The rockefeller university

An integrin found on platelets, alpha(IIb)beta(3) mediates platelet aggregation, and alpha(IIb)beta(3) antagonists are effective antithrombotic agents in the clinic. Ligands bind to integrins in part by coordinating a magnesium ion (Mg2+) located in the beta subunit metal ion-dependent adhesion site (MIDAS). Drugs patterned on the integrin ligand sequence Arg-Gly-Asp have a basic moiety that binds the alpha(IIb) subunit and a carboxyl group that coordinates the MIDAS Mg2+ in the beta(3) subunits. They induce conformational changes in the beta(3) subunit that may have negative consequences such as exposing previously hidden epitopes and inducing the active conformation of the receptor. We recently reported an inhibitor of alpha(IIb)beta(3) (RUC-1) that binds exclusively to the alpha(IIb) subunit; here, we report the structure-based design and synthesis of RUC-2, a RUC-1 derivative with a similar to 100-fold higher affinity. RUC-2 does not induce major conformational changes in beta(3) as judged by monoclonal antibody binding, light scattering, gel chromatography, electron microscopy, and a receptor priming assay. X-ray crystallography of the RUC-2-alpha(IIb)beta(3) headpiece complex in 1 mM calcium ion (Ca2+)/5 mM Mg2+ at 2.6 angstrom revealed that RUC-2 binds to aIIb the way RUC-1 does, but in addition, it binds to the beta(3) MIDAS residue glutamic acid 220, thus displacing Mg2+ from the MIDAS. When the Mg2+ concentration was increased to 20 mM, however, Mg2+ was identified in the MIDAS and RUC-2 was absent. RUC-2's ability to inhibit ligand binding and platelet aggregation was diminished by increasing the Mg2+ concentration. Thus, RUC-2 inhibits ligand binding by a mechanism different from that of all other alpha(IIb)beta(3) antagonists and may offer advantages as a therapeutic agent.

Exchange of macromolecules between the nucleus and cytoplasm is a key regulatory event in the expression of a cell's genome. This exchange requires a dedicated transport system: (1) nuclear pore complexes (NPCs), embedded in the nuclear envelope and composed of proteins termed nucleoporins (or "Nups"), and (2) nuclear transport factors that recognize the cargoes to be transported and ferry them across the NPCs. This transport is regulated at multiple levels, and the NPC itself also plays a key regulatory role in gene expression by influencing nuclear architecture and acting as a point of control for various nuclear processes. Here we summarize how the yeast Saccharomyces has been used extensively as a model system to understand the fundamental and highly conserved features of this transport system, revealing the structure and function of the NPC; the NPC's role in the regulation of gene expression; and the interactions of transport factors with their cargoes, regulatory factors, and specific nucleoporins.

A search is reported for the standard model Higgs boson in the H -> ZZ -> l(+)l(-)tau(+)tau(-) decay mode, where l = mu or e, in proton-proton collisions at root s = 7 TeV, corresponding to an integrated luminosity of 4.7 fb(-1) collected with the CMS detector at the LHC. No evidence is found for a significant deviation from the background expectation. An upper limit four to twelve times larger than the predicted value is set at 95% confidence level for the product of the standard model Higgs boson production cross section and decay branching fraction in the mass range 190 < m(H) < 600 GeV.

DEC205/CD205 is a C-type multilectin receptor, expressed highly in dendritic cells (DCs). Previous efforts to generate anti-human DEC205 (anti-hDEC205) monoclonal antibodies (mAbs) from mice immunized with subdomain proteins of hDEC205 resulted in a few mAbs. Recently, we expressed and utilized a full-length extracellular domain protein of hDEC205 to successfully generate 5 strong anti-hDEC205 mAbs from mice. In this study, DEC205 knockout (KO) mice were immunized with this full-length extracellular domain protein of hDEC205. One of the 3 immunized DEC205 KO mice was chosen for the highest anti-hDEC205 titer by flow cytometric analysis of serum samples on CHO cells stably expressing hDEC205 (CHO/hDEC205 cells) and used for hybridoma fusion. From a single fusion, more than 400 anti-hDEC205 hybridomas were identified by flow cytometric screen with CHO/hDEC205 cells, and a total of 115 hybridomas secreting strong anti-hDEC205 mAb were saved and named HD1 through HD115. To characterize in detail, 10 HD mAbs were chosen for superior anti-hDEC205 reactivity and further subjected to cloning and purification. Interestingly, out of those 10 chosen anti-hDEC205 HD mAbs, 5 mAbs were also strongly reactive to mouse DEC205 while 8 mAbs were found to stain DEC205(+) DCs on monkey spleen sections. In addition, we also identified that HD83, one of the 10 chosen HD mAbs, stains DEC205(+) DCs in rat spleen and lymph node. Therefore, by immunizing DEC205 KO mice with a full-length extracellular domain protein of hDEC205, we generated a large number of strong anti-hDEC205 mAbs many of which are cross-species reactive and able to visualize DEC205(+) DCs in lymphoid tissues of other mammals. (C) 2012 Elsevier B.V. All rights reserved.

