Long-term ex vivo maintenance and expansion of transplantable human hematopoietic stem cells

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 a's 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 strom al cell line, caused expansion of cells with CD34(+) thy-1(+) phenotype. Ad dition of other cytokines, including interleukin-3 (IL-3), IL-6, granulocyt e-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 mult ilineage differentiation, including myeloid, T; and B Cells, when transplan ted 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, su ggesting action through the murine stroma. Furthermore, another human HSC c andidate, CD34(+) CD38(-) cells, shows a similar pattern of proliferative r esponse. This suggests that ex vivo expansion of transplantable human stem cells under this in vitro culture system is a general phenomenon and not ju st specific for CD34(+) thy-1(+) cells. (C) 1999 by The American Society of Hematology.