65 research outputs found
Endoglin potentiates nitric oxide synthesis to enhance definitive hematopoiesis.
During embryonic development, hematopoietic cells develop by a process of endothelial-to hematopoietic transition of a specialized population of endothelial cells. These hemogenic endothelium (HE) cells in turn develop from a primitive population of FLK1(+) mesodermal cells. Endoglin (ENG) is an accessory TGF-β receptor that is enriched on the surface of endothelial and hematopoietic stem cells and is also required for the normal development of hemogenic precursors. However, the functional role of ENG during the transition of FLK1(+) mesoderm to hematopoietic cells is ill defined. To address this we used a murine embryonic stem cell model that has been shown to mirror the temporal emergence of these cells in the embryo. We noted that FLK1(+) mesodermal cells expressing ENG generated fewer blast colony-forming cells but had increased hemogenic potential when compared with ENG non-expressing cells. TIE2(+)/CD117(+) HE cells expressing ENG also showed increased hemogenic potential compared with non-expressing cells. To evaluate whether high ENG expression accelerates hematopoiesis, we generated an inducible ENG expressing ES cell line and forced expression in FLK1(+) mesodermal or TIE2(+)/CD117(+) HE cells. High ENG expression at both stages accelerated the emergence of CD45(+) definitive hematopoietic cells. High ENG expression was associated with increased pSMAD2/eNOS expression and NO synthesis in hemogenic precursors. Inhibition of eNOS blunted the ENG induced increase in definitive hematopoiesis. Taken together, these data show that ENG potentiates the emergence of definitive hematopoietic cells by modulating TGF-β/pSMAD2 signalling and increasing eNOS/NO synthesis.The authors thank Dr Zúñiga-Pflücker (University of Toronto) for the ENG-/- and +/- murine ES cells. This work was supported by grants from the National Health and Medical Research Council of Australia, Australian Research Council and the Dr Tom Bee Stem Cell Research Fund to JEP, Cancer Research UK to VK and GL and the BBSRC, Leukaemia and Lymphoma Research, The Leukaemia and Lymphoma Society, Cancer Research UK, and core support grants by the Wellcome Trust to the Cambridge Institute for Medical Research and Wellcome Trust - MRC Cambridge Stem Cell Institute to BG.This is the final version of the article. It first appeared from the Company of Biologists via http://dx.doi.org/10.1242/​bio.01149
Integrative Genomics Identifies the Molecular Basis of Resistance to Azacitidine Therapy in Myelodysplastic Syndromes
Myelodysplastic syndromes and chronic myelomonocytic leukemia are blood disorders characterized by ineffective hematopoiesis and progressive marrow failure that can transform into acute leukemia. The DNA methyltransferase inhibitor 5-azacytidine (AZA) is the most effective pharmacological option, but only ∼50% of patients respond. A response only manifests after many months of treatment and is transient. The reasons underlying AZA resistance are unknown, and few alternatives exist for non-responders. Here, we show that AZA responders have more hematopoietic progenitor cells (HPCs) in the cell cycle. Non-responder HPC quiescence is mediated by integrin α5 (ITGA5) signaling and their hematopoietic potential improved by combining AZA with an ITGA5 inhibitor. AZA response is associated with the induction of an inflammatory response in HPCs in vivo. By molecular bar coding and tracking individual clones, we found that, although AZA alters the sub-clonal contribution to different lineages, founder clones are not eliminated and continue to drive hematopoiesis even in complete responders.The authors acknowledge funding from the National Health and
Medical Research Council (NHMRC), Leukaemia Foundation, Anthony Rothe
Foundation, Cancer Institute for New South Wales, South Eastern Area Laboratory Services (SEALS), Wellcome Trust, Leukemia and Lymphoma Society,
Medical Research Council (UK), Swedish Cancer Society, Cancer Society in
Stockholm, Swedish Research Council, Bloodwise UK, and the NIHR Biomedical Research Centre, Oxford
DKC1 is a transcriptional target of GATA1 and drives upregulation of telomerase activity in normal human erythroblasts
Telomerase is a ribonucleoprotein complex that maintains the length and integrity of telomeres, and thereby enables cellular proliferation. Understanding the regulation of telomerase in hematopoietic cells is relevant to the pathogenesis of leukemia, in which telomerase is constitutively activated, as well as bone marrow failure syndromes that feature telomerase insufficiency. Past studies showing high levels of telomerase in human erythroblasts and a prevalence of anemia in disorders of telomerase insufficiency provide the rationale for investigating telomerase regulation in erythroid cells. Here it is shown for the first time that the telomerase RNA-binding protein dyskerin (encoded by DKC1) is dramatically upregulated as human hematopoietic stem and progenitor cells commit to the erythroid lineage, driving an increase in telomerase activity in the presence of limiting amounts of TERT mRNA. It is also shown that upregulation of DKC1 was necessary for expansion of glycophorin A+ erythroblasts and sufficient to extend telomeres in erythroleukemia cells. Chromatin immunoprecipitation and reporter assays implicated GATA1-mediated transcriptional regulation of DKC1 in the modulation of telomerase in erythroid lineage cells. Together these results describe a novel mechanism of telomerase regulation in erythroid cells which contrasts with mechanisms centered on transcriptional regulation of TERT that are known to operate in other cell types. This is the first study to reveal a biological context in which telomerase is upregulated by DKC1 and to implicate GATA1 in telomerase regulation. The results from this study are relevant to hematopoietic disorders involving DKC1 mutations, GATA1 deregulation and/or telomerase insufficiency
HBO1 is required for the maintenance of leukaemia stem cells.
