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  1. Article ; Online: Isolation of human bone marrow stromal cells from bone marrow biopsies for single-cell RNA sequencing

    Hélène F.E. Gleitz / Inge A.M. Snoeren / Stijn N.R. Fuchs / Nils B. Leimkühler / Rebekka K. Schneider

    STAR Protocols, Vol 2, Iss 2, Pp 100538- (2021)

    2021  

    Abstract: Summary: Bone marrow (BM) mesenchymal stromal cells play an important role in regulating stem cell quiescence and homeostasis; they are also key contributors to various hematological malignancies. However, human bone marrow stromal cells are difficult to ...

    Abstract Summary: Bone marrow (BM) mesenchymal stromal cells play an important role in regulating stem cell quiescence and homeostasis; they are also key contributors to various hematological malignancies. However, human bone marrow stromal cells are difficult to isolate and prone to damage during isolation. This protocol describes a combination of mechanical and enzymatic isolation of BM stromal cells from human BM biopsies, followed by FACS sorting to separate stromal sub-populations including mesenchymal stromal cells, fibroblasts, and Schwann cells for single-cell RNA sequencing.For complete details on the use and execution of this protocol, please refer to Leimkühler et al. (2020).
    Keywords Cell Biology ; Cell isolation ; Flow Cytometry/Mass Cytometry ; Cancer ; Science (General) ; Q1-390
    Language English
    Publishing date 2021-06-01T00:00:00Z
    Publisher Elsevier
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  2. Article ; Online: Using human iPSC-derived kidney organoids to decipher SARS-CoV-2 pathology on single cell level

    Katharina C. Reimer / Jitske Jansen / Gijs J. Overheul / Pascal Miesen / Ronald P. van Rij / Sergio H. Triana / Bart Smeets / Rebekka K. Schneider / Rafael Kramann

    STAR Protocols, Vol 3, Iss 3, Pp 101612- (2022)

    2022  

    Abstract: Summary: We describe a protocol for single-cell RNA sequencing of SARS-CoV-2-infected human induced pluripotent stem cell (iPSC)-derived kidney organoids. After inoculation of kidney organoids with virus, we use mechanical and enzymatic disruption to ... ...

    Abstract Summary: We describe a protocol for single-cell RNA sequencing of SARS-CoV-2-infected human induced pluripotent stem cell (iPSC)-derived kidney organoids. After inoculation of kidney organoids with virus, we use mechanical and enzymatic disruption to obtain single cell suspensions. Next, we process the organoid-derived cells into sequencing-ready SARS-CoV-2-targeted libraries. Subsequent sequencing analysis reveals changes in kidney cells after virus infection. The protocol was designed for kidney organoids cultured in a 6-well transwell format but can be adapted to organoids with different organ backgrounds.For complete details on the use and execution of this protocol, please refer to Jansen et al. (2022). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
    Keywords Cell biology ; Single cell ; Microbiology ; Stem cells ; Organoids ; Science (General) ; Q1-390
    Language English
    Publishing date 2022-09-01T00:00:00Z
    Publisher Elsevier
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  3. Article ; Online: Mapping the cardiac vascular niche in heart failure

    Fabian Peisker / Maurice Halder / James Nagai / Susanne Ziegler / Nadine Kaesler / Konrad Hoeft / Ronghui Li / Eric M. J. Bindels / Christoph Kuppe / Julia Moellmann / Michael Lehrke / Christian Stoppe / Michael T. Schaub / Rebekka K. Schneider / Ivan Costa / Rafael Kramann

    Nature Communications, Vol 13, Iss 1, Pp 1-

    2022  Volume 20

    Abstract: The cardiac vascular niche is of major importance in homeostasis and disease, but knowledge of its complexity in response to injury remains limited. Here we combine lineage tracing with single cell RNA sequencing to show alterations in fibroblasts, ... ...

    Abstract The cardiac vascular niche is of major importance in homeostasis and disease, but knowledge of its complexity in response to injury remains limited. Here we combine lineage tracing with single cell RNA sequencing to show alterations in fibroblasts, endothelial and mural cells in hypertrophic remodeling.
    Keywords Science ; Q
    Language English
    Publishing date 2022-05-01T00:00:00Z
    Publisher Nature Portfolio
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  4. Article ; Online: Non-canonical Hedgehog signaling mediates profibrotic hematopoiesis-stroma crosstalk in myeloproliferative neoplasms

    Jessica E. Pritchard / Juliette E. Pearce / Inge A.M. Snoeren / Stijn N.R. Fuchs / Katrin Götz / Fabian Peisker / Silke Wagner / Adam Benabid / Niklas Lutterbach / Vanessa Klöker / James S. Nagai / Monica T. Hannani / Anna K. Galyga / Ellen Sistemich / Bella Banjanin / Niclas Flosdorf / Eric Bindels / Kathrin Olschok / Katharina Biaesch /
    Nicolas Chatain / Neha Bhagwat / Andrew Dunbar / Rita Sarkis / Olaia Naveiras / Marie-Luise Berres / Steffen Koschmieder / Ross L. Levine / Ivan G. Costa / Hélène F.E. Gleitz / Rafael Kramann / Rebekka K. Schneider

    Cell Reports, Vol 43, Iss 1, Pp 113608- (2024)

    1481  

    Abstract: Summary: The role of hematopoietic Hedgehog signaling in myeloproliferative neoplasms (MPNs) remains incompletely understood despite data suggesting that Hedgehog (Hh) pathway inhibitors have therapeutic activity in patients. We aim to systematically ... ...

