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Leukemic Stem Cell Culture in Cytokine-Free Medium

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Part of the Methods in Molecular Biology book series (MIMB,volume 2185)

Abstract

Leukemia-initiating cells, also known as leukemic stem cells (LSCs), are experimentally defined by their ability to engraft immunocompromised mice and are believed to be a major cause of relapse in acute myeloid leukemia (AML). Despite the aggressive characteristics of acute leukemia, AML blasts are difficult to culture once removed from the patient, and LSCs, which are a minor fraction of the blast population, are especially difficult to transplant after culture. This impedes development of new therapies for AML that target LSCs. Here, we present a simple strategy to culture LSCs in cytokine-free medium and to perform flow cytometric analysis of the resulting cell population for the characterization of LSCs maintenance and differentiation.

Key words

  • Acute myeloid leukemia (AML)
  • Leukemia-initiating cells
  • Leukemic stem cells
  • Cytokine-free culture medium
  • GSK-3
  • mTOR
  • Flow cytometry

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  • DOI: 10.1007/978-1-0716-0810-4_15
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References

  1. Pearce DJ, Taussig D, Zibara K, Smith LL, Ridler CM, Preudhomme C, Young BD, Rohatiner AZ, Lister TA, Bonnet D (2006) AML engraftment in the NOD/SCID assay reflects the outcome of AML: implications for our understanding of the heterogeneity of AML. Blood 107(3):1166–1173. https://doi.org/10.1182/blood-2005-06-2325

    CAS  CrossRef  PubMed  PubMed Central  Google Scholar 

  2. Woiterski J, Ebinger M, Witte KE, Goecke B, Heininger V, Philippek M, Bonin M, Schrauder A, Rottgers S, Herr W, Lang P, Handgretinger R, Hartwig UF, Andre MC (2013) Engraftment of low numbers of pediatric acute lymphoid and myeloid leukemias into NOD/SCID/IL2Rcgammanull mice reflects individual leukemogenecity and highly correlates with clinical outcome. Int J Cancer 133(7):1547–1556. https://doi.org/10.1002/ijc.28170

    CAS  CrossRef  PubMed  Google Scholar 

  3. van Gosliga D, Schepers H, Rizo A, van der Kolk D, Vellenga E, Schuringa JJ (2007) Establishing long-term cultures with self-renewing acute myeloid leukemia stem/progenitor cells. Exp Hematol 35(10):1538–1549. https://doi.org/10.1016/j.exphem.2007.07.001

    CAS  CrossRef  PubMed  Google Scholar 

  4. Klco JM, Spencer DH, Lamprecht TL, Sarkaria SM, Wylie T, Magrini V, Hundal J, Walker J, Varghese N, Erdmann-Gilmore P, Lichti CF, Meyer MR, Townsend RR, Wilson RK, Mardis ER, Ley TJ (2013) Genomic impact of transient low-dose decitabine treatment on primary AML cells. Blood 121(9):1633–1643. https://doi.org/10.1182/blood-2012-09-459313

    CAS  CrossRef  PubMed  PubMed Central  Google Scholar 

  5. Griessinger E, Anjos-Afonso F, Pizzitola I, Rouault-Pierre K, Vargaftig J, Taussig D, Gribben J, Lassailly F, Bonnet D (2014) A niche-like culture system allowing the maintenance of primary human acute myeloid leukemia-initiating cells: a new tool to decipher their chemoresistance and self-renewal mechanisms. Stem Cells Transl Med 3(4):520–529. https://doi.org/10.5966/sctm.2013-0166

    CAS  CrossRef  PubMed  PubMed Central  Google Scholar 

  6. Pabst C, Krosl J, Fares I, Boucher G, Ruel R, Marinier A, Lemieux S, Hebert J, Sauvageau G (2014) Identification of small molecules that support human leukemia stem cell activity ex vivo. Nat Methods 11(4):436–442. https://doi.org/10.1038/nmeth.2847

    CAS  CrossRef  PubMed  Google Scholar 

  7. Bhavanasi D, Wen KW, Liu X, Vergez F, Danet-Desnoyers G, Carroll M, Huang J, Klein PS (2017) Signaling mechanisms that regulate ex vivo survival of human acute myeloid leukemia initiating cells. Blood Cancer J 7(12):636. https://doi.org/10.1038/s41408-017-0003-1

    CrossRef  PubMed  PubMed Central  Google Scholar 

  8. Huang J, Nguyen-McCarty M, Hexner EO, Danet-Desnoyers G, Klein PS (2012) Maintenance of hematopoietic stem cells through regulation of Wnt and mTOR pathways. Nat Med 18(12):1778–1785. https://doi.org/10.1038/nm.2984

    CAS  CrossRef  PubMed  PubMed Central  Google Scholar 

  9. Roederer M (2001) Spectral compensation for flow cytometry: visualization artifacts, limitations, and caveats. Cytometry 45(3):194–205

    CAS  CrossRef  PubMed  Google Scholar 

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Correspondence to Peter S. Klein .

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Liu, X., Klein, P.S. (2021). Leukemic Stem Cell Culture in Cytokine-Free Medium. In: Cobaleda, C., Sánchez-García, I. (eds) Leukemia Stem Cells. Methods in Molecular Biology, vol 2185. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0810-4_15

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  • DOI: https://doi.org/10.1007/978-1-0716-0810-4_15

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0809-8

  • Online ISBN: 978-1-0716-0810-4

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