Genistein-induced apoptosis of prostate cancer cells is preceded by a specific decrease in focal adhesion kinase activity.
Genistein (5,7,4'-trihydroxyisoflavone), an isoflavinoid found in soy beans, has been identified as potentially causal for the low incidence of metastatic prostate cancer (PCa) in certain countries. Although genistein-induced PCa cell adhesion has been identified as a possible causative mechanism, direct growth inhibition by genistein has been reported and also could be causal. If in vivo growth inhibition was significant, then growth inhibition should occur at concentrations attained with dietary consumption, the mechanism of growth inhibition should be relevant to PCa, and genistein (a broad-spectrum in vitro protein-tyrosine kinase inhibitor) should have relatively specific kinase inhibitory effects in vivo. These considerations were investigated by measuring growth inhibitory activity in a variety of PCa cell lines. Growth inhibitory effects were shown not to occur with concentrations below the low micromolar range (i.e., 3 logs above that attained in serum). In-depth mechanistic studies with the PC3-M metastatic variant cell line demonstrated that growth inhibition was independent of genistein's estrogenic effects. Genistein was shown to decrease the viability of nonadherent cells, suggesting a lack of dependence on cell adhesion for growth inhibition. However, important molecular and kinetic differences between genistein's effects on growth in adherent versus nonadherent cells were identified. Specific suppression of focal adhesion kinase activity (without global decreases in phosphotyrosine) was shown to precede induction of apoptosis, which was responsible for growth inhibition in adherent cells. These findings do not support an in vivo growth inhibitory role by genistein consumed in quantities associated with a soy-based diet. They do, however, identify genistein as a potential therapeutic agent for PCa and as a tool with which to study the control of apoptosis in PCa.


Neisseria gonorrhoeae and Chlamydia trachomatis infection in HIV-1-infected women taking antiretroviral therapy: a prospective cohort study from Burkina Faso.
Neisseria gonorrhoeae (NG) and Chlamydia trachomatis (CT) are common sexually transmitted infections (STI). We assessed the cumulative risk of NG and CT in a cohort of HIV-1-infected high-risk women taking antiretrovirals over 4 years in Burkina Faso. Between March 2007 and February 2011, participants were followed every 3-6 months. At each visit, participants underwent a gynaecological examination with collection of cervical and vaginal swabs. Random-effects logistic regression models were used to analyse associations of NG and CT infection with behavioural and biological factors. 172 women had samples tested for NG and CT during the study period, in a total of 1135 visits. NG was detected in 6.4% of women (11/172, 95% CI 2.7 to 10.1) at a rate of 2.76 cases (95% CI 1.53 to 4.99) per 100 person-years. CT was detected in 1.7% (3/172, 95% CI 0 to 3.7) of women at a rate of 0.75 cases (95% CI 0.24 to 2.34) per 100 person-years. The majority of women were asymptomatic (9/14). In the multivariable model, the presence of NG or CT was associated with tobacco use (aOR=11.85, 95% CI 1.13 to 124.17), and concurrent genital HIV-1 RNA shedding (aOR=4.78, 95% CI 1.17 to 19.46). Higher levels of education (aOR=0.17, 95% CI 0.03 to 0.92), and age greater than 35 years (aOR=0.07, 95% CI 0.01 to 0.92) were associated with lower odds of infection. The risk of NG or CT infection remains low among high-risk women in Bobo-Dioulasso. This provides some evidence that antiretroviral use does not contribute to behavioural disinhibition. The asymptomatic nature of most infections underscores the need for regular screening and treatment of STIs in core groups.


Conjugate addition reactions combined with free-radical cross-linking for the design of materials for tissue engineering.
PEG-diacrylamide was synthesized and utilized to make materials for tissue engineering. Acrylamide groups readily react with thiol groups, and peptides containing a single thiol group were coupled to the PEG-diacrylamide in aqueous solution at room temperature in about 2 h. If only a portion of the acrylamide groups were targeted for reaction with peptide, sufficient amounts of PEG-diacrylamide remained to be polymerized by free-radical mechanisms via photoinitiation. The photopolymerization step can be performed in contact with cells, providing a means to produce bioactive scaffolds for tissue engineering. The photopolymerization conditions and precursor composition greatly affect the stiffness of the materials, which subsequently affected cell spreading. The kinetics and extent of cell spreading on the bioactive materials were measured and compared to cell spreading on tissue culture polystyrene. Although the PEG materials resist protein adsorption, the experiments suggest that the cells can secrete extracellular matrix that can adhere to the gels.


Phenotype sequencing: identifying the genes that cause a phenotype directly from pooled sequencing of independent mutants.
Random mutagenesis and phenotype screening provide a powerful method for dissecting microbial functions, but their results can be laborious to analyze experimentally. Each mutant strain may contain 50-100 random mutations, necessitating extensive functional experiments to determine which one causes the selected phenotype. To solve this problem, we propose a "Phenotype Sequencing" approach in which genes causing the phenotype can be identified directly from sequencing of multiple independent mutants. We developed a new computational analysis method showing that 1. causal genes can be identified with high probability from even a modest number of mutant genomes; 2. costs can be cut many-fold compared with a conventional genome sequencing approach via an optimized strategy of library-pooling (multiple strains per library) and tag-pooling (multiple tagged libraries per sequencing lane). We have performed extensive validation experiments on a set of E. coli mutants with increased isobutanol biofuel tolerance. We generated a range of sequencing experiments varying from 3 to 32 mutant strains, with pooling on 1 to 3 sequencing lanes. Our statistical analysis of these data (4099 mutations from 32 mutant genomes) successfully identified 3 genes (acrB, marC, acrA) that have been independently validated as causing this experimental phenotype. It must be emphasized that our approach reduces mutant sequencing costs enormously. Whereas a conventional genome sequencing experiment would have cost $7,200 in reagents alone, our Phenotype Sequencing design yielded the same information value for only $1200. In fact, our smallest experiments reliably identified acrB and marC at a cost of only $110-$340.


Nuclear suppression of mitochondrial defects in cells without the ND6 subunit.
Previously, we characterized a mouse cell line, 4A, carrying a mitochondrial DNA mutation in the subunit for respiratory complex I, NADH dehydrogenase, in the ND6 gene. This mutation abolished the complex I assembly and disrupted the respiratory function of complex I. We now report here that a galactose-resistant clone, 4AR, was isolated from the cells carrying the ND6 mutation. 4AR still contained the homoplasmic mutation, and apparently there was no ND6 protein synthesis, whereas the assembly of other complex I subunits into complex I was recovered. Furthermore, the respiratory activity and mitochondrial membrane potential were fully recovered. To investigate the genetic origin of this compensation, the mitochondrial DNA (mtDNA) from 4AR was transferred to a new nuclear background. The transmitochondrial lines failed to grow in galactose medium. We further transferred mtDNA with a nonsense mutation at the ND5 gene to the 4AR nuclear background, and a suppression for mitochondrial deficiency was observed. Our results suggest that change(s) in the expression of a certain nucleus-encoded factor(s) can compensate for the absence of the ND6 or ND5 subunit.


Quantitative correlational study of microbubble-enhanced ultrasound imaging and magnetic resonance imaging of glioma and early response to radiotherapy in a rat model.
Radiotherapy remains a major treatment method for malignant tumors. Magnetic resonance imaging (MRI) is the standard modality for assessing glioma treatment response in the clinic. Compared to MRI, ultrasound imaging is low-cost and portable and can be used during intraoperative procedures. The purpose of this study was to quantitatively compare contrast-enhanced ultrasound (CEUS) imaging and MRI of irradiated gliomas in rats and to determine which quantitative ultrasound imaging parameters can be used for the assessment of early response to radiation in glioma. Thirteen nude rats with U87 glioma were used. A small thinned skull window preparation was performed to facilitate ultrasound imaging and mimic intraoperative procedures. Both CEUS and MRI with structural, functional, and molecular imaging parameters were performed at preradiation and at 1 day and 4 days postradiation. Statistical analysis was performed to determine the correlations between MRI and CEUS parameters and the changes between pre- and postradiation imaging. Area under the curve (AUC) in CEUS showed significant difference between preradiation and 4 days postradiation, along with four MRI parameters, T2, apparent diffusion coefficient, cerebral blood flow, and amide proton transfer-weighted (APTw) (all p < 0.05). The APTw signal was correlated with three CEUS parameters, rise time (r = - 0.527, p < 0.05), time to peak (r = - 0.501, p < 0.05), and perfusion index (r = 458, p < 0.05). Cerebral blood flow was correlated with rise time (r = - 0.589, p < 0.01) and time to peak (r = - 0.543, p < 0.05). MRI can be used for the assessment of radiotherapy treatment response and CEUS with AUC as a new technique and can also be one of the assessment methods for early response to radiation in glioma.


Childhood tuberculosis in southern Taiwan, with emphasis on central nervous system complications.
Childhood tuberculosis (TB) continues to be a major public health problem in Taiwan. Taiwan remains a highly endemic area despite neonatal Bacillus Calmette-Guérin (BCG) vaccination and the availability of anti-TB therapy. The presentation is highly variable and it is often difficult to make an accurate diagnosis. This study was designed to evaluate the demographic, clinical, and laboratory findings and outcomes of TB in children with emphasis on central nervous system (CNS) complications. The medical records of 80 children diagnosed with TB at a medical center in southern Taiwan over the past 24 years (1988-2012) were reviewed. Among them, 48.8% (39/80) had pulmonary TB, 27.5% (22/80) had isolated extrapulmonary TB, and 23.7% (19/80) had disseminated TB. Most infected cases were aged either < 4 years or > 12 years. TB contact history was found in 42.5% (34/80) cases. Fourteen (17.5%) of the cases had CNS involvement. The most common presentations were fever (85.7%), signs of increased intracranial pressure (71.4%), drowsiness (64.3%), and focal neurological signs (57.1%). The major radiological findings were tuberculoma (50%), basilar enhancement (41.6%), infarction (41.6%), hydrocephalus (16.6%), and transverse myelitis (16.6%). The case fatality of CNS TB was 14.3% and 21.4% had neurologic sequelae. Findings suggest that positive exposure history and suspicious clinical presentations are important clues for further confirmatory laboratory and image studies in childhood TB. CNS TB usually presented as part of disseminated TB in children. Early diagnosis and treatment may lead to favorable outcomes in CNS TB.


