Dinitrosyl-iron triggers apoptosis in Jurkat cells despite overexpression of Bcl-2

Cells expressing the cytokine-inducible NO synthase are known to trigger apoptosis in neighboring cells. Paramagnetic dinitrosyl nonheme iron complexes (DNIC) were found in tumor tissue about 40 years ago; however, the role of these NO(+)-bearing species is not completely understood. In the human Jurkat leukemia cell line, the application of the model complex DNIC-thiosulfate (50-200 microM) induced apoptosis (defined by phosphatidylserine externalization) in a concentration- and time-dependent manner. In Jurkat cells, the pan-caspase inhibitor, zVADfmk (50 microM), and/or stable transfection of antiapoptotic protein, Bcl-2, was unable to afford protection against DNIC-induced apoptosis. Th…

NFATc2 and NFATc3 transcription factors play a crucial role in suppression of CD4+ T lymphocytes by CD4+ CD25+ regulatory T cells

The phenotype of NFATc2(-/-) c3(-/-) (double knockout [DKO]) mice implies a disturbed regulation of T cell responses, evidenced by massive lymphadenopathy, splenomegaly, and autoaggressive phenomena. The population of CD4(+) CD25(+) T cells from DKO mice lacks regulatory capacity, except a small subpopulation that highly expresses glucocorticoid-induced tumor necrosis factor receptor family-related gene (GITR) and CD25. However, neither wild-type nor DKO CD4(+) CD25(+) regulatory T cells (T reg cells) are able to suppress proliferation of DKO CD4(+) CD25(-) T helper cells. Therefore, combined NFATc2/c3 deficiency is compatible with the development of CD4(+) CD25(+) T reg cells but renders c…

Observation of electroweak W±Z boson pair production in association with two jets in pp collisions at s=13 TeV with the ATLAS detector

An observation of electroweak W±Z production in association with two jets in proton–proton collisions is presented. The data collected by the ATLAS detector at the Large Hadron Collider in 2015 and 2016 at a centre-of-mass energy of s=13 TeV are used, corresponding to an integrated luminosity of 36.1fb−1 . Events containing three identified leptons, either electrons or muons, and two jets are selected. The electroweak production of W±Z bosons in association with two jets is measured with an observed significance of 5.3 standard deviations. A fiducial cross-section for electroweak production including interference effects and for a single leptonic decay mode is measured to be σWZjj−EW=0.57−0…

Search for light resonances decaying to boosted quark pairs and produced in association with a photon or a jet in proton–proton collisions at s=13TeV with the ATLAS detector

This Letter presents a search for new light resonances decaying to pairs of quarks and produced in association with a high- pT photon or jet. The dataset consists of proton–proton collisions with an integrated luminosity of 36.1 fb −1 at a centre-of-mass energy of s=13 TeV recorded by the ATLAS detector at the Large Hadron Collider. Resonance candidates are identified as massive large-radius jets with substructure consistent with a particle decaying into a quark pair. The mass spectrum of the candidates is examined for local excesses above background. No evidence of a new resonance is observed in the data, which are used to exclude the production of a lepto-phobic axial-vector Z′ boson.

Guidelines for the use of flow cytometry and cell sorting in immunological studies (second edition)

All authors: Andrea Cossarizza Hyun‐Dong Chang Andreas Radbruch Andreas Acs Dieter Adam Sabine Adam‐Klages William W. Agace Nima Aghaeepour Mübeccel Akdis Matthieu Allez Larissa Nogueira Almeida Giorgia Alvisi Graham Anderson Immanuel Andrä Francesco Annunziato Achille Anselmo Petra Bacher Cosima T. Baldari Sudipto Bari Vincenzo Barnaba Joana Barros‐Martins Luca Battistini Wolfgang Bauer Sabine Baumgart Nicole Baumgarth Dirk Baumjohann Bianka Baying Mary Bebawy Burkhard Becher Wolfgang Beisker Vladimir Benes Rudi Beyaert Alfonso Blanco Dominic A. Boardman Christian Bogdan Jessica G. Borger Giovanna Borsellino Philip E. Boulais Jolene A. Bradford Dirk Brenner Ryan R. Brinkman Anna E. S. Broo…

In vivo fate mapping with SCL regulatory elements identifies progenitors for primitive and definitive hematopoiesis in mice.

10 páginas, 6 figuras.-- et al.

Human CD25+ regulatory T cells: two subsets defined by the integrins alpha 4 beta 7 or alpha 4 beta 1 confer distinct suppressive properties upon CD4+ T helper cells.

