Silver nanoparticles induce neurotoxicity in a human embryonic stem cell-derived neuron and astrocyte network.
Silver nanoparticles (AgNPs) are among the most extensively used nanoparticles and are found in a variety of products. This ubiquity leads to inevitable exposure to these particles in everyday life. However, the effects of AgNPs on neuron and astrocyte networks are still largely unknown. In this study, we used neurons and astrocytes derived from human embryonic stem cells as a cellular model to study the neurotoxicity that is induced by citrate-coated AgNPs (AgSCs). Immunostaining with the astrocyte and neuron markers, glial fibrillary acidic protein and microtubule-associated protein-2 (MAP2), respectively, showed that exposure to AgSCs at the concentration of 0.1 µg/mL increased the astrocyte/neuron ratio. In contrast, a higher concentration of AgSCs (5.0 µg/ml) significantly changed the morphology of astrocytes. These results suggest that astrocytes are sensitive to AgSC exposure and that low concentrations of AgSCs promote astrogenesis. Furthermore, our results showed that AgSCs reduced neurite outgrowth, decreased the expression of postsynaptic density protein 95 and synaptophysin, and induced neurodegeneration in a concentration-dependent manner. Our findings additionally suggest that the expression and phosphorylation status of MAP2 isoforms, as modulated by the activation of the Akt/glycogen synthase kinase-3/caspase-3 signaling pathway, may play an important role in AgSC-mediated neurotoxicity. We also found that AgNO3 exposure only slightly reduced neurite outgrowth and had little effect on MAP2 expression, suggesting that AgSCs and AgNO3 have different neuronal toxicity mechanisms. In addition, most of these effects were reduced when the cell culture was co-treated with AgSCs and the antioxidant ascorbic acid, which implies that oxidative stress is the major cause of AgSC-mediated astrocytic/neuronal toxicity and that antioxidants may have a neuroprotective effect.


Menopausal complaints in Slovak midlife women and the impact of CYP1B1 polymorphism on their incidence.
A wide variety of symptoms have been attributed to menopause, negatively influencing women's physical and psychological health. In addition to lifestyle parameters and personal history, genetic factors are considered to be the main source of this variation. This study aims to investigate the incidence of menopausal symptoms among midlife women according to their menopausal status, and to evaluate the contribution to their manifestation from CYP1B1 Leu432Val polymorphism as a predisposing factor for menopausal symptoms. The studied cohort consisted of 299 women ranging from 39 to 59 years of age. Women were recruited from the western and middle parts of Slovakia, and all participants completed a menopause-specific questionnaire and provided blood or saliva samples for genotyping. Our results indicated that all women are at risk of typical menopausal symptoms, but there is a higher number of postmenopausal women affected than premenopausal ones. Regression analysis showed that the CYP1B1 Leu/Leu genotype can increase the experience of bloated stomach and facial hair increase in all the sampled women, while the Leu/Leu genotype may increase experience of palpitations and involuntary urination in the premenopausal women. The Leu/Leu genotype may increase the experience of nausea, bloated stomach, and vaginal dryness in peri- and postmenopausal women. We determined that women with the Leu/Leu, or Leu/Val genotypes were approximately five times more likely to suffer from vaginal dryness than the Val/Val women (OR = 4.948; 95% CI, 1.259-19.447). We therefore suggest that CYP1B1 Leu432Val polymorphism could be involved in individual susceptibility to menopausal symptoms in Slovak midlife women.


HIV-1 Molecular Epidemiology, Transmission Clusters and Transmitted Drug Resistance Mutations in Central Brazil.
We aimed to characterize HIV-1 molecular epidemiology and transmission clusters among heterosexual (HET) and men who have sex with men (MSM) individuals, as well as transmitted drug resistance mutations (TDRM) in Central-Western Brazil. This cross-sectional survey was conducted among 190 antiretroviral naïve HIV-1 infected individuals. Proviral DNA was extracted, and nested PCR amplified partial polymerase gene (PR/RT). After sequencing, subtypes were assigned, and the sequences were analyzed for the occurrence of possible transmission networks. Calibrated Population Resistance (CPR) tool from Stanford HIV Database was used to investigate the presence of TDRM. Among 150 individuals whose samples were successfully sequenced, the most prevalent HIV-1 subtype was B, followed by recombinant forms. The occurrence of twenty transmission clusters composed by at least two sequences was verified, suggesting the existence of transmission clusters among individuals from the same or distinct sexual orientations. Intermediate level of TDRM (12%) was found in the study population, and almost half of the subjects with TDRM had more than one resistance mutation. No correlations between sexual orientation and the presence of TDRM, HIV-1 subtypes/recombinants forms were verified. Taken together, the necessity of the continuous monitoring of the TDRM to verify the importance of pre-genotyping and to delineate future strategies in primary antiretroviral therapy. Likewise, the knowledge of the HIV-1 transmission networks in Brazil would allow the implementation of effective HIV-1 prevention strategies in local settings.


The Mitotic Spindle in the One-Cell C. elegans Embryo Is Positioned with High Precision and Stability.
Precise positioning of the mitotic spindle is important for specifying the plane of cell division, which in turn determines how the cytoplasmic contents of the mother cell are partitioned into the daughter cells, and how the daughters are positioned within the tissue. During metaphase in the early Caenorhabditis elegans embryo, the spindle is aligned and centered on the anterior-posterior axis by a microtubule-dependent machinery that exerts restoring forces when the spindle is displaced from the center. To investigate the accuracy and stability of centering, we tracked the position and orientation of the mitotic spindle during the first cell division with high temporal and spatial resolution. We found that the precision is remarkably high: the cell-to-cell variation in the transverse position of the center of the spindle during metaphase, as measured by the standard deviation, was only 1.5% of the length of the short axis of the cell. Spindle position is also very stable: the standard deviation of the fluctuations in transverse spindle position during metaphase was only 0.5% of the short axis of the cell. Assuming that stability is limited by fluctuations in the number of independent motor elements such as microtubules or dyneins underlying the centering machinery, we infer that the number is ∼1000, consistent with the several thousand of astral microtubules in these cells. Astral microtubules grow out from the two spindle poles, make contact with the cell cortex, and then shrink back shortly thereafter. The high stability of centering can be accounted for quantitatively if, while making contact with the cortex, the astral microtubules buckle as they exert compressive, pushing forces. We thus propose that the large number of microtubules in the asters provides a highly precise mechanism for positioning the spindle during metaphase while assembly is completed before the onset of anaphase.


Predictions of single-nucleotide polymorphism differentiation between two populations in terms of mutual information.
Mutual information (I) provides a robust measure of genetic differentiation for the purposes of estimating dispersal between populations. At present, however, there is little predictive theory for I. The growing importance in population biology of analyses of single-nucleotide and other single-feature polymorphisms (SFPs) is a potent reason for developing an analytic theory for I with respect to a single locus. This study represents a first step towards such a theory. We present theoretical predictions of I between two populations with respect to a single haploid biallelic locus. Dynamical and steady-state forecasts of I are derived from a Wright-Fisher model with symmetrical mutation between alleles and symmetrical dispersal between populations. Analytical predictions of a simple Taylor approximation to I are in good agreement with numerical simulations of I and with data on I from SFP analyses of dispersal experiments on Drosophila fly populations. The theory presented here also provides a basis for the future inclusion of selection effects and extension to multiallelic loci.


Adeno-associated virus serotype rh.10 displays strong muscle tropism following intraperitoneal delivery.
Recombinant adeno-associated virus (rAAV) is an attractive tool for basic science and translational medicine including gene therapy, due to the versatility in its cell and organ transduction. Previous work indicates that rAAV transduction patterns are highly dependent on route of administration. Based on this relationship, we hypothesized that intraperitoneal (IP) administration of rAAV produces unique patterns of tissue tropism. To test this hypothesis, we investigated the transduction efficiency of 12 rAAV serotypes carrying an enhanced green fluorescent protein (EGFP) reporter gene in a panel of 12 organs after IP injection. Our data suggest that IP administration emphasizes transduction patterns that are different from previously reported intravascular delivery methods. Using this approach, rAAV efficiently transduces the liver, pancreas, skeletal muscle, heart and diaphragm without causing significant histopathological changes. Of note, rAAVrh.10 showed excellent muscle transduction following IP administration, highlighting its potential as a new muscle-targeting vector.


Urban distribution of Rhytisma acerinum (Pers.) Fries (tar spot) on sycamore.
This study examined ecological aspects of the distribution of tar spot disease caused by the fungus Rhytisma acerinum (Pers.) Fries on Acer pseudoplatanus L. (sycamore). The host is abundant in Edinburgh and surrounding areas yet no tar spots were found in the city, although they were abundant on sycamore in the surrounding countryside, especially at sheltered sites. The vertical distribution from a localized source of leaf litter from infected sycamore showed an approximately linear relationship between tar spot index of attached leaves (TSI = no. of tar spots per 100 cm2 of leaf) and the log of height of such infected foliage above the ground. The distribution of tar spots was controlled primarily by the presence or absence of overwintered sycamore leaves infected with R. acerinum. In Edinburgh these leaves are actively removed by man and passively by wind, so reducing the potential source of inoculum by the following spring. The presence of SO2 was unimportant in the distribution of R. acerinum. An experiment to study the relative importance of current air concentrations of SO2 on the incidence of infection by R. acerinum showed that the average SO2 concentration of < 50 μg m-3 hail no effect. The observed distribution of tar spot throughout the city and surrounding countryside could be explained entirely by distance from the nearest source of inoculum without involving a role for air pollutants.