Systemic Immunity Is Required for Effective Cancer Immunotherapy.
Immune responses involve coordination across cell types and tissues. However, studies in cancer immunotherapy have focused heavily on local immune responses in the tumor microenvironment. To investigate immune activity more broadly, we performed an organism-wide study in genetically engineered cancer models using mass cytometry. We analyzed immune responses in several tissues after immunotherapy by developing intuitive models for visualizing single-cell data with statistical inference. Immune activation was evident in the tumor and systemically shortly after effective therapy was administered. However, during tumor rejection, only peripheral immune cells sustained their proliferation. This systemic response was coordinated across tissues and required for tumor eradication in several immunotherapy models. An emergent population of peripheral CD4 T cells conferred protection against new tumors and was significantly expanded in patients responding to immunotherapy. These studies demonstrate the critical impact of systemic immune responses that drive tumor rejection.


Independent signals control expression of the calcineurin inhibitory proteins MCIP1 and MCIP2 in striated muscles.
Calcineurin, a calcium/calmodulin-regulated protein phosphatase, modulates gene expression in cardiac and skeletal muscles during development and in remodeling responses such as cardiac hypertrophy that are evoked by environmental stresses or disease. Recently, we identified two genes encoding proteins (MCIP1 and MCIP2) that are enriched in striated muscles and that interact with calcineurin to inhibit its enzymatic activity. In the present study, we show that expression of MCIP1 is regulated by calcineurin activity in hearts of mice with cardiac hypertrophy, as well as in cultured skeletal myotubes. In contrast, expression of MCIP2 in the heart is not altered by activated calcineurin but responds to thyroid hormone, which has no effect on MCIP1. A approximately 900-bp intragenic segment located between exons 3 and 4 of the MCIP1 gene functions as an alternative promoter that responds to calcineurin. This region includes a dense cluster of 15 consensus binding sites for NF-AT transcription factors. Because MCIP proteins can inhibit calcineurin, these results suggest that MCIP1 participates in a negative feedback circuit to diminish potentially deleterious effects of unrestrained calcineurin activity in cardiac and skeletal myocytes. Inhibitory effects of MCIP2 on calcineurin activity may be pertinent to gene switching events driven by thyroid hormone in striated muscles. The full text of this article is available at http://www. circresaha.org.


[Metal ions in the insulin-receptor interaction in human erythrocytes].
The kinetics of insulin binding to its receptors on human erythrocytes suggests the existence of two types of receptors exhibiting negative cooperativity upon the binding of the hormone. Solubilized and purified receptors were associated to Zn++ or Cu++. The addition of these ions to erythrocytes or to purified insulin receptors from human erythrocytes resulted in an increase of specific insulin binding. Dialysis of solubilized or purified receptors against chelating agents such as EDTA or 1, 10-phenanthroline resulted in a decrease in specific binding of insulin. With the readdition of Zn++ or Cu++ to the medium an increase in specific binding was observed, and values much higher than those of the original preparations were obtained. Dialysis of purified receptors against chelating agents resulted in a decrease in the content of Zn++ and Cu++. These results suggest the possible involvement of a metal ion associated to the receptor in the formation of the insulin-receptor complex.


The CB1/CB2 receptor agonist WIN-55,212-2 reduces viability of human Kaposi's sarcoma cells in vitro.
Kaposi's sarcoma is a highly vascularized mesenchymal neoplasm arising with multiple lesions of the skin. Endogenous cannabinoids have been shown to inhibit proliferation of a wide spectrum of tumor cells. We studied the effects of cannabinoids on human Kaposi's sarcoma cell proliferation in vitro. To do so, we first investigated the presence of the cannabinoid receptors CB(1) and CB(2) mRNAs in the human Kaposi's sarcoma cell line KS-IMM by RT-PCR and, subsequently, the effects of the mixed CB(1)/CB(2) agonist WIN-55,212-2 (WIN) on cell proliferation in vitro. WIN showed antimitogenic effects on Kaposi's sarcoma cells. Western blot analysis of Kaposi's sarcoma lysates suggested that WIN treatment induced activation of both caspase-3 and -6, as well as increased phosphorylation of the stress kinase p38 and JNK, along with transient phosphorylation of ERK(1/2). To better characterize the involvement of each single CB receptor in cannabinoid-induced cell death, we incubated Kaposi's sarcoma cells with different selective cannabinoid receptor agonists, respectively ACEA (CB(1)) and JWH-133 (CB(2)). None of the agonists was able to induce KS-IMM cell apoptosis. Moreover, we co-incubated Kaposi's sarcoma cells with WIN-55,212-2 and either the CB(1) receptor antagonist AM251, the CB(2) receptor antagonist AM630, or a combination of both substances. The CB(2) receptor antagonist AM630 was able to significantly increase survival of Kaposi's sarcoma cells treated with WIN. In view of the antiproliferative effects of cannabinoids on KS-IMM cells, one could envision the cannabinoid system as a potential target for pharmacological treatment of Kaposi's sarcoma.


Canada's future healthcare: can it be better? Will it be better?
Thought leaders envisage high-performing partnerships of engaged community practitioners, informed patients and non-professional caregivers collaborating continuously, and efficiently, to improve care and outcomes for whole patient populations. These primary care health social networks would be facilitated by needs-based training, meaningful measurements, sustained funding, effective leadership and integration with available resources and processes. Broadly voiced opinion supports such integrated, community-focused partnership and data-driven healthcare models, and a province-wide implementation of the model for acute and chronic cardiac diseases in Nova Scotia has conclusively demonstrated sustained improvements in clinical and economic outcomes. A reasonable hypothesis, then, is that such strategies will be rapidly adopted to effectively manage the primary care of our increasingly aged populations, with their large and recalcitrant gaps between usual and best care. However, there are impediments to widespread adoption in the short term, not the least being disparities in various key stakeholders' level of preference, commitment, resolve and clout in making the necessary decisions to adopt and sustain the strategies. Thus, while we know things can be better in Canadian healthcare, the answers to, will they? and, when? remain less certain.


Erythema multiforme due to Mycoplasma pneumoniae infection in two children.
Mycoplasma pneumoniae is an important and highly relevant cause of bullous erythema multiforme, isolated mucositis, and Stevens-Johnson syndrome in children. In this article, we present two children with respiratory Mycoplasma pneumoniae infection and associated cutaneous findings within the spectrum of erythema multiforme. We review the literature associating these three entities with Mycoplasma pneumoniae infection and discuss controversies regarding the classification of erythema multiforme, as well as update reported infectious causes of the bullous form. Many understand the erythema multiforme spectrum to include bullous erythema multiforme, mucositis, and Stevens-Johnson syndrome in the order of increasing severity. We feel that this relationship should be reconsidered to help better understand the prognosis and outcomes. It is our opinion that bullous erythema multiforme is a separate, yet related condition that can occur in the context of Mycoplasma pneumoniae infection. With many similarities to mucositis and Stevens-Johnson syndrome, bullous erythema multiforme can be considered part of a spectrum of disease that includes Stevens-Johnson syndrome. Unlike mucositis and Stevens-Johnson syndrome, bullous erythema multiforme caused by Mycoplasma pneumoniae infection has low morbidity for the child. Mycoplasma pneumoniae-associated mucositis and Stevens-Johnson syndrome seem to occur along a spectrum with separate prognosis and potential pathogenesis compared with bullous erythema multiforme. Making the distinction between these conditions is valuable for predicting the child's prognosis. Patients who develop symptoms consistent with these conditions should be appropriately evaluated for Mycoplasma pneumoniae infection and closely monitored.


Olig1 function is required for remyelination potential of transplanted neural progenitor cells in a model of viral-induced demyelination.
Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) resulting in cumulative neurologic deficits associated with progressive myelin loss. We have previously shown that transplantation of neural progenitor cells (NPCs) into mice persistently infected with the JHM strain of mouse hepatitis virus (JHMV) results in enhanced differentiation into oligodendrocyte progenitor cells (OPCs) that is associated with remyelination and axonal sparing. The current study examines the contributions of the transcription factor Olig1 on NPC differentiation and remyelination. Under defined conditions, NPCs preferentially differentiate into oligodendroglia whereas NPCs isolated from Olig1-deficient (Olig1-/-) mice exhibit enhanced differentiation into astrocytes. Transplantation of Olig1-/- and Olig1+/+ NPCs into JHMV-infected mice resulted in similar cell survival, proliferation, and selective migration to areas of demyelination. However, only recipients of wild type NPCs exhibited extensive remyelination compared to mice receiving Olig1-/- NPCs. In vivo characterization of NPCs revealed that Olig1+/+ NPCs preferentially differentiated into NG2-positive OPCs and formed processes expressing myelin basic protein that encircled axons. In contrast, the majority of transplanted Olig1-/- NPCs differentiated into GFAP-positive cells consistent with the astrocyte lineage. These results indicate that exogenous NPCs contribute to improved clinical and histological outcome and this is associated with remyelination by this donor population. Further, these findings reveal that Olig1function is required for the remyelination potential of NPCs after transplant, through specification and/or maintenance of oligodendroglial identity.


Which, who and when? Therapeutic processes in group cognitive behaviour therapy for anxiety disorders.
The evidence for the effect of therapeutic alliance in group cognitive behaviour therapy (CBT) for anxiety disorders is unclear. Identifying whether the alliance-outcome relationship depends on (1) which components are assessed, (2) who is measuring the alliance and (3) when the alliance is measured will help to clarify the role of the client-therapist relationship in therapy. The present study explored the effects of alliance component (agreement vs. bond), rater perspective (client vs. therapist) and timing (early vs. late therapy) on the alliance-outcome relationship. Individuals with an anxiety disorder enrolled into transdiagnostic group CBT were studied, with n = 78 at early therapy and n = 57 at late therapy. Results showed that greater client-rated agreement significantly predicted improved post-treatment outcomes throughout the course of therapy, while stronger client-rated bond in late therapy predicted reduced treatment gains. In contrast, therapist perceptions of agreement and bond were not associated with post-treatment outcomes at any point in therapy. Client-reported group cohesion also was not associated with additional variance in outcome after accounting for client-rated alliance. Overall, the findings highlight the importance of prioritizing the client's perception of the client-therapist relationship in CBT for anxiety disorders, as well as distinguishing the effects of component, rater and timing in future process-outcome studies.


