Distinctive features of in vitro differentiated (tolerogenic) DC-10. (A) Protocol for in vitro differentiation of DC-10. Peripheral blood monocytes are cultured for 7 days in the presence of granulocyte–macrophage colony-stimulating factor (GM-CSF), IL-4 and IL-10. (B) DC-10 express high levels of HLA-G and immunoglobulin-like transcript (ILT) 4. DC-10 were analyzed by flow cytometry to determine the levels of expression of ILT2, ILT3, ILT4 and HLA-G. Percentages of immature dendritic cells (DCs) and DC-10 expressing the tolerogenic molecules ILT2, ILT3, ILT4 and HLA-G (mean ± standard error, n = 16) are shown. (C) Cytokine secretion profile of immature DCs and DC-10 left unstimulated (upper panel) or stimulated with lipopolysaccharide and IFNγ (lower panel). Culture supernatants were collected after 48 hours and levels of cytokines were measured by ELISA (mean ± standard deviation, n = 16). As control, immature DCs differentiated by culturing monocytes with GM-CSF and IL-4 for 7 days were used. *P < 0.05; **P < 0.005; ***P < 0.0005; ****P < 0.0001.