Chronic exposure to free fatty acid reduces pancreatic beta cell insulin content by increasing basal insulin secretion that is not compensated for by a corresponding increase in proinsulin biosynthesis translation.
The pancreatic beta cell normally maintains a stable balance among insulin secretion, insulin production, and insulin degradation to keep optimal intracellular stores of the hormone. Elevated levels of FFA markedly enhance insulin secretion; however, the effects of FFA on insulin production and intracellular stores remain unclear. In this study, twofold elevation in total circulating FFA effected by infusion of lard oil and heparin into rats for 6 h under normoglycemic conditions resulted in a marked elevation of circulating insulin levels evident after 4 h, and a 30% decrease in pancreatic insulin content after a 6-h infusion in vivo. Adding 125 muM oleate to isolated rat pancreatic islets cultured with 5.6 mM glucose caused a 50% fall in their insulin content over 24 h, coupled with a marked enhancement of basal insulin secretion. Both effects of fatty acid were blocked by somatostatin. In contrast to the stimulatory effects of oleate on insulin secretion, glucose-induced proinsulin biosynthesis was inhibited by oleate up to 24 h, but was unaffected thereafter. This result was in spite of a two- to threefold oleate-induced increase in preproinsulin mRNA levels, underscoring the importance of translational regulation of proinsulin biosynthesis in maintaining beta cell insulin stores. Collectively, these results suggest that chronically elevated FFA contribute to beta cell dysfunction in the pathogenesis of NIDDM by significantly increasing the basal rate of insulin secretion. This increase in turn results in a decrease in the beta cell's intracellular stores that cannot be offset by commensurate FFA induction of proinsulin biosynthesis.


Does caffeine consumption during pregnancy increase the risk of fetal mortality? A literature review.
The aim of this study was to evaluate the available epidemiological evidence of the effect of caffeine consumption during pregnancy on fetal mortality. A systematic qualitative review of observational studies that referred to any source of exposure to caffeine from food in pregnancy and to fetal mortality as the outcome was conducted in the databases MEDLINE and LILACS. Studies published between January 1966 and September 2004 were searched. The following descriptors were used: "caffeine", "coffee", "tea", "cola", and "cacao" to define the exposure and "fetal death", "stillbirth", "fetal demise", and "fetal loss" to define the outcome. The search strategy retrieved 32 publications, but only six met the inclusion criteria and three were included. One more article was found using "see related articles" feature in PubMed. A total of four publications were included in the review. The small number of publications addressing this subject, methodological limitations, inaccurate exposure assessment in all the studies, overall risks only marginally significant in most cases, and the possibility of publication bias preclude stating with certainty that caffeine consumption is actually associated with fetal death.


Two aspects of the clinical and humanistic burden of systemic lupus erythematosus: mortality risk and quality of life early in the course of disease.
To evaluate mortality risk and predictors among recently diagnosed systemic lupus erythematosus (SLE) patients. The vital status of 265 SLE patients and 355 controls enrolled in the Carolina Lupus Study (median time since diagnosis 13 months) was determined approximately 5 years after enrollment. We also assessed the utility of an 8-item quality of life instrument, derived from the standard 36-item Medical Outcomes Study Short Form 36, as an additional measure of disease impact. Five years after diagnosis, 9.7% of patients compared with 0.3% of controls had died (P < 0.0001). Increased mortality risk was seen among older patients (adjusted hazard ratio [HR] 1.03, 95% confidence interval [95% CI] 1.01-1.06 per 1-year increment in age) and among men, African Americans, patients with lupus nephritis, and patients with anti-double-stranded DNA antibodies (adjusted HR approximately 2.0 for each of these factors). In addition, patients who did not provide a blood sample at study enrollment experienced increased mortality risk (age-, sex-, and race-adjusted HR 3.7, 95% CI 1.5-9.1). Similar results were seen in analyses limited to time from study enrollment. Physical component scores of the quality of life measure were 7.7 points lower (P < 0.0001) and mental component scores were 1.8 points lower (P = 0.07) in patients compared with controls. The mortality risk among SLE patients is significant, particularly among African Americans, even early in the disease process and even with currently available treatments. Differences between cases and controls in health-related quality of life using the Short Form 8 also demonstrate the multidimensional burden of SLE.


Lipostar, a Comprehensive Platform-Neutral Cheminformatics Tool for Lipidomics.
To date, the main limitations for LC-MS-based untargeted lipidomics reside in the lack of adequate computational and cheminformatics tools that are able to support the analysis of several thousands of species from biological samples, enabling data mining and automating lipid identification and external prediction processes. To address these issues, we developed Lipostar, novel vendor-neutral high-throughput software that effectively supports both targeted and untargeted LC-MS lipidomics, implementing data acquisition, user-friendly multivariate analysis (to be used for model generation and new sample predictions), and advanced lipid identification protocols that can work with or without the support of preformed lipid databases. Moreover, Lipostar integrates the lipidomic processes with a full metabolite identification (MetID) procedure, essential in drug safety applications and in translational studies. Case studies demonstrating a number of Lipostar features are also presented.


Systems Genetic Analyses Highlight a TGFβ-FOXO3 Dependent Striatal Astrocyte Network Conserved across Species and Associated with Stress, Sleep, and Huntington's Disease.
Recent systems-based analyses have demonstrated that sleep and stress traits emerge from shared genetic and transcriptional networks, and clinical work has elucidated the emergence of sleep dysfunction and stress susceptibility as early symptoms of Huntington's disease. Understanding the biological bases of these early non-motor symptoms may reveal therapeutic targets that prevent disease onset or slow disease progression, but the molecular mechanisms underlying this complex clinical presentation remain largely unknown. In the present work, we specifically examine the relationship between these psychiatric traits and Huntington's disease (HD) by identifying striatal transcriptional networks shared by HD, stress, and sleep phenotypes. First, we utilize a systems-based approach to examine a large publicly available human transcriptomic dataset for HD (GSE3790 from GEO) in a novel way. We use weighted gene coexpression network analysis and differential connectivity analyses to identify transcriptional networks dysregulated in HD, and we use an unbiased ranking scheme that leverages both gene- and network-level information to identify a novel astrocyte-specific network as most relevant to HD caudate. We validate this result in an independent HD cohort. Next, we computationally predict FOXO3 as a regulator of this network, and use multiple publicly available in vitro and in vivo experimental datasets to validate that this astrocyte HD network is downstream of a signaling pathway important in adult neurogenesis (TGFβ-FOXO3). We also map this HD-relevant caudate subnetwork to striatal transcriptional networks in a large (n = 100) chronically stressed (B6xA/J)F2 mouse population that has been extensively phenotyped (328 stress- and sleep-related measurements), and we show that this striatal astrocyte network is correlated to sleep and stress traits, many of which are known to be altered in HD cohorts. We identify causal regulators of this network through Bayesian network analysis, and we highlight their relevance to motor, mood, and sleep traits through multiple in silico approaches, including an examination of their protein binding partners. Finally, we show that these causal regulators may be therapeutically viable for HD because their downstream network was partially modulated by deep brain stimulation of the subthalamic nucleus, a medical intervention thought to confer some therapeutic benefit to HD patients. In conclusion, we show that an astrocyte transcriptional network is primarily associated to HD in the caudate and provide evidence for its relationship to molecular mechanisms of neural stem cell homeostasis. Furthermore, we present a unified systems-based framework for identifying gene networks that are associated with complex non-motor traits that manifest in the earliest phases of HD. By analyzing and integrating multiple independent datasets, we identify a point of molecular convergence between sleep, stress, and HD that reflects their phenotypic comorbidity and reveals a molecular pathway involved in HD progression.


Proliferative properties of human umbilical cord blood megakaryocyte progenitor cells to human thrombopoietin.
We evaluated the effects of human thrombopoietin (TPO) alone, or in combination with other several hematopoietic factors, on megakaryocyte colony growth from human umbilical cord blood CD34+ cells in serum-depleted agar cultures. The addition of TPO alone had a concentration-dependent effect on the number of pure megakaryocyte colonies and of megakaryocytes per colony (colony size). The maximally stimulating concentration of TPO generated a greater number of megakaryocyte colonies and larger megakaryocyte colony size compared with the stimulation observed with an optimal concentration of human interleukin-3 (IL-3) or stem cell factor (SCF). At the high concentration of TPO that yielded the maximum colony numbers, a substantial proportion of megakaryocyte colonies contained 100 or more cells per colony. The combination of TPO plus SCF resulted in a synergistic enhancement of both the number and size of megakaryocyte colonies. Among the combinations of TPO plus other cytokines tested, IL-3 plus TPO had a modest effect on megakaryocyte colony numbers. The generation of megakaryocyte colonies from subpopulations of CD34+ cells was further examined. The addition of TPO alone induced a greater number of megakaryocyte colonies from CD34+CD41+ cells compared with CD34+CD41- cells, and TPO plus IL-3 exerted a synergistic effect on the number of megakaryocyte colonies only from CD34+CD41- cells. In contrast to the effects on colony numbers, TPO induced larger megakaryocyte colony sizes from CD34+CD41- cells, compared with CD34+CD41+ cells. In the case of HLA-DR expression, TPO and IL-3, administered singly or in combination, induced similar megakaryocyte colony numbers and sizes from CD34+DR+ and CD34+DR- subpopulations. Ploidy analysis revealed that the majority of megakaryocytes generated from cord blood CD34+ cells in serum-free liquid cultures containing TPO displayed 2N and 4N values, suggesting that they were immature. These results indicate that, compared with IL-3 and SCF, TPO has more potent proliferative effect on human cord blood megakaryocyte progenitors, leading to greater numbers of megakaryocyte progenitors triggered for both growth and cell division, and synergizes with SCF to enhance megakaryocyte colony growth.


Tumor-specific immunological recognition of frameshift-mutated peptides in colon cancer with microsatellite instability.
Colorectal cancers with microsatellite instability (MSI+ CRCs) caused by dysfunction of DNA mismatch repair have unique clinicopathological characteristics including good prognosis with T-cell infiltration in tumor. To identify tumor antigens that induce immune response against MSI+ CRC, SEREX (serological analysis of recombinant cDNA expression cloning) was applied. By screening a lambda phage cDNA library constructed from three MSI+ CRC cell lines with serum from a patient with MSI+ CRC with abundant T-cell infiltrates in tumor, 64 antigens were isolated. Immunogenicity of each antigen was evaluated by screening sera from patients with various cancers and from healthy individuals, and specific IgG antibodies (Abs) for 49 antigens were detected only in MSI+ CRC patients. A frameshift mutation in the repetitive G sequences (microsatellite) in the coding region of CDX2, one of the identified antigens, was found in the tumor tissue of the patient who had anti-CDX2 serum Ab. The Ab recognized both the COOH-terminal tumor-specific peptides created by the frameshift mutation and the NH(2)-terminal normal peptides of CDX2 when Western blot analysis was performed using various bacterial recombinant CDX2 proteins including the normal and altered peptides, which indicated that immune response could be raised against tumor-specific peptides generated through MSI. The anti-CDX2 Ab was detected only in the patient with the CDX2 frameshift mutation in tumor and disappeared 7 years after the curative resection, suggesting that this immune response may also be useful as a tumor marker. No altered subcellular localization and transcription ability was demonstrated in the mutated CDX2, although decreased expression was suggested in immunohistochemical analysis. Therefore, tumor-specific peptides generated by MSI may be involved in antitumor immune responses and may be useful for the development of diagnostic and therapeutic methods for patients with MSI+ CRC.


