scholarly journals The Spliceosomal Component Sf3b1 Is Essential for Hematopoietic Differentiation

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3582-3582
Author(s):  
Adriana De La Garza-Sauceda ◽  
Rosannah C. Cameron ◽  
Sara Nik ◽  
Michelle Gulfo ◽  
Sara G. Payne ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Endothelial Cells ◽  
Cell Differentiation ◽  
Progenitor Cell ◽  
Cell Development ◽  
Hematopoietic Cell ◽  
Lateral Plate Mesoderm ◽  
Lateral Plate ◽  
Hematopoietic Stem ◽  
Primitive Erythropoiesis

Abstract Myelodysplastic syndrome (MDS) is a disorder arising from hematopoietic stem and progenitor cell (HSPC) dysfunction resulting in ineffective hematopoiesis. A multitude of recurrent somatic mutations in spliceosomal components were recently identified in MDS that likely contribute to the pathogenesis of the disease. The lack of in vivo models to study cell-type specific effects of spliceosomal mutations limits our understanding of why such mutations lead to hematopoietic abnormalities. Using a zebrafish with a loss-of-function mutation in sf3b1 (sf3b1hi3394), an essential member of the spliceosome, we demonstrate hematopoietic cell differentiation and hematopoietic stem and progenitor cell (HSPC) specification are processes sensitive to spliceosomal malfunction. Primitive erythropoiesis initiates normally in sf3b1 mutants as evidenced by expression of scl in the posterior lateral plate mesoderm at 14 hours post fertilization (hpf) as well as gata1 and beta-globin at 24 hpf. Flow cytometry quantification of gata1:gfp positive erythrocytes showed sf3b1 mutants have 25% more cells at 24 hpf, but greater than 3-fold fewer cells at 36 and 48 hpf, time points when wild type erythroblasts are expanding and differentiating. At 48 hpf, we also observed decreased levels of o-dianisidine positive erythrocytes, low numbers of morphologically mature erythroblasts, and higher levels of immature erythroblasts in sf3b1 mutants. Similarly, we observed normal initiation of primitive myelopoiesis marked by stem cell leukemia (scl) expression in the anterior lateral plate mesoderm at 14 hpf, but diminished expression of more differentiated markers, l-plastin and myeloperoxidase at 24 and 28 hpf in sf3b1 mutants. Quantification of lysozyme C:dsred positivemyeloid cells using flow cytometry also showed 24-fold fewer mature myeloid cells in sf3b1 mutants at 36 hpf. Our data on primitive erythropoiesis and myelopoiesis indicate sf3b1 is required for hematopoietic cell differentiation. Additionally, sf3b1 mutants have diminished expression of the definitive HSPC marker runx1 within the aorta at 28 hpf. In contrast, we observed normal expression of the pan-endothelial marker kinase insert domain receptor-like (kdrl) and aorta-specific markers notch1b and notch3 at 24 hpf. Flow cytometry quantification of kdrl:gfp endothelial cells at 24 hpf shows no difference in the frequency of endothelial cells in sf3b1 mutants. Moreover, we observed fewer cmyb:gfp; kdrl:dsred double positive HSPCs along the dorsal aorta. The data suggest that decreased HSPC formation in sf3b1 mutants is due to a failure in hemogenic induction. From these studies, we show sf3b1 is required at specific stages of hematopoietic cell development. These results provide novel insight into the role of splicing in blood cell development, and can afford a deeper understanding of the mechanism of splicing regulation on the origins of MDS. Disclosures No relevant conflicts of interest to declare.

scholarly journals Dermomyotome-derived endothelial cells migrate to the dorsal aorta to support hematopoietic stem cell emergence

2020 ◽  
Author(s):  
Pankaj Sahai-Hernandez ◽  
Claire Pouget ◽  
Ondřej Svoboda ◽  
David Traver
Keyword(s):  
Endothelial Cells ◽  
Stem Cell ◽  
Paraxial Mesoderm ◽  
Dorsal Aorta ◽  
Similar Process ◽  
Lateral Plate Mesoderm ◽  
Lateral Plate ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells ◽  
Endothelial Precursors