Down-regulation of autoreactive T cell responses in vivo includes cell-contact-dependent as well as contact-independent mechanisms. Infectious tolerance is a contact-dependent mechanism used by naturally occurring CD25(+) T regulatory cells (Tregs) to confer suppressive activity upon conventional CD4(+) T cells thereby generating secondary T helper suppressor cells(Th(sup)), which inhibit T cell activation via soluble mediators. Here, we describe two distinct subsets of human Tregs, characterized by expression of either the alpha(4)beta(7) integrin or the alpha(4)beta(1) integrin. Upon activation, both subsets show an enhanced expression of FoxP3, recently described as a key transcription f…

Instruction of haematopoietic lineage choices, evolution of transcriptional landscapes and cancer stem cell hierarchies derived from an AML1-ETO mouse model.

The t(8;21) chromosomal translocation activates aberrant expression of the AML1-ETO (AE) fusion protein and is commonly associated with core binding factor acute myeloid leukaemia (CBF AML). Combining a conditional mouse model that closely resembles the slow evolution and the mosaic AE expression pattern of human t(8;21) CBF AML with global transcriptome sequencing, we find that disease progression was characterized by two principal pathogenic mechanisms. Initially, AE expression modified the lineage potential of haematopoietic stem cells (HSCs), resulting in the selective expansion of the myeloid compartment at the expense of normal erythro- and lymphopoiesis. This lineage skewing was foll…

Cutting Edge: TGF-β Signaling Is Required for the In Vivo Expansion and Immunosuppressive Capacity of Regulatory CD4+CD25+ T Cells

Abstract Data regarding the role of TGF-β for the in vivo function of regulatory CD4+CD25+ T cells (Treg) are controversial. A transgenic mouse model with impaired TGF-β signaling specifically in T cells was used to assess the role of endogenous TGF-β for the in vivo function of CD4+CD25+ Treg in a murine model of colitis induced by dextran sulfate. Transfer of wild-type, but not transgenic CD4+CD25+ Treg was found to suppress colitis in wild-type mice. In addition, by transferring CFSE-labeled CD4+CD25+ Treg we could demonstrate that endogenous TGF-β promotes the expansion of CD4+CD25+ Treg in vivo. Transgenic mice themselves developed reduced numbers of peripheral CD4+CD25+ Treg and were …

Lovastatin stimulates p75 TNF receptor (TNFR2) expression in primary human endothelial cells

HMG-CoA reductase inhibitors (statins) exert pleiotropic physiological effects. Among others they attenuate cellular responses to genotoxic and inflammatory stress. We investigated the effect of lovastatin on the expression level of TNF receptors (TNFR) in primary human endothelial cells (HUVEC). ELISA, FACS and immunocytochemical analyses show that lovastatin selectively increases the cell surface expression of TNFR2 without affecting the expression level of TNFR1. This effect of lovastatin is independent from inhibition of cell-cycle progression since cells both in G1- and G2-phase showed elevated levels of TNFR2 after lovastatin treatment. To analyze the physiological relevance of lovast…

Coexpression of TGF-β1 and IL-10 Enables Regulatory T Cells to Completely Suppress Airway Hyperreactivity

Abstract In allergic airway disease, Treg may play an important role in the modulation of airway hyperreactivity (AHR) and inflammation. We therefore investigated the therapeutic potential of Treg in an Ag-dependent murine asthma model. We here describe that AHR can be completely suppressed by adoptive transfer of Treg overexpressing active TGF-β1. Using mice with impaired TGF-β signaling in T cells, we could demonstrate that TGF-β signaling in recipient effector T cells or transferred Treg themselves is not required for the protective effects on AHR. However, the expression of IL-10 by Treg was found to be essential for the suppression of AHR, since Treg overexpressing active TGF-β1 but de…

Cyclic adenosine monophosphate is a key component of regulatory T cell–mediated suppression

Naturally occurring regulatory T cells (T reg cells) are a thymus-derived subset of T cells, which are crucial for the maintenance of peripheral tolerance by controlling potentially autoreactive T cells. However, the underlying molecular mechanisms of this strictly cell contact–dependent process are still elusive. Here we show that naturally occurring T reg cells harbor high levels of cyclic adenosine monophosphate (cAMP). This second messenger is known to be a potent inhibitor of proliferation and interleukin 2 synthesis in T cells. Upon coactivation with naturally occurring T reg cells the cAMP content of responder T cells is also strongly increased. Furthermore, we demonstrate that natur…

Generation and characterization of tTS-H4: a novel transcriptional repressor that is compatible with the reverse tetracycline-controlled TET-ON system

Background Conditional gene regulatory systems ensuring tight and adjustable expression of therapeutic genes are central for developing future gene therapy strategies. Among various regulatory systems, tetracycline-controlled gene expression has emerged as a safe and reliable option. Moreover, the tightness of tetracycline-regulated gene switches can be substantially improved by complementing transcriptional activators with antagonizing repressors. Methods To develop novel tetracycline-responsive transcriptional repressors, we fused various transcriptional silencing domains to the TetR (B/E) DNA-binding and dimerization domain of the Tn10-encoded tetracycline resistance operon (TetR (B/E)).…