    Abstract Summary: The role of hematopoietic Hedgehog signaling in myeloproliferative neoplasms (MPNs) remains incompletely understood despite data suggesting that Hedgehog (Hh) pathway inhibitors have therapeutic activity in patients. We aim to systematically interrogate the role of canonical vs. non-canonical Hh signaling in MPNs. We show that Gli1 protein levels in patient peripheral blood mononuclear cells (PBMCs) mark fibrotic progression and that, in murine MPN models, absence of hematopoietic Gli1, but not Gli2 or Smo, significantly reduces MPN phenotype and fibrosis, indicating that GLI1 in the MPN clone can be activated in a non-canonical fashion. Additionally, we establish that hematopoietic Gli1 has a significant effect on stromal cells, mediated through a druggable MIF-CD74 axis. These data highlight the complex interplay between alterations in the MPN clone and activation of stromal cells and indicate that Gli1 represents a promising therapeutic target in MPNs, particularly that Hh signaling is dispensable for normal hematopoiesis.
    Keywords CP: Stem cell research ; Biology (General) ; QH301-705.5
    Language English
    Publishing date 2024-01-01T00:00:00Z
    Publisher Elsevier
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  5. Article ; Online: Malignant Transformation Involving CXXC4 Mutations Identified in a Leukemic Progression Model of Severe Congenital Neutropenia

    Patricia A. Olofsen / Szabolcs Fatrai / Paulina M.H. van Strien / Julia C. Obenauer / Hans W.J. de Looper / Remco M. Hoogenboezem / Claudia A.J. Erpelinck-Verschueren / Michael P.W.M. Vermeulen / Onno Roovers / Torsten Haferlach / Joop H. Jansen / Mehrnaz Ghazvini / Eric M.J. Bindels / Rebekka K. Schneider / Emma M. de Pater / Ivo P. Touw

    Cell Reports Medicine, Vol 1, Iss 5, Pp 100074- (2020)

    2020  

    Abstract: Summary: Severe congenital neutropenia (SCN) patients treated with CSF3/G-CSF to alleviate neutropenia frequently develop acute myeloid leukemia (AML). A common pattern of leukemic transformation involves the appearance of hematopoietic clones with CSF3 ... ...

    Abstract Summary: Severe congenital neutropenia (SCN) patients treated with CSF3/G-CSF to alleviate neutropenia frequently develop acute myeloid leukemia (AML). A common pattern of leukemic transformation involves the appearance of hematopoietic clones with CSF3 receptor (CSF3R) mutations in the neutropenic phase, followed by mutations in RUNX1 before AML becomes overt. To investigate how the combination of CSF3 therapy and CSF3R and RUNX1 mutations contributes to AML development, we make use of mouse models, SCN-derived induced pluripotent stem cells (iPSCs), and SCN and SCN-AML patient samples. CSF3 provokes a hyper-proliferative state in CSF3R/RUNX1 mutant hematopoietic progenitors but does not cause overt AML. Intriguingly, an additional acquired driver mutation in Cxxc4 causes elevated CXXC4 and reduced TET2 protein levels in murine AML samples. Expression of multiple pro-inflammatory pathways is elevated in mouse AML and human SCN-AML, suggesting that inflammation driven by downregulation of TET2 activity is a critical step in the malignant transformation of SCN.
    Keywords severe congenital neutropenia ; AML ; CSF3R ; RUNX1 ; CXXC4 ; TET2 ; Medicine (General) ; R5-920
    Language English
    Publishing date 2020-08-01T00:00:00Z
    Publisher Elsevier
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  6. Article ; Online: An engineered multicomponent bone marrow niche for the recapitulation of hematopoiesis at ectopic transplantation sites

    Mónica S. Ventura Ferreira / Christian Bergmann / Isabelle Bodensiek / Kristina Peukert / Jessica Abert / Rafael Kramann / Paul Kachel / Björn Rath / Stephan Rütten / Ruth Knuchel / Benjamin L. Ebert / Horst Fischer / Tim H. Brümmendorf / Rebekka K. Schneider

    Journal of Hematology & Oncology, Vol 9, Iss 1, Pp 1-

    2016  Volume 14

    Abstract: Abstract Background Bone marrow (BM) niches are often inaccessible for controlled experimentation due to their difficult accessibility, biological complexity, and three-dimensional (3D) geometry. Methods Here, we report the development and ... ...