Comparing Palliative Care in Care Homes Across Europe (PACE): Protocol of a Cross-sectional Study of Deceased Residents in 6 EU Countries.
Although a growing number of older people are dying in care homes, palliative care has developed in these settings only recently. Cross-country representative comparative research hardly exists in this area. As part of a large EU-funded project, we aim to undertake representative comparative research in care homes in Europe, to describe and compare 6 countries in terms of (1) resident outcomes, quality and costs of palliative and end-of-life care; and (2) palliative care structures and staff knowledge and attitudes toward palliative care. We also aim to explore country, facility, staff, patient, and care characteristics related to better outcomes at resident level. To obtain a representative nationwide sample, we will conduct a large-scale cross-sectional study of deceased residents in care homes in Belgium, Finland, Italy, the Netherlands, Poland, and the United Kingdom, using proportional stratified random sampling (taking into account region, facility type and bed capacity). In each country, all participating care homes retrospectively report all deaths of residents in and outside the facilities over the previous 3-month period. For each case, structured questionnaires, including validated instruments, are sent to (1) the administrator/manager, (2) staff member most involved in care, (3) treating physician (general practitioner or elderly care physician), and (4) a closely involved relative. It is estimated that, per country, 50 care homes are needed on average to obtain a minimum of 200 deceased residents. Collected data include clinical and sociodemographic characteristics, quality of dying, quality and costs of palliative care and end-of-life care, and palliative care structures at the facility level and country level. To obtain a representative view of staff knowledge and attitudes regarding palliative care, PACE will conduct a cross-sectional study of staff working in the participating care homes. Considering the growing challenges associated with aging in all European countries, there is an urgent need to build a robust international comparative evidence base that can inform the development of policies to target improved palliative care in care homes. By describing this research protocol, we hope to inform international research in care homes on how to perform representative end-of-life care research in these settings and better understand which systems are associated with better outcomes.


Depth profiling for the identification of unknown substances and concealed content at remote distances using time-resolved stand-off Raman spectroscopy.
Time-resolved stand-off Raman spectroscopy was used to determine both the position and identity of substances relative to each other at remote distances (up to tens of meters). Spectral information of three xylene isomers, toluene, and sodium chlorate was obtained at a distance of 12 m from the setup. Pairs and triplets of these samples were placed at varying distances (10-60 cm) relative to each other. Via the photon time of flight the distance between the individual samples was determined to an accuracy of 7% (corresponding to a few cm) of the physically measured distance. Furthermore, at a distance of 40 m, time-resolved Raman depth profiling was used to detect sodium chlorate in a white plastic container that was non-transparent to the human eye. The combination of the ranging capabilities of Raman LIDAR (sample location usually determined using prior knowledge of the analyte of interest) with stand-off Raman spectroscopy (analyte detection at remote distances) provides the capability for depth profile identification of unknown substances and analysis of concealed content in distant objects. To achieve these results, a 532 nm laser with a pulse length of 4.4 ns was synchronized to an intensified charge-coupled device camera with a minimum gate width of 500 ps. For automated data analysis a multivariate curve resolution algorithm was employed.


Leptospirosis presenting as acute meningoencephalitis.
Leptospirosis in humans is a common zoonotic disease. It is often under-diagnosed, especially when associated with neurological features, resulting in significant morbidity and mortality. This subgroup of patients with neurological manifestations is often empirically treated for cerebral malaria, dengue fever, tuberculous meningitis, hepatic encephalopathy, viral encephalitis, etc. Hence it is important to be aware of uncommon manifestations of this disease. We report one such patient, which highlights the importance of considering leptospirosis as the diagnostic possibility with hepato-renal, pulmonary and nervous system involvement, particularly where diagnostic supports and resources are limited.


Balancing selection at the tomato RCR3 Guardee gene family maintains variation in strength of pathogen defense.
Coevolution between hosts and pathogens is thought to occur between interacting molecules of both species. This results in the maintenance of genetic diversity at pathogen antigens (or so-called effectors) and host resistance genes such as the major histocompatibility complex (MHC) in mammals or resistance (R) genes in plants. In plant-pathogen interactions, the current paradigm posits that a specific defense response is activated upon recognition of pathogen effectors via interaction with their corresponding R proteins. According to the "Guard-Hypothesis," R proteins (the "guards") can sense modification of target molecules in the host (the "guardees") by pathogen effectors and subsequently trigger the defense response. Multiple studies have reported high genetic diversity at R genes maintained by balancing selection. In contrast, little is known about the evolutionary mechanisms shaping the guardee, which may be subject to contrasting evolutionary forces. Here we show that the evolution of the guardee RCR3 is characterized by gene duplication, frequent gene conversion, and balancing selection in the wild tomato species Solanum peruvianum. Investigating the functional characteristics of 54 natural variants through in vitro and in planta assays, we detected differences in recognition of the pathogen effector through interaction with the guardee, as well as substantial variation in the strength of the defense response. This variation is maintained by balancing selection at each copy of the RCR3 gene. Our analyses pinpoint three amino acid polymorphisms with key functional consequences for the coevolution between the guardee (RCR3) and its guard (Cf-2). We conclude that, in addition to coevolution at the "guardee-effector" interface for pathogen recognition, natural selection acts on the "guard-guardee" interface. Guardee evolution may be governed by a counterbalance between improved activation in the presence and prevention of auto-immune responses in the absence of the corresponding pathogen.


Ferrimagnetic Large Single Domain Iron Oxide Nanoparticles for Hyperthermia Applications.
This paper describes the preparation and obtained magnetic properties of large single domain iron oxide nanoparticles. Such ferrimagnetic particles are particularly interesting for diagnostic and therapeutic applications in medicine or (bio)technology. The particles were prepared by a modified oxidation method of non-magnetic precursors following the green rust synthesis and characterized regarding their structural and magnetic properties. For increasing preparation temperatures (5 to 85 °C), an increasing particle size in the range of 30 to 60 nm is observed. Magnetic measurements confirm a single domain ferrimagnetic behavior with a mean saturation magnetization of ca. 90 Am2/kg and a size-dependent coercivity in the range of 6 to 15 kA/m. The samples show a specific absorption rate (SAR) of up to 600 W/g, which is promising for magnetic hyperthermia application. For particle preparation temperatures above 45 °C, a non-magnetic impurity phase occurs besides the magnetic iron oxides that results in a reduced net saturation magnetization.


Coupling of methylmercury uptake with respiration and water pumping in freshwater tilapia Oreochromis niloticus.
The relationships among the uptake of toxic methylmercury (MeHg) and two important fish physiological processes-respiration and water pumping--in the Nile tilapia (Oreochromis niloticus) were explored in the present study. Coupled radiotracer and respirometric techniques were applied to measure simultaneously the uptake rates of MeHg, water, and oxygen under various environmental conditions (temperature, dissolved oxygen level, and water flow). A higher temperature enhanced MeHg influx and the oxygen consumption rate but had no effect on the water uptake, indicating the influence of metabolism on MeHg uptake. The fish showed a high tolerance to hypoxia, and the oxygen consumption rate was not affected until the dissolved oxygen concentration decreased to extremely low levels (below 1 mg/L). The MeHg and water uptake rates increased simultaneously as the dissolved oxygen level decreased, suggesting the coupling of water flux and MeHg uptake. The influence of fish swimming performance on MeHg uptake was also investigated for the first time. Rapidly swimming fish showed significantly higher uptake rates of MeHg, water, and oxygen, confirming the coupling relationships among respiration, water pumping, and metal uptake. Moreover, these results support that MeHg uptake is a rate-limiting process involving energy. Our study demonstrates the importance of physiological processes in understanding mercury bioaccumulation in fluctuating aquatic environments.


Knowing when not to swing: EEG evidence that enhanced perception-action coupling underlies baseball batter expertise.
Given a decision that requires less than half a second for evaluating the characteristics of the incoming pitch and generating a motor response, hitting a baseball potentially requires unique perception-action coupling to achieve high performance. We designed a rapid perceptual decision-making experiment modeled as a Go/No-Go task yet tailored to reflect a real scenario confronted by a baseball hitter. For groups of experts (Division I baseball players) and novices (non-players), we recorded electroencephalography (EEG) while they performed the task. We analyzed evoked EEG single-trial variability, contingent negative variation (CNV), and pre-stimulus alpha power with respect to the expert vs. novice groups. We found strong evidence for differences in inhibitory processes between the two groups, specifically differential activity in supplementary motor areas (SMA), indicative of enhanced inhibitory control in the expert (baseball player) group. We also found selective activity in the fusiform gyrus (FG) and orbital gyrus in the expert group, suggesting an enhanced perception-action coupling in baseball players that differentiates them from matched controls. In sum, our results show that EEG correlates of decision formation can be used to identify neural markers of high-performance athletes.


Plasmodium falciparum Merozoite Associated Armadillo Protein (PfMAAP) Is Apically Localized in Free Merozoites and Antibodies Are Associated With Reduced Risk of Malaria.
Understanding the functional role of proteins expressed by Plasmodium falciparum is an important step toward unlocking potential targets for the development of therapeutic or diagnostic interventions. The armadillo (ARM) repeat protein superfamily is associated with varied functions across the eukaryotes. Therefore, it is important to understand the role of members of this protein family in Plasmodium biology. The Plasmodium falciparum armadillo repeats only (PfARO; Pf3D7_0414900) and P. falciparum merozoite organizing proteins (PfMOP; Pf3D7_0917000) are armadillo-repeat containing proteins previously characterized in P. falciparum. Here, we describe the characterization of another ARM repeat-containing protein in P. falciparum, which we have named the P. falciparum Merozoites-Associated Armadillo repeats protein (PfMAAP). Antibodies raised to three different synthetic peptides of PfMAAP show apical staining of free merozoites and those within the mature infected schizont. We also demonstrate that the antibodies raised to the PfMAAP peptides inhibited invasion of erythrocytes by merozoites from different parasite isolates. In addition, naturally acquired human antibodies to the N- and C- termini of PfMAAP are associated with a reduced risk of malaria in a prospective cohort analysis.


The Great Recession and Children's Mental Health in Australia.
This paper analyzes the effects of "shocks" to community-level unemployment expectations, induced by the onset of the Great Recession, on children's mental well-being. The Australian experience of the Great Recession represents a unique case study as despite little change in actual unemployment rates, levels of economic uncertainty grew. This affords us the ability to examine the effects of shocks to economic expectations independent of any actual changes to economic conditions. We draw on and link data from multiple sources, including several waves of the Longitudinal Study of Australian Children (2004⁻2010), a consumer sentiment survey, and data on local economic conditions. Using our purpose-built data set, we estimate difference-in-differences models to identify plausibly causal effects. We find, for boys, there is no detectable effect of community-level unemployment expectations shocks on mental health. For girls, however, there are modest increases in mental health problems and externalizing behaviors, as measured by the Strengths and Difficulties Questionnaire (SDQ). We additionally find no discernible change in mother's psychological distress as a result of expectations shocks. These results are stable after controlling for actual labor market conditions.


