Induction of c-kit molecules on human CD34+/c-kit < low cells: evidence for CD34+/c-kit < low cells as primitive hematopoietic stem cells.
c-kit, a receptor for stem cell factor, has been widely accepted as a distinctive marker for hematopoietic stem cells. However, the level of c-kit expression on pluripotent hematopoietic stem cells is still controversial in mice and humans. We purified CD34+/c-kit < low cells (phenotypically c-kit-negative but only detectable at the message level) from human cord blood and examined their maturational steps in relation to the expression of c-kit molecules. When the CD34+/c-kit < low cells were cultured with cytokines (flt 3 ligand, interleukin 6 and interleukin 7) plus immobilized anti-CD34 monoclonal antibody (to crosslink CD34 molecules), c-kit molecules were clearly induced within 24 h. The c-kit expression gradually increased until day 8. When CD34+/c-kit(low) or CD34+/c-kit+ cells that had been induced from CD34+/c-kit < low cells were resorted and recultured using a methylcellulose culture system, they showed the same colony-forming ability as the freshly isolated CD34+/c-kit(low) or CD34+/c-kit+ cells, respectively. Furthermore, CD34+/c-kit < low cells have a similar hematopoietic potential to CD34+/c-kit(low) cells in assays for long-term culture initiating cell and colony-forming unit culture generated from long-term cultures. These findings suggest that CD34+/c-kit < low cells mature into CD34+/c-kit(low) and CD34+/c-kit+ cells, and acquire the reactivity to various humoral hematopoietic stimuli. Moreover, CD34+/c-kit < low cells showed a low level of rhodamine 123 retention, suggesting that CD34+/c-kit < low cells have multidrug resistance. Therefore, the CD34+/c-kit < low cells without colony-forming unit-granulocyte-erythroid-macrophage-megakaryocyte activity are also a pluripotent hematopoietic stem cell population, and the expression of c-kit on c-kit < low cells is the first maturational step of hematopoiesis.
Sogo
S
S
Cellular Technology Institute, Otsuka Pharmaceutical Co., Ltd., Tokushima, Japan.
Inaba
M
M
Ogata
H
H
Hisha
H
H
Adachi
Y
Y
Mori
S
S
Toki
J
J
Yamanishi
K
K
Kanzaki
H
H
Adachi
M
M
Ikehara
S
S
D018952:Antigens, CD34 / Q000276:immunology*; D002469:Cell Separation; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D005456:Fluorescent Dyes / Q000378:metabolism; D006410:Hematopoiesis; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D006801:Humans; D019009:Proto-Oncogene Proteins c-kit / Q000096:biosynthesis* / Q000235:genetics; D012333:RNA, Messenger; D020112:Rhodamine 123; D012235:Rhodamines / Q000378:metabolism
D018952:Antigens, CD34; D005456:Fluorescent Dyes; D012333:RNA, Messenger; D012235:Rhodamines; D020112:Rhodamine 123; D019009:Proto-Oncogene Proteins c-kit