Recent advances in the management of therapy-related myelodysplastic syndromes and acute myeloid leukemia.
Therapy-related myelodysplastic syndromes and acute myelogenous leukemia (t-MDS/AML) comprise an increasingly common, poor-risk disease cohort without standard treatment options. This review describes available treatments and recent advances that may influence the clinical management of t-MDS/AML. Multiple retrospective studies have identified poor-risk cytogenetics, chemotherapy resistance, comorbidities from prior cancer and therapy, and persistence of the primary malignancy as factors that contribute to poor clinical outcomes of t-MDS/AML. Recent analyses show that t-MDS/AML can respond to standard therapy, but responses are less durable. In most cases, cure is made more likely with allogeneic stem cell transplantation. These findings suggest that improved survival may result from earlier, low-intensity nontransplant therapy, and aggressive pursuit of reduced-intensity transplant approaches in eligible individuals coupled with posttransplant relapse prevention strategies. Molecular characterization of t-MDS/AML may aid future clinical management decisions and identify targets for therapy. Data emerging from recent t-MDS/AML studies are shedding light on factors that contribute to disease biology and poor clinical outcomes. These findings can be used to develop strategies to improve the treatment and survival of patients with t-MDS/AML.


Chloride efflux inhibits single calcium channel open probability in vertebrate photoreceptors: chloride imaging and cell-attached patch-clamp recordings.
The present study uses cell-attached patch-recording techniques to study the single-channel properties of Ca2+ channels in isolated salamander photoreceptors and investigate their sensitivity to reductions in intracellular Cl-. The results show that photoreceptor Ca2+ channels possess properties similar to L-type Ca2+ channels in other preparations, including (1) enhancement of openings by the dihydropyridine agonist, (-)BayK8644; (2) suppression by a dihydropyridine antagonist, nisoldipine; (3) single-channel conductance of 22 pS with 82 mM Ba2+ as the charge carrier; (4) mean open probability of 0.1; (5) open-time distribution fit with a single exponential (tau0 = 1.1 ms) consistent with a single open state; and (6) closed time distribution fit with two exponentials (tau(c1) = 0.7 ms, tau(c2) = 25.4 ms) consistent with at least two closed states. Using a Cl- -sensitive dye to measure intracellular [Cl-], it was found that perfusion with gluconate-containing, low Cl- medium depleted intracellular [Cl-]. It was therefore possible to reduce intracellular [Cl-] by perfusion with a low Cl- solution while maintaining the extracellular channel surface in high Cl- pipette solution. Under these conditions, the single-channel conductance was unchanged, but the mean open probability fell to 0.03. This reduction can account for the 66% reduction in whole-cell Ca2+ currents produced by perfusion with low Cl- solutions. Examination of the open and closed time distributions suggests that the reduction in open probability arises from increases in closed-state dwell times. Changes in intracellular [Cl-] may thus modulate photoreceptor Ca2+ channels.


Update on Mentorship in Orthopaedic Resident Education: A Report from the American Orthopaedic Association.
Mentorship has been identified as an important element of educational and professional development for surgeons. An assessment that was conducted and reported through the American Association of Orthopaedic Surgeons (AAOS) in 2008 showed variability among U.S. residencies regarding the structure and requirements for mentorship during orthopaedic training; the assessment also demonstrated variability in residents' satisfaction with mentorship opportunities during their surgical training. An updated survey was developed and distributed via e-mail to residents attending the Resident Leadership Forum at the 2015 American Orthopaedic Association Annual Meeting to determine their views regarding the importance of mentorship, as well as their assessments of formal mentorship programs within their residencies. The updated data were compared with the prior survey results from 2008. A total of 149 (87.6%) of 170 residents responded to the survey. Of these, 34.9% (51 of 146) reported the existence of a formal mentorship program within their residency, as compared with 26.0% of residencies as stated in the 2008 report. One hundred percent of residents indicated that having a mentor during orthopaedic residency was either critical (63.7%, 93 of 146) or advantageous (36.3%, 53 of 146) to professional development as a surgeon; 74.7% (109 of 146) of residents reported currently having mentors, which appears to represent an increase from the prior report (51%, 258 of 506). However, the percentage of residents who reported being "very" satisfied (17.9%, 25 of 140) or "somewhat" satisfied (43.6%, 61 of 140) with their mentorship opportunities was almost identical to the prior report (61.9% [86 of 139] versus 61.0%, respectively). Overall, residents from programs with formal mentorship programs in place reported significantly higher satisfaction with their mentoring program/environment compared with those from programs without formal mentorship programs in place (3.98 versus 3.54, p = 0.026). Orthopaedic residents continue to overwhelmingly indicate that mentorship is an important component of residency education: 34.9% of residencies have a formal mentorship program, compared with 26.0% in the prior survey. Additionally, 74.7% of current residents reported having a mentor compared with 51% of residents in the prior study. Despite this difference, a very similar percentage of residents indicated that they were either "very" or "somewhat" satisfied with their mentorship experience. Residents from programs with formal mentorship programs reported significantly higher satisfaction with their mentorship programs compared with those without formal programs. These results support continued efforts toward improving mentorship opportunities within U.S. orthopaedic residency programs.


A multicenter study design to assess the clinical usefulness of semi-automatic measurement of flow-mediated vasodilatation of the brachial artery.
Flow-mediated vasodilatation of the brachial artery (FMD) is a marker which is related to endothelial nitric oxide bioavailability. Commercially available ultrasound machines equipped with online computer-assisted semi-automatic analysis software to measure FMD have recently become available in Japan. These devices enable more convenient examination, enhanced reproducibility of FMD measurement, and a shortened examination time. Using such devices, in the present multicenter prospective study we propose to: 1) establish standardized FMD values and determine the annual rates of FMD change in healthy subjects; 2) confirm the predictive value of FMD for future cardiovascular events in Japanese subjects; 3) evaluate the potential usefulness of a multimarker strategy, including measurements of FMD, pulse-wave velocity (PWV), ankle-brachial pressure index, biochemical markers, and proteomic biomarkers obtained by mass spectroscopic analysis to assess the prognosis of subjects with coronary artery disease; and 4) clarify the usefulness of FMD measurement to predict the rate of progression of carotid atherosclerosis, arterial stiffness and microalbuminuria in subjects with hypertension or diabetes mellitus. In total, we estimate that approximately 4000 Japanese subjects in 3 different study groups will eventually be enrolled in this prospective observational investigation. We anticipate that the present study will provide important evidence for the usefulness of FMD measurement in the risk stratification for cardiovascular disease.


Required enhancer-matrin-3 network interactions for a homeodomain transcription program.
Homeodomain proteins, described 30 years ago, exert essential roles in development as regulators of target gene expression; however, the molecular mechanisms underlying transcriptional activity of homeodomain factors remain poorly understood. Here investigation of a developmentally required POU-homeodomain transcription factor, Pit1 (also known as Pou1f1), has revealed that, unexpectedly, binding of Pit1-occupied enhancers to a nuclear matrin-3-rich network/architecture is a key event in effective activation of the Pit1-regulated enhancer/coding gene transcriptional program. Pit1 association with Satb1 (ref. 8) and β-catenin is required for this tethering event. A naturally occurring, dominant negative, point mutation in human PIT1(R271W), causing combined pituitary hormone deficiency, results in loss of Pit1 association with β-catenin and Satb1 and therefore the matrin-3-rich network, blocking Pit1-dependent enhancer/coding target gene activation. This defective activation can be rescued by artificial tethering of the mutant R271W Pit1 protein to the matrin-3 network, bypassing the pre-requisite association with β-catenin and Satb1 otherwise required. The matrin-3 network-tethered R271W Pit1 mutant, but not the untethered protein, restores Pit1-dependent activation of the enhancers and recruitment of co-activators, exemplified by p300, causing both enhancer RNA transcription and target gene activation. These studies have thus revealed an unanticipated homeodomain factor/β-catenin/Satb1-dependent localization of target gene regulatory enhancer regions to a subnuclear architectural structure that serves as an underlying mechanism by which an enhancer-bound homeodomain factor effectively activates developmental gene transcriptional programs.


Ventricular muscle-restricted targeting of the RXRalpha gene reveals a non-cell-autonomous requirement in cardiac chamber morphogenesis.
Mouse embryos lacking the retinoic acid receptor gene RXR(alpha) die in midgestation from hypoplastic development of the myocardium of the ventricular chambers and consequent cardiac failure. In this study, we address the issue of whether the RXRalpha gene is required in the cardiomyocyte lineage by generating mice that harbor a ventricular restricted deficiency in RXRalpha at the earliest stages of ventricular chamber specification. We first created a conditional ('floxed') allele of RXRalpha by flanking a required exon of the gene with loxP recombination sequences. To achieve ventricular myocardium-specific gene targeting, and to avoid potential transgenic artifacts, we employed a knock-in strategy to place cre recombinase coding sequences into the myosin light chain 2v (MLC2v) genomic locus, a gene which in the heart is expressed exclusively in ventricular cardiomyocytes at the earliest stages of ventricular specification. Crossing the MLC2v-cre allele with the floxed RXRalpha gene resulted in embryos in which approximately 80% of the ventricular cardiomyocytes lacked RXRalpha function, and yet which displayed a completely normal phenotype, without evidence of the wide spectrum of congenital heart disease phenotype seen in RXRa-/- embryos, and normal adult viability. We conclude that the RXRalpha mutant phenotype is not cell autonomous for the cardiomyocyte lineage, and suggest that RXRalpha functions in a neighboring compartment of the developing heart to generate a signal that is required for ventricular cardiomyocyte development and chamber maturation.