Tension-sensitive kinetochore phosphorylation in vitro.
Many cells have a checkpoint that detects a single misattached chromosome and delays anaphase, allowing time for error correction. Detection probably depends on tension-sensitive kinetochore protein phosphorylation. Somehow, mechanical tension, or some consequence of tension, produces a chemical change, dephosphorylation. The mechanism of tension-mediated dephosphorylation can be approached using an in vitro system. Earlier work showed that the kinetochores of washed chromosomes from a mammalian cell line can be phosphorylated in vitro simply by incubation with ATP and a phosphatase inhibitor. We confirm this for chromosomes from insect meiotic cells. Thus, kinetochores of washed chromosomes from diverse sources contain a complete phosphorylation system: a kinase, a phosphatase and the substrate protein(s). We show that phosphorylation in vitro is sensitive to tension, as it is in living cells. This makes the conditions required for phosphorylation in vitro relevant to the process in living cells. The phosphatase is ruled out as the tension-sensitive component in vitro, leaving either the kinase or the substrate as the sensitive component. We show that a kinase extracted from mammalian cells in mitosis phosphorylates the kinetochores of insect meiotic chromosomes very effectively. The mammalian kinase under-phosphorylates the kinetochore of the insect's X-chromosome, just as the native insect kinase does. This provides a clue to the evolution of a chromosome that is not detected by the checkpoint. The mammalian kinase is not tightly bound to the chromosome and thus functions primarily in solution. This suggests that the substrate's phosphorylatable groups are freely available to outside constituents, e.g. regulators, as well as to the kinetochore's own kinase and phosphatase.


Effects of hydroxyurea administration on the body weight, body composition and exercise performance of patients with sickle-cell anaemia.
1. As an ancillary study carried out during the recently completed Multicenter Study of Hydroxyurea, we examined the effect of hydroxyurea on the body weight, body composition and exercise capacity of adult patients with sickle-cell anaemia. 2. The subjects received either hydroxyurea (six males and four females) or placebo (eight males and six females). Data for each subject were generated during four separate 24 h admissions to the General Clinical Research Center. These admissions occurred at baseline and then at 6, 12 and 18 months after the start of study drug (hydroxyurea or placebo) administration. During each admission, body composition was measured by using a dual X-ray absorptiometer, and exercise testing was performed by cycle ergometry. Anaerobic performance was assessed according to a 'Wingate' protocol (20 s at maximal intensity against a cycling resistance of 7.5% body weight). Aerobic performance was examined using a steady state submaximal exercise protocol (10 min cycling time). 3. At baseline, no significant difference in any parameter was found between the hydroxyurea- and placebo-treated groups. At 18 months, the hydroxyurea-treated subjects exhibited an average weight gain of 3.16 kg. The mean weight gain in the placebo-treated subjects was 1.82 kg. Body composition analysis showed that the additional weight in both groups involved both lean and fat body mass components. In anaerobic performance, the subjects given hydroxyurea showed an increase in peak muscle power of 104.9 W. The placebo group also showed an increase, but theirs was a more modest gain of 57.7 W. The most marked improvement in anaerobic performance was observed in the hydroxyurea-treated men (P < 0.05). In aerobic performance, the hydroxyurea-treated subjects exhibited a decrease in peak heart rate response to a standardized workload of 15.2 beats/min, as compared with a decrease of only 4.3 beats/min in the placebo-treated patients. 4. Taken together, the overall weight gain, combined with increases in both anaerobic muscular performance and aerobic cardiovascular efficiency, provides objective data to support the subjective impression that hydroxyurea administration produces an improvement in the physical capacity of patients with sickle-cell anaemia.


Two Lifestyle Risks Intertwined: Parental Smoking Predicts Child Gambling Behavior at Age 12 Years.
Parental smoking can create a toxic environment for child development. A parental smoking lifestyle can predispose children to executive deficits, influencing precocious risk activities. Using a prospective birth cohort design, we examine the association between 2 lifestyle factors by estimating the relative contribution of long-term parental household smoking in predicting subsequent precocious child gambling behavior. Parents reported on the amount of household smoke exposure from ages 1.5 to 7.5 years for children from the Quebec Longitudinal Study of Child Development. The main outcome measure was children's self-report of gambling behavior (at age 12 years). Sixty percent of parents reported that their children were never exposed to secondhand smoke in the home, while 27% and 13% reported transient and continuous levels of secondhand smoke, respectively. Overall, 16% of children reported gambling participation. When compared with never-exposed children, children exposed to secondhand smoke had 18% more chances of having participated in gambling at age 12 years (odds ratio = 1.18; 95% CI = 1.080-1.293). These results are adjusted for competing explanations and possible individual and family confounders. Higher levels of early childhood household smoke exposure are associated with greater odds of reporting gambling participation at age 12 years, which is more than several years before it is normative youthful behavior. By connecting the neurotoxic influence of one lifestyle factor on another, we show a nontrivial link between 2 public health issues (smoke exposure and precocious gambling) associated with considerable individual and societal costs that are amenable to community information campaigns.


Enhancing Chinese tone recognition by manipulating amplitude envelope: implications for cochlear implants.
Tone recognition is important for speech understanding in tonal languages such as Mandarin Chinese. Cochlear implant patients are able to perceive some tonal information by using temporal cues such as periodicity-related amplitude fluctuations and similarities between the fundamental frequency (F0) contour and the amplitude envelope. The present study investigates whether modifying the amplitude envelope to better resemble the F0 contour can further improve tone recognition in multichannel cochlear implants. Chinese tone and vowel recognition were measured for six native Chinese normal-hearing subjects listening to a simulation of a four-channel cochlear implant speech processor with and without amplitude envelope enhancement. Two algorithms were proposed to modify the amplitude envelope to more closely resemble the F0 contour. In the first algorithm, the amplitude envelope as well as the modulation depth of periodicity fluctuations was adjusted for each spectral channel. In the second algorithm, the overall amplitude envelope was adjusted before multichannel speech processing, thus reducing any local distortions to the speech spectral envelope. The results showed that both algorithms significantly improved Chinese tone recognition. By adjusting the overall amplitude envelope to match the F0 contour before multichannel processing, vowel recognition was better preserved and less speech-processing computation was required. The results suggest that modifying the amplitude envelope to more closely resemble the F0 contour may be a useful approach toward improving Chinese-speaking cochlear implant patients' tone recognition.


Reducing random measurement error in assessing postural load on the back in epidemiologic surveys.
The goal of this study was to design strategies to assess postural load on the back in occupational epidemiology by taking into account the reliability of measurement methods and the variability of exposure among the workers under study. Intermethod reliability studies were evaluated to estimate the systematic bias (accuracy) and random measurement error (precision) of various methods to assess postural load on the back. Intramethod reliability studies were reviewed to estimate random variability of back load over time. Intermethod surveys have shown that questionnaires have a moderate reliability for gross activities such as sitting, whereas duration of trunk flexion and rotation should be assessed by observation methods or inclinometers. Intramethod surveys indicate that exposure variability can markedly affect the reliability of estimates of back load if the estimates are based upon a single measurement over a certain time period. Equations have been presented to evaluate various study designs according to the reliability of the measurement method, the optimum allocation of the number of repeated measurements per subject, and the number of subjects in the study. Prior to a large epidemiologic study, an exposure-oriented survey should be conducted to evaluate the performance of measurement instruments and to estimate sources of variability for back load. The strategy for assessing back load can be optimized by balancing the number of workers under study and the number of repeated measurements per worker.


First systematic experience of preimplantation genetic diagnosis for single-gene disorders, and/or preimplantation human leukocyte antigen typing, combined with 24-chromosome aneuploidy testing.
To study the feasibility, accuracy, and reproductive outcome of 24-chromosome aneuploidy testing (24-AT), combined with preimplantation genetic diagnosis (PGD) for single-gene disorders (SGDs) or human leukocyte antigen (HLA) typing in the same biopsy sample. Retrospective study. Preimplantation genetic diagnosis center. A total of 238 PGD patients, average age 36.8 years, for whom 317 combined PGD cycles were performed, involving 105 different conditions, with or without HLA typing. Whole-genome amplification product, obtained in 24-AT, was used for PGD and/or HLA typing in the same blastomere or blastocyst biopsy samples. Proportion of the embryos suitable for transfer detected in these blastomere or blastocyst samples, and the resulting pregnancy and spontaneous abortion rates. Embryos suitable for transfer were detected in 42% blastocyst and 25.1% blastomere samples, with a total of 280 unaffected, HLA-matched euploid embryos detected for transfer in 212 cycles (1.3 embryos per transfer), resulting in 145 (68.4%) unaffected pregnancies and birth of 149 healthy, HLA-matched children. This outcome is significantly different from that of our 2,064 PGD cycle series without concomitant 24-AT, including improved pregnancy (68.4% vs. 45.4%) and 3-fold spontaneous abortion reduction (5.5% vs. 15%) rates. The introduced combined approach is a potential universal PGD test, which in addition to achieving extremely high diagnostic accuracy, significantly improves reproductive outcomes of PGD for SGDs and HLA typing in patients of advanced reproductive age.


Ribosomal Database Project: data and tools for high throughput rRNA analysis.
Ribosomal Database Project (RDP; http://rdp.cme.msu.edu/) provides the research community with aligned and annotated rRNA gene sequence data, along with tools to allow researchers to analyze their own rRNA gene sequences in the RDP framework. RDP data and tools are utilized in fields as diverse as human health, microbial ecology, environmental microbiology, nucleic acid chemistry, taxonomy and phylogenetics. In addition to aligned and annotated collections of bacterial and archaeal small subunit rRNA genes, RDP now includes a collection of fungal large subunit rRNA genes. RDP tools, including Classifier and Aligner, have been updated to work with this new fungal collection. The use of high-throughput sequencing to characterize environmental microbial populations has exploded in the past several years, and as sequence technologies have improved, the sizes of environmental datasets have increased. With release 11, RDP is providing an expanded set of tools to facilitate analysis of high-throughput data, including both single-stranded and paired-end reads. In addition, most tools are now available as open source packages for download and local use by researchers with high-volume needs or who would like to develop custom analysis pipelines.