AbstractDevelopment of the dorsal aorta is a key step in the establishment of the adult blood-forming system, since hematopoietic stem and progenitor cells (HSPCs) arise from ventral aortic endothelium in all vertebrate animals studied. Work in zebrafish has demonstrated that arterial and venous endothelial precursors arise from distinct subsets of lateral plate mesoderm. Earlier studies in the chick showed that paraxial mesoderm generates another subset of endothelial cells that incorporate into the dorsal aorta to replace HSPCs as they exit the aorta and enter circulation. Here we show that a similar process occurs in the zebrafish, where a population of endothelial precursors delaminates from the somitic dermomyotome to incorporate exclusively into the developing dorsal aorta. Whereas somite-derived endothelial cells (SDECs) lack hematopoietic potential, they act as local niche to support the emergence of HSPCs from neighboring hemogenic endothelium. Thus, at least three subsets of endothelial cells (ECs) contribute to the developing dorsal aorta: vascular ECs, hemogenic ECs, and SDECs. Taken together, our findings indicate that the distinct spatial origins of endothelial precursors dictate different cellular potentials within the developing dorsal aorta.

scholarly journals Unexpected redundancy of Gpr56 and Gpr97 during hematopoietic cell development and differentiation

2021 ◽  
Vol 5 (3) ◽  
pp. 829-842
Author(s):  
Antonio Maglitto ◽  
Samanta A. Mariani ◽  
Emma de Pater ◽  
Carmen Rodriguez-Seoane ◽  
Chris S. Vink ◽  
...  
Keyword(s):  
Stem Cell ◽  
G Protein ◽  
Progenitor Cell ◽  
Cell Development ◽  
Hematopoietic Cell ◽  
Conditional Knockout ◽  
Hematopoietic Stem ◽  
Cell Fate Decisions ◽  
Development And Differentiation ◽  
G Protein Coupled

Abstract Integrated molecular signals regulate cell fate decisions in the embryonic aortic endothelium to drive hematopoietic stem cell (HSC) generation during development. The G-protein–coupled receptor 56 (Gpr56, also called Adgrg1) is the most highly upregulated receptor gene in cells that take on hematopoietic fate and is expressed by adult bone marrow HSCs. Despite the requirement for Gpr56 in hematopoietic stem/progenitor cell (HS/PC) generation in zebrafish embryos and the highly upregulated expression of GPR56 in treatment-resistant leukemic patients, its function in normal mammalian hematopoiesis remains unclear. Here, we examine the role of Gpr56 in HS/PC development in Gpr56 conditional knockout (cKO) mouse embryos and Gpr knockout (KO) embryonic stem cell (ESC) hematopoietic differentiation cultures. Our results show a bias toward myeloid differentiation of Gpr56 cKO fetal liver HSCs and an increased definitive myeloid progenitor cell frequency in Gpr56KO ESC differentiation cultures. Surprisingly, we find that mouse Gpr97 can rescue Gpr56 morphant zebrafish hematopoietic generation, and that Gpr97 expression is upregulated in mouse Gpr56 deletion models. When both Gpr56 and Gpr97 are deleted in ESCs, no or few hematopoietic PCs (HPCs) are generated upon ESC differentiation. Together, our results reveal novel and redundant functions for these 2 G-protein coupled receptors in normal mammalian hematopoietic cell development and differentiation.

Tie2 Is Essential for the Development of Lymphangiogenic Endothelial Cell during ES Cell Differentiation.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 844-844
Author(s):  
Isao Hamaguchi ◽  
Toru Morisada ◽  
Masaki Azuma ◽  
Kyoko Murakami ◽  
Yuichi Oike ◽  
...  
Keyword(s):  
Stem Cells ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Differentiation ◽  
Developmental Stage ◽  
Es Cells ◽  
Hematopoietic Cells ◽  
Cell Development ◽  
Es Cell ◽  
Hematopoietic Stem

