical samples. Sleep quality was correlated with nightmares, while sleep quality and nightmares were each correlated with SI. Sleep quality, dysfunctional beliefs, and nightmares each independently predicted SI. Our study was the first to use the four sleep measures with an adolescent psychiatric inpatient sample. It is important to develop sleep-related assessment tools in high-risk populations given the link between sleep disturbances and suicidality. Furthermore, a better understanding of the relationships between SI and sleep quality, sleep-related cognitions, and nightmares is needed to develop potential prevention and treatment options for suicidality in adolescents.


The exocytotic fusion pore modeled as a lipidic pore.
Freeze-fracture electron micrographs from degranulating cells show that the lumen of the secretory granule is connected to the extracellular compartment via large (20 to 150 nm diameter) aqueous pores. These exocytotic fusion pores appear to be made up of a highly curved bilayer that spans the plasma and granule membranes. Conductance measurements, using the patch-clamp technique, have been used to study the fusion pore from the instant it conducts ions. These measurements reveal the presence of early fusion pores that are much smaller than those observed in electron micrographs. Early fusion pores open abruptly, fluctuate, and then either expand irreversibly or close. The molecular structure of these early fusion pores is unknown. In the simplest extremes, these early fusion pores could be either ion channel like protein pores or lipidic pores. Here, we explored the latter possibility, namely that of the early exocytotic fusion pore modeled as a lipid-lined pore whose free energy was composed of curvature elastic energy and work done by tension. Like early exocytotic fusion pores, we found that these lipidic pores could open abruptly, fluctuate, and expand irreversibly. Closure of these lipidic pores could be caused by slight changes in lipid composition. Conductance distributions for stable lipidic pores matched those of exocytotic fusion pores. These findings demonstrate that lipidic pores can exhibit the properties of exocytotic fusion pores, thus providing an alternate framework with which to understand and interpret exocytotic fusion pore data.


Native cancer navigation: the state of the science.
The purpose of this literature review is to determine the current state of the science for the effectiveness of patient navigation on improving outcomes of cancer care across the continuum among Native Americans. The research will help healthcare professionals ascertain potential evidence-based practice guidelines and gaps in knowledge, which may provide direction for future research. Data synthesis included the use of Native navigation for cancer care, which has been demonstrated in limited, nonrandomized studies to improve cancer knowledge, access to care, and quality of life for Native Americans. Those studies had limitations, including small sample size, self-report of outcome measures, and lack of randomization. Evidence is insufficient to conclude that the use of Native navigation is superior to usual cancer care for Native American patients. Oncology nurses have a role in training personnel to serve as cancer navigators. Nurses need to be supportive of culturally appropriate navigation programs and know about services provided by navigators. In addition, nurse educators need to encourage Native Americans in their communities to consider choosing nursing as a profession. If an oncology nurse has an interest in research, opportunities exist to assist with or conduct research projects regarding Native cancer navigation. A particular need exists for addressing the gaps in research identified in this article.


1H,13C and 15N chemical shift assignments of the SUD domains of SARS-CoV-2 non-structural protein 3c: "the N-terminal domain-SUD-N".
Among the proteins encoded by the SARS-CoV-2 RNA, nsP3 (non-structural Protein3) is the largest multi-domain protein. Its role is multifaceted and important for the viral life cycle. Nonetheless, regarding the specific role of each domain there are many aspects of their function that have to be investigated. SARS Unique Domains (SUDs), constitute the nsP3c region of the nsP3, and were observed for the first time in SARS-CoV. Two of them, namely SUD-N (the first SUD) and the SUD-M (sequential to SUD-N), exhibit structural homology with nsP3b ("X" or macro domain); indeed all of them are folded in a three-layer α/β/α sandwich. On the contrary, they do not exhibit functional similarities, like ADP-ribose binding properties and ADP-ribose hydrolase activity. There are reports that suggest that these two SUDs may exhibit a binding selectivity towards G-oligonucleotides, a feature which may contribute to the characterization of their role in the formation of the replication/transcription viral complex (RTC) and of the interaction of various viral "components" with the host cell. While the structures of these domains of SARS-CoV-2 have not been determined yet, SUDs interaction with oligonucleotides and/or RNA molecules may provide a platform for drug discovery. Here, we report the almost complete NMR backbone and side-chain resonance assignment (1H,13C,15N) of SARS-CoV-2 SUD-N protein, and the NMR chemical shift-based prediction of the secondary structure elements. These data may be exploited for its 3D structure determination and the screening of chemical compounds libraries, which may alter SUD-N function.