Modulation of lymphocyte proliferative responses by a canine Lyme disease vaccine of recombinant outer surface protein A (OspA).
The modulation of human lymphocyte proliferative responses was demonstrated with a recombinant outer surface protein A (OspA) vaccine preparation for the prevention of Borrelia burgdorferi infection. After exposure to either the unaltered vaccine preparation or OspA prepared in saline, normal lymphocyte responses to the mitogens concanavalin A, phytohemagglutinin-M or pokeweed mitogen, or the antigen BCG were consistently reduced. Whole cell extracts of B. burgdorferi also modulated immune responses but required a much greater quantity of protein than needed for the OspA preparation. The magnitude of modulation was directly dependent on the quantity of OspA. OspA interferes with the response of lymphocytes to proliferative stimuli including a blocking of cell cycle phase progression. Future studies designed to delete the particular region or component of the OspA molecule responsible for this effect may lead to improved vaccine preparations.


Polarization converters in highly birefringent microfibers.
A novel type of polarization converters (PCs) based on highly birefringent (Hi-Bi) microfibers is presented. Analytical formulation based on the Jones Matrix method and a numerical code based on the Full Vectorial Finite Difference Beam Propagation Method are developed to analyze the polarization evolutions in such PCs. Two different design configurations, namely the "one-side" and "two-side" perturbation configurations, are studied by use of the two methods, and the results obtained agree well with each others. The PCs can be flexibly designed to have different operating wavelengths, spectral bandwidths, and devices lengths. A particular PC based on an elliptical microfiber demonstrates a bandwidth of ∼ 600 nm around 1550 nm with a device length of ∼ 150 μm.


Endocrine aspects of ACE2 regulation: RAAS, steroid hormones and SARS-CoV-2.
Coronavirus disease (COVID-19) is caused by a new strain of coronavirus, the severe acute respiratory syndrome coronavirus 2 or SARS-CoV-2. At the time of writing, SARS-CoV-2 has infected over 5 million people worldwide. A key step in understanding the pathobiology of the SARS-CoV-2 was the identification of -converting enzyme 2 (ACE2) as the receptor for SARS-CoV-2 to gain entry into host cells. ACE2 is an established component of the 'protective arm' of the renin-angiotensin-aldosterone-system (RAAS) that opposes ACE/angiotensin II (ANG II) pressor and tissue remodelling actions. Identification of ACE2 as the entry point for SARS-CoV-2 into cells quickly focused attention on the use of ACE inhibitors (ACEi), angiotensin receptor blockers (ARB) and mineralocorticoid receptor antagonists (MRA) in patients with hypertension and cardiovascular disease given that these pharmacological agents upregulate ACE2 expression in target cells. ACE2 is cleaved from the cells by metalloproteases ADAM10 and ADAM17. Steroid hormone receptors regulate multiple components of the RAAS and may contribute to the observed variation in the incidence of severe COVID-19 between men and women, and in patients with pre-existing endocrine-related disease. Moreover, glucocorticoids play a critical role in the acute and chronic management of inflammatory disease, independent of any effect on RAAS activity. Dexamethasone, a synthetic glucocorticoid, has emerged as a life-saving treatment in severe COVID-19. This review will examine the endocrine mechanisms that control ACE2 and discusses the impact of therapies targeting the RAAS, glucocorticoid and other endocrine systems for their relevance to the impact of SARS-CoV-2 infection and the treatment and recovery from COVID-19-related critical illness.


'Real-life' report on the management of chronic GvHD in the Gruppo Italiano Trapianto Midollo Osseo (GITMO).
Several guidelines have been published about management of chronic GvHD (cGvHD), but the clinical practice still remains demanding. The Gruppo Italiano Trapianto di Midollo Osseo (GITMO) has planned a prospective observational study on cGvHD, supported by a dedicated software, including the updated recommendations. In view of this study, two surveys have been conducted, focusing the management of cGvHD and ancillary therapy in cGvHD, to address the current 'real life' situation. The two surveys were sent to all 57 GITMO centers, performing allografting in Italy; the response rate was 57% and 66% of the interviewed centers, respectively. The first survey showed a great disparity especially regarding steroid-refractory cGvHD, although extracorporeal photo-apheresis resulted as the most indicated treatment in this setting. Another challenging issue was the strategy for tapering steroid: our survey showed a great variance, and this disagreement could be a real bias in evaluating outcomes in prospective studies. As for the second survey, the results suggest that the ancillary treatments are not standardized in many centers. All responding centers reported a strong need to standardize management of cGvHD and to participate in prospective trials. Before starting observational and/or interventional studies, a detailed knowledge of current practice should be encouraged.


Imaging findings of pulmonary granulomatosis with polyangiitis (Wegener's granulomatosis): lesions invading the pulmonary fissure, pleura or diaphragm mimicking malignancy.
Granulomatosis with polyangiitis (GPA, formerly Wegener's granulomatosis), in which pulmonary involvement often predominates, is a multisystem granulomatous, necrotizing vasculitis that affects small and medium-sized vessels. In this study we evaluated various radiological findings of pulmonary GPA and focused on spiculated pulmonary lesions invading the pulmonary fissure, pleura or diaphragm mimicking malignancy. This retrospective study included 48 patients, aged 28-73 (mean, 47.3) years, who showed either histopathological diagnosis of GPA (n = 39) or elevated levels of the cytoplasmic anti-neutrophilic cytoplasmic antibody serum marker (n = 9) between January 2003 and December 2013. All patients received a chest computed tomography (CT), and the types of pulmonary lesions were defined and evaluated. Among the 48 patients, 33 had abnormal pulmonary findings on CT. The most commonly detected pulmonary lesion types were nodules and masses (n = 126) observed in 24 patients. Cavitation, necrosis, spiculation and invasion of the fissure, pleura or diaphragm were observed in 14, 9, 10 and 6 patients, respectively. Consolidation was found in 14 patients and thickening of bronchial wall in 8 patients. Pulmonary lesion types of GPA have a wide spectrum, potentially mimicking a high number of diseases including malignancy, infection and noninfectious inflammatory diseases. A spiculated lung lesion invading the fissure, pleura or diaphragm is mostly present in malignancy, but it can be also seen in GPA.


Type 1 Diabetes-related Autoantibodies in Different Forms of Diabetes.
Autoantibodies against Glutamic Acid Decarboxylase (GADA), insulinoma antigen-2 (IA- 2A), insulin (IAA) and the most recently Zinc Transporter 8 (ZnT8A) are one of the most reliable biomarkers for autoimmune diabetes in both children and adults. They are today the only biomarkers that can distinguish Latent Autoimmune Diabetes in Adults (LADA) from phenotypically type 2 diabetes. As the frequency of autoantibodies at diagnosis in childhood type 1 diabetes depends on age, GADA is by far the most common in adult onset autoimmune diabetes, especially LADA. Being multiple autoantibody positive have also shown to be more common in childhood diabetes compared to adult onset diabetes, and multiple autoantibody positivity have a high predictive value of childhood type 1 diabetes. Autoantibodies have shown inconsistent results to predict diabetes in adults. Levels of autoantibodies are reported to cause heterogeneity in LADA. Reports indicate that individuals with high levels of autoantibodies have a more type 1 diabetes like phenotype and individuals with low levels of autoantibody positivity have a more type 2 diabetes like phenotype. It is also well known that autoantibody levels can fluctuate and transient autoantibody positivity in adult onset autoimmune diabetes have been reported to affect the phenotype.


Physiology of L- and M-cone inputs to H1 horizontal cells in the primate retina.
In the primate retina, H1 horizontal cells form an electrically coupled network and receive convergent input from long- (L-) and middle- (M-) wavelength-sensitive cones. Using an in vitro preparation of the intact retina to record the light-evoked voltage responses of H1 cells, we systematically varied the L- and M-cone stimulus contrast and measured the relative L- and M-cone input strength for 137 cells across 33 retinas from three Old World species (Macaca nemestrina, M. fascicularis, and Papio anubis). We found that the L- and the M-cone inputs were summed by the H1 cell in proportion to the stimulus cone contrast, which yielded a measure of what we term L- and M-cone contrast gain. The proportion of L-cone contrast gain was highly variable, ranging from 25% to 90% [mean +/- standard deviation, (60 +/- 14)%]. This variability was accounted for by retinal location within an individual, with the temporal retina showing a consistently higher percentage of L-cone gain, and by large overall variation across individuals, with the mean percentage of L-cone gain ranging from 32% to 80%. We hypothesize that the relative L- and M-cone contrast gain is determined simply by the relative number of L and M cones in the H1 cell's receptive field and that the variability in L- and M-cone contrast gain reflects a corresponding variability in the mosaic of L and M cones.


In vivo MR imaging with simultaneous RF transmission and reception.
To present a practical scheme of a simultaneous radiofrequency (RF) transmit (Tx) and receive (Rx) (STAR) system for MRI, discuss the challenges and solutions, and show preliminary in vivo MR images obtained with this new technique. A remotely controlled STAR system was built and tested with a transverse electromagnetic head coil on a 4T (Oxford, 90 cm-bore) MRI scanner equipped with an Agilent DirectDrive console (Agilent, Santa Clara, CA). In vivo head images have been acquired using continuous sweep excitation and acquisition. The bench test and MR experimental results show our STAR system to have high isolation (60 dB) between Tx and Rx, with insensitivity to load swings created by head motion. To acquire in vivo head images, ultralow RF peak power of 50 mW was used. A novel motion-insensitive STAR MRI technique was developed and experimentally tested. The first in vivo MR images using this method were acquired. Magn Reson Med 76:1932-1938, 2016. © 2016 International Society for Magnetic Resonance in Medicine.


