CCAAT/Enhancer Binding Proteins Are Not Required for HIV-1 Entry but Regulate Proviral Transcription by Recruiting Coactivators to the Long-Terminal Repeat in Monocytic Cells

AbstractCCAAT/enhancer binding proteins (C/EBP) have been shown to be required for HIV-1 transcription and replication in macrophages. However, whether these transcription factors influence the ability of virus to establish infection by altering cytokine or receptor expression or primarily regulate HIV-1 transcription has not been determined. By inhibiting endogenous C/EBP activity with a dominant-negative protein, we demonstrate that functional C/EBPs are not required for HIV-1 infection and that these factors influence replication by a transcriptional mechanism. C/EBPβ recruits coactivators to the HIV-1 long-terminal repeat (LTR) and physically interacts with histone acetyltransferase (HAT) complexes, suggesting that C/EBPs participate in remodeling the chromatin organization of the HIV-1 provirus. Furthermore, overexpression of a C/EBP dominant-negative inhibits displacement of nucleosomes located at the HIV-1 transcriptional start site. These results provide insight into the general mechanisms by which C/EBPs regulate macrophage-restricted HIV-1 transcription

Hedgehog pathway dysregulation contributes to the pathogenesis of human gastrointestinal stromal tumors via GLI-mediated activation of KIT expression.

Gastrointestinal stromal tumors (GIST) arise within the interstitial cell of Cajal (ICC) lineage due to activating KIT/PDGFRA mutations. Both ICC and GIST possess primary cilia (PC), which coordinate PDGFRA and Hedgehog signaling, regulators of gastrointestinal mesenchymal development. Therefore, we hypothesized that Hedgehog signaling may be altered in human GIST and controls KIT expression. Quantitative RT-PCR, microarrays, and next generation sequencing were used to describe Hedgehog/PC-related genes in purified human ICC and GIST. Genetic and pharmacologic approaches were employed to investigate the effects of GLI manipulation on KIT expression and GIST cell viability. We report that Hedgehog pathway and PC components are expressed in ICC and GIST and subject to dysregulation during GIST oncogenesis, irrespective of KIT/PDGFRA mutation status. Using genomic profiling, 10.2% of 186 GIST studied had potentially deleterious genomic alterations in 5 Hedgehog-related genes analyzed, including in the PTCH1 tumor suppressor (1.6%). Expression of the predominantly repressive GLI isoform, GLI3, was inversely correlated with KIT mRNA levels in GIST cells and non-KIT/non-PDGFRA mutant GIST. Overexpression of the 83-kDa repressive form of GLI3 or small interfering RNA-mediated knockdown of the activating isoforms GLI1/2 reduced KIT mRNA. Treatment with GLI1/2 inhibitors, including arsenic trioxide, significantly increased GLI3 binding to the KIT promoter, decreased KIT expression, and reduced viability in imatinib-sensitive and imatinib-resistant GIST cells. These data offer new evidence that genes necessary for Hedgehog signaling and PC function in ICC are dysregulated in GIST. Hedgehog signaling activates KIT expression irrespective of mutation status, offering a novel approach to treat imatinib-resistant GIST

A randomized, controlled trial of endovenous thermal ablation using the 810-nm wavelength laser and the ClosurePLUS radiofrequency ablation methods for superficial venous insufficiency of the great saphenous vein