A perspective on interaction effects in genetic association studies.
The identification of gene-gene and gene-environment interaction in human traits and diseases is an active area of research that generates high expectation, and most often lead to high disappointment. This is partly explained by a misunderstanding of the inherent characteristics of standard regression-based interaction analyses. Here, I revisit and untangle major theoretical aspects of interaction tests in the special case of linear regression; in particular, I discuss variables coding scheme, interpretation of effect estimate, statistical power, and estimation of variance explained in regard of various hypothetical interaction patterns. Linking this components it appears first that the simplest biological interaction models-in which the magnitude of a genetic effect depends on a common exposure-are among the most difficult to identify. Second, I highlight the demerit of the current strategy to evaluate the contribution of interaction effects to the variance of quantitative outcomes and argue for the use of new approaches to overcome this issue. Finally, I explore the advantages and limitations of multivariate interaction models, when testing for interaction between multiple SNPs and/or multiple exposures, over univariate approaches. Together, these new insights can be leveraged for future method development and to improve our understanding of the genetic architecture of multifactorial traits.


Physical-Perceptual Correspondence for Dynamic Thermal Stimulation.
Thermal displays have been applied in various haptic applications, from material simulation to interpersonal communication; however, there is insufficient knowledge about the temporal processing in human thermal sense to provide a knowledge basis for thermal display design. In this study, we investigated the physical-perceptual correspondence for dynamic thermal stimulation to shed a light on the temporal processing of human thermal sense. In the experiments, participants reported subjective timings of the temperature onset and temperature peak of continuous temperature changes applied to the thenar eminence. We found that the physical-perceptual correspondence was not consistent for warm and cold stimulations. For warm stimulation, the subjective experience always came after the corresponding physical event. On the other hand, for cold stimulation, while the subjective onset always lagged the physical onset, the subjective temperature peak preceded the physical temperature peak. We analyzed these results in the framework of linear systems theory. The results suggest that the senses of warmth and cold have distinct temporal filtering properties, with the sense of cold being more transient than the sense of warmth. These findings advance our knowledge regarding temporal processing in human thermal sense and serve as a basis for thermal display design.


Neurotoxicity of sodium fluoride in rats.
Fluoride (F) is known to affect mineralizing tissues, but effects upon the developing brain have not been previously considered. This study in Sprague-Dawley rats compares behavior, body weight, plasma and brain F levels after sodium fluoride (NaF) exposures during late gestation, at weaning or in adults. For prenatal exposures, dams received injections (SC) of 0.13 mg/kg NaF or saline on gestational days 14-18 or 17-19. Weanlings received drinking water containing 0, 75, 100, or 125 ppm F for 6 or 20 weeks, and 3 month-old adults received water containing 100 ppm F for 6 weeks. Behavior was tested in a computer pattern recognition system that classified acts in a novel environment and quantified act initiations, total times and time structures. Fluoride exposures caused sex- and dose-specific behavioral deficits with a common pattern. Males were most sensitive to prenatal day 17-19 exposure, whereas females were more sensitive to weanling and adult exposures. After fluoride ingestion, the severity of the effect on behavior increased directly with plasma F levels and F concentrations in specific brain regions. Such association is important considering that plasma levels in this rat model (0.059 to 0.640 ppm F) are similar to those reported in humans exposed to high levels of fluoride.


Antidepressants, Depression, and Venous Thromboembolism Risk: Large Prospective Study of UK Women.
Some investigators have reported an excess risk of venous thromboembolism (VTE) associated with depression and with use of antidepressant drugs. We explored these associations in a large prospective study of UK women. The Million Women Study recruited 1.3 million women through the National Health Service Breast Screening Programme in England and Scotland. Three years after recruitment, women were sent a second questionnaire that enquired about depression and regular use of medications in the previous 4 weeks. The present analysis included those who responded and did not have prior VTE, cancer, or recent surgery. Follow-up for VTE was through linkage to routinely collected National Health Service statistics. Cox regression analyses yielded adjusted hazard ratios and 95% CIs. A total of 734 092 women (mean age 59.9 years) were included in the analysis; 6.9% reported use of antidepressants, 2.7% reported use of other psychotropic drugs, and 1.8% reported being treated for depression or anxiety but not use of psychotropic drugs. During follow-up for an average of 7.3 years, 3922 women were hospitalized for and/or died from VTE. Women who reported antidepressant use had a significantly higher risk of VTE than women who reported neither depression nor use of psychotropic drugs (hazard ratio, 1.39; 95% CI, 1.23-1.56). VTE risk was not significantly increased in women who reported being treated for depression or anxiety but no use of antidepressants or other psychotropic drugs (hazard ratio, 1.19; 95% CI, 0.95-1.49). Use of antidepressants is common in UK women and is associated with an increased risk of VTE.


Urine catalytic iron and neutrophil gelatinase-associated lipocalin as companion early markers of acute kidney injury after cardiac surgery: a prospective pilot study.
Open heart surgery with cardiopulmonary bypass is recognized as a common cause of acute kidney injury (AKI). The conventional biomarker creatinine is not sensitive enough to detect AKI until a significant decline in renal filtration has occurred. Urine neutrophil gelatinase-associated lipocalin (NGAL), part of an acute response to the release of tissue iron from cells, is an early biomarker and a predictor of AKI in a variety of clinical settings. We sought to evaluate the relationship between urine catalytic iron (unbound iron) and NGAL over the course of AKI due to cardiac surgery. FOURTEEN PATIENTS WHO UNDERWENT OPEN HEART SURGERY HAD THE FOLLOWING MEASURED: serum creatinine (0, 12, 24, 48 and 72 h postoperatively), urine NGAL and urine catalytic iron (0, 8, 24 and 48 h postoperatively). Urine NGAL and urine catalytic iron were quantified by immunoassay and bleomycin-detectable iron assay, respectively. AKI was defined by the Acute Kidney Injury Network (AKIN) criteria. Urine catalytic iron increased significantly (p < 0.05) within 8 h and peaked at 24 h postoperatively in patients who developed AKI (n = 8, baseline 101.96 ± 177.48, peak 226.35 ± 238.23 nmol/l, p = 0.006), but not in non-AKI patients (n = 6, baseline 131.08 ± 116.21, peak 163.99 ± 109.62 nmol/l, p = 0.380). Urine NGAL levels also peaked at 24 h with significant increase observed only in AKI patients: AKI - baseline 34.88 ± 26.47, peak 65.50 ± 27.03 ng/ml, p = 0.043; non-AKI - baseline 59.33 ± 31.72, peak 71.00 ± 31.76 ng/ml, p = 0.100. The correlation between baseline levels of urine catalytic iron and NGAL and peak levels of urine catalytic iron and NGAL was r = 0.86, p < 0.0001. Urine catalytic iron appears to rise and fall in concert with NGAL in patients undergoing cardiac surgery and may be indicative of early AKI. Future research into the role that catalytic iron plays in acute organ injury syndromes and its potential diagnostic and therapeutic implications is warranted.


Systematic Review of Early Phase Pediatric Clinical Pharmacology Trials.
Children have generally been excluded from early-stage clinical trials owing to safety concerns based in social expectations and not data. However, the repositioning of adult therapeutics for pediatric use and the increase in the development of therapies for pediatric only conditions require the participation of children in phase 1-2 trials. Therefore, the aim of this article is to systematically review the history and current state of early phase pediatric clinical pharmacology trials in order to understand safety concerns, trends, and challenges in pediatric trials. This review analyzed the nature of early phase pediatric clinical trials conducted for nononcology conditions through a systematic search that was performed for pediatric non-oncologic phase 1 or phase 1-2 drug and vaccine studies in MEDLINE. The data show that the number of early phase pediatric clinical trials is still small relative to adults but has been on the rise in the past decade with relatively few serious adverse effects observed. The widespread concerns about children's safety when they participate in early phase clinical trials seem disproportionate, based on our findings. The data confirm that these studies can be conducted safely, and that their results can contribute significantly to pediatric pharmacotherapy.


The Power and the Promise of Cell Reprogramming: Personalized Autologous Body Organ and Cell Transplantation.
Reprogramming somatic cells to induced pluripotent stem cells (iPSCs) or direct reprogramming to desired cell types are powerful and new in vitro methods for the study of human disease, cell replacement therapy, and drug development. Both methods to reprogram cells are unconstrained by the ethical and social questions raised by embryonic stem cells. iPSC technology promises to enable personalized autologous cell therapy and has the potential to revolutionize cell replacement therapy and regenerative medicine. Potential applications of iPSC technology are rapidly increasing in ambition from discrete cell replacement applications to the iPSC assisted bioengineering of body organs for personalized autologous body organ transplant. Recent work has demonstrated that the generation of organs from iPSCs is a future possibility. The development of embryonic-like organ structures bioengineered from iPSCs has been achieved, such as an early brain structure (cerebral organoids), bone, optic vesicle-like structures (eye), cardiac muscle tissue (heart), primitive pancreas islet cells, a tooth-like structure (teeth), and functional liver buds (liver). Thus, iPSC technology offers, in the future, the powerful and unique possibility to make body organs for transplantation removing the need for organ donation and immune suppressing drugs. Whilst it is clear that iPSCs are rapidly becoming the lead cell type for research into cell replacement therapy and body organ transplantation strategies in humans, it is not known whether (1) such transplants will stimulate host immune responses; and (2) whether this technology will be capable of the bioengineering of a complete and fully functional human organ. This review will not focus on reprogramming to iPSCs, of which a plethora of reviews can be found, but instead focus on the latest developments in direct reprogramming of cells, the bioengineering of body organs from iPSCs, and an analysis of the immune response induced by iPSC-derived cells and tissues.