Charge movement in the membrane of striated muscle.
1. Non-linear polarization currents apparently due to permanent dipoles or mobile charges in the membrane can be measured by appropriate comparison of the transient currents required to produce small and large steps of membrane potential. Integration of these transient polarization currents estimates the charge transfer associated with the movement of membrane dipoles or charges. 2. Depolarization from -100 to 0 mV requires a charge transfer of 35 nC/muF in addition to the charge transfer predicted by linear extrapolation of the charge required for a small depolarization from -100 mV. Depolarizations of varying size give a charge-voltage relation which is sigmoid saturating beyond o mV and with a midpoint at about -50 mV. The ratnged depolarization reduces or removes charge movement detected by comparing currents for small and large voltage steps from -100 mV (Charge 1). However in depolarized fibres comparison of currents from a small potential step at +40 mV and a large hyperpolarizing potential step from -20 mV reveals large movements of a second charge (Charge 2). Movement of Charge 2 is less steeply dependent on voltage than movement of Charge 2 both in magnitude and in rate. 4. In size and voltage dependence these two kinds of charge movement correspond to measured voltage dependence of capacity in normally polarized and depolarized fibres (Adrian & Almers, 1976).


The poly(ADP-ribose)-dependent chromatin remodeler Alc1 induces local chromatin relaxation upon DNA damage.
Chromatin relaxation is one of the earliest cellular responses to DNA damage. However, what determines these structural changes, including their ATP requirement, is not well understood. Using live-cell imaging and laser microirradiation to induce DNA lesions, we show that the local chromatin relaxation at DNA damage sites is regulated by PARP1 enzymatic activity. We also report that H1 is mobilized at DNA damage sites, but, since this mobilization is largely independent of poly(ADP-ribosyl)ation, it cannot solely explain the chromatin relaxation. Finally, we demonstrate the involvement of Alc1, a poly(ADP-ribose)- and ATP-dependent remodeler, in the chromatin-relaxation process. Deletion of Alc1 impairs chromatin relaxation after DNA damage, while its overexpression strongly enhances relaxation. Altogether our results identify Alc1 as an important player in the fast kinetics of the NAD+- and ATP-dependent chromatin relaxation upon DNA damage in vivo.


Effects of Deep Brain Stimulation on Eye Movements and Vestibular Function.
Discovery of inter-latching circuits in the basal ganglia and invention of deep brain stimulation (DBS) for their modulation is a breakthrough in basic and clinical neuroscience. The DBS not only changes the quality of life of hundreds of thousands of people with intractable movement disorders, but it also offers a unique opportunity to understand how the basal ganglia interacts with other neural structures. An attractive yet less explored area is the study of DBS on eye movements and vestibular function. From the clinical perspective such studies provide valuable guidance in efficient programming of stimulation profile leading to optimal motor outcome. From the scientific standpoint such studies offer the ability to assess the outcomes of basal ganglia stimulation on eye movement behavior in cognitive as well as in motor domains. Understanding the influence of DBS on ocular motor function also leads to analogies to interpret its effects on complex appendicular and axial motor function. This review focuses on the influence of globus pallidus, subthalamic nucleus, and thalamus DBS on ocular motor and vestibular functions. The anatomy and physiology of basal ganglia, pertinent to the principles of DBS and ocular motility, is discussed. Interpretation of the effects of electrical stimulation of the basal ganglia in Parkinson's disease requires understanding of baseline ocular motor function in the diseased brain. Therefore we have also discussed the baseline ocular motor deficits in these patients and how the DBS changes such functions.


Effect of nandrolone decanoate administration on recovery from bupivacaine-induced muscle injury.
Although testosterone administration elicits well-documented anabolic effects on skeletal muscle mass, the enhancement of muscle regeneration after injury has not been widely examined. The purpose of this study was to determine whether anabolic steroid administration improves skeletal muscle regeneration from bupivacaine-induced injury. Male C57BL/6 mice were castrated 2 wk before muscle injury induced by an intramuscular bupivacaine injection into the tibialis anterior (TA) muscle. Control mice received an intramuscular PBS injection. Anabolic steroid [nandrolone decanoate (ND), 6 mg/kg] or sesame seed oil was administered at the time of initial injury and continued every 7 days for the study's duration. Mice were randomly assigned to one of four treatment groups for 5, 14, or 42 days of recovery, as follows: 1) control (uninjured); 2) ND only (uninjured + ND); 3) bupivacaine only (injured); or 4) bupivacaine + ND (injured + ND). TA morphology, protein, and gene expression were analyzed at 14 and 42 days after injury; protein expression was analyzed at 5 days after injury. After 14 days of recovery, the injury and injury + ND treatments induced small-diameter myofiber incidence and also decreased mean myofiber area. The increase in small-myofiber incidence was 65% greater in injury + ND muscle compared with injury alone. At 14 days, injury + ND induced a fivefold increase in muscle IGF-I mRNA expression, which was greater than injury alone. Muscle Akt activity and glycogen synthetase kinase-3beta activity were also induced by injury + ND at 14 days of recovery, but not by injury alone. ND had a main effect for increasing muscle MyoD and cyclin D1 mRNA expression at 14 days. After 42 days of recovery, injury + ND increased large-diameter myofiber incidence compared with injury only. Nandrolone decanoate (ND) administration can enhance castrated mouse muscle regeneration during the recovery from bupivacaine-induced injury.


Inclusion of minor alleles improves catalogue-based prediction of fluoroquinolone resistance in Mycobacterium tuberculosis.
Fluoroquinolone resistance poses a threat to the successful treatment of tuberculosis. WGS, and the subsequent detection of catalogued resistance-associated mutations, offers an attractive solution to fluoroquinolone susceptibility testing but sensitivities are often less than 90%. We hypothesize that this is partly because the bioinformatic pipelines used usually mask the recognition of minor alleles that have been implicated in fluoroquinolone resistance. We analysed the Comprehensive Resistance Prediction for Tuberculosis: an International Consortium (CRyPTIC) dataset of globally diverse WGS Mycobacterium tuberculosis isolates, with matched MICs for two fluoroquinolone drugs and allowed putative minor alleles to contribute to resistance prediction. Detecting minor alleles increased the sensitivity of WGS for moxifloxacin resistance prediction from 85.4% to 94.0%, without significantly reducing specificity. We also found no correlation between the proportion of an M. tuberculosis population containing a resistance-conferring allele and the magnitude of resistance. Together our results highlight the importance of detecting minor resistance-conferring alleles when using WGS, or indeed any sequencing-based approach, to diagnose fluoroquinolone resistance.


Review of Body Image and some of Contributing Factors in Iranian Population.
Body image is the perception of individuals of their own body and it can be affected by many variables such as culture, social pressure, and media, but it is not limited to these factors. Body image disturbances are important because they lead to severe physical or psychological health problems. Development of body image dissatisfaction in Eastern communities has been increased in the past years. Despite many studies in Asian societies, In Iran, studies are scattered and limited. Our goal is a better understanding of this matter in Iran. A review of scientific literature about Body Image and Iran was conducted inPubMed, Web of Science, Science Direct, Scopus, and ProQuest, also in Persian scientific databases such as Scientific Information Database (SID) and Thematic Guide to Iranian Publications (MAGIRAN). This search has been conducted in January 2018. Out of 389 articles, 44 of them were selected. In these articles, study period, sample characteristics, type of study, contributing variables, type of statistical analysis, and the main result of each article were extracted. Despite using different methods for detection of body dissatisfaction, results showed that this problem has a high prevalence in Iranian population. Contributing factors were also similar to the world findings. Review of studies regarding body dissatisfaction in Iran, revealed a high prevalence of this problem. It seems there is a need for interventional programs to prevent the negative consequences of body dissatisfaction, especially in teenagers and young people. Future studies with scientific or longitudinal design, which lead to better results, are recommended.


Framboidal ABC triblock copolymer vesicles: a new class of efficient Pickering emulsifier.
Pickering emulsions offer important advantages over conventional surfactant-stabilized emulsions, including enhanced long-term stability, more reproducible formulations and reduced foaming problems. The recent development of polymerization-induced self-assembly (PISA) offers considerable scope for the design of a wide range of block copolymer nanoparticles with tunable surface wettability that may serve as bespoke Pickering emulsifiers. In the present study, we exploit PISA to design a series of model framboidal ABC triblock copolymer vesicles with exquisite control over surface roughness. Transmission electron microscopy (TEM) and small-angle X-ray scattering (SAXS) were utilized to characterize these nanoparticles, which were subsequently used to stabilize n-dodecane emulsion droplets in water. The adsorption efficiency, Aeff, of the nanoparticles at the n-dodecane/water interface was determined as a function of increasing vesicle surface roughness using a turbidimetry assay. A strong correlation between surface roughness and Aeff was observed, with Aeff increasing from 36% up to 94%. This is a significant improvement in Pickering emulsifier efficiency compared to that reported previously for similar vesicles with smooth surfaces. In summary, nanoparticles with appreciable surface roughness are much more effective Pickering emulsifiers and this parameter can be readily fine-tuned using a highly efficient PISA formulation.


Purinergic regulation of acid/base transport in human and rat biliary epithelial cell lines.
Biliary epithelial cells (cholangiocytes) are responsible for rapid regulation of bile volume and alkalinity. Secretin and other hormones raising intracellular cyclic adenosine monophosphate (cAMP) concentrations promote biliary HCO3 secretion by stimulating apical Cl- channels and Cl-/HCO3- exchange (AE2). Cholangiocyte ion transport may also be stimulated by locally acting mediators; for example, adenosine 5'-triphosphate (ATP), a secretagogue that can be released into the bile by hepatocytes and cholangiocytes, activates Cl- conductances and Na+/H+ exchange (NHE) in cholangiocyte cell lines. To further explore the role of extracellular ATP in the paracrine regulation of carrier mechanisms regulating cholangiocyte H+/HCO3- secretion, we investigated the effects of nucleotides on intracellular pH regulation (measured by microfluorimetry with 2'7'-bis(2-carboxyethyl)-5,6,carboxyfluorescein [BCECF]) in human (MZ-ChA-1) and rat (NRC-1) cholangiocyte cell lines. In MZ-ChA-1 cells, 10 mol/L ATP, uridine 5'-triphosphate (UTP), and ATPgammas significantly increased NHE activity. The pharmacological profile of agonists was consistent with that anticipated for receptors of the P2Y2 class. ATP did not increase AE2 activity, but, when given to cells pretreated with agents raising intracellular cAMP, had a synergistic stimulatory effect that was inhibited by amiloride. To assess the polarity of purinergic receptors, monolayers of NRC-1 cells were exposed to apical or basolateral nucleotides. Apical administration of purinergic agonists, but not adenosine, increased basolateral NHE activity (ATPgammaS > UTP > ATP). Basolateral administration of purinergic agonists induced a weaker activation of NHE, which was instead strongly stimulated by adenosine and by adenosine receptor agonists (NECA = R-PIA = S-PIA). In conclusion, this study demonstrates that, consistent with the proposed role for biliary ATP in paracrine and autocrine control of cholangiocyte ion secretion, extracellular ATP stimulates cholangiocyte basolateral NHE activity through P2Y2 receptors that are predominantly expressed at the apical cell membrane.


