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Hematopoietic Colony-Forming Cell Assays

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Book cover Stem Cell Assays

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 407))

Summary

Hematopoiesis is the process by which stem cells divide and differentiate to produce the multiple types of mature cells found in blood. The process begins in early embryonic development and continues throughout adult life, primarily in the bone marrow. Various in vivo and in vitro assays have been developed to detect and assess stem cells and early multi-potential progenitors. While highly informative about primitive hematopoietic cells these assays are long and labour intensive. Alternatively, colony-forming cell (CFC) assays may be used to quantify more lineage-restricted progenitors in a simple in vitro assay. When cultured in a semi-solid medium containing the appropriate cytokines, CFCs are able to divide and differentiate into a colony of more mature cells that can be detected by light microscopy. This allows for the quantification of erythroid, myeloid, lymphoid, megakaryocytic, and multi-potential cell lineages from various cell sources. This chapter outlines the materials and methods used for the culture and assessment of CFC from humans, mice, and other species.

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References

  1. Gordon MY. (1993) Human haemopoietic stem cell assays. Blood Rev. 7(3): 190–197.

    Article  CAS  PubMed  Google Scholar 

  2. Pessina A, Albella B, Bayo M, Bueren J, Brantom P, Casati S, Croera C, Gagliardi G, Foti P, Parchment R, Parent-Massin D, Schoeters G, Sibiril Y, Van Den Heuvel R, Gribaldo L. (2003) Application of the CFU-GM assay to predict acute drug-induced neutropenia: an International Blind Trial to validate a prediction model for the maximum tolerated dose (MTD) of myelosuppressive xenobiotics. Toxicol. Sci. 75: 355–367.

    Article  CAS  PubMed  Google Scholar 

  3. McNiece I, Andrews R, Stewart M, Clarke S, Boone T, Quesenberry P. (1989) Action of interleukin-3, G-CSF, and GM-CSF on highly enriched human hematopoietic progenitor cells: synergistic interaction of GM-CSF plus G-CSF. Blood 74(1): 110–114.

    CAS  PubMed  Google Scholar 

  4. Bacigalupo A, Piaggio G, Podesta M, Figari O, Benvenuto F, Sogno G, Tedone E, Raffo MR, Grassia L, Ferrero R, et al. (1995) Influence of marrow CFU-GM content on engraftment and survival after allogeneic bone marrow transplantation. Bone Marrow Transplant. 15(2): 221–226.

    CAS  PubMed  Google Scholar 

  5. Torres A, Alonso MC, Gomez-Villagran JL, Manzanares MR, Martinez F, Gomez P, Garcia JM, Andres P, Gomez C, Torre MA, et al. (1985) No influence of number of donor CFU-GM on granulocyte recovery in bone marrow transplantation for acute leukemia. Blut 50(2): 89–94.

    Article  CAS  PubMed  Google Scholar 

  6. Larochelle A, Vormoor J, Hanenberg H, Wang JC, Bhatia M, Lapidot T, Moritz T, Murdoch B, Xiao XL, Kato I, Williams DA, Dick JE. (1996) Identification of primitive human hematopoietic cells capable of repopulating NOD/SCID mouse bone marrow: implications for gene therapy. Nat. Med. 2(12): 1329–1337.

    Article  CAS  PubMed  Google Scholar 

  7. Eaves C, Miller C, Cashman J, Conneally E, Petzer A, Zandstra P, Eaves A. (1997) Hematopoietic stem cells: inferences from in vivo assays. Stem Cells 15 (Suppl 1): 1–5.

    Article  Google Scholar 

  8. Bhatia M, Bonnet D, Murdoch B, Gan OI, Dick JE. (1998) A newly discovered class of human hematopoietic cells with SCID-repopulating activity. Nat. Med. 4(9): 1038–45.

    Article  CAS  PubMed  Google Scholar 

  9. Moore MA, Warren DJ. (1987) Synergy of interleukin 1 and granulocyte colony-stimulating factor: in vivo stimulation of stem-cell recovery and hematopoietic regeneration following 5-fluorouracil treatment of mice. Proc. Natl. Acad. Sci. U.S.A. 84(20): 7134–7138.

    Article  Google Scholar 

  10. Clarke E, Rice GC, Weeks RS, Jenkins N, Nelson R, Bianco JA, Singer JW. (1996) Lisofylline inhibits transforming growth factor beta release and enhances trilineage hematopoietic recovery after 5-fluorouracil treatment in mice. Cancer Res. 56(1): 105–112.

    Google Scholar 

  11. Helgason CD, Damen JE, Rosten P, Grewal R, Sorensen P, Chappel SM, Borowski A, Jirik F, Krystal G, Humphries RK. (1998) Targeted disruption of SHIP leads to hemopoietic perturbations, lung pathology, and a shortened life span. Genes Dev. 12(11): 1610–1620.

