IDENTIFICATION OF DENDRITIC CELL COLONY-FORMING-UNITS AMONG NORMAL HUMAN CD34(-MARROW PROGENITORS THAT ARE EXPANDED BY C-KIT-LIGAND AND YIELD PURE DENDRITIC CELL COLONIES IN THE PRESENCE OF GRANULOCYTE-MACROPHAGE COLONY-STIMULATING FACTOR AND TUMOR-NECROSIS-FACTOR-ALPHA() BONE)

Several cytokines, especially granulocyte/macrophage colony-stimulatin g factor (GM-CSF) and tumor necrosis factor alpha (TNF-alpha), have be en identified that foster the development of dendritic cells from bloo d and bone marrow precursors in suspension cultures. These precursors are reported to be infrequent or to yield small numbers of dendritic c ells in colony-forming assays. Here we readily identify dendritic cell colony-forming units (CFU-DC) that give rise to pure dendritic cell c olonies. Human CD34(+) bone marrow progenitors were expanded in semiso lid cultures with serum-replete medium containing c-kit-ligand, GM-CSF , and TNF-alpha. The addition of TNF-a to GM-CSF did not alter the num ber of typical GM colonies but did generate pure dendritic cell coloni es that accounted for similar to 40% of the total colony growth. When the two distinct types of colonies were plucked from methylcellulose a nd tested for T cell-stimulatory activity in the mixed leukocyte react ion, the potency of colony-derived dendritic cells exceeded that of CF U-GM progeny from the same cultures by at least 1.5-2 logs. Immunophen otyping and cytochemical staining of the CFU-DC-derived progeny was al so characteristic of dendritic cells. Other myeloid cells were not ide ntified in these colonies. The addition of c-kit-ligand to GM-CSF- and TNF-alpha-supplemented suspensions of CD34(+) bone marrow cells expan ded CFU-DCs almost 100-fold by 14 d. We conclude that normal human CD3 4(+) bone marrow cells include substantial numbers of clonogenic proge nitors, distinct from CFU-GMs, that can give rise to pure dendritic ce ll colonies. These CFU-DCs can be expanded for several weeks by in vit ro culture with c-kit-ligand, and their differentiation requires exoge nous TNF-alpha in addition to GM CSF. We speculate that this dendritic cell-committed pathway may in the steady state contribute cells to th e epidermis and afferent lymph, where dendritic cells are the principa l myeloid cell type, and may increase the numbers of these specialized antigen-presenting cells during T cell-mediated immune responses.