Abstract Tie2 is a receptor type tyrosine kinase, and is expressed in the hematopoietic stem cells and endothelial cells. We have recently shown that Tie2 and its ligand Angiopoietin-1 (Ang1) signal play a crucial role for maintenance of long-term repopulating hematopoietic stem cells in adult bone marrow (Cell, Vol 118, 149, 2004). Although Tie2 deficient mice showed defect of endothelial cell development, it is not clear whether Tie2 is critical for the development of hematopoietic and lymphangiogenic endothelial cells. In order to clarify the function of Tie2 during the developmental stage, we have developed the cell culture system of ES cell differentiation. After removing LIF, ES cells were cultured on collagen type IV coated plates to promote the differentiation to mesodermal lineage for two days, and the cells were cultured on OP9 stromal cells. Using our system hematopoietic and endothelial progenitors were differentiated efficiently on OP9 cells from the different ES cell straines, E14, TT2, and R1. Expression study showed that ES cell derived cells expressed Tie2 and Flk1 at day 5 of culture on OP9 cells. When we compared the cell fraction sorted by Tie2 and Flk1 mAb regarding differentiation potential, Tie2+Flk1+ fraction was revealed to be an enriched fraction of progenitors for hematopoietic cells and PECAM-1+ endothelial cells. To detect the lymphangiogenic endothelial cells derived from ES cells, we prepared the monoclonal antibody against LYVE-1, which is the receptor for extracellular matrix, glycosaminoglycan. And we confirmed that LYVE-1 was expressed in the embryonic lymphatic endothelium. By using LYVE-1 mAb, we sorted out LYVE-1+ cells from differentiated ES cells and carried out RT-PCR assay. LYVE-1+ cells expressed lymphangiogenic endothelial cell-specific genes, VEGFR-3, Podoplanin, and Prox-1, moreover LYVE-1+ cells took up the DiI-Ac-LDL. These findings indicate that LYVE-1+ cells derived from ES cells have a character of lymphangiogenic endothelial cells. When we compared the cell fraction sorted by Tie2 and Flk1 mAb, LYVE-1+ cells were differentiated from Tie2+Flk1+ fraction dominantly, but not from the other fractions, Tie2-Flk1+, Tie2+Flk1−, and Tie2-Flk1− fraction. These findings suggest that Tie2 is crucial for development of lymphangiogenic endothelial cells as well as hematopoietic cells and endothelial cells. In order to analyze the function of Tie2 during the developmental stage, we differentiated Tie2−/− ES cells using our system. The LYVE-1+ and PECAM-1+ cells derived from Tie2−/− ES cells dramatically decreased as culturing days went by, and at day6 of culture the LYVE-1+ and PECAM-1+ cells derived from Tie2−/− ES cells were one sixth and one third of Tie2+/− cells respectively. When we added 100μM of caspase inhibitor in the culture media, the number of both LYVE-1+ cells and PECAM-1+ cells were recovered. These findings suggest that developmental defect of lymphangiogenic endothelial and endothelial cells are caused by apoptosis because of the blockage of Tie2 signaling. However we could not detect abnormal development of hematopoietic cells from Tie2−/− ES cells. In conclusion, Tie2+Flk1+ fraction derived from ES cells is an enriched fraction of progenitors for lymphangiogenic endothelial cells, and Tie2 signaling is dispensable for lymphangiogenic endothelial cell development as well as endothelial cell development as an anti-apoptotic signaling during ES cell differentiation, but Tie2 is not essential for hematopoietic development.

scholarly journals Unexpected redundancy of Gpr56 and Gpr97 during hematopoietic cell development and differentiation

2020 ◽  
Author(s):  
A. Maglitto ◽  
S.A. Mariani ◽  
E. de Pater ◽  
C. Rodriguez-Seoane ◽  
C.S. Vink ◽  
...  
Keyword(s):  
Stem Cell ◽  
G Protein ◽  
Progenitor Cell ◽  
Embryonic Stem ◽  
Cell Development ◽  
Hematopoietic Cell ◽  
Conditional Knockout ◽  
Hematopoietic Stem ◽  
Development And Differentiation ◽  
G Protein Coupled