Immunopathology of pineal glands from horses with uveitis.
Pinealitis accompanying uveitis is well established in laboratory models of experimental autoimmune uveoretinitis. In naturally occurring uveitis, pinealitis has been demonstrated in the pineal gland from a mare with active uveitis and is suspected in some human uveitides. We have evaluated pineal glands from horses with various stages of uveitis for signs of immunopathology accompanying spontaneous uveitis. Pineal glands from 10 horses with uveitis and from 13 horses without uveitis were evaluated for histochemical (H&E, collagen) and immunohistochemical (MHC class II antigen expression, infiltration of T and B lymphocytes, and glial fibrillary acidic protein (GFAP) and vimentin upregulation) evidence of inflammation. Septal areas of pineal glands from horses with uveitis had clusters of MHC class II antigen-expressing cells, T lymphocytes, and enhanced collagen deposition. These changes were not as readily observed in pineal glands from horses without uveitis. B lymphocytes were detected only in the pineal gland from the one mare with active uveitis in which T and B lymphocytes were organized into follicles. No differences in GFAP or vimentin immunoreactivity were noted in pineal glands from horses with or without uveitis. These pineal gland changes suggest that the pinealitis associated with equine uveitis is transient just as the uveitis of these horses is recurrent. Study of pineal glands from horses with clinically documented uveitis allows demonstration of subtle pineal changes associated with natural uveitis. Similar changes would be difficult to document in human patient populations.


Mass Spectrometry Measurements of Neuropeptides: From Identification to Quantitation.
Neuropeptides (NPs), a unique class of neuronal signaling molecules, participate in a variety of physiological processes and diseases. Quantitative measurements of NPs provide valuable information regarding how these molecules are differentially regulated in a multitude of neurological, metabolic, and mental disorders. Mass spectrometry (MS) has evolved to become a powerful technique for measuring trace levels of NPs in complex biological tissues and individual cells using both targeted and exploratory approaches. There are inherent challenges to measuring NPs, including their wide endogenous concentration range, transport and postmortem degradation, complex sample matrices, and statistical processing of MS data required for accurate NP quantitation. This review highlights techniques developed to address these challenges and presents an overview of quantitative MS-based measurement approaches for NPs, including the incorporation of separation methods for high-throughput analysis, MS imaging for spatial measurements, and methods for NP quantitation in single neurons.


Adjunctive left anterior line ablation induced left atrial dysfunction and dyssynchrony in atrial fibrillation ablation.
We evaluated the effects of adjunctive left anterior line (LAL) ablation on LA dyssynchrony and function using real-time three-dimensional echocardiography (3DE) in connection with thromboembolic complications and tachyarrhythmia recurrence in patients with persistent atrial fibrillation (AF). We randomly and prospectively assigned consecutive persistent AF patients to the LAL (n = 52, 65 ± 7 years) and control groups (n = 50, 64 ± 10 years). In the LAL group, extensive encircling pulmonary vein isolation (EEPVI), roof line ablation, and LAL ablation regardless of the extent of the low-voltage area (LVA) were performed. The control group underwent EEPVI and roof line ablation. After ablation, 3DE demonstrated LA dyssynchrony in 23 (46%) and 4 patients (8%, P < 0.001) of the LAL and control groups, respectively. Baseline LA LVAs were relatively small in most patients and there were no significant differences in extent of LVA between control and LAL groups or between patients with and without dyssynchrony. During the follow-up periods (771 ± 121 days), patients with LA dyssynchrony in the LAL group did not show significant differences in symptomatic thromboembolic events (0%) and atrial tachyarrhythmia recurrence (39%) from patients without LA dyssynchrony in the LAL (0% and 30%) and control groups (0% and 32%, respectively). LA ejection fraction and active emptying fraction were lower by 9% on average in the LAL group than in the control group (P < 0.0001). Similarly, in the LAL group, LA ejection fraction, active emptying fraction, and expansion index were significantly lower by approximately 7%, 8%, and 15%, respectively, in LA with dyssynchrony than those in LA without dyssynchrony. In conclusion, LA dyssynchrony and LA hypofunction were induced by LAL ablation in patients with persistent AF and relatively mild LVA. LAL ablation with or without LA dyssynchrony is thought not to affect thromboembolic complications or atrial tachyarrhythmia recurrence.


Exploring health stakeholders' perceptions on moving towards comprehensive primary health care to address childhood malnutrition in Iran: a qualitative study.
Due to the multifaceted aspect of child malnutrition, a comprehensive approach, taking social factors into account, has been frequently recommended in health literature. The Alma-Ata declaration explicitly outlined comprehensive primary health care as an approach that addresses the social, economic and political causes of poor health and nutrition. Iran as a signatory country to the Alma Ata Declaration has established primary health care since 1979 with significant progress on many health indicators during the last three decades. However, the primary health care system is still challenged to reduce inequity in conditions such as child malnutrition which trace back to social factors. This study aimed to explore the perceptions of the Iranian health stakeholders with respect to the Iranian primary health care performance and actions to move towards a comprehensive approach in addressing childhood malnutrition. Health stakeholders are defined as those who affect or can be affected by health system, for example health policy-makers, health providers or health service recipients. Stakeholder analysis approach was undertaken using a qualitative research method. Different levels of stakeholders, including health policy-makers, health providers and community members were interviewed as either individuals or focus groups. Qualitative content analysis was used to interpret and compare/contrast the viewpoints of the study participants. The results demonstrated that fundamental differences exist in the perceptions of different health stakeholders in the understanding of comprehensive notion and action. Health policy-makers mainly believed in the need for a secure health management environment and the necessity for a whole of the government approach to enhance collaborative action. Community health workers, on the other hand, indicated that staff motivation, advocacy and involvement are the main challenges need to be addressed. Turning to community stakeholders, greater emphasis has been placed on community capabilities, informal link with other social sectors based on trust and local initiatives. This research provided a picture of the differences in the perceptions and values of different stakeholders with respect to primary health care concepts. The study suggests that a top-down approach, which still exists among health policy-makers, is a key obstacle that delays, and possibly worse, undermines the implementation of the comprehensive strategy codified by the Alma-Ata Declaration. A need to revitalize primary health care to use its full potential and to combine top-down and bottom-up approaches by narrowing the gap between perceptions of policy makers and those who provide and receive health-related services is crucial.


Risk of Child Maltreatment in Chinese Teenage and Young Mothers With Rapid Repeat Pregnancy: The Moderating Role of Family Cohesion and Support From Friends.
Rapid repeat pregnancy (RRP) often occurs in teenage and young mothers. Mothers with a history of RRP are more likely to experience high stress increasing their risk of child maltreatment. Despite these challenges, some mothers can continue to cope adaptively. Social support may play a role in empowering these mothers to overcome the childbearing difficulties. Although the protective effects of social support are well recognized, there has been little evidence on the relative importance of sources of support. For example, whether support from family and friends is equally important in relieving parenting stress remains unanswered. RRP, a social phenomenon encompassing various adverse living and parenting issues, provides an ideal research context to investigate the role of family and friends in preventing child maltreatment. This study examined whether family cohesion and friends' support moderated the association between RRP and child maltreatment in young mothers. We recruited 392 Chinese teenage and young mothers from a population-based integrated young mothers supporting program in Hong Kong. Questionnaires on pregnancy history, family cohesion, social support, and risk of child maltreatment were administered. Moderation analysis was conducted to examine the effect of RRP on child maltreatment as a function of family cohesion or friend support. Results showed that RRP was associated with a higher risk of child neglect (adjusted odds ratio [aOR] = 1.72, p < .05) and physical maltreatment (aOR = 1.91, p < .01), and that family cohesion was more important than friend support in mitigating the risk of child maltreatment for mothers with a history of RRP. Our findings suggest that interventions for young mothers, particularly those with a history of RRP, should be family-based so the whole family can be empowered to tackle the childrearing burden.


The Akt activation inhibitor TCN-P inhibits Akt phosphorylation by binding to the PH domain of Akt and blocking its recruitment to the plasma membrane.
Persistently hyperphosphorylated Akt contributes to human oncogenesis and resistance to therapy. Triciribine (TCN) phosphate (TCN-P), the active metabolite of the Akt phosphorylation inhibitor TCN, is in clinical trials, but the mechanism by which TCN-P inhibits Akt phosphorylation is unknown. Here we show that in vitro, TCN-P inhibits neither Akt activity nor the phosphorylation of Akt S473 and T308 by mammalian target of rapamycin or phosphoinositide-dependent kinase 1. However, in intact cells, TCN inhibits EGF-stimulated Akt recruitment to the plasma membrane and phosphorylation of Akt. Surface plasmon resonance shows that TCN, but not TCN, binds Akt-derived pleckstrin homology (PH) domain (K(D): 690 nM). Furthermore, nuclear magnetic resonance spectroscopy shows that TCN-P, but not TCN, binds to the PH domain in the vicinity of the PIP3-binding pocket. Finally, constitutively active Akt mutants, Akt1-T308D/S473D and myr-Akt1, but not the transforming mutant Akt1-E17K, are resistant to TCN and rescue from its inhibition of proliferation and induction of apoptosis. Thus, the results of our studies indicate that TCN-P binds to the PH domain of Akt and blocks its recruitment to the membrane, and that the subsequent inhibition of Akt phosphorylation contributes to TCN-P antiproliferative and proapoptotic activities, suggesting that this drug may be beneficial to patients whose tumors express persistently phosphorylated Akt.


