Establishment of mouse embryonic fibroblast cell lines that promote ex vivo expansion of human cord blood CD34+ hematopoietic progenitors.
The development of culture systems that facilitate ex vivo maintenance and expansion of transplantable hematopoietic progenitor cells (HPC) is vital to stem cell transplantation. The use of a monolayer of stromal cells on which to grow HPC in direct contact allows high efficiency ex vivo expansion of HPC. Here, we report an establishment of three murine embryonic fibroblast stromal cell lines from adherent cells of day-12 mouse embryos. Among them, HYMEQ-5 was most efficient in supporting long-term maintenance of human umbilical cord blood (CB) CD34(+) cells. Human CB CD34(+) cells cultured on HYMEQ-5 in the presence of stem cell factor (SCF), thrombopoietin, and flk-ligand (FL) showed high expansion of CD34(+)CD38(-) cells and highly proliferative potential-colony forming cells (HPP-CFC). Direct cell-to-cell contact between CD34(+) cells and HYMEQ-5 was important for this expansion. RT-PCR analysis showed that HYMEQ-5 produced FL, SCF, interleukin-6, and macrophage colony-stimulating factor (M-CSF). Expanded CB CD34(+) cells efficiently reconstituted hematopoiesis in nonobese diabetic/severe combined immunodeficient disease (NOD/SCID) mice. These findings suggest that HYMEQ-5 provides a milieu that supports long-term human hematopoiesis as well as ex vivo expansion of human CB CD34(+) HPC. This cell line may facilitate elucidation of the mechanism of cellular interactions between HPC and stromal cells.
Qiu
Huiying
H
Department of Transfusion Medicine, Hyogo College of Medicine, Nishinomiya, Japan.
Fujimori
Yoshihiro
Y
Kai
Shunro
S
Fujibayashi
Yuka
Y
Nishioka
Keisuke
K
Hara
Hiroshi
H
D000818:Animals; D018952:Antigens, CD34 / Q000096:biosynthesis*; D002455:Cell Division; D002460:Cell Line; D002478:Cells, Cultured; D016207:Cytokines / Q000096:biosynthesis / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000378:metabolism*; D005434:Flow Cytometry; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D006801:Humans; D015850:Interleukin-6 / Q000378:metabolism; D016173:Macrophage Colony-Stimulating Factor / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016513:Mice, SCID; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019089:Stem Cell Factor / Q000378:metabolism; D033581:Stem Cell Transplantation; D013234:Stem Cells; D013997:Time Factors
D018952:Antigens, CD34; D016207:Cytokines; D015850:Interleukin-6; D019089:Stem Cell Factor; D016173:Macrophage Colony-Stimulating Factor