BackgroundGreat saphenous vein (GSV) incompetence is the most common cause of superficial venous insufficiency. Radiofrequency catheter ablation (RFA) is superior to conventional ligation and stripping, and endovenous laser treatment (EVL) has emerged as an effective alternative to RFA. This randomized study evaluated RFA and EVL for superficial venous insufficiency due to GSV incompetence and compared early and 1-year results.MethodsBetween June 2006 and May 2008, patients with symptomatic primary venous insufficiency due to GSV incompetence were randomized to RFA or EVL. Patients with bilateral disease were randomized for treatment of the first leg and received the alternative method on the other. Pretreatment examination included a leg assessment using the Venous Clinical Severity Score (VCSS) and CEAP classification. Patients completed the Chronic Venous Insufficiency Questionnaire 2 (CIVIQ2). RFA was performed with the ClosurePlus system (VNUS Medical Technologies, Sunnyvale, Calif). EVL was performed with the EVLT system (AngioDynamics Inc, Queensbury, NY). Early (1-week and 1-month) postoperative results of pain, bruising, erythema, and hematoma were recorded. Duplex ultrasound (DU) imaging was used at 1 week and 1 year to evaluate vein status. VCSS scores and CEAP clinical class were recorded at each postoperative visit, and quality of life (QOL) using CIVIQ2 was assessed at 1 month and 1 year.ResultsThe study enrolled 118 patients (141 limbs): 46 (39%) were randomized to RFA and 48 (40%) to EVL, and 24 (20%) had bilateral GSV incompetence. At 1 week, one patient in the RFA group had an open GSV and was deemed a failure. More bruising occurred in the EVL group (P = .01) at 1 week, but at 1 month, there was no difference in bruising between groups. At 1 year, DU imaging showed evidence of recanalization with reflux in 11 RFA and 2 EVL patients (P = .002). The mean VCSS score change from baseline to 1 week postprocedure was higher for RFA than EVL (P = .002), but there was no difference between groups at 1 month (P = .07) and 1 year (P = .9). Overall QOL mean score improved over time for all patients (P < .001). CEAP clinical class scores of ≥3 were recorded in 21 RFA (44%) and 24 EVL patients (44%) pretreatment, but at 1-year, 9 RFA (19%) and 12 EVL patients (24%) had scores of ≥3 (P < .001). This represented a significant improvement in all patients compared with baseline.ConclusionBoth methods of endovenous ablation effectively reduce symptoms of superficial venous insufficiency. EVL is associated with greater bruising and discomfort in the perioperative period but may provide a more secure closure over the long-term than RFA

Current recommendations on the selection of measures for well-being

Measures of well-being have proliferated over the past decades. Very little guidance has been available as to which measures to use in what contexts. This paper provides a series of recommendations, based on the present state of knowledge and the existing measures available, of what measures might be preferred in which contexts. The recommendations came out of an interdisciplinary workshop on the measurement of well-being. The recommendations are shaped around the number of items that can be included in a survey, and also based on the differing potential contexts and purposes of data collection such as, for example, government surveys, or multi-use cohort studies, or studies specifically about psychological well-being. The recommendations are not intended to be definitive, but to stimulate discussion and refinement, and to provide guidance to those relatively new to the study of well-being

Ammonia Emissions from U.S. Poultry Houses: Part II—Layer Houses

An ongoing project of monitoring ammonia (NH 3 ) emissions from U.S. layer houses with two different housing and management schemes is reported in this paper. Ammonia and carbon dioxide concentration levels were collected bi-weekly from each house with portable monitoring units (PMUs). There existed substantial seasonal variations in NH 3 emission from the layer houses. Specifically, daily mean NH 3 concentrations ranged from 1 to 7 ppm in the manure belt house and from 9 to 108 ppm in the high-rise house. The NH 3 emission rates averaged 6 mg/hr-hen or 44 g/d-500kg over a 12-month monitoring period (February 2002 – March 2003) for the manure belt house and 44 mg/h-hen or 331 g/d-500kg over a 10-month monitoring period (February – November 2002) for the high-rise house. Ammonia emission rates are higher in summer than in winter, although NH 3 concentration may be much lower in summer

Crystal structure of rhodopsin bound to arrestin by femtosecond X-ray laser.

G-protein-coupled receptors (GPCRs) signal primarily through G proteins or arrestins. Arrestin binding to GPCRs blocks G protein interaction and redirects signalling to numerous G-protein-independent pathways. Here we report the crystal structure of a constitutively active form of human rhodopsin bound to a pre-activated form of the mouse visual arrestin, determined by serial femtosecond X-ray laser crystallography. Together with extensive biochemical and mutagenesis data, the structure reveals an overall architecture of the rhodopsin-arrestin assembly in which rhodopsin uses distinct structural elements, including transmembrane helix 7 and helix 8, to recruit arrestin. Correspondingly, arrestin adopts the pre-activated conformation, with a ∼20° rotation between the amino and carboxy domains, which opens up a cleft in arrestin to accommodate a short helix formed by the second intracellular loop of rhodopsin. This structure provides a basis for understanding GPCR-mediated arrestin-biased signalling and demonstrates the power of X-ray lasers for advancing the frontiers of structural biology

Refined annotation and assembly of the Tetrahymena thermophila genome sequence through EST analysis, comparative genomic hybridization, and targeted gap closure