AbstractIntegrated molecular signals regulate cell fate during embryonic hematopoietic stem cell (HSC) generation. The G-protein coupled receptor 56 (Gpr56) is the most highly-upregulated receptor gene in cells that take on hematopoietic fate and it is expressed by adult bone marrow HSCs. Although Gpr56 is required for hematopoietic stem/progenitor cell (HS/PC) generation in zebrafish embryos, its function in mammalian hematopoiesis remains unclear. Here we examine the role of Gpr56 in HS/PC development in Gpr56 conditional knockout (cKO) mouse embryos and Gpr knockout (KO) embryonic stem cell (ESC) hematopoietic differentiation cultures. Our results show a myeloid bias of Gpr56 cKO fetal liver HSCs and an increased definitive myeloid progenitor cell frequency in Gpr56KO ESC differentiation cultures. Surprisingly, we find that mouse Gpr97 rescues Gpr56 morphant zebrafish hematopoietic generation, and that Gpr97 expression is upregulated in mouse Gpr56 deletion models. When both Gpr56 and Gpr97 are deleted in ESCs, no/few HS/PCs are generated upon ESC differentiation. Together, our results reveal novel and redundant functions for these two G-protein coupled receptors in normal mammalian hematopoietic cell development and differentiation.

scholarly journals WNT9A Is a Conserved Regulator of Hematopoietic Stem and Progenitor Cell Development

Genes ◽  
2018 ◽  
Vol 9 (2) ◽  
pp. 66 ◽  
Author(s):  
Jenna Richter ◽  
Edouard Stanley ◽  
Elizabeth Ng ◽  
Andrew Elefanty ◽  
David Traver ◽  
...  
Keyword(s):  
Progenitor Cell ◽  
Cell Development ◽  
Hematopoietic Stem

scholarly journals Loss of Tie2 receptor compromises embryonic stem cell–derived endothelial but not hematopoietic cell survival

Blood ◽  
2006 ◽  
Vol 107 (3) ◽  
pp. 1207-1213 ◽  
Author(s):  
Isao Hamaguchi ◽  
Tohru Morisada ◽  
Masaki Azuma ◽  
Kyoko Murakami ◽  
Madoka Kuramitsu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Es Cells ◽  
Embryonic Stem ◽  
Hematopoietic Cell ◽  
Caspase Inhibitor ◽  
Lymphatic Endothelial Cells ◽  
Es Cell ◽  
Hematopoietic Stem ◽  
Hematopoietic System ◽  
Tie2 Receptor

AbstractTie2 is a receptor-type tyrosine kinase expressed on hematopoietic stem cells and endothelial cells. We used cultured embryonic stem (ES) cells to determine the function of Tie2 during early vascular development and hematopoiesis. Upon differentiation, the ES cell–derived Tie2+Flk1+ fraction was enriched for hematopoietic and endothelial progenitor cells. To investigate lymphatic differentiation, we used a monoclonal antibody against LYVE-1 and found that LYVE-1+ cells derived from Tie2+Flk1+ cells possessed various characteristics of lymphatic endothelial cells. To determine whether Tie2 played a role in this process, we analyzed differentiation of Tie2-/- ES cells. Although the initial numbers of LYVE-1+ and PECAM-1+ cells derived from Tie2-/- cells did not vary significantly, the number of both decreased dramatically upon extended culturing. Such decreases were rescued by treatment with a caspase inhibitor, suggesting that reductions were due to apoptosis as a consequence of a lack of Tie2 signaling. Interestingly, Tie2-/- ES cells did not show measurable defects in development of the hematopoietic system, suggesting that Tie2 is not essential for hematopoietic cell development.

scholarly journals HDAC3 is crucial in shear- and VEGF-induced stem cell differentiation toward endothelial cells

2006 ◽  
Vol 174 (7) ◽  
pp. 1059-1069 ◽  
Author(s):  
Lingfang Zeng ◽  
Qingzhong Xiao ◽  
Andriana Margariti ◽  
Zhongyi Zhang ◽  
Anna Zampetaki ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Laminar Flow ◽  
Progenitor Cell ◽  
Stem Cell Differentiation ◽  
Es Cell ◽  
Local Flow ◽  
Proliferation And Differentiation

Reendothelialization involves endothelial progenitor cell (EPC) homing, proliferation, and differentiation, which may be influenced by fluid shear stress and local flow pattern. This study aims to elucidate the role of laminar flow on embryonic stem (ES) cell differentiation and the underlying mechanism. We demonstrated that laminar flow enhanced ES cell–derived progenitor cell proliferation and differentiation into endothelial cells (ECs). Laminar flow stabilized and activated histone deacetylase 3 (HDAC3) through the Flk-1–PI3K–Akt pathway, which in turn deacetylated p53, leading to p21 activation. A similar signal pathway was detected in vascular endothelial growth factor–induced EC differentiation. HDAC3 and p21 were detected in blood vessels during embryogenesis. Local transfer of ES cell–derived EPC incorporated into injured femoral artery and reduced neointima formation in a mouse model. These data suggest that shear stress is a key regulator for stem cell differentiation into EC, especially in EPC differentiation, which can be used for vascular repair, and that the Flk-1–PI3K–Akt–HDAC3–p53–p21 pathway is crucial in such a process.