Protective role of nuclear factor of activated T cells 2 in CD8+ long-lived memory T cells in an allergy model

Background The transcriptional regulation of cytokines released and controlled by memory T cells is not well understood. Defective IFN-γ production in allergic asthma correlates in human beings with the risk of wheezing in childhood. Objective To understand the role of the transcription factor nuclear factor of activated T cells 2 (NFATc2) in memory and effector T cells in the airways in experimental allergic asthma. Methods We used murine models of allergic asthma and adoptive cell transfer of fluorescence-activated sorted cells in a disease model. Results Mice lacking NFATc2 developed an increase in airway hyperresponsiveness (AHR), remodeling, and serum IgE levels on ovalbumin sensitizat…

Search for the Higgs boson decays H → ee and H → eμ in pp collisions at s=13TeV with the ATLAS detector

Searches for the Higgs boson decays H -> ee and H -> e mu are performed using data corresponding to an integrated luminosity of 139 fb(-1) collected with the ATLAS detector in pp collisions a ...

Search for invisible Higgs boson decays in vector boson fusion at s=13TeV with the ATLAS detector

We report a search for Higgs bosons that are produced via vector boson fusion and subsequently decay into invisible particles. The experimental signature is an energetic jet pair with invariant mass of O(1)TeV and O(100)GeV missing transverse momentum. The analysis uses 36.1 fb −1 of pp collision data at s=13TeV recorded by the ATLAS detector at the LHC. In the signal region the 2252 observed events are consistent with the background estimation. Assuming a 125GeV scalar particle with Standard Model cross sections, the upper limit on the branching fraction of the Higgs boson decay into invisible particles is 0.37 at 95% confidence level where 0.28 was expected. This limit is interpreted in H…

Toxicity of ligand-dependent Cre recombinases and generation of a conditional Cre deleter mouse allowing mosaic recombination in peripheral tissues.

Ligand-activated Cre recombinases are widely used for studying gene function in vitro and in conditional mouse models. To compare ligand-dependent Cre recombinases, different Cre estrogen receptor fusions were introduced into the ROSA26 locus of embryonic stem (ES) cells and assayed for genotoxicity and recombination efficiency. Of the tested recombinases, the CreERT2 variant showed no toxicity and was highly responsive to ligand induction. To constitutively express CreERT2 in mice and also to clarify whether the CreERT2 system displays background activity, we generated a knock-in mouse line harboring the CreERT2 coding region under the control of the ROSA26 locus. Analysis of this ROSA26-…

Evidence for the production of three massive vector bosons with the ATLAS detector

A search for the production of three massive vector bosons in proton–proton collisions is performed using data at TeV recorded with the ATLAS detector at the Large Hadron Collider in the years 2015–2017, corresponding to an integrated luminosity of 79.8 fb−1. Events with two same-sign leptons ℓ (electrons or muons) and at least two reconstructed jets are selected to search for . Events with three leptons without any same-flavour opposite-sign lepton pairs are used to search for , while events with three leptons and at least one same-flavour opposite-sign lepton pair and one or more reconstructed jets are used to search for . Finally, events with four leptons are analysed to search for and .…

Test of CP invariance in vector-boson fusion production of the Higgs boson in the H → ττ channel in proton–proton collisions at s=13TeV with the ATLAS detector

A test of CP invariance in Higgs boson production via vector-boson fusion is performed in the H → ττ decay channel. This test uses the Optimal Observable method and is carried out using 36.1 fb−1 of √s = 13 TeV proton–proton collision data collected by the ATLAS experiment at the LHC. Contributions from CP-violating interactions between the Higgs boson and electroweak gauge bosons are described by an effective field theory, in which the parameter ˜ d governs the strength of CP violation. No sign of CP violation is observed in the distributions of the Optimal Observable, and ˜ d is constrained to the interval [−0.090, 0.035] at the 68% confidence level (CL), compared to an expected interval …

Tetracycline-controlled transgenic targeting from the SCL locus directs conditional expression to erythrocytes, megakaryocytes, granulocytes, and c-kit-expressing lineage-negative hematopoietic cells

The stem cell leukemia gene SCL, also known as TAL-1, encodes a basic helix-loop-helix transcription factor expressed in erythroid, myeloid, megakaryocytic, and hematopoietic stem cells. To be able to make use of the unique tissue-restricted and spatio-temporal expression pattern of the SCL gene, we have generated a knock-in mouse line containing the tTA-2S tetracycline transactivator under the control of SCL regulatory elements. Analysis of this mouse using different tetracycline-dependent reporter strains demonstrated that switchable transgene expression was restricted to erythrocytes, megakaryocytes, granulocytes, and, importantly, to the c-kit-expressing and lineage-negative cell fracti…

Possible protective role for C-reactive protein in atherogenesis: complement activation by modified lipoproteins halts before detrimental terminal sequence.