<p>Abstract</p> <p>Background</p> <p><it>Tetrahymena thermophila</it>, a widely studied model for cellular and molecular biology, is a binucleated single-celled organism with a germline micronucleus (MIC) and somatic macronucleus (MAC). The recent draft MAC genome assembly revealed low sequence repetitiveness, a result of the epigenetic removal of invasive DNA elements found only in the MIC genome. Such low repetitiveness makes complete closure of the MAC genome a feasible goal, which to achieve would require standard closure methods as well as removal of minor MIC contamination of the MAC genome assembly. Highly accurate preliminary annotation of <it>Tetrahymena</it>'s coding potential was hindered by the lack of both comparative genomic sequence information from close relatives and significant amounts of cDNA evidence, thus limiting the value of the genomic information and also leaving unanswered certain questions, such as the frequency of alternative splicing.</p> <p>Results</p> <p>We addressed the problem of MIC contamination using comparative genomic hybridization with purified MIC and MAC DNA probes against a whole genome oligonucleotide microarray, allowing the identification of 763 genome scaffolds likely to contain MIC-limited DNA sequences. We also employed standard genome closure methods to essentially finish over 60% of the MAC genome. For the improvement of annotation, we have sequenced and analyzed over 60,000 verified EST reads from a variety of cellular growth and development conditions. Using this EST evidence, a combination of automated and manual reannotation efforts led to updates that affect 16% of the current protein-coding gene models. By comparing EST abundance, many genes showing apparent differential expression between these conditions were identified. Rare instances of alternative splicing and uses of the non-standard amino acid selenocysteine were also identified.</p> <p>Conclusion</p> <p>We report here significant progress in genome closure and reannotation of <it>Tetrahymena thermophila</it>. Our experience to date suggests that complete closure of the MAC genome is attainable. Using the new EST evidence, automated and manual curation has resulted in substantial improvements to the over 24,000 gene models, which will be valuable to researchers studying this model organism as well as for comparative genomics purposes.</p

Integrated trajectories of the maternal metabolome, proteome, and immunome predict labor onset

Estimating the time of delivery is of high clinical importance because pre- and postterm deviations are associated with complications for the mother and her offspring. However, current estimations are inaccurate. As pregnancy progresses toward labor, major transitions occur in fetomaternal immune, metabolic, and endocrine systems that culminate in birth. The comprehensive characterization of maternal biology that precedes labor is key to understanding these physiological transitions and identifying predictive biomarkers of delivery. Here, a longitudinal study was conducted in 63 women who went into labor spontaneously. More than 7000 plasma analytes and peripheral immune cell responses were analyzed using untargeted mass spectrometry, aptamer-based proteomic technology, and single-cell mass cytometry in serial blood samples collected during the last 100 days of pregnancy. The high-dimensional dataset was integrated into a multiomic model that predicted the time to spontaneous labor [R = 0.85, 95% confidence interval (CI) [0.79 to 0.89], P = 1.2 × 10−40, N = 53, training set; R = 0.81, 95% CI [0.61 to 0.91], P = 3.9 × 10−7, N = 10, independent test set]. Coordinated alterations in maternal metabolome, proteome, and immunome marked a molecular shift from pregnancy maintenance to prelabor biology 2 to 4 weeks before delivery. A surge in steroid hormone metabolites and interleukin-1 receptor type 4 that preceded labor coincided with a switch from immune activation to regulation of inflammatory responses. Our study lays the groundwork for developing blood-based methods for predicting the day of labor, anchored in mechanisms shared in preterm and term pregnancies

Reliability and validity of two fitness tracker devices in the laboratory and home environment for older community-dwelling people

© 2018 The Author(s). Background: Two-thirds of older Australians are sedentary. Fitness trackers have been popular with younger people and may encourage older adults to become more active. Older adults may have different gait patterns and as such it is important to establish whether fitness trackers are valid and reliable for this population. The aim of the study was to test the reliability and validity of two fitness trackers (Fitbit Flex and ChargeHR) by step count when worn by older adults. Reliability and validity were tested in two conditions: 1) in the laboratory using a two-minute-walk-test (2MWT) and 2) in a free-living environment. Methods: Two 2MWTs were completed while wearing the fitness trackers. Participants were videoed during each test. Participants were then given one fitness tracker and a GENEactiv accelerometer to wear at home for 14-days. Results: Thirty-one participants completed two 2MWTs and 30 completed the free-living procedure. Intra Class Correlation's of the fitness trackers with direct observation of steps (criterion validity) was high (ICC:0.86,95%CI:0.76,0.93). However, both fitness trackers underestimated steps. Excellent test-retest reliability (ICC = 0.75) was found between the two 2MWTs for each device, particularly the ChargeHR devices. Good strength of agreement was found for total distance and steps (fitness tracker) and moderate-to-vigorous physical activity (GENEactiv) for the free-living environment (Spearman Rho's 0.78 and 0.74 respectively). Conclusion: Reliability and validity of the Flex and ChargeHR when worn by older adults is good, however both devices underestimated step count within the laboratory environment. These fitness trackers appear suitable for consumer use and promoting physical activity for older adults