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  • American Society of Hematology  (5)
  • 1
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    The Cytoplasmic Domain of Stem Cell Antigen CD34 Is Essential for Cytoadhesion Signaling But Not Sufficient for Proliferation Signaling (1998)
    American Society of Hematology
    Details
    Publication Date: 1998-02-15
    Description: CD34 is widely used as a marker in the identification and purification of human hematopoietic stem and progenitor cells; however, its function within hematopoiesis is largely unknown. We have investigated the contribution of cytoplasmic domain of CD34 in cytoadhesion signaling and proliferation signaling in hematopoietic cells. Engagement of particular determinants of CD34 by monoclonal antibodies leads to homotypic adhesiveness of the full-length CD34-transfected BaF3 cells. However, this homotypic adhesiveness is abrogated in BaF3 cells transfected with the truncated CD34 lacking the cytoplasmic domain. Cytoadhesion signaling through the cytoplasmic domain of CD34 cannot be restored through that of erythropoietin receptor (EPOR) or granulocyte colony-stimulating factor receptor (G-CSFR), suggesting that the cytoplasmic domain of CD34 is required for its signal transduction of cellular adhesion. In constrast, we show that replacing the cytoplasmic domain of EPOR or G-CSFR with that of CD34 abolished growth signal transduction in response to EPO or G-CSF in the chimeric receptor-transfected BaF3, 32D, and FDCP1 cells, whereas the wild-type EPOR- or G-CSFR-transfected cells responded to EPO or G-CSF growth signaling well. These results suggest that the cytoplasmic portion of CD34 may not contain the elements necessary to transduce a proliferative signal in hematopoietic cells. Thus, the function of CD34 in hematopoiesis is primarily on hematopoietic cell adhesion.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
    Topics: Biology , Medicine
    Published by American Society of Hematology
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  • 2
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    Long-Term Ex Vivo Maintenance and Expansion of Transplantable Human Hematopoietic Stem Cells (1999)
    American Society of Hematology
    Details
    Publication Date: 1999-09-01
    Description: We have developed a stromal-based in vitro culture system that facilitates ex vivo expansion of transplantable CD34+thy-1+ cells using long-term hematopoietic reconstitution in severe combined immunodeficient-human (SCID-hu) mice as an in vivo assay for transplantable human hematopoietic stem cells (HSCs). The addition of leukemia inhibitory factor (LIF) to purified CD34+ thy-1+ cells on AC6.21 stroma, a murine bone marrow–derived stromal cell line, caused expansion of cells with CD34+ thy-1+ phenotype. Addition of other cytokines, including interleukin-3 (IL-3), IL-6, granulocyte-macrophage colony-stimulating factor, and stem cell factor, to LIF in the cultures caused a 150-fold expansion of cells retaining the CD34+ thy-1+ phenotype. The ex vivo–expanded CD34+ thy-1+ cells gave rise to multilineage differentiation, including myeloid, T, and B cells, when transplanted into SCID-hu mice. Both murine LIF (cannot bind to human LIF receptor) and human LIF caused expansion of human CD34+ thy-1+ cells in vitro, suggesting action through the murine stroma. Furthermore, another human HSC candidate, CD34+ CD38− cells, shows a similar pattern of proliferative response. This suggests thatex vivo expansion of transplantable human stem cells under this in vitro culture system is a general phenomenon and not just specific for CD34+ thy-1+ cells.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
    Topics: Biology , Medicine
    Published by American Society of Hematology
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  • 3
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    Long-Term Ex Vivo Maintenance and Expansion of Transplantable Human Hematopoietic Stem Cells (1999)
    American Society of Hematology
    Details
    Publication Date: 1999-09-01
    Description: We have developed a stromal-based in vitro culture system that facilitates ex vivo expansion of transplantable CD34+thy-1+ cells using long-term hematopoietic reconstitution in severe combined immunodeficient-human (SCID-hu) mice as an in vivo assay for transplantable human hematopoietic stem cells (HSCs). The addition of leukemia inhibitory factor (LIF) to purified CD34+ thy-1+ cells on AC6.21 stroma, a murine bone marrow–derived stromal cell line, caused expansion of cells with CD34+ thy-1+ phenotype. Addition of other cytokines, including interleukin-3 (IL-3), IL-6, granulocyte-macrophage colony-stimulating factor, and stem cell factor, to LIF in the cultures caused a 150-fold expansion of cells retaining the CD34+ thy-1+ phenotype. The ex vivo–expanded CD34+ thy-1+ cells gave rise to multilineage differentiation, including myeloid, T, and B cells, when transplanted into SCID-hu mice. Both murine LIF (cannot bind to human LIF receptor) and human LIF caused expansion of human CD34+ thy-1+ cells in vitro, suggesting action through the murine stroma. Furthermore, another human HSC candidate, CD34+ CD38− cells, shows a similar pattern of proliferative response. This suggests thatex vivo expansion of transplantable human stem cells under this in vitro culture system is a general phenomenon and not just specific for CD34+ thy-1+ cells.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