Bile Acid-Based Drug Delivery Systems for Enhanced Doxorubicin Encapsulation: Comparing Hydrophobic and Ionic Interactions in Drug Loading and Release.
Doxorubicin (Dox) is a drug of choice in the design of drug delivery systems directed toward breast cancers, but is often limited by loading and control over its release from polymer micelles. Bile acid-based block copolymers present certain advantages over traditional polymer-based systems for drug delivery purposes, since they can enable a higher drug loading via the formation of a reservoir through their aggregation process. In this study, hydrophobic and electrostatic interactions are compared for their influence on Dox loading inside cholic acid based block copolymers. Poly(allyl glycidyl ether) (PAGE) and poly(ethylene glycol) (PEG) were grafted from the cholic acid (CA) core yielding a star-shaped block copolymer with 4 arms (CA-(PAGE- b-PEG)4) and then loaded with Dox via a nanoprecipitation technique. A high Dox loading of 14 wt % was achieved via electrostatic as opposed to hydrophobic interactions with or without oleic acid as a cosurfactant. The electrostatic interactions confer a pH responsiveness to the system. 50% of the loaded Dox was released at pH 5 in comparison to 12% at pH 7.4. The nanoparticles with Dox loaded via hydrophobic interactions did not show such a pH responsiveness. The systems with Dox loaded via electrostatic interactions showed the lowest IC50 and highest cellular internalization, indicating the pre-eminence of this interaction in Dox loading. The blank formulations are biocompatible and did not show cytotoxicity up to 0.17 mg/mL. The new functionalized star block copolymers based on cholic acid show great potential as drug delivery carriers.


High calorie diet triggers hypothalamic angiopathy.
Obesity, type 2 diabetes, and related diseases represent major health threats to modern society. Related pathophysiology of impaired neuronal function in hypothalamic control centers regulating metabolism and body weight has been dissected extensively and recent studies have started focusing on potential roles of astrocytes and microglia. The hypothalamic vascular system, however, which maintains the microenvironment necessary for appropriate neuronal function, has been largely understudied. We recently discovered that high fat/high sucrose diet exposure leads to increased hypothalamic presence of immunoglobulin G (IgG1). Investigating this phenomenon further, we have discovered a significant increase in blood vessel length and density in the arcuate nucleus (ARC) of the hypothalamus in mice fed a high fat/high sucrose diet, compared to matched controls fed standard chow diet. We also found a clearly increased presence of α-smooth muscle actin immunoreactive vessels, which are rarely present in the ARC and indicate an increase in the formation of new arterial vessels. Along the blood brain barrier, an increase of degenerated endothelial cells are observed. Moreover, such hypothalamic angiogenesis was not limited to rodent models. We also found an increase in the number of arterioles of the infundibular nucleus (the human equivalent of the mouse ARC) in patients with type 2 diabetes, suggesting angiogenesis occurs in the human hypothalamus of diabetics. Our discovery reveals novel hypothalamic pathophysiology, which is reminiscent of diabetic retinopathy and suggests a potential functional involvement of the hypothalamic vasculature in the later stage pathogenesis of metabolic syndrome.