Adult-onset diseases can be associated with in utero events, but mechanisms for this remain unknown(1,2). The Polycomb histone methyltransferase Ezh2 stabilizes transcription by depositing repressive marks during development that persist into adulthood(3-9), but its function in postnatal organ homeostasis is unknown. We show that Ezh2 stabilizes cardiac gene expression and prevents cardiac pathology by repressing the homeodomain transcription factor gene Six1, which functions in cardiac progenitor cells but is stably silenced upon cardiac differentiation(10). Deletion of Ezh2 in cardiac progenitors caused postnatal myocardial pathology and destabilized cardiac gene expression with activation of Six1-dependent skeletal muscle genes. Six1 induced cardiomyocyte hypertrophy and skeletal muscle gene expression. Furthermore, genetically reducing Six1 levels rescued the pathology of Ezh2-deficient hearts. Thus, Ezh2-mediated repression of Six1 in differentiating cardiac progenitors is essential for stable gene expression and homeostasis in the postnatal heart. Our results suggest that epigenetic dysregulation in embryonic progenitor cells is a predisposing factor for adult disease and dysregulated stress responses.

Background In this phase 2, randomized, double-blind, placebo-controlled, dose-ranging study, we assessed the efficacy and safety of brodalumab (AMG 827), a human anti-interleukin-17-receptor monoclonal antibody, for the treatment of moderate-to-severe plaque psoriasis. Methods We randomly assigned patients with a score of 12 or higher on the psoriasis area-and-severity index (PASI, on which scores range from 0 to 72, with higher scores indicating more severe disease) and with 10% or more of their body-surface area affected by psoriasis to receive brodalumab (70 mg, 140 mg, or 210 mg at day 1 and weeks 1, 2, 4, 6, 8, and 10 or 280 mg monthly) or placebo. The primary end point was the percentage improvement from baseline in the PASI score at week 12. Secondary end points included improvement of at least 75% and at least 90% in the PASI score and the score on the static physician's global assessment at week 12. Results A total of 198 patients underwent randomization. At week 12, the mean percentage improvements in the PASI score were 45.0% among patients receiving 70 mg of brodalumab, 85.9% among those receiving 140 mg, 86.3% among those receiving 210 mg, 76.0% among those receiving 280 mg, and 16.0% among those receiving placebo (P<0.001 for all comparisons with placebo). An improvement of at least 75% and at least 90% in the PASI score at week 12 was seen in 77% and 72%, respectively, of the patients in the 140-mg brodalumab group and in 82% and 75%, respectively, of the patients in the 210-mg group, as compared with 0% in the placebo group (P<0.001 for all comparisons). The percentage of patients with a static physician's global assessment of clear or minimal disease was 26%, 85%, 80%, and 69% with the 70-mg, 140-mg, 210-mg, and 280-mg doses, respectively, of brodalumab, as compared with 3% with placebo (P<0.01 for all comparisons with placebo). Two cases of grade 3 neutropenia were reported in the 210-mg brodalumab group. The most commonly reported adverse events in the combined brodalumab groups were nasopharyngitis (8%), upper respiratory tract infection (8%), and injection-site erythema (6%). Conclusions Brodalumab significantly improved plaque psoriasis in this 12-week, phase 2 study. (Funded by Amgen; ClinicalTrials.gov number, NCT00975637.)

BACKGROUND Antihypertensive beta-blocker use is associated with greater intervisit blood pressure variability (BPV) and with less favorable outcomes compared to other antihypertensive agents. A theoretical model demonstrated that accuracy and precision of BP measurement are affected by heart rate (HR) at a constant cuff deflation rate. We aimed to examine the empirical relationship between HR and BPV in a clinical setting. METHODS Intratracing variability in ambulatory BP monitoring (ABPM) were analyzed in search of a link between BPV and HR. BPV was expressed as standard deviation (s.d.), coefficient of variation (CV), and variability independent of the mean (VIM). RESULTS In a dataset of 4,693 subjects, HR was inversely associated with BPV and independently explained 1.3% of between-subject variation in s.d. of awake systolic BP (1.5% of CV and VIM). Linear regression suggested 0.5 mm Hg increase in s.d. of systolic BP per 10 beats per minute (bpm) decrease in HR. In a subset of 1,019 patients with available data on medications, HR was independently and inversely related with awake systolic BPV (P < 0.0001), more so in diuretic (P < 0.050) and renin-angiotensin system antagonists-treated (P < 0.050) patients. Associations of beta-blockade with increased BPV were abolished by model-adjustment for HR. In another subset of patients who were monitored twice (n = 635), HR had a mild (0.6%) but significant (P < 0.05) inverse contribution to the change in awake systolic BPV between repeated monitoring. CONCLUSIONS Ambulatory BPV is inversely related to HR and is not increased in referred patients treated with beta-blockers after correction for HR.

