DIFFERENTIATION OF HUMAN DENDRITIC CELLS FROM MONOCYTES IN-VITRO

Since either macrophages (M Phi) or dendritic cells (DC) differentiate from monocytes (MO) depending on culture conditions, we investigated the relationship of the DC and M Phi differentiation pathways. Culturi ng MO-enriched blood mononuclear cells with M Phi colony-stimulating f actor (M-CSF) or with granulocyte/M Phi (GM)-CSF induced M Phi with a different morphology and CD14/CD1a expression. In contrast, in culture s with GM-CSF and interleukin (IL)-4, cells rapidly became nonadherent and acquired DC morphology, ultrastructure, CD1a expression, and most DC markers; they lost membrane CD14 and CD64 and capacity of phagocyt osis, displayed less CD68 than M Phi but retained non-specific esteras e activity. These DC directly developed from MO without proliferation inasmuch as only day 0 FACS-sorted MO, but not small CD14(-) cells, di fferentiated into DC when cultured with GM-CSF and IL-4, or to M Phi, with M-CSF. While overall cell numbers declined, DC numbers plateaued from culture day 2 onwards, indicating that most had differentiasted b y then. This differentiation was radioresistant and occurred without [ H-3]thymidine incorporation. Commitment to differentiate into DC with GM-CSF and IL-4 was irreversible by day 2, since discontinuing IL-4 at this point did not revert cells to M Phi. Alternatively, cells rapidl y converted to DC when IL-4 was added from day 2 to cultures initiated with GM-CSF only. If cultures were initiated with M-CSF and switched to GM-CSF and IL-4 after 2 or 5 days, about half of the cells still co nverted to DC. Thus, the capacity of MO and even of M Phi to different iate into DC was conserved for at least this period. The increased cap acity to stimulate the mixed leukocyte reaction correlated with the re lative number of CD1a(+) cells at any time and under each condition te sted, a confirmation that these cells functionally qualify as DC. Thus , MO and even M Phi can be directed to differentiate into DC depending on the cytokine microenvironment.