Expansion of human and murine hematopoietic stem and progenitor cells ex vivo without genetic modification using MYC and Bcl-2 fusion proteins

• Published in: PloS one Vol. 9; no. 8; p. e105525
• Main Authors: Bird, Gregory A, Polsky, Avital, Estes, Patricia, Hanlon, Teri, Hamilton, Haley, Morton, John J, Gutman, Jonathan, Jimeno, Antonio, Turner, Brian C, Refaeli, Yosef
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 29.08.2014; Public Library of Science (PLoS)
• Subjects: Adult; Animals; Bcl-2 protein; Biology; Biology and Life Sciences; Blood; Bone marrow; Bone marrow transplantation; Bone Marrow Transplantation - methods; Cell Culture Techniques - methods; Cell Differentiation - drug effects; Cell Proliferation - drug effects; Cells (biology); Cells, Cultured; Cord blood; Dermatology; Fetal Blood - cytology; Fusion protein; Genetic modification; Granulocyte colony-stimulating factor; Hematopoiesis; Hematopoietic stem cells; Hematopoietic Stem Cells - cytology; Humans; Hydrocarbons; Interleukin Receptor Common gamma Subunit - deficiency; Interleukin Receptor Common gamma Subunit - genetics; Lymphoma; Medicine; Medicine and Health Sciences; Methylcellulose; Mice, Inbred C57BL; Mice, Inbred NOD; Mice, Knockout; Mice, SCID; Myc protein; Oncology; Peripheral blood; Populations; Progenitor cells; Proteins; Proto-Oncogene Proteins c-bcl-2 - genetics; Proto-Oncogene Proteins c-bcl-2 - metabolism; Proto-Oncogene Proteins c-bcl-2 - pharmacology; Proto-Oncogene Proteins c-myc - genetics; Proto-Oncogene Proteins c-myc - metabolism; Proto-Oncogene Proteins c-myc - pharmacology; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Recombinant Fusion Proteins - pharmacology; Reproducibility of Results; Rodents; Stem cell transplantation; Stem cells; Stem Cells - cytology; Surface markers; Taiga & tundra; Tat protein; Transplantation; United States > US; Colorado
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0105525

The long-term repopulating hematopoietic stem cell (HSC) population can self-renew in vivo, support hematopoiesis for the lifetime of the individual, and is of critical importance in the context of bone marrow stem cell transplantation. The mechanisms that regulate the expansion of HSCs in vivo and in vitro remain unclear to date. Since the current set of surface markers only allow for the identification of a population of cells that is highly enriched for HSC activity, we will refer to the population of cells we expand as Hematopoietic Stem and Progenitor cells (HSPCs). We describe here a novel approach to expand a cytokine-dependent Hematopoietic Stem and Progenitor Cell (HSPC) population ex vivo by culturing primary adult human or murine HSPCs with fusion proteins including the protein transduction domain of the HIV-1 transactivation protein (Tat) and either MYC or Bcl-2. HSPCs obtained from either mouse bone marrow, human cord blood, human G-CSF mobilized peripheral blood, or human bone marrow were expanded an average of 87 fold, 16.6 fold, 13.6 fold, or 10 fold, respectively. The expanded cell populations were able to give rise to different types of colonies in methylcellulose assays in vitro, as well as mature hematopoietic populations in vivo upon transplantation into irradiated mice. Importantly, for both the human and murine case, the ex vivo expanded cells also gave rise to a self-renewing cell population in vivo, following initial transplantation, that was able to support hematopoiesis upon serial transplantation. Our results show that a self-renewing cell population, capable of reconstituting the hematopoietic compartment, expanded ex vivo in the presence of Tat-MYC and Tat-Bcl-2 suggesting that this may be an attractive approach to expand human HSPCs ex vivo for clinical use.