Lymphoid and myeloid differentiation of single human CD34+, HLA-DR+, CD38- hematopoietic stem cells.
Multilineage differentiation of human fetal bone marrow CD34+ cell subsets was examined using a single-cell liquid culture assay. Four CD34+ cell populations, ie, (1) CD38-, HLA-DR+, (2) CD38-, HLA-DR-, (3) CD38+, HLA-DR-, and (4) CD38+, HLA-DR+ cells, were sorted as single cells into 96-well flat-bottom culture plates containing long-term culture medium supplemented with interleukin-3, interleukin-6, stem cell factor (SCF), granulocyte-macrophage colony-stimulating factor, erythropoietin, basic fibroblast growth factor (bFGF), and insulin-like growth factor-1 (IGF-1). Single CD34+, CD38-, HLA-DR+ cells had the highest replating efficiency as well as the highest replating efficiency. The cellular composition of the single-cell progeny was studied by morphologic and/or flow cytometric examination. Only the progeny of single CD34+ cells that lacked CD38 could give rise to each of the hematopoietic cell lineages. The expansion of the progeny of single CD34+, CD38-, HLA-DR+ cells was examined in more detail and showed three clearly distinguishable growth patterns: 28% (SD, +/- 10%; n = 14) of the single cells formed cell clusters/colonies; 9% (SD, +/- 4%; n = 14) formed dispersed cells; and 11% (SD, +/- 6%; n = 14) gave rise to a mixture of cell clusters and dispersed cells. The dispersed cell growth pattern was reduced when SCF or bFGF and IGF-1 was absent in the growth factor cocktail. The replating ability of the dispersed cells was considerably larger than that of cells with other growth patterns, in that 76% of the cells that gave rise to dispersed cells and 54% of the cells that gave rise to dispersed cells as well as cell clusters gave rise to a second generation, but only 7% of the cells that gave rise to cell clusters gave rise to a second generation. The second generation of cells continued to produce third and fourth generations after repetitive replating, except for the replated cells from cell clusters. In contrast with the first-generation progeny, SCF did not have an influence on the replating ability of the cells. Only in the progeny of single CD34+, CD38-, HLA-DR+ cells that gave rise to dispersed cells was each of the hematopoietic cell lineages found, ie, B lymphocytes, neutrophils, monocytes, macrophages, osteoclasts, basophils/mast cells, eosinophils, erythrocytes, megakaryocytes, and platelets.(ABSTRACT TRUNCATED AT 400 WORDS)
Huang
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Becton Dickinson Immunocytometry Systems, San Jose, CA 95131.
Terstappen
L W
LW
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D015703:Antigens, CD / Q000032:analysis*; D018952:Antigens, CD34; D000943:Antigens, Differentiation / Q000032:analysis*; D001854:Bone Marrow Cells; D002454:Cell Differentiation; D005333:Fetus / Q000166:cytology; D006684:HLA-DR Antigens / Q000032:analysis*; D006412:Hematopoietic Stem Cells / Q000276:immunology / Q000502:physiology*; D006801:Humans; D008562:Membrane Glycoproteins
D015703:Antigens, CD; D018952:Antigens, CD34; D000943:Antigens, Differentiation; D006684:HLA-DR Antigens; D008562:Membrane Glycoproteins; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D051997:ADP-ribosyl Cyclase 1