    Article  CAS  PubMed  Google Scholar 

  12. Stephenson JR, Axelrad AA, McLeod DL, Shreeve MM. (1971) Induction of colonies of hemaglobin-synthesising cells by erythropoietin in vitro. Proc. Natl. Acad. Sci. U.S.A. 68: 1542–1556.

    Article  CAS  PubMed  Google Scholar 

  13. Broxmeyer HE, Williams DE, Cooper S, Shadduch RK, Gillis S, Waheed A, Urdal DL, Bicknell DC. (1987) Comparative effects in vivo of recombinant murine interleukin 3, natural murine colony-stimulating factor-1, and recombinant murine granulocyte-macrophage colony-stimulating factor on myelopoiesis in mice. J Clin. Invest. 79(3): 721–30.

    Article  Google Scholar 

  14. Williams DE, Namen AE, Mochizuki DY, Overell RW. (1990) Clonal growth of murine pre-B colony-forming cells and their targeted infection by a retroviral vector: dependence on interleukin-7. Blood 75(5): 1132–1138.

    CAS  PubMed  Google Scholar 

  15. Banu N, Wang JF, Deng B, Groopman JE, Avraham H. (1995) Modulation of megakaryocytopoiesis by thrombopoietin: the c-Mpl ligand. Blood 86(4): 1331–1338.

    CAS  PubMed  Google Scholar 

  16. Sandmaier BM, Storb R, Santos EB, Krizanac-Bengez, Lian T, McSweeney PA, Yu C, Schuening FG, Deeg HJ, Graham T. (1996) Allogeneic transplant of canine peripheral blood stem cells mobilized by recombinant canine hematopoietic growth factors. Blood 87(8): 3508–3513.

    CAS  PubMed  Google Scholar 

  17. Moneta D, Geroni C, Valota O, Grossi P, de Jonge MJ, Brughera M, Colajori E, Ghielmini M, Sessa C. (2003) Predicting the maximum-tolerated dose of PNU-159548 (4-demethoxy-3-deamino-3-aziridinyl-4-methylsulphonyl-daunorubicin) in humans using CFU-GM clonogenic assays and prospective validation. Eur. J. Cancer. 39(5): 675–683.

    Article  Google Scholar 

  18. Masubuchi N. (2006) Risk assessment of human myelotoxicity of anticancer drugs: a predictive model and the in vitro colony forming unit granulocyte/macrophage (CFU-GM) assay. Pharmazie. Feb; 61(2): 135–9.

    Google Scholar 

  19. MacVittie TJ, Farese AM, Davis TA, Lind LB, McKearn JP. (1999) Myelopoietin, a chimeric agonist of human Interleukin 3 and granulocyte colony-stimulating factor receptors, mobilizes CD34+ cells that rapidly engraft lethally x-irradiated nonhuman primates. Exp. Hematol. 27: 1557–1568.

    Article  CAS  PubMed  Google Scholar 

  20. Tisdale JF, Hanazono Y, Sellers SE, Agricola BA, Metzger ME, Kato I, Donahue RE, Dunbar CE. (1998) Ex vivo expansion of genetically marked rhesus peripheral blood progenitor cells results in diminished long-term repopulating ability. Blood 92: 1131–1141.

    CAS  PubMed  Google Scholar 

  21. Farese AM, Herodin F, McKearn JP, Baurn C, Burton E, MacVittie TJ. (1996) Acceleration of hematopoietic reconstitution with a synthetic cytokine (SC-55494) after radiation-induced bone marrow aplasia. Blood 87: 581–591.

    CAS  PubMed  Google Scholar 

  22. Jonker M. (1990) The importance of non-human primates for preclinical testing of immunosuppressive monoclonal antibodies. Semin. Immunol. 2(6): 427–436.

    CAS  PubMed  Google Scholar 

  23. Pessina A, Malerba I, Gribaldo L. (2005) Hematotoxicity testing by cell clonogenic assay in drug development and preclinical trials. Curr. Pharm. Des. 11(8): 1055–1065.

    Article  Google Scholar 

  24. Ficoll-Paque® PLUS Instructions 71-7167-00AF, cell preparation, GE Healthcare, January 2006.

    Google Scholar 

  25. Nissen-Druey C, Tichelli A, Meyer-Monard S. (2005) Human hematopoietic colonies in health and disease. Reprint of Acta Haematoplogica (ISSN 0001-5792) 113(1) (STI cat. no. 28760).

    Google Scholar 

  26. Testa NG, Molineaux G. (ed.) (1993) Haematopoiesis: A Practical Approach. Oxford University Press, New York.

    Google Scholar 

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© 2007 Humana Press

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Pereira, C., Clarke, E., Damen, J. (2007). Hematopoietic Colony-Forming Cell Assays. In: Vemuri, M.C. (eds) Stem Cell Assays. Methods in Molecular Biology™, vol 407. Humana Press. https://doi.org/10.1007/978-1-59745-536-7_14

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  • DOI: https://doi.org/10.1007/978-1-59745-536-7_14

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-744-0

  • Online ISBN: 978-1-59745-536-7

  • eBook Packages: Springer Protocols

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