Small lesions of the dorsal or ventral hippocampus subregions are associated with distinct impairments in working memory and reference memory retrieval, and combining them attenuates the acquisition rate of spatial reference memory.
The importance of the hippocampus in spatial learning is well established, but the precise relative contributions by the dorsal (septal) and ventral (temporal) subregions remain unresolved. One debate revolves around the extent to which the ventral hippocampus contributes to spatial navigation and learning. Here, separate small subtotal lesions of dorsal hippocampus or ventral hippocampus alone (destroying 18.9 and 28.5% of total hippocampal volume, respectively) spared reference memory acquisition in the water maze. By contrast, combining the two subtotal lesions significantly reduced the rate of acquisition across days. This constitutes evidence for synergistic integration between dorsal and ventral hippocampus in mice. Evidence that ventral hippocampus contributes to spatial/navigation learning also emerged early on during the retention probe test as search preference was reduced in mice with ventral lesions alone or combined lesions. The small ventral lesions also led to anxiolysis in the elevated plus maze and over-generalization of the conditioned freezing response to a neutral context. Similar effects of comparable magnitudes were seen in mice with combined lesions, suggesting that they were largely due to the small ventral damage. By contrast, small dorsal lesions were uniquely associated with a severe spatial working memory deficit in the water maze. Taken together, both dorsal and ventral poles of the hippocampus contribute to efficient spatial navigation in mice: While the integrity of dorsal hippocampus is necessary for spatial working memory, the acquisition and retrieval of spatial reference memory are modulated by the ventral hippocampus. Although the impairments following ventral damage (alone or in combination with dorsal damage) were less substantial, a wider spectrum of spatial learning, including context conditioning, was implicated. Our results encourage the search for integrative mechanism between dorsal and ventral hippocampus in spatial learning. Candidate neural substrates may include dorsoventral longitudinal connections and reciprocal modulation via overlapping polysynaptic networks beyond hippocampus.


Analysis of sequences and loci of p44 homologs expressed by Anaplasma phagocytophila in acutely infected patients.
Anaplasma phagocytophila is an obligatory intragranulocytic bacterium that causes human granulocytic ehrlichiosis. Immunodominant 44-kDa outer membrane proteins of A. phagocytophila are encoded by a p44 multigene family. In the present study, expression profiles of p44 genes in the blood of acutely infected patients in the year 2000 were characterized. A single p44 gene was predominantly expressed in peripheral blood leukocytes from one patient, while up to 17 different p44 genes were transcribed without a single majority in the other two patients. The cDNA sequences of the central hypervariable region of several p44 genes were identical among the isolates from the three patients and a 1995 A. phagocytophila isolate. A. phagocytophila was isolated by cell culture from all of the three 2000 patients. Genomic Southern blot analysis of the three 2000 and two 1995 A. phagocytophila isolates with probes specific to the most dominant p44 transcript in each patient showed that the p44 loci in the A. phagocytophila genome were conserved. Analysis of the predicted amino acid sequences of 43 different p44 genes including 19 new sequences found in the present study, revealed that five amino acids were absolutely conserved. The hypervariable region was subdivided into five domains, including three extremely hypervariable central domains. These results suggest that variations in the sequences of p44 are not random but are restricted. Furthermore, several p44 genes are not hypermutatable in nature, based on the conservation of gene sequences and loci among isolates obtained 5 years apart.


Expression of a malarial Hsp70 improves defects in chaperone-dependent activities in ssa1 mutant yeast.
Plasmodium falciparum causes the most virulent form of malaria and encodes a large number of molecular chaperones. Because the parasite encounters radically different environments during its lifecycle, many members of this chaperone ensemble may be essential for P. falciparum survival. Therefore, Plasmodium chaperones represent novel therapeutic targets, but to establish the mechanism of action of any developed therapeutics, it is critical to ascertain the functions of these chaperones. To this end, we report the development of a yeast expression system for PfHsp70-1, a P. falciparum cytoplasmic chaperone. We found that PfHsp70-1 repairs mutant growth phenotypes in yeast strains lacking the two primary cytosolic Hsp70s, SSA1 and SSA2, and in strains harboring a temperature sensitive SSA1 allele. PfHsp70-1 also supported chaperone-dependent processes such as protein translocation and ER associated degradation, and ameliorated the toxic effects of oxidative stress. By introducing engineered forms of PfHsp70-1 into the mutant strains, we discovered that rescue requires PfHsp70-1 ATPase activity. Together, we conclude that yeast can be co-opted to rapidly uncover specific cellular activities mediated by malarial chaperones.