Acute myeloid leukaemia (AML) is a heterogeneous disease characterized by transcriptional dysregulation that results in a block in differentiation and increased malignant self-renewal. Various epigenetic therapies aimed at reversing these hallmarks of AML have progressed into clinical trials, but most show only modest efficacy owing to an inability to effectively eradicate leukaemia stem cells (LSCs)1. Here, to specifically identify novel dependencies in LSCs, we screened a bespoke library of small hairpin RNAs that target chromatin regulators in a unique ex vivo mouse model of LSCs. We identify the MYST acetyltransferase HBO1 (also known as KAT7 or MYST2) and several known members of the HBO1 protein complex as critical regulators of LSC maintenance. Using CRISPR domain screening and quantitative mass spectrometry, we identified the histone acetyltransferase domain of HBO1 as being essential in the acetylation of histone H3 at K14. H3 acetylated at K14 (H3K14ac) facilitates the processivity of RNA polymerase II to maintain the high expression of key genes (including Hoxa9 and Hoxa10) that help to sustain the functional properties of LSCs. To leverage this dependency therapeutically, we developed a highly potent small-molecule inhibitor of HBO1 and demonstrate its mode of activity as a competitive analogue of acetyl-CoA. Inhibition of HBO1 phenocopied our genetic data and showed efficacy in a broad range of human cell lines and primary AML cells from patients. These biological, structural and chemical insights into a therapeutic target in AML will enable the clinical translation of these findings
Identification of novel regulators of developmental hematopoiesis using Endoglin regulatory elements as molecular probes.
Enhancers are the primary determinants of cell identity, and specific promoter/enhancer combinations of Endoglin (ENG) have been shown to target blood and endothelium in the embryo. Here, we generated a series of embryonic stem cell lines, each targeted with reporter constructs driven by specific promoter/enhancer combinations of ENG, to evaluate their discriminative potential and value as molecular probes of the corresponding transcriptome. The Eng promoter (P) in combination with the -8/+7/+9-kb enhancers, targeted cells in FLK1 mesoderm that were enriched for blast colony forming potential, whereas the P/-8-kb enhancer targeted TIE2+/c-KIT+/CD41- endothelial cells that were enriched for hematopoietic potential. These fractions were isolated using reporter expression and their transcriptomes profiled by RNA-seq. There was high concordance between our signatures and those from embryos with defects at corresponding stages of hematopoiesis. Of the 6 genes that were upregulated in both hemogenic mesoderm and hemogenic endothelial fractions targeted by the reporters, LRP2, a multiligand receptor, was the only gene that had not previously been associated with hematopoiesis. We show that LRP2 is indeed involved in definitive hematopoiesis and by doing so validate the use of reporter gene-coupled enhancers as probes to gain insights into transcriptional changes that facilitate cell fate transitions.National Health and Medical Research Council of Australia, Australian Research Council, Dr Tom Bee Stem Cell Research Fund, Cancer Research UK, Biotechnology and Biological Sciences Research Council, Leukaemia and Lymphoma Research, The Leukaemia and Lymphoma Society, core support grants by the Wellcome Trust to the Cambridge Institute for Medical Research and Wellcome Trust - MRC Cambridge Stem Cell Institute (Grant IDs: R01 HL04880, P015PO1HL32262-32, 5P30 DK49216, 5R01 DK53298, 5U01 HL10001-05, R24 DK092760)This is the author accepted manuscript. The final version is available from the American Society of Hematology via http://dx.doi.org/10.1182/blood-2016-02-69787
Anticoagulant selection in relation to the SAMe-TT2R2 score in patients with atrial fibrillation. the GLORIA-AF registry