Elevated neuron number in the limbic thalamus in major depression.
The mediodorsal and anteroventral/anteromedial nuclei of the thalamus are brain regions of interest in the study of mood disorders because they connect subcortical limbic system structures such as the amygdala with the prefrontal, cingulate, and temporal cortices. Anatomical abnormalities have been observed both in the amygdala and in the aforementioned cortical regions in affective disorder patients. Neuroanatomical studies of the thalamus have rarely been conducted in patients with mood disorders. Postmortem tissue from the Stanley Foundation Brain Bank was obtained from subjects diagnosed with major depressive disorder, bipolar disorder, and schizophrenia as well as a nonpsychiatric comparison group (N=10-13 per group). The optical disector stereological procedure was used to count neurons in the mediodorsal and anteroventral/anteromedial nuclei of the thalamus in each brain. There were significantly more neurons in the mediodorsal (37%) and anteroventral/anteromedial (26%) nuclei in subjects with major depressive disorder relative to the nonpsychiatric comparison subjects. Neuron numbers and volumes in these limbic thalamic nuclei were normal in the schizophrenia and bipolar subjects. The data indicate that there is an elevation in total neuron number in the limbic thalamus that is specific for major depressive disorder. This represents the first report of a neuropsychiatric disorder being associated with an increase in total regional neuron number. The present findings, along with recent data, indicate that significant anatomical and functional abnormalities are present in limbic circuits in major depressive disorder.


Calcitriol, the Bioactive Metabolite of Vitamin D, Increases Ventricular K+ Currents in Isolated Mouse Cardiomyocytes.
Calcitriol, the bioactive metabolite of vitamin D, interacts with the ubiquitously expressed nuclear vitamin D receptor (VDR) to induce genomic effects, but it can also elicit rapid responses via membrane-associated VDR through mechanisms that are poorly understood. The down-regulation of K+ currents is the main origin of electrophysiological remodeling in pathological hypertrophy and heart failure (HF), which can contribute to action potential prolongation and subsequently increase the risk of triggered arrhythmias. Adult mouse ventricular myocytes were isolated and treated with 10 nM calcitriol or vehicle for 15-30 min. In some experiments, cardiomyocytes were pretreated with the Akt inhibitor triciribine. In the adult mouse ventricle, outward K+ currents involved in cardiac repolarization are comprised of three components: the fast transient outward current (Itof), the ultrarapid delayed rectifier K+ current (Ikur), and the non-inactivating steady-state outward current (Iss). K+ currents were investigated using the whole-cell or the perforated patch-clamp technique and normalized to cell capacitance to obtain current densities. Calcitriol treatment of cardiomyocytes induced an increase in the density of Itof and Ikur, which was lost in myocytes isolated from VDR-knockout mice. In addition, calcitriol activated Akt in cardiomyocytes and pretreatment with triciribine prevented the calcitriol-induced increase of outward K+ currents. In conclusion, we demonstrate that calcitriol via VDR and Akt increases both Itof and Ikur densities in mouse ventricular cardiomyocytes. Our findings may provide new mechanistics clues for the cardioprotective role of this hormone in the heart.


Arsenic exposure and incidence of type 2 diabetes in Southwestern American Indians.
Association of urinary arsenic concentration with incident diabetes was examined in American Indians from Arizona who have a high prevalence of type 2 diabetes and were screened for diabetes between 1982 and 2007. The population resides where drinking water contains arsenic at concentrations above federally recommended limits. A total of 150 nondiabetic subjects aged ≥25 years who subsequently developed type 2 diabetes were matched by year of examination and sex to 150 controls who remained nondiabetic for ≥10 years. Total urinary arsenic concentration, adjusted for urinary creatinine level, ranged from 6.6 µg/L to 123.1 µg/L, and inorganic arsenic concentration ranged from 0.1 µg/L to 36.0 µg/L. In logistic regression models adjusted for age, sex, body mass index, and urinary creatinine level, the odds ratios for incident diabetes were 1.11 (95% confidence interval (CI): 0.79, 1.57) and 1.16 (95% CI: 0.89, 1.53) for a 2-fold increase in total arsenic and inorganic arsenic, respectively. Categorical analyses suggested a positive relationship between quartiles of inorganic arsenic and incident diabetes (P = 0.056); post-hoc comparison of quartiles 2-4 with quartile 1 revealed 2-fold higher odds of diabetes in the upper quartiles (OR = 2.14, 95% CI: 1.19, 3.85). Modestly elevated exposure to inorganic arsenic may predict type 2 diabetes in American Indians. Larger studies that include measures of speciated arsenic are required for confirmation.


Exploring the needs, concerns and behaviours of people with existing respiratory conditions in relation to the H1N1 'swine influenza' pandemic: a multicentre survey and qualitative study.
People with respiratory conditions are a 'high-risk' group for H1N1 pandemic swine influenza ('swine flu'), hence they and their families may have information needs, worries and concerns regarding the condition. Health-related behaviours, including vaccination, are recommended during the pandemic; understanding uptake of these is important. To explore and compare information needs, worries and concerns, and health-related behaviours regarding swine flu in people with respiratory conditions and their family members. Mixed-methods study - cross-sectional survey (253 patients, 101 family members); one-to-one interviews (13 patients, seven family members) and focus groups (n = three groups, 30 participants). Data collected October 2009-January 2010 from hospital chest clinics (n = 7) and patient support groups (n = 10) in North West England. Most patients (P) and family members (FM) wanted more information (n = 158, 62.5% P; n = 55, 54.4% FM), but few felt completely uninformed (n = 15, 5.9% P; n = 3, 3.0% FM). Most had already received information about swine flu (n = 187, 73.9% P; n = 78, 77.2% FM), mainly via a leaflet delivered to their home (n = 125, 49.4% P; n = 55, 54.5% FM). Information received was considered helpful (n = 154, 60.9% P; n = 77, 72.6% FM), but many wanted more condition-specific information (n = 141, 55.7% P; n = 60, 59.4% FM). More patients were worried (n = 147, 58.3%) than not worried (n = 99, 39.3%) about swine flu. FM were less often concerned about personal risk (n = 47, 46.6% worried) than about risk to patients (n = 76, 77.6%). Two-thirds (n = 161, 63.6% P; 65, 65.6% FM) incorrectly believed patients had increased risk of developing swine flu, but most (n = 204, 81.0% P; 89, 89.9% FM) correctly identified patients' greater risk of developing complications. Commonly adopted preventative measures were more frequent hand-washing (107, 42.8% P; 38, 37.6% FM) and greater use of sanitising hand gel (n = 100, 40.5% P; 37, 36.6% FM). In total, 212 patients (83.8%) and 69 family members (68.3%) were very/fairly likely to take up swine flu vaccination. Qualitative data mirrored survey findings. Participants were generally well-informed about swine flu, but more targeted information would have been welcomed. Participants were not highly anxious about swine flu, but did recognise risks for patients. Behaviour change was modest, but in line with recommendations. Vaccination intent was high. The study has been registered as REC/IRAS (Ref 09/H1015/76) and NIHR CSP (Ref 32483).


Fibroblast Growth Factor 23 and Incidence of Subarachnoid Hemorrhage: Nested Case-Control Study.
Fibroblast growth factor 23 (FGF23) is a hormone with regulatory effects on mineral metabolism and arterial calcification. We aimed to study the association between FGF23 and risk of incident subarachnoid hemorrhage (SAH) in a nested case-control study. FGF23 was analyzed in stored blood samples from incident SAH cases (n=79) and control subjects (n=232), participating in the prospective, population-based Malmö Diet and Cancer Study. The association between FGF23 and SAH was studied using conditional logistic regression. High FGF23 was associated with incident SAH. In the highest compared with the lowest quartile of FGF23, the odds ratio for SAH was 3.28 (95% confidence interval, 1.34-8.00), P for trend=0.006, after adjustment for SAH risk factors. High FGF23 was associated with increased risk of incident SAH in subjects from the general population. Further studies should elucidate whether FGF23 is a causal risk factor for SAH, or could be used in risk prediction.


Multivariate genome-wide association study of rapid automatised naming and rapid alternating stimulus in Hispanic American and African-American youth.
Rapid automatised naming (RAN) and rapid alternating stimulus (RAS) are reliable predictors of reading disability. The underlying biology of reading disability is poorly understood. However, the high correlation among RAN, RAS and reading could be attributable to shared genetic factors that contribute to common biological mechanisms. To identify shared genetic factors that contribute to RAN and RAS performance using a multivariate approach. We conducted a multivariate genome-wide association analysis of RAN Objects, RAN Letters and RAS Letters/Numbers in a sample of 1331 Hispanic American and African-American youth. Follow-up neuroimaging genetic analysis of cortical regions associated with reading ability in an independent sample and epigenetic examination of extant data predicting tissue-specific functionality in the brain were also conducted. Genome-wide significant effects were observed at rs1555839 (p=4.03×10-8) and replicated in an independent sample of 318 children of European ancestry. Epigenetic analysis and chromatin state models of the implicated 70 kb region of 10q23.31 support active transcription of the gene RNLS in the brain, which encodes a catecholamine metabolising protein. Chromatin contact maps of adult hippocampal tissue indicate a potential enhancer-promoter interaction regulating RNLS expression. Neuroimaging genetic analysis in an independent, multiethnic sample (n=690) showed that rs1555839 is associated with structural variation in the right inferior parietal lobule. This study provides support for a novel trait locus at chromosome 10q23.31 and proposes a potential gene-brain-behaviour relationship for targeted future functional analysis to understand underlying biological mechanisms for reading disability.


Cerebral palsy: an overview.
The presentation of cerebral palsy can be global mental and physical dysfunction or isolated disturbances in gait, cognition, growth, or sensation. It is the most common childhood physical disability and affects 2 to 2.5 children per 1,000 born in the United States. The differential diagnosis of cerebral palsy includes metabolic and genetic disorders. The goals of treatment are to improve functionality and capabilities toward independence. Multispecialty treatment teams should be developed around the needs of each patient to provide continuously updated global treatment care plans. Complications of cerebral palsy include spasticity and contractures; feeding difficulties; drooling; communication difficulties; osteopenia; osteoporosis; fractures; pain; and functional gastrointestinal abnormalities contributing to bowel obstruction, vomiting, and constipation. Valid and reliable assessment tools to establish baseline functions and monitor developmental gains have contributed to an increasing body of evidenced-based recommendations for cerebral palsy. Many of the historical treatments for this ailment are being challenged, and several new treatment modalities are available. Adult morbidity and mortality from ischemic heart disease, cerebrovascular disease, cancer, and trauma are higher in patients with cerebral palsy than in the general population.