Safety evaluation of the food enzyme glucan 1,4-alpha-glucosidase from the genetically modified Trichoderma reesei strain DP-Nzh38.
The food enzyme glucoamylase (4-α-D-glucan glucohydrolase; EC 18.104.22.168) is produced with the genetically modified Trichoderma reesei strain DP-Nzh38 by Danisco US Inc. The genetic modifications do not give rise to safety concerns. The food enzyme is free from viable cells of the production organism and its DNA. The food enzyme is intended to be used in distilled alcohol production, starch processing for glucose syrup production, baking and brewing processes. Since residual amounts of total organic solids (TOS) are removed by distillation and by purification steps applied during the production of glucose syrups, consequently, dietary exposure was not calculated for these uses. Based on the maximum use levels recommended for baking and brewing processes and individual data from the EFSA Comprehensive European Food Consumption Database, dietary exposure to the food enzyme-TOS was estimated to be up to 5.8749 mg TOS/kg body weight per day. The toxicity studies were carried out with another glucoamylase from T. reesei (strain DP-Nzh49) considered by the Panel as a suitable substitute. Genotoxicity tests did not raise safety concern. The systemic toxicity was assessed by means of a repeated dose 90-day oral toxicity study in rats. The Panel identified no observed adverse effect level (NOAEL) at the highest dose tested of 1,149 mg TOS/kg body weight (bw) per day resulting in a margin of exposure of at least 195. Similarity of the amino acid sequence to those of known allergens was searched for and one match was found. The Panel considered that, under the intended conditions of use, the risk of allergic sensitisation and elicitation reactions by dietary exposure cannot be excluded, but the likelihood for this to occur is considered to be low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns under the intended conditions of use.


From the gut to the strut: where inflammation reigns, bone abstains.
In this issue of the JCI, Li et al. show that germ-free mice, when chemically castrated, do not lose bone - a finding that unequivocally establishes a role of gut microbiota in mediating hypogonadal bone loss. Additionally and not unexpectedly, probiotics reversed hypogonadal osteopenia in sex steroid-deficient mice by preventing the disruption of gut barrier function and dampening cytokine-induced inflammation. The authors propose that TNFα is a key mediator; however, it is very likely that other molecules - including IL-1, IL-6, IL-17, RANKL, OPG, and CCL2 - modulate probiotic action. The results of this study highlight the potential for repurposing probiotics for the therapy of osteoporosis. Future placebo-controlled clinical trials will be required to establish safety and efficacy of probiotics in reducing fracture risk in people.


Health care purchasing in Kenya: Experiences of health care providers with capitation and fee-for-service provider payment mechanisms.
Provider payment mechanisms (PPMs) play a critical role in universal health coverage due to the incentives they create for health care providers to deliver needed services, quality, and efficiency. We set out to explore public, private, and faith-based providers' experiences with capitation and fee-for-service in Kenya and identified attributes of PPMs that providers considered important. We conducted a qualitative study in two counties in Kenya. Data were collected using semistructured interviews with 29 management team members in six health providers accredited by the National Hospital Insurance Fund (NHIF). Capitation and fee-for-service payments from the NHIF and private insurers were reported as good revenue sources as they contributed to providers' overall income. The expected fee-for-service payment amounts from NHIF and private insurers were predictable while capitation funds from NHIF were not because providers did not have information on the number of enrolees in their capitation pool. Moreover, capitation payment rates were perceived as inadequate. Capitation and fee-for-service payments from NHIF and private insurers were disbursed late. Finally, public providers had lost their autonomy to access and utilise capitation and fee-for-service payments from the NHIF. Through their experiences, health care providers revealed characteristics of PPMs that they considered important.


GENCODE: producing a reference annotation for ENCODE.
The GENCODE consortium was formed to identify and map all protein-coding genes within the ENCODE regions. This was achieved by a combination of initial manual annotation by the HAVANA team, experimental validation by the GENCODE consortium and a refinement of the annotation based on these experimental results. The GENCODE gene features are divided into eight different categories of which only the first two (known and novel coding sequence) are confidently predicted to be protein-coding genes. 5' rapid amplification of cDNA ends (RACE) and RT-PCR were used to experimentally verify the initial annotation. Of the 420 coding loci tested, 229 RACE products have been sequenced. They supported 5' extensions of 30 loci and new splice variants in 50 loci. In addition, 46 loci without evidence for a coding sequence were validated, consisting of 31 novel and 15 putative transcripts. We assessed the comprehensiveness of the GENCODE annotation by attempting to validate all the predicted exon boundaries outside the GENCODE annotation. Out of 1,215 tested in a subset of the ENCODE regions, 14 novel exon pairs were validated, only two of them in intergenic regions. In total, 487 loci, of which 434 are coding, have been annotated as part of the GENCODE reference set available from the UCSC browser. Comparison of GENCODE annotation with RefSeq and ENSEMBL show only 40% of GENCODE exons are contained within the two sets, which is a reflection of the high number of alternative splice forms with unique exons annotated. Over 50% of coding loci have been experimentally verified by 5' RACE for EGASP and the GENCODE collaboration is continuing to refine its annotation of 1% human genome with the aid of experimental validation.


Tectonics, climate and the diversification of the tropical African terrestrial flora and fauna.
Tropical Africa is home to an astonishing biodiversity occurring in a variety of ecosystems. Past climatic change and geological events have impacted the evolution and diversification of this biodiversity. During the last two decades, around 90 dated molecular phylogenies of different clades across animals and plants have been published leading to an increased understanding of the diversification and speciation processes generating tropical African biodiversity. In parallel, extended geological and palaeoclimatic records together with detailed numerical simulations have refined our understanding of past geological and climatic changes in Africa. To date, these important advances have not been reviewed within a common framework. Here, we critically review and synthesize African climate, tectonics and terrestrial biodiversity evolution throughout the Cenozoic to the mid-Pleistocene, drawing on recent advances in Earth and life sciences. We first review six major geo-climatic periods defining tropical African biodiversity diversification by synthesizing 89 dated molecular phylogeny studies. Two major geo-climatic factors impacting the diversification of the sub-Saharan biota are highlighted. First, Africa underwent numerous climatic fluctuations at ancient and more recent timescales, with tectonic, greenhouse gas, and orbital forcing stimulating diversification. Second, increased aridification since the Late Eocene led to important extinction events, but also provided unique diversification opportunities shaping the current tropical African biodiversity landscape. We then review diversification studies of tropical terrestrial animal and plant clades and discuss three major models of speciation: (i) geographic speciation via vicariance (allopatry); (ii) ecological speciation impacted by climate and geological changes, and (iii) genomic speciation via genome duplication. Geographic speciation has been the most widely documented to date and is a common speciation model across tropical Africa. We conclude with four important challenges faced by tropical African biodiversity research: (i) to increase knowledge by gathering basic and fundamental biodiversity information; (ii) to improve modelling of African geophysical evolution throughout the Cenozoic via better constraints and downscaling approaches; (iii) to increase the precision of phylogenetic reconstruction and molecular dating of tropical African clades by using next generation sequencing approaches together with better fossil calibrations; (iv) finally, as done here, to integrate data better from Earth and life sciences by focusing on the interdisciplinary study of the evolution of tropical African biodiversity in a wider geodiversity context.


Akt signals through the mammalian target of rapamycin pathway to regulate CNS myelination.
Mammalian target of rapamycin (mTOR), a well known Akt substrate, regulates multiple cellular functions including cell growth and protein synthesis. The current study identifies a novel role of the Akt/mTOR pathway as a regulator of CNS myelination. Previously, we showed that overexpressing constitutively active Akt in oligodendrocytes in a transgenic mouse model induces enhanced CNS myelination, with no changes in the proliferation or survival of oligodendrocyte progenitor or mature cells. The present study focused on the signaling mechanisms regulating this hypermyelination induced by Akt. Activation of mTOR and its downstream substrates (p70S6 kinase and S6 ribosomal protein) was observed in Akt-overexpressing oligodendrocytes. When mTOR signaling was inhibited chronically in vivo with rapamycin starting at 6 weeks of age, the observed hypermyelination was reduced to approximately the amount of myelin seen in wild-type mice. mTOR inhibition had little impact on wild-type myelination between 6 and 12 weeks of age, suggesting that, in normal adults, myelination is relatively complete and is no longer regulated by mTOR signaling. However, when mTOR was chronically inhibited in young adult wild-type mice, myelination was reduced. These results suggest that, during active myelination, the major Akt signal regulating CNS myelination is the mTOR pathway.


Learning From England's Best Practice Tariff: Process Measure Pay-for-Performance Can Improve Hip Fracture Outcomes.
The objective of this study was to evaluate England's Best Practice Tariff (BPT) and consider potential implications for Medicare patients should the US adopt a similar plan. Since the beginning of the Affordable Care Act, Medicare has renewed efforts to improve the outcomes of older adults through introduction of an expanding set of alternative-payment models. Among trauma patients, recommended arrangements met with mixed success given concerns about the heterogeneous nature of trauma patients and resulting outcome variation. A novel approach taken for hip fractures in England could offer a viable alternative. Linear regression, interrupted time-series, difference-in-difference, and counterfactual models of 2000 to 2016 Medicare (US), HES-APC (England) death certificate-linked claims (≥65 years) were used to: track US hip fracture trends, look at changes in English hip fracture trends before-and-after BPT implementation, compare changes in US-versus-English mortality, and estimate total/theoretical lives saved. A total of 806,036 English and 3,221,109 US hospitalizations were included. After BPT implementation, England's 30-day mortality decreased by 2.6 percentage-points (95%CI: 1.7-3.5) from a baseline of 9.9% (relative reduction 26.3%). 90- and 365-day mortality decreased by 5.6 and 5.4 percentage-points. 30/90/365-day readmissions also declined with a concurrent shortening of hospital length-of-stay. From 2000 to 2016, US outcomes were stagnant (P > 0.05), resulting in an inversion of the countries' mortality and >38,000 potential annual US lives saved. Process measure pay-for-performance led to significant improvements in English hip fracture outcomes. As efforts to improve US older adult health continue to increase, there are important lessons to be learned from a successful initiative like the BPT.


Noncontact quantitative biomechanical characterization of cardiac muscle using shear wave imaging optical coherence tomography.
We report on a quantitative optical elastographic method based on shear wave imaging optical coherence tomography (SWI-OCT) for biomechanical characterization of cardiac muscle through noncontact elasticity measurement. The SWI-OCT system employs a focused air-puff device for localized loading of the cardiac muscle and utilizes phase-sensitive OCT to monitor the induced tissue deformation. Phase information from the optical interferometry is used to reconstruct 2-D depth-resolved shear wave propagation inside the muscle tissue. Cross-correlation of the displacement profiles at various spatial locations in the propagation direction is applied to measure the group velocity of the shear waves, based on which the Young's modulus of tissue is quantified. The quantitative feature and measurement accuracy of this method is demonstrated from the experiments on tissue-mimicking phantoms with the verification using uniaxial compression test. The experiments are performed on ex vivo cardiac muscle tissue from mice with normal and genetically altered myocardium. Our results indicate this optical elastographic technique is useful as a noncontact tool to assist the cardiac muscle studies.