scholarly journals Analyses of Avascular Mutants Reveal Unique Transcriptomic Signature of Non-conventional Endothelial Cells

2020 ◽  
Vol 8 ◽  
Author(s):  
Boryeong Pak ◽  
Christopher E. Schmitt ◽  
Woosoung Choi ◽  
Jun-Dae Kim ◽  
Orjin Han ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Molecular Mechanisms ◽  
Lineage Tracing ◽  
Molecular Characteristics ◽  
Lateral Plate Mesoderm ◽  
Rna Seq ◽  
Developmental History ◽  
Lateral Plate ◽  
Developmental Origin ◽  
Transcriptomic Signature

Endothelial cells appear to emerge from diverse progenitors. However, to which extent their developmental origin contributes to define their cellular and molecular characteristics remains largely unknown. Here, we report that a subset of endothelial cells that emerge from the tailbud possess unique molecular characteristics that set them apart from stereotypical lateral plate mesoderm (LPM)-derived endothelial cells. Lineage tracing shows that these tailbud-derived endothelial cells arise at mid-somitogenesis stages, and surprisingly do not require Npas4l or Etsrp function, indicating that they have distinct spatiotemporal origins and are regulated by distinct molecular mechanisms. Microarray and single cell RNA-seq analyses reveal that somitogenesis- and neurogenesis-associated transcripts are over-represented in these tailbud-derived endothelial cells, suggesting that they possess a unique transcriptomic signature. Taken together, our results further reveal the diversity of endothelial cells with respect to their developmental origin and molecular properties, and provide compelling evidence that the molecular characteristics of endothelial cells may reflect their distinct developmental history.

Expression of 4-Integrin Defines the Earliest Precursor of Hematopoietic Cell Lineage Diverged From Endothelial Cells

Blood ◽  
1999 ◽  
Vol 93 (4) ◽  
pp. 1168-1177 ◽  
Author(s):  
Minetaro Ogawa ◽  
Masami Kizumoto ◽  
Satomi Nishikawa ◽  
Tetsuhiro Fujimoto ◽  
Hiroaki Kodama ◽  
...  
Keyword(s):  
Stem Cells ◽  
Endothelial Cells ◽  
Cell Population ◽  
Embryonic Stem ◽  
Cell Lineage ◽  
Hematopoietic Cell ◽  
Hematopoietic Stem ◽  
Hematopoietic Cell Lineage ◽  
E Cadherin

Abstract Embryonic stem cells can differentiate in vitro into hematopoietic cells through two intermediate stages; the first being FLK1+ E-cadherin− proximal lateral mesoderm and the second being CD45− VE-cadherin+endothelial cells. To further dissect the CD45−VE-cadherin+ cells, we have examined distribution of 4-integrin on this cell population, because 4-integrin is the molecule expressed on hematopoietic stem cells. During culture of FLK1+ E-cadherin− cells, CD45− VE-cadherin+4-integrin− cells differentiate first, followed by 4-integrin+ cells appearing in both CD45− VE-cadherin+ and CD45−VE-cadherin− cell populations. In the CD45−VE-cadherin+ cell population, 4-integrin+ subset but not 4-integrin− subset had the potential to differentiate to hematopoietic lineage cells, whereas endothelial cell progenitors were present in both subsets. The CD45−VE-cadherin− 4-integrin+ cells also showed hematopoietic potential. Reverse transcription-polymerase chain reaction analyses showed that differential expression of the Gata2 and Myb genes correlated with the potential of the 4-integrin+ cells to give rise to hematopoietic cell differentiation. Hematopoietic CD45−VE-cadherin+ 4-integrin+ cells were also present in the yolk sac and embryonic body proper of 9.5 day postcoitum mouse embryos. Our results suggest that the expression of 4-integrin is a marker of the earliest precursor of hematopoietic cell lineage that was diverged from endothelial progenitors.

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