    Abstract Abstract Background Bone marrow (BM) niches are often inaccessible for controlled experimentation due to their difficult accessibility, biological complexity, and three-dimensional (3D) geometry. Methods Here, we report the development and characterization of a BM model comprising of cellular and structural components with increased potential for hematopoietic recapitulation at ectopic transplantation sites. Cellular components included mesenchymal stromal cells (MSCs) and hematopoietic stem and progenitor cells (HSPCs). Structural components included 3D β-tricalcium phosphate (β-TCP) scaffolds complemented with Matrigel or collagen I/III gels for the recreation of the osteogenic/extracellular character of native BM. Results In vitro, β-TCP/Matrigel combinations robustly maintained proliferation, osteogenic differentiation, and matrix remodeling capacities of MSCs and maintenance of HSPCs function over time. In vivo, scaffolds promoted strong and robust recruitment of hematopoietic cells to sites of ectopic transplantation, vascularization, and soft tissue formation. Conclusions Our tissue-engineered BM system is a powerful tool to explore the regulatory mechanisms of hematopoietic stem and progenitor cells for a better understanding of hematopoiesis in health and disease.
    Keywords Diseases of the blood and blood-forming organs ; RC633-647.5 ; Specialties of internal medicine ; RC581-951 ; Internal medicine ; RC31-1245 ; Medicine ; R ; Neoplasms. Tumors. Oncology. Including cancer and carcinogens ; RC254-282
    Subject code 610
    Language English
    Publishing date 2016-01-01T00:00:00Z
    Publisher BioMed Central
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  7. Article ; Online: Loss of function of TET2 cooperates with constitutively active KIT in murine and human models of mastocytosis.

    Serena De Vita / Rebekka K Schneider / Michael Garcia / Jenna Wood / Mathilde Gavillet / Benjamin L Ebert / Alexander Gerbaulet / Axel Roers / Ross L Levine / Ann Mullally / David A Williams

    PLoS ONE, Vol 9, Iss 5, p e

    2014  Volume 96209

    Abstract: Systemic Mastocytosis (SM) is a clonal disease characterized by abnormal accumulation of mast cells in multiple organs. Clinical presentations of the disease vary widely from indolent to aggressive forms, and to the exceedingly rare mast cell leukemia. ... ...

    Abstract Systemic Mastocytosis (SM) is a clonal disease characterized by abnormal accumulation of mast cells in multiple organs. Clinical presentations of the disease vary widely from indolent to aggressive forms, and to the exceedingly rare mast cell leukemia. Current treatment of aggressive SM and mast cell leukemia is unsatisfactory. An imatinib-resistant activating mutation of the receptor tyrosine kinase KIT (KIT D816V) is most frequently present in transformed mast cells and is associated with all clinical forms of the disease. Thus the etiology of the variable clinical aggressiveness of abnormal mast cells in SM is unclear. TET2 appears to be mutated in primary human samples in aggressive types of SM, suggesting a possible role in disease modification. In this report, we demonstrate the cooperation between KIT D816V and loss of function of TET2 in mast cell transformation and demonstrate a more aggressive phenotype in a murine model of SM when both mutations are present in progenitor cells. We exploit these findings to validate a combination treatment strategy targeting the epigenetic deregulation caused by loss of TET2 and the constitutively active KIT receptor for the treatment of patients with aggressive SM.
    Keywords Medicine ; R ; Science ; Q
    Subject code 610
    Language English
    Publishing date 2014-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  8. Article: (R)-2-Hydroxyglutarate Is Sufficient to Promote Leukemogenesis and Its Effects Are Reversible

    Losman, Julie-Aurore / Benjamin L. Ebert / Christine McMahon / David E. Root / Glenn S. Cowley / Peppi Koivunen / Rebekka K. Schneider / Ryan E. Looper / Sungwoo Lee / William G. Kaelin

    Science. 2013 Mar. 29, v. 339, no. 6127

    2013  

    Abstract: Focusing on the Right Metabolite A variety of human cancers, including acute leukemias and brain tumors, have mutations in the genes encoding isocitrate dehydrogenase 1 or 2 (IDH1, IDH2), which cause overproduction of a metabolite called 2- ... ...

    Abstract Focusing on the Right Metabolite A variety of human cancers, including acute leukemias and brain tumors, have mutations in the genes encoding isocitrate dehydrogenase 1 or 2 (IDH1, IDH2), which cause overproduction of a metabolite called 2-hydroxyglutarate (2HG). Losman et al. (p. 1621, published online 7 February) show that the R - but not the S -enantiomer of 2HG can transform cells and that R -2HG mediates transformation at least in part through effects on protein modifying EglN prolyl hydroxylases. Importantly, the transforming activity of R -2HG was reversible, suggesting that therapeutic strategies focusing on inhibition of R -2HG production or inhibition of EglN prolyl hydroxylases merit further investigation.
    Keywords brain ; genes ; humans ; isocitrate dehydrogenase ; metabolites ; mutation ; neoplasms
    Language English
    Dates of publication 2013-0329
    Size p. 1621-1625.
    Publishing place American Association for the Advancement of Science
    Document type Article
    ZDB-ID 128410-1
    ISSN 1095-9203 ; 0036-8075
    ISSN (online) 1095-9203
    ISSN 0036-8075
    DOI 10.1126/science.1231677
    Database NAL-Catalogue (AGRICOLA)

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