Web-based database for the management of tissue specimens in a transregional histological research facility.
In the setting of a histological research core facility sample tracking and project specific archiving of tissue specimens and communication of related data is of central importance. Over a 24-month period 10 laboratories from two transregional research centers submitted in excess of 3000 tissue samples for histological processing and evaluation to our core facility. A web based database was set up to overcome the logistical problem of managing samples with inconsistent, duplicate and missing labels and to allow for efficient sample tracking, archiving and communication with the collaborating research laboratories. The database allows the users to remotely generate unique sample identifiers and enter sample annotation prior to sample processing. Furthermore the database facilitates communication about experimental set-up results and media files such as histological images. Our newly constructed web based portal is an important tool for the management of research samples of our histological core facility and facilitates significantly interdisciplinary and transregional research. It is freely available for scientific use.


Venezuelan encephalitis virus infection in neotropical bats. II. Experimental infections.
Eighty-nine Neotropical bats of five species were inoculated subcutaneously with epizootic or enzootic strains of Venezuelan encephalitis (VE) virus. Viremia was detected in 92.5% of all bats, but no illness attributable to virus infection was observed. Detectable viremias averaged slightly over 4 days in Artibeus jamaicensis and A. lituratus, and 2.8 days in Phyllostomus discolor, and maximal viremia titers in these three species averaged 6.9, 6.6, and 4.6 log10 SMicLD50 per ml of blood, respectively. In general Artibeus developed and maintained detectable levels of both hemagglutination-inhibition (HI) and neutralizing antibody for as long as tested (up to 506 days), although HI antibody to enzootic VE virus strains disappeared in some A. lituratus. The detectable antibody response of P. discolor was slower and of lower magnitude and shorter duration than that of Artibeus, although individual P. discolor which had lost detectable HI and N antibody resisted challenge. Vertical passage of antibody was observed in three A. lituratus offspring. Artibeus jamaicensis were found to be only slightly less susceptible to VE virus infection than a U.S. subspecies of the cotton rat (Sigmodon hispidus). No virus was recovered by mouse inoculation of organ pools of bats killed as early as 2 days and as late as 299 days after the last day of detectable viremia.


Impact of body mass and body composition on circulating levels of natriuretic peptides: results from the Dallas Heart Study.
The association between higher body mass index (BMI) and lower B-type natriuretic peptide (BNP) level is thought to be mediated by expression of the natriuretic peptide clearance receptor (NPR-C) in adipose tissue. To explore this association, we tested 2 hypotheses: (1) that N-terminal (NT)-proBNP, which is not believed to bind NPR-C, would not be associated with BMI and (2) that lower BNP would be more closely associated with fat mass than with lean mass. Measurements of BNP, NT-proBNP, and body composition by direct dual energy x-ray absorptiometry (DEXA) were performed in 2707 subjects from the Dallas Heart Study. The associations between obesity and low BNP (<4 ng/L) or low NT-proBNP (lowest sex-specific quartile) were evaluated with multivariable logistic regression models stratified by sex and adjusted for age, race/ethnicity, hypertension, left ventricular mass, and end-diastolic volume. Higher BMI was independently associated with lower BNP and NT-proBNP (all P<0.001). When BMI was replaced with both DEXA-derived lean and fat mass, greater lean mass, but not fat mass, was associated with low BNP and NT-proBNP levels. In a large, population-based cohort, we confirm the previously described association between higher BMI and lower BNP and demonstrate a similar inverse association between BMI and NT-proBNP. Interestingly, both BNP and NT-proBNP are more closely associated with lean mass than with fat mass. These findings do not support the hypothesis that the lower BNP levels seen in obesity are driven by enhanced BNP clearance mediated via NPR-C.


A missing piece of the Papio puzzle: Gorongosa baboon phenostructure and intrageneric relationships.
Most authors recognize six baboon species: hamadryas (Papio hamadryas), Guinea (Papio papio), olive (Papio anubis), yellow (Papio cynocephalus), chacma (Papio ursinus), and Kinda (Papio kindae). However, there is still debate regarding the taxonomic status, phylogenetic relationships, and the amount of gene flow occurring between species. Here, we present ongoing research on baboon morphological diversity in Gorongosa National Park (GNP), located in central Mozambique, south of the Zambezi River, at the southern end of the East African Rift System. The park exhibits outstanding ecological diversity and hosts more than 200 baboon troops. Gorongosa National Park baboons have previously been classified as chacma baboons (P. ursinus). In accordance with this, two mtDNA samples from the park have been placed in the same mtDNA clade as the northern chacma baboons. However, GNP baboons exhibit morphological features common in yellow baboons (e.g., yellow fur color), suggesting that parapatric gene flow between chacma and yellow baboons might have occurred in the past or could be ongoing. We investigated the phenostructure of the Gorongosa baboons using two approaches: 1) description of external phenotypic features, such as coloration and body size, and 2) 3D geometric morphometric analysis of 43 craniofacial landmarks on 11 specimens from Gorongosa compared to a pan-African sample of 352 baboons. The results show that Gorongosa baboons exhibit a mosaic of features shared with southern P. cynocephalus and P. ursinus griseipes. The GNP baboon phenotype fits within a geographic clinal pattern of replacing allotaxa. We put forward the hypothesis of either past and/or ongoing hybridization between the gray-footed chacma and southern yellow baboons in Gorongosa or an isolation-by-distance scenario in which the GNP baboons are geographically and morphologically intermediate. These two scenarios are not mutually exclusive. We highlight the potential of baboons as a useful model to understand speciation and hybridization in early human evolution.


Averting biodiversity collapse in tropical forest protected areas.
The rapid disruption of tropical forests probably imperils global biodiversity more than any other contemporary phenomenon. With deforestation advancing quickly, protected areas are increasingly becoming final refuges for threatened species and natural ecosystem processes. However, many protected areas in the tropics are themselves vulnerable to human encroachment and other environmental stresses. As pressures mount, it is vital to know whether existing reserves can sustain their biodiversity. A critical constraint in addressing this question has been that data describing a broad array of biodiversity groups have been unavailable for a sufficiently large and representative sample of reserves. Here we present a uniquely comprehensive data set on changes over the past 20 to 30 years in 31 functional groups of species and 21 potential drivers of environmental change, for 60 protected areas stratified across the world's major tropical regions. Our analysis reveals great variation in reserve 'health': about half of all reserves have been effective or performed passably, but the rest are experiencing an erosion of biodiversity that is often alarmingly widespread taxonomically and functionally. Habitat disruption, hunting and forest-product exploitation were the strongest predictors of declining reserve health. Crucially, environmental changes immediately outside reserves seemed nearly as important as those inside in determining their ecological fate, with changes inside reserves strongly mirroring those occurring around them. These findings suggest that tropical protected areas are often intimately linked ecologically to their surrounding habitats, and that a failure to stem broad-scale loss and degradation of such habitats could sharply increase the likelihood of serious biodiversity declines.


The effect of age and cataract surgery on macular pigment optic density: a cross-sectional, comparative study.
To analyze the effects of age, cataract surgery and postoperative period on macular pigment optical density (MPOD). The study included cases referred to Ankara University Department of Ophthalmology, between April and June 2012, who had a transparent natural lens or had undergone cataract surgery at least a year ago with their best corrected visual acuity of ≥ 0.5 based on Snellen chart. Presence of an ocular disease that might affect lens, retina and optic nerve (cataract, macular degeneration, diabetic retinopathy, glaucoma etc.), cataract surgery within the previous year, light-colored iris, smoking and use of micronutrition supplementation were determined as exclusion criteria. After detailed opthalmologic examination of all patients, they were divided into three groups based on their age and their lens status as: group 1, patients < 50 years of age having a clear lens; group 2, patients > 50 years of age having a clear lens; and group 3, patients > 50 years of age who had cataract surgery. Age, gender, and postoperative period of the patients as well as the MPOD values of the eyes measured with heterochromatic flicker photometric (HFP) method (MacularMetricsTM) were included in the analysis. Sixty-eight eyes of 37 cases with a mean age of 53.4 ± 15.3 years were enrolled in the study. Group 1 included 20 eyes of 10 cases (mean age 29.4 ± 9.5); group 2 included 32 eyes of 16 cases (mean age 60.3 ± 6.8); and group 3 included 16 eyes of 11 cases (mean age 65.2 ± 9.7). The mean macular pigment optical density value of all cases was 0.511 ± 0.192 log unit, while the mean MPOD values of groups 1, 2 and 3 were 0.570 ± 170, 0.528 ± 203 and 0.400 ± 180 log units, respectively. The mean MPOD values of the patients with clear lens aged < 50 and aged > 50 years did not reveal a statisticially significant difference (p = 1). However, the mean MPOD value of the cataract surgery group (group 3) was found to be statistically significantly lower than the group 1 and group 2 (p = 0.022, p = 0.039, respectively). The correlations between MPOD values and postoperative periods of the patients in group 3 showed that a decrease in MPOD values in parallel with duration of the postoperative period and this negative correlation was found to be statistically significant (r: -0.66, p = 0.005). Our study has demonstrated that a significant correlation does not exist between age of the patients and MPOD values. MPOD values were lower than age-matched patients who had undergone cataract surgery and finally an inverse correlation existed between duration of the postoperative period after cataract surgery and MPOD values.