Background—Previous work indicated that enzymatically remodeled LDL (E-LDL) might activate complement in atherosclerotic lesions via a C-reactive protein (CRP)–dependent and CRP-independent pathway. We sought to substantiate this contention and determine whether both pathways drive the sequence to completion.Methods and Results—E-LDL was prepared by sequential treatment of LDL with a protease and cholesteryl esterase. Trypsin, proteinase K, cathepsin H, or plasmin was used with similar results. Functional tests were used to assess total complement hemolytic activity, and immunoassays were used to demonstrate C3 cleavage and to quantify C3a, C4a, C5a, and C5b-9. E-LDL preparations activated …

P38 MAP Kinase Signaling Is Required for the Conversion of CD4+CD25− T Cells into iTreg

CD4+CD25+ regulatory T cells (Treg) are important mediators of immune tolerance. A subset of Treg can be generated in the periphery by TGF-beta dependent conversion of conventional CD4+CD25− T cells into induced Treg (iTreg). In chronic viral infection or malignancy, such induced iTreg, which limit the depletion of aberrant or infected cells, may be of pathogenic relevance. To identify potential targets for therapeutic intervention, we investigated the TGF-beta signaling in Treg. In contrast to conventional CD4+ T cells, Treg exhibited marked activation of the p38 MAP kinase pathway. Inhibition of p38 MAP kinase activity prevented the TGF-beta-dependent conversion of CD4+CD25− T cells into …

Differential Regulatory Capacity of CD25+ T Regulatory Cells and Preactivated CD25+ T Regulatory Cells on Development, Functional Activation, and Proliferation of Th2 Cells

Abstract CD25+ T regulatory (Treg) cells play a central role regarding the maintenance of peripheral tolerance via suppression of autoaggressive CD4+ T cells, CD8+ T cells, and Th1 cells. In this study we demonstrate that CD25+ Treg cells can also suppress the differentiation of murine conventional CD4+ T cells toward Th2 cells in a contact-dependent manner. However, the cytokine production and proliferation of established Th2 cells could not be inhibited by freshly isolated CD25+ Treg cells, whereas a strong inhibition of differentiated Th2 cells by in vitro preactivated CD25+ Treg cells could be observed. Inhibition of both conventional CD4+ T cells and Th2 cells is accompanied by a stron…

Measurement of prompt photon production in sNN=8.16 TeV p + Pb collisions with ATLAS

The inclusive production rates of isolated, prompt photons in p+Pb collisions at sNN=8.16 TeV are studied with the ATLAS detector at the Large Hadron Collider using a dataset with an integrated luminosity of 165 nb −1 recorded in 2016. The cross-section and nuclear modification factor RpPb are measured as a function of photon transverse energy from 20 GeV to 550 GeV and in three nucleon–nucleon centre-of-mass pseudorapidity regions, (−2.83,−2.02) , (−1.84,0.91) , and (1.09,1.90) . The cross-section and RpPb values are compared with the results of a next-to-leading-order perturbative QCD calculation, with and without nuclear parton distribution function modifications, and with expectations b…

Measurement of the production cross section for a Higgs boson in association with a vector boson in the H → WW⁎ → ℓνℓν channel in pp collisions at s=13 TeV with the ATLAS detector

A measurement of the Higgs boson production cross sections via associated WH and ZH production using H -> WW* -> l nu l nu decays, where l stands for either an electron or a muon, is presente ...

Guidelines for the use of flow cytometry and cell sorting in immunological studies

The marriage between immunology and cytometry is one of the most stable and productive in the recent history of science. A rapid search in PubMed shows that, as of July 2017, using “flow cytometry immunology” as a search term yields more than 68 000 articles, the first of which, interestingly, is not about lymphocytes. It might be stated that, after a short engagement, the exchange of the wedding rings between immunology and cytometry officially occurred when the idea to link fluorochromes to monoclonal antibodies came about. After this, recognizing different types of cells became relatively easy and feasible not only by using a simple fluorescence microscope, but also by a complex and some…

Search for low-mass resonances decaying into two jets and produced in association with a photon using pp collisions at s=13 TeV with the ATLAS detector

A search is performed for localised excesses in dijet mass distributions of low-dijet-mass events produced in association with a high transverse energy photon. The search uses up to 79.8 fb−1 of LHC proton–proton collisions collected by the ATLAS experiment at a centre-of-mass energy of 13 TeV during 2015–2017. Two variants are presented: one which makes no jet flavour requirements and one which requires both jets to be tagged as b-jets. The observed mass distributions are consistent with multi-jet processes in the Standard Model. The data are used to set upper limits on the production cross-section for a benchmark Z′ model and, separately, on generic Gaussian-shape contributions to the mas…

17. Mainzer Allergie-Workshop