A murine stromal cell line promotes the expansion of CD34high+-primitive progenitor cells isolated from human umbilical cord blood in combination with human cytokines.
The in vitro expansion of CD34+ cells is important for clinical applications such as transplantation and gene therapy with CD34+ cells isolated from human umbilical cord blood. In the present study, we developed a xenogenic coculture system involving HUCB-CD34+ cells and a murine stromal cell line, HESS-5 cells, in the presence of recombinant human (rh) cytokines. We examined the effects of combinations of cytokines, such as rh-IL-3, rh-SCF, rh-granulocyte colony-stimulating factor (G-CSF), rh-granulocyte-macrophage-CSF and h-erythropoietin (EPO), on the expansion of CD34high+ cells and colony-forming progenitor cells (CFCs). The proliferation of CD34high+ cells and CFCs was dramatically promoted on coculture with HESS-5 cells, and the expansion ratio of the CD34high+ cells showed good correlation with that of high-proliferative potential colony-forming cells (HPP-CFCs). The most potent combination of cytokines in this xenogenic coculture system for the expansion of CD34high+ cells and HPP-CFCs was rh-IL-3 and rh-SCF. The proliferation of CD34high+ cells was supported in the presence of HESS-5 cells with direct cell contact, but not observed in the indirect coculture involving a microporous membrane. Furthermore, we developed a unique coculture method, designated as the bilayer coculture method, involving CD34+ cells and HESS-5 cells using a microporous membrane. This expansion system will be applicable to the expansion of the primitive progenitor cells of HUCB-CD34+ cells and is worthy of consideration for the clinical application of HUCB-CD34+ cells.
Tsuji
T
T
Division of Hematology, Pharmaceutical Frontier Research Laboratories, JT Inc., Yokohama, Kanagawa, Japan. tsuji@ikrl.jti.co.jp
Nishimura-Morita
Y
Y
Watanabe
Y
Y
Hirano
D
D
Nakanishi
S
S
Mori
K J
KJ
Yatsunami
K
K
D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism*; D018929:Cell Culture Techniques / Q000379:methods*; D002460:Cell Line; D016207:Cytokines / Q000378:metabolism*; D005312:Fetal Blood / Q000378:metabolism*; D005434:Flow Cytometry; D006412:Hematopoietic Stem Cells / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D051379:Mice; D008855:Microscopy, Electron, Scanning; D008954:Models, Biological; D013234:Stem Cells; D017154:Stromal Cells / Q000378:metabolism*
D018952:Antigens, CD34; D016207:Cytokines