Association between serum and adipose tissue resistin with dysglycemia in South Asian women.
Mechanisms of obesity-associated insulin resistance and dysglycemia in South Asians remain relatively unknown. The objective of this study was to detect subcutaneous (SAT) vs. visceral (VAT) adipose tissue characteristics and adipocytokines associated with obesity, insulin resistance, and dysglycemia in South Asian women. This was a hospital-based cross-sectional study conducted in Sri Lanka. Subjects comprised of 58 adult women who underwent routine abdominal surgeries. SAT and VAT were obtained from anterior abdominal wall and omentum, respectively. Measures of adiposity, serum insulin and glucose, SAT and VAT crown-like structures (CLS), macrophages, resistin by immunohistochemistry, mean adipocyte area (MAA), and serum adipocytokines were examined. The homeostatic model assessment for insulin resistance (HOMA-IR) score significantly correlated with age and waist circumference (WC), but not with body mass index (BMI). Although the number of CLS positively correlated with BMI, there were no significant differences between the number of CLS in women with normal fasting glucose (NFG) vs. those with impaired fasting glucose (IFG), indicating that adipose tissue macrophage infiltration is unlikely to be related to dysglycemia. In contrast, serum resistin level was on average 60% higher in women with IFG compared to ones with NFG (p < 0.05). Serum resistin levels correlated with age (r = 0.36, p < 0.05) and WC (r = 0.27, p < 0.05). There were no associations in serum levels of other adipocytokines with IFG. Adipose immunohistochemistry showed that women with IFG had a higher percentage of resistin positive adipocytes in SAT compared to ones with NFG. MAA of VAT, but not SAT, correlated with both BMI and WC. Resistin may be an important adipokine linking central adiposity and insulin resistance in South Asian women. Both systemic and adipose tissue resistin are linked to dysglycemia in these individuals and may be a potential biomarker for diabetes in this population.


[How to cope with psychiatric illness in patients with epilepsy].
Almost every kind of psychiatric problems are associated with epilepsy such as psychotic states, manic as well as depressive states and anxiety attacks. Overall, the prevalence of psychiatric comorbidities in patients with epilepsy amounts to as high as 20-30% of all cases. Acute and chronic interictal psychoses, as well as postictal psychosis (or more precisely periictal psychosis), comprise 95% of psychosis in patients with epilepsy. Prevalence of depressive states in patients with yet active epilepsy ranges from 20-55%. Prevalence in patients with controlled epilepsy ranges from 3-9%. Depressive states comprise 50-80% of psychiatric co-morbidities in patients with epilepsy. Several studies reported that PNES amounted to as high as 30% among patients considered as candidates for epilepsy surgery due to intractable epilepsy. It is of clinical use that PNES is divided into 3 groups: The first group belongs to PNES without either intellectual disability nor epilepsy; The second group suffers from intellectual disability in addition to PNES; The third group shows both epileptic seizure and PNES. These groups need to be differently treated. After temporal lobectomy for controlling pharmacoresistant TLE, severe but transient depression possibly leading to suicide can appear, especially within the first few months after surgery.


Conservative management of a case of plexiform ameloblastoma.
Ameloblastomas are locally aggressive, benign odontogenic neoplasms having a wide variety of histologic patterns. It is essential to distinguish between the three clinical types of ameloblastomas - the intra-osseous solid lesion, the unicystic type and the extra-osseous lesion, as they differ in their biological behaviour and rate of recurrence and therefore require different forms of treatment. The case presented here is of a 9-year-old boy who reported to the department with pain and swelling along the right side of the mandible of 3 months'duration. Previous histopathological examination of the lesion, performed at a local hospital, produced a picture consistent with unicystic ameloblastoma. The lesion was managed by enucleation and the patient has been followed up for the past 5 years. Radiographic and clinical examinations reveal signs of healing without recurrence. Conservative management of unicystic ameloblastomas may be justified in children provided that the patient can be followed up at regular intervals.


Porous and highly dispersible voriconazole dry powders produced by spray freeze drying for pulmonary delivery with efficient lung deposition.
Systemic administration of antifungal agents for the treatment of pulmonary aspergillosis is limited by the poor lung deposition and severe adverse effects. In contrast, pulmonary delivery allows a higher amount of drug to be delivered directly to the infection site and therefore a lower dose is required. This study aimed to develop porous and inhalable voriconazole dry powder with good lung deposition by spray freeze drying (SFD), using tert-butyl alcohol (TBA) as a co-solvent. A three-factor two-level full factorial design approach was used to investigate the effect of total solute concentration, drug content and co-solvent composition on the aerosol performance of the SFD powder. In general, the SFD voriconazole powder exhibited porous and spherical structure, and displayed crystalline characteristics. The analysis of factorial design indicated that voriconazole content was the most significant variable that could influence the aerosol performance of the SFD powders. The formulations that contained a high voriconazole content (40% w/w) and high TBA concentration in the feed solution (70% v/v) displayed the highest fine particle fraction of over 40% in the Next Generation Impactor study in which the powder was dispersed with a Breezhaler® at 100 L/min. In addition, the fine particle dose of the SFD powder showed a faster dissolution rate when compared to the unformulated voriconazole. Intratracheal administration of SFD voriconazole powder to mice resulted in a substantially higher drug concentration in the lungs when comparing to the group that received an equivalent dose of liquid voriconazole formulation intravenously, while a clinically relevant plasma drug concentration was maintained for at least two hours. Overall, an inhalable voriconazole dry powder formulation exhibiting good aerosol property and lung deposition was developed with clinical translation potential.


Combination therapy of inhibitors of epidermal growth factor receptor/vascular endothelial growth factor receptor 2 (AEE788) and the mammalian target of rapamycin (RAD001) offers improved glioblastoma tumor growth inhibition.
Malignant gliomas are highly lethal tumors that display striking genetic heterogeneity. Novel therapies that inhibit a single molecular target may slow tumor progression, but tumors are likely not dependent on a signal transduction pathway. Rather, malignant gliomas exhibit sustained mitogenesis and cell growth mediated in part through the effects of receptor tyrosine kinases and the mammalian target of rapamycin (mTOR). AEE788 is a novel orally active tyrosine kinase inhibitor that decreases the kinase activity associated with the epidermal growth factor receptor and, at higher concentrations, the vascular endothelial growth factor receptor 2 (kinase domain region). RAD001 (everolimus) is an orally available mTOR inhibitor structurally related to rapamycin. We hypothesized that combined inhibition of upstream epidermal growth factor receptor and kinase domain region receptors with AEE788 and inhibition of the downstream mTOR pathway with RAD001 would result in increased efficacy against gliomas compared with single-agent therapy. In vitro experiments showed that the combination of AEE788 and RAD001 resulted in increased rates of cell cycle arrest and apoptosis and reduced proliferation more than either agent alone. Combined AEE788 and RAD001 given orally to athymic mice bearing established human malignant glioma tumor xenografts resulted in greater tumor growth inhibition and greater increases in median survival than monotherapy. These studies suggest that simultaneous inhibition of growth factor receptor and mTOR pathways offer increased benefit in glioma therapy.


The association of early life socioeconomic conditions with prediabetes and type 2 diabetes: results from the Maastricht study.
Using cross-sectional data from The Maastricht Study, we examined the association of socioeconomic conditions in early life with prediabetes and T2DM in adulthood. We also examined potential mediating pathways via both adulthood socioeconomic conditions and adult BMI and health behaviours. Of the 3263 participants (aged 40-75 years), 493 had prediabetes and 906 were diagnosed with T2DM. By using logistic regression analyses, the associations and possible mediating pathways were examined. Participants with low early life socioeconomic conditions had a 1.56 times higher odds of prediabetes (95% confidence interval (CI) = 1.21-2.02) and a 1.61 times higher odds of T2DM (95% CI = 1.31-1.99). The relation between low early life socioeconomic conditions and prediabetes was independent of current socioeconomic conditions (OR = 1.38, 95% CI = 1.05-1.80), whereas the relation with T2DM was not independent of current socioeconomic conditions (OR = 1.10, 95% CI = 0.87-1.37). BMI party mediated the association between early life socioeconomic conditions and prediabetes. Socioeconomic inequalities starting in early life were associated with diabetes-related outcomes in adulthood and suggest the usefulness of early life interventions aimed at tackling these inequalities.


The size of the proteasomal substrate determines whether its degradation will be mediated by mono- or polyubiquitylation.
A polyubiquitin chain anchored to the substrate has been the hallmark of proteasomal recognition. However, the degradation signal appears to be more complex and to contain also a substrate's unstructured region. Recent reports have shown that the proteasome can degrade also monoubiquitylated proteins, which adds an additional layer of complexity to the signal. Here, we demonstrate that the size of the substrate is an important determinant in its extent of ubiquitylation: a single ubiquitin moiety fused to a tail of up to ∼150 residues derived from either short artificial repeats or from naturally occurring proteins, is sufficient to target them for proteasomal degradation. Importantly, chemically synthesized adducts, where ubiquitin is attached to the substrate via a naturally occurring isopeptide bond, display similar characteristics. Taken together, these findings suggest that the ubiquitin proteasomal signal is adaptive, and is not always made of a long polyubiquitin chain.


Association of coffee intake with total and cause-specific mortality in a Japanese population: the Japan Public Health Center-based Prospective Study.
Despite the rising consumption of coffee worldwide, few prospective cohort studies assessed the association of coffee intake with mortality including total and major causes of death. We aimed to investigate the association between habitual coffee drinking and mortality from all causes, cancer, heart disease, cerebrovascular disease, respiratory disease, injuries, and other causes of death in a large-scale, population-based cohort study in Japan. We studied 90,914 Japanese persons aged between 40 and 69 y without a history of cancer, cerebrovascular disease, or ischemic heart disease at the time of the baseline study. Subjects were followed up for an average of 18.7 y, during which 12,874 total deaths were reported. The association between coffee intake and risk of total and cause-specific mortality was assessed by using a Cox proportional hazards regression model with adjustment for potential confounders. We showed an inverse association between coffee intake and total mortality in both men and women. HRs (95% CIs) for total death in subjects who consumed coffee compared with those who never drank coffee were 0.91 (0.86-0.95) for <1 cup/d, 0.85 (0.81-0.90) for 1-2 cups/d, 0.76 (0.70-0.83) for 3-4 cups/d, and 0.85 (0.75-0.98) for >5 cups/d (P-trend < 0.001). Coffee was inversely associated with mortality from heart disease, cerebrovascular disease, and respiratory disease. With this prospective study, we suggest that the habitual intake of coffee is associated with lower risk of total mortality and 3 leading causes of death in Japan.