    Topics: Biology , Medicine
    Published by American Society of Hematology
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  • 4
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    A secreted and LIF-mediated stromal cell–derived activity that promotes ex vivo expansion of human hematopoietic stem cells (2000)
    American Society of Hematology
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    Publication Date: 2000-03-15
    Description: The development of culture systems that facilitate ex vivo maintenance and expansion of transplantable hematopoietic stem cells (HSCs) is vital to stem cell research. Establishment of such culture systems will have significant impact on ex vivo manipulation and expansion of transplantable stem cells in clinical applications such as gene therapy, tumor cell purging, and stem cell transplantation. We have recently developed a stromal-based culture system that facilitates ex vivo expansion of transplantable human HSCs. In this stromal-based culture system, 2 major contributors to the ex vivo stem cell expansion are the addition of leukemia inhibitory factor (LIF) and the AC6.21 stromal cells. Because the action of LIF is indirect and mediated by stromal cells, we hypothesized that LIF binds to the LIF receptor on AC6.21 stromal cells, leading to up-regulated production of stem cell expansion promoting factor (SCEPF) and/or down-regulated production of stem cell expansion inhibitory factor (SCEIF). Here we demonstrate a secreted SCEPF activity in the conditioned media of LIF-treated AC6.21 stromal cell cultures (SCM-LIF). The magnitude of ex vivo stem cell expansion depends on the concentration of the secreted SCEPF activity in the SCM-LIF. Furthermore, we have ruled out the contribution of 6 known early-acting cytokines, including interleukin-3, interleukin-6, granulocyte macrophage colony-stimulating factor, stem cell factor, flt3 ligand, and thrombopoietin, to this SCEPF activity. Although further studies are required to characterize this secreted SCEPF activity and to determine whether this secreted SCEPF activity is mediated by a single factor or by multiple growth factors, our results demonstrate that stromal cells are not required for this secreted SCEPF activity to facilitate ex vivo stem cell expansion.
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    Electronic ISSN: 1528-0020
    Topics: Biology , Medicine
    Published by American Society of Hematology
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  • 5
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    Identification of a candidate human neurohematopoietic stem-cell population (2001)
    American Society of Hematology
    Details
    Publication Date: 2001-10-15
    Description: It was recently reported that transplantation of clonally derived murine neurosphere cells into sublethally irradiated allogeneic hosts leads to a donor-derived hematopoietic reconstitution. The confirmation of the existence of a common neurohematopoietic stem cell in the human brain will have a significant effect on stem cell research and on clinical transplantation. Here, it is demonstrated that the human fetal brain contains separate but overlapping epidermal growth factor (EGF)–responsive and basic fibroblast growth factor (FGF-2)–responsive neural stem cells. The majority (〉 85%) of cells within these EGF- and/or FGF-2–generated neurospheres express characteristic neural stem/progenitor cell markers including nestin, EGF receptor, and FGF-2 receptor. These neural stem cells can be continuously passaged in vitro, and demonstrate a constant 20-fold expansion in every passage for up to the fifth passage (the longest period that has been carried out in the authors' laboratory). These neural stem cells are multipotential for neurons, astrocytes, and oligodendrocytes. After transplantation into SCID-hu mice, all neural stem cells, regardless of passages, culture conditions, and donors, are able to establish long-term hematopoietic reconstitution in the presence of an intact human bone marrow microenvironment.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
    Topics: Biology , Medicine
    Published by American Society of Hematology
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