Methanotrophic Bacterial Biomass as Potential Mineral Feed Ingredients for Animals.
Microorganisms play an important role in animal nutrition, as they can be used as a source of food or feed. The aim of the study was to determine the nutritional elements and fatty acids contained in the biomass of methanotrophic bacteria. Four bacterial consortia composed of Methylocystis and Methylosinus originating from Sphagnum flexuosum (Sp1), S. magellanicum (Sp2), S. fallax II (Sp3), S. magellanicum IV (Sp4), and one composed of Methylocaldum, Methylosinus, and Methylocystis that originated from coalbed rock (Sk108) were studied. Nutritional elements were determined using the flame atomic absorption spectroscopy technique after a biomass mineralization stage, whereas the fatty acid content was analyzed with the GC technique. Additionally, the growth of biomass and dynamics of methane consumption were monitored. It was found that the methanotrophic biomass contained high concentrations of K, Mg, and Fe, i.e., approx. 9.6-19.1, 2.2-7.6, and 2.4-6.6 g kg-1, respectively. Consequently, the biomass can be viewed as an appropriate feed and/or feed additive for supplementation with macroelements and certain microelements. Moreover, all consortia demonstrated higher content of unsaturated acids than saturated ones. Thus, methanotrophic bacteria seem to be a good solution, in natural supplementation of animal diets.


Relationships between Staphylococcus aureus genetic background, virulence factors, agr groups (alleles), and human disease.
The expression of most Staphylococcus aureus virulence factors is controlled by the agr locus, which encodes a two-component signaling pathway whose activating ligand is an agr-encoded autoinducing peptide (AIP). A polymorphism in the amino acid sequence of the AIP and of its corresponding receptor divides S. aureus strains into four major groups. Within a given group, each strain produces a peptide that can activate the agr response in the other member strains, whereas the AIPs belonging to different groups are usually mutually inhibitory. We investigated a possible relationship between agr groups and human S. aureus disease by studying 198 S. aureus strains isolated from 14 asymptomatic carriers, 66 patients with suppurative infection, and 114 patients with acute toxemia. The agr group and the distribution of 24 toxin genes were analyzed by PCR, and the genetic background was determined by means of amplified fragment length polymorphism (AFLP) analysis. The isolates were relatively evenly distributed among the four agrgroups, with 61 strains belonging to agr group I, 49 belonging to group II, 43 belonging to group III, and 45 belonging to group IV. Principal coordinate analysis performed on the AFLP distance matrix divided the 198 strains into three main phylogenetic groups, AF1 corresponding to strains of agr group IV, AF2 corresponding to strains of agr groups I and II, and AF3 corresponding to strains of agr group III. This indicated that the agr type was linked to the genetic background. A relationship between genetic background, agr group, and disease type was observed for several toxin-mediated diseases: for instance, agr group IV strains were associated with generalized exfoliative syndromes, and phylogenetic group AF1 strains with bullous impetigo. Among the suppurative infections, endocarditis strains mainly belonged to phylogenetic group AF2 and agr groups I and II. While these results do not show a direct role of the agr type in the type of human disease caused by S. aureus, the agr group may reflect an ancient evolutionary division of S. aureus in terms of this species' fundamental biology.


GNP2 Encodes a High-Specificity Proline Permease in Candida albicans.
The tight association of Candida albicans with the human host has driven the evolution of mechanisms that permit metabolic flexibility. Amino acids, present in a free or peptide-bound form, are abundant carbon and nitrogen sources in many host niches. In C. albicans, the capacity to utilize certain amino acids, like proline, is directly connected to fungal morphogenesis and virulence. Yet the precise nature of proline sensing and uptake in this pathogenic fungus has not been investigated. Since C. albicans encodes 10 putative orthologs of the four Saccharomyces cerevisiae proline transporters, we tested deletion strains of the respective genes and identified Gnp2 (CR_09920W) as the main C. albicans proline permease. In addition, we found that this specialization of Gnp2 was reflected in its transcriptional regulation and further assigned distinct substrate specificities for the other orthologs, indicating functional differences of the C. albicans amino acid permeases compared to the model yeast. The physiological relevance of proline uptake is exemplified by the findings that strains lacking GNP2 were unable to filament in response to extracellular proline and had a reduced capacity to damage macrophages and impaired survival following phagocytosis. Furthermore, GNP2 deletion rendered the cells more sensitive to oxidative stress, illustrating new connections between amino acid uptake and stress adaptation in C. albicans. IMPORTANCE The utilization of various nutrients is of paramount importance for the ability of Candida albicans to successfully colonize and infect diverse host niches. In this context, amino acids are of special interest due to their ubiquitous availability, relevance for fungal growth, and direct influence on virulence traits like filamentation. In this study, we identify a specialized proline transporter in C. albicans encoded by GNP2. The corresponding amino acid permease is essential for proline-induced filamentation, oxidative stress resistance, and fungal survival following interaction with macrophages. Altogether, this work highlights the importance of amino acid uptake for metabolic and stress adaptation in this fungus.


PI 3-kinase activation in BCR/abl-transformed hematopoietic cells does not require interaction of p85 SH2 domains with p210 BCR/abl.
BCR/abl is a chimeric oncogene implicated in the pathogenesis of human chronic myelogenous leukemia. Expression of the BCR/abl gene induces hematologic malignancies in transgenic mice and transformation of interleukin-3-dependent hematopoietic cells. The mechanism of BCR/abl-mediated transformation of hematopoietic cells is poorly understood and involves activation of at least two signaling pathways, p21ras and PI 3-kinase. Here we report that PI 3,4-P2 and PI 3,4,5-P3, the enzymatic products of PI 3-kinase, accumulate in metabolically labeled transformed hematopoietic cells, in contrast to our previous report on the lack of accumulation of PI 3-kinase products in nontransformed NIH 3T3 fibroblasts that express p210 BCR/abl. Transformed cells also have increased PI 3-kinase activity in total cell extracts and membrane fractions. Activation of PI 3-kinase occurs by occupancy of SH2 domains of PI 3-kinase regulatory subunit, p85, by phosphorylated YXXM motifs. Therefore, we investigated whether BCR/abl binds to p85 and whether this binding is mediated by interaction of p85 SH2 domains with YXXM motif of BCR/abl. Association of p210 BCR/abl with p85 in immune complexes and with p85 SH2 domains was evident in hematopoietic cells that express the wt p210 BCR/abl. However, the binding of BCR/abl to p85 SH2 domains was abolished in cells expressing mutant, temperature-sensitive (ts) p210 BCR/abl in which the tyrosine in the YXXM motif of p210 BCR/abl was replaced by histidine. Despite lack of direct interaction with p85 SH2 domains, expression of ts p210 BCR/abl resulted in rapid, time-dependent activation of total and membrane-associated PI 3-kinase and increased PI 3-kinase activity in anti-P-tyr and anti-abl immunoprecipitates. These data suggest that BCR/abl-induced activation of PI 3-kinase in hematopoietic cells does not require binding of p85 SH2 domains to BCR/abl gene product and involves interaction with other tyrosine phosphorylated intermediate proteins.


Transgenic expression of human complement regulatory proteins in mice results in diminished complement deposition during organ xenoperfusion.
Complement activation is an essential step in the hyperacute rejection of a vascularized xenograft. Endothelial cell-associated complement regulatory proteins limit complement activation in most settings, but are not able to limit the extensive complement activation that occurs in xenografts, at least in part due to their species specificity. To overcome this problem we and others have sought to express human complement regulatory proteins in the organs of potential donor animals. As an initial step toward evaluating this concept we tested organs from transgenic mice expressing human CD59 and/or decay-accelerating factor (DAF) in two in vitro perfusion systems for the ability to control activation of heterologous complement. In the first system, mouse hearts were perfused on a Langendorff circuit with 50% human plasma. Immunopathologic analysis of heart biopsies revealed deposition of human IgG, IgM, and C4 in both control and transgenic organs. The hearts from mice transgenic for human CD59 had substantially less and in some cases no membrane attack complex (MAC) and hearts from CD59/DAF transgenic mice had substantially less or no C5b and MAC. In the second system, mouse hearts were perfused with baboon blood through arterial lines inserted into baboons. Immunopathologic analysis of serial biopsies revealed the deposition of IgG, IgM, and C4 in control and transgenic hearts. Compared with controls, less MAC was deposited in many CD59-expressing hearts and less C5b and MAC in DAF-expressing hearts. These results demonstrate that human complement regulatory proteins expressed in a xenogeneic organ are able to contribute to the control of complement activation in that organ and support the concept that expression of these human molecules would help protect a xenogeneic organ transplanted into a human.


Protein folding in vivo revisited.
Protein folding in vivo is extremely intricate and challenging to examine or predict because the conformational changes, including folding, misfolding, and aggregation, are largely influenced by the cellular environment. Traditionally, cellular protein folding has been considered predominantly in the context of the Anfinsen postulate and molecular chaperones. However, accumulating evidence reveals that these models have limitations. In this review we revisit these models, and discuss co-translational folding, binding partner-mediated folding, and RNA-mediated folding as alternative or supplementary folding helpers. In addition, we discuss the folding helper systems mediated by macromolecules (e.g., ribosomes, membranes, and prefolded domains in multidomain proteins) that are tightly linked to newly synthesized polypeptides during protein biogenesis. These cis-acting folding helper systems, conceptually different from the trans-acting molecular chaperones, could play a crucial role in protein folding in vivo. Importantly, there is increasing evidence that the surface charges and excluded volume of macromolecules are important factors for stabilizing their connected polypeptides against aggregation. This stabilizing mechanism suggests that macromolecules including RNAs and proteins, let alone molecular chaperones, have an intrinsic ability to exert chaperoning function on their connected polypeptides independent of the linkage type between them. As an effective way to overcome the adverse effect of macromolecular crowding on protein folding, here we suggest that nascent polypeptide chains utilize the crowded environment in favor of productive folding by interacting with macromolecules.