Aim: The SAMe-TT2R2 score helps identify patients with atrial fibrillation (AF) likely to have poor anticoagulation control during anticoagulation with vitamin K antagonists (VKA) and those with scores >2 might be better managed with a target-specific oral anticoagulant (NOAC). We hypothesized that in clinical practice, VKAs may be prescribed less frequently to patients with AF and SAMe-TT2R2 scores >2 than to patients with lower scores. Methods and results: We analyzed the Phase III dataset of the Global Registry on Long-Term Oral Antithrombotic Treatment in Patients with Atrial Fibrillation (GLORIA-AF), a large, global, prospective global registry of patients with newly diagnosed AF and ≥1 stroke risk factor. We compared baseline clinical characteristics and antithrombotic prescriptions to determine the probability of the VKA prescription among anticoagulated patients with the baseline SAMe-TT2R2 score >2 and ≤ 2. Among 17,465 anticoagulated patients with AF, 4,828 (27.6%) patients were prescribed VKA and 12,637 (72.4%) patients an NOAC: 11,884 (68.0%) patients had SAMe-TT2R2 scores 0-2 and 5,581 (32.0%) patients had scores >2. The proportion of patients prescribed VKA was 28.0% among patients with SAMe-TT2R2 scores >2 and 27.5% in those with scores ≤2. Conclusions: The lack of a clear association between the SAMe-TT2R2 score and anticoagulant selection may be attributed to the relative efficacy and safety profiles between NOACs and VKAs as well as to the absence of trial evidence that an SAMe-TT2R2-guided strategy for the selection of the type of anticoagulation in NVAF patients has an impact on clinical outcomes of efficacy and safety. The latter hypothesis is currently being tested in a randomized controlled trial. Clinical trial registration: URL: https://www.clinicaltrials.gov//Unique identifier: NCT01937377, NCT01468701, and NCT01671007
"Ain't nobody got time for that!": Framing and stereotyping in legacy and social media
Background Social media can be powerful tools for rallying support for a social cause, political mobilization, and social commentary. They can also greatly contribute to incendiary discourses and social stereotypes-often through memes. This article explores the case of one American, known by the moniker of "Sweet Brown," whose interview about a local fire made her an overnight celebrity in 2012. Analysis A frame analysis of her portrayals in legacy and social media is conducted, and reveals that social media platforms facilitate and even encourage a reductionist approach to messaging. Conclusion and implications Sweet Brown's appearance, which conjures gender, race, and socio-economic class, became a powerful tool for circulating stereotypes. The interplay between legacy and social media can serve to reproduce stereotypes and marginalization, as is evident in the case of Sweet Brown
‘Liking and sharing’ the stigmatization of poverty and social welfare: Representations of poverty and welfare through internet memes on social media
Internet memes play an important role in the reproduction, reinforcement and cir-culation of social stereotypes, including those about people who live in poverty. In this paper we investigate the relationship between Internet memes and stereotypes about poverty by examining a set of memes that make claims about one particular aspect of poverty in high-income countries – receipt of social assistance in the form of welfare cheques, medical cov-erage and food. We apply critical discourse analysis to a set of widely circulated poverty memes to identify how notions of individual responsibility and deservedness surface in these messages. Whereas the memes in our sample consist of both visual and textual elements, we found that the text was decidedly more important for reproducing stereotypes linked to the abstract neoliberal values of individual responsibility and participation in market econo-my
Seal hunts in Canada and on Twitter: Exploring the tensions between indigenous rights and animal rights with #sealfie
Background In 2014, a Twitter discussion of seal hunting, using the hashtag #sealfie, spurred a digital conflict between two rights movements-Indigenous rights in Canada and animal rights. This digital controversy touches on race, class, and geography. Analysis The hashtag's life on Twitter obscures the two movements' shared challenges: the undeniably neoliberal context consisting of ongoing economic struggles in northern and remote communities, and the continued loss of wildlife habitat. Conclusions and implications The authors analyze the #sealfie Twitter content generated between 2014 and 2017, exploring the tensions between the claims of the Indigenous rights and animal rights movements. They probe the failure of Twitter, and more generally social media, to generate a climate of genuine debate, and they consider how such digital platforms can serve as echo chambers for stereotypes and discriminatory discourse
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