We present a search for the standard model Higgs boson production in association with a W boson in proton-antiproton collisions (p (p) over bar -> W+ H -> lvb (b) over bar) at a center of mass energy of 1.96 TeV. The search employs data collected with the CDF II detector which correspond to an integrated luminosity of approximately 2.7 fb(-1). We recorded this data with two kinds of triggers. The first kind required high-p(T) charged leptons and the second required both missing transverse energy and jets. The search selects events consistent with a signature of a single lepton (e(+/-)/mu(+/-)),missing transverse energy, and two jets. Jets corresponding to bottom quarks are identified with a secondary vertex tagging method and a jet probability tagging method. Kinematic information is fed in an artificial neural network to improve discrimination between signal and background. The search finds that both the observed number of events and the neural network output distributions are consistent with the standard model background expectations, and sets 95% confidence level upper limits on the production cross section times branching ratio. The limits are expressed as a ratio to the standard model production rate. The limits range from 3.6 (4.3 expected) to 61.1 (43.2 expected) for Higgs masses from 100 to 150 GeV/c(2), respectively.

Limbic endocannabinoid signaling is known to be sensitive to chronic stress; however, studies investigating the impact of prolonged exposure to glucocorticoid hormones have been limited by the concurrent exposure to the stress of daily injections. The present study was designed to examine the effects of a noninvasive approach to alter plasma corticosterone (CORT) on the endocannabinoid system. More precisely, we explored the effects of a 4-week exposure to CORT dissolved in the drinking water of mice (100 mu g/ml) and measured cannabinoid CB1 receptor binding, endocannabinoid content, activity of the endocannabinoid degrading enzyme fatty acid amide hydrolase (FAAH), and mRNA expression of both the CB1 receptor and FAAH in both the hippocampus and amygdala. Our data demonstrate that CORT decreases CB1 receptor binding site density in both the hippocampus and amygdala and also reduced anandamide (AEA) content and increased FAAH activity within both structures. These changes in both CB1 receptor binding and FAAH activity were not accompanied by changes in mRNA expression of either the CB1 receptor or FAAH in either brain region. Interestingly, our CORT delivery regimen significantly increased 2-AG concentrations within the hippocampus, but not the amygdala. Collectively, these data demonstrate that the confounder of injection stress is sufficient to conceal the ability of protracted exposure to glucocorticoids to reduce CB1 receptor density and augment AEA metabolism within limbic structures. This article is part of a Special Issue entitled: Stress, Emotional Behavior and the Endocannabinoid System. (C) 2011 IBRO. Published by Elsevier Ltd. All rights reserved.

Natural killer (NK) cells are circulating cytotoxic lymphocytes that exert potent and nonredundant antiviral activity and antitumoral activity in the mouse; however, their function in host defense in humans remains unclear. Here, we investigated 6 related patients with autosomal recessive growth retardation, adrenal insufficiency, and a selective NK cell deficiency characterized by a lack of the CD56(dim) NK subset. Using linkage analysis and fine mapping, we identified the disease-causing gene, MCM4, which encodes a component of the MCM2-7 helicase complex required for DNA replication. A splice-site mutation in the patients produced a frameshift, but the mutation was hypomorphic due to the creation of two new translation initiation methionine codons downstream of the premature termination codon. The patients' fibroblasts exhibited genomic instability, which was rescued by expression of WT MCM4. These data indicate that the patients' growth retardation and adrenal insufficiency likely reflect the ubiquitous but heterogeneous impact of the MCM4 mutation in various tissues. In addition, the specific loss of the NK CD56(dim) subset in patients was associated with a lower rate of NK CD56(bright) cell proliferation, and the maturation of NK CD56(bright) cells toward an NK CD56(dim) phenotype was tightly dependent on MCM4-dependent cell division. Thus, partial MCM4 deficiency results in a genetic syndrome of growth retardation with adrenal insufficiency and selective NK deficiency.